Catastrophic chromosome fragmentation probes the nucleoid structure and dynamics in Escherichia coli.

Escherichia coli cells treated with a combination of cyanide (CN) and hydrogen peroxide (HP) succumb to catastrophic chromosome fragmentation (CCF), detectable in pulsed-field gels as >100 double-strand breaks per genome equivalent. Here we show that CN + HP-induced double-strand breaks are independent of replication and occur uniformly over the chromosome,-therefore we used CCF to probe the nucleoid structure by measuring DNA release from precipitated nucleoids. CCF releases surprisingly little chromosomal DNA from the nucleoid suggesting that: (i) the nucleoid is a single DNA-protein complex with only limited stretches of protein-free DNA and (ii) CN + HP-induced breaks happen within these unsecured DNA stretches, rather than at DNA attachments to the central scaffold.

Potentiation of hydrogen peroxide toxicity: From catalase inhibition to stable DNA-iron complexes.

Hydrogen peroxide (HO) is unique among general toxins, because it is stable in abiotic environments at ambient temperature and neutral pH, yet rapidly kills any type of cells by producing highly-reactive hydroxyl radicals. This life-specific reactivity follows the distribution of soluble iron, Fe(II) (which combines with HO to form the famous Fenton's reagent),Fe(II) is concentrated inside cells, but is virtually absent outside them. Because of the immediate danger of HO, all cells have powerful HO scavengers, the equally famous catalases, which enable cells to survive thousand-fold higher concentrations of HO and, in combination with adequate movement of HO across membranes, make the killing HO concentrations virtually impractical to generate in vivo.

Prompt repair of hydrogen peroxide-induced DNA lesions prevents catastrophic chromosomal fragmentation.

Iron-dependent oxidative DNA damage in vivo by hydrogen peroxide (H2O2, HP) induces copious single-strand(ss)-breaks and base modifications. HP also causes infrequent double-strand DNA breaks, whose relationship to the cell killing is unclear. Since hydrogen peroxide only fragments chromosomes in growing cells, these double-strand breaks were thought to represent replication forks collapsed at direct or excision ss-breaks and to be fully reparable.

Static and Dynamic Factors Limit Chromosomal Replication Complexity in Escherichia coli, Avoiding Dangers of Runaway Overreplication.

We define chromosomal replication complexity (CRC) as the ratio of the copy number of the most replicated regions to that of unreplicated regions on the same chromosome. Although a typical CRC of eukaryotic or bacterial chromosomes is 2, rapidly growing Escherichia coli cells induce an extra round of replication in their chromosomes (CRC = 4). There are also E.

Cyanide enhances hydrogen peroxide toxicity by recruiting endogenous iron to trigger catastrophic chromosomal fragmentation.

Hydrogen peroxide (HP) or cyanide (CN) are bacteriostatic at low-millimolar concentrations for growing Escherichia coli, whereas CN + HP mixture is strongly bactericidal. We show that this synergistic toxicity is associated with catastrophic chromosomal fragmentation. Since CN alone does not kill at any concentration, while HP alone kills at 20 mM, CN must potentiate HP poisoning.