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 PAR2 (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 ( approximately 3.7-fold) and the PAR2-activating peptides, SLIGKV-NH(2) ( approximately 2.5-fold) and SLIGRL-NH(2) ( approximately 2.8-fold), but an increased sensitivity toward the selective PAR2 agonist, trans-cinnamoyl-LIGRLO-NH(2) ( approximately 4-fold). Increased sensitivity was also observed toward the selective PAR-1 agonist, TFLLR-NH(2) ( approximately 7-fold), but not to other PAR-1 agonists tested. Furthermore, we found that TLIGRL-NH(2) and a PAR4-derived peptide, trans-cinnamoyl-YPGKF-NH(2), were selective PAR(2)F240S agonists. By introducing the F240S mutation into rat PAR2, we observed shifts in agonist potencies that mirrored the human PAR(2)F240S, suggesting that Phe(240) is involved in determining agonist specificity of PAR2. 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.