Mitochondria, apoptosis and the Bcl-2 family

描述(申请人提供)：bcl2家族蛋白在控制细胞凋亡中起关键作用。特别是，在小鼠身上进行的基因打靶研究表明，Bax和Bak(以此类推，可能还有Bok)是关键的效应器；在缺乏这些蛋白质的情况下，细胞在多种形式的凋亡中表现出缺陷。我们的重点是这些Bcl2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一个主要事件。MOMP导致细胞色素c和其他来自线粒体的凋亡触发蛋白的移位！内膜间隙进入细胞质；这些蛋白反过来调节caspase的激活和细胞凋亡的执行阶段。然而，即使caspase不活跃或不存在，MOMP似乎仍然通过启动关键线粒体功能的丧失和活性氧的产生来注定大多数细胞的死亡。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Bcl2家族蛋白在MOMP中的作用。我们将使用两种无细胞系统来梳理这些蛋白质的作用机制，以及全细胞和活体方法，这将把这些研究扩展到更具生理学的背景下。Bc l-2家族有三个主要亚群：“BH1-4”蛋白，它们是抗凋亡蛋白；“BH1-3”蛋白，它包括促凋亡家族成员Bax、Bak和Bok，以及“BH3-only”蛋白，它们也是促凋亡蛋白。只有BH3的蛋白数量更多，在不同的细胞应激背景下，通过转录和翻译后机制特异性地激活，并似乎调节其他两个亚家族。我们的目标依次针对BH1-2家族中的每一类，第一，探索仅BH3-蛋白调节Bax-型蛋白激活的机制；第二，研究Bax-型蛋白的膜通透性机制；第三，了解BH1-4类别的成员Bcl-xl如何既能预防MOMP，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.