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- Title
Isoform specificity of human Na<sup>+</sup>,K<sup>+</sup>-ATPase localization and aldosterone regulation in mouse kidney cells.
- Authors
Summa, Vanessa; Camargo, Simone M. R.; Bauch, Christian; Zecevic, Marija; Verrey, François
- Abstract
Short-term aldosterone coordinately regulates the cell-surface expression of luminal epithelial sodium channels (ENaC) and of basolateral Na+ pumps (Na+,K+-ATPaseα1–β1) in aldosterone-sensitive distal nephron (ASDN) cells. To address the question of whether the subcellular localization of the Na+,K+-ATPase and its regulation by aldosterone depend on subunit isoform-specific structures, we expressed the cardiotonic steroid-sensitive humanα isoforms 1–3 by retroviral transduction in mouse collecting duct mpkCCDc14 cells. Each of the three exogenous human isoforms could be detected by Western blotting. Immunofluorescence indicated that the exogenousα1 subunit to a large extent localizes to the basolateral membrane or close to it, whereas much of theα2 subunit remains intracellular. An ouabain-sensitive current carried by exogenous pumps could be detected in apically amphotericin B-permeabilized epithelia expressing humanα1 andα2 subunits, but not theα3 subunit. This current displayed a higher apparent Na+ affinity in pumps containing humanα2 subunits (10 mm) than in pumps containing humanα1 (33.2 mm) or endogenous (cardiotonic steroid-resistant) mouseα1 subunits (mean: 16.3 mm). A very low mRNA level of the Na+,K+-ATPaseγ subunit (FXYD2) in mpkCCDc14 cells suggested that this ancillary gene product is not responsible for the relatively low apparent Na+ affinity measured for a1 subunit-containing pumps. Aldosterone increased the pump current carried by endogenous pumps and by pumps containing the humanα1 subunit. In contrast, the current carried by pumps with a humanα2 subunit was not stimulated by the same treatment. In summary, quantitative basolateral localization of the Na+,K+-ATPase and its responsiveness to aldosterone requireα1 subunit-specific sequences that differentiate this isoform from theα2 andα3 subunit isoforms.
- Publication
Journal of Physiology, 2004, Vol 555, Issue 2, p355
- ISSN
0022-3751
- Publication type
Academic Journal
- DOI
10.1113/jphysiol.2003.054270