Mitochondrial GPx1 decreases induced but not basal oxidative damage to mtDNA in T47D cells

The production of oxyradicals by mitochondria (mt) is a source of oxidative damage to mtDNA such as 8-oxo-dG lesions that may lead to mutations and mi tochondrial dysfunction. The potential protection of mtDNA by glutathione p eroxidase-1 (GPx1) was investigated in GPx1-proficient (GPx-2) and GPx1-def icient (Hygro-3) human breast T47D cell transfectants. GPx activity and GPx 1-like antigen concentration in mitochondria were respectively at least 100 -fold and 20- to 25-fold higher in GPx2 than Hygro-3 cells. In spite of thi s large difference in peroxide-scavenging capacity, the basal 8-oxo-dG freq uency in mtDNA, assessed by carefully controlled postlabeling assay, was st rikingly similar in both cell lines. In contrast, in response to menadione- mediated oxidative stress, induction of 8-oxo-dG and DNA strand breaks was much lower in the GPx1-proficient mitochondria (e.g., +14% 8-oxo-dG versus +54% in Hygro-3 after 1-h exposure to 25 mu M menadione, P < 0.05). Our dat a indicate that the mitochondrial glutathione/GPx1 system protected mtDNA a gainst damage induced by oxidative stress, but did not prevent basal oxidat ive damage to mtDNA, which, surprisingly, appeared independent of GPx1 stat us in the T47D model. (C) 2000 Academic Press.