Characterization of cell GBV-C envelope glycoprotein interactions

描述（由申请人提供）： 与大多数研究HIV复制的体外系统不同，感染HIV的人同时感染多种致病性和非致病性微生物。合并感染可能影响任一感染的临床结果。例如，在HAART之前进行的所有研究中，HIV感染加速丙型肝炎病毒（HCV）相关肝病的进程，而GB病毒C型（GBV-C）的持续感染与HIV感染个体的生存期延长相关。微生物合并感染相互作用以改变疾病进程的机制提供了对HIV自然史的深入了解，并可能确定新的治疗靶点或协助制定预防策略。GBV-C是黄病毒科的成员，GBV-C是人类的常见感染。病毒在人外周血单核细胞（PBMC）中复制，包括B和T淋巴细胞（CD 4+和CD 8+亚群）。PBMCs与GBV-C和HIV的共感染导致HIV复制的抑制，这部分是通过HIV辅助受体的下调和可溶性抗病毒因子的诱导介导的。GBV-C抑制嗜性CCR 5和CXCR 4 HIV分离株的复制，并抑制来自世界各地的多种HIV分支。此外，GBV-C感染似乎阻断接受IL-2治疗的人中的CD 4扩增，并且与通过几种标志物（CD 38、CCR 5、CD 25、CD 69）的表面表达测量的T细胞活化降低相关。加入包膜糖蛋白E2可重现其中一些作用（CCR 5调节、HIV抑制），尽管尚未对其进行彻底研究。由于GBV-C E2蛋白在不依赖于GBV-C复制而加入细胞中时介导HIV复制抑制，因此这可能是由E2与其细胞受体之间的相互作用介导的。为了验证这一假设，我们计划表征GBV-C E2与其细胞受体之间的相互作用，鉴定和验证候选E2受体，确定E2上与细胞受体相互作用的结构域，并表征GBV-C E2如何调节细胞细胞因子，趋化因子和受体。初步数据已经确定了候选受体，五个初始候选受体中有两个是淋巴细胞表面蛋白，它们在HIV复制和宿主免疫反应中都很重要。我们提出了四个目标，以进一步确定候选受体，验证这些受体与E2相互作用，我们将评估E2和GBV-C颗粒与淋巴细胞（复制和不）的相互作用对趋化因子受体表达，T细胞活化和诱导抗HIV趋化因子的影响。最后，我们将开始表征E2内负责结合和调节T细胞受体表达的结构域。这些信息有可能促进基于细胞的抗HIV和免疫调节治疗的发展。 公共卫生相关性： VA医疗保健系统是美国艾滋病毒感染者的最大提供者，尽管艾滋病毒现在可以通过抗逆转录病毒疗法（ART）进行管理，但ART价格昂贵，副作用显著，并且很容易产生耐药性，特别是在不严格遵守治疗的患者中。GB病毒C是一种常见的人类病毒，似乎不会引起任何疾病，研究表明它与HIV在同一细胞中生长，从而抑制HIV复制。该申请旨在研究GBV-C病毒蛋白（E2）之一如何修饰T细胞，使HIV不能生长，并将作为开发HIV治疗新方法的基础。

DESCRIPTION (provided by applicant): Unlike most in vitro systems developed to study HIV replication, humans infected with HIV are co- infected with a variety of pathogenic and nonpathogenic microbes. Coinfections may influence the clinical outcome of either infection. For example, HIV infection accelerates the course of hepatitis C virus (HCV) related hepatic disease while persistent infection with GB virus type C (GBV-C) is associated with prolonged survival in HIV-infected individuals in all studies conducted prior to HAART. The mechanism(s) by which microbial coinfections interact to alter disease course provide insight into HIV natural history, and may identify novel therapeutic targets or assist in the development of prevention strategies. GBV-C is a member of the family Flaviviridae and GBV-C is a common infection of humans. The virus replicates in human peripheral blood mononuclear cells (PBMCs) including B and T lymphocytes (CD4+ and CD8+ subsets). Co-infection of PBMCs with GBV-C and HIV results in inhibition of HIV replication, and this is mediated in part by downregulation of HIV coreceptors and by induction of soluble antiviral factors. GBV-C inhibits the replication of both CCR5- and CXCR4-tropic HIV isolates, and inhibits diverse HIV clades from around the world. In addition, GBV-C infection appears to block CD4 expansion in people receiving IL-2 therapy and is associated with decreased T cell activation as measured by surface expression of several markers (CD38, CCR5, CD25, CD69). Addition of the envelope glycoprotein E2 reproduces some of these effects (CCR5 regulation, HIV inhibition) although it has not been thoroughly studied. Since GBV-C E2 protein, when added to cells independent of GBV-C replication mediates HIV replication inhibition, it is likely that this is mediated by interactions between E2 and its cellular receptor or receptors. To examine this hypothesis, we plan to characterize interactions between GBV-C E2 and it's cellular receptor(s), identify and validate candidate E2 receptors, determine the domains on E2 that interact with cellular receptors, and characterize how GBV-C E2 modulates cellular cytokines, chemokines and receptors. Preliminary data have identified candidate receptors, and two of the five initial candidates are lymphocyte surface proteins that are important in both HIV replication and host immune responses. We propose four aims to further identify candidate receptors, validate that these receptors interact with E2, and we will assess the effect of E2 and GBV-C particle interaction with lymphocytes (both replicating and not) on chemokine receptor expression, T cell activation, and induction of anti-HIV chemokines. Finally, we will begin characterization of the domain within E2 that is responsible for binding and modulating T cell receptor expression. This information has the potential to facilitate development of cellular-based, anti-HIV and immunomodulatory treatments. PUBLIC HEALTH RELEVANCE: The VA healthcare system is the largest provider for HIV-infection people in the United States, and although HIV can now be managed by antiretroviral therapy (ART), ART is expensive, has significant side effects, and resistance develops easily, particularly in patients who are not strictly adherent to therapy. GB virus C is a common human virus that does not appear to cause any disease, and studies show that it grows in the same cells as HIV resulting in inhibition of HIV replication. This application proposes to study how one of the GBV-C virus proteins (E2) modifies T cells so that HIV does not grow as well, and will serve as the basis for developing new approaches to HIV therapy.