Host protein targets of HIV-1 Vpr in gene expression, cell cycle and innate immunity

HIV-1 Vpr 在基因表达、细胞周期和先天免疫中的宿主蛋白靶标

基本信息

批准号：
10681282
负责人：
Paul D. Bieniasz
金额：
$ 42.38万
依托单位：
ROCKEFELLER UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-09-17 至 2025-08-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10681282
关键词：
African Green Monkey Biological Cell Cycle Cell Cycle Arrest Cell Cycle Progression Cell Cycle Regulation Cell Death Cell Death Induction Cells Chromatin Chromosomes Cis-Acting Sequence Clustered Regularly Interspaced Short Palindromic Repeats Complex Cullin Proteins DNA DNA Damage Dependence Enhancers Gene Expression HIV HIV-1 Human Hybrids Innate Immune Response Integration Host Factors Investigation Lentivirus M cell Macaca mulatta Macrophage Maps Mediating Mediator Messenger RNA Methods Mitotic Molecular Natural Immunity Nuclear Primates Proteins Proteomics RNA Interference Regulation Reporter Reporting Repression Role SIV Series Viral Viral Genes Viral Proteins Virus Virus Integration Yeasts cell type cofactor fascinate gene repression insight integration site knock-down mutant novel therapeutic intervention prevent promoter response screening sensor ubiquitin-protein ligase vector viral DNA

项目摘要

The role and mechanism of action of Vpr (Viral Protein R), an accessory protein encoded by HIV- 1, has been enigmatic for decades. Vpr causes cell cycle arrest at G2/M, triggers a DNA damage response, and enhances viral gene expression. It exerts these activities by targeting host protein(s) for degradation, hijacking cullin4-based E3 ubiquitin ligase complex (CRL4) to induce their depletion. We recently identified a host protein CCDC137, also known as cPERP-B, as a key target protein depleted by Vpr in a CRL4 complex dependent manner. Specifically, CCDC137 depletion by RNA interference recapitulates the aforementioned effects of Vpr on host and virus. In this project we seek to study the molecular details of how CCDC137 represses HIV-1 gene expression as well as how it controls cell cycle progression and the DNA damage response. In Aim 1, we will determine whether CCDC137 depletion is a conserved feature of Vpr proteins from diverse HIV and SIV strains, map the CCDC137 determinants required for Vpr-induced depletion, and assess the effect of Vpr from diverse viruses on viral gene expression. In addition, we will define host proteins required for CCDC137 depletion by Vpr. Aim 2 is centered on the mechanisms of CCDC137-mediated repression of HIV-1 gene expression. We will delineate cis- acting sequences required for CCDC137-mediated repression and evaluate the effect of integration and integration site selection on the Vpr/CCDC137-regulated HIV-1 gene expression. We will also combine screening methods (proteomics, yeast 2-hybrid, and CRISPR functional screens) to identify CCDC137 interacting cofactor(s) to illuminate the mechanism of how CCDC137 inhibits HIV-1 gene expression. In Aim 3 we will investigate how CCDC137 prevents DNA damage response and controls cell cycle progression. In particular, we will determine whether CCDC137 protects chromosomal DNA and delineate host factor(s) cooperating with CCDC137 to modulate the DNA damage response.

Vpr（病毒蛋白 R）的作用和作用机制，一种由 HIV 编码的辅助蛋白 1、几十年来一直是个谜。 Vpr 导致细胞周期停滞在 G2/M 期，引发 DNA 损伤反应，并增强病毒基因表达。它通过靶向宿主来发挥这些活动蛋白质降解，劫持基于 cullin4 的 E3 泛素连接酶复合物 (CRL4) 诱导他们的耗尽。我们最近鉴定了一种宿主蛋白 CCDC137，也称为 cPERP-B，作为 Vpr 以 CRL4 复合物依赖性方式耗尽关键靶蛋白。具体来说，CCDC137 RNA 干扰造成的耗竭概括了上述 Vpr 对宿主和病毒的影响。在这个项目中，我们寻求研究 CCDC137 如何抑制 HIV-1 基因的分子细节表达以及它如何控制细胞周期进程和 DNA 损伤反应。在目标 1，我们将确定 CCDC137 耗尽是否是 Vpr 蛋白的保守特征不同的 HIV 和 SIV 毒株，绘制 Vpr 诱导消耗所需的 CCDC137 决定簇，并评估不同病毒的 Vpr 对病毒基因表达的影响。此外，我们将定义 Vpr 消耗 CCDC137 所需的宿主蛋白。目标 2 的中心是 CCDC137介导的HIV-1基因表达抑制机制。我们将划定顺式 CCDC137 介导的抑制所需的作用序列并评估其效果 Vpr/CCDC137 调节的 HIV-1 基因表达的整合和整合位点选择。我们还将结合筛选方法（蛋白质组学、酵母 2-杂交和 CRISPR 功能）筛选）来识别 CCDC137 相互作用的辅因子，以阐明其机制 CCDC137 抑制 HIV-1 基因表达。在目标 3 中，我们将研究 CCDC137 如何预防 DNA 损伤反应并控制细胞周期进程。特别是，我们将确定 CCDC137 是否保护染色体 DNA 并描述与其合作的宿主因子 CCDC137 调节 DNA 损伤反应。