Oxidative Stress Disrupts Glucocorticoid Hormone-dependent Transcription of the Amiloride-sensitive Epithelial Sodium Channel α-Subunit in Lung Epithelial Cells through ERK-dependent and Thioredoxin-sensitive Pathways

• Published in: The Journal of biological chemistry 2000-03, Vol.275 (12), p.8600-8609
• Main Authors: Wang, Hao-Chien, Zentner, Mark D., Deng, Hong-Tao, Kim, Kwang-Jin, Wu, Reen, Yang, Pan-Chyr, Ann, David K.
• Format: Article
• Language: English
• Subjects: aENaC gene; Amiloride; Butadienes - pharmacology; Dexamethasone - metabolism; Epithelial Cells - cytology; Epithelial Cells - metabolism; Epithelial Sodium Channels; ERK protein; Gene Expression Regulation; Glucocorticoids - metabolism; Humans; Hydrogen Peroxide - metabolism; JNK Mitogen-Activated Protein Kinases; Lung - cytology; Lung - metabolism; MAP Kinase Kinase Kinase 1; Mitogen-Activated Protein Kinases - antagonists & inhibitors; Mitogen-Activated Protein Kinases - metabolism; Models, Biological; Nitriles - pharmacology; Oxidation-Reduction; Oxidative Stress - physiology; Protein-Serine-Threonine Kinases - metabolism; ras Proteins - metabolism; Receptor Cross-Talk; Receptors, Glucocorticoid - metabolism; Respiratory Mucosa - cytology; Respiratory Mucosa - metabolism; Response Elements; Signal Transduction; Sodium Channels - genetics; Thioredoxin; Thioredoxins - metabolism; Transcriptional Activation; Tumor Cells, Cultured; Up-Regulation
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.275.12.8600

The amiloride-sensitive epithelial Na+ channel (ENaC) plays a critical role in the maintenance of alveolar fluid balance. It is generally accepted that reactive oxygen and nitrogen species can inhibit ENaC activity and aggravate acute lung injury; however, the molecular mechanism for free radical-mediated ENaC inhibition is unclear. Previously, we showed that the expression of the α-subunit of ENaC,α-ENaC, which is indispensable for ENaC activity, is repressed by Ras activation in salivary epithelial cells. Here, we investigated whether exogenous H2O2 modulatesα-ENaC gene expression in lung epithelial cells through a similar molecular mechanism. Utilizing transient transfection reporter assays and site-directed mutagenesis analyses, we found that the glucocorticoid response element (GRE), located at −1334 to −1306 base pairs of the α-ENaC 5′-flanking region, is the major enhancer for the stimulated α-ENaC expression in A549 lung epithelial cells. We further demonstrate that the presence of an intact GRE is necessary and sufficient for oxidants to repressα-ENaC expression. Consistent with our hypothesis, exogenous H2O2-mediated repression ofα-ENaC GRE activity is partially blocked by either a specific inhibitor for extracellular signal-regulated kinase (ERK) pathway activation, U0126, or dominant negative ERK, suggesting that, in part, activated ERK may mediate the repressive effects of H2O2 on α-ENaC expression. In addition, overexpression of thioredoxin restored glucocorticoid receptor action on the α-ENaC GRE in the presence of exogenous H2O2. Taken together, we hypothesize that oxidative stress impairs Na+ transport activity by inhibiting dexamethasone-dependent α-ENaC GRE activation via both ERK-dependent and thioredoxin-sensitive pathways. These results suggest a putative mechanism whereby cellular redox potentials modulate the glucocorticoid receptor/dexamethasone effect on α-ENaC expression in lung and other tight epithelia.