Laboratory of Bioimaging and Pathologies (LBP), UMR 7021, Faculty of pharmacy, University of Strasbourg, 67400 Illkirch, France.
Expression génétique microbienne, UMR 8261, CNRS, Université de Paris, Institut de biologie physico-chimique, 13 rue Pierre et Marie Curie, 75005 Paris, France.
Nucleic Acids Res. 2020 Sep 18;48(16):9218-9234. doi: 10.1093/nar/gkaa659.
The HIV-1 Gag protein playing a key role in HIV-1 viral assembly has recently been shown to interact through its nucleocapsid domain with the ribosomal protein L7 (RPL7) that acts as a cellular co-factor promoting Gag's nucleic acid (NA) chaperone activity. To further understand how the two proteins act together, we examined their mechanism individually and in concert to promote the annealing between dTAR, the DNA version of the viral transactivation element and its complementary cTAR sequence, taken as model HIV-1 sequences. Gag alone or complexed with RPL7 was found to act as a NA chaperone that destabilizes cTAR stem-loop and promotes its annealing with dTAR through the stem ends via a two-step pathway. In contrast, RPL7 alone acts as a NA annealer that through its NA aggregating properties promotes cTAR/dTAR annealing via two parallel pathways. Remarkably, in contrast to the isolated proteins, their complex promoted efficiently the annealing of cTAR with highly stable dTAR mutants. This was confirmed by the RPL7-promoted boost of the physiologically relevant Gag-chaperoned annealing of (+)PBS RNA to the highly stable tRNALys3 primer, favoring the notion that Gag recruits RPL7 to overcome major roadblocks in viral assembly.
HIV-1 Gag 蛋白在 HIV-1 病毒组装中发挥关键作用，最近的研究表明，它通过核衣壳结构域与核糖体蛋白 L7（RPL7）相互作用，RPL7 作为一种细胞辅助因子，促进 Gag 的核酸（NA）伴侣活性。为了进一步了解这两种蛋白质如何共同作用，我们分别研究了它们的作用机制以及它们在促进 dTAR（病毒转录激活元件的 DNA 版本）与其互补 cTAR 序列之间退火中的协同作用，以 HIV-1 序列为模型。研究发现，单独的 Gag 或与 RPL7 形成复合物都可以作为 NA 伴侣，通过两步途径破坏 cTAR 茎环结构，并通过茎端促进其与 dTAR 的退火。相比之下，单独的 RPL7 作为 NA 退火酶，通过其 NA 聚集特性，通过两条平行途径促进 cTAR/dTAR 退火。值得注意的是，与分离的蛋白质相比，它们的复合物有效地促进了高度稳定的 dTAR 突变体与 cTAR 的退火。这一点通过 RPL7 促进生理相关的 Gag 伴侣的 (+)PBS RNA 与高度稳定的 tRNALys3 引物的退火得到了证实，这表明 Gag 招募 RPL7 来克服病毒组装中的主要障碍。
