Modulation of kidney urea transporter UT-A3 activity by alpha2,6-sialylation

• Type of Material: Article
• Authors: Xiaoqian Qian, Emory University; Jeff Sands, Emory University; Xiang Song, Harbin Medical University; Guangping Chen, Emory University
• Language: English
• Date: 2016-07-01
• Publisher: Springer Verlag
• Publication Version: Author Accepted Manuscript (After Peer Review)
• Copyright Statement: © 2016, Springer-Verlag Berlin Heidelberg.
• Final Published Version (URL): http://dx.doi.org/10.1007/s00424-016-1802-0
• Title of Journal or Parent Work: Pflügers Archiv European Journal of Physiology
• Volume: 468
• Issue: 7
• Start Page: 1161
• End Page: 1170
• Grant/Funding Information: This work was supported by NIH grants R01-DK087838 (to G. Chen), R01-DK89828 (to J. Sands), and by Chinese National Natural Science Foundation Project 81570358, 81300248 (to X. Song), Heilongjiang Science and Technology funding LBH-Q13113 (to X. Song).
• Keywords: Science & Technology; MICE LACKING; MEMBRANE; CONCENTRATING DEFECT; COLLECTING DUCT; Glycosylation; PROTEIN; SIALYLATION; ACCUMULATION; Protein kinase C; Physiology; Urea transporter; UT-B; PHOSPHORYLATION; GLYCOSYLATION; Sialyltransferase; Life Sciences & Biomedicine
• Research Categories: Biology, Cell; Biology, Physiology

Abstract

Two urea transporters, UT-A1 and UT-A3, are expressed in the kidney terminal inner medullary collecting duct (IMCD) and are important for the production of concentrated urine. UT-A1, as the largest isoform of all UT-A urea transporters, has gained much attention and been extensively studied; however, the role and the regulation of UT-A3 are less explored. In this study, we investigated UT-A3 regulation by glycosylation modification. A site-directed mutagenesis verified a single glycosylation site in UT-A3 at Asn279. Loss of the glycosylation reduced forskolin-stimulated UT-A3 cell membrane expression and urea transport activity. UT-A3 has two glycosylation forms, 45 and 65 kDa. Using sugar-specific binding lectins, the UT-A3 glycosylation profile was examined. The 45-kDa form was pulled down by lectin concanavalin A (Con A) and Galant husnivalis lectin (GNL), indicating an immature glycan with a high amount of mannose (Man), whereas the 65-kDa form is a mature glycan composed of acetylglucosamine (GlcNAc) and poly-N-acetyllactosame (poly-LacNAc) that was pulled down by wheat germ agglutinin (WGA) and tomato lectin, respectively. Interestingly, the mature form of UT-A3 glycan contains significant amounts of sialic acid. We explored the enzymes responsible for directing UT-A3 sialylation. Sialyltransferase ST6GalI, but not ST3GalIV, catabolizes UT-A3 α2,6-sialylation. Activation of protein kinase C (PKC) by PDB treatment promoted UT-A3 glycan sialylation and membrane surface expression. The PKC inhibitor chelerythrine blocks ST6GalI-induced UT-A3 sialylation. Increased sialylation by ST6GalI increased UT-A3 protein stability and urea transport activity. Collectively, our study reveals a novel mechanism of UT-A3 regulation by ST6GalI-mediated sialylation modification that may play an important role in kidney urea reabsorption and the urinary concentrating mechanism.