GENETIC-ANALYSIS OF THE U5-PBS OF A NOVEL HIV-1 REVEALS MULTIPLE INTERACTIONS BETWEEN THE TRANSFER-RNA AND RNA GENOME REQUIRED FOR INITIATION OF REVERSE TRANSCRIPTION

A novel HIV-1 genome that stably utilizes tRNA(His) rather than tRNA(L ys,3) to initiate reverse transcription was used to study features for the interaction between the tRNA and viral RNA genome, In addition to a primer binding site (PBS) complementary to tRNA(His), this virus co ntains a six-nucleotide sequence in U5 complementary to the anticodon- loop of tRNA(His) and three additional substitutions: U-174-to-G, G(18 1)-to-A, and U-200-to-C [HXB2(His-AC-GAC)]. Mutations in these three n ucleotides resulted in viruses with three different genotypes: one gro up maintained a PBS complementary to tRNA(His) with restored G(174)A(1 81)C(200) or G(174)A(181)U(200) configurations, one group reverted to a Pas complementary to tRNA(Lys,3) and one group contained two or more PBSs complementary to different tRNAs on the same viral genome, Chara cterization of a previously identified virus with additional C-152-to- A and C-160-to-U substitutions [HXB2(His-AC-A(152)U(160)-GAC)] reveale d that this virus maintained a PBS complementary to tRNA(His), whereas a mutant HXB2(His-AC-U(152)A(160)-GAC) reverted after culture to cont ain dual PBS complementary to tRNA(Lys,3) and tRNA(His), respectively. Our results demonstrate that regions in U5 act in concert with the PB S to promote use of the tRNA primer for initiation of reverse transcri ption, These results are discussed with respect to structural models f or the U5-PBS interactions with tRNA.