Dysregulated DnaB unwinding induces replisome decoupling and daughter strand gaps that are countered by RecA polymerization

Repair of DNA damage begins with the elicitation of targeted cellular responses to restore the genome. In E. coli, major products of DNA damage result in the buildup of single-stranded DNA (ssDNA) that is rapidly bound by cooperative filamentation of RecA to initiate the SOS response. The replicative helicase, DnaB, is a central component of the replisome, unwinding duplex DNA in concert with Pol III template dependent synthesis. Interestingly, helicase unwinding is heavily regulated, and the unwinding rate can be reduced by over 10-fold if DnaB becomes decoupled from Pol III. However, if DnaB is dysregulated by mutations that enforce a faster more constricted conformation, unwinding can continue independently, generating excess ssDNA resulting in severe cellular stress. This surplus ssDNA can stimulate RecA recruitment for recombinational repair or activation of SOS to increase the available repair protein pool. To better understand the consequences of dysregulated unwinding, we combined targeted dnaB mutations with an inducible plasmid-based RecA filament inhibition strategy to examine the dependencies on RecA in counteracting decoupling. We find that RecA filamentation is instrumental for processing daughter strand gaps left behind from decoupled unwinding and synthesis to prevent DNA breaks. Without functional RecA filaments, dnaB mutant strains had a greater burden from endogenous damage but without a compensatory increase in mutagenesis. Overall, RecA plays a critical role in strain survival by processing DNA gaps and protecting from breaks caused by dysregulated or interrupted helicase activity in vivo. ### Competing Interest Statement The authors have declared no competing interest.