RAD18 gene product of yeast Saccharomyces cerevisiae controls mutagenesis induced by hydrogen peroxide

Abstract

Oxygen and various types of peroxides generated in the process of cell metabolism are important sources of DNA damage. Peroxides can interact with different DNA groups, which may ultimately result in damage to bases and apurine/apyrimidine (AP) sites or the breaks of DNA strands [1]. In addition to direct DNA oxidation, bases damaged by peroxides can be incor porated into this molecule as a result of replication synthesis. An example of this process is the incorpora tion of 8 hydroxyguanine (8 oxoG) in DNA while using the oxidized derivative of guanine, namely 8 oxodeoxyguanine triphosphate, by DNA poly merases. This lesion has a high mutation potential due to the frequent pairing of 8 oxoG with adenine during replication [2, 3]. It is not surprising that during the evolution many organisms gained enzyme systems that protect their genome from 8 oxoG incorporation.The OGG1 gene of S. cerevisiae possesses DNA gly cosylase activity excising oxidized purines from DNA [4]. Deletion of the OGG1 gene has little effect on the cell sensitivity to DNA damaging agents, such as UV light, hydrogen peroxide or methylmethanesulfonate (MMS), and displays a 50 fold increase in spontane ously occurring GC TA transversions compared to wild type strain. The other base substitutions are not affected by the disruption of the OGG1 gene [5].The oxidation induced DNA damage is a substrate for the mismatch repair (MMR) system of yeast [6,7]. Genetic analysis showed that ogg1 msh2 and ogg1 msh6 double mutants provide a synergetic increase in spon taneous GC TA transversions compared to single mutants [7]. These results indicate that MMR excises adenine incorporated opposite 8 oxoG during the replication.The RAD6 and RAD18 genes play a critical role in postreplication repair (PRR) [8,9]. The Rad6-Rad18 complex possesses ubiquitin conjugating activity that catalyzes the monoubiquitylation of PCNA at lysine 164, which is a signal for PRR launch and attracts spe cialized DNA polymerases to the site of DNA damage [10]. Mutants of these genes are highly sensitive to the various genotoxic agents, such as gamma and UV rays, and alkylating agents [8,9]. They reduce the level of mutagenesis induced by various mutagens, in this case the efficiency of mutagenesis suppression depends on the type of damage and mutation system used for the quantification of mutations [11]. The rad6 and rad18 mutants are of the mutator phenotype and enhance only the rate of spontaneous GC TA transversions [12]. The research group headed by Fabre showed that during UV irradiation of rad18 mutant cells the fre quency of mutations, arising as a result of events related to PRR of pyrimidine dimers and 6,4 photo products (targeted mutagenesis), dramatically reduced, but increased significantly during bypass of some other types of DNA damage (nontargeted mutagenesis) [13].We hypothesized that "nontargeted" mutations that are stimulated during UV irradiation by mutations in rad6 and rad18 genes, arise from error prone syn thesis on a matrix DNA strand containing 8 oxoG that was ...