EP1- and FP-mediated cross-desensitization of the alpha (alpha) and beta (beta) isoforms of the human thromboxane A2 receptor.

1. Heterologous desensitization or intermolecular cross-talk plays a critical role in regulating intracellular signalling by diverse members of the G-protein-coupled receptor superfamily. We have previously established that the alpha and beta isoforms of the human thromboxane A(2) receptor (TP) undergo differential desensitization of signalling in response to 17 phenyl trinor prostaglandin (PG)E(2), an agonist of the EP(1) subtype of the PGE(2) receptor (EP) family. 2. Herein, we investigated the molecular basis of TPalpha and TPbeta desensitization in human embryonic kidney (HEK) 293 cells and in renal mesangial cells in response to 17 phenyl trinor PGE(2) and in response to the PGF(2alpha) receptor (FP) agonist PGF(2alpha), and sought to identify the target site(s) of those desensitizations. 3. Our results demonstrated that TPalpha and TPbeta receptors are subject to desensitization in response to both EP(1) and FP receptor activation and that these effects are mediated by direct protein kinase (PK)C phosphorylation of the individual TP isoforms within their unique carboxyl-terminal (C)-tail domains. 4. Moreover, deletion/site-directed mutagenesis and metabolic labelling studies identified Thr(337), within TPalpha, and Thr(399), within TPbeta, as the specific target residues for PKC phosphorylation and EP(1)- and FP-mediated desensitization of TPalpha and TPbeta signalling, respectively. 5. Hence, in conclusion, while the TPalpha and TPbeta diverge within their C-tail domains, they have evolved to share a similar mechanism of PKC-induced phosphorylation and desensitization in response to EP(1) and FP receptor activation, though it occurs at sites unique to the individual TP isoforms.