The alpha, but not the beta, isoform of the human thromboxane A2 receptor is a target for prostacyclin-mediated desensitization.
In this study, we examined the effects the prostacyclin receptor (IP) agonist cicaprost exhibited on U46619-mediated thromboxane A(2) receptor (TP) signaling in platelets and compared it to that which occurs in human embryonic kidney (HEK) 293 cells stably overexpressing the individual TPalpha or TPbeta isoforms. Consistent with previous studies, cicaprost abrogated U46619-mediated platelet aggregation and mobilization of intracellular calcium ([Ca(2+)](i)). In HEK 293 cells, signaling by TPalpha, but not TPbeta, was subject to IP-mediated desensitization in a protein kinase A-dependent, protein kinase C-independent manner. Desensitization of TPalpha signaling was independent of the nature of the IP agonist used, the level of IP expression, or the subtype of G(q) protein. Signaling by TP(Delta)(328), a truncated variant of TP devoid of the divergent residues of the TPs, or by TPalpha(S329A), a site-directed mutant of TPalpha, were insensitive to IP agonist activation. Whole cell phosphorylations established that TPalpha, but not TPbeta or TPalpha(S329A), is subject to IP-mediated phosphorylation and that TPalpha phosphorylation is inhibited by H-89. Thus, we conclude that TPalpha, but not TPbeta, is subject to cross-desensitization by IP mediated through direct protein kinase A phosphorylation at Ser(329) and propose that TPalpha may be the isoform physiologically relevant to TP:IP-mediated vascular hemostasis.
Walsh
M T
MT
Department of Biochemistry, Conway Institute of Biomolecular and Biomedical Research, Merville House, University College Dublin, Belfield, Dublin 4, Ireland.
Foley
J F
JF
Kinsella
B T
BT
D019796:15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid / Q000494:pharmacology; D001792:Blood Platelets / Q000187:drug effects; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors; D011464:Epoprostenol / Q000031:analogs & derivatives / Q000494:pharmacology*; D006801:Humans; D007295:Inositol Phosphates / Q000378:metabolism; D007546:Isoquinolines / Q000494:pharmacology; D009154:Mutation; D010766:Phosphorylation; D010974:Platelet Aggregation / Q000187:drug effects; D020033:Protein Isoforms / Q000378:metabolism; D044006:Receptors, Epoprostenol; D011982:Receptors, Prostaglandin / Q000819:agonists; D017482:Receptors, Thromboxane / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D013449:Sulfonamides; D014662:Vasoconstrictor Agents / Q000494:pharmacology
D007295:Inositol Phosphates; D007546:Isoquinolines; D020033:Protein Isoforms; D044006:Receptors, Epoprostenol; D011982:Receptors, Prostaglandin; D017482:Receptors, Thromboxane; D013449:Sulfonamides; D014662:Vasoconstrictor Agents; D019796:15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; D011464:Epoprostenol; D017868:Cyclic AMP-Dependent Protein Kinases; C063509:N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide; C043867:cicaprost; D002118:Calcium