Negative regulation of Gq-mediated pathways in platelets by G(12/13) pathways through Fyn kinase

J Biol Chem. 2011 Jul 8;286(27):24170-9. doi: 10.1074/jbc.M110.212274. Epub 2011 May 18.

Abstract

Platelets contain high levels of Src family kinases (SFKs), but their functional role downstream of G protein pathways has not been completely understood. We found that platelet shape change induced by selective G(12/13) stimulation was potentiated by SFK inhibitors, which was abolished by intracellular calcium chelation. Platelet aggregation, secretion, and intracellular Ca(2+) mobilization mediated by low concentrations of SFLLRN or YFLLRNP were potentiated by SFK inhibitors. However, 2-methylthio-ADP-induced intracellular Ca(2+) mobilization and platelet aggregation were not affected by PP2, suggesting the contribution of SFKs downstream of G(12/13), but not G(q)/G(i), as a negative regulator to platelet activation. Moreover, PP2 potentiated YFLLRNP- and AYPGKF-induced PKC activation, indicating that SFKs downstream of G(12/13) regulate platelet responses through the negative regulation of PKC activation as well as calcium response. SFK inhibitors failed to potentiate platelet responses in the presence of G(q)-selective inhibitor YM254890 or in G(q)-deficient platelets, indicating that SFKs negatively regulate platelet responses through modulation of G(q) pathways. Importantly, AYPGKF-induced platelet aggregation and PKC activation were potentiated in Fyn-deficient but not in Lyn-deficient mice compared with wild-type littermates. We conclude that SFKs, especially Fyn, activated downstream of G(12/13) negatively regulate platelet responses by inhibiting intracellular calcium mobilization and PKC activation through G(q) pathways.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Adenosine Diphosphate / analogs & derivatives
  • Adenosine Diphosphate / pharmacology
  • Animals
  • Blood Platelets / enzymology*
  • Calcium / metabolism
  • Enzyme Activation / drug effects
  • Enzyme Activation / genetics
  • GTP-Binding Protein alpha Subunits, G12-G13 / genetics
  • GTP-Binding Protein alpha Subunits, G12-G13 / metabolism*
  • GTP-Binding Protein alpha Subunits, Gq-G11 / genetics
  • GTP-Binding Protein alpha Subunits, Gq-G11 / metabolism*
  • Humans
  • Mice
  • Mice, Mutant Strains
  • Oligopeptides / pharmacology
  • Peptide Fragments / pharmacology
  • Platelet Aggregation / drug effects
  • Platelet Aggregation / physiology*
  • Protein Kinase C / genetics
  • Protein Kinase C / metabolism
  • Proto-Oncogene Proteins c-fyn / genetics
  • Proto-Oncogene Proteins c-fyn / metabolism*
  • Thionucleotides / pharmacology

Substances

  • Oligopeptides
  • Peptide Fragments
  • Thionucleotides
  • thrombin receptor peptide (42-47)
  • tyrosyl-phenylalanyl-leucyl-leucyl-arginyl-asparaginyl-arginine
  • methylthio-ADP
  • Adenosine Diphosphate
  • FYN protein, human
  • Fyn protein, mouse
  • Proto-Oncogene Proteins c-fyn
  • Protein Kinase C
  • GTP-Binding Protein alpha Subunits, G12-G13
  • GTP-Binding Protein alpha Subunits, Gq-G11
  • Calcium