Tu Shine, McStay Gavin P, Boucher Louis-Martin, Mak Tak, Beere Helen M, Green Douglas R
Division of Cellular Immunology, La Jolla Institute for Allergy and Immunology, 10355 Science Center Dr., San Diego, CA 92121, USA.
Nat Cell Biol. 2006 Jan;8(1):72-7. doi: 10.1038/ncb1340. Epub 2005 Dec 18.
Activation of 'initiator' (or 'apical') caspases-2, -8 or -9 (refs 1-3) is crucial for induction of apoptosis. These caspases function to activate executioner caspapses that, in turn, orchestrate apoptotic cell death. Here, we show that a cell-permeable, biotinylated pan-caspase inhibitor (bVAD-fmk) both inhibited and 'trapped' the apical caspase activated when apoptosis was triggered. As expected, only caspase-8 was trapped in response to ligation of death receptors, whereas only caspase-9 was trapped in response to a variety of other apoptosis-inducing agents. Caspase-2 was exclusively activated in heat shock-induced apoptosis. This activation of caspase-2 was also observed in cells protected from heat-shock-induced apoptosis by Bcl-2 or Bcl-xL. Reduced sensitivity to heat-shock-induced death was observed in caspase-2(-/-) cells. Furthermore, cells lacking the adapter molecule RAIDD failed to activate caspase-2 after heat shock treatment and showed resistance to apoptosis in this setting. This approach unambiguously identifies the apical caspase activated in response to apoptotic stimuli, and establishes caspase-2 as a proximal mediator of heat shock-induced apoptosis.
“起始”（或“顶端”）半胱天冬酶-2、-8或-9（参考文献1-3）的激活对于诱导细胞凋亡至关重要。这些半胱天冬酶的作用是激活执行性半胱天冬酶，而执行性半胱天冬酶反过来又协调细胞凋亡性死亡。在此，我们表明一种可透过细胞的、生物素化的泛半胱天冬酶抑制剂（bVAD-fmk）在细胞凋亡被触发时既能抑制又能“捕获”被激活的顶端半胱天冬酶。正如预期的那样，在死亡受体被连接时只有半胱天冬酶-8被捕获，而在对多种其他细胞凋亡诱导剂产生反应时只有半胱天冬酶-9被捕获。半胱天冬酶-2仅在热休克诱导的细胞凋亡中被激活。在通过Bcl-2或Bcl-xL免受热休克诱导的细胞凋亡的细胞中也观察到了半胱天冬酶-2的这种激活。在半胱天冬酶-2基因敲除（-/-）细胞中观察到对热休克诱导的死亡的敏感性降低。此外，缺乏衔接分子RAIDD的细胞在热休克处理后未能激活半胱天冬酶-2，并且在这种情况下表现出对细胞凋亡的抗性。这种方法明确地鉴定了对细胞凋亡刺激产生反应而被激活的顶端半胱天冬酶，并确立了半胱天冬酶-2作为热休克诱导的细胞凋亡的近端介质。
