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Mutation affecting the conserved acidic WNK1 motif causes inherited hyperkalemic hyperchloremic acidosis
Hélène Louis-Dit-Picard, … , Juliette Hadchouel, Xavier Jeunemaitre
Hélène Louis-Dit-Picard, … , Juliette Hadchouel, Xavier Jeunemaitre
Published August 13, 2020
Citation Information: J Clin Invest. 2020;130(12):6379-6394. https://doi.org/10.1172/JCI94171.
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Research Article Genetics Nephrology

Mutation affecting the conserved acidic WNK1 motif causes inherited hyperkalemic hyperchloremic acidosis

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Abstract

Gain-of-function mutations in with no lysine (K) 1 (WNK1) and WNK4 genes are responsible for familial hyperkalemic hypertension (FHHt), a rare, inherited disorder characterized by arterial hypertension and hyperkalemia with metabolic acidosis. More recently, FHHt-causing mutations in the Kelch-like 3–Cullin 3 (KLHL3-CUL3) E3 ubiquitin ligase complex have shed light on the importance of WNK’s cellular degradation on renal ion transport. Using full exome sequencing for a 4-generation family and then targeted sequencing in other suspected cases, we have identified new missense variants in the WNK1 gene clustering in the short conserved acidic motif known to interact with the KLHL3-CUL3 ubiquitin complex. Affected subjects had an early onset of a hyperkalemic hyperchloremic phenotype, but normal blood pressure values”Functional experiments in Xenopus laevis oocytes and HEK293T cells demonstrated that these mutations strongly decrease the ubiquitination of the kidney-specific isoform KS-WNK1 by the KLHL3-CUL3 complex rather than the long ubiquitous catalytically active L-WNK1 isoform. A corresponding CRISPR/Cas9 engineered mouse model recapitulated both the clinical and biological phenotypes. Renal investigations showed increased activation of the Ste20 proline alanine–rich kinase–Na+-Cl– cotransporter (SPAK-NCC) phosphorylation cascade, associated with impaired ROMK apical expression in the distal part of the renal tubule. Together, these new WNK1 genetic variants highlight the importance of the KS-WNK1 isoform abundance on potassium homeostasis.

Authors

Hélène Louis-Dit-Picard, Ilektra Kouranti, Chloé Rafael, Irmine Loisel-Ferreira, Maria Chavez-Canales, Waed Abdel-Khalek, Eduardo R. Argaiz, Stéphanie Baron, Sarah Vacle, Tiffany Migeon, Richard Coleman, Marcio Do Cruzeiro, Marguerite Hureaux, Nirubiah Thurairajasingam, Stéphane Decramer, Xavier Girerd, Kevin O’Shaugnessy, Paolo Mulatero, Gwenaëlle Roussey, Ivan Tack, Robert Unwin, Rosa Vargas-Poussou, Olivier Staub, Richard Grimm, Paul A. Welling, Gerardo Gamba, Eric Clauser, Juliette Hadchouel, Xavier Jeunemaitre

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Figure 9

Expression of ROMK, BK channel, and NKCC2.

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Expression of ROMK, BK channel, and NKCC2.
(A) ROMK protein abundance. N...
(A) ROMK protein abundance. No change in cortical ROMK expression was observed in Wnk1+/delE631 (n = 4) vs. littermate Wnk1+/+ (n = 4) mice, despite the latter group having significantly higher plasma K+ levels (4.4 ± 0.08 mM, n = 20 vs. 5.1 ± 0.12 mM, n = 20). Conversely, a more than 2-fold increase was observed in Wnk1+/+ mice treated by amiloride (25 mg/kg/d for 4 days) achieving a similar rise in plasma potassium (Wnk1+/+ vehicle: 3.8 ± 0.06 mM, n = 4 vs. Wnk1+/+ amiloride: 5.0 ± 0.11 mM). Quantification of cortical ROMK expression (n = 4 per group). *P < 0.05, t test. (B) ROMK immunofluorescence in the distal tubule. Left panel: immunolocalization of ROMK in the DCT2 of Wnk1+/+ and Wnk1+/delE631 mice. Right panel: analysis of membrane labeling intensity showed no change in ROMK apical expression in the DCT2 and CNT of Wnk1+/delE631 mice (n = 4 animals per genotype). Scale bar: 10 microns. (C) Basal BK α channel protein abundance in Wnk1+/delE631 mice and Wnk1+/+ littermates. BK α immunoblots (left panel) and quantification (right panel) demonstrate that cortical BK α expression is unchanged between Wnk1+/+ (n = 4) and Wnk1+/delE631 mice (n = 5). (D) Basal NKCC2 and P-NKCC2 protein abundance in Wnk1+/delE631 mice and Wnk1+/+ littermates. (n = 7 per group). NKCC2 and pNKCC2 immunoblots (left panel) and quantification (right panel) demonstrate that NKCC expression is unchanged between Wnk1+/+ (n = 7) and Wnk1+/delE631 mice (n = 7), but pNKCC2 is significantly increased in Wnk1+/delE631 mice compared with Wnk1+/+. ****P < 0.0001, unpaired Student’s t test.

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ISSN: 0021-9738 (print), 1558-8238 (online)

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