Cell Biology of Presenilin 1 and Associated Proteins

早老素 1 和相关蛋白的细胞生物学

• 批准号: 6755908
• 负责人: GOPAL THINAKARAN
• 金额: $ 32.63万
• 依托单位: UNIVERSITY OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-05-01 至 2007-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6755908
• 关键词: amyloid proteins; biochemistry; cell biology; cell line; endopeptidases; enzyme activity; genetically modified animals; glycoproteins; immunoaffinity chromatography; immunocytochemistry; mass spectrometry; membrane proteins; presenilin; protein biosynthesis; protein localization; protein protein interaction; protein transport

DESCRIPTION (provided by applicant): This project seeks to clarify the molecular and cellular mechanisms by which Presenilins (PS1 and PS2) and associated molecules mediate protein trafficking, gamma-secretase activity and AB production. Recent studies have revealed that high molecular weight protein complexes containing PS play a critical role in AB production. However, the molecular composition of PS1 complexes has not been elucidated. In this regard, a type I membrane protein, termed nicastrin, was recently identified as a stoichiometric component of the PS1 complex. Experimental mutants of nicastrin effect AB production and Notch 1 cleavage, suggesting an accessory role for this molecule in regulating gamma secretase activity. Under Aim 1, we propose to characterize the cellular and subcellular distributions of nicastrin, the interaction of nicastrin and PS1 using deletion mutagenesis strategies, and assessment of the role of PS1-nicastrin interactions on gamma-secretase activity. Finally, we will investigate the co-distribution of APP, APP B-CTF, PS1 and nicastrin by biochemical and immunocytochemical methods. In addition to its role in mediating intramembranous gamma-secretase cleavage of APP and Notch, PS1 is also known to regulate the trafficking of several membrane proteins, including APP and the neurotrophin receptor, TrkB. Our Preliminary Results show that experimental PS1 mutants that effect APP processing also alter the subcellular distributions of APP, APP CTFs, APLP2 and TrkC. Studies under Aim 2 will extend these investigations to examine the role of PS1 on the trafficking of APP, APP homologues, neurotrophin receptors, low-density lipoprotein receptor-related protein (LRP), transferrin receptor, and AMPA receptors. Under Aim 3, we outline biochemical strategies to identify and characterize the molecular components of PS1 complexes and our approaches to characterize gamma-secretase activity in vitro. These latter studies will also facilitate the identification of novel substrates of gamma-secretase. Collectively, the studies proposed will provide new insights into: the functional relationship between nicastrin and PS1 in modulation of gamma-secretase activity; the identity of components contained within PS1-resident high molecular weight complexes; and the function of PS1 in regulating intracellular trafficking and metabolism of membrane proteins.

描述(由申请人提供)：本项目旨在澄清 早老素(PS1和PS2)和 相关分子介导蛋白质运输、伽马分泌酶活性和 AB生产。最近的研究表明，高分子量 含有PS的蛋白质复合体在AB的产生中起着至关重要的作用。 然而，PS1复合体的分子组成尚不清楚。 在这方面，一种被称为尼古丁的I型膜蛋白最近被发现 被鉴定为PS1复合体的化学计量成分。实验 尼古丁影响AB产生和Notch 1裂解的突变体，表明 该分子在调节伽马分泌酶活性中起辅助作用。 在目标1下，我们建议描述细胞和亚细胞的特征 尼氏胃泌素的分布及其与PS1相互作用的研究 缺失突变策略和PS1-尼古丁的作用评估 相互作用对γ-分泌酶活性的影响。最后，我们将调查 APP、APP B-CTF、PS1和NICASTIN的生化和免疫共分布 免疫细胞化学方法。除了它在调解方面的作用之外 APP和Notch、PS1的膜内伽马分泌酶裂解也是已知的 调节几种膜蛋白的运输，包括APP和 神经营养素受体TrkB。我们的初步结果显示， 影响APP处理的实验性PS1突变体也改变了亚细胞 APP、APP CTF、APLP2和TrkC的分发。目标2下的研究将 扩大这些调查以审查PS1在贩运毒品方面的作用 APP、APP同系物、神经营养素受体、低密度脂蛋白受体相关 蛋白质(LRP)、转铁蛋白受体和AMPA受体。在目标3下， 我们概述了鉴定和表征分子的生化策略。 PS1复合体的组成及其表征伽马分泌酶的方法 体外活性。后者的研究也将有助于 γ-分泌酶新底物的鉴定。总体而言， 建议的研究将为以下方面提供新的见解： 尼古丁与PS1在调节γ-分泌酶中的关系 活动；PS1中包含的组件的标识-驻留高 分子量复合体以及PS1在调节中的作用 膜蛋白在细胞内的运输和代谢。