Expression of Glycogen Phosphorylase Isoforms in Cultured Muscle from Patients with McArdle's Disease Carrying the p.R771PfsX33 PYGM Mutation

BACKGROUND Mutations in the PYGM gene encoding skeletal muscle glycogen phosphorylase (GP) cause a metabolic disorder known as McArdle's disease. Previous studies in muscle biopsies and cultured muscle cells from McArdle patients have shown that PYGM mutations abolish GP activity in skeletal muscle, but that the enzyme activity reappears when muscle cells are in culture. The identification of the GP isoenzyme that accounts for this activity remains controversial. METHODOLOGY/PRINCIPAL FINDINGS In this study we present two related patients harbouring a novel PYGM mutation, p.R771PfsX33. In the patients' skeletal muscle biopsies, PYGM mRNA levels were ∼60% lower than those observed in two matched healthy controls; biochemical analysis of a patient muscle biopsy resulted in undetectable GP protein and GP activity. A strong reduction of the PYGM mRNA was observed in cultured muscle cells from patients and controls, as compared to the levels observed in muscle tissue. In cultured cells, PYGM mRNA levels were negligible regardless of the differentiation stage. After a 12 day period of differentiation similar expression of the brain and liver isoforms were observed at the mRNA level in cells from patients and controls. Total GP activity (measured with AMP) was not different either; however, the active GP activity and immunoreactive GP protein levels were lower in patients' cell cultures. GP immunoreactivity was mainly due to brain and liver GP but muscle GP seemed to be responsible for the differences. CONCLUSIONS/SIGNIFICANCE These results indicate that in both patients' and controls' cell cultures, unlike in skeletal muscle tissue, most of the protein and GP activities result from the expression of brain GP and liver GP genes, although there is still some activity resulting from the expression of the muscle GP gene. More research is necessary to clarify the differential mechanisms of metabolic adaptations that McArdle cultures undergo in vitro.