PHOSPHATASE 2A PARTICIPATES IN INTERFERON-GAMMA-S INDUCED UP-REGULATION OF C1 INHIBITOR MESSENGER-RNA EXPRESSION

C1 inhibitor (C1 INH) is the major inhibitor of the proteolytically ac tive subcomponents of C1, kallikrein, activated forms of factor XII, a nd factor XIa in plasma. We determined the mechanism(s) how interferon -gamma (IFN-gamma) regulates C1 INH mRNA expression in HepG2 cells. Cy cloheximide or aniso-mycin treatment alone did not increase C1 INH mRN A nor did it potentiate C1 INH mRNA expression after IFN-gamma stimula tion. C1 INH mRNA levels on Northern blot from untreated and IFN-gamma -treated cells did not change for more than 20 hours after actinomycin D treatment. Actinomycin D and 5,6-dichloro-1-beta-ribofuranosylbenzi midazole abolished IFN-gamma-induced C1 INH mRNA expression. Relativel y more C1 INH mRNA precursor (heterogeneous nuclear RNA [hnRNA]) was d etected in total RNA from IFN-gamma-treated HepG2 cells than unstimula ted cells. Treatment of HepG2 cells with the phosphatase 1 and 2A inhi bitors, okadaic acid (greater than or equal to 50 nmol/L) and calyculi n (greater than or equal to 25 nmol/L), decreased IFN-gamma's ability to upregulate C1 INH mRNA. The phosphatase 2A inhibitor, cantharidin ( greater than or equal to 10 mu mol/L), also blocked the IFN-gamma indu ction of the C1 INH gene. In HepG2 cells total phosphatase 2A activity was significantly increased by C6 ceramide but not IFN-gamma. However , C6 ceramide itself did not increase C1 INH mRNA expression. These da ta indicate that phosphatase 2A is required to dephosphorylate a subst rate in order for IFN-gamma to induce the transcriptional upregulation of C1 INH mRNA, but phosphatase 2A is not a direct stimulator of C1 I NH gene expression. (C) 1996 by The American Society of Hematology.