Stimulation of oxygen consumption promotes mitochondrial calcium accumulation, a process associated with, and causally linked to, enhanced formation of tert-butylhydroperoxide-induced DNA single-strand breaks.
The NADH-linked substrates pyruvate, L-glutamine, and beta-hydroxybutyrate, while enhancing the rate of oxygen consumption, also increased the formation of DNA single-strand breaks induced by tert-butylhydroperoxide in intact U937 cells. A cause-effect relationship between these two parameters was established by showing that: (a) rotenone, an inhibitor of complex I, abolished respiration and prevented the enhancement of the DNA-damaging response under all the above circumstances; (b) the membrane-impermeant, complex I-activating substrate L-malate gave similar results in permeabilized cells; and (c) none of the NADH-linked substrates affected the DNA-damaging response to tert-butylhydroperoxide in respiration-deficient cells. Stimulation of electron transport potentiated the DNA-cleaving ability of tert-butylhydroperoxide via a process involving enforced mitochondrial calcium accumulation in the absence of a discernible elevation in the cytosolic concentration of free Ca2+. Finally, mitochondrial calcium was found to promote the mitochondrial formation of DNA-damaging levels of hydrogen peroxide. In conclusion, the data herein presented define a previously unexpected role of respiratory substrates in the control of the deleterious effects of an organic hydroperoxide at the level of genomic DNA. The enhanced DNA cleavage mediated by NADH-linked substrates in response to tert-butylhydroperoxide would appear to depend on a sequence of events involving stimulation of electron transport, mitochondrial accumulation of Ca2+, and mitochondrial formation of DNA-damaging levels of hydrogen peroxide via a Ca(2+)-dependent process.
Guidarelli
A
A
Istituto di Farmacologia e Farmacognosia, Università di Urbino, Italy.
Brambilla
L
L
Clementi
E
E
Sciorati
C
C
Cantoni
O
O
D020155:3-Hydroxybutyric Acid; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone / Q000494:pharmacology; D002374:Catalase / Q000494:pharmacology; D004249:DNA Damage; D004277:DNA, Single-Stranded / Q000187:drug effects*; D004579:Electron Transport / Q000187:drug effects; D005947:Glucose / Q000494:pharmacology; D005973:Glutamine / Q000494:pharmacology; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism / Q000633:toxicity; D006885:Hydroxybutyrates / Q000494:pharmacology; D007700:Kinetics; D007951:Leukemia, Myeloid; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism*; D009243:NAD / Q000378:metabolism; D010101:Oxygen Consumption / Q000187:drug effects / Q000502:physiology*; D010545:Peroxides / Q000633:toxicity*; D011773:Pyruvates / Q000494:pharmacology; D017382:Reactive Oxygen Species; D012402:Rotenone / Q000494:pharmacology*; D014407:Tumor Cells, Cultured; D020122:tert-Butylhydroperoxide
D004277:DNA, Single-Stranded; D006885:Hydroxybutyrates; D010545:Peroxides; D011773:Pyruvates; D017382:Reactive Oxygen Species; D012402:Rotenone; D005973:Glutamine; D009243:NAD; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; D002110:Caffeine; D020122:tert-Butylhydroperoxide; D006861:Hydrogen Peroxide; D002374:Catalase; D005947:Glucose; D002118:Calcium; D020155:3-Hydroxybutyric Acid