title: KLHL3 regulates paracellular chloride transport in the kidney by ubiquitination of claudin-8.
abstract: A rare Mendelian syndrome--pseudohypoaldosteronism type II (PHA-II)--features hypertension, hyperkalemia, and metabolic acidosis. Genetic linkage studies and exome sequencing have identified four genes--with no lysine kinase 1 (wnk1), wnk4, Kelch-like 3 (KLHL3), and Cullin 3 (Cul3)--mutations of which all caused PHA-II phenotypes. The previous hypothesis was that the KLHL3-Cul3 ubiquitin complex acted on the wnk4-wnk1 kinase complex to regulate Na(+)/Cl(-) cotransporter (NCC) mediated salt reabsorption in the distal tubules of the kidney. Here, we report the identification of claudin-8 as a previously unidentified physiologic target for KLHL3 and provide an alternative explanation for the collecting duct's role in PHA-II. Using a tissue-specific KO approach, we have found that deletion of claudin-8 in the collecting duct of mouse kidney caused hypotension, hypokalemia, and metabolic alkalosis, an exact mirror image of PHA-II. Mechanistically, the phenotypes in claudin-8 KO animals were caused by disruption of the claudin-8 interaction with claudin-4, the paracellular chloride channel, and delocalization of claudin-4 from the tight junction. In mouse collecting duct cells, knockdown of KLHL3 profoundly increased the paracellular chloride permeability. Mechanistically, KLHL3 was directly bound to claudin-8, and this binding led to the ubiquitination and degradation of claudin-8. The dominant PHA-II mutation in KLHL3 impaired claudin-8 binding, ubiquitination, and degradation. These findings have attested to the concept that the paracellular pathway is physiologically regulated through the ubiquitination pathway, and its deregulation may lead to diseases of electrolyte and blood pressure imbalances.
authors.affiliation: Departments of Internal Medicine-Renal Division and Center for Investigation of Membrane Excitability Diseases, Washington University Medical School, St. Louis, MO 63110.
authors.forename: Yongfeng
authors.initials: Y
authors.lastname: Gong
authors.affiliation: Departments of Internal Medicine-Renal Division and Center for Investigation of Membrane Excitability Diseases, Washington University Medical School, St. Louis, MO 63110.
authors.forename: Jinzhi
authors.initials: J
authors.lastname: Wang
authors.affiliation: Departments of Internal Medicine-Renal Division and Center for Investigation of Membrane Excitability Diseases, Washington University Medical School, St. Louis, MO 63110.
authors.forename: Jing
authors.initials: J
authors.lastname: Yang
authors.affiliation: Neurology and.
authors.forename: Ernie
authors.initials: E
authors.lastname: Gonzales
authors.affiliation: Neurology and.
authors.forename: Ronaldo
authors.initials: R
authors.lastname: Perez
authors.affiliation: Departments of Internal Medicine-Renal Division and Center for Investigation of Membrane Excitability Diseases, Washington University Medical School, St. Louis, MO 63110 jhou@wustl.edu.
authors.forename: Jianghui
authors.initials: J
authors.lastname: Hou
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D002712:Chlorides / Q000737:chemistry*; D057167:Claudins / Q000502:physiology*; D057809:HEK293 Cells; D006801:Humans; D006973:Hypertension / Q000378:metabolism; D007473:Ion Channels / Q000737:chemistry; D007668:Kidney / Q000378:metabolism*; D008297:Male; D051379:Mice; D018345:Mice, Knockout; D008840:Microfilament Proteins / Q000502:physiology*; D009154:Mutation; D010539:Permeability; D010641:Phenotype; D011485:Protein Binding; D019108:Tight Junctions; D025801:Ubiquitin / Q000737:chemistry*
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D002712:Chlorides; D057167:Claudins; D007473:Ion Channels; C000598025:KLHL3 protein, mouse; D008840:Microfilament Proteins; D025801:Ubiquitin; C502011:claudin 8
keywords: chloride; ion channel; nephrology; tight junction; ubiquitination
references.citation: Proc Natl Acad Sci U S A. 2013 May 7;110(19):7838-43
references.pmid: 23576762
references.citation: Cell Rep. 2013 Mar 28;3(3):858-68
references.pmid: 23453970
references.citation: Proc Natl Acad Sci U S A. 2014 Sep 9;111(36):E3766-74
references.pmid: 25157135
references.citation: Proc Natl Acad Sci U S A. 2014 Oct 28;111(43):15556-61
references.pmid: 25313067
references.citation: J Clin Invest. 2014 Nov;124(11):4723-36
references.pmid: 25250572
references.citation: Semin Cell Dev Biol. 2014 Dec;36:204-12
references.pmid: 25263012
references.citation: Science. 2001 Aug 10;293(5532):1107-12
references.pmid: 11498583
references.citation: J Clin Invest. 2003 Apr;111(7):1039-45
references.pmid: 12671053
references.citation: Proc Natl Acad Sci U S A. 2004 Mar 30;101(13):4690-4
references.pmid: 15070779
references.citation: Am J Physiol Renal Physiol. 2004 Jun;286(6):F1063-71
references.pmid: 14722018
references.citation: J Am Soc Nephrol. 2004 Sep;15(9):2276-88
references.pmid: 15339977
references.citation: Proc Natl Acad Sci U S A. 2004 Oct 12;101(41):14877-82
references.pmid: 15465913
references.citation: Kidney Int. 1981 May;19(5):716-27
references.pmid: 7026872
references.citation: Hypertension. 1986 Feb;8(2):93-102
references.pmid: 3002982
references.citation: Proc Natl Acad Sci U S A. 1993 Dec 15;90(24):11663-7
references.pmid: 8265605
references.citation: Nature. 1994 Feb 3;367(6462):463-7
references.pmid: 8107805
references.citation: Mech Dev. 2005 Jun;122(6):805-19
references.pmid: 15908185
references.citation: Nat Genet. 2006 Oct;38(10):1124-32
references.pmid: 16964266
references.citation: Am J Hum Genet. 2006 Nov;79(5):949-57
references.pmid: 17033971
references.citation: J Biol Chem. 2006 Nov 24;281(47):36117-23
references.pmid: 17018523
references.citation: Proc Natl Acad Sci U S A. 2007 Mar 6;104(10):4020-4
references.pmid: 17360470
references.citation: Cell Metab. 2007 May;5(5):331-44
references.pmid: 17488636
references.citation: J Am Soc Nephrol. 2007 Jun;18(6):1679-87
references.pmid: 17475815
references.citation: J Neurochem. 2008 Jan;104(1):147-54
references.pmid: 17971126
references.citation: J Clin Invest. 2008 Feb;118(2):619-28
references.pmid: 18188451
references.citation: J Cell Biol. 2008 May 19;181(4):683-95
references.pmid: 18474622
references.citation: Biochem Biophys Res Commun. 2009 Jan 2;378(1):45-50
references.pmid: 19000657
references.citation: J Cell Sci. 2009 Apr 1;122(Pt 7):985-94
references.pmid: 19295125
references.citation: Proc Natl Acad Sci U S A. 2009 Sep 8;106(36):15350-5
references.pmid: 19706394
references.citation: Am J Physiol Renal Physiol. 2010 Jan;298(1):F24-34
references.pmid: 19759267
references.citation: Proc Natl Acad Sci U S A. 2010 Oct 19;107(42):18010-5
references.pmid: 20921420
references.citation: J Am Soc Nephrol. 2010 Nov;21(11):1942-51
references.pmid: 20947633
references.citation: Cell Metab. 2011 Sep 7;14(3):352-64
references.pmid: 21907141
references.citation: Hypertension. 2011 Nov;58(5):888-94
references.pmid: 21896937
references.citation: Nature. 2012 Feb 2;482(7383):98-102
references.pmid: 22266938
references.citation: Nat Genet. 2012 Apr;44(4):456-60, S1-3
references.pmid: 22406640
references.citation: J Cell Biochem. 2012 Jul;113(7):2415-23
references.pmid: 22389112
references.citation: Hum Mol Genet. 2014 Oct 1;23(19):5052-60
references.pmid: 24821705