FIGURE 7.

Phosphorylation of PDE3A increases binding to 14-3-3 proteins. Platelets were stimulated with the indicated agonists for 5 min (SFLLRN for 30 s, A), incubated with the indicated compounds for 30 min before treatment with 0.1 units/ml thrombin for 5 min (B) or stimulated with 0.1 units/ml thrombin for 5 min (C, D) prior to extraction. PDE3A was immunoprecipitated using anti-PDE3A covalently coupled to Sepharose (A-D). Immunoprecipitates were untreated (A, B) or incubated with 1 mm free peptide for 30 min at room temperature to compete off bound 14-3-3 (C, D). PDE3A immunoprecipitates were subsequently analyzed by immunoblotting with the indicated antibodies (A-C) or assayed for 3′-cAMP phosphodiesterase activity (D).