Overexpression of the Na⁺,K⁺-ATPase α₁ subunit increases Na⁺,K⁺-ATPase function in A549 cells

We hypothesized that viral mediated transfer of Na⁺,K⁺-ATPase subunit genes to alveolar epithelial cells to overexpress Na⁺,K⁺-ATPase could increase Na⁺,K⁺-ATPase function. We produced replication-deficient human type 5 adenoviruses that contained cytomegalovirus (CMV)-driven cDNAs for the rat α₁, and β₁ subunits of Na⁺,K⁺-ATPase (AdMRCMVα₁ and AdMRCMVβ₁, respectively). These viruses were used to transduce human adenocarcinoma cells (A549) in culture. Na⁺,K⁺-ATPase function was increased by 2.5-fold in the AdMRCMVα₁-infected cells. Sham and AdMRCMVβ₁-infected cells, and cells infected by a CMV-driven β-galactosidase-expressing adenovirus, had no increases in Na⁺,K⁺-ATPase activity. A549 cells infected with multiplicities of infection of 10-200 of AdMRCMVα₁ demonstrated expression of a rat α₁ mRNA and increased α₁ protein; no change in β₁ message or protein was noted. Ouabain sensitivity was measured in A549 cells following infection with AdMRCMVα₁. In contrast to controls, AdMRCMVα₁-infected cells demonstrated two IC₅₀s. The first was similar to the IC₅₀s of the controls: the second IC₅₀ was 2 logs greater than the first, consistent with the presence of both the rat and human α₁ isozymes. These results demonstrate for the first time that adenoviruses can be used to augment Na⁺,K⁺-ATPase function.