Shoshan-Barmatz Varda, Keinan Nurit, Abu-Hamad Salah, Tyomkin Dalia, Aram Lior
Department of Life Sciences, Ben-Gurion University of the Negev, Beer-Sheva 84105, Israel.
Biochim Biophys Acta. 2010 Jun-Jul;1797(6-7):1281-91. doi: 10.1016/j.bbabio.2010.03.003. Epub 2010 Mar 6.
Mitochondria, central to basic life functions due to their generation of cellular energy, also serve as the venue for cellular decisions leading to apoptosis. A key protein in mitochondria-mediated apoptosis is the voltage-dependent anion channel (VDAC), which also mediates the exchange of metabolites and energy between the cytosol and the mitochondria. In this study, the functions played by the N-terminal region of VDAC1 and by VDAC1 oligomerization in the release of cytochrome c, Smac/Diablo and apoptosis-inducing factor (AIF) and subsequent apoptosis were addressed. We demonstrate that cells undergoing apoptosis induced by STS or cisplatin and expressing N-terminally truncated VDAC1 do not release cytochrome c, Smac/Diablo or AIF. Ruthenium red (RuR), AzRu, DIDS and hexokinase-I (HK-I), all known to interact with VDAC, inhibited the release of cytochrome c, Smac/Diablo and AIF, while RuR-mediated inhibition was not observed in cells expressing RuR-insensitive E72Q-VDAC1. These findings suggest that VDAC1 is involved in the release of not only cytochrome c but also of Smac/Diablo and AIF. We also demonstrate that apoptosis induction is associated with VDAC oligomerization, as revealed by chemical cross-linking and monitoring in living cells using Bioluminescence Resonance Energy Transfer. Apoptosis induction by STS, H2O2 or selenite augmented the formation of VDAC oligomers several fold. The results show VDAC1 to be a component of the apoptosis machinery and offer new insight into the functions of VDAC1 oligomerization in apoptosis and of the VDAC1 N-terminal domain in the release of apoptogenic proteins as well as into regulation of VDAC by anti-apoptotic proteins, such as HK and Bcl2.
线粒体因其产生细胞能量而对基本生命功能至关重要，同时也是导致细胞凋亡的细胞决策场所。线粒体介导的细胞凋亡中的一种关键蛋白是电压依赖性阴离子通道（VDAC），它还介导细胞质和线粒体之间代谢物与能量的交换。在本研究中，探讨了VDAC1的N端区域以及VDAC1寡聚化在细胞色素c、Smac/Diablo和凋亡诱导因子（AIF）释放及随后的细胞凋亡中所起的作用。我们证明，经历STS或顺铂诱导的细胞凋亡并表达N端截短的VDAC1的细胞不会释放细胞色素c、Smac/Diablo或AIF。钌红（RuR）、叠氮钌（AzRu）、二异硫氰酸二苯乙烯（DIDS）和己糖激酶-I（HK-I），所有这些都已知与VDAC相互作用，抑制了细胞色素c、Smac/Diablo和AIF的释放，而在表达对RuR不敏感的E72Q-VDAC1的细胞中未观察到RuR介导的抑制作用。这些发现表明，VDAC1不仅参与细胞色素c的释放，还参与Smac/Diablo和AIF的释放。我们还证明，细胞凋亡诱导与VDAC寡聚化有关，这通过化学交联和使用生物发光共振能量转移在活细胞中进行监测得以揭示。STS、H2O2或亚硒酸盐诱导的细胞凋亡使VDAC寡聚物的形成增加了几倍。结果表明VDAC1是细胞凋亡机制的一个组成部分，并为VDAC1寡聚化在细胞凋亡中的功能以及VDAC1 N端结构域在凋亡蛋白释放中的功能，以及抗凋亡蛋白如HK和Bcl2对VDAC的调节提供了新的见解。
