NOX2 (gp91) is a predominant O2 sensor in a human airway chemoreceptor cell line: biochemical, molecular, and electrophysiological evidence

Buttigieg J, Pan J, Yeger H, Cutz E. NOX2 (gp91) is a predominant O2 sensor in a human airway chemoreceptor cell line: biochemical, molecular, and electrophysiological evidence. Am J Physiol Lung Cell Mol Physiol 303: L598–L607, 2012. First published August 1, 2012; doi:10.1152/ajplung.00170.2012.—Pulmonary neuroepithelial bodies (NEBs), composed of clusters of amine [serotonin (5-HT)] and peptide-producing cells, are widely distributed within the airway mucosa of human and animal lungs. NEBs are thought to function as airway O2-sensors, since they are extensively innervated and release 5-HT upon hypoxia exposure. The small cell lung carcinoma cell line (H146) provides a useful model for native NEBs, since they contain (and secrete) 5-HT and share the expression of a membrane-delimited O2 sensor [classical NADPH oxidase (NOX2) coupled to an O2-sensitive K channel]. In addition, both native NEBs and H146 cells express different NADPH oxidase homologs (NOX1, NOX4) and its subunits together with a variety of O2-sensitive voltage-dependent K channel proteins (Kv) and tandem pore acid-sensing K channels (TASK). Here we used H146 cells to investigate the role and interactions of various NADPH oxidase components in O2-sensing using a combination of coimmunoprecipitation, Western blot analysis (quantum dot labeling), and electrophysiology (patchclamp, amperometry) methods. Coimmunoprecipitation studies demonstrated formation of molecular complexes between NOX2 and Kv3.3 and Kv4.3 ion channels but not with TASK1 ion channels, while NOX4 associated with TASK1 but not with Kv channel proteins. Downregulation of mRNA for NOX2, but not for NOX4, suppressed hypoxia-sensitive outward current and significantly reduced hypoxia -induced 5-HT release. Collectively, our studies suggest that NOX2/Kv complexes are the predominant O2 sensor in H146 cells and, by inference, in native NEBs. Present findings favor a NEB cell-specific plasma membrane model of O2sensing and suggest that unique NOX/K channel combinations may serve diverse physiological functions.