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：
10203782
负责人：
Akira Ono
金额：
$ 53.78万
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：
2007
资助国家：
美国
起止时间：
2007-02-01 至 2022-06-30
项目状态：
已结题

项目摘要

Title: Mechanisms that determine subcellular sites of HIV-1 assembly Summary/Abstract: 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, in vitro and cell-based studies 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 new host factor that regulates MA-membrane interactions. However, molecular 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 elucidated. Moreover, how these interactions begin and what effect these interactions have on the property of progeny virions at the end are poorly understood. 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 is likely to create accumulation of acidic lipids, but its impact on the ability of progeny virions to spread to uninfected cells remains unknown. 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 molecular determinants for interaction of MA HBR with tRNAs and its reversal by PI(4,5)P2; 2) identify tRNAs that suppress PI(4,5)P2-independent membrane binding but allow PI(4,5)P2-mediated reversal; 3) examine the possibility that Gag associates with tRNA during translation; and 4) examine the effect of acidic lipid accumulation at virus assembly sites on the virion properties. 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.

标题：决定 HIV-1 组装亚细胞位点的机制摘要/摘要： HIV-1（艾滋病的病原体）的病毒颗粒组装发生在细胞质膜 (PM) 上大多数细胞类型，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。然后- Gag 的末端矩阵 (MA) 结构域决定 Gag 的定位，从而决定病毒在 PM 的组装。硕士通过 N 端肉豆蔻酰部分和结合的高碱性区域 (HBR) 介导 Gag 的膜结合酸性脂质。 HBR 与 PM 特异性酸性磷脂 PI(4,5)P2 的结合对于 Gag 的 PM 定位至关重要和高效的病毒释放。值得注意的是，体外和细胞研究表明 MA HBR 还与 tRNA，抑制 Gag 与非 PI(4,5)P2 酸性脂质的结合，表明 tRNA 作为新的宿主因子调节 MA 膜相互作用。然而，tRNA-MA HBR 相互作用的分子决定因素及其通过与 PI(4,5)P2 相互作用的逆转，两者的组合调节 PM 特异性 Gag 定位，仍有待阐明。此外，这些相互作用如何开始以及这些相互作用对对于子代病毒体最终的特性知之甚少。 tRNA 与 MA HBR 的结合最有可能由于翻译机器之外 tRNA 的可用性有限，因此发生在翻译位点。然而，很少众所周知，Gag 翻译的亚细胞位点是 Gag 开始向 PM 移动的地方。下午，堵嘴多聚化可能会造成酸性脂质的积累，但它会影响子代病毒颗粒的能力传播到未感染的细胞仍然未知。我们的长期目标是阐明决定 HIV-1 组装位点的机制并利用制定抗病毒策略的知识。我们在此应用中的中心假设是 MA HBR 与 tRNA 的相互作用（在翻译过程中开始）以及与酸性脂质的相互作用决定了亚细胞 Gag 子代病毒颗粒的定位和特性。为了检验这个假设，我们计划：1）识别分子 MA HBR 与 tRNA 相互作用及其被 PI(4,5)P2 逆转的决定因素； 2) 识别 tRNA 抑制 PI(4,5)P2 独立的膜结合，但允许 PI(4,5)P2 介导的逆转； 3）检查 Gag 在翻译过程中与 tRNA 结合的可能性； 4) 检查酸性脂质的影响病毒组装位点的积累对病毒粒子特性的影响。从实验中获得的知识概述该提案中的内容可能会帮助我们制定针对 Gag 调节机制的抗病毒策略定位到PM，从而抑制细胞外病毒的释放和传播。