Specific MicroRNAs Modulate Embryonic Stem Cell–Derived Neurogenesis

Harvard University, Cambridge, Massachusetts, United States
Stem Cells (Impact Factor: 6.52). 05/2006; 24(4):857-64. DOI: 10.1634/stemcells.2005-0441
Source: PubMed


MicroRNAs (miRNAs) are recently discovered small non-coding transcripts with a broad spectrum of functions described mostly in invertebrates. As post-transcriptional regulators of gene expression, miRNAs trigger target mRNA degradation or translational repression. Although hundreds of miRNAs have been cloned from a variety of mammalian tissues and cells and multiple mRNA targets have been predicted, little is known about their functions. So far, a role of miRNA has only been described in hematopoietic, adipocytic, and muscle differentiation; regulation of insulin secretion; and potentially regulation of cancer growth. Here, we describe miRNA expression profiling in mouse embryonic stem (ES) cell- derived neurogenesis in vitro and show that a number of miRNAs are simultaneously co-induced during differentiation of neural progenitor cells to neurons and astrocytes. There was a clear correlation between miRNA expression profiles in ES cell-derived neurogenesis in vitro and in embryonal neurogenesis in vivo. Using both gain-of-function and loss-of-function approaches, we demonstrate that brain-specific miR-124a and miR-9 molecules affect neural lineage differentiation in the ES cell-derived cultures. In addition, we provide evidence that signal transducer and activator of transcription (STAT) 3, a member of the STAT family pathway, is involved in the function of these miRNAs. We conclude that distinct miRNAs play a functional role in the determination of neural fates in ES cell differentiation.

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Available from: Kenneth S Kosik
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    • "miR-9 is also encoded by three genes (miR-9-1, miR-9-2, miR-9-3), but in contrast to miR-124, where only one mature miRNA exists, each of the miR-9 genes gives rise to two mature miRNAs, miR-9 (which is the 5' strand, miR-9-5p) and miR-9* (which is the 3' strand, miR-9-3p) (Conaco et al., 2006). Gain of function experiments with miR-124 alone or in combination with miR-9/9* in embryonic stem cells (ESCs) (Krichevsky et al., 2006; Maiorano and Mallamaci, 2009) and NPCs (Cheng et al., 2009; Maiorano and Mallamaci, 2009), show that miR-124 promotes neuronal differentiation, while inhibiting glial genes. Moreover, miR-124-9-9*, has been shown to reprogram fibroblasts (Yoo et al., 2011), and miR-124 alone can increase retinoic acid-induced neurogenesis in P19 stem cells (Makeyev et al., 2007). "
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    ABSTRACT: The Müller glia of fish provide a source for neuronal regeneration after injury, but they do not do so in mammals. We previously showed that lentiviral gene transfer of the transcription factor Achaete-scute homolog 1 (Ascl1/Mash1) in murine Müller glia cultures resulted in partial reprogramming of the cells to retinal progenitors. The microRNAs (miRNAs) miR-124-9-9* facilitate neuronal reprogramming of fibroblasts, but their role in glia reprogramming has not been reported. The aim of this study was to test whether (1) lentiviral gene transfer of miR-124-9-9* can reprogram Müller glia into retinal neurons and (2) miR-124-9-9* can improve Ascl1-induced reprogramming. Primary Müller glia cultures were generated from postnatal day (P) 11/12 mice, transduced with lentiviral particles, i.e., miR-124-9-9*-RFP, nonsense-RFP, Ascl1-GFP, or GFP-control. Gene expression and immunofluorescence analyses were performed within 3 weeks after infection. Overexpression of miR-124-9-9* induced the expression of the proneural factor Ascl1 and additional markers of neurons, including TUJ1 and MAP2. When Ascl1 and miR-124-9-9* were combined, 50 to 60% of Müller glia underwent neuronal reprogramming, whereas Ascl1 alone results in a 30 to 35% reprogramming rate. Analysis of the miR-124-9-9* treated glial cells showed a reduction in the level of Ctdsp1 and Ptbp1, indicating a critical role for the REST pathway in the repression of neuronal genes in Müller glia. Our data further suggest that miR-124-9-9* and the REST complex may play a role in regulating the reprogramming of Müller glia to progenitors that underlies retinal regeneration in zebrafish. GLIA 2016.
    Full-text · Article · Jan 2016 · Glia
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    • "While Ngn1 could both activate the neurogenic program and simultaneously inhibit the astrogliogenic program (Figure 3—figure supplement 1C), miR-9 appeared to be only involved in the inhibition of glial fate. We confirmed that transfection of mouse NPCs with exogenous miR-9 duplex blocked phosphorylation of Stat1/3 without altering their protein levels (Figure 3B), which is in agreement with the work by Krichevsky et al. (2006). To explore how miR-9 inhibits Stat phosphorylation, we scanned the 3′ UTRs of three upstream components of the Jak-Stat pathway ( "
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    ABSTRACT: It has been postulated that a proneural factor, neurogenin 1 (Ngn1), simultaneously activates the neurogenic program and inhibits the alternative astrogliogenic program when specifying the neuronal fate. While Ngn1 substantially suppresses the activation of the astrogliogenic Jak-Stat pathway, the underlying molecular mechanism was unknown. Here, by employing in vivo and in vitro approaches, we report that Ngn1 binds to the promoter of a brain-enriched microRNA, miR-9, and activates its expression during neurogenesis. Subsequently, our in vitro study showed that miR-9 directly targets mRNAs of Lifr-beta, Il6st (gp130), and Jak1 to down-regulate these critical upstream components of the Jak-Stat pathway, achieving inhibition of Stat phosphorylation and consequently, suppression of astrogliogenesis. This study revealed Ngn1 modulated non-coding RNA epigenetic regulation during cell fate specifications.
    Full-text · Article · Aug 2015 · eLife Sciences
    • "Two such miRNAs are miR-9 and miR-124a, which are highly expressed in neurons and astrocytes of the brain. Together, these miRNAs indirectly modulate the phosphorylation status of STAT3, an important intracellular signaling molecule mediating the inhibition of neuronal terminal differentiation (Krichevsky et al. 2006; Delaloy et al. 2010). Inhibition of miR-9 increases STAT3 phosphorylation, resulting in reduced neuronal development , whereas overexpression of miR-9 and miR-124a decreases STAT3 phosphorylation, limiting development of the astrocytic lineage. "
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    ABSTRACT: The field of miRNA biology is relatively young, but its impact on our understanding of the regulation of a wide array of cell functions is far-reaching. The importance of miRNAs in development has become nearly ubiquitous, with miRNAs contributing to development of most cells and organs. Although miRNAs are clearly interwoven into known regulatory networks that control cell development, the specific modalities by which they intersect are often quite distinct and cleverly achieved. The frequently emerging theme of feed-back and feed-forward loops to either counterbalance or reinforce the gene programs that they influence is a common thread. Many of these examples of miRNAs as developmental regulators are presently found in organs with different miRNAs and targets, whereas novel, unexpected themes emerge in the context of mouse development as we learn more about this rapidly developing area of biology. Copyright © 2015 Cold Spring Harbor Laboratory Press; all rights reserved.
    No preview · Article · Jul 2015 · Cold Spring Harbor perspectives in biology
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