Localization of GFP-tagged concentrative nucleoside transporters in a renal polarized epithelial cell line

GFP-tagged concentrative nucleoside transporters in a renal polarized epithelial cell line. Am J Physiol Renal Physiol 280: F879–F885, 2001.—Many nucleosides undergo active reab-sorption within the kidney, probably via nucleoside trans-porters. To date, two concentrative nucleoside transporters have been cloned, the sodium-dependent purine-selective nu-cleoside transporter (SPNT) and concentrative nucleoside transporter 1 (CNT1). We report the stable expression of green fluorescence protein (GFP)-tagged SPNT and CNT1 in Madin-Darby canine kidney (MDCK) cells, a polarized renal epithelial line. We demonstrate that the GFP tag does not alter the substrate selectivity and only modestly affects the kinetic activity of the transporters. By using confocal micros-copy and functional studies, both SPNT and CNT1 are local-ized primarily to the apical membrane of MDCK and LLC