Role of Tetraspan Glycoproteins (CD63) in HIV Entry

四跨糖蛋白 (CD63) 在 HIV 进入中的作用

• 批准号: 6590953
• 负责人: WILLIAM AUSTIN O'BRIEN
• 金额: $ 33.19万
• 依托单位: UNIVERSITY OF TEXAS MED BR GALVESTON
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-12-15 至 2005-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6590953
• 关键词: HIV infections; SDS polyacrylamide gel electrophoresis; biological signal transduction; cell membrane; clinical research; confocal scanning microscopy; cytokine; cytokine receptors; endocytosis; flow cytometry; glycoproteins; human immunodeficiency virus 1; human tissue; immunoprecipitation; macrophage; monoclonal antibody; polymerase chain reaction; virion; virus infection mechanism; virus receptors

DESCRIPTION (provided by applicant): HIV infection of primary human cells typically requires interaction with both the primary receptor CD4 and a chemokine coreceptor, either CCR5 or CXCR4. There may be additional cellular factors involved in determining the efficiency of HIV entry, however, which may explain (in part) the marked differences in susceptibility to HIV infection among primary cells from different donors. During studies to identify H1V cofactors in macrophages (Mphi), we found that four independently generated monoclonal antibodies (mAb) to a tetraspan membrane glycoprotein, CD63, could block HIV entry into Mphi, but not T-cells. The function of CD63 is not well understood, but another tetraspan protein, CD81, has been proposed as a receptor for Hepatitis C virus, and CD9 has been implicated in infection by feline immunodeficiency virus (FIV). CD63 has previously been recognized as a platelet activation marker, as well as a marker for malignant melanoma, both seemingly unrelated to retroviral infection. Our experiments will investigate the role of CD63 in HIV entry into MO). These studies may lead to development of novel antiretroviral therapies targeting HIV entry. We will pursue these Aims over four years: Specific Aim 1. To test the hypothesis that CD63 has a specific role in facilitating HIV entry into Mphi, we will identify potential mechanisms of CD63-mediated HIV entry. Experiments will be prioritized based on initial studies that more precisely identify the step of HIV entry involved in anti-CD63 inhibition, directing initial studies toward late entry events, such as the role of CD63 in CD4 turnover or virus/receptor endocytosis and signal transduction, or early events, such as anti-CD63 interactions at the cell surface including binding and CCR5-dependent entry pathways. Specific Aim 2. To test the hypothesis that cell-specific differences between HIV target cells affect the role of CD63 in facilitating HIV entry. We will control for and test effects of various levels of receptor/coreceptor/CD63 expression and activation markers on susceptibility to anti-CD63 inhibition in primary CD4+ cells. We will also test cell line models of anti-CD63 inhibition, which would facilitate cell manipulation and the consistency of inhibitory effects in these studies. Specific Aim 3. To test the hypothesis that infection of Mphi by diverse primary HIV-1 strains can be inhibited by anti- CD63 mAb. This will include both subtype B and non-subtype B strains, including some that exclusively use CCR5 as a coreceptor for efficient infection (R5), as well as dual-tropic strains that use either CXCR4 or CCR5 (R5X4), and X4 strains able to replicate in Me. Achieving these Aims will provide important new insights into the mechanism of HIV-1 entry, and may well provide important new targets for antiretroviraI therapy.

描述（由申请人提供）： 原代人类细胞的 HIV 感染通常需要与原代受体 CD4 和趋化因子辅助受体（CCR5 或 CXCR4）相互作用。然而，在决定 HIV 进入效率时可能还涉及其他细胞因素，这可能（部分）解释了来自不同供体的原代细胞对 HIV 感染的易感性的显着差异。在识别巨噬细胞 (Mphi) 中 H1V 辅因子的研究中，我们发现四种独立生成的针对四跨膜糖蛋白 CD63 的单克隆抗体 (mAb) 可以阻止 HIV 进入 Mphi，但不能阻止 T 细胞。 CD63 的功能尚不清楚，但另一种四跨蛋白 CD81 已被认为是丙型肝炎病毒的受体，而 CD9 与猫免疫缺陷病毒 (FIV) 的感染有关。 CD63 先前被认为是血小板活化标记物以及恶性黑色素瘤的标记物，两者似乎与逆转录病毒感染无关。我们的实验将研究 CD63 在 HIV 进入 MO 中的作用。这些研究可能会导致针对艾滋病毒进入的新型抗逆转录病毒疗法的开发。我们将在四年内实现这些目标： 具体目标 1. 为了检验 CD63 在促进 HIV 进入 Mphi 方面具有特定作用的假设，我们将确定 CD63 介导的 HIV 进入的潜在机制。实验将根据初步研究确定优先顺序，这些研究更准确地确定参与抗 CD63 抑制的 HIV 进入步骤，将初步研究引导至晚期进入事件，例如 CD63 在 CD4 周转或病毒/受体内吞作用和信号转导中的作用，或早期事件，例如细胞表面的抗 CD63 相互作用，包括结合和 CCR5 依赖性进入途径。具体目标 2. 检验 HIV 靶细胞之间的细胞特异性差异影响 CD63 在促进 HIV 进入中的作用这一假设。我们将控制并测试不同水平的受体/辅助受体/CD63 表达和激活标记物对原代 CD4+ 细胞中抗 CD63 抑制的敏感性的影响。我们还将测试抗 CD63 抑制的细胞系模型，这将有助于这些研究中的细胞操作和抑制效果的一致性。具体目标 3. 检验以下假设：抗 CD63 mAb 可以抑制不同原发性 HIV-1 毒株对 Mphi 的感染。这将包括 B 亚型和非 B 亚型菌株，包括一些专门使用 CCR5 作为有效感染的辅助受体 (R5) 的菌株，以及使用 CXCR4 或 CCR5 (R5X4) 的双向菌株，以及能够在我身上复制。实现这些目标将为 HIV-1 进入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目标。