Interferon-gamma modulates the lipopolysaccharide-induced expression of AP-1 and NF-kappa B at the mRNA and protein level in human monocytes.
Interferon-gamma (IFN-gamma) modulates the expression of several cytokines by human monocytes at the transcriptional level. In view of these findings, we analyzed the effects of IFN-gamma on the expression of different transcription factors in activated human monocytes. Priming of human monocytes with IFN-gamma resulted in the down regulation of c-fos and c-jun mRNA in response to stimulation with lipopolysaccharide (LPS) compared to the effects of LPS alone. Not only was this effect observed at the mRNA level, but activator protein-1 (AP-1) DNA binding capacity was affected as well, A strong reduction was observed in the LPS-induced DNA-binding activity of AP-1 in the presence of IFN-gamma. LPS-stimulated monocytes showed an increased expression of p105 mRNA, the precursor of the p50 subunit of the transcription factor nuclear factor-kappa B (NF-kappa B), while no effect was noticed on the expression of p65 mRNA. In contrast, IFN-gamma priming did not affect the expression of p105 transcripts but enhanced the expression of p65 mRNA (two-fold). Priming with IFN-gamma followed by LPS stimulation resulted in a further increase in the expression of p65 mRNA. This was due to an increase in the half-life of p65 mRNA (75 vs 150 minutes). Electrophoretic mobility shift assays (EMSAs) demonstrated that unstimulated monocytes predominantly expressed p50 NF-kappa B. Stimulation with LPS or IFN-gamma resulted in the expression of p50 and p65 subunits, while the combination of IFN-gamma plus LPS caused a further increase in the expression of NF-kappa B. With Western blotting, it was shown that nuclear extracts from monocytes contained p50 and p65 protein in response to LPS and IFN-gamma stimulation. However, the combined stimulation did not result in enhanced p50 and p65 protein expression. The effects of IFN-gamma on the transcription factors were specific, since no change was observed in the expression of NF-IL-6 or I kappa B alpha, the inhibitor of NF-kappa B. We conclude that the effects of IFN-gamma on the expression of the transcription factors AP-1 and NF-kappa B may be important for the modulatory effects of IFN-gamma on the cytokine expression in activated human monocytes.
de Wit
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Division of Hematology, Department of Medicine, University of Groningen, The Netherlands.
Hoogstraten
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Halie
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Vellenga
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D001483:Base Sequence; D005786:Gene Expression Regulation / Q000187:drug effects*; D016762:Genes, fos / Q000187:drug effects; D016758:Genes, jun / Q000187:drug effects; D006801:Humans; D007371:Interferon-gamma / Q000494:pharmacology*; D015850:Interleukin-6 / Q000096:biosynthesis* / Q000235:genetics; D008070:Lipopolysaccharides / Q000494:pharmacology; D008969:Molecular Sequence Data; D009000:Monocytes / Q000187:drug effects* / Q000378:metabolism; D016328:NF-kappa B / Q000096:biosynthesis* / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000187:drug effects; D014176:Protein Biosynthesis / Q000187:drug effects*; D012333:RNA, Messenger / Q000096:biosynthesis* / Q000235:genetics; D018808:Transcription Factor AP-1 / Q000096:biosynthesis* / Q000235:genetics; D014158:Transcription, Genetic / Q000187:drug effects*
D015850:Interleukin-6; D008070:Lipopolysaccharides; D016328:NF-kappa B; D012333:RNA, Messenger; D018808:Transcription Factor AP-1; D007371:Interferon-gamma