Cathepsin S causes inflammatory pain via biased agonism of PAR2 and TRPV4

Abstract

Serine proteases such as trypsin and mast cell tryptase cleave protease-activated receptor-2 (PAR2) at R36↑S37and reveal a tethered ligand that excites nociceptors, causing neurogenic inflammation and pain. Whether proteases that cleave PAR2at distinct sites are biased agonists that also induce inflammation and pain is unexplored. Cathepsin S (Cat-S) is a lysosomal cysteine protease of antigen-presenting cells that is secreted during inflammation and which retains activity at extracellular pH.We observed that Cat-S cleaved PAR2at E56↑T57, which removed the canonical tethered ligand and prevented trypsin activation. In HEK and KNRK cell lines and in nociceptive neurons of mouse dorsal root ganglia, Cat-S and a decapeptide mimicking the Cat-S-revealed tethered ligand-stimulated PAR2coupling to Gαs and formation of cAMP. In contrast to trypsin, Cat-S did not mobilize intracellular Ca2+, activate ERK1/2, recruit β-arrestins, or induce PAR2endocytosis. Cat-S caused PAR2-dependent activation of transient receptor potential vanilloid 4 (TRPV4) in Xenopus laevis oocytes, HEK cells and nociceptive neurons, and stimulated neuronal hyperexcitability by adenylyl cyclase and protein kinase A-dependent mechanisms. Intraplantar injection of Cat-S caused inflammation and hyperalgesia in mice that was attenuated by PAR2or TRPV4 deletion and adenylyl cyclase inhibition. Cat-S and PAR2antagonists suppressed formalin-induced inflammation and pain, which implicates endogenous Cat-S and PAR2in inflammatory pain. Our results identify Cat-S as a biased agonist of PAR2that causes PAR2- and TRPV4-dependent inflammation and pain. They expand the role of PAR2 as a mediator of protease-driven inflammatory pain.