Molecular dissection of Oncoprotein-induced apoptosis

描述（由申请人提供）：Myc癌蛋白诱导细胞凋亡被认为是防止瘤形成的主要先天机制。然而，尽管许多不同的蛋白质都被认为是关键的下游凋亡效应物，但Myc与凋亡机制的基础联系的分子机制在很大程度上是未知的。Myc使细胞对由广泛的损伤诱导的凋亡敏感，这表明Myc起着引发凋亡决策的一些中心决定因素的作用。与此相一致，我们的工作直接涉及Myc促凋亡作用的快速和直接靶向的peption。基于这一观察结果，我们使用了一种有偏见的蛋白质组学方法来识别Bcl-xL相互作用蛋白，这些蛋白在人类和啮齿动物成纤维细胞中急性Myc激活后丰度或迁移率迅速变化。已经确定了三个候选者-已知的促凋亡BH 3蛋白Bim，一种具有称为PDCD 10的共有BH 3结构域的新蛋白，最近显示由Myc靶基因编码，以及小G蛋白Rab 7。我们建议通过分子、细胞和分子生物学和遗传学策略的组合来验证这些候选Myc凋亡效应子中的每一个，无论是在体外还是在体内的几个组织中，使用一组独特的可切换Myc小鼠转基因模型。最后，我们建议扩展我们最初的蛋白质组学研究，以建立一个地图的线粒体蛋白质组学的变化，迅速伴随Myc激活，特别强调限制蛋白质的外线粒体隔室最有可能靶向Myc凋亡效应。这些研究将提供新的见解的分子机制连接失调增殖与其先天肿瘤抑制凋亡故障安全，以及如何在癌细胞中切断这种联系。

DESCRIPTION (provided by applicant): The induction of apoptosis by the Myc oncoprotein is considered to be a major innate mechanism preventing the emergence of neoplasia. However, although many, diverse proteins have each been proposed as key downstream apoptotic effectors, the molecular mechanisms linking Myc with the basal the apoptotic machinery are largely unknown. Myc sensitizes cells to apoptosis induced by a wide range of insults, suggesting that Myc acts to prime some central determinant of apoptotic decision-making. Consistent with this, our work has directly implicated the mitochondrion as rapid and immediate target of Myc pro-apoptotic action. Based upon this observation, we have used a biased proteomic approach to identify Bcl-xL-interacting proteins that rapidly change in abundance or mobility following acute Myc activation in human and rodent fibroblasts. Three candidates have been identified - the known pro-apoptotic BH3 protein Bim, a novel protein with a consensus BH3 domain called PDCD10 recently shown to be encoded by a Myc target gene, and the small G protein Rab7. We propose to validate each of these candidate Myc apoptotic effectors by a combination of molecular, cell and molecular biological and genetics strategies, both in vitro and in several tissues in vivo using a unique set of switchable Myc mouse transgenic models. Finally, we propose to extend our initial proteomic study to establish a map of the proteomic changes in mitochondria that rapidly accompany Myc activation, with particular emphasis on the restricted proteins in the outer mitochondrial compartment most likely targeted by Myc apoptotic effectors. These studies will provide new insights into the molecular machinery linking deregulated proliferation with its innate tumor suppressive apoptotic failsafe, and how that linkage is severed in cancer cells.