Evidence for a role for Galphai1 in mediating weak agonist-induced platelet aggregation in human platelets: reduced Galphai1 expression and defective Gi signaling in the platelets of a patient with a chronic bleeding disorder.
We have examined platelet functional responses and characterized a novel signaling defect in the platelets of a patient suffering from a chronic bleeding disorder. Platelet aggregation responses stimulated by weak agonists such as adenosine diphosphate (ADP) and adrenaline were severely impaired. In comparison, both aggregation and dense granule secretion were normal following activation with high doses of collagen, thrombin, or phorbol-12 myristate-13 acetate (PMA). ADP, thrombin, or thromboxane A2 (TxA2) signaling through their respective Gq-coupled receptors was normal as assessed by measuring either mobilization of intracellular calcium, diacylglycerol (DAG) generation, or pleckstrin phosphorylation. In comparison, Gi-mediated signaling induced by either thrombin, ADP, or adrenaline, examined by suppression of forskolin-stimulated rise in cyclic AMP (cAMP) was impaired, indicating dysfunctional Galphai signaling. Immunoblot analysis of platelet membranes with specific antiserum against different Galpha subunits indicated normal levels of Galphai2,Galphai3,Galphaz, and Galphaq in patient platelets. However, the Galphai1level was reduced to 25% of that found in normal platelets. Analysis of platelet cDNA and gDNA revealed no abnormality in either the Galphai1 or Galphai2 gene sequences. Our studies implicate the minor expressed Galphai subtype Galphai1 as having an important role in regulating signaling pathways associated with the activation of alphaIIbbeta3 and subsequent platelet aggregation by weak agonists.
Patel
Yatin M
YM
Department of Haematology, Canterbury Health Laboratories, Christchurch, New Zealand. yatin.m.patel@kcl.ac.uk
Patel
Kirti
K
Rahman
Salman
S
Smith
Mark P
MP
Spooner
Gillian
G
Sumathipala
Rushika
R
Mitchell
Michael
M
Flynn
Geraldine
G
Aitken
Alexandra
A
Savidge
Geoffrey
G
D000244:Adenosine Diphosphate / Q000494:pharmacology; D000328:Adult; D016718:Arachidonic Acid / Q000097:blood; D001791:Blood Platelet Disorders / Q000209:etiology* / Q000235:genetics; D001792:Blood Platelets / Q000187:drug effects / Q000502:physiology; D002118:Calcium / Q000097:blood; D002250:Carbon Radioisotopes; D002462:Cell Membrane / Q000737:chemistry; D005576:Colforsin / Q000494:pharmacology; D003094:Collagen / Q000494:pharmacology; D000242:Cyclic AMP / Q000097:blood; D018076:DNA, Complementary / Q000737:chemistry; D004075:Diglycerides / Q000097:blood; D004837:Epinephrine / Q000494:pharmacology; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000097:blood / Q000235:genetics* / Q000502:physiology*; D006801:Humans; D008297:Male; D009000:Monocytes / Q000737:chemistry; D010761:Phosphorus Radioisotopes; D010974:Platelet Aggregation / Q000187:drug effects; D017422:Sequence Analysis, DNA; D012701:Serotonin / Q000097:blood; D015398:Signal Transduction; D013917:Thrombin / Q000494:pharmacology; D013929:Thromboxane B2 / Q000097:blood; D014316:Tritium
D002250:Carbon Radioisotopes; D018076:DNA, Complementary; D004075:Diglycerides; D010761:Phosphorus Radioisotopes; D014316:Tritium; D005576:Colforsin; D016718:Arachidonic Acid; D012701:Serotonin; D013929:Thromboxane B2; D000244:Adenosine Diphosphate; D003094:Collagen; D000242:Cyclic AMP; D013917:Thrombin; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D002118:Calcium; D004837:Epinephrine