miR-124, -128, and -137 Orchestrate Neural Differentiation by Acting on Overlapping Gene Sets Containing a Highly Connected Transcription Factor Network

Abstract

The ventricular-subventricular zone harbors neural stem cells (NSCs) that can differentiate into neurons, astrocytes, and oligodendrocytes. This process requires loss of stem cell properties and gain of characteristics associated with differentiated cells. miRNAs function as important drivers of this transition; miR-124, -128, and -137 are among the most relevant ones and have been shown to share commonalities and act as proneurogenic regulators. We conducted biological and genomic analyses to dissect their target repertoire during neurogenesis and tested the hypothesis that they act cooperatively to promote differentiation. To map their target genes, we transfected NSCs with antagomiRs and analyzed differences in their mRNA profile throughout differentiation with respect to controls. This strategy led to the identification of 910 targets for miR-124, 216 for miR-128, and 652 for miR-137. The target sets show extensive overlap. Inspection by gene ontology and network analysis indicated that transcription factors are a major component of these miRNAs target sets. Moreover, several of these transcription factors form a highly interconnected network. Sp1 was determined to be the main node of this network and was further investigated. Our data suggest that miR-124, -128, and -137 act synergistically to regulate Sp1 expression. Sp1 levels are dramatically reduced as cells differentiate and silencing of its expression reduced neuronal production and affected NSC viability and proliferation. In summary, our results show that miRNAs can act cooperatively and synergistically to regulate complex biological processes like neurogenesis and that transcription factors are heavily targeted to branch out their regulatory effect. STEM CELLS 2016;34:220-232 SIGNIFICANCE STATEMENTWe demonstrate here, perhaps for the first time, that miRNAs (miR-124, -128 and -137) can act cooperatively to regulate a complex biological problem such as neurogenesis. They do so by targeting overlapping sets of genes. Moreover, our genomic analyses determined that transcription factors are the main component of their target sets. Our results indicate additional associations between the three analyzed miRNA and the targeted TFs. Based on described ChIP analysis, we suggest that miR-124, -128 and -137 and their targeted TFs could act on overlapping and related target sets producing different functional outcomes that ultimately would influence the balance self-renewal/differentiation.

MiR-124 inhibits the migration and invasion of human hepatocellular carcinoma cells by suppressing integrin αV expression

“…Through the luciferase reporter assay and binding site mutation, the results showed that miR-124 directly targeted Sp1 mRNA. MiR-124 may act together with miR-137 and miR-128 synergistically to regulate neural cells25. In this study we observed that miR-124 alone down-regulated the expression of Sp1 efficiently.…”

Exosome Mediated Delivery of miR-124 Promotes Neurogenesis after Ischemia

“…We produced the ischemic model and injected RVG-Exos-miR124 and RVG-Exos-Scr at 1 dpi. Then we harvested the ischemic tissue at 3 dpi and examined the mRNA expression of E2F1, Gli3, Stat3, and TCF12, which were previously predicted under the regulation of miR-124 and participated in neural differentiation17, 18, 19, 20, 21 (Figure 3B). The results of qRT-PCR showed that there was no significant difference of mRNA expression of E2F1 and TCF12, while significant decrease of Gli3 and Stat3 upon exosomal miR-124 injection, indicating the functional regulation of target genes by exosomal miR-124.…”

Synergic Functions of miRNAs Determine Neuronal Fate of Adult Neural Stem Cells

“…The proposed synergy of miRNA actions refers to the “convergence” or “cooperativity” of miRNAs as a rapidly emerging theme in neurobiology, and has recently been proposed for embryonic neurogenesis (Barca-Mayo and De Pietri Tonelli, 2014), the adult SVZ (Santos et al., 2016), and apoptosis in the adult DG (Schouten et al., 2015). Consistent with this idea, we identified 26 putative targets of the 11 miRNAs that did not share immediate involvement in any pathway, but synergistically regulate biological processes such as neurogenesis, nervous system development, and neuronal differentiation.…”