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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：
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：确定HIV-1组装亚细胞位点的机制总结/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。N- Gag的末端基质（MA）结构域决定Gag定位于PM并因此决定病毒组装于PM。马通过N-末端肉豆蔻酰基部分和结合Gag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，体外和基于细胞的研究表明，MA HBR也与 tRNA抑制Gag与非PI（4，5）P2酸性脂质的结合，表明tRNA是一种新的宿主因子调节MA-膜相互作用。然而，tRNA-MA HBR相互作用的分子决定因素以及通过与PI（4，5）P2相互作用的逆转，其组合调节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末端的子代病毒体的性质知之甚少。tRNA与MA HBR的结合最有可能由于翻译机器外的tRNA的可用性有限，因此在翻译位点发生。不过小已知Gag翻译的亚细胞位点，其中Gag开始向PM移动。下午，加格多聚化可能会产生酸性脂质的积累，但其对子代病毒粒子的能力的影响，扩散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）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）鉴定抑制PI（4，5）P2非依赖性膜结合，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;以及4）检查酸性脂质的影响在病毒装配位点的积累对病毒体性质的影响。从实验中获得的知识概述了这项提议可能会帮助我们开发针对调节Gag机制的抗病毒策略，定位于PM，从而抑制细胞外病毒释放和传播。