Rap1 activation in platelets stimulated with PAR4p requires CalDAG-GEFI and P2Y12-mediated signaling. (A) Time course of Rap1 activation in WT (± 5 μg/mL Ro31-8220) or CalDAG-GEFI^−/− platelets stimulated with 1.25 mM of PAR4p. (B) Rap1-GTP detected in lysates from CalDAG-GEFI–deficient platelets at t = 10 minutes after addition of PAR4p. Rap1 activation was completely inhibited in the presence of Ro31-8220. (C) Effect of 2-MesAMP on Rap1 activation in WT or CalDAG-GEFI-deficient platelets. Platelets were pretreated with 50 μM 2-MesAMP for 5 minutes followed by stimulation with 1.25 mM of PAR4p.