Optical biosensor differentiates signaling of endogenous PAR(1 )and PAR(2 )in A431 cells

BACKGROUND: Protease activated receptors (PARs) consist of a family of four G protein-coupled receptors. Many types of cells express several PARs, whose physiological significance is mostly unknown. RESULTS: Here, we show that non-invasive resonant waveguide grating (RWG) biosensor differentiates signaling of endogenous protease activated receptor subtype 1 (PAR(1)) and 2 (PAR(2)) in human epidermoid carcinoma A431 cells. The biosensor directly measures dynamic mass redistribution (DMR) resulted from ligand-induced receptor activation in adherent cells. In A431, both PAR(1 )and PAR(2 )agonists, but neither PAR(3 )nor PAR(4 )agonists, trigger dose-dependent Ca(2+ )mobilization as well as G(q)-type DMR signals. Both Ca(2+ )flux and DMR signals display comparable desensitization patterns upon repeated stimulation with different combinations of agonists. However, PAR(1 )and PAR(2 )exhibit distinct kinetics of receptor re-sensitization. Furthermore, both trypsin- and thrombin-induced Ca(2+ )flux signals show almost identical dependence on cell surface cholesterol level, but their corresponding DMR signals present different sensitivities. CONCLUSION: Optical biosensor provides an alternative readout for examining receptor activation under physiologically relevant conditions, and differentiates the signaling of endogenous PAR(1 )and PAR(2 )in A431.