Figure 5.

ROCK increase MRLC phosphorylation upon EGF stimulation. (A) ROCK but not MLCK phosphorylates MRLC upon EGF stimulation. aPKCζ^−/− and control cells were preincubated with ML-7, Y-27632, or both, stimulated with EGF for 4 min, lysed in SDS-PAGE sample buffer, divided into 2 samples that were subjected to Western blot analysis, One sample was probed with anti-MRLC antibodies and the other with anti-pMRLC antibodies. Shown are representative Western blots. The relative amounts of pMRLC to total MRLC are shown. Values represent the mean ± SEM for 5 independent experiments, ** p < 0.01, values are compared with control cells. (B) aPKCζ is required for EGF-dependent phosphorylation of MYPT. aPKCζ^−/− and control cells were stimulated with EGF, and at the indicated time point cells were lysed in SDS-PAGE sample buffer and subjected to Western blot analysis with anti-pMYPT and anti-actin antibodies. Shown are representative Western blots. (B) Densitometry analysis of Western blots. The relative amounts of pMYPT to total actin are shown. Values represent the mean ± SEM for n = 3.