Sialylation of N-linked glycans mediates apical delivery of endolyn in MDCK cells via a galectin-9-dependent mechanism

• Published in: Molecular biology of the cell 2012-09, Vol.23 (18), p.3636-3646
• Main Authors: Mo, Di, Costa, Simone A, Ihrke, Gudrun, Youker, Robert T, Pastor-Soler, Nuria, Hughey, Rebecca P, Weisz, Ora A
• Format: Article
• Language: English
• Subjects: Alkaloids - pharmacology; Animals; Dogs; Electrophoresis, Polyacrylamide Gel; Endolyn - genetics; Endolyn - metabolism; Galectins - genetics; Galectins - metabolism; Gene Expression; Kidney - cytology; Kidney - drug effects; Kidney - metabolism; Kidney Tubules - metabolism; Lysosomes - metabolism; Madin Darby Canine Kidney Cells; Microscopy, Confocal; N-Acetylglucosaminyltransferases - genetics; N-Acetylglucosaminyltransferases - metabolism; Polysaccharides - metabolism; Rats; Reverse Transcriptase Polymerase Chain Reaction; RNA Interference; Sialic Acids - metabolism; Sialyltransferases - genetics; Sialyltransferases - metabolism
• ISSN: 1059-1524; 1939-4586
• DOI: 10.1091/mbc.E12-04-0329

The sialomucin endolyn is implicated in adhesion, migration, and differentiation of various cell types. Along rat kidney tubules, endolyn is variously localized to the apical surface and endosomal/lysosomal compartments. Apical delivery of newly synthesized rat endolyn predominates over direct lysosomal delivery in polarized Madin-Darby canine kidney cells. Apical sorting depends on terminal processing of a subset of lumenal N-glycans. Here we dissect the requirements of N-glycan processing for apical targeting and investigate the underlying mechanism. Modulation of glycan branching and subsequent polylactosamine elongation by knockdown of N-acetylglucosaminyltransferase III or V had no effect on apical delivery of endolyn. In contrast, combined but not individual knockdown of sialyltransferases ST3Gal-III, ST3Gal-IV, and ST6Gal-I, which together are responsible for addition of α2,3- and α2,6-linked sialic acids on N-glycans, dramatically decreased endolyn surface polarity. Endolyn synthesized in the presence of kifunensine, which blocks terminal N-glycan processing, reduced its interaction with several recombinant canine galectins, and knockdown of galectin-9 (but not galectin-3, -4, or -8) selectively disrupted endolyn polarity. Our data suggest that sialylation enables recognition of endolyn by galectin-9 to mediate efficient apical sorting. They raise the intriguing possibility that changes in glycosyltransferase expression patterns and/or galectin-9 distribution may acutely modulate endolyn trafficking in the kidney.