Mechanism and role of nuclear entry in HIV-1 replication

描述(由申请人提供)：HIV-1利用细胞机制进入核的能力使病毒能够感染静止的CD4+T细胞和组织巨噬细胞，这两种不分裂的细胞类型被认为在HIV-1的传播、持久性和发病机制中发挥关键作用。然而，HIV-1复制过程中核进入的确切机制和具体要求仍不清楚。衣壳是HIV-1感染未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的依赖性与它们感染未分裂细胞的能力之间存在完美的相关性。本项目验证的假设是，衣壳蛋白通过协助细胞内病毒复合体劫持依赖TNPO3的细胞机制来调节HIV-1的核进入，这一能力对HIV-1在体内的复制至关重要。在目标1中，我们将确定HIV-1衣壳蛋白决定HIV-1对TNPO3的利用，以及TNPO3和其他宿主因素如何促进HIV-1核进入。接下来，我们将定义这种依赖于TNPO3的核进入对HIV-1在体内复制的具体要求。在目标2中，我们将研究一例携带人类白细胞抗原B27的患者中HIV-1衣壳演变的独特案例。初步数据表明，这些患者反复选择依赖TNPO3的病毒。在目标3中，我们将通过使用猴免疫缺陷病毒(SIV)将我们的基因数据与HIV-1猕猴模型相结合，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于TNPO3的SIV突变体，它们要么保留要么失去感染未分裂细胞的能力。这项拟议研究的成功完成将有助于我们了解艾滋病毒-1利用核运输机制最大限度地传播它的机制。 与公共卫生相关：艾滋病毒-1是艾滋病的病原体，它主动将病毒DNA运送到细胞核中的能力将其细胞范围扩大到非分裂细胞类型，从而使艾滋病毒-1可以感染静止的CD4+T细胞和组织巨噬细胞。该项目的目标是确定核进入在HIV-1复制中的机制和作用。我们预计，我们的发现可能对开发针对病毒衣壳蛋白和核进入的新型抗病毒药物具有实际意义，并将为研究静息的CD4+T细胞和巨噬细胞如何在体内对HIV-1感染的多个方面做出贡献提供新的见解。

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. PUBLIC HEALTH RELEVANCE: The ability of HIV-1, the causative agent of AIDS, to actively transport viral DNA into the nucleus expands its cellular range to non-dividing cell types such that HIV-1 can infect resting CD4+ T cells and tissue macrophages. The goal of this project is to determine the mechanism and role of nuclear entry in HIV-1 replication. We expect that our findings could have practical implications for the development of novel antiviral drugs targeting viral capsid protein and nuclear entry and will provide novel insights into how resting CD4+ T cells and macrophages contribute to multiple aspects of HIV-1 infection in vivo.