EFFECT OF DNA CYTOSINE METHYLATION UPON DEAMINATION-INDUCED MUTAGENESIS IN A NATURAL TARGET SEQUENCE IN DUPLEX DNA

Are 5-methylcytosine residues in DNA hot spots for transition mutagene sis? Numerous studies identify 1) structural changes induced by DNA me thylation, 2) high percentages of human mutations that result from GC to AT transition pathways, and 3) differences between G.C and G.mC bas e pairs in susceptibility to nonenzymatic deamination. However, invest igations of chemical stability necessarily involve non-physiological c onditions for chemical analysis of deamination. Here we describe an ex periment that compares rates of deamination-induced mutagenesis betwee n a G.C and G.mC base pair, when both are present in duplex DNA, incub ated at 37 degrees C and pH 7.4, within identical sequence contexts, i n a natural mutational target (the Escherichia coli lacZ alpha gene) t hat selects for mutagenesis at the specific site under investigation. Under these conditions the rate of spontaneous deamination at G.mC exc eeds that at G.C by more than 21-fold. Our data implicate differences in chemical stability toward deamination as a major causal factor rela ting DNA cytosine methylation to spontaneous mutagenesis.