Molecular mechanism of Presenilin-1 (PS1) linked Alzheimer's Disease pathology

Presenilin-1 (PS1) 与阿尔茨海默病病理学相关的分子机制

• 批准号: 7844858
• 负责人: OKSANA BEREZOVSKA
• 金额: $ 23.48万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-05-01 至 2011-12-01
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7844858
• 关键词: AP40; APP-PS1; Active Sites; Adopted; Affect; Age of Onset; Alzheimer' s Disease; Amyloid beta-Protein Precursor; Biochemical; Biological; Biological Assay; Biology; Brain; Catalytic Domain; Cell surface; Cells; Cleaved cell; Complex; Data; Dendritic Spines; Deposition; Disease; Drug Design; Elements; Fluorescence; Fluorescence Resonance Energy Transfer; Future; Generations; Genetic; Goals; Image; Integral Membrane Protein; Lead; Life; Link; Measures; Mediating; Methodology; Methods; Microscopy; Modeling; Molecular; Molecular Conformation; Monitor; Mus; Mutation; N-Methylaspartate; Neurons; Non-Steroidal Anti-Inflammatory Agents; Pathology; Peptide Hydrolases; Peptides; Pharmaceutical Preparations; Phenotype; Phosphotransferases; Physiological; Play; Presynaptic Terminals; Production; Property; Protein Conformation; Relative (related person); Research Personnel; Resolution; Role; Site; Stimulus; Structure; Substrate Interaction; Synapses; Techniques; Testing; Therapeutic; base; design; familial Alzheimer disease; fluorophore; insight; interest; member; mutant; neuronal cell body; notch protein; novel; presenilin; presenilin-1; presynaptic; programs; protein complex; research study; response; secretase; therapeutic target; trafficking

DESCRIPTION (provided by applicant): The long term goal of this application is to understand the molecular basis of presenilin (PS)-linked production of Ap40 and Ap42 in familial Alzheimer's disease (FAD). Very little is known about the structure of the PS/y-secretase complex due to the complexity and difficulty of crystallizing multipass transmembrane proteins. We hypothesize that PS/y-secretase exists in different conformations, leading to different alignments of APP with the catalytic site, resulting in the production of different AB species. Our observations, using a novel FRET based technique in intact cells as well as biochemical approaches; provide methods to examine PS1/y-secretase conformation in intact cells as well as to determine proximity of PS1 and y-secretase substrates, including APP and Notch. Our observations support a model with the following important, testable features: mature PS1 associates with other y-secretase complex members and adopts a conformation with N- and C-termini (NT and CT) close together; and the specific presentation of APP to the y-secretase active site changes under conditions favoring AB40 vs. AB42. We will test this model from several clinically important perspectives. FAD-linked PS1 mutations lead to elevations in Ap42/4o ratio, but the mechanism is unknown. Our preliminary data suggest that FAD PS1 mutations bring PS1 NT and CT even closer together, and that this represents a pathogenic change in conformation associated with an alteration in the alignment of the active site of PS1/y-secretase with APP, favoring cleavage at Ap42. We will test several models to define the precise molecular mechanism that leads to this observed change in conformation (Aiml). Some nonsteroidal anti-inflammatory drugs have been shown to selectively lower AB42, a phenotype opposite to FAD mutations. We will test the hypothesis that pharmacological agents that selectively alter the AB42/40 ratio can do so by allosterically modulating the conformation of PS1/y-secretase and PS1/APP interactions in the opposite way to FAD mutations. We will explore whether Ap42 lowering allosteric modulators of the y-secretase can "fix" the pathogenic FAD mutant PS1 conformation (Aim2). Finally, we propose to test the hypothesis that physiologic stimuli that activate y-secretase and/or its substrate selectively change PS1 /y-secretase interactions with substrates in neurons. We will take advantage of the high spatial resolution of the new FRET technique, Fluorescence Lifetime Imaging Microscopy (FLIM), which is uniquely suited for monitoring relative conformational changes of proteins and complexes in intact and/or live cells, to show in what subcellular compartments in intact cells these changes occur (Aim3), and ultimately aim to extend these studies to the mouse brain. The results obtained will provide important new insights into conformational changes in the PS1/y-secretase complexes and its interaction with substrates within a cellular context, and thus will help in the design of allosteric modulators of y-secretase function as a therapeutic approach. More generally, the assay of monitoring PS1 conformation and y-secretase -substrate proximity will help to help develop methodologies that may prove useful in understanding disease-related protein conformation changes.

描述（由申请人提供）：本申请的长期目标是了解家族性阿尔茨海默病（FAD）中早老素（PS）相关的Ap 40和Ap 42产生的分子基础。由于多通道跨膜蛋白结晶的复杂性和困难性，对PS/γ-分泌酶复合物的结构知之甚少。我们假设PS/γ-分泌酶以不同的构象存在，导致APP与催化位点的不同对齐，从而产生不同的AB物种。我们的观察，使用一种新的FRET为基础的技术在完整的细胞以及生化方法;提供的方法来检查PS1/γ-分泌酶构象在完整的细胞，以及确定接近PS1和γ-分泌酶底物，包括APP和Notch。我们的观察结果支持一个模型，具有以下重要的，可测试的功能：成熟的PS1协会与其他Y-分泌酶复合物的成员，并采取了构象与N-和C-末端（NT和CT）靠近在一起;和特定的介绍APP的Y-分泌酶活性位点的变化有利于AB 40与AB 42的条件下。我们将从几个临床重要的角度来测试这个模型。FAD连锁的PS1突变导致Ap 42/4 o比值升高，但机制尚不清楚。我们的初步数据表明，FAD PS1突变使PS1 NT和CT甚至更接近，这代表了一种致病性的构象变化，与PS1/γ-分泌酶的活性位点与APP的比对改变相关，有利于在Ap 42处裂解。我们将测试几种模型，以确定导致这种观察到的构象变化（Aiml）的精确分子机制。一些非甾体抗炎药已被证明选择性降低AB 42，一种与FAD突变相反的表型。我们将测试的假设，药理学代理，选择性地改变AB 42/40的比例可以这样做，通过变构调制的构象PS1/γ-分泌酶和PS1/APP的相互作用，在相反的方式FAD突变。我们将探讨是否Ap 42降低变构调节剂的γ-分泌酶可以“固定”致病FAD突变体PS1构象（Aim 2）。最后，我们建议测试的假设，生理刺激，激活γ-分泌酶和/或其基板选择性地改变PS1 /γ-分泌酶的相互作用，在神经元中的基板。我们将利用高空间分辨率的新FRET技术，荧光寿命成像显微镜（FLIM），这是唯一适合于监测完整和/或活细胞中蛋白质和复合物的相对构象变化，以显示在什么样的亚细胞区室在完整细胞中发生这些变化（Aim 3），并最终旨在将这些研究扩展到小鼠大脑。所获得的结果将提供重要的新的见解，在PS1/γ-分泌酶复合物的构象变化及其与底物的相互作用在细胞内的情况下，因此将有助于在设计的变构调节剂的γ-分泌酶功能作为一种治疗方法。更一般地说，监测PS1构象和γ-分泌酶-底物接近度的测定将有助于开发可能有助于理解疾病相关蛋白质构象变化的方法。

项目成果

N-cadherin-based adhesion enhances Abeta release and decreases Abeta42/40 ratio.

基于 N-钙粘蛋白的粘附增强 Abeta 释放并降低 Abeta42/40 比率。

• DOI: 10.1111/j.1471-4159.2008.05760.x
• 发表时间: 2009
• 期刊: Journal of neurochemistry
• 影响因子: 4.7
• 作者: Uemura,Kengo;Lill,ChristinaM;Banks,Mary;Asada,Megumi;Aoyagi,Nobuhisa;Ando,Koichi;Kubota,Masakazu;Kihara,Takeshi;Nishimoto,Takaaki;Sugimoto,Hachiro;Takahashi,Ryosuke;Hyman,BradleyT;Shimohama,Shun;Berezovska,Oksana;Kinoshita,A
• 通讯作者: Kinoshita,A