Na,K-ATPase β-subunit cis homo-oligomerization is necessary for epithelial lumen formation in mammalian cells

Na,K-ATPase is a hetero-oligomer of an α- and a β-subunit. The α-subunit (Na,K-α) possesses the catalytic function, whereas the bsubunit (Na,K-β) has cell-cell adhesion function and is localized to the apical junctional complex in polarized epithelial cells. Earlier, we identified two distinct conserved motifs on the Na,K-β₁ transmembrane domain that mediate protein-protein interactions: a glycine zipper motif involved in the cis homo-oligomerization of Na,K-β₁ and a heptad repeat motif that is involved in the hetero-oligomeric interaction with Na,K-α₁. We now provide evidence that knockdown of Na,K-β₁ prevents lumen formation and induces activation of extracellular regulated kinases 1 and 2 (ERK1/2) mediated by phosphatidylinositol 3-kinase in MDCK cells grown in three-dimensional collagen cultures. These cells sustained cell proliferation in an ERK1/2-dependent manner and did not show contact inhibition at high cell densities, as revealed by parental MDCK cells. This phenotype could be rescued by wild-type Na,K-β₁ or heptad repeat motif mutant of Na,K-β₁, but not by the glycine zipper motif mutant that abrogates Na,K-β₁ cis homo-oligomerization. These studies suggest that Na,K-β₁ cis homo-oligomerization rather than hetero-oligomerization with Na,K-α₁ is involved in epithelial lumen formation. The relevance of these findings to pre-neoplastic lumen filling in epithelial cancer is discussed.