4 publications mentioning mtr-MIR164c

Open access articles that are associated with the species Medicago truncatula and mention the gene name MIR164c. Click the [+] symbols to view sentences that include the gene name, or the word cloud on the right for a summary.

1

Expression of a miR164-Resistant form of MtNAM Leads to a Breakdown in Carpel Margin Fusion and Other Developmental Fusion Events in Medicago truncatula FlowersTo test the role of the NAM/miR164 developmental module on carpel closure in M. truncatula, we produced transgenic plants expressing genomic constructs of MtNAM (MtNAMg-m4 and MtNAMg-wt), respectively, with or without four point mutations in their predicted miR164 -binding sites, identical to those present in the CUC2g-m4 construct (Nikovics et al., 2006).

These transcription factors are expressed at organ margins and tissue boundaries, and their down-regulation by miR164 consequently facilitates organ outgrowth and/or developmental fusion.

These expression data reveal several underlying similarities in the expression of miR164-regulated NAM orthologs between A. thaliana and M. truncatula.

Given the role of the NAM/miR164 module in carpel closure in both A. thaliana and M. truncatula (Figures 2 and 4), and the gene expression differences we have noted between the congenitally and post-genitally fused carpel margins of these two respective species, it would be interesting to compare the expression of NAM orthologs in a range of Fabales that show different spatial and temporal patterns of carpel closure.

To test the role of the NAM/miR164 developmental module on carpel closure in M. truncatula, we produced transgenic plants expressing genomic constructs of MtNAM (MtNAMg-m4 and MtNAMg-wt), respectively, with or without four point mutations in their predicted miR164 -binding sites, identical to those present in the CUC2g-m4 construct (Nikovics et al., 2006).

Accordingly, we furthermore hypothesize, based on our gene expression analyses (Figure 3), that the origin of plicate carpels in various later-emerging angiosperm lineages may have depended on subtle modifications to the NAM/miR164 module that allowed a limited level of early expression of NAM orthologs in the carpel margins, as occurs in present-day M. truncatula.

Expression of NAM Orthologs is Absent or Reduced During Carpel Margin Fusion in Arabidopsis thaliana and Medicago truncatulaThe observation that the NAM/miR164 module regulates developmental closure events in the gynoecium in both syncarpous and monocarpous genotypes of A. thaliana led us to speculate that this molecular mechanism might be wi dely conserved within the angiosperms.

Comparison of these data strongly suggests that the NAM/miR164 expression balance in A. thaliana lies heavily in favor of miR164 from the earliest stages of gynoecium development.

Notably, the NAM/miR164 expression balance at very early stages of carpel development may be important in determining whether carpel margins will fuse congenitally or postgenitally.

Thus, the different balances of NAM and miR164 expression observed at very early stages of A. thaliana and M. truncatula carpel development (Figure 3; Galbiati et al., 2013) correlate closely with the different timings of carpel closure observed in these species (Smyth et al., 1990; Benlloch et al., 2003).

Expression of a miR164-Resistant form of MtNAM Leads to a Breakdown in Carpel Margin Fusion and Other Developmental Fusion Events in Medicago truncatula Flowers.

A detailed comparison of gene expression patterns suggests that fine-tuning of the NAM/miR164 module may regulate species–specific differences in the timing of carpel margin fusion.

Thus, it appears reasonable to postulate that the NAM/miR164 module operates in favor of the expression of miR164, and against that of NAM orthologs, from the earliest stages of gynoecium development in A. trichopoda, as it does in A. thaliana.

Carpel fusion in A. thaliana is regulated by a genetic module, generically termed here the NAM/miR164 module, which consists of a subset of NAC -family (NAC for NAM, ATAF and CUC; Aida et al., 1997) transcription factors and their post-transcriptional regulator miR164 (Mallory et al., 2004).

Plants expressing a miR164-resistant CUC2 gene reveal the importance of post-meristematic maintenance of phyllotaxy in Arabidopsis.

Thus, disruption of the NAM/miR164 developmental module in monocarpous mutant gynoecia of A. thaliana causes the failure of developmental closure in these structures in a similar manner to the disruption of carpel fusion in syncarpous, wild-type gynoecia.

Despite the differences observed, we concluded that the presence of MtNAM expression in M. truncatula carpel margins suggested that the NAM/miR164 module may be involved in the fusion of these structures, leading us to test this hypothesis experimentally.

The observation that the NAM/miR164 module regulates developmental closure events in the gynoecium in both syncarpous and monocarpous genotypes of A. thaliana led us to speculate that this molecular mechanism might be wi dely conserved within the angiosperms.

Prior to initiating functional experiments in M. truncatula, we used in situ hybridization to examine the conservation of expression of NAM orthologs in flower tissues between A. thaliana and M. truncatula and thereby ascertain the likelihood that the NAM/miR164 module might function in carpel closure in the latter species.

Thus, the establishment of negative regulation by SPT of a miR164-regulated NAM gene in a common ancestor of the angiosperms may have been a crucial step in the evolution of the closed carpel.

Redundancy and specialization among plant microRNAs: role of the MIR164 family in developmental robustness.

Accordingly, in miR164 triple mutants or CUC2g-m4 transformants, the two carpels of the A. thaliana gynoecium emerge separately and remain unfused and open throughout development.

An important test of this hypothesis will depend on the development of plant transformation strategies in basally diverging angiosperms, which would allow, for example, the transformation of A. trichopoda with a miR164-resistant form of AtrNAM.

Like the NAM/miR164 module, it seems that SPT may have conserved its function in carpel development from the earliest stages of angiosperm evolution (Reymond et al., 2012).

Accordingly, we discuss the possibility that the activity of the NAM/miR164 module may be conserved in carpel development throughout the angiosperms, while subtle modulations to this mechanism may determine the distinction between congenital and post-genital carpel margin fusion events in specific angiosperm groups.

Such experiments, in quite closely related species showing marked differences in gynoecium anatomy, could provide strong correlative evidence of a role for the subtle modulation of gynoecium development by changes to the balance of the NAM/miR164 module.

These data indicate that the NAM/miR164 module has conserved a role in developmental fusion events between carpel margins at least since the MRCA of the eurosids.

The NAM/miR164 in Arabidopsis thaliana Plays a Role in Both Syncarpy and the Closure of Single CarpelsAs the NAM/miR164 developmental module is necessary for carpel fusion in wild-type, syncarpous A. thaliana (Nikovics et al., 2006; Sieber et al., 2007), we aimed to discover whether this mechanism could also contribute to the closure of single carpels in this species.

Loss of miR164 function through mutations to all three MIR164 paralogs in A. thaliana (Sieber et al., 2007), or genetic transformation of A. thaliana with a miR164-resistant version of CUC2 (CUC2g-m4; Nikovics et al., 2006), results in a breakdown of carpel fusion.

In the present work, we show that the role of the NAM/miR164 module in carpel margin fusion was conserved during this developmental transition.

As the NAM/miR164 developmental module is necessary for carpel fusion in wild-type, syncarpous A. thaliana (Nikovics et al., 2006; Sieber et al., 2007), we aimed to discover whether this mechanism could also contribute to the closure of single carpels in this species.

These data indicate a range of roles of the NAM/miR164 developmental module in fusion events in the corolla, androecium, and gynoecium of M. truncatula flowers.

As its genetic components are present in both gymnosperms and angiosperms (Axtell and Bartel, 2005; Larsson et al., 2012), the NAM/miR164 genetic module is clearly of ancient origin in seed plants.

From the above observations, we hypothesize that the NAM/miR164 module may have played a role in the fusion of carpel margins in the MRCA of the living angiosperms, as it does in present-day mo del angiosperms.

The NAM/miR164 Module Maintained its Role in Carpel Margin Fusion During a Transition from Syncarpy to Monocarpy in an Ancestor of Fabales.

A 1.2-kb fragment containing the miR164 -binding site of MtNAM was released from a sub-cloned BAC DNA fragment by cleavage with SstI and re-ligated into the pGEM T-Easy vector.

We show that disruption of the NAM/miR164 module in both A. thaliana aux1-22 mutants (Figure 2) and a wild-type background of M. truncatula (Figure 4) produces single carpels that are no longer completely fused at their margins.

From the results of these experiments, we conclude that the NAM/miR164 module has conserved a role in carpel margin fusion, at least since the most recent common ancestor (MRCA) of living eurosids.

In this study, we show that a previously characterized developmental module involving the post-transcriptional regulation of NAM orthologs by miR164 is involved not only in carpel fusion in syncarpous A. thaliana (Nikovics et al., 2006; Sieber et al., 2007), but also in the closure of the single carpels present in two species whose lineages diverged at the base of the eurosid clade, some 114–113 MYA.

A Possible Role for the NAM/miR164 Module in the Timing of Carpel FusionIn A. thaliana, the gynoecium forms as a radially symmetrical cylinder that later differentiates to show the positions of the carpel margins.

A Role of the NAM/miR164 Module in the Fusion of Carpel Margins has Been Conserved at Least Since the MRCA of the Eurosids.

A Role of the NAM/miR164 Module in the Fusion of Carpel Margins has Been Conserved at Least Since the MRCA of the EurosidsIn this study, we show that a previously characterized developmental module involving the post-transcriptional regulation of NAM orthologs by miR164 is involved not only in carpel fusion in syncarpous A. thaliana (Nikovics et al., 2006; Sieber et al., 2007), but also in the closure of the single carpels present in two species whose lineages diverged at the base of the eurosid clade, some 114–113 MYA.

The NAM/miR164 in Arabidopsis thaliana Plays a Role in Both Syncarpy and the Closure of Single Carpels.

The NAM/miR164 Module Maintained its Role in Carpel Margin Fusion During a Transition from Syncarpy to Monocarpy in an Ancestor of FabalesCharacter-state mapping (Figure 1) further indicates that the monocarpy present in Fabales (including M. truncatula) is a derived condition that occurred by reversion from syncarpy, present in earlier eurosids.

These oligonucleotides generate the same four-base mismatch (shown above in lower case) present in the miR164 -binding site of CUC2g-m4 (Nikovics et al., 2006).

Analysis of the pathway linking SPT, and its cofactors such as the HECATE transcription factors (Schuster et al., 2015), with the NAM/miR164 module in mo del angiosperms could provide insights into this possibility, and thus potentially indicate a molecular mechanism for the enclosure of the ovule with the carpel in the first angiosperms.

The Role of the NAM/miR164 Module in Carpel Evolution.

A Possible Role for the NAM/miR164 Module in the Timing of Carpel Fusion.

In this work, we hypothesized that the role of the NAM/miR164 module in syncarpous fusion in A. thaliana might reflect a more general role in the fusion of carpel margins in angiosperms.

The Role of the NAM/miR164 Module in Carpel EvolutionAs its genetic components are present in both gymnosperms and angiosperms (Axtell and Bartel, 2005; Larsson et al., 2012), the NAM/miR164 genetic module is clearly of ancient origin in seed plants.

We further speculate on mechanisms acting upstream of the NAM/miR164 module that may have contributed to the origin of the carpel in the first flowering plants.

Thus, our study strongly suggests that the NAM/miR164 module provides an underlying mechanism that is necessary for fusion events at the carpel margins of both syncarpous and monocarpous eurosids.

FIGURE 2Gynoecium morphology of A. thaliana aux1-22 mutants transformed with miR164-resistant (CUC2g-m4) or un-mutated control (CUC2g-wt) constructs.

The action of the NAM/miR164 module in the A. thaliana leaf margin has been mo deled and found to generate, via effects on the auxin efflux carrier PINFORMED1 (PIN1), an alternating series of auxin maxima and minima that, respectively, generate regions of higher and lower marginal growth (Bilsborough et al., 2011).

2

miR164 has been reported to regulate root development by a homeostatic mechanism to clear NAC1 mRNA, leading to down-regulation of auxin signals.

Therefore, it is conceivable that the suppression of miR164 expression may contribute to the increase in root/shoot ratio under drought stress.

Conversely, 10 members belonging to 6 miRNA families, i. e., miR164, miR169, miR171, miR396, miR398 and miR1510, were down-regulated in response to drought stress (Figure 3b and 3c).

It has been shown that decreases in NAC1 mRNA levels due to inducible expression of miR164, thus resulting in reduction in lateral root emergence in Arabidopsis [9].

In the present study, we found that the known or predicted targets of miR164, miR169, miR171, miR396, miR1510 and miR5558 were either transcription factors or F-box proteins (Table 3, 4).

3

Many of them, like miR160, miR164, miR167, miR390 and miR393, directly or indirectly regulate genes related to auxin signaling [10].

Other miRNAs, like miR164, miR166 or miR396, were shown to play indirect roles in nodule development or mycorrhizal symbiosis due to their global impact on auxin responses and/or tissue patterning in roots [13– 15].

4

D'Haeseleer et al. (2011) reported that over -expression of miR164, a conserved miRNA targeting NAC1 TF in roots, affected nodule organogenesis in M. truncatula, presumably through deregulation of auxin responses.