P-SELECTIN MEDIATES ADHESION OF THE HUMAN-MELANOMA CELL-LINE NKI-4 - IDENTIFICATION OF GLYCOPROTEIN LIGANDS

Activated endothelial cells and stimulated platelets express the cell adhesion molecule P-selectin (CD62P), which mediates adhesion to vario us leukocytes and certain types of cancer cells. In this study, we sho w Ca2+-dependent binding of P-selectin to NKI-4 cells, a cell line der ived from a human melanoma. The binding is inhibited by P7 (a leukocyt e adhesion blocking mAb against P-selectin), but not by PL5 (a leukocy te adhesion blocking mAb against P-selectin glycoprotein ligand-1; PSG L-1). Further, expression of PSGL-1 could not be detected on NKI-4 cel ls by either PL5 mAb or an Ab against a synthetic peptide correspondin g to a portion of the PSGL-1 sequence. P-selectin affinity chromatogra phy of lysates from in vivo [H-3]-glucosamine-labeled NKI-4 cells resu lted in the isolation of three glycoproteins, with apparent molecular masses of similar to 250, similar to 110, and similar to 100 kDa under reducing conditions and similar to 230, similar to 105, and similar t o 85 kDa under nonreducing conditions. These molecules could be precip itated by P-selectin, but not by E-selectin. EDTA and the P7 mAb. but not the PL5 mAb, inhibited the binding of P-selectin to the purified l igands. Surprisingly, we found that sodium chlorate, a sulfation inhib itor, did not inhibit the binding of P-selectin to NKI-4 cells and tha t [S-35]-sulfate did not label the NKI-4 cell ligands. We conclude tha t P-selectin-dependent adhesion of the human melanoma cell line NKI-4 is mediated by a novel class of glycoprotein ligands.