Am. J. Respir. Cell Mol. Biol., Volume 18, Number 6, June, 1998 741-749

Overexpression of the Na⁺,K⁺-ATPase ₁ Subunit Increases Na⁺,K⁺-ATPase Function in A549 Cells

Phillip Factor, Christina Senne, Vidas Dumasius, Karen Ridge, H. Ari Jaffe, Bruce Uhal, Zahn Gao, and J. Iasha Sznajder

Division of Pulmonary and Critical Care Medicine, Department of Medicine, Michael Reese Hospital, Chicago; and Division of Respiratory and Critical Care Medicine, Department of Medicine, University of Illinois at Chicago, Chicago, Illinois

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 ₁ and ₁ subunits of Na⁺,K⁺-ATPase (AdMRCMV₁ and AdMRCMV₁, 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 AdMRCMV₁-infected cells. Sham and AdMRCMV₁-infected cells, and cells infected by a CMV-driven -galactosidase-expressing adenovirus, had no increases in Na⁺,K⁺-ATPase activity. A549 cells infected with multiplicities of infection of 10-200 of AdMRCMV₁ demonstrated expression of a rat ₁ mRNA and increased ₁ protein; no change in ₁ message or protein was noted. Ouabain sensitivity was measured in A549 cells following infection with AdMRCMV₁. In contrast to controls, AdMRCMV₁-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 ₁ isozymes. These results demonstrate for the first time that adenoviruses can be used to augment Na⁺,K⁺-ATPase function.

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