Mechanisms that determine subcellular sites of HIV-1 assembly

艾滋病的病原体HIV-1的病毒颗粒组装发生在质膜上 在大多数细胞类型中，包括自然宿主T细胞。这一过程是由病毒结构蛋白Gag驱动的。这个 Gag的N-末端矩阵(MA)结构域决定了Gag对PM的定位，从而决定了病毒在PM的组装 MA通过N-末端肉豆蔻基部分和高碱性区域(HBr)介导GAG的膜结合 结合酸性脂类的物质。HBr与质膜特异性酸性磷脂PI(4，5)P2的结合是质膜的关键 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tRNAs结合，抑制Gag与非PI(4，5)P2酸性脂类的结合，提示tRNAs是宿主 调节MA-膜相互作用的因子。然而，tRNA-MA HBR的结构决定因素 与PI(4，5)P2的相互作用及其逆转 GAG的本地化，还有待研究。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的可获得性有限，这些站点的数量很少。然而，人们对此知之甚少。 GAG翻译的亚细胞位置，在那里GAG开始向PM移动在首相面前，恶作剧 多聚体和随后的酸性脂积累可能会促进宿主的招募 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 无细胞和细胞间传输的环境。 我们的长期目标是阐明确定HIV-1组装位点的机制，并使用 开发抗病毒策略的知识。我们在此应用程序中的中心假设是MA HBR 在翻译过程中与tRNAs的相互作用，以及与酸性脂质的相互作用，决定了亚细胞Gag 子代病毒粒子的定位和性质。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)识别抑制 PI(4，5)P2不依赖膜结合，但允许PI(4，5)P2介导的逆转；3)了解其影响 以及4)检测宿主跨膜蛋白的影响。 在病毒颗粒中形成病毒性突触和病毒与细胞的接触。《知识》 从本提案中概述的实验中获得的信息可能有助于我们开发针对以下目标的抗病毒策略 调节GAG定位到PM的机制，从而抑制细胞外病毒的释放和 散开。 相关性(请参阅说明)： 病毒成分在细胞中的正确定位对于有效生产病毒颗粒和 从感染的细胞传播到未感染的细胞。这项拟议研究的目标是阐明这种机制。 该GAG是HIV-1的一种结构蛋白，可导致艾滋病，并将其定向到病毒表达的适当位置 细胞。从拟议的研究中获得的知识将帮助我们开发新的抗病毒策略，这些策略可能 要么直接抑制HIV-1颗粒的组装和释放，要么阻止该病毒向未感染的人传播 宿主细胞

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