Aldosterone mediates activation of the thiazide-sensitive Na-Cl cotransporter through an SGK1 and WNK4 signaling pathway
David J. Rozansky, … , Chao-Ling Yang, David H. Ellison
David J. Rozansky, … , Chao-Ling Yang, David H. Ellison
Published August 17, 2009
Citation Information: J Clin Invest. 2009;119(9):2601-2612. https://doi.org/10.1172/JCI38323.
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Research Article Nephrology

Aldosterone mediates activation of the thiazide-sensitive Na-Cl cotransporter through an SGK1 and WNK4 signaling pathway

Abstract

Aldosterone regulates volume homeostasis and blood pressure by enhancing sodium reabsorption in the kidney’s distal nephron (DN). On the apical surface of these renal epithelia, aldosterone increases expression and activity of the thiazide-sensitive Na-Cl cotransporter (NCC) and the epithelial sodium channel (ENaC). While the cellular mechanisms by which aldosterone regulates ENaC have been well characterized, the molecular mechanisms that link aldosterone to NCC-mediated Na+/Cl– reabsorption remain elusive. The serine/threonine kinase with-no-lysine 4 (WNK4) has previously been shown to reduce cell surface expression of NCC. Here we measured sodium uptake in a Xenopus oocyte expression system and found that serum and glucocorticoid–induced kinase 1 (SGK1), an aldosterone-responsive gene expressed in the DN, attenuated the inhibitory effect of WNK4 on NCC activity. In addition, we showed — both in vitro and in a human kidney cell line — that SGK1 bound and phosphorylated WNK4. We found one serine located within an established SGK1 consensus target sequence, and the other within a motif that was, to our knowledge, previously uncharacterized. Mutation of these target serines to aspartate, in order to mimic phosphorylation, attenuated the effect of WNK4 on NCC activity in the Xenopus oocyte system. These data thus delineate what we believe to be a novel mechanism for aldosterone activation of NCC through SGK1 signaling of WNK4 kinase.

Authors

David J. Rozansky, Tonya Cornwall, Arohan R. Subramanya, Shaunessy Rogers, Yong-Feng Yang, Larry L. David, Xiaoman Zhu, Chao-Ling Yang, David H. Ellison

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Figure 1

The active form of SGK1 reverses WNK4 inhibition of NCC-mediated Na+ uptake in Xenopus oocytes.

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The active form of SGK1 reverses WNK4 inhibition of NCC-mediated Na+ upt...
(A) Schematic diagram of the DCT, outlining the hypothesis of where SGK1 acts to influence WNK4 inhibition of Na+/Cl– cotransport (shown by thick arrows). The question mark and dashed line indicate the area of interest for this study. Arrows denote SGK1 movement among inactive, active, and degraded forms. Stimulatory and inhibitory effects are indicated by filled circles and blunt-headed arrows, respectively. Phosphorylation steps are denoted by “P.” (B) Representative sample of formaldehyde/agarose gel stained with ethidium bromide showing equivalent amounts of SGK/S422D and SGK/K127M cRNA and no obvious degradation prior to injection into oocytes. Lanes were run on the same gel, which was split to maintain the sample order in C. (C) Relative to NCC alone, WNK4 reduced NCC-mediated Na+ flux by 60%. The addition of constitutively active SGK1/S422D reversed that effect, whereas addition of kinase-dead SGK1/K127M continued to reduce Na+ flux. n = 3 for each condition (± SEM). Significance (by ANOVA) is indicated.