Mutagenesis at 3,N4 -ethenocytosine (C), a nonpairing mutagenic lesion, is significantly enhanced in Escherichia coli cells pretreated with UV, alkylating agents, or H 2 O 2 . This effect, termed UVM (for UV modulation of mutagenesis), is distinct from known DNA damage-inducible responses, such as the SOS response, the adaptive response to alkylating agents, or the oxyR-mediated response to oxidative agents. Here, we have addressed the hypothesis that UVM results from transient depletion of a mismatch repair activity that normally acts to reduce mutagenesis. To test whether the loss of mismatch repair activities results in the predicted constitutive UVM phenotype, E. coli cells defective for methyl-directed mismatch repair, for veryshort-patch repair, or for the N-glycosylase activities MutY and MutM were treated with the UVM-inducing agent 1-methyl-3-nitro-1-nitrosoguanidine, with subsequent transfection of M13 viral single-stranded DNA bearing a site-specific C lesion. Survival of the M13 DNA was measured as transfection efficiency, and mutation fixation at the lesion was characterized by multiplex sequencing technology. The results showed normal UVM induction patterns in all the repair-defective strains tested. In addition, normal UVM induction was observed in cells overexpressing MutH, MutL, or MutS. All strains displayed UVM reactivation, the term used to describe the increased survival of C-containing DNA in UVM-induced cells. Taken together, these results indicate that the UVM response is independent of known mismatch repair systems in E. coli and may thus represent a previously unrecognized misrepair or misreplication pathway.The term inducible mutagenesis has been used to describe the transient mutator phenotype observed in cells exposed to UV or other mutagens. In the well-studied Escherichia coli model, a set of inducible genes constituting the SOS response is presumed to mediate this phenotype. It is widely accepted that induced SOS proteins are required for facilitating translesion synthesis at blocks to replication created by DNA damage (7, 9, 32) and that this translesion synthesis is accomplished by lowering the fidelity of DNA replication, accounting for so-called SOS mutagenesis. However, the biochemical mechanism that facilitates the replication of damaged DNA remains undescribed.Our investigation of the mutagenic properties of the nonpairing lesion 3,N 4 -ethenocytosine (εC) has revealed the existence of a novel UV-inducible mutagenic response in E. coli, termed UVM (for UV modulation of mutagenesis) (34). The UVM effect is observed as a significant increase in mutagenesis at an εC residue borne on M13 viral single-stranded DNA (ssDNA) in E. coli cells pretreated with UV, alkylating agents, or H 2 O 2 . This response has been shown to be distinct from known DNA damage-inducible responses, such as the SOS response, the adaptive response to alkylating agents, or the oxyR-mediated response to oxidative agents (38,51,52). εC, the model lesion used in our studies, is inflicted by metabolites of se...

UVM (ultraviolet modulation of mutagenesis) is a recently described recA-independent, inducible mutagenic phenomenon in which prior UV irradiation of Escherichia coli cells strongly enhances mutation fixation at a site-specific 3-N4-ethenocytosine (epsilon C) lesion borne on a transfected single-stranded M13 DNA vector. Subsequent studies demonstrated that UVM is also induced by alkylating agents, and is distinct from both the SOS response and the adaptive response to alkylation damage. Because of the increasing significance being attributed to oxidative DNA damage, it is interesting to ask whether this class of DNA damage can also induce UVM. By transfecting M13 vector DNA bearing a site-specific epsilon C lesion into cells pretreated with inducing agents, we show here that the oxidative agent H2O2 is a potent inducer of UVM, and that the induction of UVM by H2O2 does not require oxyR-regulated gene expression. UVM induction by H2O2 appears to be mediated by DNA damage, as indicated by the observation of a concomitant reduction in cellular toxicity and UVM response in OxyRc cells. Available evidence suggests that UVM represents a generalized cellular response to a broad range of chemical and physical genotoxicants, and that DNA damage constitutes the most likely signal for its induction.

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