G.P.11: SMCHD1 mutations cause FSHD type 2 and act as modifiers of disease severity

Abstract

Facioscapulohumeral muscular dystrophy (FSHD) is an autosomal dominant muscular disorder of wide clinical variability. Contractions of the D4Z4 macro-satellite repeat on the permissive chromosome 4qA haplotype are the molecular basis of the disease. D4Z4 repeats with less than 11 copies are associated with chromatin decondensation and hypomethylation allowing for expression of the flanking DUX4 gene which is highly cytotoxic and causes skeletal muscle cell death. Recently, mutations in the SMCHD1 gene have been identified in a subset of patients without D4Z4 repeat contractions (FSHD2). Haploinsufficiency of the SMCHD1 protein results in hypomethylation of D4Z4 and a more open chromatin structure permitting activation of DUX4. We have screened phenotypic FSHD patients from 95 unrelated families for mutations in SMCHD1 by next generation sequencing (NGS). We identified mutations (deletions, missense, splice site and nonsense) in nine index patients without D4Z4 contractions. We developed a pyrosequencing assay to determine the methylation status of the D4Z4 repeat array and found significantly lower methylation levels for FSHD2 patients than for healthy controls and FSHD1 patients. SMCHD1 mutations were also found in two patients with moderately contracted D4Z4 alleles thus suffering from FSHD1 and 2. Comparing the phenotype of patients, all FSHD2 patients were relatively mildly affected while patients with FSHD1+2 were much more severely affected than expected from their D4Z4 copy number. Our findings confirm the digenic inheritance of FSHD2 with mutations in SMCHD1 and hypomethylation of D4Z4. In cases of “double trouble” with FSHD1 and 2, SMCHD1 mutations seem to act as a modifier of disease severity. With SMCHD1 mutations found in 16.4% of phenotypic FSHD patients, FSHD2 cannot be considered to be excessively rare. Hence, we suggest including sequencing of SMCHD1, haplotyping and methylation analysis in the workflow of molecular FSHD diagnostics.