FIG. 1.
Effect of PD 184352 on MEK phosphorylation by Raf in vitro and in vivo. (A) Kinase-inactive MEK was incubated with Raf in the presence of increasing concentrations of PD 184352 in an in vitro kinase assay as described in Materials and Methods. PD 184352 did not inhibit Raf-mediated phosphorylation of MEK under these conditions. (B) KBALB cells were serum starved, treated for 30 min with the noted concentrations of PD 184352, and then stimulated for 5 min with PDGF. Lysates were prepared and immunoblotted for levels of phosphorylated MEK (pMEK), total MEK (MEK), and phosphorylated MAPK (pp44MAPK1 and pp42MAPK2). Samples are shown in duplicate.