Document Detail


beta-Subunit overexpression alters the stoicheometry of assembled Na-K-ATPase subunits in MDCK cells.
MedLine Citation:
PMID:  18701620     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
In eukaryotic cells, the apparent maintenance of 1:1 stoicheometry between the Na-K-ATPase alpha- and beta-subunits led us to question whether this was alterable and thus if some form of regulation was involved. We have examined the consequences of overexpressing Na-K-ATPase beta1-subunits using Madin-Darby canine kidney (MDCK) cells expressing flag-tagged beta1-subunits (beta1flag) or Myc-tagged beta1-subunits (beta1myc) under the control of a tetracycline-dependent promoter. The induction of beta1flag subunit synthesis in MDCK cells, which increases beta1-subunit expression at the plasma membrane by more than twofold, while maintaining stable alpha1 expression levels, revealed that all mature beta1-subunits associate with alpha1-subunits, and no evidence of "free" beta1-subunits was obtained. Consequently, the ratio of assembled beta1- to alpha1-subunits is significantly increased when "extra" beta-subunits are expressed. An increased beta1/alpha1 stoicheometry is also observed in cells treated with tunicamycin, suggesting that the protein-protein interactions involved in these complexes are not dependent on glycosylation. Confocal images of cocultured beta1myc-expressing and beta1flag-expressing MDCK cells show colocalization of beta1myc and beta1flag subunits at the lateral membranes of neighboring cells, suggesting the occurrence of intercellular interactions between the beta-subunits. Immunoprecipitation using MDCK cells constitutively expressing beta1myc and tetracycline-regulated beta1flag subunits confirmed beta-beta-subunit interactions. These results demonstrate that the equimolar ratio of assembled beta1/alpha1-subunits of the Na-K-ATPase in kidney cells is not fixed by the inherent properties of the interacting subunits. It is likely that cellular mechanisms are present that regulate the individual Na-K-ATPase subunit abundance.
Authors:
Rebecca J Clifford; Jack H Kaplan
Publication Detail:
Type:  Journal Article; Research Support, N.I.H., Extramural     Date:  2008-08-13
Journal Detail:
Title:  American journal of physiology. Renal physiology     Volume:  295     ISSN:  1931-857X     ISO Abbreviation:  Am. J. Physiol. Renal Physiol.     Publication Date:  2008 Nov 
Date Detail:
Created Date:  2008-11-06     Completed Date:  2009-02-09     Revised Date:  2013-06-05    
Medline Journal Info:
Nlm Unique ID:  100901990     Medline TA:  Am J Physiol Renal Physiol     Country:  United States    
Other Details:
Languages:  eng     Pagination:  F1314-23     Citation Subset:  IM    
Affiliation:
Department of Biochemistry and Molecular Genetics, University of Illinois at Chicago, Chicago, IL 60607-7170, USA.
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MeSH Terms
Descriptor/Qualifier:
Adenosine Triphosphatases / metabolism
Animals
Blotting, Western
Cell Line
Cell Membrane / metabolism
Dogs
Electrophoresis, Polyacrylamide Gel
Endoplasmic Reticulum / metabolism
Epithelial Cells / cytology,  metabolism*
Gene Expression
Glycosylation / drug effects
Golgi Apparatus / metabolism
Humans
Immunoprecipitation
Intracellular Membranes / metabolism
Protein Binding
Protein Subunits / chemistry,  genetics,  metabolism
Rats
Sheep
Sodium-Potassium-Exchanging ATPase / chemistry,  genetics,  metabolism*
Species Specificity
Transfection
Tunicamycin / pharmacology
Grant Support
ID/Acronym/Agency:
GM-39500/GM/NIGMS NIH HHS
Chemical
Reg. No./Substance:
0/Protein Subunits; 11089-65-9/Tunicamycin; EC 3.6.1.-/Adenosine Triphosphatases; EC 3.6.3.9/Na K ATPase beta2 subunit, rat; EC 3.6.3.9/Sodium-Potassium-Exchanging ATPase
Comments/Corrections
Comment In:
Am J Physiol Renal Physiol. 2008 Nov;295(5):F1313   [PMID:  18768585 ]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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