Association of Cell Surface Mucins with Galectin-3 Contributes to the Ocular Surface Epithelial Barrier

• DOI: 10.1074/jbc.m109.033332
• Publication Date: 2009-06-26T00:12:43Z
• Authors: Pablo Argüeso; Ana Guzman-Aranguez; Flavio Mantelli; Zhiyi Cao; Jessica Ricciuto; Noorjahan Panjwani
• Keywords: Radioimmunoprecipitation Assay; 0303 health sciences; Mucous Membrane; Galectin 3; Blotting, Western; Mucin-1; Mucins; Cell Polarity; Membrane Proteins; Epithelial Cells; In Vitro Techniques; Eye; Galactosyltransferases; Chromatography, Affinity; Cell Line; 03 medical and health sciences; CA-125 Antigen; Humans; Biotinylation; Protein Binding

Maintenance of an intact mucosal barrier is critical to preventing damage to and infection of wet-surfaced epithelia. The mechanism of defense has been the subject of much investigation, and there is evidence now implicating O-glycosylated mucins on the epithelial cell surface. Here we investigate a new role for the carbohydrate-binding protein galectin-3 in stabilizing mucosal barriers through its interaction with mucins on the apical glycocalyx. Using the surface of the eye as a model system, we found that galectin-3 colocalized with two distinct membrane-associated mucins, MUC1 and MUC16, on the apical surface of epithelial cells and that both mucins bound to galectin-3 affinity columns in a galactose-dependent manner. Abrogation of the mucin-galectin interaction in four different mucosal epithelial cell types using competitive carbohydrate inhibitors of galectin binding, beta-lactose and modified citrus pectin, resulted in decreased levels of galectin-3 on the cell surface with concomitant loss of barrier function, as indicated by increased permeability to rose bengal diagnostic dye. Similarly, down-regulation of mucin O-glycosylation using a stable tetracycline-inducible RNA interfering system to knockdown c1galt1 (T-synthase), a critical galactosyltransferase required for the synthesis of core 1 O-glycans, resulted in decreased cell surface O-glycosylation, reduced cell surface galectin-3, and increased epithelial permeability. Taken together, these results suggest that galectin-3 plays a key role in maintaining mucosal barrier function through carbohydrate-dependent interactions with cell surface mucins.