title: Modulation of kidney urea transporter UT-A3 activity by alpha2,6-sialylation.
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.
authors.affiliation: Department of Physiology, Emory University, Atlanta, GA, 30322, USA.|Cardiovascular Center, The 4th Affiliated Hospital, Harbin Medical University, 31 Yinhang Street, Nangang District, Harbin, Heilongjiang, 150001, China.
authors.forename: Xiaoqian
authors.initials: X
authors.lastname: Qian
authors.affiliation: Department of Physiology, Emory University, Atlanta, GA, 30322, USA.|Renal Division, Department of Medicine, Emory University, Atlanta, GA, 30322, USA.
authors.forename: Jeff M
authors.initials: JM
authors.lastname: Sands
authors.affiliation: Cardiovascular Center, The 4th Affiliated Hospital, Harbin Medical University, 31 Yinhang Street, Nangang District, Harbin, Heilongjiang, 150001, China. song761231@sina.com.
authors.forename: Xiang
authors.initials: X
authors.lastname: Song
authors.affiliation: Department of Physiology, Emory University, Atlanta, GA, 30322, USA.|Renal Division, Department of Medicine, Emory University, Atlanta, GA, 30322, USA.
authors.forename: Guangping
authors.initials: G
authors.lastname: Chen
mesh_terms: D001692:Biological Transport / Q000502:physiology; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D006031:Glycosylation; D057809:HEK293 Cells; D006801:Humans; D007679:Kidney Medulla / Q000378:metabolism*; D007685:Kidney Tubules, Collecting / Q000378:metabolism*; D026901:Membrane Transport Proteins / Q000378:metabolism*; D011134:Polysaccharides / Q000378:metabolism; D011493:Protein Kinase C / Q000378:metabolism; D014508:Urea / Q000378:metabolism*; D000096982:Urea Transporters
chemical_list: D026901:Membrane Transport Proteins; D011134:Polysaccharides; D011493:Protein Kinase C; D014508:Urea; D000096982:Urea Transporters
keywords: Glycosylation; Protein kinase C; Sialyltransferase; Urea transporter