AUTOANTIGEN CLEAVAGE DURING APOPTOSIS AND NECROSIS

细胞凋亡和坏死过程中自身抗原的裂解

基本信息

批准号：
6511148
负责人：
Carlos A. Casiano
金额：
$ 13.46万
依托单位：
LOMA LINDA UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
1998
资助国家：
美国
起止时间：
1998-04-01 至 2003-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/6511148
关键词：
antinuclear autoantibody apoptosis autoantigens cell free system endopeptidases enzyme activity human tissue laboratory mouse mercury monoclonal antibody necrosis proteolysis

项目摘要

Apoptosis and necrosis are two fundamental cell death pathways that may be the end result of response to physiologic activators, physical trauma, or environmental toxins and chemicals. These modes of cell death are associated with a wide range of human pathological conditions, including systemic autoimmunity, cancer, AIDS, and neurodegenerative disorders. Emerging evidence indicates that activation of cysteine proteases of the ICE/CED-3 family is a central mechanism in the execution of apoptosis. Although the mechanisms driving cell necrosis are still obscure, recent studies also point to activation of proteases as a key event in this cell death process. However, the nature of these proteases and their substrates remains largely unknown. Using human antinuclear autoantibodies as probes, we have demonstrated that both apoptosis and necrosis involve the selective, but distinctively different, proteolytic cleavage of a specific set of nuclear protein autoantigens. It is proposed that different proteolytic mechanisms underlie the execution of apoptosis and necrosis, and that dissecting these mechanisms should be helpful for establishing a clear distinction between these cell death processes. The broad, long term objectives of this research is to dissect and differentiate events associated with these mechanisms. Three specific aims are proposed to test our hypothesis. (1) To use a panel of antinuclear autoantibodies where the antigen targets are well characterized nuclear proteins, as probes to define and differentiate patterns of substrate proteolysis associated with Fas-mediated apoptosis and with necrosis induced by the toxic xenobiotic mercury. (2) To characterize proteolytic activities involved in the cleavage of specific autoantigens in both cell death processes. Cell free systems of apoptosis and necrosis will be used in these studies to analyze the inhibition profile of autoantigen cleavage and to determine whether activation of ICE/CED-3 proteases occurs during necrosis. The ability of purified proteases to cleave specific antigens and the identify of specific cleavage sites will be also investigated. (3) To exploit the differences in autoantigen cleavage observed in apoptosis and necrosis for the development of monoclonal antibodies which react specifically with epitopes associated with either apoptotic or necrotic cell death. It is anticipated that these studies should contribute to enhancing our understanding of cell death in molecular terms and establishing additional criteria that would help to differentiate between apoptosis and necrosis.

细胞凋亡和坏死是细胞死亡的两种基本途径是对生理激活剂、物理激活剂反应的最终结果外伤、或环境毒素和化学物质。细胞的这些模式死亡与多种人类病理状况有关，包括系统性自身免疫、癌症、艾滋病和神经退行性疾病失调。新出现的证据表明半胱氨酸的激活 ICE/CED-3家族的蛋白酶是一个核心机制细胞凋亡的执行。尽管驱动细胞坏死的机制仍然不清楚，最近的研究也指出蛋白酶的激活作为细胞死亡过程中的关键事件。然而，这些的性质蛋白酶及其底物仍然很大程度上未知。利用人类抗核自身抗体作为探针，我们已经证明两者细胞凋亡和坏死涉及选择性但独特的一组特定核蛋白的不同蛋白水解切割自身抗原。据推测，不同的蛋白水解机制是细胞凋亡和坏死执行的基础，并且解剖这些机制应该有助于建立明确的区分这些细胞死亡过程之间。广泛、长期的目标这项研究的目的是剖析和区分与这些机制。提出了三个具体目标来测试我们的假设。 (1) 使用一组抗核自身抗体，其中抗原靶标是已明确表征的核蛋白，作为探针定义和区分相关底物蛋白水解的模式 Fas 介导的细胞凋亡和毒性物质诱导的坏死外源汞。 (2) 表征所涉及的蛋白水解活性在两个细胞死亡过程中特定自身抗原的裂解。细胞凋亡和坏死的无细胞系统将用于这些研究分析自身抗原裂解的抑制谱以确定 ICE/CED-3 蛋白酶的激活是否发生在坏死。纯化的蛋白酶切割特定抗原的能力还将研究特定切割位点的识别。 (3) 利用观察到的自身抗原裂解的差异细胞凋亡和坏死用于单克隆抗体的开发与与凋亡相关的表位发生特异性反应或坏死细胞死亡。预计这些研究应该有助于增强我们对分子死亡的理解条款并建立有助于区分细胞凋亡和坏死。