The expression of the interleukin 6 gene is induced by the human immunodeficiency virus 1 TAT protein.
Human immunodeficiency virus 1 (HIV1) infection is associated with severe psoriasis, B cell lymphoma, and Kaposi's sarcoma. A deregulated production of interleukin 6 (IL-6) has been implicated in the pathogenesis of these diseases. The molecular mechanisms underlying the abnormal IL-6 secretion of HIV1-infected cells may include transactivation of the IL-6 gene by HIV1. To test this hypothesis, we used the pIL6Pr-chloramphenicol acetyltransferase (CAT) plasmid, an IL-6 promoter-CAT construct, as a target of the transactivating function of the HIV1 TAT protein. By cotransfecting the pIL6Pr-CAT and the tat-expressing pSVT8 plasmid in MC3 B-lymphoblastoid or in HeLa epithelial cells, we observed that TAT transactivates the human IL-6 promoter. These results were confirmed when pIL6Pr-CAT was transfected in MC3 or HeLa cells that constitutively expressed the tat gene in a sense (pSVT8 cells) or antisense (pSVT10 cells) orientation. 5' deletion plasmids of pIL6Pr-CAT, in which regions at -658, -287, and -172 were inserted 5' to the cat gene, were transiently transfected in pSVT10 and pSVT8 cells and showed that TAT-induced activation of the IL-6 promoter required a minimal region located between -287 and -54 bp. Moreover, experiments with plasmids carrying the -658, -287, and -172 bp regions of the IL-6 promoter inserted downstream to a TAR-deleted HIV1-LTR identified the sequence of -172 to -54 as the minimal region of the IL-6 promoter required for TAT to transactivate the TAR-deleted HIV1-LTR. By DNA-protein binding experiments, tat-transfected cells expressed a consistent increase in kappa B and nuclear factor (NF)-IL-6 binding activity. Accordingly, the pDRCAT and IL-1REK9CAT, carrying tandem repeats of NF-kappa B or NF-IL6 binding motifs, respectively, were activated in TAT-expressing cells. The biological relevance of the TAT-induced IL-6 secretion was addressed by generating 7TD1 cells, an IL-6-dependent mouse cell line, stably expressing the tat gene. These tat-positive cells expressed the endogenous IL-6 gene, secreted high amounts of murine IL-6, and grew efficiently in the absence of exogenous IL-6. Moreover, the tat-positive 7TD1 cells sustained the growth of parental 7TD1 cells and showed a dramatic increase in their tumorigenic potency. These results suggest that TAT protein may play a role in the pathogenesis of some HIV1-associated diseases by modulating the expression of host cellular genes.
Scala
G
G
Department of Biochemistry and Biomedical Technology, Medical School, University Federico II, Naples, Italy.
Ruocco
M R
MR
Ambrosino
C
C
Mallardo
M
M
Giordano
V
V
Baldassarre
F
F
Dragonetti
E
E
Quinto
I
I
Venuta
S
S
D000818:Animals; D001402:B-Lymphocytes; D001483:Base Sequence; D002460:Cell Line; D002461:Cell Line, Transformed; D002471:Cell Transformation, Neoplastic; D015500:Chloramphenicol O-Acetyltransferase / Q000096:biosynthesis / Q000378:metabolism; D017931:DNA Primers; D005260:Female; D015870:Gene Expression; D015696:Gene Products, tat / Q000096:biosynthesis / Q000378:metabolism*; D015756:Genes, tat; D015497:HIV-1 / Q000235:genetics* / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D015850:Interleukin-6 / Q000096:biosynthesis* / Q000235:genetics; D007700:Kinetics; D051379:Mice; D008819:Mice, Nude; D008969:Molecular Sequence Data; D010957:Plasmids; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D015533:Transcriptional Activation; D014162:Transfection; D054322:tat Gene Products, Human Immunodeficiency Virus
D017931:DNA Primers; D015696:Gene Products, tat; D015850:Interleukin-6; D054322:tat Gene Products, Human Immunodeficiency Virus; D015500:Chloramphenicol O-Acetyltransferase