Absolute dependence on kappa B responsive elements for initiation and Tat-mediated amplification of HIV transcription in blood CD4 T lymphocytes.
The role of NF-kappa B-dependent signals in activating the transcriptional activity of the HIV regulatory region (LTR) was analyzed by systematic comparison of HIV LTR activity in human CD4 T cells purified from peripheral blood and a transformed lymphoblastoid T cell line. In normal CD4 T cells we also analyzed the role played by the viral kappa B responsive elements in HIV replication. Analysis of nuclear extracts of resting, normal T lymphocytes revealed the presence of the p50, but not the p65, NF-kappa B subunit and the induction by phorbol esters of bona fide (p50-p65) NF-kappa B complexes. In parallel, we observed clear enhancer-dependent HIV LTR transactivation comparable in intensity with that observed in lymphoblastoid cells. We show that unstimulated CD4 T lymphocytes offer a cellular environment of very low permissivity to HIV LTR functioning. This was in sharp contrast to the high spontaneous LTR activity observed in lymphoblastoid T cells, where LTR activity was essentially independent of kappa B-responsive elements. Due to the low basal LTR activity in resting T lymphocytes, NF-kappa B-dependent transactivation was a sine qua non event for induction of the HIV LTR. Surprisingly, even the function of HIV Tat in resting CD4 T lymphocytes was found to be absolutely dependent on LTR kappa B responsive elements. The relevance of these observations obtained in transient transfections was confirmed by the incapacity of blood CD4 T lymphocytes infected with an HIV infectious provirus carrying critical point mutations in the kappa B responsive elements to show any detectable transcriptional activity upon cell activation and prolonged culture in vitro.(ABSTRACT TRUNCATED AT 250 WORDS)
Alcamí
J
J
Servicio de Microbiología, Hospital 12 de Octubre, Madrid, Spain.
Laín de Lera
T
T
Folgueira
L
L
Pedraza
M A
MA
Jacqué
J M
JM
Bachelerie
F
F
Noriega
A R
AR
Hay
R T
RT
Harrich
D
D
Gaynor
R B
RB
D001483:Base Sequence; D015496:CD4-Positive T-Lymphocytes / Q000502:physiology / Q000821:virology*; D002461:Cell Line, Transformed; D002478:Cells, Cultured; D004279:DNA, Viral / Q000096:biosynthesis; D004742:Enhancer Elements, Genetic; D015967:Gene Expression Regulation, Viral; D015696:Gene Products, tat / Q000096:biosynthesis / Q000378:metabolism*; D006678:HIV / Q000235:genetics* / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D016328:NF-kappa B / Q000378:metabolism*; D011533:Proviruses / Q000235:genetics / Q000502:physiology; D012333:RNA, Messenger / Q000096:biosynthesis; D012367:RNA, Viral / Q000096:biosynthesis; D015398:Signal Transduction; D013601:T-Lymphocytes / Q000502:physiology / Q000821:virology*; D014158:Transcription, Genetic; D014162:Transfection; D054322:tat Gene Products, Human Immunodeficiency Virus
D004279:DNA, Viral; D015696:Gene Products, tat; D016328:NF-kappa B; D012333:RNA, Messenger; D012367:RNA, Viral; D054322:tat Gene Products, Human Immunodeficiency Virus