9 publications mentioning hsa-mir-4516

Open access articles that are associated with the species Homo sapiens and mention the gene name mir-4516. Click the [+] symbols to view sentences that include the gene name, or the word cloud on the right for a summary.

1

Finally, based on the cellular mechanisms of miRNAs, which is the regulation of gene expression by the sequence-selective targeting of mRNA leading to their cleavage or a reduction in translational efficiency, we checked the protein expression level of PVRL1 by a WB assay and also found that PVRL1 protein expression was sharply declined in 293T cells transfected with the miR-4516-up plasmid (Figure 4D).

The protein expression levels were slightly and dramatically decreased in EV-A71- and CV-A16-infected cells (P < 0.05), respectively, whereas the protein expression levels were remarkably decreased with miR-4516-overexpression pretreatment in an EV-A71 infection and apparently recovered with a miR-4516-knockdown pretreatment in a CV-A16 infection (P < 0.05).

Among these miRNAs, miR-4516, which was down-regulated in EV-A71 infection and up-regulated in CV-A16 infection, presented the most significant change (Figure 4A).

Our WB and IF observations showed that nectin1 was slightly lower and miR-4516 was down-regulated in EV-A71-infected cells, while nectin1 was notably lower and miR-4516 was up-regulated in CV-A16-infected cells, but these alterations of nectin1 were completely reversed in EV-A71- and CV-A16-infected cells pretreated with the miR-4516-up and miR-4516-down plasmids, respectively.

miRNA -negative control (NC) plasmid, miR-4516-overexpression plasmid, 3′-UTR NC plasmid, PVRL1 3′-UTR plasmid, PVRL1 3′UTR-Mutant plasmid and TRAF6 3′-UTR plasmid (which was demonstrated that it was a direct target of miR-4516) used in this study were synthesized by the Shanghai Genechem Co.

Figure 6Junctional proteins expression in 16HBE cells infected with EV-A71 or EV-A71 pre -treated with a miR-4516-overexpression plasmid, and CV-A16, or CV-A16 pre -treated with a miR-4516-knockdown plasmid.

In the present study, based on results from our previous study, we screened the miR-4516 and its target PVRL1 (also known as nectin1) which might be the key regulators contributed to the different impairments of the epithelial barrier because of their opposite expression pattern during EV-A71 and CV-A16 infections.

Collectively, we proposed that differentially expressed miR-4516 induced by EV-A71 and CV-A16 infections might lead to the disruption of cellular junctions by targeting nectin1, which ultimately resulted in perturbation of barrier functions of 16HBE cells.

Furthermore, the expression level of PVRL1, a predicted target of miR-4516, was remarkably increased in EV-A71 infection and markedly decreased in CV-A16 infection (Figure 4B).

PVRL1-3′UTR is the direct target of miR-4516 which was the key miRNA involved in regulation of epithelial barrier function.

Transfection with the miR-4516-overexpression plasmid resulted in a significant reduction of PVRL1 protein expression in 16HBE cells.

Therefore, these observations indicated that EV-A71 and CV-A16 could interfere with AJ and TJ assembly, which was intimately linked to the breakdown of the epithelial barrier, and miR-4516 was differentially expressed following infection with EV-A71 or CV-A16, which might be important regulators involved in the degradation of AJ- and TJ-related proteins.

In contrast, in EV-A71 infected cells that overexpressed miR-4516, most showed a substantial reduction or even the disappearance of the nectin1 staining, while in CV-A16 infected miR-4516-knockdown cells, we observed stronger nectin1 staining at the cell-cell contact sites.

In conclusion, for the first time, this study demonstrated that miR-4516 regulated the epithelial permeability barrier functions by targeting PVRL1 and might contribute to the differences in the pathogenesis of EV-A71 and CV-A16 infections.

Figure 4PVRL1 was speculated as the direct target of miR-4516.

Therefore, it was confirmed that PVRL1 3′-UTR was the direct target of miR-4516.

It is noteworthy that EV-A71 and CV-A16 both actually broke the epithelial barrier, but comparatively, an opposite miR-4516 expression pattern resulted from EV-A71 and CV-A16 infection in 16HBE cells, which might regulate cell-cell TJs and AJs and consequently cause different degrees of impairment of the epithelial barrier that further facilitate viral infection, especially CV-A16 infection.

Once cells reached approximately 80% confluence, co-transfection, including miRNA-NC plasmid or miR-4516- overexpression plasmid with the other four plasmids, respectively, was carried out using the FuGENE® HD Transfection Reagent (Promega, USA) according to the manufacturer's instructions.

Detection of adhesion-related molecules and miR-4516 expressions by qRT-PCR.

The light blue arrows represent that the EV-A71-infected cells with miR-4516-overexpression plasmid pretreatment showed remarkably lower levels of these proteins.

miR-4516 destroys the location of junctional proteins and decreases the expression levels of junctional proteins.

It has been reported that high expression levels of miR-4516 were associated with the infiltrative growth of the follicular variant of papillary thyroid carcinomas (Borrelli et al., 2017), but its role in EV-A71 and CV-A16 infections remains unclear.

The saffron arrows indicate that CV-A16-infected cells pre -treated with the miR-4516-knockdown plasmid show less recovery of these proteins.

Additionally, the regulation mechanism of miR-4516 on PVRL1 was further detected by WB in 293T cells subjected to miR-4516-up transfection.

The si-control and miR-4516-knockdown plasmids contain a GFP tag (green color).

First, we investigated the distribution of the direct target of miR-4516.

The total RNA from each sample was obtained and then subjected to qRT-PCR for all adhesion-related genes and miR-4516 by a One-Step SYBR® PrimeScript™ PLUS RT-PCR Kit (TAKARA, Japan) as previously described.

The miR-4516 was chosen for further study due to its most significant change.

U6 snRNA and GAPDH are served as an appropriate calibrator housekeeping gene for miR-4516 and adhesion-related genes quantitative analysis, respectively.

Therefore, we investigated the role of the opposite expression of miR-4516 in EV-A71 and CV-A16 infections on the distribution of the AJ and TJ-related proteins.

2

S8 TableAll targets of both miR-601 and miR-4516 are listed with scores according to both TargetScan and microRNA.

To better understand mechanism(s) by which these miRNAs are acting, mRNA targets of all novel miRNAs, including miR-601 and miR-4516 were identified using TargetScan [79] and microRNA.

S10 Table Ten genes with the lowest scores and highest probability of being targeted by miR-4516 according to TargetScan are listed with their known gene functions.

Although miR-601 appears to be a major player in predictive ability of salvage radiation response, miR-4516 and miR-601 together appear to be correlated (Pearson r = 0.56, p-value <0.0001) and target pathways of both STAT3 and NF-kB pathways that are critical regulators of the inflammatory responses in cancer [89].

In March of 2014, the only cancer-related study for miR-4516 was published in which Chowdhari et al. showed that miR-4516 downregulates STAT3 and mediates UV -induced apoptosis in a human keratinocyte cell line [45].

Additional putative targets of both miR-601 and miR-4516 are shown in S8– S10 Tables and proteins in multiple pathways including apoptosis and cell cycle regulation were identified.

The two miRNAs that were significant within the predictive mo del were miR-601 and miR-4516; both of which were upregulated in patients who experienced biochemical failure post-salvage RT.

Top 10 putative gene targets and function for miR-4516.

Putative targets of miRNAs, miR-601 and miR-4516, within our predictive mo del of biochemical recurrence post-salvage RT (second biochemical recurrence).

Predictive effect of miR-601 + miR-4516 & Development of a miRNA -based Predictive Salvage RT Mo del.

0118745.g004 Fig 4 A K-M plot was generated using the miR-4516 + miR-601 + Gleason score + lymph node status mo del for biochemical recurrence post salvage radiation therapy.

Interestingly, each miRNA alone (miR-601 AUC = 0.77 and miR-4516 AUC = 0.68) or together (miR-601 + miR-4516 AUC = 0.76) (Fig. 3) had a better predictive capability than the combination of positive lymph nodes together with Gleason score (AUC = 0.66) (Fig. 3).

Thus, we have identified 3 novel PCa miRNAs (miR-1193, miR-4516, and miR-626) associated with second biochemical recurrence after salvage RT.

Using the risk score generated by this mo del (miR-4516, miR-601, Gleason score, and lymph node status), patients were classified as high or low risk groups based on median, the probabilities in recurrence post-salvage RT of the two groups were found to be significantly different (log-rank, p<0.001) as shown in Fig. 4. To ensure that hormonal therapy did not confound the predictive mo del, patients that received hormonal treatment at any point were removed from the mo del and the mo del still retained its specificity and sensitivity with a log-rank p-value of 0.016 and AUC value of 0.83 (S1 Fig. ).

A K-M plot was generated using the miR-4516 + miR-601 + Gleason score + lymph node status mo del for biochemical recurrence post salvage radiation therapy.

Because these miRNAs are novel to PCa, we wanted to investigate the putative mRNA targets of miR-1193, miR-4516, and miR-626.

The two miRNAs, miR-601 and miR-4516 alone and together with Gleason score and lymph node status were selected by the mo del.

An AUC for lymph node and Gleason score was found to be 0.66, the addition of miR-601 and miR-4516 increased the AUC to 0.83 (Fig. 3).

miR-4516 was not listed in microRNA.

These future analyses will help elucidate the roles that both miR-4516 and miR-601 have in the clinical response to salvage-RT post-prostatectomy.

We showed that miRNAs alone predicted biochemical recurrence post-salvage RT and adding two miRNAs, miR-601 and miR-4516, to lymph node status and Gleason score, greatly improved the predictive ability of these clinical factors alone.

Currently, there are no publications regarding miR-4516 and PCa.

Gleason score, lymph node status, hsa-miR-4516, and hsa-miR-601 (red) performs the best with an AUC of 0.83 followed by a mo del containing hsa-miR-601-alone (green) (AUC = 0.77), hsa-miR-4516-alone (blue) (AUC = 0.68), and lastly, Gleason score and lymph node status (grey) (AUC = 0.66).

A new predictive mo del for biochemical recurrence after salvage radiation therapy was developed; this mo del consisted of miR-4516 and miR-601 together with, Gleason score, and lymph node status.

3

Chowdhari et al. show that overexpression of hsa-miR-4516 downregulates STAT3, p-STAT3, CDK6, and UBE2N proteins that are consistently upregulated in psoriasis and induces apoptosis in HaCaT cells [63].

One of the 37 miRNAs where the expression was upregulated in brucellosis is miR-4516.

The expression of miR-4516 was 4.35 fold and 3.74 fold in chronic group and acute infection groups respectively in comparison to the control group.

Asahchop et al. showed that miR-4516, which is a plasma-derived miRNA, was differentially expressed in HIV/AIDS patients with HIV -associated neurocognitive disorder [75].

Thus we suggest that, miR-4516 may involve effector responses of CD8+ T cells in these patients through reducing STAT3, CDK6, and UBE2N expression.

4

MicroRNA inhibitors were purchased from Exiqon (Woburn, MA, USA) including miRCURY LNA Power Inhibitor Control (199020-04), miRCURY LNA Power Inhibitor hsa-miR-494 (427173-04), miRCURY LNA Power Inhibitor has-miR-1246 (426697-04) and a custom miRCURY LNA Power Inhibitor hsa-miR-4516.

miR-4488 levels were very low and not detected by qRT-PCR and miR-494, miR-1246 and miR-4516 were significantly upregulated in cells treated with 2DG+TRAIL relative to controls treated with TRAIL or 2DG alone, validating the array data.

Knockdown of miR-4516 in the colon cancer cells did not abrogate tumor cell apoptosis upon treatment with 2DG+TRAIL.

MiR-494, miR-4488 and miR-4516 were upregulated by 2.5-, 5.7- and 2.9-fold, respectively, under conditions of 2DG+TRAIL treatment compared with TRAIL alone, and 4.5-, 15.0- and 3.4-fold compared with 2DG alone.

MicroRNA mimics were purchased from Qiagen, including Syn-hsa-miR-494 (MSY0002816), Syn-hsa-miR-1246 (MSY0005898) and Syn-hsa-miR-4516 (MSY0019053).

This sensitivity was not enhanced further when combining the miR-494 mimic with those of miR-1246 and miR-4516.

5

According to previous reports, rRNA-contained miRNAs such as miR-663, miR-1275, miR-3648, miR-3656, miR-3687, miR-4417, and miR-4516 are associated with tumor suppression, carcinomas, neuronal differentiation, breast cancer, breast cancer/neuronal differentiation, breast cancer, and regulation of signal transducer and activator of transcription 3, respectively [103– 108].

Subsequently, seventeen RNA alignments identical to human mature miRNAs, namely, miR-663a, miR-663b-3p, miR-1268a, miR-1268b, miR-1275, miR-3648, miR-3656-3p, miR-3687-3p, miR-4417, miR-4466, miR-4488, miR-4492-3p, miR-4508, miR-4516, miR-4532, miR-6087, and miR-6724, were detected from the human rRNA sequences (Table 1).

6

On the other hand, the expression levels of naked miRNAs hsa-miR-320e and hsa-miR-4516 were particularly high in the unfractionated urine, indicating that their absence in exosomes was not due to the detection failure.

We also found 15 naked miRNAs including miRNAs hsa-miR-320e, hsa-miR-4516, hsa-miR-451a and hsa-miR-194–5p, which were not detected in any of the exosomal RNA preparations, but present in at least 5 out of 6 total urinary RNAs.

7

Eleven miRNAs (miRNA-4530, miRNA-4739, miRNA-762, miRNA-4787-5p, miRNA-940, miRNA-3676-5p, miRNA-6090, miRNA-150-5p, miRNA-4516, miRNA-4284, miRNA-3656) demonstrated a similar expression trend in both the acute and chronic groups.

8

The change from the C allele to the T allele of this SNP deletes the regulation of EPHB1 by miR-4434 and miR-4516.

9

Hsa-miR-1246, hsa-miR-4488, hsa-miR-4508, hsa-miR-4492 and hsa-miR-4516 were identified as the 5 most exosomal enriched miRNAs (Fig 2C).

Hsa-miR-1246, hsa-miR-4488, hsa-miR-4508, hsa-miR-4492 and hsa-miR-4516 were identified as the 5 most abundant miRNA types in exosomes derived from hNSC producers and had the highest ratio of exosomal to cellular miRNA abundance.