2024
Structural bases for Na+-Cl− cotransporter inhibition by thiazide diuretic drugs and activation by kinases
Zhao Y, Schubert H, Blakely A, Forbush B, Smith M, Rinehart J, Cao E. Structural bases for Na+-Cl− cotransporter inhibition by thiazide diuretic drugs and activation by kinases. Nature Communications 2024, 15: 7006. PMID: 39143061, PMCID: PMC11324901, DOI: 10.1038/s41467-024-51381-y.Peer-Reviewed Original ResearchConceptsNa+-Cl- cotransporterFamilial hyperkalemic hypertensionRenal salt retentionThiazide diuretic drugsNa+-Cl-Cotransporter inhibitionNCC activitySalt reabsorptionDiuretic drugsBlood pressureBalanced electrolyteTreat hypertensionIon translocation pathwayIon translocationThiazideHypertensionSalt retentionOrthosteric siteCo-structureCarboxyl-terminal domainKinase cascadeEdemaChlorthalidoneCotransporterTranslocation
2021
The structural basis of function and regulation of neuronal cotransporters NKCC1 and KCC2
Zhang S, Zhou J, Zhang Y, Liu T, Friedel P, Zhuo W, Somasekharan S, Roy K, Zhang L, Liu Y, Meng X, Deng H, Zeng W, Li G, Forbush B, Yang M. The structural basis of function and regulation of neuronal cotransporters NKCC1 and KCC2. Communications Biology 2021, 4: 226. PMID: 33597714, PMCID: PMC7889885, DOI: 10.1038/s42003-021-01750-w.Peer-Reviewed Original ResearchConceptsCryo-electron microscopy structureHuman NKCC1Microscopy structureEssential residuesFunctional characterizationKCC transportersPlasma membraneStructural basisTransepithelial saltTransport activityMechanistic understandingTransportersStructural studiesCritical roleCotransporter NKCC1Computational analysisIon transportWater transportNeuronal excitabilityNKCC1PhosphorylationCell volumeNKCCKCC2Residues
2013
Functional Expression of Human NKCC1 from a Synthetic Cassette-Based cDNA: Introduction of Extracellular Epitope Tags and Removal of Cysteines
Somasekharan S, Monette MY, Forbush B. Functional Expression of Human NKCC1 from a Synthetic Cassette-Based cDNA: Introduction of Extracellular Epitope Tags and Removal of Cysteines. PLOS ONE 2013, 8: e82060. PMID: 24339991, PMCID: PMC3855340, DOI: 10.1371/journal.pone.0082060.Peer-Reviewed Original ResearchConceptsNa-K-Cl cotransporterEpitope tagYFP tagPlasma membraneMost animal cellsRemoval of cysteineOpen reading frameSilent restriction sitesHuman NKCC1Regulation of NKCC1Na-K-Cl cotransport activityHEK-293 cellsCellular homeostasisBiosynthetic machinerySynthetic cassetteAnimal cellsNovel cDNACysteine mutantsReading frameSynthetic cDNACysteine mutationsEndoplasmic reticulumFunctional expressionCotransport activityCDNA
2012
Loop Diuretic and Ion-binding Residues Revealed by Scanning Mutagenesis of Transmembrane Helix 3 (TM3) of Na-K-Cl Cotransporter (NKCC1)*
Somasekharan S, Tanis J, Forbush B. Loop Diuretic and Ion-binding Residues Revealed by Scanning Mutagenesis of Transmembrane Helix 3 (TM3) of Na-K-Cl Cotransporter (NKCC1)*. Journal Of Biological Chemistry 2012, 287: 17308-17317. PMID: 22437837, PMCID: PMC3366785, DOI: 10.1074/jbc.m112.356014.Peer-Reviewed Original ResearchConceptsTransmembrane helix 3Na-K-Cl cotransporterTranslocation pathwayHelix 3Homology modelTryptophan-scanning mutagenesisMutation of residuesStructural homology modelEpithelial salt transportExtracellular gateCellular chloride homeostasisScanning mutagenesisOpen conformationIntracellular endPore residuesFunctional roleIon translocationTranslocation rateResiduesMutagenesisCentral roleChloride homeostasisMutationsPathwayLarge effect
2011
Regulatory Activation Is Accompanied by Movement in the C Terminus of the Na-K-Cl Cotransporter (NKCC1)*
Monette MY, Forbush B. Regulatory Activation Is Accompanied by Movement in the C Terminus of the Na-K-Cl Cotransporter (NKCC1)*. Journal Of Biological Chemistry 2011, 287: 2210-2220. PMID: 22121194, PMCID: PMC3265899, DOI: 10.1074/jbc.m111.309211.Peer-Reviewed Original ResearchConceptsC-terminusFluorescence resonance energy transferNa-K-Cl cotransporterFRET decreasesSame C-terminusMost vertebrate cellsKey structural roleEmbryonic kidney cell lineYellow fluorescent proteinHuman embryonic kidney cell lineRegulation of NKCC1Vertebrate cellsKidney cell linePlasma membraneNKCC1 regulationN-terminusFluorescent proteinStructural roleRegulatory activationTransporter activationConformational changesTerminusTransport activityResonance energy transferHEK cellsRare mutations in the human Na-K-Cl cotransporter (NKCC2) associated with lower blood pressure exhibit impaired processing and transport function
Monette MY, Rinehart J, Lifton RP, Forbush B. Rare mutations in the human Na-K-Cl cotransporter (NKCC2) associated with lower blood pressure exhibit impaired processing and transport function. American Journal Of Physiology. Renal Physiology 2011, 300: f840-f847. PMID: 21209010, PMCID: PMC3074999, DOI: 10.1152/ajprenal.00552.2010.Peer-Reviewed Original ResearchConceptsHEK-293 cellsNa-K-Cl cotransporterTransport functionMajor salt transport pathwayPlasma membrane localizationHEK cellsLow transport activitySequence conservationMembrane localizationProtein functionHeterologous expressionXenopus laevis oocytesImportant residuesMutantsRenal salt reabsorptionMolecular mechanismsIndependent mutationsConstitutive activityTransport activityBlood pressureFunctional consequencesImpaired transport functionSuch mutationsProcessing defectsLaevis oocytes