AN EXTRACELLULAR HUMAN INHIBITOR OF APOPTOSIS

细胞外人类细胞凋亡抑制剂

• 批准号: 6073837
• 负责人: PAUL Clifton LEAVIS
• 金额: $ 10.59万
• 依托单位: ANALYTICAL BIOTECHNOLOGY SERVICES
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-03-15 至 2000-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6073837
• 关键词: apoptosis; biological signal transduction; extracellular; genetic library; heat shock proteins; high performance liquid chromatography; inhibitor /antagonist; molecular cloning; oligonucleotides; protein purification; protein sequence; tissue /cell culture

The broad, long term objective of this project is to develop therapeutic approaches to block apoptosis. The major inducible heat shock protein Hsp72 protects cells from most stresses and thus represents a generalized repair/stress resistance component. Hsp72 not only refolds damaged proteins but also interferes with the cell death signaling pathway thus providing cells with time to repair the damage. A protein factor (HIF) from human ascites fluid activated Hsp72 synthesis in cell cultures. HIF protected cells from apoptosis caused by heat shock, mutagen treatment, and oxidative damage. Isolation and cloning of this extracellular apoptosis inhibitor is the goal of this study. The specific aims of this study are: 1. We will purify the anti-apoptotic (HIF). A number of chromatographic approaches will be used in a bioassay-driven purification of human HIF. Expression of Hsp72 by fibroblasts will be used as a test The purity of active fractions will be tested by SDS-PAGE and 2-D electrophoresis. 2. We will clone the HIF gene. When HIF is purified, we will use a partial protein sequence to prepare degenerate oligonucleotides to identify the gene in a human genomic library. The clone will be partially sequenced, and a specific probe will be used to screen a cDNA library. The coding region will be completely sequenced, and this will provide useful information regarding the nature of the factor. Significance. Establishing the nature of a systemic apoptotic inhibitor that promotes cell repair will lay the basis for learning how an organism controls the fate of its cells. Relevance to human health. A natural extracellular inhibitor of apoptosis will serve as a basis for developing effective therapies for treating such stress- related conditions as ischemia and neurodegenerative disorders, where apoptosis in response to damage is an important factor in disease. PROPOSED COMMERCIAL APPLICATIONS: The research will lead to the identification of a natural human inhibitor of apoptosis that will be used to develop therapies against a number of diseases, such as ischemia and neurodegenerative disorders.

该项目的长远目标是开发阻断细胞凋亡的治疗方法。主要的诱导热休克蛋白Hsp72保护细胞免受大多数应激，因此代表了一个广泛的修复/抗逆性成分。Hsp72不仅可以重新折叠受损蛋白，还可以干扰细胞死亡信号通路，从而为细胞修复损伤提供时间。人腹水蛋白因子（HIF）在细胞培养中激活Hsp72合成。HIF保护细胞免受热休克、诱变剂处理和氧化损伤引起的细胞凋亡。分离和克隆这种细胞外细胞凋亡抑制剂是本研究的目的。本研究的具体目的是：1。我们将纯化抗凋亡细胞（HIF）。许多色谱方法将用于生物测定驱动的人类HIF纯化。成纤维细胞表达Hsp72作为检测，活性组分的纯度将通过SDS-PAGE和2-D电泳检测。2. 我们将克隆HIF基因。当HIF被纯化后，我们将使用部分蛋白序列制备退化寡核苷酸，以在人类基因组文库中鉴定该基因。将对克隆进行部分测序，并使用特定探针筛选cDNA文库。编码区将被完全测序，这将提供有关因子性质的有用信息。的意义。建立促进细胞修复的系统性凋亡抑制剂的性质将为了解生物体如何控制其细胞的命运奠定基础。与人类健康有关。一种天然的细胞外细胞凋亡抑制剂将成为开发有效疗法的基础，用于治疗诸如缺血和神经退行性疾病等与应激相关的疾病，其中细胞凋亡对损伤的反应是疾病的重要因素。拟议的商业应用：该研究将导致鉴定一种天然的人类细胞凋亡抑制剂，该抑制剂将用于开发针对许多疾病的治疗方法，如缺血和神经退行性疾病。