Abstract
An autosomal dominant protein aggregate myopathy, characterized by high plasma creatine kinase and calsequestrin-1 (CASQ1) accumulation in skeletal muscle, has been recently associated with a missense mutation in CASQ1 gene. The mutation replaces an evolutionarily-conserved aspartic acid with glycine at position 244 (p.D244G) of CASQ1, the main sarcoplasmic reticulum (SR) Ca2+ binding and storage protein localized at the terminal cisternae of skeletal muscle cells. Here, immunocytochemical analysis of myotubes, differentiated from muscle-derived primary myoblasts, shows that sarcoplasmic vacuolar aggregations positive for CASQ1 are significantly larger in CASQ1-mutated cells than control cells. A strong co-immuno staining of both RyR1 and CASQ1 was also noted in the vacuoles of myotubes and muscle biopsies derived from patients. Electrophysiological recordings and sarcoplasmic Ca2+ measurements provide evidence for less Ca2+ release from the SR of mutated myotubes when compared to that of controls. These findings further clarify the pathogenic nature of the p.D244G variant and point out defects in sarcoplasmic Ca2+ homeostasis as a mechanism underlying this human disease, which could be distinctly classified as “CASQ1-couplonopathy”.
Highlights
A surplus of endogenous proteins within circumscribed areas of muscle fibers often characterizes protein aggregate myopathies
Densitometric analysis of the immunoblot bands of protein extracted from myotube cultures showed that CASQ1 values were higher in patient samples compared to controls (1.12 ±0.21 vs 0.62±0.48, respectively), whereas, values for RyR1 bands were similar (0.26±0.22 vs 0.21±0.11, respectively)
These data indicate that patient-derived cultured myotubes recapitulate distinct morphological features of diseased adult muscle fibers, namely, a doi:10.1371/journal.pone.0155516.g001
Summary
A surplus of endogenous proteins within circumscribed areas of muscle fibers often characterizes protein aggregate myopathies. Chronically elevated plasma creatine kinase (CK) is a common feature of these myopathies and may precede clinical presentation. Patients from several Italian families with hyperCKaemia and clinical myopathy, whose muscle biopsies showed pathological features and sarcoplasmic inclusions positive for the SR Ca2+ binding/.
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