Excessive increases in intracellular [Ca2+] in skeletal muscle fibres cause failure of excitation-contraction (EC-) coupling by disrupting communication between the dihydropyridine receptors (DHPRs) in the transverse tubular (T-) system and the Ca2+ release channels (RyRs) in the sarcoplasmic reticulum (SR), but the exact mechanism is unknown. Previous work suggested a possible role of Ca2+-dependent proteolysis in this uncoupling process, but found no proteolysis of the DHPRs, RyRs or triadin. Junctophilin 1 (JP1) (~90 kDa) stabilizes close apposition of the T-system and SR membranes in adult skeletal muscle; its C-terminal end is embedded in the SR and its N-terminal associates with the T-system membrane. Exposure of skeletal muscle homogenates to precisely set [Ca2+] revealed that JP1 undergoes Ca2+-dependent proteolysis over the physiological [Ca2+] range in tandem with autolytic activation of endogenous µ calpain. JP1 cleavage occurs close to the C-terminal, yielding a ~75 kDa diffusible fragment and a fixed ~15 kDa fragment. Depolarization-induced force responses in rat skinned fibres were abolished following 1 min exposure to 40µM Ca2+, with accompanying loss of full-length JP1. Supra-physiological stimulation of rat skeletal muscle in vitro by repeated tetanic stimulation in 30 mM caffeine also produced marked proteolysis of JP1 (and not RyR1). In dystrophic mdx mice, JP1 proteolysis is seen in limb muscles at 4 and not at 10 weeks of age. Junctophilin-2 in cardiac and skeletal muscle also undergoes Ca2+-dependent proteolysis and JP2 levels are reduced following cardiac ischaemia-reperfusion. Junctophilin proteolysis may contribute to skeletal muscle weakness and cardiac dysfunction in a range of circumstances.
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