Loss of calpain 3 dysregulates store-operated calcium entry and its exercise response in mice

Abstract

ABSTRACTLimb-Girdle Muscular Dystrophy 2A (LGMD2A) is caused by mutations in theCAPN3gene encoding Calpain 3, a skeletal-muscle specific, Ca2+-dependent protease. Localization of Calpain 3 within the triad suggests it contributes to Ca2+homeostasis. Through live-cell Ca2+measurements, muscle mechanics, immunofluorescence, and electron microscopy (EM) inCapn3deficient (C3KO) and wildtype (WT) mice, we determined if loss of Calpain 3 altered Store-Operated Calcium Entry (SOCE) activity. Direct Ca2+influx measurements revealed loss ofCapn3elicits elevated resting SOCE and increased resting cytosolic Ca2+, supported by high incidence of calcium entry units (CEUs) observed by EM. C3KO and WT mice were subjected to a single bout of treadmill running to elicit SOCE. Within 1HR post-treadmill running, C3KO mice exhibited diminished force production inextensor digitorum longusmuscles and a greater decay of Ca2+transients inflexor digitorum brevismuscle fibers during repetitive stimulation. Striking evidence for impaired exercise-induced SOCE activation in C3KO mice included poor colocalization of key SOCE proteins, stromal-interacting molecule 1 (STIM1) and ORAI1, combined with disappearance of CEUs in C3KO muscles. These results demonstrate that Calpain 3 is a key regulator of SOCE in skeletal muscle and identify SOCE dysregulation as a contributing factor to LGMD2A pathology.