A distal region in the interferon-gamma gene is a site of epigenetic remodeling and transcriptional regulation by interleukin-2.
Interferon-gamma (IFN-gamma) is a multifunctional cytokine that defines the development of Th1 cells and is critical for host defense against intracellular pathogens. IL-2 is another key immunoregulatory cytokine that is involved in T helper differentiation and is known to induce IFN-gamma expression in natural killer (NK) and T cells. Despite concerted efforts to identify the one or more transcriptional control mechanisms by which IL-2 induces IFN-gamma mRNA expression, no such genomic regulatory regions have been described. We have identified a DNase I hypersensitivity site approximately 3.5-4.0 kb upstream of the transcriptional start site. Using chromatin immunoprecipitation assays we found constitutive histone H3 acetylation in this distal region in primary human NK cells, which is enhanced by IL-2 treatment. This distal region is also preferentially acetylated on histones H3 and H4 in primary Th1 cells as compared with Th2 cells. Within this distal region we found a Stat5-like motif, and in vitro DNA binding assays as well as in vivo chromosomal immunoprecipitation assays showed IL-2-induced binding of both Stat5a and Stat5b to this distal element in the IFNG gene. We examined the function of this Stat5-binding motif by transfecting human peripheral blood mononuclear cells with -3.6 kb of IFNG-luciferase constructs and found that phorbol 12-myristate 13-acetate/ionomycin-induced transcription was augmented by IL-2 treatment. The effect of IL-2 was lost when the Stat5 motif was disrupted. These data led us to conclude that this distal region serves as both a target of chromatin remodeling in the IFNG locus as well as an IL-2-induced transcriptional enhancer that binds Stat5 proteins.
Bream
Jay H
JH
Lymphocyte and Cell Biology Section, Molecular Immunology and Inflammation Branch, National Institute of Arthritis and Musculoskeletal and Skin Diseases, National Institutes of Health (NIH), Bethesda, Maryland 20892-1820, USA. jbream@jhsph.edu
Hodge
Deborah L
DL
Gonsky
Rivkah
R
Spolski
Rosanne
R
Leonard
Warren J
WJ
Krebs
Stephanie
S
Targan
Stephan
S
Morinobu
Akio
A
O'Shea
John J
JJ
Young
Howard A
HA
D020816:Amino Acid Motifs; D002455:Cell Division; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D002843:Chromatin / Q000378:metabolism; D003001:Cloning, Molecular; D004268:DNA-Binding Proteins / Q000378:metabolism; D003850:Deoxyribonuclease I / Q000378:metabolism; D004742:Enhancer Elements, Genetic; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D006657:Histones / Q000378:metabolism; D006801:Humans; D007371:Interferon-gamma / Q000235:genetics* / Q000378:metabolism; D018664:Interleukin-12 / Q000378:metabolism; D007376:Interleukin-2 / Q000378:metabolism*; D015759:Ionomycin / Q000494:pharmacology; D007963:Leukocytes, Mononuclear / Q000378:metabolism; D008894:Milk Proteins / Q000378:metabolism; D011233:Precipitin Tests; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000378:metabolism; D050799:STAT5 Transcription Factor; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D018417:Th1 Cells / Q000378:metabolism; D018418:Th2 Cells / Q000378:metabolism; D015534:Trans-Activators / Q000378:metabolism; D014158:Transcription, Genetic; D014162:Transfection; D025521:Tumor Suppressor Proteins
D002843:Chromatin; D004268:DNA-Binding Proteins; D006657:Histones; D007376:Interleukin-2; D008894:Milk Proteins; D012333:RNA, Messenger; D050799:STAT5 Transcription Factor; C494092:STAT5A protein, human; C494095:STAT5B protein, human; D015534:Trans-Activators; D025521:Tumor Suppressor Proteins; D018664:Interleukin-12; D015759:Ionomycin; D007371:Interferon-gamma; D003850:Deoxyribonuclease I; D013755:Tetradecanoylphorbol Acetate