Endogenous NO regulates superoxide production at low oxygen concentrations by modifying the redox state of cytochrome c oxidase.
We have investigated in whole cells whether, at low oxygen concentrations ([O(2)]), endogenous nitric oxide (NO) modulates the redox state of the mitochondrial electron transport chain (ETC), and whether such an action has any signaling consequences. Using a polarographic-and-spectroscopic-coupled system, we monitored redox changes in the ETC cytochromes b(H), cc(1), and aa(3) during cellular respiration. The rate of O(2) consumption (VO(2)) remained constant until [O(2)] fell below 15 microM, whereas the onset of reduction of cytochromes aa(3), part of the terminal ETC enzyme cytochrome c oxidase, occurred at approximately 50 microM O(2). Incubation of the cells with an inhibitor of NO synthase lowered significantly (P < 0.05) the [O(2)] at which reduction of the cytochromes occurred. We also measured intracellular superoxide (O(2)(-)) production at different [O(2)] and found there was no increase in O(2)(-) generation in control cells, or those treated with the NO synthase inhibitor, when incubated at 21% O(2). However, after 30-min exposure of control cells to 3% O(2), an increase in O(2)(-) generation was observed, accompanied by translocation to the nucleus of the transcription factor NF-kappa B. Both of these responses were diminished by NO synthase inhibition. Our results suggest that endogenous NO, by enhancing the reduction of ETC cytochromes, contributes to a mechanism by which cells maintain their VO(2) at low [O(2)]. This, in turn, favors the release of O(2)(-), which initiates the transcriptional activation of NF-kappa B as an early signaling stress response.
Palacios-Callender
Miriam
M
Wolfson Institute for Biomedical Research, University College London, Cruciform Building, Gower Street, London WC1E 6BT, United Kingdom.
Quintero
Marisol
M
Hollis
Veronica S
VS
Springett
Roger J
RJ
Moncada
Salvador
S
D000818:Animals; D019069:Cell Respiration / Q000502:physiology; D003576:Electron Transport Complex IV / Q000378:metabolism*; D000860:Hypoxia / Q000378:metabolism*; D051379:Mice; D009000:Monocytes; D016328:NF-kappa B / Q000378:metabolism; D009569:Nitric Oxide / Q000378:metabolism*; D010084:Oxidation-Reduction; D013053:Spectrophotometry; D013481:Superoxides / Q000378:metabolism*
D016328:NF-kappa B; D013481:Superoxides; D009569:Nitric Oxide; D003576:Electron Transport Complex IV