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Am J Physiol Renal Physiol 286: F979-F987, 2004. First published January 6, 2004; doi:10.1152/ajprenal.00334.2003
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The basolateral expression of mUT-A3 in the mouse kidney

Gavin S. Stewart,1 Robert A. Fenton,2 Weidong Wang,3 Tae-Hwan Kwon,3 Stanley J. White,4 Valerie M. Collins,4 Gordon Cooper,5 Søren Nielsen,3 and Craig P. Smith1

1School of Biological Sciences, University of Manchester, Manchester M13 9PT; 4School of Biomedical Sciences, University of Leeds, Leeds LS2 9JT; 5Department of Biomedical Sciences, University of Sheffield, Sheffield S10 2TN, United Kingdom; 2Laboratory of Kidney and Electrolyte Metabolism, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland 20892; and 3The Water and Salt Research Center, University of Aarhus, DK-8000 Aarhus C, Denmark

Submitted 16 September 2003 ; accepted in final form 2 January 2004

Facilitative UT-A urea transporters play a central role in the urinary concentrating mechanism. There are three major UT-A isoforms found in the mouse kidney: mUT-A1, mUT-A2, and mUT-A3. The major aim of this study was to identify the location and function of mUT-A3. UT-A proteins were investigated using three novel mouse UT-A-targeted antibodies: ML446, MQ2, and ML194. ML446 detected mUT-A1 and mUT-A3. ML194 detected mUT-A1 and mUT-A2. Importantly, MQ2 was found to be selective for mUT-A3. MQ2 detected a 45- to 65-kDa signal in the mouse kidney inner medulla, which was deglycosylated to a 40-kDa protein band. Immunolocalization studies showed that mUT-A3 was strongly detected in the papillary tip, mainly in the basolateral regions of inner medullary collecting duct (IMCD) cells. Immunoblotting of subcellular fractions of inner medullary protein suggested that in mouse kidney mUT-A3 was present in plasma membranes. Consistent with this, immunoelectron microscopy demonstrated that mUT-A3 was predominantly localized at the basal plasma membrane domains of the IMCD cells in mouse kidney. Heterologous expression of mUT-A3-enhanced green fluorescent protein in Madin-Darby canine kidney cells showed that the protein localized to the basolateral membrane. In conclusion, our study indicates that mUT-A3 is a basolateral membrane transporter expressed in IMCD cells.

inner medullary collecting duct cells; fluid homeostasis


Address for reprint requests and other correspondence: C. P. Smith, School of Biological Sciences, Univ. of Manchester, G.38 Stopford Bldg., Oxford Road, Manchester M13 9PT, UK (E-mail: cpsmith{at}man.ac.uk).




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