RNA Interference Screening for Genes Regulating Drosophila Muscle Morphogenesis

Kaya-Çopur, Aynur; Schnorrer, Frank

doi:10.1007/978-1-4939-8897-6_20

Cited by 11 publications

(8 citation statements)

References 52 publications

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“…We have previously reported a similar variation in phenotypic strength after knockdown with multiple RNAi hairpins targeting other RBPs, notably SF1 , Hrb87F , snf , Prp19 , and others ( Kao et al, 2021 ). These results illustrate the experimental importance of testing multiple, independent RNAi constructs to distinguish hypomorphic from null phenotypes, beyond merely addressing off-target effects ( Mohr & Perrimon, 2012 ; Kaya-Çopur & Schnorrer, 2019 ; Neumeier & Meister, 2020 ). Moreover, tuning the level of knockdown can offer insight into regulatory events that are sensitive to RBP activity level.…”

Section: Discussionmentioning

confidence: 79%

Rbfox1 is required for myofibril development and maintaining fiber type–specific isoform expression in Drosophila muscles

et al. 2022

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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.

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Section: Discussionmentioning

confidence: 79%

Rbfox1 is required for myofibril development and maintaining fiber type–specific isoform expression in Drosophila muscles

et al. 2022

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“…To bypass this early larval lethality, we expressed each DmCFL RNAi construct with the muscle Gal4 line, Mhc-Gal4, that drives expression later in embryogenesis and at lower levels relative to Dmef2-Gal4 ( Kaya-Çopur and Schnorrer, 2019 ; Viswanathan et al, 2015 ; Zhang and Bernstein, 2001 ). At the 3 rd -instar stage, DmCFL RNAi (1) expressing larvae showed slower locomotion, disrupted muscle structure with Zasp-containing aggregates, and death after the larval stages ( Figures 1G , 1H , and S1F ).…”

Section: Resultsmentioning

confidence: 99%

Cofilin Loss in Drosophila Muscles Contributes to Muscle Weakness through Defective Sarcomerogenesis during Muscle Growth

Balakrishnan¹,

Yu²,

Chin³

et al. 2020

Cell Reports

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SUMMARY Sarcomeres, the fundamental contractile units of muscles, are conserved structures composed of actin thin filaments and myosin thick filaments. How sarcomeres are formed and maintained is not well understood. Here, we show that knockdown of Drosophila cofilin ( DmCFL ), an actin depolymerizing factor, disrupts both sarcomere structure and muscle function. The loss of DmCFL also results in the formation of sarcomeric protein aggregates and impairs sarcomere addition during growth. The activation of the proteasome delays muscle deterioration in our model. Furthermore, we investigate how a point mutation in CFL2 that causes nemaline myopathy (NM) in humans affects CFL function and leads to the muscle phenotypes observed in vivo . Our data provide significant insights to the role of CFLs during sarcomere formation, as well as mechanistic implications for disease progression in NM patients.

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“…Disease models in the fly have proven highly informative for many genes implicated in human muscle disease, for example CELF1, MBNL1, SMN (Survival motor neuron) and DMD (Dystrophin) [42][43][44]. Flies have a particularly diverse and powerful genetic toolbox that can be employed in vivo, including well-established tools to regulate spatial and temporal expression and to fine-tune levels of both RNAi-mediated knockdown and transgene expression [45][46][47]. These characteristics make the fly an attractive model to both screen for RBPs with muscle phenotypes as well as to investigate conserved molecular mechanisms of RBP function.…”

Section: Drosophila As a Model To Identify And Study Muscle-specific Rbp Functionmentioning

confidence: 99%

A Candidate RNAi Screen Reveals Diverse RNA-Binding Protein Phenotypes in Drosophila Flight Muscle

Kao

Nikonova

Chaabane

et al. 2021

Cells

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The proper regulation of RNA processing is critical for muscle development and the fine-tuning of contractile ability among muscle fiber-types. RNA binding proteins (RBPs) regulate the diverse steps in RNA processing, including alternative splicing, which generates fiber-type specific isoforms of structural proteins that confer contractile sarcomeres with distinct biomechanical properties. Alternative splicing is disrupted in muscle diseases such as myotonic dystrophy and dilated cardiomyopathy and is altered after intense exercise as well as with aging. It is therefore important to understand splicing and RBP function, but currently, only a small fraction of the hundreds of annotated RBPs expressed in muscle have been characterized. Here, we demonstrate the utility of Drosophila as a genetic model system to investigate basic developmental mechanisms of RBP function in myogenesis. We find that RBPs exhibit dynamic temporal and fiber-type specific expression patterns in mRNA-Seq data and display muscle-specific phenotypes. We performed knockdown with 105 RNAi hairpins targeting 35 RBPs and report associated lethality, flight, myofiber and sarcomere defects, including flight muscle phenotypes for Doa, Rm62, mub, mbl, sbr, and clu. Knockdown phenotypes of spliceosome components, as highlighted by phenotypes for A-complex components SF1 and Hrb87F (hnRNPA1), revealed level- and temporal-dependent myofibril defects. We further show that splicing mediated by SF1 and Hrb87F is necessary for Z-disc stability and proper myofibril development, and strong knockdown of either gene results in impaired localization of kettin to the Z-disc. Our results expand the number of RBPs with a described phenotype in muscle and underscore the diversity in myofibril and transcriptomic phenotypes associated with splicing defects. Drosophila is thus a powerful model to gain disease-relevant insight into cellular and molecular phenotypes observed when expression levels of splicing factors, spliceosome components and splicing dynamics are altered.

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RNA Interference Screening for Genes Regulating Drosophila Muscle Morphogenesis

Cited by 11 publications

References 52 publications

Rbfox1 is required for myofibril development and maintaining fiber type–specific isoform expression in Drosophila muscles

Rbfox1 is required for myofibril development and maintaining fiber type–specific isoform expression in Drosophila muscles

Cofilin Loss in Drosophila Muscles Contributes to Muscle Weakness through Defective Sarcomerogenesis during Muscle Growth

A Candidate RNAi Screen Reveals Diverse RNA-Binding Protein Phenotypes in Drosophila Flight Muscle

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