Figure 5

Effect of gemcitabine on the MAPK pathway in CK2 subunits-depleted human PANC-1 cells. A. Total cellular proteins (80 μg) from exponentially growing cells treated as indicated in the figure were subjected to Western blot analysis with antibodies directed against the proteins or their phosphorylated form as indicated. β-actin was applied as control for equal loading. p44/42MAPK: the upper band is p44MAPK, the lower one is p42MAPK. B. Whole lysates (1 mg) from cells treated as indicated in the figure were subjected to a non-radioactive kinase activity assay in the presence of a GST-c-Jun fusion protein linked to glutathione agarose beads. The precipitated complex was subjected to a phosphorylation assay as described in the materials and methods section. Where indicated, the JNK inhibitor SP600125 was used at a concentration of 20 μM. The protein and phosphorylation levels of c-Jun were detected by Western blot with the indicated antibodies. Representative results from three independent experiments are shown.