P-selectin glycoprotein ligand-1 mediates rolling of mouse bone marrow-derived mast cells on P-selectin but not efficiently on E-selectin.
It has been shown recently that mast cells play an essential role as a source of tumor necrosis factor-alpha production during neutrophil recruitment to sites of bacterial infection. Increased numbers of mast cells are indeed noted at sites of wound healing and inflammation. These cells are either recruited from the bone marrow or proliferate locally under cytokine stimulation. Little is known about how mast cell progenitors extravasate into tissue. Using antibody-like fusion proteins of mouse E-selectin and P-selectin, we have analyzed the ability of immature mouse bone marrow-derived mast cells (BMMC) to interact with the endothelial selectins. The P-selectin glycoprotein ligand-1 (PSGL-1) was affinity-isolated from detergent extracts of surface biotinylated BMMC with both selectin-IgG fusion proteins. However, only P-selectin-IgG, but not E-selectin-IgG showed significant interaction with intact BMMC as tested by flow cytometry and cell attachment assays with the immobilized fusion proteins under flow and non-flow conditions at physiological shear stress. Thus, in spite of carrying the necessary carbohydrate modifications which enable solubilized PSGL-1 to bind avidly to E-selectin, PSGL-1 on the surface of BMMC is presented in a way that prevents it from interacting efficiently with E-selectin. Affinity-purified rabbit antibodies against mouse PSGL-1 almost completely blocked the interaction of BMMC with P-selectin-IgG in flow cytometry as well as in cell adhesion assays under static and under flow conditions. Our data reveal that PSGL-1 is the major binding site for P-selectin on mouse BMMC progenitors, but does not support efficient interactions with E-selectin.
Steegmaier
M
M
Institute of Cell Biology, ZMBE, University of Münster, Germany.
Blanks
J E
JE
Borges
E
E
Vestweber
D
D
D000818:Animals; D001666:Binding Sites, Antibody; D001853:Bone Marrow / Q000276:immunology; D001854:Bone Marrow Cells; D002448:Cell Adhesion / Q000276:immunology; D019040:E-Selectin / Q000276:immunology* / Q000378:metabolism; D005260:Female; D007074:Immunoglobulin G / Q000378:metabolism; D008024:Ligands; D008407:Mast Cells / Q000276:immunology* / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000276:immunology / Q000302:isolation & purification / Q000502:physiology*; D051379:Mice; D019007:P-Selectin / Q000276:immunology* / Q000378:metabolism; D011233:Precipitin Tests; D011485:Protein Binding / Q000276:immunology; D012995:Solubility
D019040:E-Selectin; D007074:Immunoglobulin G; D008024:Ligands; D008562:Membrane Glycoproteins; D019007:P-Selectin; C084882:P-selectin ligand protein