EFFECTS OF DNA METHYLATION ON TOPOISOMERASE-I AND TOPOISOMERASE-II CLEAVAGE ACTIVITIES

DNA methylation is deregulated during oncogenesis. Since several major anti-cancer drugs act on topoisomerases, we investigated the effects of cytosine methylation on topoisomerase cleavage activities. Both top oisomerase I and II cleavage patterns were modified by CpG methylation in c-myc gene DNA fragments. Topoisomerase II changes, mainly cleavag e reduction, occurred for methylation sites within 7 base pairs from t he topoisomerase II breaks and were different for VM-26 and azatoxin. For topoisomerase I, cleavage enhancement as well as suppression were observed. Using synthetic methylated oligonucleotides, we show that he mimethylation is sufficient to alter topoisomerase I activity. Cytosin e methylation on the scissile strand within the topoisomerase I consen sus sequence had strong effects. Cleavage was stimulated by methylatio n at position -4 and was strongly inhibited by methylation at position -3 (with position -1 being the enzyme linked nucleotide). This inhibi tory effect was attributed to the presence of a methyl group in the ma jor groove, since the transition uracil to thymine also inhibited clea vage. Altogether these results suggest an interaction of topoisomerase I with the DNA major grove at positions -3 and -4. In addition, DNA m ethylation may have profound effects on the activity of topoisomerases and may alter the distribution of cleavage sites produced by anticanc er drugs in chromatin.