Neurogenesis is a tightly regulated process in time and space both in the developing embryo and in adult neurogenic niches. A drastic change in the transcriptome and proteome of radial glial cells or neural stem cells towards the neuronal state is achieved due to sophisticated mechanisms of epigenetic, transcriptional, and post-transcriptional regulation. Understanding these neurogenic mechanisms is of major importance, not only for shedding light on very complex and crucial developmental processes, but also for the identification of putative reprogramming factors, that harbor hierarchically central regulatory roles in the course of neurogenesis and bare thus the capacity to drive direct reprogramming towards the neuronal fate. The major transcriptional programs that orchestrate the neurogenic process have been the focus of research for many years and key neurogenic transcription factors, as well as repressor complexes, have been identified and employed in direct reprogramming protocols to convert non-neuronal cells, into functional neurons. The post-transcriptional regulation of gene expression during nervous system development has emerged as another important and intricate regulatory layer, strongly contributing to the complexity of the mechanisms controlling neurogenesis and neuronal function. In particular, recent advances are highlighting the importance of specific RNA binding proteins that control major steps of mRNA life cycle during neurogenesis, such as alternative splicing, polyadenylation, stability, and translation. Apart from the RNA binding proteins, microRNAs, a class of small non-coding RNAs that block the translation of their target mRNAs, have also been shown to play crucial roles in all the stages of the neurogenic process, from neural stem/progenitor cell proliferation, neuronal differentiation and migration, to functional maturation. Here, we provide an overview of the most prominent post-transcriptional mechanisms mediated by RNA binding proteins and microRNAs during the neurogenic process, giving particular emphasis on the interplay of specific RNA binding proteins with neurogenic microRNAs. Taking under consideration that the molecular mechanisms of neurogenesis exert high similarity to the ones driving direct neuronal reprogramming, we also discuss the current advances in in vitro and in vivo direct neuronal reprogramming approaches that have employed microRNAs or RNA binding proteins as reprogramming factors, highlighting the so far known mechanisms of their reprogramming action.

The miRNA miR-124 has been employed supplementary to neurogenic TFs and other miRNAs to enhance direct neurogenic conversion by suppressing multiple non-neuronal targets. Aim of the study was to investigate whether miR-124 is sufficient to drive direct reprogramming of astrocytes to induced-neurons (iNs) on its own and elucidate its independent mechanism of reprogramming action. Our data show that miR-124 is a potent driver of the reprogramming switch of astrocytes towards an immature neuronal fate, by directly targeting the RNA-binding protein Zfp36l1 implicated in ARE-mediated mRNA decay and subsequently de-repressing Zfp36l1 neurogenic interactome. To this end miR-124 contribution in iNs’ production largely recapitulates endogenous neurogenesis pathways, being further enhanced upon addition of the neurogenic compound ISX9, which greatly improves both miR-124-induced reprogramming efficiency and iNs’ functional maturation. Importantly, miR-124 is potent to guide direct conversion of reactive astrocytes to immature iNs of cortical identity in vivo following cortical trauma, confirming its ‘master’ reprogramming capacity within the injured cortical microenvironment, while ISX9 supplementation confers a survival advantage to newly produced iNs.

The microRNA miR-124 promotes neuronal identity by acting globally, through its multiple targets, at the epigenetic, transcriptional and post-transcriptional levels. We have previously shown that miR-124 acts as a potent driver of the astrocytic fate switch toward an immature neuronal identity, while its supplementation with the neurogenic compound ISX9 enhances in vitro neuronal maturation. Nevertheless, additional cues are needed to enhance the in vivo neurogenic reprogramming capacity of miR-124/ISX9 following neurodegeneration or neurotrauma. In this study, we constructed the core transcriptional regulatory network regulated by miR-124 and ISX9 during astrocyte-to-neuron conversion. Our analysis revealed that the DNA demethylase TET1 is a pivotal transcriptional regulator of the miR-124/ISX9 neurogenic reprogramming process. Silencing of Tet1 impaired the miR-124-mediated neuronal conversion of astrocytes, as well as the ISX9-reinforced differentiation of iNs. We also identified the DNA/RNA binding protein LIN28A as the top mediator of ISX9 neurogenic action and provide evidence that it acts as a coregulator of the expression of synaptic genes, along with TET1. Taken together, our data suggest that TET1 and LIN28A are potent candidates for amplifying miR-124/ISX9 combined in vivo reprogramming action and enhance iNs’ differentiation state.