title: Myeloid differentiation factor-2 interacts with Lyn kinase and is tyrosine phosphorylated following lipopolysaccharide-induced activation of the TLR4 signaling pathway.
abstract: Stimulation with LPS induces tyrosine phosphorylation of numerous proteins involved in the TLR signaling pathway. In this study, we demonstrated that myeloid differentiation factor-2 (MD-2) is also tyrosine phosphorylated following LPS stimulation. LPS-induced tyrosine phosphorylation of MD-2 is specific; it is blocked by the tyrosine kinase inhibitor, herbimycin A, as well as by an inhibitor of endocytosis, cytochalasin D, suggesting that MD-2 phosphorylation occurs during trafficking of MD-2 and not on the cell surface. Furthermore, we identified two possible phospho-accepting tyrosine residues at positions 22 and 131. Mutant proteins in which these tyrosines were changed to phenylalanine had reduced phosphorylation and significantly diminished ability to activate NF-κB in response to LPS. In addition, MD-2 coprecipitated and colocalized with Lyn kinase, most likely in the endoplasmic reticulum. A Lyn-binding peptide inhibitor abolished MD-2 tyrosine phosphorylation, suggesting that Lyn is a likely candidate to be the kinase required for MD-2 tyrosine phosphorylation. Our study demonstrated that tyrosine phosphorylation of MD-2 is important for signaling following exposure to LPS and underscores the importance of this event in mediating an efficient and prompt immune response.
authors.affiliation: Division of Pediatric Infectious Diseases and Immunology, Cedars-Sinai Medical Center, Los Angeles, CA 90048, USA.
authors.forename: Pearl
authors.initials: P
authors.lastname: Gray
authors.forename: Jargalsaikhan
authors.initials: J
authors.lastname: Dagvadorj
authors.forename: Kathrin S
authors.initials: KS
authors.lastname: Michelsen
authors.forename: Constantinos
authors.initials: C
authors.lastname: Brikos
authors.forename: Altan
authors.initials: A
authors.lastname: Rentsendorj
authors.forename: Terrence
authors.initials: T
authors.lastname: Town
authors.forename: Timothy R
authors.initials: TR
authors.lastname: Crother
authors.forename: Moshe
authors.initials: M
authors.lastname: Arditi
mesh_terms: D004797:Enzyme-Linked Immunosorbent Assay; D057809:HEK293 Cells; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D008070:Lipopolysaccharides / Q000276:immunology; D051218:Lymphocyte Antigen 96 / Q000276:immunology / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D019000:Phosphotyrosine / Q000378:metabolism*; D015398:Signal Transduction / Q000276:immunology*; D051197:Toll-Like Receptor 4 / Q000276:immunology* / Q000378:metabolism; D019061:src-Family Kinases / Q000276:immunology / Q000378:metabolism*
chemical_list: C495377:LY96 protein, human; D008070:Lipopolysaccharides; D051218:Lymphocyte Antigen 96; C495345:TLR4 protein, human; D051197:Toll-Like Receptor 4; D019000:Phosphotyrosine; C060902:lyn protein-tyrosine kinase; D019061:src-Family Kinases