title: L-selectin from human, but not from mouse neutrophils binds directly to E-selectin.
abstract: L-Selectin on neutrophils as well as inducible E- and P-selectin on endothelium are involved in the recruitment of neutrophils into inflamed tissue. Based on cell attachment assays, L-selectin was suggested to function as a carbohydrate presenting ligand for E- and P-selectin. However, previous affinity isolation experiments with an E-selectin-Ig fusion protein had failed to detect L-selectin among the isolated E-selectin ligands from mouse neutrophils. We show here that L-selectin from human neutrophils, in contrast to mouse neutrophils, can be affinity-isolated as a major ligand from total cell extracts using E-selectin-Ig as affinity probe. Binding of human L-selectin to E-selectin was direct, since purified L-selectin could be reprecipitated with E-selectin-Ig. Recognition of L-selectin was abolished by sialidase-treatment, required Ca2+, and was resistant to treatment with endoglycosidase F. Binding of L-selectin to a P-selectin-Ig fusion protein was not observed. In agreement with the biochemical data, the anti-L-selectin mAb DREG56 inhibited rolling of human neutrophils on immobilized E-selectin-Ig but not on P-selectin-Ig. No such inhibitory effect was seen with the anti-mouse L-selectin mAb MEL14 on mouse neutrophils. Rolling of E-selectin transfectants on purified and immobilized human L-selectin was inhibited by mAb DREG56. We conclude that L-selectin on human neutrophils is a major glycoprotein ligand among very few glycoproteins that can be isolated by an E-selectin affinity matrix. The clear difference between human and mouse L-selectin suggests that E-selectin-binding carbohydrate moieties are attached to different protein scaffolds in different species.
authors.affiliation: Institute of Cell Biology, ZMBE, University of Münster, Germany.
authors.forename: O
authors.initials: O
authors.lastname: Zöllner
authors.forename: M C
authors.initials: MC
authors.lastname: Lenter
authors.forename: J E
authors.initials: JE
authors.lastname: Blanks
authors.forename: E
authors.initials: E
authors.lastname: Borges
authors.forename: M
authors.initials: M
authors.lastname: Steegmaier
authors.forename: H G
authors.initials: HG
authors.lastname: Zerwes
authors.forename: D
authors.initials: D
authors.lastname: Vestweber
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism; D001854:Bone Marrow Cells; D016466:CHO Cells; D006224:Cricetinae; D019040:E-Selectin / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007074:Immunoglobulin G / Q000235:genetics / Q000378:metabolism; D019041:L-Selectin / Q000378:metabolism*; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase / Q000378:metabolism; D051379:Mice; D009504:Neutrophils / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012794:Sialic Acids / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; D019040:E-Selectin; D007074:Immunoglobulin G; D011993:Recombinant Fusion Proteins; D012794:Sialic Acids; D019041:L-Selectin; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase