Abstract
ABSTRACTIt is well established that DNA double-strand break (DSB) repair is required to underpin chromosomal DNA replication. Because DNA replication forks are prone to breakage, faithful DSB repair and correct replication fork restart are critically important. Cells, where the proteins required for DSB repair are absent or altered, display characteristic disturbances to genome replication. In this review, we analyze how bacterial DNA replication is perturbed in DSB repair mutant strains and explore the consequences of these perturbations for bacterial chromosome segregation and cell viability. Importantly, we look at how DNA replication and DSB repair processes are implicated in the striking recent observations of DNA amplification and DNA loss in the chromosome terminus of various mutant Escherichia coli strains. We also address the mutant conditions required for the remarkable ability to copy the entire E. coli genome, and to maintain cell viability, even in the absence of replication initiation from oriC, the unique origin of DNA replication in wild type cells. Furthermore, we discuss the models that have been proposed to explain these phenomena and assess how these models fit with the observed data, provide new insights and enhance our understanding of chromosomal replication and termination in bacteria.
Highlights
In bacteria with circular chromosomes, DNA replication initiates at a fixed position called the origin and proceeds bidirectionally to reach the opposite side of the chromosome, where replication forks meet and replication terminates
Classical and recent studies point to D-loops and R-loops being implicated in STABLE DNA REPLICATION (SDR) and suggest that SDR is intimately associated with terminus DNA amplification
The formation of a D-loop is not sufficient to set up SDR since most D-loops are expected to be resolved to restore normal DNA replication or an intact chromosome (Fig. 1)
Summary
In bacteria with circular chromosomes, DNA replication initiates at a fixed position called the origin (oriC in E. coli) and proceeds bidirectionally to reach the opposite side of the chromosome, where replication forks meet and replication terminates. Similar losses of viability for these mutants were reported in an earlier study (Capaldo, Ramsey and Barbour 1974) These observations led to the proposal that a subpopulation of growing E. coli cells experience the formation of replication dependent DSBs that can be reset by RecBCD-RecA mediated recombination or, in the absence of RecA, the broken chromosomal ends can be degraded by RecBCD exonuclease activity to generate intact chromosomes. An attempt to map specific loci that can exhibit oriM1 and oriM2 activity in a heterologous system was not successful, suggesting that these increases in copy number are the direct outcome of random chromosome DSB generation and repair by RecBCD-RecA mediated homologous recombination and not due to any sequence-specific origin induction This was supported by the observation that a recD mutation (which induces hyper-recombination) enhances iSDR after thymine starvation and nalidixic acid treatment (Magee & Kogoma 1990; Magee et al 1992; Asai et al 1993). A high amount of R-loops and of cSDR were observed in topA topB null cells and the growth phenotype could be suppressed either by RNase HI overproduction, recA mutation (RecA is required to repair the resulting double-strand breaks) or dnaT mutation (required for replication restart) (Masse and Drolet 1999; Martel et al 2015; Kouzminova, Kadyrov and Kuzminov 2017; Brochu et al 2018; Drolet and Brochu 2019)
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