Sap6 Initiates Oral Mucosal Inflammation the Protease Activated Receptor PAR2

Sap6, a secreted aspartyl protease (Sap), contributes to fungal virulence in oral candidiasis. Beside its protease activity, Sap6 contains RGD (RGDRGD) motif required for its binding to host integrins. Sap6 activates immune cells to induce proinflammatory cytokines, although its ability to interact and activate human oral epithelial cells (OECs) remain unknown. Addition of purified recombinant Sap6 (rSap6) to OECs resulted in production of IL-1β and IL-8 cytokines similar to live hyphal . OECs exposed to rSap6 showed phosphorylation of p38 and MKP1 and expression of c-Fos not found with , heat-inactivated Sap6, or rSap6 . Heat inactivated rSap6 was able to induce IL-1β but not IL-8 in OECs, while rSap6 induced IL-8 but not IL-1β suggesting parallel signaling pathways. hyphae increased surface expression of Protease Activated Receptors , and , while rSap6 increased expression exclusively. Pretreatment of OECs with a PAR2 antagonist blocked rSap6-induced p38 MAPK signaling and IL-8 release, while rSap6 had reduced MKP1 signaling and IL-1β release independent from PAR2. OECs exposed to rSap6 exhibited loss of barrier function as measured by TEER and reduction in levels of E-cadherin and occludin junctional proteins that was prevented by pretreating OECs with a PAR2 antagonist. OECs treated with PAR2 antagonist also showed reduced rSap6-mediated invasion by cells. Thus, Sap6 may initiate OEC responses mediated both through protease activation of PAR2 and by its RGD domain. This novel role of PAR2 suggests new drug targets to block oral infection.