ABSTRACT
All retroviral genomes contain a nucleotide sequence designated as the primer binding site (PBS) which is complementary to the tRNA used for initiation of reverse transcription. For human immunodeficiency virus type 1 (HIV-1), all naturally occurring genomes have a PBS complementary to tRNA3
Lys. However, within HIV-1 virions, there are approximately equal amounts of tRNA1
Lys, tRNA2
Lys, and tRNA3
Lys. We have used an endogenous reverse transcription-PCR technique specific for the tRNA species within isolated HIV-1 virions to demonstrate that in addition to tRNA3
Lys, tRNA1
Lys and tRNA2
Lys could be used for initiation of HIV-1 reverse transcription. Using a single-round infection assay which employed an HIV-1 genome with a gpt gene encoding xanthine-guanine phosphoribosyl transferase in place of the env gene, we generated cell lines resistant to mycophenolic acid. Analysis of the U5-PBS from single-cell clones revealed PBS complementary to tRNA3
Lys, not tRNA1
Lys or tRNA2
Lys. A mutant HIV-1 genome was then created which would favor the completion of reverse transcription with tRNA1,2
Lys. Using this provirus in the complementation system, we again found only genomes with a PBS complementary to tRNA3
Lys from proviral DNA isolated fromgpt-resistant single-cell colonies. Finally, infection of cells with a mutant HIV genome with a PBS complementary to tRNA1,2
Lys resulted in gpt- resistant cell colonies which contained integrated provirions with a PBS complementary to tRNA1,2
Lys. The results of these studies suggest that the selection of tRNA3
Lys for initiation of HIV-1 reverse transcription occurs both at the initiation and at a postinitiation step in reverse transcription prior to integration of the proviral DNA.
Distribution and Quantification of Human Immunodeficiency Virus Type 1, Strain JC499, Proviral DNA in Tissues from an Infected Chimpanzee
