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Mitochondrial reactive oxygen species generated at the complex-II matrix or intermembrane space microdomain have distinct effects on redox signaling and stress sensitivity in caenorhabditis elegans
journal contribution
posted on 2019-01-01, 00:00 authored by Adam Trewin, Laura L Bahr, Anmol Almast, Brandon J Berry, Alicia Y Wei, Thomas H Foster, Andrew P WojtovichAims: How mitochondrial reactive oxygen species (ROS) impact physiological function may depend on the quantity of ROS generated or removed, and the subcellular microdomain in which this occurs. However, pharmacological tools currently available to alter ROS production in vivo lack precise spatial and temporal control.
Results: We used CRISPR/Cas9 to fuse the light-sensitive ROS-generating protein, SuperNova to the C-terminus of mitochondrial complex II succinate dehydrogenase subunits B (SDHB-1::SuperNova) and C (SDHC-1::SuperNova) in Caenorhabditis elegans to localize SuperNova to the matrix-side of the inner mitochondrial membrane, and to the intermembrane space (IMS), respectively. The presence of the SuperNova protein did not impact complex II activity, mitochondrial respiration, or C. elegans development rate under dark conditions. ROS production by SuperNova protein in vitro in the form of superoxide (O2•-) was both specific and proportional to total light irradiance in the 540-590 nm spectra, and was unaffected by varying the buffer pH to resemble the mitochondrial matrix or IMS environments. We then determined using SuperNova whether stoichiometric ROS generation in the mitochondrial matrix or IMS had distinct effects on redox signaling in vivo. Phosphorylation of PMK-1 (a p38 MAPK homolog) and transcriptional activity of SKN-1 (an Nrf2 homolog) were each dependent on both the site and duration of ROS production, with matrix-generated ROS having more prominent effects. Furthermore, matrix- but not IMS-generated ROS attenuated susceptibility to simulated ischemia reperfusion injury in C. elegans.
Innovation and Conclusion: Overall, these data demonstrate that the physiological output of ROS depends on the microdomain in which it is produced.
Results: We used CRISPR/Cas9 to fuse the light-sensitive ROS-generating protein, SuperNova to the C-terminus of mitochondrial complex II succinate dehydrogenase subunits B (SDHB-1::SuperNova) and C (SDHC-1::SuperNova) in Caenorhabditis elegans to localize SuperNova to the matrix-side of the inner mitochondrial membrane, and to the intermembrane space (IMS), respectively. The presence of the SuperNova protein did not impact complex II activity, mitochondrial respiration, or C. elegans development rate under dark conditions. ROS production by SuperNova protein in vitro in the form of superoxide (O2•-) was both specific and proportional to total light irradiance in the 540-590 nm spectra, and was unaffected by varying the buffer pH to resemble the mitochondrial matrix or IMS environments. We then determined using SuperNova whether stoichiometric ROS generation in the mitochondrial matrix or IMS had distinct effects on redox signaling in vivo. Phosphorylation of PMK-1 (a p38 MAPK homolog) and transcriptional activity of SKN-1 (an Nrf2 homolog) were each dependent on both the site and duration of ROS production, with matrix-generated ROS having more prominent effects. Furthermore, matrix- but not IMS-generated ROS attenuated susceptibility to simulated ischemia reperfusion injury in C. elegans.
Innovation and Conclusion: Overall, these data demonstrate that the physiological output of ROS depends on the microdomain in which it is produced.
History
Journal
Antioxidants & redox signalingVolume
31Issue
9Pagination
594 - 607Publisher
Mary Ann LiebertLocation
New Rochelle, N.Y.Publisher DOI
ISSN
1523-0864eISSN
1557-7716Language
engPublication classification
C1 Refereed article in a scholarly journalCopyright notice
2019, Mary Ann LiebertUsage metrics
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SuperNovaischemia reperfusion injurymitohormesisoptogeneticsphotosensitizerreactive oxygen speciessuperoxideScience & TechnologyLife Sciences & BiomedicineBiochemistry & Molecular BiologyEndocrinology & MetabolismOXIDATIVE STRESSHYDROGEN-PEROXIDESUPEROXIDE DISMUTASESOPTOGENETIC CONTROLMAP-KINASESC-ELEGANSSKN-1ROSINACTIVATIONACCUMULATION
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