Mitochondria, apoptosis and the Bcl-2 family

描述（申请人提供）：bcl -2家族蛋白在细胞凋亡的控制中起关键作用。特别是，在老鼠身上进行的基因靶向研究表明，Bax和Bak（类似的，也许还有Bok）是关键的效应物；在缺乏这些蛋白质的情况下，细胞在许多形式的凋亡中表现出缺陷。我们的重点是这些bcl -2家族蛋白如何调节线粒体外膜通透性（MOMP），这是许多细胞死亡途径中的主要事件。MOMP导致细胞色素c和线粒体中其他凋亡触发蛋白的易位！内膜间隙进入细胞质；这些蛋白反过来调节半胱天冬酶的激活和细胞凋亡的执行阶段。然而，即使半胱天冬酶失活或缺失，MOMP似乎仍然会导致大多数细胞死亡，通过启动关键线粒体功能的丧失以及活性氧的产生。因此，这一过程的调控和机制至关重要。在这里，我们提出的研究将有助于阐明bcl -2家族蛋白在MOMP中的作用。我们将使用无细胞系统来梳理这些蛋白质的作用机制，以及全细胞和体内方法，这将把这些研究扩展到更生理的背景下。Bcl-2家族有三个主要亚群：抗凋亡的“BH1-4”蛋白；“BH1-3”蛋白，包括促凋亡家族成员Bax， Bak和Bok，以及“BH3-only”蛋白，也具有促凋亡作用。BH3-only蛋白数量更多，在不同的细胞应激背景下通过转录和翻译后机制被特异性激活，并且似乎调节其他两个亚家族。我们的目标是依次解决Bcl-2家族的每个类别，首先，探索BH3-only蛋白调节bax型蛋白激活的机制；二是探讨bax型蛋白的膜透性机制；第三，了解BH1-4类Bcl-xL如何既能防止MOMP发生，又能在MOMP发生后重新密封MOM。

DESCRIPTION (provided by applicant): Bcl-2-family proteins play a key role in the control of apoptosis. In particular, gene targeting studies in mice have shown that Bax and Bak (and by analogy, perhaps Bok) are critical effectors; in the absence of these proteins, cells show deficiencies in many forms of apoptosis. Our focus is on how these Bcl-2-family proteins regulate mitochondrial outer membrane permeabilization (MOMP), a primary event in many cell death pathways. MOMP leads to the translocation of cytochrome c and other apoptotic trigger proteins from the mitochondria! inner membrane space into the cytoplasm; these proteins in turn regulate caspase activation and the execution phase of apoptosis. However, even if caspases are inactive or absent, MOMP nevertheless appears to doom most cells to die, through initiating a loss of key mitochondrial functions as well as the generation of reactive oxygen species. Thus, the regulation and mechanism of this process are of critical importance. Here we propose studies that will help elucidate the roles of Bcl-2-family proteins in MOMP. We will use both cell-free systems, to tease apart the mechanisms of action of these proteins, and whole-cell and in vivo approaches, which will extend these investigations to a more physiological context. There are three principal subgroups of the Bcl-2 family: "BH1-4" proteins, which are anti-apoptotic; "BH1-3" proteins, which include the pro-apoptotic family members Bax, Bak and Bok, and the "BH3-only" proteins, which are also pro-apoptotic. The BH3-only proteins are more numerous, are activated specifically through transcriptional and post-translational mechanisms in the context of different cellular stresses, and appear to regulate the other two subfamilies. Our aims, which address each category of the Bcl-2 family in turn, are first, to explore the mechanisms through which the BH3-only proteins regulate the activation of Bax-type proteins; second, to investigate the mechanism of membrane permeabilization by Bax-type proteins; and third, to understand how Bcl-xL, a member of the BH1-4 category, can both prevent MOMP and also reseal the MOM after MOMP has occurred.