Modulation of Herpes Simplex Virus Pathogenesis by Leucine Rich Repeat Kinase 2

项目摘要 在美国，1型单纯疱疹病毒（HSV-1）感染了一半以上的成年人口，是世界上最严重的疱疹病毒感染者。 病毒性脑炎和传染性失明的主要原因。然而，造成最严重的 HSV-1的严重表现仍不清楚。炎性小体，最近发现的抗病毒先天 免疫防御和两种炎性细胞因子（白细胞介素（IL）-1 β和IL-18）的产生是至关重要的 HSV-1控制和发病机制。然而，在HSV-1感染期间炎性小体是如何调节的， 不太了解。最近，富含亮氨酸的重复序列激酶2（LRRK 2）已经成为一种可能的调节剂， 炎性小体信号传导。LRRK 2已被广泛研究，因为特定的等位基因与 帕金森病、克罗恩病、癌症和麻风病。尽管有疾病的关联， LRRK 2的功能及其如何介导疾病尚不清楚。在免疫反应中发挥作用的证据 包括LRRK 2在免疫细胞上高度表达，在感染应答中上调，并被 最近显示出响应细菌感染调节NLRC 4炎性体。我们最近 NLRP 3在HSV-1的炎性小体激活中起关键作用，我们的初步数据表明， LRRK 2增强NLRP 3信号传导。我们发现，常见的LRRK 2突变G2019 S增强HSV-1， 1病理学而不改变病毒复制。这些结果指出LRRK 2在以下方面的新作用： 炎症体信号传导和HSV-1发病机制。我们假设LRRK 2调节炎性小体， G2019 S突变进一步增强了炎症小体， 体内激活，导致HSV-1后更严重的眼睛、皮肤和中枢神经系统（CNS）病理 感染LRRK 2尚未在人类病毒感染中进行过研究，这些研究代表了第一个定义LRRK 2的研究。 其在常见嗜神经性人类病毒感染的发病机制中的作用。在目标1中，我们将确定如何 LRRK 2与NLRP 3相关，NLRP 3是HSV-1感染中的关键炎性体适配器，以及LRRK 2突变如何影响HSV-1感染。 LRRK 2在HSV-1感染的背景下改变炎性体信号传导。我们将通过使用 在炎性体信号转导的转染模型中的靶向诱变和通过研究炎性体 HSV-1感染后稳定表达LRRK 2突变体的细胞系的活化。对于目标2，我们将确定 LRRK 2如何改变使用LRRK 2转基因小鼠和具有LRRK 2的小鼠对HSV-1的炎性体应答 炎性体基因破坏。此外，PI将获得研究所需的关键技能和专业知识， 对HSV-1和其他疱疹病毒的先天免疫反应。这些学习目标将会实现 通过正式的课程，科学的编程，以及病毒和鼠类专家的直接指导， 免疫学、疱疹病毒学和生物统计学。拟议的职业发展计划和研究目标将 提供了一个途径，作为一个独立的研究人员研究疱疹病毒之间的相互作用的职业生涯 和先天免疫系统。

PROJECT SUMMARY In the United States, herpes simplex virus type 1 (HSV-1) infects more than half the adult population and is the leading cause of viral encephalitis and infectious blindness. Yet, the host factors contributing to the most severe manifestations of HSV-1 remain unclear. Inflammasomes, a recently discovered antiviral innate immune defense, and the production of two inflammasome cytokines (interleukin (IL)-1and IL-18) are critical in HSV-1 control and pathogenesis. However, how inflammasomes are regulated during HSV-1 infection is poorly understood. Recently, leucine-rich repeat kinase 2 (LRRK2) has emerged as a possible regulator of inflammasome signaling. LRRK2 has been studied extensively because specific alleles are associated with Parkinson’s disease, Crohn’s disease, cancer, and leprosy. Despite the disease associations, the precise function of LRRK2 and how it mediates disease are unknown. Evidence for a role in the immune response include that LRRK2 is highly expressed on immune cells, is upregulated in response to infection, and was recently shown to modulate the NLRC4 inflammasome in response to bacterial infection. We recently demonstrated that NLRP3 is critical in inflammasome activation by HSV-1, and our preliminary data indicate that LRRK2 enhances NLRP3 signaling. We found that the common LRRK2 mutation G2019S enhances HSV- 1 pathology in mice without altering viral replication. These results point to a novel role for LRRK2 in inflammasome signaling and HSV-1 pathogenesis. We hypothesize that LRRK2 modulates inflammasome signaling in response to HSV-1 infection and that the G2019S mutation further enhances inflammasome activation in vivo, leading to more severe eye, skin, and central nervous system (CNS) pathology after HSV-1 infection. LRRK2 has not been studied in human viral infection, and these represent the first studies to define its role in the pathogenesis of a common neurotropic human viral infection. In Aim 1, we will identify how LRRK2 associates with NLRP3, the critical inflammasome adapter in HSV-1 infection, and how mutations in LRRK2 alter inflammasome signaling in the context of HSV-1 infection. We will accomplish this by using targeted mutagenesis in a transfection model of inflammasome signaling and by studying inflammasome activation after HSV-1 infection of cell lines stably expressing LRRK2 mutants. For Aim 2, we will determine how LRRK2 alters the inflammasome response to HSV-1 using LRRK2 transgenic mice and mice with inflammasome gene disruptions. Additionally, the PI will gain critical skills and expertise necessary to study the innate immune response to HSV-1 and other herpesviruses. These learning objectives will be accomplished through formal coursework, scientific programing, and direct mentorship by experts in viral and murine immunology, herpesvirology, and biostatistics. The proposed career development plan and research aims will provide a pathway to a career as an independent investigator studying the interactions between herpesviruses and the innate immune system.