Coupling of thromboxane A2 receptor isoforms to Gα13:: effects on ligand binding and signalling

Previous subtyping of thromboxane A(2) (TXA(2)) receptors in platelets and vascular smooth muscle cells was based on pharmacological criteria. Two distinct carboxy-terminal splice variants for TXA(2) receptors exist and they couple to several different G protein alpha subunits including G alpha(13), but it has not been established whether either or both isoforms interact with and signal through it. We sought to determine: (1) which TXA(2) receptor isoforms exist in vascular smooth muscle, (2) if G alpha(13) is present in vascular smooth muscle and (3) if G alpha(13) interacts with either or both of the two TXA(2) receptor isoforms as determined by changes in ligand binding properties and generation of intracellular signals. Both TXA(2) receptor isoforms and G alpha(13) were found in vascular smooth muscle cells. Both the alpha and beta isoforms of the TXA(2) receptors were transiently transfected with or without G alpha(13) into COS-7 (radioligand binding assays) or CHO cells (agonist induced Na+/H+ exchange). Co;expression of each receptor isoform with G alpha(13) significantly (P < 0.05) increased the affinity of each receptor for the two agonists, I-BOP and ONO11113, and decreased the affinity of the receptor for the antagonists, SQ29,548 and L657,925. I-BOP stimulated Na+/H+ exchange in vascular smooth muscle cells. Co-expression of G alpha(13) with each TXA(2) receptor isoform in CHO cells resulted in a significant (P < 0.04) agonist induced increase in Na+/H+ exchange compared to cells not transfected with G alpha(13). The results support the possibility that the previous classification of TXA(2) receptor subtypes based on pharmacological criteria reflect unique interactions with specific G protein alpha subunits. (C) 1999 Elsevier Science B.V. All rights reserved.