Prostate Apoptosis Response 4 (Par-4), a Novel Substrate of Caspase-3 during Apoptosis Activation
Prostate Apoptosis Response 4 (Par-4), a Novel Substrate of Caspase-3 during Apoptosis Activation
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DOI：
10.1128/mcb.06321-11
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发表时间：
2012-02-01
影响因子：
5.3
通讯作者：
Asselin, Eric
中科院分区：
文献类型：
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作者：
Chaudhry, Parvesh;Singh, Mohan;Asselin, Eric
前列腺凋亡反应4（Par-4）是一种广泛表达的促凋亡肿瘤抑制蛋白。在这里，我们第一次表明，Par-4是一种新的底物caspase-3在细胞凋亡过程中。我们发现Par-4在顺铂诱导的人正常和癌细胞系凋亡过程中被切割。Par-4切割产生类似于25 kDa的C-末端片段，并且Par-4的切割被半胱天冬酶-3抑制剂完全抑制，表明半胱天冬酶-3直接参与Par-4的切割。Caspase-3缺陷型MCF-7细胞对顺铂处理不显示Par-4裂解，并且MCF-7细胞中Caspase-3的恢复产生Par-4水平的降低，并出现裂解片段。此外，Par-4的敲低降低了半胱天冬酶-3活化和凋亡诱导。定点突变显示，caspase-3对Par-4的切割发生在非常规位点，EEPD 131向下箭头G。有趣的是，野生型Par-4的过表达而不是Par-4 D131 A突变体使细胞对顺铂诱导的凋亡敏感。在半胱天冬酶-3切割后，Par-4的切割片段在细胞核中积累并显示出增加的凋亡活性。Par-4切割片段的过表达抑制I κ B α磷酸化并阻断NF-κ B核转位。我们已经确定了一个新的特定的caspase-3裂解位点的Par-4，裂解片段的Par-4保留促凋亡活性。
Prostate apoptosis response 4 (Par-4) is a ubiquitously expressed proapoptotic tumor suppressor protein. Here, we show for the first time, that Par-4 is a novel substrate of caspase-3 during apoptosis. We found that Par-4 is cleaved during cisplatin-induced apoptosis in human normal and cancer cell lines. Par-4 cleavage generates a C-terminal fragment of similar to 25 kDa, and the cleavage of Par-4 is completely inhibited by a caspase-3 inhibitor, suggesting that caspase-3 is directly involved in the cleavage of Par-4. Caspase-3-deficient MCF-7 cells do not show Par-4 cleavage in response to cisplatin treatment, and restoration of caspase-3 in MCF-7 cells produces a decrease in Par-4 levels, with the appearance of a cleaved fragment. Additionally, knockdown of Par-4 reduces caspase-3 activation and apoptosis induction. Site-directed mutagenesis reveals that Par-4 cleavage by caspase-3 occurs at an unconventional site, EEPD131 down arrow G. Interestingly, overexpression of wild-type Par-4 but not the Par-4 D131A mutant sensitizes cells to cisplatin-induced apoptosis. Upon caspase-3 cleavage, the cleaved fragment of Par-4 accumulates in the nucleus and displays increased apoptotic activity. Overexpression of the cleaved fragment of Par-4 inhibits I kappa B alpha phosphorylation and blocks NF-kappa B nuclear translocation. We have identified a novel specific caspase-3 cleavage site in Par-4, and the cleaved fragment of Par-4 retains proapoptotic activity.