Calpain-3 Is Not a Sodium Dependent Protease and Simply Requires Calcium for Activation

Area of Science:

• Muscle physiology
• Protease biochemistry

Background:

• Calpain-3 (CAPN3) is a muscle-specific protease.
• Previous studies suggested Na+ could activate CAPN3, but under non-physiological conditions.

Purpose of the Study:

• To investigate the activation mechanisms of CAPN3 under physiological conditions.
• To determine the effects of CAPN3 activation on muscle proteins like RyR1 and titin.

Main Methods:

• Autolysis assays of human muscle homogenates with varying Ca²⁺ and Na⁺ concentrations.
• Analysis of protein degradation products using SDS-PAGE and Western blotting.

Main Results:

• CAPN3 autolysis is primarily Ca²⁺-dependent, requiring lower concentrations than CAPN1.
• High Na⁺ did not activate CAPN3 under physiological K⁺ conditions.
• Autolysed CAPN3 dissociates from titin, but does not degrade RyR1.
• CAPN1 activation by Ca²⁺ degraded junctophilin but not RyR1.

Conclusions:

• CAPN3 activation in muscle relies on Ca²⁺, not Na⁺.
• CAPN3's interaction with titin is regulated by its autolytic state.
• CAPN3 and CAPN1 have distinct substrate specificities, with neither degrading RyR1 under tested conditions.