Different pathways for activation of extracellular signal-regulated kinase through thromboxane A2 receptor isoforms

Thromboxane A2 receptor (TP) consists of two alternatively spliced isoforms, TPa and TPP, which differ in their cytoplasmic tails. In the present study, we examined the difference in signal transcluction of TP alpha and TP beta, using stably expressing cells of TPa and TPP. The cells expressing TP alpha (TP alpha-SC2) and TP beta (TP beta-SC15) were selected based on the similar binding sites of [H-3]-SQ29548, a TP antagonist. U46619, a TP agonist, elicited phosphoinositide hydrolysis in TPa-SC2 and TPP-SC15 cells with a similar concentration-dependency. U46619 also caused the phosphorylation of extracellular signal-regulated kinase (ERK1/2) in both TPCr-SC2 and TPJ5-SC15 cells. While the peak of the phosphorylation of ERKI/2 was observed 5 min after addition of U46619 in TlPm-SC2 cells, the long lasting phosphorylation up to 60 min was in TPP-SC 15 cells. U46619-induced phosphorylation of ERK1/2 at 5 min was inhibited toy pertussis toxin in both cells, suggesting that G, is involved in the phosphorylation mediated via both TP isoforms. Interfering G(12/13) activity by overexpression of p115-RGS reduced U46619induced ERK1/2 phosphorylation in TPP-SC15 cells, but not in TPCrSC2 cells. H89, an inhibitor of protein kinase A (PKA), reduced U46619-induced ERK1/2 phosphorylation in TP alpha-SC2 cells, but not in TP beta-SC15 cells. These results indicate that Gi may be involved in TP-mediated ERK1/2 phosphorylation in both isoforms. In addition, H89-sensitive kinase and G(12/13) may be involved in TP-mediated ERK1/2 phosphorylation in TPa and TPP, respectively.