THE ROLE OF HIV-1 VPU IN THE REGULATION OF CD40

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 本研究的目的是了解HIV-1蛋白VPU在调节细胞因子受体CD40中的作用。我们以前的研究表明，HIV-1感染的内皮细胞(EC)支持非霍奇金淋巴瘤(NHL)B细胞的牢固附着。这一机制被追溯到CD40的病毒增强，允许CD40依赖的VCAM-1的诱导。在体内，HIV感染的EC可能因此有助于艾滋病非霍奇金淋巴瘤的结外表现。HIV-1辅助蛋白VPU被发现通过转录后途径诱导CD40。虽然VPU改变了CD4和某些其他细胞蛋白的表达，但其影响的全部范围尚未确定。VPU诱导的免疫调节蛋白可能会对艾滋病相关疾病产生严重影响，因此值得进一步研究。我们假设VPU与正常的CD40调节通路相交，从而增加了细胞表面功能性CD40受体的水平。令人惊讶的是，无论病毒调节剂的影响如何，人们对CD40在细胞中的正常更新途径知之甚少。通过研究VPU对CD_4~+和CD_4~+细胞中CD40的调节作用，我们的目标是：1)明确正常的CD40调节途径；2)阐明VPU在白细胞和非白细胞靶点中的作用；3)评价这些发现的临床意义。在目标1中，我们将使用生化和成像技术来比较在VPU存在和不存在的情况下，CD40在EC中的生物合成和转化途径。在目标2中，我们将使用一系列VPU突变蛋白来确定已知的结构功能基序是否在CD40调节中发挥作用，或者是否涉及独特的基序和机制。在目标3中，我们将把这些研究扩大到包括巨噬细胞，这是一个主要的CD4+/CD40+HIV-1靶点。成功地完成这些目标将使人们对CD40调节和VPU功能有更深入的了解，并可能识别具有免疫调节和/或抗病毒意义的靶点。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The goal of this study is to understand the role of the HIV-1 protein Vpu in the regulation of the cytokine receptor CD40. Our previous studies demonstrated that HIV-1-infected endothelial cells (EC) support the firm attachment of non-Hodgkin's lymphoma (NHL) B cells. This mechanism was traced to viral enhancement of CD40, allowing for CD40-dependent induction of VCAM-1. In vivo, HIV-infected EC may thus contribute to the characteristic extranodal presentation of the AIDS-NHL. The HIV-1 accessory protein Vpu was found to be responsible for CD40 induction via a post-transcriptional pathway. While Vpu alters the expression of CD4 and certain other cellular proteins, the full scope of its influence has yet to be determined. Vpu-induction of immune regulatory proteins could have severe implications for AIDS-related conditions and is thus deserving of further study. We hypothesize that Vpu intersects with normal CD40 regulation pathways to increase the levels of functional CD40 receptor at the cell surface. Surprisingly little is known about the normal turnover pathways of CD40 in cells, regardless of the influence of a viral modulator. By studying Vpu-modulation of CD40 in CD4-negative as well as CD4-positive cells, we aim to i) clearly define normal CD40 regulation pathways, ii) clarify the role of Vpu in leukocyte and non-leukocyte targets, and iii) appreciate the clinical significance of these findings. In Aim 1 we will use biochemical and imaging techniques to compare the biosynthetic and turnover pathways of CD40 in EC in the presence and absence of Vpu. In Aim 2 we will use a series of Vpu mutant proteins to determine whether known structure-function motifs play a role in CD40 regulation, or if unique motifs and mechanisms are involved. In Aim 3 we will extend these studies to include macrophages, a major CD4+/CD40+ HIV-1 target. Successful completion of these Aims will lead to an increased understanding of both CD40 regulation and Vpu function, and may identify targets of immune modulatory and/or anti-viral significance.