Junctophilin-2
positive regulation of ryanodine-sensitive calcium-release channel activity
S165F mutation of junctophilin 2 affects Ca2+ signalling in skeletal muscle.
JPs (junctophilins) contribute to the formation of junctional membrane complexes in muscle cells by physically linking the t-tubule (transverse-tubule) and SR (sarcoplasmic reticulum) membranes. In humans with HCM (hypertrophic cardiomyopathy), mutations in JP2 are linked to altered Ca2+ signalling in cardiomyocytes; however, the effects of these mutations on skeletal muscle function have not been examined. In the present study, we investigated the role of the dominant-negative JP2-S165F mutation (which is associated with human HCM) in skeletal muscle. Consistent with the hypertrophy observed in human cardiac muscle, overexpression of JP2-S165F in primary mouse skeletal myotubes led to a significant increase in myotube diameter and resting cytosolic Ca2+ concentration. Single myotube Ca2+ imaging experiments showed reductions in both the excitation-contraction coupling gain and RyR (ryanodine receptor) 1-mediated Ca2+ release from the SR. Immunoprecipitation assays revealed defects in the PKC (protein kinase C)-mediated phosphorylation of the JP2-S165F mutant protein at Ser165 and in binding of JP2-S165F to the Ca2+ channel TRPC3 (transient receptor potential cation canonical-type channel 3) on the t-tubule membrane. Therefore both the hypertrophy and altered intracellular Ca2+ signalling in the JP2-S165F-expressing skeletal myotubes can be linked to altered phosphorylation of JP2 and/or altered cross-talk among Ca2+ channels on the t-tubule and SR membranes.