PUBLICATION
Rbfox-regulated alternative splicing is critical for zebrafish cardiac and skeletal muscle functions
- Authors
- Gallagher, T.L., Arribere, J.A., Geurts, P.A., Exner, C.R., McDonald, K.L., Dill, K.K., Marr, H.L., Adkar, S.S., Garnett, A.T., Amacher, S.L., and Conboy, J.G.
- ID
- ZDB-PUB-110921-28
- Date
- 2011
- Source
- Developmental Biology 359(2): 251-61 (Journal)
- Registered Authors
- Amacher, Sharon, Dill, Kariena, Garnett, Aaron
- Keywords
- alternative splicing, zebrafish, muscle, rbfox, a2bp1l, rbm9
- MeSH Terms
-
- Alternative Splicing*
- Animals
- Animals, Genetically Modified
- Base Sequence
- Embryo, Nonmammalian/embryology
- Embryo, Nonmammalian/metabolism
- Embryo, Nonmammalian/ultrastructure
- Female
- Gene Expression Regulation, Developmental
- Gene Knockdown Techniques
- Green Fluorescent Proteins/genetics
- Green Fluorescent Proteins/metabolism
- Green Fluorescent Proteins/ultrastructure
- Heart/embryology
- Heart/physiology*
- Immunohistochemistry
- In Situ Hybridization
- Male
- Microscopy, Confocal
- Microscopy, Electron
- Molecular Sequence Data
- Muscle, Skeletal/embryology
- Muscle, Skeletal/metabolism
- Muscle, Skeletal/physiology*
- Myocardium/metabolism
- Protein Isoforms/genetics
- Protein Isoforms/metabolism
- RNA-Binding Proteins/genetics
- RNA-Binding Proteins/metabolism
- RNA-Binding Proteins/physiology*
- Reverse Transcriptase Polymerase Chain Reaction
- Zebrafish/embryology
- Zebrafish/genetics
- Zebrafish/physiology*
- Zebrafish Proteins/genetics
- Zebrafish Proteins/metabolism
- Zebrafish Proteins/physiology*
- PubMed
- 21925157 Full text @ Dev. Biol.
Citation
Gallagher, T.L., Arribere, J.A., Geurts, P.A., Exner, C.R., McDonald, K.L., Dill, K.K., Marr, H.L., Adkar, S.S., Garnett, A.T., Amacher, S.L., and Conboy, J.G. (2011) Rbfox-regulated alternative splicing is critical for zebrafish cardiac and skeletal muscle functions. Developmental Biology. 359(2):251-61.
Abstract
Rbfox RNA binding proteins are implicated as regulators of phylogenetically-conserved alternative splicing events important for muscle function. To investigate the function of rbfox genes, we used morpholino-mediated knockdown of muscle-expressed rbfox1l and rbfox2 in zebrafish embryos. Single and double morphant embryos exhibited changes in splicing of overlapping sets of bioinformatically-predicted rbfox target exons, many of which exhibit a muscle-enriched splicing pattern that is conserved in vertebrates. Thus, conservation of intronic Rbfox binding motifs is a good predictor of Rbfox-regulated alternative splicing. Morphology and development of single morphant embryos were strikingly normal; however, muscle development in double morphants was severely disrupted. Defects in cardiac muscle were marked by reduced heart rate and in skeletal muscle by complete paralysis. The predominance of wavy myofibers and abnormal thick and thin filaments in skeletal muscle revealed that myofibril assembly is defective and disorganized in double morphants. Ultra-structural analysis revealed that although sarcomeres with electron dense M- and Z-bands are present in muscle fibers of rbfox1l/rbox2 morphants, they are substantially reduced in number and alignment. Importantly, splicing changes and morphological defects were rescued by expression of morpholino-resistant rbfox cDNA. Additionally, a target-blocking MO complementary to a single UGCAUG motif adjacent to an rbfox target exon of fxr1 inhibited inclusion in a similar manner to rbfox knockdown, providing evidence that Rbfox regulates the splicing of target exons via direct binding to intronic regulatory motifs. We conclude that Rbfox proteins regulate an alternative splicing program essential for vertebrate heart and skeletal muscle functions.
Genes / Markers
Expression
Phenotype
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Fish
Orthology
Engineered Foreign Genes
Mapping