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 (PAR₂F240S) characterized by a phenylalanine to serine mutation at residue 240 within extracellular loop 2, with allelic frequencies of 0.916 (Phe²⁴⁰) and 0.084 (Ser²⁴⁰) for the wild-type and mutant alleles, respectively. Elevations in intracellular calcium were measured in permanently transfected cell lines expressing the receptors. PAR₂F240S displayed a significant reduction in sensitivity toward trypsin (∼3.7-fold) and the PAR2-activating peptides, SLIGKV-NH₂ (∼2.5-fold) and SLIGRL-NH₂(∼2.8-fold), but an increased sensitivity toward the selective PAR2 agonist,trans-cinnamoyl-LIGRLO-NH₂(∼4-fold). Increased sensitivity was also observed toward the selective PAR-1 agonist, TFLLR-NH₂ (∼7-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 PAR₂F240S agonists. By introducing the F240S mutation into rat PAR2, we observed shifts in agonist potencies that mirrored the human PAR₂F240S, suggesting that Phe²⁴⁰ 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.