DNA damage checkpoint activation is blocked by mutations in YneA and the septal peptidoglycan polymerase FtsW

During normal DNA replication, all cells encounter damage to their genetic material. As a result, organisms from bacteria to humans have developed response pathways that provide time for the cell to complete DNA repair before cell division resumes. In Bacillus subtilis, it is well established that the SOS-induced cell division inhibitor YneA blocks cell division after exposure to genotoxic stress; however, it remains unclear how YneA enforces the checkpoint. In this study, we used genetic approaches to identify mutations that disrupt YneA activity and mutations that are refractory to the YneA-induced checkpoint. We isolated YneA C-terminal truncation mutants and point mutants in or near the LysM peptidoglycan binding domain that rendered YneA incapable of checkpoint enforcement. These results show that the LysM domain is required to elicit checkpoint activation. In addition, we developed a genetic method which isolated mutations in the ftsW gene that completely bypassed checkpoint enforcement. Characterization of an ftsW variant resulted in considerably shorter cells during the DNA damage response indicative of a hyperactive form of the peptidoglycan polymerase FtsW. With our results, we present a model where YneA binds peptidoglycan, facilitated by its LysM domain, inhibiting septal cell wall synthesis by FtsW preventing cell division.