FIG. 2.

p38 mediates stress-induced EGFR phosphorylation. (A) Time course assay of p38 phosphorylation dependent on EGFR kinase function. Cos-7 cells were pretreated with AG1478 (250 nM) or an equal volume of empty vehicle (dimethyl sulfoxide [DMSO]) for 20 min and stimulated with 0.3 M sorbitol or 200 μM hydrogen peroxide for the indicated periods. Cell extracts were immunoblotted with anti-phospho-p38 antibody, and the same filters were reprobed with polyclonal anti-p38 antibody. (B) Stress-induced EGFR activation depends on p38 but not ERK activity in Cos-7 cells. Cos-7 cells were pretreated with PD98059 (50 μM), SB202190 (10 μM), or an equal volume of empty vehicle (dimethyl sulfoxide) for 30 min and stimulated with 0.3 M sorbitol and 200 μM hydrogen peroxide for 10 min and the GPCR agonist LPA (10 μM) or EGF (2 ng/ml) for 3 min as a positive control. Cell extracts were assayed for EGFR tyrosine phosphorylation content. (C) Stress-induced EGFR activation depends on p38 activity in TCC-Sup carcinoma cells. TCC-Sup cells were pretreated as described for panel A and stimulated with 0.3 M sorbitol and 200 μM hydrogen peroxide for 10 min and EGF (2 ng/ml) for 3 min as a positive control. After lysis cell extracts were assayed for EGFR tyrosine phosphorylation content.