title: The Toll-like receptor 2 (TLR2) ligand FSL-1 is internalized via the clathrin-dependent endocytic pathway triggered by CD14 and CD36 but not by TLR2.
abstract: Little is known of how Toll-like receptor (TLR) ligands are processed after recognition by TLRs. This study was therefore designed to investigate how the TLR2 ligand FSL-1 is processed in macrophages after recognition by TLR2. FSL-1 was internalized into the murine macrophage cell line, RAW264.7. Both chlorpromazine and methyl-beta-cyclodextrin, which inhibit clathrin-dependent endocytosis, reduced FSL-1 uptake by RAW264.7 cells in a dose-dependent manner but nystatin, which inhibits caveolae- and lipid raft-dependent endocytosis, did not. FSL-1 was co-localized with clathrin but not with TLR2 in the cytosol of RAW264.7 cells. These results suggest that internalization of FSL-1 is clathrin dependent. In addition, FSL-1 was internalized by peritoneal macrophages from TLR2-deficient mice. FSL-1 was internalized by human embryonic kidney 293 cells transfected with CD14 or CD36 but not by the non-transfected cells. Also, knockdown of CD14 or CD36 in the transfectants reduced FSL-1 uptake. In this study, we suggest that (i) FSL-1 is internalized into macrophages via a clathrin-dependent endocytic pathway, (ii) the FSL-1 uptake by macrophages occurs irrespective of the presence of TLR2, and (iii) CD14 and CD36 are responsible for the internalization of FSL-1.
authors.affiliation: Laboratory of Oral Molecular Microbiology, Department of Oral Pathobiological Science, Hokkaido University Graduate School of Dental Medicine, Japan.
authors.forename: Haque M
authors.initials: HM
authors.lastname: Shamsul
authors.forename: Akira
authors.initials: A
authors.lastname: Hasebe
authors.forename: Mitsuhiro
authors.initials: M
authors.lastname: Iyori
authors.forename: Makoto
authors.initials: M
authors.lastname: Ohtani
authors.forename: Kazuto
authors.initials: K
authors.lastname: Kiura
authors.forename: Diya
authors.initials: D
authors.lastname: Zhang
authors.forename: Yasunori
authors.initials: Y
authors.lastname: Totsuka
authors.forename: Ken-ichiro
authors.initials: K
authors.lastname: Shibata
mesh_terms: D000818:Animals; D018955:CD36 Antigens / Q000235:genetics / Q000276:immunology*; D002460:Cell Line; D002746:Chlorpromazine / Q000494:pharmacology; D002966:Clathrin / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D004075:Diglycerides / Q000276:immunology / Q000378:metabolism / Q000494:pharmacology*; D018492:Dopamine Antagonists / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004705:Endocytosis / Q000187:drug effects / Q000235:genetics / Q000276:immunology*; D006801:Humans; D007476:Ionophores / Q000494:pharmacology; D018950:Lipopolysaccharide Receptors / Q000235:genetics / Q000276:immunology*; D017737:Macrophages, Peritoneal / Q000276:immunology* / Q000378:metabolism; D021962:Membrane Microdomains / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D009761:Nystatin / Q000494:pharmacology; D009842:Oligopeptides / Q000276:immunology / Q000378:metabolism / Q000494:pharmacology*; D051195:Toll-Like Receptor 2 / Q000235:genetics / Q000276:immunology* / Q000378:metabolism
chemical_list: D018955:CD36 Antigens; D002966:Clathrin; D004075:Diglycerides; D018492:Dopamine Antagonists; C460864:FSL-1 lipoprotein, synthetic; D007476:Ionophores; D018950:Lipopolysaccharide Receptors; D009842:Oligopeptides; C495342:TLR2 protein, human; C493486:Tlr2 protein, mouse; D051195:Toll-Like Receptor 2; D009761:Nystatin; D002746:Chlorpromazine