# Role of RelA-synthesized (p)ppGpp and ROS-induced mutagenesis in de novo acquisition of antibiotic resistance in E. coli

## TL;DR

This study shows how the bacterial response to stress, involving (p)ppGpp and ROS, helps E. coli develop antibiotic resistance, and how blocking these processes could reduce resistance.

## Contribution

The study reveals a novel mechanism linking (p)ppGpp and ROS to the de novo acquisition of antibiotic resistance in E. coli.

## Key findings

- Deletion of relA or rpoS slows the development of resistance to bactericidal antibiotics.
- Resistant strains show mutations in genes related to antibiotic resistance and stress responses.
- Knockout of relA reduces ROS and DNA damage, weakening DNA repair in resistant strains.

## Abstract

The stringent response of bacteria to starvation and stress also fulfills a role in addressing the threat of antibiotics. Within this stringent response, (p)ppGpp, synthesized by RelA or SpoT, functions as a global alarmone. However, the effect of this (p)ppGpp on resistance development is poorly understood. Here, we show that knockout of relA or rpoS curtails resistance development against bactericidal antibiotics. The emergence of mutated genes associated with starvation and (p)ppGpp, among others, indicates the activation of stringent responses. The growth rate is decreased in ΔrelA-resistant strains due to the reduced ability to synthesize (p)ppGpp and the persistence of deacylated tRNA impeding protein synthesis. Sluggish cellular activity causes decreased production of reactive oxygen species (ROS), thereby reducing oxidative damage, leading to weakened DNA mismatch repair, potentially reducing the generation of mutations. These findings offer new targets for mitigating antibiotic resistance development, potentially achieved through inhibiting (p)ppGpp or ROS synthesis.

•Deletion of relA or rpoS results in a decelerated acquisition of resistance to bactericidal antibiotics•Resistant strains exhibit mutations in genes linked to antibiotic resistance, oxidative stress, the SOS response, and the stringent response•Knockout of relA leads to reduced ROS production and mitigated DNA oxidative damage•Transcription levels of genes involved in DNA repair are diminished in ΔrelA-resistant strains exposed to bactericidal antibiotics

Deletion of relA or rpoS results in a decelerated acquisition of resistance to bactericidal antibiotics

Resistant strains exhibit mutations in genes linked to antibiotic resistance, oxidative stress, the SOS response, and the stringent response

Knockout of relA leads to reduced ROS production and mitigated DNA oxidative damage

Transcription levels of genes involved in DNA repair are diminished in ΔrelA-resistant strains exposed to bactericidal antibiotics

Molecular biology; Microbiology