Sialyl Lewis(x)-mediated, PSGL-1-independent rolling adhesion on P-selectin.
Selectin-mediated cell adhesion is an essential component of the inflammatory response. In an attempt to unambiguously identify molecular features of ligands that are necessary to support rolling adhesion on P-selectin, we have used a reconstituted ("cell-free") system in which ligand-coated beads are perfused over soluble P-selectin surfaces. We find that beads coated with the saccharides sialyl Lewis(x) (sLe(x)), sialyl Lewis(a) (sLe(a)), and sulfated Lewis(x) (HSO(3)Le(x) support rolling adhesion on P-selectin surfaces. Although it has been suggested that glycosylation and sulfation of P-selectin glycoprotein ligand-1 (PSGL-1) is required for high-affinity binding and rolling on P-selectin, our findings indicate that sulfation of N-terminal tyrosine residues is not required for binding or rolling. However, beads coated with a tyrosine-sulfated, sLe(x)-modified, PSGL-1-Fc chimera support slower rolling on P-selectin than beads coated with sLe(x) alone, suggesting that sulfation improves rolling adhesion by modulating binding to P-selectin. In addition, we find it is not necessary that P-selectin carbohydrate ligands be multivalent for robust rolling to occur. Our results demonstrate that beads coated with monovalent sLe(x), exhibiting a more sparse distribution of carbohydrate than a similar amount of the multivalent form, are sufficient to yield rolling adhesion. The relative abilities of various ligands to support rolling on P-selectin are quantitatively examined among themselves and in comparison to human neutrophils. Using stop-time distributions, rolling dynamics at video frame rate resolution, and the average and variance of the rolling velocity, we find that P-selectin ligands display the following quantitative trend, in order of decreasing ability to support rolling adhesion on P-selectin: PSGL-1-Fc > sLe(a) approximately sLe(x) > HSO(3)Le(x).
Department of Chemical Engineering and Institute for Medicine and Engineering, University of Pennsylvania, Philadelphia, Pennsylvania 19107, USA.
S D
SD
Rodgers
R T
RT
Camphausen
D A
DA
Hammer
D000328:Adult; D000818:Animals; D016466:CHO Cells; D002448:Cell Adhesion; D006224:Cricetinae; D006801:Humans; D066298:In Vitro Techniques; D016256:Lewis X Antigen / Q000737:chemistry; D008562:Membrane Glycoproteins / Q000737:chemistry* / Q000502:physiology; D009504:Neutrophils / Q000502:physiology*; D009844:Oligosaccharides / Q000737:chemistry*; D019007:P-Selectin / Q000737:chemistry* / Q000502:physiology; D011994:Recombinant Proteins / Q000737:chemistry; D000080506:Sialyl Lewis X Antigen; D013314:Stress, Mechanical; D014162:Transfection
D016256:Lewis X Antigen; D008562:Membrane Glycoproteins; D009844:Oligosaccharides; D019007:P-Selectin; C084882:P-selectin ligand protein; D011994:Recombinant Proteins; D000080506:Sialyl Lewis X Antigen