RNA Dominance in Human Disease

DESCRIPTION (provided by applicant): Myotonic dystrophy (DM) is caused by the expansion of CTG and CCTG microsatellites in the non-coding regions of two different genes, DMPK and ZNF9/CNBP. Upon transcription, these repeat expansions fold into stable RNA hairpin structures that are toxic because they alter the activities of two families of alternative splicing factors, the muscleblind-like (MBNL) and CUGBP/ETR-3-like factor (CELF) proteins. Although this RNA-mediated pathogenesis model accounts for many aspects of adult-onset DM, the molecular events underlying congenital DM (CDM) are less clear since this form of the disease is only associated with very large CTG repeat expansions in the DMPK gene. While searching for factors involved in CDM pathogenesis, we discovered that Mbnl3 expression is restricted to embryonic muscle development and postnatal muscle regeneration. Based on these, and additional, results, this proposal is designed to test the hypothesis that Mbnl3 activity is essential for normal muscle development and regeneration and loss of this activity leads to delayed myogenesis in CDM followed by muscle weakness/wasting in adults. To test this hypothesis, Mbnl3 conditional knockout mice have been generated to test the effects of Mbnl3 loss on the development and regeneration of skeletal muscle. Additionally, RNA targets for Mbnl3 will be identified in both developing and regenerating muscle followed by assays designed to determine how loss of Mbnl3 leads to mis-regulation of these targets and specific pathological outcomes. Since we have discovered that Mbnl3 encodes both nuclear and cytoplasmic proteins, functional differences between these isoforms will be investigated and the hypothesis that MBNL proteins form different types of complexes on CUG versus CCUG expansion RNAs will be tested. Finally, we will generate Mbnl double and triple muscle-specific knockout mice to test our model that myotonic dystrophy is an MBNL loss-of-function disease. The goal of these studies is to understand the molecular events which lead to CDM and adult-onset DM and use this knowledge for the development of novel disease therapies.