Permanent occupancy of the human immunodeficiency virus type 1 enhancer by NF-kappa B is needed for persistent viral replication in monocytes.
This work aimed to ascertain the role of kappaB-responsive elements of the human immunodeficiency virus type 1 (HIV-1) enhancer not only in early initiation but also in long-term maintenance of proviral transcription in cells of the monocytic lineage. For this purpose, we used three main approaches. The first was to abruptly terminate tumor necrosis factor-induced NF-kappaB binding to the enhancer sequences in U1 monocytic cells, using a short pulse of exogenous tumor necrosis factor. This resulted in concomitant decrease in nuclear NF-kappaB DNA-binding activity and endogenous long terminal repeat transcriptional activity. The second was to suppress the permanent NF-kappaB translocation induced by HIV-1 replication itself in chronically infected U937 cells, using a specific proteasome inhibitor (Z-LLL-H). As early as 2 h after addition of the inhibitor to the culture medium, there was an inhibition of both constitutive activation of NF-kappaB and HIV-1 genome expression. The third approach was to monitor the replication competence in U937 cells of an infectious HIV-1 provirus carrying point mutations in the kappaB-responsive elements of both long terminal repeats. Compared with its wild-type counterpart, this mutated provirus showed a profoundly decreased, Z-LLL-H-insensitive transcriptional and replicative activity in U937 monocytes. Together, our results indicate that occupancy of the viral enhancer by NF-kappaB (p50/p65) heterodimers is required for ongoing transcription of integrated HIV provirus in monocytes, even in cells chronically infected and permanently producing functional HIV Tat protein. Thus, the ability of HIV-1 replication to activate NF-kappaB is crucial to the intense self-perpetuated viral transcription observed in cells of the monocytic lineage.
Jacqué
J M
JM
Unité d'Immunologie Virale, Institut Pasteur, Paris, France.
Fernández
B
B
Arenzana-Seisdedos
F
F
Thomas
D
D
Baleux
F
F
Virelizier
J L
JL
Bachelerie
F
F
D001483:Base Sequence; D001665:Binding Sites; D002460:Cell Line; D002467:Cell Nucleus / Q000502:physiology; D003546:Cysteine Endopeptidases / Q000378:metabolism; D003593:Cytoplasm / Q000502:physiology; D017931:DNA Primers; D004742:Enhancer Elements, Genetic; D016325:HIV Long Terminal Repeat; D015497:HIV-1 / Q000235:genetics / Q000502:physiology*; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D009000:Monocytes / Q000502:physiology / Q000821:virology*; D009097:Multienzyme Complexes / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism*; D009838:Oligodeoxyribonucleotides; D016376:Oligonucleotides, Antisense; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D046988:Proteasome Endopeptidase Complex; D011533:Proviruses / Q000502:physiology; D014158:Transcription, Genetic; D014779:Virus Replication
D017931:DNA Primers; D009097:Multienzyme Complexes; D016328:NF-kappa B; D009838:Oligodeoxyribonucleotides; D016376:Oligonucleotides, Antisense; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex