G proteins of the G12 family are activated via thromboxane A2 and thrombin receptors in human platelets.
Using subtype-specific antisera, we were able to identify the recently described alpha subunits of G12 and G13 in platelet membranes as 43-kDa proteins. Activation of the thromboxane A2 and the thrombin receptors in platelet membranes led to increased incorporation of the photoreactive GTP analogue [alpha-32P]GTP azidoanilide into immunoprecipitated alpha 12 and alpha 13, indicating that both receptors couple to G12 and G13. In addition, both activated receptors were demonstrated to couple to one or more members of the Gq family. In the absence of receptor agonists, incorporation of [alpha-32P]GTP azidoanilide into alpha 12 and alpha 13 was low over a long time period (up to 45 min) due to an obviously low basal nucleotide exchange rate, whereas an agonist-stimulated photolabeling of alpha 12 and alpha 13 could be observed after 4-8 min and reached a maximum after 30-45 min. Effective activation of G12 and G13 via the thromboxane A2 and the thrombin receptors was not dependent on the presence of GDP. Our results provide evidence that G12 and G13 play a functional role in transmembrane signal transduction and suggest that both proteins are involved in pathways leading to platelet activation.
Offermanns
S
S
Institut für Pharmakologie, Freie Universität Berlin, Germany.
Laugwitz
K L
KL
Spicher
K
K
Schultz
G
G
D019796:15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; D000345:Affinity Labels; D000595:Amino Acid Sequence; D001386:Azides / Q000378:metabolism; D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D008566:Membranes / Q000378:metabolism; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010777:Photochemistry; D011450:Prostaglandin Endoperoxides, Synthetic / Q000494:pharmacology; D018179:Receptors, Thrombin / Q000378:metabolism*; D017482:Receptors, Thromboxane / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D013917:Thrombin / Q000494:pharmacology; D013928:Thromboxane A2 / Q000031:analogs & derivatives / Q000494:pharmacology
D000345:Affinity Labels; D001386:Azides; D011450:Prostaglandin Endoperoxides, Synthetic; D018179:Receptors, Thrombin; D017482:Receptors, Thromboxane; D013928:Thromboxane A2; C031907:GTP gamma-4-azidoanilide; D019796:15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; D006160:Guanosine Triphosphate; D013917:Thrombin; D019204:GTP-Binding Proteins