The C‐Terminus of the Prostaglandin‐E‐Receptor EP3 Subtype is Essential for Activation of GTP‐Binding Protein

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Abstract
Three isoforms of the mouse prostaglandin‐E‐receptor EP3 subtype (EP3), EP3α, EP3β, and EP3γ, with different C‐termini, which are produced through alternative splicing, showed different efficiencies with respect to heterotrimeric GTP‐binding protein activation and adenylate cyclase inhibition [Sugimoto, Y., Negishi, M., Hayashi, Y., Namba, T., Honda, A., Watabe, A., Hirata, M., Narumiya, S. & Ichikawa, A. (1993) J. Biol. Chem. 268, 2712–2718; Irie, A., Sugimoto, Y., Namba, T., Harazono, A., Honda, A., Watabe, A., Negishi, M., Narumiya, S. & Ichikawa, A. (1993) Eur. J. Biochem. 217, 313–318]. To assess the role of the C‐terminus in GTP‐binding protein coupling, we truncated the C‐terminus of EP3 at an alternative splicing site and expressed the mutant receptor. The truncated receptor retained the ability to physically associate with Gi2, forming an agonist/receptor/Gi2 ternary complex, and to undergo the characteristic conversion of its agonist‐binding affinity, mediated by a guanine nucleotide from a low‐affinity state to a high‐affinity state. However, sulprostone, an EP3 agonist, failed not only to inhibit the forskolin‐induced CAMP accumulation in the mutant receptor‐expressing cells but also to stimulate the GTPase activity in the mutant receptor‐expressing cell membrane. These results indicated that the C‐terminus of EP3 is essential for the activation of GTP‐binding protein.