Trafficking and Proteolysis of Notch and Other Gamma-Secretase Substrates

Notch 和其他γ-分泌酶底物的运输和蛋白水解

• 批准号: 7808761
• 负责人: Mark E Fortini
• 金额: $ 31.74万
• 依托单位: THOMAS JEFFERSON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-05-01 至 2013-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7808761
• 关键词: Affect; Alzheimer' s Disease; Amyloid; Amyloid beta-Protein Precursor; Animals; Aspartic Endopeptidases; Biochemical; Biogenesis; Biological; Biology; Brain; Cell Adhesion Molecules; Cell physiology; Cell surface; Cells; Chromosomes; Cleaved cell; Complex; Data; Development; Disease; Drosophila genus; Early Endosome; Endocytosis; Endosomes; Event; Excision; Family; Genes; Genetic; Genetic Screening; Goals; Human; Immune system; Integral Membrane Protein; Intracellular Membranes; Ions; Lead; Ligand Binding; Ligands; Light; Lysosomes; Malignant Neoplasms; Mammalian Cell; Mediating; Membrane; Membrane Protein Traffic; Molecular; Molecular Cloning; Mutation; Neurodegenerative Disorders; Neurons; Pathogenesis; Pathway interactions; Peptides; Play; Process; Production; Proteins; Proteolysis; Proteolytic Processing; Receptor Signaling; Recovery; Recycling; Research Proposals; Role; Route; Screening Result; Signal Pathway; Signal Transduction; Sorting - Cell Movement; T-Cell Leukemia; T-Lymphocyte; Water; amyloid precursor protein processing; brain tissue; cell fate specification; gamma secretase; human disease; human tissue; interest; member; mutant; notch protein; novel; public health relevance; secretase; solute; trafficking; tumorigenesis; water channel

DESCRIPTION (provided by applicant): Many integral membrane proteins have been identified as cleavage substrates for the intramembrane aspartyl protease complex termed ??secretase. These ??secretase substrates include receptors for signaling pathways, adhesion molecules, and other factors that are localized at the cell surface and whose activities are regulated by ??secretase-mediated cleavage. Understanding the molecular details of ??secretase substrate biology is important due to the prominent role that many substrates play in normal cellular physiology and in human disease. For example, the Notch receptor, a well- characterized ??secretase substrate, is essential for proper cell-fate specification and cellular differentiation during the development of animal and human tissues. Inappropriate activation of the Notch pathway in the human immune system causes T-cell leukemia and dysregulated Notch activity has been implicated in other cancers and developmental diseases. Furthermore, processing of the Amyloid Precursor Protein (APP) by ??secretase contributes directly to the production of secreted amyloid-2 peptide in human brain tissue, a crucial event in the pathogenesis of Alzheimer's disease. A key feature of ??secretase substrate cleavage is its tight control by ligand binding, ectodomain removal, and other regulatory processes. A growing body of data indicates that cleavage also depends upon intracellular membrane trafficking of ??secretase substrates and their sorting within specific trafficking compartments. In the case of both Notch and APP, ??secretase-mediated cleavage is associated with endocytosis of the substrate from the cell surface and its entry into endosomes. We have found that proper biogenesis of the early endosome compartment requires the function of an aquaporin channel protein, defining a specialized endosome route for efficient ??secretase-dependent Notch trafficking and signaling. In Specific Aim 1, we propose to characterize further the role of this aquaporin in endosome biogenesis and Notch trafficking. These studies will shed light on the cell biological aspects of ?? secretase substrate trafficking and their relationship to ??secretase substrate cleavage, recycling, and degradation. In Specific Aim 2, we propose to conduct a forward genetic screen for new factors involved in the intracellular membrane trafficking of Notch and other ??secretase substrates. Preliminary results from this screen have already proven to be very promising, including the recovery of many potentially new genes needed for various steps of Notch intracellular trafficking. Further analysis and molecular cloning of several of these new genes is now underway. The main goal of this project is to advance our understanding of ??secretase substrate trafficking and its relevance to secretase- dependent processes in tumorigenesis and neurodegenerative disease. PUBLIC HEALTH RELEVANCE: Proteins cleaved by ??secretase, including the Notch receptor and amyloid precursor protein, are important for developmental signaling and neuronal function, and their dysregulation contributes to cancer and neurodegenerative disease. This research proposal seeks to identify cellular factors that control the production, trafficking, and proteolytic processing of ?? secretase targets. These studies might lead to the identification of new proteins and intracellular pathways that could be therapeutically targeted in cancer and neurodegenerative disorders.

描述（由申请人提供）：许多整合膜蛋白已被鉴定为膜内天冬氨酰蛋白酶复合物（称为β分泌酶）的切割底物。这些分泌酶底物包括信号传导途径的受体、粘附分子和位于细胞表面的其他因子，其活性受到分泌酶介导的裂解的调节。由于许多底物在正常细胞生理学和人类疾病中发挥着突出作用，因此了解分泌酶底物生物学的分子细节非常重要。例如，Notch 受体是一种经过充分表征的分泌酶底物，对于动物和人体组织发育过程中正确的细胞命运规范和细胞分化至关重要。人类免疫系统中 Notch 通路的不当激活会导致 T 细胞白血病，而 Notch 活性失调与其他癌症和发育性疾病有关。此外，分泌酶对淀粉样蛋白前体蛋白 (APP) 的加工直接有助于人脑组织中分泌型淀粉样蛋白 2 肽的产生，这是阿尔茨海默病发病机制中的一个关键事件。分泌酶底物裂解的一个关键特征是其通过配体结合、胞外域去除和其他调控过程的严格控制。越来越多的数据表明，裂解还取决于分泌酶底物的细胞膜内运输及其在特定运输区室中的分类。就Notch和APP而言，分泌酶介导的裂解与细胞表面的底物的内吞作用及其进入内涵体相关。我们发现早期内体区室的正确生物发生需要水通道蛋白通道蛋白的功能，从而定义了一种专门的内体途径，以实现有效的分泌酶依赖性Notch运输和信号传导。在具体目标 1 中，我们建议进一步表征该水通道蛋白在内体生物合成和 Notch 运输中的作用。这些研究将揭示 ?? 的细胞生物学方面。分泌酶底物运输及其与分泌酶底物裂解、回收和降解的关系。在具体目标 2 中，我们建议对参与 Notch 和其他分泌酶底物细胞内膜运输的新因子进行正向遗传筛选。该筛选的初步结果已被证明非常有希望，包括恢复了 Notch 细胞内运输各个步骤所需的许多潜在新基因。目前正在进行对其中几个新基因的进一步分析和分子克隆。该项目的主要目标是增进我们对分泌酶底物运输及其与肿瘤发生和神经退行性疾病中分泌酶依赖性过程的相关性的理解。公共健康相关性：由分泌酶裂解的蛋白质，包括 Notch 受体和淀粉样前体蛋白，对于发育信号传导和神经元功能非常重要，它们的失调会导致癌症和神经退行性疾病。该研究计划旨在确定控制 ?? 的生产、运输和蛋白水解加工的细胞因素。分泌酶目标。这些研究可能会导致新蛋白质和细胞内途径的鉴定，这些新蛋白质和细胞内途径可以作为癌症和神经退行性疾病的治疗靶点。