Role of Tetraspan Glycoproteins (CD63) in HIV Entry

EXCEED提供的空间。HIV感染原代人类细胞通常需要与主要受体CD4和趋化运动受体CCRs或CXCR4相互作用。我们已经发现了CD63在HIV感染M0中的潜在作用，因为针对CD63的单抗可以阻止HIV进入MO，但不能阻止T细胞。CD63是一种四聚糖膜糖蛋白，在血小板上被称为A1激活标志，在人类细胞中几乎无处不在，与整合素密切相关。本实验旨在探讨CD63在HIV进入M0过程中的作用。这些研究可能导致针对HIV进入的新的抗逆转录病毒疗法的开发。我们将在五年内实现这些目标：具体目标1.为了检验抗CD63单抗可以抑制不同原发HIV-1毒株感染M0的假设，这将包括B亚型和非B亚型毒株，包括一些使用CCRs作为辅助受体的毒株(R5)，以及使用CXCR4或CCRs的双嗜性毒株(RSX4)，以及能够在M0中复制的X4毒株。将对不依赖CD4和CCRYCXCR4相互作用进入的两性MLV-Env假型病毒进行评估，以确定抗CD63抗体是否在结合逆转录病毒进入事件后起作用。具体目的2.验证HIV靶细胞之间的细胞特异性差异影响CD63在促进HIV进入中的作用的假设。我们将控制和测试不同水平的受体/辅受体/CD63的表达和激活标记01对抗CD63抑制的敏感性。我们还将测试抗CD63抑制的细胞系模型，这将有助于在这些研究中的细胞操纵和抑制效应的一致性，在目标3中的机制研究中。特定目的3.为了验证CD63在促进HIV进入巨噬细胞方面具有特定作用的假说，我们将确定CD63介导的HIV进入的潜在机制，包括(A)受体/辅受体与CD63的相互作用或共存，(B)CD63与gp120的结合，(C)CD63在CD4周转或病毒/受体内吞中的作用，以及(D)信号转导。这些实验将基于MLV-Env伪型病毒(和X4 HIV-1毒株)的抗CD63敏感性而优先考虑，这将指导我们专注于晚期事件，如果抑制不依赖于CD4和CCRs相关事件。实现这些目标将为了解HIV-1进入的机制提供重要的新见解，并很可能为抗逆转录病毒治疗提供重要的新靶点。PERFORMANCESITE(========================================Section End===========================================

EXCEEDTHE SPACE PROVIDED. HIV infection of primary human cells typically requires interaction with both the primary receptor CD4 and a chemokinecoreceptor either CCRS or CXCR4. We have discovered a potential role for CD63 in HIV infection of M0, as monoclonal antibodies (mAb specific for CD63 can prevent HIV entry into MO, but not T-cells. CD63 is a tetraspan membrane glycoprotein, best known as a1 activation marker on platelets, that is nearly ubiquitous on human cells and is closely associated with integrins. Our experimentswil investigate the role of CD63 in HIV entry into M0. These studies may lead to developmentof novel antiretroviraltherapies targetini HIV entry. We will pursue these Aims over five years: Specific Aim 1. To test the hypothesis that infection of M0by diverse primary HIV-1 strains can be inhibited by anti-CD63 mAb This will include both subtype B and non-subtypeB strains, includingsome that use CCRS as a coreceptor (R5), as well as dual-tropic strains that use either CXCR4 or CCRS (RSX4), and X4 strains able to replicate in M0. Amphotropic MLV-Env-pseudotyped virus which enters independent of CD4 and CCRYCXCR4 interactions, will be assessed to determine if anti-CD63 irhbition acts at post binding retroviral entry events. Specific Aim 2. To test the hypothesis that cell-specific differences between HIV target cells affect the role of CD63 in facilitatini HIV entry. We will control for and test effects of various levels of receptor/coreceptor/CD63 expression and activation markers 01 susceptibility to anti-CD63 inhibition. We will also test cell line models of anti-CD63 inhibition, which would facilitate cel manipulation and the consistencycif lnhibitoryeffects in these studies,and in the mechanistic studiesin Aim 3. Specific Aim 3. To test the hypothesis that CD63 has a specific role in facilitating HIV entry into macrophages, we will identify potential mechanisms of CD63-mediatedHIV entry, including (a) receptor/coreceptor interactions or colocalization with CD63, (b) CD63 binding to gp120, (c) the role of CD63 in CD4 turnover or virus/receptor endocytosis and (d) signal transduction. These experiments will be prioritized based on anti-CD63 susceptibility of MLV-Env pseudotyped virus (and X4 HIV-1 strains), which would direct us to focus on late events if inhibition is not dependent on CD4 and CCRS-associated events. Achieving these Aims will provide important new insights into the mechanism of HIV-1 entry, and may well provide important new targets for antiretroviraltherapy. PERFORMANCESITE( ========================================Section End===========================================