Introduction, distribution, and removal of 7-methylguanine in different liver chromatin fractions of young and old mice.

The distribution and elimination of 7-methylguanine (m7Gua) from different liver DNA chromatin fractions has been studied after treating young and old mice with N-methyl-N-nitrosourea (MNU). Guanine methylation kinetics was first studied in total liver DNA following intraperitoneal injections of 25 mg/kg and 50 mg/kg MNU does. MNU-induced DNA alkylation, as measured by m7Gua levels, was dose-dependent in liver tissues of young (9-11 month) and old mice (28-29 month). However, liver DNA in old mice incurred approximately 50% more damage than young mice for weight-normalized doses of MNU. The kinetics of adduct removal from total DNA was biphasic. A more rapid phase of m7Gua removal was observed during the first 24 h to 48 h following MNU administration; thereafter, the remaining m7Gua adducts were hydrolyzed much more slowly. Similar amounts of m7Gua were removed by 24 h at both the 25 mg/kg and 50 mg/kg MNU doses for a single age-group, but old liver tissue removed significantly more m7Gua than young liver tissue during this initial phase. Following a single injection of carcinogen (50 mg/kg), liver nuclei were isolated and chromatin was sheared by limited Micrococcal nuclease digestion. Chromatin was separated into nuclease-soluble, low-salt, high-salt and nuclear matrix fractions. All four fractions of young liver chromatin were methylated to the same degree. In contrast, there were differences in m7Gua levels between old liver chromatin fractions. DNA in the nuclease-sensitive fraction was most heavily alkylated, whereas nuclear matrix sequences were modified the least. Removal of m7Gua occurred at relatively uniform rates in all chromatin fractions regardless of age, indicating that m7Gua was not preferentially repaired in different nuclease-susceptible regions of chromatin. These results suggest that the N-methylpurine-DNA glycosylase responsible for eliminating m7Gua from the mammalian genome is not deficient in senescent liver tissue. However, there may be age-related changes in chromatin composition or structure that make some genomic sequences more accessible to alkylating agents in liver tissue of older animals.