Mitochondrial p32 regulation of the Mdm2-p53 tumor suppression signaling and apop

描述（由申请人提供）：我们的实验室最近发现了一种线粒体定位蛋白p32，作为肿瘤抑制蛋白p14ARF的结合伴侣。初步数据表明，p32对于p14ARF至关重要，p14ARF是将高增殖的致癌应激信号转导到Mdm2-p53肿瘤抑制途径的关键介质，定位到线粒体并诱导p53依赖性细胞凋亡。重要的是，破坏p32结合的人类癌症源性p14ARF突变会损害这两种功能。最近，我们的初步研究表明，p32实际上是多种凋亡刺激诱导的细胞凋亡所必需的。提出的研究背后的总体假设是，p32指定了一个监视系统的必要因素，该系统监测线粒体功能的完整性，并促进细胞凋亡，以应对不可修复的线粒体损伤。该假设的基本原理基于以下观察结果。(1) p32是线粒体蛋白。(2)敲低p32可使细胞对多种凋亡刺激诱导的凋亡脱敏。(3)与细胞色素c一样，p32在细胞凋亡过程中在细胞质中积累。(4)异位表达、胞质定位的p32诱导细胞凋亡。(5)据报道p32在氧化磷酸化中起作用。本课题的实验重点是p32定位的动态和结果，p32在调节细胞凋亡中的功能和机制，以及p32在代谢调节和肿瘤发生中的遗传学和生物学。基于这些观察结果，本研究将通过生物化学、细胞和遗传方法的结合，重点研究p32在调节细胞凋亡中的功能和机制。具体目的是评估p32在多种凋亡条件下调节细胞凋亡的作用，确定p32亚细胞定位的动力学，研究p32促进凋亡细胞死亡的机制，并利用p32条件敲除小鼠研究p32的生理功能。如果成功，该研究将赋予p32新的功能。它还将有助于我们理解细胞凋亡死亡——这是一个在发育过程中以及在癌症、风湿性关节炎和神经退行性疾病等疾病的发病机制中至关重要的过程——并可能最终导致在治疗这些疾病时控制细胞凋亡的额外药物靶点。

DESCRIPTION (provided by applicant): A mitochondria-localized protein, p32, was recently identified by our lab as a binding partner for the tumor suppressor protein p14ARF. Preliminary data indicated that p32 is essential for p14ARF, a critical mediator for transducing hyperproliferative, oncogenic stress signals to the Mdm2-p53 tumor suppression pathway, to localize to mitochondria and to induce p53-dependent apoptosis. Importantly, human cancer-derived p14ARF mutations that disrupt p32 binding can impair both of these functions. Recently, our preliminary studies have shown that p32 is in fact essential for apoptosis induced by a broad range of apoptotic stimuli. The overall hypothesis behind the proposed research is that p32 specifies an essential factor for a surveillance system that monitors the integrity of mitochondrial function and promotes apoptosis in response to irreparable mitochondrial damage. The rationale for the hypothesis is based on the following observations. (1) p32 is a mitochondrial protein. (2) Knockdown p32 desensitizes cells to apoptosis induced by a broad range of apoptotic stimuli. (3) Like cytochrome c, p32 accumulates in the cytoplasm during apoptosis. (4) Ectopically expressed, cytoplasm- localized p32 induces apoptosis. (5) p32 is reported to have a role in oxidative phosphorylation. The experimental focus of the proposal is on dynamics and outcome of p32 localization, function and mechanism of p32 in regulating apoptosis, and genetics and biology of p32 in metabolic regulation and tumorigenesis. Based on these observations, the proposed research will focus on characterizing the function and mechanism of p32 in regulating apoptosis by a combination of biochemical, cellular, and genetic approaches. The specific aims are designed to assess p32's role in regulating apoptosis under a broad range of apoptotic conditions and define the dynamics of p32 subcellular localization, to Investigate mechanisms by which p32 promotes apoptotic cell death, and to investigate p32's physiological function using p32 conditional knockout mice. If successful, the proposed study will ascribe new functions to p32. It will also aid in our understanding of apoptotic cell death - a process that is critical during development and in the pathogenesis of diseases such as cancer, rheumatoid arthritis, and neurodegenerative diseases - and may eventually lead to additional drug targets for controlling apoptosis in treatment of these diseases.