Regions of the mouse CD14 molecule required for toll-like receptor 2- and 4-mediated activation of NF-kappa B.
Regions of mouse CD14 required for Toll-like receptor 2 (TLR2)- and TLR4-mediated activation of NF-kappaB were studied in transiently transfected 293 cells. Wild-type CD14 enhanced lipopolysaccharide (LPS)-induced NF-kappaB-dependent reporter activity in cells expressing TLR4/MD-2, and deletion of amino acid regions 35-44, 144-153, 235-243, and 270-275 impaired the TLR4-mediated activation. Unlike human CD14, mouse CD14 truncated at amino acid 151 lost the activity. Deletion of amino acids 35-44 or 235-243 also abrogated TLR2-mediated activation of NF-kappaB, whereas mutants lacking 144-153 and 270-275 retained the activity. Deletion and alanine substitution experiments revealed that amino acids 151-153 and 273-275 were required for the TLR4-mediated activation. Both deletion mutants lacking amino acids 35-44 and 235-243 and alanine substitution mutants in regions 151-153 and 273-275 were expressed on the cell surface and retained the ability to associate with TLR4. A cross-linking study with photoreactive LPS showed that the labeling intensities to CD14 mutants/TLR4/MD-2 were paralleled by the ability of CD14 mutants to increase TLR4-mediated activation. These results indicate that different regions of mouse CD14 are required for TLR4- and TLR2-mediated activation of NF-kappaB and suggest that amino acids 35-44, 151-153, 235-243, and 273-275 of mouse CD14 play an important role in LPS binding and its transfer to TLR4/MD-2.
Muroi
Masashi
M
Division of Microbiology, National Institute of Health Sciences, 1-18-1 Kamiyoga, Setagaya, Tokyo 158-8501, Japan.
Ohnishi
Takahiro
T
Tanamoto
Ken-Ichi
K
D000818:Animals; D002460:Cell Line; D029721:Drosophila Proteins; D006801:Humans; D018950:Lipopolysaccharide Receptors / Q000737:chemistry* / Q000502:physiology; D008070:Lipopolysaccharides / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000502:physiology*; D051379:Mice; D016328:NF-kappa B / Q000378:metabolism*; D011956:Receptors, Cell Surface / Q000502:physiology*; D013045:Species Specificity; D013329:Structure-Activity Relationship; D051195:Toll-Like Receptor 2; D051197:Toll-Like Receptor 4; D051193:Toll-Like Receptors
D029721:Drosophila Proteins; D018950:Lipopolysaccharide Receptors; D008070:Lipopolysaccharides; D008562:Membrane Glycoproteins; D016328:NF-kappa B; D011956:Receptors, Cell Surface; C495342:TLR2 protein, human; C495345:TLR4 protein, human; D051195:Toll-Like Receptor 2; D051197:Toll-Like Receptor 4; D051193:Toll-Like Receptors