Department of Biochemistry and Molecular Pharmacology; Program in Molecular Medicine
Adenosine Triphosphatases; Alkylating Agents; DNA Helicases; *DNA Methylation; DNA Polymerase I; *DNA Repair; Escherichia coli; Escherichia coli Proteins; Genes, Bacterial; Methyl Methanesulfonate; Methylnitronitrosoguanidine; Mutation; *Recombination, Genetic
Life Sciences | Medicine and Health Sciences
Methylating agents such as N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) and methyl methane sulfonate (MMS) produce a wide variety of N- and O-methylated bases in DNA, some of which can block replication fork progression. Homologous recombination is a mechanism by which chromosome replication can proceed despite the presence of lesions. The two major recombination pathways, RecBCD and RecFOR, which repair double-strand breaks (DSBs) and single-strand gaps respectively, are needed to protect against toxicity with the RecBCD system being more important. We find that recombination-deficient cell lines, such as recBCD recF, and ruvC recG, are as sensitive to the cytotoxic effects of MMS and MNNG as the most base excision repair (BER)-deficient (alkA tag) isogenic mutant strain. Recombination and BER-deficient double mutants (alkA tag recBCD) were more sensitive to MNNG and MMS than the single mutants suggesting that homologous recombination and BER play essential independent roles. Cells deleted for the polA (DNA polymerase I) or priA (primosome) genes are as sensitive to MMS and MNNG as alkA tag bacteria. Our results suggest that the mechanism of cytotoxicity by alkylating agents includes the necessity for homologous recombination to repair DSBs and single-strand gaps produced by DNA replication at blocking lesions or single-strand nicks resulting from AP-endonuclease action.
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Citation: Nucleic Acids Res. 2006 May 2;34(8):2258-68. Print 2006. Link to article on publisher's site