Mechanisms of PSGL-1 restriction of HIV virion infectivity

项目摘要/摘要 PSGL-1抑制HIV病毒粒子感染性的机制 限制因子是宿主先天免疫的重要组成部分。抗HIV作用机制的研究 限制因素是理解病毒-宿主相互作用的核心。这些机制还可能提供新的 灭活病毒库的治疗策略，以实现艾滋病毒的持久缓解。最近，我们有 发现了一种新的HIV限制因子PSGL-1(P-选择素糖蛋白配体-1)，它可以灭活 从HIV产生细胞释放的HIV病毒粒子的传染性。PSGL-1是一种粘蛋白样二聚体，分子量为120-kD 主要在淋巴和髓系细胞表面表达的糖蛋白。PSGL-1与P-，L-， 和E-选择素，并介导白细胞在内皮上的捆绑和滚动，使白细胞迁移到 发炎的组织。PSGL-1也是一个受干扰素-γ调节的因子，参与Th1介导的抗病毒活性。我们的 初步的机制研究进一步表明，PSGL-1被整合到病毒颗粒中，从而阻止 HIV Env掺入，并使病毒粒子附着到目标CD4T细胞以进行感染的能力失效。在……里面 此外，我们还发现VPU和Nef通过表面下调来拮抗PSGL-1。基于 在这些初步研究中，我们假设：(1)PSGL-1介导的HIV感染性限制涉及其 (2)PSGL-1可能通过竞争性排斥环境来限制HIV的感染性 病毒组装过程中的掺入和Env与细胞受体结合的空间位阻(3)Nef介导 PSGL-1的下调很可能是通过将PSGL-1连接到依赖于笼蛋白的成分上来实现的 贩运路径。在本申请中，我们将追求以下目标：具体目标1是表征 PSGL-1用于灭活HIV传染性。我们建议确定限制HIV-1的关键PSGL-1结构域。我们 将确定PSGL-1的结构-功能关系，定义PSGL-1的二聚化作用，N- 末端糖基化和酪氨酸硫化，N-末端十聚体重复序列，以及 限制艾滋病病毒。具体目标2是对PSGL-1灭活HIV病毒的机制进行研究 传染性。我们推测PSGL-1可能通过两种可能的机制限制HIV的传染性：(1) 病毒组装过程中Env掺入的竞争性排斥；(2)残留Env结合的空间位阻 到细胞感受器。我们将检验这两个假说，以确定其作用机制。具体目标3是 研究Nef介导的PSGL-1表面下调的机制。我们将确定功能 Nef和PSGL-1的结构域参与PSGL-1的下调。NEF介导的PSGL-1 下调调控可能会促进病毒在免疫细胞中的传播。

Project Summary/Abstract Mechanisms of PSGL-1 restriction of HIV virion infectivity Restriction factors are an important component of host innate immunity. Studying the anti-HIV mechanisms of restriction factors is central to understanding virus-host interaction. These mechanisms may also offer new therapeutic strategies to inactivate viral reservoirs to achieve lasting HIV remission. Recently, we have identified a new HIV restriction factor, PSGL-1 (P-selectin glycoprotein ligand-1), that can inactivate the infectivity of HIV virions released from HIV producing cells. PSGL-1 is a dimeric mucin-like 120-KD glycoprotein that is primarily expressed on the surface of lymphoid and myeloid cells. PSGL-1 binds to P-, L-, and E-selectin, and mediates leukocyte tethering and rolling on endothelium for leukocyte migration into inflamed tissues. PSGL-1 is also an INF-γ-regulated factor involved in Th1-mediated anti-viral activity. Our preliminary mechanistic studies further revealed that PSGL-1 is incorporated into viral particles, which blocks HIV Env incorporation and disables the ability of virions to attach to target CD4 T cells for infection. In addition, we found that PSGL-1 is antagonized by Vpu and Nef through surface down-regulation. Based on these preliminary studies, we hypothesize that: (1) PSGL-1-mediated restriction of HIV infectivity involves its specific domains; (2) PSGL-1 restricts HIV infectivity likely through competitive exclusion of Env incorporation during viral assembly and steric hindrance of Env binding to cell receptors (3) Nef-mediated PSGL-1 down-regulation is likely achieved through linking PSGL-1 to components of clathrin-dependent trafficking pathways. In this application, we will pursue the following aims: Specific Aim 1 is to characterize PSGL-1 for inactivating HIV infectivity. We propose to identify PSGL-1 domains key to restricting HIV-1. We will determine the structure-function relationship of PSGL-1, defining the roles of PSGL-1 dimerization, N- terminal glycosylation and tyrosine sulfation, N-terminal decameric repeats, and the polybasic region in restricting HIV. Specific Aim 2 is to perform mechanistic studies of PSGL-1 inactivation of HIV viral infectivity. We hypothesized that PSGL-1 restricts HIV infectivity likely through two possible mechanisms: (1) competitive exclusion of Env incorporation during viral assembly; (2) steric hindrance of residual Env binding to cell receptors. We will test these two hypotheses to determine the mechanisms of action. Specific Aim 3 is to study the mechanism of Nef-mediated surface down-regulation of PSGL-1. We will identify functional domains of Nef and PSGL-1 for their involvement in PSGL-1 down-regulation. Nef-mediated PSGL-1 downregulation may facilitate viral spread in immune cells.