Hypercapnia Impairs Na,K-ATPase Function by Inducing Endoplasmic Reticulum Retention of the β-Subunit of the Enzyme in Alveolar Epithelial Cells.

Vitalii Kryvenko,Jacob I Sznajder,Werner Seeger,Susanne Herold,Rory E Morty,Laura A Dada,Miriam Wessendorf,Olga Vagin,István Vadász

doi:10.3390/ijms21041467

Abstract

Alveolar edema, impaired alveolar fluid clearance, and elevated CO2 levels (hypercapnia) are hallmarks of the acute respiratory distress syndrome (ARDS). This study investigated how hypercapnia affects maturation of the Na,K-ATPase (NKA), a key membrane transporter, and a cell adhesion molecule involved in the resolution of alveolar edema in the endoplasmic reticulum (ER). Exposure of human alveolar epithelial cells to elevated CO2 concentrations caused a significant retention of NKA-β in the ER and, thus, decreased levels of the transporter in the Golgi apparatus. These effects were associated with a marked reduction of the plasma membrane (PM) abundance of the NKA-α/β complex as well as a decreased total and ouabain-sensitive ATPase activity. Furthermore, our study revealed that the ER-retained NKA-β subunits were only partially assembled with NKA α-subunits, which suggests that hypercapnia modifies the ER folding environment. Moreover, we observed that elevated CO2 levels decreased intracellular ATP production and increased ER protein and, particularly, NKA-β oxidation. Treatment with α-ketoglutaric acid (α-KG), which is a metabolite that has been shown to increase ATP levels and rescue mitochondrial function in hypercapnia-exposed cells, attenuated the deleterious effects of elevated CO2 concentrations and restored NKA PM abundance and function. Taken together, our findings provide new insights into the regulation of NKA in alveolar epithelial cells by elevated CO2 levels, which may lead to the development of new therapeutic approaches for patients with ARDS and hypercapnia.

Highlights

Na,K-ATPase (NKA) is a heterodimeric enzyme and a member of the P-type ATPase family
Our results revealed that α-ketoglutaric acid (α-KG) treatment rescues the effects of hypercapnia on the levels of NKA-β in the endoplasmic reticulum (ER), which was associated with an increased expression of NKA-β at the plasma membrane (PM)
We show that, in alveolar epithelial cells, hypercapnia downregulates NKA function by promoting oxidation of NKA-β, which, thereby, impairs its interaction with NKA-α and retains the misfolded NKA-β in the ER

Summary

Introduction

Na,K-ATPase (NKA) is a heterodimeric enzyme and a member of the P-type ATPase family. A γ-subunit has been identified, which represents a family of single-span transmembrane proteins containing the FXYD motif that is not an integral part of the transporter but rather regulates the activity and membrane abundance of the enzyme [3,4]. The NKA β-subunit, which is a type II membrane glycoprotein, has a pivotal role in delivery and appropriate insertion of the NKA-α subunit in the PM [1]. Mice deficient in the NKA-β subunit in alveolar epithelial cells have reduced alveolar fluid clearance, which results in aggravation of acute lung injury (ALI) and further underlies the pivotal role of NKA-β in the overall transporter function [5]. Numerous reports have shown that the function of NKA-β is not limited to regulation of NKA-α, but is centrally involved in establishing epithelial cell polarity, formation of adherens junctions, and regulation of paracellular permeability, which are key for maintaining a functional epithelial barrier [6,7,8,9,10]

Methods

Results

Conclusion