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Mechanisms that determine subcellular sites of HIV-1 assembly

决定 HIV-1 组装亚细胞位点的机制

基本信息

批准号：
10617799
负责人：
Akira Ono
金额：
$ 68.28万
依托单位：
UNIVERSITY OF MICHIGAN AT ANN ARBOR
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-07-01 至 2027-06-30
项目状态：
未结题

项目摘要

Virus particle assembly of HIV-1, the causative agent of AIDS, takes place at the plasma membrane (PM) in most cell types including natural host T cells. This process is driven by a viral structural protein Gag. The N-terminal matrix (MA) domain of Gag determines Gag localization to and hence virus assembly at the PM. MA mediates membrane binding of Gag via N-terminal myristoyl moiety and a highly basic region (HBR) that binds acidic lipids. Binding of HBR to a PM-specific acidic phospholipid PI(4,5)P2 is critical for PM localization of Gag and efficient virus release. Notably, we and others showed that MA HBR also interacts with tRNAs, which suppress binding of Gag to non-PI(4,5)P2 acidic lipids, suggesting tRNAs as a host factor that regulates MA-membrane interactions. However, structural determinants for the tRNA-MA HBR interaction and its reversal by the interaction with PI(4,5)P2, combination of which regulates PM-specific Gag localization, remain to be examined. Binding of tRNAs to MA HBR is most likely to occur at translation sites due to limited availability of tRNAs outside of the translation machinery. However, little is known about subcellular sites of Gag translation, where Gag begins its movement to the PM. At the PM, Gag multimerization and subsequent accumulation of acidic lipids are likely to promote recruitment of host transmembrane proteins, but their effects on virus spread to uninfected cells remains to be determined in the context of cell-free and cell-to-cell transmission. Our long-term goal is to elucidate mechanisms that determine sites of HIV-1 assembly and to use the knowledge for developing antiviral strategies. Our central hypothesis in this application is that MA HBR interactions with tRNAs, which begin during translation, and with acidic lipids determine subcellular Gag localization and the properties of progeny virions. To test this hypothesis, we plan to: 1) identify structural determinants for interactions of MA HBR with tRNAs and acidic lipids; 2) identify tRNAs that suppress PI(4,5)P2-independent membrane binding but allow PI(4,5)P2-mediated reversal; 3) understand the effects of Gag translation sites on the fate of Gag; and 4) examine the effects of host transmembrane proteins incorporated into virus particles on formation of virological synapse and virus-cell contact. The knowledge gained from experiments outlined in this proposal will likely help us develop antiviral strategies that target mechanisms regulating Gag localization to the PM, thereby inhibiting extracellular virus release and spread. RELEVANCE (See instructions): Proper localization of viral components in cells is critical for efficient production of virus particles and spread from infected to uninfected cells. The goal of the proposed research is to elucidate the mechanisms that direct Gag, a structural protein of HIV-1 that causes AIDS, to the proper sites within virus-expressing cells. Knowledge obtained from proposed studies will help us develop novel antiviral strategies that may either directly suppress assembly and release of HIV-1 particles or inhibit spread of this virus to uninfected host cells

艾滋病的病原体HIV-1的病毒颗粒组装发生在质膜（PM）上在大多数细胞类型中，包括天然宿主T细胞。这个过程是由病毒结构蛋白Gag驱动的。的 Gag的N-末端基质（MA）结构域决定Gag定位于PM并因此决定病毒组装于PM。 MA通过N-末端肉豆蔻酰基部分和高碱性区域（HBR）介导Gag的膜结合与酸性脂质结合HBR与PM特异性酸性磷脂PI（4，5）P2的结合对于PM至关重要 Gag的定位和有效的病毒释放。值得注意的是，我们和其他人表明，MA HBR也相互作用与抑制Gag与非PI（4，5）P2酸性脂质结合的tRNA，表明tRNA作为宿主调节MA-膜相互作用的因子。然而，tRNA-MA HBR的结构决定因素通过与PI（4，5）P2的相互作用及其逆转，其组合调节PM特异性窒息定位，有待检查。tRNA与MA HBR的结合最有可能发生在翻译时由于翻译机器之外的tRNA的可用性有限，因此这些位点不存在。然而，人们对 Gag翻译的亚细胞位点，其中Gag开始向PM移动。下午，加格酸性脂质的多聚化和随后的积累可能促进宿主的募集跨膜蛋白，但它们对病毒传播到未感染细胞的影响仍有待确定，无细胞和细胞到细胞传输的环境。我们的长期目标是阐明决定HIV-1组装位点的机制，并使用开发抗病毒策略的知识。我们在这个应用中的中心假设是，MA HBR 在翻译过程中开始与tRNA的相互作用，以及与酸性脂质的相互作用决定了亚细胞Gag 定位和子代病毒体的性质。为了验证这一假设，我们计划：1）确定结构 MA HBR与tRNA和酸性脂质相互作用的决定因素; 2）鉴定抑制 PI（4，5）P2独立膜结合，但允许PI（4，5）P2介导的逆转; 3）了解影响 Gag翻译位点对Gag命运的影响;以及4）检查宿主跨膜蛋白的影响在形成病毒突触和病毒-细胞接触时掺入病毒颗粒。知识从这项提议中概述的实验中获得的信息可能有助于我们开发针对调节Gag定位于PM的机制，从而抑制细胞外病毒释放，传播. 相关性（参见说明）：病毒组分在细胞中的适当定位对于病毒颗粒的有效产生和从受感染的细胞传播到未受感染的细胞。这项研究的目的是阐明它将导致艾滋病的HIV-1的结构蛋白Gag引导到病毒表达细胞内的适当位点，细胞从拟议的研究中获得的知识将有助于我们开发新的抗病毒策略，直接抑制HIV-1颗粒的组装和释放，或抑制这种病毒向未感染者的传播宿主细胞