title: Epithelial junctions depend on intercellular trans-interactions between the Na,K-ATPase β₁ subunits.
abstract: N-Glycans of the Na,K-ATPase β₁ subunit are important for intercellular adhesion in epithelia, suggesting that epithelial junctions depend on N-glycan-mediated interactions between the β₁ subunits of neighboring cells. The level of co-immunoprecipitation of the endogenous β₁ subunit with various YFP-linked β₁ subunits expressed in Madin-Darby canine kidney cells was used to assess β₁-β₁ interactions. The amount of co-precipitated endogenous dog β₁ was greater with dog YFP-β₁ than with rat YFP-β₁, showing that amino acid-mediated interactions are important for β₁-β₁ binding. Co-precipitation of β₁ was also less with the unglycosylated YFP-β₁ than with glycosylated YFP-β₁, indicating a role for N-glycans. Mixing cells expressing dog YFP-β₁ with non-transfected cells increased the amount of co-precipitated β₁, confirming the presence of intercellular (YFP-β₁)-β₁ complexes. Accordingly, disruption of intercellular junctions decreased the amount of co-precipitated β₁ subunits. The decrease in β₁ co-precipitation both with rat YFP-β₁ and unglycosylated YFP-β₁ was associated with decreased detergent stability of junctional proteins and increased paracellular permeability. Reducing N-glycan branching by specific inhibitors increased (YFP-β₁)-β₁ co-precipitation and strengthened intercellular junctions. Therefore, interactions between the β₁ subunits of neighboring cells maintain integrity of intercellular junctions, and alterations in the β₁ subunit N-glycan structure can regulate stability and tightness of intercellular junctions.
authors.affiliation: Department of Physiology, School of Medicine, UCLA and Veterans Administration Greater Los Angeles Health Care System, Los Angeles, California 90073, USA.
authors.forename: Elmira
authors.initials: E
authors.lastname: Tokhtaeva
authors.forename: George
authors.initials: G
authors.lastname: Sachs
authors.forename: Puneet
authors.initials: P
authors.lastname: Souda
authors.forename: Sara
authors.initials: S
authors.lastname: Bassilian
authors.forename: Julian P
authors.initials: JP
authors.lastname: Whitelegge
authors.forename: Liora
authors.initials: L
authors.lastname: Shoshani
authors.forename: Olga
authors.initials: O
authors.lastname: Vagin
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002448:Cell Adhesion; D002460:Cell Line; D004285:Dogs; D004847:Epithelial Cells / Q000166:cytology*; D006801:Humans; D007365:Intercellular Junctions / Q000378:metabolism*; D010539:Permeability; D011134:Polysaccharides / Q000378:metabolism; D011485:Protein Binding; D051381:Rats; D000254:Sodium-Potassium-Exchanging ATPase / Q000737:chemistry / Q000378:metabolism*
chemical_list: C503554:Atp1b1 protein, rat; D011134:Polysaccharides; D000254:Sodium-Potassium-Exchanging ATPase