The endoplasmic reticulum chaperone glycoprotein GRP94 with Ca2+-binding and antiapoptotic properties is a novel proteolytic target of calpain during etoposide-induced apoptosis
The endoplasmic reticulum chaperone glycoprotein GRP94 with Ca2+-binding and antiapoptotic properties is a novel proteolytic target of calpain during etoposide-induced apoptosis
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DOI：
10.1074/jbc.274.40.28476
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发表时间：
1999-10-01
影响因子：
4.8
通讯作者：
Lee, AS
中科院分区：
文献类型：
--
作者：
Reddy, RK;Lu, J;Lee, AS
GRP 94是一种分子量为94-kDa的分子伴侣糖蛋白，具有钙离子结合特性。我们在这里报告，在拓扑异构酶II抑制剂依托泊苷诱导的细胞凋亡过程中，与内质网膜相关的GRP 94的一部分经历了特定的蛋白水解裂解，与caspase CPP 32的激活和DNA片段化的启动相一致。在体内，能够阻断依托泊苷诱导的细胞凋亡的半胱天冬酶抑制剂只能部分保护GRP 94免受蛋白水解裂解，而用钙蛋白酶抑制剂I观察到完全抑制，但用蛋白酶体抑制剂则没有。在体外，GRP 94不是CPP 32的底物;相反，它可以被钙蛋白酶完全切割，钙蛋白酶是一种Ca 2+调节的蛋白酶。钙蛋白酶对GRP 94的切割是Ca 2+依赖的，并产生80 kDa的离散多肽。相反，钙蛋白酶对其他应激蛋白如GRP 78或HSP 70没有影响。此外，免疫组织化学染色揭示了特异性共定位的GRP 94与钙蛋白酶在核周区域后依托泊苷治疗。我们进一步表明，减少GRP 94的反义减少依托泊苷处理的Jurkat细胞的细胞活力，我们的研究提供了新的证据，细胞保护GRP 94，在抗凋亡蛋白Bcl-2的情况下，可以在凋亡过程中蛋白水解裂解本身的目标。
GRP94 is a 94-kDa chaperone glycoprotein with Ca2+-binding properties. We report here that during apoptosis induced by the topoisomerase II inhibitor etoposide, a fraction of GRP94 associated with the endoplasmic reticulum membrane undergoes specific proteolytic cleavage, coinciding with the activation of the caspase CPP32 and initiation of DNA fragmentation. In vivo, inhibitors of caspases able to block etoposide-induced apoptosis can only partially protect GRP94 from proteolytic cleavage, whereas complete inhibition is observed with calpain inhibitor I but not with the proteasome inhibitor. In vitro, GRP94 is not a substrate for CPP32; rather, it can be completely cleaved by calpain, a Ca2+-regulated protease, The cleavage of GRP94 by calpain is Ca2+-dependent and generates a discrete polypeptide of 80 kDa, In contrast, calpain has no effect on other stress proteins such as GRP78 or HSP70. Further, immunohistochemical staining reveals specific co-localization of GRP94 with calpain in the perinuclear region following etoposide treatment. We further showed that reduction of GRP94 by antisense decreased cell viability in etoposide-treated Jurkat cells, Our studies provide new evidence that the cytoprotective GRP94, as in the case of the antiapoptotic protein Bcl-2, can be targets of proteolytic cleavage themselves during the apoptotic process.