Enhancer role of STAT5 in CD2 activation of IFN-gamma gene expression.
IFN-gamma is an important immunoregulatory protein with tightly controlled expression in activated T and NK cells. Three potential STAT binding regions have been recognized within the IFN-gamma promoter: 1) an IL-12-mediated STAT4 binding site at -236 bp; 2) a newly identified IL-2-induced STAT5 binding element at -3.6 kb; and 3) CD2-mediated STAT1 and STAT4 binding to an intronic element in mucosal T cells. However, functional activation of these sites remains unclear. In this study we demonstrate CD2-mediated activation of the newly characterized -3.6-kb IFN-gamma STAT5 binding region. CD2 signaling of human PBMC results in activation of the -3.6-kb IFN-gamma promoter, whereas mutation of the -3.6-kb STAT5 site attenuates promoter activity. Functional activation is accompanied by STAT5A but little STAT5B nucleoprotein binding to the IFN-gamma STAT5 site, as determined by competition and supershift assays. STAT5 activation via CD2 occurs independent of IL-2. Western and FACS analysis shows increased phospho-STAT5 following CD2 signaling. AG490, a tyrosine kinase inhibitor affecting Jak proteins, inhibits CD2-mediated IFN-gamma mRNA expression, secretion, and nucleoprotein binding to the IFN-gamma STAT5 site in a dose-dependent fashion. This report is the first to describe CD2-mediated activation of STAT5 and supports STAT5 involvement in regulation of IFN-gamma expression.
Gonsky
Rivkah
R
Inflammatory Bowel Disease Research Center, Cedars-Sinai Medical Center, 8700 Beverly Boulevard, Los Angeles, CA 90048, USA.
Deem
Richard L
RL
Bream
Jay
J
Young
Howard A
HA
Targan
Stephan R
SR
D018801:CD2 Antigens / Q000502:physiology*; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D004742:Enhancer Elements, Genetic / Q000276:immunology*; D006801:Humans; D007371:Interferon-gamma / Q000037:antagonists & inhibitors / Q000096:biosynthesis* / Q000235:genetics*; D053613:Janus Kinase 1; D008894:Milk Proteins / Q000235:genetics / Q000378:metabolism; D009687:Nuclear Proteins / Q000378:metabolism; D010766:Phosphorylation; D011401:Promoter Regions, Genetic / Q000276:immunology; D011485:Protein Binding / Q000235:genetics / Q000276:immunology; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors; D020218:Response Elements / Q000276:immunology; D050799:STAT5 Transcription Factor; D015398:Signal Transduction / Q000235:genetics* / Q000276:immunology*; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D015533:Transcriptional Activation / Q000276:immunology*; D025521:Tumor Suppressor Proteins; D020032:Tyrphostins / Q000494:pharmacology; D015854:Up-Regulation / Q000276:immunology
D018801:CD2 Antigens; D004268:DNA-Binding Proteins; D008894:Milk Proteins; D009687:Nuclear Proteins; D047428:Protein Kinase Inhibitors; D050799:STAT5 Transcription Factor; C494092:STAT5A protein, human; C494095:STAT5B protein, human; D015534:Trans-Activators; D025521:Tumor Suppressor Proteins; D020032:Tyrphostins; C095512:alpha-cyano-(3,4-dihydroxy)-N-benzylcinnamide; D007371:Interferon-gamma; D011505:Protein-Tyrosine Kinases; C507904:JAK1 protein, human; D053613:Janus Kinase 1