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 分子和细胞机制，presenilins（PS1和PS2）和 相关分子介导蛋白质运输，γ-分泌酶活性和 AB生产。最近的研究表明，高分子量 含有PS的蛋白质复合物在AB生产中起关键作用。 但是，尚未阐明PS1复合物的分子组成。 在这方面，最近称为尼卡斯特林的I型膜蛋白是 被识别为PS1复合物的化学计量分量。实验 尼加斯特林的突变体产生AB的产生和Notch 1裂解，表明 该分子在调节伽马神分泌活性中的辅助作用。 在AIM 1下，我们建议表征细胞和亚细胞 尼加斯特林的分布，尼加斯特林和PS1的相互作用 删除诱变策略和评估PS1-Nicastrin的作用 γ-分泌酶活性的相互作用。最后，我们将调查 生化和 免疫细胞化学方法。除了它在调解中的作用 APP和Notch的膜内γ-分泌酶切割，PS1也已知 调节几种膜蛋白的贩运，包括应用和 神经营养蛋白受体TRKB。我们的初步结果表明 影响应用程序处理的实验PS1突变体也会改变亚细胞 应用程序，应用CTF，APLP2和TRKC的分布。 AIM 2的研究将 扩展这些调查以检查PS1在贩运方面的作用 APP，APP同源物，神经营养蛋白受体，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AIM 3下， 我们概述了识别和表征分子的生化策略 PS1复合物的组成部分和我们表征γ-分泌酶的方法 体外活性。这些后一项研究也将促进 伽马分泌酶的新底物的鉴定。集体， 提出的研究将提供新的见解：功能 在调节伽马分泌酶的调节中，尼卡斯特林和PS1之间的关系 活动; PS1居民高中包含的组件的身份 分子量复合物； PS1在调节中的功能 膜蛋白的细胞内运输和代谢。