Bacterial Chromosome Replication and DNA Repair During the Stringent Response

ABSTRACT Balancing of reducing equivalents is a fundamental issue in bacterial metabolism and metabolic engineering. Mutations in the key metabolic genes ldhA and pflB of Escherichia coli are known to stall anaerobic growth and fermentation due to a buildup of intracellular NADH. We observed that the rate of spontaneous mutation in E. coli BW25113 (ΔldhA ΔpflB) was an order of magnitude higher than that in wild-type (WT) E. coli BW25113. We hypothesized that the increased mutation frequency was due to an increased NADH/NAD+ ratio in this strain. Using several redox-impaired strains of E. coli and different redox conditions, we confirmed a significant correlation (P < 0.01) between intracellular-NADH/NAD+ ratio and mutation frequency. To identify the genetic basis for this relationship, whole-genome transcriptional profiles were compared between BW25113 WT and BW25113 (ΔldhA ΔpflB). This analysis revealed that the genes involved in DNA repair were expressed at significantly lower levels in BW25113 (ΔldhA ΔpflB). Direct measurements of the extent of DNA repair in BW25113 (ΔldhA ΔpflB) subjected to UV exposure confirmed that DNA repair was inhibited. To identify a direct link between DNA repair and intracellular-redox ratio, the stringent-response-regulatory gene relA and the global-stress-response-regulatory gene rpoS were deleted. In both cases, the mutation frequencies were restored to BW25113 WT levels.

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Blocking the Trigger: Inhibition of the Initiation of Bacterial Chromosome Replication as an Antimicrobial Strategy

Antibiotics ◽

10.3390/antibiotics8030111 ◽

2019 ◽

Vol 8 (3) ◽

pp. 111

Author(s):

Julia E. Grimwade ◽

Alan C. Leonard

Keyword(s):

Drug Targets ◽

Antimicrobial Agents ◽

Current Knowledge ◽

Chromosome Replication ◽

Bacterial Chromosome ◽

Resistant Bacteria ◽

Bacterial Cells ◽

Drug Resistant Bacteria ◽

New Antibiotics ◽

Novel Drug

All bacterial cells must duplicate their genomes prior to dividing into two identical daughter cells. Chromosome replication is triggered when a nucleoprotein complex, termed the orisome, assembles, unwinds the duplex DNA, and recruits the proteins required to establish new replication forks. Obviously, the initiation of chromosome replication is essential to bacterial reproduction, but this process is not inhibited by any of the currently-used antimicrobial agents. Given the urgent need for new antibiotics to combat drug-resistant bacteria, it is logical to evaluate whether or not unexploited bacterial processes, such as orisome assembly, should be more closely examined for sources of novel drug targets. This review will summarize current knowledge about the proteins required for bacterial chromosome initiation, as well as how orisomes assemble and are regulated. Based upon this information, we discuss current efforts and potential strategies and challenges for inhibiting this initiation pharmacologically.

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Faculty Opinions recommendation of Frequent exchange of the DNA polymerase during bacterial chromosome replication.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.727464346.793532119 ◽

2017 ◽

Author(s):

Luca Pellegrini

Keyword(s):

Dna Polymerase ◽

Chromosome Replication ◽

Bacterial Chromosome ◽

Frequent Exchange

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oriC-encoded instructions for the initiation of bacterial chromosome replication

Frontiers in Microbiology ◽

10.3389/fmicb.2014.00735 ◽

2015 ◽

Vol 5 ◽

Cited By ~ 32

Author(s):

Marcin WolaÅ„ski ◽

RafaÅ‚ Donczew ◽

Anna Zawilak-Pawlik ◽

Jolanta Zakrzewska-CzerwiÅ„ska

Keyword(s):

Chromosome Replication ◽

Bacterial Chromosome

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Surviving chromosome replication: the many roles of the S-phase checkpoint pathway

Philosophical Transactions of the Royal Society B Biological Sciences ◽

10.1098/rstb.2011.0071 ◽

2011 ◽

Vol 366 (1584) ◽

pp. 3554-3561 ◽

Cited By ~ 65

Author(s):

Karim Labib ◽

Giacomo De Piccoli

Keyword(s):

Dna Damage ◽

Dna Repair ◽

Cell Cycle Progression ◽

Reductase Activity ◽

Critical Role ◽

Chromosome Replication ◽

S Phase ◽

Replication Forks ◽

Checkpoint Pathway ◽

S Phase Checkpoint

Checkpoints were originally identified as signalling pathways that delay mitosis in response to DNA damage or defects in chromosome replication, allowing time for DNA repair to occur. The ATR (ataxia- and rad-related) and ATM (ataxia-mutated) protein kinases are recruited to defective replication forks or to sites of DNA damage, and are thought to initiate the DNA damage response in all eukaryotes. In addition to delaying cell cycle progression, however, the S-phase checkpoint pathway also controls chromosome replication and DNA repair pathways in a highly complex fashion, in order to preserve genome integrity. Much of our understanding of this regulation has come from studies of yeasts, in which the best-characterized targets are the stimulation of ribonucleotide reductase activity by multiple mechanisms, and the inhibition of new initiation events at later origins of DNA replication. In addition, however, the S-phase checkpoint also plays a more enigmatic and apparently critical role in preserving the functional integrity of defective replication forks, by mechanisms that are still understood poorly. This review considers some of the key experiments that have led to our current understanding of this highly complex pathway.

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