Molecular and Cellular Mechanisms of Herpes Simplex Transport and Egress Pathway in Neurons

项目摘要/摘要 阿尔法疱疹病毒，包括人类重要的病原体单纯疱疹病毒1型(HSV-1)，是 在极少数进化了神经元特异性病毒机制的病毒中， 神经细胞生物学。在疾病的自然病程中，阿尔法疱疹病毒感染感觉或自主神经。 神经元，并在周围神经系统建立终身潜伏感染。在重新激活后，病毒 可以返回到周围组织，引起疱疹或带状疱疹病变，也可以扩散到中枢神经 导致严重的疱疹脑炎。这项建议集中在HSV-1的三个主要步骤 复制周期，其中病毒颗粒必须与专门的神经系统相互作用：1.远距离后- 进入轴突运输；2.子代颗粒复制后向轴突位置的长距离运输 通过轴突、树突或细胞体的胞吐作用而极化的病毒出口。在目标1中，我们将确定 HSV-1颗粒胞吐在神经元中的位置及HSV-1的分子/细胞机制 轴突、树突或细胞体的出口。在目标2中，我们将研究HSV-1的结构生物学 冷冻电子显微镜观察轴突运输。我以前的教育和研究经验是 专注于从分子和细胞生物学的角度研究病毒，特别是研究 包膜病毒与宿主膜系统的相互作用。在我的研究生和博士后课程中 在研究中，我在使用专门的显微镜方法对时空进行解剖方面积累了专业知识 病毒复制周期中的特定步骤。在我职业生涯的博士后阶段，我发展了 一种新的基于特殊荧光的活细胞荧光显微镜检测病毒出口的方法 显微镜方法和一种新型的pH敏感的荧光探针。此外，我目前正在开发新的 研究病毒颗粒轴突运输的低温电子显微镜断层扫描方法。这项提案旨在延长 这些方法可以实现所提出的目标。我的科学背景使我非常适合进行 建议的研究。这个K22职业转折奖将帮助我实现我的科学和职业生涯 目标，包括过渡到一个独立的职位，作为助理教授，并建立一个 由美国国立卫生研究院资助的独立研究项目专注于神经病毒学。

Project Summary/Abstract The alpha herpesviruses, which includes important human pathogen herpes simplex virus 1 (HSV-1), are among the very few viruses that have evolved neuron-specific viral mechanisms to exploit highly-specialized neuronal cell biology. During the natural course of disease, alpha herpesviruses infect sensory or autonomic neurons, and establish a life-long latent infection in the peripheral nervous system. Upon reactivation, the virus can return to peripheral tissues, causing herpetic or zosteriform lesions, or it can spread to the central nervous system, causing severe herpes encephalitis. This proposal focuses on three major steps in the HSV-1 replication cycle, where virus particles must interact with specialized neuronal systems: 1. long-distance post- entry axonal transport; 2. long-distance post-replication transport of progeny particles towards axonal sites of egress; 3. polarized viral egress via exocytosis from axons, dendrites, or cell bodies. In Aim 1, we will identify sites of HSV-1 particle exocytosis in neurons, and determine the molecular/cellular mechanisms of HSV-1 egress from axons, dendrites, or cell bodies. In Aim 2, we will investigate the structural biology of HSV-1 axonal transport by cryo electron microscopy (cryoEM). My prior education and research experience has focused on studying viruses from a molecular and cell biological perspective, in particular, studying the interactions of enveloped viruses with host membrane systems. In the course of my graduate and postdoctoral research, I have developed expertise in using specialized microscopy methods to spatiotemporally dissect particular steps in the virus replication cycle. In the current postdoctoral phase of my career, I have developed a novel live-cell fluorescence microscopy assay of virus egress based on a specialized fluorescence microscopy method and a novel pH-sensitive fluorescent probe. In addition, I am currently developing new cryoEM tomography methods to investigate axonal transport of virus particles. This proposal seeks to extend these methods to accomplish the proposed aims. My scientific background makes me well suited to carry out the proposed research. This K22 Career Transition Award will help me to achieve my scientific and career goals, which include transitioning to an independent position as an assistant professor, and establishing an independent NIH-funded research program focused on neurovirology.