Chemical Probes and Chaperone-Accelerated Turnover of Tau

描述(申请人提供)：tau是一种微管结合蛋白，本质上是无序的，在神经元中大量表达。某些翻译后修饰形式的tau，如过度磷酸化和蛋白分解片段，已被发现在超过15种神经退行性疾病中积累，包括阿尔茨海默病(AD)。这些“异常的”tau变体特别容易聚集，它们的积累会导致蛋白毒性、神经元丢失和认知障碍。因此，加速去除异常tau的策略有望预防tau病。细胞蛋白质质量控制(PQC)途径，包括分子伴侣，已被证明通过控制tau的翻译后修饰、与微管的结合和其周转来调节tau的动态平衡。然而，这一过程的分子细节尚不清楚。伴侣蛋白如何区分正常的tau和易于聚集的tau？哪些监护人和共同监护人对分诊决定至关重要？在tau降解的最早阶段，哪些分子事件是至关重要的？这个项目的主要目标是了解PQC的分子伴侣和其他成分是如何决定降解tau及其变体的。初步证据表明，热休克蛋白Hsp70和Hsp90与它们的一些相关辅助伴侣结合在一起，可以检测到容易聚集的tau的出现，然后做出分类决定来移除这些蛋白质。基于这些研究，我们假设伴侣和辅助伴侣的特定组合结合tau，区分变体，然后制定降解计划。为了探索这一模型，我们提出了两个具体的目标：(1)对正常的、循环中的tau和被精确定位于蛋白酶体降解的tau进行免疫共沉淀和质谱学研究，以确定与降解命运特定相关的蛋白质；(2)测试特定的Hsp40辅助伴侣是否选择性地将Hsp70招募到易于聚集的tau变体。这些研究是及时的，因为他们利用了新的化学探针，通过泛素-蛋白酶体途径触发tau的快速(~10分钟)和戏剧性(~80%)降解。Gestwicki实验室在过去五年里一直在开发、表征和优化这套化合物，并与Dickey团队一起，在细胞和动物模型中展示了它们的活性。我们建议，由于它们异常快速的活动，这些化学探针将首次允许 时间，洞察最早阶段的决定，降级牛磺酸。因此，我们期望这些 AIMS将深入了解伴侣复合体如何区分tau变体，以及蛋白质质量控制组件如何穿梭异常tau进行降解。这项工作的长期目标是通过了解控制tau周转的关键途径，找到治疗AD和其他tau病的新方法。

DESCRIPTION (provided by applicant): Tau is a microtubule-binding protein that intrinsically disordered and abundantly expressed in neurons. Certain post-translationally modified forms of tau, such as hyper-phosphorylated and proteolyzed fragments, have been found to accumulate in more than fifteen neurodegenerative disorders, including Alzheimer's disease (AD). These "abnormal" tau variants are particularly prone to aggregation and their accumulation leads to proteotoxicity, neuron loss and cognitive impairment. Thus, strategies for accelerating the removal of abnormal tau are expected to protect against tauopathies. Cellular protein quality control (PQC) pathways, including molecular chaperones, have been shown to regulate tau homeostasis by controlling its post-translational modification, binding to microtubules and its turnover. However, the molecular details of this process are not yet clear. How do the chaperones distinguish between normal tau and aggregation-prone tau? Which chaperones and co-chaperones are critical for the triage decisions? What molecular events are critical during the earliest stages of tau degradation? The major goal of this project is to understand how the molecular chaperones and other components of PQC make the decision to degrade tau and its variants. Preliminary evidence suggests that the heat shock protein Hsp70 and Hsp90, in combination with some of their associated co-chaperones, detect the appearance of aggregation-prone tau and then make triage decisions to remove these proteins. Based on these studies, we hypothesize that a specific combination of chaperones and co-chaperones bind tau, distinguish between variants and then enact degradation programs. Towards probing that model, we propose two specific aims: (1) perform co-immunoprecipitation and mass spectrometry studies on normal, cycling tau and tau that has been acutely targeted for proteasomal degradation to identify proteins that are specifically associated with the degradation fate and (2) test whether specific Hsp40 co-chaperones selectively recruit Hsp70 to aggregation-prone tau variants. These studies are timely because they make use of new chemical probes that trigger the rapid (~ 10 min) and dramatic (~80%) degradation of tau by the ubiquitin-proteosome pathway. The Gestwicki laboratory has spent the past five years developing, characterizing and optimizing this suite of compounds and, together with the Dickey group, we have shown their activity in cellular and animal models of tauopathy. We propose that, because of their unusually rapid activities, these chemical probes will permit, for the first time, insight into the very earliest stages of the "decision" to degrade tau. Thus, we expect these aims will provide insight into how chaperone complexes discriminate between tau variants and how protein quality control components shuttle abnormal tau for degradation. The long-term goal of this work is to find new ways of treating AD and other tauopathies by understanding the critical pathways that control tau turnover.