Cloning of a novel human prostaglandin receptor with characteristics of the pharmacologically defined EP₂ subtype

A cDNA that when expressed has the binding and functional characteristics of the pharmacologically defined EP₂ prostaglandin (PG) receptor [Cardiovasc. Drug Rev. 11:165-179 (1993)] has been cloned from a human placenta library. This clone, known as Hup-4, encodes a protein of 358 amino acids that has only ~30% overall identity with other PG receptors, including mouse and human clones that have been designated as EP₂ receptors [J. Biol. Chem. 268:7759-7762 (1993); Biochem. Biophys. Res. Commun. 197:263-270 (1993)]. In COS-7 cells transfected with Hup-4, PGE₂ stimulated the formation of cAMP with an EC₅₀ of ~50 nM. The EP₂-selective agonists AH13205 and butaprost were also active, with EC₅₀ values in the range of 2- 6 μM. The order of potency of PGs for competition with binding of [³H]PGE₂ to membranes prepared from COS-7 cells transfected with Hup-4 was PGE₂ ≥ PGE₁ > 16,16-dimethyl-PGE₂ ≥ 11-deoxy-PGE₁ > butaprost > AH13205 > 19(R)- OH-PGE₂. Natural PGs and analogues that are selective for the FP (PGF(2α)), DP (PGD₂), EP₁ (sulprostone), EP₃ (MB 28767), and EP₄ (1-OH-PGE₁) receptors were inactive or competed poorly with the binding of [³H]PGE₂ (<50% displacement of specific binding at 10 μM). Northern blot analysis showed the presence of a Hup-4 message of ~3.1 kilobases in mRNA from human lung and placenta. Reverse transcription-polymerase chain reaction studies also indicated that Hup-4 is probably expressed in human uterus and in HL-60 (human promyelocytic leukemia) cells. Our findings suggest that Hup-4 encodes the pharmacologically defined EP₂ receptor, whereas the mouse and human cDNAs previously classified as EP₂ may represent another EP receptor subtype or the recently defined EP₄ subtype [Prostaglandins 47:151-168 (1994)].