Calcium binding of ARC mediates regulation of caspase 8 and cell death
Calcium binding of ARC mediates regulation of caspase 8 and cell death
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DOI：
10.1128/mcb.24.22.9763-9770.2004
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发表时间：
2004-11-01
影响因子：
5.3
通讯作者：
Jung, YK
中科院分区：
文献类型：
--
作者：
Jo, DG;Jun, JI;Jung, YK
具有CARD的凋亡抑制因子（ARC）不仅具有阻断caspase 8活化的能力，而且具有调节与细胞死亡相关的caspase非依赖性线粒体事件的能力。然而，目前还不知道ARC如何调节半胱天冬酶依赖性和半胱天冬酶非依赖性细胞死亡。在这里，我们报告说，ARC是一个Ca 2+依赖的caspase 8和细胞死亡的调节。我们发现，在Ca 2+覆盖和所有的实验中，ARC蛋白通过C-末端富含脯氨酸/谷氨酸（P/E-rich）的结构域与Ca 2+结合。ARC的表达不仅减少胞浆Ca 2+瞬变，但毒胡萝卜素，A23187，和离子霉素，其中ARC的Ca 2+结合结构域是必不可少的细胞毒性作用。相反，通过靶向ARC直接干扰内源性ARC合成增强了这种Ca 2+介导的细胞死亡。此外，结合和免疫沉淀分析表明，ARC和caspase 8之间的蛋白质-蛋白质相互作用的减少，在体外的Ca 2+浓度的增加和在体内的HEK 293细胞与毒胡萝卜素的治疗。Caspase 8的激活也是毒胡萝卜素诱导的细胞死亡所必需的，并被ARC的异位表达所抑制。这些结果表明，钙结合介导的半胱天冬酶8和细胞死亡的ARC的调节。
Apoptosis repressor with CARD (ARC) possesses the ability not only to block activation of caspase 8 but to modulate caspase-independent mitochondrial events associated with cell death. However, it is not known how ARC modulates both caspase-dependent and caspase-independent cell death. Here, we report that ARC is a Ca2+-dependent regulator of caspase 8 and cell death. We found that in Ca2+ overlay and Stains-all assays, ARC protein bound to Ca2+ through the C-terminal proline/glutamate-rich (P/E-rich) domain. ARC expression reduced not only cytosolic Ca2+ transients but also cytotoxic effects of thapsigargin, A23187, and ionomycin, for which the Ca2+-binding domain of ARC was indispensable. Conversely, direct interference of endogenous ARC synthesis by targeting ARC enhanced such Ca2+-mediated cell death. In addition, binding and immunoprecipitation analyses revealed that the protein-protein interaction between ARC and caspase 8 was decreased by the increase of Ca2+ concentration in vitro and by the treatment of HEK293 cells with thapsigargin in vivo. Caspase 8 activation was, also required for the thapsigargin-induced cell death and suppressed by the ectopic expression of ARC. These results suggest that calcium binding mediates regulation of caspase 8 and cell death by ARC.