Open AccessSecreted MD-2 is a large polymeric protein that efficiently confers lipopolysaccharide sensitivity to Toll-like receptor 4
Author(s) -
Alberto Visintin,
Alessandra Mazzoni,
Jessica A. Spitzer,
David M. Segal
Publication year - 2001
Publication title -
proceedings of the national academy of sciences of the united states of america
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.011
H-Index - 771
eISSN - 1091-6490
pISSN - 0027-8424
DOI - 10.1073/pnas.211445098
Subject(s) - endoplasmic reticulum , toll like receptor , tlr4 , receptor , secretion , lipopolysaccharide , golgi apparatus , transfection , microbiology and biotechnology , hek 293 cells , extracellular , transmembrane protein , transmembrane domain , epitope , biology , chemistry , biochemistry , antibody , innate immune system , gene , immunology
Toll-like receptor 4 (TLR4), the principal signaling receptor for lipopolysaccharide (LPS) in mammals, requires the binding of MD-2 to its extracellular domain for maximal responsiveness. MD-2 contains a leader sequence but lacks a transmembrane domain, and we asked whether it is secreted into the medium as an active protein. As a source of secreted MD-2 (sMD-2), we used culture supernatants from cells stably transduced with epitope-tagged human MD-2. We show that sMD-2 exists as a heterogeneous collection of large disulfide-linked oligomers formed from stable dimeric subunits and that concentrations of sMD-2 as low as 50 pM enhance the responsiveness of TLR4 reporter cells to LPS. An MD-2-like activity is also released by monocyte-derived dendritic cells from normal donors. When coexpressed, TLR4 indiscriminately associates in the endoplasmic reticulum/cis Golgi with different-sized oligomers of MD-2, and excess MD-2 is secreted into the medium. We conclude that normal and transfected cells secrete a soluble form of MD-2 that binds with high affinity to TLR4 and that could play a role in regulating responses to LPS and other pathogen-derived substances in vivo.