SOS and UVM pathways have lesion-specific additive and competing effects on mutation fixation at replication-blocking DNA lesions

Escherichia coli cells have multiple mutagenic pathways that are induced in response to environmental and physiological stimuli. Unlike the well-inves tigated classical SOS response, little is known about newly recognized path ways such as the UVM (UV modulation of mutagenesis) response, In this study , we compared the contributions of the SOS and UVM pathways on mutation fix ation at two representative noninstructive DNA lesions: 3,N-4-ethenocytosin e (epsilon C) and abasic (AP) sites, Because both SOS and UVM responses are induced by DNA damage, and defined UVM-defective E. coli strains are not y et available, we first constructed strains in which expression of the SOS m utagenesis proteins UmuD' and UmuC (and also RecA in some cases) is uncoupl ed from DNA damage by being placed under the control of a heterologous lac- derived promoter, M13 single-stranded viral DNA bearing site-specific lesio ns was transfected into cells induced for the SOS or UVM pathway. Survival effects were determined from transfection efficiency, and mutation fixation at the lesion was analyzed by a quantitative multiplex sequence analysis p rocedure, Our results suggest that induction of the SOS pathway can indepen dently elevate mutagenesis at both lesions, whereas the UVM pathway signifi cantly elevates mutagenesis at EC in an SOS-independent fashion and at AP s ites in an SOS-dependent fashion, Although mutagenesis at EC appears to be elevated by the induction of either the SOS or the UVM pathway, the mutatio nal specificity profiles for EC under SOS and UVM pathways are distinct. In terestingly, when both pathways are active, the UVM effect appears to predo minate over the SOS effect on mutagenesis at eC, but the total mutation fre quency is significantly increased over that observed when each pathway is i ndividually induced. These observations suggest that the UVM response affec ts mutagenesis not only at class 2 noninstructive lesions (EC) but also at classical SOS-dependent (class 1) lesions such as AP sites. Our results add new layers of complexity to inducible mutagenic phenomena: DNA damage acti vates multiple pathways that have lesion-specific additive as well as suppr essive effects on mutation fixation, and some of these pathways are not dir ectly regulated by the SOS genetic network.