MUC1 Regulates Epithelial Inflammation and Apoptosis by Polyl:C through Inhibition of Toll/IL-1 Receptor-Domain-Containing Adapter-Inducing IFN-β (TRIF) Recruitment to Toll-like Receptor 3

MUC1/Muc1 (MUC1 in humans, Muc1 in animals) is a membrane-tethered mucin expressed by airway epithelial cells and plays an antiinflammatory role during airway bacterial infection. We previously demonstrated that MUC1/Muc1 is a negative regulator of Toll-like receptor (TLR) inflammatory signaling mediated through the myeloid differentiation primary response gene 88 (MyD88) adaptor protein. In the present study, we determined whether MUC1 regulates MyD88-independent TLR signaling mediated through the TLR3-Toll/IL-1 receptor-domain-containing adapter-inducing IFN-beta (TRIF) pathway in response to poly(I:C). Compared with MUC1/Muc1-expressing controls, cells deficient in MUC1/Muc1 were more prone to poly(I:C)-induced apoptosis; had increased poly(I:C)-driven activation of caspase-3, caspase-8, IFN regulatory factor-3, and NF-kappa B; and displayed heightened IFN-beta gene expression. MUC1 overexpression by these cells had the opposite effects. Reciprocal coimmunoprecipitation experiments established constitutive TLR3/MUC1-CT (cytoplasmic tail) protein interaction in human embryonic kidney (HEK) 293T cells overexpressing the two proteins and in lung epithelial cells expressing the endogenous proteins, the latter of which was confirmed by immunofluorescence colocalization of TLR3 with MUC1-CT. Coimmunoprecipitation studies also revealed that MUC1 overexpression by HEK293T cells reduced poly(I:C)-induced TLR3/TRIF protein interaction. Finally, MUC1 overexpression had no effect on TRIF-dependent auto-activation of TLR3 signaling, suggesting that the site of action of the MUC1-CT in TLR3 signaling is not downstream of TRIF. These data indicate that MUC1-CT counter-regulates apoptotic and inflammatory responses of airway epithelial cell through constitutive association with TLR3, thereby inhibiting poly(I:C)-induced recruitment of TRIF to TLR3.