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.
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
Jay H
JH
Bream
Deborah L
DL
Hodge
Rivkah
R
Gonsky
Rosanne
R
Spolski
Warren J
WJ
Leonard
Stephanie
S
Krebs
Stephan
S
Targan
Akio
A
Morinobu
John J
JJ
O'Shea
Howard A
HA
Young
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