Signal Requirements for Interleukin 4 Promoter Activation in Human T Cells

We have studied the signal requirements for human IL-4 promoter activation in Jurkat T cells by the use of DNA transfection assays with vectors carrying the IL-4 promoter linked to a reporter gene. Stimulation with calcium (Ca2+) ionophores (ionomycin), but not with phorbol esters (phorbol myristate acetate, PMA) or cyclic AMP elevating agents (prostaglandin E2, PGE2), induced the transcriptional activity of the IL-4 promoter by ∼3-fold. Costimulation with ionomycin and PGE2resulted in the same level of IL-4 promoter activity as the stimulation with ionomycin alone. In contrast, costimulation with ionomycin and PMA decreased the activity of the IL-4 promoter by ∼40% compared to stimulation with ionomycin alone. Induction of IL-4 promoter by ionomycin was partially inhibited (∼50% inhibition) in the presence of as high as 2 μg/ml cyclosporin A (CsA), an inhibitor of the Ca+/calmodulin-dependent phosphatase calcineurin. Under the same conditions, only 0.1 μg/ml of CsA inhibited by >95% the transactivation of the IL-2 promoter in response to ionomycin and PMA. Transfection of a deletion mutant of the calcineurin catalytic subunit (ΔCaM-AI) known to have Ca2+-independent, constitutive phosphatase activity increased IL-4 promoter activity by ∼14-fold. Stimulation with ionomycin of cells transfected with low doses of ΔCaM-AI, further induced IL-4 promoter activity by ∼2-fold. These results identify the Ca2+-signaling system as a key component of the signal transduction pathway leading to IL-4 promoter activation in Jurkat T cells and suggest a major role of calcineurin in its regulation.