IFN-gamma-induced TNF-alpha expression is regulated by interferon regulatory factors 1 and 8 in mouse macrophages.
We have previously described that IFN-gamma induces cyclooxygenase 2 and inducible NO synthase expression by a mechanism that involved endogenously produced TNF-alpha. In this study, we report that TNF-alpha production is induced by IFN-gamma treatment in the murine macrophage cell line RAW 264.7. TNF-alpha mRNA levels are increased in cells treated with IFN-gamma in a time-dependent manner and IFN-gamma also increased human TNF-alpha promoter-dependent transcription. Two regions in the TNF-alpha promoter seem to be responsible for the IFN-gamma response: a distal region between -1311 and -615 bp of the human TNF-alpha promoter, and a proximal region located between -95 and -36 bp upstream of the transcriptional start. In contrast, IFN-gamma stimulation induces the expression of the transcription factors IRF-1 and IRF-8. Overexpression of these transcription factors produces an increase in the transcriptional activity of the human TNF-alpha promoter. There is a correlation between the regions of the TNF-alpha promoter responsible of the transcriptional activation elicited by IRF-1 and IRF-8 and those required for IFN-gamma response. In addition, IRF-1 and IRF-8 are recruited to the TNF-alpha promoter in IFN-gamma-treated RAW 264.7 cells, as demonstrated by chromatin immunoprecipitation assays. Moreover, overexpression of IRF-1 and IRF-8 induces TNF-alpha production in unstimulated RAW 264.7 macrophages, comparable to the production of TNF-alpha elicited by IFN-gamma stimulation, and silencing of IRF-1 and/or IRF-8 with specific small interfering RNAs, decreases IFN-gamma-elicited TNF-alpha production. In summary, IFN-gamma treatment induces TNF-alpha expression at transcriptional level requiring the coordinate action of IRF-1 and IRF-8.
Vila-del Sol
Virginia
V
Centro de Biología Molecular Severo Ochoa, Consejo Superior de Investigaciones Científicas-Universidad Autónoma de Madrid, c/Nicolás Cabrera, 1 Campus Cantoblanco, Universidad Autónoma de Madrid, Madrid, Spain.
Punzón
Carmen
C
Fresno
Manuel
M
D000818:Animals; D002460:Cell Line; D005786:Gene Expression Regulation / Q000276:immunology*; D006801:Humans; D050856:Interferon Regulatory Factor-1 / Q000235:genetics / Q000502:physiology*; D050835:Interferon Regulatory Factors / Q000235:genetics / Q000502:physiology*; D007371:Interferon-gamma / Q000502:physiology*; D008264:Macrophages / Q000276:immunology* / Q000378:metabolism*; D051379:Mice; D011401:Promoter Regions, Genetic / Q000276:immunology; D014158:Transcription, Genetic / Q000276:immunology; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis* / Q000235:genetics; D000099185:Interferon Regulatory Factor-8
D050856:Interferon Regulatory Factor-1; D050835:Interferon Regulatory Factors; D007371:Interferon-gamma; D014409:Tumor Necrosis Factor-alpha; D000099185:Interferon Regulatory Factor-8