Toll-like Receptor 5 (TLR5) is expressed on the cell surface and specifically senses bacterial flagellin, a major structural protein in flagella of motile bacteria. Flagellin-stimulated TLR5 recruits the adaptor protein MyD88 and activates the NF-kB pathway, resulting in the production of proinflammatory cytokines such as IL-6 and TNF-α. TLR5 also delivers flagellin into endolysosomes via receptor internalization for presentation of flagellin-derived peptides on MHC class II and the subsequent activation of cognate CD4+ T cells, which is required for production of flagellin-specific antibodies.
We previously reported that the cell surface localization of TLR5 depends on UNC93B1, whose function was thought to be limited to delivery of nucleotide-sensing TLRs, including TLR3, 7, 8, and 9, into endolysosomes. Because all other UNC93B1-dependent TLRs mediate the type I interferon (IFN) production and some cell-surface TLRs are able to initiate signaling for type I IFN induction upon internalization into endolysosomes, we tested whether flagellin-mediated TLR5 activation stimulates secretion of type I IFNs. In both human and mouse cell lines, we found that flagellin induced IFN-β secretion and type I IFN receptor signaling in a TLR5-dependent manner. When injected into mice, flagellin rapidly induced IFN-β expression in monocytes and neutrophils, resulting in elevated serum IFN-β levels. TLR5-mediated IFN-β production depended on MyD88, but not TRIF, and required the receptor endocytosis and endosomal signaling. Furthermore, induction of fecal anti-flagellin antibodies by flagellin immunization was abrogated in the type I IFN receptor-deficient mice, indicating that TLR5-mediated secretion of type I IFN is required for production of mucosal anti-flagellin antibodies.