Effect of agent and host factors on alpha-synuclein strain pathogenesis

蛋白质错误折叠疾病，或蛋白质病，是一组总是致命的神经退行性疾病 影响了超过680万美国人。在多系统萎缩（MSA）和其他突触核蛋白病 在患者中，蛋白质α-突触核蛋白（α-syn）错误折叠成自我模板构象，通过朊病毒传播， 类似的方式在整个身体，包括中枢神经系统（CNS）。据推测， 构象，或应变，α-syn错误折叠成编码有关临床症状的信息， 病人会出现的神经病变虽然以前的研究集中在生物化学差异上， 在α-syn菌株之间，这些差异如何编码不同的生物表型的机制， 疾病知之甚少。我们研究的长期目标是确定 有助于在突触核蛋白病中观察到的各种临床表现。在本提案中，我们将测试 聚集体转运和神经解剖学扩散的菌株特异性差异有助于 发病机理在目标1中，我们将使用alexa荧光标记的α-syn聚集体来研究速率和 体外和体内轴突运输的方向。确定α-syn的分子机制 运输，我们将使用化学和遗传工具来破坏微管聚合，动力蛋白运动活性， 和动力蛋白货物衔接子结合，并量化对α-syn轴突运输的应变特异性影响。在目标2中， 我们将确定跨突触扩散在α-syn菌株发病机制中的作用。严格执行这些 研究中，我们将首先在体外和体内测定三种不同α-syn菌株的滴度。然后我们将使用 坐骨神经注射模型，有和没有神经横断，以确定是否α-syn神经侵袭依赖于 如果神经通路有助于疾病的发病机制， 注射相同滴度的每种菌株。最后，我们将进行彻底的疾病发病机制研究，以建立 菌株特异性α-syn扩散的时空图。这项工作是创新的，因为它是第一个研究， 研究宿主和菌株之间的相互作用如何影响疾病进展， 体外和体内α-syn滴度。这项工作意义重大，因为它是第一个调查如何相互作用 宿主和应变之间的相互作用有助于轴突运输和跨突触扩散的机制 疾病。重要的是，通过识别负责α-syn传播的细胞和分子机制， 这些实验的结果将导致有希望的研究的新领域。

Protein misfolding diseases, or proteinopathies, are a group of invariably fatal neurodegenerative disorders affecting more than 6.8 million Americans. In multiple system atrophy (MSA) and other synucleinopathy patients, the protein α-synuclein (α-syn) misfolds into a self-templating conformation that spreads via a prion- like manner throughout the body, including the central nervous system (CNS). It is hypothesized that the conformation, or strain, that α-syn misfolds into encodes information about the clinical symptoms and neuropathologies a patient will develop. While previous studies focused on the biochemical differences between α-syn strains, the mechanism of how those differences encode distinct biological phenotypes of disease is poorly understood. The long-term goal of our research is to identify the agent and host factors that contribute to the varied clinical presentations observed across synucleinopathies. In this proposal, we will test the hypothesis that strain-specific differences in aggregate transport and neuroanatomical spread contribute to disease pathogenesis. In Aim 1, we will use alexa fluor-labeled α-syn aggregates to investigate the rate and direction of axonal transport in vitro and in vivo. To determine the molecular mechanisms responsible for α-syn transport, we will use chemical and genetic tools to disrupt microtubule polymerization, dynein motor activity, and dynein cargo adaptor binding, and quantify the strain-specific effects on α-syn axonal transport. In Aim 2, we will determine the role of trans-synaptic spread on α-syn strain pathogenesis. To rigorously perform these studies, we will first determine the titer of three different α-syn strains both in vitro and in vivo. We will then use the sciatic nerve injection model, with and without nerve transection, to determine if α-syn neuroinvasion relies exclusively on trans-synaptic spread, of if extraneural pathways contribute to disease pathogenesis when the same titer of each strain is injected. Finally, we will perform a thorough disease pathogenesis study to establish a temporal-spatial map of strain-specific α-syn spread. This work is innovative because it is the first study to investigate how interactions between the host and strain impact disease progression, and to establish between in vitro and in vivo α-syn titers. This work is significant because it is the first to investigate how interactions between host and strain contribute to the mechanisms underlying axonal transport and trans-synaptic spread of disease. Critically, by identifying the cellular and molecular machinery responsible for α-syn propagation, the results of these experiments will lead to new areas of promising investigation.