Abstract
Bacteria, yeast and human cancer cells possess mechanisms of mutagenesis upregulated by stress responses. Stress-inducible mutagenesis potentially accelerates adaptation, and may provide important models for mutagenesis that drives cancers, host pathogen interactions, antibiotic resistance and possibly much of evolution generally. In Escherichia coli repair of double-strand breaks (DSBs) becomes mutagenic, using low-fidelity DNA polymerases under the control of the SOS DNA-damage response and RpoS general stress response, which upregulate and allow the action of error-prone DNA polymerases IV (DinB), II and V to make mutations during repair. Pol IV is implied to compete with and replace high-fidelity DNA polymerases at the DSB-repair replisome, causing mutagenesis. We report that up-regulated Pol IV is not sufficient for mutagenic break repair (MBR); damaged bases in the DNA are also required, and that in starvation-stressed cells, these are caused by reactive-oxygen species (ROS). First, MBR is reduced by either ROS-scavenging agents or constitutive activation of oxidative-damage responses, both of which reduce cellular ROS levels. The ROS promote MBR other than by causing DSBs, saturating mismatch repair, oxidizing proteins, or inducing the SOS response or the general stress response. We find that ROS drive MBR through oxidized guanines (8-oxo-dG) in DNA, in that overproduction of a glycosylase that removes 8-oxo-dG from DNA prevents MBR. Further, other damaged DNA bases can substitute for 8-oxo-dG because ROS-scavenged cells resume MBR if either DNA pyrimidine dimers or alkylated bases are induced. We hypothesize that damaged bases in DNA pause the replisome and allow the critical switch from high fidelity to error-prone DNA polymerases in the DSB-repair replisome, thus allowing MBR. The data imply that in addition to the indirect stress-response controlled switch to MBR, a direct cis-acting switch to MBR occurs independently of DNA breakage, caused by ROS oxidation of DNA potentially regulated by ROS regulators.
Highlights
Spontaneous mutations drive development of most cancers and their resistance to therapy, aging, pathogen escape from the immune response and antibiotics, and evolution generally
Mutagenesis mechanisms upregulated by stress responses promote de novo antibiotic resistance and cross resistance in bacteria, anti-fungal-drug resistance in yeasts, and genome instability in cancer cells under hypoxic stress
A widely useful model mechanism is mutagenic DNA break repair in Escherichia coli, in which activation of two stress responses allows error-prone DNA polymerase in the break-repair replisome and introduce misincorporations, later fixed as mutations. Both stress responses upregulate the error-prone mutagenic DNA polymerase Pol IV (DinB), suggesting that the regulation of mutagenesis to times of stress is accomplished by indirect gene upregulation, followed by DNA polymerase competition
Summary
Spontaneous mutations drive development of most cancers and their resistance to therapy, aging, pathogen escape from the immune response and antibiotics, and evolution generally. That DNA damage underlies much of spontaneous mutagenesis was supported by discoveries that yeast antimutator mutants, i.e., mutants with lower-than-normal spontaneous mutation rate, carry mutations in DNA-damage-survival genes [17]. These genes encode alternative error-prone DNA polymerases or proteins that assist them, allowing survival of DNA damage by replication over or extension from otherwise-replication-inhibiting damaged DNA bases [18], implying that most spontaneous mutagenesis in yeast results from use of errorprone DNA polymerases during DNA-damage survival. The only mechanistic detail available on spontaneous mutagenesis mechanisms is that about half of all spontaneous base-substitutions and small insertion/deletions (indels) in starving Escherichia coli form dependently on the proteins used in stress-inducible mutagenic DNA break repair (MBR) [7]
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