Distinct mechanisms for regulation of the interleukin-8 gene involve synergism and cooperativity between C/EBP and NF-kappa B.
The interleukin-8 promoter is transcriptionally activated by interleukin-1, tumor necrosis factor alpha, phorbol myristate acetate, or hepatitis B virus X protein through a sequence located between positions -91 and -71. This region contains an NF-kappa B-like and a C/EBP-like binding site. We show here that several members of the NF-kappa B family, including p65, p50, p52, and c-Rel, can bind to this region, confirming an authentic NF-kappa B binding site in the interleukin-8 promoter. Further, C/EBP binds only weakly to the interleukin-8 promoter site. Electrophoretic mobility shift assays with proteins overexpressed in COS cells and with nuclear extracts from tumor necrosis factor alpha-stimulated HeLa cells demonstrated a strong cooperative binding of C/EBP to its site when NF-kappa B is bound to its adjacent binding site. Transfection studies lead to a model that suggests a highly complex regulation of interleukin-8 gene expression at multiple levels: independent binding of C/EBP and NF-kappa B to their respective sites, cooperative binding of C/EBP and NF-kappa B to DNA, and positive synergistic activation through the C/EBP binding site and inhibition through the NF-kappa B binding site by combinations of C/EBP and NF-kappa B. Thus, the ultimate regulation of interleukin-8 gene expression depends on the ratio of cellular C/EBP and NF-kappa B.
Stein
B
B
Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill 27599.
Baldwin
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AS
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D022762:CCAAT-Enhancer-Binding Proteins; D018236:Carcinoma, Embryonal; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000187:drug effects; D016762:Genes, fos; D006367:HeLa Cells; D006801:Humans; D007375:Interleukin-1 / Q000494:pharmacology; D016209:Interleukin-8 / Q000096:biosynthesis* / Q000235:genetics*; D007700:Kinetics; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D016328:NF-kappa B / Q000378:metabolism*; D009687:Nuclear Proteins / Q000378:metabolism*; D009838:Oligodeoxyribonucleotides / Q000138:chemical synthesis / Q000378:metabolism; D017354:Point Mutation; D011401:Promoter Regions, Genetic / Q000187:drug effects; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D015534:Trans-Activators / Q000494:pharmacology; D014157:Transcription Factors / Q000378:metabolism; D014162:Transfection; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D054334:Viral Regulatory and Accessory Proteins
D022762:CCAAT-Enhancer-Binding Proteins; D004268:DNA-Binding Proteins; D007375:Interleukin-1; D016209:Interleukin-8; D016328:NF-kappa B; D009687:Nuclear Proteins; D009838:Oligodeoxyribonucleotides; D015534:Trans-Activators; D014157:Transcription Factors; D014409:Tumor Necrosis Factor-alpha; D054334:Viral Regulatory and Accessory Proteins; C057628:hepatitis B virus X protein; D013755:Tetradecanoylphorbol Acetate