PDF(Pubmed)
Abstract：
Limb-Girdle Muscular Dystrophy R1/2A (LGMD R1/2A) is caused by mutations in the CAPN3 gene 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) in Capn3 deficient (C3KO) and wild-type (WT) mice, we determined whether loss of Calpain 3 altered Store-Operated Calcium Entry (SOCE) activity. Direct Ca2+ influx measurements revealed loss of Capn3 elicits 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 in extensor digitorum longus muscles and a greater decay of Ca2+ transients in flexor digitorum brevis muscle 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 LGMD R1/2A pathology.
摘要：
肢带肌营养不良R1/2A（LGMDR1/2A）是由编码骨骼肌特异性Calpain3的CAPN3基因突变引起的，Ca2+-依赖性蛋白酶。钙蛋白酶3在三联体中的定位表明它有助于Ca2稳态。通过活细胞Ca2+测量，肌肉力学，免疫荧光，Capn3缺陷（C3KO）和野生型（WT）小鼠的电子显微镜（EM），我们确定了钙蛋白酶3的丢失是否改变了储存操作的钙输入（SOCE）活性。直接的Ca2流入测量显示Capn3的损失引起静息SOCE升高和静息胞浆Ca2增加，由EM观察到的钙进入单位（CEU）的高发生率支持。对C3KO和WT小鼠进行一次跑步机跑步以引起SOCE。在跑步机后运行的1HR内，在重复刺激期间，C3KO小鼠在指长伸肌中的力产生减少，而在指短屈肌肌纤维中的Ca2瞬变衰减更大。C3KO小鼠运动诱导的SOCE激活受损的惊人证据包括关键SOCE蛋白的共定位不良，基质相互作用分子1（STIM1）和ORAI1，并伴有C3KO肌肉中CEU的消失。这些结果表明，钙蛋白酶3是骨骼肌中SOCE的关键调节剂，并将SOCE失调鉴定为LGMDR1/2A病理的促成因素。
