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448 publications mentioning hsa-mir-34c (showing top 100)

Open access articles that are associated with the species Homo sapiens and mention the gene name mir-34c. Click the [+] symbols to view sentences that include the gene name, or the word cloud on the right for a summary.

1
[+] score: 385
Other miRNAs from this paper: hsa-mir-34a, hsa-mir-34b
Western blot analysis (Figure 2 A ) showed that transfection of miR-34 mimics downregulated expression of target genes, Bcl-2, Notch1 and Notch2 at the protein level, but had no effect on Bcl-xL and Mcl-1 expression, indicating the target gene knock-down by miR-34 mimics affects transcripts harbouring miR-34 target sites. [score:15]
Interestingly, the Notch2 expression inhibition at the protein level by miR-34 was not accompanied by inhibition at the mRNA level, in agreement with previous reports that miRNA inhibits target gene expression post-transcriptionally, with or without mRNA degradation [11], [20]. [score:13]
Our results show that miR-34 restoration in human pancreatic cancer MiaPaCa2 and BxPC3 cells inhibited the expression of target genes, Bcl-2, Notch1 and Notch2; significantly inhibited clonogenic cell growth and invasion; induced apoptosis and G1 and G2/M arrest; and sensitized the cells to chemotherapy and radiation. [score:9]
Restoration of miR-34 down-regulates target genes' expression. [score:8]
This strategy was explored in the current study, where p53 downstream target miR-34 was restored in p53-mutant pancreatic cancer MiaPaCa2 cells with a high level of Bcl-2 and low levels of miR-34s, resulting in downregulation of Bcl-2 and Notch1-2, together with the inhibited clonogenic cell growth and invasion; increased apoptosis and G1 and G2/M arrest in cell cycle; and sensitization to chemotherapy and radiation. [score:8]
Our results are consistent with the notion that Bcl-2 is a direct target of miR-34 and miR-34 potently inhibits Bcl-2 expression. [score:8]
This multi-mode action of miR-34 provides a therapeutic advantage over the siRNA -based therapies in that miR-34 has multiple targets, can work on multiple cell signalling pathways at the same time, leading to synergistic effects which may translate into improved clinical efficacy for pancreatic cancer patients with p53 deficiency and advanced disease. [score:7]
Figure 2 B shows the qRT-PCR analysis of the potential target genes; miR-34 mimics potently inhibited BCL2 and Notch1 gene expression, consistent with the Western blot data. [score:7]
Our results demonstrate that miR-34 is involved in MiaPaCa2 cell growth; miR-34 restoration inhibits the clonogenic growth, and inhibition of endogenous miR-34 by miR-34 inhibitors promotes the growth. [score:7]
We have recently shown that expression of miR-34s is dramatically reduced in p53-mutant gastric cancer cells and that the restoration of miR-34 expression inhibited the cancer cell growth [6]. [score:7]
These data indicate that the CD44+/CD133+ cells were the target cells of miR-34, i. e., miR-34 exerts its tumor-suppressing activity via inhibiting the CD44+/CD133+ cells. [score:7]
However, our study is the first report showing that miRNA miR-34 inhibits pancreatic CD44+/CD133+ CSC, potentially via inhibiting downstream target “stem cell genes” such as Notch and Bcl-2. Interestingly, miR-34a and miR-34b are among the short-list of the stem cell-specific miRNAs discovered by Dr. [score:7]
A, miR-34 restoration down-regulates target proteins Bcl-2, Notch1 and Notch2, no effects on Mcl-1. MiaPaCa2 cells were transfected with miR-34 mimics or non-specific control miRNA mimic (NC mimic) (100 pmol per well in 6-well plates by Lipofectamine 2000) for 48 hours, then collected for Western blot analysis. [score:6]
0006816.g002 Figure 2 A, miR-34 restoration down-regulates target proteins Bcl-2, Notch1 and Notch2, no effects on Mcl-1. MiaPaCa2 cells were transfected with miR-34 mimics or non-specific control miRNA mimic (NC mimic) (100 pmol per well in 6-well plates by Lipofectamine 2000) for 48 hours, then collected for Western blot analysis. [score:6]
Restoration of miR-34 expression in the pancreatic cancer cells by either transfection of miR-34 mimics or infection with lentiviral miR-34-MIF downregulated Bcl-2 and Notch1/2. [score:6]
The report from He et al. indicated that ectopic expression of miR-34 induces cell cycle arrest in both primary and tumor-derived cell lines, which is consistent with the ability of miR-34 to downregulate a program of genes promoting cell cycle progression [11]. [score:6]
B, qRT-PCR analysis shows that the target gene Bcl-2 is downregulated in miR-34-MIF clone. [score:6]
Recently, the three members of the miR-34 family were found to be directly regulated by p53 and the functional activity of miR-34 indicated a potential role as a tumor suppressor [6], [7], [8], [9], [10], [11], [12]. [score:5]
MiaPaCa2 and BxPC3 cells have very low expression levels of both primary and mature miR-34a,b,c but high levels of the miR-34 target genes BCL2 and Notch1, and different levels of Notch2–4 (Figure 1 ). [score:5]
We have also shown that the CD44+/CD133+ tumorsphere-forming and tumor-initiating cells have high Bcl-2 and loss of miR-34 expression, indicating that miR-34 and its target Bcl-2 might be involved in cancer stem cells. [score:5]
qRT-PCR was performed to determine the expression levels of potential miR-34 target genes [6]. [score:5]
Restoration of miR-34 inhibits the clonogenic growth of MiaPaCa2 cells, whereas inhibition of miR-34 promotes cell growth. [score:5]
We also assessed in parallel the expression of presumptive miR-34-regulated target genes and proteins, using the primers and methods as we described recently [6]. [score:5]
The miR-34 -mediated reduction of the CD44+/CD133+ CSC is associated with the potent and simultaneous inhibition of its downstream target genes Notch and Bcl-2, genes involved in stem cells self-renewal and survival, so-called “stem cell genes” or “stemness genes” [6], [41], [42], [43]. [score:5]
C, qRT-PCR analysis of the expression levels of miR-34 target genes in human pancreatic cancer cell lines as well as normal human fibroblast WI-38 cells. [score:5]
More significantly, miR-34 restoration inhibits the CD44+/CD133+ tumor-initiating cells or CSC, accompanied by significant inhibition of tumorsphere growth in vitro and tumor formation in vivo. [score:5]
The transfected miR-34 mimics inhibited the luciferase reporter gene expression, which is controlled by Bcl-2 3′UTR in the promoter region (Figure 2 C ). [score:5]
Our above results have shown that miR-34 potently inhibits Bcl-2 expression and cell growth and increases cell death and response to chemo-/radiotherapy in the overall population of MiaPaCa-2 cells. [score:5]
Here again, miR-34 shows the advantage of its multi-target potential, as both stem cell genes Notch and Bcl-2 are inhibited by miR-34 at the same time, a potent synergy may be achieved in blocking both Notch signalling pathway and the anti-apoptotic function of Bcl-2 in tumor-initiating cells or cancer stem cells. [score:5]
We also examined the effect of inhibition of endogenous miR-34 on cell growth by mi RIDIAN miR-34 inhibitors. [score:5]
Next, we carried out qRT-PCR analysis of the sorted MiaPaCa2 cells to assess whether there is any difference in these populations as to the expression levels of miR-34 and its target genes. [score:5]
As shown in Figure 3 A–B, miR-34 restoration significantly inhibited the clonogenic cell growth, with miR-34a mimic inducing >80% inhibition of colony formation compared to NC mimic (18.3±3.8 colonies/well vs. [score:4]
Our data support the view that miR-34 may be involved in pancreatic cancer stem cell self-renewal, potentially via the direct modulation of downstream targets Bcl-2 and Notch, implying that miR-34 may play an important role in pancreatic cancer stem cell self-renewal and/or cell fate determination. [score:4]
Our data support the view that miR-34 may be involved in pancreatic cancer stem cell self-renewal, potentially via the direct modulation of downstream targets Bcl-2 and Notch, implying that miR-34 may play an important role in pancreatic cancer stem cell self-renewal and/or cell fate determination, at least in the p53-mutant MiaPaCa2 mo del. [score:4]
Among the target proteins regulated by miR-34 are Notch pathway proteins and Bcl-2, suggesting the possibility of a role for miR-34 in the maintenance and survival of cancer stem cells. [score:4]
Our data provide the first evidence that miR-34 is involved in pancreatic CSC self-renewal, potentially via the direct modulation of downstream targets Notch and Bcl-2. Our results provide novel insight into how miR-34 works in pancreatic cancer cells with p53 loss of function. [score:4]
The results demonstrate that the transfected miR-34s are functional and confirm that Bcl-2 is a direct target of miR-34, consistent with earlier reports [8], [10], [21]. [score:4]
In addition, because more than 50% of primary human cancers have mutations inactivating p53 function, the findings provided impetus to explore the functional restoration of miR-34 as a novel approach to inhibit cancers with p53 loss-of-function. [score:4]
CD44+/CD133+ cells are tumorsphere-forming and tumor-initiating cells with high Bcl-2 and loss of miR-34 expression. [score:3]
There is an inverse correlation in the expression levels of miR-34 and Bcl-2 in Q2 versus Q3, e. g., Q2 cells (with enriched cancer stem cells) have high Bcl-2 and low miR-34, Q3 cells (non-tumorigenic cells) have low Bcl-2 and high miR-34 levels. [score:3]
miR-34 restoration inhibits the MiaPaCa2 tumor formation in nude mice. [score:3]
In conclusion, our study demonstrates that miR-34 may restore, at least in part, the tumor suppressing function of p53 in p53 -deficient human pancreatic cancer cells. [score:3]
The miR-34 expression data were normalized to that of Actin and the relative levels are shown (set unsorted cells = 1). [score:3]
miR-34 restoration leads to a significant reduction of CD44+/CD133+ cells and inhibition of tumorsphere growth. [score:3]
Our results show that miR-34 restoration caused an 87% reduction of the CD44+/CD133+ tumorsphere-forming and tumor-initiating CSC in MiaPaCa2 cells with p53 loss of function, accompanied by a significant inhibition of tumorsphere growth in vitro and tumor formation in vivo. [score:3]
By modulating CSC, the restoration of tumor suppressor miR-34 may provide a novel therapeutic approach for p53 -deficient pancreatic cancer. [score:3]
We are currently carrying out more detailed mechanism studies to delineate the involvement of Notch signaling pathway in miR-34 -induced inhibition of pancreatic CSC and its role in chemo/radiosensitization of pancreatic cancer with p53 loss of function. [score:3]
More significantly, we show that miR-34 restoration led to an 87% reduction of the CD44+/CD133+ CSC, accompanied by significant inhibition of tumorsphere growth in vitro as well as tumor formation in vivo. [score:3]
mi RIDIAN miRNA miR34a,b,c mimics and negative control miRNA mimic (NC mimic), mi RIDIAN miR-34 inhibitors and negative controls were obtained from Dharmacon (Chicago, IL) [6]. [score:3]
Our data are consistent with the reported tumor suppressor function of miR-34 [6], [8], [9], [11], [22]. [score:3]
miR-34 restoration inhibits the MiaPaCa2 tumor initiation in nude mice. [score:3]
We previously reported that the Bcl-2 protein is regulated directly by miR-34 [10]. [score:3]
Our data provide the first evidence that miR-34 is able to inhibit CD44+/CD133+ tumorsphere-forming and tumor-initiating cancer stem cells in p53-mutant pancreatic cancer, implying that miR-34 might play a role in the self-renewal of pancreatic cancer stem cells. [score:3]
miR-34 restoration inhibits MiaPaCa2 cell clonogenic growth and leads to caspase-3 activation and apoptosis. [score:3]
Twenty-four hr after miR-34 mimic transfection of MiaPaCa2 cells with miR-34 mimics (100 pmol per well in 6-well plates), the expression of potential target genes was measured by qRT-PCR with SYBR Green PCR System (TaqMan). [score:3]
Transcription of the three miRNA miR-34 family members was recently found to be directly regulated by p53. [score:3]
Our results demonstrate that miR-34 may restore, at least in part, the tumor suppressing function of the p53 in p53 -deficient human pancreatic cancer cells. [score:3]
Our study demonstrates that miR-34 may restore, at least in part, the tumor suppressing function of p53 in p53 -deficient cancer cells. [score:3]
Next, we examined whether miR-34 restoration could sensitize the pancreatic cancer cells with a high level of endogenous Bcl-2 expression to chemo- and radiotherapy. [score:3]
Figure S3Restoration of miR-34 by MIF lentiviral system inhibited MiaPaCa2 tumorspheres. [score:3]
More significantly, miR-34 restoration led to an 87% reduction of the tumor-initiating cell population, accompanied by significant inhibition of tumorsphere growth in vitro and tumor formation in vivo. [score:3]
This effect on cell cycle is similar to that of p53 restoration as we previously reported [23], [24], [27], [28], indicating that miR-34 restoration can exert effects akin to restoration of p53 tumor suppressor function, at least in part, in the cells with p53 loss of function. [score:3]
miR-34 significantly inhibited the invasion potential of MiaPaCa2 cells (Figure S2). [score:3]
Restoration of miR-34 by MIF lentiviral system decreases the CD44+/CD133+ MiaPaCa2 cells and inhibits tumorspheres from the sorted CD44+/CD133+ cells. [score:3]
At present, the linkages between p53, the downstream target miR-34 and presumptive pancreatic cancer stem cells are unknown. [score:3]
Another important finding from the current study is that our data provide a potential link between the tumor suppressor miR-34 and the tumor-initiating cells or cancer stem cells. [score:3]
0006816.g003 Figure 3MiaPaCa2 cells were transfected with miR-34 mimics or inhibitors, 24 hr later the cells were seeded in 6-well plates (200 cells/well, in triplicates). [score:3]
Taken together, the published studies suggest miR-34 family members may have tumor suppressor function downstream of p53. [score:3]
More importantly, our results demonstrate for the first time that the CD44+/CD133+ tumor-initiating cells, or pancreatic cancer stem cells, have a low level of miR-34 accompanied by a high level of Bcl-2, suggesting a potential link of miR-34 and its target Bcl-2 to pancreatic cancer stem cells. [score:3]
MiaPaCa2 cells were transfected with miR-34 mimics or inhibitors, 24 hr later the cells were seeded in 6-well plates (200 cells/well, in triplicates). [score:3]
miR-34 restoration significantly inhibited clonogenic cell growth and invasion, induced apoptosis and G1 and G2/M arrest in cell cycle, and sensitized the cells to chemotherapy and radiation. [score:3]
They are single-stranded chemically enhanced oligonucleotides that can effectively inhibit the endogenous mature miR-34. [score:3]
Figure S2 Restoration of miR-34 inhibits the invasion of MiaPaCa2 cells. [score:3]
Restoration of miR-34 may hold significant promise as a novel molecular therapy for human pancreatic cancer with loss of p53–miR34, potentially via inhibiting pancreatic cancer stem cells. [score:3]
It has been reported that miR-34 targets Notch, c-Met and Bcl-2, genes involved in the self-renewal and survival of cancer stem cells [10], [11], [14]. [score:3]
B, Quantitative real-time PCR analysis of the potential target genes' mRNA levels after miR-34 mimic transfection in MiaPaCa2 cells. [score:3]
Since p53 tumor suppressor function is mediated in part via induction of apoptosis [23], [24], we examined the effect of miR-34 restoration on apoptosis-induction in MiaPaCa2 cells transfected with miR-34 mimics. [score:3]
miR-34 restoration could thus re-build, at least in part, the p53 tumor suppressing signalling network in pancreatic cancer cells lacking functional p53. [score:3]
Delineating the role of miR-34 in regulation of cell growth and tumor progression, and its potential link to tumor-initiating cells or cancer stem cells may provide a basis for exploring its potential as a novel treatment strategy. [score:2]
miR-34 inhibitors induced an almost 20% increase in clonogenic growth as compared with control (120.3±2.9 colonies/well vs. [score:2]
E, Colony formation assay shows the miR-34a inhibits clonogenic growth of the miR-34-MIF. [score:2]
However, mutation in the Bcl-2 3′UTR complementary to the miR-34 seed sequence abolished this effect, indicating that the observed activity is sequence-specific. [score:2]
miR-34a, miR-34b and miR-34c mimics all had similar activities. [score:1]
We identified that CD44+/CD133+ MiaPaCa2 cells are enriched with tumorsphere-forming and tumor-initiating cells or cancer stem/progenitor cells with high levels of Notch/Bcl-2 and loss of miR-34. [score:1]
A, miR-34 restoration sensitizes the cells to chemotherapeutic agents. [score:1]
Sharp's group in their pioneer miRNA study [47], supporting the link of miR-34 to CSC. [score:1]
C, Restoration of miR-34 leads to caspase-3 activation. [score:1]
Restoration of miR-34 sensitizes MiaPaCa2 cells to chemotherapy and radiation. [score:1]
MiaPaCa2 CD44+/CD133+ cells are tumorsphere-forming cells that have high Bcl-2 and loss of miR-34. [score:1]
Another potential role for miR-34 in cancer initiation and progression may be a link to tumor-initiating cells or cancer stem cells (CSC). [score:1]
For cell cycle and apoptosis analysis by flow cytometry, MiaPaCa2 cells were transfected with miR-34 mimics or NC mimic in 6-well plates, trypsinized 24 hr later and washed with phosphate-buffered saline, and fixed in 70% ethanol on ice. [score:1]
We examined the roles of miR-34 in p53-mutant human pancreatic cancer cell lines MiaPaCa2 and BxPC3, and the potential link to pancreatic cancer stem cells. [score:1]
To investigate the potential role of miR-34 in pancreatic cancer stem cells, we examined whether miR-34 restoration could inhibit the CD44+/CD133+ cells and their self-renewal potential. [score:1]
Cells were also co -transfected with 100 pmol of each miR-34 mimic or NC mimic, as indicated, using Lipofectamine 2000. [score:1]
As shown in Figure 3 C, transient transfection of miR-34 mimics resulted in significantly increased activation of caspase-3, a key indication of the cells undergoing apoptosis [25]. [score:1]
To evaluate the long-term effects of the miR-34 restoration, we also employed a lentiviral system to express miR-34a. [score:1]
miR-34 restoration by transfection of MiaPaCa2 cells with miR-34 mimics. [score:1]
These findings suggest that miR-34 mimics may hold significant promise as a novel molecular therapy for human pancreatic cancer with loss of p53–miR34, potentially via modulating pancreatic cancer stem cells. [score:1]
miR-34 restoration sensitizes MiaPaCa2 cells to chemo- and radiotherapy. [score:1]
Our data demonstrate that miR-34 restoration can overcome chemo-/radioresistance of the pancreatic cancer cells that have high levels of Bcl-2 and low basal levels of miR-34s, and are dependent on Bcl-2 for survival and resistance to therapy. [score:1]
MiaPaCa2 cells were transfected with miR-34 mimic or NC mimic for 24 hr, plated in 96-well plates (5,000 cells/well), and treated with serially diluted chemotherapeutic agents, in triplicates. [score:1]
B, miR-34 restoration increases caspase-3 activation induced by gemcitabine or X-ray radiation in MiaPaCa2 cells. [score:1]
0006816.g004 Figure 4 A, miR-34 restoration sensitizes the cells to chemotherapeutic agents. [score:1]
Loss of miR-34 has been linked to chemoresistance of cancer [13]. [score:1]
In the current study, we have examined the effects of functional restoration of miR-34 by miR-34 mimics and lentiviral miR-34a on human p53-mutant pancreatic cancer MiaPaCa2 cells, as well as the potential link to the pancreatic cancer stem cell self-renewal. [score:1]
MiaPaCa2 cells were co -transfected with the Bcl-2 3′UTR Luciferase Reporter or its mutant, b-gal vector, together with either miR-34 mimics or NC mimic. [score:1]
miR-34 mimics resulted in significant G1 and G2/M arrest and a reduction of cells in S phase (Figure 3 D ), consistent with other reports on miR-34 restoration in various cancer mo dels [6], [7], [8], [10], [11], [21], [22], [26]. [score:1]
Transfection of miR-34 mimics. [score:1]
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2
[+] score: 370
Other miRNAs from this paper: hsa-mir-34a, hsa-mir-34b
Consistent with these C. elegans results, of a published GEO dataset for hippocampus of wild-type adult male C57BL/6 mice 44 revealed upregulation of genes related to extracellular matrix, cell adhesion, basement membrane and anti-apoptosis when mir-34 was knocked down by adeno -associated viral (AAV)- delivered mir-34 sponges (180 upregulated and 36 downregulated genes, FDR < 0.01) (Table S3). [score:11]
We observed reduced stress resistance in both mir-34 mutants and overexpressors, supporting the role of this feedback inhibition loop in regulation of mir-34 and DAF-16 levels to reduce the fluctuations in daf-16 and myc network target expression levels under stress conditions. [score:10]
We conclude that mir-34 upregulation is necessary for inducing developmental arrest with correct morphogenesis and enhanced survival of dauers, and that this role of mir-34 relies on a functional insulin signaling receptor, DAF-2. mir-34 is regulated by DAF-16, PQM-1 and DAF-12The insulin signaling pathway regulates dauer-related phenotypes and responses to stress conditions by regulating nuclear localization of its downstream target transcription factor, DAF-16/FOXO 32 33. [score:10]
According to this regulatory loop, if miR-34 becomes upregulated above threshold levels, mir-34 expression is lowered via the feedback inhibition of DAF-16, which results in reduced stress resistance. [score:9]
Supplementary Table 1. Supplementary Table 2. Supplementary Table 3. P mir-34 [2.2kb] ::gfp is expressed in various tissues during development of C. elegans and its expression is upregulated in dauers. [score:9]
We conclude that mir-34 upregulation is necessary for inducing developmental arrest with correct morphogenesis and enhanced survival of dauers, and that this role of mir-34 relies on a functional insulin signaling receptor, DAF-2. The insulin signaling pathway regulates dauer-related phenotypes and responses to stress conditions by regulating nuclear localization of its downstream target transcription factor, DAF-16/FOXO 32 33. [score:9]
A mir-34 rescue strain, which has a single copy insertion of mir-34 and restores expression of miR-34 to 70% of the wild type level (Fig. 2A), as well as a mir-34 overexpression strain (mir-34OE), which has 4 copies of mir-34 and expresses miR-34 3-fold higher than wild type (Fig. 2A), rescue these morphological defects (Fig. S1). [score:7]
Genes that were expressed higher in the absence of mir-34 were significantly enriched for DAF-16 binding elements (DBE) and AGO-CLIP supported miR-34 targets, whereas genes which were expressed higher in the wild-type dauers were significantly depleted for DAF-16 binding but enriched for PQM-1 binding the DAE (Fig. 4B). [score:7]
These data suggest that differential expression of genes between wild-type and mutant dauers is the result of both direct regulation by miR-34 and indirect regulation via DAF-16/PQM-1 binding. [score:7]
We observed a large overlap between class 1 genes, dauer related genes, and genes that were up-regulated in mir-34(gk437) dauers, and between class 2 genes, non-dauer genes, and genes that were down-regulated in mir-34(gk437) dauers (Fig. S4). [score:7]
The same expression patterns were observed in dauers of the transgenic P mir-34 [5kb] ::gfp reporter 26, suggesting that all crucial regulatory elements for dauer related upregulation of mir-34 were located within the 2.2 kb upstream region. [score:7]
If miR-34 expression helps in establishing the stress response program, then gene expression changes when mir-34 is overexpressed under normal conditions should overlap with stress response genes that are observed in WT animals grown under heat stress. [score:7]
Although the observed changes in daf-16 expression upon miR-34 overexpression are not large, combined with the experimental AGO-CLIP data 39 and DAF-16::GFP reporter analysis results, they suggest direct regulation of daf-16 by miR-34. [score:7]
In mir-34OE and mir-34 mutant dauers a large number of genes were differentially expressed compared to WT dauers (1157 and 4652, respectively; Fig. 4B), consistent with the observed phenotypes and upregulated mir-34 expression pattern in dauers. [score:7]
GO term analysis of genes that were upregulated in the mir-34(gk437) dauer background and had ALG-1 binding sites and/or MIRZA scores higher than 100 revealed upregulation of genes encoding glycoproteins, cytoskeletal genes, intermediate filaments, extracellular matrix proteins, transmembrane and transport proteins (Table S2). [score:7]
In mutants that enhance temperature -induced dauer formation, the P mir-34 [2.2kb] ::gfp transgene expression patterns were similar to those seen in the wild-type (WT) background, implying that high mir-34 expression derived from differential gene expression at the dauer stage, and not from starvation conditions (Fig. 1C). [score:7]
mir-34 expression is regulated by the dauer larva gene expression program. [score:6]
Stress -induced upregulation of mir-34 expression was observed in starved worms, dauers and older adults 26 31, which was recapitulated by our P mir-34 [2.2kb] ::gfp transgenic line (Fig. 1A–D). [score:6]
Further evidence for a DAF-16- mir-34 feedback inhibition loopFinally, we sought additional evidence for daf-16 regulation by miR-34 in a daf-16::gfp reporter strain and in gene expression data. [score:6]
In this study, we demonstrated that miR-34 levels are upregulated to sustain a gene expression program that is associated with morphological and metabolic adaptation of stress. [score:6]
mir-34 expression is regulated by the dauer larva gene expression programTo study the relationship between mir-34, cell cycle arrest and stress, we focused on the dauer stage of C. elegans, which is the stress-resistant diapause stage that forms under harsh environmental conditions such as crowding, high temperatures and starvation 28 29. [score:6]
The highest expression levels were observed with insulin signaling pathway mutant dauers (Fig. 1C ii and iii), suggesting the possibility for the involvement of DAF-16/FOXO in the regulation of mir-34 expression. [score:6]
Furthermore, in N2 animals shift from 20 °C to 25 °C does not significantly change daf-16 levels (6% change, P = 0.523) but overexpression of miR-34 at 25 °C leads to a 25% downregulation of daf-16 compared to WT at 20 °C (P = 0.012). [score:5]
However, several other autophagy-related genes (lgg-1, atg-13, atg-16.2, unc-51, bec-1) were downregulated in mir-34OE dauers compared to mir-34 mutants (Table S1), which may suggest autophagy inhibition by mir-34 as was proposed in the aforementioned study 57. [score:5]
Detailed MIRZA alignment of the miR-34 target in daf-16 mRNA shown in panel C. (E) Expression of DAF-16::GFP is elevated with temperature in amphid neurons (indicated by arrows) in mir-34(gk437) mutants but not in wild-type animals. [score:5]
Small internal promoter deletions in the region bound by these TFs revealed that DAF-12 binding elements, insulin response element (IRE) and GA-repeats were required for mir-34 expression in hypodermis and seam cells (Fig. S2), and DAF-16 was necessary for activation of P mir-34 [2.2kb] ::gfp expression in dauers (Fig. 1C and Fig. S3A) and in amphid neurons, especially AWC neurons of adults (Fig. S3D). [score:5]
A good target would have a MIRZA score above 50, and Table S1 also lists how many miR-34 targets were found in a gene, and their cumulative MIRZA score. [score:5]
Effects of mir-34 deletion or overexpression on gene expression under various conditions. [score:5]
miR-34 targets daf-16To understand the molecular programs underlying phenotypic changes observed in the mir-34 mutant and overexpression strains, we identified experimentally supported targets of miR-34 using Argonaute crosslinking and immunoprecipitation (AGO-CLIP) data generated by Grosswendt et al. 39, in combination with calculated by MIRZA software 40. [score:5]
These results suggest that mir-34 upregulation is dependent upon DAF-16 in the dauer stage. [score:4]
We also observed that upregulation of P mir-34 [2.2kb] ::gfp in dauers is abolished by mutating this region, and in the daf-16(mu86) background. [score:4]
While in wild-type animals temperature shift from 20 °C to 25 °C resulted in 1891 and 2425 down- and up- regulated genes respectively (Fig. 4D), the number of differentially expressed genes was smaller in mir-34(gk437) (1192/1709 genes) and mir-34OE (1008/1442) backgrounds (Fig. 4D). [score:4]
Thus, our results suggest that mir-34 is involved in a feedback inhibition loop that includes the daf-16 and myc networks to regulate a stress response program in C. elegans (Fig. S6). [score:4]
Furthermore, analysis of P mir-34 [2.2kb] ::gfp in excretory gland cells in various mutant background showed a direct correlation between DAF-16 levels and reporter expression (Fig. S3B,C). [score:4]
Finally, we sought additional evidence for daf-16 regulation by miR-34 in a daf-16::gfp reporter strain and in gene expression data. [score:4]
Additionally, mdl-1 and mxl-3, from the Myc-like interaction network in C. elegans, showed mir-34 dependent downregulation under high temperature growth, suggesting that the myc network is a part of the stress response pathway that is modulated by daf-16 and mir-34. [score:4]
We showed that mir-34 mutation results in morphogenesis defects of dauers, which correlates with our transcriptome analysis results that shows deregulation of cell adhesion, cytoskeleton, ECM and basement membrane related gene categories both in of mir-34 mutant dauers and mir-34 knockdown mouse hippocampus. [score:4]
Therefore, we think that upregulation of mir-34 is necessary for correct morphogenesis of tissues to ensure long survival of dauers. [score:4]
Additionally, ATG9A did not show a significant change in expression levels in mir-34 knockdown in male mouse hippocampus (Table S3). [score:4]
We identified the minimal promoter region responsible for mir-34 upregulation by generating several P mir-34 [2.2kb] ::gfp strains with shorter upstream regions relative to the initial 2.2 kb promoter (Fig. 3A). [score:4]
mir-34 is regulated by DAF-16 and targets daf-16. [score:4]
Upregulation of mir-34 in hypodermis and seam cells and the morphological defects of mir-34(gk437) mutants correlate with these GO terms. [score:4]
miR-34 targets daf-16. [score:3]
Construction of mir-34 rescue and overexpression strains. [score:3]
The predicted miR-34 target region is located in the last coding exon of daf-16, not far from the stop codon (Fig. 3C). [score:3]
The mdl-1 gene promoter is bound by DAF-16 and PQM-1, according to modENCODE data 35, and the mdl-1 mRNA is targeted by miR-34 according to AGO-CLIP data 39 and MIRZA prediction, although the MIRZA score is modest (Table S1). [score:3]
qPCR validation of miR-34 expression. [score:3]
Furthermore, according to our combined analysis of MIRZA scores and AGO-CLIP data, one of the top predicted miR-34 targets is daf-16/FOXO. [score:3]
We observed higher DAF-16::GFP levels in mir-34(gk437) mutants grown at high temperatures (P = 0.0175, t test), accompanied by higher levels of nuclear localization of the translational fusion protein in amphid neurons, however, there were no significant differences under normal growth conditions (Fig. 3E,F). [score:3]
P mir-34 [2.2kb] ::gfp levels were similar to WT levels in daf-2(e1370);daf-16(mu86) background (Fig. S3D), suggesting that other factors were also involved in mir-34 induction upon inhibition of insulin signaling pathway. [score:3]
miR-34 expression is necessary for inducing stress response programs. [score:3]
The number of differentially expressed genes was highly diminished in mir-34(gk437) mutants when the comparison was done in the daf-2(e1370) background, where nuclear DAF-16 levels are saturated (Fig. 4C). [score:3]
Construction of mir-34 rescue and overexpression strainsP mir-34 [2.2kb]:: mir-34 was cloned into MosSCI plasmid and isolated plasmid was microinjected into N2 worms together with marker plasmids. [score:3]
The levels of daf-16 mRNA decreased by 12% and 8%, respectively, in adults and dauers overexpressing miR-34 at 20 °C, although the statistical significance of these changes is low (adjusted P value 0.41 and 0.58 respectively). [score:3]
We demonstrated DAF-16 dependent changes in the transcriptomes of animals that lack and overexpress mir-34. [score:3]
To understand the molecular programs underlying phenotypic changes observed in the mir-34 mutant and overexpression strains, we identified experimentally supported targets of miR-34 using Argonaute crosslinking and immunoprecipitation (AGO-CLIP) data generated by Grosswendt et al. 39, in combination with calculated by MIRZA software 40. [score:3]
At 25 °C, overexpression of miR-34 in adults results in a 17% decrease of daf-16 levels (adjusted P = 0.161). [score:3]
However, we found that in C. elegans atg-9 mRNA expression was lower in mir-34 and daf-2;mir-34 backgrounds, and did not observe a significant change in atg-9 transcript levels in adult stages (Table S1). [score:3]
mir-34 regulates dauer morphogenesis and survival dependent upon the insulin signaling pathwayWe investigated the role of mir-34 upregulation in dauers by studying dauer morphogenesis and survival in mir-34 mutants. [score:3]
Native levels of mir-34 expression are required for correct morphogenesis of dauers and dauer survival. [score:3]
In predauers, P mir-34 [2.2kb] ::gfp expression was increased in amphid neurons, especially in AWC neurons. [score:3]
In line with these findings, our results suggest that mir-34 has an evolutionarily conserved function in orchestrating responses to stresses, by modulating expression levels of DAF-16/FOXO and the Myc network. [score:3]
Additionally, glucose supplementation reduced P mir-34 [2.2kb] ::gfp levels (Fig. S3C), and prolonged stress conditions resulted in reduction of P mir-34 [2.2kb] ::gfp expression in many tissues of the worms. [score:3]
Further evidence for a DAF-16- mir-34 feedback inhibition loop. [score:3]
There were only 28 such genes for which expression increased with temperature in WT animals but decreased in mir-34(gk437) animals (Fig. 4H), including mdl-1 and mxl-3. mdl-1 is a basic helix-loop-helix (bHLH) transcription factor that acts as a part of the Myc-like interaction network in C. elegans. [score:3]
The analysis of these strains indicated that sequences between 0.5 kb and 1.2 kb upstream of mir-34 gene are essential for its regulation (Fig. 3B). [score:2]
These results suggest that indeed mir-34 plays a direct role in establishing the stress response program. [score:2]
The combination of AGO-CLIP evidence and highly-scoring MIRZA prediction suggests that daf-16 is very likely regulated by miR-34 and suggests existence of a negative feedback-loop between daf-16 and mir-34. [score:2]
mir-34 is regulated by DAF-16, PQM-1 and DAF-12. [score:2]
Other transcription factors, which showed same pattern as mdl-1 in terms of sensitivity to mir-34 levels included nhr-23, egl-13 and zip-7. NHR-23 is a critical co-regulator of functionally linked genes involved in growth and molting. [score:2]
The survival defect of mir-34 mutants required a functional insulin signaling pathway, where DAF-16 nuclear localization levels are not saturated, and probably DAF-16 activity is more prone to regulation by mir-34. [score:2]
Additionally, compared to WT dauers, mir-34(gk437) mutants had a shorter body size, and worms that overexpress mir-34 had a slightly longer body size (Fig. 2B). [score:2]
Additionally, the mir-34(gk437) mutation enhanced dauer formation in daf-7(e1372) mutant background but it did not have an effect on dauer formation in daf-2(e1370) mutants (Fig. 2D). [score:2]
mir-34 regulates dauer morphogenesis and survival dependent upon the insulin signaling pathway. [score:2]
We investigated the role of mir-34 upregulation in dauers by studying dauer morphogenesis and survival in mir-34 mutants. [score:2]
Since daf-16(mu86) null mutants cannot form dauers, this mutation was introduced into the daf-7(e1372);P mir-34 [2.2kb] ::gfp strain, which is dauer constitutive at 25 °C, in order to investigate whether the high expression of mir-34 in daf-7(e1372) mutant dauers was dependent upon DAF-16. [score:2]
For example, mir-34 deletion mutants do not show any abnormal morphological, developmental or biological phenotypes under standard laboratory culture conditions. [score:2]
According to a previous study the autophagy-related mRNA ATG9A was regulated by mir-34 in mammalian cells 57. [score:2]
Which genes might be particularly sensitive to miR-34 levels in this stress response program? [score:1]
To study the relationship between mir-34, cell cycle arrest and stress, we focused on the dauer stage of C. elegans, which is the stress-resistant diapause stage that forms under harsh environmental conditions such as crowding, high temperatures and starvation 28 29. [score:1]
Thus, a strong daf-2/dauer transcriptional signature is present in mir-34(gk437) mutant dauers, as it is also evident from our transcriptome analysis (Fig. S5A (i, ii)). [score:1]
In line with these results, both mir-34(gk437) and mir-34OE adults were more sensitive to hypoxia, heat stress, and starvation. [score:1]
Although mir-34 mutant dauers exhibit more a dauer-related transcriptional signature, changes in gene category representation are accompanied by the body defects and short survival rates of mir-34 mutants. [score:1]
This suggests that precise levels of miR-34 are required to elicit proper response to heat stress, and that deviations from these levels impair the stress response program. [score:1]
qPCR was performed on N2, mir-34(gk437) and N2 and mir-34(gk437) worms carrying P mir-34 [2.2kb]:: mir-34 transgene by using TaqMan kit. [score:1]
This finding was in line with the lower amount of phenotypic changes observed between daf-2(e1370) and daf-2(e1370);mir-34(gk437) mutant worms. [score:1]
These transcription factors may also be responsible for miR-34 dependent transcriptome and phenotypic changes under stress conditions. [score:1]
At the same time, there was no significant difference between daf-2(e1370) and daf-2(e1370);mir-34(gk437) worms in terms of body size and survival (data not shown). [score:1]
Around 80% (250 dauers tested) of mir-34(gk437) dauers that were selected from starved plates had locomotion defects, and were rolling along their body axis. [score:1]
How to cite this article: Isik, M. et al. MicroRNA mir-34 provides robustness to environmental stress response via the DAF-16 network in C. elegans. [score:1]
Transcriptome analysis reveals the crosstalk between DAF-16 and mir-34To investigate the possible crosstalk between mir-34 and DAF-16, we performed microarray gene expression analysis for several genetic backgrounds and stress conditions (Table S1, Fig. 4). [score:1]
It has a MIRZA score of 625, and exhibits perfect complementarity to nucleotides 1–8 of the mature miR-34 sequence (Fig. 3D). [score:1]
Transcriptome analysis reveals the crosstalk between DAF-16 and mir-34. [score:1]
This suggests that the C. elegans myc network may play an important role in modulating a stress response program downstream of mir-34. [score:1]
By engineering promoter truncations we showed that an IRE sequence, which binds DAF-16, is present in mir-34 promoter. [score:1]
P mir-34 [2.2kb]:: mir-34 was cloned into MosSCI plasmid and isolated plasmid was microinjected into N2 worms together with marker plasmids. [score:1]
To investigate the possible crosstalk between mir-34 and DAF-16, we performed microarray gene expression analysis for several genetic backgrounds and stress conditions (Table S1, Fig. 4). [score:1]
MIRZA, miR-34 target predictions calculated by MIRZA. [score:1]
ALG – presence of AGO-CLIP regions from Grosswendt et al. 39, overlapping miR-34 MIRZA predictions. [score:1]
Furthermore, mir-34(gk437) mutant dauers exhibited a lower survival rate (at both 20 °C and 25 °C) than WT dauers (Fig. 2C). [score:1]
However, miR-34 is critical in the DNA damage response in both mammals and C. elegans. [score:1]
To address this question, we looked for genes that responded oppositely to heat stress in WT and mir-34(gk437) animals. [score:1]
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The full-length blot images are presented in Supplementary Figure  3. Reciprocal regulation between Notch1 and miR-34cAs overexpressing N1ICD decreases miR-34c expression during PSCs development (Fig.   1) and miR-34c mimics decreases Notch1 gene expression in the proliferation period (Fig.   2), differentiation day 1and differentiation day 7 (Fig.   4), these results suggest a regulatory feedback may exist between Notch1 and miR-34c. [score:10]
The full-length blot images are presented in Supplementary Figure  3. As overexpressing N1ICD decreases miR-34c expression during PSCs development (Fig.   1) and miR-34c mimics decreases Notch1 gene expression in the proliferation period (Fig.   2), differentiation day 1and differentiation day 7 (Fig.   4), these results suggest a regulatory feedback may exist between Notch1 and miR-34c. [score:9]
Since our in vitro study has shown that overexpressing miR-34 inhibits muscle development, we believe the miR-34c overexpression experiment alone would be sufficient to demonstrate the role of miR-34c in vivo. [score:8]
Overexpressing N1ICD decreases miR-34c expression in vitroTo explore the regulations of Notch1 signaling in PSCs development, we constructed the constitutively activated N1ICD PSCs. [score:7]
Our study ascertains that miR-34c inhibits PSCs proliferation by inhibiting Notch1 expression. [score:7]
These results demonstrate we have successfully overexpressed N1ICD in PSCs and N1ICD decreased miR-34c expression in all three periods of PSCs development. [score:6]
Figure 1Overexpressing N1ICD decreases miR-34c expression during PSCs development. [score:6]
But miR-34c inhibitor had no effect on Myod gene expression on differentiation day 7 (Fig.   5D and E). [score:5]
Figure 2Overexpressing miR-34c mimics inhibits PSCs proliferation. [score:5]
The full-length blot images are presented in Supplementary Figure  3. Next, miR-34c inhibitor or Control (miR-34c inhibitor and Control are both synthetic oligonucleotide sequences; see Table  S1 for details) was transfected into PSCs, and the cells were induced to differentiation for 1 day and 7 days. [score:5]
Overexpressing miR-34c inhibits PSCs proliferation in vitro. [score:5]
Our results not only established Notch1 is the target gene of miR-34c but also discovered that Notch1, in turn, directly regulates miR-34c in PSCs. [score:5]
Figure 3Overexpressing miR-34c inhibitor increase PSCs proliferation. [score:5]
But miR-34c inhibited the skeletal muscle satellite cell proliferation by targeting Notch1, and miR-34c reduced the number of satellite cells to be fused to existing myofibers, resulting in smaller muscle fiber diameter after miR-34c injection. [score:5]
In this study, we used miR-34c mimics and inhibitor to manipulate the miR-34c level in transfected PSCs and that elevated miR-34c not only inhibits PSCs proliferation but also promotes PSCs differentiation. [score:5]
The full-length blot images are presented in Supplementary Figure  3. Next, miR-34c inhibitor or Control (miR-34c inhibitor and Control are both synthetic oligonucleotide sequences; see Table  S1 for details) was transfected into PSCs, and the cells were induced to differentiation for 1 day and 7 days. [score:5]
Figure 5Overexpressing miR-34c inhibitor reduces PSCs differentiation. [score:5]
The expression of miR-34c was decreased in the N1ICD overexpressed cells (p < 0.01; Fig.   1C). [score:5]
From the RNA-seq data, we discovered that many miRNAs were differently expressed in the N1ICD overexpressing PSCs, including miR-34c [29]. [score:5]
Since N1ICD expression was decreased after PSCs transfected with miR-34c mimics (Figs  2 C and 4B), Notch1 may be a potential target gene of miR-34c. [score:5]
Overexpressing N1ICD decreases miR-34c expression in vitro. [score:5]
For all the experimental data in this study, including miR-34c inhibiting PSCs proliferation, miR-34c promoting PSCs differentiation and miR-34c repressing mice muscle development, we are comparing only the control group to the treatment group; thus we used the student’s t-test for the statistical analyses (SPSS 18.0, Chicago, IL, USA). [score:4]
In conclusion, our research demonstrates that miR-34c inhibits PSCs proliferation but promotes PSCs differentiation in vitro, and miR-34c represses pig muscle development in vivo. [score:4]
These results suggest that Notch1 is the direct target gene of miR-34c. [score:4]
However, there is no report regarding the role of miR-34c on PSCs development; therefore, the objective of this study was to define the role of miR-34c on PSCs development and ascertain whether there is a regulatory relationship between miR-34c and N1ICD. [score:4]
This reciprocal regulatory loop formed by miR-34c and Notch1 that controls skeletal muscle development is novel, and this information expands our understanding of the mechanisms involved in muscle development. [score:4]
To establish N1ICD regulates miR-34c expression, N1ICD was transfected into PSCs in primary culture. [score:4]
Furthermore, the objective of this particular experiment was to ascertain whether the miR-34c inhibits muscle development in vivo. [score:4]
Taken together, these results demonstrate miR-34c inhibits PSCs proliferation in vitro. [score:3]
Another miR-34 family member miR-34c also has been shown to inhibit rat vascular smooth muscle cell proliferation [27]. [score:3]
This study would be better to have the miR-34c inhibitor group in the in vivo study. [score:3]
PSCs transfected with pCDNA3.1- N1ICD, miR-34c inhibitor, miR-34c mimics or Control by Lipofectamine 2000 (Invitrogen, Carlsbad, CA, USA), according to the manufacturer’s instructions. [score:3]
We have also established Notch1 is a direct target gene of miR-34c using a dual-luciferase reporter assay. [score:3]
These results demonstrate there exists a regulatory loop between Notch1 and miR-34c in PSCs development. [score:3]
However, after considerable deliberations, we decided to forgo the miR-34c inhibitor group for the following reason: the mice used in this study are 4-week-old. [score:3]
Representative images of immunofluorescent staining of PSCs transfected with miR-34c inhibitor. [score:3]
The mRNA level of myogenin, myosin, and MyHC was elevated by the miR-34c overexpression on differentiation day 7 (p < 0.01, Fig.   4C), and protein level of myogenin and myosin follows the same pattern as the mRNA (p < 0.05, Fig.   4D). [score:3]
PSCs transfected with miR-34c mimics, miR-34c inhibitor or Control and maintained in growth medium. [score:3]
However, on differentiation day 7, overexpression of miR-34c showed no significant influence on MyoD mRNA and protein levels (Fig.   4C and D). [score:3]
Figure 4Overexpressing miR-34c mimics promotes PSCs differentiation. [score:3]
The full-length blot images are presented in Supplementary Figure  2. Next, we used miR-34c inhibitor to further ascertain the role of miR-34c on PSCs proliferation. [score:3]
Therefore, injection of LV-miR-34c inhibitor may not obtain additional muscle growth; thus, the value of using more mice may be questionable following the principle of a minimal number of animals to be used in any experiments. [score:3]
Overexpressing miR-34c reduced the percentage of Edu positive cells (p < 0.05, Fig.   2A). [score:3]
The full-length blot images are presented in Supplementary Figure  2. Next, we used miR-34c inhibitor to further ascertain the role of miR-34c on PSCs proliferation. [score:3]
MiR-34c inhibitor reduced the endogenous miR-34c level (Fig.   3B). [score:3]
But miR-34c inhibitor had no effect (p > 0.05) on myogenin and MyHC mRNA level on differentiation day 1 (Fig.   5A). [score:3]
Overexpression of miR-34c reduced Notch1 mRNA, and elevated MyoD mRNA and myosin mRNA on differentiation day 1 (p < 0.05, Fig.   4A). [score:3]
PSCs transfected with miR-34c inhibitor increased the mRNA and protein levels of N1ICD on both differentiation day 1 and day 7 (Fig.   5). [score:3]
On differentiation day 7, western blot result shown miR-34c inhibitor decreased (p < 0.05) myogenin and myosin protein levels. [score:3]
The full-length blot images are presented in Supplementary Figure  1. Overexpressing miR-34c inhibits PSCs proliferation in vitroFirst, we investigated the role of miR-34 in PSCs proliferation. [score:3]
After transfected with miR-34c inhibitor, the mRNA level of Notch1, CCNB, CCND and CCNE and PCNA were increased and p21 was decreased (p < 0.05, Fig.   3B). [score:3]
As for protein level of these genes, western blot result showed overexpression of miR-34c reduced Notch1, and elevated MyoD and myogenin on differentiation day 1 (p < 0.05, Fig.   4B). [score:3]
On differentiation day 1, western blot result shown miR-34c inhibitor decreased (p < 0.05) MyoD and myosin protein levels, but had no effect on myogenin (Fig.   5B). [score:3]
Overexpressing miR-34c promotes PSCs differentiation. [score:3]
We injected miR-34c into the gastrocnemius muscle of the mice to establish the changes in muscle growth and gene expressions. [score:3]
We measured the expression of Notch1 and the myogenic marker genes after PSCs were transfected with miR-34c inhibitor. [score:3]
Finally, through miR-34c lentivirus injection into mice gastrocnemius muscle, we confirmed miR-34c represses mice muscle development. [score:2]
Using, we showed that CSL-N1ICD complex binds directly to the −3631~−3625 sites of miR-34c. [score:2]
As the miR-34c level is reduced when N1ICD was overexpressed during PSCs development (Fig.   1A) and a CSL-N1ICD complex binding site (GTGGGAA) exists at upstream of the miR-34c genomic site (Fig.   6C), we measured the DNA fragment binding with CSL-N1ICD complex by ChIP. [score:2]
MiR-34c mimics and inhibitor were (see Table  S1) purchased from GENEWIZ (Suzhou, China). [score:2]
But, the role of miR-34 plays in pig skeletal muscle development has not been reported. [score:2]
These findings establish there exists a regulatory loop between Notch1 and miR-34c. [score:2]
These results demonstrate that injecting LV-miR-34c miR-34c represses muscle development in vivo. [score:2]
MiR-34c inhibitor decreased MyoD and myosin mRNA level on differentiation day 1 (p < 0.05, Fig.   5A). [score:2]
Through the dual-luciferase reporter assay, we found N1ICD decreased the pGL3-basic-miR-34 upstream recombinant vector relative luciferase activity, but this inhibition was abolished by the mutated CSL-N1ICD complex binding site (GTGGGAA) (Fig.   6F). [score:2]
Reciprocal regulation between Notch1 and miR-34c. [score:2]
Consistent with this result is the average area of myofibers decreased (p < 0.01, Fig.   7C), which means miR-34c repressed muscle development in vivo. [score:2]
This result indicates CSL-N1ICD complex may regulate miR-34c transcription. [score:2]
In our study, we used Edu assay to confirm miR-34c inhibits PSCs proliferation, and our qRT-PCR and western blot results show miR-34c is positively correlated with p21, and negatively correlated with CCNB, CCND, and CCNE. [score:2]
Thus the presence of virus affected muscle development, which explains the gastrocnemius muscle weight of the injected virus groups (LV-Control and LV-miR-34c) was lower than that of the Normal group. [score:2]
MiR-34c has been shown to be a new modulator of VSMC proliferation through targeting SCF [27]. [score:2]
MiR-34c inhibitor increased the percentage of Edu positive cells (p < 0.01, Fig.   3A). [score:2]
The CSL-N1ICD complex binds directly to the -3631~-3625 sites of miR-34c. [score:2]
MiR-34c represses muscle development in vivoTo evaluate the function of miR-34c in vivo, we injected lentivirus expressing miR-34c mimics or Control into mice gastrocnemius muscle. [score:2]
To our knowledge, there is no report about the miR-34c function on skeletal muscle development. [score:2]
MiR-34c inhibitor decreased (p < 0.05) myogenin, myosin and MyHC genes mRNA level on differentiation day 7 (Fig.   5D). [score:2]
In human, three miR-34 precursors are produced from two transcriptional units, miR-34a precursor is transcribed from chromosome 1, and miR-34b and miR-34c precursors are co-transcribed from a region on chromosome 11 [23]. [score:1]
The miR-34 family members (miR-34a, miR-34b, and miR-34c) were discovered computationally [20] and later verified by experiment 21, 22. [score:1]
Either pGL3-basic-miR-34c upstream-mut or pGL3-control was used as a control for pGL3-basic-miR-34c upstream. [score:1]
But only miR-34a and miR-34c are found in pig 24, 25. [score:1]
Thus, to ascertain the miR-34c function observed in our cell culture studies, we conducted the in vivo study in mice. [score:1]
To evaluate the function of miR-34c in vivo, we injected lentivirus expressing miR-34c mimics or Control into mice gastrocnemius muscle. [score:1]
We found relative luciferase activity was decreased (p < 0.01; Fig.   6B) when HEK-293T cells were co -transfected with miR-34c mimics and pmirGLO- Notch1-3′UTR. [score:1]
Mice were purchased from Guangdong Medical Lab Animal Center, and lentivirus containing miR-34c mimics or Control were purchased from Shanghai JiKai Gene Chemical Technology Co. [score:1]
MiR-34c or microRNA-control was delivered by a lentiviral vector (LV). [score:1]
We measured the expression of Notch1 and the myogenic marker genes after PSCs were transfected with miR-34c mimics. [score:1]
As shown in Fig.   6E the miR-34 upstream of its genomic site (about 4600 bp) is inserted into the pGL3-basic vector. [score:1]
Body weights were no difference among all groups, but the weights of gastrocnemius muscle were decreased in the LV-miR-34c treatment group and Normal groups (p < 0.05, Fig.   7D). [score:1]
*Indicates a difference between Control and miR-34c mimics. [score:1]
However, miR-34c mimics had no effect on CCNB mRNA and protein levels (Fig.   2B and C). [score:1]
Western blot confirmed that miR-34c mimics reduced (p < 0.05) Notch1, CCND and CCNE protein level, and increased (p < 0.01) p21 protein level (Fig.   2C). [score:1]
So we constructed the pGL3-basic-miR-34 upstream recombinant vector (pGL3-basic-miR-34 upstream). [score:1]
The miR-34c mimics decreased Notch1, CCND, CCNE and PCNA mRNA levels and increased p21 mRNA level (p < 0.05, Fig.   2B). [score:1]
A ~4600 bp sequence upstream of miR-34c in genomic DNA was amplified and inserted into pGL3-basic Vector (Ambion, Carlsbad, CA, USA). [score:1]
qRT-PCR result showed miR-34c mimics increased (p < 0.001) cellular miR-34c level at 24 h after transfection (Fig.   2B). [score:1]
Figure 6Negative feedback between miR-34c and Notch1. [score:1]
Total fibers in one field were increased after injection with LV-miR-34c. [score:1]
CCND1 mRNA level was decreased after LV-miR-34c injection, but mRNA levels of CCNB1 and p21 were increased (p < 0.05, Fig.   7E). [score:1]
MiR-34c represses muscle development in vivo. [score:1]
After injecting LV-miR-34c, MyoD protein level was decreased (Fig.   7F) and myogenin protein level was increased (Fig.   7F), but their mRNA levels were not different (Fig.   7E). [score:1]
Mice were injected with physiological saline, LV-Control or LV-miR-34c. [score:1]
Both miR-34c mimics and Control are synthetic oligonucleotide sequences delivered by the lentiviral vector. [score:1]
N1ICD with pGL3-basic-miR-34 upstream recombinant vector relative or pGL3-basic-miR-34 upstream (mut) recombinant vector were transfected into HEK-293T cells respectively. [score:1]
Notch1 mRNA level was decreased in the LV-miR-34c injection group (p < 0.05, Fig.   7F). [score:1]
Using dual-luciferase reporter assay and Chromatin immunoprecipitation (ChIP), we demonstrated there is a regulatory loop between Notch1 and miR-34c. [score:1]
These results indicate elevated miR-34c promotes PSCs differentiation. [score:1]
A fragment about 400 bp long located at -3631 upstream of miR-34c was amplified (Fig.   6D). [score:1]
LV-miR-34c injection also increased myosin mRNA level (p < 0.05, Fig.   7E). [score:1]
Lanes 1–7 represent DL 10000, pGL3-basic, double digested pGL3-basic, double digested miR-34 upstream of its genomic site, pGL3-basic-miR-34 upstream recombinant vector, double digested pGL3-basic-miR-34 upstream recombinant vector, DL5000, respectively. [score:1]
The full-length blot images are presented in Supplementary Figure  2. The medium was changed to differentiation medium for 1 and 7 days to study the function of miR-34c on PSCs differentiation. [score:1]
In Fig.   6D *Indicates differences between Normal and LV-Control or Normal and LV-miR-34c in mice gastrocnemius muscle. [score:1]
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More importantly, other potential miR-34 target genes were inhibited in addition to Bcl-2. As shown in Figure 4, Notch1 and HMGA2 were inhibited by all three miR-34a, b, c mimics, while miR-34b mimic inhibited Notch2 and 4, and miR-34c mimic inhibited Notch1-4. Notch1-2 knockdown by miR-34 mimics has been confirmed by Western blot (data not shown). [score:12]
As shown in Figure 3, revealed that transfection of miR-34 mimics downregulated target gene Bcl-2 expression at the protein level, but had no obvious effect on Bcl-xL and Mcl-1 expression, indicating that the Bcl-2 knockdown by miR-34 mimics was sequence-specific. [score:11]
Figure 4 Quantitative real-time PCR shows that restoration of miR-34 by downregulates target gene expression. [score:8]
Figure 3 Restoration of miR-34 by downregulates target gene Bcl-2 expression. [score:8]
This strategy was explored in the current study, where p53 downstream target miR-34 was restored in p53-mutant gastric cancer Kato III cells with a high level of Bcl-2 and low levels of miR-34, resulting in downregulation of Bcl-2 and Notch/HMGA2, tumor cell growth inhibition and accumulation in G1 phase, and chemosensitization and Caspase-3 activation/apoptosis. [score:8]
Bcl-2 is a direct target of miR-34, and our data have shown that miR-34 restoration inhibits Bcl-2 expression. [score:8]
The expression of miR-34 is dramatically reduced in 6 of 14 (43%) non-small cell lung cancers (NSCLC) and the restoration of miR-34 expression inhibits growth of NSCLC cells [10]. [score:7]
As a target of miR-34, Bax was also downregulated by miR-34. [score:6]
He et al. reported that ectopic expression of miR-34 induces cell cycle arrest in both primary and tumor-derived cell lines, which is consistent with the observed ability of miR-34 to downregulate a program of genes promoting cell cycle progression [12]. [score:6]
The mechanism of miR-34 -mediated suppression of self-renewal appears to be related to the direct modulation of downstream targets Bcl-2, Notch, and HMGA2, indicating that miR-34 may be involved in gastric cancer stem cell self-renewal/differentiation decision-making. [score:6]
The mechanism of miR-34 -mediated suppression of self-renewal might be related to the direct modulation of downstream targets Bcl-2, Notch, and HMGA2, indicating that miR-34 may be involved in gastric cancer stem cell self-renewal/differentiation decision-making. [score:6]
The mechanism of miR-34 -mediated suppression of gastric cancer cell self-renewal might be related to the direct modulation of downstream targets Bcl-2, Notch, and HMGA2, implying that miR-34 may be involved in gastric cancer stem cell self-renewal/differentiation decision-making. [score:6]
Since miR-34 is a downstream target of the p53 pathway and Bcl-2 is a direct target of miR-34, our data with Kato III are consistent with the cells' p53-mutant status, i. e., Kato III has mutant p53, the lowest level of miR-34, and the highest level of Bcl-2. Therefore, we focused on this cell line for the current study of the effect of miR-34 restoration. [score:6]
MicroRNA miR-34 was recently found to be a direct target of p53, functioning downstream of the p53 pathway as a tumor suppressor. [score:6]
Expression of miR-34 and target genes in human gastric cancer cell lines. [score:5]
This multi-mode action of miR-34 provides a therapeutic advantage over other molecular therapies, in that miR-34 has multiple targets and can work on multiple cell signalling pathways simultaneously, leading to synergistic effects that may translate into improved clinical efficacy for gastric cancer patients with p53 deficiency and multidrug resistance. [score:5]
Recently, miRNA miR-34 was identified as a p53 target and a potential tumor suppressor [4, 8- 12]. [score:5]
More significantly, miR-34 potently inhibits tumorsphere formation and growth in p53-mutant human gastric cancer cells, providing the first proof-of-concept that there is a potential link between the tumor suppressor miR-34 and gastric cancer cell self-renewal, which involves the presumed gastric cancer stem cells. [score:5]
We next examined these gastric cancer cell lines for the expression level of miR-34 and target genes using qRT-PCR. [score:5]
As shown in Figure 5, the transfected miR-34 mimics effectively inhibited luciferase reporter gene expression, which is controlled by Bcl-2 3'UTR in the promoter region. [score:5]
Quantitative real-time PCR was performed to determine the expression levels of potential miR-34 target genes. [score:5]
Human gastric cancer Kato III cells with miR-34 restoration reduced the expression of target genes Bcl-2, Notch, and HMGA2. [score:5]
In this study, we examined the effects of miR-34 restoration on p53-mutant human gastric cancer cells and potential target gene expression. [score:5]
Bommer et al. reported that the abundance of the three-member miRNA34 family is directly regulated by p53 in cell lines and tissues, and the Bcl-2 protein is regulated directly by miR-34 [10]. [score:5]
Our results demonstrate that in p53 -deficient human gastric cancer cells, restoration of functional microRNA miR-34 inhibits cell growth, induces apoptosis, and leads to chemosensitization, indicating that miR-34 may restore, at least in part, the p53 tumor-suppressing function. [score:5]
The results demonstrate that the transfected miR-34a, b, c are functional, and confirm that Bcl-2 is a direct target of miR-34, consistent with earlier reports [8, 10, 16]. [score:4]
Delineating the role of miR-34 in regulation of cell growth and tumor progression, as well as its potential relationship to cancer stem cells, will help us better understand the p53 tumor suppressor signalling network, facilitate our research in carcinogenesis and cancer therapy, and serve as a basis for our exploration of novel strategies in cancer diagnosis, treatment, and prevention. [score:4]
miR-34 targets Notch, HMGA2, and Bcl-2, genes involved in the self-renewal and survival of cancer stem cells. [score:3]
miR-34 restoration chemosensitizes gastric cancer cells with a high level of Bcl-2. miR-34 restoration inhibits gastric cancer cell growth. [score:3]
Another important implication from the current study is that our data provide a potential link between tumor suppressor miR-34 and the presumed gastric cancer stem cells. [score:3]
Fold increase was calculated by dividing the normalized target gene expression of the treated sample with that of the untreated control, with the value from the NC mimic set as 1. For cell cycle analysis by flow cytometry, Kato III cells were transfected with miR-34 mimics or NC mimic in 6-well plates, trypsinized 24 hours later and washed with phosphate-buffered saline, and fixed in 70% ethanol on ice. [score:3]
However, our data indicate that miR-34 restoration inhibits tumorspheres from p53-mutant gastric cancer cells, suggesting that miR-34 might be involved in the self-renewal of the presumed gastric cancer stem cells. [score:3]
Our study suggests that restoration of the tumor-suppressor miR-34 may provide a novel molecular therapy for p53-mutant gastric cancer. [score:3]
Since part of the p53 tumor-suppressing function is via promoting apoptosis [19, 20], we next examined the effect of miR-34 restoration on apoptosis. [score:3]
Our data provide the first evidence that miR-34 is able to inhibit tumorsphere formation and growth in p53-mutant gastric cancer cells, implying that miR-34 might play a role in the self-renewal of gastric cancer cells, presumably gastric cancer stem cells. [score:3]
of the potential miR-34 target protein Bcl-2 48 hours after of Kato III cells (100 pmol per well in 6-well plates). [score:3]
Taken together, these published studies establish that miR-34 is a new tumor suppressor functioning downstream of the p53 pathway, and provide impetus to explore the functional restoration of miR-34 as a novel therapy for cancers lacking p53 signalling. [score:3]
Our results demonstrate that in p53 -deficient human gastric cancer cells, restoration of functional miR-34 inhibits cell growth and induces chemosensitization and apoptosis, indicating that miR-34 may restore p53 function. [score:3]
Our study suggests that restoration of the tumor suppressor miR-34 may provide a novel molecular therapy for p53-mutant gastric cancer. [score:3]
Restoration of miR-34 inhibits tumorsphere formation and growth, which is reported to be correlated to the self-renewal of cancer stem cells. [score:3]
Figure 9 Restoration of miR-34 by MIF lentiviral system inhibits Kato III tumorspheres. [score:3]
miR-34 restoration inhibits gastric cancer tumorspheres. [score:3]
miR-34 restoration could thus rebuild, at least in part, the p53 tumor-suppressing signalling network in gastric cancer cells lacking p53 function. [score:3]
It has been reported that miR-34 targets Notch, HMGA2, and Bcl-2, genes involved in the self-renewal and survival of cancer stem cells [10, 12, 14]. [score:3]
miR-34 restoration inhibits tumorsphere formation and growth, which is reported to be correlated to the self-renewal of cancer stem cells. [score:3]
Bcl-2 3'UTR reporter assay showed that the transfected miR-34s were functional and confirmed that Bcl-2 is a direct target of miR-34. [score:3]
Restoration of miR-34 chemosensitized Kato III cells with a high level of Bcl-2, but not MKN-45 cells with a low level of Bcl-2. miR-34 impaired cell growth, accumulated the cells in G1 phase, increased caspase-3 activation, and, more significantly, inhibited tumorsphere formation and growth. [score:3]
However, mutation in the Bcl-2 3'UTR complimentary to the miR-34 root sequence abolished this effect, indicating that the observed reporter activity is miR-34 sequence-specific. [score:2]
Human gastric cancer cells were transfected with miR-34 mimics or infected with the lentiviral miR-34-MIF expression system, and validated by miR-34 reporter assay using Bcl-2 3'UTR reporter. [score:2]
As shown in Figure 9, restoration of miR-34 by MIF lentiviral system inhibited Kato III tumorsphere formation and growth; the stable cells with functional miR-34a restoration had significantly fewer tumorspheres, and the formed tumorspheres were significantly smaller, as compared with that of the MIF control (P < 0.001, Student's t-test, n = 3). [score:2]
The role of miR-34 in gastric cancer has not been reported previously. [score:1]
and using human primary gastric cancer tissues to identify the true side population of the assumed gastric cancer stem cells, and to delineate the role of miR-34 in these tumor-initiating cells. [score:1]
Cells in each well were also co -transfected with 100 pmol of each miR-34 mimics or NC mimic as indicated, using Lipofectamine 2000. [score:1]
This effect on cell cycle is similar to that of p53 restoration as we previously reported [18- 23], indicating that miR-34 restoration can restore p53 signalling, at least in part, in the cells lacking a functional p53 pathway. [score:1]
miR-34 mimic transfection. [score:1]
A. of Kato III cells after miR-34 restoration. [score:1]
As shown in Figure 6A, the miR-34 mimics induced an accumulation of Kato III cells in G1 phase and a reduction of cells in S phase, consistent with other reports on miR-34 restoration in various tumor mo dels [4, 8, 10, 12, 13, 16, 17]. [score:1]
Transfection of miR-34 mimics in p53-mutant gastric cancer cells. [score:1]
miR-34 restoration results in Kato III cell accumulation in G1 phase and caspase-3 activation. [score:1]
For miR-34 restoration, we transfected the Kato III cells with miR-34 mimics. [score:1]
Figure 6 Restoration of miR-34 in Kato III cells resulted in G1 block and caspase-3 activation. [score:1]
Since no cellular markers for gastric cancer stem cells have been wi dely accepted thus far, in the current study we employed tumorsphere culture to explore whether there is any link between miR-34 and tumorsphere-forming cells. [score:1]
In the current study, we examined the effects of functional restoration of miR-34 by miR-34 mimics and lentiviral miR-34a on human gastric cancer cells, and the effect of miR-34 on tumorsphere formation and growth of p53-mutant gastric cancer cells. [score:1]
was performed 24 hours after transfected with miR-34 mimics or negative control mimic (NC mimic). [score:1]
KATO3 cells were transfected with Bcl-2 3'UTR luciferase reporter plasmid or its mutant, plus the control β-galactosidase plasmid and 100 pmol of each miR-34 mimic or NC mimic. [score:1]
Cells in each well were also co -transfected with 100 pmol of each miR-34 mimic or NC mimic as indicated, using Lipofectamine 2000. [score:1]
Figure 8 Restoration of miR-34 in Kato III cells delays cell growth. [score:1]
Our data suggest that miR-34 may hold significant promise as a novel molecular therapy for human gastric cancer, potentially for gastric cancer stem cells. [score:1]
miRNA miR-34a, b, c mimics, antagonists, and negative control miRNA mimic (NC mimic) were obtained from Dharmacon (Chicago, IL) with the sequences for hsa-miR-34a: 5'- uggcagugucuuagcugguugu-3' ; hsa-miR-34b: 5'- caaucacuaacuccacugccau-3' ; hsa-miR-34c: 5'- aggcaguguaguuagcugauugc-3'. [score:1]
Briefly, Kato III cells were transfected with miR-34 mimics or NC mimic for 24 h, plated in 96-well plates (5,000 cells/well), and treated with serially diluted chemotherapeutic agents in triplicate. [score:1]
As shown in Figure 6B, transient transfection of miR-34 mimics resulted in significantly increased caspase-3 activation, a key indication of the cells undergoing apoptosis. [score:1]
Our data demonstrate that miR-34 restoration can chemosensitize those gastric cancer cells that have high levels of Bcl-2 and low basal levels of miR-34, which are dependent on Bcl-2 for survival and drug resistance. [score:1]
However, for gastric cancer MKN-45 cells that have a low level of Bcl-2 and a high level of miR-34, miR-34 restoration showed no chemosensitization (Figure 7B). [score:1]
To evaluate the long-term effects of miR-34 restoration, we have employed a lentiviral system to express miR-34a and have generated stable cells. [score:1]
Thus far, there is limited study on miRNA and gastric cancer; the link between p53 downstream target miR-34 and gastric cancer has not been established; and the role of miR-34 in gastric cancer remains to be investigated. [score:1]
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We detected modest upregulation of cMyc, E2f3, Met and Sirt1 in miR-34 -deficient MEFs, while Bcl2 was expressed at similar levels in wild-type and mutant cells (Figure 3K). [score:6]
Expression of members of the miR-34 family was similarly upregulated in response to p53 stabilization (Figure 3G). [score:6]
In addition, miR-449 expression is not substantially increased in miR-34 -null mice, and activation of the p53 pathway does not lead to significant upregulation of miR-449 (Figure S8). [score:6]
We also examined the consequences of miR-34 loss in MEFs on the expression of a subset of its previously reported direct targets [17], [20], [23], [25]. [score:6]
Thus, although a longer follow-up of miR-34 [T KO/T KO] mice may be needed to uncover very subtle defects in tumor suppression, we conclude that loss of miR-34 expression does not lead to a substantial increase in spontaneous tumorigenesis. [score:5]
Because previous work has relied on the use of miRNA antagonists to inhibit miR-34 function, it is possible that some of the previous observations reflected miR-34-independent off-target effects. [score:5]
With respect to the potential tumor suppressive role of miR-34, our experiments indicate that loss of miR-34 expression does not lead to an obvious increase in tumor incidence in mice and does not cooperate with Myc in the context of B cell lymphomagenesis. [score:5]
Importantly, in these three tissues, miR-34 expression is almost entirely p53-independent (Figure 1B–1D and [58]), a finding that suggests that additional transcription factors control the expression of this family of miRNAs in the absence of genotoxic or oncogenic stresses. [score:5]
Consistent with a possible tumor-suppressor role, loss of expression of members of the miR-34 family has been reported in human cancers. [score:5]
Here, we probe the tumor suppressive functions of the miR-34 family in vivo by generating mice carrying targeted deletion of the entire miR-34 family. [score:5]
The upregulation of Myc and E2f3 might contribute to the increased proliferation rate we have observed in miR-34 deficient MEFs. [score:4]
Consistent with previous reports indicating that miR-34a expression is under the direct control of p53 [13], [17], [18], we detected reduced levels of this miRNA in a subset of p53 -deficient tissues (heart, small and large intestine, liver and kidney), but the levels of both miR-34a and miR-34b∼c remained high in the brains, testes and lungs (Figure 1B–1D) of p53 [−/−] mice, a finding that suggests that p53-independent mechanisms determine basal miR-34 transcription in these tissues. [score:4]
Many of the predicted miR-34 target genes encode for proteins that are involved in cell cycle regulation, apoptosis, and growth factor signaling. [score:4]
Our results show that complete loss of miR-34 expression is compatible with normal development and that the p53 pathway is apparently intact in miR-34 -deficient mice. [score:4]
miR-34 and tumor suppression in vivo To extend our analysis to an in vivo setting, we next examined whether miR-34 inactivation is sufficient to accelerate spontaneous and oncogene -induced transformation in mice. [score:3]
Although our observation that single KO and miR-34 [T KO/T KO] mice produce viable offspring argues against an essential role for miR-34 in these processes, members of the related miR-449 family, that are particularly highly expressed in the testis (Figure S8), could partially compensate for miR-34 loss in this context. [score:3]
Despite the growing body of evidence supporting this hypothesis, previous studies on miR-34 have been done in vitro or using non-physiologic expression levels of miR-34. [score:3]
p53 -dependent and p53-independent miR-34 expression in vivo. [score:3]
Consistent with these results, doxorubicin treatment caused similar activation of p53 and of its downstream targets in wild-type and miR-34 [T KO/T KO] MEFs (Figure 3E and 3F). [score:3]
To test whether miR-34 plays a role in this context, we ectopically expressed oncogenic K-Ras in wild-type, miR-34 [T KO/T KO], and p53 [−/−] MEFs. [score:3]
Next, we sought to determine whether loss of miR-34 expression affects the p53 response in vitro. [score:3]
Complete loss of miR-34 expression in miR-34 [T KO/T KO] animals was confirmed by Northern blot and qPCR (Figure 2D). [score:3]
Figure S1 Relative miR-34 expression in mouse tissues upon irradiation. [score:3]
We have reported the generation of mice carrying targeted deletion of miR-34a, miR-34b and miR-34c, and we have investigated the consequences of loss of miR-34 expression on p53 -dependent responses in vitro and in vivo. [score:3]
Although these observations point towards an important role for miR-34 members as critical downstream effectors of p53 and potential tumor suppressors, these hypotheses have not been formally tested using miR-34 -deficient animals and cells. [score:3]
Our results show that the miR-34 family is not required for tumor suppression in vivo, and they suggest p53-independent functions for this family of miRNAs. [score:3]
In humans, for example, loss of miR-34 expression has been reported in a large fraction of primary melanomas, prostatic adenocarcinomas and small cell lung cancers [27], [28], among others. [score:3]
However, the tumor suppressive function of miR-34 might be restricted to specific tissues and loss of miR-34 might cooperate with specific oncogenic lesions. [score:3]
Ectopic expression of members of the miR-34 family is sufficient to induce cell cycle arrest or apoptosis, depending on the cellular context [14], [17]– [21]. [score:3]
Members of the miR-34 family (miR-34a, miR-34b, and miR-34c) have been wi dely speculated to be important tumor suppressors and mediators of p53 function. [score:3]
To determine whether loss of miR-34 expression leads to increased spontaneous tumorigenesis, we aged a cohort of 14 miR-34 [T KO/T KO] and 12 wild-type mice. [score:3]
miR-34 and tumor suppression in vitro. [score:3]
We show that under basal conditions the expression of both miR-34 loci is particularly elevated in the testes and, to a lesser extent, in the brains and lungs of mice. [score:3]
miR-34 and tumor suppression in vitro The p53 pathway provides a crucial barrier against the neoplastic transformation of primary cells [40]. [score:3]
In addition, inactivation of miR-34 expression has been recently shown to lead to accelerated neurodegeneration and ageing in Drosophila melanogaster [64]. [score:3]
miR-34 and tumor suppression in vivo. [score:3]
Introducing the miR-34 -null alleles we have generated into mouse mo dels of these types of human cancers will be important to fully explore the tumor suppressive potential of this family of miRNAs. [score:3]
These results show that while miR-34 alone is not required for p53 -mediated tumor suppression in MEFs, its loss might cooperate with inactivation of the Rb pathway in promoting cellular transformation. [score:3]
P53 -dependent cell cycle arrest in miR-34 [T KO/T KO] MEFsNext, we sought to determine whether loss of miR-34 expression affects the p53 response in vitro. [score:3]
Complete loss of miR-34a and miR-34c expression was further confirmed in MEFs by qPCR (lower panel). [score:3]
p53 -dependent and p53-independent miR-34 expression in vivo To investigate the biological functions of miR-34, we first examined the expression of this family of miRNAs under basal conditions and in response to p53 activation in vivo. [score:3]
However, even in this context complete loss of miR-34 expression was not sufficient to accelerate tumor formation. [score:3]
Consistent with this mo del is our observation that while loss of miR-34 expression alone does not allow the transformation of primary cells by oncogenic K-Ras, it slightly increases the efficiency of transformation when combined with inactivation of the Rb pathway by E1A (Figure 5A, 5B). [score:3]
Recent reports have also implicated miR-34 in neuronal development and behavior [60], [61] and a role for miR-34c in learning and memory [62], as well as in stress -induced anxiety [63], has been reported. [score:2]
First, in the tissues and cells used in our experiments, the expression of miR-449 members is much lower compared to miR-34a and miR-34c, as judged by multiple independent methods including qPCR, Northern blotting and high throughput sequencing (Figure S8 and data not shown). [score:2]
It is also possible that other miRNAs sharing sequence similarities with miR-34 may compensate for miR-34 loss in the knock-out animals. [score:2]
However, when MEFs were co-transduced with oncogenic K-Ras and E1A, which binds to and inhibits the retinoblastoma protein (pRb) [42], we observed a slight increase in the number of foci formed in miR-34 [T KO/T KO] MEFs compared to wild-type cells (Figure 5A, 5B). [score:2]
MiR-34 expression in wild-type and p53 [−/−] mouse tissues. [score:2]
Although we detected a remarkable induction of miR-34a and miR-34c expression in late-passage wild-type MEFs compared to early-passage MEFs (Figure 3A), miR-34 -deficient MEFs became senescent with a kinetic identical to wild-type MEFs (Figure 3B). [score:2]
To exclude the possibility that tissue culture conditions may have masked a physiologic role of miR-34 in modulating the p53 response, we next examined the consequences of p53 activation in miR-34 -deficient tissues directly in vivo. [score:2]
In particular, three highly related miRNAs—miR-34a, miR-34b, and miR-34c (Figure 1A)—are directly induced upon p53 activation in multiple cell types and have been proposed to modulate p53 function [13]– [20]. [score:2]
Generation of miR-34 constitutive and conditional knockout mice. [score:2]
Age range of the cohorts is 359–521 days (mean: 464 days) for wild-type and 359–521 days (mean: 445 days) for miR-34 [T KO/T KO]. [score:1]
Although it will be important to follow a larger cohort of animals over a more prolonged period, these results suggest that miR-34 does not provide a potent barrier to tumorigenesis in response to genotoxic stress in vivo. [score:1]
Although as predicted, p53 -null cells failed to arrest in G1 in response to doxorubicin treatment, the response of miR-34 [T KO/T KO] MEFs was indistinguishable from that of wild-type cells (Figure 3H–3I). [score:1]
Thymocytes were isolated from sex-matched, age-matched wild-type, miR-34 [T KO/T KO], and p53 [−/−] mice and seeded at a density of 1×10 [6] cells/ml in MEF medium. [score:1]
For example, p53 has been proposed to modulate autophagy [55] and stem cell quiescence [56], [57] and we cannot exclude that miR-34 plays an important role in these contexts. [score:1]
1002797.g005 Figure 5Oncogene -induced transformation in miR-34 [T KO/T KO] fibroblasts and mice. [score:1]
Age- and sex-matched wild-type, miR-34 [T KO/T KO] and p53 [−/−] mice were exposed to 10 Gy of ionizing radiation and euthanized 6 hours later. [score:1]
P53 -dependent cell cycle arrest in miR-34 [T KO/T KO] MEFs. [score:1]
Both wild-type and miR-34 -deficient mice appeared healthy throughout the follow-up period (Figure S7), in striking contrast with the ∼15 weeks reported median tumor-free survival of irradiated p53 [−/−] mice [52]. [score:1]
To extend our analysis to an in vivo setting, we next examined whether miR-34 inactivation is sufficient to accelerate spontaneous and oncogene -induced transformation in mice. [score:1]
Furthermore, loss-of-function studies using miR-34 antagonists have provided some evidence that this miRNA family is required for p53 function [13], [18], [22]– [24]. [score:1]
The incidence and latency of B cell lymphomas was virtually identical in Eμ-Myc;miR-34 [T KO/T KO] and Eμ-Myc;miR-34 [+/+] mice (Figure 5C) and the resulting tumors displayed similar histopathological features and extent of spontaneous apoptosis (Figure 5D–5E). [score:1]
As expected, p53 [−/−] thymocytes were almost entirely resistant to irradiation -induced apoptosis; however, wild-type and miR-34 -deficient cells were equally sensitive to DNA damage -induced apoptosis, as judged by dose-response and time-course experiments (Figure 4A, 4B). [score:1]
We also observed modest but significant miR-34c induction in the thymus, small and large intestine of irradiated mice, but not in the other tissues examined. [score:1]
Representative pictures of miR-34a [−/−] (E), miR-34b∼c [−/−] (F), and miR-34 [T KO/T KO] (G) males at 4 weeks of age. [score:1]
Samples obtained from sex- and age-matched adult (age range 3–16 months) wild-type and miR-34 [T KO/T KO] mice were subjected to a standard panel of serum chemistry tests to determine liver and kidney function (n≥5 per genotype). [score:1]
More difficult, however, is to reconcile our findings with previous reports of impaired p53-function in cells treated with miR-34 antagonists. [score:1]
To generate mice carrying deletion of the miR-34b∼c bicistronic cluster, we used recombineering to replace a 1.3 kbp DNA region in BAC RP-23-281F13 containing pre-miR-34b and pre-miR-34c with a frt-Neo-frt cassette. [score:1]
Figure S5Serum chemistry of age- and sex-matched wild-type and miR-34 -deficient mice. [score:1]
Wild-type, miR-34 [T KO/T KO], p53 [−/−] MEFs were seeded at 70% confluence and infected with virus. [score:1]
For the irradiation experiments, 150,000 wild-type, miR-34 [T KO/T KO] and p53 [−/−] MEFs were seeded into each well of a 6-well culture plate and starved for 72 hours. [score:1]
For the miR-34 [T KO] allele (G), double heterozygous mice were inter-crossed. [score:1]
We next sought to determine whether loss of miR-34 might accelerate tumor formation in response to genotoxic stress. [score:1]
Based on these results we conclude that miR-34 function is not required for p53 -induced cell-cycle arrest and apoptosis in response to genotoxic stresses. [score:1]
Age range of the cohorts is 298–425 days (mean: 333 days) for wild-type and 387–425 days (mean: 401 days) for miR-34 [T KO/T KO]. [score:1]
An additional issue raised by the results presented in this manuscript relates to possible p53-independent functions of miR-34. [score:1]
Future studies using the miR-34 -deficient animals we have generated will be needed to test these possibilities. [score:1]
These findings highlight likely redundancies among p53's downstream effectors, show that the miR-34 family is largely dispensable for p53 function in vivo, and suggest possible p53-independent functions. [score:1]
The animals were monitored for at least 12 months (wild-type = 359 days; miR-34 [T KO/T KO] = 359 days) and up to 17.3 months (wild-type = 521 days; miR-34 [T KO/T KO] = 521 days). [score:1]
Wild-type and miR-34 [T KO/T KO] MEFs were seeded into a 6-well plate (40,000 cells/well) and counted every day for the growth curves. [score:1]
Here we report the generation of mice carrying targeted deletion of all three members of the miR-34 family and systematically investigate the impact of miR-34 loss on the p53 pathway. [score:1]
To examine the consequences of complete loss of miR-34 function, we crossed miR-34a [−/−] and miR-34b∼c [−/−] mice to generate compound mutant animals carrying homozygous deletion of all three family members (miR-34 [T KO/T KO]). [score:1]
One notable exception is a recent elegant paper by Choi and colleagues demonstrating that miR-34 -deficient MEFs are more susceptible to reprogramming [30]. [score:1]
The results presented in this paper do not necessarily conflict with previous experiments showing that ectopic expression of miR-34 can induce many of the most characteristic consequences of p53 activation; here we have tested whether miR-34 is necessary for p53 function and not whether it is sufficient. [score:1]
Epigenetic silencing of miR-34 members has also been reported in human cancers. [score:1]
We next examined the role of miR-34 in the response to the DNA damaging agent doxorubicin. [score:1]
Peripheral blood samples obtained from sex- and age-matched adult (age range 3–16 months) wild-type (WT) and miR-34 -null (T KO) mice were subjected to complete blood cell count (n≥5 per genotype). [score:1]
For BrdU cell cycle analysis, wild-type, miR-34 [T KO/T KO], and p53 [−/−] MEFs were plated in complete medium at 70% confluence, treated with varying doses of doxorubicin for 16 hours or treated at different time points, and pulsed with 10 µM BrdU for one hour. [score:1]
Finally, we sought to determine whether genetic ablation of miR-34 could contribute to tumor formation in cooperation with a defined oncogenic lesion. [score:1]
Response to p53 activation in miR-34 [T KO/T KO] mouse embryonic fibroblasts (MEFs). [score:1]
In particular, members of the miR-449 family (miR-449a, b and c) have the same “seed” sequence as miR-34, and miR-34 antagonists could in principle impair their function as well. [score:1]
The experiments described above were performed on asynchronously growing early-passage MEFs and as such may not be sensitive enough to detect a modest effect of miR-34 loss on the S-phase checkpoint. [score:1]
Oncogene -induced transformation in miR-34 [T KO/T KO] fibroblasts and mice. [score:1]
Probes specific for miR-34a and miR-34c were used. [score:1]
This interpretation is also consistent with the faster proliferation rate displayed by miR-34 -deficient MEFs (Figure 3B, 3C) and with the observation by Lal and colleagues that miR-34a is involved in modulating the cellular response to growth factors [38]. [score:1]
RNAs from miR-34 [T KO/T KO] tissues were included to control for cross-hybridization. [score:1]
Five Eμ-Myc;miR-34 [+/+] tumors and and four Eμ-Myc;miR-34 [T KO/T KO] tumors were analyzed. [score:1]
The most logical interpretation of these results is that miR-34 -deficient MEFs, rather than being more resistant to irradiation -induced cell cycle arrest, possess a slightly faster basal proliferation or more rapid re-entry into the cell cycle following serum starvation. [score:1]
P53 -dependent apoptosis in miR-34 [T KO/T KO] cells and mice Having established that miR-34 is not required for cell cycle arrest in response to genotoxic stress in MEFs, we next sought to determine whether this miRNA family might contribute to p53 -induced apoptosis. [score:1]
Ionizing radiation induced similar activation of the p53 pathway and of its downstream effectors in wild-type and miR-34 [T KO/T KO] mice (Figure 4C). [score:1]
The standard 3T3 protocol was followed to determine the cumulative population doublings of wild-type, miR-34 [T KO/T KO], and p53 [−/−] MEFs. [score:1]
To investigate the biological functions of miR-34, we first examined the expression of this family of miRNAs under basal conditions and in response to p53 activation in vivo. [score:1]
1002797.g003 Figure 3Response to p53 activation in miR-34 [T KO/T KO] mouse embryonic fibroblasts (MEFs). [score:1]
A role for miR-34c in spermatogenesis and in controlling the first zygotic cleavage has been recently proposed [58], [59]. [score:1]
The sequence similarity between the three miR-34 family members (Figure 1A), which share the same “seed”, suggests that they may be functionally redundant. [score:1]
MiR-34 wild-type and miR-34 [T KO/T KO] MEF lines were also verified by qPCR. [score:1]
The blots were then hybridized with [32]P-labeled probes specific for miR-34a, miR-34c, and U6. [score:1]
However, the consequences of miR-34 loss on p53 function were not examined in detail. [score:1]
Our observation that inactivation of miR-34 does not impair p53 -mediated responses in vitro and in vivo is particularly relevant because a key role for miR-34 in the p53 pathway had been previously proposed by a number of independent groups. [score:1]
Having established that miR-34 is not required for cell cycle arrest in response to genotoxic stress in MEFs, we next sought to determine whether this miRNA family might contribute to p53 -induced apoptosis. [score:1]
Figure S4 Complete blood cell count of age- and sex-matched wild-type and miR-34 -deficient mice. [score:1]
The results are representatitve of two independent experiments performed on a total of four wild-type and four miR-34 [T KO/T KO] MEF lines. [score:1]
Experiments were performed on three independent wild-type and three independent miR-34 [T KO/T KO] MEF lines. [score:1]
To investigate the physiologic functions of the miR-34 family and to determine the extent to which its induction is required for p53 function, we generated mice carrying targeted deletion of both miR-34a and miR-34b∼c loci (Figure 2A–2C). [score:1]
Figure S7Overall survival of wild-type and miR-34 [T KO/T KO] cohorts. [score:1]
An analysis of the major myeloid and lymphoid populations of the bone marrow, spleen and thymus also did not reveal any statistically significant difference between wild-type and miR-34 [T KO/T KO] mice (Figure S6). [score:1]
Representative images of hematoxylin and eosin staining of heart, kidney, liver, lung, small intestine, ovary, testis, and spleen (black scale bar, 200 µm), brain (green scale bar, 2000 µm), and colon (red scale bar, 100 µm) from wild-type and miR-34 [T KO/T KO] mice. [score:1]
Analogous to what we observed in thymocytes in vitro, the apoptotic response was equally dramatic in wild-type and in miR-34 -deficient mice, while it was virtually absent in p53 [−/−] animals (Figure 4D–4G). [score:1]
P53 -dependent apoptosis in miR-34 [T KO/T KO] cells and mice. [score:1]
We therefore exposed a cohort of 14 miR-34 [T KO/T KO] and 11 wild-type mice to 1 Gy of ionizing radiation soon after birth and monitored them for 42–60 weeks. [score:1]
Notice the loss of signal for miR-449b in the miR-34 [T KO/T KO] lung and testis samples, which likely reflects cross-hybridization of the miR-449b probe to miR-34. [score:1]
Generation of miR-34 -deficient mice. [score:1]
miR-34 [T KO/T KO] embryos were obtained by intercrossing miR-34 mutant mice. [score:1]
We therefore examined the effects of DNA damage on thymocytes from wild-type, p53 [−/−], and miR-34 [T KO/T KO] mice. [score:1]
Figure S6Bone marrow, spleen and thymus analysis of age- and sex-matched wild-type and miR-34 [T KO/T KO] mice. [score:1]
A conclusive test for this hypothesis will require the generation of compound miR-34 and miR-449 mutant animals, but several lines of evidence suggest that this explanation is not particularly likely. [score:1]
A full histological examination (Figure S3), complete blood cell count (Figure S4), and serum chemistry analysis (Figure S5) did not detect any statistically significant defects in adult miR-34 [T KO/T KO] mice of both sexes. [score:1]
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6
[+] score: 197
From this analysis, three differentially expressed microRNAs were revealed, namely, miR-214 (permutation p value 3.7×10 [−3]), expressed specifically in human PBMCs exposed to either HMGB1 [+/+] or HMGB1 [−/−] lysates, and miR-34c (permutation p value 6×10 [−4]), expressed in PBMCs exposed to HMGB1 [+/+] lysates alone (Table 1). [score:9]
There was an increase in fold expression from 2.1±1.97 to 15.9±1.85 (log2-transformed fold expression values) in cells pre -transfected with pre-miR-34c and exposed to HMGB1 [−/−] lysates (Fig. 4B), suggesting targeting of the seed sequence, preventing miR-34c -mediated degradation of IKK mRNA. [score:7]
In our study, miR-34a was significantly down-regulated in human PBMCs exposed to damaged lysates, in contrast to miR-34c upregulation. [score:7]
Levels of miR-34c Expression (and Pro-inflammatory Cytokines) Decreased After Pre-incubation with the Inflammasome Inhibitor, Glybenclamide. [score:5]
We hypothesize that hsa-miR-34c may be required for fine-tuning expression of IKKγ, a key signal transduction intermediate in the expression of multiple immunity or inflammation associated genes. [score:5]
Together these data suggest that hsa-miR-214 expression is a general “DAMPmiR” expressed in human PBMCs exposed to damaged cells, while hsa-miR-34c is a miRNA that is sensitive to the presence of HMGB1 in damaged cells. [score:5]
Levels of hsa-miR-34 expression in pre-miR-34c transfected PBMCs were confirmed by the increase in fold expression of this miR using TaqMan microRNA real-time RT-PCR. [score:5]
C: Decreased protein expression of IKKγ after transfection with pre-miR-34c (lane 3) or increased protein expression of IKKγ after transfection with anti-miR-34c (lane 2) and exposure to damaged HMGB1 [+/+] cell lysates for 24 hrs. [score:5]
These results support the observation that both hsa-miR-34c and hsa-miR-214 are upregulated when human PBMCs are exposed to damaged or necrotic cells, where hsa-miR34c appears to be responsive to the presence of HMGB1. [score:4]
Differential expression of TNFα and hsa-miR-34c in human donor PBMCs following exposure to wild-type (wt) HCT116 or HMGB1 stable knock-down (kd) lysates. [score:4]
uk/cgi-bin/targets/) for hsa-miR-34c is the regulatory non- enzymatic scaffold protein NEMO (NF-kappa B essential modulator also known as IKKγ (or Ik Kinase gamma). [score:4]
Hierarchical clustering analysis of microRNA profiling confirmed that hsa-miR-34c is preferentially upregulated in PBMCs exposed to HMGB1-containing lysates but not HMGB1 [−/−] lysates (Figure 1A, upper panel). [score:4]
These findings support the notion that IKKγ may be a direct target of hsa-miR-34c. [score:4]
Here, we report that when human PBMCs are exposed to damaged HMGB1 [+/+] cell lysates, or conditioned media from serum-starved and glucose-deprived cells, both hsa-miR-34c and hsa-miR-214 are upregulated. [score:4]
Figure 1C shows the fold expression changes (as log 2-transformed values) for hsa-miR-34a, miR-34b, miR-34c, miR-214 and miR-155 under these conditions. [score:3]
Our findings clearly indicate that miR-34c and miR-214 are specifically expressed in human PBMCs following exposure to sterile cell lysates or conditioned media from stressed cells, but not when exposed to PAMPs as TLR ligands. [score:3]
Expression of Hsa-miR-34c and Hsa-miR-214 does not Increase in Human PBMCs Stimulated with Specific Pathogen-activated Molecular Pattern Molecules (PAMPs) or TLR Ligands. [score:3]
The computational binding energy level of hsa-miR-34c to IKKγ 3′ untranslated region (3′-UTR) is extremely low, about −22 kcal/mol (see Table S1), indicating the binding potential between the two sequences is very high. [score:3]
We also demonstrate that one of the functional targets for miR-34c could be IKKγ an essential signaling intermediate of the NFκB inflammatory pathway. [score:3]
Fig. 4C shows a significant reduction in the amount of IKKγ protein expressed in PBMCs pre -transfected with pre-miR-34c and exposed to HMGB1-containing lysates for 24 hrs. [score:3]
miR-34c and miR-214 are Differentially Expressed in Human PBMCs Following Exposure to Damaged/necrotic Cell Lysates. [score:3]
Expression levels of miR-34c and miR-214 were assessed in conditioned media from stressed (hypoxia, serum starvation) cells. [score:3]
Levels of miR-34c and miR-214 Expression (and Pro-inflammatory Cytokine Release) Increased After Exposure of Donor PBMCs to Conditioned Media from Serum-starved and Glucose-deprived Cells. [score:3]
Our findings indicate that miR-34c expression is due to the inflammatory response in human PBMCs and partly dependent on the presence of HMGB1 in cells from which damaged lysates or conditioned media were obtained. [score:3]
Changes in miR-34c, miR-214 and miR-155 expression in PMBCs pre-incubated with 50 µM glybenclamide (Glyb) for 30 minutes before being exposed to conditioned media (MEF CM) for 48 hrs were used to assess the inflammasome pathway. [score:3]
B: Changes in miR-34c, miR-214 and miR-155 expression in PBMCs from another donor exposed to conditioned media. [score:3]
When PBMC cultures were pre-incubated with 50 µM glybenclamide for 30 minutes, and exposed to conditioned media from serum-starved and glucose-deprived cells with heat shock, levels of miR-34c expression decreased significantly in both donor PBMC cultures (Fig. 5). [score:3]
Increased expression of miR-34c has been reported in Duchenne Muscular Dystrophy, where muscle damage occurs at large scales [18], indicating that miR-34c may be a diagnostic biomarker of internal tissue damage. [score:3]
C: Fold changes in expression (as log-2-transformed RQ values) of hsa-miR-34c and hsa-miR-214 in donor PBMCs exposed to the indicated HCT116 necrotic lysates for 48 hrs. [score:3]
Both TNFα release and hsa-miR-34c expression increased significantly following exposure to HMGB1 [+/+] lysates with respect to HMGB1 [−/−] lysates (Figure 2). [score:3]
To determine whether any PAMPs or TLR ligands are associated with miR-34c or miR-214 expression changes in donor PBMCs, we stimulated the cells with various PAMPS or known TLR ligands. [score:3]
Differential Expression of miR-34c in Donor PBMCs Exposed to a Necrotic Human Carcinoma Cell Lysate. [score:3]
Changes in IKKγ mRNA and protein expression levels in human PBMCs pre -transfected with pre-miR-34c or anti-miR-34c and exposed to HMGB1 [+/+] or HMGB1. [score:3]
C: Differential expression of hsa-miR-34a, miR-34b, miR-34c and other miRs when donor PBMCs are exposed to HMGB1 [+/+] or HMGB1 [−/−] lysates for 8 hrs. [score:3]
The fold expression changes for hsa-miR-34c in donor PBMCs exposed to HMGB1 [−/−] lysates varied from 0.1 to 0.78 fold, and 2.0 to 4.5 fold following exposure to HMGB1 [+/+] lysates. [score:3]
A: Changes in fold expression (as log 2-transformed RQ values) of IKKγ mRNA levels in human PBMCs transfected with pre-miR-34c-5p or anti-miR-34c-5p and exposed to damaged HMGB1 [+/+] or HMGB1− /− lysates for 8 hrs. [score:3]
Expression of hsa-miR-34c and hsa-miR-214 is a hallmark of human PBMCs exposed to necrotic cell lysates. [score:3]
IKKγ is a Potential Functional Target of Hsa-miR-34c. [score:3]
Fig. 3 shows the fold expression changes (as log2-transformed values) of miR-34a, miR-34c, miR-214, and miR-155 after stimulation of donor PBMCs with various concentrations of TLR ligands. [score:3]
Table S1 Some computational targets of hsa-miR-34c with very low binding energies. [score:3]
Expression of miR-34c and miR-214 was negligible in all samples stimulated with the various TLR ligands. [score:3]
hsa-miR-34c and hsa-miR-214 are expressed at negligible levels in human PBMCs stimulated with various PAMPS or TLR ligands. [score:3]
Expression levels of miR-34c and miR-214 are changed when donor PBMCs are exposed to conditioned media from dying cells. [score:3]
0038899.g005 Figure 5 A: Changes in miR-34c, miR-214 and miR-155 expression in PMBCs (from one donor) exposed to conditioned media from HMGB1 [+/+] and HMGB1 [−/−] MEF cells. [score:3]
However, in PBMCs exposed to HMGB1 [−/−] cell lysates, the levelsof hsa-miR-34c expressed are significantly less. [score:3]
Donor PBMCs seeded at 15×106 cells/2 mls/well in 6-well plates were transfected with 5 nM (final concentration) of pre-miR negative control oligos (AM17110), miR-34c-5p precursor (PM11039), or anti-miR inhibitor oligos (AM11039), (Applied Biosystems, Foster City, CA), using siPORT Lipid transfection reagent (Applied Biosystems/Ambion, Austin, TX). [score:3]
Here, we demonstrate that NEMO is a functional target of an inflammation -associated miR, miR-34c. [score:3]
A: Changes in miR-34c, miR-214 and miR-155 expression in PMBCs (from one donor) exposed to conditioned media from HMGB1 [+/+] and HMGB1 [−/−] MEF cells. [score:3]
0038899.g004 Figure 4Changes in IKKγ mRNA and protein expression levels in human PBMCs pre -transfected with pre-miR-34c or anti-miR-34c and exposed to HMGB1 [+/+] or HMGB1 − /− lysates. [score:3]
We show that miR-34c expression in human PBMCs is dependent on the presence of HMGB1 within cells serving as a source of lysates or conditioned media from stressed cells. [score:3]
For quantification of IKKγ mRNA or hsa-miR-34c expression after 48 hrs of transfection, total RNA (with microRNA) was isolated using the miRNeasy mini kit (Qiagen) after exposure to damaged HMGB1 [+/+] or HMGB1 [−/−] lysates for 8 hrs. [score:3]
The fold increase in hsa-miR-34c expression was from an average of 3.4 fold (in donors exposed to HMGB1 [−/−] lysates), compared to an average of 5.7 fold (in donors exposed to HMGB1 [+/+] lysates). [score:2]
As shown in Fig. 5, both miR-34c and miR-214 were significantly expressed in cultures exposed to the conditioned media, compared to untreated cultures. [score:2]
Total mRNA was isolated from donor PBMCs and Taqman miR PCR was carried out for miR-34c, mir-214, miR-155 and the endogenous nucleolar control RNA, RNU48. [score:1]
d., of two independent experiments and normalized to the untreated (UT) samples transfected either with control miR, pre-miR-34c or anti-miR-34c, where ***indicates p<0.001, by paired Student’s t test. [score:1]
Changes in IKKγ mRNA and protein expression levels in human PBMCs pre- transfected with pre-miR-34c or anti-miR-34c and exposed to HMGB1 [+/+] or HMGB1 [−/−] lysates were evaluated. [score:1]
Transfection of Human PBMCs with Pre-miR-34c or Anti-miR-34c Oligos. [score:1]
From the microRNA profiling data, fold expression values (as log 2–transformed RQ values) for the statistically significant microRNAs (hsa-miR-34a, miR-34c, miR-214, and miR-155) were calculated for each donor after exposure to lysates or LPS (Figure 1B). [score:1]
Interestingly, miR34c seed region sequence is highly conserved in humans and chimpanzees, as shown in Fig. S4, suggesting a possible major alteration in relatively recent evolutionary time. [score:1]
B: Data shown are 48 hrs after transfection of pre- miR-34c and negative control precursor oligos into donor PBMCs. [score:1]
Using a Taqman microRNA profiling low-density PCR array we identified several microRNA genes, including miR-34c, miR-214, miR-210, miR-125b and miR-10b in human PBMCs, which are involved in the inflammatory response to damaged cells. [score:1]
Figure S4 Sequence alignment of miR-34c seed region in various species. [score:1]
This indicates that the inflammasome, shown to be activated when immune cells are exposed to necrotic/damaged cells [14], is important for the activation of miR-34c. [score:1]
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[+] score: 185
In the 29 ex vivo lung samples used for miRNA microarray profiling, qRT-PCR confirmed that expression of both SERPINE1 (fold-change >2.2) and HNF4A (fold-change >1.3) had generally higher expression in association with lower miR-34c expression; however only SERPINE1 (p = 0.05) was significantly differentially expressed between mild and moderate emphysema (Figure  3) in this cohort, and it was correspondingly upregulated in more severe disease in both TPCH-KCO (fold change >1.2, p < 0.05) and Spira (fold change >1.13) datasets (Figure  2). [score:14]
Their findings support our study, since there was down-regulation of miR-34c expression in their COPD patients, compared to controls, and also down-regulation in our moderate emphysema patients, compared to mild emphysema, indicating a similar continuum of expression with increasing disease severity. [score:11]
Among the 2,152 genes, fifty were predicted as miR-34c targets by the databases Pictar and TargetScan-and they were significantly down-regulated (>1.8 fold down-regulation) in the transfectants. [score:11]
In vitro upregulation of miR-34c in respiratory cells led to down-regulation of predicted target mRNAs, including SERPINE1, MAP4K4, ZNF3, ALDOA and HNF4A. [score:9]
Furthermore, increasing the expression of miR-34c (the miRNA with the largest fold change between the mild and moderate emphysema patient groups) in respiratory cells resulted in decreased expression of predicted mRNA targets in vitro, providing additional functional expression data. [score:9]
Increasing miR-34c expression in vitro in respiratory cell lines decreased the expression of its predicted targets, consistent with miRNA -mediated regulation of mRNA. [score:8]
Identification of miR-34c predicted targetsThe genes differentially expressed between miR-34c transfected and non -transfected cells were compared to the predicted targets of miR-34c from the TargetScan and PicTar databases. [score:8]
Of the miRNAs identified as differentially expressed in lung between moderate vs mild emphysema patients in this study, miR-34c is particularly relevant to human lung disease, as its expression is increased during normal lung development [34]. [score:8]
In the setting of reduced miR-34c expression in moderate emphysema lung (vs mild), we were able to confirm increased gene expression of two of its predicted mRNA targets, HNF4 and SERPINE1. [score:7]
Effect of miR-34c-5p over -expression on its predicted mRNA targets in respiratory cells in vitro using gene expression microarrays. [score:7]
Class comparison analysis in BRB-ArrayTools V4.2 identified 2,152 elements (48 expected by chance) affected by over -expression of miR-34c (p < 0.001) in BEAS-2B and HFL1 cell lines, the vast majority of which (90%) were down-regulated by elevated miR-34c levels. [score:6]
The genes differentially expressed between miR-34c transfected and non -transfected cells were compared to the predicted targets of miR-34c from the TargetScan and PicTar databases. [score:6]
Van Pottelberge et al. also found greater than three-fold downregulation of miR-34c in sputum from smokers with COPD, compared to those without COPD, and a direct correlation between miR-34c expression and percent predicted FEV [1][24]. [score:6]
Figure 2 Histogram of in vitro (i) and ex vivo (ii) expression of genes predicted and modulated by miR-34c over -expression in BEAS-2B and HFL1 cell lines. [score:5]
In both of these datasets, five genes (MAP4K4, SERPINE1, ALDOA, HNF4A and ZNF3) were expressed at higher levels in lung from patients with moderate or severe emphysema compared with lung from normal subjects or subjects with mild emphysema, whereas expression of miR-34c was reciprocal (Figure  2), consistent with the possibility that these five genes may be regulated by miR-34c in emphysema. [score:5]
The fold change in ex-vivo expression of all five predicted target genes inversely correlated with that of miR-34c in emphysematous lung, but this relationship was strongest for SERPINE1 (p = 0.05). [score:5]
Additionally, we have shown by qRT-PCR that SERPINE1 expression is increased in lung from patients with increasing severity of emphysema, in whom miR-34c expression is low. [score:5]
Identifying genes modulated by miR-34c using in vitro techniquesmRNAs targets of altered miRNA expression in commonly used lung cell lines were identified using in vitro techniques. [score:5]
Candidate target genes whose expression were negatively correlated to that of miR-34c in vitro (in cell lines) and ex vivo (in lung of TPCH-KCO and Spira et al. [19] subjects) were identified. [score:5]
Pre-miR™ miRNA precursor molecules (Invitrogen by Life Sciences, Carlsbad, CA) for miR-34c-5p were used to increase the expression of the miR-34c in HFL1 and BEAS-2B lung cells and the expected increase in expression of miR-34c was confirmed using TaqMan microRNAs assays (Invitrogen by Life Sciences, Carlsbad, CA). [score:4]
MiR-34c expression exhibited the greatest difference between groups with 0.3 fold lower expression in the moderate severity group. [score:4]
MiR-34c modulates expression of its putative target gene, SERPINE1, in vitro in respiratory cell lines and ex vivo in emphysematous lung tissue. [score:4]
Illumina HumanHT12 V3 transcriptome microarrays were then used to examine the expression of mRNAs in miR-34c -transfected cells, relative to cells transfected with scrambled sequence controls. [score:3]
We next examined ex vivo expression patterns of the genes found to be miR-34c responsive in vitro. [score:3]
Five miRNAs (miR-34c, miR-34b, miR-149, miR-133a and miR-133b) were significantly down-regulated in lung from patients with moderate compared to mild emphysema as defined by gas transfer (p < 0.01). [score:3]
Future studies should aim to directly demonstrate miR-34c -mediated SERPINE1 regulation in emphysema and evaluate the pathogenetic role of other miRNAs, with the goal of identifying potentially important targets for more effective treatment of COPD. [score:3]
Class comparison identified five miRNAs (miR-34c, miR-34b, miR-149, miR-133a and miR-133b) that were significantly differentially expressed between mild and moderate emphysema (p < 0.01, Additional file 1: Figure S2 & Table  2). [score:3]
Identification of miR-34c predicted targets. [score:3]
qRT-PCR confirmed similar fold differences in expression to microarray results for the two miRNAs, miR-34c and miR-133a tested (Additional file 1: Figure S3). [score:3]
We found decreased expression of miR-34c in lungs of patients with moderate emphysema, compared to mild emphysema. [score:2]
Figure 3 Histogram of qRT-PCR measurements of miR-34c regulated targets in TPCH-KCO dataset. [score:2]
Dysregulation of SERPINE1 by miR-34c could therefore be a potential mechanism involved in emphysema severity and progression. [score:2]
Our data suggest modulation of SERPINE1 by miR-34c in vitro and ex vivo. [score:1]
The probe sequence represented on the microarray was derived from the miR-34c-5p sequence. [score:1]
A microRNA (miR-34c) that technically validated and displayed high fold change difference between emphysema classes was chosen for this purpose. [score:1]
The-5p isoform of miR-34c-5p represents the 5′ arm of the hairpin precursor of the mature miRNA from which the mature sequence has been excised. [score:1]
Identifying genes modulated by miR-34c using in vitro techniques. [score:1]
The correlation of miR34c expression determined by microarrays in lung tissues to its consequent KCO measurements were not significant (p > 0.05). [score:1]
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[+] score: 182
Other miRNAs from this paper: hsa-mir-34a, hsa-mir-1-2, hsa-mir-1-1, hsa-mir-34b
In addition, we confirmed the targeting of E2F1 by miR-34 and transactivation of miR-34 by p53, and thus revealed the dual mechanisms by which miR-34 controls expression of h-eag1: directly repressing h-eag1 at the post-transcriptional level and indirectly downregulating h-eag1 at the transcriptional level through repressing E2F1. [score:10]
Downregulation of miR-34 should produce the opposite changes and upregulation of h-eag1 may mediate the cell growth-promoting effect of miR-34 downregulation. [score:10]
p53 activates miR-34 transcription; upregulation of miR-34 represses E2F1 and h-eag1; repression of E2F1 downregulates expression of h- eag1. [score:9]
Indeed, downregulation of miR-34 has been found in a wide spectrum of tumors [36] in one hand and upregulation of h-eag1 in cancer tissues on the other hand, consistent with h-eag1 being a CNS-localized voltage-gated K [+] channel that is ectopically expressed in a majority of extracranial solid tumors [39]. [score:9]
Transfection of miR-34a markedly suppressed the luciferase activities and the effect was reversed by their multiple-target anti-miRNA antisense oligonucleotides (MT-AMO) (Fig. 2A ; Supporting Figures online; Figure S5), a single oligomer capable of targeting all three members of the miR-34 subfamily [28]. [score:7]
Moreover, in the presence of p53 inhibitor, exogenously applied miR-34a retained the full ability to downregulate E2F1 (Fig. 4C & 4D ) and h-eag1 (Fig. 4E & 4F ), suggesting that miR-34 mediates the regulatory role of p53 on E2F1 and h-eag1. [score:7]
Therefore, p53 negatively regulates h-eag1 expression by a negative feed-forward mechanism through the p53− miR-34−E2F1 pathway and inactivation of p53 activity as it is the case in many cancers can thus cause oncogenic overexpression of h-eag1 by relieving the negative feed-forward regulation. [score:7]
We first demonstrated that activation of p53 by nutlin-3 induced a cell growth arrest in SHSY5Y cells, and overexpression of E2F1 alleviated the cell growth inhibition and so did transfection with the MT-AMO to knock down miR-34 (Fig. 7A & 7B ). [score:6]
Our study herein revealed that miR-34, a known transcriptional target of p53, is an important negative regulator of h-eag1 through dual mechanisms by direct repression at the post-transcriptional level and indirect silencing at the transcriptional level via post-transcriptionally repressing E2F1 that we have established to be a transactivator of h- eag1. [score:6]
Intriguingly, it has been demonstrated that in vertebrates miR-34 is initially expressed widespread throughout the brain in early stage of development and expression becomes limited to the anterior region of the hindbrain in later stages [37]. [score:6]
The major finding includes identification of E2F1 as a key transcriptional activator of h- eag1 and miR-34 as an important translational inhibitor of h-eag1. [score:5]
When p53 activity increases in response to environmental and cellular stresses, miR-34 is deemed to increase, and the increased miR-34 will decrease E2F1 to diminish h- eag1 gene transcription and will also repress h-eag1 protein translation as well; diminishment of h-eag1 expression and function then results in a shut-down of cell proliferation or a cell cycle arrest. [score:5]
0020362.g004 Figure 4(A & B) Effects of p53 activation by Mdm2 inhibitor nutlin-3 (1 µM) on expression of miR-34, E2F1 and h- eag1 at mRNA and protein levels. [score:5]
miR-34 has been known to be a direct transcriptional target of p53 [33]– [36] and to mediate the apoptotic action of p53. [score:4]
Figure S5The multiple-target anti-miRNA antisense oligonucleotide fragment (MT-AMO) used to knock down all three different isoforms of has- miR-34 (miR-34a, miR-34b and miR-34c). [score:4]
These results indicate that miR-34 regulates h- eag1 expression through at least two mechanisms. [score:4]
Cells were pretreated with nutlin-3 to activate p53 and then transfected with the plasmid carrying E2F1 cDNA for overexpression (E2F1-P) or MT-AMO to knockdown miR-34; control cells (Ctl/Lipo) were mock -treated with lipofectamine 2000. [score:4]
It appears that h-eag1 is a terminal effecter component in the p53− miR-34−E2F1 pathway for expression regulation and functional signaling. [score:4]
We reasoned that if E2F1 and miR-34 are indeed important in the expression regulation of h-eag1, then we should see a positive correlation between E2F1 and eag1 levels and an inverse relationship between miR-34 and h- eag1 levels. [score:4]
These findings not only help us understand the molecular mechanisms for oncogenic overexpression of h-eag1 in tumorigenesis but also uncover the cell-cycle regulation through the p53− miR-34−E2F1−h-eag1 pathway. [score:4]
Figure S6Effects of miR-34b and miR-34c on expression of E2F1 (A) and h-eag1 (B) at the protein level in SHSY5Y cells, assessed by. [score:3]
Finally, we demonstrated that cell growth controls at the level of p53, miR-34 or E2F1 were related to h-eag1 expression. [score:3]
To experimentally establish miR-34:h- eag1 interaction, we inserted a fragment of 3′UTR of h- eag1 containing the miR-34 target sites into the position downstream the luciferase gene in the pMIR-REPORTTM vector. [score:3]
And we identified multiple binding sites for a tumor-suppressor miRNA subfamily miR-34 (including miR-34a, miR-34b and miR-34c) in the 3′UTR of h- eag1 mRNA (Figure S4). [score:3]
RA: retinoic acid, which has been shown to enhance miR-34 expression; E2F1/3: E2F1 and E2F3. [score:3]
Figure S9Expression correlations between E2F1 and h-eag1 and between miR-34 and h-eag1. [score:3]
Moreover, this low level of miR-34 may also explain the enriched expression of E2F1 in brain [40]. [score:3]
Indeed, p53 activation by Mdm2 inhibitor nutlin-3 (1 µM) increased miR-34 level (Fig. 4A ), and simultaneously decreased E2F1 and h- eag1 mRNA concentrations (Fig. 4A ) and protein levels (Fig. 4B ). [score:3]
miR-34a, miR-34b, and miR-34c (Figure S4), and their antisense inhibitor oligonucleotides (MT-AMO) (Figure S5) were synthesized by Integrated DNA Technologies, Inc. [score:3]
Ctl: cells transfected with the luciferase vector alone; MT-AMO: the multiple-target anti-miRNA antisense oligonucleotides to miR-34a, miR-34b and miR-34c, co -transfected with the luciferase vector and miR-34a or miR-34c. [score:3]
This low expression of miR-34 may partially underlie the high abundance of eag1 in brain [39]. [score:3]
First, miR-34 directly represses h-eag1 protein. [score:2]
Moreover, these findings place h-eag1 in the p53− miR-34−E2F1−h-eag1 pathway with h-eag as a terminal effecter component and with miR-34 (and E2F1) as a linker between p53 and h-eag1. [score:1]
Figure S8Multiple complementary motifs between each of the three isoforms of has- miR-34 and the 3′UTR of E2F1 mRNA. [score:1]
This implies that h-eag1 executes the cell-cycle checkpoint signal from p53 transmitting along the p53− miR-34−E2F1−h-eag1 pathway (Fig. 6 ). [score:1]
miR-34 as a post-transcriptional repressor of h-eag1. [score:1]
The same observations were expanded to miR-34b and miR-34c and to MCF-7 cells (Figure S6 & S7). [score:1]
Thus, changes of p53 activity are deemed to change the level of miR-34 thereby those of E2F1 and h-eag1 as well. [score:1]
0020362.g003 Figure 3 miR-34 as a post-transcriptional repressor of E2F1. [score:1]
The MT-AMO tested in this study was designed to integrate the AMOs against miR-34a, miR-34b and miR-34c into one AMO unit. [score:1]
Effects of the p53− miR-34−E2F1−h-eag1 pathway on cell proliferation. [score:1]
MT-AMO: an antisense oligomer to miR-34a, miR-34b and miR-34c; miR+AMO: co-transfection of miR-34a and MT-AMO; NC-miR: scrambled negative control miRNA. [score:1]
These above data allowed us to propose a new signaling pathway p53− miR-34−E2F1−h-eag1 (Fig. 6 ). [score:1]
0020362.g006 Figure 6Proposed mo del of the p53− miR-34−E2F1−h-eag1 signaling pathway. [score:1]
Figure S4Multiple complementary motifs between each of the three isoforms of has- miR-34 and the 3′UTRs of h -eag1 mRNA (A) and h- erg1 mRNA (B). [score:1]
MT-AMO: an antisense oligomer to miR-34a, miR-34b and miR-34c; +MT-AMO: co-app;lication of miR-34c and MT-AMO. [score:1]
Proposed mo del of the p53− miR-34−E2F1−h-eag1 signaling pathway. [score:1]
Second, miR-34 represses E2F1 protein, leading to reduced transcription of h- eag1. [score:1]
This latter effect also explains partially the effectiveness of miR-34 to decrease h- eag1 mRNA. [score:1]
0020362.g007 Figure 7Effects of the p53− miR-34−E2F1−h-eag1 pathway on cell proliferation. [score:1]
These findings may be extended to h-erg-1 K [+] channel (or HERG): h-erg1 may also be a component of the p53− miR-34−E2F1 pathway, according to our data shown in Figures S3, S8 and S9. [score:1]
We analyzed the summation of the two bands to represent the total h-eag1 protein level and both bands were found affected by miR-34 and MT-AMO. [score:1]
Figure S7 miR-34 as a post-transcriptional repressor of h-eag1 in MCF-7 human breast cancer cells. [score:1]
0020362.g002 Figure 2 miR-34 as a post-transcriptional repressor of h-eag1. [score:1]
miR-34 as a post-transcriptional repressor of E2F1. [score:1]
Based on our findings, we were able to establish a novel signaling pathway: p53− miR-34−E2F1− h-eag1. [score:1]
We confirmed that transfection of miR-34a reduced E2F1 protein levels by ∼68% in SHSY5Y cells (Fig. 3A ) and the same results were obtained with miR-34b and miR-34c (Figure S6 & S8). [score:1]
In adult, miR-34 is absent from forebrain and midbrain and present only in the caudal ventral and lateral isthmus and hindbrain nuclei [38]. [score:1]
Recent studies have shown that members of the miR-34 family possess anti-proliferative potential and induce cell cycle arrest, senescence, and/or apoptosis [31]– [36]. [score:1]
[1 to 20 of 59 sentences]
9
[+] score: 171
Other miRNAs from this paper: hsa-mir-21, hsa-mir-29b-1, hsa-mir-29b-2, hsa-mir-34a, hsa-mir-34b
The fact that miR-34 family is up-regulated in human cancer and its up-regulation is associated with poor prognosis, seems to be conflicted with tumor suppressive role of miR-34 demonstrated in previous reports. [score:9]
Previously, Roy et al. demonstrated that the expression of miR-34a and miR-34c were down-regulated in colon cancer specimens compared to adjacent colonic mucosa in small number expression analysis (n = 10) [27]. [score:7]
In addition, the association between miR-34 expression and TP53 mutation, global gene expression was examined to clarify possible mechanisms of miR-34 regulation. [score:7]
The average expression of miR-34b and miR-34c was used to analyze the association of miR-34b/c expression and global mRNA expression. [score:7]
Considering the tumor suppressive function of miR-34 as shown previously, we had expected that miR-34 family was suppressed in human colon cancer and decreased expression was associated with poor prognosis when we started the present study. [score:7]
Although it has been well known that miR-34 was direct target of p53, a recent study demonstrated that mouse mo dels that contain deletions for miR-34a, miR-34b and miR-34c still retain p53 function and the expression of these microRNAs is largely independent of p53 status [38]. [score:6]
3) The expression of miR-34 family was not associated with TP53 mutational status although pathway analysis showed association between miR-34 expression and TP53 transcriptional activity. [score:6]
Ingenuity Pathway Analysis (IPA) predicts transcription regulators that potentially associated with miR-34 up-regulation. [score:5]
One possible explanation of our findings is that miR-34 up-regulation in stromal tissues might reflect inflammation of tumor tissues, as it is well known that inflammation contributes to the development of cancer, including colon cancer [35]. [score:5]
On the basis of these findings, we analyzed epithelial and stromal expression of miR-34 separately using laser microdissection technique to examine if cancer cells express miR-34. [score:5]
To examine where the miR-34 family is expressed in, we extracted RNA from cancer epithelium, cancer stroma, normal adjacent epithelium and normal adjacent stroma separately using laser microdissection for miR-34 expression analysis (S3 Fig). [score:5]
Several converging evidence demonstrated that ectopic expression of miR-34 family induces apoptosis, senescence, cell cycle arrest and inhibits migration and invasion [9, 13]. [score:5]
Expression of miR-34 is lost in colon cancer which can be re-expressed by a novel agent CDF. [score:5]
Considering the fact that miR-34 was expressed in stromal tissues predominantly, miR-34 detected in macro-dissected cancer tissues seems to be mainly regulated by cancer stromal tissues harboring wild-type p53. [score:4]
To address this, we examined whether miR-34 expression was associated with TP53 mutational status. [score:4]
That may be a plausible explanation that the expression of miR-34 family was not associated with TP53 mutational status of cancer. [score:4]
As shown in Fig 3, the expression of miR-34 family was not associated with TP53 mutational status. [score:4]
Although previous reports have demonstrated that down-regulation of miR-34 family members was associated with poor prognosis in several types of malignancies [17– 20], the prognostic value of miR-34 family in colon cancer has not been systematically studied. [score:4]
The association between miR-34 expression and TP53 mutation. [score:4]
However, the reason for the up-regulation of miR-34 in stromal tissue is still unclear. [score:4]
Contrary to our expectations, the expression of miR-34 family was not associated with TP53 mutational status in either the American and Chinese cohorts (Fig 3A and 3B). [score:4]
TP53 mutation is not associated with the expression of miR-34 family. [score:4]
In addition, the expression levels of miR-34 seemed to be the highest in cancer stromal tissues. [score:3]
Correlation analysis between Affymetrix microarray data and miR-34 family expression in 56 colon tumors. [score:3]
There was no significant correlation between tumor/stroma ratio and expression levels of miR-34 (S5 Fig). [score:3]
The finding may demonstrate one more possibility that different level of miR-34 expression could be significant for miR-34 -mediated interaction between human colon cancer and stroma. [score:3]
MiR-34b and miR-34c are co-transcribed from a bicistronic transcript and their expression should be highly correlated [9]. [score:3]
Correlation between miR-34 expression and survival. [score:3]
Ingenuity Pathway Analysis (IPA 8.0, Redwood city, CA) was performed to identify molecules and pathways potentially involved with the expression of miR-34 family. [score:3]
In colorectal cancer, prior studies have focused on a tumor suppressive function for miR-34 family as shown in many kinds of malignancy [9]. [score:3]
The comparison of miR-34 expression between colon epithelium and stroma with laser microdissection (LMD) technique. [score:3]
As a matter of fact, the present data is consistent with several recent reports analyzing miR-34 expression and its prognostic value. [score:3]
To our knowledge, this is the largest and systematic study analyzing miR-34 expression in human colon cancer. [score:3]
miR-34 family tends to be expressed predominantly in stromal tissue. [score:3]
Since miR-34a, miR-34b, and miR-34c were each systematically altered in colon tumors, we next attempted to determine possible reasons for this altered expression. [score:3]
Therefore, miR-34 family is thought to be important mediator of p53’s tumor-suppressive activities. [score:3]
The miR-34 family is largely considered to be a tumor suppressor miRNA [9]. [score:2]
We performed TaqMan qRT-PCR of miR-34a, miR-34b and miR-34c in colon tumors and adjacent non-tumor tissues from both the American and the Chinese cohorts. [score:1]
To evaluate the issue, we set out to systematically study the expression of the miR-34 family in two large, independent cohorts of colon cancer patients. [score:1]
*Cases with low miR-34b and/or low miR-34c. [score:1]
The miR-34 family does not associated with increased stromal area. [score:1]
Further analysis is needed to clarify the function and miR-34 -mediated interaction between human colon cancer and stroma. [score:1]
In addition, we have performed H&E on 82 of the tissues from the American cohort to examine if there was an association with increased stromal area and any of the miR-34 family. [score:1]
In order to investigate potential reasons for the altered expression of the miR-34 family, we used Affymetrix microarray data to identify genes that correlated with miR-34a and -34b/c and examined these genes with Ingenuity Pathway Analysis (IPA). [score:1]
**Cases with both high miR-34b and high miR-34c. [score:1]
Several studies indicate that p53 can bind to the promoter and activate the transcription of miR-34 family [10– 12]. [score:1]
This family consists of three members: miR-34a, generated from a transcriptional unit on the human chromosome 1p36, and miR-34b and miR-34c, which are generated by processing of a bicistronic transcript from chromosome 11q23. [score:1]
In conclusion, we showed novel aspects of miR-34 family in human colon cancer. [score:1]
The demonstrated findings imply that increased miR-34 in stromal tissues may have important roles in colon cancer progression. [score:1]
[1 to 20 of 49 sentences]
10
[+] score: 160
Other miRNAs from this paper: hsa-mir-34a, hsa-mir-34b
In particular, downregulated miR-34c is a critical factor that contributes to malignancy in human laryngeal carcinoma, while its overexpression inhibits cell proliferation and induces apoptosis via targeting of c-Met (11). [score:10]
miR-34c-3p overexpression reduces expression of Notch2, but miR-34c-5p overexpression does not. [score:7]
Overexpression of either miR-34c-3p or miR-34c-5p in U251 cells caused inhibition of proliferation, cell apoptosis, S phase arrest and cell invasion suppression. [score:7]
Previous studies have demonstrated that miR-34 family overexpression regulated proliferation and senescence through inhibition of E2F3, BCL-2 and MYC (16, 21, 22). [score:6]
In the present study, it was found that the value of Notch2 was downregulated after transfection with miR-34c-3p, indicating that the effect of miR-34c-3p on cell proliferation inhibition occurred via the Notch2 pathway. [score:6]
By contrast, Notch2 expression was not downregulated following transfection with miR-34c-5p mimics (Fig. 6B). [score:6]
Our experiments demonstrated that miR-34c-3p or miR-34c-5p overexpression inhibited the invasive ability of U251 and U87 glioblastoma cells in vitro. [score:5]
Although Luan et al have demonstrated that miR-34a overexpression inhibited cell migration and invasion in the glioma cell line, U251 (18), the effect of miR-34c on gliomas is unknown. [score:5]
In the present study, we also predicted that Notch2 was the precise intracellular target of miR-34c by using miRanda, TargetScan and PICTAR databases, which was different from that in a previous study (14). [score:5]
Although miR-34c-3p and miR-34c-5p have been established as tumor suppressors in a variety of tumors (13, 14), their targets and functions in glioma are largely unknown. [score:5]
It is well-documented that the mature miRNA-34 family, as tumor suppressors, shows a global decrease in expression in many different human cancers, including laryngeal carcinoma (11), prostate cancer (16) and cervical carcinoma (17). [score:5]
Several online softwares, including miRanda, TargetScan and PICTAR, predicted that the sequence between nucleotides 2045 and 3073 is likely targeted by miR-34c-3p and miR-34c-5p. [score:5]
To confirm apoptosis following miR-34c-3p or miR-34c-5p overexpression in glioma cell lines, transfected cells were analyzed for Annexin V expression by flow cytometry. [score:5]
The S phase fraction of normal and NC cells was 20.66 and 13.61%, respectively, and this increased significantly with miR-34c-3p overexpression to 48.83%; however, this effect did not occur with miR-34c-5p overexpression (11.04%). [score:5]
These data suggested that the tumor suppressor activity of miR-34c-3p in glioblastoma cells may be regulated by the Notch pathway, but that of miR-34c-5p is not. [score:4]
miR-34c-3p and miR-34c-5p are downregulated in malignant glioma. [score:4]
A significant difference in proliferation inhibition was only observed in U87 cells transfected with pre-miR-34c-3p (72.37%) against cells transfected with pre-miR-34c-5p (94.89%), suggesting alternative regulatory pathways are involved (Fig. 2). [score:4]
However, transfection with miR-34c-5p only inhibited the cell proliferation to 80.84% in U251 cell lines (P<0.05 vs. [score:3]
However, the present study showed that expression of miR-34c-3p or miR-34c-5p in U251 and U87 cells caused S phase arrest, indicating that the function of miR-34c was different but complementary to that of miR-34a. [score:3]
Furthermore, miR-34c-3p expression clearly produced cell apoptosis and S phase arrest, while no apoptosis or major cell cycle changes were observed with miR-34c-5p. [score:3]
However, the G2/M phase fraction in normal and NC cells were 4.4 and 4.31%, respectively, while miR-34c-3p and miR-34c-5p overexpression increased this fraction to 21.6 and 21.83%, respectively. [score:3]
While miR-34c-3p overexpression decreased the G0/G1 phase to 44.05%, transfected miR-34c-5p did not affect the G0/G1 phase (79.07%). [score:3]
The proliferation inhibition effect of miR-34c-5p was not observed. [score:3]
miR-34c-3p and miR-34c-5p may target different mRNAs and thus display different results through dissimilar pathways in U87 cells. [score:3]
miR-34c-3p inhibits U251 and U87 cell proliferation, but miR-34c-5p only affects U251 cells. [score:3]
miR-34c-3p and miR-34c-5p overexpression inducesapoptosis in glioma cell lines. [score:3]
These results implied that miR-34c-3p and miR-34c-5p may function as tumor suppressors in glioma cells in vitro. [score:3]
Expression of miR-34c-3p and miR-34c-5p were quantified by miR-qRT PCR using SYBR Green RealtimePCR master mix (Toyobo Co. [score:3]
In the present study, we profiled miR-34c-3p and miR-34c-5p expression in glioblastoma and normal brain tissue. [score:3]
These findings suggested that loss of miR-34c-3p or miR-34c-5p expression may be critical in glioblastoma pathogenesis. [score:3]
Transfection of U251 and U87 cells with miR-34c-3p mimics resulted in 78.42 and 72.37% growth inhibition, compared with normal and NC groups (Fig. 2; P<0.05; 96 h after transfection). [score:2]
miR-34c-3p mimics reduced Notch2 expression by ~15% compared to normal or NC cells (Fig. 6A) in U251 and U87 cell lines. [score:2]
Hence, we hypothesize that the mechanism of miR-34c-5p may be different from that of miR-34c-3p. [score:1]
These results suggested that transfection with miR-34c-3p or miR-34c-5p in U251 cells and with miR-34c-3p in U87 cells produced S phase arrest with G0/G1 reduction. [score:1]
In the present study, we analyzed the function of miR-34c-3p and miR-34c-5p in human glioblastoma. [score:1]
miR-34c has two identified mature miRNAs: miR-34c-3p and miR-34c-5p (12). [score:1]
Spearman’s rank correlation test was used for association analysis between endogenous miR-34c-3p and miR-34c-5p levels and pathological grading. [score:1]
Transfection of either pre-miR-34c-3p or pre-miR-34c-5p mimics in U251 and U87 cells significantly increased their respective levels, while transfection with scrambled miRNA negative control (NC) had no effect (Fig. 1B). [score:1]
To evaluate the impact of miR-34c-3p and miR-34c-5p expression on cell invasion, U251 and U87 cells were treated with oligonucleotides and placed on 8 μm-pore size insert chambers coated with a mixture of extracellular matrix proteins. [score:1]
Many investigators have demonstrated that expression of the miR-34 family resulted in G0/G1 cell cycle arrest in diverse cellular contexts (19, 20). [score:1]
To the best of our knowledge, this study provided the first evidence of miR-34c-3p and miR-34c-5p values in glioma patients’ tissues. [score:1]
Notably, miR-34c-3p and miR-34c-5p treatment produced significant levels of apoptosis relative to those of the controls, while no significant sub-G0 population was observed. [score:1]
Our findings suggest important roles of miR-34c-3p and miR-34c-5p in glioma etiology and provide potential candidates for treating malignant glioma. [score:1]
Effect of miR-34c-3p and miR-34c-5p on the cell cycle of U251 and U87 cell lines. [score:1]
Similarly, U251 and U87 cell lines also exhibited a small amount of endogenous miR-34c-3p or miR-34c-5p (Fig. 1B), indicating an etiological role of miR-34c-3p and miR-34c-5p reduction in glioma progression. [score:1]
Although, no significant changes were observed with miR-34c-5p transfection in U87 cells or in the normal or NC groups. [score:1]
However, only miR-34c-3p -treated cells underwent apoptosis in U87 cell lines (3.56%), but no apoptosis was observed in miR-34c-5p -treated U87 (1.91%), normal (1.53%) or NC (1.68%) cells (Fig. 3B). [score:1]
The miR-34c-3p and miR-34c-5p expression levels in malignant glioma were measured. [score:1]
The miR-34 family (miR-34a, miR-34b and miR-34c), which is described as a p53 effector, has antiproliferative and pro-apoptotic functions (9, 10). [score:1]
Administration of miR-34c-3p and miR-34c-5p mimics decreased the percentage of cells in G0/G1 phase to 44.24 and 43.58%, respectively. [score:1]
However, the level of Notch2 was not significantly different between the miR-34c-5p and control groups. [score:1]
U251 and U87 glioblastoma cell lines were transfected with miR-34c-3p or miR-34c-5p mimics. [score:1]
Cell invasive ability is depressed by miR-34c-3p and miR-34c-5p mimic transfection. [score:1]
The number of U251 cells invading through the Matrigel following miR-34c-3p and miR-34c-5p mimic treatment was 66.75±10.63 and 60.63±15.32 cells per field respectively, which was lower than that of the normal (90.88±10.38 cells per field) and the NC (104.75±12.06 cells per field) groups (Fig. 5A). [score:1]
More experiments are required to further validate the effect of miR-34c-3p on the Notch2 pathway. [score:1]
[1 to 20 of 55 sentences]
11
[+] score: 149
Other miRNAs from this paper: hsa-mir-34a, hsa-mir-34b
The re -expression of miR-34 led to a marked reduction in the expression of its target gene, Notch-1. The loss of expression of miR-34 in colon cancer is in part due to promoter hypermethylation of miR-34, which can be re-expressed with our novel agent CDF, suggesting that CDF could be a novel demethylating agent for restoring the expression of miR-34 family, and thus CDF could become a newer therapeutic agent for the treatment of colon cancer. [score:13]
This down-regulation was attributed to promoter hypermethylation, because we found that the treatment of colon cancer cells with 5-aza-2´-deoxycytidine, a methyltransferase inhibitor, markedly induced the levels of miR-34a and miR-34c expression. [score:8]
Our current observation on colorectal cancer tissues and those reported by others show that miR-34 is downregulated in colorectal cancer, suggesting that downregulation of this microRNA may partly contribute to the unregulated cellular growth and drug resistance that occurs in colorectal cancer. [score:8]
We found that the expression of miR-34a and miR-34c was down-regulated in colon cancer specimens compared to normal colonic mucosa and the loss of expression was also consistent with data from colon cancer cell lines. [score:7]
Indeed, up-regulation of miR-34 has been shown to induce cell-cycle arrest, inhibition of invasion and migration and p53 induced apoptosis [16, 17]. [score:6]
Figure 2 CDF upregulates the expression of miR-34a and miR-34c in different colon cancer cell lines. [score:6]
Our current observation that CDF induces the expression of miR-34a and miR-34c in chemo resistant and p53 defficient colon cancer cells, which suggests that CDF is effective in re -expressing miR-34 and could be a potential therapeutic agent for colorectal cancer. [score:5]
Numerous cellular events, including deregulated expression of microRNAs (miRNAs), specifically the family of miR-34 consisting of miR-34a, b and c, is known to regulate the processes of growth and metastasis. [score:5]
Emerging evidence suggests that p53 acts as a transcription factor to increase the expression of the miR-34 family members which, in turn, modulate cell cycle progression, senescence and apoptosis, inhibition of invasion and migration [12, 13]. [score:5]
Likewise, CDF was very effective in the re -expression of miR-34a and miR-34c, which was consistent with inhibition of cell growth of both chemo-sensitive and chemo-resistant colon cancer cells. [score:5]
Figure 1 miR-34a and miR34c are downregulated in colon cancer. [score:4]
miR-34a and miR-34c are down-regulated in colon cancer. [score:4]
Moreover, we also assessed the expression of miR-34 in colon cancer cell lines treated with our newly developed synthetic analogue of curcumin referred as difluorinated curcumin (CDF) compared to well known inhibitor of methyl transferase. [score:4]
It is essential to develop the strategy for restoring the expression of miRs specifically the family of miR-34 which are dysregulated in cancer. [score:4]
The levels of miR-34a and miR-34c in histologically normal and colon cancer tissues, as determined by quantitative real time RT-PCR revealed that both miR-34a and miR-34c were significantly down-regulated in colon cancer (Figure  1A, B). [score:4]
In view of this, it is tempting to speculate that p53 -mediated processes of apoptosis in colon cancer cells could be affected by down-regulation of miR-34. [score:4]
CDF up-regulates miR-34a and miR-34c in different colon cancer cell lines. [score:4]
Members of the miR-34 family act as a tumor suppressor, hence their reduction or loss in colonic mucosa leads to malignancy as reported in other cancers [30]. [score:3]
As shown in Figure  2C and 2D, both CDF (experimental agent) and Aza-dC (control agent), led to increased expression of miR-34a and miR-34c in SW620 cells. [score:3]
Our current data demonstrated that CDF significantly induced the expression of both miR-34a and miR-34c in HCT116CR, HCT116p53−/− and SW620 that were either in chemo-resistant or p53-defficient (Figure  2). [score:3]
Family of miR-34 that includes 34a, b and c has been reported to inhibit CSLCs [9]. [score:3]
Quantitative real-time RT PCR was performed with RNA isolated from formalin fixed paraffin embedded (FFPE) normal and colon cancer tissues (n = 10 for each group) to determine the expression of miR34a (A) and miR34c (B). [score:3]
In conclusion, our findings demonstrate that the expression of miR-34a and miR-34c is greatly reduced in colon cancer. [score:3]
Additionally, we used HCT116 CR cells to examine how CDF acts for miR34 expression in drug resistance. [score:3]
Silencing of miR-34 expression due to its promoter hypermethylation of the CpG site has been documented in colon cancer [28, 31], suggesting that the use of demethylating agents like azacitidine (Aza-dC) and decitabine could be useful for the treatment of solid tumors [32] although these agents show unacceptable side effects. [score:3]
Interestingly, re -expression of miR34a and miR34c levels was found to be greater in HCT116p53−/− cells indicating that CDF may act independent of p53 status (Figure  2A, 2B). [score:3]
We further extended our study for CpG methylation analysis of the miR-34a promoter only, since its expression was found to be relatively higher than miR-34b, miR-34c in all human tissues except lung [33]. [score:3]
Our primary objective was, therefore to determine whether, CDF would modulate miR-34 expression in colon cancer cells. [score:3]
Therefore, demthylation is likely to enhance the expression of miR-34. [score:3]
So far, no studies have been performed to determine whether of miR-34 could be expressed in colon cancer by any novel agent(s). [score:3]
Archival formalin-fixed paraffin-embedded tissues from normal colonic mucosa and colon tumors were obtained from the Pathology Service of the John D. Dingell VA Medical Center; Detroit through the Wayne State University IRB approved protocol to isolated RNA for assessing the expression of miR-34 family. [score:3]
The family of miR-34, that includes miR-34a, b and c, has been known to regulate several cellular events, including cell cycle, cell migration and apoptosis [16, 17]. [score:2]
Assuming that CDF may act as a demethylating agent, we compared the effects of CDF with Aza-dc on the expression of miR-34a and miR-34c in colon cancer SW620 cells. [score:2]
To investigate whether the effect of CDF on miR-34 expression is p53 dependent, we extended our study using HCT116p53−/− and SW620 (p53 mutant, where G > A mutation in codon 273 of the p53 gene results in an Arg > His substitution) cell lines. [score:2]
Although the reason is unclear, why CDF could not induce miR-34 in HCT116Wt cell, one possibility could be the over growth making CDF less available to induce miR-34. [score:1]
However, little is known whether agent(s) that modulates colon CSLCs would also modulate the family of miR-34 in colon cancer cells or not. [score:1]
To determine the miRNA-34 levels, RNA isolated from FFPE tissues using miRNeasy FFPE Kit (Qiagen) and from cultured cells using miREasy kit (Qiagen) was utilized. [score:1]
Although the precise mechanism(s) by which CDF induces miR-34a and miR-34c has not been fully elucidated, our current data suggest that demethylation of the respective promoter of miR-34a and miR-34c by CDF could be one possibility. [score:1]
This relevant information prompted us to determine whether CDF could be utilized to modulate the family of miR-34, and if so, whether CDF -induced modulation of miR-34 could in part be attributed to epigenetic alterations, specifically the methylation status of the promoter of miR-34. [score:1]
[1 to 20 of 39 sentences]
12
[+] score: 141
Thrombospondin 1 (THBS1) expression was analysed since this transcript was predicted to be a target of the miR-34b-5p both by the TargetScan and by the MiRDB databases, and furthermore, the microarray data showed that THBS1 expression was reduced in miR-34b transfected cells, but up-regulated in cells transfected with miR-34c. [score:12]
We found that 17% of the transcripts down-regulated by miR-34c were predicted miR-34a/c-5p family targets, whereas only 0.9% of the up-regulated transcripts were predicted miR-34c targets. [score:11]
Only one transcript had opposite changed expression level, THBS1 that were downregulated by miR-34b and upregulated by miR-34c (Fig.   2a). [score:9]
A number of validated miR-34 family mRNA targets, such as cyclin -dependent kinase 4 (CDK4) [8], cyclin -dependent kinase 6 (CDK6), lymphoid enhancer -binding factor 1 (LEF1) [16], Met Proto-oncogene (MET) [17] and NOTCH1 [18] were among the 305 transcripts that were downregulated by both mimics. [score:6]
Nevertheless, similarities in the transcriptome changes and repression of CDK4 and CCND1 expression are observed, indicating that the annotated miR-34b still recognises some miR-34 target genes. [score:5]
We first assessed the expression of three selected miR-34 targets by qPCR. [score:5]
Functional analyses demonstrated that the miR-34b expressed in the MDA-MB-231 cells had tumour suppressive capacity resembling that of miR-34c, while the annotated miR-34b did not. [score:5]
We observed changed expression of 101 genes known to be involved in cell migration upon introduction of miR-34c (p = 1.78E-08), while only 78 (p = 9.44E-07) genes were affected by over -expression of miR-34b. [score:5]
To further validate the gene lists, we compared the transcripts that were changed by miR-34 mimics with genes identified as predicted targets of the miR-34 family using TargetScan software (version 7.1). [score:4]
35 transcripts were regulated by both miR-34b and miR-34c, all of these were changed in the same direction. [score:3]
CDK4 and cyclin D1 (CCND1) were chosen since these are validated miR-34 targets 8, 22. [score:3]
We found that miR-34c was expressed at a higher level than miR-34b in control -transfected MDA-MB-231 cells (342 versus 19 reads, respectively, Supplementary data  S5). [score:3]
15 transcripts were regulated by both miR-34b and miR-34c, all of these were regulated in a similar manner. [score:3]
In conclusion, the miR-34 family is an important group of miRNAs with anti-tumorigenic effects, and the precise knowledge of the expressed isomiRs and their function is critical. [score:3]
We first examined the global transcription response to each miR-34 mimic by mRNA expression profiling using microarray 48 hours after transfection. [score:3]
Overexpression of any of the miR-34 isoforms decreased the transcript levels CDK4 and CCND1 48 h after transfection (Fig.   4b). [score:3]
These levels are thought to be below the functional level [32], so it was expected that MDA-MB-231 cells could be targeted by miR-34 replacement therapy. [score:3]
The miR-34 miRNAs are tumour suppressors and are critical mediators in the p53 pathway 8, 9. In particular, it has been shown that the miR-34 family members reduce cell growth, induce apoptosis and affect cell migration 10, 11. [score:3]
Interestingly, the exposed nucleotides for miR-34b pairs perfectly to the predicted miR-34 binding site in the 3′ UTR of THBS1 mRNA, while as a result of the shift in the exposed nucleotides for miR-34b-5p’, this seed sequence is no longer complementary to the target mRNA. [score:3]
The IPA analyses revealed that miR-34c affected the expression levels of 50 genes involved in cell morphology while miR-34b only affected 30 (p = 3.46E-07 and p = 5.09E-03, respectively). [score:3]
52 transcripts were regulated by both miR-34b and miR-34c, all of these were regulated in a similar manner. [score:3]
MiR-34b and miR-34c are encoded from the same locus situated on chromosome 11q23, and expressed as a bicistronic transcript. [score:3]
Gene expression analyses of miR-34b and miR-34c transfected cells identified a number of genes involved in cellular proliferation (165 and 212 genes, p = 1.49E-08 and p = 1.71E-10, respectively). [score:3]
The analysis predicted changes in several biological functions known to be regulated by the miR-34 family, such as cell growth, apoptosis, cell morphology and cell migration (Fig.   2b and Supplementary data  S3). [score:2]
76 mRNAs were regulated by both miR-34b and miR-34c, all of these were changed in a similar manner. [score:2]
IPA analysis revealed that miR-34c regulated 167 transcripts implicated in apoptosis, while miR-34b affected 105 transcripts involved in the same process (p = 8.39E-14 and p = 1.10E-04, respectively). [score:2]
We separately introduced synthetic mimics representing human annotated versions of the miR-34b-5p (miR-34b) and miR-34c-5p (miR-34c) into the breast cancer cell line MDA-MB-231. [score:1]
Cells were transiently transfected with mimics representing miR-34b or miR-34c or a negative control mimic. [score:1]
Furthermore, miR34b-5p’ reduced the growth of MDA-MB-231 cells more efficiently than miR-34b, although not quite as strong as observed for miR-34c (Fig.   4c). [score:1]
The arrow indicates cells with changed morphology in miR-34c transfected cells. [score:1]
More specifically, in contrast to the spindle-shaped, mesenchymal-like morphology of untreated and control treated cells, the cells transfected with miR-34c appeared rounder, with a more transparent cytoplasm. [score:1]
MiR-34b-5p and miR-34c-5p exert different functions in MDA-MB-231 cells. [score:1]
Moreover, miR-34b-5p’, unlike the mimic representing the annotated miR-34b, induced apoptosis as indicated by increased caspase 3/7 activity, at even higher level than observed for cells transfected with miR-34c (Fig.   4d). [score:1]
The annotated miR-34c-5p was in accordance with the observed miR-34c sequences, both at the 5′ and the 3′ end (95%). [score:1]
The human miR-34 family consists of three members, miR-34a, miR-34b, and miR-34c. [score:1]
We observed a reduced cell growth capacity after transfection with miR-34c, while miR-34b only had a minor, delayed effect on cell growth (Fig.   2c). [score:1]
In humans miR-34b-5p has an additional base at the 5′ end, shifting its seed sequence by one base, relative to the other miR-34 family members as annotated in databases like miRBase and miRNAMap 2.0 and found in scientific reviews 13– 15. [score:1]
Loss of miR-34 is strongly associated with cancer and miR-34 replacement therapy is currently in clinical trials for treatment of primary liver cancer and other selected cancer types with liver metastasis [12]. [score:1]
The miR-34 family seed sequence is highlighted in red. [score:1]
Degradation would be consistent with the observation that the levels of miR-34b in most of the samples are lower than miR-34c, even though they originate from the same transcript. [score:1]
In line with this, we observed that miR-34c had a profound effect on the cellular morphology of MDA-MB-231 cells 72 hours after transfection, whereas no clear morphological changes were observed for miR-34b transfected cells (Fig.   2e). [score:1]
Interestingly, the miR-34b-5p’ seed sequence was identical to those annotated for miR-34a and miR-34c. [score:1]
Figure 1 Mature human miR-34 family sequence. [score:1]
Both the 5p and the 3p miRNA arms were detected for endogenous miR-34b and miR-34c, with the 5p arm as the major product, accounting for 76% and 99.7%, respectively (Supplementary data  S5). [score:1]
Importantly, we observed that the majority of transcripts with significantly changed levels were specific for each miR-34 isoform. [score:1]
To identify the common and unique effects of the bicistronic miR-34b and miR-34c we introduced miR-34b and miR-34c mimics into the breast cancer cell line MDA-MB-231. [score:1]
Figure 2Comparing effects of introducing miR-34b-5p and miR-34c-5p mimics into MDA-MB-231 cells. [score:1]
This cell line is derived from a highly aggressive metastatic breast cancer with low levels of endogenous miR-34. [score:1]
MiR-34b-5p’ function closely resembles that of miR-34c. [score:1]
The synthetic pre-miRNA precursors (Ambion, Grand Island, USA) hsa-miR-34b-5p (PM10743), hsa-miR-34c-5p (PM 11039), hsa-miR-34b-5p’ (AM17103) and Negative Control #2 oligos (AM 17111) were transiently transfected into the cells at a final concentration of 18 nM using the lipidic transfection agent INTERFERin (PolyPLUS, Illkirch, France), according to the manufacturer’s protocol. [score:1]
The analysis revealed that the levels of 777 and 1001 transcripts significantly changed upon introduction of miR-34b and miR-34c, respectively (Supplementary data  S1). [score:1]
These findings were in conflict with published data comparing miR-34 family members [8] and led us to sequence the endogenous miR-34b. [score:1]
In our study, we found that mimics representing the annotated versions of human miR-34b-5p (miR-34b) and miR-34c-5p (miR-34c) affected different gene sets, and importantly, resulted in different cellular effects. [score:1]
Sequence alignment of the mature miR-34a-5p, miR-34b-5p and miR-34c-5p molecules, as annotated by miRBase, the primary repository for miRNA sequences. [score:1]
Our experiments showed that caspase-3/7 activity was induced in miR-34c -transfected cells approximately 72 hours after transfection, while transfection with the miR-34b mimic did not induce caspase-3/7 activity (Fig.   2d). [score:1]
In addition, unlike cells transfected with miR-34b, cells transfected with miR-34b-5p’ changed morphology, closely resembling cells transfected with miR-34c (Fig.   4e). [score:1]
Interestingly, in contrast to the miR-34b mimic, which as predicted reduced the mRNA levels of THBS1, an increase of mRNA levels was observed following introduction of either miR-34b-5p’ or miR-34c mimic. [score:1]
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13
[+] score: 139
Number of embryos analyzed: miR-34c inhibitor: 5, miR-34c mimic: 5, miR-204 inhibitor: 6, miR-204 mimic: 7, control inhibitor: 9, control mimic: 8. Small RNA sequencing of embryonic porcine cortex across fetal development revealed a remarkable shift in expression levels of a distinct set of miRNAs from embryonic day 60–80 (E60–E80), which is the period of cortical folding in the pig brain (manuscript in preparation). [score:10]
Although we have shown that DCX is a target for both miR-34c and miR-204, and that over -expression of these miRNAs in IUE experiments significantly inhibit neuron migration in the mouse embryo, we cannot conclude that this migration effect is directly caused by inhibition of DCX. [score:10]
Number of embryos analyzed: miR-34c inhibitor: 5, miR-34c mimic: 5, miR-204 inhibitor: 6, miR-204 mimic: 7, control inhibitor: 9, control mimic: 8. Cortical folding is a complex process orchestrated by spatially controlled gene expression in subsections of the brain, and miRNA is likely to play key roles in the process. [score:9]
miR-34c and miR-204 mimics are shown to reduce DCX 3′UTR luciferase reporter expression, and miR-34c inhibitor increases DCX 3′UTR luciferase reporter expression. [score:7]
In accordance with being cognate targets, mimics of both miR-204 and mir-34c were able to significantly reduce luciferase expression, whereas miR-34c inhibitor increased the production from the DCX 3′UTR reporter. [score:7]
Both miR-34c and miR-204 are predicted by TargetScan to target DCX mRNA through 1 and 2 target sites, respectively (Table 1). [score:7]
The most strongly upregulated miRNAs, miR-34c, and miR-204, were shown to alter the extent of neuronal migration in embryonic mouse brain, and their large expression increase during cortical folding in pig, suggests a pivotal role in mediating correct cortical morphogenesis of gyrencephalic animals. [score:6]
MiR-204 and miR-34c are particularly notable upregulated miRNAs, due their almost exclusive expression in E80 cortex tissue (Figure 1B). [score:6]
Our finding, that increased miR-34c and miR-204 expression reduces neuronal migration, implies that suppression of EMT related factors could be a contributing mechanism controlling neuronal migration. [score:5]
Both the miR-34 family and miR-204 miRNAs are well known tumor suppressors in several cancers (He et al., 2007; Li et al., 2016) and have been linked to suppression of epithelial to mesenchymal transition (EMT; Hahn et al., 2013; Morizane et al., 2014; Li et al., 2016; Liu et al., 2016). [score:5]
To confirm that the predicted target sites are indeed functional targets of miR-34c and miR-204, the DCX 3′UTR was cloned downstream of a luciferase reporter and analyzed in HEK293-H cells. [score:5]
This suggests that controlled upregulation of miR-34 family miRNAs is important, both in embryonic and adult animals. [score:4]
The authors also find DCX to be a direct target of the miR-34-5p/449-5p family. [score:4]
Our finding of tightly controlled expression of miR-34c and miR-204 in brain development seems to extrapolate into neuroprotective effects in the aging animal. [score:4]
GFP signal indicates neurons transfected with miR-34c inhibitor (A) or mimic (B). [score:3]
Detection of miR-204 and miR-34c by in situ hybridization (ISH) at E60 and E80 in the porcine cortex showed clearly increased expression at E80 for both miRNAs (Figure 2A). [score:3]
Table 1 supports the shared function of miR-34a and miR-34c, given that both miRNAs show large expression increases from E60 to E80 of 50 and 180-fold, respectively. [score:3]
Analysis of migration of GFP -positive neurons in the motor cortex revealed that inhibition of miR-34c induced a robust increase in the percentage of GFP -positive cells in the cortical plate (CP; P = 0.006) and a reduction in both the intermediate zone (IZ; P = 0.007) and the subventricular zone (SVZ; P = 0.042) (Figures 3A,C). [score:3]
The striking finding that miR-204 and miR-34c exhibit specific expression increases in excess of 100 fold at the time of porcine gyration implies that they have key roles in the timing of the process. [score:3]
The authors found that miR-34 over -expression ameliorated age-related neurodegeneration and increased median lifespan. [score:3]
In a study by Liu et al. (2012), Drosophila miR-34 was found to display increased brain expression, specifically in old animals. [score:3]
Three psicheck2 vectors with inserted fragment were made from WT sequences originating from: DcxUp-3′UTR_WT: chrX:110543495-110543999       505 bp (The miR-34c target site). [score:3]
Three psicheck2 vectors with inserted fragment were made from WT sequences originating from: DcxUp-3′UTR_WT: chrX:110543495-110543999       505 bp (The miR-34c target site). [score:3]
Together, these data strongly suggest that miR-34c and miR-204 regulate neuronal migration in the embryonic cortex, in line with our expression and reporter assay experiments. [score:3]
MiR-204 and miR-34c sequences and their predicted targets in DCX 3′UTR are conserved, implying that the effects of miR-34c and miR-204 on the migration of neurons in the mouse is most likely conserved in higher mammals, such as pigs and humans. [score:3]
Mutating the predicted miR-34c, miR-204 and miR-15a target sites individually in the luciferase reporters abolished the observed effects (Figure 2C). [score:3]
Figure 3 miR-34c and miR-204 regulate cortical neuron migration. [score:2]
Altogether, every member of the miR-34-5p/449-5p family of miRNAs, all sharing identical seed sequences, have been found to affect brain development. [score:2]
miR-34c attenuates epithelial-mesenchymal transition and kidney fibrosis with ureteral obstruction. [score:1]
Of the 6 miRNAs in this miRNA seed family (miR-34a,b,c, and miR-449a,b,c) only miR-34a and miR-34c are annotated in pig, in agreement with the detected miRNAs in our profiling experiment. [score:1]
Thus, embryonic functions of miRNAs, such as miR-34c and miR-204, which affect cortical morphogenesis, are also likely to affect epilepsy susceptibility later in life. [score:1]
The microRNA miR-34 modulates ageing and neurodegeneration in Drosophila. [score:1]
Although miR-34a and miR-34c have identical seed sequences, they have a substantial 5 nt sequence dissimilarity outside the seed, ensuring that miR-34a and miR-34c can readily be distinguished in ISH experiments via specific LNA probes. [score:1]
Here we unveil a functional role for two of these miRNAs in neuronal migration, miR-34c and miR-204. [score:1]
Both miR-34c and miR-204 are conserved between mouse and pig and despite the fact that mice have a non-gyrated brain, the underlying process of neuronal migration is conserved between lissencephalic and gyrencephalic species (Kerjan and Gleeson, 2007). [score:1]
Mir-34a, a close relative of miR-34c, was recently found to play an active role in neural cell differentiation and was shown to affect migration of neuroblasts. [score:1]
The alkaline phosphatase-labeled probes used for miRNAs were 22 nts and 23 nts in length for miR-204 and miR-34c, respectively. [score:1]
miR-34c and miR-204 affect neuronal migration. [score:1]
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14
[+] score: 138
About half the mRNAs down-regulated by miR-34b or miR-34c were also down-regulated by miR-34a, but less than a fifth (91 of 482) of the genes down-regulated after miR-34a overexpression were down-regulated by miR-34b or miR-34c (Fig 2A), suggesting that individual miR-34 miRNAs regulate unique targets. [score:18]
Activation of p53 by cellular stress leads to transcription of miR-34 miRNAs, which in turn can enhance p53 function by: (1) miR-34a -mediated inhibition of multiple negative regulators of p53 to further increase p53 transcriptional activity; and (2) miR-34a -mediated increase of p53 protein stability (miR-34a feed-forward loops); or inhibit p53 function by: (3) direct miR-34a -mediated inhibition of TP53; and (4) direct miR-34 inhibition of many p53-activated genes (negative feedback loops). [score:12]
Although mature miR-34b and miR-34c have sequences almost identical to miR-34a even outside the seed, over -expression of miR-34b or miR-34c, unlike over -expression of miR-34a, had little effect on p53 promoter activity and only weakly up-regulated the mRNA levels of p53 transcriptional targets. [score:10]
Consistent with this result, induction of 6 p53 transcriptional targets in HCT116 cells was significantly less after miR-34b or miR-34c overexpression than after miR-34a overexpression (Fig 1D), despite highly elevated miRNA overexpression (S1A Fig). [score:9]
Functional Annotation Analysis of downregulated genes in HCT116 cells overexpressing miR-34 using DAVID Bioinformatics tool. [score:6]
Genes down-regulated by miR-34 over -expression in HCT116 cells. [score:6]
482, 163 and 29 mRNAs were significantly down-regulated (fold decrease ≥ 1.5 fold relative to miRNA control) after miR-34a, miR-34b or miR-34c overexpression, respectively (Fig 2A and S1 Table). [score:6]
miR-34b-5p (hereafter designated miR-34b) overexpression had a modest, but significant, effect on 2 of the 4 promoters, while miR-34c did not significantly increase activity of any (Fig 1C), even though it was over-expressed more than a hundred fold above its endogenous level after genotoxic stress (data not shown). [score:5]
As expected, miR-34 overexpression decreased the miR-34a target gene CDK6 and increased the p53-activated gene CDKN1A, assessed as controls. [score:5]
We set out to study how the different miR-34 miRNAs contribute to p53 function, analyze whether they regulate overlapping sets of targets and determine if miR-34 is essential for p53 -mediated function in human cells. [score:4]
To determine whether the miR-34 family might regulate non-overlapping mRNAs, we performed gene microarray analysis of HCT116 cells overexpressing each family member (S1B Fig). [score:4]
In addition, the lack of a strong effect of genetic deletion of miR-34a could also be secondary to functional redundancy provided by the other miR-34 members or other p53-regulated tumor suppressor miRNAs [45– 49] or by the p53-independent miR-449 family, which shares a seed sequence with miR-34 [50]. [score:4]
These data are consistent with a previous report showing differing proteomics profiles in HeLa cells over -expressing miR-34a or miR-34c [41]. [score:3]
A better knowledge of the mutual functional dependence between miR-34 and p53 will help to understand miR-34 tumor suppressor function. [score:3]
We next used luciferase reporter promoter assays, in p53-sufficient HCT116 cells, to assess whether miR-34 overexpression enhanced promoter activities of a sequence of 13 tandem repeats of the p53 binding site (pG13-luc) [16] or the promoters of p53-regulated genes, PUMA, CDKN1A (the gene encoding p21/WAF1) and BAX. [score:3]
Our observation that only miR-34a overexpression enhances p53 -mediated transcription was surprising since the miR-34 family active strands are highly homologous—the seed (residues 2–9) and residues 11–17 and 19–21 are identical (Fig 1C). [score:3]
Analysis of miR-34 levels in miR-34 over -expressing samples. [score:3]
Of note, for both experiments, miR-34c expression is ~ 9 fold less than in miR-34b transfected samples. [score:3]
0132767.g002 Fig 2 (A) Overlap of genes down-regulated ≥ 1.5 fold in miR-34 OE HCT116 cells compared to control -transfected cells. [score:3]
An unexpected finding of this study was the weak effect of miR-34b or miR-34c over -expression on p53 function. [score:3]
Single colonies were tested for miR-34 expression by qRT-PCR and negative colonies were verified by sequencing. [score:3]
Since their initial identification as p53 transcriptional targets, the three members of the miR-34 family have been considered crucial mediators of the p53 response [39]. [score:2]
However, miR-34c is still highly expressed, even compared to DOX -treated HCT116-WT cells. [score:2]
Multiple miRNAs, including the miR-34 family, are transcriptionally activated by p53. [score:1]
miR-34c is increased in miR-34c transfected samples by 100X and 285X, relative to DOX treated HCT116-WT cells, respectively (compare S1A and S1B Fig to Fig 6C). [score:1]
Our results showing that miR-34a is not essential for the p53 mediated response to stress are in agreement with data published by Concepcion et al reporting intact p53 function in miR-34 deficient mice [12]. [score:1]
Genome-wide transcriptome analysis of miR-34 OE HCT116 cells. [score:1]
Thus sequence determinants outside the seed might profoundly affect miR-34 family function by an unknown mechanism that is worth exploring. [score:1]
Future experiments with miR-34 -deficient human cells should address the contribution of miR-34 in these other scenarios. [score:1]
Mean +/- SD of three independent experiments is shown in cells transfected with miR-34 family or cel-miR-67 (M-control) mimics. [score:1]
WT cells have <1 copy/cell of miR-34b and miR-34c, which only increases to 5 and 10 copies/cell, respectively, after DOX (Fig 6C). [score:1]
Thus miR-34 -mediated increased p53 transcription is largely limited to miR-34a. [score:1]
miR-34—a microRNA replacement therapy is headed to the clinic. [score:1]
Normalized Firefly luciferase activity, relative to Renilla luciferase activity, after miR-34 transfection is plotted as fold change relative to control miRNA -transfected sample. [score:1]
Alignment of the miR-34 family with the seed sequence highlighted in red is shown at top. [score:1]
miR-34 levels in transfected samples from Fig 1D (A) and Fig 2 (B), analyzed by qRT-PCR. [score:1]
In this regard, it has been shown recently that somatic cells from miR-34 deficient mice can be reprogrammed more efficiently [51]. [score:1]
Three chromosome 5q11.2 miRNAs (miR-449a/b/c) share a seed sequence with miR-34, and have a tissue distribution similar to that of miR-34b/c [6, 7]. [score:1]
Here, we investigated in detail how the different miR-34 family members contribute to p53 function, the miR-34a targets that are relevant for its contribution and how much p53 relies on miR-34a. [score:1]
Although antagonizing miR-34a in human cells impairs p53 function in a few studies [4, 10, 11], mice genetically deficient in all miR-34 family genes have unimpaired stress responses [12]. [score:1]
The miR-34 family consists of 3 miRNAs—miR-34a on human chromosome 1p36 and miR-34b/c, co-transcribed on human chromosome 11q23. [score:1]
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[+] score: 136
Other miRNAs from this paper: hsa-mir-34a, hsa-mir-34b
0067581.g002 Figure 2(a) qRT-PCR showing PDGFR-α and PDGFR-β mRNAs downregulation in Calu-6 and H1703 cells after miR-34 and miR-34c but not miR-34b enforced expression (b) miR-34a and miR-34c enforced expression decreases endogenous levels of PDGFR-α/β protein levels in H1703 and Calu-6 NSCLC. [score:8]
In 2007, reports from several laboratories showed that members of the miR-34 family are direct p53 targets, and that their upregulation induced apoptosis and cell-cycle arrest [16, 17]. [score:7]
PDGFR-α/β downregulation by miR-34a and miR-34c inhibits migration and invasiveness of NSCLC cells. [score:6]
Since PDGFR-α and PDGFR-β regulate the PI3K/Akt and ERK1/2 pathways [23, 24, 25], we next examined, by immunostaining, the expression and/or activation of some of the proteins involved in these pathways following miR-34a and miR-34c enforced expression or PDGFR-α/β silencing by siRNAs. [score:6]
Indeed, overexpression of miR-34a and miR-34c or downregulation of PDGFR-α/β by siRNAs, highly increased the response of semi-resistant NSCLC cells to TRAIL -induced apoptosis. [score:6]
In this study, we show that miR-34a and miR-34c, are strongly downregulated in NSCLC cells and lung tumors whereas they are highly expressed in normal lung tissues. [score:6]
Enforced expression of miR-34a and miR-34c downregulated PDGFR-α and PDGFR-β mRNA and protein levels. [score:6]
Among the miRNAs, miR-34 family members play important tumor suppressive roles, as they are directly regulated by p53 and compose the p53 network [16, 17]. [score:5]
MiR-34a or miR-34c and not miR-34b forced expression decreases PDGFRβ expression levels and reduces the activation of the ERK1/2. [score:5]
Intriguingly, we observed a significant decrease of the migratory and invasive capabilities of Calu-6 and H1703 cells after miR-34a or miR-34c overexpression (Figure 6a) as well after PDGFR-α and PDGFR-β downregulation (Figure 6b), confirmed also by scratch-wound assay (Figure 6c). [score:5]
MiR-34a and miR-34c are inversely related to PDGFR-α/β expression in vitro and in vivo Next, we analyzed the consequences of the ectopic expression of miR-34a and -34c in Calu-6 and H1703 cells. [score:5]
Taken together the results indicate that miR-34a and miR-34c and not miR-34b directly target PDGFR-α and PDGFR-β. [score:4]
Luciferase and western blot experiments demonstrated that PDGFR-α and PDGFR-β are direct targets of miR-34a and miR-34c but not of miR-34b. [score:4]
Interestingly, increased expression of miR-34a and miR-34c, and not miR-34b, upon transfection, confirmed by qRT-PCR (data not shown), significantly decreased luciferase activity, indicating a direct interaction between the miRNAs and PDGFRα and PDGFRβ 3’ UTRs (Figure 1c,d). [score:4]
Here, we reported that miR-34a and miR-34c overexpression or PDGFR-α/β silencing inhibited the migration and invasion capacity of Calu-6 and H1703 cells, compared to cells transfected with a scrambled miR or siRNA control. [score:4]
MiR-34a and miR-34c overcome TRAIL resistance of NSCLC cells through PDGFR-α and PDGFR-β downregulation. [score:4]
Here, we report that not only miR-34a but miR-34c also downregulates PDGFR-α in NSCLC cells. [score:4]
MiR-34a and miR-34c overexpression or PDGFR-α/β silencing decreases migratory and invasive capacity of NSCLC cells. [score:3]
The expression levels of the four PDGF ligands (PDGFA, PDGFB, PDGFC, PDGFD) after miR-34a and miR-34c enforced expression were also evaluated in both Calu-6 and H1703 cells. [score:3]
Moreover, the promoter region of miR-34a, miR-34b and miR-34c contains CpG islands and aberrant CpG methylation reduces miR-34 family expression in multiple types of cancer [18, 19, 20]. [score:3]
MiR-34a and miR-34c are inversely related to PDGFR-α/β expression in vitro and in vivo. [score:3]
Figure S3, Enforced expression of miR-34a and miR-34c or PDGFR-α /β silencing increases the response to TRAIL -induced apoptosis and reduces tumorigenicity of NSCLC cell. [score:3]
MiR-34a and miR-34c are downregulated in the tumors compared to the normal lung tissues. [score:3]
Increased expression of miR-34a and miR-34c upon transfection was confirmed by qRT-PCR (data not shown) and then the effects on PDGFR-α and PDGFR-β mRNA and protein levels were analyzed by qRT-PCR and western blot. [score:3]
We previously demonstrated that the PI3K/AKT pathway plays a key role in TRAIL -induced apoptosis [26], therefore the effects of miR-34a and miR-34c overexpression on cell survival and TRAIL resistance of NSCLC were examined. [score:3]
MiR-34a and miR-34c target PDGFR-α and PDGFR-β 3’ UTRs. [score:3]
MiR-34a and miR-34c overexpression or PDGFR-α/β silencing increases the response of NSCLC cells to TRAIL -induced apoptosis. [score:3]
Moreover, miR-34a and miR-34c, by targeting PDGFR-α and PDGFR-β, increase TRAIL -induced apoptosis and decrease invasiveness of lung cancer cells. [score:3]
As shown in Figure 3a and b, miR-34a and miR-34c expression was lower in the tumor compared to the normal samples (Figure 3a,b). [score:2]
First, we performed a proliferation assay on Calu-6 and H1703 TRAIL semi-resistant cells after enforced expression of miR-34a and miR-34c. [score:2]
Intriguingly, overexpression of PDGFR-α or PDGFR-β (using two plasmids containing only the coding sequences and not the 3’ UTRs of PDGFR-α/β) along with miR-34a or miR-34c, decreased the sensitivity to TRAIL -induced apoptosis, as assessed by both MTT and caspase 3/7 assay (Figure S4 in File S1). [score:2]
As shown in Figure 5a, phosphorylation levels of ERKs decreased after miR-34a and miR-34c enforced expression compared to cells transfected with the control miR. [score:2]
In mammalians, the miR-34 family comprises three processed miRNAs that are encoded by two different genes: miR-34a is encoded by its own transcript, whereas miR-34b and miR-34c share a common primary transcript. [score:1]
A previous study indicated that miR-34 methylation was present in NSCLC and was significantly related to an unfavorable clinical outcome [20]. [score:1]
Briefly, 2x 10 [5] Calu-6 cells transfected with pcDNA-PDGFR-α, pcDNA-PDGFR-β and/or miR-34a and miR-34c in RPMI supplemented with 1% FBS were plated into upper chambers of with a 8-um pore size-polycarbonate membrane. [score:1]
Cells were transfected with either scrambled, miR-34a or miR-34c for 72h. [score:1]
Briefly, Calu-6 cells were transfected with pcDNA-PDGFR-α, pcDNA-PDGFR-β and/or hsa-miR-34a and miR-34c, respectively. [score:1]
MiR-34a and miR-34c reduce PDGFR-α and PDGFR-β mRNA and protein levels. [score:1]
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[+] score: 134
data are expressed as relative quantification using miR-30b as normalizer, normalized on average of control expression Fig. 3miR-31, miR-708, and miR-34c targeted 3′UTR sequences of NOS1 gene. [score:7]
RQ are obtained using average of values of all miRNA for normalizer miR-708-5p, miR-31-5p, and miR-34c-5p target 3′UTR sequences of NOS1 geneTo select miRNAs that could modulate nNOS expression, the total sequence of the 3′UTR of the NOS1 gene (NOS1-3′UTR) was submitted to two predictive software, i. e., TargetScan Human and microRNA. [score:7]
In DMDd45-52 cells, we demonstrated that the inhibition of miR-708 and miR-34c increased nNOS expression, confirming that both miRNAs can modulate nNOS expression in human myoblasts. [score:7]
The reduction in the nNOS level was confirmed by Western blot experiments showing a decrease of about 30% of nNOS expression in cells overexpressing miR-708 or miR-34c, while no significant decrease could be observed in overexpressing miR-31 (Fig.   5c). [score:7]
Fig. 5miR-708 and miR-34c overexpression inhibit nNOS expression in transfected control human myoblasts. [score:7]
These results strongly suggest that miR-708 and miR-34c, overexpressed in dystrophic context, are new actors involved in the regulation of nNOS expression in dystrophic muscle. [score:6]
We show here that inhibitors of miR-708 and/or miR-34c could also be considered as therapeutic targets to rescue these defects by increasing the expression of nNOS. [score:6]
Fig. 6Inhibition of miR-708 and miR-34c increased nNOS expression in transfected DMDd45-52 human myoblasts. [score:5]
The complexity of the mechanisms modulating NOS1 transcription indicates that the nNOS isoform expressed in myoblasts and regulated by miR-34c and miR-708 has not been precisely identified and that information on the transcriptional regulation of its gene remains to be thorough. [score:5]
Nevertheless, no decrease of the reporter gene was observed when miR-212 was co -transfected with the parts #1, #2, #3, nor #4. These results demonstrated that miR31, miR-708, and miR34c, but not miR-212, were able to target NOS1-3′UTR sequences leading to a decrease of the reporter gene Firefly luciferase expression. [score:5]
We thus demonstrated that miR-708 and miR-34c could decrease nNOS expression in human healthy myoblasts and that their inhibition led to an increase of this protein in DMDd45-52 human cells. [score:5]
First, we showed a decrease of nNOS expression when miR-708 or miR-34c were overexpressed in control myoblasts. [score:5]
Fig. 4miR-31, miR-708, miR-34c, and nNOS expression in DMDd45-52 myoblasts. [score:3]
miR-708-5p, miR-31-5p, and miR-34c-5p target 3′UTR sequences of NOS1 gene. [score:3]
Our data were in the same way as miR-34c is overexpressed in BMDd45-55 muscle biopsies, in DMDd45-52 myoblasts, and in mdx mice (data not shown). [score:3]
Altogether, these results demonstrated that miR-708 or miR-34c could modulate nNOS expression in human healthy myoblasts. [score:3]
Concerning miR-708 and miR-34c, our results showed an effect of these two miRNAs on nNOS expression in human healthy and DMDd45-52 myoblasts. [score:3]
Thus, a therapeutic strategy combining the inhibition of miR-708 and miR-34c with the restoration of dystrophin will most likely be a benefit for the improvement of phenotype of DMD and BMD patients. [score:3]
Analysis of the pictures showed a decrease of the nNOS labelling in the nuclei of cells overexpressing miR-708 or miR-34c. [score:3]
In the present study, we could not exclude a link between nuclear nNOS location, HDAC2 nitrosylation, and the modulation of the miR-31, miR-708, and/or miR-34c expression. [score:3]
Furthermore, by analyzing the sequence of the NOS1-3′UTR regarding the 4 selected miRNAs, we identified 5 sequences as potential targets of miR-31, 5 for miR-708, 9 for miR-34c, and 3 for miR-212 (Additional file  3: Table S1 and Fig.   3a). [score:3]
A luciferase reporter study validated the targeting of NOS1-3′UTR by miR-31, miR-708, and miR-34c. [score:3]
miR-31, miR-708, and miR-34c effect on nNOS expression in human myoblasts. [score:3]
The inhibition of miR-708 or the miR-34c levels by their antimiRNAs was validated by RT-qPCR experiments (Fig.   6a). [score:3]
Overall, these results were consistent with those obtained on BMDd45-55 muscle biopsies, namely a higher level of miR-31, miR-708, and miR-34c and a decrease in the expression of nNOS, thus allowing the use of these DMDd45-52 myoblasts as a suitable in vitro cellular mo del. [score:3]
Graph represents average of relative quantification of miRNA normalized on SNORD44 expression of 5 (miR-31) or 3 (miR-708 and miR-34c) independent experiments. [score:3]
For miR-34c, several studies described it as overexpressed in mdx mice and in DMD patients [23, 35]. [score:3]
The present study revealed that the nNOS expression could be modulated by miR-708 and miR-34c. [score:3]
Quantification by RT-qPCR confirmed a higher level of expression for miR-31, miR-708, and miR-34c in DMDd45-52 cells compared to control with a fold change of 2.2, 2.2, and 3.8, respectively (Fig.   4a). [score:2]
A higher level of expression of the 4 miRNAs was detected in BMDd45-55 compared to control muscles with a fold change of 6.6, 4.4, 10.1, and 3.3 for miR-31, miR-708, miR-34c, and miR-212, respectively, confirming the results obtained by TLDA (Fig.   2b, Additional file  2). [score:2]
Immunofluorescence experiments showed an increased staining in the nuclei of cells in which the miR-708 or the miR-34c were inhibited compared to cells transfected with a non-specific control miRNA (Fig.   6b). [score:2]
Among them, only the overexpression of miR-31, miR-708, and miR-34c led to a decrease of luciferase activity in an NOS1-3′UTR-luciferase assay, confirming their interaction with the NOS1-3′UTR. [score:2]
Our data showed a significant decrease of luciferase activity when the part #2 was co -transfected with the miR-31 and the part #3 with the miR-708 and when the parts #1, or #3, or #4 were co -transfected with the miR-34c. [score:1]
miR-708 n = 7, miR-31 n = 7, and miR-34c n = 8. b nNOS immunoblot in control and DMDd45-52 cells. [score:1]
Cells were transfected with 12.5 pg of either miR -negative control (AM17111, Ambion); miR-31, miR-34c, or miR-708 (AM17100, Ambion); or antimiR-34c or antimiR-708 (AM17000, Ambion) using lipofectamine 2000 diluted in Optimem reduced medium. [score:1]
We then confirmed that DMDd45-52 cells displayed an endogenous increased of miR-31, miR-708, and miR-34c and a decreased of nNOS expression, the same characteristics observed in BMDd45-55 biopsies. [score:1]
Each 3′UTR construction (24.5 ng) was co -transfected in 293T-HEK cells with 25 pg of either miR -negative control (AM17111, Ambion) or miR-212, miR-31, miR-34c, or miR-708 (AM17100, Ambion) using lipofectamine 2000 diluted in Optimem reduced medium. [score:1]
We selected 4 miRNAs (i. e., miR-31, miR-708, miR-34c, and miR-212) since they were overexpressed in muscular biopsies of BMDd45-55 patients compared to healthy subjects or in muscular biopsies of patients with severe phenotypes compared to other patients. [score:1]
[1 to 20 of 38 sentences]
17
[+] score: 124
Other miRNAs from this paper: hsa-mir-34a, hsa-mir-34b, hsa-mir-449a, hsa-mir-449b, hsa-mir-449c
By looking more precisely at the three major families of regulators of small GTPases expressed during HAECs differentiation or after miR-34/449 transfection, we detected 23 distinct ARHGAPs, including ARHGAP1; we also detected 22 distinct ARHGEF transcripts but none of them were predicted as direct miR-34/449 targets; regarding the three known mammalian ARHGDI transcripts 40, only ARHGDIB was further analysed, as ARHGDIA expression levels did not change during HAECs differentiation or after miR-34/449 transfection and ARHGDIG was not detected (Supplementary Fig. 3b, see also GEO GSE22147). [score:9]
Considering that R-Ras activity can be increased by Notch pathway activation, as previously observed in another cellular mo del 39, and considering the early repression of Notch signalling by miR-34/449 during vertebrate multiciliogenesis 9, miR-34/449 is therefore able to control R-Ras function at two distinct levels: (1) directly, via the inhibition of RRAS expression, and (2) indirectly, via the inhibition of R-Ras activity through the repression of the Notch pathway. [score:9]
Previously, we knocked down miR-449 in Xenopus MCCs by injecting a cocktail of morpholino antisense oligonucleotides against miR-449a/b/c (449-MOs) into the prospective epidermis at the eight-cell stage 9. In Xenopus, miR-34b was detected in MCCs (Supplementary Fig. 2d–f), and in situ hybridization (ISH) experiments also revealed that 449-MOs not only blocked the expression of miR-449 but also blocked the expression of miR-34b (Supplementary Fig. 2f), suggesting that 449-MOs collectively inhibit miR-34/449 miRNAs (Supplementary Fig. 2e,f). [score:8]
Specifically, we looked for relevant miR-34/449 mRNA targets that were repressed during MCC differentiation and after overexpression of miR-34/449 in proliferating HAECs (Gene Expression Omnibus (GEO) data set GSE22147). [score:7]
RRAS2 expression remained at a very low level during MCC differentiation and in response to miR-449 overexpression, and was not altered by miR-34/449 knockdown or PO- RRAS protection (HAECs, see GEO GSE22147; Xenopus, Supplementary Fig. 4c). [score:6]
MiR-34/449 may initially downregulate the expression of several cell cycle-regulated genes and members of the Notch pathway to promote entry into differentiation. [score:6]
In a bid to identify such additional factors, we applied several miRNA target prediction tools 58 to identify putative miR-34/449 targets among the small GTPase pathways. [score:5]
Next, we focused on the three most significantly regulated miR-34/449 targets ARHGAP1, ARHGDIB and RRAS. [score:4]
We thus looked for other possible targets of miR-34/449 that would be directly related to actin dynamics. [score:4]
Of note, in Xenopus, the percentage of cells exhibiting defective apical actin meshwork was higher in miR-34/449 morphants (Fig. 2b) than in PO- Dll1 morphants (Fig. 2d), suggesting that miR-34/449 may affect additional targets. [score:3]
In control embryos and in miR-34/449 morphants, almost all rGBD-GFP -positive cells expressed the early MCC differentiation marker α-tubulin (Fig. 3c). [score:3]
Thus, following the experimental inhibition of miR-34/449 in Xenopus, RhoA activation can still be detected in MCCs unable to grow cilia. [score:3]
These results establish ARHGAP1, ARHGDIB and RRAS transcripts as bona fide targets of miR-34/449. [score:3]
In conclusion, our data further document how the miR-34/449 family can participate to multiciliogenesis through the repression of several important targets. [score:3]
The huge induction of miR-449 expression at early ciliogenesis (stage EC) suggests that early effects were mainly mediated through miR-449, without excluding a later role for miR-34 (Supplementary Fig. 2a). [score:3]
Incidentally, the existence of five miR-34/449 -binding sites in the 3′-UTR of ARHGAP1 made elusive the assessment of a target protection of ARHGAP1 against a miR-34/449 action in primary HAEC cultures. [score:3]
This could also explain the lack of compensation of miR-34 in our antago-449 conditions, which do not alter the expression of miR-34. [score:3]
miR-34/449 targets components of the small GTPase pathways. [score:3]
These data posit the absence of R-Ras in miR-34/449 -expressing MCCs as a conserved feature across tetrapods. [score:3]
Collectively, these results suggest that, although the modulation of RhoA activity by miR-34/449 may play a role in the control of apical actin polymerization, other miR-34/449 targets contribute to the profound disruption of the actin cap observed after miR-34/449 inactivation. [score:3]
As miR-34 and miR-449 miRNAs share the same targets, we only used miR-449 in the rest of this study. [score:3]
We used target protection assays (cholesterol-conjugated modified oligonucleotides in HAECs or morpholino oligonucleotides in frog epidermis) to compete with the binding of miR-34/449 on sites identified within the human and Xenopus 3′-UTRs of RRAS mRNA. [score:2]
At the dose used for this assay, injection of MO-ATG- rras alone had no significant effect on either apical actin meshwork formation or multiciliogenesis (Fig. 7a–d), suggesting that rras expression may be already repressed by the presence of endogenous miR-34/449 in those MO-ATG- rras maturing MCCs. [score:2]
To invalidate miR-449 or miR-34b/c activity in human, we transfected HAECs with a cholesterol-conjugated antagomiR directed against miR-449 (Antago-449) or against miR-34 (Antago-34) and assessed MCC differentiation. [score:2]
In addition, miR-34/449 knockdown or protection of RRAS mRNA from miR-34/449 in frog epidermis led to an increase in rras transcript levels (Supplementary Fig. 4c). [score:2]
Collectively, these assays unambiguously establish that RRAS transcripts were specifically targeted by miR-34/449 in human and Xenopus MCCs. [score:2]
We designed target protection assays in which cholesterol-conjugated modified oligonucleotides were transfected in differentiating HAECs to compete with the binding of miR-34/449 on the site identified within the human 3′-UTR of ARHGDIB mRNA (PO- ARHGDIB). [score:2]
Thus, our data show that the miR-34/449 family clearly contributes to actin cytoskeleton remo delling in several independent mo dels. [score:1]
MiR-449 -mediated silencing of either ARHGAP1, ARHGDIB or RRAS was respectively abolished when miR-34/449-predicted binding sites were mutated (Fig. 4c). [score:1]
In human (Fig. 6a,b) and in Xenopus (Fig. 7a,d), protection of the RRAS transcript from miR-34/449 binding led to a strong reduction in apical actin meshwork and motile cilia formation. [score:1]
We also noticed a reduction of miR-449 levels when preventing miR-34/449 binding on Notch1 in differentiating HAECs (Fig. 1f) and on Dll1 in frog epidermis (Fig. 2f). [score:1]
We examined whether the miR-34/449 family can control the formation of the apical actin network, a prerequisite for basal body anchoring and cilium elongation. [score:1]
miR-34/449 control apical actin network assembly in MCCs. [score:1]
How to cite this article: Chevalier, B. et al. miR-34/449 control apical actin network formation during multiciliogenesis through small GTPase pathways. [score:1]
Our data unambiguously indicate that the repression of RRAS at a late step of MCC differentiation by miR-34/449 is required for apical actin network assembly and multiciliogenesis in human (Fig. 6) as well as in frog (Fig. 7). [score:1]
We observed that preventing the binding of miR-34/449 on Notch1 (PO- Notch1 in HAECs, Fig. 1a,b) or on the Notch ligand Dll1 (PO- Dll1 in Xenopus, Fig. 2c–f) with protector oligonucleotides coordinately blocked multiciliogenesis and apical actin network formation. [score:1]
These results establish that the miR-34/449 family interferes with MCC apical actin meshwork formation in both mo dels. [score:1]
Next, we addressed the precise mode of action of miR-34/449 in the construction of the apical actin network in MCCs. [score:1]
This is consistent with the need for an early repression of the Notch pathway by miR-34/449, to allow MCC differentiation. [score:1]
These observations point to the participation of miR-34/449 and R-Ras to actin network reorganization and their capacity to alter the RhoA activity. [score:1]
As the miR-34/449 family represses the Notch pathway during MCC differentiation 9, we assessed the contribution of the Notch signal to the actin web reorganization. [score:1]
miR-34/449 control apical actin assembly by repressing R-Ras. [score:1]
However, we cannot rule out discrete changes in the sub-cellular localization of activated RhoA in miR-34/449 -deficient embryos. [score:1]
Its repression by miR-34/449 appears to affect centriole maturation, but not apical actin network assembly 48. [score:1]
In a tentative mo del (Fig. 8d), we propose that the silencing of R-Ras by miR-34/449 in MCCs affects the interaction between R-Ras and FLNA, and favours a redistribution of FLNA in a cytoskeletal components involved into the anchoring of basal bodies. [score:1]
[1 to 20 of 45 sentences]
18
[+] score: 121
Other miRNAs from this paper: hsa-mir-34a, mmu-mir-34c, mmu-mir-34b, mmu-mir-34a, hsa-mir-34b
The relative intensity was normalized to the expression of the control sample Fig. 6 a Western blot analysis after Ant34 treatment (50 nM for 7 days as described above), b miR34 transduction and c p63 expression after Numb overexpression. [score:7]
In an opposite way as the inhibition, miR34 overexpression induces a downmodulation of cKit, Notch, and hey-1 expression (Fig.   5b). [score:7]
The mRNA expression on different LNA34 -treated CDCs showed that miR34 inhibition causes an increase of cKit, Notch-1 and hey-1 expression (Fig.   5a). [score:7]
Protein expression analysis after mir34 inhibition, overexpression or Numb transduction. [score:7]
The relative intensity was normalized to the expression of the control sample a Western blot analysis after Ant34 treatment (50 nM for 7 days as described above), b miR34 transduction and c p63 expression after Numb overexpression. [score:7]
It has been demonstrated in mouse mo del that miR34 inhibition reduces cardiac dysfunction 6, 7. Boon and coworkers [7] found that miR34 is implicated in cardiac aging and its downmodulation through LNA inhibition supports cardiac repair in mice after AMI. [score:5]
We observed an increased Numb expression by miR34 inhibition (Fig.   6a). [score:5]
It is worthy to note that LNA34 treatment enhances not only Notch mRNA, but also hey-1 expression, indicating that the Notch pathway is activated after miR34 inhibition. [score:5]
Gene expression analysis after mir34 inhibition or transduction. [score:5]
MiR34 expression directly correlates with age in human biopsies (r = 0.125037328, p = 0.010672365) On the basis of this evidence, we silenced miR34 using an LNA 8mer (Ant34). [score:4]
This indicates an indirect correlation between miR34 expression and proliferation in these cell populations. [score:4]
Our data demonstrate not only that Numb is regulated by miR34 in cardiac stem cells, but also that Numb overexpression itself induces an increase in cardiac progenitors growth (revised by Wu and Li [19] in other mo dels as mouse and drosophila). [score:4]
To confirm the regulative activity of miR34, we overexpressed in CDCs mature miR34 with lentiviral vector. [score:4]
It is possible to assess that in the dividing population miR34 is less expressed compared with the quiescent population (result of three independent experiments on five different CDCs populations p ≤ 0.05) We tested in our specimens the relation between age and miR34 relative expression [7]. [score:4]
Our study represents the first evidence that miR34 inhibition in human cardiac progenitor/stem cells could be proficiently employed in new therapeutic interventions for human cardiac pathologies. [score:3]
It is possible to observe a clear incorporation of Numb after overexpression, Rab5 was used as exosomal protein normalizer A recent report has demonstrated that the miR34 downmodulation after MI induces a functional recovery and an increase in the scar reduction in mice [7]. [score:3]
The two separated populations were examined for the ability to form spheres in culture (Fig.   1b) and for miR34 relative expression (Fig.   1c). [score:3]
Growth rate after mir34 inhibition. [score:3]
Moreover, by overexpressing miR34 we observed that Numb was also specifically downmodulated (Fig.   6b). [score:3]
Boon et al. [7] have shown that miR34 is involved in cardiac aging and its downmodulation through LNA inhibition supports cardiac repair in mice after AMI (acute myocardial infarction). [score:3]
The indication of this downmodulation by miR34 prompted us to test if the role of miR34 can be due to the Numb overexpression after Ant34 treatment. [score:3]
The ability of miR34 to downmodulate c-Kit has been demonstrated in colon cancer cells [16], where it has been linked to p53 expression. [score:3]
It is possible to assess that in the dividing population miR34 is less expressed compared with the quiescent population (result of three independent experiments on five different CDCs populations p ≤ 0.05) Real-time PCR of total RNA from 32 biopsies. [score:2]
Mir34 expression in cardiac human specimens vs age. [score:2]
MiR34 has been reported to target different genes in various cellular system that can account its function. [score:2]
In these cells, miR34 modulates, in vitro, various genes that are clearly involved in cardiac development and/or repair. [score:2]
As already found in cancer stem cells [17], miR34 plays an apparent bimodal role, regulating as Notch as Numb pairwise. [score:2]
MiR34 expression in Cardiac stem cells subpopulation. [score:2]
We observed that the less proliferating (CSFE++) cells had a higher expression of miR34 compared with the more proliferating cells (CSFE--). [score:2]
Our results show that miR34 has a complex role in human cardiac progenitor cells, where its downmodulation induces a cascade essential for cardiac repair, as assessed in mouse mo dels [7]. [score:1]
First, we tried to evaluate the miR34 expression in proliferating CSs cells. [score:1]
MiR34 expression directly correlates with age in human biopsies (r = 0.125037328, p = 0.010672365) a FACS analysis to evaluate Ant34a 5' FITC-LNA’s ability to enter into human CDCs/CSs by gymnosis after treatment (50 nM for 24 h). [score:1]
One of the most promising miRNAs in this regard is miR34 (reviewed by Li et al. [4]), which acts as a controller in reprogramming efficiency, while miR34 ablation shows a higher susceptibility to induced progenitor stem cells (iPSC) generation without compromising self-renewal and differentiation [5]. [score:1]
We observed, after miR34 downmodulation, an increase of Notch and its downstream-activated gene hey-1. In cardiac progenitor cells Notch-1 activation, with the nuclear translocation of Notch-1 intracellular domain (N1ICD), stimulates proliferative signaling such as G1/S cyclins and p38 (revised by Li and coworkers [24]) in vitro, induces myocytes differentiation [25], MAPK activity and promotes immature cardiomyocytes expansion. [score:1]
In particular, our results indicate that miR34 downmodulation plays a role in human cardiac progenitor proliferation. [score:1]
The authors deduced that the repair activity could be also due to the increased vascularization in the infarcted zone in in vivo mouse mo del, where LNA34 antisense (Ant34) treatment induces angiogenesis and promotes proliferation in endothelial progenitor cells and Human Umbilical Vein Endothelial Cells (HUVECs) 7, 8. The aim of this work was to establish whether the cardiac repair activity of miR34 inhibition could be used for human heart treatment, evaluating in vitro the influences of Ant34 in human heart cardiac stem cells. [score:1]
Our study indicates, for the first time, to the best of our knowledge, that the role of miR34 downmodulation in cardiac repair can also be held by Numb, which has been found to be important in cardiac morphogenesis [19]. [score:1]
[1 to 20 of 37 sentences]
19
[+] score: 119
Down-regulation of miR-34c and up-regulation of miR-183 and miR-210 were identified in cancer groups. [score:7]
Despite a limited statistical data, high expression group of miR-34c also showed poor overall survival than low expression one, especially in EGFR exon 19 mutation lung cancer patients. [score:6]
The miR-34 family is composed of three miRNAs (miR-34a, miR-34b, and miR-34c) that are direct transcriptional targets of p53, so their expression is induced by p53 in response to DNA damage and oncogenic stress [18, 19]. [score:6]
In EGFR exon 19 mutation group, miR-34c high expression group showed poor overall survival than low expression one (p = 0.035, Fig.   3). [score:6]
Compared to normal control lung tissue, down-regulation of miR-34c and up-regulation of miR-183 and miR-210 were identified in caner groups (p < 0.05 for each). [score:6]
In EGFR exon 19 mutation group, miR-34c high expression group showed poor overall survival than low expression one by univariate Kaplan-Meier method. [score:6]
But miR-34c expression levels were similar between two groups, and miR-210 showed not significant but higher expression tendency in EGFR mutation groups than wild type. [score:6]
We found that miR34c showed significant down expression in lung adenocarcinoma tissue than normal lung, indicating tumor suppressor role of miR-34c. [score:5]
Fig. 3Comparison of overall survivals according to miR-34c expression level in lung adenocarcinomas showing EGFR exon 19 mutation Despite the development of early detection and surgical techniques, lung cancer is still the leading cause of cancer-related death. [score:5]
miR-34c expression levels were similar between EGFR mutation group and wild type group. [score:4]
In this context, it can be a reasonable excuse that high expression group of miR-34c shows poor overall survival in EGFR exon 19 mutation patients. [score:4]
Fig. 3Comparison of overall survivals according to miR-34c expression level in lung adenocarcinomas showing EGFR exon 19 mutation Microarray analysis was used to detect some physiologically relevant miRNAs in lung adenocarcinoma. [score:4]
miR-34c expression was repressed in lung cancer cell lines [20] and other human malignancies, including colorectal, breast, melanoma, head and neck, and prostate cancers [21]. [score:3]
There were no proven correlations between miR-34c expression and tumor stage, pleural involvement, lymphovascular invasion, smoking history in this study. [score:3]
The chi-square test was used to assess miR-34c, miR-183, miR-210 expression with respect to clinicopathological parameters. [score:3]
To investigate the potential biologic roles of three miRNAs, we further subclassified lung adenocarcinomas into “high” and “low” groups in the expression of miR-34c, miR-183, miR-210, based on the mean of expression after normalization [8], and performed survival analysis by univariate Kaplan-Meier method. [score:3]
Herein, we identified that miR-34c, miR-183, and miR-210 showed significantly altered expression in some lung adenocarcinomas. [score:3]
Tumor differentiation was significantly associated with miR-34c, miR-183, and miR-210 expression levels (p = 0.037, 0.029, and 0.003, respectively). [score:3]
We identified three microRNAs (miR-34c, miR-183, and miR-210) which showed significantly altered expression in all groups of lung adenocarcinoma by microarray study. [score:3]
Three miRNAs, miR-34c, miR-183, and miR-210, which with showed statistical differences in their expression among three groups, were selected. [score:3]
Expression levels of miR-34c, miR-183, and miR-210 were significantly different between normal control group and cancer groups (p = 0.034, <0.000, and 0.036, respectively). [score:3]
In spite of limited results, we assume that miR-34c acts like-a tumor suppressor gene in tumorigenesis, but in a particular condition, it may contribute tumor progression. [score:3]
We identified three microRNAs (miR-34c, miR-183, and miR-210) which showed significant altered expression in all groups of lung adenocarcinoma by microarray study. [score:3]
Expression levels of miR-34c, miR-183, and miR-210 were significantly different between normal control group and cancer groups (p = 0.034, <0.000, and 0.036, respectively; Fig.   2a). [score:3]
In conclusion, we show that miR-34c may act as a potential tumor suppressor gene and miR-183 and miR-210 have a potential oncogenic role, using FFPE samples with various histopathologic parameters. [score:3]
Paradoxically, however, there was a positive correlation between miR-34c expression level and poor tumor differentiation. [score:3]
Here, we show that miR-34c may act as a potential tumor suppressor gene and miR-183 and miR-210 have a potential oncogenic role in pulmonary adenocarcinoma. [score:3]
It seems that there is no relationship between miR-34c and EGFR mutation provisionally, but further studies are needed from now on. [score:2]
Catuogno et al. demonstrated that miR-34c-5p may confer resistance to caspase-8 -induced apoptosis by silencing of Bmf (Bcl-2-modifying factor), and miR-34c-5p controls cell proliferation and apoptosis by acting on p53 and c-myc [22]. [score:1]
There is no definite evidence, but the action of miR-34c may depend on biologic circumstance. [score:1]
The transcriptional start site of miR-34c is adjacent to a predicted p53 binding site, transcriptional activation of miR-34c by p53 induces post-transcriptional gene silencing of cyclin -dependent kinase(CDK)-4, CDK-6, cyclinE2, and Bcl2 [19]. [score:1]
miRNA miR-34c miR-183 miR-210 NSCLC Epidermal growth factor receptor Lung cancer is the major leading cause of cancer mortality [1], and non-small cell lung cancer (NSCLC) occupies about 80 % of lung cancer. [score:1]
Thus, we consider that miR-34c may have opposite role in a particular condition. [score:1]
We presume that miR-34c acts differently depending on p53 status. [score:1]
It is an intriguing question that which mechanisms determine miR-34c to have opposite roles on cell survival or apoptosis. [score:1]
[1 to 20 of 35 sentences]
20
[+] score: 93
Using the median expression values for miR-34c and miR-422b as respective cutoffs, high grade serous carcinomas were separated into high -expression group (expression values > median value) and low -expression group (expression values ≤ median value) for Kaplan-Meier survival analysis (Figure 4). [score:11]
Based on microarray-derived expression values, the group with lower level miR-34c expression had decreased recurrence survival (p = 0.049) and decreased disease-specific survival (p = 0.019) compared to the group with higher level miR-34c expression. [score:8]
However, the lack of significant association between miR-34c downregulation and p53 mutation in our current series indicate that additional influences are likely important in regulating miR-34c expression in high grade serous carcinomas. [score:8]
When qRT-PCR derived expression values were used for miR-34c, a similar trend between decreased miR-34c expression and decreased disease-specific survival was seen though statistical significance was not reached (p = 0.06). [score:7]
miR-34c was among the 9 miRNA that were found to be downregulated in high grade serous carcinomas compared to fallopian tubes and high grade serous carcinomas with greater downregulation of miR-34c were associated with more aggressive clinical behavior. [score:6]
With regard to the prognostic significance of miR-34c, analysis based on the microarray data demonstrated significant association between low-level miR-34c expression and decreased disease-specific survival, while analysis based on qRT-PCR data showed a similar trend but did not reach statistical significance with a p-value of 0.06. [score:5]
Further study will be needed to verify the prognostic significance of miR-34c downregulation in high grade serous carcinoma. [score:4]
Functionally, miR-34c expression is known to be regulated by p53 and low level of miR-34c was observed in p53 deficient ovarian carcinoma cell lines [34], [35], [52]. [score:4]
In this study, the downregulation of miR-34c in high grade serous carcinomas shown by microarray analysis was confirmed by qRT-PCR analysis but inter-method variability appears to have affected the reproducibility of demonstration of a statistically significant miR-34c prognostic association, indicating borderline prognostic significance, based on this cohort. [score:4]
Quantitative RT-PCR (qRT-PCR) analysis was performed for selected miRNA (miR-34c, miR-143, miR-145, miR-29a and miR-29b) on the same series of high grade serous carcinoma and normal fallopian tube samples, and the findings are depicted in Figure 3. In accordance with the microarray data, miR-34c, miR-143 and miR-145 showed significant downregulation in high grade serous carcinomas compared to normal fallopian tubes by qRT-PCR analysis. [score:3]
0007314.g003 Figure 3 A) miR-34c expression levels for 32 high grade serous carcinomas (HG serous ca) and 3 fallopian tubes (sample STT5049 was not included in the analysis due to insufficient material). [score:3]
miR-34c was found to be the sole independent predictor of recurrence-free survival (HR = 0.29, 95% CI = 0.10–0.84; p = 0.02) and miR-422b was found to be the sole independent predictor of disease-specific survival (HR = 0.21, 95% CI = 0.08–0.53; p = 0.001). [score:3]
These findings point to a tumor suppressor role for miR-34c in high grade serous carcinomas. [score:3]
Prognostically, lower level miR-422b and miR-34c in high grade serous carcinomas were both associated with decreased disease-specific survival by Kaplan-Meier analysis (p<0.05). [score:3]
There was no significant difference in miR-34c levels between mutation -positive and mutation -negative tumors by either microarray or qRT-PCR analyses. [score:3]
A) miR-34c expression levels for 32 high grade serous carcinomas (HG serous ca) and 3 fallopian tubes (sample STT5049 was not included in the analysis due to insufficient material). [score:3]
Because of the known functional link between p53 and miR-34c [34], [35], [36], p53 mutation status was assessed and interpretable data was available for 20 of the 33 high grade serous carcinomas. [score:2]
High grade serous carcinomas as a group exhibit significant miRNA dysregulation in comparison to tubal epithelium and the levels of miR-34c and miR-422b appear to be prognostically important. [score:2]
Comparison between high grade serous carcinomas and normal tubal epithelium revealed several dysregulated miRNA, including miR-34c and miR-422b, the levels of which are both associated with prognostic importance in our current series. [score:2]
Comparison of 33 high grade serous carcinomas to 3 normal fallopian tube samples identified several dysregulated miRNAs (false discovery rate <5%), including miR-422b and miR-34c. [score:2]
Prognostically, the level of miR-422b and the level of miR-34c were both found to be positive predictors of patient survival by microarray analysis. [score:1]
Both miR-34c and miR-422b were also among the miRNAs found to be significantly dysregulated in high grade serous carcinomas compared to fallopian tubes (Table 2). [score:1]
There was no statistically significant correlation between the levels of miR-34c or miR-422b and the initial tumor stages. [score:1]
Prognostic significance of miR-34c and miR-422b high grade ovarian serous carcinomas. [score:1]
0007314.g004 Figure 4 A) Kaplan-Meier survival curves for low miR-422b level group (dashed line) and high miR-422b level group (solid line) based on microarray analysis, B) Kaplan-Meier survival curves for low miR-34c level group (dashed line) and high miR-34c level group (solid line) based on microarray analysis, C) Kaplan-Meier survival curves for low miR-34c level group (dashed line) and high miR-34c level group (solid line) based on qRT-PCR analysis. [score:1]
A) Kaplan-Meier survival curves for low miR-422b level group (dashed line) and high miR-422b level group (solid line) based on microarray analysis, B) Kaplan-Meier survival curves for low miR-34c level group (dashed line) and high miR-34c level group (solid line) based on microarray analysis, C) Kaplan-Meier survival curves for low miR-34c level group (dashed line) and high miR-34c level group (solid line) based on qRT-PCR analysis. [score:1]
Quantitative RT-PCR analysis of miR-34c, miR-143, miR-145, miR-29a and miR-29b was performed on the current series of high grade serous carcinomas and fallopian tube samples, using the same extracted total RNA used for the microarray analysis. [score:1]
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[+] score: 85
Overexpression of hsa-miR-34c-5p, led to attenuation of JEV induced TNF and IL-6 expression, both at mRNA and protein level, but this overexpression has no effect on JEV replication as no change in viral specific NS1 protein expression was observed in miR-34c-5p overexpressed cells as compared to control mimic transfected cells (Fig. 9D, lower panel). [score:10]
Our study reveal that Notch pathway targeted miRNA (miR-34c-5p) is suppressed during JEV infection and their upregulation impair JEV induced inflammatory cytokine production. [score:8]
We identify miR-34c-5p which is suppressed during JEV infection and overexpression of this miRNA modulates JEV induced proinflammatory cytokine production possibly through downregulating Notch activation. [score:8]
To understand if the miR-34c suppression is JEV specific, we infected the microglia cells with JEV having varying MOI and miR-34c-5p expression was examined after 48 h pi. [score:5]
Overexpression of miR-34c-5p attenuates JEV induced proinflammatory cytokine expression. [score:5]
Expression of all miRNAs (miR-145-5p, miR-26b-5p, miR-34c-5p, and miR-374b-5p) was suppressed in varying degree with JEV infection (Fig. 3F–I). [score:5]
Further, we took miR-34c-5p as it can target multiple gene (Notch1, Dll1, JAG1, Hes1) expression in Notch signaling pathway. [score:5]
In order to understand that these miRNAs are really affect Notch1 pathway, we have checked Notch1, Dll1, JAG1 and Hes1 gene expression after overexpressing the miR-34c-5p by transfecting miRNA specific mimic followed by JEV infection. [score:5]
No significant change was observed in miR-34c-5p expression in neuronal cells infected with two different strains, but significant suppression was observed in mouse microglia as well as mouse brain infected with JEV (Fig. 7C). [score:5]
JEV infection specifically suppressed hsa-miR-34c-5p expression in microglial cells. [score:5]
To check further if this suppression is related to JEV infection, we compared miR-34c-5p expression in WNV infected cells. [score:4]
WNV infection induced miR-34c-5p upregulation was observed in mouse embryonic Fibroblast cells, but no change was noted in MEFs infected with JEV (Fig. 7B). [score:4]
This further suggest that suppression pattern of miR-34c is specific for JEV and might linked to JEV specific pathogenesis. [score:3]
A gradual repression of miR-34c-5p expression was observed with increasing MOI (Fig. 7A). [score:3]
Further, we provided experimental evidence suggesting miR-34c-5p probably binds 3’UTR of Notch gene as overexpression of miR-34c-5p repress luciferase activity in the HEK293T cells when co -transfected with Notch 3’UTR pMirTraget Luciferase vector (Fig. 9B). [score:3]
Our results suggested that strain specific variation may be marginal, but cell type specific variation may exist, especially, we observed microglial cell type specific expression pattern for miR-34c-5p (Fig. 7c). [score:3]
Interestingly, miR-34c-5p highly expressed in WNV infected cells which is totally opposite to the phenomenon that we observed during JEV infection (Fig. 7B). [score:3]
The CHME3 cells were infected with JEV (P20778) with different MOI (0.5, 1, 3 and 5 MOI) for 48 h pi and miR-34c-5p expression was measured by. [score:1]
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[+] score: 82
In this study, we demonstrated that the expression of miR-34 family members is generally down-regulated in late-gestational fetal KCs by NGS. [score:6]
Although the potential target genes of miR-34b-5p are TGF-βRII and SMAD3, as predicted by miRanda, miR-34b-5p may have regulatory effects on the miR-34c-5p potential target genes TGF-β3, TGF-βRII, SMAD4 and SAR1A. [score:6]
6. The miRNA-34 Family is Down-regulated in Late-gestational Fetal KCs and Extensively Targets the TGF-β Pathway. [score:6]
The overexpression of the novel miRNA candidates and miRNA-34 family members in early- to mid-gestational fetal KCs may contribute to scarless wound healing by targeting the TGF-β pathway. [score:5]
The overexpression of miR-34 family members may suppress the TGF-β signal pathway and leads to a loss of control of miR-34a. [score:5]
Moreover, the significantly differentially expressed miRNAs, including some novel miRNA candidates and miR-34 family members, extensively target the TGF-β pathway. [score:5]
We considered that the expression of miRNA-34 family members is generally down-regulated in late-gestational fetal KCs. [score:5]
The expression levels of miR-34a-3p, miR-34b-5p, and miR-34c-3p were changed by more than 2.0-fold, suggesting that these miRNAs are expressed at significantly lower levels. [score:5]
We considered that the overexpression of miR-34 family members may contribute to scarless wound healing in mid-gestational fetal KCs by targeting the TGF-β pathway. [score:4]
Furthermore, we predicted that miR-34 family members may extensively suppress the TGF-β signal pathway. [score:3]
To predict potential target genes of miRNA-34 family members, we used the miRanda online software. [score:3]
Both miR-34a-5p and miR-34c-5p, which have the same seed sequence, targeted TGF-β3, TGF-βRII, SMAD4 and SAR1A. [score:3]
Thirty-three differentially expressed miRNAs and miR-34 family members are correlated with the transforming growth factor-β (TGF-β) pathway. [score:3]
The potential target gene of miR-34a-3p and miR-34c-3p was SMAD4. [score:3]
The results showed that the miRNA-34 family may extensively suppress genes that play important roles in the TGF-β pathway, including TGF-β3, TGF-βRI, TGF-βRII, SMAD3, SMAD4 and SAR1A (Fig 4b and 4c). [score:3]
With the exception of miR-34a-5p, the expression levels of miR-34 family members in late-gestational fetal KCs were significantly lower (p value <0.05). [score:3]
With the exception of miR-34a-5p (p value = 0.078), the expression levels of miR-34 family members in late-gestational fetal KCs were significantly lower than those in mid-gestational fetal KCs (Fig 4b). [score:3]
However, the regulatory effects between miR-34 family members and TGF-β signal pathway are markedly more complicated. [score:2]
Therefore, there are six mature miR-34 miRNAs: miR-34a-3p, miR-34a-5p, miR-34b-3p and miR-34b-5p, miR-34c-3p and miR-34c-5p. [score:1]
Nucleotides 2–10 from the sequence of miR-34b-5p are perfectly matched to nucleotides 1–9 from the sequence of miR-34c-5p. [score:1]
MiR-34a-3p, miR-34b-5p, and miR-34c-3p changed by more than 2.0-fold. [score:1]
Nucleotides 1–10 from the sequence of miR-34c-3p and nucleotides 2–11 from the sequence of miR-34b-3p are matched perfectly. [score:1]
Nucleotides 2–9 from the sequences of miR-34a-5p and miR-34c-5p are matched perfectly. [score:1]
Furthermore, the mature sequences of miR-34 are highly similar to each other. [score:1]
The human miR-34 miRNA precursor family consists of three members encoded by two different transcripts: miR-34a, which is encoded by its own transcript, and miR-34b and miR-34c, which share a common primary transcript. [score:1]
Of all the miR-34 family members, miR-34a has been well studied. [score:1]
Although the mature sequences of miRNA-34 family members were highly similar to each others’, mature miR-34b does not have the same seed sequences as the others. [score:1]
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[+] score: 81
Other miRNAs from this paper: hsa-mir-34a, hsa-mir-34b
Systemic overexpression of miR-34, which is broadly expressed and regulates physiological processes, could target genes in healthy tissues and cause side effects such as cardiovascular disease, although this may be minimised by the use of the particular liposome formulation [92]. [score:10]
In addition, since a considerable number of oncogenes are direct targets of miR-34, and cancer is now considered a multipathway disease [34- 36], this therapeutic approach would allow the use of only one bullet to hit more than one pathway deregulated by the loss of miR-34. [score:6]
In addition, the correlation between miR-34 family expression and patient survival would not always support its tumor suppressor role. [score:5]
Ectopic expression of the members of the miR-34 family can recapitulate some biological functions of p53 such as apoptosis [20, 45] and cell cycle arrest [46, 47], at least in some cell types, although other studies have failed to demonstrate an apoptotic effect of overexpressed miR-34 [44, 48]. [score:5]
Despite the lack of spontaneous tumours in miR-34 knockout mice, there is evidence, at least in some cancers, for miR-34 dysregulation. [score:3]
Moreover, miR-34 knockout mice are born with the normal Men delian ratio, are fertile, and are not, as might be expected, a phenocopy of the p53 knockout. [score:3]
miR34 expression in human cancer. [score:3]
miR-34 Targets list in Cancer. [score:3]
The miR-34 family, which consists of miR-34a, b and c, has attracted a lot of attention since it plays a key role as a tumor suppressor in several cancers [16- 18]. [score:3]
Therefore, we would predict that the reduction of miR-34 expression is associated with poor prognosis and survival. [score:3]
In particular, the miR-34 family binds to the 3'-UTRs of genes such as CDK4 and CDK6 [50, 51] (cell cycle) [19], Bcl-2 [24, 52] (apoptosis), SNAIL [29, 32] (epithelial mesenchymal transition) [53] and CD44 (migration and metastasis) [54], and the miR-34 family thus represses their expression. [score:3]
Thus, while, miR-34 expression is reduced in some tissues in p53 null mice, in others it remains unaffected, confirming the promiscuity and cell context dependency referred to above. [score:3]
In human ovarian cancer (83 samples) miR-34 family expression was found to be reduced when compared to six (apparently mouse) ovarian surface epithelium cell samples. [score:2]
The last 7 years of studies have clearly shown that the miR-34 family is a master regulator of tumor biology. [score:2]
Figure 1 summarizes regulators and functions of the miR-34 family. [score:2]
The miR-34 family acts on apoptosis and cell cycle through the repression of many proteins involved in the regulation of these two biological processes. [score:2]
Clearly, future studies are required in order to have a more coherent picture of the miR-34 family regulation in cancer and whether this family can be used as a prognostic biomarker [80, 81]. [score:2]
The miR-34 Family: origin, regulation and function. [score:2]
Thus, miR34 null mice do not develop spontaneous tumors like p53 knockout mice [55]. [score:2]
Although, as mentioned above, the miR-34 family is regulated by p53, it would be more correct to say the p53 family [42]. [score:2]
The miR-34 family regulators and their functions. [score:2]
As a result of these and other studies, a miR-34 analogue has become the first microRNA to enter the clinic after a surprisingly swift 6 years passage from the bench to bedside. [score:1]
Although this is at first sight surprising, this correlation analysis of the human hepatocellular carcinoma dataset is in agreement with a previous report, and may be further evidence for the cell context dependency of the biological effects of miR-34. [score:1]
In mammals, the miR-34 family consists of three homologous transcripts miR-34a, miR-34b and miR-34c. [score:1]
On the other hand, the phase 1 clinical trail that has recently started represents an important step forward not only for miR-34 itself, but forms a valuable proof of principle study for the rationale of using miRNAs as anticancer drugs. [score:1]
Moreover, systemic delivery of miR-34 in a mouse mo del of hepatocellular carcinoma resulted in a reduced tumor burden and prolonged survival [33]. [score:1]
In particular, miR-34 null mice do not show increased spontaneous or irradiation -induced tumorigenesis, and show only small and subtle differences from wild-type mice in other p53 -dependent functions such as replicative senescence and the DNA damage response [55, 56]. [score:1]
However, the genes coding for both miR-34b and miR-34c map to chromosome 11q23.1 and are located within intron 1 and exon 2 respectively, of the same primary transcript. [score:1]
Survival correlation of miR-34 family in several human cancer datasets. [score:1]
miR-34 family survival analysis in cancer. [score:1]
The miR-34 gene was first identified in C. elegans where it encodes a single miR that is evolutionarily conserved in several invertebrates [37, 38]. [score:1]
However, it should be remembered that miR-34 and p53 have independent functions. [score:1]
Indeed, although the endpoint of this clinical trial at this stage is to investigate the safety, pharmacokinetics and pharmacodynamics of the miR-34 mimetic in patients with unresectable primary liver cancer, it might shed light on two main challenges for miRNA -based therapies: i) delivery system and ii) potential off-target effects. [score:1]
There is also some evidence for the involvement of the miR-34 family, again particularly miR-34a, in cancer stem cells (CSCs). [score:1]
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[+] score: 81
Moreover, by carefully examining the negatively correlated expression of the individual miRNAs and target genes, we found that PLCXD3, a phospholipase that hydrolyzes phospholipids into fatty acids, may be a novel target of miR-34c-3p. [score:7]
The cells (GC-1, TM4 and NCM460 cells) were transfected with the indicated miRNA mimics and inhibitors (hsa-miR-34c-3p mimic: 5′-aaucacuaaccacacggccagg-3′, hsa-miR-34c-3p mimic inhibitor: 5′-ccuggccgugugguuagugauu-3′; negative control mimic: 5′-uuuguacuacacaaaaguacug-3′; negative control inhibitor: 5′-caguacuuuuguguaguacaaa-3′). [score:7]
The downregulation of miR-34c in our microarray results may affect germinal lineage differentiation, which closely correlates with the low sperm concentration in SO disease. [score:6]
This downregulation was efficiently prevented by the miR-34c-3p inhibitor. [score:6]
miR-34c-3p downregulates PLCXD3 expression. [score:6]
The pmirGLO Dual-Luciferase miRNA Target Expression Vector (Promega, USA) was used to confirm the function of the putative miR-34c-3p binding site in the 3′-UTR of PLCXD3. [score:5]
Recently, miR-34c was reported to regulate a number of targets, including Nanos2 [20], p53 [21] and RARg [22], which are mostly involved in the biological process of male germ cell differentiation. [score:4]
Subsequently, we discovered and constructed a post-transcriptional regulatory network with the miRNA-target gene pair miR-34c-3p and PLCXD3 (Phosphatidylinositol-Specific Phospholipase C, X Domain Containing 3) to shed light on the interplay between mRNAs and miRNAs. [score:4]
These results supported the hypothesis that miR-34c-3p directly targeted the PLCXD3 3′-UTR. [score:4]
Furthermore, the luciferase activity and the level of the PLCXD3 protein were downregulated by the miR-34c-3p mimic. [score:4]
The correlation between the expression levels of miR-34c-3p and PLCXD3 provides valuable data for the analysis of spermatogenic dysfunction in male infertility. [score:3]
Confirmation of PLCXD3 as the target gene of miR-34c-3p. [score:3]
This finding suggests that PLCXD3 is a target of miR-34c-3p. [score:3]
Based on the miRNA-mRNA interaction analysis and the results of the PIAT, RNAhybrid, and DIANA predictions, PLCXD3 was a potential target gene of miR-34c-3p (Table 2). [score:3]
Moreover, the western blots suggested that miR-34c-3p repressed the expression of PLCXD3 at the protein level (Figure 8). [score:3]
In this study, we observed an inverse relationship between the expression of miR-34c-3p and PLCXD3 in SO patients. [score:3]
Mouse spermatogonia GC-1 (A), Mouse Sertoli TM4 cells (B), and human colonic epithelial NCM460 cells (C) were transfected with miR-34c-3p mimic/inhibitor (50 nM, 100 nM, respectively), or negative control mimic/inibitor (50 nM, 100 nM, respectively) and PLCXD3 protein levels assayed 48 h post-transfection by Western blot (two technical replicates). [score:2]
Our results are the first to indicate that PLCXD3 was regulated by miR-34c-3p and played an important role in testicular failure. [score:2]
Figure 8Mouse spermatogonia GC-1 (A), Mouse Sertoli TM4 cells (B), and human colonic epithelial NCM460 cells (C) were transfected with miR-34c-3p mimic/inhibitor (50 nM, 100 nM, respectively), or negative control mimic/inibitor (50 nM, 100 nM, respectively) and PLCXD3 protein levels assayed 48 h post-transfection by Western blot (two technical replicates). [score:2]
We also found that the 3′-UTR region of the PLCXD3 region contains one highly conserved miR-34c-3p -binding site (Figure 7A). [score:1]
The blank vector (pmirGLO-Control) has no seed -binding site, and thus firefly luciferase activity was not affected by miR-34c-3p. [score:1]
miR-34c was observed in the late stages of meiosis (pachytene spermatocytes and round spermatids) and is likely to influence the germinal phenotype [19, 20]. [score:1]
Verification of posttranscriptional repression of PLCXD3 by miR-34c-3p in multiple cell types. [score:1]
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[+] score: 72
The miR-34a and the miR-34b/c loci are direct transcriptional targets of the TP53 tumor suppressor (Chang et al., 2007; He et al., 2007; Raver-Shapira et al., 2007; Tarasov et al., 2007), the levels of which, in turn, are indirectly augmented by miR-34. [score:7]
MiRNAs also modulate inflammatory pathways mediated by the transcription factors NFΚB and STAT3 by directly inhibiting IL-6 (via Let-7 miRNAs, which are inhibited by LIN28B) or the IL-6 receptor (via miR-34 and miR-125b). [score:6]
KITLG is a novel target of miR-34c that is associated with the inhibition of growth and invasion in colorectal cancer cells. [score:5]
First, miR-34 directly represses Mdm4 (HDM4 in humans), which encodes a RING-finger protein that binds to TP53 and blocks its ability to activate target genes (Okada et al., 2014). [score:4]
This relationship might be integral to the tumor-suppressive properties of miR-34 miRNAs; the negative regulation of Wnt signaling might also mediate miR-34 -driven repression of intestinal stem cell fate (see below). [score:4]
Known relationships between canonical Wnt signaling and miRNAs are illustrated in Fig.  3. Also operating upstream of Wnt, the miR-34 family (miR-34a/b/c) directly targets and represses multiple effectors of Wnt signaling, including WNT1, WNT3, LRP6 (a Wnt ligand co-receptor), β-catenin and LEF1 (an HMG-box transcription factor that, like TCF4, interacts with β-catenin) (Kim et al., 2011). [score:4]
Known relationships between canonical Wnt signaling and miRNAs are illustrated in Fig.  3. Also operating upstream of Wnt, the miR-34 family (miR-34a/b/c) directly targets and represses multiple effectors of Wnt signaling, including WNT1, WNT3, LRP6 (a Wnt ligand co-receptor), β-catenin and LEF1 (an HMG-box transcription factor that, like TCF4, interacts with β-catenin) (Kim et al., 2011). [score:4]
p53 regulates nuclear GSK-3 levels through miR-34 -mediated Axin2 suppression in colorectal cancer cells. [score:4]
These miRNAs regulate mRNAs involved in the cell cycle (Ebner and Selbach, 2014), growth (Kress et al., 2011), DNA damage (Takeda and Venkitaraman, 2015) and apoptosis (Ebner and Selbach, 2014); these interactions are likely to be associated with the tumor-suppressive properties of miR-34 miRNAs and their ability to induce apoptosis and senescence (Tazawa et al., 2007). [score:4]
p53 and microRNA-34 are suppressors of canonical Wnt signaling. [score:3]
A positive feedback between p53 and miR-34 miRNAs mediates tumor suppression. [score:3]
Lastly, several miRNAs have effects on EMT in CRC tumorigenesis, with miR-15/16 and miR-34 (which are transcriptionally activated by TP53) inhibiting this process, while miR-21 enhances EMT. [score:3]
Aside from acting as an effector of TP53, miR-34a expression downstream of the canonical oncogenic transcription factors, MYC and STAT3, described above, provides negative feedback on tumor cell proliferation, survival and metastasis, which highlights the multifaceted mechanisms by which miR-34 represses tumorigenesis. [score:3]
Thus, both MYC and TP53 can promote miR-34 expression. [score:3]
Studies using cultured human CRC cells (Bu et al., 2013) and mouse mo dels (Bu et al., 2016) indicate that miR-34 also regulates IESC and CRC stem cell division. [score:2]
In summary, many miRNAs act as regulators of the Wnt pathway at multiple levels of the signaling cascade, and some miRNAs, such as miR-34, are capable of restraining both Wnt and Notch signaling pathways. [score:2]
Quantitative proteomic analysis of gene regulation by miR-34a and miR-34c. [score:2]
Second, miR-34 promotes modest (and probably indirect) stimulation of the TP53 promoter (Gao et al., 2015). [score:2]
The transcription of miR-34 is directly stimulated by TP53, providing insight into how TP53 can repress Wnt signaling (Kim et al., 2011). [score:2]
Independent of TP53, miR-34 is induced by FOXO3A (forkhead box O3a transcription factor) in a feedback loop involving MK5 and MYC. [score:1]
The miR-34 family consists of miR-34a, transcribed at one locus, and miR-34b and miR-34c, co-transcribed at another locus. [score:1]
The connection of miR-34 with TNFα and IL-6 also highlights the key role of inflammation, which increases the risk and fuels the progression of CRC (Lasry et al., 2016). [score:1]
Repression of c-Kit by p53 is mediated by miR-34 and is associated with reduced chemoresistance, migration and stemness. [score:1]
The miR-34 anti-oncomiR family and TP53. [score:1]
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[+] score: 61
While miR-34b and let-7i inhibitor/mimic treatments had no substantial effects on PLK4 transcript and protein expression (data not shown), miR-34c mimic considerably up-regulated PLK4 transcript (Figure 6A) and protein expression (Figure 6B), as well as influenza NP levels (Figure 6C). [score:10]
Expression of PLK4 transcript and protein were significantly modulated by miR-34c inhibitor/mimic and these also had a significant impact on viral replication (Figure 6C) suggesting that PLK4 is an important miR-34c target during influenza replication. [score:7]
MiR-34c (and also miR-149*) expression is driven by p53 activation during influenza infection [104] to negatively regulate the activity of the transcription factor, Myc, which regulates S phase progression and DNA replication [106], [107]. [score:5]
Thus, we hypothesize that during influenza virus infection, NS1 mediated p53 up-regulation triggers miR-34c activity to regulate cell cycle through Myc, PLK4 and NEK8 (Figure S6). [score:5]
Thus, a miR-34c mimic transfection can cause S phase cell cycle arrest and promote translation of target transcripts. [score:5]
While miR-34c is believed to primarily target c-Myc [108], and the findings from this study show that miR-34c positively regulates PLK4, other miR-34c targets cannot be discounted and warrant investigation. [score:4]
Inhibition of miR-34c accelerates S phase, promotes excessive DNA synthesis, and affects the Myc-regulated gene, Bcl2, which is important for cell cycle control. [score:4]
Inhibition of miR-34c accelerates cell cycling and increased degradation of PLK4 protein and decreased influenza replication. [score:3]
Analysis of existing literature suggests that miR-34c alters PLK4 activity by modulating the activity of either p53 or by stabilizing PLK4 translation. [score:3]
miR-34c mimic arrests cells in G2/S phase by suppressing Myc leading to increased influenza replication. [score:3]
miRNA regulators of PLK4 (A) A549 cells were transfected with 25 nM of miR-34c inhibitor/mimic for 48 hrs followed by RNA extraction and RT-qPCR with PLK4 specific primers. [score:3]
Additionally, members of the miR-34 family, e. g. miR-34a/b-5, have been shown to enhance translation of neuronal tissue-specific polyadenylated transcript of β-actin [107]. [score:3]
miR-34c suppresses Myc mediated entry into S phase. [score:3]
miR-34c arrest of S phase cell cycle would prevent degradation of PLK4 transcript and protein, and thus help to explain the observation of increased PLK4 transcript and protein following miR-34c mimic treatment. [score:1]
The most significant effects were observed for miR-34c and the PLK4 gene. [score:1]
Interestingly, miR-34c-3p has been shown to be a major miRNA induced during infection by H1N1 and H5N1 influenza viruses [50], although its role in influenza biology is presently incompletely understood. [score:1]
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[+] score: 59
In addition, we also postulate a counteracting pathway in which maintains p53 expression and, indirectly, the expression of miR-34a, providing a substantial protective axis against the loss of let-7. Further studies will be aimed at identifying the mechanism of promotion of p53 expression and its mechanism of negatively regulating let-7. In 2007, several groups identified the miR-34 family of miRNAs (miR-34a, b, and c) as a direct transcriptional target of the key tumor suppressor p53 [26]– [29], [36], [37]. [score:14]
One possible mechanism for the sensitization of cells to -mediated apoptosis is suggested by data showing that miR-34 targets CD44, which has been shown to inhibit -mediated apoptosis by directly binding the region required forL engagement. [score:6]
expression affects the ability of cells to upregulate miR-34 in response to genotoxic stress. [score:6]
In 2007, several groups identified the miR-34 family of miRNAs (miR-34a, b, and c) as a direct transcriptional target of the key tumor suppressor p53 [26]– [29], [36], [37]. [score:6]
The importance of miR-34a and miR-34b is highlighted by their loss of expression in more than a dozen different cancers [37], [38] suggesting a crucial role for miR-34 in suppressing tumorigenesis. [score:5]
It appears that the mere presence of p53 or can affect expression levels of miRNAs, although stimulation through did not have a major effect on the expression of either let-7 or miR-34 (data not shown). [score:5]
Moreover a CD44 variant (CD44v6) binds the miR-34 target, c-Met, and this interaction is required for c-Met signaling [42]. [score:3]
miR-34a is ubiquitously expressed, whereas in most tissues miR-34b and miR-34c are minor species [37]. [score:3]
Although miR-34b and miR-34c are minor species in these cells, these miRNAs also displayed an enhanced response to etoposide treatment in overexpressing cells (Figure 3B ). [score:3]
Induction of pluripotency by Oct24, Sox2, Klf4, and c-Myc in mouse embryonic fibroblasts induces all three miR-34 species to cooperatively inhibit reprogramming by repressing Nanog, Sox2, and N-myc [63]. [score:3]
miR-34 is a selective marker for cancer cells that are sensitive to -mediated apoptosis. [score:1]
Such miRNAs include the let-7 [20], miR-200 [60], and miR-34 families of miRNAs [61]– [64]. [score:1]
In addition, pancreatic cancer cells frequently display miR-34 loss due to epigenetic silencing by methylation. [score:1]
Let-7. p53 and miR-34. [score:1]
Taking into account the role of miR-34 as an effector for p53, it is not surprising that miR-34 also participates in protecting both stem cells themselves and the organism from pluripotent cells that have turned rogue. [score:1]
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[+] score: 56
The unions of targets predicted by the 9 algorithms at FDR<10% cutoff were used as experimental verified miRNA targets, except for miR-34c-5p, which target FDR was taken at 15% due to a weaker inhibition effect, observed in our transfection experiment (Table S14). [score:9]
To assess whether miRNA with expression divergence on the human lineage might be associated with human cognitive functions, we investigated the expression of genes targeted by five miRNA showing human-specific expression, according to multiple methodologies: miR-184, miR-487a, miR-383, miR-34c-5p and miR-299-3p (Figure 2). [score:7]
Functionally, miR-34c-5p was previously shown to be down-regulated in cancer and Parkinson disease [41]– [44]. [score:6]
Table S16 This table contains GO terms and KEGG pathways enriched in experimental verified targets of miRNA showing human-specific expression, hsa-miR-34c-5p. [score:5]
Putative functions of miR-34c-5p targets were determined using CORNA [65], using experimentally verified target genes of miR-34c-5p as predicted by the 9 aforementioned algorithms, at FDR = 15%. [score:5]
These findings indicate that changes in miR-34c-5p expression on the human evolutionary linage might have resulted in gene expression changes affecting cognitive functions. [score:5]
Thus, although indirectly, these results indicate that the change in miR-34c-5p with human-specific expression might have taken place after the separation of the human and the Neanderthal evolutionary lineages. [score:4]
Requiring significant support by at least two out of three methodologies (sequencing, microarrays and Q-PCR), expression changes in five miRNA (miR-184, miR-299-3p, miR-487a, miR-383 and miR-34c-5p) could be assigned to the human evolutionary lineage and two (miR-375 and miR-154*) to the chimpanzee evolutionary lineage (Figure 2). [score:3]
We indeed found a significant excess of human derived SNPs, indicating the presence of positive selection on the human evolution linage after the human-Neanderthal split, in the upstream regions of one out of five miRNA with human-specific gene expression: miR-34c-5p (Fisher's exact test, Bonferroni corrected p<0.05, ). [score:3]
Signature of positive selection found in the enhancer region of the miRNA, miR-34c-5p, further indicates that this change might have had adaptive significance. [score:1]
On the DNA sequence level, these miRNA tend to be conserved: miR-184 mature miRNA sequence is evolutionarily conserved from insects to humans, with only one nucleotide different at 3′end of mature sequence, while miR-383 and miR-34c-5p are classified as broadly conserved and miR-299-3p - as conserved among animal species [25], [31]. [score:1]
Excess of human derived SNPs in the upstream region of hsa-miR-34c. [score:1]
The plot shows 150kb region upstream of human miR-34c. [score:1]
1002327.g007 Figure 7The plot shows 150kb region upstream of human miR-34c. [score:1]
Notably, for miR-34c-5p signature of positive selection was located in the putative enhancer region approximately 100kb upstream of the miRNA gene (Figure 7). [score:1]
Four windows in an upstream region of miR-34c-5p were significant at Bonferroni corrected p<0.05. [score:1]
We further characterized possible functions of miR-34c-5p in the human brain, based on target genes experimentally verified in cell lines. [score:1]
We chose three types of miRNA differences: (1) consistent by both methodologies: miR-383 and miR-34c-5p; (2) significant according to sequencing, but unconfirmed in the microarray experiment: miR-143 and miR-499; (3) significant according to sequencing, but not detected or masked on the microarrays: miR-184 and miR-299-3p. [score:1]
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[+] score: 55
The qPCR confirmed significant a up-regulation of eight miRNAs, b down-regulation of miR-1303 in both the DOX-Day2 and DOX-Day6 groups, and c prolonged up-regulation of miR-182-5p, miR-4423-3p and miR-34c-5p in the drug washout groups at day 14. [score:10]
Enriched GOs of verified gene targets of miR-34c-5p included those such as the sarcomere and muscle contraction and verified down-regulated gene targets including ion channels such as KCNN2, SCN5A, CACNA2D2 and KCNK3. [score:8]
These observations suggest that up-regulation of miR-34c-3p and miR-34c-5p may be early indicators of cardiac dysfunction, the development of cardiac pathologies and future heart failure. [score:5]
Consistent with these observations, our data demonstrated early up-regulation of miR-34c-3p and miR-34c-5p during DOX exposure. [score:4]
qPCR also confirmed the prolonged up-regulation of miR-182-5p, miR-4423-3p and miR-34c-5p in DOX-Day2WO and DOX-Day6WO groups. [score:4]
DOX -induced up-regulation of miR-34b and miR-34c was reported in a mouse mo del (Desai et al. 2014) and in rat hearts (Vacchi-Suzzi et al. 2012). [score:4]
Similarly, miR-34b and miR-34c (miR-34 family members) are up-regulated during rat cardiac hypertrophy (Feng et al. 2014), in the aged hearts of mice (Boon et al. 2013), in mouse hearts with myocardial infarction (Bernardo et al. 2012) and in diabetic ischaemic heart failure patients (Greco et al. 2012). [score:4]
In the present study, an early up-regulation of miR-34 family members was identified, which indicated that apoptosis could be a very early molecular event that induces cardiac cell loss in hiPSC-CMs after repeated exposure to DOX. [score:4]
Furthermore, miR-34c-5p showed persistent up-regulation after the drug washout. [score:4]
Verified gene targets of miR-187-3p, miR-486-5p, miR-34a, miR-212-3p, miR-34c-3p, miR-675-5p and miR-3911 were not enriched for GO terms related to cardiac and general toxicity responses. [score:3]
The miR-34 family members are involved in cardiac ageing, cardiac diseases and in cardiac apoptotic events. [score:3]
Validation of miRNA microarray data using qPCR confirmed deregulation of miR-187-3p, miR-182-5p, miR-486-3p, miR-486-5p, miR-34a-3p, miR-4423-3p, miR-34c-3p, miR-34c-5p and miR-1303 in both DOX-Day2 and DOX-Day6 groups. [score:2]
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[+] score: 54
Strikingly, 13 of the 22 upregulated genes contained 3′UTR miR-34 ‘seed’ matches and were predicted targets of the miR-34 family (Fig. 5 A–B). [score:6]
We performed a microRNA -expression screening and identified 5 members of the miR-34 family (miR-34bc and miR-449abc) as highly expressed from late meiosis to the sperm stage. [score:5]
The same onset of expression in the adult was observed with sustained miR-34c expression detected throughout meiosis and spermiogenesis (Fig. 2F). [score:5]
From the 9 validated miR-34 target genes identified, the forkhead transcription factor FoxJ2 merits special interest as it contains two highly conserved miR-34 binding sites and has been shown that transgenic levels of FoxJ2 overexpression are incompatible with male fertility [40]. [score:5]
The word corresponding to seed matching miR-34 family (Red) is enriched in the up-regulated genes. [score:4]
The expression of the miR-34 family members is summarized from the array data. [score:3]
The spatial expression of miR-34c (Green) is shown by in situ hybridization on sections of 14 dpp mouse testis, the section were counterstained with anti-γH2AX (Red) antibodies to precisely identify the meiotic stage. [score:3]
For the miR-34bc loss of function allele, the targeting strategy allows for Cre -mediated deletion of the hairpins that encode both miR-34b and miR-34c. [score:3]
Representative data is shown from two independent experiments in panel C and D. (E) The onset of miR-34c expression in early pachytene spermatocytes during the first wave of spermatogenesis. [score:3]
miRNA in situ coupled with immunostaining of γH2AX as a meiotic marker revealed the onset of miR-34c expression in early pachytene spermatocytes within the first wave (Fig. 2E). [score:3]
Thus in combination with the histological analysis we can conclude that the miR-34 family has multiple functions during spermatogenesis both in regulating meiosis as well as the later stages of spermiogenesis (Fig. 4F). [score:2]
This unbiased approach revealed a highly significant enrichment (p = 2.44×10 [−9]) for the complementary seed match of miR-34 family (CACTGCC) in the cohort of most unregulated genes (Fig. 5D). [score:2]
Representative images from one of three independent experiments are shown for panel E and F. The miR-34 family genes are proven important regulators of cell fate and physiology. [score:2]
The miR34a locus also regulates cardiac function upon aging, however none of the individual miR-34 family gene disruptions affects fertility in mice (Fig. S2) [32], [34]– [36]. [score:2]
The miR-34b and miR-34c miRNAs are derived from a single non-coding transcriptional unit. [score:1]
The miR-34b/c miRNAs are part of a miR-34 family encompassing six miRNAs (miR-34a, b, c and 449a, b, c) encoded by three distinct loci (miR-34a, miR-34b/c and miR-449) (Fig. 2B). [score:1]
Position of the DNA encoding the pre-miR-34b and pre-miR-34c are indicated. [score:1]
Also indicated is the gene function as well as number of miR-34 binding sites. [score:1]
Representative images from one of three independent experiments are shown for panel E and F. (A) qRT-PCR of miR-34a, miR-34b, miR-34c and miR-449a from control (Ctl) and miR-34bc [−/−];449 [−/−] adult testis. [score:1]
In situ hybridization was performed using LNA-probes with 3′-DIG label (Exiqon) for mir-34c. [score:1]
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[+] score: 49
Thus, by computing the correlation coefficient using the NCI-60 expression profiling data, we quantified the correlation strength between miR-34 familiy and telomerase reverse transcriptase (hTERT) expression profile in different cancer cell lines (except neurologic cancer cells). [score:5]
Our data showed that miR-34a, miR-34b and miR-34c were underexpressed in more than half HCC samples compared with the adjacent tissues, suggesting that down-regulation of miR-34 famlily might be involved in the hepatic carcinogenesis (P < 0.05, Figure 1A–1B). [score:5]
Expression correlation between miR-34 family and hTERT or TP53 expression profile were analyzed using those two database. [score:5]
Kaplan–Meier curves showed that patients with underexpressed miR-34a and miR-34b had poorer overall survival and higher recurrence rates than those with higher expression (P < 0.05), whereas no substantial difference was observed for miR-34c (P > 0.05, Figure 1C–1D). [score:5]
The miR-34 family is frequently downregulated in cancer partly due to the inactivation of p53 [19]. [score:4]
miR-34 family is frequently down-regulated in HCC and associates with poor prognosis. [score:4]
To gain insight into the biological role of miR-34 family in human HCC development, we examined the expression levels of miR-34 family in 75 paired HCC samples by qRT-PCR. [score:4]
Recently, the miR-34 family (a, b and c) has gained attention as they were identified as p53 targets and regulate p53 -mediated cycle arrest and apoptosis [18]. [score:4]
Figure 2 (A, B) Relationship between miR-34 family levels and hTERT mRNA expression in NCI-60 cell lines. [score:3]
Figure 1 (A, B) miR-34a, miR-34b and miR-34c expression were significantly decreased in HCC compared with the corresponding adjacent tissues using qRT-PCR analyses. [score:2]
We then examined the relationship between miR-34 family and telomerase activity in 75 HCC samples by qRT-PCR. [score:1]
Correlation of miR-34 family levels with telomerase activity. [score:1]
Previous reports showed that miR-34 -induced senescence in cancer cell is all in the form of telomere-independent cell cycle arrest. [score:1]
To further explore the potential roles of miR-34 family in affecting malignant characteristics, the expression levels of miR-34 family in tumor tissues were used to build a signature of prognosis. [score:1]
miR-34 family expression is associated with malignant characteristics in patients with HCC. [score:1]
Previous studies demonstrated that miR-34 family could induce cellular senescence by participating in cell cycle arrest. [score:1]
Until recently, it was unknown whether miR-34 family could induce cellular senescence in HCC in a telomere -dependent way. [score:1]
By analyzing nutlin-3a -treated cells, Kumamoto et al. firstly demonstrated that miR-34 family was involved in the p53 -dependent senescence pathway [28]. [score:1]
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[+] score: 48
Top 10 up-regulated and down regulated genes after over expression of mir-34c, mir-98, mir-424 and let-7f in UCI-101 cell line. [score:7]
miR-424 and let-7f were downregulated in both tissues and cell lines (Figure 3A ), while miR-34c and miR-98 were found downregulated in ovarian cancer tissues and in cell lines, respectively. [score:7]
Top 10 up-regulated and down regulated genes after over expression of mir-34c, mir-98, mir-424 and let-7f in BG-1 cell line. [score:7]
To further study the potential targets of key ovarian miRNAs, we overexpressed miR-34c, miR-98, miR-424, and let-7f in 2 ovarian cancer cell lines (BG-1 and UCI-101). [score:5]
This analysis therefore suggests that while the general pathways affected by miR-34c, miR-98, miR-424, and let-7f expression in these two cell lines are related to cancer, the exact molecular pathways targeted are variable and depend on the cell line and on the miRNAs. [score:5]
miR-34c has recently been shown to be a transcriptional target of p53 [41], [42] and can suppress proliferation and colony formation in soft agar in neoplastic epithelial ovarian cells [42]. [score:5]
0002436.g004 Figure 4Pre- miR-34c, Pre-miR-98, Pre- miR-424, Pre- let-7f were overexpressed in BG-1 and UCI-101. [score:3]
There was little consistency in the canonical pathways identified through IPA and except for integrin signaling (found following miR34c or miR98 expression in BG-1), none of the canonical pathways was found more than once in Tables 5 and 6. [score:3]
Pre- miR-34c, Pre-miR-98, Pre- miR-424, Pre- let-7f were overexpressed in BG-1 and UCI-101. [score:3]
Interestingly, we found miR-34c deregulated in ovarian cancer tissues and this finding suggests that miR-34c may play a role in ovarian tumorigenesis through its role in the p53 pathway. [score:2]
In order to experimentally investigate the changes in mRNA levels, we over-expressed 4 different miRNA candidates (miR-98, miR-424, miR-34c and let-7f) in 2 cell lines and assessed transcript levels using an Illumina oligonucleotide array. [score:1]
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[+] score: 44
The strong tumor-suppressive effects observed for miR-34 are likely due to the combined modulation of several target mRNAs involved in different oncogenic processes, rather than regulation of a single target, since none of the miR-34 target mRNAs alone can fully recapitulate the miR-34 loss-of-function phenotype (Kaller et al, 2011). [score:10]
Notably, p53, a well-known tumor suppressor that plays a key role in suppressing cancer by regulating cell cycle, apoptosis and DNA repair, has been shown to transcriptionally activate the expression of all miR-34 family members (Chang et al, 2007). [score:8]
Indeed, the seed -targeting 8-mer LNA was effective in inhibiting all three miR-34 family members in two different cardiac stress mo dels and attenuated cardiac remo deling and atrial enlargement, whereas inhibition of miR-34a alone with a 15-mer LNA -modified provided no benefit in the MI mo del (Bernardo et al, 2012). [score:7]
On the other hand, miR-34 can stimulate p53 activity by targeting and down -regulating SIRT1, an NAD [+] -dependent lysine deacetylase that removes protective acetyl groups on p53, causing p53 ubiquitylation and proteasome -mediated degradation (Yamakuchi et al, 2008), thereby establishing a positive feedback loop. [score:4]
The miR-34 family of miRNAs is consistently down-regulated in a broad range of malignancies. [score:4]
Based on its strong tumor-suppressive effects in vitro, many efforts have focused on increasing miR-34 levels in cancer cells by usings. [score:3]
The second study asked whether inhibition of the miR-34 family (miR-34a, -34b and -34c) by a subcutaneously delivered 8-mer LNA could provide a therapeutic benefit in mice with preexisting pathological cardiac remo deling and dysfunction due to MI (Bernardo et al, 2012). [score:3]
In most cases, the miR-34 mimic was delivered directly by intratumoral injections, which is only therapeutically feasible for easily accessible and localized tumors that have not yet metastasized (Wiggins et al, 2010). [score:2]
The miR-34 family has been shown to control cellular proliferation, cell cycle and apoptosis. [score:1]
In May 2013, Mirna Therapeutics announced the commencement of a phase 1 study of the liposome-formulated miR-34 mimic -based drug, designated as MRX34, in patients with primary liver cancer or metastatic cancer with liver involvement. [score:1]
miR-34 -based cancer therapeutics. [score:1]
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[+] score: 43
Kumamoto K Nutlin-3a activates p53 to both down-regulate inhibitor of growth 2 and up-regulate mir-34a, mir-34b, and mir-34c expression, and induce senescenceCancer Res. [score:11]
Recent reports demonstrate that inhibition of the miR-34 family does not promote tumorigenesis, supporting the potential for therapeutic suppression of this family as a treatment for BPD [56]. [score:5]
To address whether miR-34 expression was required and sufficient for the hyperoxia -induced lung injury and inflammation leading to the BPD pulmonary phenotype, we next asked whether only miR-34a overexpression itself was sufficient, in the absence of hyperoxia i. e., in RA. [score:5]
Given that the miR-34 family has been implicated in the p53 tumor suppressor network, and that p53 pathway defects are common features of human cancer [25], miR-34 inhibition therapy is considered a promising therapeutic approach [26]. [score:4]
Of note, miR-34 family members also have been recognized as tumor suppressor miRNAs. [score:3]
miR-34 overexpression in RA restores the BPD phenotype. [score:3]
Bernardo BC Therapeutic inhibition of the miR-34 family attenuates pathological cardiac remo deling and improves heart functionProc. [score:3]
a Representative graphs showing miR-34 expression in WT NB mice exposed to hyperoxia for 2, 4, and 7 days after birth and in the BPD mo del. [score:3]
In addition, in the PN7 HALI mo del, Ang1 treatment showed improved Ki67 staining levels similar to that of the miR-34 (−/−) mice lungs (Supplementary Fig.   7). [score:1]
b The wild-type Ang1 3′ UTR reporter vector was co -transfected into the MLE12 cells with either the N. C. mimic or miR-34a mimic c The WT Tie2 3′ UTR reporter vector was co -transfected into the MLE12 cells with either the N. C. mimic or miR-34-a mimic. [score:1]
Choi YJ miR-34 miRNAs provide a barrier for somatic cell reprogrammingNat. [score:1]
Concepcion CP Intact p53 -dependent responses in miR-34 -deficient micePLoS Genet. [score:1]
In addition, miR34a−/− mice [71] and conditional miR-34 [fl/fl] [72] (JAX laboratory) and SPC-CreER (gift from Brigid Hogan, PhD, Duke University, USA) were housed in the Yale and Drexel Universities Animal Care Facilities (New Haven, CT and Phila delphia, PA, respectively). [score:1]
Representative bar graph showing tamoxifen deletion of miR-34a in Spc CRE positive miR-34 KO lungs (T2-miR34a [−/−]). [score:1]
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[+] score: 41
MiRNA families such as miR-200 (cfa-miR-200a, cfa-miR-200b and cfa-miR-200c), Mirlet-7 (cfa-let-7a, cfa-let-7b, cfa-let-7c, cfa-let-7g and cfa-let-7f), miR-125 (cfa-miR-125a and cfa-miR-125b), miR-146 (cfa-miR-146a and cfa-miR-146b), miR-34 (cfa-miR-34a, cfa-miR-34b and cfa-miR-34c), miR-23 (cfa-miR-23a and cfa-miR-23b), cfa-miR-184, cfa-miR-214 and cfa-miR-141 were significantly up-regulated with testicular RA intervention via administration of CYP26B1 inhibitor and all-trans-RA (Figure 5). [score:6]
Up-regulated miRNAs (cfa-let-7, cfa-miR-200, cfa-miR-125, cfa-miR-34, cfa-miR-23, cfa-miR-146 clusters, cfa-miR-184 and cfa-miR-214) in adult canine testis treated with DMSO, RA or CYP26B1 inhibitor. [score:6]
In dogs, cfa-miR-34a, cfa-miR-34b and cfa-miR-34c are transcribed from regions of chromosome 5. In murine testis, miR-34a, miR-34b and miR-34c are expressed from post-natal day 11 (P11) and the expression is increased steadily to its highest enrichment at P60. [score:5]
In the present investigation, cfa-miR-34 clusters have been up-regulated significantly by exogenous RA and CYP26B1 inhibitor in canine testis. [score:4]
It has also been observed that enhancement of miR-34 cluster is higher with the CYP26B1 inhibitor treatment than direct RA administration. [score:4]
Mir-21, mir-34c and mir-221/222 control self-renewal of undifferentiated spermatogonia [19], Mirc1, Mirc3 and Mirlet7 regulate spermatogonial differentiation [9], [20], mir-15a and mir-184 mediate differentiation of spermatocytes [18], [19], miR-18, miR-34b, miR-34c, miR-184, miR-383, miR-449 and miR-469 mediate meiotic division of spermatocytes to spermatids [21], [23]– [25] and miR-469, miR-34c regulate differentiation of spermatid to form spermatozoa [24], [25]. [score:3]
Target genes and their biological functions for miR-34 and miR-125 clusters are given in Table S2. [score:3]
Among dysregulated miRNAs in this study, an association network was created for let-7, miR-200, miR-34 and miR-125 families. [score:2]
Additionally, a number of previous studies demonstrated that several miRNA species including miR-34 cluster regulate murine spermatogenesis. [score:2]
Table S2 Associated genes and their biological function for miR34 and miR125 clusters (with references). [score:1]
0099433.g007 Figure 7 Let-7, miR-200, miR-34 and miR-125 clusters are chosen to create networks. [score:1]
These previous findings together support the association of miR-34 clusters, RA signaling and murine spermatogenesis. [score:1]
The results of the current study support the role of cfa-miR-34 clusters with RA -induced spermatogenesis in dogs. [score:1]
In mammals, miR-34 miRNA family members were discovered computationally and then verified experimentally. [score:1]
Let-7, miR-200, miR-34 and miR-125 clusters are chosen to create networks. [score:1]
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[+] score: 39
Other miRNAs from this paper: hsa-mir-34a, mmu-mir-34c, mmu-mir-34b, mmu-mir-34a, hsa-mir-34b
A key regulator of tumor suppression, miR-34 is a direct transcriptional target of the tumor suppressor p53, given that the miR-34a promoter region contains a p53 -binding site [17]. [score:9]
Given that miR-34 was a candidate regulator, we determined PRKD1 mRNA expression and protein translation levels following ectopic expression of miR-34a, miR-34b, and miR-34c. [score:8]
Different miRNAs are involved in the formation and regulation of human BCSCs [7], with previous studies reporting that ectopic expression of miR-34c suppressed epithelial–mesenchymal transition and reduced self-renewal capacity in BCSCs [8]. [score:6]
Expression levels of miR-34b and miR-34c were also detected, however, no significant downregulation of either variant in MCF-7-ADR cells was observed (Supplementary Figure 1A, 1B). [score:6]
Although miR-34a, miR-34b, and miR-34c have the same seed sequence, the results indicated that PKD/PKCμ was downregulated only by miR-34a (Figure 1B). [score:4]
β-actin was used as the loading control and qRT-PCR was performed to validate PRKD1 mRNA and miR-34 variant expression. [score:3]
HEK293T cells were transiently transfected with 3′-UTR reporter constructs (1.5 μg/well in 6-well plates) and 15 nM of miR-34 family precursors (Ambion), using Lipofectamine 2000 (Invitrogen). [score:1]
The seed sequences of miR-34 from PRKD1 were mutated using PCR -based methods and the reporter constructs were verified by sequencing. [score:1]
B. Proteins, mRNAs and totalRNAs were obtained after 48-h transfection of miRNA-34 variants. [score:1]
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[+] score: 39
Cai and colleagues identified an inverse relationship between the expression of miR-34c and c-Met, in 10-paired fresh samples from tumor tissues and adjacent normal tissues of laryngeal carcinoma, showing that down-regulated miR-34c is a critical factor that contributes to malignancy in human laryngeal carcinoma by targeting of c-Met [23]. [score:8]
Cai K. M. Bao X. L. Kong X. H. Jinag W. Mao M. R. Chu J. S. Huang Y. J. Zhao X. J. Hsa-miR-34c suppresses growth and invasion of human laryngeal carcinoma cells via targeting c-Met Int. [score:5]
Moreover, it was reported by the same authors that miR-34 inhibits cell invasion, proliferation and tumorigenesis, whereas c-Met over -expression partially reversed the cell death and cell cycle arrest induced by miR-34 in brain tumors and glioma [20, 21]. [score:5]
Particularly in glioblastoma and ovarian cancer, miR-34 family members’ a–b–c expression was inversely correlated with c-Met expression [19, 20, 21]. [score:5]
Corney D. C. Hwang C. I. Matoso A. Vogt M. Flesken-Nikitin A. Godwin A. K. Kamat A. A. Sood A. K. Ellenson L. H. Hermeking H. Frequent downregulation of miR-34 family in human ovarian cancers Clin. [score:4]
Hereafter, c-Met was established as a bona fide miR-34 target in different tumors such as melanoma, lung, colon, breast and gastric cancer cells [18]. [score:3]
Indeed He and colleagues demonstrated for the first time the direct interaction between miR-34 and c-Met in mouse embryonic fibroblasts (MEF) cells [17]. [score:2]
Several studies pointed out miR-34 as one of the main miR -regulating c-Met. [score:2]
They found that miR-34 was able to overcome HGF -induced gefitinib resistance in HCC827 and PC-9 cells by modulating c-Met and downstream pathway molecules, suggesting a new strategy for reversing HGF -induced resistance to gefitinib in lung cancers [24]. [score:1]
Importantly, miR-34 blocked the phosphorylation signal cascade of c-Met, Akt, ERK and compromised c-Met -driven invasion [18]. [score:1]
MiR-34a is transcribed from chromosome 1, a locus deleted in neuroblastoma, breast, thyroid, and cervical cancer [13, 14, 15, 16], while miR-34b and miR-34c are co-transcribed from a region on chromosome 11. [score:1]
The miR-34 family, consist of miR-34a, miR-34b and miR-34c that are frequently silenced in a variety of tumors, indicating their role in tumorigenesis. [score:1]
2.1. miR-34 Family Members. [score:1]
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[+] score: 39
At 30 weeks of age, the expression of miR-216 (p-value = 0.016), miR-217 (p-value = 0.0078), miR-150 (p-value =0.023), Let-7b (p-value = 0.031,) and miR-96 were significantly downregulated, whereas the expression of miR-146b (p-value = 0.0078), miR-205, (p-value - 0.0078), miR-21, miR-195 (p-value = 0.031), and miR-34c (p-value = 0.063) were significantly upregulated in KC animals compared to control animals (Figure 2B). [score:10]
At 40 weeks of age, the expression of miR-216, miR-217, miR-223, miR-141, miR-483-3p (p-value = 0.031), miR-195, Let-7b (p-value = 0.063) and miR-96 were significantly downregulated; on the other hand, the expression of miR-21, miR-205, miR-146b (p-value = 0.031), and miR-34c (p-value = 0.063) were upregulated in KC mice compared to the control animals (Figure 2C). [score:10]
Notably, the expression of miR-34c is activated by p53 following DNA damage and serves as an important regulator of c-Myc expression, acting downstream to the p38 MAPK/MK2 pathway [70]. [score:6]
The expressions of miR-216 and miR-217 were also progressively reduced in KC mice, but the expressions of miR-21, miR-205, miR-146b, miR-34c, and miR-223 progressively increased (Figure 1A, 2A– 2D). [score:5]
We observed downregulation of miR-146b, miR-34c, and miR-223 at 10 weeks of age; however, their expression increased with the progression of PC in KC animals compared to control animals (Figure 2A– 2D). [score:5]
On the other hand, miR-146b, miR-34c, miR-223, miR-195 (p-value = 0.031) and miR-216 (p-value = 0.063) were downregulated in KC mice compared to control littermates. [score:3]
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[+] score: 37
The miR-34 family is worth special attention because two of its members, mmu-miR-34a and mmu-miR-34b-5p, were significantly up-regulated one day after ENU exposure and maintained increased expression at the 5 subsequent time points up to PTD 30, while another family member, mmu-miR-34c, displayed significant over -expression at multiple time points from PTD 3 to 30. [score:8]
The miR34 family genes are the direct transcription targets of tumor suppressor p53 [32, 38]. [score:6]
Their expressions were enhanced by 3.21-fold (miR-34a), 3.11-fold (miR-34b) and 2.37-fold (miR-34c) on PTD 1 and the fold changes continued to increase and peaked at PTDs 7 or 15. [score:3]
TaqMan qPCR confirmation of the temporal expression changes of miR-34 family miRNAs. [score:3]
Confirmation of the temporal expression changes of three miR-34 family miRNAs and one miR-762 family miRNA by individual TaqMan assays. [score:2]
Figure 3 The temporal expression changes of three miR-34 family members and one miR-762 family member as determined by PCR arrays and individual TaqMan assays. [score:2]
TaqMan MicroRNA Assays were used to confirm the temporal expression changes of 3 miR-34 family members, mmu-miR-34a, mmu-miR-34b-5p, and mmu-miR-34c, as well as a miR-762 family member, mmu-miR-762. [score:2]
A comparison of miR-34 family miRNA expression measured by the two different platforms is shown in Figure 3. The results from the two real-time PCR assay platforms are very consistent and show similar temporal kinetics of miRNA expression for miR-34 family miRNAs, rising from day 1 or 3, reaching peaks at day 15, and decreasing until the end of observation, day 120. [score:2]
Among these miRNAs, the miR-34 family is worth special attention. [score:1]
miR-34b and miR-34c are encoded by the same primary transcript from chromosome 11 in human or chromosome 9 in mouse while miR-34a is located in a different chromosome [35]. [score:1]
Interestingly, our results found that miR-34b and miR-34c changed in correlated manner at all the sampling time points (Figure 3). [score:1]
miR-34c + + + + + +Induction of cell cycle arrest by joining p53 network [35]. [score:1]
Our results indicate that the miR-34 family of miRNAs seems to have the potential to be valuable biomarkers for toxicological application. [score:1]
These biological processes controlled by miRNAs in the miR-34 family are related to ENU cytotoxicity, genotoxicity, and carcinogenicity. [score:1]
Moreover, miRNAs in the miR-34 family worth further study to explore their potential as biomarkers for exposure of genotoxic carcinogens. [score:1]
miRNAs in miR-34 family play important roles in various p53-initiated biological processes. [score:1]
Another miRNA, mmu-miR-762 that is not similar with miR-34 family miRNAs in sequence, were also examined to confirm the array data. [score:1]
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[+] score: 34
By comprehensively analyzing the experimentally validated and all potential target genes of these 6 miRNAs, our study suggested miR-34a-5p, miR-34c-5p and miR-302b-3p seemed to be particularly important in inhibition of lung cancer metastasis by curcumin because their target genes (e. g. CCND1, WNT1, MYC and LEF1) were significantly enriched in metastasis related pathways (Wnt signaling pathway and Focal adhesion). [score:7]
Further, 6 miRNAs (miR-302b-3p, miR-335-5p, miR-338-3p, miR-34c-5p, miR-29c-3p and miR-34a-35p) with more verified target genes and TFs than others in lung cancer review literatures were screened, suggesting these 6 miRNAs might play critical roles in the suppression of lung cancer metastasis by curcumin. [score:5]
In addition, miR-34 was also demonstrated to regulate the TFs followed by the target genes of TFs. [score:4]
However, in our study, we found miR-34c was significantly downregulated (FC = 0.37) after treatment with 10 μM curcumin, indicating miR-34c may be a proto-oncogene. [score:4]
miR-34a-5p, miR-34c-5p and miR-302b can regulate the target gene CCND1, Wnt family member 1 (WNT1) and MYC via the LEF1 transcription factor (Fig 3). [score:4]
This conclusion seemed to be supported by a recent study which indicated forced expression of miR-34c may contribute to resistance to caspase-8 -induced apoptosis in lung cancer cells [37]. [score:3]
Therefore, we believe miR-34a-5p/miR-34c-5p/miR-302b-3p —LEF1—CCND1/WNT1/MYC axis may be a crucial mechanism in inhibition of lung cancer metastasis by curcumin. [score:3]
Nevertheless, it is essential to further explore how miR-34c, miR-34a and miR-302b collectively to regulate LEF1 followed by Wnt signaling related genes in lung cancer [38]. [score:2]
As a member of miR-34 conserved family, miR-34c plays a similar role with miR-34a theoretically. [score:1]
Our present study provides some novel, underlying mechanisms of curcumin (miR-34a-5p/miR-34c-5p/miR-302b-3p— LEF1—CCND1/WNT1/MYC axis) on lung cancer metastasis. [score:1]
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[+] score: 33
The down-regulation of miR-34 upregulates MET and BCL2, which leads to cell proliferation [64, 65, 66]. [score:7]
Decreased expression of miR-34 in lung cancer induces increased -expression of miR-34 target genes, such as BCL2, MET, PDGFRA, and PDGFRB, which leads to TNF-related apoptosis-inducing ligand (TRAIL) -induced cell death. [score:7]
MiR-34, which is transcribed by TP53, directly binds to the PD-L1 3′ untranslated region and downregulates it. [score:6]
A recent study found that tumor PD-L1 expression is regulated by TP53 via miR-34 [67]. [score:4]
Bommer G. T. Gerin I. Feng Y. Kaczorowski A. J. Kuick R. Love R. E. Zhai Y. Giordano T. J. Qin Z. S. Moore B. B. p53 -mediated activation of miRNA34 candidate tumor-suppressor genesCurr. [score:3]
MiR-34 is an important component of TP53 tumor suppressor function [30]. [score:2]
MiR-34 MiR-34 is directly transcribed by TP53, responding to DNA damage and oncogenic stress. [score:1]
MiR-34 is directly transcribed by TP53, responding to DNA damage and oncogenic stress. [score:1]
Kasinski A. L. Slack F. J. miRNA-34 prevents cancer initiation and progression in a therapeutically resistant K-ras and p53 -induced mouse mo del of lung adenocarcinomaCancer Res. [score:1]
The identified TP53/ miR-34/PD-L1 axis deserves consideration for the improvement of emerging immunotherapy. [score:1]
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[+] score: 32
Furthermore, seven miRNAs were expressed more highly in C57BL/6J mice and were mainly downregulated across the time course (miR-92b-3p, miR-34b-5p, miR-672-5p, miR-31-5p, miR-34c-5p, miR-34b-3p, and miR-182-5p; listed in descending order according to the heat map in Figure 5). [score:6]
Five of these [miR-34b-3p, miR-34c-5p, miR-34b-5p, miR-92b-3p, and miR-182-5p; as well as miR-31-5p, which was identified through literature search (41)] belonged to the aforementioned seven miRNAs which were expressed more highly in the C57BL/6J mice and downregulated throughout the time course. [score:6]
Expression of 75 miRNAs, including miRNAs of the miR-21, miR-223, miR-34, and miR-449 correlated with both HA mRNA expression and any of the hematological parameters. [score:5]
Indeed, changes in expression of several of these 20 miRNAs (miR-147-3p, miR-155-3p, miR-223-3p, as well as the miR-34 and miR-449 families) correlate with IAV virulence (14, 15, 17, 64). [score:3]
These 20 miRNAs (which included miR34 families, which are strongly associated with regulation of apoptosis and the PI3k-Akt pathway [e. g., Figure 6]) thus constituted part of a highly regulated response that can predominate in either strain, depending on the time after infection, and is likely to play a role in host susceptibility. [score:3]
Many miRNAs whose expression differed between DBA/2J and C57BL/6J mice during infection belong to the miR-467, miR-449, and miR-34 families. [score:3]
Higher abundance of antiapoptotic (e. g., miR-467 family) and lower abundance of proapoptotic miRNAs (e. g., miR-34 family) and those regulating the PI3K-Akt pathway (e. g., miR-31-5p) were associated with the more susceptible DBA/2J strain. [score:2]
The miR-34 and miR-449 families control epithelial barrier repair (65) and regulate multiciliogenesis via the Delta/Notch pathway (66, 67), which might help transport virions out of the respiratory tract (68) and reduce end-organ damage. [score:2]
Of note, miR-31-5p, miR-379-5p, miR-7a-5p, as well as some members of the miR-449 (-5p) and miR-34 (-5p) families were moderately to highly abundant (>10 CPM), making it more likely that they would bind to a biologically relevant number of viral RNAs. [score:1]
Using the ViTa Database, the human homologs of miR-135b-5p, miR-147-3p, miR-31-5p, miR-379-5p, miR-7a-5p, as well as the miR-449 (-5p) and miR-34 (-5p) families, were predicted to bind to viral RNA segments of influenza A/Puerto Rico/8/34/Mount Sinai (H1N1). [score:1]
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[+] score: 31
Other miRNAs from this paper: hsa-let-7a-1, hsa-let-7a-2, hsa-let-7a-3, hsa-let-7b, hsa-let-7c, hsa-let-7d, hsa-let-7e, hsa-let-7f-1, hsa-let-7f-2, hsa-mir-15a, hsa-mir-16-1, hsa-mir-17, hsa-mir-20a, hsa-mir-23a, hsa-mir-24-1, hsa-mir-24-2, hsa-mir-26a-1, hsa-mir-26b, hsa-mir-29a, hsa-mir-30a, hsa-mir-93, hsa-mir-101-1, hsa-mir-29b-1, hsa-mir-29b-2, hsa-mir-107, hsa-mir-16-2, mmu-let-7g, mmu-let-7i, mmu-mir-15b, mmu-mir-23b, mmu-mir-29b-1, mmu-mir-30a, mmu-mir-30b, mmu-mir-101a, mmu-mir-124-3, mmu-mir-125a, mmu-mir-130a, mmu-mir-9-2, mmu-mir-135a-1, mmu-mir-136, mmu-mir-138-2, mmu-mir-140, mmu-mir-144, mmu-mir-145a, mmu-mir-146a, mmu-mir-149, mmu-mir-152, mmu-mir-10b, mmu-mir-181a-2, mmu-mir-182, mmu-mir-183, mmu-mir-185, mmu-mir-24-1, mmu-mir-191, mmu-mir-193a, mmu-mir-195a, mmu-mir-200b, mmu-mir-204, hsa-mir-30c-2, hsa-mir-30d, mmu-mir-30e, hsa-mir-7-1, hsa-mir-7-2, hsa-mir-7-3, hsa-mir-10a, hsa-mir-10b, hsa-mir-34a, hsa-mir-181a-2, hsa-mir-181b-1, hsa-mir-181c, hsa-mir-182, hsa-mir-183, hsa-mir-204, hsa-mir-181a-1, hsa-mir-221, hsa-mir-222, hsa-mir-200b, mmu-mir-301a, mmu-mir-34c, mmu-mir-34b, mmu-let-7d, mmu-mir-130b, hsa-let-7g, hsa-let-7i, hsa-mir-15b, hsa-mir-23b, hsa-mir-30b, hsa-mir-124-1, hsa-mir-124-2, hsa-mir-124-3, hsa-mir-130a, hsa-mir-135a-1, hsa-mir-135a-2, hsa-mir-138-2, hsa-mir-140, hsa-mir-144, hsa-mir-145, hsa-mir-152, hsa-mir-191, hsa-mir-9-1, hsa-mir-9-2, hsa-mir-9-3, hsa-mir-125a, hsa-mir-136, hsa-mir-138-1, hsa-mir-146a, hsa-mir-149, hsa-mir-185, hsa-mir-193a, hsa-mir-195, hsa-mir-320a, mmu-mir-30c-1, mmu-mir-30c-2, mmu-mir-30d, mmu-let-7a-1, mmu-let-7a-2, mmu-let-7b, mmu-let-7c-1, mmu-let-7c-2, mmu-let-7e, mmu-let-7f-1, mmu-let-7f-2, mmu-mir-15a, mmu-mir-16-1, mmu-mir-16-2, mmu-mir-20a, mmu-mir-23a, mmu-mir-24-2, mmu-mir-26a-1, mmu-mir-26b, mmu-mir-29a, mmu-mir-29c, mmu-mir-93, mmu-mir-34a, mmu-mir-330, mmu-mir-339, mmu-mir-340, mmu-mir-135b, mmu-mir-101b, hsa-mir-200c, hsa-mir-181b-2, mmu-mir-107, mmu-mir-10a, mmu-mir-17, mmu-mir-200c, mmu-mir-181a-1, mmu-mir-320, mmu-mir-26a-2, mmu-mir-221, mmu-mir-222, mmu-mir-29b-2, mmu-mir-135a-2, mmu-mir-124-1, mmu-mir-124-2, mmu-mir-9-1, mmu-mir-9-3, mmu-mir-138-1, mmu-mir-181b-1, mmu-mir-181c, mmu-mir-7a-1, mmu-mir-7a-2, mmu-mir-7b, hsa-mir-29c, hsa-mir-30c-1, hsa-mir-101-2, hsa-mir-34b, hsa-mir-301a, hsa-mir-130b, hsa-mir-30e, hsa-mir-26a-2, hsa-mir-361, mmu-mir-361, hsa-mir-376a-1, mmu-mir-376a, hsa-mir-340, hsa-mir-330, hsa-mir-135b, hsa-mir-339, hsa-mir-335, mmu-mir-335, mmu-mir-181b-2, mmu-mir-376b, mmu-mir-434, mmu-mir-467a-1, hsa-mir-376b, hsa-mir-485, hsa-mir-146b, hsa-mir-193b, hsa-mir-181d, mmu-mir-485, mmu-mir-541, hsa-mir-376a-2, hsa-mir-320b-1, hsa-mir-320c-1, hsa-mir-320b-2, mmu-mir-301b, mmu-mir-674, mmu-mir-146b, mmu-mir-467b, mmu-mir-669c, mmu-mir-708, mmu-mir-676, mmu-mir-181d, mmu-mir-193b, mmu-mir-467c, mmu-mir-467d, hsa-mir-541, hsa-mir-708, hsa-mir-301b, mmu-mir-467e, mmu-mir-467f, mmu-mir-467g, mmu-mir-467h, hsa-mir-320d-1, hsa-mir-320c-2, hsa-mir-320d-2, mmu-mir-467a-2, mmu-mir-467a-3, mmu-mir-467a-4, mmu-mir-467a-5, mmu-mir-467a-6, mmu-mir-467a-7, mmu-mir-467a-8, mmu-mir-467a-9, mmu-mir-467a-10, hsa-mir-320e, hsa-mir-676, mmu-mir-101c, mmu-mir-195b, mmu-mir-145b, mmu-let-7j, mmu-mir-130c, mmu-mir-30f, mmu-let-7k, mmu-mir-9b-2, mmu-mir-124b, mmu-mir-9b-1, mmu-mir-9b-3
Regarding miRNA targeting of these hub genes (Figure  7B), miR-34c-5p and let-7g-5p are main regulatory candidates based on significant expression pattern correlations (FDR < 0.10) with Syt11, Snx-7, Tom1 (both let-7g-5p and miR-34c-5p), and Bim1 (miR-34c-5p). [score:6]
The central role of miR-34c-5p while targeting additional genes in the brown module is again evident in the expression correlation network shown in Additional file 1: Figure S5B (note the higher connectivity of this particular miRNA). [score:5]
The central role played by miR-34c-5p and let-7g-5p while targeting additional genes in the red module is also evident in the expression correlation network shown in Additional file 1: Figure S4 and S5 (note the higher connectivity of these particular miRNAs). [score:5]
Regarding miRNA regulation of this module, the central role of miR-34c-5p and let-7g-5p is yet again evident in the expression correlation network shown in Additional file 1: Figure S6B (note the higher connectivity of these particular miRNAs). [score:4]
MicroRNA miR-34c-5p appeared again as a main modulator of hub genes in this module, namely Ppm1a and Hsp90ab1, as detected by significant correlation among respective miRNA and mRNA expression profiles. [score:3]
The miRNA families that change expression in both mouse and human were: let-7, miR-7, miR-15, miR-101, miR-140, miR-152 (all validated by qPCR, P < 0.05), as well as miR-17, miR-34, miR-135, miR-144, miR-146, miR-301, miR-339, miR-368 (qPCR not performed). [score:3]
This suggests that miR-34c-5p and let-7g-5p play an important role in the attempted regulation of this module. [score:2]
MicroRNAs miR-34c-5p and let-7g-5p, in particular, appear to be central regulators of hub genes in the most significant ethanol-responsive gene modules. [score:2]
50E-0367mmu-miR-339-5pmir-3390.206.807.92E-037.53E-028mmu-miR-34c-5pmir-340.246.689.54E-066.88E-0477mmu-miR-34a-5pmir-340.179.541.17E-029.66E-0245mmu-miR-340-5pmir-3400.178.511.71E-032.45E-0217mmu-miR-361-5pmir-3610.247.887.74E-052.90E-0319mmu-miR-376b-3pmir-3680.268.451.05E-043.50E-0356mmu-miR-376a-3pmir-3680.1910.215.63E-036.40E-0223mmu-miR-434-3pmir-4340.2210.461.76E-044. [score:1]
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Other miRNAs from this paper: hsa-mir-29a, hsa-mir-34a, hsa-mir-34b
Interestingly, emerging data support that down-regulation of miR34 genes, including miR34c, are also implicated in EMT through a negative feedback loop with SNAIL, which comes in line with our findings [103]. [score:4]
According to reports, miR34c along with a panel of additional miRs up-regulated at day 6, are related to senescence (Additional file  15: Table S6). [score:4]
miR-34c expression analysis in OFF, ON and “ESCAPED” HBEC CDC6 Tet-ON cells utilizing: a) qRT-PCR and b) miRseq analysis, tpm (transcripts per million). [score:3]
Step 1; miR34c detection: Before proceeding with ISH of miR34c, its expression following CDC6 induction was confirmed by qRT-PCR (Additional file  14: Figure S9). [score:3]
Prompted by the miRseq analysis we selected miR34c as a miR target. [score:3]
For in situ co-detection we followed a three-step immuno-fluorescence process: i) Fluorescence in situ hybridization (FISH) for miR34c, followed by ii) GL13 staining to spot senescent cells, and finally iii) detection of 53BP1 foci. [score:1]
In addition, miR-34s, including miR34c, were reported to trigger senescence in various human lung settings [104, 105]. [score:1]
Two miRs were of particular interest: miR34c and miR29a. [score:1]
Notably, miR34c was not detected in the “escaped” cells (Fig.   9), which is in agreement with the miRseq and qRT-PCR analysis (Additional file  14: Figure S9), probably because of declined p53 levels in these cells (Fig.   3c). [score:1]
The specificity of each individual probe/antibody was tested by omitting sequentially the following reagents: miR34c probe, GL13 and anti-53BP1 antibodies at 6d ON cells. [score:1]
Fig. 8 In situ detection of miR34c in senescent cells. [score:1]
Step 1: Fluorescence FISH of miR34c employing a double-DIG-labeled LNA probe, visualized as green emission in the cytoplasm, using TSA plus Fluorescein (emitting at 518 nm). [score:1]
Detection of miR34c in senescent cells employing the HBEC CDC6 Tet-ON system. [score:1]
Scale bar: 20 μmAs depicted in Figs.   8 and 9 following a two (steps 1 and 2) and a three step (steps 1, 2 and 3) process we successfully co-detected miR34c in senescent cells that clearly showed evidence of DDR activation (53BP1 foci). [score:1]
Co-detection of miR34c and 53BP1 in senescent cells, employing the HBEC CDC6 Tet-ON system and our proposed protocol (see Additional file  17 and Additional file  18: Figure S10). [score:1]
Expression of miR-34c was calculated relative to U6snRNA levels according to the comparative method of 2- [ΔΔCT]. [score:1]
For each step a parallel experiment took place, omitting the primary reagent (miR34c probe, GL13 and anti-53BP1 antibody), to exclude false positive staining from the secondary antibodies. [score:1]
Scale bar: 20 μm As depicted in Figs.   8 and 9 following a two (steps 1 and 2) and a three step (steps 1, 2 and 3) process we successfully co-detected miR34c in senescent cells that clearly showed evidence of DDR activation (53BP1 foci). [score:1]
Step 1: miR34c FISH employing a double-DIG-labeled LNA probe, visualized as green emission in the cytoplasm, using TSA plus Fluorescein (emission at 518 nm). [score:1]
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This response leads to the activation of Snail1, that along with the upregulation of other genes responsible for loss of adherence, represents a signal promoting cell migration and metastasis (present work, Figure 4 adherens junction); (iii) miR-34 downregulation contributes to the abnormal expression of Snail1, which is normally antagonized by miR-34 and whose pathological expression has been linked to cancer cell epithelial-mesenchymal transition; (iv) miR-200 downregulation contributes to the epithelial-mesenchymal transition as well. [score:14]
All these data strengthen the correlation between high zinc levels, Sanil1 upregulation, miR-34, and miR-200 family members downregulation. [score:7]
Two major miRNAs were downregulated in our samples: a miR-34 family member (−1.1 fold change) and a miR-200 family member (−1.2 fold change). [score:4]
Namely, in the absence of a functional p53 and of a decrease of miRNA-34, Snail1 is upregulated, as we found in our samples. [score:4]
Very interestingly, a decrease in miR-34, which normally antagonizes Snail1, was recently described as part of the p53/miRNA-34 axis. [score:1]
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[+] score: 29
Other miRNAs from this paper: hsa-let-7a-1, hsa-let-7a-2, hsa-let-7a-3, hsa-let-7b, hsa-let-7c, hsa-let-7d, hsa-let-7e, hsa-let-7f-1, hsa-let-7f-2, hsa-mir-17, hsa-mir-18a, hsa-mir-19a, hsa-mir-19b-1, hsa-mir-19b-2, hsa-mir-20a, hsa-mir-21, hsa-mir-24-1, hsa-mir-24-2, hsa-mir-27a, hsa-mir-29a, hsa-mir-30a, hsa-mir-92a-1, hsa-mir-92a-2, hsa-mir-93, hsa-mir-100, hsa-mir-29b-1, hsa-mir-29b-2, hsa-mir-103a-2, hsa-mir-103a-1, hsa-mir-107, mmu-let-7g, mmu-let-7i, mmu-mir-1a-1, mmu-mir-23b, mmu-mir-27b, mmu-mir-29b-1, mmu-mir-30a, mmu-mir-30b, mmu-mir-125a, mmu-mir-9-2, mmu-mir-133a-1, mmu-mir-136, mmu-mir-138-2, mmu-mir-181a-2, mmu-mir-24-1, mmu-mir-191, hsa-mir-196a-1, hsa-mir-148a, hsa-mir-30c-2, hsa-mir-30d, mmu-mir-122, mmu-mir-143, mmu-mir-30e, hsa-mir-34a, hsa-mir-181a-2, hsa-mir-196a-2, hsa-mir-181a-1, mmu-mir-296, mmu-mir-298, mmu-mir-34c, mmu-let-7d, mmu-mir-130b, hsa-let-7g, hsa-let-7i, hsa-mir-1-2, hsa-mir-23b, hsa-mir-27b, hsa-mir-30b, hsa-mir-122, hsa-mir-133a-1, hsa-mir-133a-2, hsa-mir-138-2, hsa-mir-143, hsa-mir-191, hsa-mir-9-1, hsa-mir-9-2, hsa-mir-9-3, hsa-mir-125a, hsa-mir-136, hsa-mir-138-1, mmu-mir-19b-2, mmu-mir-30c-1, mmu-mir-30c-2, mmu-mir-30d, mmu-mir-148a, mmu-mir-196a-1, mmu-mir-196a-2, mmu-let-7a-1, mmu-let-7a-2, mmu-let-7b, mmu-let-7c-1, mmu-let-7c-2, mmu-let-7e, mmu-let-7f-1, mmu-let-7f-2, mmu-mir-18a, mmu-mir-20a, mmu-mir-21a, mmu-mir-24-2, mmu-mir-29a, mmu-mir-29c, mmu-mir-27a, mmu-mir-92a-2, mmu-mir-93, mmu-mir-34a, mmu-mir-103-1, mmu-mir-103-2, mmu-mir-330, mmu-mir-346, hsa-mir-1-1, mmu-mir-1a-2, mmu-mir-107, mmu-mir-17, mmu-mir-19a, mmu-mir-100, mmu-mir-181a-1, mmu-mir-29b-2, mmu-mir-19b-1, mmu-mir-92a-1, mmu-mir-9-1, mmu-mir-9-3, mmu-mir-138-1, hsa-mir-29c, hsa-mir-30c-1, hsa-mir-296, hsa-mir-130b, hsa-mir-30e, hsa-mir-375, hsa-mir-381, mmu-mir-375, mmu-mir-381, hsa-mir-330, mmu-mir-133a-2, hsa-mir-346, hsa-mir-196b, mmu-mir-196b, hsa-mir-18b, hsa-mir-20b, hsa-mir-146b, hsa-mir-519d, hsa-mir-501, hsa-mir-503, mmu-mir-20b, mmu-mir-503, hsa-mir-92b, mmu-mir-146b, mmu-mir-669c, mmu-mir-501, mmu-mir-718, mmu-mir-18b, mmu-mir-92b, hsa-mir-298, mmu-mir-1b, hsa-mir-103b-1, hsa-mir-103b-2, hsa-mir-718, mmu-mir-21b, mmu-let-7j, mmu-mir-21c, mmu-mir-30f, mmu-let-7k, mmu-mir-9b-2, mmu-mir-9b-1, mmu-mir-9b-3
Of the 14 downregulated miRNAs, miR-669c and miR-34c, tended to be downregulated further during the maturation of white adipocytes compared to browns. [score:6]
Of these 10 miRNAs, only mir-21, mir34c and mir-143 were differentially expressed between mature adipocytes and preadipocytes (p < 0.05), see Table 1 and Figure 2. Mir-34c upregulation during brown adipocyte maturation was in contrast to our microarray data and this result must be treated with caution. [score:6]
Two miRNA, miR-669c and miR-34c, demonstrated a trend for downregulation during the maturation of white but not brown adipocytes. [score:4]
Nevertheless, the miRNAs mir-34c, mir-143, mir-24, mir-720 and mir-21 showed robust expression in the adipocyte cultures, and these 5 miRNAs were thus profiled in subcutaneous adipose tissue from healthy humans with different BMIs to examine their regulation in adipose tissue expansion. [score:4]
Figure 2 Expression levels of mir-21, mir-34c and mir-143 in primary murine brown and white preadipocytes and mature adipocytes (n = 3+3 for each tissue). [score:3]
The miRNAs targeting Sirt1 include miR-143, miR-23b miR-34c as well as mir-34a [51, 52]. [score:3]
Five miRNAs (mir-21, mir-143, mir-34c, mir-24 and mir-720) were profiled in subcutaneous adipose tissue from healthy humans with varying degrees of obesity. [score:1]
Figure 4 Expression levels of mir-21, mir-24, mir-34c, mir-143 and mir-720 were measured in subcutaneous adipose tissue of obese (BMI >30, n = 10) and non-obese (BMI <30, n = 10) healthy persons. [score:1]
Of the 10 miRNAs that showed expression in the adipocyte cultures, we chose a subset of 5 miRNAs (mir-34c, mir-143, mir-24, mir-720 and mir-21) to measure in human adipose tissue RNA samples from obese persons (BMI >30, n = 10) and non-obese persons (BMI <30, n = 10). [score:1]
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[+] score: 29
The log2 -based background fluorescence intensity is between 5 and 6, so tumor lung and non-lung normal human tissues have essentially no miR-34b expression and some minimally detectable miR-34c, consistent with the TaqMan [®] -based results in the panel A. D, expression of the 17 "core" genes in specimens from the Database 1; blue bar, genes with higher expression in SCC; red bar, genes with higher expression in AD; *, two genes with higher expression in normal lung; scale bar represents fold change while gray in the heat map indicates missing data. [score:11]
A search of miRNAs preferentially expressed in normal lung in our previously published dataset found a group of 3 miRNAs (miR-34b, miR-34c, and miR-449) that had approximately a thousand copies or less each cell in testes, fallopian tubes, lung, and trachea, while the rest of tissues examined had no or barely detectable levels of expression [9] (Figure 2A). [score:5]
One is from our previous published miRNA expression profiles in the NCI-60 panel of cell lines derived from human cancers that used real-time PCR for quantitation [22], and the expression of miR-34b and miR-34c in 9 cell lines derived from lung was compared with that in normal lung tissue obtained from our body map data [9] (Figure 3, and Addition file 2 and its Table 1). [score:4]
Another dataset used a bead -based technology to quantitate miRNAs [8], and the expression of both miR-34b and miR-34c is again significantly higher in normal lung than in 6 lung tumor specimens (Figure 2C, p = 0.004 and 0.002, respectively, by t-test). [score:3]
Expression of the four miRNA sequences quantitated in this study (miR-34b/34bN and miR-34c/34cN) in normal lung is from 90-fold to over 1,300-fold higher than in any of the lung cancer cell lines tested (Figure 2B). [score:3]
The third report did not have information for miR-34c but showed increased expression of miR-34b in lung [19]. [score:3]
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[+] score: 29
Using miR-34 direct up-regulation by doxorubicin, we show here that p53 induction results in the down-regulation of Dll1 via miR-34 transcriptional control. [score:8]
For this reason, we performed additional Dll1 3’-UTR reporter activity assays using miR-34b- and miR-34c-containing expression constructs, and showed that both miR-34b and miR-34c down-regulate Dll1 3’-UTR to the same levels as those seen with miR-34a (Fig. S5D). [score:5]
An additional question was raised whether other miR-34 family members can have synergistic actions on Dll1 down-regulation. [score:4]
The MiR-34 family is directly regulated by the transcription factor p53 [9], [10], [11], and all of the members of this family (miR-34a, mi-R34b and miR-34c) share high sequence similarities [12]. [score:3]
We thus asked whether by targeting Dll1, miR-34 can impair the proliferation rate of MB cells. [score:3]
These data provide further supporting evidence that the whole miR-34 family (miR-34a, miR-34b and miR-34c) can regulate Notch signaling through Dll1 in MB. [score:2]
This activation can be explained by the relatively high expression of miR-34 in this clone, as compared to clone #2 (Fig. S1C). [score:2]
This evidence led to a mo del for the potential therapeutic use of miR-34 as a radio-sensitizing agent in p53-mutant breast cancer [14]. [score:1]
Several studies have confirmed that the miR-34 family is required for normal cell responses to DNA damage following irradiation in vivo. [score:1]
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c-Myc overexpression partially rescued RhoA expression (Fig. 5A, compare lane 4 with 3) and miR-34 -induced suppression of invasion (Fig. 5B), suggesting that miR-34a inhibits invasion, at least partially, via RhoA reduction by targeting c-Myc. [score:11]
c-Met reversed miR-34 -induced suppression of invasion, indicating that miR-34a inhibits invasion, at least partially, by targeting c-Met (Figs. 7B and C). [score:7]
p53 has been found to target the miR-34 family [4], [5], [6] and the ectopic expression of miR-34 genes has drastic effects on cell proliferation and survival. [score:5]
miR-34c has been shown to negatively regulate c-Myc in response to DNA damage and to inhibit c-Myc -induced DNA synthesis [14]. [score:4]
The results clearly revealed that miR-34 reduced invasion of PC-3 cells to 20% of that of controls (Figs. 2A and B). [score:1]
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[+] score: 28
The relationship between miR-71 or miR-34 expression levels and mf counts tended to follow a similar pattern. [score:3]
In C. elegans, both miR-71 and miR-34 showed stage-specific up-regulation in dauer larvae compared to late second-stage larvae [57]. [score:3]
Five highly abundant mature nematode-derived miRNA sequences (bma-miR-100d_R+1, bma-miR-100c_R+1_1ss12CT, asu-miR-71, cel-miR-34-5p_R+1_1ss1AT, and bma-miR-228), were selected for amplification by RT-qPCR. [score:1]
In contrast, neither miR-71 nor miR-34 was detected in samples from four uninfected dogs. [score:1]
Both miR-71 and miR-34 were detected in all four samples from dogs infected with D. immitis (Table 3). [score:1]
Like miR-71, miR-34 promotes longevity in C. elegans [55]. [score:1]
Number of positive RT-qPCR reactions for miR-71, miR-34 and miR-223. [score:1]
The sample ranking was highly similar for miR-34, with Dim4 and Dim1 alternating with the highest copy numbers. [score:1]
Further efforts to validate D. immitis/ B. pahangi miR-71 and miR-34 as diagnostic candidates are necessary, as the resulting copy numbers displayed extreme variation. [score:1]
Isoforms of miR-100 were the most abundant, followed by miR-71, miR-34, miR-228, miR-50, and miR-57. [score:1]
A possible explanation is that miR-34 may be released in higher proportions by mf than by adults. [score:1]
As miR-71 and miR-34 were previously reported in B. pahangi whole-worm extracts [28], we attempted their amplification from two B. pahangi-infected dogs. [score:1]
The most abundant of the detected miRNAs was miR-71, followed by miR-34 and miR-223. [score:1]
0002971.g001 Figure 1A: miR-71, only D. immitis and B. pahangi-infected samples; B: miR-34, only D. immitis and B. pahangi-infected samples; C: miR-223, all samples. [score:1]
Table S8 Detailed RT-qPCR information for miR-34. [score:1]
In terms of absolute copy numbers, miR-34 was approximately 12 times more abundant in infected dog plasma than in the whole adult extract [46]. [score:1]
A: miR-71, only D. immitis and B. pahangi-infected samples; B: miR-34, only D. immitis and B. pahangi-infected samples; C: miR-223, all samples. [score:1]
In contrast, the maximum copy number variation between experiments using the same sample was 1.7-fold for miR-34 and 3.8-fold for miR-223. [score:1]
Black bars = experiment 1; gray bars = experiment 2; white bars = experiment 3. A: miR-71; B: miR-34. [score:1]
0002971.g002 Figure 2A: miR-71; B: miR-34. [score:1]
Figure 2 shows the relationship between mf counts per ml dog blood (D. immitis and B. pahangi) and miR-71 and miR-34 copy numbers (from one experiment only; gray bars in Figure 1) per ml dog plasma. [score:1]
The miR-34 family has been described in a number of species. [score:1]
Both miR-71 and miR-34 were detected in B. pahangi-infected dog plasma. [score:1]
Similarly, >13-fold difference in miR-34 levels (2.5*10 [5]–3.4*10 [6] copy number per ml plasma) was observed for the same four samples showing comparable mf counts. [score:1]
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[+] score: 28
miR-15b, miR-16, miR-181b and miR-34 have the same downstream target, B-cell lymphoma 2 (BCL-2), which exhibits an antiapoptotic function; overexpression of these miRNAs inhibits the expression of BCL-2 and induces apoptosis. [score:9]
miR-34 molecules act as tumor suppressors by regulating the expression of the corresponding targets. [score:8]
Restoration of miR-34b and miR-34c promotes the repression of cell growth, which demonstrates that miR-34b and miR-34c function as tumor-suppressor genes (46). [score:3]
The miR-34 family, itself targeted by p53 via a positive feedback mechanism, is universally inactivated in various types of cancer. [score:3]
Therefore, the aberrant methylation of miR-34b and miR-34c may be a diagnostic or predictive biomarker, and the re -expression of miR-34b and miR-34c using demethylation drugs may be a novel therapeutic strategy for GC or a useful preventative measure. [score:1]
Recently, Suzuki et al (47) found that aberrant methylated miR-34b and miR-34c could be an important predictive biomarker of metachronous GC risk. [score:1]
In addition to the miR-124a family, miR-34b and miR-34c have also been observed to be silenced by aberrant promoter -associated CGI methylation in the majority of GC cell lines and tissues. [score:1]
miR-124a and miR-34b/miR-34c. [score:1]
The authors also demonstrated that DNA methylation of miR-34b and miR-34c was associated with H. pylori infection in normal individuals, and that the methylation levels of miR-34b and miR-34c in the non-cancerous gastric mucosae of patients with multiple GC were higher than those of patients with single GC. [score:1]
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MicroRNA Expression in cancer Function Mechanism of deregulation Targets Let-7a-2 Down in breast, lung, colon, ovarian, and stomach cancer Tumor suppressor Repressed by MYC KRAS, HMGA2, MYC, DICER, BCLXL, IMP-1, CDC34, IL6 miR-15/16 Down in CLL, prostate cancer, and pituitary adenomas Tumor suppressor Genomic loss, mutated, activated by p53 BCL2, COX2, CHECK1, CCNE1, CCND1, CCND2, BMI-1, FGF2, FGFR1, VEGF, VEGFR2, CDC25a miR-29 family Down in AML, CLL, lung and breast cancer, lymphoma, hepatocarcinoma, rhabdomyosarcoma Tumor suppressor Genomic loss, activated by p53, repressed by MYC CDK6, MCL1, TCL1, DNMT1, DNMT3a, DNMT3b miR-34 family Down in colon, lung, breast, kidney, and bladder cancer Tumor suppressor Repressed by MYC SIRT1, BCL2, NOTCH, HMGA2, MYC, MET, AXL. [score:14]
Frequent downregulation of miR-34 family in human ovarian cancers. [score:4]
In accordance with the loss of TP53, miR-34 members (miR-34a, b, c) have been found strongly repressed in ovarian cancer: miR-34a expression is decreased in 100% and miR-34b*/c in 72% of EOC with p53 mutation (Corney et al., 2010). [score:4]
An interesting example of multi mechanism control of miRNA expression in ovarian cancer is represented by the miR-34 family. [score:3]
miR-34 clusters are part of the transcriptional program activated by p53 (He et al., 2007). [score:1]
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[+] score: 26
As a direct target of p53, miR-34 targeting can potentially have effects on any number of cancer types. [score:6]
Thus, its downregulation in the respective cancers in those organs would likely benefit the greatest from a miR-34 mimetic like MXR34. [score:4]
0060301) 19053174 Corney DC Hwang CI Matoso A Vogt M Flesken-Nikitin A Godwin AK Kamat AA Sood AK Ellenson LH Hermeking H 2010 Frequent downregulation of miR-34 family in human ovarian cancers. [score:4]
The miR-34a is a member of miR-34 family that target TGF-β/Smad4 signaling in T-regulatory (Treg) cell tumor recruitment. [score:4]
Some of the roles of miR-34 have already been discussed, as one of its targets is Smad4. [score:3]
Analysis of miR-34 in human epithelial ovarian cancer showed that there was a 100% decrease in miR-34, and a 72% decrease in miR-34b*/c in the context of p53 mutation (Corney et al. 2010). [score:2]
Wild type p53 correlated with a 93% decrease in miR-34 in ovarian cancer cells. [score:1]
Clinically, stage III and stage IV tumors were analyzed where reduced miR-34 was significant (P = 0.0029, P = 0.0171, respectively). [score:1]
In these phase I studies, miR mimetics have been used to restore miR-34 (MRX34) and miR-16 (TargomiRs) activity. [score:1]
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[+] score: 26
Other miRNAs from this paper: hsa-mir-34a, hsa-mir-34b
Indeed, the miR-34 family members function as tumor suppressors, inducing apoptosis, cell cycle arrest and senescence, in part, through their interaction with the p53 tumor suppressor network [33, 37– 39]. [score:5]
The miR-34 family, most notably miR-34a, is frequently lost or down-regulated in human malignancies including neuroblastoma, breast, lung, and colorectal carcinomas, and osteosarcoma. [score:4]
Among the miRNAs implicated in cancer development and progression, the miR-34 family has been intensively studied and data indicate family members function as tumor suppressors in a variety of human cancers [25, 26]. [score:4]
Indeed, a variety of miRNA formulations and target-specific delivery strategies have accelerated the clinical development of miR-34 mimics, Miravirsen (Santaris Pharma) and MRX34 (Mirna Therapeutics) which recently entered first-in-human phase I clinical trials (NCT01829971) in patients with advanced solid tumors [50– 52]. [score:4]
Furthermore, p53 -mediated transcriptional regulation of the miR-34 family is conserved across different cell types [33– 37]. [score:2]
MiR-34 genes exhibited minimal deletions, loss of heterozygosity (LOH), and epigenetic inactivation in human OSA tumor tissues, demonstrating that other genetic and epigenetic mechanisms may account for the observed decrease expression [30]. [score:2]
The mature miR-34a sequence is located within the second exon of its non-coding host gene whereas miR-34b and miR-34c are co-transcribed and located within a single non-coding precursor (miR-34b/c) [25]. [score:1]
Deletions of the gene regions harboring these transcripts or CpG promoter methylation with miR-34 gene silencing are frequently observed in human malignancies including neuroblastoma, glioma, breast cancer, non-small cell lung cancer, colorectal cancer, and osteosarcoma [27– 32]. [score:1]
Effects of miR-34 on OSA cell line migration and invasiveness appear to be at least partially mediated through repression of CD44, the receptor for hyaluronic acid and a well-established marker of cancer cell stemness [43]. [score:1]
The miR-34 family consists of three evolutionarily conserved miRNAs: MiR-34a, MiR-34b and MiR-34c. [score:1]
miR-34: from bench to bedside. [score:1]
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[+] score: 25
Other miRNAs from this paper: hsa-mir-34a, mmu-mir-34c, mmu-mir-34b, mmu-mir-34a, hsa-mir-34b
The aims of the current work were to examine whether miR-34 family members regulate MAT2A and MAT2B expression and whether SAMe and MTA target this axis in multiple human cancers where miR-34a has been reported to be down-regulated. [score:9]
All three family members are direct transcriptional targets of p53 and many of the targets of the miR-34 family members are involved in cell cycle, apoptosis, invasion and migration [2, 4]. [score:6]
MiR-34 family members are transcriptional targets of p53 and p53 was shown to inhibit CRC metastasis by inducing miR-34a [5]. [score:5]
Most of the published literature shows miR-34 family members as tumor suppressors [2, 4]. [score:3]
MiR-34a is part of a family that includes miR-34b and miR-34c, with miR-34a having its own transcript while the other two share a common primary transcript [4]. [score:1]
R KO (C) and SW620 (D) cells were treated with 250 μM SAMe or MTA, overexpression of miR-34a or miR-34b as described in Methods for 24 hours and were processed for apoptosis, growth by BrdU, miR-34 and miR-34b transfection efficiency measurements. [score:1]
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[+] score: 25
Other miRNAs from this paper: hsa-let-7a-1, hsa-let-7a-2, hsa-let-7a-3, hsa-let-7b, hsa-let-7c, hsa-let-7d, hsa-let-7e, hsa-let-7f-1, hsa-let-7f-2, hsa-mir-15a, hsa-mir-16-1, hsa-mir-17, hsa-mir-18a, hsa-mir-19a, hsa-mir-20a, hsa-mir-21, hsa-mir-22, hsa-mir-23a, hsa-mir-26a-1, hsa-mir-26b, hsa-mir-27a, hsa-mir-29a, hsa-mir-30a, hsa-mir-31, hsa-mir-33a, hsa-mir-92a-1, hsa-mir-92a-2, hsa-mir-93, hsa-mir-96, hsa-mir-99a, hsa-mir-101-1, hsa-mir-29b-1, hsa-mir-29b-2, hsa-mir-106a, hsa-mir-16-2, hsa-mir-192, hsa-mir-199a-1, hsa-mir-148a, hsa-mir-30c-2, hsa-mir-30d, hsa-mir-139, hsa-mir-7-1, hsa-mir-7-2, hsa-mir-7-3, hsa-mir-10a, hsa-mir-10b, hsa-mir-34a, hsa-mir-181a-2, hsa-mir-181b-1, hsa-mir-181c, hsa-mir-182, hsa-mir-183, hsa-mir-199a-2, hsa-mir-199b, hsa-mir-203a, hsa-mir-210, hsa-mir-181a-1, hsa-mir-214, hsa-mir-215, hsa-mir-219a-1, hsa-mir-221, hsa-mir-222, hsa-mir-223, hsa-mir-224, hsa-mir-200b, hsa-let-7g, hsa-let-7i, hsa-mir-15b, hsa-mir-23b, hsa-mir-27b, hsa-mir-30b, hsa-mir-122, hsa-mir-124-1, hsa-mir-124-2, hsa-mir-124-3, hsa-mir-125b-1, hsa-mir-128-1, hsa-mir-130a, hsa-mir-132, hsa-mir-133a-1, hsa-mir-133a-2, hsa-mir-135a-1, hsa-mir-135a-2, hsa-mir-140, hsa-mir-142, hsa-mir-143, hsa-mir-145, hsa-mir-153-1, hsa-mir-153-2, hsa-mir-191, hsa-mir-9-1, hsa-mir-9-2, hsa-mir-9-3, hsa-mir-125a, hsa-mir-125b-2, hsa-mir-126, hsa-mir-134, hsa-mir-136, hsa-mir-146a, hsa-mir-150, hsa-mir-185, hsa-mir-190a, hsa-mir-194-1, hsa-mir-195, hsa-mir-206, hsa-mir-200c, hsa-mir-155, hsa-mir-181b-2, hsa-mir-128-2, hsa-mir-194-2, hsa-mir-29c, hsa-mir-30c-1, hsa-mir-200a, hsa-mir-101-2, hsa-mir-219a-2, hsa-mir-34b, hsa-mir-99b, hsa-mir-296, hsa-mir-130b, hsa-mir-30e, hsa-mir-26a-2, hsa-mir-370, hsa-mir-373, hsa-mir-374a, hsa-mir-375, hsa-mir-376a-1, hsa-mir-151a, hsa-mir-148b, hsa-mir-331, hsa-mir-338, hsa-mir-335, hsa-mir-423, hsa-mir-18b, hsa-mir-20b, hsa-mir-429, hsa-mir-491, hsa-mir-146b, hsa-mir-193b, hsa-mir-181d, hsa-mir-517a, hsa-mir-500a, hsa-mir-376a-2, hsa-mir-92b, hsa-mir-33b, hsa-mir-637, hsa-mir-151b, hsa-mir-298, hsa-mir-190b, hsa-mir-374b, hsa-mir-500b, hsa-mir-374c, hsa-mir-219b, hsa-mir-203b
Microarray profiling studies showed reduction in miRNAs expression specific of HCV and HBV -associated cases: down-regulation of miR-190, miR-134, and miR-151 occurs in HCV cases, and of miR-23a, miR-142-5p, miR-34c, in HBV cases (Ura et al., 2009). [score:6]
Izzotti et al. (2009a, b) have monitored the expression of 484 miRNAs in the lungs of mice exposed to cigarette smoking, the most remarkably downregulated miRNAs belonged to several miRNA families, such as let-7, miR-10, miR-26, miR-30, miR-34, miR-99, miR-122, miR-123, miR-124, miR-125, miR-140, miR-145, miR-146, miR-191, miR-192, miR-219, miR-222, and miR-223. [score:6]
As quoted above, up-regulation of miRNAs, including miR-17-92 cluster, miR-106a, and miR-34, occurs during tamoxifen -induced hepatocarcinogenesis in female rats (Pogribny et al., 2007), also long-term-administration of 2-AAF resulted in disruption of regulatory miR-34a-p53 feed-back loop (Pogribny et al., 2009). [score:5]
In rats, tamoxifen up-regulate miR-17-92 cluster, miR-106a, and miR-34 (Pogribny et al., 2007). [score:4]
4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), a tobacco carcinogen, down-regulate miR-34, miR-101, miR-126, and miR-199 (Kalscheuer et al., 2008). [score:4]
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[+] score: 25
Other miRNAs from this paper: hsa-let-7a-1, hsa-let-7a-2, hsa-let-7a-3, hsa-let-7b, hsa-let-7c, hsa-let-7e, hsa-mir-17, hsa-mir-18a, hsa-mir-19a, hsa-mir-20a, hsa-mir-21, hsa-mir-92a-1, hsa-mir-92a-2, hsa-mir-93, hsa-mir-101-1, hsa-mir-106a, hsa-mir-107, hsa-mir-192, hsa-mir-34a, hsa-mir-204, hsa-mir-205, hsa-mir-214, hsa-mir-215, hsa-mir-222, hsa-mir-223, hsa-mir-1-2, hsa-mir-15b, hsa-mir-125b-1, hsa-mir-141, hsa-mir-191, hsa-mir-125a, hsa-mir-125b-2, hsa-mir-126, hsa-mir-127, hsa-mir-149, hsa-mir-184, hsa-mir-186, hsa-mir-200c, hsa-mir-1-1, hsa-mir-200a, hsa-mir-101-2, hsa-mir-34b, hsa-mir-339, hsa-mir-146b, hsa-mir-548a-1, hsa-mir-548b, hsa-mir-548a-2, hsa-mir-548a-3, hsa-mir-548c, hsa-mir-624, hsa-mir-650, hsa-mir-651, hsa-mir-548d-1, hsa-mir-548d-2, hsa-mir-449b, hsa-mir-1185-2, hsa-mir-1283-1, hsa-mir-1185-1, hsa-mir-708, hsa-mir-548e, hsa-mir-548j, hsa-mir-1285-1, hsa-mir-1285-2, hsa-mir-548k, hsa-mir-548l, hsa-mir-548f-1, hsa-mir-548f-2, hsa-mir-548f-3, hsa-mir-548f-4, hsa-mir-548f-5, hsa-mir-548g, hsa-mir-548n, hsa-mir-548m, hsa-mir-548o, hsa-mir-548h-1, hsa-mir-548h-2, hsa-mir-548h-3, hsa-mir-548h-4, hsa-mir-548p, hsa-mir-548i-1, hsa-mir-548i-2, hsa-mir-548i-3, hsa-mir-548i-4, hsa-mir-1283-2, hsa-mir-548q, hsa-mir-548s, hsa-mir-548t, hsa-mir-548u, hsa-mir-548v, hsa-mir-548w, hsa-mir-548x, hsa-mir-548y, hsa-mir-548z, hsa-mir-548aa-1, hsa-mir-548aa-2, hsa-mir-548o-2, hsa-mir-548h-5, hsa-mir-548ab, hsa-mir-548ac, hsa-mir-548ad, hsa-mir-548ae-1, hsa-mir-548ae-2, hsa-mir-548ag-1, hsa-mir-548ag-2, hsa-mir-548ah, hsa-mir-548ai, hsa-mir-548aj-1, hsa-mir-548aj-2, hsa-mir-548x-2, hsa-mir-548ak, hsa-mir-548al, hsa-mir-548am, hsa-mir-548an, hsa-mir-548ao, hsa-mir-548ap, hsa-mir-548aq, hsa-mir-548ar, hsa-mir-548as, hsa-mir-548at, hsa-mir-548au, hsa-mir-548av, hsa-mir-548aw, hsa-mir-548ax, hsa-mir-548ay, hsa-mir-548az, hsa-mir-548ba, hsa-mir-548bb, hsa-mir-548bc
Strongly down-regulated tumor suppressing miRNAs included miR-1, miR-34c, let-7a, let-7b, miR-127, with a percentile rank of inhibition of 96.2, 84.2, 69.1, 61.6, 59.3, respectively. [score:8]
Other down-regulated tumor suppressors were let-7c, miR-101, let-7e, miR-125b, miR-141, miR-126, miR-34a, miR-34c and miR-200a. [score:6]
These microRNAs, that included miR-15b, miR-205, miR-34a, miR-34c, miR-192, miR-200a and miR-107 target p53 inhibitors (such as MDM2), cell cycle genes (CDK4/6, E2F, CCNE1/2, CDC7), oncogenes (MET), growth factors (IGF1), antiapoptotic genes (BCL2), metabolic genes (LDHA), stemness genes (NANOG, SOX2, KLF4, OCT4, CD44), proangiogenic genes (ANRT) [27– 29] or genes involved in proliferation and metastasis (LAMC1). [score:5]
miR-449b and miR34 which block the p53 repressors SIRT1 and HDAC1 were down-regulated. [score:4]
MicroRNA-34b and MicroRNA-34c are targets of p53 and cooperate in control of cell proliferation and adhesion-independent growth. [score:2]
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58
[+] score: 25
Other miRNAs from this paper: hsa-let-7b, hsa-mir-15a, hsa-mir-19a, hsa-mir-19b-1, hsa-mir-19b-2, hsa-mir-27a, hsa-mir-28, hsa-mir-30a, hsa-mir-100, hsa-mir-30c-2, hsa-mir-30d, hsa-mir-181a-2, hsa-mir-210, hsa-mir-181a-1, hsa-mir-221, hsa-mir-1-2, hsa-mir-15b, hsa-mir-30b, hsa-mir-122, hsa-mir-132, hsa-mir-141, hsa-mir-191, hsa-mir-9-1, hsa-mir-9-2, hsa-mir-9-3, hsa-mir-125a, hsa-mir-195, hsa-mir-200c, hsa-mir-1-1, hsa-mir-30c-1, hsa-mir-34b, hsa-mir-30e, hsa-mir-371a, hsa-mir-372, hsa-mir-373, hsa-mir-375, hsa-mir-151a, hsa-mir-429, hsa-mir-449a, hsa-mir-483, hsa-mir-193b, hsa-mir-520e, hsa-mir-520f, hsa-mir-520a, hsa-mir-520b, hsa-mir-520c, hsa-mir-520d, hsa-mir-520g, hsa-mir-520h, hsa-mir-548a-1, hsa-mir-548b, hsa-mir-548a-2, hsa-mir-548a-3, hsa-mir-548c, hsa-mir-548d-1, hsa-mir-548d-2, hsa-mir-449b, hsa-mir-151b, hsa-mir-320b-1, hsa-mir-320b-2, hsa-mir-891a, hsa-mir-935, hsa-mir-1233-1, hsa-mir-548e, hsa-mir-548j, hsa-mir-548k, hsa-mir-548l, hsa-mir-548f-1, hsa-mir-548f-2, hsa-mir-548f-3, hsa-mir-548f-4, hsa-mir-548f-5, hsa-mir-548g, hsa-mir-548n, hsa-mir-548m, hsa-mir-548o, hsa-mir-548h-1, hsa-mir-548h-2, hsa-mir-548h-3, hsa-mir-548h-4, hsa-mir-1275, hsa-mir-548p, hsa-mir-548i-1, hsa-mir-548i-2, hsa-mir-548i-3, hsa-mir-548i-4, hsa-mir-1973, hsa-mir-548q, hsa-mir-548s, hsa-mir-548t, hsa-mir-548u, hsa-mir-548v, hsa-mir-548w, hsa-mir-548x, hsa-mir-1233-2, hsa-mir-548y, hsa-mir-548z, hsa-mir-548aa-1, hsa-mir-548aa-2, hsa-mir-548o-2, hsa-mir-548h-5, hsa-mir-548ab, hsa-mir-548ac, hsa-mir-548ad, hsa-mir-548ae-1, hsa-mir-548ae-2, hsa-mir-548ag-1, hsa-mir-548ag-2, hsa-mir-548ah, hsa-mir-548ai, hsa-mir-548aj-1, hsa-mir-548aj-2, hsa-mir-548x-2, hsa-mir-548ak, hsa-mir-548al, hsa-mir-548am, hsa-mir-548an, hsa-mir-371b, hsa-mir-548ao, hsa-mir-548ap, hsa-mir-548aq, hsa-mir-548ar, hsa-mir-548as, hsa-mir-548at, hsa-mir-548au, hsa-mir-548av, hsa-mir-548aw, hsa-mir-548ax, hsa-mir-548ay, hsa-mir-548az, hsa-mir-548ba, hsa-mir-548bb, hsa-mir-548bc
When focusing on the miRNA differential expression patterns in NOA, three studies pinpointed four miRNAs that are involved in spermatogenesis [36, 52, 55]: hsa-miR-34b*, hsa-miR-34c-5p and hsa-miR-122, which are downregulated [52, 55], and hsa-miR-429, which is upregulated [36, 52]. [score:9]
Moreover, it has been reported that in severe oligozoospermia, hsa-miR-34c-3p downregulates Phosphatidylinositol-Specific Phospholipase C, X Domain containing 3 (PLCXD3). [score:4]
Then, Abu-Halima et al. showed that hsa-miR-34b*, hsa-miR-34b, hsa-miR-34c-5p, hsa-miR-449a and hsa-miR-449b, which are structurally similar, are downregulated in testes of infertile men with different histopathologic patterns (the most common feature of male-factor infertility). [score:4]
Specifically, the percentage of good quality embryos at day 3 was higher in the group with higher hsa-miR-34c expression in spermatozoa. [score:3]
Among them, hsa-miR-15a, hsa-miR-15b, hsa-miR-30, hsa-miR-34b, hsa-miR-34c-5p and hsa-miR-193b-5p have a direct role in spermatogenesis [51, 53]. [score:2]
The most abundant miRNA in human sperm is hsa-miR-34c [63] and it is involved in spermatogenesis. [score:1]
Recently, Cui et al. showed that hsa-miR-34c levels in spermatozoa are correlated with ICSI outcomes [64]. [score:1]
This finding suggests that hsa-miR-34c could become a predictive biomarker to identify defective spermatozoa. [score:1]
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59
[+] score: 24
In turn, miR34 also acts as a tumor suppressor gene, via inhibition of bcl2, notch1/2 and c-met, all central pro-survival and pro-proliferative molecules [70, 71]. [score:5]
Re -expression of miR34 in this population triggers a loss of the CSC population, as demonstrated by a marked reduction in the CD44+/CD133+ subset, as well as inhibition of spherule formation and in vivo tumor growth. [score:5]
Studies examining the effects of tumor suppressor genes on miRNAs have demonstrated that p53 activates the transcription of the miR34 family of miRNAs. [score:3]
Bommer G. T. Gerin I. Feng Y. Kaczorowski A. J. Kuick R. Love R. E. Zhai Y. Giordano T. J. Qin Z. S. Moore B. B. p53 -mediated activation of miRNA34 candidate tumor-suppressor genesCurr. [score:3]
Importantly, overexpression of miR34 in these cells restored their chemo- and radiotherapy sensitivity. [score:3]
These observations suggest that miR34 is an important negative regulator of cancer stem cell renewal and may hold promise as a form of molecular therapy in pancreatic cancer [72]. [score:2]
Using an adenoviral- delivery system, these authors demonstrate that miR34 can drive neural differentiation of tumor neurospheres and impair tumor growth in a mouse mo del of medulloblastoma. [score:1]
CD44+/CD133+ cancer stem cell populations in p53-mutated pancreatic cancer cell lines contain low levels of miR34 and high levels of bcl2 and notch1/2. [score:1]
miR34 in p53-Mutated Pancreatic Cancer Stem Cells. [score:1]
[1 to 20 of 9 sentences]
60
[+] score: 24
Recent data have shown that genes coding for the miR-34 family are direct transactivational targets of p53 and their over -expression results in the induction of apoptosis, cell cycle arrest and senescence [13- 16] (Fig. 1). [score:6]
Taken together, these data indicate that p53 directly activates the expression of miR-34 genes, which play an important role in p53 -mediated apoptotic pathway. [score:4]
Prominent among them is the finding that miR-34 family is a direct transactivational target of p53 and it induces apoptosis, cell cycle arrest and senescence [12- 15]. [score:4]
Bommer et al. [17] showed that miR-34 family members may be tissue specific with miR-34a being expressed at higher level than miR-34b/c with the exception of the lungs. [score:3]
A direct molecular explanation of how miR-34 interferes in the p53 pathway and apoptosis is highlighted in the result published by Lowenstein and colleagues [18]. [score:2]
In mammals, the miR-34 family members are made up of three miRNAs encoded by two different genes: miR-34b/c (who share a common primary transcript) and miR-34a (encoded on its own. [score:1]
Paradoxically, miR-29 acted in a fashion different from miR-34 since it is an apoptotic inducer only in the presence of wild type p53 gene. [score:1]
The miR-34 story set the precedence and a number of papers have now been published showing that other miRNAs interfere the p53 pathway, in a p53-independent manner (miR-34), partial p53 -dependent manner (miR-192, miR-194, miR-215, and miR-21) or a p53 -dependent manner (miR-29). [score:1]
Furthermore, miRNAs is implicated in every aspect of cellular outcome of p53 activation: apoptosis (miR-34 and miR-29), cell cycle arrest (miR-192, miR-194, and miR-215), and senescence (miR-34). [score:1]
miR-34 Family and Apoptosis. [score:1]
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61
[+] score: 24
One of the differentially expressed miRNAs, miR-34c, was previously identified to be highly expressed in the hippocampus of patients with AD and in animal mo dels of AD [50]. [score:5]
When miR-34c is targeted for removal, learning and memory is restored. [score:3]
Interestingly, our data examining miRNAs differentially expressed in the progression of Lewy bodies from limbic to neocortical, also identified miR-34c and 34b as significantly altered. [score:3]
One of the mRNA targets for miR-34c is SIRT1, involved in synaptic plasticity and memory formation [64]. [score:3]
The same group linked miR-34c as a negative regulator of memory consolidation [50]. [score:2]
Interestingly, SIRT1 can also be regulated by miR-34c (below). [score:2]
miR-34c was found in our study to be upregulated in PDD patients compared with PD patients and in AD patients compared to control subjects. [score:2]
Zovoilis et al. [50] found high levels of miR-34c in hippocampus of AD patients and in animal mo dels of AD. [score:1]
They observed that, when miR-34c is elevated, memory consolidation is impaired. [score:1]
The authors confirmed that elevated miR-34c correlated with decrease in SIRT1 in tissue samples. [score:1]
The hypothesis that elevated levels of miR-34c is related to cognitive decline holds true in our data from patient serum samples. [score:1]
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62
[+] score: 24
Upregulation of miR-34c significantly inhibited migration and invasion of HEC1-B cells via reduction of E2F3 protein [36]. [score:6]
According to a recent report, it was shown that overexpression of miR-34c significantly inhibited proliferation of endometrial carcinoma cells by reducing E2F3 [36]. [score:5]
In endometrial carcinoma (EC), miR-34c was significantly reduced and E2F3 was reduced after up-regulation of miR-34c in the HEC-1-B cell, suggesting that miR-34c functions via reduction of E2F3 protein [36]. [score:4]
Li F et al. suggested that miR-34c acted as a tumor suppressor via induction of apoptosis via E2F3 [36]. [score:3]
Meanwhile, miR-449a/b and miR-34 was reported to induce inhibition of E2F1 and E2F3 in a negative feedback loopy, respectively [80]. [score:3]
The miR-34 family consists of miR-34a, 34b and 34c, which are dysregulated in various human cancers. [score:2]
Also, transfection of miR-34c could be found to decrease E2F3 protein levels in head and neck cancer cells [72]. [score:1]
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63
[+] score: 23
Moreover, restoring expression of miR-15a/16-1 indirectly affects expression of miR-34 family by modulating p53 expression and downregulation of miR-29 and miR-181b in aggressive CLL contributes to overexpression of Tcl1 [43]. [score:13]
Besides, miR-15a/16-1 target TP53 while miR-34 targets ZAP-70 mRNA expression [32]. [score:7]
MiR-34 family members are involved in a fine-regulated feedback circuitry with p53 and miR-15a/16-1 in 13q deleted CLL, suggesting bidirectional interplay between microRNAs and genes. [score:3]
[1 to 20 of 3 sentences]
64
[+] score: 23
For miR-277–3p, miR-987–5p and miR-34–5p their cognate miRNA precursors also increase upon Dis3 knockdown, suggesting that Dis3 normally affects their expression via indirect means, perhaps by increasing the expression of specific transcription factors. [score:7]
[37] Wing phenotypes are not always seen when miRNAs are overexpressed in the wing; for example, no phenotype is seen when miR-34 is overexpressed using MS1096-GAL4. [score:5]
In contrast, the levels of the pri/pre-miRNAs are increased in levels in Dis3 -depleted cells for miR-277–3p, miR-34–5p, miR-317–5p and miR-987–5p suggesting that these miRNAs are not direct targets of Dis3. [score:4]
Using this stringent filtering method, we identified 6 miRNAs whose expression increased ≥2-fold in the knockdown samples compared to both parental controls (miR-277–3p, miR-987–5p, miR-252–5p, miR-34–5p, and miR-7–3p and miR-317–5p). [score:3]
The set of miRNAs that increase ≥2-fold; by RNA-seq upon Dis3 depletion are miR-277–3p, miR-987–5p, miR-252–5p, miR-34–5p and miR-317–5p (Table 2). [score:1]
* miR-317 is located in close proximity to miR-34 and miR-277. [score:1]
The microRNA miR-34 modulates ageing and neurodegeneration in Drosophila. [score:1]
A similar effect is seen for miR-34–5p although the changes in levels are not so pronounced. [score:1]
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65
[+] score: 23
An Exiqon microRNA assay was used to analyze the expression of these miRNAs after treatment with 0, 10, 20, 40 μmol/L PQ for 24 h. Using independent procedures, we could confirm the microarray data (Figure 6), that is, a clear concentration -dependent induction of hNPCs differentiation-related miRNA (hsa-mir-10a, hsa-let-7f and hsa-mir-34c) and a significant down-regulation of the expression of hNPCs proliferation-maintained miRNA (hsa-mir-124) in comparison to the solvent control. [score:7]
Known tumor-suppressor hsa-mir-34c, which was recently found to be involved in brain development and spermatogenesis [14], was significantly upregulated in hNPCs treated with PQ. [score:7]
In addition, it is reported that mir-34c is a target of p53 and plays a role in control of cell proliferation and adhesion-independent growth [14]. [score:3]
Corney D. C. Flesken-Nikitin A. Godwin A. K. Wang W. Nikitin A. Y. MicroRNA-34b and MicroRNA-34c are targets of p53 and cooperate in control of cell proliferation and adhesion-independent growth Cancer Res. [score:2]
As mentioned above, miRNAs such as hsa-mir-124, hsa-mir-10a, hsa-mir-34c, hsa-mir-9, hsa-mir-34c and hsa-let-7 family were previously proved to be involved in embryonic/adult neurogenesis during brain development [14]. [score:2]
The results from our study suggest that PQ affects neural proliferation and differentiation processes are specific for mir-124, mir-10a, let-7d/f/g and mir-34c but not mir-9. The quantification of neural proliferation/differentiation related genes using real time RT-PCR further confirmed the impacts of PQ on these miRNAs. [score:1]
The results from our study suggest that PQ affects neural proliferation and differentiation processes are specific for mir-124, mir-10a, let-7d/f/g and mir-34c but not mir-9. The quantification of neural proliferation/differentiation related genes using real time RT-PCR further confirmed the impacts of PQ on these miRNAs. [score:1]
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66
[+] score: 22
In contrast, miR-34c-5p had a different pattern of expression in human tissues, suggesting that the two transcripts may be independently regulated (Fig 5C). [score:4]
MiR-34b and miR-34c are transcribed from the same locus on chromosome 11 (cytogenetic band 11q23.1; Fig 5A), and their expression is regulated by epigenetics, such as CpG island methylation [29], and p53 function [32]. [score:4]
In this analysis, only four miRNAs were differentially expressed in both human prostate cancer cell lines and tumor samples from TRAMP mice, including miR-34b-3p, miR-34c-5p, miR-138, and miR-224 (Fig 2C and 2D). [score:3]
B, C) miR-34b-3p and miR-34c-5p expression in the indicated samples of normal prostate (pool of 10 specimens), benign hyperplasia (BPH), prostate cancer (PCa) or non-neoplastic (RWPE-1), hyperplastic (BPH-1) or tumor (LNCaP, DU145) prostate cell lines. [score:3]
The miR-34 family of miRNAs has been previously reported to suppress tumorigenesis by different mechanisms, including modulation of cell cycle transitions, EMT, metastasis, or cancer stemness [33]. [score:3]
Expression analysis followed by unsupervised hierarchical clustering D) of miR-224, -34b-3p, -138 and miR-34c-5p reveals that androgen-independent prostate cancer cells are more similar to TRAMP tissues than to androgen -dependent or non-tumorigenic prostate human cells. [score:3]
MicroRNA-34b and MicroRNA-34c are targets of p53 and cooperate in control of cell proliferation and adhesion-independent growth. [score:2]
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67
[+] score: 21
In a previous study we quantified the expression levels of 742 miRNAs in 30 cancer and 20 normal endometrium samples, among these, the commonly down-regulated miRNA mir-34c showed no significant down-regulation in the EAC samples, and mir-101 were up-regulated in the EAC samples compared to normal [11]. [score:11]
The miRNA mir-34c is a known tumor suppressor in other cancer forms, however it is not confirmed in EAC, mir-34c was shown to be low expressed in endometrial cancer cell lines [29], however no studies confirm this in primary tumors, including our material [11]. [score:5]
The Ishikawa and HEK293 cell lines were transfected with mir-34a inhibitor, mir-34 mimic and their respective negative controls by using the Lipofectamine® RNAiMAX Transfection Reagent (Life Technologies) in antibiotic-free Opti-MEM medium (Life Technologies) according to the manufacturer’s protocol at a final concentration of 100 μM. [score:3]
MiRNA-34a belongs to the family miRNA-34, which also includes mir-34b and mir-34c. [score:1]
Mir-34b and mir-34c are transcribed from a common miRNA gene located on HSA11, while mir-34a is located on HSA1. [score:1]
[1 to 20 of 5 sentences]
68
[+] score: 20
miRNAs Deregulated in OS Expression Levels Compared to the Controls Overall Function miR-382 Down-regulated Poor survival outcome and metastasis marker[70, 83] miR-154 Down-regulated Poor survival outcome[70] miR-33a Up-regulated Chemoresistance[84] miR-34c Down-regulated Chemoresistance[85] Most forms of human cancer have changes in the epigenome compared to the normal cellular counterparts from which they are derived. [score:14]
One of the key miRNAs in the TP53 pathway is miR-34, which is significantly downregulated in many osteosarcoma tumors that affects the cell cycle and proliferation [77, 78, 79]. [score:4]
Xu M. Jin H. Xu C. X. Bi W. Z. Wang Y. MiR-34c inhibits osteosarcoma metastasis and chemoresistance Med. [score:2]
[1 to 20 of 3 sentences]
69
[+] score: 20
RES increased the expression of miR-34 which resulted in the decreased expression of E2F3 and SIRT1 [68]. [score:5]
The expression of miR-34c is increased by RES in CRC and this expression was in part responsible for the effects of RES. [score:5]
miR-34a and miR-34c were determined to be down-regulated in CRC patient samples compared to normal colonic mucosa. [score:3]
Histone modifying enzymes have been shown to be regulated by CUR and miR-34. [score:2]
Ingenuity Pathway Analysis revealed that CUR and miR-34 could be regulators of the histone-modifying genes in ALL [157]. [score:2]
CDF can induce miR-34 which is normally silenced in CRC. [score:1]
In this xenograft system, RES increased miR-34 levels and also reduced the level of IL-6 which normally promotes CRC progression. [score:1]
miRs such as: miR-21, miR-22, miR-34, miR-101, miR-146a miR-200 and let-7 have been associated with the CSC phenotype. [score:1]
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70
[+] score: 20
Considering that the aberrant overexpression of RUNX2 correlates to resistance to chemotherapy [166], it is indicative that miR-34c contributes to the improvement of chemo-sensitivity of drug-resistant osteosarcoma cells through the down-regulation of RUNX2. [score:6]
van der Deen et al. described that p53 -mediated stimulation of miR-34c expression causes a massive decrease in RUNX2 and reduces the metastatic potential of osteosarcoma cells [165]. [score:3]
Ji et al. reported that miR-34 family members have tumor-suppressive function downstream of p53, and their restoration renders p53-mutated pancreatic cancer cells 2-3-fold more sensitive to GEM [159]. [score:3]
It has been shown that the additional miRNAs (miR-23a, miR-34c, miR-133a, miR-135a, miR-205 and miR-217) also attenuate osteogenesis by targeting RUNX2 [162, 163]. [score:3]
Ji Q Hao X Zhang M Tang W Yang M Li L Xiang D Desano JT Bommer GT Fan D Fearon ER Lawrence TS Xu L MicroRNA miR-34 inhibits human pancreatic cancer tumor-initiating cellsPLoS One. [score:3]
CCR-10-2619 21159887 165. van der Deen M Taipaleenmäki H Zhang Y Teplyuk NM Gupta A Cinghu S Shogren K Maran A Yaszemski MJ Ling L Cool SM Leong DT Dierkes C Zustin J Salto-Tellez M Ito Y Bae SC Zielenska M Squire JA Lian JB Stein JL Zambetti GP Jones SN Galindo M Hesse E Stein GS van Wijnen AJ MicroRNA-34c inversely couples the biological functions of the runt-related transcription factor RUNX2 and the tumor suppressor p53 in osteosarcomaJ Biol Chem. [score:2]
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71
[+] score: 20
Other miRNAs from this paper: hsa-let-7a-1, hsa-let-7a-2, hsa-let-7a-3, hsa-let-7b, hsa-let-7c, hsa-let-7d, hsa-let-7e, hsa-let-7f-1, hsa-let-7f-2, hsa-mir-16-1, hsa-mir-17, hsa-mir-18a, hsa-mir-19a, hsa-mir-21, hsa-mir-22, hsa-mir-24-1, hsa-mir-24-2, hsa-mir-25, hsa-mir-26a-1, hsa-mir-26b, hsa-mir-27a, hsa-mir-30a, hsa-mir-31, hsa-mir-98, hsa-mir-99a, hsa-mir-101-1, hsa-mir-16-2, hsa-mir-192, hsa-mir-197, hsa-mir-199a-1, hsa-mir-208a, hsa-mir-30c-2, hsa-mir-30d, hsa-mir-10a, hsa-mir-10b, hsa-mir-34a, hsa-mir-187, hsa-mir-199a-2, hsa-mir-199b, hsa-mir-203a, hsa-mir-211, hsa-mir-219a-1, hsa-mir-221, hsa-mir-222, hsa-mir-223, hsa-mir-224, hsa-mir-200b, hsa-let-7g, hsa-let-7i, hsa-mir-27b, hsa-mir-30b, hsa-mir-122, hsa-mir-124-1, hsa-mir-124-2, hsa-mir-124-3, hsa-mir-125b-1, hsa-mir-128-1, hsa-mir-132, hsa-mir-133a-1, hsa-mir-133a-2, hsa-mir-138-2, hsa-mir-140, hsa-mir-142, hsa-mir-143, hsa-mir-144, hsa-mir-145, hsa-mir-191, hsa-mir-9-1, hsa-mir-9-2, hsa-mir-9-3, hsa-mir-125a, hsa-mir-125b-2, hsa-mir-126, hsa-mir-138-1, hsa-mir-146a, hsa-mir-200c, hsa-mir-155, hsa-mir-128-2, hsa-mir-29c, hsa-mir-30c-1, hsa-mir-200a, hsa-mir-101-2, hsa-mir-219a-2, hsa-mir-34b, hsa-mir-99b, hsa-mir-30e, hsa-mir-26a-2, hsa-mir-375, hsa-mir-328, hsa-mir-337, hsa-mir-338, hsa-mir-339, hsa-mir-384, hsa-mir-424, hsa-mir-429, hsa-mir-449a, hsa-mir-485, hsa-mir-146b, hsa-mir-494, hsa-mir-497, hsa-mir-498, hsa-mir-520a, hsa-mir-518f, hsa-mir-499a, hsa-mir-509-1, hsa-mir-574, hsa-mir-582, hsa-mir-606, hsa-mir-629, hsa-mir-449b, hsa-mir-449c, hsa-mir-509-2, hsa-mir-874, hsa-mir-744, hsa-mir-208b, hsa-mir-509-3, hsa-mir-1246, hsa-mir-1248, hsa-mir-219b, hsa-mir-203b, hsa-mir-499b
Targets of the most remarkably down-regulated miRNAs (let-7, miR-10, miR-26, miR-30, miR-34, miR-99, miR-122, miR-123, miR-124, miR-125, miR-140, miR-145, miR-146, miR-191, miR-192, miR-219, miR-222, and miR-223) regulate proliferation, gene expression, stress response, apoptosis, and angiogenesis. [score:9]
Microarray analysis of nasal mucosa identified several miRNAs with altered expression in acute RSV -positive infants (down-regulated miR-34b, miR-34c, miR-125b, miR-29c, mir125a, miR-429 and miR-27b and up-regulated miR-155, miR-31, miR-203a, miR-16 and let-7d) as compared to healthy infants [86]. [score:8]
Interestingly, the members of the miR-34/449 family were highly repressed in asthma; in vivo experiments have shown that their expression levels are also decreased after IL-13 treatment. [score:3]
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72
[+] score: 20
Other miRNAs from this paper: hsa-let-7a-1, hsa-let-7a-2, hsa-let-7a-3, hsa-let-7b, hsa-let-7c, hsa-let-7d, hsa-let-7e, hsa-let-7f-1, hsa-let-7f-2, hsa-mir-15a, hsa-mir-16-1, hsa-mir-17, hsa-mir-18a, hsa-mir-19a, hsa-mir-19b-1, hsa-mir-19b-2, hsa-mir-20a, hsa-mir-21, hsa-mir-25, hsa-mir-26a-1, hsa-mir-26b, hsa-mir-27a, hsa-mir-29a, hsa-mir-30a, hsa-mir-31, hsa-mir-92a-1, hsa-mir-93, hsa-mir-98, hsa-mir-101-1, hsa-mir-29b-1, hsa-mir-29b-2, hsa-mir-106a, hsa-mir-16-2, hsa-mir-192, hsa-mir-196a-1, hsa-mir-197, hsa-mir-7-1, hsa-mir-7-2, hsa-mir-7-3, hsa-mir-10b, hsa-mir-34a, hsa-mir-181a-2, hsa-mir-182, hsa-mir-183, hsa-mir-196a-2, hsa-mir-205, hsa-mir-181a-1, hsa-mir-221, hsa-mir-222, hsa-mir-200b, hsa-let-7g, hsa-let-7i, hsa-mir-1-2, hsa-mir-15b, hsa-mir-122, hsa-mir-124-1, hsa-mir-124-2, hsa-mir-124-3, hsa-mir-125b-1, hsa-mir-128-1, hsa-mir-133a-1, hsa-mir-133a-2, hsa-mir-135a-1, hsa-mir-135a-2, hsa-mir-137, hsa-mir-140, hsa-mir-141, hsa-mir-143, hsa-mir-145, hsa-mir-152, hsa-mir-191, hsa-mir-9-1, hsa-mir-9-2, hsa-mir-9-3, hsa-mir-125a, hsa-mir-125b-2, hsa-mir-126, hsa-mir-146a, hsa-mir-150, hsa-mir-194-1, hsa-mir-206, hsa-mir-200c, hsa-mir-1-1, hsa-mir-155, hsa-mir-128-2, hsa-mir-194-2, hsa-mir-106b, hsa-mir-29c, hsa-mir-200a, hsa-mir-101-2, hsa-mir-34b, hsa-mir-301a, hsa-mir-26a-2, hsa-mir-372, hsa-mir-374a, hsa-mir-375, hsa-mir-328, hsa-mir-133b, hsa-mir-20b, hsa-mir-429, hsa-mir-449a, hsa-mir-486-1, hsa-mir-146b, hsa-mir-494, hsa-mir-503, hsa-mir-574, hsa-mir-628, hsa-mir-630, hsa-mir-449b, hsa-mir-449c, hsa-mir-708, hsa-mir-301b, hsa-mir-1827, hsa-mir-486-2
miR-34 and miR-449 are also involved in the inhibition of NSCLC cell migration and invasion by suppression of AXL and SNAIL-1, respectively a tyrosine-kinase receptor and a zinc-finger protein involved in cellular migration, proliferation and cancer cell epithelial-mesenchymal transition [123- 124]. [score:5]
Interestingly, Trang et al. used synthetic tumor suppressors miR-34 and let-7 mimics complexed with a novel neutral lipid emulsion to target a KRAS-activated mouse mo del of NSCLC. [score:5]
The miR-34 family consists of tumor-suppressive miRNAs (miR-34a, miR-34b, miR-34c) reported to be under p53 regulation and involved in controlling apoptosis and G1 cell cycle arrest. [score:4]
Landi et al. identified a five-miRNA signature (miR-25, miR-34c-5p, miR-191, let-7e, miR-34a) that strongly differentiated squamous cell carcinoma from adenocarcinoma (global P< 0.0001), and reported that the lower expression level of this signature correlated with the poor overall survival rates of squamous cell carcinoma patients [168]. [score:3]
MiR-34 reduced expression has been found in various cancers, including NSCLC [120- 121]. [score:2]
The miR-449 cluster (miR-449a, miR-449b, miR-449c) belongs to the same family as miR-34 (p53- responsive microRNAs) [122]. [score:1]
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73
[+] score: 19
Other miRNAs from this paper: hsa-mir-34a, hsa-mir-34b
The miR-34 family has been identified as a p53 target and plays a key role as regulator of tumor suppression in many cancers controlling cell cycle arrest and apoptosis [20]– [22]. [score:6]
In p53-mutant pancreatic cancer cells, restoration of miR-34 expression significantly inhibited cell growth inducing apoptosis and cell cycle arrest [24]. [score:5]
In U2-OS and U2-OS175 cells, miR-34a promoter was unmethylated in both gene alleles, while MG63 and Saos-2 showed CpG methylation of the two alleles in accordance with very low expression levels and lack of miR-34 induction after etoposide exposure. [score:3]
This suggested that recruitment of p53 by miR-34 was not impaired by expression of dominant negative p53. [score:3]
Previous studies wi dely validated the action of p53 on the target miR-34a using a primer for pri-miR and for pre-miR-34 as well as for mature miR-34 [13], [14]. [score:2]
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74
[+] score: 19
Other miRNAs from this paper: hsa-let-7a-1, hsa-let-7a-2, hsa-let-7a-3, hsa-let-7b, hsa-let-7c, hsa-let-7d, hsa-let-7e, hsa-let-7f-1, hsa-let-7f-2, hsa-mir-15a, hsa-mir-16-1, hsa-mir-17, hsa-mir-18a, hsa-mir-19a, hsa-mir-19b-1, hsa-mir-19b-2, hsa-mir-20a, hsa-mir-21, hsa-mir-24-1, hsa-mir-24-2, hsa-mir-92a-1, hsa-mir-92a-2, hsa-mir-106a, hsa-mir-16-2, hsa-mir-181a-2, hsa-mir-181b-1, hsa-mir-181a-1, hsa-mir-221, hsa-mir-222, hsa-mir-223, hsa-let-7g, hsa-let-7i, hsa-mir-15b, hsa-mir-23b, hsa-mir-27b, hsa-mir-122, hsa-mir-125b-1, hsa-mir-140, hsa-mir-125b-2, hsa-mir-136, hsa-mir-146a, hsa-mir-150, hsa-mir-206, hsa-mir-155, hsa-mir-181b-2, hsa-mir-106b, hsa-mir-302a, hsa-mir-34b, hsa-mir-302b, hsa-mir-302c, hsa-mir-302d, hsa-mir-367, gga-let-7i, gga-let-7a-3, gga-let-7b, gga-let-7c, gga-mir-125b-2, gga-mir-155, gga-mir-222a, gga-mir-221, gga-mir-92-1, gga-mir-19b, gga-mir-20a, gga-mir-19a, gga-mir-18a, gga-mir-17, gga-mir-16-1, gga-mir-15a, gga-mir-1a-2, gga-mir-206, gga-mir-223, gga-mir-106, gga-mir-302a, gga-mir-181a-1, gga-mir-181b-1, gga-mir-16-2, gga-mir-15b, gga-mir-140, gga-let-7g, gga-let-7d, gga-let-7f, gga-let-7a-1, gga-mir-146a, gga-mir-181b-2, gga-mir-181a-2, gga-mir-1a-1, gga-mir-1b, gga-let-7a-2, gga-mir-34b, gga-mir-34c, gga-let-7j, gga-let-7k, gga-mir-23b, gga-mir-27b, gga-mir-24, gga-mir-122-1, gga-mir-122-2, hsa-mir-429, hsa-mir-449a, hsa-mir-146b, hsa-mir-507, hsa-mir-455, hsa-mir-92b, hsa-mir-449b, gga-mir-146b, gga-mir-302b, gga-mir-302c, gga-mir-302d, gga-mir-455, gga-mir-367, gga-mir-429, gga-mir-449a, hsa-mir-449c, gga-mir-21, gga-mir-1458, gga-mir-1576, gga-mir-1612, gga-mir-1636, gga-mir-449c, gga-mir-1711, gga-mir-1729, gga-mir-1798, gga-mir-122b, gga-mir-1811, gga-mir-146c, gga-mir-15c, gga-mir-449b, gga-mir-222b, gga-mir-92-2, gga-mir-125b-1, gga-mir-449d, gga-let-7l-1, gga-let-7l-2, gga-mir-122b-1, gga-mir-122b-2
MiR-1a, miR-140 and miR-449 were significantly up-regulated in both tissues, while miR-455, miR-34b and miR-34c were only up-regulated with AIV infection in tracheae. [score:7]
Cluster mir-34b-mir-34c was up-regulated in tracheae. [score:4]
And the mir-34b-mir-34c cluster was the only significantly up-regulated cluster in the AIV infected trachea. [score:4]
MiR-34b and miR34c, whose target genes are B-cell CLL-pymphoma 2 & 11, might be involved in the B-cell differentiation. [score:3]
We hypothesize that miR-34b, miR-34c, miR-206, miR-1458 and miR-1612 might be some of the most important miRNAs associated with AIV infection. [score:1]
[1 to 20 of 5 sentences]
75
[+] score: 19
This pattern included the two created target sites for ssc-miR-34a and ssc-miR-34c, both predicted by TargetScan and PACMIT in SLA-1 (Figure 3 A); the disrupted target site for ssc-miR-148a in HSPA1A predicted by PACMIT and TargetSpy (Figure 3 B); the ssc-miR-133b (TargetScan and PACMIT), ssc-miR-133a-3p (TargetScan) and ssc-miR-323 (TargetSpy) created target sites in RNF5 (Figure 3 C); and the disrupted site for ssc-miR-2320 predicted by TargetSpy in SLA-1 (Figure 3D). [score:19]
[1 to 20 of 1 sentences]
76
[+] score: 19
For example, the expression of tumor suppressor genes, like PTEN [26] and P53 [27], which are the targets of miR-21 and miR-34c, were significantly reduced in the premalignant foci and HCC nodules [28]. [score:7]
Similarly, the expression of oncogenic miRNAs like miR-21, miR-10b, let-7i, miR-34c, were increased more than 2 fold in EpCAM [+] liver cancer cells; whereas miR-125b, miR-200a, miR-148b were most down-regulated. [score:6]
miR-21, miR-10b and miR-34c have been reported to be upregulated in various types of cancers, including HCC [25]. [score:4]
Our previous report also revealed that the relative expression levels of miR-92b, miR-21, miR-34c, miR-10b, and let-7i in EpCAM [+] liver cancer cells compared to fetal liver cells were increased (P<0.05). [score:2]
[1 to 20 of 4 sentences]
77
[+] score: 19
miR-34-5p (B), miR-410-3p (C), miR-449-5p (D) and miR-203 (E) expression, determined by Real-time PCR, was down-regulated in HPCx tumor tissues from gemcitabine -treated mice (p < 0.05). [score:6]
Thus, we identified potential miRNAs related to gemcitabine resistance in a human pancreatic cancer xenograft (HPCx) with miRNA microarray analysis and showed that miR-34-5p, miR-410-3p, miR-449-5p and miR-203 were significantly down-regulated in HPCx tumor tissues from gemcitabine -treated mice. [score:4]
Real-time PCR confirmed that miR-34-5p (Figure 1B), miR-410-3p (Figure 1C), miR-449-5p (Figure 1D) and miR-203 (Figure 1E) were down-regulated in HPCx tumor tissues from gemcitabine -treated mice (P < 0.05). [score:4]
Real-time PCR was used to detect the expression levels of miR-34-5p, miR-410-3p, miR-449-5p, miR-203, HMGB1, ARFIP1, GRIA2, CPEB4, NDFIP2, KLF6, PARG, OTX2, TMEFF2, TRPC1 and KLHL5. [score:3]
MiR-15a [11], miR-21 [12, 13], miR-34 [14], members of the miR-200 family [12, 15], miR-214 [11], miR-221 [16], members of the let7 family [15], and miR-320c [17] have been reported to play roles in gemcitabine chemoresistance in pancreatic cancer. [score:1]
In contrast, the chemoresistance to gemcitabine was merely slightly repressed in human PDAC cells treated with miR-34-5p or miR-203 mimics (Supplementary Figure 2). [score:1]
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78
[+] score: 18
The tumor-suppressive function of QKI-5 is described in more detail in Section 4.3. miR-34 family is highly conserved in the evolutionary context. [score:3]
Corney D. C. Flesken-Nikitin A. Godwin A. K. Wang W. Nikitin A. Y. MicroRNA-34b and microRNA-34c are targets of p53 and cooperate in control of cell proliferation and adhesion-independent growthCancer Res. [score:3]
The tumor-suppressive function of QKI-5 is described in more detail in Section 4.3. miR-34 family is highly conserved in the evolutionary context. [score:3]
Bommer G. T. Gerin I. Feng Y. Kaczorowski A. J. Kuick R. Love R. E. Zhai Y. Giordano T. J. Qin Z. S. Moore B. B. P53 -mediated activation of miRNA34 candidate tumor-suppressor genesCurr. [score:3]
Because of these broad anti-oncogenic function, miR-34 has been considered to be a novel therapeutic target, and MRX34, miR-34a mimics, becomes the first miRNA mimics to reach clinical trial for cancer therapy [131, 132]. [score:2]
Bader A. G. miR-34—A microRNA replacement therapy is headed to the clinicFront. [score:1]
In vertebrates, miR-34 family consists of three members: miR-34a, miR-34b and miR-34c, which are generated from two distinct genomic loci. [score:1]
miR-34a is encoded by its own transcript located within the chromosome 1 p 36, while miR-34b and miR-34c are generated by processing a bicistronic transcript from chromosome 11q23. [score:1]
Hermeking H. The miR-34 family in cancer and apoptosisCell Death Differ. [score:1]
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79
[+] score: 18
In this case, ectopic expression of miR-34 in colorectal cancer cells inhibited cell proliferation and sensitized cancer cells to 5-FU. [score:5]
Similar to the down-regulation of the miR-15/16-1 cluster, another miRNA frequently lost in cancer is represented by the miR-34 family. [score:4]
Recently, miR-34 family knock-out mice have been generated, and no obvious developmental or pathological abnormalities were observed at up to 12 months of age (30). [score:3]
However, miR-34 inhibition has been shown to enhance the apoptotic response to bortezomib in Myc-transformed B-cells, clearly demonstrating that miRNA involvement in chemotherapy is context -dependent (88). [score:3]
Like miR-21, miR-34 has also been shown to modulate cancer cell sensitivity to 5-FU (87). [score:1]
However, the authors showed that miR-34 -deficient mouse embryonic fibroblasts accelerate the reprogramming, not as expected through cell proliferation, but instead, at least partly, by post-transcriptional derepression of pluripotency genes, such as Sox2, N-Myc, and Nanog (30). [score:1]
The miR-34 family has been shown to be transcriptionally activated by p53, and it represents one of the main effectors of p53 -induced apoptosis, senescence, and cell cycle arrest. [score:1]
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80
[+] score: 18
Therefore, the p53 network suppressed tumor formation through a number of coordinated interactions and several transcriptional targets including the role played by miR-34 family members in inhibiting unregulated cell proliferation and tumor development [58, 59]. [score:9]
The authors compared microRNA expression profile of wild-type and p-53 -deficient cells and found that the expression of microRNA family members (miR-34a-c) reflected the p53 status and the genes encoding miR-34 family members were transcriptional targets of p53 in vivo and in vitro. [score:6]
The first microRNAs involved in the p-53 tumor suppressor network were reported in 2007 and they belong to the miR-34 family, these being miR-34a, miR-34b, and miR-34c [57]. [score:3]
[1 to 20 of 3 sentences]
81
[+] score: 17
mir-200a, mir-34, mir-195, and mir-381-3p are usually downregulated in presence of SIRT1 expression, and vice versa low expression of SIRT1 relates to miRNAs upregulation [9, 11, 17, 18, 26– 28]. [score:11]
Also, upregulation of mir-34c-5p and mir-381-3p that target Nampt, an enzyme involved in the production of NAD, the cofactor for SIRT1 [29], diminishes SIRT1 actions. [score:6]
[1 to 20 of 2 sentences]
82
[+] score: 17
Interestingly, several miRNAs differentially expressed in neurodegenerative diseases such as, miR-29 family, and miR-34 family are considered to be potential tumor suppressor miRNAs (Table 1). [score:6]
miRNA expression profiles revealed decreased expression of miR-34b and miR-34c in brain areas with variable neuropathological affectation at clinical stages of PD. [score:5]
miR-34b and miR-34c are also reported to be direct targets of p53 and silenced by aberrant CpG island methylation in colorectal cancer (Toyota et al., 2008). [score:4]
miR-34 b/34c. [score:1]
Depletion of miR-34b or miR-34c in differentiated SH-SY5Y dopaminergic neuronal cells resulted in a moderate reduction in cell viability that was accompanied by altered mitochondrial function and dynamics, oxidative stress and reduction in total cellular adenosine triphosphate content. [score:1]
[1 to 20 of 5 sentences]
83
[+] score: 17
Interestingly, several reports indicate that miR-34 family members are direct transcriptional targets of the tumor suppressor p53 and suggest that some cellular roles of p53, including those related to the regulation of cellular proliferation and apoptosis, could be mediated by these miRNAs [55, 56]. [score:7]
Corney D. C. Hwang C. I. Matoso A. Vogt M. Flesken-Nikitin A. Godwin A. K. Kamat A. A. Sood A. K. Ellenson L. H. Hermeking H. Frequent downregulation of mir-34 family in human ovarian cancers Clin. [score:4]
Decreased expression of miR-34 has been observed in lung, ovarian, CLL and colorectal cancer [50, 51, 52, 53]. [score:3]
The miR-34 family, which comprises miR-34a, b and c, has also received considerable attention for its potential role as tumor suppressor in several human malignancies. [score:3]
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84
[+] score: 17
miR-34 cooperates with p53 in suppression of prostate cancer by joint regulation of stem cell compartment. [score:4]
p53 is deleted in many cancer types and p53 promotes transcription of miR-34, hence its low expression. [score:3]
miR-34 also becomes an important tumor suppressor in many sarcoma types. [score:3]
A positive feedback between p53 and miR-34 miRNAs mediates tumor suppression. [score:3]
In fact, miR-34 replacement therapy has shown promise against cancer cell survival, stemness, metastasis, and chemoresistance in various cancer cell types and animal mo dels and is under Phase I clinical trial undertaken by MiRNA Therapeutics (Bader, 2012). [score:1]
miR-34–a microRNA replacement therapy is headed to the clinic. [score:1]
Another commonly decreased level miRNA found in various cancers is miR-34 (Cheng et al., 2014; Okada et al., 2014). [score:1]
For instance, miR-34 levels inversely correlate with poor patient survival outcomes indicating its potential role as a diagnostic marker in (Marino et al., 2014). [score:1]
[1 to 20 of 8 sentences]
85
[+] score: 17
Other miRNAs from this paper: hsa-mir-34a, hsa-mir-34b
The miR-34b and miR-34c were found frequently downregulated by aberrant methylation in MM, resulting in the loss of tumor-suppressive p53 function and the acquisition of a malignant phenotype [62]. [score:6]
The inhibition of these endogenous miRNAs by using antagomiRs dramatically increased Met expression and, conversely, transfection of miR-34b and miR-34c impaired Met signaling and the invasive growth program in cells of lung carcinoma and other cancers [63]. [score:5]
Tanaka N. Toyooka S. Soh J. Tsukuda K. Shien K. Furukawa M. Muraoka T. Maki Y. Ueno T. Yamamoto H. Downregulation of microRNA-34 induces cell proliferation and invasion of human mesothelial cells Oncol. [score:4]
This suggests a key role of the balance between miR-34 family members and Met in the early carcinogenic process of MM [64]. [score:1]
The importance of Met in the process of HM transformation to MM and acquisition of the invasive phenotype was also confirmed by the role played by two microRNA-34 (miRNA-34) family members. [score:1]
[1 to 20 of 5 sentences]
86
[+] score: 17
Other miRNAs from this paper: hsa-mir-139, hsa-mir-182, hsa-mir-486-1, hsa-mir-4423, hsa-mir-486-2
The hsa-miR-486-3p, hsa-miR-182-5p, and hsa-miR-139-5p showed dose -dependent upregulation in fold expression value, whereas hsa-miR-34c-5p and hsa-miR-4423-3p remained twofold upregulated with no significant deviation in fold expression at all three test concentration. [score:11]
Moreover we also found upregulation in 5 miRNAs (has-miR-486-3p, has-miR-34c-5p, has-miR-4423-3p, has-miR-182-5p, and has-miR-139-5p) which play role in muscle contraction, Arginine and proline metabolism and Hypertrophic cardiomyopathy (HCM). [score:4]
In addition, we also identified 5 out of 14 miRNAs; hsa-miR-486-3p, hsa-miR-34c-5p, hsa-miR-4423-3p, hsa-miR-182-5p, and hsa-miR-139-5p deregulated after ETP treatment (Fig.   2e). [score:2]
[1 to 20 of 3 sentences]
87
[+] score: 17
miRNAExpression change [a] miRbase accession numberExpression change [b] miR-132-5p Down MIMAT0004594 DownLi et al., 2013 miR-125b-1-3p Down MIMAT0004592 DownLi et al., 2013; Mar-Aguilar et al., 2013a miR-34c-5p Down MIMAT0000686 DownYang et al., 2013 miR-382-3p Down MIMAT0022697 DownLi et al., 2013; Mar-Aguilar et al., 2013b miR-485-5p Down MIMAT0002175 DownAnaya-Ruiz et al., 2013 miR-323b-3p Down MIMAT0015050 NA NA miR-598-3p Down MIMAT0003266 NA NA miR-224-5p Up MIMAT0000281 UpHuang et al., 2012 miR-1246 Up MIMAT0005898 UpPigati et al., 2010 miR-184 Up MIMAT0000454 NA NA a Expression change in this study. [score:7]
MicroRNA-34 suppresses breast cancer invasion and metastasis by directly targeting Fra-1. Oncogene 32, 4294– 4303 10.1038/onc. [score:5]
For example, the previous studies showed that miR-132 (Li et al., 2013), miR-125b (Zhang et al., 2011), miR-34c (Yang et al., 2013), and miR-485 (Anaya-Ruiz et al., 2013), functioning as suppressors, played an important role in breast cancer by suppressing cell proliferation and migration. [score:5]
[1 to 20 of 3 sentences]
88
[+] score: 17
Other miRNAs from this paper: hsa-let-7a-1, hsa-let-7a-2, hsa-let-7a-3, hsa-let-7b, hsa-let-7c, hsa-let-7d, hsa-let-7e, hsa-let-7f-1, hsa-let-7f-2, hsa-mir-15a, hsa-mir-26b, hsa-mir-29a, hsa-mir-30a, hsa-mir-29b-1, hsa-mir-29b-2, hsa-mir-106a, mmu-let-7g, mmu-let-7i, mmu-mir-15b, mmu-mir-29b-1, mmu-mir-30a, mmu-mir-30b, mmu-mir-125a, mmu-mir-125b-2, mmu-mir-130a, mmu-mir-138-2, mmu-mir-181a-2, mmu-mir-182, hsa-mir-30c-2, hsa-mir-30d, mmu-mir-30e, hsa-mir-10a, hsa-mir-34a, hsa-mir-181a-2, hsa-mir-181b-1, hsa-mir-181c, hsa-mir-182, hsa-mir-181a-1, mmu-mir-297a-1, mmu-mir-297a-2, mmu-mir-301a, mmu-mir-34c, mmu-mir-34b, mmu-let-7d, mmu-mir-106a, mmu-mir-106b, hsa-let-7g, hsa-let-7i, hsa-mir-15b, hsa-mir-30b, hsa-mir-125b-1, hsa-mir-130a, hsa-mir-138-2, hsa-mir-125a, hsa-mir-125b-2, hsa-mir-138-1, mmu-mir-30c-1, mmu-mir-30c-2, mmu-mir-30d, mmu-let-7a-1, mmu-let-7a-2, mmu-let-7b, mmu-let-7c-1, mmu-let-7c-2, mmu-let-7e, mmu-let-7f-1, mmu-let-7f-2, mmu-mir-15a, mmu-mir-26b, mmu-mir-29a, mmu-mir-29c, mmu-mir-34a, rno-mir-301a, rno-let-7d, rno-mir-344a-1, mmu-mir-344-1, rno-mir-346, mmu-mir-346, rno-mir-352, hsa-mir-181b-2, mmu-mir-10a, mmu-mir-181a-1, mmu-mir-29b-2, mmu-mir-138-1, mmu-mir-181b-1, mmu-mir-181c, mmu-mir-125b-1, hsa-mir-106b, hsa-mir-29c, hsa-mir-30c-1, hsa-mir-34b, hsa-mir-301a, hsa-mir-30e, hsa-mir-362, mmu-mir-362, hsa-mir-369, hsa-mir-374a, mmu-mir-181b-2, hsa-mir-346, rno-let-7a-1, rno-let-7a-2, rno-let-7b, rno-let-7c-1, rno-let-7c-2, rno-let-7e, rno-let-7f-1, rno-let-7f-2, rno-let-7i, rno-mir-10a, rno-mir-15b, rno-mir-26b, rno-mir-29b-2, rno-mir-29a, rno-mir-29b-1, rno-mir-29c-1, rno-mir-30c-1, rno-mir-30e, rno-mir-30b, rno-mir-30d, rno-mir-30a, rno-mir-30c-2, rno-mir-34b, rno-mir-34c, rno-mir-34a, rno-mir-106b, rno-mir-125a, rno-mir-125b-1, rno-mir-125b-2, rno-mir-130a, rno-mir-138-2, rno-mir-138-1, rno-mir-181c, rno-mir-181a-2, rno-mir-181b-1, rno-mir-181b-2, rno-mir-181a-1, hsa-mir-449a, mmu-mir-449a, rno-mir-449a, mmu-mir-463, mmu-mir-466a, hsa-mir-483, hsa-mir-493, hsa-mir-181d, hsa-mir-499a, hsa-mir-504, mmu-mir-483, rno-mir-483, mmu-mir-369, rno-mir-493, rno-mir-369, rno-mir-374, hsa-mir-579, hsa-mir-582, hsa-mir-615, hsa-mir-652, hsa-mir-449b, rno-mir-499, hsa-mir-767, hsa-mir-449c, hsa-mir-762, mmu-mir-301b, mmu-mir-374b, mmu-mir-762, mmu-mir-344d-3, mmu-mir-344d-1, mmu-mir-673, mmu-mir-344d-2, mmu-mir-449c, mmu-mir-692-1, mmu-mir-692-2, mmu-mir-669b, mmu-mir-499, mmu-mir-652, mmu-mir-615, mmu-mir-804, mmu-mir-181d, mmu-mir-879, mmu-mir-297a-3, mmu-mir-297a-4, mmu-mir-344-2, mmu-mir-466b-1, mmu-mir-466b-2, mmu-mir-466b-3, mmu-mir-466c-1, mmu-mir-466e, mmu-mir-466f-1, mmu-mir-466f-2, mmu-mir-466f-3, mmu-mir-466g, mmu-mir-466h, mmu-mir-493, mmu-mir-504, mmu-mir-466d, mmu-mir-449b, hsa-mir-374b, hsa-mir-301b, rno-mir-466b-1, rno-mir-466b-2, rno-mir-466c, rno-mir-879, mmu-mir-582, rno-mir-181d, rno-mir-182, rno-mir-301b, rno-mir-463, rno-mir-673, rno-mir-652, mmu-mir-466l, mmu-mir-669k, mmu-mir-466i, mmu-mir-669i, mmu-mir-669h, mmu-mir-466f-4, mmu-mir-466k, mmu-mir-466j, mmu-mir-1193, mmu-mir-767, rno-mir-362, rno-mir-504, rno-mir-582, rno-mir-615, mmu-mir-3080, mmu-mir-466m, mmu-mir-466o, mmu-mir-466c-2, mmu-mir-466b-4, mmu-mir-466b-5, mmu-mir-466b-6, mmu-mir-466b-7, mmu-mir-466p, mmu-mir-466n, mmu-mir-344e, mmu-mir-344b, mmu-mir-344c, mmu-mir-344g, mmu-mir-344f, mmu-mir-374c, mmu-mir-466b-8, hsa-mir-466, hsa-mir-1193, rno-mir-449c, rno-mir-344b-2, rno-mir-466d, rno-mir-344a-2, rno-mir-1193, rno-mir-344b-1, hsa-mir-374c, hsa-mir-499b, mmu-mir-466q, mmu-mir-344h-1, mmu-mir-344h-2, mmu-mir-344i, rno-mir-344i, rno-mir-344g, mmu-let-7j, mmu-mir-30f, mmu-let-7k, mmu-mir-692-3, rno-let-7g, rno-mir-15a, rno-mir-762, mmu-mir-466c-3, rno-mir-29c-2, rno-mir-29b-3, rno-mir-344b-3, rno-mir-466b-3, rno-mir-466b-4
Such a situation occurred for miR-26b, miR-30, and miR-374 downregulation, and for miR-34, miR-301, and miR-352 upregulation [121]. [score:7]
Similarly, miR-34, an established p53 effector that is typically downregulated in malignant lung cancer [105], was upregulated in microadenomas but not in adenomas, as demonstrated in the present study. [score:7]
Thus, maintenance of miR-34 expression is a prerequisite to avoid the passage from benign to malignant cancer lesions in lung tissue. [score:3]
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89
[+] score: 16
miR-34c is downregulated in laryngeal premalignant epithelial lesions [98] and lung cancer, associated with the use of tobacco [109], and upregulated in two studies related to alcohol consumption [30, 97]. [score:7]
In premalignant lesions of the larynx, miR-10a levels are reduced and miR-34c increases gradually by severity of the dysplasia, wherein miR-34c expression levels are higher in patients who consume alcohol [98]. [score:3]
Hu Y. Liu H. MicroRNA-10a-5p and microRNA-34c-5p in laryngeal epithelial premalignant lesions: Differential expression and clinicopathological correlation Eur. [score:3]
Tobacco and alcohol seem to regulate miR-34c and miR-223 through the signaling pathways of apoptosis and the cell cycle [12, 30, 70]. [score:2]
According to the studies addressed in this review, the major miRNAs associated with tobacco and alcohol are miR-21, miR-34a, miR-34c [30, 97, 98, 109], miR-223 [12, 70], miR-375, and miR-210 [63, 64], which are involved in many signaling pathways, such as proliferation [52], transformation [9, 13], inflammation [9, 13], angiogenesis [13], apoptosis, and the cell cycle [12, 30, 70, 71, 98, 109]. [score:1]
[1 to 20 of 5 sentences]
90
[+] score: 16
[31] The p53-miR-34 regulatory axis is another example of how transcriptional factor regulates miRNA expression to mediate tumor suppressive function. [score:7]
[31] The p53-miR-34 regulatory axis is another example of how transcriptional factor regulates miRNA expression to mediate tumor suppressive function. [score:7]
Similar to p53 -mediated phenotypes, miR-34 family including miR-34a/b/c promotes cell-cycle arrest, cell senescence and apoptosis in cancer, [33] implying p53 and miR-34 are in the same regulatory pathway. [score:2]
[1 to 20 of 3 sentences]
91
[+] score: 16
Members of the miR-34 family of miRNAs are direct targets of p53 and function as tumor suppressors, inhibiting reprogramming through the repression of pluripotency genes such as Nanog, Sox2, and N-myc (Choi et al., 2011) (Figure 2). [score:8]
Evidence that let-7 and miR-34 family members are tumor suppressor miRNAs (Takamizawa et al., 2004; Johnson et al., 2005; Tazawa et al., 2007) suggests that stem cell-specific miRNAs play important roles in tumor initiation and development. [score:4]
The expression levels of miR-34 and let-7 family members increase during differentiation. [score:3]
miR-34 miRNAs provide a barrier for somatic cell reprogramming. [score:1]
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92
[+] score: 16
Since we observed that PI3K-C2β regulates activation of the transcription factor STAT3, which in turn can also be involved in miR-34 family regulation [44], we then tested whether STAT3 could be involved in PI3K-C2β -dependent regulation of miR-449a in MCF7 cells. [score:4]
miR-449a belongs to the miR-34 family, which is expressed at a low level in several cancer cell lines and solid tumors including breast cancer [38]. [score:3]
Upregulation of miR-34b (Supplementary Figure S3D), miR-34c (Supplementary Figure S3E) and miR-449b (Supplementary Figure S3F) was also confirmed in T47D lacking PI3K-C2β compared to control cells, although the levels of these miRs were much lower compared to the levels of miR449-a. Taken together these data indicated that PI3K-C2β regulates miR levels, in particular it down modulates miR-449a levels. [score:3]
This analysis revealed a selective upregulation of 16 miRs in T47D cells lacking PI3K-C2β compared to control cells, including 5 miRs belonging to the same family: miR-449a, miR-449b, miR-34b, miR-34b*, miR-34c-5p (Figure 3D). [score:3]
Interestingly, members of the miR-34 family have been shown to regulate cyclin B1 levels [20, 21] therefore, they could potentially be involved in the PI3K-C2β -mediated regulation of cyclin B1. [score:3]
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93
[+] score: 16
miR-34 is involved in regulating the p53 pathway and inhibits cancer cell growth by directly targeting oncogenes such as Myc, c-Met, Bcl-2, CDK4, CDK6, Cyclin D1, and Cyclin E2 [42, 43, 46]. [score:7]
The expression levels of miR-34 are decreased in most human cancers [42– 44], including several epithelial cancers, melanomas, neuroblastomas, leukemias and sarcoma [45]. [score:3]
MRX34 is a liposome-formulated mimic of the tumor suppressor miR-34 (Mirna Therapeutics, Austin, TX). [score:3]
Anti-miR-122 therapy against chronic hepatitis C. miR-34 mimics as a therapeutic against primary and metastatic liver cancer. [score:1]
In addition to the current success of anti-miR122 therapy against chronic hepatitis C and the ongoing studies of miR-34 mimics against liver cancers in human clinical trials, the results of preclinical studies will likely lead to human clinical trials in the near future. [score:1]
In addition to miravirsen, a clinical trial of MRX34 as a mimic of miR-34 is ongoing. [score:1]
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94
[+] score: 15
For example, to re-introduce miR-34 and its tumor suppressor capabilities, transfection with miR-34 mimics into cancer cells was shown to block the cell cycle in the G1 phase, significantly increasing activation of caspase-3, and knocked down its downfield targets of bcl-2, Notch, and HMGA2 [127]. [score:6]
In this sense, a lentiviral system restored the tumor suppressor effect of miR-34 in pancreatic cancer stem cells [128]. [score:3]
Therapeutic delivery of synthetic mi -RNA, using a neutral lipid emulsion (NLE), exhibited tumor -inhibitory effects of let-7 and miR-34 formulations in an autochthonous transgenic mouse mo del of lung cancer. [score:3]
The miRNA mimic, is therefore restored miR-34 with its tumor suppressor potential; however, the transfection of the miR-34 mimics can only last a couple of days and the long-term biological effects were not observed very effectively. [score:3]
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95
[+] score: 15
The remarkable role of miR-34 in development and disease (Rokavec et al., 2014) is explained by the existence of the p53/miR-34 axis. [score:4]
The p53/miR-34 axis in development and disease. [score:4]
Also, HS leads to down -expression of mir-277 cluster (mir-34, mir-317) and mir-306 cluster (mir-9c, mir-9b, mir-79). [score:3]
Therefore, miR34 is involved in negative regulation of aging and death of neurons (Ghosh et al., 2008; Maciotta et al., 2013; Feng et al., 2014). [score:2]
As show in this study, additional binding site for mir-34 is created by the insertion of S-element from Tc1/mariner family in agn ts3 (Figure S3). [score:1]
A biomarker for AD is miR-34. [score:1]
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96
[+] score: 15
At present, it was found that p53 regulated PDL-1 via miR-34, which directly binded to the PDL-1 3′untranslated region in mo dels of NSCLC [98]. [score:5]
P53 regulated PDL-1 via miR-34, and miR-34 enhanced T cell activation via targeting diacylglycerol kinase ζ. [score:4]
What's more, it was also reported that miR-34 enhanced T cell activation via targeting diacylglycerol kinase ζ [99] (Figure 2A). [score:3]
In addition, lncRNAs could be precursors of miRNAs and act as ceRNAs to alter the distribution of miRNA molecules on their targets [6, 101] (Figure 2B), for example, it was found that lncRNA ARSR acted as a ceRNA for miR-34 and miR-449 and finally promoted Sunitinib resistance in renal cancer [102]. [score:3]
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97
[+] score: 15
Other miRNAs from this paper: hsa-let-7a-1, hsa-let-7a-2, hsa-let-7a-3, hsa-let-7b, hsa-let-7c, hsa-let-7d, hsa-let-7e, hsa-let-7f-1, hsa-let-7f-2, hsa-mir-15a, hsa-mir-16-1, hsa-mir-17, hsa-mir-19b-1, hsa-mir-19b-2, hsa-mir-20a, hsa-mir-21, hsa-mir-24-1, hsa-mir-24-2, hsa-mir-26a-1, hsa-mir-26b, hsa-mir-29a, hsa-mir-30a, hsa-mir-98, hsa-mir-101-1, hsa-mir-29b-1, hsa-mir-29b-2, hsa-mir-106a, hsa-mir-16-2, hsa-mir-192, hsa-mir-148a, hsa-mir-30c-2, hsa-mir-30d, hsa-mir-7-1, hsa-mir-7-2, hsa-mir-7-3, hsa-mir-10a, hsa-mir-10b, hsa-mir-34a, hsa-mir-210, hsa-mir-215, hsa-mir-200b, hsa-let-7g, hsa-let-7i, hsa-mir-1-2, hsa-mir-30b, hsa-mir-124-1, hsa-mir-124-2, hsa-mir-124-3, hsa-mir-125b-1, hsa-mir-133a-1, hsa-mir-133a-2, hsa-mir-137, hsa-mir-138-2, hsa-mir-143, hsa-mir-144, hsa-mir-145, hsa-mir-152, hsa-mir-9-1, hsa-mir-9-2, hsa-mir-9-3, hsa-mir-125a, hsa-mir-125b-2, hsa-mir-126, hsa-mir-127, hsa-mir-138-1, hsa-mir-146a, hsa-mir-193a, hsa-mir-194-1, hsa-mir-206, hsa-mir-320a, hsa-mir-200c, hsa-mir-1-1, hsa-mir-155, hsa-mir-194-2, hsa-mir-106b, hsa-mir-29c, hsa-mir-30c-1, hsa-mir-200a, hsa-mir-302a, hsa-mir-101-2, hsa-mir-34b, hsa-mir-30e, hsa-mir-26a-2, hsa-mir-302b, hsa-mir-302c, hsa-mir-302d, hsa-mir-369, hsa-mir-371a, hsa-mir-340, hsa-mir-335, hsa-mir-133b, hsa-mir-146b, hsa-mir-519e, hsa-mir-519c, hsa-mir-519b, hsa-mir-519d, hsa-mir-519a-1, hsa-mir-519a-2, hsa-mir-499a, hsa-mir-504, hsa-mir-421, hsa-mir-320b-1, hsa-mir-320c-1, hsa-mir-320b-2, hsa-mir-190b, hsa-mir-301b, hsa-mir-302e, hsa-mir-302f, hsa-mir-320d-1, hsa-mir-320c-2, hsa-mir-320d-2, hsa-mir-320e, hsa-mir-371b, hsa-mir-499b
As well as representing transcriptional targets of p53, miRNAs miR-34a, miR-34b and miR-34c target several key mRNAs in DNA damage response itself, including transcripts involved in cell cycle arrest in the G1-phase or cellular apoptosis such as CDK4 and CCNE2. [score:5]
Studies in p53 deficient mouse embryonic fibroblast cells have indicated that one family of miRNAs, miR34a—miR34c has particular importance and are direct targets of p53 [20]. [score:4]
In addition to its role in regulation of DNA damage response, the miR-34 family may also regulate telomere length; miR-34 levels were associated with telomere length in a series of gall bladder adenocarcinomas [34]. [score:3]
Jin K. Xiang Y. Tang J. Wu G. Li J. Xiao H. Li C. Chen Y. Zhao J. miR-34 is associated with poor prognosis of patients with gallbladder cancer through regulating telomere length in tumor stem cells Tumour Biol. [score:2]
Similarly, silencing of the miR-124a, miR-34, miR-9 and miR-200 gene families by DNA methylation or histone modifications have been noted in several studies [38, 39, 40, 41]. [score:1]
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98
[+] score: 15
The expression of miR-34c-5p was upregulated during the T cell receptor (TCR) signaling -mediated activation of naïve CD4 [+] T-cells, which altered the expression levels of multiple genes involved in HIV-1 replication. [score:8]
Interestingly, the enhancement of HIV-1 replication in T-cells overexpressing miR-34c-5p and the downregulation of endogenous miR-34c-5p upon HIV-1 infection led the authors to speculate that the latter is an antiviral host response. [score:6]
Another member of the cellular miR-34 family, the miR-34c-5p was also recently shown to promote HIV-1 infection in CD4 [+] T-cells [191]. [score:1]
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99
[+] score: 14
In particular, the following target proteins were downregulated: PTEN which is known to be targeted by miR-21 [28], [29], cyclin D1 which is known to be targeted by miR-100, miR-99a and miR-223 [30] and Bcl-2 which is known to be targeted directly by miR-34, miR-181b and miR-16 [31], [32], [33] or indirectly modulated by miR-21 [34]. [score:14]
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100
[+] score: 14
The mir-34 genes induce cell cycle arrest, cellular senescence, and apoptosis when ectopically expressed (Bommer et al., 2007; He et al., 2007; Welch et al., 2007) through the downregulation of multiple target genes such as Bcl-2, Cyclin D1, Cyclin E2, CDK4, CDK6, c-Myc, and c-Met (Hermeking, 2010). [score:8]
p53 -mediated activation of miRNA34 candidate tumor-suppressor genes. [score:3]
MicroRNA-34b and MicroRNA-34c are targets of p53 and cooperate in control of cell proliferation and adhesion-independent growth. [score:2]
The miR-34 family in cancer and apoptosis. [score:1]
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