Akt activation in platelets depends on Gi signaling pathways.
The serine-threonine kinase Akt has been established as an important signaling intermediate in regulating cell survival, cell cycle progression, as well as agonist-induced platelet activation. Stimulation of platelets with various agonists including thrombin results in Akt activation. As thrombin can stimulate multiple G protein signaling pathways, we investigated the mechanism of thrombin-induced activation of Akt. Stimulation of platelets with a PAR1-activating peptide (SFLLRN), PAR4-activating peptide (AYPGKF), and thrombin resulted in Thr308 and Ser473 phosphorylation of Akt, which results in its activation. This phosphorylation and activation of Akt were dramatically inhibited in the presence of AR-C69931MX, a P2Y12 receptor-selective antagonist, or GF 109203X, a protein kinase C inhibitor, but Akt phosphorylation was restored by supplemental Gi or Gz signaling. Unlike wild-type mouse platelets, platelets from Galphaq-deficient mice failed to trigger Akt phosphorylation by thrombin and AYPGKF, whereas Akt phosphorylation was not affected by these agonists in platelets from mice that lack P2Y1 receptor. However, ADP caused Akt phosphorylation in Galphaq- and P2Y1-deficient platelets, which was completely blocked by AR-C69931MX. In contrast, ADP failed to cause Akt phosphorylation in platelets from mice treated with clopidogrel, and thrombin and AYPGKF induced minimal phosphorylation of Akt, which was not affected by AR-C69931MX in these platelets. These data demonstrate that Gi, but not Gq or G12/13, signaling pathways are required for activation of Akt in platelets, and Gi signaling pathways, stimulated by secreted ADP, play an essential role in the activation of Akt in platelets.
Kim
Soochong
S
Department of Physiology, Temple University School of Medicine, Philadelphia, Pennsylvania 19140, USA.
Jin
Jianguo
J
Kunapuli
Satya P
SP
D000244:Adenosine Diphosphate / Q000494:pharmacology; D000818:Animals; D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism*; D000077144:Clopidogrel; D004789:Enzyme Activation / Q000187:drug effects; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000097:blood* / Q000172:deficiency / Q000235:genetics; D006801:Humans; D066298:In Vitro Techniques; D051379:Mice; D018345:Mice, Knockout; D009842:Oligopeptides / Q000494:pharmacology; D010446:Peptide Fragments / Q000494:pharmacology; D010766:Phosphorylation; D011493:Protein Kinase C / Q000097:blood; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins / Q000097:blood*; D051057:Proto-Oncogene Proteins c-akt; D018048:Receptors, Purinergic P2 / Q000097:blood / Q000172:deficiency / Q000235:genetics; D058472:Receptors, Purinergic P2Y1; D015398:Signal Transduction; D013917:Thrombin / Q000494:pharmacology; D013988:Ticlopidine / Q000031:analogs & derivatives* / Q000494:pharmacology
D009842:Oligopeptides; C546382:P2RY1 protein, human; C546383:P2ry1 protein, mouse; D010446:Peptide Fragments; D011518:Proto-Oncogene Proteins; D018048:Receptors, Purinergic P2; D058472:Receptors, Purinergic P2Y1; C455901:alanyl-tyrosyl-prolyl-glycyl-lysyl-phenylalanine; C082835:thrombin receptor peptide (42-47); D000244:Adenosine Diphosphate; D000077144:Clopidogrel; C494918:AKT1 protein, human; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D011493:Protein Kinase C; D013917:Thrombin; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D013988:Ticlopidine