REGULATION OF VIRUS INDUCED PROGRAMMED CELL DEATH

病毒诱导的程序性细胞死亡的调节

• 批准号: 6137202
• 负责人: PAUL D FRIESEN
• 金额: $ 19.78万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-01-01 至 2001-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6137202
• 关键词: Baculoviridae; apoptosis; endopeptidases; gene mutation; protease inhibitor; recombinant proteins; site directed mutagenesis; tissue /cell culture; transfection; virus infection mechanism; virus protein

Programmed cell death or apoptosis is a built-in, signal-induced process by which a cell self-destructs. It is a mechanism that provides control of cell numbers, and is therefore critical for normal development and tissue homeostasis. In humans, aberrant programmed cell death is associated with tumorigenesis, neurodegenerative diseases, immunodeficiency, and viral pathogenesis. Although the apoptotic pathway is highly conserved between diverse organisms and involves central death regulators, the cellular mechanisms involved in execution and regulation of apoptosis are largely unknown. Animal viruses that induce apoptosis can block cell death by the expression of novel apoptotic suppressors. The mechanisms for viral intervention of host apoptosis have therefore provided key insight into the cell death program. It is the long term objective of this proposal to define the molecular mechanisms by which apoptosis is regulated through the study of the baculovirus-encoded apoptotic regulators, p35 and iap. Our approach focuses on the use of baculovirus-infected insect cells as a powerful, yet convenient system for molecular analysis of the induction and suppression of apoptosis. In an integrated set of experiments that build on recent advances in programmed cell death, we use biochemical, genetic, and cell biology approaches to determine the molecular mechanism of P35 and vIAP anti- apoptotic activity. An extensive collection of loss-of-function p35 mutations will be used to characterize the molecular interaction between P35 and the CED-3/ICE-related death proteases, for which P35 is a potent inhibitor. By focusing on the invertebrate CED-3/ICE-like protease(s) induced by baculovirus infection, we also investigate the mechanisms involved in activation and inhibition of these critical cell death enzymes. In genetic studies, we use novel p35- and iap-expressing baculovirus recombinants, as well as stably transfected insect cell lines, to definitively place the anti-apoptotic activity of P35 and vIAP into the conserved death pathway. Lastly, we use p35 and iap as tools to define the mechanisms by which large DNA viruses trigger apoptosis in their host cell and contribute to pathogenesis. Collectively, these studies are expected to yield important insight into the mechanisms by which programmed cell death is regulated and thereby facilitate the long term design of novel therapeutic strategies for treatment of apoptosis- linked human disorders.

程序性细胞死亡或细胞凋亡是一个内置的、信号诱导的过程 细胞由此自我毁灭。它是一种提供控制的机制 细胞数量，因此对于正常发育和 组织稳态。 在人类中，异常的程序性细胞死亡是 与肿瘤发生、神经退行性疾病相关， 免疫缺陷和病毒发病机制。虽然凋亡途径 在不同生物体之间高度保守并涉及中枢死亡 调节器，参与执行和调节的细胞机制 细胞凋亡的发生很大程度上是未知的。诱导细胞凋亡的动物病毒 可以通过表达新型细胞凋亡抑制剂来阻止细胞死亡。 因此，病毒干预宿主细胞凋亡的机制 提供了对细胞死亡程序的关键见解。这是长期的 该提案的目标是定义分子机制 通过杆状病毒编码的研究来调节细胞凋亡 凋亡调节因子 p35 和 iap。我们的方法侧重于使用 杆状病毒感染的昆虫细胞是一个强大而方便的系统 用于诱导和抑制细胞凋亡的分子分析。在 一组基于最新进展的综合实验 程序性细胞死亡，我们使用生化、遗传和细胞生物学 确定 P35 和 vIAP 抗-分子机制的方法 凋亡活性。大量丧失功能的 p35 突变将用于表征之间的分子相互作用 P35 和 CED-3/ICE 相关死亡蛋白酶，其中 P35 是一种有效的 抑制剂。关注无脊椎动物 CED-3/ICE 样蛋白酶 由杆状病毒感染引起的，我们还研究了其机制 参与这些关键细胞死亡的激活和抑制 酶。在遗传学研究中，我们使用新型 p35 和 iap 表达 杆状病毒重组体以及稳定转染的昆虫细胞 系，以确定 P35 和 vIAP 的抗凋亡活性 进入保守的死亡途径。最后，我们使用 p35 和 iap 作为工具 定义大DNA病毒触发细胞凋亡的机制 它们的宿主细胞并有助于发病机制。总的来说，这些 研究预计将通过以下方式对机制产生重要的见解： 程序性细胞死亡受到调节，从而促进长期 治疗细胞凋亡的新治疗策略的术语设计- 相关的人类疾病。