Elucidating the biophysics of pre-fibrillar, toxic tau oligomers: from amino acid motifs to neuronal dysfunction

阐明前原纤维有毒 tau 寡聚体的生物物理学：从氨基酸基序到神经元功能障碍

• 批准号: 10489810
• 负责人: Jonathan N Sachs
• 金额: $ 52.94万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-30 至 2024-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10489810
• 关键词: AMPA Receptors; Affect; Alzheimer' s Disease; Alzheimer' s disease related dementia; Amino Acid Motifs; Amino Acid Sequence; Binding; Biochemical; Biological Assay; Biophysics; Biosensor; Brain; CASP2 gene; Catalogs; Cell model; Cells; Cognitive; Cognitive deficits; Complex; Cryoelectron Microscopy; Data; Dendritic Spines; Disease; Distress; Electrostatics; Enzymes; Event; Feedback; Fluorescence; Fluorescence Resonance Energy Transfer; Goals; Human; Image; Investigation; Label; Lead; Length; Medicine; Molecular Structure; Monitor; Morphologic artifacts; Mus; Nature; Neurobiology; Neurofibrillary Tangles; Neuronal Dysfunction; Neurons; Paper; Pathologic; Pathology; Phosphorylation; Post-Translational Protein Processing; Proteins; Reagent; Research; Research Personnel; Resolution; Structure; Tauopathies; Technology; Testing; Therapeutic; Toxic effect; Translating; Work; alpha synuclein; base; biophysical analysis; comorbidity; design; dopaminergic neuron; experimental study; induced pluripotent stem cell; inhibitor; mutant; neuron loss; non-Native; novel; novel strategies; orientation selectivity; paired helical filament; prevent; protein protein interaction; rational design; small molecule; small molecule inhibitor; tau Proteins; tau aggregation; tau interaction; tau mutation; tau phosphorylation; therapeutic target; tool; virtual

Abstract One of the most pressing questions in the study of Alzheimer’s disease (AD) and related dementias (ADRD) is how alterations in the amino-acid sequence of tau, along with post-translational modifications (PTMs) such as phosphorylation and cleavage, lead the protein to misfold and disrupt normal neuronal function. While much has been learned over decades of rigorous and focused research, there are currently no disease modifying therapies to treat AD or related tauopathies. Recently, the field has begun a complicated but promising shift from targeting large tau fibrils (e.g. PHFs and NFTs) to disrupting smaller, non-fibrillar tau oligomers. While late-stage tau fibrils have been studied extensively—including a flurry of recent high-resolution cryo- EM structures—there are few tools to study early-stage oligomers, especially in cells. As a result, almost nothing is known about 1) early misfolding events that produce toxic, non-fibrillar tau oligomers; nor 2) how these oligomers co-opt protein machinery to cause cellular distress. To begin to fill this void, our 2019 Alzheimer’s & Dementia paper established a set of high-resolution, lifetime-FRET based biosensors that monitor full-length tau oligomers in cells. Here, we present compelling preliminary data showing that these biosensors can delineate which folding motifs in the fibril structures, as well as PTMs, most affect early-stage tau oligomers. These biosensors have also enabled us to study two distinct pathological tau interactions in cells. First, co- Investigators Karen Ashe and Kathryn Nelson’s 2016 Nature Medicine paper showed that cleavage of tau by caspase-2 (Casp2) causes tau to mislocalize to dendritic spines, shut down AMPA receptors and promote cognitive defects in mice. We show intriguing evidence to suggest a complex feedback loop between cleavage, oligomerization and toxicity. Second, tau and alpha-Synuclein (aSyn) have well-known co-morbidity in multiple Alzheimer’s Disease related dementias, but the biophysics of their interaction in early-stage misfolding is poorly understood. We provide preliminary evidence of a preferred binding orientation between tau and aSyn, suggesting a stable and hence targetable binding motif. The two major goals of this proposal are to: 1) determine which structural motifs revealed in the available tau fibril structures, and which PTMs, contribute most to early-stage oligomerization in cells, and to pathology; and 2) to characterize and inhibit two pathogenic tau interactions: tau/Casp2 and tau/aSyn. In Aim 1, we analyze the recently available fibril structures and ask: how can these structures be used to unravel otherwise elusive structural details of non-fibrillar tau oligomers? Additionally, to deepen the impact of our investigations, and with the help of co-Investigator Shauna Yuan, we will develop new lines of iPSC-derived human cortical dopaminergic neurons expressing our biosensors. Then, in Aims 2 and 3, we study the biophysical interplay between tau oligomerization and toxicity of tau/Casp2 and tau/aSyn respectively. In each case, we will also perform high- throughput small-molecule screens to identify potent inhibitors of these two pathological, oligomeric assemblies.

抽象的 阿尔茨海默氏病（AD）和相关痴呆症（ADRD）研究中最紧迫的问题之一 是如何在Tau的氨基酸序列以及翻译后修饰（PTM）的改变（例如 磷酸化和裂解，导致蛋白质折叠并破坏正常的神经元功能。虽然有很多 数十年来的严格和集中研究学会了，目前尚无疾病修改疗法 治疗广告或相关的tauopathies。最近，该领域已经开始一个复杂但有望从目标转变 大的tau原纤维（例如PHF和NFTS）破坏了较小的非纤维tau低聚物。 虽然后期tau纤维已经进行了广泛研究，包括一系列最近的高分辨率冷冻 EM结构 - 很少有研究早期低聚物，尤其是在细胞中的工具。结果，几乎没有 已知大约1）产生有毒的非纤维tau低聚物的早期错误折叠事件；也不2）如何 低聚物的同事蛋白质机制会导致细胞窘迫。为了开始填补这一空白，我们的2019年阿尔茨海默氏症＆ 痴呆症纸建立了一组高分辨率的基于寿命的生物传感器，可监视全长tau 细胞中的低聚物。在这里，我们提供了引人入胜的初步数据，表明这些生物传感器可以描述 在原纤维结构以及PTM中，哪些折叠基序都会影响早期Tau低聚物。 这些生物传感器还使我们能够研究细胞中的两种不同的病理TAU相互作用。首先，共同 调查人员凯伦·阿什（Karen Ashe）和凯瑟琳·尼尔森（Kathryn Nelson）的2016年自然医学论文表明，tau的裂解 caspase-2（CASP2）导致tau误倾至树突状刺，关闭AMPA接收器并促进 小鼠的认知缺陷。我们展示了有趣的证据，以暗示乳沟之间的复杂反馈回路， 寡聚和毒性。其次，tau和α-突触核蛋白（ASYN）在多个中具有众所周知的合并症 阿尔茨海默氏病与痴呆症相关，但在早期错误折叠中相互作用的生物物理学很差 理解。我们提供了tau和asyn之间首选结合方向的初步证据， 提出稳定的，因此是可靶向的结合基序。 该提案的两个主要目标是：1）确定可用tau中揭示的哪些结构图案 原纤维结构和PTMS在细胞和病理学中对早期寡聚的贡献最大。和 2）表征和抑制两种致病性tau相互作用：tau/casp2和tau/asyn。在AIM 1中，我们分析了 最近可用的原纤维结构，并询问：如何使用这些结构来解开其他难以捉摸的 非纤维tau低聚物的结构细节？此外，要加深我们调查的影响，并 在共同投资者Shauna Yuan的帮助下，我们将开发IPSC衍生的人皮质多巴胺能的新线条 表达我们的生物传感器的神经元。然后，在目标2和3中，我们研究了tau之间的生物物理相互作用 tau/casp2和tau/asyn的寡聚和毒性。在每种情况下，我们还将执行高 吞吐量的小分子筛选，以鉴定这两种病理性的寡聚组件的潜在抑制剂。

项目成果

Proteasomal Stimulation by MK886 and Its Derivatives Can Rescue Tau-Induced Neurite Pathology.

• DOI: 10.1007/s12035-023-03417-5
• 发表时间: 2023-10
• 期刊: MOLECULAR NEUROBIOLOGY
• 影响因子: 5.1
• 作者: Liao, Elly E.;Yang, Mu;Kochen, Noah Nathan;Vunnam, Nagamani;Braun, Anthony R.;Ferguson, David M.;Sachs, Jonathan N.
• 通讯作者: Sachs, Jonathan N.