Global regulation of alternative splicing during myogenic differentiation

Department of Pathology and Immunology, Interdepartmental Program in Cell and Molecular Biology, Baylor College of Medicine, Houston, TX 77030, USA.
Nucleic Acids Research (Impact Factor: 9.11). 11/2010; 38(21):7651-64. DOI: 10.1093/nar/gkq614
Source: PubMed


Recent genome-wide analyses have elucidated the extent of alternative splicing (AS) in mammals, often focusing on comparisons of splice isoforms between differentiated tissues. However, regulated splicing changes are likely to be important in biological transitions such as cellular differentiation, or response to environmental stimuli. To assess the extent and significance of AS in myogenesis, we used splicing-sensitive microarray analysis of differentiating C2C12 myoblasts. We identified 95 AS events that undergo robust splicing transitions during C2C12 differentiation. More than half of the splicing transitions are conserved during differentiation of avian myoblasts, suggesting the products and timing of transitions are functionally significant. The majority of splicing transitions during C2C12 differentiation fall into four temporal patterns and were dependent on the myogenic program, suggesting that they are integral components of myogenic differentiation. Computational analyses revealed enrichment of many sequence motifs within the upstream and downstream intronic regions near the alternatively spliced regions corresponding to binding sites of splicing regulators. Western analyses demonstrated that several splicing regulators undergo dynamic changes in nuclear abundance during differentiation. These findings show that within a developmental context, AS is a highly regulated and conserved process, suggesting a major role for AS regulation in myogenic differentiation.

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    • "During muscle differentiation, myoblasts give rise to multinucleated myotubes through a temporally ordered series of changes in alternative splicing that generates alternate isoforms of key myogenic factors (Bland et al., 2010; Trapnell et al., 2010). Several candidate ciselements (Sugnet et al., 2006; Bland et al., 2010) and trans-acting splicing factors involved in regulating muscle-specific alternative splicing have been described. These include Mbnl (Pascual et al., 2006), Rbfox (Kuroyanagi, 2009), Celf (Dasgupta and Ladd, 2012) and Ptb families of proteins (Romanelli et al., 2013). "
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    ABSTRACT: Among Mef2 family of transcription factors, Mef2D is unique in that it undergoes tissue-specific splicing to generate an isoform essential for muscle differentiation. However, the mechanisms mediating this muscle-specific processing of Mef2D remain unknown. Using bioinformatics, we identified Rbfox proteins as putative modulators of Mef2D muscle-specific splicing. Accordingly, we found direct and specific Rbfox1 and 2 binding to Mef2D pre-mRNA in vivo. Gain and loss of function experiments demonstrated that Rbfox1 and 2 cooperate in promoting Mef2D splicing and subsequent myogenesis. Thus, our findings reveal a novel role for Rbfox1/2 proteins in regulating myogenesis through activation of essential muscle-specific splicing events.
    Journal of Cell Science 01/2015; 128(4). DOI:10.1242/jcs.161059 · 5.43 Impact Factor
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    • "Motif enrichments were calculated using 100 bp of the flanking exons and the complete sequence of the cassette exons. For introns, we used maximum intronic flanks of 250 nt, removing SS context to avoid BP, SS and PPT signals, 9 nt at donor side and 30 nt at the acceptor side (Bland et al, 2010 "
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    ABSTRACT: Matrin3 is an RNA- and DNA-binding nuclear matrix protein found to be associated with neural and muscular degenerative diseases. A number of possible functions of Matrin3 have been suggested, but no widespread role in RNA metabolism has yet been clearly demonstrated. We identified Matrin3 by its interaction with the second RRM domain of the splicing regulator PTB. Using a combination of RNAi knockdown, transcriptome profiling and iCLIP, we find that Matrin3 is a regulator of hundreds of alternative splicing events, principally acting as a splicing repressor with only a small proportion of targeted events being co-regulated by PTB. In contrast to other splicing regulators, Matrin3 binds to an extended region within repressed exons and flanking introns with no sharply defined peaks. The identification of this clear molecular function of Matrin3 should help to clarify the molecular pathology of ALS and other diseases caused by mutations of Matrin3. © 2015 The Authors. Published under the terms of the CC BY 4.0 license.
    The EMBO Journal 01/2015; 34(5). DOI:10.15252/embj.201489852 · 10.43 Impact Factor
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    • "In contrast, only 6% of co-regulation was observed on a different set of 47 genes implicated in cancer [65]. While MBNL1 and RBFOX1 have been individually implicated in modulating splicing decisions during muscle and heart development [2], [66], our results suggest that MBNL1 and RBFOX proteins converge to regulate the splicing of a common subset of genes involved in muscle function. Recent work suggests that MBNL1 and RBFOX2 also cooperate to implement a splicing program associated with the differentiation of human stem cells [67]. "
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    ABSTRACT: With the goal of identifying splicing alterations in myotonic dystrophy 1 (DM1) tissues that may yield insights into targets or mechanisms, we have surveyed mis-splicing events in three systems using a RT-PCR screening and validation platform. First, a transgenic mouse model expressing CUG-repeats identified splicing alterations shared with other mouse models of DM1. Second, using cell cultures from human embryonic muscle, we noted that DM1-associated splicing alterations were significantly enriched in cytoskeleton (e.g. SORBS1, TACC2, TTN, ACTN1 and DMD) and channel (e.g. KCND3 and TRPM4) genes. Third, of the splicing alterations occurring in adult DM1 tissues, one produced a dominant negative variant of the splicing regulator RBFOX1. Notably, half of the splicing events controlled by MBNL1 were co-regulated by RBFOX1, and several events in this category were mis-spliced in DM1 tissues. Our results suggest that reduced RBFOX1 activity in DM1 tissues may amplify several of the splicing alterations caused by the deficiency in MBNL1.
    PLoS ONE 09/2014; 9(9):e107324. DOI:10.1371/journal.pone.0107324 · 3.23 Impact Factor
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