The monoclonal antibody CHO-131 binds to a core 2 O-glycan terminated with sialyl-Lewis x, which is a functional glycan ligand for P-selectin

Bruce Walcheck, Anne Leppanen, Richard D. Cummings, Randall N. Knibbs, Lloyd M. Stoolman, Shelia R. Alexander, Polly E. Mattila, Rodger P. McEver

Research output: Contribution to journalArticle

42 Scopus citations

Abstract

Core 2 O-glycans terminated with sialyl-Lewis x (sLe x) are functionally important oligosaccharides that endow particular macromolecules with high-affinity glycan ligands for the selectin family. To date, antibodies that recognize these structures on leukocytes have not been described. We characterize such a monoclonal antibody (mAb) here (CHO-131). The binding specificity of CHO-131 was directly examined by means of synthetic glycopeptides containing precise O-glycan structures. CHO-131 bound to sLe x extended from a core 2 branch (C2-O- sLe x), but CHO-131 demonstrated no reactivity if this oligosaccharide lacked fucose or if sLe x was extended from a core 1 branch. Using transfected cell lines, we found that CHO-131 binding required the functional activity of the glycosyltransferases α2,3-sialyltransferase, α1,3-fucosyltransferase-VII, and core 2 β1,6 N-acetylglucosaminyltransferase (C2GnT). The C2-O-sLe x motif occurs primarily on sialomucins and has been directly shown to contribute to high-affinity P-selectin glycoprotein ligand-1 binding by P-selectin. Indeed, CHO-131 staining of neutrophils was diminished following sialomucin removal by O-glycoprotease, and its reactivity with transfected hematopoietic cell lines correlated with the expression of P-selectin ligands. CHO-131 also stained a small population of lymphocytes that were primarily CD3 +, CD4 +, and CD45RO + and represented a subset (37.8% ± 18.3%) of cutaneous lymphocyte-associated antigen (CLA) T cells, distinguished by the mAb HECA-452, which detects sLe x-related glycans. Unlike anti-sLe x mAbs, CHO-131 binding also indicates C2GnT activity and demonstrates that CLA T cells are heterogeneous based on the glycan structures they synthesize. These findings support evidence that differential C2GnT activity results in T-cell subsets that express ligands for E-selectin, P-selectin, or both.

Original languageEnglish (US)
Pages (from-to)4063-4069
Number of pages7
JournalBlood
Volume99
Issue number11
DOIs
StatePublished - Jun 1 2002

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