# Bacillus subtilis DinG 3′⟶5′ Exo(ribo)nuclease: A Helpmate to Mitigate Replication Stress

## TL;DR

This paper explores how the enzyme DinG in Bacillus subtilis helps reduce replication stress by degrading RNA-DNA hybrids and working with other proteins during DNA repair.

## Contribution

The study reveals DinG's role in degrading R-loops and its physical interaction with RecA and TLS polymerases to mitigate replication stress.

## Key findings

- DinG degrades RNA strands in RNA–DNA hybrids and removes genomic R-loops.
- DinG interacts with RecA and TLS DNA polymerases to respond to DNA damage.
- DinG forms foci at replication forks under stress conditions like MMS treatment.

## Abstract

Bacillus subtilis DinG/XPD-like paralogues, DinG and YpvA, have been implicated in overcoming replication stress. DinG possesses a DEDD exonuclease and DNA helicase domains, whereas YpvA lacks the DEDD exonuclease domain. We report that DinG·Mg2+ (hereafter referred to as DinG) degrades linear single-stranded (lss) DNA with 3′→5′ polarity and binds lssDNA with higher affinity than its exonuclease-deficient mutant DinG D10A E12A. DinG’s ssDNA-dependent ATPase activity neither stimulates nor inhibits DNA degradation. When bound to the 3′-end of forked DNA, DinG destabilises and degrades the substrate; however, in the presence of ATP, DinG dissociates before reaching the duplex junction. DinG degrades the RNA strand within RNA–DNA hybrids but does not cleave lssRNA unless complexed with Mn2+. DinG removes genomic R-loops, as RnhC and PcrA do. DinG physically interacts with RecA and PolA and functions in the same pathway as translesion synthesis (TLS) DNA polymerases (DNAPs) to respond to both spontaneous and methyl methanesulphonate (MMS)-induced mutagenesis. DinG-mGold forms spontaneous foci at or near replication forks, which become enriched following MMS or rifampicin treatment. We propose that DinG contributes to mitigating replication stress by degrading R-loop barriers and facilitating TLS, potentially via RecA-linked mechanisms.