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  • Title: In virio SHAPE analysis of tRNA(Lys3) annealing to HIV-1 genomic RNA in wild type and protease-deficient virus.
    Author: Seif E, Niu M, Kleiman L.
    Journal: Retrovirology; 2015 May 16; 12():40. PubMed ID: 25981241.
    Abstract:
    BACKGROUND: tRNA(Lys3) annealing to the viral RNA of human immunodeficiency virus type-1 (HIV-1) is an essential step in the virus life cycle, because this tRNA serves as the primer for initiating reverse transcription. tRNA(Lys3) annealing to viral RNA occurs in two steps. First, Gag promotes annealing of tRNA(Lys3) to the viral RNA during cytoplasmic HIV-1 assembly. Second, mature nucleocapsid (NCp7), produced from the processing of Gag by viral protease during viral budding from the cell, remodels the annealed complex to form a more stable interaction between the viral RNA and tRNA(Lys3), resulting in a more tightly bound and efficient primer for reverse transcription. RESULTS: In this report, we have used in virio SHAPE analysis of both the 5´-untranslated region in HIV-1 RNA and the annealed tRNA(Lys3) to determine structural differences of the annealed complex that occur between protease-negative (Pr-) and wild type viruses. Our results indicate that the weaker binding of tRNA(Lys3) annealed by Gag in Pr- virions reflects both missing interactions of tRNA(Lys3) with viral RNA regions in the upper PBS stem, and a weaker interaction with the internal stem-loop found within the unannealed primer binding site in viral RNA. CONCLUSIONS: We propose secondary structure models for the tRNA(Lys3)/viral RNA annealed complexes in PR- and wild type viruses that support the two-step annealing model by showing that Gag promotes a partial annealing of tRNA(Lys3) to HIV-1 viral RNA, followed by a more complete annealing by NCp7.
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