Zebrafish mbnl mutants model molecular and physical phenotypes of human MBNL loss of function disorders

Abstract The muscleblind RNA binding proteins (MBNL1, MBNL2, and MBNL3) are highly conserved across vertebrate species and are important regulators of alternative splicing and other aspects of RNA metabolism. Loss of MBNL protein function through sequestration by CUG or CCUG RNA repeats is partially responsible for the phenotypes of two important human genetic disorders, myotonic dystrophy (DM) and Fuchs endothelial corneal dystrophy (FECD). We generated the first stable zebrafish (Danio rerio) models of MBNL loss of function disorders through CRISPR-based mutation of the three zebrafish mbnl genes. In contrast to mouse models, zebrafish double and triple homozygous mbnl mutants were viable to adulthood. Zebrafish mbnl mutants displayed many DM- and FECD-associated alternative splicing changes in disease relevant tissues. In general, mbnl1 and mbnl2 mutants displayed more pronounced alternative splicing defects than mbnl3 mutants, and compound mbnl mutants exhibited larger splicing changes than single mbnl mutants. Splicing changes in zebrafish mbnl mutants were greatest in adult heart and skeletal muscle. Zebrafish mbnl mutants displayed disease relevant physical phenotypes including decreased sized and impaired movement. The high fecundity and larval optical transparency of these new zebrafish models will make them useful for studying DM- and FECD- related phenotypes and how individual Mbnl proteins contribute to them. Summary Statement Zebrafish mbnl mutants that were generated to model myotonic dystrophy and Fuchs endothelial corneal dystrophy exhibited disease-relevant phenotypes including alternative splicing changes, decreased body size, and impaired movement.