Mechanism of HIV-1 gRNA-Gag interaction leading to selective genome packaging

 描述（由申请人提供）：人类免疫缺陷病毒1型（HIV-1）RNA转录物用于翻译病毒蛋白，也可选择性地掺入nascen病毒体中作为遗传物质。HIV-1能够包装二聚体全长基因组RNA（gRNA）的单个拷贝，确保其遗传信息的正确传播。尽管进行了广泛的研究，但对HIV-1选择性掺入gRNA的机制仍然知之甚少。病毒Gag蛋白负责通过与其5 ′非翻译区（5 ′ UTR）的相互作用包装gRNA，特别是Psi包装信号。已知5 'UTR采用多种构象并二聚化，并且这已被提出影响gRNA包装。Gag还显示出采用多种构象，取决于磷脂和/或核酸的存在，在弯曲构象和直构象之间波动。目前解释基因组包装的模型是Gag的核衣壳（NC）结构域识别gRNA并通过Gag基质（MA）结构域将其靶向质膜（PM），使进一步的Gag寡聚化成核并导致病毒颗粒形成。一些Gag分子可能以仅NC构象与gRNA相互作用以允许MA结合PM。然而，缺乏对导致RNA包装的Gag构象和特异性gRNA相互作用的清楚理解。我假设，含Psi的二聚体gRNA以组装能力模式促进Gag相互作用，导致其优先掺入病毒体。以下实验将阐明可以改变Gag的结合模式和构象的RNA序列和构象，以确定Gag如何以导致其选择性结合的方式与这些RNA相互作用。该提议试图回答以下悬而未决的问题：1）在特异性结合模式中Gag相互作用需要什么gRNA序列和构象？2)gRNA序列和构象如何影响Gag构象平衡？3)这些相同的gRNA序列和构象是否会导致感染期间的选择性gRNA包装？阐明Gag成功靶向和包装其gRNA所采用的分子机制将有助于未来利用新型抗逆转录病毒疗法靶向这种相互作用的努力。

 DESCRIPTION (provided by applicant): Human immunodeficiency virus type-1 (HIV-1) RNA transcripts are used to translate viral proteins, and are also selectively incorporated into nascen virions as the genetic material. HIV-1 is capable of packaging a single copy of dimeric, full-length genomic RNA (gRNA), ensuring correct propagation of its genetic information. Despite extensive study, the mechanism governing the selective incorporation of gRNA by HIV-1 remains poorly understood. The viral Gag protein is responsible for packaging gRNA via interactions with its 5´ untranslated region (5´UTR), specifically the Psi packaging signal. The 5´UTR is known to adopt multiple conformations and dimerize, and this has been proposed to affect gRNA packaging. Gag has also been shown to adopt multiple conformations, fluctuating between bent and straight conformers depending on the presence of phospholipids and/or nucleic acids. The current model explaining genome packaging is that the nucleocapsid (NC) domain of Gag recognizes gRNA and targets it to the plasma membrane (PM) via the Gag matrix (MA) domain, nucleating further Gag oligomerization and leading to virus particle formation. A few Gag molecules likely interact with gRNA in an NC-only conformation to allow MA to bind the PM. However, a clear understanding of Gag conformation and specific gRNA interactions that lead to RNA packaging is lacking. I hypothesize that Psi-containing, dimeric gRNAs promote Gag interaction in an assembly-competent mode, leading to their preferential incorporation into virions. The following experiments will elucidate the RNA sequence and conformations that can alter Gag's binding mode and conformation, in order to establish how Gag interacts with these RNAs in a manner that leads to their selective encapsidation. This proposal seeks to answer the following outstanding questions: 1) What gRNA sequence and conformation are required for Gag interactions in the specific binding mode? 2) How does gRNA sequence and conformation affect the Gag conformational equilibrium? 3) Do these same gRNA sequences and conformations lead to selective gRNA packaging during infection? Elucidation of the molecular mechanism employed by Gag to successfully target and package its gRNA will facilitate future efforts at targeting this interaction with novel antiretroviral therapies.