Stress-Induced Mutagenesis, Gambler Cells, and Stealth Targeting Antibiotic-Induced Evolution

Abstract

Mechanisms of evolution and evolution of antibiotic resistance are both fundamental and world health problems. Stress-induced mutagenesis defines mechanisms of mutagenesis upregulated by stress responses, which drive adaptation when cells are maladapted to their environments-when stressed. Work in mutagenesis induced by antibiotics had produced tantalizing clues but not coherent mechanisms. We review recent advances in antibiotic-induced mutagenesis that integrate how reactive oxygen species (ROS), the SOS and general stress responses, and multichromosome cells orchestrate a stress response-induced switch from high-fidelity to mutagenic repair of DNA breaks. Moreover, while sibling cells stay stable, a mutable "gambler" cell subpopulation is induced by differentially generated ROS, which signal the general stress response. We discuss other evolvable subpopulations and consider diverse evolution-promoting molecules as potential targets for drugs to slow evolution of antibiotic resistance, cross-resistance, and immune evasion. An FDA-approved drug exemplifies "stealth" evolution-slowing drugs that avoid selecting resistance to themselves or antibiotics.

Keywords: antibiotic resistance; antibiotics; antievolvability drugs; cell subpopulations; drug resistance evolution; evolution; evolvability; stress-induced mutagenesis.

FIG 1

Temporal regulation of mutagenesis by stress responses in E. coli mutagenic break repair. (Step 1) DNA double-strand breaks (DSBs) are generated by various processes and can then be repaired by homologous or microhomologous repair mechanisms. (Top) During homology-directed DSB repair (HR) (reviewed in references and 75), ssDNA exposed at the DSB ends base pairs with complementary sequence in a sister chromosome, promoting repair DNA synthesis. (Step 2) DSBs also induce the SOS response, which transcriptionally upregulates the error-prone DNA polymerases (Pols) IV, V, and II (82); however, repair remains accurate unless another stressor induces the general stress (σ^S) response (44, 45). (Step 3) The σ^S response induces two kinds of switches to mutagenic DSB repair. In cells that are also SOS induced, the σ^S response, by unknown means, allows the use of, or persistence of errors made by, the error-prone DNA Pols in repair, causing base substitutions (45, 84) and indels (43, 44, 83, 166, 167). σ^S also downregulates mismatch repair (37–40, 88), which allows errors in DNA synthesis to persist. (Bottom) Less frequently, microhomologous MBR of a DSB occurs. It is SOS independent and requires (step 2) the σ^S response and DNA Pol I for template switching to regions containing microhomology (49, 50) (few complementary bases). The repair replication creates genome rearrangements. A duplicated chromosome segment is shown (blue arrows). Parallel lines represent base-paired DNA strands, and half arrowheads represent the 3′ DNA ends.

FIG 2

Pathway to and potential intervention points in formation of mutable gambler cells. cipro-induced DSBs activate the SOS response, which slows aerobic respiration (140). We suggest that increased autoxidation of reduced ubiquinone leads to (as observed in reference 57) a cell subpopulation with high levels of reactive oxygen species (ROS). ROS activate σ^S by upregulating transcription of sRNAs DsrA and ArcZ (57), which, with the Hfq RNA chaperone, increase translation of rpoS (σ^S) mRNA in the cells with high ROS (57). This σ^S-high “gambler” cell subpopulation allows mutagenic DNA break repair (MBR) (Fig. 1, top) and produces antibiotic cross-resistant mutants induced by cipro (57). The gambler subpopulation is transiently mutable (57). The SOS response also upregulates the SulA inhibitor of cell division, which promotes formation of multichromosome cells (168), which facilitate cipro-induced MBR (57). Potential antievolvability drug targets (depicted as “–| target,” in which “–|” indicates inhibition) have been identified by the discovery of the various steps of gambler cell formation and the action illustrated. The FDA-approved human drug edaravone inhibits gambler cell formation by quenching ROS (57), which does not select antievolvability drug resistance. Previous and proposed drugs to target the SOS response (155–157), DSB repair (158), any other DNA repair (154) and/or the error-prone DNA polymerases do reduce fitness in the antibiotic and so select resistance.