Immunoregulatory Activities of HSV gD Binding to its Entry Receptors

描述(申请人提供)：单纯疱疹病毒(HSV)是一种常见的人类病原体，可导致终生感染。单纯疱疹病毒感染并在上皮细胞中复制，然后扩散到感觉神经元，在那里它建立了终生潜伏期，其间不时出现重新激活事件。先天免疫系统和获得性免疫系统的细胞毒细胞在限制病毒在上皮细胞中的原发和复发传播和复制方面发挥着关键作用。此外，细胞毒性 CD8+T细胞控制着潜伏感染神经元中病毒重新激活的程度。我的目标是了解HSV如何通过改变其两个主要进入受体：HVEM和Nectin-1的关键免疫调节功能来影响细胞毒细胞的功能。值得注意的是，这两种受体也参与了复杂的相互作用网络，平衡了细胞溶解细胞的激活和抑制。在抗原识别过程中，HVEM处于T细胞的TNFR和CD28共调节网络的十字路口。Nectin-1是NK细胞激活受体CD96的配体，但其在先天免疫中的作用尚不清楚。我证明了病毒配体HSV gD的结合可以通过两种方式影响HVEM和Nectin-1的功能：通过与天然配体竞争和通过诱导受体内吞。因此，我的一般假设是，HSV使用gD作为非正则配体来改变宿主的防御。在这项资助中，我将比较和对比GD如何影响CTL和NK细胞对感染细胞的功能。拟议的目标将侧重于分子相互作用(目标1)、目标识别(目标2)和功能激活(目标3)。目标1是基于我的初步数据，比较了Hvem和Nectin-1与Gd或它们各自的天然配体结合的结构。GD结合部位重叠，但与受体的功能部位不同，提示GD可与天然配体竞争并诱导特异性反应。在目标2中，我将研究GD如何通过分别影响NK和CTL免疫突触上的Nectin-1和Hvem来扰乱细胞溶解细胞与其靶细胞之间的联系。为了解决AIM 3中的NK和T细胞功能问题，我与宾夕法尼亚大学的J.Orange和J.Riley博士以及洛桑大学的D.Nardelli Haefliger博士建立了合作关系。我将研究在存在或不存在抗原提呈的情况下，GD如何影响NK细胞的激活，并调节CD8 T细胞的细胞毒作用。这项研究的成功完成将确定作为关键进入糖蛋白的gD如何通过靶向先天和获得性免疫反应的关键调节因子而在HSV的两种新的免疫逃避策略中发挥作用。我提出了可检验的假说，以了解gD在HSV感染过程中的免疫学作用，并将其作为一种工具来解开复杂的免疫调节网络，涉及到NK和T细胞的一般细胞溶解活性。这种“分子免疫学”方法将为在活体内解决这些功能提供可能。此外，这项研究的数据可能会为设计新的抗病毒干预措施提供建议。

DESCRIPTION (provided by applicant): Herpes simplex virus (HSV) is a common human pathogen causing lifelong infection. HSV infects and replicates in epithelial cells before spreading to sensory neurons where it establishes a lifelong latency punctuated by frequent reactivation events. Cytotoxic cells of the innate and adaptive immune systems play critical roles in limiting primary and recurrent viral spread and replication in epithelia. Furthermore, cytotoxic CD8+ T cells control the extent of viral reactivation from latently infected neurons. My goal is t learn how HSV affects the function of cytotoxic cells by altering key immunoregulatory functions of its two main entry receptors: HVEM and nectin-1. It is remarkable that both receptors are also involved in complex networks of interactions that balance the activation and inhibition of cytolyti cells. HVEM is at the crossroad between TNFR and CD28 co-modulatory networks of T cells during antigen recognition. Nectin-1 is a ligand for the NK cell activating receptor CD96 but its role in innate immunity remains relatively unclear. I showed that binding of the viral ligand, HSV gD, can affect the function of HVEM and nectin-1 in two ways: by competing with natural ligands and by inducing receptor endocytosis. Thus, my general hypothesis is that HSV uses gD as a non-canonical ligand to alter the host's defenses. In this grant I will compare and contrast how gD affects the function of CTL and NK cells towards infected cells. The proposed aims will focus on molecular interactions (aim 1), target recognition (aim 2), and functional activation (aim 3). Aim 1 is based on my preliminary data that compare the structures of HVEM and nectin-1 bound to gD or to their respective natural ligands. The gD binding sites overlap but differ from the functional sites of the receptors suggesting that gD can compete with natural ligands and induce a specific response. In Aim 2, I will study how gD perturbs the contacts between cytolytic cells and their targets by affecting nectin-1 and HVEM at the NK and CTL immunological synapses respectively. To address NK and T cell function in Aim 3, I established collaborations with Drs J. Orange and J. Riley (both at PENN) and Dr D. Nardelli Haefliger (U. of Lausanne). I will investigate how gD affects the activation of NK cells and modulates CD8 T cell cytotoxicity in the presence or absence of antigen presentation. Successful completion of this study will define how gD, a key entry glycoprotein functions in two novel immune evasion strategies for HSV by targeting key regulators of the innate and adaptive immune responses. I have proposed testable hypotheses to understand the immunological role of gD during HSV infection and to use it as a tool to unravel the complex immunoregulatory networks involved in cytolytic activity of NK and T cells in general. This "molecular immunology" approach will open possibilities to address these functions in vivo. Furthermore, data from this study may suggest ways to design novel antiviral interventions.