FIG. 2.

Mechanism of cyclic nucleotide-induced ERK5 inhibition. (A) Cells cultured as described for Fig. 1 were treated with 50 μg/ml of cycloheximide for 30 min, then with DMSO or 10 μM forskolin and 50 μM IBMX for 5, 15, or 30 min, and then for 15 min with 10 ng/ml EGF. ERK5 (top panel) and ERK1/2 (bottom panel) activities were monitored as described for Fig. 1. (B) Cells were first treated with either 5 μM H-89 for 1 h (top panel) or 15 μM myr-PKI peptide (bottom panel) for 4 h or overnight, then with DMSO or 10 μM forskolin and 50 μM IBMX for 15 min, and then for 15 min with 10 ng/ml EGF. Results are representative of at least three independent experiments.