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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    Full-text · Article · Jan 2015 · Journal of Cell Science
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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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    • "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.
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