title: Costimulation by B7-1 and LFA-3 targets distinct nuclear factors that bind to the interleukin-2 promoter: B7-1 negatively regulates LFA-3-induced NF-AT DNA binding.
abstract: We have characterized the regulation of nuclear factors involved in transcriptional control of the interleukin-2 (IL-2) promoter-enhancer activity in Jurkat T cells stimulated with superantigen presented on HLA-DR transfectants combined with the ligands LFA-3 (CD58) and B7-1 (CD80). Gel shift analyses showed that NF-AT was strongly induced in LFA-3-costimulated Jurkat T cells, suggesting that NF-AT is a key target nuclear factor for the CD2-LFA-3 pathway. Studies using HLA-DR-B7-1-LFA-3 triple transfectants showed that the LFA-3-induced NF-AT DNA binding activity was negatively regulated by B7-1 costimulation. In contrast, induction of a CD28 response complex containing only c-Rel proteins was seen after B7-1 costimulation. Both LFA-3 costimulation and B7-1 costimulation induced the AP-1 and NF-kappaB nuclear factors. Distinct compositions of the NF-AT complexes were seen in B7-1- and LFA-3-costimulated cells. LFA-3 induced primarily Jun-D, Fra-1, and Fra-2, while B7-1 induced June-D-Fos complexes. In contrast, AP-1 and NF-kappaB complexes induced in B7-1- and LFA-3-costimulated T cells showed similar contents. Transient transfection of Jurkat T cells with a construct encoding the IL-2 enhancer-promoter region (position -500 to +60) linked to a luciferase reporter gene revealed that B7-1 costimulation was required to induce strong transcriptional activity. Combined B7-1-LFA-3 costimulation resulted in a synergistic increase in IL-2 transcriptional activity. Multimers of the AP-1, NF-AT, NF-kappaB, and CD28 response elements showed distinct kinetics and activity after LFA-3 and B7-1 costimulation and revealed that B7-1 and LFA-3 converge to superinduce transcriptional activity of the AP-1, NF-AT, and CD28 response elements. Transcriptional studies with an IL-2 enhancer-promoter carrying a mutation in the CD28 response element site revealed that the activity was reduced by 80% after B7-1 and B7-1-LFA-3 costimulation whereas the transcriptional activity induced by LFA-3 was unaffected. Our data strongly suggest a selectivity in induction of nuclear factors by the CD2-LFA-3 and CD28-B7-1 pathways. This selectivity may contribute to regulation of the levels of IL-2 induced by LFA-3 and B7-1 costimulation and favor autocrine and paracrine T-cell responses, respectively.
authors.affiliation: Department of Cell and Molecular Biology, University of Lund, Sweden.
authors.forename: E
authors.initials: E
authors.lastname: Parra
authors.forename: M
authors.initials: M
authors.lastname: Varga
authors.forename: G
authors.initials: G
authors.lastname: Hedlund
authors.forename: T
authors.initials: T
authors.lastname: Kalland
authors.forename: M
authors.initials: M
authors.lastname: Dohlsten
mesh_terms: D018122:B7-1 Antigen / Q000235:genetics / Q000502:physiology*; D018801:CD2 Antigens / Q000502:physiology; D018106:CD28 Antigens / Q000502:physiology; D018968:CD58 Antigens / Q000235:genetics / Q000502:physiology*; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004768:Enterotoxins / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000276:immunology; D006684:HLA-DR Antigens / Q000235:genetics / Q000502:physiology; D006801:Humans; D007376:Interleukin-2 / Q000235:genetics*; D019169:Jurkat Cells; D007700:Kinetics; D050778:NFATC Transcription Factors; D009687:Nuclear Proteins; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic
chemical_list: D018122:B7-1 Antigen; D018801:CD2 Antigens; D018106:CD28 Antigens; D018968:CD58 Antigens; D004268:DNA-Binding Proteins; D004768:Enterotoxins; D006684:HLA-DR Antigens; D007376:Interleukin-2; D050778:NFATC Transcription Factors; D009687:Nuclear Proteins; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; C049463:enterotoxin E, Staphylococcal; D004247:DNA