title: Surface-expressed TLR6 participates in the recognition of diacylated lipopeptide and peptidoglycan in human cells.
abstract: Recognition of microbial components by TLR2 requires cooperation with other TLRs. TLR6 has been shown to be required for the recognition of diacylated lipoproteins and lipopeptides derived from mycoplasma and to activate the NF-kappaB signaling cascade in conjunction with TLR2. Human TLR2 is expressed on the cell surface in a variety of cells, including monocytes, neutrophils, and monocyte-derived, immature dendritic cells (iDCs), whereas the expression profile of TLR6 in human cells remains obscure. In this study we produced a function-blocking mAb against human TLR6 and analyzed TLR6 expression in human blood cells and cell lines and its participation in ligand recognition. TLR6 was expressed, although at a lower level than TLR2, on the cell surface in monocytes, monocyte-derived iDCs, and neutrophils, but not on B, T, or NK cells. Confocal microscopic analysis revealed that TLR6 was colocalized with TLR2 at the plasma membrane of monocytes. Importantly, TLR2/6 signaling did not require endosomal maturation, and anti-TLR6 mAb inhibited cytokine production in monocytes and iDCs stimulated with synthetic macrophage-activating lipopeptide-2 or peptidoglycan, indicating that TLR6 recognized diacylated lipopeptide and peptidoglycan at the cell surface. In addition, TLR2 mutants C30S and C36S (Cys(30) and Cys(36) in TLR2 were substituted with Ser), which were expressed intracellularly in HEK293 cells, failed to induce NF-kappaB activation upon macrophage-activating lipopeptide-2 stimulation even in the presence of TLR6. Thus, coexpression of TLR2 and TLR6 at the cell surface is crucial for recognition of diacylated lipopeptide and peptidoglycan and subsequent cellular activation in human cells.
authors.affiliation: Department of Immunology, Osaka Medical Center for Cancer and Cardiovascular Diseases, Osaka, Japan.
authors.forename: Yoshiya
authors.initials: Y
authors.lastname: Nakao
authors.forename: Kenji
authors.initials: K
authors.lastname: Funami
authors.forename: Satomi
authors.initials: S
authors.lastname: Kikkawa
authors.forename: Mitsue
authors.initials: M
authors.lastname: Taniguchi
authors.forename: Miyuki
authors.initials: M
authors.lastname: Nishiguchi
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Fukumori
authors.forename: Tsukasa
authors.initials: T
authors.lastname: Seya
authors.forename: Misako
authors.initials: M
authors.lastname: Matsumoto
mesh_terms: D000215:Acylation; D000818:Animals; D019138:Antibodies, Blocking / Q000494:pharmacology; D000911:Antibodies, Monoclonal / Q000378:metabolism; D002460:Cell Line; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000276:immunology / Q000378:metabolism; D016207:Cytokines / Q000037:antagonists & inhibitors / Q000096:biosynthesis; D003713:Dendritic Cells / Q000276:immunology / Q000378:metabolism; D006801:Humans; D008024:Ligands; D055666:Lipopeptides; D008074:Lipoproteins / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000096:biosynthesis* / Q000097:blood / Q000276:immunology / Q000378:metabolism; D051379:Mice; D009000:Monocytes / Q000276:immunology / Q000378:metabolism; D009504:Neutrophils / Q000276:immunology / Q000378:metabolism; D009842:Oligopeptides / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000494:pharmacology; D010457:Peptidoglycan / Q000378:metabolism* / Q000494:pharmacology; D011956:Receptors, Cell Surface / Q000096:biosynthesis* / Q000097:blood / Q000276:immunology / Q000378:metabolism; D051195:Toll-Like Receptor 2; D051216:Toll-Like Receptor 6; D051193:Toll-Like Receptors; D014162:Transfection
chemical_list: D019138:Antibodies, Blocking; D000911:Antibodies, Monoclonal; D016207:Cytokines; D008024:Ligands; D055666:Lipopeptides; D008074:Lipoproteins; D008562:Membrane Glycoproteins; D009842:Oligopeptides; D010457:Peptidoglycan; D011956:Receptors, Cell Surface; C495342:TLR2 protein, human; C495368:TLR6 protein, human; C495369:Tlr6 protein, mouse; D051195:Toll-Like Receptor 2; D051216:Toll-Like Receptor 6; D051193:Toll-Like Receptors; C106335:macrophage stimulatory lipopeptide 2