A polymorphic protease-activated receptor 2 (PAR2) displaying reduced sensitivity to trypsin and differential responses to PAR agonists

Protease-activated receptor 2 (PAR2) is a trypsin-activated member of a family of G-protein-coupled PARs. We have identified a polymorphic form of human PARS (PAR(2)F240S) characterized by a phenylalanine to serine mutation at residue 240 within extracellular loop 2, with allelic frequencies of 0.916 (Phe(240)) and 0.084 (Ser(240)) for the wild-type and mutant alleles, respectively. Elevations in intracellular calcium were measured in permanently transfected cell lines expressing the receptors. PAR(2)F240S displayed a significant reduction in sensitivity toward trypsin (similar to3.7-fold) and the PARS-activating peptides, SLIGKV-NH2 (similar to2.5-fold) and SLIGRL-NH2 (similar to2.8-fold), but an increased sensitivity toward the selective PARS agonist, trans- cinnamoyl-LIGRLO-NH2 (similar to4-fold). Increased sensitivity was also observed toward the selective PAR-1 agonist, TFLLR-NH2 (similar to7-fold), but not to other PAR-1 agonists tested. Furthermore, we found that TLIGRL-NH2 and a PAR4-derived peptide, trans-cinnamoyl-YPGKF-NH2, were selective PAR(2)F240S agonists. By introducing the F240S mutation into rat PARS, we observed shifts in agonist potencies that mirrored the human PAR,F240S, suggesting that Phe240 is involved in determining agonist specificity of PARS. Finally, differences in receptor signaling were paralleled in a cell growth assay. We suggest that the distinct pharmacological profile induced by this polymorphism will have important implications for the design of PAR-targeted agonists/antagonists and may contribute to, or be predictive of, an inflammatory disease.