Role of Calpain-mediated Cleavage of Calcineurin in Excitotoxic Neurodegeneration

钙调神经磷酸酶和钙调蛋白分别是一种钙/钙调素依赖的蛋白磷酸酶和一种钙依赖的半胱氨酸蛋白酶，它们通过独立的级联反应介导神经细胞死亡。在本研究中，我们发现在神经兴奋性毒性过程中，钙调神经磷酸酶A(CNA)在体外和体内都会被Calain直接切割，导致该酶转化为活性形式。质谱学鉴定了CNA中的三个裂解位点，其中两个是结构性活性形式。裂解的CNA过表达可诱导caspase活性和神经细胞死亡。钙蛋白酶抑制剂和膜透性的钙调蛋白多肽不仅能阻断CNA的切割，而且在体内外对兴奋性毒性神经细胞的死亡也有保护作用。这些结果表明，CNA是钙蛋白酶的一个重要靶点，而钙蛋白酶介导的CNA激活触发了兴奋性毒性神经变性。本研究建立了钙调神经磷酸酶与钙调神经磷酸酶之间的分子联系，从而揭示了钙调神经磷酸酶在某些病理状态下对钙调神经磷酸酶的蛋白水解性调节的新机制。

Calcineurin and calpain, a Ca2+/calmodulin-dependent protein phosphatase and a Ca2+-dependent cysteine protease, respectively, mediate neuronal cell death through independent cascades. In this research, we showed that during neuroexcitotoxicity, calcineurin A (CnA) is directly cleaved by calpain in vitro and in vivo, resulting in the enzyme being converted to an active form. Mass spectrometry identified three cleavage sites in CnA, two of which were constitutively active forms. Overexpression of the cleaved CnA induced caspase activity and neuronal cell death. Calpain inhibitors and membrane-permeable calpastatin peptides not only blocked the cleavage of CnA, but also protected against excitotoxic neuronal cell death in vitro and in vivo. These results indicate that CnA is a crucial target for calpain, and the calpain-mediated activation of CnA triggers excitotoxic neurodegeneration. This study established a molecular link between calpain and calcineurin, thereby demonstrating a new mechanism for proteolytical regulation of calcineurin by calpain in response to certain pathological states.