title: The amino-terminal region of toll-like receptor 4 is essential for binding to MD-2 and receptor translocation to the cell surface.
abstract: Toll-like receptor 4 (TLR4) and MD-2 are pivotal components that elicit inflammatory responses to lipopolysaccharide (LPS). They have been shown to form a physical complex on the cell surface that responds directly to LPS. However, the functional region of TLR4 required for association with MD-2 and LPS responsiveness is poorly understood. To identify the region of TLR4, we created a series of mutants with deletions in the extracellular domain and examined their activities in human embryonic kidney 293 cells. A mutant with a 317-amino acid deletion from the membrane proximal region of TLR4 was capable of associating with MD-2, while only a 9-amino acid truncation of the N terminus severely impaired the interaction. The association between the two molecules was well correlated with TLR4 maturation into an endoglycosidase H-resistant form and the cell surface expression. Mouse MD-2 bound to human TLR4, but its activity to facilitate the cell surface expression of TLR4 and confer LPS responsiveness was much weaker than that of human MD-2, indicating species specificity. A chimeric receptor composed of the N-terminal region of human TLR4 and the adjacent region of mouse TLR4 showed preference for human MD-2 in its transport to the cell surface and responsiveness to LPS. Taken together, the N-terminal region of TLR4 is essential for association with MD-2, which is required for the cell surface expression and hence the responsiveness to LPS.
authors.affiliation: Department of Molecular and Cellular Biochemistry, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Takeshi
authors.initials: T
authors.lastname: Fujimoto
authors.forename: Soh
authors.initials: S
authors.lastname: Yamazaki
authors.forename: Akiko
authors.initials: A
authors.lastname: Eto-Kimura
authors.forename: Koichiro
authors.initials: K
authors.lastname: Takeshige
authors.forename: Tatsushi
authors.initials: T
authors.lastname: Muta
mesh_terms: D000818:Animals; D000950:Antigens, Ly / Q000235:genetics / Q000378:metabolism*; D000954:Antigens, Surface / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism*; D006801:Humans; D008070:Lipopolysaccharides / Q000276:immunology*; D051218:Lymphocyte Antigen 96; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology / Q000378:metabolism*; D051379:Mice; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021381:Protein Transport / Q000502:physiology; D011956:Receptors, Cell Surface / Q000737:chemistry / Q000235:genetics* / Q000276:immunology / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology; D051197:Toll-Like Receptor 4; D051193:Toll-Like Receptors
chemical_list: D000950:Antigens, Ly; D000954:Antigens, Surface; C495377:LY96 protein, human; D008070:Lipopolysaccharides; C411728:Ly96 protein, mouse; D051218:Lymphocyte Antigen 96; D008562:Membrane Glycoproteins; D011956:Receptors, Cell Surface; D011993:Recombinant Fusion Proteins; C495345:TLR4 protein, human; D051197:Toll-Like Receptor 4; D051193:Toll-Like Receptors