INCORPORATION OF EXCESS WILD-TYPE AND MUTANT TRNA(3)(LYS) INTO HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1

Human immunodeficiency virus (HIV) particles produced in COS-7 cells transfected with HIV type 1 (HIV-1) proviral DNA contain 8 molecules of tRNA(3)(Lys) per 2 molecules of genomic RNA and 12 molecules of tRNA(1,2)(Lys) per 2 molecules of genomic RNA. When COS-7 cells are transfected with a plasmid containing both HIV-1 proviral DNA and a human tRNA(3)(Lys) gene, there is a large increase in the amount of cytoplasmic tRNA(3)(Lys) per microgram of total cellular RNA, and the tRNA(3)(Lys) content in the virus increases from 8 to 17 molecules per 2 molecules of genomic RNA. However, the total number of tRNA(Lys) molecules per 2 molecules of genomic RNA remains constant at 20; i.e., the viral tRNA(1,2)(Lys); content decreases from 12 to 3 molecules per 2 molecules of genomic RNA. All detectable tRNA(3)(Lys) is aminoacylated in the cytoplasm of infected cells and deacylated in the virus. When COS-7 cells are transfected with a plasmid containing both HIV-1 proviral DNA and a mutant amber suppressor tRNA(3)(Lys) gene (in which the anticodon is changed from TTT to CTA), there is also a large increase in the relative concentration of cytoplasmic tRNA(3)(Lys), and the tRNA(3)(Lys) content in the virus increases from 8 to 15 molecules per 2 molecules of genomic RNA, with a decrease in viral tRNA(1,2)(Lys) from 12 to 5 molecules per 2 molecules of genomic RNA. Thus, the total number of molecules of tRNA(Lys) in the virion remains at 20. The alteration of the anticodon has little effect on the viral packaging of this mutant tRNA in spite of the fact that it no longer contains the modified base mcm (5)s(2)U at position 34 and its ability to be aminoacylated is significantly impaired compared with that of wild-type tRNA(3)(Lys). Viral particles which have incorporated either excess wild-type tRNA(3)(Lys) or mutant suppressor tRNA(3)(Lys) show no differences in viral infectivity compared with wild-type HIV-1.