ABSOLUTE DEPENDENCE ON KAPPA-B RESPONSIVE ELEMENTS FOR INITIATION ANDTAT-MEDIATED AMPLIFICATION OF HIV TRANSCRIPTION IN BLOOD CD4 T-LYMPHOCYTES

The role of NF-kappa B-dependent signals in activating the transcripti onal activity of the HIV regulatory region (LTR) was analyzed by syste matic comparison of HIV LTR activity in human CD4 T cells purified fro m peripheral blood and a transformed lymphoblastoid T cell line. In no rmal 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 o f bona fide (p50-p65) NF-kappa B complexes. In parallel, we observed c lear enhancer-dependent HIV LTR transactivation comparable in intensit y with that observed in lymphoblastoid cells. We show that unstimulate d CD4 T lymphocytes offer a cellular environment of very low permissiv ity to HIV LTR functioning. This was in sharp contrast to the high spo ntaneous LTR activity observed in lymphoblastoid T cells, where LTR ac tivity was essentially independent of kappa B responsive elements. Due to the low basal LTR activity in resting T lymphocytes, NF-kappa B-de pendent transactivation was a sine qua non event for induction of the HIV LTR. Surprisingly, even the function of HIV Tat in resting CD4 T l ymphocytes was found to be absolutely dependent on LTR kappa B respons ive elements. The relevance of these observations obtained in transien t transfections was confirmed by the incapacity of blood CD4 T lymphoc ytes infected with an HIV infectious provirus carrying critical point mutations in the kappa B responsive elements to show any detectable tr anscriptional activity upon cell activation and prolonged culture in v itro. Our observations emphasize the importance of analyzing the funct ioning of HIV regulatory domains in the natural environment provided b y normal CD4 T lymphocytes for HIV infection, and demonstrate an absol ute requirement for NF-kappa B responsive elements for Tat-dependent a nd Tat-independent HIV transcription in blood CD4 T lymphocytes.