Ordering the cytochrome c-initiated caspase cascade: hierarchical activation of caspases-2, -3, -6, -7, -8, and -10 in a caspase-9-dependent manner.
Ordering the cytochrome c-initiated caspase cascade: hierarchical activation of caspases-2, -3, -6, -7, -8, and -10 in a caspase-9-dependent manner.
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DOI：
10.1083/jcb.144.2.281
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发表时间：
1999-01-25
期刊：
影响因子：
--
通讯作者：
Martin SJ
中科院分区：
文献类型：
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作者：
Slee EA;Harte MT;Kluck RM;Wolf BB;Casiano CA;Newmeyer DD;Wang HG;Reed JC;Nicholson DW;Alnemri ES;Green DR;Martin SJ
细胞色素c从线粒体进入胞质溶胶的退出已经被认为是细胞凋亡中的重要步骤。在胞质溶胶中，细胞色素c与CED-4同系物Apaf-1结合，从而触发Apaf-1介导的半胱天冬酶-9活化。Caspase-9被认为通过触发其他caspase激活事件传播死亡信号，其细节仍然不清楚。在这里，我们报告说，6个额外的半胱天冬酶（半胱天冬酶-2，3，-6，7，-8和-10）的无细胞提取物中处理响应细胞色素c，和其他三个（半胱天冬酶-1，4和-5）未能在相同的条件下被激活。体外结合试验证实，caspase-9选择性结合Apaf-1，而caspase-1，-2，-3，-6，-7，-8，和-10没有。细胞提取物中caspase-9的耗尽消除了caspase-2、-3、-6、-7、-8和-10的细胞色素c诱导的活化，表明caspase-9是所有这些下游caspase活化事件所必需的。从细胞提取物中免疫耗竭半胱天冬酶-3、-6和-7使我们能够将半胱天冬酶-9下游的半胱天冬酶激活事件的顺序排序，并揭示分支半胱天冬酶级联的存在。在该途径中，胱天蛋白酶-3是激活其他四种胱天蛋白酶（-2、-6、-8和-10）所必需的，并且还参与涉及胱天蛋白酶-9的反馈放大环。
Exit of cytochrome c from mitochondria into the cytosol has been implicated as an important step in apoptosis. In the cytosol, cytochrome c binds to the CED-4 homologue, Apaf-1, thereby triggering Apaf-1–mediated activation of caspase-9. Caspase-9 is thought to propagate the death signal by triggering other caspase activation events, the details of which remain obscure. Here, we report that six additional caspases (caspases-2, -3, -6, -7, -8, and -10) are processed in cell-free extracts in response to cytochrome c, and that three others (caspases-1, -4, and -5) failed to be activated under the same conditions. In vitro association assays confirmed that caspase-9 selectively bound to Apaf-1, whereas caspases-1, -2, -3, -6, -7, -8, and -10 did not. Depletion of caspase-9 from cell extracts abrogated cytochrome c–inducible activation of caspases-2, -3, -6, -7, -8, and -10, suggesting that caspase-9 is required for all of these downstream caspase activation events. Immunodepletion of caspases-3, -6, and -7 from cell extracts enabled us to order the sequence of caspase activation events downstream of caspase-9 and reveal the presence of a branched caspase cascade. Caspase-3 is required for the activation of four other caspases (-2, -6, -8, and -10) in this pathway and also participates in a feedback amplification loop involving caspase-9.