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Negative regulation of Toll-like receptor-4 signaling through the binding of glycosylphosphatidylinositol-anchored glycoprotein, CD14, with the sialic acid-binding lectin, CD33.
- Source :
-
The Journal of biological chemistry [J Biol Chem] 2014 Sep 05; Vol. 289 (36), pp. 25341-50. Date of Electronic Publication: 2014 Jul 24. - Publication Year :
- 2014
-
Abstract
- When monocyte-derived immature dendritic cells (imDCs) were stimulated with LPS in the presence of anti-CD33/Siglec-3 mAb, the production of IL-12 and phosphorylation of NF-κB decreased significantly. The cell surface proteins of imDCs were chemically cross-linked, and CD33-linked proteins were analyzed by SDS-PAGE and immunoblotting. It was CD14 that was found to be cross-linked with CD33. A proximity ligation assay also indicated that CD33 was colocalized with CD14 on the cell surface of imDCs. Sialic acid-dependent binding of CD33 to CD14 was confirmed by a plate assay using recombinant CD33 and CD14. Three types of cells (HEK293T cells expressing the LPS receptor complex (Toll-like receptor (TLR) cells), and the LPS receptor complex plus either wild-type CD33 (TLR/CD33WT cells) or mutated CD33 without sialic acid-binding activity (TLR/CD33RA cells)) were prepared, and then the binding and uptake of LPS were investigated. Although the level of LPS bound on the cell surface was similar among these cells, the uptake of LPS was reduced in TLR/CD33WT cells. A higher level of CD14-bound LPS and a lower level of TLR4-bound LPS were detected in TLR/CD33WT cells compared with the other two cell types, probably due to reduced presentation of LPS from CD14 to TLR4. Phosphorylation of NF-κB after stimulation with LPS was also compared. Wild-type CD33 but not mutated CD33 significantly reduced the phosphorylation of NF-κB. These results suggest that CD14 is an endogenous ligand for CD33 and that ligation of CD33 with CD14 modulates with the presentation of LPS from CD14 to TLR4, leading to down-regulation of TLR4-mediated signaling.<br /> (© 2014 by The American Society for Biochemistry and Molecular Biology, Inc.)
- Subjects :
- Antibodies, Monoclonal immunology
Antibodies, Monoclonal pharmacology
Blotting, Western
Cell Membrane metabolism
Dendritic Cells drug effects
Dendritic Cells metabolism
HEK293 Cells
Humans
Interleukin-2 metabolism
Ligands
Lipopolysaccharide Receptors genetics
Lipopolysaccharides metabolism
Microscopy, Confocal
Models, Biological
Mutation
NF-kappa B metabolism
Phosphorylation drug effects
Protein Binding
Sialic Acid Binding Ig-like Lectin 3 genetics
Sialic Acid Binding Ig-like Lectin 3 immunology
Toll-Like Receptor 4 genetics
Lipopolysaccharide Receptors metabolism
Sialic Acid Binding Ig-like Lectin 3 metabolism
Signal Transduction
Toll-Like Receptor 4 metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 1083-351X
- Volume :
- 289
- Issue :
- 36
- Database :
- MEDLINE
- Journal :
- The Journal of biological chemistry
- Publication Type :
- Academic Journal
- Accession number :
- 25059667
- Full Text :
- https://doi.org/10.1074/jbc.M113.523480