ALTERATIONS OF DNA METHYLATION BY GLUTATHIONE DEPLETION

One of the most consistent findings in cancer cells is an overall decr ease of 5-methylcytosine content in DNA. The causes that lead to this alteration are not known. We have shown in a recent study that the met hyl-donor, methionine (Met), can easily be depleted and that O- and S- methylation can be impaired in response to glutathione (GSH) depletion . This is because mammalian cells are capable of resynthesizing GSH af ter GSH is depleted, and GSH turnover occurs at the expense of Met. An extensive utilization of Met for the resynthesis of GSH causes Met de pletion and impairment in methylation. In the present study we now dem onstrate that GSH depletion has a significant impact on DNA methylatio n. An i.p. dose of a model GSH-depleting hepatotoxin, bromobenzene (BB ), caused a progressive impairment in genomic DNA methylation in the S yrian hamster. The administration of a single i.p. dose of Met labeled with [(CH3)-C-14]Met to BB-treated hamsters at either 1, 3, 5.5 or 9 h after BB resulted in an increase of methyl-group incorporation into liver genomic DNA at 24 h after BB. With respect to the time points ch osen for Met administration, methyl-group incorporation found in the B B + Met groups were 1-, 2-, 4- and 12-fold of the controls that receiv ed only Met. We further employed an in vitro methylation assay using s pecific bacterial SssI CPG methylase as the catalyzing enzyme to demon strate that BB caused a progressive increase of unmethylated C(p)G sit es in genomic DNA. Interestingly, the time response curve of global DN A methylation in vitro showed an identical pattern to that observed in the in vivo experiment. The results provide strong evidence that GSH- depleting agents significantly impair cytosine methylation. Thus, alte rations in gene expression could result from a high dose and/or prolon ged exposure to GSH-depleting agents, e.g. medications, chemotherapeut ic agents and environmental toxins. (C) 1997 Elsevier Science Ireland Ltd.