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 phosphatase 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 inhibitors 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 provide a convenient and sensitive tool to study NFAT regulation.
Li
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Department of Pharmacology, Yale University School of Medicine, New Haven, Connecticut 06520-8066, USA.
Handschumacher
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D000469:Alkaline Phosphatase / Q000096:biosynthesis; D003994:Bucladesine / Q000494:pharmacology; D002999:Clone Cells; D005576:Colforsin / Q000494:pharmacology; D016572:Cyclosporine / Q000494:pharmacology; D004268:DNA-Binding Proteins / Q000096:biosynthesis / Q000502:physiology*; D015232:Dinoprostone / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004988:Ethers, Cyclic / Q000494:pharmacology; D005260:Female; D006801:Humans; D007166:Immunosuppressive Agents / Q000494:pharmacology; D007700:Kinetics; D008213:Lymphocyte Activation; D008387:Marine Toxins; D050778:NFATC Transcription Factors; D009687:Nuclear Proteins; D019319:Okadaic Acid; D010080:Oxazoles / Q000494:pharmacology; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors; D010920:Placenta; D011247:Pregnancy; D054645:Protein Phosphatase 1; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism; D013601:T-Lymphocytes / Q000187:drug effects / Q000276:immunology / Q000502:physiology*; D016559:Tacrolimus / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D014157:Transcription Factors / Q000096:biosynthesis / Q000502:physiology*; D014162:Transfection; D014407:Tumor Cells, Cultured
D004268:DNA-Binding Proteins; D004791:Enzyme Inhibitors; D004988:Ethers, Cyclic; D007166:Immunosuppressive Agents; D008387:Marine Toxins; D050778:NFATC Transcription Factors; D009687:Nuclear Proteins; D010080:Oxazoles; D011994:Recombinant Proteins; D014157:Transcription Factors; D005576:Colforsin; D019319:Okadaic Acid; D003994:Bucladesine; C059041:calyculin A; D016572:Cyclosporine; D000469:Alkaline Phosphatase; D010749:Phosphoprotein Phosphatases; D054645:Protein Phosphatase 1; D015232:Dinoprostone; D013755:Tetradecanoylphorbol Acetate; D016559:Tacrolimus