Calpain-dependent cleavage of cain/cabin1 activates calcineurin to mediate calcium-triggered cell death
Calpain-dependent cleavage of cain/cabin1 activates calcineurin to mediate calcium-triggered cell death
复制标题
DOI：
10.1073/pnas.152336999
复制
发表时间：
2002-07-23
影响因子：
11.1
通讯作者：
Jung, YK
中科院分区：
文献类型：
--
作者：
Kim, MJ;Jo, DG;Jung, YK
Cain/cabin 1是钙调神经磷酸酶（Cn）的内源性抑制剂，钙调神经磷酸酶是一种钙依赖性丝氨酸/苏氨酸磷酸酶，参与包括凋亡在内的多种细胞功能。我们在这里表明，在细胞凋亡过程中，cain/cabin 1被钙蛋白酶在羧基末端裂解，产生一个裂解产物的分子量为32 kDa的一个必要的步骤，导致Cn介导的细胞死亡。用钙蛋白酶体外底物筛选法从小鼠胸腺cDNA文库中鉴定出cain/cabin 1。暴露的Jurkat细胞的钙离子载体，A23187，诱导cain/cabin 1裂解和细胞死亡，伴随着激活的钙蛋白酶和Cn。钙蛋白酶抑制剂calpeptin和zLLY抑制A23187诱导的cain/cabin 1裂解和Cn激活，表明Cn激活和cain/cabin 1裂解是钙蛋白酶依赖性的。cain/cabin 1或无催化活性的Cn突变体[CnAbeta（2）（1-401/H160 N）]的表达和FK 506处理减少了A23187诱导的细胞死亡。在体外钙蛋白酶切割和免疫沉淀试验与cain/cabin 1的缺失突变体表明，切割发生在Cn结合结构域的cain/cabin 1，表明在其C末端的切割钙蛋白酶阻止cain/cabin 1从结合到Cn。此外，体外结合试验表明cain/cabin 1与Cn A的Cn B结合结构域结合。总之，这些结果表明，钙蛋白酶切割cain/cabin 1的钙调神经磷酸酶结合结构域，以激活Cn并引起钙触发的细胞死亡。
Cain/cabin1 is an endogenous inhibitor of calcineurin (Cn), a calcium-dependent serine/threonine phosphatase involved in various cellular functions including apoptosis. We show here that during apoptosis cain/cabin1 is cleaved by calpain at the carboxyl terminus to generate a cleavage product with a molecular mass of 32 kDa as a necessary step leading to Cn-mediated cell death. Mouse cain/cabin1 was identified from a thymus cDNA library by an in vitro substrate-screening assay with calpain. Exposure of Jurkat cells to the calcium ionophore, A23187, induced cain/cabin1 cleavage and cell death, accompanied by activation of calpain and Cn. The calpain inhibitors, calpeptin and zLLY, suppressed both A23187-induced cain/cabin1 cleavage and Cn activation, indicating that Cn activation and cain/cabin1 cleavage are calpain-dependent. Expression of cain/cabin1 or a catalytically inactive Cn mutant [CnAbeta(2)(1-401/H160N)] and treatment with FK506 reduced A23187-induced cell death. In vitro calpain cleavage and immunoprecipitation assays with deletion mutants of cain/cabin1 showed that cleavage occurred in the Cn-binding domain of cain/cabin1, indicating that the cleavage at its C terminus by calpain prevented cain/cabin1 from binding to Cn. In addition, in vitro binding assays showed that cain/cabin1 bound to the Cn B-binding domain of Cn A. Taken together, these results indicate that calpain cleaves the calcineurin-binding domain of cain/cabin1 to activate Cn and elicit calcium-triggered cell death.