HIV-1 evolution driven by intracellular defenses

HIV-1进化由细胞内防御驱动

• 批准号: 7006413
• 负责人: Viviana A Simon
• 金额: $ 34.48万
• 依托单位: AARON DIAMOND AIDS RESEARCH CENTER
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-06-01 至 2010-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7006413
• 关键词: HIV infections; amination; aminohydrolases; binding sites; biochemical evolution; clinical research; drug resistance; enzyme activity; fluorescent dye /probe; gene expression; gene induction /repression; gene mutation; host organism interaction; human immunodeficiency virus 1; human tissue; molecular genetics; protein degradation; virulence; virus DNA; virus cytopathogenic effect; virus genetics; virus infection mechanism

DESCRIPTION (provided by applicant): Retroviruses have devised a number of strategies to evade cellular mechanisms aimed at preventing retroviral infection. HIV-1 expresses Vif, a protein that counteracts the antiviral activity of the cytidine deaminases APOBEC3G and APOBEC3F. The nucleotide composition of the HIV-1 genome suggests, however, that protection from host-mediated viral cDNA deamination may not be absolute. In preliminary studies, we showed that vif genes encoding proteins that fail to degrade APOBEC3G, APOBEC3F or both can be detected in vivo. The loss of Vif function was mapped to single nucleotide substitutions. These studies indicate that natural variation in Vif function may profoundly impact the extent and direction of viral sequence evolution within HIV-1 infected individuals. The experiments proposed herein will determine the extent to which host mechanisms aimed to prevent retroviral infection, in fact, contribute to viral diversification and pathogenesis. We will analyze the fitness of viruses expressing Vif proteins that are closely related but differ in their ability to neutralize APOBEC3G or APOBEC3F activities. We will also test if variation in Vif function may be beneficial for viral adaptation under certain circumstances (e.g., in the presence of antiretroviral drugs) by determining whether cytidine deamination by APOBEC3G or APOBEC3F selects for certain drug resistance mutations. The pattern of hypermutations associated with partial protection from cytidine deamination will be correlated to HIV fitness. Since APOBEC3 enzymes induce hypermutations in different dinucleotide contexts the understanding of how activity against one enzyme but not the other is maintained is relevant for viral evolution. We will conduct structure function studies to determine whether domains other than the Vif SOCS box motif are necessary and essential for Vif mediated specific neutralization of APOBEC3 enzymes. Finally, we will assess the impact of reverse transcription and APOBECS-driven mutagenesis on loss of Vif function. We will determine the rate of Vif inactivation as result of either reverse transcriptase or cytidine deamination induced mutations on a single cell level using assays based on fluorescence tagged APOBEC3G degradation. Variation in Vif function may influence the pathogenicity of HIV-1 by rendering its genome more or less resistant to deaminase activity and these studies have the potential to reveal how Vif mediated protection from cytidine deamination is modulated in vivo.

描述（由申请人提供）：逆转录病毒已经设计出多种策略来逃避旨在预防逆转录病毒感染的细胞机制。 HIV-1 表达 Vif，这是一种抵消胞苷脱氨酶 APOBEC3G 和 APOBEC3F 抗病毒活性的蛋白质。然而，HIV-1 基因组的核苷酸组成表明，对宿主介导的病毒 cDNA 脱氨基作用的保护可能不是绝对的。在初步研究中，我们表明可以在体内检测到编码不能降解 APOBEC3G、APOBEC3F 或两者的蛋白质的 vif 基因。 Vif 功能的丧失被映射为单核苷酸取代。这些研究表明，Vif 功能的自然变异可能深刻影响 HIV-1 感染者体内病毒序列进化的程度和方向。 本文提出的实验将确定旨在预防逆转录病毒感染的宿主机制实际上在多大程度上有助于病毒多样化和发病机制。我们将分析表达 Vif 蛋白的病毒的适应性，这些蛋白密切相关，但中和 APOBEC3G 或 APOBEC3F 活性的能力不同。我们还将通过确定 APOBEC3G 或 APOBEC3F 的胞苷脱氨是否选择某些耐药突变来测试 Vif 功能的变化是否有利于在某些情况下（例如，在存在抗逆转录病毒药物的情况下）的病毒适应。与胞苷脱氨的部分保护相关的超突变模式将与 HIV 适应性相关。由于 APOBEC3 酶在不同的二核苷酸环境中诱导超突变，因此了解如何维持针对一种酶而不是另一种酶的活性与病毒进化相关。我们将进行结构功能研究，以确定 Vif SOCS 盒基序以外的结构域对于 Vif 介导的 APOBEC3 酶的特异性中和是否是必需的和必需的。最后，我们将评估逆转录和 APOBECS 驱动的诱变对 Vif 功能丧失的影响。我们将使用基于荧光标记的 APOBEC3G 降解的测定法，确定单细胞水平上逆转录酶或胞苷脱氨诱导突变导致的 Vif 失活率。 Vif 功能的变化可能会通过使其基因组或多或少地抵抗脱氨酶活性来影响 HIV-1 的致病性，这些研究有可能揭示体内如何调节 Vif 介导的胞苷脱氨保护。