Figure 1.

Inhibition of IL-6 activation of Stat3 by PMA is reversed by MEK inhibitors. MM6 cells (A and B) or 293T and CHO cells (C) were incubated in complete medium with PMA for 5 min (A) or 15 min (B and C), followed by a 12-min stimulation with IL-6 (20 ng/ml). Cells were preincubated with PD98059 (20 μM) or U0126 (10 μM) for 45 min. Cell extracts were assayed for binding to the hSIE oligonucleotide by using EMSA as described (17). In A, the same extracts were analyzed by immunoblotting with specific Stat3 antiserum or were immunoprecipitated with ERK2 antibodies. Seventy-five percent of each immunoprecipitate was used in a kinase assay with MBP substrate, and 25% was subjected to immunoblotting with ERK2 antibodies. IB, immunoblot; PD, PD98059; U, U0126.