GAG Protein/RNA Interactions in the HIV Lifecycle

GAG蛋白是HIV-1病毒体核心的主要成分，它以多种对病毒生命周期至关重要的方式与病毒RNA相互作用。 与HIV-1基因组中的5'包装（或psi）区域的特异性Gag结合对于将该RNA靶向到新生病毒颗粒中是必不可少的，并且有助于启动基因组二聚体的形成。 Gag/RNA相互作用还促进tRNA引发、逆转录、病毒体组装和其他关键过程，因此为新的抗病毒疗法提供了特别有希望的靶点。 我们的目标是在体外解剖Gag与HIV-1 RNA的各种分子相互作用，并评估它们在活病毒中的遗传功能。 我们以前的特点是在psi位点的GAG结合和RNA二聚体接触的网站。 我们现在建议详细探讨这些位点之间的功能相互作用，绘制其他网站相信位于上游，并寻找其他基因组中的其他地方，可能有助于病毒体组装和成熟。 我们将试图描绘所有的顺式作用信号所需的HIV-1包装在体内。 我们早期的研究揭示了一类新的HIV-1突变体，它们在RNA二聚化中具有选择性缺陷，与显著降低的感染性相关。 通过分析这些突变体，我们将评估基因组二聚化对包装，逆转录，序列重组，RNA稳定性和HIV-1核心内基因组构象的精确贡献。 计划在小鼠白血病病毒和人端粒酶复合物中进行比较研究。 我们还确定了一种异源RNA配体，结合HIV-1 Gag具有高亲和力，并在体内具有包装活性，我们现在建议研究这种配体是否可以作为一个有效的竞争性抑制剂Gag/psi结合和HIV-1复制。 总之，这些研究将阐明特定的Gag/RNA相互作用是脆弱的药物靶点，并将提供新的体外试验，利用这些靶点在抗病毒药物的发现。

GAG protein is the major constituent of the HIV-1 virion core, and it interacts with viral RNA in a number of ways that are critical for the viral lifecycle. Specific Gag binding to the 5' packaging (or psi) region in the HIV-1 genome is essential for targeting this RNA into nascent viral particles, and helps initiate formation of the genomic dimer. Gag/RNA interactions also promote tRNA priming, reverse transcription, virion assembly, and other crucial processes, and so offer an especially promising target for new antiviral therapies. Our goal is to dissect the various molecular interactions of Gag with HIV-1 RNA in vitro, and to evaluate their functions genetically in the live virus. We have previously characterized sites of Gag-binding and RNA dimer contact in the psi locus. We now propose to explore the functional interactions among those sites in detail, to map additional sites believed to lie upstream, and to search for others elsewhere in the genome that may contribute to virion assembly and maturation. We will attempt to delineate all of the cis-acting signals required for HIV-1 packaging in vivo. Our earlier studies revealed a new class of HIV-1 mutants that have a selective defect in RNA dimerization, associated with markedly reduced infectivity. By analyzing those mutants, we will evaluate the precise contributions of genomic dimerization to packaging, reverse transcription, sequence recombination, RNA stability, and the conformation of the genome within the HIV-1 core. Comparative studies in murine leukemia virus and the human telomerase complex are planned. We have also identified a heterologous RNA ligand that binds HIV-1 Gag with high affinity and has packaging activity in vivo; we now propose to investigate whether this ligand can act as an effective competitive inhibitor of Gag/psi binding and of HIV-1 replication. Together, these studies will illuminate particular Gag/RNA interactions that are vulnerable drug targets, and will provide novel in vitro assays for exploiting those targets in antiviral drug discovery.