Abstract
Protein isoform transitions confer muscle fibers with distinct properties and are regulated by differential transcription and alternative splicing. RNA-binding Fox protein 1 (Rbfox1) can affect both transcript levels and splicing, and is known to contribute to normal muscle development and physiology in vertebrates, although the detailed mechanisms remain obscure. In this study, we report that Rbfox1 contributes to the generation of adult muscle diversity in Drosophila Rbfox1 is differentially expressed among muscle fiber types, and RNAi knockdown causes a hypercontraction phenotype that leads to behavioral and eclosion defects. Misregulation of fiber type-specific gene and splice isoform expression, notably loss of an indirect flight muscle-specific isoform of Troponin-I that is critical for regulating myosin activity, leads to structural defects. We further show that Rbfox1 directly binds the 3'-UTR of target transcripts, regulates the expression level of myogenic transcription factors myocyte enhancer factor 2 and Salm, and both modulates expression of and genetically interacts with the CELF family RNA-binding protein Bruno1 (Bru1). Rbfox1 and Bru1 co-regulate fiber type-specific alternative splicing of structural genes, indicating that regulatory interactions between FOX and CELF family RNA-binding proteins are conserved in fly muscle. Rbfox1 thus affects muscle development by regulating fiber type-specific splicing and expression dynamics of identity genes and structural proteins.
Highlights
Muscles are an ideal model to understand the strategies involved in the generation of diversity within a tissue, as they are developmentally patterned with distinct morphologies and diverse contractile properties (Spletter & Schnorrer, 2014)
We found that Myocyte enhancer factor 2 (Mef2)-Gal4 driven knockdown with RNA-binding Fox protein 1 (Rbfox1)-IRKK110518 was pupal lethal, and larval lethal when driven with Act5c-Gal4, which expresses in all cells, or when combined with UAS-Dicer2 (Dcr2), which is reported to increase the efficiency of RNAi knockdown (Dietzl et al, 2007) (Fig 2A and B)
We show that Rbfox1 functions in a fiber type–specific manner to modulate both fibrillar and tubular muscle development
Summary
Muscles are an ideal model to understand the strategies involved in the generation of diversity within a tissue, as they are developmentally patterned with distinct morphologies and diverse contractile properties (Spletter & Schnorrer, 2014). Composite muscle fiber profiles are a result of coordinated regulation of gene expression (Firulli & Olson, 1997; Black & Olson, 1998; Majesky, 2007), co-integrated with protein isoform transitions facilitated by alternative splicing (Smith et al, 1989; Guo et al, 2010; Kalsotra & Cooper, 2011; Nikonova et al, 2020), accompanied by post-translational modifications (Anthony et al, 2002; Michele & Campbell, 2003; Wells et al, 2003; Nayak & Amrute-Nayak, 2020). It is critically important to understand RBP function in detail
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