Novel host proteins in the HIV-1 preintegration complexes

描述(由申请人提供)：HIV-1前整合复合体(PIC)由HIV-1全长DNA、病毒蛋白和宿主蛋白组成，对病毒整合和复制至关重要。以前的研究已经确定并鉴定了几种与HIV-1整合酶相互作用并在病毒感染中发挥重要作用的人类蛋白。然而，在获得足够数量的催化活性PIC方面的技术限制阻碍了这一进程。 与HIV-1 PIC相关的宿主蛋白的全面鉴定。我们最近开发了一种新的方法，并鉴定了18种新的人类蛋白，这些蛋白与从受感染的CD4+T细胞中分离的HIV-1 PIC特异相关。我们的初步研究表明，其中一种蛋白质，非POU结构域的八聚体结合蛋白(NONO)，是在CD4+T细胞中有效感染HIV-1所必需的。然而，目前尚不清楚我们鉴定的其他17种PIC相关宿主蛋白是否能影响HIV-1感染。NONO是一种多功能核蛋白，参与细胞内转录调控、mRNA剪接和DNA修复，但其影响HIV-1感染的机制尚不清楚。我们试图在这份R21提案中解决这两个重要问题。我们假设，HIV-1感染细胞中与催化活性PIC特异相关的宿主蛋白(如NONO)在病毒整合和感染中发挥作用。为了验证这一假说，我们设计了以下两个特定目的：目的1.确定新型PIC相关宿主蛋白在HIV-1感染中的作用。在目标1中，除了NONO外，我们还可能在这18个候选蛋白中确定其他与PIC相关的宿主蛋白，这些蛋白对HIV-1感染有显著影响。我们将选择对病毒感染有最显著影响的2-3个候选蛋白来进一步研究它们的作用机制。目的2.探讨PIC相关宿主蛋白影响HIV-1感染的机制。我们将首先研究NONO影响HIV-1感染的机制，然后我们将进行类似的研究，以检查目标1中确定的其他候选蛋白。该项目的长期目标是了解PIC相关宿主蛋白在HIV-1整合或感染中的功能和机制，并开发新的治疗策略。一种新的抗HIV治疗策略是阻断HIV-1整合酶与其关键细胞辅助因子之间的相互作用。我们提议的研究有可能确定阻止HIV-1整合和感染的新药物靶点。 与公共卫生相关：全世界有超过3300万艾滋病毒携带者，美国每年约有56,300人新感染艾滋病毒。虽然抗逆转录病毒疗法可以控制感染者中的HIV-1感染，但需要新的抗HIV药物靶点来提高疗效和防止耐药性。该项目旨在确定和表征对艾滋病毒整合过程至关重要的新宿主蛋白。拟议研究的结果将有助于确定阻止艾滋病毒整合和感染的新药物靶点。

DESCRIPTION (provided by applicant): The HIV-1 pre-integration complex (PIC) that comprises of HIV-1 full-length DNA, viral proteins, and host proteins is essential for viral integration and replication. Previous studies have identified and characterized several human proteins that interact with HIV-1 integrase and play important roles in viral infection. However, the technical limitations in acquiring sufficient amounts of catalytically active PICs has hindered a comprehensive identification of host proteins associated with the HIV-1 PICs. We recently developed a novel approach and identified 18 new human proteins that are specifically associated with the HIV-1 PICs isolated from infected CD4+ T cells. Our preliminary study suggested that one of these proteins, non-POU domain-containing octamer-binding protein (NonO), is required for efficient HIV-1 infection in CD4+ T cells. However, it is unclear whether the other 17 PIC- associated host proteins that we identified can affect HIV-1 infection. NonO is a multifunctional nuclear protein involved in transcription regulation, mRNA splicing, and DNA repair in the cell, but the mechanism by which NonO affects HIV-1 infection is not known. We seek to address these two significant questions in this R21 proposal. We hypothesize that host proteins (such as NonO) specifically associated with catalytically active PICs in HIV-1-infected cells play a role in viral integration and infection. The following two specific aims are designed o test this hypothesis: Aim 1. To determine the role of novel PIC-associated host proteins in HIV-1 infection. In addition to NonO, we may identify other PIC-associated host proteins among these 18 candidates that can significantly affect HIV-1 infection in Aim 1. We will select 2-3 protein candidates that have the most significant effect on viral infection to further study their mechanisms of action. Aim 2. To investigate the mechanisms by which PIC-associated host proteins affect HIV-1 infection. We will first investigate the mechanism by which NonO affects HIV-1 infection, and we will then perform similar studies to examine other protein candidates identified in Aim 1. The long-term goal of this project is to understand the function and mechanism of PIC-associated host proteins in HIV-1 integration or infection and to develop novel therapeutic strategies. An emerging anti-HIV therapeutic strategy is to block the interactions between HIV-1 integrase and its critical cellular cofactors. Our proposed study has the potential to identify novel drug targets to block HIV-1 integration and infection. PUBLIC HEALTH RELEVANCE: There are over 33 million people living with HIV worldwide and approximately 56,300 new HIV infections each year in the US. Although antiretroviral therapy can control HIV-1 infection in infected individuals, new anti-HIV drug targets are needed to increase efficacy and prevent drug resistance. This project aims to identify and characterize novel host proteins that are essential for the HIV integration process. The findings from the proposed studies will help identify novel drug targets for blocking HIV integration and infection.