Mechanism of VCP interaction and processing of neurodegenerative tau fibrils

项目摘要 这项建议的目的是研究AAA ATPase相互作用的结构和动力学， 含Valosin蛋白(VCP)和疾病相关tau纤维用于确定VCP介导的疾病 进展和治疗潜力。阿尔茨海默病(AD)和皮质基底膜变性(CBD)是 以不同形式的纤维状tau聚集体为特征的20多种“tauopathy”。牛黄的程度 尸检病理与认知能力下降有关，但神经毒性、疾病的机制 它的发生、发展和疾病进展仍有待阐明。聚集性疾病 表明蛋白质平衡功能障碍--蛋白质组的调节和平衡。VCP扮演着至关重要的角色 通过从复合体、膜和聚集体(如形成的那些)中去除蛋白质而在蛋白质稳定中发挥作用 从而指出VCP在蛋白抑制功能障碍中发挥着重要作用，并且是一种潜在的药物。 紧张症的靶子。 VCP有多种辅助因子，使其能够执行一系列重要的细胞。Ufd1和Npl4 辅因子(UN)共同作用使VCP能够识别多泛素化底物。Ad tau原纤维有 泛素化的比率很高，可能是因为细胞已经标记了这些纤维，以便被VCP等蛋白质去除。 此外，VCP内的突变与神经退行性疾病有关。VCP基因低活性突变先例 被爱德华·李实验室描述为痴呆症，并已被证明促进tau纤维 病理学。低活性的VCP突变被认为降低了VCP功能，导致tau的积累 纤维，导致神经退行性疾病。然而，VCP的类似机制不能充分清除tau 在散发性肌萎缩侧索硬化症的病例中可能存在纤维。VCP/UN，突变型或野生型，也可能被粘住 Tau原纤维(可能是由于原纤维的高稳定性)，不能分解或释放原纤维，减少 通过将VCP捕获在原纤维上，有效地发挥VCP功能。这在26S中已经看到了类似的AAA ATPase 蛋白酶体和神经退行性集合体。 我认为，对于生产性解聚酶活性，VCP/UN在 然后，tau纤维一次只能拉出一种tau单体。我还建议，行动机制不会改变，但 由于原纤维结构的不同，tau纤维之间的化学计量比和反应速度会发生变化。 泛素化。为了测试这个模型，在目标1中，我将从结构上描述VCP/UN如何与大脑相互作用- 衍生的AD和CBD tau纤维。从结构上理解VCP/UN如何与tau纤维相互作用将有所帮助 阐明对VCP/UN功能至关重要的蛋白质区域，并了解这种相互作用如何变化 纤维类型将有助于理解VCP/UN纤维相互作用的概括性。在《目标2》中，我会 描述VCP/UN-tau相互作用的动力学。了解VCP/UN互动的动态 这些纤维将进一步阐明VCP/UN与疾病相关的tau纤维的机制。

Project Summary The goal of this proposal is to study the structure and dynamics of the interaction of the AAA+ ATPase, valosin-containing protein (VCP) with disease-relevant tau fibrils to determine VCP-mediated disease progression and therapeutic potential. Alzheimer's Disease (AD) and Corticobasal degeneration (CBD) are 2 of more than 20 “tauopathies” characterized by different forms of fibrillar tau aggregates. The degree of tau pathology on autopsy correlates with cognitive decline, yet the mechanism of neurotoxicity, disease development, and disease progression in tauopathies remains to be elucidated. Diseases of aggregation indicate a dysfunction in proteostasis – the regulation and balance of the proteome. VCP plays an essential role in proteostasis by removing proteins from complexes, membranes, and aggregates such as those formed by fibrillar tau, thus pointing to VCP as an important player in proteostatic dysfunction and as a potential drug target for tauopathies. VCP has a variety of cofactors that enable it to perform an array of vital cellular. Ufd1 and Npl4 cofactors (UN) together specifically enable VCP to recognize poly-ubiquitinated substrates. AD tau fibrils have high rates of ubiquitination, likely because cells have marked these fibrils for removal by proteins such as VCP. Additionally, mutations within VCP are linked to neurodegenerative disease. A hypoactive VCP mutation first described by the Edward Lee lab clinically presents as dementia and has been shown to promote tau fibril pathology. The hypoactive VCP mutation is thought to reduce VCP function causing the accumulation of tau fibrils, leading to neurodegenerative disease. However, a similar mechanism of VCP inadequately clearing tau fibrils may be present in cases of sporadic tauopathies. VCP/UN, mutant or wild-type, may also get stuck on tau fibrils (possibly due to fibrils' high stability) and be unable to break down the fibril or be released, reducing effective VCP function by trapping it on the fibril. This has been seen with a similar AAA+ ATPase in the 26S proteasome and neurodegenerative aggregates. I propose that for productive disaggregase activity VCP/UN binds unstructured ubiquitin at either end of tau fibrils then pulls off one tau monomer at a time. I also propose the mechanism of action will not change but the stoichiometry and rate of reaction will change between tau fibrils due to differences in fibril structure and ubiquitination. To test this model, in Aim 1 I will structurally characterize how VCP/UN interacts with brain- derived AD and CBD tau fibrils. Structurally understanding how VCP/UN interacts with tau fibrils will help elucidate the protein regions that are crucial to VCP/UN function, and knowing how this interaction changes with fibril type will help understand the generalizability of the VCP/UN fibril interaction. In Aim 2, I will characterize the dynamics of the VCP/UN-tau interaction. Understanding the dynamics of VCP/UN interaction with these fibrils will further elucidate the mechanism of VCP/UN in relation to disease-relevant tau fibrils.