Several acute and chronic lung diseases are associated with alveolar hypoventilation leading to accumulation of CO2 (hypercapnia). The beta-subunit of the Na,K-ATPase plays a pivotal role in maintaining epithelial integrity by functioning as a cell adhesion molecule and regulating cell surface stability of the catalytic alpha-subunit of the transporter, thereby, maintaining optimal alveolar fluid balance. Here, we identified the E3 ubiquitin ligase for the Na,K-ATPase beta-subunit, which promoted polyubiquitination, subsequent endocytosis and proteasomal degradation of the protein upon exposure of alveolar epithelial cells to elevated CO2 levels, thus impairing alveolar integrity. Ubiquitination of the Na,K-ATPase beta-subunit required lysine 5 and 7 and mutating these residues (but not other lysines) prevented trafficking of Na,K-ATPase from the plasma membrane and stabilized the protein upon hypercapnia. Furthermore, ubiquitination of the Na,K-ATPase beta-subunit was dependent on prior phosphorylation at serine 11 by protein kinase C (PKC)-zeta. Using a protein microarray, we identified the tumor necrosis factor receptor-associated factor 2 (TRAF2) as the E3 ligase driving ubiquitination of the Na,K-ATPase beta-subunit upon hypercapnia. Of note, prevention of Na,K-ATPase beta-subunit ubiquitination was necessary and sufficient to restore the formation of cell-cell junctions under hypercapnic conditions. These results suggest that a hypercapnic environment in the lung may lead to persistent epithelial dysfunction in affected patients. As such, the identification of the E3 ligase for the Na,K-ATPase may provide a novel therapeutic target, to be employed in patients with acute or chronic hypercapnic respiratory failure, aiming to restore alveolar epithelial integrity.
- Gabrielli, N. M.
- Mazzocchi, L. C.
- Kryvenko, V.
- Tello, K.
- Herold, S.
- Morty, R. E.
- Grimminger, F.
- Dada, L. A.
- Seeger, W.
- Sznajder, J. I.
- Vadasz, I.
Keywords
- K-ATPase beta-subunit
- Na
- PKC-zeta
- Traf2
- adherens junction
- carbon dioxide
- endocytosis
- hypercapnia
- ubiquitination
- commercial or financial relationships that could be construed as a potential
- conflict of interest.