RecBCD, SbcCD and ExoI process a substrate created by convergent replisomes to complete DNA replication

Abstract

The accurate completion of DNA replication on the chromosome requires RecBCD and structure specific SbcCD and ExoI nucleases. However, the substrates and mechanism by which this reaction occurs remains unknown. Here we show that these completion enzymes operate on plasmid substrates containing two replisomes, but are not required for plasmids containing one replisome. Completion on the two-replisome plasmids requires RecBCD, but does not require RecA and no broken intermediates accumulate in its absence, indicating that the completion reaction occurs normally in the absence of any double-strand breaks. Further, similar to the chromosome, we show that when the normal completion reaction is prevented, an aberrant RecA-mediated recombination process leads to amplifications that drive most of the instabilities associated with the two-replisome substrates. The observations imply that the substrate SbcCD, ExoI and RecBCD act upon in vivo is created specifically by two convergent replisomes, and demonstrate that the function of RecBCD in completing replication is independent of double-strand break repair, and likely promotes joining of the strands of the convergent replication forks.