Nef Function in HIV Infectivity

Nef 在 HIV 感染中的作用

• 批准号: 7554896
• 负责人: HEINRICH GOTTLINGER
• 金额: $ 39.97万
• 依托单位: UNIV OF MASSACHUSETTS MED SCH WORCESTER
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-07-01 至 2012-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7554896
• 关键词: Acquired Immunodeficiency Syndrome; Address; Affect; Antibodies; Binding; Cell membrane; Cell physiology; Cells; Clathrin; Cultured Cells; Development; Disease; Dominant-Negative Mutation; Dynamin 2; Dynamin I; Endocytic Vesicle; Goals; Guanosine Triphosphate Phosphohydrolases; HIV; HIV-1; Human; Individual; Link; Mediating; Primate Lentiviruses; Protein Isoforms; Proteins; Public Health; RNA Interference; Role; Small Interfering RNA; Viral; Virion; Virulence Factors; Virus; coated pit; in vivo; nef Protein; novel; particle; trafficking

DESCRIPTION (provided by applicant): Nef is a virulence factor of HIV-1 and other primate lentiviruses that is crucial for rapid progression to AIDS. In cell culture, Nef increases the infectivity of HIV-1 progeny virions by an unknown mechanism. We have recently identified dynamin 2 (Dyn2), a key regulator of clathrin-mediated trafficking, as a novel Nef binding partner that is required for its ability to increase viral infectivity. Dominant-negative Dyn2 or the depletion of Dyn2 by RNAi potently inhibited the effect of Nef on HIV-1 infectivity. Furthermore, in Dyn2-depleted cells this function of Nef could be rescued by ectopically expressed Dyn2 but not by Dyn1, a closely related isoform that does not bind Nef. The infectivity enhancement by Nef also depended on clathrin, since it was diminished in clathrin- depleted cells and profoundly inhibited in cells expressing the clathrin-binding domain of AP180, which blocks clathrin-coated pit formation. This application focuses on the role of the Nef-Dyn2 interaction in the enhancement of HIV-1 infectivity and replication, because this function of Nef remains poorly understood. One goal is to elucidate what enables Nef to interact with Dyn2 but not with closely related isoforms. In particular, since the two Dyn2 domains which confer Nef binding mediate Dyn2 assembly, we propose to determine whether Nef binding and Dyn2 assembly are linked. Another goal is to understand the relevance of Dyn2 and its interaction with Nef for HIV-1 infectivity and replication. Our preliminary results also indicate that Nef affects the neutralization sensitivity of HIV-1 virions, which could have important implications for HIV-1 replication in vivo. We therefore propose to examine the relationship between the effects of Nef on HIV-1 infectivity and neutralization, the breadth of the effect of Nef on neutralization, and whether Nef affects antibody binding to the Env trimer. PUBLIC HEALTH RELEVANCE The Nef protein of HIV-1 is crucial for the development of AIDS in infected individuals. Nef directly enhances the infectivity of viral particles by an unknown mechanism, and we have recently shown that a human protein called dynamin 2 binds to Nef and is required for its ability to enhance virus infectivity. Since this activity of Nef is likely to affect the ability of HIV-1 to spread in people, we propose to examine what enables Nef to bind dynamin 2 but not closely related proteins, and to investigate the relevance of this interaction for the ability of HIV-1 to infect human cells and to cause disease.

描述（由申请人提供）：NEF是HIV-1和其他灵长类动病毒的毒力因子，对于快速发展至艾滋病至关重要。在细胞培养中，NEF通过未知机制增加了HIV-1后代病毒体的感染性。我们最近确定了Dynatin 2（Dyn2），这是网格蛋白介导的运输的关键调节剂，是一种新型的NEF结合伴侣，这是其提高病毒感染性的能力所必需的。 RNAi的主要阴性DYN2或DYN2的耗竭可有效抑制NEF对HIV-1感染性的影响。此外，在dyn2耗尽的细胞中，这种NEF的功能可以通过异位表达的DYN2挽救，而不是由Dyn1（一种密切相关的同工型），不结合NEF。 NEF的感染性增强也取决于网格蛋白，因为它在网格蛋白耗尽的细胞中降低，并在表达AP180的网格蛋白结合结构域的细胞中严重抑制，这阻断了网格蛋白涂层的坑形成。该应用的重点是NEF-DYN2相互作用在增强HIV-1感染性和复制方面的作用，因为NEF的这种功能仍然很少了解。一个目标是阐明什么使NEF与DYN2相互作用，但与密切相关的同工型相互作用。特别是，由于赋予NEF结合的两个DYN2域介导Dyn2组装，因此我们建议确定NEF结合和DYN2组件是否连接起来。另一个目标是了解Dyn2的相关性及其与NEF与HIV-1感染性和复制的相互作用。我们的初步结果还表明，NEF会影响HIV-1病毒体的中和敏感性，这可能对体内HIV-1复制具有重要意义。因此，我们建议检查NEF对HIV-1感染性和中和的影响之间的关系，NEF对中和的影响的广度以及NEF是否影响与Env Trimer的抗体结合。公共卫生相关性HIV-1的NEF蛋白对于受感染个体的艾滋病的发展至关重要。 NEF通过未知机制直接增强了病毒颗粒的感染性，我们最近表明，一种称为Dynamin 2的人蛋白与NEF结合，这对于增强病毒感染的能力是必需的。由于NEF的这种活性可能会影响HIV-1在人中传播的能力，因此我们建议检查什么使NEF结合Dynamin 2但不密切相关的蛋白质，并研究这种相互作用对于HIV-1感染人类细胞并引起疾病的能力的相关性。