Smac induces cytochrome c release and apoptosis independently from Bax/Bcl-xL in a strictly caspase-3-dependent manner in human carcinoma cells
Smac induces cytochrome c release and apoptosis independently from Bax/Bcl-xL in a strictly caspase-3-dependent manner in human carcinoma cells
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DOI：
10.1038/sj.onc.1207594
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发表时间：
2004-06-03
期刊：
影响因子：
8
通讯作者：
Daniel, PT
中科院分区：
文献类型：
--
作者：
Hasenjäger, A;Gillissen, B;Daniel, PT
线粒体凋亡途径通过向细胞质中释放各种促凋亡因子，包括细胞色素c和半胱天冬酶的第二种线粒体激活剂Smac，介导细胞死亡。Smac先前被证明可以抑制IAP蛋白，并在细胞色素c释放时促进caspase级联的启动。为了研究Smac在细胞凋亡过程中的功能，并探索Smac作为实验性癌症治疗药物的作用，我们在cis提供的Tet-off系统的控制下，构建了基于单个腺病毒载体的Smac表达系统。无论Bax缺失或BCl-X-L过表达，条件表达Smac诱导人HCT116和DU145癌细胞凋亡。然而，Smac诱导的细胞凋亡与细胞色素c的释放和线粒体膜电位的破坏有关。这表明Smac在细胞凋亡启动过程中独立于Bax和BCl-X-L起作用，并触发一个正反馈循环，导致线粒体不依赖Bax/BCl-X-L的激活。在caspase精通的细胞中，smac诱导的细胞凋亡可以通过细胞渗透性LEHD （caspase-9抑制剂）和DEVD （caspase-3抑制剂）肽部分抑制。此外，在携带caspase-3零突变的MCF-7细胞中，caspase-3的表达缺失完全消除了对smac诱导的凋亡或非凋亡、坏死样细胞死亡的敏感性，而caspase-3的重新表达则赋予了敏感性。总之，caspase-3激活而非caspase-9激活是执行smac诱导的细胞死亡所必需的。值得注意的是，在caspase-3缺失的情况下，Smac不诱导caspase-9加工。因此，在smac诱导的细胞凋亡过程中，caspase-9加工发生在caspase-3激活之后。总之，Smac能够规避线粒体凋亡信号中的缺陷，如Bax的缺失或BCl-X-L的过表达，这些缺陷在抗癌治疗耐药的肿瘤细胞中经常观察到。因此，Smac似乎是一个有希望的抗癌治疗靶点。
The mitochondrial apoptosis pathway mediates cell death through the release of various pro-apoptotic factors including cytochrome c and Smac, the second mitochondrial activator of caspases, into the cytosol. Smac was shown previously to inhibit IAP proteins and to facilitate initiation of the caspase cascade upon cytochrome c release. To investigate Smac function during apoptosis and to explore Smac as an experimental cancer therapeutic, we constructed an expression system based on a single adenoviral vector containing Smac under control of the Tet-off system supplied in cis. Conditional expression of Smac induced apoptosis in human HCT116 and DU145 carcinoma cells regardless of the loss of Bax or overexpression of BCl-X-L. Nevertheless, apoptosis induced by Smac was associated with cytochrome c release and breakdown of the mitochondrial membrane potential. This indicates that Smac acts independently of Bax and BCl-X-L during initiation of apoptosis and triggers a positive feedback loop that results in Bax/BCl-X-L-independent activation of mitochondria. In caspase-proficient cells, Smac-induced apoptosis could be inhibited partially by cell-permeable LEHD (caspase-9 inhibitor) and DEVD (caspase-3 inhibitor) peptides. Furthermore, loss of caspase-3 expression in MCF-7 cells carrying a caspase-3 null mutation completely abrogated the sensitivity for Smac-induced apoptotic or nonapoptotic, necrosis-like cell death, while re-expression of caspase-3 conferred sensitivity. Altogether, caspase-3 but not caspase-9 activation was necessary for execution of Smac-induced cell death. Notably, Smac did not induce caspase-9 processing in the absence of caspase-3. Thus, caspase-9 processing occurs secondary to caspase-3 activation during Smac-induced apoptosis. Altogether, Smac is capable of circumventing defects in mitochondrial apoptosis signaling such as loss of Bax or overexpression of BCl-X-L that are frequently observed in tumor cells resistant to anticancer therapy. Consequently, Smac appears to be a promising therapeutic target in anticancer treatment.