title: Dual Oxidase 2 (Duox2) Regulates Pannexin 1-mediated ATP Release in Primary Human Airway Epithelial Cells via Changes in Intracellular pH and Not H2O2 Production.
abstract: Human airway epithelial cells express pannexin 1 (Panx1) channels to release ATP, which regulates mucociliary clearance. Airway inflammation causes mucociliary dysfunction. Exposure of primary human airway epithelial cell cultures to IFN-γ for 48 h did not alter Panx1 protein expression but significantly decreased ATP release in response to hypotonic stress. The IFN-γ-induced functional down-regulation of Panx1 was due to the up-regulation of dual oxidase 2 (Duox2). Duox2 suppression by siRNA led to an increase in ATP release in control cells and restoration of ATP release in cells treated with IFN-γ. Both effects were reduced by the pannexin inhibitor probenecid. Duox2 up-regulation stoichiometrically increases H2O2 and proton production. H2O2 inhibited Panx1 function temporarily by formation of disulfide bonds at the thiol group of its terminal cysteine. Long-term exposure to H2O2, however, had no inhibitory effect. To assess the role of cellular acidification upon IFN-γ treatment, fully differentiated airway epithelial cells were exposed to ammonium chloride to alkalinize the cytosol. This led to a 2-fold increase in ATP release in cells treated with IFN-γ that was also inhibited by probenecid. Duox2 knockdown also partially corrected IFN-γ-mediated acidification. The direct correlation between intracellular pH and Panx1 open probability was shown in oocytes. Therefore, airway epithelial cells release less ATP in response to hypotonic stress in an inflammatory environment (IFN-γ exposure). Decreased Panx1 function is a response to cell acidification mediated by IFN-γ-induced up-regulation of Duox2, representing a novel mechanism for mucociliary dysfunction in inflammatory airway diseases.
authors.affiliation: From the Division of Pulmonary, Allergy, Critical Care, and Sleep Medicine and.
authors.forename: Stefanie
authors.initials: S
authors.lastname: Krick
authors.affiliation: Department of Physiology and Biophysics, University of Miami, Miami, Florida 33136 and.
authors.forename: Junjie
authors.initials: J
authors.lastname: Wang
authors.affiliation: From the Division of Pulmonary, Allergy, Critical Care, and Sleep Medicine and.
authors.forename: Melissa
authors.initials: M
authors.lastname: St-Pierre
authors.affiliation: the Interdisciplinary Center for Neuroscience of Valparaíso, Universidad de Valparaíso, Valparaíso, 2362735, Chile.
authors.forename: Carlos
authors.initials: C
authors.lastname: Gonzalez
authors.affiliation: Department of Physiology and Biophysics, University of Miami, Miami, Florida 33136 and.
authors.forename: Gerhard
authors.initials: G
authors.lastname: Dahl
authors.affiliation: From the Division of Pulmonary, Allergy, Critical Care, and Sleep Medicine and msalathe@med.miami.edu.
authors.forename: Matthias
authors.initials: M
authors.lastname: Salathe
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D001980:Bronchi / Q000166:cytology; D002478:Cells, Cultured; D017630:Connexins / Q000378:metabolism*; D000074623:Dual Oxidases; D004790:Enzyme Induction; D004847:Epithelial Cells / Q000201:enzymology*; D006801:Humans; D006861:Hydrogen Peroxide; D006863:Hydrogen-Ion Concentration; D007371:Interferon-gamma / Q000502:physiology; D008564:Membrane Potentials; D019255:NADPH Oxidases / Q000502:physiology*; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D009865:Oocytes / Q000201:enzymology; D061251:Primary Cell Culture; D020545:Respiratory Mucosa / Q000166:cytology; D014981:Xenopus
chemical_list: D017630:Connexins; D009419:Nerve Tissue Proteins; C492764:PANX1 protein, human; D007371:Interferon-gamma; D000255:Adenosine Triphosphate; D006861:Hydrogen Peroxide; D000074623:Dual Oxidases; D019255:NADPH Oxidases; C431515:DUOX2 protein, human
keywords: ATP; acidosis; dual oxidase 2; epithelial cell; hydrogen peroxide; pannexin
references.citation: J Physiol. 2004 Oct 15;560(Pt 2):519-32
references.pmid: 15308676
references.citation: Proc Natl Acad Sci U S A. 2008 Nov 4;105(44):17169-74
references.pmid: 18957549
references.citation: Br J Pharmacol. 1991 Jul;103(3):1649-56
references.pmid: 1718521
references.citation: Am J Physiol. 1992 May;262(5 Pt 1):C1313-23
references.pmid: 1590365
references.citation: Am J Respir Cell Mol Biol. 1993 Sep;9(3):315-22
references.pmid: 8398169
references.citation: Am J Physiol. 1995 Jun;268(6 Pt 1):L1021-8
references.pmid: 7611424
references.citation: J Physiol. 1995 Jul 1;486 ( Pt 1):123-30
references.pmid: 7562628
references.citation: Biophys J. 1996 Mar;70(3):1294-302
references.pmid: 8785285
references.citation: Am J Physiol. 1998 Apr;274(4 Pt 1):C904-13
references.pmid: 9575786
references.citation: J Neurosci. 1998 Sep 1;18(17):6740-7
references.pmid: 9712645
references.citation: Biophys J. 1998 Nov;75(5):2323-31
references.pmid: 9788927
references.citation: J Biol Chem. 1999 Feb 26;274(9):5581-7
references.pmid: 10026174
references.citation: J Gen Physiol. 1999 May;113(5):721-42
references.pmid: 10228184
references.citation: J Physiol. 2004 Nov 15;561(Pt 1):183-94
references.pmid: 15345749
references.citation: J Biol Chem. 2005 Mar 18;280(11):10646-54
references.pmid: 15615707
references.citation: FEBS Lett. 2005 Aug 29;579(21):4911-7
references.pmid: 16111680
references.citation: J Biol Chem. 2009 Jul 3;284(27):18143-51
references.pmid: 19416975
references.citation: Am J Respir Cell Mol Biol. 2009 Nov;41(5):525-34
references.pmid: 19213873
references.citation: Free Radic Biol Med. 2009 Nov 15;47(10):1450-8
references.pmid: 19703552
references.citation: Proc Natl Acad Sci U S A. 2010 Sep 7;107(36):15681-6
references.pmid: 20724658
references.citation: J Gen Physiol. 2010 Nov;136(5):515-27
references.pmid: 20937692
references.citation: Subcell Biochem. 2011;55:95-138
references.pmid: 21560046
references.citation: J Biol Chem. 2011 Jun 3;286(22):19830-9
references.pmid: 21454692
references.citation: Channels (Austin). 2011 May-Jun;5(3):193-7
references.pmid: 21532340
references.citation: J Biol Chem. 2011 Jul 29;286(30):26277-86
references.pmid: 21606493
references.citation: Plant Cell. 2012 Jun;24(6):2546-61
references.pmid: 22730405
references.citation: Inflamm Allergy Drug Targets. 2012 Aug 1;11(4):292-7
references.pmid: 22856520
references.citation: Sci Signal. 2013 Jun 11;6(279):ra46
references.pmid: 23757023
references.citation: Am J Respir Cell Mol Biol. 2013 Oct;49(4):672-9
references.pmid: 23742099
references.citation: Am J Physiol Lung Cell Mol Physiol. 2014 Mar 1;306(5):L453-62
references.pmid: 24414257
references.citation: Biochim Biophys Acta. 2014 May;1840(5):1596-604
references.pmid: 24060746
references.citation: Channels (Austin). 2014;8(2):118-23
references.pmid: 24576994
references.citation: Glia. 2014 Sep;62(9):1486-501
references.pmid: 24839011
references.citation: PLoS One. 2014;9(8):e106269
references.pmid: 25170954
references.citation: J Physiol. 2014 Nov 1;592(21):4747-62
references.pmid: 25172944
references.citation: Am J Respir Crit Care Med. 2015 Jul 15;192(2):182-90
references.pmid: 25909230
references.citation: J Biol Chem. 2015 Oct 16;290(42):25710-6
references.pmid: 26338706
references.citation: Cell Calcium. 2001 Dec;30(6):361-71
references.pmid: 11728131
references.citation: Chest. 2003 Nov;124(5):1909-15
references.pmid: 14605067
references.citation: J Pharmacol Exp Ther. 2009 Feb;328(2):409-18
references.pmid: 19023039
references.citation: Am J Physiol Cell Physiol. 2009 Feb;296(2):C250-5
references.pmid: 18945939
references.citation: Proc Natl Acad Sci U S A. 2003 Nov 11;100(23):13644-9
references.pmid: 14597722
references.citation: Circ Res. 2004 Feb 6;94(2):215-22
references.pmid: 14699011
references.citation: FEBS Lett. 2004 Aug 13;572(1-3):65-8
references.pmid: 15304325
references.citation: J Biol Chem. 2004 Aug 27;279(35):36454-61
references.pmid: 15210697
references.citation: FEBS Lett. 2006 Jan 9;580(1):239-44
references.pmid: 16364313
references.citation: Proc Natl Acad Sci U S A. 2006 May 16;103(20):7655-9
references.pmid: 16682648
references.citation: J Biol Chem. 2006 Aug 11;281(32):22992-3002
references.pmid: 16754672
references.citation: Inflamm Allergy Drug Targets. 2006 Dec;5(4):253-6
references.pmid: 17168796
references.citation: FEBS Lett. 2007 Jul 10;581(17):3241-6
references.pmid: 17586501
references.citation: J Cell Sci. 2007 Nov 1;120(Pt 21):3772-83
references.pmid: 17925379
references.citation: Antioxid Redox Signal. 2008 Apr;10(4):705-14
references.pmid: 18177232
references.citation: Methods Mol Med. 2005;107:183-206
references.pmid: 15492373