title: NOX2 (gp91phox) is a predominant O2 sensor in a human airway chemoreceptor cell line: biochemical, molecular, and electrophysiological evidence.
abstract: 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 O(2)-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 O(2) sensor [classical NADPH oxidase (NOX2) coupled to an O(2)-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 O(2)-sensitive voltage-dependent K(+) channel proteins (K(v)) 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 O(2)-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 K(v)3.3 and K(v)4.3 ion channels but not with TASK1 ion channels, while NOX4 associated with TASK1 but not with K(v) 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/K(v) complexes are the predominant O(2) sensor in H146 cells and, by inference, in native NEBs. Present findings favor a NEB cell-specific plasma membrane model of O(2)-sensing and suggest that unique NOX/K(+) channel combinations may serve diverse physiological functions.
authors.affiliation: Division of Pathology, Dept. of Paediatric Laboratory Medicine, The Hospital for Sick Children, Toronto, Ontario, Canada.
authors.forename: Josef
authors.initials: J
authors.lastname: Buttigieg
authors.forename: Jie
authors.initials: J
authors.lastname: Pan
authors.forename: Herman
authors.initials: H
authors.lastname: Yeger
authors.forename: Ernest
authors.initials: E
authors.lastname: Cutz
mesh_terms: D045744:Cell Line, Tumor; D002628:Chemoreceptor Cells / Q000502:physiology*; D015536:Down-Regulation; D006801:Humans; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000502:physiology*; D000074662:NADPH Oxidase 2; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases / Q000737:chemistry / Q000502:physiology*; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000502:physiology; D046568:Neuroepithelial Bodies / Q000502:physiology*; D010100:Oxygen / Q000502:physiology*; D024683:Potassium Channels, Tandem Pore Domain / Q000737:chemistry / Q000502:physiology; D012701:Serotonin / Q000378:metabolism; D051672:Shal Potassium Channels / Q000737:chemistry / Q000502:physiology; D051670:Shaw Potassium Channels / Q000737:chemistry / Q000502:physiology
chemical_list: C489594:KCNC3 protein, human; D008562:Membrane Glycoproteins; D009419:Nerve Tissue Proteins; D024683:Potassium Channels, Tandem Pore Domain; D051672:Shal Potassium Channels; D051670:Shaw Potassium Channels; C420132:potassium channel subfamily K member 3; D012701:Serotonin; C056558:CYBB protein, human; D000074662:NADPH Oxidase 2; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases; C498910:NOX4 protein, human; D010100:Oxygen