AUTOANTIGEN CLEAVAGE DURING APOPTOSIS AND NECROSIS

细胞凋亡和坏死是两条基本的细胞死亡途径，可能 是对生理激活剂反应的最终结果，物理的 创伤，或环境毒素和化学物质。这些模式的细胞 死亡与广泛的人类病理状况有关， 包括系统性自身免疫、癌症、艾滋病和神经退行性变 精神错乱。新出现的证据表明，半胱氨酸的激活 ICE/CED-3家族的蛋白水解酶是 执行细胞凋亡。尽管导致细胞坏死的机制 仍然不清楚，最近的研究也指出了蛋白酶的激活 作为细胞死亡过程中的一个关键事件。然而，这些问题的性质 蛋白酶及其底物在很大程度上仍不为人所知。使用人类 抗核自身抗体作为探针，我们已经证明了两者 细胞凋亡和坏死涉及选择性的，但明显的 一组特定核蛋白的不同蛋白水解性切割 自身抗原。不同的蛋白质降解机制被提出。 细胞凋亡和坏死的执行，以及解剖 这些机制应该有助于建立明确的区分 在这些细胞死亡过程之间。的广泛、长期目标 这项研究是为了剖析和区分与 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 抗原靶标是具有良好特性的核蛋白，作为探针 定义和区分与底物蛋白分解相关的模式 Fas介导的细胞凋亡和毒素诱导的坏死 异种生物汞。(2)鉴定所涉及的蛋白分解活性 在两个细胞死亡过程中特定的自身抗原的裂解。 无细胞的细胞凋亡和坏死系统将用于这些 分析自身抗原裂解抑制率的研究 确定ICE/CED-3蛋白水解酶是否在 坏死。纯化的蛋白水解酶裂解特定抗原的能力 还将对特定切割位点的鉴定进行调查。 (3)利用观察到的自身抗原裂解的差异 细胞凋亡和坏死与单抗的形成 它们与与两种细胞凋亡相关的表位发生特异性反应 或坏死性细胞死亡。预计这些研究应 有助于加深我们对分子中细胞死亡的理解 条款和建立其他标准，这将有助于 区分细胞凋亡和坏死。

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.