Identification of 45 novel mutations in the nebulin gene associated with autosomal recessive nemaline myopathy

Abstract

Nemaline myopathy (NM) is a clinically and genetically heterogeneous disorder of skeletal muscle caused by mutations in at least five different genes encoding thin filament proteins of the striated muscle sarcomere. We have previously described 18 different mutations in the last 42 exons of the nebulin gene (NEB) in 18 families with NM. Here we report 45 novel NEB mutations detected by denaturing high-performance liquid chromatography (dHPLC) and sequence analysis of all 183 NEB exons in NM patients from 44 families. Altogether we have identified, including the deletion of exon 55 identified in the Ashkenazi Jewish population, 64 different mutations in NEB segregating with autosomal recessive NM in 55 families. The majority (55%) of the mutations in NEB are frameshift or nonsense mutations predicted to cause premature truncation of nebulin. Point mutations (25%) or deletions (3%) affecting conserved splice signals are predicted in the majority of cases to cause in-frame exon skipping, possibly leading to impaired nebulin-tropomyosin interaction along the thin filament. Patients in 18 families had one of nine missense mutations (14%) affecting conserved amino acids at or in the vicinity of actin or tropomyosin binding sites. In addition, we found the exon 55 deletion in four families. The majority of the patients (in 49/55 families) were shown to be compound heterozygous for two different mutations. The mutations were found in both constitutively and alternatively expressed exons throughout the NEB gene, and there were no obvious mutational hotspots. Patients with more severe clinical pictures tended to have mutations predicted to be more disruptive than patients with milder forms.

Nebulin (NEB) mutations in a childhood onset distal myopathy with rods and cores uncovered by next generation sequencing

“…This in-frame deletion predicted to result in a reduced protein molecular mass, was initially identified as a founder mutation in the Ashkenazi population but has also recently identified in other populations with less welldefined ancestry. 10,19 These findings demonstrate the strength of targeted NGS as a powerful technique for the detection of point mutations as well as for larger copy number changes.…”

Reduced thin filament length in nebulin-knockout skeletal muscle alters isometric contractile properties

“…In addition, NEB is required for normal muscle contractility, which has been demonstrated by altered Ca 2ϩ sensitivity (55) and inferior isometric stress production (3) in NEB-deficient muscle. NEB is of clinical interest because 50% or more of nemaline myopathy cases are caused by mutations in its gene (21,35,48,49).To assess the role of NEB, NEB knockout (NEB-KO) mice were compared with wild-type (WT) mice. NEB deficiency is a neonatal lethal mutation; NEB-KO mice have abnormally short thin filaments, exhibit rapid postnatal myofibrillar disorganization and muscle degeneration, and die after ϳ1-2 postnatal weeks (3, 55).…”

“…In addition, NEB is required for normal muscle contractility, which has been demonstrated by altered Ca 2ϩ sensitivity (55) and inferior isometric stress production (3) in NEB-deficient muscle. NEB is of clinical interest because 50% or more of nemaline myopathy cases are caused by mutations in its gene (21,35,48,49).…”

Clinical utility gene card for: Nemaline myopathy – update 2015

“…The majority of the variants are either frameshift or nonsense variants, but also missense variants, and point variants and deletions affecting splice sites are known. 6,7 An in-frame deletion of exon 55 is present in the Ashkenazi Jewish population at a carrier frequency of~1 in 108. 8 Some patients present with a distal myopathy, with their skeletal muscle biopsies containing nemaline bodies, both nemaline bodies and cores, or no nemaline bodies.…”