title: Eosinophilic inflammation: mechanisms regulating IL-5 transcription in human T lymphocytes.
abstract: BACKGROUND
  Interleukin (IL)-5 is a key regulator of eosinophilia in allergic inflammation and parasite infections but the mechanisms regulating IL-5 expression in activated human T lymphocytes are poorly understood. From studies on mouse cells, the activation protein (AP)-1 and GATA-3 sites in the proximal promoter region appear to be important in IL-5 regulation but the significance of an adjacent Ets/nuclear factor of activated T cell (NFAT) site has been less clear.
  
  
  METHODS
  Interleukin-5 transcriptional activity was measured by transfection of reporter genes into the human HSB-2 cells and normal T lymphocytes. Expression vectors encoding transcription factors were used for transactivation studies and IL-5 expression measured using reporter genes and mRNA levels. Transcription factor binding was shown with chromatin immunoprecipitation (ChIP).
  
  
  RESULTS
  HSB-2 cells showed high inducible expression of IL-5 mRNA. Mutation of reporter gene plasmids showed the Ets/NFAT site was of equal importance to the AP-1 and GATA-3 sites in regulating IL-5 transcription. Transactivation by Ets1 increased luciferase expression 15-fold, in the absence of stimulation, and AP-1 (c-Fos/c-Jun) and GATA-3 gave transactivations of 85-fold, and 100-fold, respectively. Synergistic interactions were demonstrated between Ets1, GATA-3 and AP-1. Dominant-negative AP-1 inhibited IL-5 transcription. Transactivation by GATA-3 and synergy between GATA-3, Ets1 and AP-1 were verified measuring IL-5 mRNA levels. Chromatin immunoprecipitation showed increased binding of Ets1 and GATA-3 to the IL-5 promoter after stimulation. The importance of the Ets1 site and of synergistic interactions between the three transcription factors were verified with primary human T cells.
  
  
  CONCLUSIONS
  Ets1, GATA-3 and AP-1 synergize to regulate IL-5 transcription in human T cells.
authors.affiliation: Division of Molecular Bioscience, John Curtin School of Medical Research, Australian National University, Canberra, Australia.
authors.forename: J
authors.initials: J
authors.lastname: Wang
authors.forename: I G
authors.initials: IG
authors.lastname: Young
mesh_terms: D000818:Animals; D001665:Binding Sites; D047369:Chromatin Immunoprecipitation / Q000379:methods; D004804:Eosinophils / Q000276:immunology*; D050990:GATA3 Transcription Factor / Q000378:metabolism; D005786:Gene Expression Regulation; D017930:Genes, Reporter / Q000235:genetics; D006801:Humans; D007249:Inflammation / Q000235:genetics* / Q000276:immunology; D015848:Interleukin-5 / Q000096:biosynthesis / Q000235:genetics*; D008156:Luciferases / Q000235:genetics; D008213:Lymphocyte Activation; D051379:Mice; D011401:Promoter Regions, Genetic; D050784:Proto-Oncogene Protein c-ets-1 / Q000378:metabolism*; D013601:T-Lymphocytes / Q000276:immunology*; D018808:Transcription Factor AP-1 / Q000378:metabolism*; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014407:Tumor Cells, Cultured
chemical_list: C494041:ETS1 protein, human; D050990:GATA3 Transcription Factor; D015848:Interleukin-5; D050784:Proto-Oncogene Protein c-ets-1; D018808:Transcription Factor AP-1; D008156:Luciferases