THE KAPPA-B SITES IN THE HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 LONG TERMINAL REPEAT ENHANCE VIRUS-REPLICATION YET ARE NOT ABSOLUTELY REQUIREDFOR VIRAL GROWTH

The dependence of human immunodeficiency virus type 1 (HIV-1) on its N F-kappa B binding sites (kappa B sites) for replication in transformed and primary T-cell targets was examined by infecting cells with HIV-1 reporter viruses containing kappa B Site enhancer mutations, Viral tr anscription was measured either with luciferase-expressing HIV-1 that infects for a single round off by flow cytometric analyses with HIV-1 expressing placental alkaline phosphatase (FLAP) or green-fluorescent] protein (GFP). Both PLAP- and GFP-expressing viruses spread from cell to cell and allowed analysis of viral gene expression patterns in sing le cells, Infection of a panel of T-cell lines with different basal le vels of NF-kappa B demonstrated a direct correlation between the amoun t of constitutive nuclear NF-kappa B and the degree to which a wild-ty pe virus outperformed kappa B Site mutants, One T-cell line with a con stitutively high level of nuclear NF-kappa B, PM1, showed a 20-fold de crease in transcription when its kappa B sites were mutated, In contra st, in a T-cell line with a low basal level of NF-kappa B, SupT1, muta tion of the kappa B Site in the enhancer had no effect on viral transc ription or growth rate, Phytohemagglutinin-activated peripheral blood mononuclear cells showed a large dependence on the kappa B sites for o ptimal virus growth. Viruses without marker genes corroborated the fin ding that mutations to the kappa B sites impair virus production in ce lls with a high basal level of NF-kappa B, These data show that in T c ells, HIV-1 can use NF-kappa B to enhance its growth but the virus is clearly able to grow in its absence.