Covalent linkage between DNA and proteins produces highly toxic lesions and can be caused by commonly used chemotherapeutic agents, by internal and external chemicals and by radiation. In this study, using Escherichia coli, we investigate the consequences of 5-azacytidine (5-azaC), which traps covalent complexes between itself and the Dcm cytosine methyltransferase protein. DNA protein crosslink-dependent effects can be ascertained by effects that arise in wild-type but not in dcmΔ strains. We find that 5-azaC induces the bacterial DNA damage response and stimulates homologous recombination, a component of which is Dcm-dependent. Template-switching at an imperfect inverted repeat ("quasipalindrome", QP) is strongly enhanced by 5-azaC and this enhancement was entirely Dcm-dependent and independent of double-strand break repair. The SOS response helps ameliorate the mutagenic effect of 5-azaC but this is not a result of SOS-induced DNA polymerases since their induction, especially PolIV, seems to stimulate QP-associated mutagenesis. Cell division regulator SulA was also required for recovery of QP mutants induced by 5-azaC. In the absence of Lon protease, Dcm-dependent QP-mutagenesis is strongly elevated, suggesting it may play a role in DPC tolerance. Deletions at short tandem repeats, which occur likewise by a replication template-switch, are elevated, but only modestly, by 5-azaC. We see evidence for Dcm-dependent and-independent killing by 5-azaC in sensitive mutants, such as recA, recB, and lon; homologous recombination and deletion mutations are also stimulated in part by a Dcm-independent effect of 5-azaC. Whether this occurs by a different protein/DNA crosslink or by an alternative form of DNA damage is unknown.
Keywords: DNA damage response; DNA repair; Genetic rearrangements; Genome instability; Homologous recombination.
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