FIG. 5.

SEB stimulates the ERK1/2 and p38 MAPK pathways. (A) MSIE cells were treated with SEB (10 μg/ml) for the times indicated, and total protein lysates were harvested. As positive controls, cells were stimulated with anisomycin (ANISO, 10 μg/ml) or phorbol-12,13-myristate acetate (PMA) for 30 min. Activated stress kinases were determined by using antibodies to the phosphospecific forms of the kinases as described in Materials and Methods. Images shown are representative of the results from three separate experiments. Approximately equal protein loading was observed on blots probed for total MAPK levels (data not shown). (B) MSIE cells were pretreated with the MEK-1 inhibitor, PD98059 (PD; 50 μM), the p38 inhibitor, SB203580 (SB; 30 μM), or both (PD + SB) for 2 h prior to treatment with SEB (10 μg/ml, 24 h) or saline vehicle (−). Hsc73 was used as a loading control. Images shown are representative of the results from three separate experiments.