Hypercapnia-induced calcium dysregulation in the endoplasmic reticulum impairs Na,K-ATPase maturation in precision-cut lung slices and alveolar epithelial cells

Elevated carbon dioxide levels (hypercapnia) are frequently observed in patients with acute and chronic lung diseases. Na,K-ATPase is a sodium transporter and junctional molecule that is critically required for the maintenance of optimal alveolar fluid balance and thus gas exchange. Here, we explored how hypercapnia affects maturation of the Na,K-ATPase in the endoplasmic reticulum (ER). Murine precision-cut lung slices and human alveolar epithelial cells exposed to elevated CO2 levels showed a rapid and dose-dependent reduction of the Na,K-ATPase β-subunit in the ER and decreased Na,K-ATPase plasma membrane abundance. By combining chemical and genetic approaches, we identified that hypercapnia activated inositol-requiring enzyme 1α (IRE1α) and initiated a specific signaling pathway that resulted in ER-associated degradation (ERAD) of the Na,K-ATPase β-subunit. ERAD of the β-subunit prevented assembly with the α-subunit of the enzyme and thus downregulated formation of functional transporter molecules as well as delivery of the complex to the cell surface. Moreover, we observed that IRE1α activation was triggered by changes in ER and cytosolic calcium levels, and were dependent on the activity of the inositol trisphosphate ER receptor (IP3R). Treatment with the IP3R inhibitor, 2-aminoethoxydiphenyl borate (2-APB) prevented calcium dysregulation and increased plasma membrane abundance of the Na,K-ATPase α/β-complex. Accordingly, our results suggest that elevated CO2 levels decrease Na,K-ATPase plasma membrane abundance by altering ER calcium homeostasis and enhancing degradation of the Na,K-ATPase β-subunit in the ER.