MODULATION OF LEVELS OF A NEGATIVE TRANSCRIPTION FACTOR FOR IL-2 BY 12-O-TETRADECANOYL PHORBOL-13-ACETATE AND OKADAIC ACID

A negatively acting transcription factor, negative regulatory element- A (NREA) that suppresses the transcription of interleukin 2 (IL-2) mRN A, has been described previously. We found that treatment of primary b ovine lymphocytes with 12-O-tetradecanoyl phorbol-13-acetate (TPA), an activator of protein kinase C, for at least 18 h both increased the l evels of the factor and blocked concanavalin A (Con Al-induced prolife ration. In contrast, treatments of less than 18 h with TPA decreased N REA levels and increased Con A-induced proliferation. NREA binding act ivity also increased over basal levels during the first 4 h of stimula tion of lymphocytes with Con A in the absence of TPA; after 4 h, NREA levels fell. Phosphorylation of the NREA protein was required for bind ing to its DNA consensus sequence. Furthermore, incubation of lymphocy tes with okadaic acid (OKA), a phosphatase inhibitor, led to increased levels of NREA binding activity and to decreased cell proliferation. Because exposure of lymphocytes to either OKA or TPA should lead to an increase in the phosphorylation and binding of the NREA protein, and a decrease in IL-2 production, proliferation should be decreased. Incu bation of lymphocytes with either TPA or OKA inhibited proliferation. However, the mechanisms of action of OKA and TPA appeared to be differ ent because exogenous IL-2 reversed the inhibition of proliferation ca used by TPA but not by OKA. (C) 1996 Academic Press Limited.