Fig. 2.

ERK1/2 selectively affects CME activities in cancer cells. (A) Representative Western blots used to measure the efficiencies of kinase inhibitors in reducing ERK1/2 phosphorylation in control, ERK1/2 inhibitor (FR180204 and SCH772984; 10 μM)-treated or MEK1/2 inhibitor (GSK1120212; 10 μM)-treated ARPE-19 and H1299 cells. See SI Appendix, Fig. S2A for quantitation. (B) Endocytosis of TfnR was measured in ARPE-19 and H1299 cells untreated (Ctrl) or incubated with the indicated inhibitors, as described above. All data were normalized to the percentage of TfnR internalized after 10 min in control cells and represent mean ± SEM (n = 3). See Fig. 1B for examples of absolute kinetics of TfnR uptake, which vary among cell lines. (C and D) Rates of CCP initiation (Left), median lifetimes (Middle) and average lifetime distribution (Right) of all bona fide CCPs in control and inhibitor-treated ARPE-19 cells (C) and H1299 cells (D). Data in C and D were obtained from at least 15 cells per condition (>10,000 CCPs per condition). The box plots represent median, 25th, and 75th percentiles, and error bars indicate the outermost data points. Two-tailed Student’s t tests were used to assess statistical significance for comparison with controls. **P < 0.005, ***P < 0.0005.