Quantitative microarray analysis of intact glycolipid-CD1d interaction and correlation with cell-based cytokine production.
The protein CD1d binds self and foreign glycolipids for presentation to CD1-restricted T cells by means of TCR recognition and activates T(H)1 and T(H)2 chemokine release. In this study, a variety of glycolipid ligands were attached to a microarray surface and their binding with dimeric CD1d was investigated. An alpha-galactosyl ceramide (alpha-GalCer) bearing a carbamate group at the 6'-OH position was tethered to the surface, and the dissociation constant on surface with CD1d was determined to reflect the multivalent interaction. Competition assays were then used to determine the dissociation constants (Ki) of new and intact glycolipids in solution. The 4-fluorophenyloctanoyl-modified alpha-GalCer (18) was found to bind most strongly with CD1d (Ki 0.21 microM), 2 orders of magnitude stronger than alpha-GalCer and more than three times more selective than alpha-GalCer for IFN-gamma release from NKT cells. Various alpha-GalCer analogues were analyzed, and the results showed that the binding affinity of glycolipids to CD1d correlates well with IFN-gamma production but poorly with IL-4 secretion by NKT cells, suggesting that tighter binding ligands could bias cytokine release through the T(H)1 pathway.
Liang
Pi-Hui
PH
The Genomics Research Center, Academia Sinica, 128 Academia Road, Section 2, Nankang Dist., Taipei, Taiwan 11529.
Imamura
Masakazu
M
Li
Xiangming
X
Wu
Douglass
D
Fujio
Masakazu
M
Guy
Richard T
RT
Wu
Bing-Ching
BC
Tsuji
Moriya
M
Wong
Chi-Huey
CH
D017951:Antigen Presentation; D018949:Antigens, CD1 / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism; D055610:Antigens, CD1d; D001667:Binding, Competitive; D005699:Galactosylceramides / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D007371:Interferon-gamma / Q000096:biosynthesis* / Q000276:immunology; D015847:Interleukin-4 / Q000096:biosynthesis* / Q000276:immunology; D007694:Killer Cells, Natural / Q000276:immunology; D007700:Kinetics; D046228:Microarray Analysis / Q000379:methods*; D013050:Spectrometry, Fluorescence; D013601:T-Lymphocytes / Q000276:immunology; D018417:Th1 Cells / Q000276:immunology
D018949:Antigens, CD1; D055610:Antigens, CD1d; C526877:CD1D protein, human; D005699:Galactosylceramides; C493154:alpha-galactosylceramide; D015847:Interleukin-4; D007371:Interferon-gamma