of work: Based upon our previous finding that there is an increase with age in the activity of a mitochondrial DNA glycosylase/endonuclease (mtODE) which is specific for 8-OH deoxyguanosine (8-OHdG), we have hypothesized that there is an induction of this enzyme of DNA repair in response to chronic oxidative stress. This project tests this hypothesis. Rats were subjected to regimens described as causing oxidative damage to liver mitochondrial DNA (mt-DNA) and mitochondrial extracts were tested for activity in the incision of oligonucleotides containing a single 8-OHdG. Treatment with AZT for 6 weeks elicited no increase in mt-ODE activity, but also gave no increase in TBARS (a product of lipid peroxidation) or in superoxide dismutase (SOD) activity. Provision of 8% ethanol as the sole source of drinking water for periods of 6 weeks or 15 weeks, a procedure which has been reported to raise liver mt-DNA 8-OHdG content, also gave no change in mt-ODE activity relative to controls. Again, there was no change in TBARS or SOD activity, questioning the severity of the oxidative insult. Thus the role of oxidative stress in the induction of this enzyme activity remains to be resolved. However, we have extended and strengthened our original finding of an increase in mt-ODE activity with aging. In liver mitochondrial extracts, there was a highly significant increase in activity between 6 months and 23 months of age, with the activity being at a maximum at an intermediate age (12 months). The same pattern was seen in extracts of heart mitochondria. By contrast, two other mitochondrial enzymes of DNA metabolism which are not specifically involved in the repair of oxidative damage, viz. uracil DNA glycosylase (mtUDG) and AP endonuclease, were found to be either unchanged or minimally changed with aging. This lends credence to the idea that there is a specific induction of mt-ODE in aging, likely in response to oxidative stress. This project is being broadened to measure actual rates of DNA repair in cells which have been exposed briefly to light and methylene blue, to generate 8-OHdG in the mitochondrial genome. Experiments thus far have used primary cultures of hepatocytes from young and old rats. However, viability is much affected by methylene blue and other regimens for introducing 8-OHdG are being examined.
| Scheibye-Knudsen, Morten; Croteau, Deborah L; Bohr, Vilhelm A (2013) Mitochondrial deficiency in Cockayne syndrome. Mech Ageing Dev 134:275-83 |
| Ramamoorthy, Mahesh; Sykora, Peter; Scheibye-Knudsen, Morten et al. (2012) Sporadic Alzheimer disease fibroblasts display an oxidative stress phenotype. Free Radic Biol Med 53:1371-80 |
| Venø, Susanne T; Kulikowicz, Tomasz; Pestana, Cezar et al. (2011) The human Suv3 helicase interacts with replication protein A and flap endonuclease 1 in the nucleus. Biochem J 440:293-300 |
| Gredilla, Ricardo; Garm, Christian; Holm, Rikke et al. (2010) Differential age-related changes in mitochondrial DNA repair activities in mouse brain regions. Neurobiol Aging 31:993-1002 |
| Canugovi, Chandrika; Maynard, Scott; Bayne, Anne-Cécile V et al. (2010) The mitochondrial transcription factor A functions in mitochondrial base excision repair. DNA Repair (Amst) 9:1080-9 |
| de Souza-Pinto, Nadja C; Maynard, Scott; Hashiguchi, Kazunari et al. (2009) The recombination protein RAD52 cooperates with the excision repair protein OGG1 for the repair of oxidative lesions in mammalian cells. Mol Cell Biol 29:4441-54 |
| Maynard, Scott; Schurman, Shepherd H; Harboe, Charlotte et al. (2009) Base excision repair of oxidative DNA damage and association with cancer and aging. Carcinogenesis 30:2-10 |
| de Souza-Pinto, Nadja C; Wilson 3rd, David M; Stevnsner, Tinna V et al. (2008) Mitochondrial DNA, base excision repair and neurodegeneration. DNA Repair (Amst) 7:1098-109 |
| Bohr, V A; Ottersen, O P; Tonjum, T (2007) Genome instability and DNA repair in brain, ageing and neurological disease. Neuroscience 145:1183-6 |
| Weissman, L; de Souza-Pinto, N C; Stevnsner, T et al. (2007) DNA repair, mitochondria, and neurodegeneration. Neuroscience 145:1318-29 |
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