Determining the mechanism of alpha-synuclein dependent innate immune responses in the brain

项目摘要 神经系统传统上被认为独立于有机体的免疫反应而活动，并被 一个“免疫特权站点”。越来越多的科学证据表明，神经系统也不能幸免 拥有特权，但却拥有独特的免疫反应，这对维持动态平衡至关重要，对 中枢神经系统(CNS)功能。神经炎症领域的许多工作都集中在 小胶质细胞的功能，但对神经元在调节神经炎性反应中的作用知之甚少 在中枢神经系统。五年前，我们发现神经元蛋白--α-突触核蛋白(ASYN)在 保护神经元免受病毒感染。我们已经扩展了这些数据，以表明Ayn调制类型1 干扰素(T1IFN)信号传导。Asyn被认为是帕金森氏病(PD)的原因之一，并被认为是 在神经退行性疾病、创伤性脑损伤和其他不同的中枢神经系统疾病中调节失调。尽管 ASYN在中枢神经系统疾病中的重要性，ASYN表达的功能作用还不是很清楚。 我们发现ASYN的表达是支持刺激的特异性干扰素表达所必需的 在T1干扰素信号传递过程中大脑中的基因(ISGs)，不依赖于小胶质细胞的激活。使用诱导式 多能干细胞(IPSC)和CRISPR介导的SNCA缺失建立异体KO人多巴胺能 对于神经元，我们发现在ASYN表达存在的情况下，神经元中的病毒生长受到抑制，并且病毒- 诱导的ISGs如Ifit1、OAS1和TRIM25 Exbibit可降低ASYN KO神经元的表达。我们是下一个 发现在用PolyI：C或1型处理后，asyn KO神经元表现出广泛的ISG表达缺失 干扰素(2)治疗由于失去了异步化依赖的STAT2激活和异步化核定位。已被占用 总而言之，我们的数据首次表明，ASYN功能支持神经元中的干扰素反应。这个 这项建议的目的是确定非同步依赖的先天免疫反应的特定机制 神经元。我们假设ASYN是中枢神经系统固有免疫反应的一种新的神经元内源性调节因子。 我们将从三个方面检验我们的假设。目标1将使用ASYN KO和WT人类神经元来定义特定的 神经元中ASYN、干扰素信号和囊泡运输之间的相互作用。目标2将定义 使用可诱导的Nestin-Cre-lox对大脑中固有T细胞反应的神经元内源性ASYN的产生 在小鼠中敲除asyn基因(SNCA)。AIM 3将评估PD特定的和物种特定的变化 这可能会影响其在神经元中的天然功能。综上所述，拟议的研究将显著 促进我们对神经元对中枢神经系统固有免疫反应的内在控制的理解，并提供 对导致不同人类中枢神经系统疾病的潜在免疫病理机制的新见解。

Project Summary The nervous system was traditionally thought to act independently of an organism’s immune response and be an “immune privileged site”. Increasing scientific evidence has shown that the nervous system is not immune privileged but instead has a unique immune response that is critical for maintaining homeostasis and critical for central nervous system (CNS) function. Much of the work in the field of neuroinflammation has focused on the function of microglia, but little is known about the role of neurons in modulating neuroinflammatory responses in the CNS. Five years ago, we discovered that the neuronal protein, alpha-synuclein(asyn), was critical in protecting neurons from viral infection. We have extended these data to show that ayn modulates type 1 interferon (T1IFN) signaling. Asyn is known as a cause of Parkinson’s disease (PD) and is known to be dysregulated in neurodegenerative diseases, traumatic brain injury, and other diverse CNS diseases. Despite the importance of asyn in CNS disease states, the functional role of asyn expression is not well understood. We have discovered that asyn expression is necessary to support expression of specific interferon stimulated genes (ISGs) in the brain during T1IFN signaling, independent of microglia activation. Using induced pluripotent stem cells (iPSC) and CRISPR-mediated SNCA deletion to create asyn KO human dopaminergic neurons, we found that viral growth in neurons is inhibited in the presence of asyn expression and that viral- induced ISGs such as IFIT1, OAS1, and TRIM25 exbibit decreased expression in asyn KO neurons. We next found that asyn KO neurons exhibit a broad loss of ISG expression following treatment with poly I:C or type 1 interferon (2) treatment due to loss of asyn-dependent STAT2 activation and asyn nuclear localization. Taken together, our data show for the first time that asyn functions to support interferon responses in neurons. The goal of this proposal is to determine the specific mechanism of asyn-dependent innate immune responses in neurons. We hypothesize that asyn is a novel neuron-intrinsic regulator of the CNS innate immune response. We will test our hypothesis in three aims. Aim 1 will use asyn KO and WT human neurons to define the specific interactions between asyn, interferon signaling, and vesicle transport in neurons. Aim 2 will define the role of neuron-intrinsic asyn production on the innate T-cell response in the brain using an inducible, nestin-Cre-lox knockout of the asyn gene (Snca) in mice. Aim 3 will evaluate PD-specific and species specific changes in asyn that may influence its native function in neurons. Taken together, the proposed studies will significantly advance our understanding of neuron-intrinsic control of the innate immune response in the CNS and provide novel insight into the underlying immunopathogenesis that contributes to diverse human diseases of the CNS.