Abstract
Duplication of bacterial chromosomes is initiated via the assembly of two replication forks at a single defined origin. Forks proceed bi-directionally until they fuse in a specialised termination area opposite the origin. This area is flanked by polar replication fork pause sites that allow forks to enter but not to leave. The precise function of this replication fork trap has remained enigmatic, as no obvious phenotypes have been associated with its inactivation. However, the fork trap becomes a serious problem to cells if the second fork is stalled at an impediment, as replication cannot be completed, suggesting that a significant evolutionary advantage for maintaining this chromosomal arrangement must exist. Recently, we demonstrated that head-on fusion of replication forks can trigger over-replication of the chromosome. This over-replication is normally prevented by a number of proteins including RecG helicase and 3’ exonucleases. However, even in the absence of these proteins it can be safely contained within the replication fork trap, highlighting that multiple systems might be involved in coordinating replication fork fusions. Here, we discuss whether considering the problems associated with head-on replication fork fusion events helps us to better understand the important role of the replication fork trap in cellular metabolism.
Highlights
Chromosome replication in all cells studied is regulated by recruitment of the replication machinery to specific initiation sites where two forks are established and move in opposite directions until they meet either an opposing fork or the end of a chromosome
Genes transcribedGenes at verytranscribed high levels are relatively distant from the fork trap area, forks escaping the termination area progress with little the fork trap area, forks escaping the termination area progress with little indication of problems, and, indication problems, one and,replisome in addition, one replisome from entering opposite in addition,ofpreventing frompreventing entering the opposite replichore does not the explain why replichore does not explain why in coli the replication fork trap covers almost of the in E. coli the replication fork trap covers almost 50% of the chromosome
An increasing number of experimental results suggest that a number of proteins such as RecG and 3’ exonucleases have an important function in preventing chromosome over-replication triggered by head-on fork fusion events, and it is significant that this over-replication in cells lacking RecG is blocked by mutations that suppress many features of the recG mutant phenotype [100]
Summary
Chromosome replication in all cells studied is regulated by recruitment of the replication machinery to specific initiation sites (origins) where two forks are established and move in opposite directions until they meet either an opposing fork or the end of a chromosome. We showed that the doubling time of ∆oriC oriZ cells in LB broth was, at 40 min or longer, twice as long as in wild type cells, and suppressor mutations rapidly accumulated, demonstrating that the initiation of synthesis from an ectopic location causes severe problems for cells [10] It was shown before both in E. coli and B. subtilis that the most severe problems for ongoing replication arise at the highly transcribed rrn operons, which slow or block progressing replication forks, and it was shown that RecBCD processing of dsDNA ends is required to allow replication to restart [45,46,47,48]. Were re‐plotted from [18]
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