MD-2 and TLR4 N-linked glycosylations are important for a functional lipopolysaccharide receptor
- PMID: 11706042
- DOI: 10.1074/jbc.M109910200
MD-2 and TLR4 N-linked glycosylations are important for a functional lipopolysaccharide receptor
Abstract
The lipopolysaccharide (LPS) receptor is a multi-protein complex that consists of at least three proteins, CD14, TLR4, and MD-2. Because each of these proteins is glycosylated, we have examined the functional role of N-linked carbohydrates of both MD-2 and TLR4. We demonstrate that MD-2 contains 2 N-glycosylated sites at positions Asn(26) and Asn(114), whereas the amino-terminal ectodomain of human TLR4 contains 9 N-linked glycosylation sites. Site-directed mutagenesis studies showed that cell surface expression of MD-2 did not depend on the presence of either N-linked site, whereas in contrast, TLR4 mutants carrying substitutions in Asn(526) or Asn(575) failed to be transported to the cell surface. Using a UV-activated derivative of Re595 LPS (ASD-Re595 LPS) in cross-linking assays, we demonstrated a critical role of MD-2 and TLR4 carbohydrates in LPS cross-linking to the LPS receptor. The ability of the various glycosylation mutants to support cell activation was also evaluated in transiently transfected HeLa cells. The double mutant of MD-2 failed to support LPS-induced activation of an interleukin-8 (IL-8) promoter-driven luciferase reporter to induce IL-8 secretion or to activate amino-terminal c-Jun kinase (JNK). Similar results were observed with TLR4 mutants lacking three or more N-linked glycosylation sites. Surprisingly, the reduction in activation resulting from expression of the Asn mutants of MD-2 and TLR4 can be partially reversed by co-expression with CD14. This suggests that the functional integrity of the LPS receptor depends both on the surface expression of at least three proteins, CD14, MD-2, and TLR4, and that N-linked sites of both MD-2 and TLR4 are essential in maintaining the functional integrity of this receptor.
Similar articles
-
Endotoxin recognition and signal transduction by the TLR4/MD2-complex.Microbes Infect. 2004 Dec;6(15):1361-7. doi: 10.1016/j.micinf.2004.08.015. Microbes Infect. 2004. PMID: 15596121 Review.
-
Interaction of soluble form of recombinant extracellular TLR4 domain with MD-2 enables lipopolysaccharide binding and attenuates TLR4-mediated signaling.J Immunol. 2004 Dec 1;173(11):6949-54. doi: 10.4049/jimmunol.173.11.6949. J Immunol. 2004. PMID: 15557191
-
Endotoxin recognition molecules, Toll-like receptor 4-MD-2.Semin Immunol. 2004 Feb;16(1):11-6. doi: 10.1016/j.smim.2003.10.007. Semin Immunol. 2004. PMID: 14751758 Review.
-
N-linked glycosylations at Asn(26) and Asn(114) of human MD-2 are required for toll-like receptor 4-mediated activation of NF-kappaB by lipopolysaccharide.J Immunol. 2001 Sep 15;167(6):3354-9. doi: 10.4049/jimmunol.167.6.3354. J Immunol. 2001. PMID: 11544325
-
Lipopolysaccharide is in close proximity to each of the proteins in its membrane receptor complex. transfer from CD14 to TLR4 and MD-2.J Biol Chem. 2001 Jun 15;276(24):21129-35. doi: 10.1074/jbc.M009164200. Epub 2001 Mar 26. J Biol Chem. 2001. PMID: 11274165
Cited by
-
Revisiting the immunopathology of congenital disorders of glycosylation: an updated review.Front Immunol. 2024 Mar 14;15:1350101. doi: 10.3389/fimmu.2024.1350101. eCollection 2024. Front Immunol. 2024. PMID: 38550576 Free PMC article. Review.
-
A pleiotropic role of sialidase in the pathogenicity of Porphyromonas gingivalis.Infect Immun. 2024 Mar 12;92(3):e0034423. doi: 10.1128/iai.00344-23. Epub 2024 Feb 20. Infect Immun. 2024. PMID: 38376159
-
Core fucosylation and its roles in gastrointestinal glycoimmunology.World J Gastrointest Oncol. 2023 Jul 15;15(7):1119-1134. doi: 10.4251/wjgo.v15.i7.1119. World J Gastrointest Oncol. 2023. PMID: 37546555 Free PMC article. Review.
-
Anterograde trafficking of Toll-like receptors requires the cargo sorting adaptors TMED-2 and 7.Traffic. 2023 Nov;24(11):508-521. doi: 10.1111/tra.12912. Epub 2023 Jul 26. Traffic. 2023. PMID: 37491993 Free PMC article.
-
Exploration of the Molecular Basis of Forsythia Fruit in the Prevention and Treatment of Cholestatic Liver Injury through Network Pharmacology and Molecular Docking.Nutrients. 2023 Apr 25;15(9):2065. doi: 10.3390/nu15092065. Nutrients. 2023. PMID: 37432229 Free PMC article.
Publication types
MeSH terms
Substances
Grants and funding
LinkOut - more resources
Full Text Sources
Other Literature Sources
Molecular Biology Databases
Research Materials
Miscellaneous