Proton channel blockers inhibit Duox activity independent of Hv1 effects
| Autor: | Adam Jaffe, Monica Valencia Gattas, Juliana Barahona, Gregory E. Conner |
|---|---|
| Rok vydání: | 2020 |
| Předmět: |
0301 basic medicine
Nigericin Clinical Biochemistry Bronchial epithelium Respiratory Mucosa Biochemistry Ion Channels Superoxide dismutase Duox 03 medical and health sciences chemistry.chemical_compound 0302 clinical medicine Extracellular Homeostasis Humans Channel blocker Secretion lcsh:QH301-705.5 lcsh:R5-920 NADPH oxidase biology Chemistry Superoxide Organic Chemistry NADPH Oxidases Epithelial Cells Hydrogen Peroxide Hydrogen-Ion Concentration Cell biology Zinc HEK293 Cells 030104 developmental biology lcsh:Biology (General) biology.protein Calcium lcsh:Medicine (General) Hv1 030217 neurology & neurosurgery Intracellular Research Paper |
| Zdroj: | Redox Biology Redox Biology, Vol 28, Iss, Pp-(2020) |
| ISSN: | 2213-2317 |
| DOI: | 10.1016/j.redox.2019.101346 |
| Popis: | The NADPH oxidase reaction produces protons. In the case of the NADPH oxidase, NOX2, activity depends on secretion of these protons and is inhibited by blockade of the voltage-gated proton channel (Hv1). Duox1 and Duox2 activities similarly produce intracellular protons but synthesize hydrogen peroxide directly instead of superoxide. Hv1 contributes to acid secretion in some epithelia that express Duox. To test the hypothesis that Duox activity is also sensitive to Hv1 channel blockers, Duox was assayed in the presence of either Zn2+ or 5-chloro-2-guanidinobenzimidazole (ClGBI). Both compounds inhibited Duox activity in normal human bronchial epithelial cells but with an IC50 over 10-fold higher than that reported for Hv1 (IC50 Zn2+ = 0.68 mM; IC50 ClGBI = 0.07–0.14 mM). Homogenized HEK293T cells expressing either Duox1 or Duox2 showed similar IC50 values for ClGBI suggesting these compounds inhibit the enzymes through alternate mechanisms independent of Hv1 proton secretion. Inclusion of superoxide dismutase did not restore Duox hydrogen peroxide synthesis. Addition of nigericin to eliminate any possible transmembrane pH gradients in intracellular membrane-localized Duox did not alter activity in HEK293T homogenates. Extracellular Zn2+ blocked intracellular Ca2+ increases needed for Duox activity. Together the data suggest that Duox enzyme activities in epithelia are inhibited by compounds that block Hv1 but inhibition occurs through Hv1-independent mechanisms and support the idea that Hv1 is not required for Duox activity. Graphical abstract Image 1 Highlights • Hv1 proton channel inhibitors block Duox in differentiated bronchial epithelial cells. • Zinc blocks Duox activity concurrently with reduction of calcium transients. • ClGBI, an inhibitor of Hv1, blocks Duox activity in homogenates of cells lacking Hv1. • In differentiated bronchial epithelia, Hv1 blockers did not alter intracellular pH. • H+/K+ ATPase inhibition acidified cytoplasm but did not block Duox activity. |
| Databáze: | OpenAIRE |
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