Gene Regulation Mechanisms by the APP/Fe65/TIP60 Complex

APP/Fe65/TIP60 复合物的基因调控机制

• 批准号: 6887390
• 负责人: JOHN A.A. LADIAS
• 金额: $ 29.75万
• 依托单位: BETH ISRAEL DEACONESS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-05-01 至 2008-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6887390
• 关键词: X ray crystallography; acyltransferase; amyloid proteins; androgen receptor; biological signal transduction; clinical research; coenzyme A; confocal scanning microscopy; conformation; enzyme mechanism; estrogen receptors; gel mobility shift assay; gene expression; genetic regulation; immunoprecipitation; molecular genetics; protein protein interaction; protein structure; proteolysis; site directed mutagenesis; tissue /cell culture; transcription factor; transfection

DESCRIPTION (provided by applicant): Our long-term objective is to elucidate the molecular mechanisms of transcriptional regulation of human gene expression in response to Amyloid Precursor Protein (APP) signaling. APP is a transmembrane protein that is the precursor of the beta-amyloid peptide (Abeta), the major component of amyloid plaques that characterize Alzheimer's disease. This peptide is generated from APP through proteolysis by the beta- and gamma-secretases. Following proteolysis, the released 47-residue cytoplasmic domain of APP (APP-C) interacts with the proteins Fe65 and TIP60, forming a nuclear complex that regulates gene expression. However, despite the central importance of the APP-C in human physiology and Alzheimer's disease pathogenesis, the sequence-specific DNA-binding transcription factors that recruit the TIP60/Fe65/APP-C complex onto the promoters of human genes and the molecular mechanisms underlying the transcriptional regulation by this complex remain elusive. TIP60 has histone acetyltransferase (HAT) activity with unique substrate specificity and interacts with class I nuclear receptors, supporting the hypothesis that these transcription factors may mediate APP-C signaling in the nucleus. This proposal will test this hypothesis and will focus on a mechanistic analysis of the APP-C signaling in gene regulation. The Specific Aims are: 1) To dissect the molecular mechanisms of gene regulation by the TIP60/Fe65/APP-C complex through its association with the androgen and estrogen receptors, using protein-protein interaction assays, in combination with site-directed mutagenesis and transient transfection experiments. 2) To elucidate the structural basis of the TIP60 HAT substrate specificity and the structural determinants of the TIP60 interaction with the androgen and estrogen receptors. 3) To determine the three-dimensional conformational changes induced on the TIP60/Fe65 complex by the APP-C binding, using X-ray crystallography. These studies will provide high-resolution three-dimensional structures of the TIP60, Fe65, and APP-C proteins, and will provide mechanistic insights into the regulation of human gene expression in response to APP signaling. This information could be used for the development of novel strategies in the treatment of Alzheimer's disease.

描述（由申请人提供）：我们的长期目标是阐明响应淀粉样前体蛋白（APP）信号传导的人类基因表达转录调控的分子机制。 APP 是一种跨膜蛋白，是 β-淀粉样肽 (Abeta) 的前体，β-淀粉样肽是阿尔茨海默病特征性淀粉样蛋白斑的主要成分。该肽是通过 β 和 γ 分泌酶的蛋白水解作用从 APP 生成的。 蛋白水解后，释放的 APP (APP-C) 的 47 个残基胞质结构域与蛋白质 Fe65 和 TIP60 相互作用，形成调节基因表达的核复合物。然而，尽管 APP-C 在人类生理学和阿尔茨海默氏病发病机制中具有核心重要性，但将 TIP60/Fe65/APP-C 复合物募集到人类基因启动子上的序列特异性 DNA 结合转录因子及其潜在的分子机制该复合物的转录调控仍然难以捉摸。 TIP60 具有组蛋白乙酰转移酶 (HAT) 活性，具有独特的底物特异性，并与 I 类核受体相互作用，支持这些转录因子可能介导细胞核中 APP-C 信号传导的假设。该提案将检验这一假设，并将重点关注基因调控中 APP-C 信号传导的机制分析。具体目标是： 1) 使用蛋白质-蛋白质相互作用分析，结合定点诱变和瞬时转染实验，剖析 TIP60/Fe65/APP-C 复合物通过与雄激素和雌激素受体结合进行基因调控的分子机制。 2) 阐明TIP60 HAT底物特异性的结构基础以及TIP60与雄激素和雌激素受体相互作用的结构决定因素。 3) 使用X射线晶体学测定APP-C结合在TIP60/Fe65复合物上诱导的三维构象变化。 这些研究将提供 TIP60、Fe65 和 APP-C 蛋白的高分辨率三维结构，并将为响应 APP 信号传导而调节人类基因表达提供机制见解。该信息可用于开发治疗阿尔茨海默病的新策略。