Molecular Basis of Type 2 Polysaccharide Storage Myopathy

Polysaccharide Storage Myopathy (PSSM) in horses is characterized by accumulation of excess and abnormal skeletal muscle glycogen and rhadbomyolysis. A mutation in GYS1 causes one form of PSSM (PSSM1); however, a significant portion of PSSM cannot be attributed to GYS1. Evidence points to a second heritable form of PSSM (PSSM2); however the mutation responsible is unknown. GWAS was performed in 104 PSSM2 Quarter Horse (QH) cases and 124 QH controls, genotyped with the Equine SNP50 Beadchip.  Association tests identified a 350 kb region highly associated with PSSM2 that encompasses 2 annotated genes and a novel-protein coding mRNA. Attempts to sequence key functional elements in the region by Sanger sequence were not entirely successful and were hampered by apparent poor annotation and assembly, segments of highly repetitive sequence, and the likely presence of a large structural variant(s). Therefore we have evaluated sequence in this region using next-generation sequencing (NGS) and de novo assembly of BAC clones and long-range PCR amplicons from 11 PSSM2 cases and 11 controls, as well as whole genome NGS of 3 PSSM2 cases and 3 controls (100 bp paired-end, 12x coverage). PCR amplicons and whole genome reads were mapped to EquCab2.0 without discovering significant differences between cases and controls. Thus a de novo assembly was created from BAC clone which contains 233kb through the promoter and 5’ UTR of one gene of interest and encompasses 4 gaps present in the reference genome. Reads are being mapped to the new contig to identify variants in the region.