Aspects of HIV-1 assembly that promote primer tRNALys3 annealing to viral RNA

The major cellular tRNALys isoacceptors are tRNALys1,2 and tRNALys3. During the replication of human immunodeficiency virus 1 (HIV-1), tRNALys3 is used to prime reverse transcription of the viral RNA genome into double-stranded DNA, which is then integrated into the host genome. The annealing of tRNALys3 to 5′-terminal sequences of viral RNA is multi-staged, with an initial poor quality, cytoplasmic annealing promoted by the Gag precursor protein, followed by a more effective annealing imposed upon the Gag-annealed tRNALys3 that occurs after viral protein processing, and that is facilitated by mature nucleocapsid (NCp7). The initial annealing by Gag is assisted by the architecture of an early viral assembly intermediate we term the “tRNALys3 annealing complex” whose composition includes Gag, GagPol, viral RNA, lysyl-tRNA synthetase (LysRS), and the tRNALys isoacceptors. Our model proposes that the reverse transcriptase sequences in GagPol bind all tRNAs non-specifically, and that the cytoplasmic tRNA population to which GagPol is exposed is enriched in tRNALys isoacceptors due to a specific interaction between Gag and LysRS. We further predict a protein conformation within the annealing complex that not only promotes this tRNALys enrichment, but that also facilitates the transfer of tRNALys3 from GagPol to the viral RNA where annealing is carried out by nucleocapsid sequences within Gag.

► During HIV-1 assembly, tRNALys3, the primer for reverse transcription in HIV-1, is selectively concentrated and annealed to the 5′-terminal region of viral RNA. ► Annealing is facilitated by Gag before viral protein processing, and after protein processing, is fine-tuned by re-annealing with mature nucleocapsid. ► A pre-processed assembly intermediate promotes the annealing by Gag.