Probing the HIV-1 Genomic RNA Trafficking Pathway and Dimerization by Genetic Recombination and Single Virion Analyses

Once transcribed, the nascent full-length RNA of HIV-1 must travel to the appropriate host cell sites to be translated or to find a partner RNA for copackaging to form newly generated viruses. In this report, we sought to delineate the location where HIV-1 RNA initiates dimerization and the influence of the RNA transport pathway used by the virus on downstream events essential to viral replication. Using a cell-fusion-dependent recombination assay, we demonstrate that the two RNAs destined for copackaging into the same virion select each other mostly within the cytoplasm. Moreover, by manipulating the RNA export element in the viral genome, we show that the export pathway taken is important for the ability of RNA molecules derived from two viruses to interact and be copackaged. These results further illustrate that at the point of dimerization the two main cellular export pathways are partially distinct. Lastly, by providing Gag in trans, we have demonstrated that Gag is able to package RNA from either export pathway, irrespective of the transport pathway used by the gag mRNA. These findings provide unique insights into the process of RNA export in general, and more specifically, of HIV-1 genomic RNA trafficking.

经转录生成后，人类免疫缺陷病毒1型（HIV-1）的新生全长核糖核酸（RNA）需转运至相应的宿主细胞位点，以进行翻译，或寻找配对RNA完成共包装，进而形成新生病毒颗粒。本研究旨在明确HIV-1 RNA启动二聚化的位点，以及病毒所采用的RNA转运通路对病毒复制必需的下游事件的影响。本研究采用细胞融合依赖重组检测法，证实拟被共包装至同一病毒粒子中的两条RNA，其相互识别主要发生在细胞质内。此外，通过操控病毒基因组中的RNA输出元件，本研究显示病毒所选用的输出通路，对于两种病毒来源的RNA分子相互作用并完成共包装至关重要。上述结果进一步表明，在RNA二聚化阶段，宿主细胞的两条主要RNA输出通路存在部分差异。最后，通过反式提供Gag蛋白（Gag），本研究证实Gag蛋白可包装任意一条输出通路来源的RNA，且不受gag mRNA所采用的转运通路影响。本研究结果为通用的RNA输出过程，特别是HIV-1基因组RNA的转运过程，提供了独特的见解。