IDENTIFICATION OF A SPECIFIC GLYCOPROTEIN LIGAND FOR P-SELECTIN (CD62) ON MYELOID CELLS

P-selectin (CD62, GMP-140, PADGEM), a Ca2+-dependent lectin on activated platelets and endothelium, functions as a receptor for myeloid cells by interacting with sialylated, fucosylated lactosaminoglycans. P-selectin binds to a limited number of protease-sensitive sites on myeloid cells, but the protein(s) that carry the glycans recognized by P-selectin are unknown. Blotting of neutrophil or HL-60 cell membrane extracts with [I-125]P-selectin and affinity chromatography of [H-3]glucosamine-labeled HL-60 cell extracts were used to identify P-selectin ligands. A major ligand was identified with an almost-equal-to 250,000 M(r) under nonreducing conditions and almost-equal-to 120,000 under reducing conditions. Binding of P-selectin to the ligand was Ca2+ dependent and was blocked by mAbs to P-selectin. Brief sialidase digestion of the ligand increased its apparent molecular weight; however, prolonged digestion abolished binding of P-selectin. Peptide:N-glycosidase F treatment reduced the apparent molecular weight of the ligand by almost-equal-to 3,000 but did not affect P-selectin binding. Western blot and immunodepletion experiments indicated that the ligand was not lamp-1, lamp-2, or L-selectin, which carry sialyl Le(x), nor was it leukosialin, a heavily sialylated glycoprotein of similar molecular weight. The preferential interaction of the ligand with P-selectin suggests that it may play a role in adhesion of myeloid cells to activated platelets and endothelial cells.