Biosynthesis of L-Selectin Ligands:  Sulfation of Sialyl Lewis x-Related Oligosaccharides by a Family of GlcNAc-6-sulfotransferases†

C2GnT-I [core2 β(1,6)-N-acetyglucosaminyltransferase-I] and FucT-VII [α(1,3)-fucosyltransferase-VII] are the key enzymes for the biosynthesis of sialyl-Lewis x determinants on selectin ligands and therefore they represent good drug targets for the treatment of inflammatory disorders and other pathologies involving selectins. In the present study, we examined the importance of N-glycosylation for the ability of C2GnT-I and FucT-VII to generate functional selectin ligands, particularly the PSGL-1 (P-selectin glycoprotein ligand-1). We found that (i) both enzymes have their two N-glycosylation sites occupied, (ii) for C2GnT-I, the N-glycan chain linked to Asn-95 significantly contributes to the synthesis of functional PSGL-1 and is required to localize the enzyme to the cis/medial-Golgi compartment, (iii) all N-glycosylation-deficient proteins of FucT-VII displayr a dramatic impairment of their in vitro enzymatic activities, but retain their ability to fucosylate the core2-modified PSGL-I and to generate P- and L-selectin binding, and (iv) the glycomutants of FucT-VII fail to synthesize sialyl-Lewis x or to generate E-selectin binding unless core2-modified PSGL-1 is present. All combined, our results show a differential functional impact of N-glycosylation on C2GnT-1 and FucT-VII and disclose that a strongly reduced FucT-VII activity retains the ability to fucosylate PSGL-1 on the core2-based binding site(s) for the three selectins.

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Endothelial Sulfated Sialyl Lewis x Glycans, Putative L-Selectin Ligands, Are Preferentially Expressed in Bronchial Asthma but Not in Other Chronic Inflammatory Lung Diseases

American Journal of Respiratory Cell and Molecular Biology ◽

10.1165/ajrcmb.23.4.4113 ◽

2000 ◽

Vol 23 (4) ◽

pp. 492-498 ◽

Cited By ~ 28

Author(s):

Sanna Toppila ◽

Timo Paavonen ◽

Annika Laitinen ◽

Lauri A. Laitinen ◽

Risto Renkonen

Keyword(s):

Bronchial Asthma ◽

Lung Diseases ◽

Sialyl Lewis X ◽

Lewis X ◽

Sialyl Lewis ◽

Selectin Ligands

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Selectin glycoprotein ligands.

Acta Biochimica Polonica ◽

10.18388/abp.2000_4019 ◽

2000 ◽

Vol 47 (2) ◽

pp. 393-412 ◽

Cited By ~ 19

Author(s):

I Zak ◽

E Lewandowska ◽

W Gnyp

Keyword(s):

Biosynthetic Pathway ◽

Structure And Function ◽

Cell Interactions ◽

Sialyl Lewis X ◽

Lewis X ◽

Sialyl Lewis ◽

Selectin Ligands ◽

The Common ◽

And Function ◽

Lymphatic Organs

Lectin selectins and their counter-receptors participate in discontinuous cell-cell interactions concurrent with leukocyte tethering and rolling on endothelium, which, in consequence, leads to leukocyte penetration to lymphatic organs and generation of inflammation sites. Counter-receptors are glycoproteins in which carbohydrate units, the direct selectin ligands, are built into the polypeptide framework. In this review, the distribution, structure and function of the main ligands and counter-receptors for P-, L- and E-selectins known so far, have been discussed. The common biosynthetic pathway of sialyl-Lewis x and sulpho-sialyl-Lewis x determinants of selectin ligands has been described.

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Sialyl-Lewis x/a -decorated selectin ligands in head and neck tumours

Journal of Cancer Research and Clinical Oncology ◽

10.1007/s004320050318 ◽

1999 ◽

Vol 125 (10) ◽

pp. 569-576 ◽

Cited By ~ 7

Author(s):

Jutta Renkonen ◽

Antti Mäkitie ◽

Timo Paavonen ◽

Risto Renkonen

Keyword(s):

Head And Neck ◽

Sialyl Lewis X ◽

Head And Neck Tumours ◽

Lewis X ◽

Sialyl Lewis ◽

Selectin Ligands

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Discordant expression of selectin ligands and sialyl Lewis x–related epitopes on murine myeloid cells

Blood ◽

10.1182/blood-2002-06-1799 ◽

2002 ◽

Vol 100 (13) ◽

pp. 4485-4494 ◽

Cited By ~ 38

Author(s):

Marcin M. A. Kobzdej ◽

Anne Leppänen ◽

Vishwanath Ramachandran ◽

Richard D. Cummings ◽

Rodger P. McEver

Keyword(s):

Monoclonal Antibodies ◽

Sialic Acid ◽

Fluid Phase ◽

Sialyl Lewis X ◽

Lewis X ◽

Intact Cells ◽

Sialyl Lewis ◽

Selectin Ligands ◽

Low Levels ◽

Myeloid Leukocytes

Murine leukocytes are thought to express α2-3-sialylated and α1-3-fucosylated selectin ligands such as sialyl Lewis x (sLex), although monoclonal antibodies (mAbs) to sLex or Lex reportedly do not bind to murine leukocytes. We observed that P- and E-selectin bound to pronase-sensitive ligands on murine monocytic WEHI-3 cells and murine neutrophils, indicating that the ligands for both selectins are glycoproteins. CSLEX-1, HECA-452, and other widely used mAbs to sLex and Lex did not bind to WEHI-3 cells and bound at very low levels to murine neutrophils. Only the anti-sLex mAbs 2H5 and KM93, which also recognize nonfucosylated glycans, bound to WEHI-3 cells. 2H5 and KM93 bound to pronase-resistant structures, indicating that the mAbs did not identify selectin ligands. Treatment of WEHI-3 cells with glycosidases or chlorate demonstrated that sialic acid modifications, α1-3-galactosylation, or sulfation did not mask epitopes for mAbs to sLex or Lex. Compared to human promyelocytic HL-60 cells, WEHI-3 cells and murine neutrophils expressed low α1-3-fucosyltransferase activities. Consistent with very low endogenous fucosylation, forced fucosylation of intact WEHI-3 cells or murine neutrophils by exogenous α1-3-fucosyltransferase FTVI and GDP-fucose created many new epitopes for anti-sLexmAbs such as HECA-452 and CSLEX-1. Nevertheless, forced fucosylation of intact cells did not significantly augment their ability to bind to fluid-phase P- or E-selectin or to roll on immobilized P- or E-selectin under flow. These data suggest that murine myeloid leukocytes fucosylate only a few specific glycans, which interact preferentially with P- and E-selectin.

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