FIGURE 3.

Sustained ERK signaling is required for hyperinduction of cyclin D1 in PMA-treated IEC-18 cells. A, experimental scheme for PMA and U0126 treatment. Cells were treated with PMA or vehicle (EtOH) for a total of 6 h, with the addition of 10 μm U0126 to inhibit ERK activity 1, 2, 3, or 4 h into the PMA treatment. The duration of ERK activity after PMA addition is indicated on the right. B, effect of U0126 on cyclin D1 recovery and hyperinduction. IEC-18 cells were treated with PMA (P) or vehicle (C), and U0126 (10 μm) or vehicle (DMSO) was added at the times indicated in A. After 6 h of PMA treatment, cells were harvested, and protein extracts were subjected to Western blot analysis for cyclin D1, phospho-ERK, and total ERK. Data are representative of at least three independent experiments.