MutS2 Promotes Homologous Recombination in Bacillus subtilis

Bacterial MutS proteins are subdivided into two families, MutS1 and MutS2. MutS1 family members recognize DNA replication errors during their participation in the well-characterized mismatch repair (MMR) pathway. In contrast to the well-described function of MutS1, the function of MutS2 in bacteria has remained less clear. InHelicobacter pylori andThermus thermophilus , MutS2 has been shown to suppress homologous recombination. The role of MutS2 is unknown in the Gram-positive bacteriumBacillus subtilis . In this work, we investigated the contribution of MutS2 to maintaining genome integrity inB. subtilis . We found that deletion ofmutS2 rendersB. subtilis sensitive to the natural antibiotic mitomycin C (MMC), which requires homologous recombination for repair. We demonstrate that the C-terminals mallM utS-r elated (Smr) domain is necessary but not sufficient for tolerance to MMC. Further, we developed a CRISPR/Cas9 genome editing system to test if the inducible prophage PBSX was the underlying cause of the observed MMC sensitivity. Genetic analysis revealed that MMC sensitivity was dependent on recombination and not on nucleotide excision repair or a symptom of prophage PBSX replication and cell lysis. We found that deletion ofmutS2 resulted in decreased transformation efficiency using both plasmid and chromosomal DNA. Further, deletion ofmutS2 in a strain lacking the Holliday junction endonuclease generecU resulted in increased MMC sensitivity and decreased transformation efficiency, suggesting that MutS2 could function redundantly with RecU. Together, our results support a model whereB. subtilis MutS2 helps to promote homologous recombination, demonstrating a new function for bacterial MutS2.IMPORTANCE Cells contain pathways that promote or inhibit recombination. MutS2 homologs are Smr-endonuclease domain-containing proteins that have been shown to function in antirecombination in some bacteria. We present evidence thatB. subtilis MutS2 promotes recombination, providing a new function for MutS2. We found that cells lackingmutS2 are sensitive to DNA damage that requires homologous recombination for repair and have reduced transformation efficiency. Further analysis indicates that the C-terminal Smr domain requires the N-terminal portion of MutS2 for functionin vivo . Moreover, we show that amutS2 deletion is additive with arecU deletion, suggesting that these proteins have a redundant function in homologous recombination. Together, our study shows that MutS2 proteins have adapted different functions that impact recombination.