SIALOSYL-FUCOSYL POLY-LACNAC WITHOUT THE SIALOSYL-LE(X) EPITOPE AS THE PHYSIOLOGICAL MYELOID CELL LIGAND IN E-SELECTIN-DEPENDENT ADHESION -STUDIES UNDER STATIC AND DYNAMIC FLOW CONDITIONS

The majority of E-and P-selectin ligands in leukocytes and myelocytic or monocytic leukemia cells are carried by transmembrane glycoproteins having a tandem repeat mucin-like domain through which O-linked carbo hydrate ligands are carried. However, determination of structure and a dhesive function of carbohydrates in glycoproteins is extremely diffic ult because of the extensive structural heterogeneity and the scarcity of material for functional analysis. We have overcome this difficulty through use of poly-LacNAc gangliosides isolated from a large quantit y of (similar to 1.2 L packed) HL60 cells [Stroud, M. R., Handa, K., S alyan, M. E. K., Ito, K., Levery, S. B., Hakomori, S., Reinhold, B. B. , & Reinhold, V. N. (1996) Biochemistry 35, 758-769, 770-778]. We iden tified two major types of poly-LacNAc gangliosides without the sialosy l-Le(x) epitope as being capable of binding to E-selectin: (i) those h aving a single alpha 1-->3 fucosylation at internal GlcNAcs but not at the penultimate GlcNAc and (ii) those having double alpha 1-->3 fucos ylation at internal GlcNAcs, excluding the penultimate GlcNAc. Ganglio sides from group i above did not show any adhesion under static condit ions, but showed strong adhesion under dynamic flow conditions. Gangli osides from group ii above showed adhesion under both static and dynam ic conditions, as did sialosyl-Le(x) (SLe(x))-containing structures in previous studies. However, SLe(x)-containing poly LacNAc gangliosides are virtually absent or present in only trace quantities in leukocyte s and HL60 cells. Poly-LacNAc gangliosides from groups i and ii above, lacking SLe(x) structure, are the major membrane components of leukoc ytes and HL60 cells. These carbohydrates, bound to lipid or to protein , may therefore be the physiological epitope for E selectin-dependent binding of these cells, particularly under dynamic flow conditions.