Interactions of herpes simplex virus with nectin-1 at cell junctions

描述（由申请人提供）：单纯疱疹病毒（HSV）是一种非常常见的人类病原体，在传播至感觉神经元之前感染上皮细胞和皮肤，在那里建立终身感染。HSV的进入是一个复杂的分子过程。一个必要的步骤是HSV糖蛋白D（gD）与细胞受体的相互作用，以诱导病毒包膜与质膜或内体膜的融合。神经元和上皮细胞上的主要gD受体之一是细胞粘附分子nectin-1。这项研究的长期目标是解释病毒如何利用细胞途径到达细胞连接处的nectin-1，刺激内吞作用并与宿主膜融合。为实现这些目标，提出了两个具体目标。在目标1中，将定义在HSV进入期间当gD结合至连接蛋白-1时被激活的细胞途径。将表达Nectin-1的细胞暴露于膜结合或可溶形式的gD，以鉴定参与受体内化的细胞应答。将gD与nectin-1结合引起的信号与天然配体nectin-3的信号进行比较，nectin-3在细胞连接处和突触处与nectin-1相互作用。最后，参与受体内吞作用的途径将与HSV进入期间激活的途径相关。在目标2中，将定义HSV进入对细胞连接完整性的影响。由于gD结合影响nectin-1介导的细胞粘附，因此将研究gD对细胞连接结构的影响。将定义HSV颗粒在质膜上冲浪并在细胞连接处接近nectin-1的病毒和细胞需求。将分析这种运动进入极化上皮细胞的重要性。该项目将扩展HSV附着和结合受体的研究，以研究病毒诱导的即时细胞反应的新方面。将使用最先进的技术和新的试剂组来探索HSV与细胞表面结合所触发的细胞机制。在这个项目中开发的创新方法将允许HSV进入细胞连接处的研究直接观察病毒和细胞成分在真实的时间使用活细胞共聚焦显微镜。这项研究将增加我们对HSV感染早期步骤的了解，并确定抗病毒治疗作用的新靶点。单纯疱疹病毒1型和2型（HSV-1和HSV-2）分别感染80%和20%的美国成年人，目前的治疗不能减少HSV再激活和传播的发生，因此需要开发新的治疗策略。这项研究将确定病毒与细胞结合的直接后果，并确定涉及病毒进入的细胞机制。通过确定病毒进入宿主的新要求，这项研究将为干扰这一过程的新治疗方法提供靶点。

DESCRIPTION (provided by applicant): Herpes simplex virus (HSV) is a very common human pathogen that infects epithelia and skin before spreading to sensory neurons where it establishes a lifelong infection. Entry of HSV is a complex molecular process. One essential step is the interaction of HSV glycoprotein D (gD) with a cellular receptor to induce fusion of the viral envelope with the plasma membrane or an endosomal membrane. One of the main gD-receptor on neurons and epithelial cells is the cell adhesion molecule nectin-1. The long-term objectives of this study are to explain how the virus exploits cellular pathways to reach nectin-1 at cell junctions, stimulate endocytosis and fuse with the host membrane. Two specific aims are proposed to achieve these objectives. In aim 1 the cellular pathways that are activated when gD binds to nectin-1 during HSV entry will be defined. Nectin-1 expressing cells will be exposed to either membrane bound or soluble forms of gD to identify cellular responses involved in receptor internalization. The signals elicited by gD binding to nectin-1 will be compared to those of a natural ligand, nectin-3, which interacts with nectin-1 at cell junctions and at synapses. Finally, pathways involved in receptor endocytosis will be correlated with pathways activated during HSV entry. In aim 2 the effect of HSV entry on the integrity of cellular junctions will be defined. Since gD binding affects nectin-1-mediated cell adhesion, the effects of gD on the architecture of cell junctions will be studied. The viral and cellular needs for HSV particles to surf on the plasma membrane and access nectin-1 at cellular junctions will be defined. The importance of this movement for entry into polarized epithelial cells will be analyzed. This project will extend studies of HSV attachment and binding to receptors toward novel aspects of the immediate cellular responses induced by the virus. State of the art technology and new sets of reagents will be used to explore the cellular mechanisms triggered by HSV binding to the cell surface. Innovative approaches developed in this project will allow studies of HSV entry at cellular junctions by direct observation of viruses and cell components in real time using live cell confocal microscopy. This study will increase our understanding of the early steps of HSV infection and identify new targets for antiviral therapeutic actions. Herpes simplex viruses 1 and 2 (HSV-1 and HSV-2) infect 80% and 20% of all American adults respectively and current treatments do not reduce the occurrence of HSV reactivation and transmission, thus new therapeutic strategies need to be developed. This study will determine the immediate consequences of the virus binding to the cell and identify cellular mechanisms involved in virus entry. By identifying new requirements for virus entry in the host, this study will provide targets for novel therapeutic approaches to interfere with this process.