Mechanism and role of nuclear entry in HIV-1 replication

描述（由申请人提供）：HIV-1利用细胞机制进入细胞核的能力使病毒能够感染静息的CD4+ T细胞和组织巨噬细胞，这两种非分裂细胞类型被认为在HIV-1的传播、持续和发病中起关键作用。然而，HIV-1复制中核进入的确切机制和具体要求尚不清楚。衣壳是HIV-1感染非分裂细胞的主要决定因素，但也决定了HIV-1对TNPO3的依赖性，TNPO3是一种最近与HIV-1核进入有关的宿主分子。我们的初步研究发现，HIV-1衣壳突变体对TNPO3的依赖性与其感染非分裂细胞的能力之间存在完美的相关性。在这个项目中测试的假设是，衣壳蛋白通过协助细胞内病毒复合物劫持依赖tnpo3的细胞机制来调节HIV-1的核进入，这种能力对HIV-1在体内的复制至关重要。在Aim 1中，我们将确定HIV-1衣壳蛋白如何决定HIV-1对TNPO3的使用，以及TNPO3和其他宿主因子如何促进HIV-1进入核。接下来，我们将定义这种依赖tnpo3的核进入对HIV-1体内复制的具体要求。在Aim 2中，我们将研究携带HLA-B27的患者中HIV-1衣壳进化的独特案例。初步资料显示，这些患者反复选择依赖TNPO3的病毒。在Aim 3中，我们将通过使用猴免疫缺陷病毒（SIV）将我们的遗传数据与经过充分研究的HIV-1猕猴模型结合起来，直接解决体内核进入的重要性。在这里，我们将产生和利用不依赖tnpo3的SIV突变体，这些突变体要么保留要么失去感染非分裂细胞的能力。这项研究的成功完成将有助于我们了解HIV-1利用核转运机制最大化其传播的机制。

DESCRIPTION (provided by applicant): The ability of HIV-1 to utilize the cellular machinery for nuclear entry allows the virus to infect resting CD4+ T cells and tissue macrophages, the two non-dividing cell types believed to play critical roles in HIV-1 transmission, persistence and pathogenesis. However, the precise mechanism and specific requirement of nuclear entry in HIV-1 replication remains unclear. Capsid is the major determinant for HIV-1 infection of non-dividing cells but also dictates the dependence of HIV-1 on TNPO3, a host molecule recently implicated in HIV-1 nuclear entry. Our preliminary studies found a perfect correlation between the dependence of HIV-1 capsid mutants on TNPO3 and their ability to infect non-dividing cells. The hypothesis being tested in this project is that the capsid protein regulates HIV-1 nuclear entry by assisting intracellular virus complexes in hijacking the TNPO3-dependent cellular machinery and this ability is critical for HIV-1 replication in vivo. In Aim 1, we will determine hw the HIV-1 capsid protein determines TNPO3 usage by HIV-1 and how TNPO3 and other host factors promote HIV-1 nuclear entry. We will next define the specific requirement of this TNPO3-dependent nuclear entry for HIV-1 replication in vivo. In Aim 2, we will study a unique case of HIV-1 capsid evolution in patients carrying the HLA-B27. Preliminary data suggests recurrent selection of TNPO3- dependent viruses in these patients. In Aim 3, we will combine our genetic data with a well-studied macaque model of HIV-1 by using simian immunodeficiency virus (SIV) to directly address the importance of nuclear entry in vivo. Here we will generate and utilize TNPO3-independent SIV mutants, which either retain or lose the ability to infect non-dividing cells. Successful completion of this proposed research will help us understand the mechanism by which HIV-1 exploits the nuclear transport machinery to maximize its propagation.