Date of Award
Doctor of Philosophy (PhD)
The ectopic expression of two brain-enriched microRNAs (miRNAs), miR-9/9* and miR-124 (miR-9/9*-124), can robustly and efficiently reprogram human skin fibroblasts into neurons. The miRNAs act as repressors of non-neuronal genes in fibroblasts for the induction of the neuronal program. This process is analogous to neurogenesis in vivo when the expression of miR-9/9* and miR-124 represses anti-neurogenic genes such as REST or NRSF (neuron-restrictive silencer factor/repressor element-1 silencing transcription factor). Although we have some mechanistic insights into how miR-9/9*-124 drives fate conversion by acting as negative regulators of gene expression, little remained understood of the role of miRNAs as positive regulators of gene expression for the activation of neuronal fate. In this thesis, we present our current understanding of how miR-9/9*-124 drives neuronal reprogramming as well as the mechanistic insights into how miR-9/9*-124 can promote expression of neuronal genes. Based on Argonuate (AGO) HITS-CLIP, we uncovered that AGO is bound to neuronal transcripts that are progressively upregulated during reprogramming. Such observation suggests that contrary to the canonical repressive roles of miRNAs, miR-9/9*-124 may be playing a positive role in the expression of bound neuronal transcripts. Using PTB family of RNA-binding proteins as an example, we delineate a mechanism by which miR-124 can simultaneously repress PTBP1, the non-neuronal PTB homolog, while promoting the upregulation of PTBP2, the neuronal PTB homolog, in neurons. This process requires the synergism of miRNA targeting as well as a family of neuronal ELAVL proteins (nELAVLs). We further showed that this miRNA- and nELAVL-mediated upregulation of PTBP2 is neither unique to the conversion process nor PTBP2 transcript alone, but also in primary human neurons and is likely a mechanism used to regulate other neuronal transcripts. With PTBP2 expression induced in neurons, PTBP2 is involved in the alternative splicing of numerous neuronal transcripts. Such transcript includes a subunit of the chromatin remodeling complex, DPF1. Although detailed function of DPF1 remains unclear in neurons, different PTBP2-mediated DPF1 spliced isoforms are likely involved in the interaction of different modified histones for the establishment of a neuronal chromatin landscape.
Chair and Committee
Andrew S. Yoo
Shiming Chen, Harrison Gabel, Shin-ichro Imai, Kristen Kroll,
Lu, Ya-Lin, "Delineating the Role of MiR-124 for the Activation of Neuronal Program" (2020). Arts & Sciences Electronic Theses and Dissertations. 2329.