IL-4-activated STAT-6 inhibits IFN-gamma-induced CD40 gene expression in macrophages/microglia.
The antagonism between the cytokines IFN-gamma and IL-4 is well documented, but the mechanism by which IL-4 inhibits IFN-gamma-induced gene expression is not clearly understood. CD40 is a type I transmembrane protein that is critical for proper functioning of the immune system. We have previously shown that IFN-gamma is the most potent inducer of CD40 expression by macrophages and microglia. In this report, we describe the molecular mechanisms by which IL-4 inhibits IFN-gamma-induced CD40 expression. IL-4 suppresses IFN-gamma-induced CD40 gene expression in both macrophages and microglia, and such inhibition is dependent on the activation of STAT-6. Nuclear run-on and transfection studies indicate that IL-4-mediated repression is at the transcriptional level. Furthermore, IL-4 inhibition of IFN-gamma-induced CD40 expression is specific, since IL-4 does not inhibit IFN-gamma-induced IFN-responsive factor-1 gene expression. Site-directed mutagenesis studies demonstrate that two STAT binding sites, named proximal and distal IFN-gamma-activated sequences, in the human CD40 promoter are important for IL-4 inhibition of IFN-gamma-induced CD40 promoter activity. Moreover, EMSAs indicate that IL-4-activated STAT-6 binds to these two STAT binding sites. These results suggest that IL-4 inhibition of IFN-gamma-induced CD40 gene expression is mediated by direct STAT-6 binding to the CD40 promoter.
Nguyen
V T
VT
Department of Cell Biology, University of Alabama, Birmingham, AL 35294, USA.
Benveniste
E N
EN
D000818:Animals; D019013:CD40 Antigens / Q000096:biosynthesis / Q000235:genetics*; D002460:Cell Line; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation / Q000276:immunology*; D006801:Humans; D007166:Immunosuppressive Agents / Q000494:pharmacology*; D007371:Interferon-gamma / Q000037:antagonists & inhibitors* / Q000494:pharmacology; D015847:Interleukin-4 / Q000494:pharmacology*; D008264:Macrophages / Q000201:enzymology / Q000276:immunology* / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D008809:Mice, Inbred C3H; D017628:Microglia / Q000201:enzymology / Q000276:immunology* / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000276:immunology; D011505:Protein-Tyrosine Kinases / Q000378:metabolism / Q000502:physiology; D012333:RNA, Messenger / Q000096:biosynthesis / Q000378:metabolism; D020218:Response Elements / Q000276:immunology; D050800:STAT6 Transcription Factor; D015398:Signal Transduction / Q000235:genetics / Q000276:immunology; D015534:Trans-Activators / Q000378:metabolism / Q000502:physiology*; D014158:Transcription, Genetic / Q000276:immunology
D019013:CD40 Antigens; D004268:DNA-Binding Proteins; D007166:Immunosuppressive Agents; D012333:RNA, Messenger; D050800:STAT6 Transcription Factor; C494101:STAT6 protein, human; C494100:Stat6 protein, mouse; D015534:Trans-Activators; D015847:Interleukin-4; D007371:Interferon-gamma; D011505:Protein-Tyrosine Kinases