FIG. 5.

FLNa Ser²¹⁵² phosphorylation is stimulated by agonists of the MAP kinase pathway and specifically inhibited by UO126. (A) Rat 3Y1 cells were serum starved for 24 h. The cells were stimulated with 50 ng of EGF ml⁻¹ or 100 ng of phorbol myristate acetate (PMA) ml⁻¹ for 15 min, with 10% serum or 10 μM forskolin for 10 min, or with 100 nM insulin for 20 min. Lysates were blotted with phospho-Ser²¹⁵²-FLNa (p-FLNa), total FLNa, phospho-Ser³⁸⁰-RSK (p-RSK), and total ERK1/2 antibodies. (B) Rat 3Y1 cells were serum starved and then pretreated with one of the following inhibitors for 30 min prior to EGF stimulation: DMSO control, 10 μM UO126, 10 μM H-89, or 100 nM wortmannin. Lysates were subjected to Western blot analysis using p-FLNa, total FLNa, p-RSK, and phosphorylated ERK1/2 (p-ERK 1/2) antibodies.