Characterization of the Prostaglandin EP₃-Receptor from Porcine Heart

Abstract

Protection of the myocardium from ischemic injury by prostacyclins and PGE₁ has been repeatedly reported. Part of the antiischemic effects may be mediated by cardiac sarcolemmal EP₃-receptors which have been characterized earlier in this laboratory by ligand binding studies. Of particular notice is the presence of multiple EP₃ isoforms, e. g. in mouse, cattle and man which are generated by differential splicing. Here we describe the cloning and subtype characterization of the porcine EP₃-receptor. We obtained two different isoforms of the receptor by screening a porcine heart cDNA library. The predicted receptor proteins consisted of 395 and 387 amino acids, respectively, with differences only in the C-terminal part of the protein. Several overlapping genomic clones were obtained from a porcine genomic library. Sequence comparison of the cDNA with the genomic DNA indicated that an intron was located in the region corresponding to the sixth transmembrane domain of the receptor protein. This position exactly matches the corresponding intron in the human TXA₂-, the human PGD₂- and the mouse PGE-receptor (EP₁-subtype). In further experiments, we expressed the porcine EP₃-receptor in COS-7 cells and performed ligand-binding-studies. In addition the expression of the porcine EP₃-receptor mRNA in different tissues was demonstrated by RT-PCR.