Molecular Basis of the Tau Aggregation Pathway

Tau 聚集途径的分子基础

基本信息

批准号：
9895602
负责人：
Songi Han
金额：
$ 52.95万
依托单位：
UNIVERSITY OF CALIFORNIA SANTA BARBARA
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-04-01 至 2022-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9895602
关键词：
Address Alternative Splicing Alzheimer&apos s Disease Alzheimer&apos s disease pathology Antibodies Appearance Automobile Driving Binding Biological Biophysics Cell model Cell surface Cells Clinical Complex Coupled Cytoplasm Cytoplasmic Granules Data Deposition Detection Disease Electron Microscopy Electron Spin Resonance Spectroscopy Electrostatics Exposure to Foundations Goals Grain Heparin Human Hydrophobicity Immune In Vitro Inclusion Bodies Kinetics Knowledge Label Laboratories Learning Length Liquid substance Microtubule Stabilization Modeling Molecular Molecular Chaperones Molecular Conformation Molecular Probes Monoclonal Antibodies Morphology Mutation Nature Nerve Degeneration Neurodegenerative Disorders Neurons Pathologic Pathology Pathway interactions Phase Physiologic pulse Physiological Population Post-Translational Protein Processing Protein Conformation Proteins RNA RNA-Binding Proteins Research Research Personnel Rest Role Route Seeds Site Sodium Chloride Solvents Spin Labels Stress Structure Sulfate Surface System Tauopathies Testing Transfer RNA Translations Treatment Efficacy Variant Veins Work aggregation pathway base beta pleated sheet conformer design genetic variant guided inquiry heparin proteoglycan in silico in vivo induced pluripotent stem cell innovation knowledge base mRNA Differential Displays microscopic imaging molecular dynamics mutant nanometer novel sarkosyl self assembly shape analysis simulation tau Proteins tau aggregation tau conformation tau interaction tau mutation tool

项目摘要

PROJECT SUMMARY Tau is a microtubule-stabilizing protein that is abundant in neurons. It is a highly soluble, intrinsically disordered protein (IDP) with little tendency for aggregation under native conditions. However, under several experimental conditions and in a variety of neurodegenerative disorders including Alzheimer’s disease, Tau can spread from cell to cell and aggregates as intra-cellular β-sheet fibrilar deposits. Our laboratories have critical new data concerning the temporal, structural and cell biological details of Tau misfolding and fluid-phase assembly—the basis of this proposal. Our research team consists of a cell biologist, a physical chemist, and a theoretical biophysicist. Working together closely in an iterative manner we intend to determine the pathway from normal Tau to disease-related Tau fibrils. The tools for this analysis include (a) cellular systems capable of addressing in vivo Tau interactions, and indirectly its conformational state based on a variety of molecular probes;; (b) site- directed spin labeling, electron paramagnetic resonance (EPR) line shape analysis and pulsed dipolar EPR to determine conformational signatures of Tau;; and (c) fully atomistic modeling of IDP conformations, their populations and energetics, and coarse-grained simulation of higher-order assemblies of Tau. The conceptual flow of the proposal begins with a remarkable observation from the Han lab: When exposed to sub-stoichiometric amounts of heparin, segments of Tau dramatically extend by a nanometer to solvent-expose the hydrophobic PHF6(*) segment capable of stacking into neat β-sheets. This observation correlates with the appearance of fibrils, and thus we refer to this initiating step as “on pathway” seeding. In vivo, Tau is known to populate a vast conformational landscape controlled by alternative splicing, mutations and post-translational modifications. We propose that the IDP Tau populates an ensemble of different conformations with different aggregation propensities, fibril morphologies and interaction partners, depending on the exact Tau variant. However, the defining and specific conformational signatures within this ensemble are unknown. Determining the conformational signatures of aggregation-prone Tau variants is our core objective, while a missing puzzle piece in connecting Tau conformation to cellular interactions is the existence and nature of aggregation intermediates. In this vein, the Han lab discovered that RNA induces liquid-liquid phase separation of Tau in vitro into protein droplets held together by weak electrostatic forces. At the in vivo cellular level, the Kosik lab discovered Tau- tRNA complexes, thereby adding Tau to the growing list of RNA-binding proteins involved in neurodegeneration, and capable of establishing liquid-liquid phase separation in the cytoplasm. The Tau-tRNA complexes may be a physiologic or pathological entity—we will obtain clues by determining their loci in neuronal cells. Finally, we intend to learn whether the conformation of Tau, as modulated by disease mutations or co-factors, influences the stability and in vivo locality of the Tau-tRNA complexes. Our goal is to discover a detailed route from soluble Tau to fibrils, from the nanometer to the cellular level, and discover the pathological entities of Tau aggregation.

项目概要 Tau 是一种微管稳定蛋白，在神经元中含量丰富。它是一种高度可溶的、本质上无序的蛋白。蛋白质（IDP）在天然条件下几乎没有聚集倾向。然而，在几个实验下在包括阿尔茨海默病在内的各种神经退行性疾病中，Tau 蛋白可以从我们的实验室拥有重要的新数据。关于 Tau 错误折叠和液相组装的时间、结构和细胞生物学细节 - 我们的研究团队由一名细胞生物学家、一名物理化学家和一名理论家组成。生物物理学家以迭代的方式密切合作，我们打算确定正常的途径。 Tau 到疾病相关的 Tau 原纤维。用于此分析的工具包括 (a) 能够处理的细胞系统。体内 Tau 相互作用，以及基于各种分子探针的间接构象状态；(b) 位点- 定向自旋标记、电子顺磁共振 (EPR) 线形分析和脉冲偶极 EPR 确定 Tau 的构象特征；以及 (c) IDP 构象的完全原子建模及其种群和能量学，以及 Tau 高阶组装的粗粒度模拟。该提案的流程始于汉实验室的一个显着观察：当暴露于亚化学计量时在一定量的肝素作用下，Tau 片段显着延伸一纳米，从而使疏水性暴露在溶剂中 PHF6(*) 片段能够堆积成整齐的 β-折叠，这一观察结果与的外观相关。纤维，因此我们将这一起始步骤称为“途径”播种在体内，Tau 蛋白会大量繁殖。由选择性剪接、突变和翻译后修饰控制的构象景观。提出 IDP Tau 填充具有不同聚合的不同构象的集合倾向、原纤维形态和相互作用伙伴，具体取决于 Tau 变体。该集合中的定义和具体构象特征是未知的。易于聚集的 Tau 变体的构象特征是我们的核心目标，同时也是一个缺失的拼图。将 Tau 构象与细胞相互作用联系起来的是聚集中间体的存在和性质。在这方面，Han 实验室发现 RNA 在体外诱导 Tau 液-液相分离成蛋白质。 Kosik 实验室在体内细胞水平上发现了 Tau-，通过微弱的静电力将液滴结合在一起。 tRNA 复合物，从而将 Tau 添加到参与神经退行性变的不断增长的 RNA 结合蛋白列表中，并能够在细胞质中建立液-液相分离。 Tau-tRNA 复合物可以是 a. 生理或病理实体——我们将通过确定它们在神经元细胞中的位点来获得线索。打算了解受疾病突变或辅助因子调节的 Tau 构象是否会影响 Tau-tRNA 复合物的稳定性和体内定位我们的目标是发现可溶性的详细途径。 Tau到原纤维，从纳米到细胞水平，发现Tau聚集的病理实体。