Regulation of the Nuclear Factor of Activated T Cells in Stably Transfected Jurkat Cell Clones

Two Jurkat cell clones have been stably transfected with a reporter vector for the nuclear factor of activated T cells (NFAT). Upon stimulation, they express high levels of secreted heat stable placental alkaline phosphatase. With these clones, we demonstrated that NFAT activation induced by phorbol 12-myristate 13-acetate and ionomycin was inhibited by both cyclosporin A (CsA) (IC50= 8 nM) and FK506 (IC50= 160 pM), presumably by inhibition of calcineurin activity. Selective phosphatase inhibitors for protein phophatase 1 (PP1) and 2A (PP2A) that do not inhibit calcineurin, such as okadaic acid and calyculin A, also inhibited NFAT activation with IC50s of 87 nM and 4 nM, respectively, suggesting that okadaic acid and related inhibitiors may block NFAT activation through the inhibition of PP1, instead of PP2A. NFAT activation was also inhibited by agents that increase cAMP concentrations such as dibutyryl cAMP, forskolin and prostaglandin E2. These stable Jurkat cell clones proved a convenient and sensitive tool to study NFAT regulation.