HIV Nef Function and Early Events in Macrophage and Microglia Infection

HIV Nef 功能以及巨噬细胞和小胶质细胞感染的早期事件

• 批准号: 7594513
• 负责人: Jon W Marsh
• 金额: $ 63.66万
• 依托单位: NATIONAL INSTITUTE OF MENTAL HEALTH
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7594513
• 关键词: AIDS neuropathy; Acquired Immunodeficiency Syndrome; Affect; Antigen Presentation; Biochemical; Biochemical Pathway; Brain; Brain Diseases; Brain Pathology; CD4 Positive T Lymphocytes; Cells; Cessation of life; Characteristics; DNA; Development; Event; Family; Goals; HIV; HIV Infections; HIV-1; Human; Individual; Infection; Inflammation; Inflammatory; Knowledge; Lead; Macaca; Mediating; Messenger RNA; Microglia; Neurologic Dysfunctions; Neurons; Pathogenesis; Pathology; Pathway interactions; Peripheral; Process; Production; Proteins; RNA Splicing; Retroviridae; Reverse Transcription; Role; SIV; Site; Stream; Structure; T-Cell Receptor; T-Lymphocyte; Transcript; Viral; Virulence Factors; chemokine; cytokine; genetic regulatory protein; in vivo; macrophage; member; monocyte; nef Genes; nef Protein; p21 activated kinase; response

The objective of this study is to elucidate the intracellular interactions of the HIV regulatory protein Nef with host cell activities. Early in the HIV-1 infection of CD4 T cells this retrovirus expresses regulatory proteins, with the Nef transcript representing nearly 80% of total viral mRNA. In vivo infectivity by SIV and HIV is achieved in the absence of Nef expression, but there is an absolute requirement for Nef in the production of high viral titers. In the absence of Nef, there is loss of development of the immunological and neurological dysfunctions characteristic of AIDS. The essential activity of the Nef protein in the development of AIDS has not been defined. We have discovered that expression of Nef in human CD4 T cells results in a lowering of the activation threshold. Since activation is essential for viral synthesis, Nef thus has a direct role in increasing viral production. Evidence suggests that the biochemical site affected by Nef is early in the T cell receptor pathway with down-stream enhancement of the Erk cascade. We had previously established that the Nef protein associates with a member of the p21-activated kinase (PAK) family, which is known to initiate activation pathways as well as alter cytoskeletal structures. Our exploration of early infection of HIV in quiescent CD4 T cells has lead to the discovery that prior to integration, reverse-transcribed HIV DNA generates multiply spliced transcripts that encode the HIV regulatory proteins. These expressed proteins prime quiescent cells for activation and enhanced viral production. NeuroAIDS is characterized by neuronal death, although HIV does not infect neurons. Infection is largely restricted to the peripheral monocyte-derived brain macrophages and microglia. These cells are responsible for antigen presentation and immunological functions in the brain. There is limited knowledge of the role of Nef in macrophage and microglia, although there is evidence that Nef contributes to proinflammatory processes. It is presently believed that this inflammation, along with other secreted HIV gene products, contributes to neuron death. Infection of macaques by Nef-negative SIV results in no pathology--including no brain disease. We are presently examining biochemical pathways in macrophage and microglial cells that are parallel to those affected by Nef expression in T cells. These pathways in macrophages and microglia are responsible for secretion of inflammatory cytokines and chemokines. An understanding of the biochemical activity of this virulence factor could lead to new options for treating HIV-infected individuals. The recruitment of inflammatory cells in the brain following HIV infection is preceded by chemokine induction. This is mediated by the infection process of the resident macrophage/microglia cells, and appears to be a required event for AIDS-related brain pathologies. We are presently identifying the earliest events that trigger a specific chemokine release. Reverse transcription of the HIV RNA, but not viral replication, in the macrophage/microglia cells appears to be required. Given the contributions of these chemokines to the eventual brain disease, and given that existing HIV treatments affect only viral replication, understanding these mechanisms is critical.

本研究的目的是阐明 HIV 调节蛋白 Nef 与宿主细胞活性的细胞内相互作用。在 HIV-1 感染 CD4 T 细胞的早期，这种逆转录病毒表达调节蛋白，其中 Nef 转录本占病毒总 mRNA 的近 80%。 SIV和HIV的体内感染性是在没有Nef表达的情况下实现的，但在产生高病毒滴度时对Nef有绝对的要求。如果缺乏 Nef，艾滋病特有的免疫和神经功能障碍的发展就会受到阻碍。 Nef 蛋白在艾滋病发展中的基本活性尚未确定。我们发现人类 CD4 T 细胞中 Nef 的表达会导致激活阈值降低。由于激活对于病毒合成至关重要，因此 Nef 在增加病毒产量方面具有直接作用。有证据表明，受 Nef 影响的生化位点位于 T 细胞受体途径的早期，并具有 Erk 级联的下游增强作用。我们之前已经确定 Nef 蛋白与 p21 激活激酶 (PAK) 家族的成员相关，该家族已知可以启动激活途径并改变细胞骨架结构。我们对静态 CD4 T 细胞中 HIV 早期感染的探索发现，在整合之前，逆转录的 HIV DNA 会产生编码 HIV 调节蛋白的多重剪接转录本。这些表达的蛋白质使静止细胞活化并增强病毒产生。 尽管艾滋病毒不感染神经元，但神经艾滋病的特征是神经元死亡。感染主要限于外周单核细胞来源的脑巨噬细胞和小胶质细胞。 这些细胞负责大脑中的抗原呈递和免疫功能。尽管有证据表明 Nef 有助于促炎症过程，但对 Nef 在巨噬细胞和小胶质细胞中的作用的了解有限。 目前认为这种炎症与其他分泌的艾滋病毒基因产物一起导致神经元死亡。 Nef 阴性 SIV 感染猕猴不会导致任何病理变化，包括脑部疾病。 我们目前正在研究巨噬细胞和小胶质细胞中的生化途径，这些途径与 T 细胞中受 Nef 表达影响的途径平行。巨噬细胞和小胶质细胞中的这些途径负责炎症细胞因子和趋化因子的分泌。 了解这种毒力因子的生化活性可能会为治疗艾滋病毒感染者提供新的选择。 HIV感染后大脑中炎症细胞的募集先于趋化因子诱导。 这是由常驻巨噬细胞/小胶质细胞的感染过程介导的，并且似乎是艾滋病相关脑部病变的必要事件。 我们目前正在确定触发特定趋化因子释放的最早事件。 巨噬细胞/小胶质细胞中似乎需要 HIV RNA 的逆转录，但不需要病毒复制。 考虑到这些趋化因子对最终脑部疾病的贡献，并且考虑到现有的艾滋病毒治疗仅影响病毒复制，了解这些机制至关重要。