# Germination and Outgrowth of Bacillus subtilis Spores Deficient in BER and DisA Unveil Alternative Genetic Checkpoints

## TL;DR

This study explores how Bacillus subtilis spores manage DNA damage during germination and outgrowth, revealing alternative genetic checkpoints beyond DisA.

## Contribution

The study identifies DisA-independent DNA damage checkpoints in Bacillus subtilis spores during germination and outgrowth.

## Key findings

- Spores lacking Nfo, ExoA, and Nth showed delayed outgrowth even when DisA was disrupted.
- DisA-deficient spores exhibited higher oxidative DNA lesions and chromosome segregation issues.
- DisA promotes low-fidelity DNA repair and replication events, as shown by DNA gel electrophoresis and mutagenesis.

## Abstract

During Bacillus subtilis spore germination/outgrowth, the rehydration of the spore core and activation of aerobic metabolism can generate reactive oxygen species (ROS)-promoted DNA lesions that are repaired via the base excision repair pathway (BER). Accordingly, spores deficient in the AP-endonucleases (APEs) Nfo and ExoA exhibit a delayed outgrowth that is suppressed following disruption of the checkpoint protein DisA. Here, we report that DisA-independent DNA damage checkpoints operate during B. subtilis spore outgrowth. Consistent with this notion, spores lacking Nfo, ExoA, and Nth, which functions as an APE, did not suppress delayed outgrowth following disA disruption. Furthermore, in reference to the ∆nfo ∆exoA ∆nth spores, spores deficient for these APEs and DisA displayed a significantly higher number of oxidative genetic lesions and failed to properly segregate its chromosome during the first round of replication in the outgrowth stage. Finally, we found that DisA promotes low-fidelity repair and replication events, as revealed by DNA-alkaline gel electrophoresis (AGE) as well as spontaneous and H2O2-promoted RifR mutagenesis. Overall, our results unveil the existence of DisA-independent DNA damage checkpoint(s) that are activated by genomic lesions of an oxidative nature during spore germination and outgrowth, ensuring a proper transition to vegetative growth.