524V Poster Online - Virtual Posters
Wednesday April 06, 4:00 PM - 7:00 PM

Authors:
Elena Nikonova ¹; Marc Canela Grimau ²; Tobias Straub ¹; Maria Spletter ¹

Affiliations:
1) Ludwig-Maximilians-University Munich; 2) University of Barcelona

Keywords:
g. alternative splicing; j. muscle

Animals have different types of muscle fibers with distinct morphologies and contractile properties. These characteristics are established during development through regulation of alternative splicing and gene expression. Patterns of mRNA isoform expression differ between myofiber types, change as muscles differentiate, mature and age, and are often disrupted in muscle disease, suggesting that RNA regulation and in particular alternative splicing helps define muscle functional properties. It is therefore important to examine how RNA processing functions in healthy muscle, to better understand how misregulation contributes to muscle disease. Misregulation of the RNA-binding protein CELF1 is thought to underly myotonic dystrophy (DM1), but its function in muscle development is incompletely understood. Using Drosophila melanogaster as a tractable genetic model, we have previously shown using RNAi knockdown that the CELF1 homologue Bruno-1 (Bru1) controls flight muscle specific alternative splicing, regulating both sarcomere growth and myosin contractility. We generated new CRISPR-mediated alleles in Bru1 that affect all Bru1 isoforms, resulting in stronger phenotypes in mutant flies that revealed structural defects at the earliest stages of myofibril formation, notably disorganization of the actin cytoskeleton that adversely affects myofibrillogenesis in differentiating IFM. Using temporally-restricted RNAi knockdown and rescue experiments, we demonstrate that there are distinct requirements for Bru1 mediated splicing during early and late stages of myofibril formation. After sarcomere formation, aberrant actin incorporation arrests growth in thin-filament length, but promotes lateral growth leading to formation of hollow myofibrils. We performed mRNA-Seq and mass spectrometry and identified misregulation of both sarcomeric proteins and other RNA-processing factors. Moreover, our temporal transcriptomics data reveal a progressive misregulation of gene expression and splicing as IFM development proceeds. Taken together, our data indicate that during muscle differentiation, Bru1 regulates cytoskeletal rearrangement necessary for myofibril formation as well as the balance in length versus lateral growth of the thin-filament. Defective RNA processing thus causes sarcomeric structural defects and progressive malfunction of dystrophic muscle.