Natural ligands of the B cell adhesion molecule CD22β can be masked by 9-O-Acetylation of sialic acids

Abstract

CD22β is a B cell-restricted phospho-protein expressed on the surface of mature resting B cells. It mediates interactions with other cells partly or exclusively via recognition of α2-6-linked sialic acids on glycoconjugates. The sialylated N-linked oligosaccharides recognized best by CD22β are common to many glycoproteins, suggesting that additional regulatory mechanisms may exist. Since the exocyclic side chain of sialic acid is required for recognition, we explored the effects of a naturally occurring modification of the side chain, 9-O-acetylation. Semisynthetic N-linked oligosaccharides terminating with 9-O-acetylated, α2-6-linked sialic acids showed markedly reduced binding to CD22β relative to their non-O-acetylated counterparts. Murine lymphoid cells were probed for natural CD22β ligands that might be O-acetylated using recombinant soluble forms of CD22β (CD22βRg) and influenza C esterase (CHE-Fc, which specifically removes 9-O-acetyl esters from sialic acids). By flow cytometry analysis, CD22βRg binding to splenic B cells and a subset of T cells was increased by pretreatment with CHE-Fc, indicating that some potential CD22β ligands are naturally "masked" by 9-O-acetylation. Unmasking of these CD22β ligands by removal of 9-O-acetyl esters from intact splenocytes substantially increases their CD22β-dependent adhesion in an in vitro adhesion assay. Probing of murine lymphoid tissue sections by CD22βRg and CHE-Fc treatment demonstrates regionally restricted and differentially expressed patterns of distribution between masked and unmasked ligands. For example, lymph node-associated follicular B cells express high levels of CD22β ligands, none of which are masked by 9-O-acetylation. In contrast, the ligands on lymph node-associated dendritic cells are almost completely masked by 9-O-acetylation, suggesting that masking may regulate interactions between CD22β-positive B cells and dendritic cells. In the thymus, only medullary cells express CD22β ligands, and a significant portion of these are masked by 9-O-acetylation, particularly at the cortical-medullary junction. Thus, 9-O-acetylation of sialic acids on immune cells is in a position to negatively regulate CD22β adhesion events in a manner depending on both cell type and tissue localization.