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。多系统萎缩(MSA)和其他突触核素病 在患者身上，蛋白质α-突触核蛋白(α-SYN)错误地折叠成一种自我模板构象，这种构象通过普恩病毒传播- 类似的方式遍及全身，包括中枢神经系统(CNS)。它被假设为 构象或菌株，α-SYN错误折叠成关于临床症状和 患者会发展成神经病理。虽然之前的研究主要集中在生化差异上 在α-SYN菌株之间，这些差异如何编码不同的生物学表型的机制 人们对疾病知之甚少。我们研究的长期目标是确定代理人和宿主因素 有助于观察到不同的联体核病的临床表现。在这份提案中，我们将测试 假设菌株在聚集运输和神经解剖学传播方面的特定差异有助于 疾病发病机制。在目标1中，我们将使用Alexa荧光标记的α-SYN聚集体来研究 体外和体内轴突运输的方向。确定α-SYN发病的分子机制 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Dynein货物接头结合，并量化菌株特异性对α-SYN轴突运输的影响。在目标2中， 我们将确定跨突触扩散在α-SYN株发病中的作用。严格执行这些任务 首先，我们将在体外和体内测定三种不同的α-SYN株的滴度。然后我们将使用 坐骨神经注射模型，有无神经横断，以确定α-SYN神经侵袭是否依赖于 仅限于跨突触扩散，当神经外通路在疾病发病机制中起作用时 注射每种菌株的相同效价。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传播的时间-空间图。这项工作具有创新性，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的进展，并建立 体外和体内α-SYN滴度。这项工作意义重大，因为它是第一次研究相互作用是如何 宿主和菌株之间的关系有助于轴突运输和跨突触扩散的机制 疾病的威胁。关键的是，通过确定负责α-SYN传播的细胞和分子机制， 这些实验的结果将带来新的有希望的研究领域。

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.