Identification of Genetic and Molecular Pathways in Congenital Rare Disorders Affecting the Brain and Muscle

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ABSTRACT Congenital muscular dystrophies (CMDs) are a group of heterogeneous pediatric disorders leading to motor and developmental delay, and childhood mortality. CMDs have variable presentation often affecting multiple organs, such as the eyes and brain. While this clinical heterogeneity initially hindered genetic analyses, the ad- vent of next-generation sequencing and whole exome studies greatly increased gene identification reaching up to 30 genes, with eight genes identified in 2012-2013, three of which by the PI. Each new gene has provided a different piece of a complex puzzle, not only providing a genetic diagnosis for the affected individuals, but also informing us on how the mutated genes converge onto shared molecular pathways such as protein glycosyla- tion. Yet, our group and others have found that mutations in each gene are only present in a small portion of cases, and 30-40% of cases remain unexplained. In parallel, the large number of known genes has hindered therapy development as it remains unclear how cases with different genotypes and phenotypes can be grouped for treatment. The PI has focused the past decade on studying the genetics of CMD and developing zebrafish models to define how disease-causing mutations affect muscle and brain development. Through these studies, we have devel- oped our central hypotheses that most CMD genes regulate interactions with the extracellular matrix (ECM) through glycosylation, and that unexplained cases will either carry mutations in novel genes involved in cell- ECM interaction or noncoding variants in already known CMD genes. The proposed studies will test these hypotheses through two independent and complementary Specific Aims. Specific Aim 1 will leverage the multi- ple zebrafish models we have developed for known CMD genes to test whether increasing glycosylation will restore cell-ECM interactions in the muscle and brain in different genetic models of CMD. We will 1) define which biochemical deficits are shared by different genetic mutations in muscle cells and neurons, and 2) test whether changes in glycosylation can be beneficial in different forms of CMD. This workflow can then be ex- panded to future therapeutic interventions and novel disease genes can be rapidly taken from gene identifica- tion to therapy evaluation. Specific Aim 2 will close the gap in gene discovery in CMDs by testing the hypothe- sis that undiagnosed CMD cases are caused by a combination of rare mutations in novel genes and non- coding mutations in intronic/regulatory regions of known CMD genes. We have developed a next generation sequencing and bioinformatic pipeline that will integrate data from exome, genome, and transcriptome to iden- tify coding, splicing, and regulatory variants to fully unravel the genetics of CMD. These studies will directly impact the CMD fields by both discovering how different disease genes contribute to pathogenesis and by developing novel genetic tests for the global patient population.