FIGURE 9.

Iso-dependent VASP phosphorylation of PDE4-null MEFs. MEFs deficient in PDE4B (4B) and PDE4D (4D) along with wild type controls were stimulated with Iso for the times indicated and then harvested in SDS buffer containing phosphatase inhibitors. Equal amounts of protein were separated by SDS-PAGE, and Western blotting was performed using an antibody against phosphorylated Ser-157 of VASP. A, representative blot showing the time course of phosphorylated VASP detection in MEFs stimulated with 100 nm Iso in the indicated genotypes. B, quantification of VASP phosphorylation time course as a ratio of maximum wild type signal. C, representative blot showing the increased Ser-157 phosphorylation in response to increasing concentrations of Iso after 3 min of stimulation. D, quantification of the Iso dose response. Data were corrected for loading with tubulin and represent the mean ± S.E. of four experiments. EC₅₀ values are as follows: WT, 5.57 ± 2.36 nm; PDE4B^−/−, 1.54 ± 1.85 nm and PDE4D^−/−, 1.74 ± 2.50 nm (mean ± S.E.; n = 4). Error bars, S.E.