Acquisition of P-selectin binding activity by en bloc transfer of sulfo Le(x) trisaccharide to the cell surface: Comparison to a sialyl Le(x) tetrasaccharide transferred on the cell surface

Sialyl Le(x), NeuNAc alpha 2 --> 3Gal beta 1 --> 4(Fuc alpha 1 --> 3)GlcNAc beta --> R, is known to be a ligand for E-selectin in various assays. The sulfated counterpart of sialyl Le(x), sulfo Le(x), (Sulfo --> 3) Gal beta 1 --> 4 (Fuc alpha 1 --> 3) GlcNAc beta --> R, was also shown to be a ligand for E-selectin in solid- phase assays employing immobilized oligosaccharid es, In order to determine whether sulfo Le(x) structure on the cell surface also works as E-selectin or P-selectin ligand, a novel approach for in vit ro transfer of oligosaccharides (S. Tsuboi, Y. Isogai, N. Hada, J. K. King, O. Hindsgaul, and M. Fukuda (1996) J. Biol. Chem. 271, 27213-27216) was ut ilized. A synthetic GDP-fucose harboring sialyl Le(x) or sulfo Le(x) oligos accharide was enzymatically transferred to Chinese hamster ovary (CHO) cell s with a milk fucosyltransferase. The resultant cells, CHO-sialyl Le(x) and CHO-sulfo Le(x) were tested for adhesion to E-selectin IgG or P-selectin I gG: chimeric protein coated on plates. The results indicate that CHO-sialyl Le(x) adhered efficiently to E-selectin, while adhesion of CHO-sulfo Le(x) was very poor despite the fact that near equal number of the ligands had b een attached to the cell surface. In contrast, CHO-sulfo Le(x) adhered effi ciently to P-selectin, while CHO-sialyl Le(x) adhered modestly to P-selecti n. These results demonstrate that sialyl Le(x) and sulfo Le(x) structures o n the cell surface differ substantially in their ability to adhere to E- an d P-selectin. (C) 2000 Academic Press.