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.

程序性细胞死亡或凋亡是一个内置的，信号诱导的过程 细胞自我毁灭。这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，编程的异常细胞死亡是 与肿瘤发生，神经退行性疾病有关 免疫缺陷和病毒发病机理。虽然凋亡途径 在各种生物之间高度保守，涉及中央死亡 调节器，执行和调节涉及的细胞机制 凋亡的凋亡在很大程度上是未知的。诱导凋亡的动物病毒 可以通过新型凋亡抑制子的表达来阻止细胞死亡。 因此，宿主凋亡病毒干预的机制具有 提供了对细胞死亡计划的关键见解。这是长期的 该提议定义分子机制的目的 细胞凋亡是通过对杆状病毒编码的研究来调节的 凋亡调节剂P35和IAP。我们的方法着重于使用 杆状病毒感染的昆虫细胞是一个强大但方便的系统 用于分子分析凋亡的诱导和抑制。在 一组集成的实验集，基于最新进展 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。广泛的功能丧失P35 突变将用于表征分子相互作用 p35和CED-3/与冰有关的死亡蛋白酶，p35是有效的 抑制剂。通过专注于无脊椎动物CED-3/冰样蛋白酶（S） 由杆状病毒感染诱导，我们还研究了机制 参与激活和抑制这些关键细胞死亡 酶。在遗传研究中，我们使用新型p35和IAP表达 杆状病毒重组和稳定转染的昆虫细胞 线条，确切地放置p35和VIAP的抗凋亡活性 进入保守的死亡道路。最后，我们使用p35和iap作为工具 定义大型DNA病毒触发凋亡的机制 它们的宿主细胞并有助于发病机理。总的来说，这些 期望研究将对机制产生重要的见解 哪些程序性细胞死亡受到调节，从而有助于长期 凋亡治疗的新型治疗策略的术语设计 - 联系的人类疾病。