P- and E-selectins recognize sialyl 6-sulfo Lewis X, the recently identified L-selectin ligand

Recently we identified sialyl 6-sulfo Le(x) as a major L-selectin ligand on high endothelial venules of human peripheral lymph nodes. In this study we investigated the ligand activity of sialyl g-sulfo Le(x) to E-and P-select ins and compared it with the binding activity of conventional sialyl Le(x), by using cultured human lymphoid cells expressing both carbohydrate determ inants. The results of the recombinant selectin binding studies and the non static monolayer cell adhesion assays indicated that both sialyl 6-sulfo Le (x) and conventional sialyl Le(x) served as ligand for E-and P-selectins, w hile L-selectin was quite specific to sialyl 6-sulfo Le(x). Anti-PSGL-1 ant ibodies as well as O-sialoglycoprotein endopeptidase treatment almost compl etely abrogated the binding of P-selectin but barely affected the binding o f E-selectin, indicating that these carbohydrate determinants carried by O- glycans of PSGL-1 selectively serves as a ligand for P-selectin, while the ligand for E-selectin is not restricted to PSGL-1 nor to O-sialoglycoprotei n endopeptidase-sensitive glycans, The binding of L-selectin was markedly r educed by O-sialoglyco-protein endopeptidase treatment but only minimally a ffected by anti-PSGL-1 antibodies, indicating that O-glycans carrying sialy l 6-sulfo Le(x) were the major L-selectin ligands, while PSGL-1 was only a minor core protein for L-selectin in these cells. These results indicated t hat each member of the selectin family has a distinct ligand binding specif icity. (C) 2000 Academic Press.