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. 1993 Nov 1;178(5):1483–1496. doi: 10.1084/jem.178.5.1483

Two essential regulatory elements in the human interferon gamma promoter confer activation specific expression in T cells

PMCID: PMC2191230  PMID: 8228802

Abstract

Like interleukin 2 (IL-2), interferon gamma (IFN-gamma) is an early response gene in T cells and both are prototypical T helper cell type 1 (Th-1) lymphokines. Yet IL-2 and IFN-gamma production are independently regulated, as demonstrated by their differential expression in certain T cell subsets, suggesting that the regulatory elements in these two genes must differ. To explore this possibility, the 5' flank of the human IFN-gamma gene was analyzed. Expression of IFN-gamma promoter- driven beta-galactosidase reporter constructs containing 538 bp of 5' flank was similar to that by constructs driven by the IL-2 promoter in activated Jurkat T cells; expression nearly as great was observed with the construct containing only 108 bp of IFN-gamma 5' flank. These IFN- gamma promoter constructs faithfully mirrored expression of the endogenous gene, in that expression required activation both with ionomycin and PMA, was inhibited by cyclosporin A, and was not observed in U937 or THP-1 cells. The region between -108 and -40 bp in the IFN- gamma promoter was required for promoter function and contained two elements that are conserved across species. Deletion of 10 bp within either element reduced promoter function by 70%, whereas deletions in nonconserved portions of this region had little effect on promoter function. The distal conserved element (-96 to -80 bp) contained a consensus GATA motif and a potential regulatory motif found in the promoter regions of the GM-CSF and macrophage inflammatory protein (MIP) genes. Factors binding to this element, including GATA-3, were found in Jurkat nuclear extracts by electromobility shift assays and two of the three complexes observed were altered in response to activation. One or both of these motifs are present in the 5' flank of multiple, other lymphokine genes, including IL-3, IL-4, IL-5, and GM- CSF, but neither is present in the promoter of the IL-2 gene. The proximal conserved element (-73 to -48 bp) shares homology with the NFIL-2A element in the IL-2 promoter; these elements compete for binding of factors in Jurkat nuclear extracts, although the NFIL-2A element but not the IFN-gamma element binds Oct-1. Factors binding to this element in the IFN-gamma gene were present in extracts from resting and activated Jurkat T cells. However, by in vivo footprinting of intact cells, this element was protected from methylation only with activation.(ABSTRACT TRUNCATED AT 400 WORDS)

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Selected References

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  1. Biassoni R., Verdiani S., Cambiaggi A., Romeo P. H., Ferrini S., Moretta L. Human CD3-CD16+ natural killer cells express the hGATA-3 T cell transcription factor and an unrearranged 2.3-kb TcR delta transcript. Eur J Immunol. 1993 May;23(5):1083–1087. doi: 10.1002/eji.1830230516. [DOI] [PubMed] [Google Scholar]
  2. Bikoff E. K., Jaffe L., Ribaudo R. K., Otten G. R., Germain R. N., Robertson E. J. MHC class I surface expression in embryo-derived cell lines inducible with peptide or interferon. Nature. 1991 Nov 21;354(6350):235–238. doi: 10.1038/354235a0. [DOI] [PubMed] [Google Scholar]
  3. Brown D. A., Kondo K. L., Wong S. W., Diamond D. J. Characterization of nuclear protein binding to the interferon-gamma promoter in quiescent and activated human T cells. Eur J Immunol. 1992 Sep;22(9):2419–2428. doi: 10.1002/eji.1830220935. [DOI] [PubMed] [Google Scholar]
  4. Brown D. A., Nelson F. B., Reinherz E. L., Diamond D. J. The human interferon-gamma gene contains an inducible promoter that can be transactivated by tax I and II. Eur J Immunol. 1991 Aug;21(8):1879–1885. doi: 10.1002/eji.1830210815. [DOI] [PubMed] [Google Scholar]
  5. Budd R. C., Cerottini J. C., MacDonald H. R. Selectively increased production of interferon-gamma by subsets of Lyt-2+ and L3T4+ T cells identified by expression of Pgp-1. J Immunol. 1987 Jun 1;138(11):3583–3586. [PubMed] [Google Scholar]
  6. Chrivia J. C., Wedrychowicz T., Young H. A., Hardy K. J. A model of human cytokine regulation based on transfection of gamma interferon gene fragments directly into isolated peripheral blood T lymphocytes. J Exp Med. 1990 Aug 1;172(2):661–664. doi: 10.1084/jem.172.2.661. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Ciccarone V. C., Chrivia J., Hardy K. J., Young H. A. Identification of enhancer-like elements in human IFN-gamma genomic DNA. J Immunol. 1990 Jan 15;144(2):725–730. [PubMed] [Google Scholar]
  8. Clipstone N. A., Crabtree G. R. Identification of calcineurin as a key signalling enzyme in T-lymphocyte activation. Nature. 1992 Jun 25;357(6380):695–697. doi: 10.1038/357695a0. [DOI] [PubMed] [Google Scholar]
  9. Durand D. B., Bush M. R., Morgan J. G., Weiss A., Crabtree G. R. A 275 basepair fragment at the 5' end of the interleukin 2 gene enhances expression from a heterologous promoter in response to signals from the T cell antigen receptor. J Exp Med. 1987 Feb 1;165(2):395–407. doi: 10.1084/jem.165.2.395. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Durand D. B., Shaw J. P., Bush M. R., Replogle R. E., Belagaje R., Crabtree G. R. Characterization of antigen receptor response elements within the interleukin-2 enhancer. Mol Cell Biol. 1988 Apr;8(4):1715–1724. doi: 10.1128/mcb.8.4.1715. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Ehlers S., Smith K. A. Differentiation of T cell lymphokine gene expression: the in vitro acquisition of T cell memory. J Exp Med. 1991 Jan 1;173(1):25–36. doi: 10.1084/jem.173.1.25. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. English B. K., Hammond W. P., Lewis D. B., Brown C. B., Wilson C. B. Decreased granulocyte-macrophage colony-stimulating factor production by human neonatal blood mononuclear cells and T cells. Pediatr Res. 1992 Mar;31(3):211–216. doi: 10.1203/00006450-199203000-00004. [DOI] [PubMed] [Google Scholar]
  13. Eustice D. C., Feldman P. A., Colberg-Poley A. M., Buckery R. M., Neubauer R. H. A sensitive method for the detection of beta-galactosidase in transfected mammalian cells. Biotechniques. 1991 Dec;11(6):739-40, 742-3. [PubMed] [Google Scholar]
  14. Fiering S., Northrop J. P., Nolan G. P., Mattila P. S., Crabtree G. R., Herzenberg L. A. Single cell assay of a transcription factor reveals a threshold in transcription activated by signals emanating from the T-cell antigen receptor. Genes Dev. 1990 Oct;4(10):1823–1834. doi: 10.1101/gad.4.10.1823. [DOI] [PubMed] [Google Scholar]
  15. Fox H. S., Bond B. L., Parslow T. G. Estrogen regulates the IFN-gamma promoter. J Immunol. 1991 Jun 15;146(12):4362–4367. [PubMed] [Google Scholar]
  16. Geballe A. P., Mocarski E. S. Translational control of cytomegalovirus gene expression is mediated by upstream AUG codons. J Virol. 1988 Sep;62(9):3334–3340. doi: 10.1128/jvi.62.9.3334-3340.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Gray P. W., Goeddel D. V. Structure of the human immune interferon gene. Nature. 1982 Aug 26;298(5877):859–863. doi: 10.1038/298859a0. [DOI] [PubMed] [Google Scholar]
  18. Hardy K. J., Manger B., Newton M., Stobo J. D. Molecular events involved in regulating human interferon-gamma gene expression during T cell activation. J Immunol. 1987 Apr 1;138(7):2353–2358. [PubMed] [Google Scholar]
  19. Hardy K. J., Peterlin B. M., Atchison R. E., Stobo J. D. Regulation of expression of the human interferon gamma gene. Proc Natl Acad Sci U S A. 1985 Dec;82(23):8173–8177. doi: 10.1073/pnas.82.23.8173. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Ho I. C., Vorhees P., Marin N., Oakley B. K., Tsai S. F., Orkin S. H., Leiden J. M. Human GATA-3: a lineage-restricted transcription factor that regulates the expression of the T cell receptor alpha gene. EMBO J. 1991 May;10(5):1187–1192. doi: 10.1002/j.1460-2075.1991.tb08059.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Horton R. M., Cai Z. L., Ho S. N., Pease L. R. Gene splicing by overlap extension: tailor-made genes using the polymerase chain reaction. Biotechniques. 1990 May;8(5):528–535. [PubMed] [Google Scholar]
  22. Jain J., Valge-Archer V. E., Rao A. Analysis of the AP-1 sites in the IL-2 promoter. J Immunol. 1992 Feb 15;148(4):1240–1250. [PubMed] [Google Scholar]
  23. Joulin V., Bories D., Eléouet J. F., Labastie M. C., Chrétien S., Mattéi M. G., Roméo P. H. A T-cell specific TCR delta DNA binding protein is a member of the human GATA family. EMBO J. 1991 Jul;10(7):1809–1816. doi: 10.1002/j.1460-2075.1991.tb07706.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Kang S. M., Beverly B., Tran A. C., Brorson K., Schwartz R. H., Lenardo M. J. Transactivation by AP-1 is a molecular target of T cell clonal anergy. Science. 1992 Aug 21;257(5073):1134–1138. doi: 10.1126/science.257.5073.1134. [DOI] [PubMed] [Google Scholar]
  25. Klausner R. D., Samelson L. E. T cell antigen receptor activation pathways: the tyrosine kinase connection. Cell. 1991 Mar 8;64(5):875–878. doi: 10.1016/0092-8674(91)90310-u. [DOI] [PubMed] [Google Scholar]
  26. Ko L. J., Yamamoto M., Leonard M. W., George K. M., Ting P., Engel J. D. Murine and human T-lymphocyte GATA-3 factors mediate transcription through a cis-regulatory element within the human T-cell receptor delta gene enhancer. Mol Cell Biol. 1991 May;11(5):2778–2784. doi: 10.1128/mcb.11.5.2778. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Lewis D. B., Larsen A., Wilson C. B. Reduced interferon-gamma mRNA levels in human neonates. Evidence for an intrinsic T cell deficiency independent of other genes involved in T cell activation. J Exp Med. 1986 Apr 1;163(4):1018–1023. doi: 10.1084/jem.163.4.1018. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Lewis D. B., Prickett K. S., Larsen A., Grabstein K., Weaver M., Wilson C. B. Restricted production of interleukin 4 by activated human T cells. Proc Natl Acad Sci U S A. 1988 Dec;85(24):9743–9747. doi: 10.1073/pnas.85.24.9743. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Lewis D. B., Wilson C. B. Gamma-interferon: an immunoregulatory lymphokine with immunotherapeutic potential. Pediatr Infect Dis J. 1990 Sep;9(9):642–651. [PubMed] [Google Scholar]
  30. Lewis D. B., Yu C. C., Meyer J., English B. K., Kahn S. J., Wilson C. B. Cellular and molecular mechanisms for reduced interleukin 4 and interferon-gamma production by neonatal T cells. J Clin Invest. 1991 Jan;87(1):194–202. doi: 10.1172/JCI114970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Liu J., Farmer J. D., Jr, Lane W. S., Friedman J., Weissman I., Schreiber S. L. Calcineurin is a common target of cyclophilin-cyclosporin A and FKBP-FK506 complexes. Cell. 1991 Aug 23;66(4):807–815. doi: 10.1016/0092-8674(91)90124-h. [DOI] [PubMed] [Google Scholar]
  32. Marine J., Winoto A. The human enhancer-binding protein Gata3 binds to several T-cell receptor regulatory elements. Proc Natl Acad Sci U S A. 1991 Aug 15;88(16):7284–7288. doi: 10.1073/pnas.88.16.7284. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Martin D. I., Orkin S. H. Transcriptional activation and DNA binding by the erythroid factor GF-1/NF-E1/Eryf 1. Genes Dev. 1990 Nov;4(11):1886–1898. doi: 10.1101/gad.4.11.1886. [DOI] [PubMed] [Google Scholar]
  34. Miyatake S., Shlomai J., Arai K., Arai N. Characterization of the mouse granulocyte-macrophage colony-stimulating factor (GM-CSF) gene promoter: nuclear factors that interact with an element shared by three lymphokine genes--those for GM-CSF, interleukin-4 (IL-4), and IL-5. Mol Cell Biol. 1991 Dec;11(12):5894–5901. doi: 10.1128/mcb.11.12.5894. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Mosmann T. R., Schumacher J. H., Street N. F., Budd R., O'Garra A., Fong T. A., Bond M. W., Moore K. W., Sher A., Fiorentino D. F. Diversity of cytokine synthesis and function of mouse CD4+ T cells. Immunol Rev. 1991 Oct;123:209–229. doi: 10.1111/j.1600-065x.1991.tb00612.x. [DOI] [PubMed] [Google Scholar]
  36. Mueller P. R., Wold B. In vivo footprinting of a muscle specific enhancer by ligation mediated PCR. Science. 1989 Nov 10;246(4931):780–786. doi: 10.1126/science.2814500. [DOI] [PubMed] [Google Scholar]
  37. Nakao M., Nomiyama H., Shimada K. Structures of human genes coding for cytokine LD78 and their expression. Mol Cell Biol. 1990 Jul;10(7):3646–3658. doi: 10.1128/mcb.10.7.3646. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Nimer S. D., Morita E. A., Martis M. J., Wachsman W., Gasson J. C. Characterization of the human granulocyte-macrophage colony-stimulating factor promoter region by genetic analysis: correlation with DNase I footprinting. Mol Cell Biol. 1988 May;8(5):1979–1984. doi: 10.1128/mcb.8.5.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Nimer S., Fraser J., Richards J., Lynch M., Gasson J. The repeated sequence CATT(A/T) is required for granulocyte-macrophage colony-stimulating factor promoter activity. Mol Cell Biol. 1990 Nov;10(11):6084–6088. doi: 10.1128/mcb.10.11.6084. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Northrop J. P., Crabtree G. R., Mattila P. S. Negative regulation of interleukin 2 transcription by the glucocorticoid receptor. J Exp Med. 1992 May 1;175(5):1235–1245. doi: 10.1084/jem.175.5.1235. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Pfeifer G. P., Steigerwald S. D., Mueller P. R., Wold B., Riggs A. D. Genomic sequencing and methylation analysis by ligation mediated PCR. Science. 1989 Nov 10;246(4931):810–813. doi: 10.1126/science.2814502. [DOI] [PubMed] [Google Scholar]
  42. Rubinstein M., Novick D., Fischer D. G. The human interferon-gamma receptor system. Immunol Rev. 1987 Jun;97:29–50. doi: 10.1111/j.1600-065x.1987.tb00515.x. [DOI] [PubMed] [Google Scholar]
  43. Sanders M. E., Makgoba M. W., Sharrow S. O., Stephany D., Springer T. A., Young H. A., Shaw S. Human memory T lymphocytes express increased levels of three cell adhesion molecules (LFA-3, CD2, and LFA-1) and three other molecules (UCHL1, CDw29, and Pgp-1) and have enhanced IFN-gamma production. J Immunol. 1988 Mar 1;140(5):1401–1407. [PubMed] [Google Scholar]
  44. Stimac E., Lyons S., Pious D. Transcription of HLA class II genes in the absence of B-cell-specific octamer-binding factor. Mol Cell Biol. 1988 Sep;8(9):3734–3739. doi: 10.1128/mcb.8.9.3734. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Swain S. L., Bradley L. M., Croft M., Tonkonogy S., Atkins G., Weinberg A. D., Duncan D. D., Hedrick S. M., Dutton R. W., Huston G. Helper T-cell subsets: phenotype, function and the role of lymphokines in regulating their development. Immunol Rev. 1991 Oct;123:115–144. doi: 10.1111/j.1600-065x.1991.tb00608.x. [DOI] [PubMed] [Google Scholar]
  46. Tocci M. J., Matkovich D. A., Collier K. A., Kwok P., Dumont F., Lin S., Degudicibus S., Siekierka J. J., Chin J., Hutchinson N. I. The immunosuppressant FK506 selectively inhibits expression of early T cell activation genes. J Immunol. 1989 Jul 15;143(2):718–726. [PubMed] [Google Scholar]
  47. Ullman K. S., Flanagan W. M., Edwards C. A., Crabtree G. R. Activation of early gene expression in T lymphocytes by Oct-1 and an inducible protein, OAP40. Science. 1991 Oct 25;254(5031):558–562. doi: 10.1126/science.1683003. [DOI] [PubMed] [Google Scholar]
  48. Weiss A. Structure and function of the T cell antigen receptor. J Clin Invest. 1990 Oct;86(4):1015–1022. doi: 10.1172/JCI114803. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Young H. A., Komschlies K. L., Ciccarone V., Beckwith M., Rosenberg M., Jenkins N. A., Copeland N. G., Durum S. K. Expression of human IFN-gamma genomic DNA in transgenic mice. J Immunol. 1989 Oct 1;143(7):2389–2394. [PubMed] [Google Scholar]
  50. Young H. A., Ortaldo J. R. One-signal requirement for interferon-gamma production by human large granular lymphocytes. J Immunol. 1987 Aug 1;139(3):724–727. [PubMed] [Google Scholar]

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