Mechanisms of Bax Activation

Bax 激活机制

• 批准号: 7018585
• 负责人: Xu Luo
• 金额: $ 24.26万
• 依托单位: UNIVERSITY OF NEBRASKA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-02-01 至 2011-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7018585
• 关键词: BCL2 gene /protein; Bax gene /protein; HeLa cells; SDS polyacrylamide gel electrophoresis; apoptosis; biological signal transduction; mitochondria; protein protein interaction; protein structure; protein structure function; protein transport; site directed mutagenesis; small interfering RNA; transcription factor; transfection; western blottings

DESCRIPTION (provided by applicant): Apoptosis, or programmed cell death, is a biochemical pathway critical to normal development and tissue homeostasis in multi-cellular organisms. Aberrant suppression of apoptosis is a requisite contributor to human cancer. The long-term objective of this application is to elucidate the intracellular pathways and the molecular mechanisms that lead to apoptosis. The Bcl-2 family proteins are major regulators of apoptosis. Genetic and biochemical studies establish that two of the Bcl-2 family proteins, Bax and Bak, are requisite mediators of the mitochondria-dependent apoptotic pathway. In order to initiate apoptosis, Bax needs to undergo an activation process, which involves its mitochondrial translocation, the formation of Bax homo-oligomers, and the subsequent damage of mitochondria, a hallmark of apoptosis. The mechanisms of this activation process remain poorly understood. Using techniques of biochemistry and molecular biology, this proposal aims at determining the molecular mechanisms of Bax activation during apoptosis. In Aim 1, we will determine the mechanism of mitochondrial translocation of Bax. The role of mitochondrial targeting sequence(s), conformational changes, and dimeric states of Bax in its mitochondrial translocation will be examined. In Aim 2, the helices and amino acid residues critical for the oligomerization of Bax will be defined. Mutational study and biochemical assays will be used to examine the role of homo-oligomerization in the apoptotic activity of Bax. In Aim 3, we will determine the mechanism of Bax activation by upstream activators. In addition, loss-of-function studies using siRNA will be combined with in vitro studies to examine the role of other Bcl-2 family proteins in the activation of Bax during apoptosis. These studies will determine the molecular mechanisms of Bax activation and will help elucidate the pathways that lead to apoptosis. Upon a successful completion, this proposal will not only provide basic knowledge on the regulation of apoptosis, but may also provide new insights on the design of therapeutic agents against cancer.

描述（由申请人提供）：细胞凋亡或程序性细胞死亡是对多细胞生物体的正常发育和组织稳态至关重要的生化途径。细胞凋亡的异常抑制是人类癌症的必要因素。该应用的长期目标是阐明导致细胞凋亡的细胞内途径和分子机制。 Bcl-2 家族蛋白是细胞凋亡的主要调节因子。遗传和生化研究表明，Bcl-2 家族蛋白中的两种蛋白 Bax 和 Bak 是线粒体依赖性细胞凋亡途径的必要介质。为了启动细胞凋亡，Bax 需要经历一个激活过程，该过程涉及其线粒体易位、Bax 同源寡聚物的形成以及随后的线粒体损伤（细胞凋亡的标志）。这种激活过程的机制仍然知之甚少。该提案旨在利用生物化学和分子生物学技术，确定细胞凋亡过程中 Bax 激活的分子机制。在目标 1 中，我们将确定 Bax 线粒体易位的机制。将检查线粒体靶向序列、Bax 构象变化和二聚体状态在线粒体易位中的作用。在目标 2 中，将定义对 Bax 寡聚化至关重要的螺旋和氨基酸残基。突变研究和生化检测将用于检查同源寡聚化在 Bax 凋亡活性中的作用。在目标 3 中，我们将确定上游激活剂激活 Bax 的机制。此外，使用 siRNA 的功能丧失研究将与体外研究相结合，以检查其他 Bcl-2 家族蛋白在细胞凋亡过程中 Bax 激活中的作用。这些研究将确定 Bax 激活的分子机制，并有助于阐明导致细胞凋亡的途径。一旦成功完成，该提案不仅将提供细胞凋亡调控的基础知识，还可能为癌症治疗药物的设计提供新的见解。