COSTIMULATION BY B7-1 AND LFA-3 TARGETS DISTINCT NUCLEAR FACTORS THATBIND TO THE INTERLEUKIN-2 PROMOTER - B7-1 NEGATIVELY REGULATES LFA-3-INDUCED NF-AT DNA-BINDING

We have characterized the regulation of nuclear factors involved in tr anscriptional control of the interleukin-2 (IL-2) promoter-enhancer ac tivity in Jurkat T cells stimulated with superantigen presented on HLA -DR transfectants combined,vith the ligands LFA-3 (CD58) and B7-1 (CD8 0), 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 nu clear 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 w as seen after B7-1 costimulation, Both LFA-3 costimulation and B7-1 co stimulation induced the AP-1 and NF-kappa B nuclear factors, Distinct compositions of the NF-AT complexes were seen in B7-1- and LFA-3-costi mulated 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-kappa B co mplexes induced in B7-1- and LFA-3-costimulated T cells showed similar contents, Transient transfection of Jurkat T cells with a construct e ncoding the IL-2 enhancer-promoter region (position -500 to +60) linke d to a luciferase reporter gene revealed that B7-1 costimulation was r equired to induce strong transcriptional activity, Combined B7-1-LFA-3 costimulation resulted in a synergistic increase in IL-2 transcriptio nal activity, Multimers of the AP-I, NF-AT, NF-kappa B, and CD28 respo nse elements showed distinct kinetics and activity after LFA-3 and B7- 1 costimulation and revealed that B7-1 and LFA-3 converge to superindu ce transcriptional activity of the AP-1, NF-AT, and CD28 response elem ents, Transcriptional studies with an IL-2 enhancer-promoter carrying a mutation in the CD28 response element site revealed that the activit y was reduced by 80% after B7-1 and B7-1-LFA-3 costimulation whereas t he 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 r egulation of the levels of IL-2 induced by LFA-3 and B7-1 costimulatio n and favor autocrine and paracrine T-cell responses, respectively.