Role of alternative splicing factor MBNL1 in pathogenesis of myotonic dystrophy

Myotonic dystrophy (DM) is an RNA-mediated disease caused by a non-coding CTG repeat expansion in the DMPK gene (DM type 1, DM1) or a CCTG expansion in ZNF9 (DM type 2, DM2). Pathogenesis of DM involves dysregulation of alternative splicing factors such as the MBNL1 protein. MBNL1 is sequestered by mutant transcripts containing C(C)UG repeat RNAs which form discrete nuclear RNA foci that, in turn, leads to the loss of MBNL1 function. Because MBNL1 is an RNA binding protein that regulates alternative splicing of a specific subset of pre-mRNAs during postnatal development, loss of this splicing factor due to its sequestration results in missplicing of MBNL1 target pre-mRNAs and perturbation of developmental signals. DM is a systemic disease that affects multiple organs and key features of this disease include myotonia (hyperexcitability of muscle), myopathy (muscle weakness), cardiac conduction defects and subcapsular cataracts. However DM1 is also remarkably variable in severity and penetrance mainly due to somatic mosaicism of CTG repeat size but also due to the variability in genetic background between individuals. We hypothesized that MBNL1 loss of function due to sequestration is a primary pathogenic event in DM and is responsible for disease-associated phenotypes owing to the failure of developmental transitions as a result of mistakes in alternative splicing. In this study, we show that overexpression of Mbnl1 in vivo mediated by transduction of skeletal muscle with a recombinant adeno-associated viral vector is sufficient to rescue myotonia and missplicing in the HSALR poly(CUG) mouse model for DM, suggesting that loss of MBNL1 activity is primarily responsible for disease pathogenesis. We also report that Mbnl1 deficiency leads to defects not only in skeletal muscle but also other organs like thymus and skin in Mbnl1 knockout mice depending on the genetic background. This observation indicates that MBNL1 has a broad role in developmental pathways. Furthermore, using congenic Mbnl1 knockout mice, we provide evidence that functional and structural muscle abnormalities in DM may be separable from myotonia and may be attributed to the altered splicing of genes important for calcium homeostasis, including the ryanodine receptor. Our results suggest a fundamental role of MBNL1 in development and disease and provide a theoretical and experimental basis for the development of novel therapies for this neuromuscular disease.