Yang J C, Cortopassi G A
Department of Molecular Biosciences, University of California, Davis 95616, USA.
Free Radic Biol Med. 1998 Mar 1;24(4):624-31. doi: 10.1016/s0891-5849(97)00367-5.
It was recently reported that the mitochondrial protein cytochrome c is required for the induction of apoptosis, and that the overexpression of Bcl-2 caused increased retention of this apoptogenic factor by mitochondria. Several cellular toxins, including H2O2, tBOOH and Ca++, induce the Mitochondrial Permeability Transition (MPT); we tested the possibility that MPT is an intracellular sensor of toxicity that results in the release of cytochrome c. We observe that the release of cytochrome c from purified mitochondria is stimulated by the classical inducers of MPT, and is inhibited by the classical inhibitor of MPT, cyclosporin A (CsA). After induction of MPT, mitochondrial supernatants gained the activity to induce cleavage of caspase 3 (CPP32) in cytosolic extracts, and this gain of activity was inhibited by CsA pretreatment of mitochondria, and was cancelled by immunodepletion of cytochrome c from the supernatants. After induction of MPT, mitochondrial supernatants mixed with or without cytosolic extract gained the activity to ladder nuclei, and this gain of activity was inhibited by CsA pretreatment of mitochondria, and cancelled by immunodepletion of cytochrome c from the supernatants. These results demonstrate that the induction of MPT causes release of cytochrome c from mitochondria, which is required for the hallmarks of cytosolic and nuclear apoptosis, caspase 3 activation and nuclear laddering, and identify the MPT as a potential intracellular sensor of oxidants and other toxins, and as a target for the pharmacological inhibition of apoptosis.
最近有报道称，线粒体蛋白细胞色素c是诱导细胞凋亡所必需的，并且Bcl-2的过表达导致线粒体对这种凋亡诱导因子的保留增加。几种细胞毒素，包括H2O2、叔丁基过氧化氢和Ca++，可诱导线粒体通透性转换（MPT）；我们测试了MPT是否是一种导致细胞色素c释放的细胞内毒性传感器的可能性。我们观察到，纯化线粒体中细胞色素c的释放受到MPT经典诱导剂的刺激，并受到MPT经典抑制剂环孢素A（CsA）的抑制。诱导MPT后，线粒体上清液获得了在胞质提取物中诱导半胱天冬酶3（CPP32）裂解的活性，这种活性的增加受到线粒体CsA预处理的抑制，并通过从上清液中免疫去除细胞色素c而消除。诱导MPT后，与有或没有胞质提取物混合的线粒体上清液获得了使细胞核形成梯形条带的活性，这种活性的增加受到线粒体CsA预处理的抑制，并通过从上清液中免疫去除细胞色素c而消除。这些结果表明，MPT的诱导导致细胞色素c从线粒体释放，这是胞质和细胞核凋亡的标志、半胱天冬酶3激活和细胞核梯形条带所必需的，并确定MPT是氧化剂和其他毒素的潜在细胞内传感器，也是凋亡药理学抑制的靶点。
