Mature N-linked glycans facilitate UT-A1 urea transporter lipid raft compartmentalization

• Authors: Guangping Chen, Emory University; Ashley G. Howe, Emory University; Gang Xu, Emory University; Otto Fröhlich, Emory University; Janet D Klein, Emory University; Jeff M Sands, Emory University
• Journal Title: FASEB Journal
• Volume: Volume 25, Number 12
• Publisher: Federation of American Society of Experimental Biology (FASEB) | 2011-12, Pages 4531-4539
• Type of Work: Article | Post-print: After Peer Review
• Publisher DOI: http://doi.org/10.1096/fj.11-185991
• Subjects: Psychology, Physiological
• Keywords: glycosylation; apical membrane; trafficking; lectin; sucrose gradient centrifugation

Abstract

The UT-A1 urea transporter is a glycoprotein with two different glycosylated forms of 97 and 117 kDa. In this study, we found the 117-kDa UT-A1 preferentially resides in lipid rafts, suggesting that the glycosylation status may interfere with UT-A1 lipid raft trafficking. This was confirmed by a site-directed mutagenesis study in MDCK cells. The nonglycosylated UT-A1 showed reduced localization in lipid rafts. By using sugar-specific binding lectins, we further found that the UT-A1 in nonlipid rafts contained a high amount of mannose, as detected by concanavalin A, while the UT-A1 in lipid rafts was the mature N-acetylglucosamine-containing form, as detected by wheat germ agglutinin. In the inner medulla (IM) of diabetic rats, the more abundant 117-kDa UT-A1 in lipid rafts was the mature glycosylation form, with high amounts of N-acetylglucosamine and sialic acid. In contrast, in the IM of normal rats, the predominant 97-kDa UT-A1 was the form enriched in mannose. Functionally, inhibition of glycosylation by tunicamycin or elimination of the glycosylation sites by mutation significantly reduced UT-A1 activity in oocytes. Taken together, our study reveals a new role of N-linked glycosylation in regulating UT-A1 activity by promoting UT-A1 trafficking into membrane lipid raft subdomains.—Chen, G., Howe, A. G., Xu, G., Fröhlich, O., Klein, J. D., Sands, J. M. Mature N-linked glycans facilitate UT-A1 urea transporter lipid raft compartmentalization.