Phase Transition-Mediated Tau Function and Dysfunction

项目摘要 相变介导的Tau功能和功能障碍 这项拟议的研究将破译液-液两相的分子机制。 分离(LLP)介导的Tau功能和功能障碍。分相Tau液滴 丰富微管蛋白和促进微管组装(Tau功能)，而持续液滴 导致蛋白质聚集。 利用之前在无序蛋白质系统方面的丰富经验，研究人员将 确定LLP如何在三个阶段将Tau功能丧失和毒性获得功能障碍联系起来： 它们将描述病理性的翻译后修饰(PTM； 过度磷酸化和高乙酰化)调节Tau LLPs介导的微管 组装和蛋白质聚集(目标1)；他们将跟踪对两者都至关重要的Tau构象 LLP介导的Tau功能和功能障碍(目标2)；并且，他们将决定如何共同 聚集蛋白质引发/促进/协同牛磺酸聚集(目标3)。 第一个目标将利用整体光谱学和时间推移显微镜技术。 了解PTMS在LLPs介导的功能和功能障碍中的作用。相变 将生成不同Tau变体的地图，以表征不同的PTM如何改变Tau 有限责任公司。Tau异构体在液滴或纤维聚集体中共分配对Tau功能的影响 还将对功能障碍进行研究。 第二个目标是利用超灵敏的荧光光谱技术来追踪 作为单体Tau形成凝聚液滴和作为液滴的构象转换 转变为固态，即功能性(在微管组装的细丝中)和/或 功能障碍状态(淀粉样原纤维状态)。 第三个目标将确定共聚集蛋白在Tau LLP和Tau LLP中的作用 随后的蛋白质聚集。共相分离蛋白之间的协同作用 将被描述为。聚合交叉播种的机制将对蛋白质有深入的了解 共同错误折叠。

Project Summary Phase Transition-Mediated Tau Function and Dysfunction The proposed research will decipher the molecular mechanism of liquid-liquid phase separation (LLPS)-mediated Tau function and dysfunction. Phase-separated Tau droplets enrich tubulin and facilitate microtubule assembly (Tau function), whereas persistent droplets lead to protein aggregation. Utilizing prior extensive experience with disordered protein systems, the investigators will determine how LLPS links Tau loss-of-function and gain-of-toxic dysfunction in three stages: They will characterize how pathologic post-translational modifications (PTMs; hyperphosphorylation and hyperacetylation) modulate Tau LLPS-mediated microtubule assembly and protein aggregation (Aim 1); they will track Tau conformations that are key to both LLPS-mediated Tau function and dysfunction (Aim 2); and, they will determine how co- aggregating proteins initiate/ facilitate/ synergize Tau aggregation (Aim 3). The first Aim will utilize ensemble spectroscopy and time-lapse microscopy techniques to understand the role of PTMs in LLPS-mediated function and dysfunction. Phase transition maps of different Tau variants will be generated to characterize how different PTMs alter Tau LLPS. Effects of Tau variant co-partitioning in droplets or fibrillar aggregates on Tau function and dysfunction will also be studied. The second Aim will utilize ultrasensitive fluorescence spectroscopy techniques to track conformational conversions as monomeric Tau forms condensed droplets and as liquid droplets transform to solid states, i.e., functional (in microtubule assembled filaments) and/or dysfunctional states (amyloid fibril states). The third Aim will determine the role of co-aggregating proteins in Tau LLPS and subsequent protein aggregation. Synergistic interaction between co-phase separating proteins will be characterized. Mechanism of aggregate cross-seeding will provide insights into protein co-misfolding.