Synergistic activation of interleukin-8 gene transcription by all-trans-retinoic acid and tumor necrosis factor-alpha involves the transcription factor NF-kappaB.
Induction of interleukin-8 (IL-8) by IL-1 or tumor necrosis factor (TNF), and repression by interferons or glucocorticoids have been shown to involve sequences between nucleotides -94 and -71 of the 5'-flanking region, and the transcription factors NF-IL-6 and NF-kappaB. The A3 cell line was derived from the human melanoma cell line G-361 by stable transfection with part of the IL-8 promoter (nucleotides -101 to +40 from transcription start) fused to the luciferase coding region. These regulatory sequences were sufficient for transcriptional activation by all-trans-retinoic acid (ATRA), 9-cis-retinoic acid, IL-1beta, or TNF-alpha. Simultaneous treatment of A3 cells with ATRA and TNF-alpha resulted in a dose- and time-dependent synergistic increase in luciferase expression and IL-8 mRNA levels. Transient transfections of the parental cell line demonstrated that the NF-kappaB binding site is essential for this synergistic transactivation. Electrophoretic mobility shift assays with nuclear extracts of A3 cells showed that stimulation with ATRA and TNF-alpha for more than 16 h resulted in enhanced NF-kappaB binding compared to that induced by TNF-alpha alone. The simultaneous treatment with ATRA and TNF-alpha also resulted in changes in the composition of NF-kappaB complexes bound to the IL-8 NF-kappaB site, preventing the formation of two TNF-alpha-inducible binding activities. We suggest that these complexes consist of repressive factors which, when removed, allow enhanced binding of NF-kappaB to its cognate site.
Harant
H
H
Sandoz Research Institute, A-1235 Vienna, Austria.
de Martin
R
R
Andrew
P J
PJ
Foglar
E
E
Dittrich
C
C
Lindley
I J
IJ
D004357:Drug Synergism; D006801:Humans; D018836:Inflammation Mediators / Q000378:metabolism; D016209:Interleukin-8 / Q000235:genetics*; D008545:Melanoma / Q000378:metabolism / Q000473:pathology; D016328:NF-kappa B / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D012333:RNA, Messenger / Q000235:genetics; D014158:Transcription, Genetic / Q000187:drug effects*; D014212:Tretinoin / Q000494:pharmacology*; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D015854:Up-Regulation
D018836:Inflammation Mediators; D016209:Interleukin-8; D016328:NF-kappa B; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D014212:Tretinoin