Regulation of apoptosis by Bcl-XL, Bak and Bax

Bcl-XL、Bak 和 Bax 对细胞凋亡的调节

基本信息

批准号：
7921271
负责人：
Chi Li
金额：
$ 5万
依托单位：
UNIVERSITY OF LOUISVILLE
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-09-01 至 2010-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7921271
关键词：
Address Affect Agonist Animals Apoptosis Apoptotic Bax protein Biochemical Caspase Cell Line Cell Lineage Cell Membrane Permeability Cell physiology Cells Cessation of life Characteristics Development Embryo Endoplasmic Reticulum Fibroblasts Gene Expression Genes Genetic Global Change Goals Hematopoietic Homeostasis ITPR1 gene In Vitro Intracellular Membranes Ion Channel Maintenance Malignant Neoplasms Mediating Membrane Microarray Analysis Mitochondria Molecular Molecular Profiling Mus Mutation Nuclear Oligonucleotides Organelles Pathway interactions Pattern Play Property Protein Family Proteins RNA Interference Recombinants Regulation Research Research Project Grants Resistance Role Signal Transduction Stimulus Tissues bak protein bcl-xlong protein cell suicide cell type design neoplastic cell novel overexpression patch clamp pro-apoptotic protein programs reconstitution research study response

项目摘要

DESCRIPTION (provided by applicant): The development of cancer involves genetic changes leading to alterations in the control of tissue development and homeostatic maintenance, which is tightly regulated by the cellular suicide program, apoptosis. Many tumor cells are resistant to intrinsic and extrinsic apoptotic stimuli, probably due to impaired apoptotic pathways in these cells. Bcl-2 family proteins are major regulators of the cellular response to apoptotic signals. Among them, anti-apoptotic Bcl-XLand pro-apoptotic Bak and Bax have-been shown to play an essential role during apoptosis. Initial studies suggest that in addition to their functions at mitochondria, Bcl-2 proteins also localize on the endoplasmic reticulum (ER) and function to regulate a parallel pathway of apoptosis. Experiments in this proposal are designed to elucidate the detailed molecular mechanisms of Bcl-2 proteins regulating apoptosis. The following specific aims are proposed: 1) Investigate the electrophysiological properties of Bcl-XL, Bak and Bax on the ER membrane and their contribution to the induction of membrane permeabilization during apoptosis. A novel patch-clamp approach to record single ion channel activity will be used to characterize Bcl-2 proteins on the ER membrane. The electrophysiological properties of purified recombinant as well as reexpressed Bcl-XL, Bak and Bax proteins on the ER membrane will be studied. The hypothesis that Bak and Bax form lipidic pores on membranes during apoptosis will also be examined; 2) Study the distinct apoptotic pathways mediated by Bcl-2 family proteins localized on different organelles. Efforts will be made to better characterize caspase pathways mediated by Bak or Bcl-XL localized on the ER or mitochondria using a RNAi approach. The importance of intracellular localizations of pro- and anti-apoptotic Bcl-2 proteins for the regulation of apoptosis will be studied by selectively targeting these proteins to specific organelles. The possible role of Ca2+ in the regulation of distinct apoptotic pathways will be investigated; and 3) Determine whether the expression of Bak and Bax affects cellular physiology and thus cellular response to death signals. Oligonucleotide-based microarray analysis will be performed to examine the changes in gene expression patterns in response to Bak or Bax expression. Genes whose expression may be involved in apoptosis regulation will be further studied.

描述（由申请人提供）：癌症的发展涉及导致组织发育和稳态维持的控制发生改变的遗传变化，而组织发育和稳态维持受到细胞自杀程序（细胞凋亡）的严格调节。许多肿瘤细胞对内在和外在的凋亡刺激具有抵抗力，可能是由于这些细胞中的凋亡途径受损。 Bcl-2 家族蛋白是细胞对凋亡信号反应的主要调节因子。其中，抗凋亡的Bcl-XL和促凋亡的Bak和Bax已被证明在细胞凋亡过程中发挥重要作用。初步研究表明，除了在线粒体中发挥作用外，Bcl-2 蛋白还定位在内质网 (ER) 上，并发挥调节细胞凋亡平行途径的作用。本提案中的实验旨在阐明 Bcl-2 蛋白调节细胞凋亡的详细分子机制。提出以下具体目标： 1) 研究内质网膜上 Bcl-XL、Bak 和 Bax 的电生理特性及其在细胞凋亡过程中诱导膜透化的贡献。一种记录单离子通道活性的新型膜片钳方法将用于表征 ER 膜上的 Bcl-2 蛋白。将研究在内质网膜上纯化的重组蛋白以及重新表达的 Bcl-XL、Bak 和 Bax 蛋白的电生理特性。还将检验 Bak 和 Bax 在细胞凋亡过程中在膜上形成脂质孔的假设； 2) 研究位于不同细胞器上的Bcl-2家族蛋白介导的不同细胞凋亡途径。我们将努力使用 RNAi 方法更好地表征由位于 ER 或线粒体上的 Bak 或 Bcl-XL 介导的 caspase 通路。将通过选择性地将这些蛋白质靶向特定细胞器来研究促凋亡和抗凋亡 Bcl-2 蛋白的细胞内定位对于细胞凋亡调节的重要性。将研究 Ca2+ 在调节不同细胞凋亡途径中的可能作用； 3) 确定 Bak 和 Bax 的表达是否影响细胞生理学，从而影响细胞对死亡信号的反应。将进行基于寡核苷酸的微阵列分析，以检查基因表达模式响应 Bak 或 Bax 表达的变化。其表达可能参与细胞凋亡调控的基因将进一步研究。