title: Differential effects of TNF-alpha and IFN-gamma on gene transcription mediated by NF-kappaB-Stat1 interactions.
abstract: Regulation of gene transcription by the cytokines tumor necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma) involves complex interactions between NF-kappaB and Stat families of transcription factors. The purpose of this study was to identify the spatial promoter requirements that govern cytokine synergy for gene transcription regulated by NF-kappaB and Stat factors. Using a set of transcription reporter-luciferase constructs, we show that the relative orientation of juxtaposed NF-kappaB-Stat (SIE) cis-elements determines the ability of TNF-alpha and IFN- gamma to induce gene transcription. Further, NF-kappaB and Stat1 proteins directly regulate transcription by interacting cooperatively on NF-kappaB-SIE DNA binding in response to TNF-alpha plus IFN-gamma. Coimmunoprecipitation provides evidence for a direct NF-kappaB/Stat1 protein-protein interaction. In contrast, IFN-gamma inhibits TNF-alpha-induced transcription of an NF-kappaB reporter gene in a Stat1-dependent mechanism in 2fTGH fibroblasts. Similarly, Stat1 is inhibitory to NF-kappaB overexpression-induced transcription. IFN-gamma and Stat1-dependent inhibition of NF-kappaB transcription occurs independent of TNF-alpha-induced NF-kappaB DNA binding. Interestingly, IFN-gamma pretreatment of 2fTGH fibroblasts potentiates TNF-alpha induction of Stat1 DNA binding. Further, ChIP analysis was applied to detect cytokine-induced in vivo binding and transcriptional regulation of the human inducible nitric oxide synthase (iNOS) gene by NF-kappaB and Stat1. These data demonstrate complex transcriptional regulatory mechanisms elicited by TNF-alpha and IFN-gamma and have potentially important implications for other genes differentially controlled by cytokines.
authors.affiliation: Department of Surgery, University of Pittsburgh School of Medicine, PA 15261, USA.
authors.forename: Raymond W
authors.initials: RW
authors.lastname: Ganster
authors.forename: Zhong
authors.initials: Z
authors.lastname: Guo
authors.forename: Lifang
authors.initials: L
authors.lastname: Shao
authors.forename: David A
authors.initials: DA
authors.lastname: Geller
mesh_terms: D045744:Cell Line, Tumor; D002843:Chromatin / Q000737:chemistry; D047369:Chromatin Immunoprecipitation; D016207:Cytokines / Q000378:metabolism; D004247:DNA / Q000737:chemistry; D005347:Fibroblasts / Q000378:metabolism; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D006801:Humans; D047468:Immunoprecipitation; D007249:Inflammation; D007371:Interferon-gamma / Q000378:metabolism*; D008156:Luciferases / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism*; D009841:Oligonucleotides / Q000737:chemistry; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D050794:STAT1 Transcription Factor / Q000378:metabolism*; D015534:Trans-Activators; D014158:Transcription, Genetic; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*
chemical_list: D002843:Chromatin; D016207:Cytokines; D016328:NF-kappa B; D009841:Oligonucleotides; D050794:STAT1 Transcription Factor; C494079:STAT1 protein, human; D015534:Trans-Activators; D014409:Tumor Necrosis Factor-alpha; D007371:Interferon-gamma; D004247:DNA; D008156:Luciferases