Calpain 3 Is a Rapid-Action, Unidirectional Proteolytic Switch Central to Muscle Remodeling

Calpain 3 (CAPN3) is a cysteine protease that when mutated causes Limb Girdle Muscular Dystrophy 2A. It is thereby the only described Calpain family member that genetically causes a disease. Due to its inherent instability little is known of its substrates or its mechanism of activity and pathogenicity. In this investigation we define a primary sequence motif underlying CAPN3 substrate cleavage. This motif can transform non-related proteins into substrates, and identifies >300 new putative CAPN3 targets. Bioinformatic analyses of these targets demonstrate a critical role in muscle cytoskeletal remodeling and identify novel CAPN3 functions. Among the new CAPN3 substrates are three E3 SUMO ligases of the Protein Inhibitor of Activated Stats (PIAS) family. CAPN3 can cleave PIAS proteins and negatively regulates PIAS3 sumoylase activity. Consequently, SUMO2 is deregulated in patient muscle tissue. Our study thus uncovers unexpected crosstalk between CAPN3 proteolysis and protein sumoylation, with strong implications for muscle remodeling.钙蛋白酶3（Calpain 3, CAPN3）是一种半胱氨酸蛋白酶（cysteine protease），其突变可引发肢带型肌营养不良2A（Limb Girdle Muscular Dystrophy 2A）。它是目前已报道的唯一一种在遗传学上与疾病相关的钙蛋白酶家族成员。由于其固有的不稳定性，学界对其底物、活性机制及致病机理的认知仍十分有限。本研究明确了CAPN3底物切割所依赖的一级序列基序，该基序可将非同源蛋白转化为CAPN3底物，并鉴定出超过300个新的潜在CAPN3靶标。对这些靶标的生物信息学分析显示，CAPN3在肌肉细胞骨架重塑中发挥关键作用，并揭示了其此前未被报道的新功能。在新鉴定的CAPN3底物中，包含激活信号转导子与转录激活子蛋白抑制剂（Protein Inhibitor of Activated Stats, PIAS）家族的3种E3 SUMO连接酶（E3 SUMO ligases）。CAPN3可切割PIAS家族蛋白，并负向调控PIAS3的SUMO连接酶活性。由此，SUMO2在患者肌肉组织中出现调控紊乱。本研究揭示了CAPN3蛋白水解与蛋白质SUMO化修饰之间此前未被发现的交叉调控通路，该发现对肌肉重塑研究具有重要的启示意义。