Bibliography.bib

@article{Lepore2019a,
   abstract = {Cell biology is increasingly dependent on quantitative methods resulting in the need for microscopic labelling technologies that are highly sensitive and specific. Whilst the use of fluorescent proteins has led to major advances, they also suffer from their relatively low brightness and photo-stability, making the detection of very low abundance proteins using fluorescent protein-based methods challenging. Here, we characterize the use of the self-labelling protein tag called HaloTag, in conjunction with an organic fluorescent dye, to label and accurately count endogenous proteins present in very low numbers (<7) in individual {{\textit{Escherichia coli}}} cells. This procedure can be used to detect single molecules in fixed cells with conventional epifluorescence illumination and a standard microscope. We show that the detection efficiency of proteins labelled with the HaloTag is ≥80%, which is on par or better than previous techniques. Therefore, this method offers a simple and attractive alternative to current procedures to detect low abundance molecules.},
   author = {Alessia Lepore and Hannah Taylor and Dirk Landgraf and Burak Okumus and Sebastián Jaramillo-Riveri and Lorna McLaren and Somenath Bakshi and Johan Paulsson and M. El Karoui},
   doi = {10.1038/s41598-019-44278-0},
   issn = {20452322},
   issue = {1},
   journal = {Scientific Reports},
   keywords = {Cellular microbiology,Single,cell imaging},
   month = {12},
   pages = {1-9},
   pmid = {31133640},
   publisher = {Nature Publishing Group},
   title = {Quantification of very low-abundant proteins in bacteria using the HaloTag and epi-fluorescence microscopy},
   volume = {9},
   url = {https://doi.org/10.1038/s41598-019-44278-0},
   year = {2019}
}
@article{Yu2006,
   abstract = {We directly observed real-time production of single protein molecules in individual {{\textit{Escherichia coli}}} cells. A fusion protein of a fast-maturing yellow fluorescent protein (YFP) and a membrane-targeting peptide was expressed under a repressed condition. The membrane-localized YFP can be detected with single-molecule sensitivity. We found that the protein molecules are produced in bursts, with each burst originating from a stochastically transcribed single messenger RNA molecule, and that protein copy numbers in the bursts follow a geometric distribution. The quantitative study of low-level gene expression demonstrates the potential of single-molecule experiments in elucidating the workings of fundamental biological processes in living cells.},
   author = {Ji Yu and Jie Xiao and Xiaojia Ren and Kaiqin Lao and X. Sunney Xie},
   issn = {00368075},
   issue = {5767},
   journal = {Science},
   month = {3},
   pages = {1600-1603},
   pmid = {16543458},
   publisher = {American Association for the Advancement of Science},
   title = {Probing gene expression in live cells, one protein molecule at a time},
   volume = {311},
   url = {https://www.science.org/doi/10.1126/science.1119623},
   year = {2006}
}
@article{Elf2007,
   abstract = {Transcription factors regulate gene expression through their binding to {DNA}. In a living {{\textit{Escherichia coli}}} cell, we directly observed specific binding of a lac repressor, labeled with a fluorescent protein, to a chromosomal lac operator. Using single-molecule detection techniques, we measured the kinetics of binding and dissociation of the repressor in response to metabolic signals. Furthermore, we characterized the nonspecific binding to {DNA}, one-dimensional (1D) diffusion along {DNA} segments, and 3D translocation among segments through cytoplasm at the single-molecule level. In searching for the operator, a lac repressor spends ∼90% of time nonspecifically bound to and diffusing along {DNA} with a residence time of <S milliseconds. The methods and findings can be generalized to other nucleic acid binding proteins.},
   author = {Johan Elf and Gene Wei Li and X. Sunney Xie},
   issn = {00368075},
   issue = {5828},
   journal = {Science},
   month = {5},
   pages = {1191-1194},
   pmid = {17525339},
   publisher = {American Association for the Advancement of Science},
   title = {Probing transcription factor dynamics at the single-molecule level in a living cell},
   volume = {316},
   url = {https://www.science.org/doi/10.1126/science.1141967},
   year = {2007}
}
@article{Wiktor2021,
   abstract = {Homologous recombination is essential for the accurate repair of double-stranded {DNA} breaks (DSBs)1. Initially, the {{RecB}CD} complex2 resects the ends of the DSB into 3′ single-stranded {DNA} on which a {RecA} filament assembles3. Next, the filament locates the homologous repair template on the sister chromosome4. Here we directly visualize the repair of DSBs in single cells, using high-throughput microfluidics and fluorescence microscopy. We find that, in {{\textit{Escherichia coli}}}, repair of DSBs between segregated sister loci is completed in 15 ± 5 min (mean ± s.d.)&nbsp;with minimal fitness loss. We further show that the search takes less than 9 ± 3 min (mean&nbsp;±&nbsp;s.d)&nbsp;and is mediated by a thin, highly dynamic {RecA} filament that stretches throughout the cell. We propose that the architecture of the {RecA} filament effectively reduces search dimensionality. This model predicts a search time that is consistent with our measurement and is corroborated by the observation that the search time does not depend on the length of the cell or the amount of {DNA}. Given the abundance of {RecA} homologues5, we believe this model to be&nbsp;widely conserved across living organisms. Observations of rapid repair of double-stranded {DNA} breaks in sister choromosomes in {{\textit{Escherichia coli}}} are consistent with a reduced-dimensionality-search model of {RecA}-mediated repair.},
   author = {Jakub Wiktor and Arvid H. Gynnå and Prune Leroy and Jimmy Larsson and Giovanna Coceano and Ilaria Testa and Johan Elf},
   doi = {10.1038/s41586-021-03877-6},
   issn = {1476-4687},
   issue = {7876},
   journal = {Nature 2021 597:7876},
   keywords = {{DNA} recombination,Homologous recombination,Super,resolution microscopy},
   month = {9},
   pages = {426-429},
   pmid = {34471288},
   publisher = {Nature Publishing Group},
   title = {{RecA} finds homologous {DNA} by reduced dimensionality search},
   volume = {597},
   url = {https://www.nature.com/articles/s41586-021-03877-6},
   year = {2021}
}
@article{Sinha2018,
   abstract = {It was recently reported that the recBC mutants of {{\textit{Escherichia coli}}}, deficient for {DNA} double-strand break (DSB) repair, have a decreased copy number of their terminus region. We previously showed that this deficit resulted from {DNA} loss after post-replicative breakage of one of the two sister-chromosome termini at cell division. A viable cell and a dead cell devoid of terminus region were thus produced and, intriguingly, the reaction was transmitted to the following generations. Using genome marker frequency profiling and observation by microscopy of specific {DNA} loci within the terminus, we reveal here the origin of this phenomenon. We observed that terminus {DNA} loss was reduced in a recA mutant by the double-strand {DNA} degradation activity of {{RecB}CD}. The terminus-less cell produced at the first cell division was less prone to divide than the one produced at the next generation. {DNA} loss was not heritable if the chromosome was linearized in the terminus and occurred at chromosome termini that were unable to segregate after replication. We propose that in a recB mutant replication fork breakage results in the persistence of a linear {DNA} tail attached to a circular chromosome. Segregation of the linear and circular parts of this “σ-replicating chromosome” causes terminus {DNA} breakage during cell division. One daughter cell inherits a truncated linear chromosome and is not viable. The other inherits a circular chromosome attached to a linear tail ending in the chromosome terminus. Replication extends this tail, while degradation of its extremity results in terminus {DNA} loss. Repeated generation and segregation of new σ-replicating chromosomes explains the heritability of post-replicative breakage. Our results allow us to determine that in {{\textit{E. coli}}} at each generation, 18% of cells are subject to replication fork breakage at dispersed, potentially random, chromosomal locations.},
   author = {Anurag Kumar Sinha and Christophe Possoz and Adeline Durand and Jean Michel Desfontaines and François Xavier Barre and David R.F. Leach and Bénédicte Michel},
   doi = {10.1371/JOURNAL.PGEN.1007256},
   isbn = {1111111111},
   issn = {1553-7404},
   issue = {3},
   journal = {PLOS Genetics},
   keywords = {Cell cycle and cell division,Chromosome structure and function,Circular {DNA},{DNA} recombination,{DNA} repair,{DNA} replication,Genetic loci,Homologous recombination},
   month = {3},
   pages = {e1007256},
   pmid = {29522563},
   publisher = {Public Library of Science},
   title = {Broken replication forks trigger heritable {DNA} breaks in the terminus of a circular chromosome},
   volume = {14},
   url = {https://journals.plos.org/plosgenetics/article?id=10.1371/journal.pgen.1007256},
   year = {2018}
}
@article{Wilkinson2016,
   abstract = {Our previous paper (Wilkinson et al, 2016) used high-resolution cryo-electron microscopy to solve the structure of the {{\textit{Escherichia coli}}} {{RecB}CD} complex, which acts in both the repair of double-stranded {DNA} breaks and the degradation of bacteriophage {DNA}. To counteract the latter activity, bacteriophage λ encodes a small protein inhibitor called Gam that binds to {{RecB}CD} and inactivates the complex. Here, we show that Gam inhibits {{RecB}CD} by competing at the {DNA}-binding site. The interaction surface is extensive and involves molecular mimicry of the {DNA} substrate. We also show that expression of Gam in {{\textit{E. coli}}} or Klebsiella pneumoniae increases sensitivity to fluoroquinolones; antibacterials that kill cells by inhibiting topoisomerases and inducing double-stranded {DNA} breaks. Furthermore, fluoroquinolone-resistance in K. pneumoniae clinical isolates is reversed by expression of Gam. Together, our data explain the synthetic lethality observed between topoisomerase-induced {DNA} breaks and the {{RecB}CD} gene products, suggesting a new co-antibacterial strategy.},
   author = {Martin Wilkinson and Lucy Troman and Wan AK Wan Nur Ismah and Yuriy Chaban and Matthew B Avison and Mark S Dillingham and Dale B Wigley},
   doi = {10.7554/ELIFE.22963},
   journal = {eLife},
   month = {12},
   publisher = {eLife Sciences Publications, Ltd},
   title = {Structural basis for the inhibition of {{RecB}CD} by Gam and its synergistic antibacterial effect with quinolones},
   volume = {5},
   year = {2016}
}
@article{Odsbu2014,
   abstract = {The nucleoids of undamaged {{\textit{Escherichia coli}}} cells have a characteristic shape and number, which is dependent on the growth medium. Upon induction of the {SOS} response by a low dose of {UV} irradiation an extensive reorganization of the nucleoids occurred. Two distinct phases were observed by fluorescence microscopy. First, the nucleoids were found to change shape and fuse into compact structures at midcell. The compaction of the nucleoids lasted for 10-20 min and was followed by a phase where the {DNA} was dispersed throughout the cells. This second phase lasted for ~1 h. The compaction was found to be dependent on the recombination proteins {RecA}, {RecO} and {RecR} as well as the {SOS}-inducible, SMC (structural maintenance of chromosomes)- like protein {RecN}. {RecN} protein is produced in high amounts during the first part of the {SOS} response. It is possible that the {RecN}-mediated 'compact {DNA}' stage at the beginning of the {SOS} response serves to stabilize damaged {DNA} prior to recombination and repair. © 2014 The Authors.},
   author = {Ingvild Odsbu and Kirsten Skarstad},
   issn = {14652080},
   issue = {PART 5},
   journal = {Microbiology (United Kingdom)},
   month = {5},
   pages = {872-882},
   pmid = {24615185},
   publisher = {Microbiology Society},
   title = {{DNA} compaction in the early part of the {SOS} response is dependent on {RecN} and {RecA}},
   volume = {160},
   url = {https://www.microbiologyresearch.org/content/journal/micro/10.1099/mic.0.075051-0},
   year = {2014}
}
@article{Yu1998,
   abstract = {The recombinational hot spot X modulates the nuclease and helicase activities of the {{RecB}CD} enzyme, leading to generation of an early {DNA} intermediate for homologous recombination. Here we identify the subunit location of the nuclease active site in {{RecB}CD}. The isolated {RecB} protein cleaves circular single-stranded M13 phage {DNA}, but {RecB}1-929, comprising only the 100 {kDa} N-terminal domain of {RecB}, does not. We reported previously that the reconstituted {RecB}1-929CD enzyme also is not a nuclease, suggesting that the C-terminal 30 {kDa} domain of {RecB} is a non-specific ss{DNA} endonuclease. However, we were unable to detect nuclease activity with the subtilisin-generated C-terminal 30 {kDa} fragment of {RecB}. Since the subtilisin-generated fragment did not bind to a ss{DNA}-agarose column, we designed a chimeric enzyme by attaching the C-terminal 30 {kDa} domain of {RecB} to the gene 32 protein of T4 phage, a ss{DNA} binding protein that does not have strand scission ability. In addition, Asp427 in the chimeric enzyme (Asp1080 in {RecB}), a residue that is conserved among several {RecB} homologs, was substituted to alanine (the D427A mutant). The wild-type chimeric enzyme cleaves the M13 {DNA} and the D427A mutation abolishes the endonuclease activity of the chimeric enzyme but does not affect its {DNA} binding ability. This finding indicates an unusual bipartite nature in the structural organization of {RecB}, in which the {DNA}-binding function is located in the N-terminal 100 {kDa} domain and the nuclease catalytic domain is located in the C-terminal 30 {kDa} domain. The purified {RecB}({D1080A})CD mutant is a processive helicase but not a nuclease, demonstrating that {{RecB}CD} has a single nuclease active site in the C terminal 30 {kDa} domain of {RecB}.},
   author = {Misook Yu and Jehanne Souaya and Douglas A. Julin},
   doi = {10.1006/JMBI.1998.2127},
   issn = {0022-2836},
   issue = {4},
   journal = {Journal of Molecular Biology},
   keywords = {Chi sequence,Endonuclease,Gene 32 protein,{{RecB}CD},Recombination},
   month = {11},
   pages = {797-808},
   pmid = {9790841},
   publisher = {Academic Press},
   title = {Identification of the nuclease active site in the multifunctional {{RecB}CD} enzyme by creation of a chimeric enzyme},
   volume = {283},
   year = {1998}
}
@article{Ivancic-Bace_2003,
   abstract = {The {RecA} loading activity of the {{RecB}CD} enzyme, together with its helicase and 5′ → 3′ exonuclease activities, is essential for recombination in {{\textit{Escherichia coli}}}. One particular mutant in the nuclease catalytic center of {RecB}, i.e., recB1080, produces an enzyme that does not have nuclease activity and is unable to load {RecA} protein onto single-stranded {DNA}. There are, however, previously published contradictory data on the recombination proficiency of this mutant. In a recF- background the recB1080 mutant is recombination deficient, whereas in a recF+ genetic background it is recombination proficient. A possible explanation for these contrasting phenotypes may be that the {{RecF}OR} system promotes {RecA}-single-strand {DNA} filament formation and replaces the {RecA} loading defect of the {RecB}1080CD enzyme. We tested this hypothesis by using three in vivo assays. We compared the recombination proficiencies of recB1080, rec0, recR, and {RecF} single mutants and recB1080 rec0, recB1080 recR, and recB1080 recF double mutants. We show that {{RecF}OR} functions rescue the repair and recombination deficiency of the recB1080 mutant and that {RecA} loading is independent of {{RecF}OR} in the recB1080 recD double mutant where this activity is provided by the {RecB}1080C(D-) enzyme. According to our results as well as previous data, three essential activities for the initiation of recombination in the recB1080 mutant are provided by different proteins, i.e., helicase activity by {RecB}1080CD, 5′ → 3′ exonuclease by {RecJ}- and {RecA}-single-stranded {DNA} filament formation by {{RecF}OR}.},
   author = {Ivana Ivančić-Baće and Petra Peharec and Sunčana Moslavac and Nikolina Škrobot and Erika Salaj-Šmic and Krunoslav Brčić-Kostić},
   doi = {10.1093/GENETICS/163.2.485},
   issn = {00166731},
   issue = {2},
   journal = {Genetics},
   month = {2},
   pages = {485-494},
   pmid = {12618388},
   publisher = {Oxford Academic},
   title = {{{RecF}OR} Function Is Required for {DNA} Repair and Recombination in a {RecA} Loading-Deficient recB Mutant of {{\textit{Escherichia coli}}}},
   volume = {163},
   url = {https://dx.doi.org/10.1093/genetics/163.2.485},
   year = {2003}
}
@article{Stracy2021,
   abstract = {To understand how {DNA}-binding proteins find their target sites, Stracy et al. tracked the motion of 11 diverse proteins in living {{\textit{Escherichia coli}}}. By comparing protein behavior in normal cells and cells without chromosomes, they showed that the {DNA}-binding proteins spend most of their search time bound to {DNA}.},
   author = {Mathew Stracy and Jakob Schweizer and David J. Sherratt and Achillefs N. Kapanidis and Stephan Uphoff and Christian Lesterlin},
   doi = {10.1016/J.MOLCEL.2021.01.039},
   issn = {1097-2765},
   issue = {7},
   journal = {Molecular Cell},
   keywords = {Chromosome-crowding,chromosome-free cells,single-molecule tracking,target search of bacterial {DNA}-binding proteins},
   month = {4},
   pages = {1499-1514.e6},
   pmid = {33621478},
   publisher = {Cell Press},
   title = {Transient non-specific {DNA} binding dominates the target search of bacterial {DNA}-binding proteins},
   volume = {81},
   year = {2021}
}
@article{Krasin1977,
   abstract = {A method was devised for extracting, from cells of {{\textit{Escherichia coli}}} K12, {DNA} molecules which sedimented on neutral sucrose gradients as would be expected for free {DNA} molecules approaching the genome in size. Gamma ray irradiation of oxygenated cells produced 0.20 {DNA} double-strand breaks per kilorad per 109 daltons. Incubation after irradiation of cells grown in K medium, with four to five genomes per cell, showed repair of the double-strand breaks. No repair of double-strand breaks was found in cells grown in aspartate medium, with only 1.3 genomes per cell, although {DNA} single-strand breaks were still efficiently repaired. Cells which were recA- or recA- recB- also did not repair double-strand breaks. These results suggest that repair of {DNA} double-strand breaks may occur by a recombinational event involving another {DNA} double helix with the same base sequence. © 1977.},
   author = {Frank Krasin and Franklin Hutchinson},
   doi = {10.1016/0022-2836(77)90120-6},
   issn = {0022-2836},
   issue = {1},
   journal = {Journal of Molecular Biology},
   month = {10},
   pages = {81-98},
   pmid = {338918},
   publisher = {Academic Press},
   title = {Repair of {DNA} double-strand breaks in {{\textit{Escherichia coli}}}, which requires recA function and the presence of a duplicate genome},
   volume = {116},
   year = {1977}
}
@article{Dillingham2008,
   abstract = {The {{RecB}CD} enzyme of {{\textit{Escherichia coli}}} is a helicase-nuclease that initiates the repair of double-stranded {DNA} breaks by homologous recombination. It also degrades linear double-stranded {DNA}, protecting the bacteria from phages and extraneous chromosomal {DNA}. The {{RecB}CD} enzyme is, however, regulated by a cis-acting {DNA} sequence known as Chi (crossover hotspot instigator) that activates its recombination-promoting functions. Interaction with Chi causes an attenuation of the {{RecB}CD} enzyme's vigorous nuclease activity, switches the polarity of the attenuated nuclease activity to the 5' strand, changes the operation of its motor subunits, and instructs the enzyme to begin loading the {RecA} protein onto the resultant Chi-containing single-stranded {DNA}. This enzyme is a prototypical example of a molecular machine: the protein architecture incorporates several autonomous functional domains that interact with each other to produce a complex, sequence-regulated, {DNA}-processing machine. In this review, we discuss the biochemical mechanism of the {{RecB}CD} enzyme with particular emphasis on new developments relating to the enzyme's structure and {DNA} translocation mechanism.},
   author = {Mark S. Dillingham and Stephen C. Kowalczykowski},
   issn = {1092-2172},
   issue = {4},
   journal = {Microbiology and Molecular Biology Reviews},
   month = {12},
   pages = {642-671},
   pmid = {19052323},
   publisher = {American Society for Microbiology},
   title = {{{RecB}CD} Enzyme and the Repair of Double-Stranded {DNA} Breaks},
   volume = {72},
   url = {https://journals.asm.org/doi/10.1128/mmbr.00020-08},
   year = {2008}
}
@article{Kohanski2010,
   abstract = {Bacterial responses to antibiotics are complex and involve many genetic and biochemical pathways. This Review describes the effects of bactericidal antibiotics on bacterial cellular processes, the associated responses that contribute to killing and recent insights into these processes revealed through the study of biological networks. Antibiotic drug–target interactions, and their respective direct effects, are generally well characterized. By contrast, the bacterial responses to antibiotic drug treatments that contribute to cell death are not as well understood and have proven to be complex as they involve many genetic and biochemical pathways. In this Review, we discuss the multilayered effects of drug–target interactions, including the essential cellular processes that are inhibited by bactericidal antibiotics and the associated cellular response mechanisms that contribute to killing. We also discuss new insights into these mechanisms that have been revealed through the study of biological networks, and describe how these insights, together with related developments in synthetic biology, could be exploited to create new antibacterial therapies.},
   author = {Michael A. Kohanski and Daniel J. Dwyer and James J. Collins},
   doi = {10.1038/nrmicro2333},
   issn = {1740-1534},
   issue = {6},
   journal = {Nature Reviews Microbiology 2010 8:6},
   keywords = {Antibiotics,Bacterial infection,Mechanism of action},
   month = {5},
   pages = {423-435},
   pmid = {20440275},
   publisher = {Nature Publishing Group},
   title = {How antibiotics kill bacteria: from targets to networks},
   volume = {8},
   url = {https://www.nature.com/articles/nrmicro2333},
   year = {2010}
}
@article{Wiktor2018,
   abstract = {The formation of 3 single-stranded {DNA} overhangs is a first and essential step during homology-directed repair of double-stranded breaks (DSB) of {DNA}, a task that in {{\textit{Escherichia coli}}} is performed by {{RecB}CD}. While this protein complex has been well characterized through in vitro single-molecule studies, it has remained elusive how end resection proceeds in the crowded and complex environment in live cells. Here, we develop a two-color fluorescent reporter to directly observe the resection of individual inducible DSB sites within live {{\textit{E. coli}}} cells. Realtime imaging shows that {{RecB}CD} during end resection degrades {DNA} with remarkably high speed (i"1.6 kb/s) and high processivity (>7sim;100 kb). The results show a pronounced asymmetry in the processing of the two {DNA} ends of a DSB, where much longer stretches of {DNA} are degraded in the direction of terminus. The microscopy observations are confirmed using quantitative polymerase chain reaction measurements of the {DNA} degradation. Deletion of the recD gene drastically decreased the length of resection, allowing for recombination with short ectopic plasmid homologies and significantly increasing the efficiency of horizontal gene transfer between strains.We thus visualized and quantified {DNA} end resection by the {{RecB}CD} complex in live cells, recorded {DNA}-degradation linked to end resection and uncovered a general relationship between the length of end resection and the choice of the homologous recombination template.},
   author = {Jakub Wiktor and Marit Van Der Does and Lisa Büller and David J. Sherratt and Cees Dekker},
   doi = {10.1093/NAR/GKX1290},
   issn = {0305-1048},
   issue = {4},
   journal = {Nucleic Acids Research},
   month = {2},
   pages = {1821-1833},
   pmid = {29294118},
   publisher = {Oxford Academic},
   title = {Direct observation of end resection by {{RecB}CD} during double-stranded {DNA} break repair in vivo},
   volume = {46},
   url = {https://dx.doi.org/10.1093/nar/gkx1290},
   year = {2018}
}
@article{Taylor1992,
   abstract = {During its unidirectional unwinding of {DNA}, {{RecB}CD} enzyme cuts one {DNA} strand near a properly oriented Chi site, a hotspot of homologous genetic recombination in {{\textit{Escherichia coli}}}. We report here th...},
   author = {Andrew F. Taylor and Gerald R. Smith},
   doi = {10.1073/PNAS.89.12.5226},
   issn = {00278424},
   issue = {12},
   journal = {Proceedings of the National Academy of Sciences},
   month = {6},
   pages = {5226-5230},
   pmid = {1535156},
   publisher = {Proceedings of the National Academy of Sciences},
   title = {{{RecB}CD} enzyme is altered upon cutting {DNA} at a chi recombination hotspot.},
   volume = {89},
   url = {https://www.pnas.org/doi/abs/10.1073/pnas.89.12.5226},
   year = {1992}
}
@article{Churchill2000,
   abstract = {Genetic recombination in {{\textit{Escherichia coli}}} is stimulated by the recombination hotspot Chi (χ), a regulatory element that modifies the activities of the {{RecB}CD} enzyme and leads to loading of the {DNA} strand exchange protein, {RecA}, onto the χ-containing {DNA} strand. The {RecB}C enzyme, which lacks the RecD subunit, loads {RecA} protein constitutively, in a manner that is independent of χ. Using a truncated {RecB}C enzyme lacking the 30 {kDa} C-terminal domain of the {RecB} subunit, we show that this domain is necessary for {RecA} protein-loading. We propose that this domain harbors a site that interacts with {RecA} protein, recruiting it to single-stranded {DNA} during unwinding. This ability of a translocating enzyme to deliver material ({RecA} protein) to a specific target site (the χ sequence) parallels that of other cellular motor proteins. (C) 2000 Academic Press.},
   author = {Jason J. Churchill and Stephen C. Kowalczykowski},
   doi = {10.1006/JMBI.2000.3590},
   issn = {0022-2836},
   issue = {3},
   journal = {Journal of Molecular Biology},
   keywords = {Chi sequence,Helicase,{RecA} protein,{{RecB}CD} enzyme,Recombination},
   month = {3},
   pages = {537-542},
   pmid = {10731409},
   publisher = {Academic Press},
   title = {Identification of the {RecA} protein-loading domain of {{RecB}CD} enzyme},
   volume = {297},
   year = {2000}
}
@article{Spies2006,
   abstract = {{{RecB}CD} enzyme facilitates loading of {RecA} protein onto ss{DNA} produced by its helicase/nuclease activity. This process is essential for {{RecB}CD}-mediated homologous recombination. Here, we establish that the C-terminal nuclease domain of the {RecB} subunit ({RecB}nuc) forms stable complexes with {RecA}. Interestingly, {RecB}nuc also interacts with and loads noncognate {DNA} strand exchange proteins. Interaction is with a conserved element of the {RecA}-fold, but because the binding to noncognate proteins decreases in a phylogenetically consistent way, species-specific interactions are also present. {RecB}nuc does not impede activities of {RecA} that are important to {DNA} strand exchange, consistent with its role in targeting of {RecA}. Modeling predicts the interaction interface for the {RecA}-{{RecB}CD} complex. Because a similar interface is involved in the binding of human Rad51 to the conserved BRC repeat of BRCA2 protein, the {RecB}-domain may be one of several structural domains that interact with and recruit {DNA} strand exchange proteins to {DNA}. ©2006 Elsevier Inc.},
   author = {Maria Spies and Stephen C. Kowalczykowski},
   doi = {10.1016/j.molcel.2006.01.007},
   issn = {10972765},
   issue = {4},
   journal = {Molecular Cell},
   keywords = {{DNA}},
   month = {2},
   pages = {573-580},
   pmid = {16483938},
   publisher = {Elsevier},
   title = {The {RecA} binding locus of {{RecB}CD} is a general domain for recruitment of {DNA} strand exchange proteins},
   volume = {21},
   url = {http://www.cell.com/article/S1097276506000086/fulltext http://www.cell.com/article/S1097276506000086/abstract https://www.cell.com/molecular-cell/abstract/S1097-2765(06)00008-6},
   year = {2006}
}
@article{Taylor1999,
   abstract = {We report here an unusual mechanism for enzyme regulation: the disassembly of all three subunits of {{RecB}CD} enzyme after its interaction with a Chi recombination hot spot. The enzyme, which is essential for the major pathway of recombination in {{\textit{Escherichia coli}}}, acts on linear double-stranded {DNA} bearing a Chi site to produce single-stranded {DNA} substrates for strand exchange by {RecA} protein. We show that after reaction with {DNA} bearing Chi sites, {{RecB}CD} enzyme is inactivated and the three subunits migrate as separate species during glycerol gradient ultracentrifugation or native gel electrophoresis. This Chi-mediated inactivation and disassembly of purified {{RecB}CD} enzyme can account for the previously reported Chi-dependent loss of Chi activity in {{\textit{E. coli}}} cells containing broken {DNA}. Our results support a model of recombination in which Chi regulates one {{RecB}CD} enzyme molecule to make a single recombinational exchange ('one enzyme-one exchange' hypothesis).},
   author = {Andrew F. Taylor and Gerald R. Smith},
   journal = {Genes and development},
   keywords = {Chi sites,{{\textit{Escherichia coli}}},[Key Words: {{RecB}CD} enzyme,disassembly],genetic recombination},
   title = {Regulation of homologous recombination: Chi inactivates {{RecB}CD} enzyme by disassembly of the three subunits},
   url = {https://genesdev.cshlp.org/content/13/7/890.short},
   year = {1999}
}
@article{Michel1997,
   abstract = {We report here that {DNA} double‐strand breaks (DSBs) form in {{\textit{Escherichia coli}}} upon arrest of replication forks due to a defect in, or the inhibition of, replicative {DNA} helicases. The formation of D...},
   author = {Bénédicte Michel and S. Dusko Ehrlich and Marilyne Uzest},
   doi = {10.1093/EMBOJ/16.2.430},
   issn = {02614189},
   issue = {2},
   journal = {The EMBO Journal},
   keywords = {{DnaB},{{RecB}CD},Rep,helicase,homologous recombination},
   month = {1},
   pages = {430-438},
   pmid = {9029161},
   publisher = {John Wiley \& Sons, LtdChichester, UK},
   title = {{DNA} double‐strand breaks caused by replication arrest},
   volume = {16},
   url = {https://www.embopress.org/doi/10.1093/emboj/16.2.430},
   year = {1997}
}
@article{Seigneur1998,
   abstract = {Replication arrest leads to the occurrence of {DNA} double-stranded breaks (DSB). We studied the mechanism of DSB formation by direct measure of the amount of in vivo linear {DNA} in {{\textit{Escherichia coli}}} cells that lack the {{RecB}CD} recombination complex and by genetic means. The {RuvAB}C proteins, which catalyze migration and cleavage of Holliday junctions, are responsible for the occurrence of DSBs at arrested replication forks. In cells proficient for {RecB}C, {RuvAB} is uncoupled from RuvC and DSBs may be prevented. This may be explained if a Holliday junction forms upon replication fork arrest, by annealing of the two nascent strands. {{RecB}CD} may act on the double-stranded tail prior to the cleavage of the {RuvAB}-bound junction by RuvC to rescue the blocked replication fork without breakage.},
   author = {Marie Seigneur and Vladimir Bidnenko and S. Dusko Ehrlich and Bénédicte Michel},
   doi = {10.1016/S0092-8674(00)81772-9},
   issn = {00928674},
   issue = {3},
   journal = {Cell},
   month = {10},
   pages = {419-430},
   pmid = {9814711},
   publisher = {Elsevier B.V.},
   title = {{RuvAB} acts at arrested replication forks},
   volume = {95},
   url = {http://www.cell.com/article/S0092867400817729/fulltext http://www.cell.com/article/S0092867400817729/abstract https://www.cell.com/cell/abstract/S0092-8674(00)81772-9},
   year = {1998}
}
@article{Michel2001,
   abstract = {{DNA} synthesis is an accurate and very processive phenomenon; nevertheless, replication fork progression on chromosomes can be impeded by {DNA} lesions, {DNA} secondary structures, or {DNA}-bound proteins...},
   author = {B. Michel and M. J. Flores and E. Viguera and G. Grompone and M. Seigneur and V. Bidnenko},
   doi = {10.1073/PNAS.111008798},
   issn = {00278424},
   issue = {15},
   journal = {Proceedings of the National Academy of Sciences},
   month = {7},
   pages = {8181-8188},
   pmid = {11459951},
   publisher = {The National Academy of Sciences},
   title = {Rescue of arrested replication forks by homologous recombination},
   volume = {98},
   url = {https://www.pnas.org/doi/abs/10.1073/pnas.111008798},
   year = {2001}
}
@article{Amarh2018,
   abstract = {Chromosomal replication is the major source of spontaneous {DNA} double-strand breaks (DSBs) in living cells. Repair of these DSBs is essential for cell viability, and accuracy of repair is critical to avoid chromosomal rearrangements. Repair of replication-dependent DSBs occurs primarily by homologous recombination with a sister chromosome. However, this reaction has never been visualized at a defined chromosomal locus, so little is known about its spatial or temporal dynamics. Repair of a replication-independent DSB generated in {{\textit{Escherichia coli}}} by a rare-cutting endonuclease leads to the formation of a bundle of {RecA} filaments. In this study, we show that in contrast, repair of a replication-dependent DSB involves a transient {RecA} focus localized in the central region of the cell in which the {DNA} is replicated. The recombining loci remain centrally located with restricted movement before segregating with little extension to the period of postreplicative sister-chromosome cohesion. The spatial and temporal efficiency of this reaction is remarkable.},
   author = {Vincent Amarh and Martin A. White and David R.F. Leach},
   doi = {10.1083/JCB.201803020},
   issn = {0021-9525},
   issue = {7},
   journal = {Journal of Cell Biology},
   keywords = {chromosomes,dna,double-stranded dna breaks,recombination, genetic},
   month = {7},
   pages = {2299-2307},
   pmid = {29789437},
   publisher = {The Rockefeller University Press},
   title = {Dynamics of {RecA}-mediated repair of replication-dependent {DNA} breaks},
   volume = {217},
   url = {https://doi.org/10.1083/jcb.201803020},
   year = {2018}
}
@article{Horii1968,
   abstract = {Abstract— Degradation of the {DNA} of a rec‐ mutant of {{\textit{Escherichia coli}}} K12 (JC1569 b) induced by u.v. light was investigated. The rate of degradation was much larger by growing bacteria than by stationary cells. When growing bacteria were starved for amino acids, their {DNA} became resistant to irradiation. The mode of u.v.‐induced degradation was investigated by comparing the time course of release from the acid‐insoluble fraction of the label for two growing cultures; the one was pulse‐labeled with 3H‐thymidine and the other was pulse‐labeled and chased thereafter for 12 min. It was found that the label incorporated into the former culture begins to be lost from the acid‐insoluble fraction prior to the loss of the label incorporated into the latter culture. It was concluded that breakdown of the replicating point precedes degradation of the bulk of the {DNA}. This result suggested that the replicating point is a sensitive site to irradiation and the u.v.‐induced degradation of {DNA} seemed to be influenced by the state of chromosome at the time of irradiation. Experiments of centrifugation of lysed spheroplasts of bacteria uniformly labeled with 3H‐thymidine in alkaline sucrose demonstrated that {DNA} of low molecular weight appeared after irradiation with only 5 ergs/ mm2, and that the molecular weight could not be restored by post‐irradiation incubation. Considering these results, an hypothesis is proposed concerning the initiation of induced degradation of the {DNA} of the rec‐ mutant. Copyright © 1968, Wiley Blackwell. All rights reserved},
   author = {Z. I. Horii and K. Suzuki},
   doi = {10.1111/J.1751-1097.1968.TB05850.X},
   issn = {1751-1097},
   issue = {2},
   journal = {Photochemistry and Photobiology},
   month = {8},
   pages = {93-105},
   publisher = {John Wiley \& Sons, Ltd},
   title = {Degradation of the {DNA} of {{\textit{Escherichia coli}}} K12 rec- (JC1569b) after irradiation with ultraviolet light},
   volume = {8},
   url = {https://onlinelibrary.wiley.com/doi/full/10.1111/j.1751-1097.1968.tb05850.x https://onlinelibrary.wiley.com/doi/abs/10.1111/j.1751-1097.1968.tb05850.x https://onlinelibrary.wiley.com/doi/10.1111/j.1751-1097.1968.tb05850.x},
   year = {1968}
}
@article{Chow2007,
   abstract = {Chow, K-H. and Courcelle, J. {{RecB}CD} and {RecJ}/{RecQ} Initiate {DNA} Degradation on Distinct Substrates in {UV}-Irradiated {{\textit{Escherichia coli}}}. Radiat. Res. 168, 499–506 (2007).After {UV} irradiation, recA mutants fail to recover replication, and a dramatic and nearly complete degradation of the genomic {DNA} occurs. Although the {{RecB}CD} helicase/nuclease complex is known to mediate this catastrophic {DNA} degradation, it is not known how or where this degradation is initiated. Previous studies have speculated that {{RecB}CD} targets and initiates degradation from the nascent {DNA} at replication forks arrested by {DNA} damage. To test this question, we examined which enzymes were responsible for the degradation of genomic {DNA} and the nascent {DNA} in {UV}-irradiated recA cells. We show here that, although {{RecB}CD} degrades the genomic {DNA} after {UV} irradiation, it does not target the nascent {DNA} at arrested replication forks. Instead, we observed that the nascent {DNA} at arrested replication forks in recA cultures is degraded by {RecJ}/{RecQ}, similar to what occurs in wild-type cultures. These findings indicate that the genomic {DNA} degradation and nascent {DNA} degradation in {UV}-irradiated recA mutants are mediated separately through {{RecB}CD} and {RecJ}/{RecQ}, respectively. In addition, they demonstrate that {{RecB}CD} initiates degradation at a site(s) other than the arrested replication fork directly.},
   author = {Kin-Hoe Chow and Justin Courcelle},
   doi = {10.1667/RR1033.1},
   issn = {0033-7587},
   issue = {4},
   journal = {Radiation Research},
   month = {10},
   pages = {499-506},
   publisher = {Radiation Research Society},
   title = {{{RecB}CD} and {RecJ}/{RecQ} Initiate {DNA} Degradation on Distinct Substrates in {UV}-Irradiated {{\textit{Escherichia coli}}}},
   volume = {168},
   url = {https://bioone.org/journals/radiation-research/volume-168/issue-4/RR1033.1/{{RecB}CD}-and-{RecJ}-{RecQ}-Initiate-{DNA}-Degradation-on-Distinct-Substrates/10.1667/RR1033.1.full https://bioone.org/journals/radiation-research/volume-168/issue-4/RR1033.1/{{RecB}CD}-and-{RecJ}-{RecQ}-Initiate-{DNA}-Degradation-on-Distinct-Substrates/10.1667/RR1033.1.short},
   year = {2007}
}
@article{Wang2000,
   abstract = {The {{RecB}CD} enzyme of {{\textit{Escherichia coli}}} is an ATP-dependent {DNA} exonuclease and a helicase. Its exonuclease activity is subject to regulation by an octameric nucleotide sequence called χ. In this study, site-directed mutations were made in the carboxyl-terminal nuclease domain of the {RecB} subunit, and their effects on {{RecB}CD}'s enzymatic activities were investigated. Mutation of two amino acid residues, Asp1067 and Lys1082, abolished nuclease activity on both single- and double-stranded {DNA}. Together with Asp1080, these residues compose a motif that is similar to one shown to form the active site of several restriction endonucleases. The nuclease reactions catalyzed by the {{RecB}CD} enzyme should therefore follow the same mechanism as these restriction endonucleases. Furthermore, the mutant enzymes were unable to produce χ-specific fragments that are thought to result from the 3'-5' and 5'-3' single-stranded exonuclease activities of the enzyme during its reaction with χ-containing double-stranded {DNA}. The results show that the nuclease active site in the {RecB} C-terminal 30-{kDa} domain is the universal nuclease active site of {{RecB}CD} that is responsible for {DNA} degradation in both directions during the reaction with double-stranded {DNA}. A novel explanation for the observed nuclease polarity switch and {{RecB}CD}-{DNA} interaction is offered.},
   author = {Jingdi Wang and Ruiwu Chen and Douglas A. Julin},
   doi = {10.1074/jbc.275.1.507},
   issn = {00219258},
   issue = {1},
   journal = {Journal of Biological Chemistry},
   month = {1},
   pages = {507-513},
   pmid = {10617645},
   publisher = {Elsevier},
   title = {A single nuclease active site of the {{\textit{Escherichia coli}}} {{RecB}CD} enzyme catalyzes single-stranded {DNA} degradation in both directions},
   volume = {275},
   url = {http://www.jbc.org/article/S0021925819528927/fulltext http://www.jbc.org/article/S0021925819528927/abstract https://www.jbc.org/article/S0021-9258(19)52892-7/abstract},
   year = {2000}
}
@article{Anderson1999,
   abstract = {Homologous recombination and double-stranded {DNA} break repair in {{\textit{Escherichia coli}}} are initiated by the multifunctional {{RecB}CD} enzyme. After binding to a double-stranded {DNA} end, the {{RecB}CD} enzyme unwinds and degrades the {DNA} processively. This processing is regulated by the recombination hot spot, Chi (χ: 5'-GCTGGTGG-3'), which induces a switch in the polarity of {DNA} degradation and activates {{RecB}CD} enzyme to coordinate the loading of the {DNA} strand exchange protein, {RecA}, onto the single-stranded {DNA} products of unwinding. Recently, a single mutation in {RecB}, Asp-1080 → Ala, was shown to create an enzyme ({RecB}({D1080A})CD) that is a processive helicase but not a nuclease. Here we show that the {RecB}({D1080A})CD enzyme is also unable to coordinate the loading of the {RecA} protein, regardless of whether χ sites are present in the {DNA}. However, the {RecB}({D1080A})CD enzyme does respond to χ sites by inactivating in a χ-dependent manner. These data define a locus of the {{RecB}CD} enzyme that is essential not only for nuclease function but also for the coordination of {RecA} protein loading.},
   author = {Daniel G. Anderson and Jason J. Churchill and Stephen C. Kowalczykowski},
   doi = {10.1074/jbc.274.38.27139},
   issn = {00219258},
   issue = {38},
   journal = {Journal of Biological Chemistry},
   month = {9},
   pages = {27139-27144},
   pmid = {10480929},
   publisher = {Elsevier},
   title = {A single mutation, {RecB}({D1080A}), eliminates {RecA} protein loading but not Chi recognition by {{RecB}CD} enzyme},
   volume = {274},
   url = {http://www.jbc.org/article/S0021925819551365/fulltext http://www.jbc.org/article/S0021925819551365/abstract https://www.jbc.org/article/S0021-9258(19)55136-5/abstract},
   year = {1999}
}
@article{Huang2021,
   abstract = {The widely used quinolone antibiotics act by trapping prokaryotic type IIA topoisomerases, resulting in irreversible topoisomerase cleavage complexes (TOPcc). Whereas the excision repair pathways of TOPcc in eukaryotes have been extensively studied, it is not known whether equivalent repair pathways for prokaryotic TOPcc exist. By combining genetic, biochemical, and molecular biology approaches, we demonstrate that exonuclease {VII} (Exo{VII}) excises quinolone-induced trapped {DNA} gyrase, an essential prokaryotic type IIA topoisomerase. We show that Exo{VII} repairs trapped type IIA TOPcc and that Exo{VII} displays tyrosyl nuclease activity for the tyrosyl-{DNA} linkage on the 5′-{DNA} overhangs corresponding to trapped type IIA TOPcc. Exo{VII}-deficient bacteria fail to remove trapped {DNA} gyrase, consistent with their hypersensitivity to quinolones. We also identify an Exo{VII} inhibitor that synergizes with the antimicrobial activity of quinolones, including in quinolone-resistant bacterial strains, further demonstrating the functional importance of Exo{VII} for the repair of type IIA TOPcc.},
   author = {Shar Yin N. Huang and Stephanie A. Michaels and Brianna B. Mitchell and Nadim Majdalani and Arnaud Vanden Broeck and Andres Canela and Yuk Ching Tse-Dinh and Valerie Lamour and Yves Pommier},
   issn = {23752548},
   issue = {10},
   journal = {Science Advances},
   month = {3},
   pmid = {33658195},
   publisher = {American Association for the Advancement of Science},
   title = {Exonuclease {VII} repairs quinolone-induced damage by resolving {DNA} gyrase cleavage complexes},
   volume = {7},
   url = {https://www.science.org/doi/10.1126/sciadv.abe0384},
   year = {2021}
}
@article{Wentzell2000,
   abstract = {Quinolone drugs can inhibit bacterial {DNA} replication, via interaction with the type II topoisomerase {DNA} gyrase. Using a {DNA} template containing a preferred site for quinolone-induced gyrase cleavage, we have demonstrated that the passage of the bacteriophage {T7} replication complex is blocked in vitro by the formation of a gyrase-drug-{DNA} complex. The majority of the polymerase is arrested approximately 10 bp upstream of this preferred site, although other minor sites of blocking have been observed. The ability of mutant gyrase proteins to arrest {DNA} replication in vitro has been investigated. Gyrase containing mutations in the A subunit at either the active-site tyrosine (Tyr122) or Ser83 (a residue known to be involved in quinolone interaction) failed to halt the progress of the polymerase. A low-level, quinolone-resistant mutation in the B subunit of gyrase showed reduced blocking compared to wild-type. We have demonstrated that {DNA} cleavage and replication blocking occur on similar time-scales and we conclude that formation of the cleavable complex is a prerequisite for polymerase blocking. Additionally, we have shown that collision of the replication proteins with the gyrase-drug-{DNA} complex is not sufficient to render this complex irreversible and that further factors must be involved in processing this stalled complex. © 2000 Academic Press.},
   author = {L. M. Wentzell and A. Maxwell},
   doi = {10.1006/JMBI.2000.4266},
   issn = {0022-2836},
   issue = {5},
   journal = {Journal of Molecular Biology},
   keywords = {Ciprofloxacin,Supercoiling,Topoisomerase},
   month = {12},
   pages = {779-791},
   pmid = {11124026},
   publisher = {Academic Press},
   title = {The Complex of {DNA} Gyrase and Quinolone Drugs on {DNA} Forms a Barrier to the {T7} {DNA} Polymerase Replication Complex},
   volume = {304},
   year = {2000}
}
@article{Drlica2008,
   abstract = {The fluoroquinolones are broad-spectrum antibacterial agents that are becoming increasingly popular as bacterial resistance erodes the effectiveness of other agents (fluoroquin-olone sales accounted for 18% of the antibacterial market in 2006) (41). One of the attractive features of the quinolones is their ability to kill bacteria rapidly, an ability that differs widely among the various derivatives. For example, quinolones differ in rate and extent of killing, in the need for aerobic metabolism to kill cells, and in the effect of protein synthesis inhibitors on quinolone lethality. Understanding the mechanisms underlying these differences could lead to new ways for identifying the most bactericidal quinolone derivatives. Before describing the types of damage caused by the quin-olones, it is useful to define lethal activity. Operationally, it is the ability of drug treatment to reduce the number of viable cells, usually measured as CFU on drug-free agar after treatment. This assay is distinct from measurements that detect inhibition of growth (e.g., MIC), since with the latter bacteria are exposed to drug throughout the measurement. The distinction between killing and blocking growth is important because it allows susceptibility determinations to be related to particular biological processes. For example, inhibition of growth is typically reversed by the removal of drug, while cell death is not. Thus, biochemical events associated with blocking growth should be readily reversible, while those responsible for cell death should be difficult to reverse. Reversibility can be used to distinguish among quinolone derivatives and assign functions to particular aspects of drug structure. Moreover, protective functions, such as repair and stress responses, can be distinguished by whether their absence affects inhibition of growth, killing, or both. The intracellular targets of the quinolones are two {DNA} topoisomerases: gyrase and topoisomerase IV. Gyrase tends to be the primary target in gram-negative bacteria, while topo-isomerase IV is preferentially inhibited by most quinolones in gram-positive organisms (28). Both enzymes use a double-strand {DNA} passage mechanism, and it is likely that quinolone biochemistry is similar for both. However, physiological differences between the enzymes exist, some of which may bear on quinolone lethality. In the present minireview we consider cell death through a two-part "poison" hypothesis in which the quinolones form reversible drug-topoisomerase-{DNA} complexes that subsequently lead to several types of irreversible (lethal) damage. Other consequences of quinolone treatment, such as depletion of gyrase and topoisomerase IV activity, are probably less immediate (42). To provide a framework for considering quin-olone lethality, we begin by briefly describing the drug-topo-isomerase-{DNA} complexes. Readers interested in a more comprehensive discussion of quinolones are referred to a previously published work (28). QUINOLONE-TOPOISOMERASE-{DNA} COMPLEXES As a normal part of their reaction mechanism, gyrase and topoisomerase IV introduce a pair of staggered, single-strand breaks (nicks) into {DNA} and become covalently bound to the 5 ends of the cleaved {DNA} (55, 57). Quinolones bind rapidly to enzyme-{DNA} complexes (35), probably before {DNA} cleav-age occurs (Fig. 1, step b 1); drug binding occurs with mutant gyrase (gyrA) or topoisomerase IV (parC) that fails to cleave {DNA} (9, 54). After drug binding, a slower, {DNA} cleavage-associated step occurs (35) that results in drug-mediated inhibition of religation of the {DNA} ends by topoisomerases (1). In a sense, quinolones trap the bacterial type II topoisomerases on {DNA} (17, 23, 73, 75) (Fig. 1, step b 2). The resulting structures have been termed cleavable, cleavage, and cleaved complexes. In the present study we refer to them as cleaved complexes to emphasize that phosphodiester bonds in the {DNA} moiety are broken (the term ternary complex is reserved for the early step in drug-enzyme-{DNA} complex formation in which the {DNA} is unbroken). A variety of quinolone-mediated phenomena follow from formation of cleaved complexes. To better understand lethal processes, we briefly describe key features of the {DNA} and protein components of the complexes. Since crystal structures have not been reported for cleaved complexes, tentative inferences concerning the relative positions of drug, protein, and {DNA} are drawn from biochemical experiments, partial structures of the bacterial topoisomerases, and complete structures of eukaryotic topoisomerase II. Evidence for the {DNA} being cleaved derives from the recovery of {DNA} fragments when cleaved complexes are treated with protein denaturants, such as sodium dodecyl sulfate (SDS) (Fig. 1, step g). These {DNA} fragments are covalently bound to the GyrA or ParC proteins (55, 57). While cleaved complexes preferentially form at particular sites on {DNA} (44,},
   author = {Karl Drlica and Muhammad Malik and Robert J. Kerns and Xilin Zhao},
   issn = {00664804},
   issue = {2},
   journal = {Antimicrobial Agents and Chemotherapy},
   month = {2},
   pages = {385-392},
   pmid = {17724149},
   publisher = {American Society for Microbiology},
   title = {Quinolone-mediated bacterial death},
   volume = {52},
   url = {https://journals.asm.org/doi/10.1128/aac.01617-06},
   year = {2008}
}
@article{Zhao2006,
   abstract = {Inhibition of {DNA} replication in an {{\textit{Escherichia coli}}} dnaB-22 mutant failed to block quinolone-mediated lethality. Inhibition of protein synthesis by chloramphenicol inhibited nalidixic acid lethality and, to a lesser extent, ciprofloxacin lethality in both dnaB-22 and wild-type cells. Thus, major features of quinolone-mediated lethality do not depend on ongoing replication. Copyright © 2006, American Society for Microbiology. All Rights Reserved.},
   author = {Xilin Zhao and Muhammad Malik and Nymph Chan and Alex Drlica-Wagner and Jian Ying Wang and Xinying Li and Karl Drlica},
   issn = {00664804},
   issue = {1},
   journal = {Antimicrobial Agents and Chemotherapy},
   month = {1},
   pages = {362-364},
   pmid = {16377712},
   publisher = {American Society for Microbiology},
   title = {Lethal action of quinolones against a temperature-sensitive dnaB replication mutant of {{\textit{Escherichia coli}}}},
   volume = {50},
   url = {https://journals.asm.org/doi/10.1128/aac.50.1.362-364.2006},
   year = {2006}
}
@article{Lesterlin2013,
   abstract = {{RecA} bundles are shown to be important for the pairing of homologous loci that have segregated to opposite ends of the cell during {DNA} double-strand break repair in vivo in {{\textit{Escherichia coli}}}. Although bacterial {RecA} protein functions as a filament during {DNA} strand exchange, early studies of {RecA} also noted that, in vivo, it formed bundles. These bundles were inactive in vitro, so were thought to be a way of storing {RecA} until it was needed. David Sherratt and colleagues now show that {RecA} bundles do have an important function in vivo. Super-resolution microscopy imaging shows that bundles are excluded from the bulk of the nucleoid and locate to the cell periphery where they facilitate the pairing of homologous loci that have segregated to opposite ends of the cell. After sister locus pairing, {RecA} bundles disassemble. {DNA} double-strand break (DSB) repair by homologous recombination has evolved to maintain genetic integrity in all organisms1. Although many reactions that occur during homologous recombination are known1,2,3, it is unclear where, when and how they occur in cells. Here, by using conventional and super-resolution microscopy, we describe the progression of DSB repair in live {{\textit{Escherichia coli}}}. Specifically, we investigate whether homologous recombination can occur efficiently between distant sister loci that have segregated to opposite halves of an {{\textit{E. coli}}} cell. We show that a site-specific DSB in one sister can be repaired efficiently using distant sister homology. After {{RecB}CD} processing of the DSB, {RecA} is recruited to the cut locus, where it nucleates into a bundle that contains many more {RecA} molecules than can associate with the two single-stranded {DNA} regions that form at the DSB. Mature bundles extend along the long axis of the cell, in the space between the bulk nucleoid and the inner membrane. Bundle formation is followed by pairing, in which the two ends of the cut locus relocate at the periphery of the nucleoid and together move rapidly towards the homology of the uncut sister. After sister locus pairing, {RecA} bundles disassemble and proteins that act late in homologous recombination are recruited to give viable recombinants 1–2-generation-time equivalents after formation of the initial DSB. Mutated {RecA} proteins that do not form bundles are defective in sister pairing and in DSB-induced repair. This work reveals an unanticipated role of {RecA} bundles in channelling the movement of the {DNA} DSB ends, thereby facilitating the long-range homology search that occurs before the strand invasion and transfer reactions.},
   author = {Christian Lesterlin and Graeme Ball and Lothar Schermelleh and David J. Sherratt},
   doi = {10.1038/nature12868},
   issn = {1476-4687},
   issue = {7487},
   journal = {Nature 2013 506:7487},
   keywords = {Chromosomes,{DNA} recombination,Double,strand {DNA} breaks},
   month = {12},
   pages = {249-253},
   pmid = {24362571},
   publisher = {Nature Publishing Group},
   title = {{RecA} bundles mediate homology pairing between distant sisters during {DNA} break repair},
   volume = {506},
   url = {https://www.nature.com/articles/nature12868},
   year = {2013}
}
@article{Badrinarayanan2015,
   abstract = {Double-strand breaks (DSBs) can lead to the loss of genetic information and cell death. Although DSB repair via homologous recombination has been well characterized, the spatial organization of this process inside cells remains poorly understood, and the mechanisms used for chromosome resegregation after repair are unclear. In this paper, we introduced site-specific DSBs in Caulobacter crescentus and then used time-lapse microscopy to visualize the ensuing chromosome dynamics. Damaged loci rapidly mobilized after a DSB, pairing with their homologous partner to enable repair, before being resegregated to their original cellular locations, independent of {DNA} replication. Origin-proximal regions were resegregated by the ParABS system with the ParA structure needed for resegregation assembling dynamically in response to the DSB-induced movement of an origin-associated ParB away from one cell pole. Origin-distal regions were resegregated in a ParABS-independent manner and instead likely rely on a physical, spring-like force to segregate repaired loci. Collectively, our results provide a mechanistic basis for the resegregation of chromosomes after a DSB.},
   author = {Anjana Badrinarayanan and Tung B.K. Le and Michael T. Laub},
   doi = {10.1083/JCB.201505019},
   issn = {0021-9525},
   issue = {3},
   journal = {Journal of Cell Biology},
   keywords = {double strand break repair,double-stranded dna breaks},
   month = {8},
   pages = {385-400},
   pmid = {26240183},
   publisher = {The Rockefeller University Press},
   title = {Rapid pairing and resegregation of distant homologous loci enables double-strand break repair in bacteria},
   volume = {210},
   url = {www.jcb.org/cgi/doi/10.1083/jcb.201505019},
   year = {2015}
}
@article{Kidane2005,
   abstract = {We show that {RecN} protein is recruited to a defined {DNA} double strand break (DSB) in Bacillus subtilis cells at an early time point during repair. Because {RecO} and {RecF} are successively recruited to DSBs, it is now clear that dynamic DSB repair centers (RCs) exist in prokaryotes. {RecA} protein was also recruited to RCs and formed highly dynamic filamentous structures, which we term threads, across the nucleoids. Formation of {RecA} threads commenced ∼30 min after the induction of DSBs, after {RecN} recruitment to RCs, and disassembled after 2 h. Time-lapse microscopy showed that the threads rapidly changed in length, shape, and orientation within minutes and can extend at 1.02μm/min. The formation of {RecA} threads was abolished in recJ addAB mutant cells but not in each of the single mutants, suggesting that {RecA} filaments can be initiated via two pathways. Contrary to proteins forming RCs, {DNA} polymerase I did not form foci but was present throughout the nucleoids (even after induction of DSBs or after {UV} irradiation), suggesting that it continuously scans the chromosome for {DNA} lesions. © The Rockefeller University Press.},
   author = {Dawit Kidane and Peter L. Graumann},
   issn = {00219525},
   issue = {3},
   journal = {Journal of Cell Biology},
   keywords = {bacillus subtilis,burkina faso,chromosomes,dna,dna polymerase i,dna, double-stranded,double-stranded dna breaks,prokaryotic cells,rec a recombinases},
   month = {9},
   pages = {357-366},
   pmid = {16061691},
   publisher = {The Rockefeller University Press},
   title = {Dynamic formation of {RecA} filaments at {DNA} double strand break repair centers in live cells},
   volume = {170},
   url = {http://www.jcb.org/cgi/},
   year = {2005}
}
@article{Renzette2005,
   abstract = {{RecA} is important in recombination, {DNA} repair and repair of replication forks. It functions through the production of a protein-{DNA} filament. To study the localization of {RecA} in live {{\textit{Escherichia coli}}} cells, the {RecA} protein was fused to the green fluorescence protein ({GFP}). Strains with this gene have recombination/{DNA} repair activities three- to tenfold below wild type (or about 1000-fold above that of a recA null mutant). {RecA}-{GFP} cells have a background of green fluorescence punctuated with up to five foci per cell. Two types of foci have been defined: 4,6-diamidino-2-phenylindole (DAPI)-sensitive foci that are bound to {DNA} and DAPI-insensitive foci that are {DNA}-less aggregates/storage structures. In log phase cells, foci were not localized to any particular region. After {UV} irradiation, the number of foci increased and they localized to the cell centre. This suggested colocalization with the {DNA} replication factory. recA, recB and recF strains showed phenotypes and distributions of foci consistent with the predicted effects of these mutations. © 2005 Blackwell Publishing Ltd.},
   author = {Nicholas Renzette and Nathan Gumlaw and Jared T. Nordman and Marlee Krieger and Su Ping Yeh and Edward Long and Richard Centore and Ruethairat Boonsombat and Steven J. Sandler},
   doi = {10.1111/J.1365-2958.2005.04755.X},
   isbn = {4135451578},
   issn = {1365-2958},
   issue = {4},
   journal = {Molecular Microbiology},
   month = {8},
   pages = {1074-1085},
   pmid = {16091045},
   publisher = {John Wiley \& Sons, Ltd},
   title = {Localization of {RecA} in {{\textit{Escherichia coli}}} {K-12} using {RecA}–{GFP}},
   volume = {57},
   url = {https://onlinelibrary.wiley.com/doi/full/10.1111/j.1365-2958.2005.04755.x https://onlinelibrary.wiley.com/doi/abs/10.1111/j.1365-2958.2005.04755.x https://onlinelibrary.wiley.com/doi/10.1111/j.1365-2958.2005.04755.x},
   year = {2005}
}
@article{Centore2007,
   abstract = {{RecA} is important for recombination, {DNA} repair, and {SOS} induction. In Escherichia colt, {{RecB}CD}, {{RecF}OR}, and {{RecJ}Q} prepare {DNA} substrates onto which {RecA} binds. {UvrD} is a 3′-to-5′ helicase that participates in methyl-directed mismatch repair and nucleotide excision repair. uvrD deletion mutants are sensitive to {UV} irradiation, hypermutable, and hyper-rec. In vitro, {UvrD} can dissociate {RecA} from single-stranded {DNA}. Other experiments suggest that {UvrD} removes {RecA} from {DNA} where it promotes unproductive reactions. To test if {UvrD} limits the number and/or the size of {RecA}-{DNA} structures in vivo, an uvrD mutation was combined with recA-gfp. This recA allele allows the number of {RecA} structures and the amount of {RecA} at these structures to be assayed in living cells. uvrD mutants show a threefold increase in the number of {RecA}-{GFP} foci, and these foci are, on average, nearly twofold higher in relative intensity. The increased number of {RecA}-green fluorescent protein foci in the uvrD mutant is dependent on recF, recO, recR, recJ, and recQ. The increase in average relative intensity is dependent on recO and recQ. These data support an in vivo role for {UvrD} in removing {RecA} from the {DNA}. Copyright © 2007, American Society for Microbiology. All Rights Reserved.},
   author = {Richard C. Centore and Steven J. Sandler},
   issn = {00219193},
   issue = {7},
   journal = {Journal of Bacteriology},
   month = {4},
   pages = {2915-2920},
   pmid = {17259317},
   publisher = {American Society for Microbiology},
   title = {{UvrD} limits the number and intensities of recA-green fluorescent protein structures in {{\textit{Escherichia coli}}} {K-12}},
   volume = {189},
   url = {https://journals.asm.org/doi/10.1128/jb.01777-06},
   year = {2007}
}
@article{Liu2013,
   abstract = {The bacterial {{RecB}CD} helicase/nuclease shows broad, and apparently static, heterogeneity in the unwinding rate manifest by individual molecules: here it is shown that transiently halting an enzyme during processive translocation allows for a change, most likely conformational, such that the velocity of the molecule after pausing can fall anywhere within the spectrum of rates seen for a population. The bacterial {{RecB}CD} helicase/nuclease shows broad heterogeneity in unwinding rates of individual molecules. Stephen Kowalczykowski and colleagues have investigated this behaviour and find that transiently stopping the enzyme during processive translocation allows for a change, probably conformational, such that the velocity of the molecule after pausing can fall anywhere within the spectrum of rates seen for a population. The interpretation is that ligand binding stabilizes a particular equilibrium conformational sub-state that can persist for the duration of unwinding of the substrates used, and that determines the rate for that molecule. Single-molecule studies can overcome the complications of asynchrony and ensemble-averaging in bulk-phase measurements, provide mechanistic insights into molecular activities, and reveal interesting variations between individual molecules1,2,3. The application of these techniques to the {{RecB}CD} helicase of {{\textit{Escherichia coli}}} has resolved some long-standing discrepancies, and has provided otherwise unattainable mechanistic insights into its enzymatic behaviour4,5,6. Enigmatically, the {DNA} unwinding rates of individual enzyme molecules are seen to vary considerably6,7,8, but the origin of this heterogeneity remains unknown. Here we investigate the physical basis for this behaviour. Although any individual {{RecB}CD} molecule unwound {DNA} at a constant rate for an average of approximately 30,000 steps, we discover that transiently halting a single enzyme–{DNA} complex by depleting Mg2+-ATP could change the subsequent rates of {DNA} unwinding by that enzyme after reintroduction to ligand. The proportion of molecules that changed rate increased exponentially with the duration of the interruption, with a half-life of approximately 1 second, suggesting that a conformational change occurred during the time that the molecule was arrested. The velocity after pausing an individual molecule was any velocity found in the starting distribution of the ensemble. We suggest that substrate binding stabilizes the enzyme in one of many equilibrium conformational sub-states that determine the rate-limiting translocation behaviour of each {{RecB}CD} molecule. Each stabilized sub-state can persist for the duration (approximately 1 minute) of processive unwinding of a {DNA} molecule, comprising tens of thousands of catalytic steps, each of which is much faster than the time needed for the conformational change required to alter kinetic behaviour. This ligand-dependent stabilization of rate-defining conformational sub-states results in seemingly static molecule-to-molecule variation in {{RecB}CD} helicase activity, but in fact reflects one microstate from the equilibrium ensemble that a single molecule manifests during an individual processive translocation event.},
   author = {Bian Liu and Ronald J. Baskin and Stephen C. Kowalczykowski},
   doi = {10.1038/nature12333},
   issn = {1476-4687},
   issue = {7463},
   journal = {Nature 2013 500:7463},
   keywords = {Double,Kinetics,Single,molecule biophysics,strand {DNA} breaks},
   month = {7},
   pages = {482-485},
   pmid = {23851395},
   publisher = {Nature Publishing Group},
   title = {{DNA} unwinding heterogeneity by {{RecB}CD} results from static molecules able to equilibrate},
   volume = {500},
   url = {https://www.nature.com/articles/nature12333},
   year = {2013}
}
@article{Joo2006,
   abstract = {{RecA} and its homologs help maintain genomic integrity through recombination. Using single-molecule fluorescence assays and hidden Markov modeling, we show the most direct evidence that a {RecA} filament grows and shrinks primarily one monomer at a time and only at the extremities. Both ends grow and shrink, contrary to expectation, but a higher binding rate at one end is responsible for directional filament growth. Quantitative rate determination also provides insights into how {RecA} might control {DNA} accessibility in vivo. We find that about five monomers are sufficient for filament nucleation. Although ordinarily single-stranded {DNA} binding protein (SSB) prevents filament nucleation, single {RecA} monomers can easily be added to an existing filament and displace SSB from {DNA} at the rate of filament extension. This supports the proposal for a passive role of {RecA}-loading machineries in SSB removal. © 2006 Elsevier Inc. All rights reserved.},
   author = {Chirlmin Joo and Sean A. McKinney and Muneaki Nakamura and Ivan Rasnik and Sua Myong and Taekjip Ha},
   doi = {10.1016/j.cell.2006.06.042},
   issn = {00928674},
   issue = {3},
   journal = {Cell},
   month = {8},
   pages = {515-527},
   pmid = {16901785},
   publisher = {Elsevier B.V.},
   title = {Real-Time Observation of {RecA} Filament Dynamics with Single Monomer Resolution},
   volume = {126},
   url = {http://www.cell.com/article/S0092867406009470/fulltext http://www.cell.com/article/S0092867406009470/abstract https://www.cell.com/cell/abstract/S0092-8674(06)00947-0},
   year = {2006}
}
@article{Galletto2006,
   abstract = {{{\textit{Escherichia coli}}} {RecA} is essential for the repair of {DNA} double-strand breaks by homologous recombination1. Repair requires the formation of a {RecA} nucleoprotein filament. Previous studies have indicated a mechanism of filament assembly whereby slow nucleation of {RecA} protein on {DNA} is followed by rapid growth2,3,4,5,6,7. However, many aspects of this process remain unclear, including the rates of nucleation and growth and the involvement of ATP hydrolysis, largely because visualization at the single-filament level is lacking. Here we report the direct observation of filament assembly on individual double-stranded {DNA} molecules using fluorescently modified {RecA}. The nucleoprotein filaments saturate the {DNA} and extend it ∼1.6-fold. At early time points, discrete {RecA} clusters are seen, permitting analysis of single-filament growth from individual nuclei. Formation of nascent {RecA} filaments is independent of ATP hydrolysis but is dependent on the type of nucleotide cofactor and the {RecA} concentration, suggesting that nucleation involves binding of ∼4–5 ATP–{RecA} monomers to {DNA}. Individual {RecA} filaments grow at rates of 3–10 nm s-1. Growth is bidirectional and, in contrast to nucleation, independent of nucleotide cofactor, suggesting addition of ∼2–7 monomers s-1. These results are in accord with extensive genetic and biochemical studies, and indicate that assembly in vivo is controlled at the nucleation step. We anticipate that our approach and conclusions can be extended to the related eukaryotic counterpart, Rad51 (see ref.8), and to regulation by assembly mediators9,10,11.},
   author = {Roberto Galletto and Ichiro Amitani and Ronald J. Baskin and Stephen C. Kowalczykowski},
   doi = {10.1038/nature05197},
   issn = {1476-4687},
   issue = {7113},
   journal = {Nature 2006 443:7113},
   keywords = {Humanities and Social Sciences,Science,multidisciplinary},
   month = {9},
   pages = {875-878},
   pmid = {16988658},
   publisher = {Nature Publishing Group},
   title = {Direct observation of individual {RecA} filaments assembling on single {DNA} molecules},
   volume = {443},
   url = {https://www.nature.com/articles/nature05197},
   year = {2006}
}
@article{Handa2009,
   abstract = {Fluorescent fusion proteins are exceedingly useful for monitoring protein localization in situ or visualizing protein behavior at the single molecule level. Unfortunately, some proteins are rendered inactive by the fusion. To circumvent this problem, we fused a hyperactive {RecA} protein ({RecA}803 protein) to monomeric red fluorescent protein (mRFP1) to produce a functional protein ({RecA}-RFP) that is suitable for in vivo and in vitro analysis. In vivo, the {RecA}-RFP partially restores{UV}resistance, conjugational recombination, and {SOS} induction to recA- cells. In vitro, the purified {RecA}-RFP protein forms a nucleoprotein filament whose kcat for single-stranded {DNA}-dependent ATPase activity is reduced ∼3-fold relative to wild-type protein, and which is largely inhibited by single-stranded {DNA}-binding protein. However, {RecA} protein is also a dATPase; dATP supports {RecA}-RFP nucleoprotein filament formation in the presence of single-stranded {DNA}-binding protein. Furthermore, as for the wild-type protein, the activities of {RecA}-RFP are further enhanced by shifting thepHto 6.2. As a consequence, {RecA}-RFP is proficient for {DNA} strand exchange with dATP or at lower pH. Finally, using single molecule visualization, {RecA}-RFP was seen to assemble into a continuous filament on duplex {DNA}, and to extend the {DNA} ∼1.7-fold. Consistent with its attenuated activities, {RecA}-RFP nucleates onto double-stranded {DNA} ∼3-fold more slowly than the wild-type protein, but still requires ∼3 monomers to form the rate-limited nucleus needed for filament assembly. Thus, {RecA}-RFP reveals that its attenuated biological functions correlate with a reduced frequency of nucleoprotein filament nucleation at the single molecule level. © 2009 by The American Society for Biochemistry and Molecular Biology, Inc.},
   author = {Naofumi Handa and Ichiro Amitani and Nathan Gumlaw and Steven J. Sandler and Stephen C. Kowalczykowski},
   doi = {10.1074/jbc.M109.004895},
   issn = {00219258},
   issue = {28},
   journal = {Journal of Biological Chemistry},
   month = {7},
   pages = {18664-18673},
   pmid = {19419960},
   publisher = {Elsevier},
   title = {Single molecule analysis of a red fluorescent {RecA} protein reveals a defect in nucleoprotein filament nucleation that relates to its reduced biological functions},
   volume = {284},
   url = {http://www.jbc.org/article/S0021925819811997/fulltext http://www.jbc.org/article/S0021925819811997/abstract https://www.jbc.org/article/S0021-9258(19)81199-7/abstract},
   year = {2009}
}
@article{Forget2012,
   abstract = {The search for {DNA} homology is vital to recombinational {DNA} repair and occurs by intersegment contact sampling wherein the three-dimensional conformational state of the double-stranded {DNA} target and the length of the homologous {RecA}–single-stranded {DNA} filament have important roles. During {DNA} repair by homologous recombination, the molecule containing a double-strand break must find an undamaged, intact, exact copy of the sequence at the break to initiate a process known as strand exchange. This is facilitated by cooperative binding of a {RecA} family strand-exchange protein to a single-strand tail of the broken {DNA}. But how does the {RecA}–{DNA} filament find its matching sequence, which represents a tiny fraction of the total {DNA} content? In a new study, Anthony Forget and Stephen Kowalczykowski show that weak, transient contacts by the {RecA}–{DNA} filament with duplex {DNA} allow it to sample three-dimensional space to accelerate the recognition of homologous sequence. {DNA} breaks can be repaired with high fidelity by homologous recombination. A ubiquitous protein that is essential for this {DNA} template-directed repair is {RecA}1. After resection of broken {DNA} to produce single-stranded {DNA} (ss{DNA}), {RecA} assembles on this ss{DNA} into a filament with the unique capacity to search and find {DNA} sequences in double-stranded {DNA} (ds{DNA}) that are homologous to the ss{DNA}. This homology search is vital to recombinational {DNA} repair, and results in homologous pairing and exchange of {DNA} strands. Homologous pairing involves {DNA} sequence-specific target location by the {RecA}–ss{DNA} complex. Despite decades of study, the mechanism of this enigmatic search process remains unknown. {RecA} is a {DNA}-dependent ATPase, but ATP hydrolysis is not required for {DNA} pairing and strand exchange2,3, eliminating active search processes. Using dual optical trapping to manipulate {DNA}, and single-molecule fluorescence microscopy to image {DNA} pairing, we demonstrate that both the three-dimensional conformational state of the ds{DNA} target and the length of the homologous {RecA}–ss{DNA} filament have important roles in the homology search. We discovered that as the end-to-end distance of the target ds{DNA} molecule is increased, constraining the available three-dimensional (3D) conformations of the molecule, the rate of homologous pairing decreases. Conversely, when the length of the ss{DNA} in the nucleoprotein filament is increased, homology is found faster. We propose a model for the {DNA} homology search process termed ‘intersegmental contact sampling’, in which the intrinsic multivalent nature of the {RecA} nucleoprotein filament is used to search {DNA} sequence space within 3D domains of {DNA}, exploiting multiple weak contacts to rapidly search for homology. Our findings highlight the importance of the 3D conformational dynamics of {DNA}, reveal a previously unknown facet of the homology search, and provide insight into the mechanism of {DNA} target location by this member of a universal family of proteins.},
   author = {Anthony L. Forget and Stephen C. Kowalczykowski},
   doi = {10.1038/nature10782},
   issn = {1476-4687},
   issue = {7385},
   journal = {Nature 2012 482:7385},
   keywords = {Homologous recombination,Single,Structural biology,molecule biophysics},
   month = {2},
   pages = {423-427},
   pmid = {22318518},
   publisher = {Nature Publishing Group},
   title = {Single-molecule imaging of {DNA} pairing by {RecA} reveals a three-dimensional homology search},
   volume = {482},
   url = {https://www.nature.com/articles/nature10782},
   year = {2012}
}
@article{Ragunathan2012,
   abstract = {During homologous recombination, {RecA} forms a helical filament on a single stranded (ss) {DNA} that searches for a homologous double stranded (ds) {DNA} and catalyzes the exchange of complementary base pairs to form a new heteroduplex. Using single molecule fluorescence imaging tools with high spatiotemporal resolution we characterized the encounter complex between the {RecA} filament and ds{DNA}. We present evidence in support of the 'sliding model' wherein a {RecA} filament diffuses along a ds{DNA} track. We further show that homology can be detected during sliding. Sliding occurs with a diffusion coefficient of approximately 8000 bp2/s allowing the filament to sample several hundred base pairs before dissociation. Modeling suggests that sliding can accelerate homology search by as much as 200 fold. Homology recognition can occur for as few as 6 nt of complementary basepairs with the recognition efficiency increasing for higher complementarity. Our data represents the first example of a {DNA} bound multi-protein complex which can slide along another {DNA} to facilitate target search. © Ragunathan et al.},
   author = {Kaushik Ragunathan and Cheng Liu and Taekjip Ha},
   doi = {10.7554/ELIFE.00067},
   issn = {2050084X},
   issue = {1},
   journal = {eLife},
   month = {12},
   pmid = {23240082},
   publisher = {eLife Sciences Publications Ltd},
   title = {{RecA} filament sliding on {DNA} facilitates homology search},
   volume = {2012},
   year = {2012}
}
@article{Spies2003,
   abstract = {{{RecB}CD} enzyme is a heterotrimeric helicase/nuclease that initiates homologous recombination at double-stranded {DNA} breaks. Several of its activities are regulated by the {DNA} sequence χ (5′-GCTGGTGG-3′), which is recognized in cis by the translocating enzyme. When {{RecB}CD} enzyme encounters χ, the intensity and polarity of its nuclease activity are changed, and the enzyme gains the ability to load {RecA} protein onto the χ-containing, unwound single-stranded {DNA}. Here, we show that interaction with χ also affects translocation by {{RecB}CD} enzyme. By observing translocation of individual enzymes along single molecules of {DNA}, we could see {{RecB}CD} enzyme pause precisely at χ. Furthermore, and more unexpectedly, after pausing at χ, the enzyme continues translocating but at approximately one-half the initial rate. We propose that interaction with χ results in an enzyme in which one of the two motor subunits, likely the RecD motor, is uncoupled from the holoenzyme to produce the slower translocase.},
   author = {Maria Spies and Piero R. Bianco and Mark S. Dillingham and Naofumi Handa and Ronald J. Baskin and Stephen C. Kowalczykowski},
   doi = {10.1016/S0092-8674(03)00681-0},
   issn = {0092-8674},
   issue = {5},
   journal = {Cell},
   month = {9},
   pages = {647-654},
   pmid = {13678587},
   publisher = {Cell Press},
   title = {A Molecular Throttle: The Recombination Hotspot χ Controls {DNA} Translocation by the {{RecB}CD} Helicase},
   volume = {114},
   year = {2003}
}
@article{Ghodke2019,
   abstract = {The {RecA} protein orchestrates the cellular response to {DNA} damage via its multiple roles in the bacterial {SOS} response. Lack of tools that provide unambiguous access to the various {RecA} states within the cell have prevented understanding of the spatial and temporal changes in {RecA} structure/function that underlie control of the damage response. Here, we develop a monomeric C-terminal fragment of the l repressor as a novel fluorescent probe that specifically interacts with {RecA} filaments on single-stranded {DNA} ({RecA}*). Single-molecule imaging techniques in live cells demonstrate that {RecA} is largely sequestered in storage structures during normal metabolism. Upon {DNA} damage, the storage structures dissolve and the cytosolic pool of {RecA} rapidly nucleates to form early {SOS}-signaling complexes, maturing into {DNA}-bound {RecA} bundles at later time points. Both before and after {SOS} induction, {RecA}* largely appears at locations distal from replisomes. Upon completion of repair, {RecA} storage structures reform.},
   author = {Harshad Ghodke and Bishnu P. Paudel and Jacob S. Lewis and Slobodan Jergic and Kamya Gopal and Zachary J. Romero and Elizabeth A. Wood and Roger Woodgate and Michael M. Cox and Antoine M.Van Oijen},
   doi = {10.7554/ELIFE.42761},
   issn = {2050084X},
   journal = {eLife},
   month = {2},
   pmid = {30719973},
   publisher = {eLife Sciences Publications Ltd},
   title = {Spatial and temporal organization of reca in the escherichia coli dna-damage response},
   volume = {8},
   year = {2019}
}
@article{Renzette2007,
   abstract = {{RecA} plays a central role in recombination, {DNA} repair and {SOS} induction through forming a {RecA}-{DNA} helical filament. Biochemical observations show that at low ratios to {RecA}, {DinI} and {RecX} stabilize and destabilize {RecA}-{DNA} filaments, respectively, and that the C-terminal 17 residues of {RecA} are important for {RecX} function. {RecA}-{DNA} filament formation was assayed in vivo using {RecA}-{GFP} foci formation in log-phase and {UV}-irradiated cells. In log-phase cells, dinI mutants have fewer foci than wild type and that recX mutants have more foci than wild type. A recA Δ17::gfp mutant had more foci like a recX mutant. dinI recX double mutants have the same number of foci as dinI mutants alone, suggesting that dinI is epistatic to recX. After {UV} treatment, the dinI, recX and dinI recX mutants differed in their ability to form foci. All three mutants had fewer foci than wild type. The dinI mutant's foci persisted longer than wild-type foci. Roles of {DinI} and {RecX} after {UV} treatment differed from those during log-phase growth and may reflect the different {DNA} substrates, population of proteins or amounts during the {SOS} response. These experiments give new insight into the roles of these proteins. © 2006 The Authors.},
   author = {Nicholas Renzette and Nathan Gumlaw and Steven J. Sandler},
   doi = {10.1111/J.1365-2958.2006.05496.X},
   isbn = {4135451578},
   issn = {1365-2958},
   issue = {1},
   journal = {Molecular Microbiology},
   month = {1},
   pages = {103-115},
   pmid = {17163974},
   publisher = {John Wiley \& Sons, Ltd},
   title = {{DinI} and {RecX} modulate {RecA}–{DNA} structures in {{\textit{Escherichia coli}}} {K-12}},
   volume = {63},
   url = {https://onlinelibrary.wiley.com/doi/full/10.1111/j.1365-2958.2006.05496.x https://onlinelibrary.wiley.com/doi/abs/10.1111/j.1365-2958.2006.05496.x https://onlinelibrary.wiley.com/doi/10.1111/j.1365-2958.2006.05496.x},
   year = {2007}
}
@article{Klimova2020,
   abstract = {{RecA} is essential for double-strand-break repair (DSBR) and the {SOS} response in {{\textit{Escherichia coli}}} {K-12}. {RecN} is an {SOS} protein and a member of the Structural Maintenance of Chromosomes family of proteins thought to play a role in sister chromatid cohesion/interactions during DSBR. Previous studies have shown that a plasmid-encoded recA4190 (Q300R) mutant had a phenotype similar to DrecN (mitomycin C sensitive and {UV} resistant). It was hypothesized that {RecN} and {RecA} physically interact, and that recA4190 specifically eliminated this interaction. To test this model, an epistasis analysis between recA4190 and DrecN was performed in wild-type and recBC sbcBC cells. To do this, recA4190 was first transferred to the chromosome. As single mutants, recA4190 and DrecN were Rec+ as measured by transductional recombination, but were 3-fold and 10-fold decreased in their ability to do I-SceIinduced DSBR, respectively. In both cases, the double mutant had an additive phenotype relative to either single mutant. In the recBC sbcBC background, recA4190 and DrecN cells were very {UV}S (sensitive), Rec2, had high basal levels of {SOS} expression and an altered distribution of {RecA}-{GFP} structures. In all cases, the double mutant had additive phenotypes. These data suggest that recA4190 (Q300R) and DrecN remove functions in genetically distinct pathways important for {DNA} repair, and that {RecA} Q300 was not important for an interaction between {RecN} and {RecA} in vivo. recA4190 (Q300R) revealed modest phenotypes in a wild-type background and dramatic phenotypes in a recBC sbcBC strain, reflecting greater stringency of {RecA}'s role in that background.},
   author = {Anastasiia N. Klimova and Steven J. Sandler},
   doi = {10.1534/GENETICS.120.303476},
   issn = {19432631},
   issue = {2},
   journal = {Genetics},
   keywords = {Bacteria,{DNA} repair,Homologous recombination,{SOS} response},
   month = {10},
   pages = {381-393},
   pmid = {32816866},
   publisher = {Oxford Academic},
   title = {An Epistasis Analysis of recA and recN in {{\textit{Escherichia coli}}} {K-12}},
   volume = {216},
   url = {https://dx.doi.org/10.1534/genetics.120.303476},
   year = {2020}
}
@article{Lepore2023,
   abstract = {Efficient {DNA} repair is crucial for maintaining genome integrity and ensuring cell survival. In {{\textit{Escherichia coli}}} , {{RecB}CD} plays a crucial role in processing {DNA} ends following a {DNA} double-strand break (DSB) to initiate repair by homologous recombination. While {{RecB}CD} has been extensively studied in vitro , less is known about how it contributes to rapid and efficient repair in living bacteria. Here, we perform single-molecule microscopy to investigate {DNA} repair in real-time in {{\textit{E. coli}}} . We quantify {RecB} single-molecule mobility and monitor the induction of the {DNA} damage response ({SOS} response) in individual cells. We show that {RecB} binding to broken {DNA} ends leads to efficient repair without {SOS} induction. In contrast, in a {RecB} mutant with modified activities leading to the activation of an alternative repair pathway, repair is less efficient and leads to high {SOS} induction. Our findings reveal how subtle alterations in {RecB} activity profoundly impact the efficiency of {DNA} repair in {{\textit{E. coli}}} .

### Competing Interest Statement

The authors have declared no competing interest.},
   author = {Alessia Lepore and Daniel Thédié and Lorna McLaren and Benura Azeroglu and Oliver J. Pambos and Achillefs N. Kapanidis and Meriem El Karoui},
   doi = {10.1101/2023.12.22.573010},
   isbn = {10.1101/2023.12.2},
   journal = {bioRxiv},
   month = {12},
   pages = {2023.12.22.573010},
   publisher = {Cold Spring Harbor Laboratory},
   title = {In vivo single-molecule imaging of {RecB} reveals efficient repair of {DNA} damage in {{\textit{Escherichia coli}}}},
   url = {https://www.biorxiv.org/content/10.1101/2023.12.22.573010v1 https://www.biorxiv.org/content/10.1101/2023.12.22.573010v1.abstract},
   year = {2023}
}
@article{Baharoglu2014,
   abstract = {The presence of an abnormal amount of single-stranded {DNA} in the bacterial cell constitutes a genotoxic alarm signal that induces the {SOS} response, a broad regulatory network found in most bacterial species to address {DNA} damage. The aim of this review was to point out that beyond being a repair process, {SOS} induction leads to a very strong but transient response to genotoxic stress, during which bacteria can rearrange and mutate their genome, induce several phenotypic changes through differential regulation of genes, and sometimes acquire characteristics that potentiate bacterial survival and adaptation to changing environments. We review here the causes and consequences of {SOS} induction, but also how this response can be modulated under various circumstances and how it is connected to the network of other important stress responses. In the first section, we review articles describing the induction of the {SOS} response at the molecular level. The second section discusses consequences of this induction in terms of {DNA} repair, changes in the genome and gene expression, and sharing of genomic information, with their effects on the bacteria's life and evolution. The third section is about the fine tuning of this response to fit with the bacteria's 'needs'. Finally, we discuss recent findings linking the {SOS} response to other stress responses. Under these perspectives, {SOS} can be perceived as a powerful bacterial strategy against aggressions.},
   author = {Zeynep Baharoglu and Didier Mazel},
   doi = {10.1111/1574-6976.12077},
   issn = {0168-6445},
   issue = {6},
   journal = {FEMS Microbiology Reviews},
   keywords = {Antibiotic resistance,Bacteria,{DNA} repair,{SOS},Stress,bacteria},
   month = {11},
   pages = {1126-1145},
   pmid = {24923554},
   publisher = {Oxford Academic},
   title = {{SOS}, the formidable strategy of bacteria against aggressions},
   volume = {38},
   url = {https://dx.doi.org/10.1111/1574-6976.12077},
   year = {2014}
}
@article{Bos2015,
   abstract = {Bacteria can rapidly evolve resistance to antibiotics via the {SOS} response, a state of high-activity {DNA} repair and mutagenesis. We explore here the first steps of this evolution in the bacterium {{\textit{Escherichia coli}}}. Induction of the {SOS} response by the genotoxic antibiotic ciprofloxacin changes the {{\textit{E. coli}}} rod shape into multichromosome-containing filaments. We show that at subminimal inhibitory concentrations of ciprofloxacin the bacterial filament divides asymmetrically repeatedly at the tip. Chromosome-containing buds are made that, if resistant, propagate nonfilamenting progeny with enhanced resistance to ciprofloxacin as the parent filament dies. We propose that the multinucleated filament creates an environmental niche where evolution can proceed via generation of improved mutant chromosomes due to the mutagenic {SOS} response and possible recombination of the new alleles between chromosomes. Our data provide a better understanding of the processes underlying the origin of resistance at the single-cell level and suggest an analogous role to the eukaryotic aneuploidy condition in cancer.},
   author = {Julia Bos and Qiucen Zhang and Saurabh Vyawahare and Elizabeth Rogers and Susan M. Rosenberg and Robert H. Austin},
   issn = {10916490},
   issue = {1},
   journal = {Proceedings of the National Academy of Sciences of the United States of America},
   keywords = {Antibiotic resistance,Evolution,Filamentation,Mutation,Sos response},
   month = {1},
   pages = {178-183},
   pmid = {25492931},
   publisher = {National Academy of Sciences},
   title = {Emergence of antibiotic resistance from multinucleated bacterial filaments},
   volume = {112},
   url = {https://www.pnas.org/doi/abs/10.1073/pnas.1420702111},
   year = {2015}
}
@article{Otsu1979,
   author = {N Otsu},
   doi = {10.1371/JOURNAL.PONE.0035550},
   journal = {IEEE Trans SMC},
   pages = {62-},
   title = {A threshold selection method from gray-level histograms.},
   volume = {9},
   url = {https://cir.nii.ac.jp/crid/1370567187454221059},
   year = {1979}
}
@article{Ollion2021,
   abstract = {We propose a novel self-supervised image blind denoising approach in which two neural networks jointly predict the clean signal and infer the noise distribution. Assuming that the noisy observations are independent conditionally to the signal, the networks can be jointly trained without clean training data. Therefore, our approach is particularly relevant for biomedical image denoising where the noise is difficult to model precisely and clean training data are usually unavailable. Our method significantly outperforms current state-of-the-art self-supervised blind denoising algorithms, on six publicly available biomedical image datasets. We also show empirically with synthetic noisy data that our model captures the noise distribution efficiently. Finally, the described framework is simple, lightweight and computationally efficient, making it useful in practical cases.},
   author = {Jean Ollion and Charles Ollion and Elisabeth Gassiat and Luc Lehéricy and Sylvain Le Corff and Laboratoire J A Dieudonné},
   journal = {arxiv},
   month = {2},
   title = {Joint self-supervised blind denoising and noise estimation},
   url = {https://arxiv.org/abs/2102.08023v1},
   year = {2021}
}
@article{Kobayashi2020,
   abstract = {We propose a general framework for solving inverse problems in the presence of noise that requires no signal prior, no noise estimate, and no clean training data. We only require that the forward model be available and that the noise be statistically independent across measurement dimensions. We build upon the theory of "J-invariant" functions (Batson & Royer, 2019) and show how self-supervised denoising la Noise2Self is a special case of learning a noise-tolerant pseudo-inverse of the identity. We demonstrate our approach by showing how a convolutional neural network can be taught in a self-supervised manner to deconvolve images and surpass in image quality classical inversion schemes such as Lucy-Richardson deconvolution.},
   author = {Hirofumi Kobayashi and Ahmet Can Solak and Joshua Batson and Loic A Royer},
   journal = {arxiv},
   title = {Image Deconvolution via Noise-Tolerant Self-Supervised Inversion},
   year = {2020}
}
@article{Ollion2019,
   abstract = {The analysis of bacteria at the single-cell level is essential to characterization of processes in which cellular heterogeneity plays an important role. BACMMAN (bacteria mother machine analysis) is a software allowing fast and reliable automated image analysis of high-throughput 2D or 3D time-series images from experiments using the ‘mother machine’, a very popular microfluidic device allowing biological processes in bacteria to be investigated at the single-cell level. Here, we describe how to use some of the BACMMAN features, including (i) segmentation and tracking of bacteria and intracellular fluorescent spots, (ii) visualization and editing of the results, (iii) configuration of the image-processing pipeline for different datasets and (iv) BACMMAN coupling to data analysis software for visualization and analysis of data subsets with specific properties. Among software specifically dedicated to the analysis of mother machine data, only BACMMAN allows segmentation and tracking of both bacteria and intracellular spots. For a single position, single channel with 1,000 frames (2-GB dataset), image processing takes ~6 min on a regular computer. Numerous implemented algorithms, easy configuration and high modularity ensure wide applicability of the BACMMAN software. BACMMAN software is used to automate image analysis of high-throughput 2D or 3D time-series images from experiments using the ‘mother machine’, a microfluidic device that allows growth and division of single bacterial cells to be followed.},
   author = {Jean Ollion and Marina Elez and Lydia Robert},
   doi = {10.1038/s41596-019-0216-9},
   issn = {1750-2799},
   issue = {11},
   journal = {Nature Protocols 2019 14:11},
   keywords = {Bioinformatics,Image processing,Software},
   month = {9},
   pages = {3144-3161},
   pmid = {31554957},
   publisher = {Nature Publishing Group},
   title = {High-throughput detection and tracking of cells and intracellular spots in mother machine experiments},
   volume = {14},
   url = {https://www.nature.com/articles/s41596-019-0216-9},
   year = {2019}
}
@article{Wertman1986,
   abstract = {A class of recombinant phage λ clones are recovered from human genomic libraries on {{\textit{Escherichia coli}}} recB21 recC22 sbcB15 cells, which fail to form plaques on wild-type cells. We report experiments which address the mechanism of this inhibition. The introduction of the recombination-stimulating sequence chi into one such clone allows growth of this phage on Rec+ cells. In addition, the insertion of λ gam+ gene into a rec+ -inhibited clone results in the ability of the phage to form plaques on wild-type cells. Since λ Gam protein is an inhibitor of host {RecB}C enzyme, we tested a collection of such phage for growth on a variety of hosts altered in {RecB}C function. Host permissiveness correlated with the inactivation of the {RecB}C nucleolytic activities and not with the recombinational activities. These observations suggest that the inserted {DNA} sequences of these phage limit the production of packageable chromosomes. This conclusion is easily reconciled with our current knowledge of the interaction of the host recombination systems with λ replication and encapsidation. Based on these experiments we have constructed strains, both recombination-proficient and recombination-deficient, which serve as improved hosts for the recovery of genomic sequences which are otherwise inhibitory to the growth of phage L. © 1986.},
   author = {Kenneth F. Wertman and Arlene R. Wyman and David Botstein},
   doi = {10.1016/0378-1119(86)90286-6},
   issn = {0378-1119},
   issue = {2},
   journal = {Gene},
   keywords = {{RecB}C enzyme,Recombinant {DNA},Spi- phenotype,chi site,genomic libraries,palindromes,rec+ -inhibited phenotype,recD},
   month = {1},
   pages = {253-262},
   pmid = {2952553},
   publisher = {Elsevier},
   title = {Host/vector interactions which affect the viability of recombinant phage lambda clones},
   volume = {49},
   year = {1986}
}
@article{Capaldo1975,
   abstract = {Using a method to separate dividing from non-dividing cells of Rec- strains of {{\textit{Escherichia coli}}} K12 (Capaldo & Barbour, 1973), we found: 1. (1) Non-dividing cells of recA-, recB-recC- and recA-recB-recC- strains are defective in {DNA} synthesis. 2. (2) recA- non-dividing cells contain little or no {DNA}; recB-recC- and recA-recB-recC- non-dividing cells contain normal amounts of {DNA}. 3. (3) {DNA} synthesized in cultures of the recB-recC- and recA-recB-recC- strains is made in the dividing cells and then segregated irreversibly, as a function of subsequent cell division, into the non-dividing cells. 4. (4) Old {DNA} in recA-recB-recC- cells contains an increased number of single-strand breaks compared to newly synthesized {DNA} of the same strain or to both old and new {DNA} from rec+ cells; no additional breaks in old {DNA} of the recB-recC- strain have been detected. We propose that recA+, recB+, recC+ gene products are involved in repair of {DNA} chain breaks that arise during cell growth. According to this view, mutation in the rec genes results in defective repair. This may lead to failure to replicate {DNA}, loss of viability, degradation of the {DNA} in the case of the recA- strain, and accumulation of {DNA} chain breaks in the recA-recB-recC- strain. © 1978.},
   author = {Florence N. Capaldo and Stephen D. Barbour},
   doi = {10.1016/0022-2836(75)90371-X},
   issn = {0022-2836},
   issue = {1},
   journal = {Journal of Molecular Biology},
   month = {1},
   pages = {53-66},
   pmid = {1102696},
   publisher = {Academic Press},
   title = {{DNA} content, synthesis and integrity in dividing and non-dividing cells of rec- strains of {{\textit{Escherichia coli}}} K12},
   volume = {91},
   year = {1975}
}
@article{Skarstad1993,
   abstract = {Rapidly growing wild-type {{\textit{Escherichia coli}}} cells contain two, four, or eight fully replicated chromosomes after treatment with rifampin, reflecting that all replication origins are initiated simult...},
   author = {K. Skarstad and E. Boye},
   doi = {10.1128/JB.175.17.5505-5509.1993},
   issn = {00219193},
   issue = {17},
   journal = {Journal of Bacteriology},
   pages = {5505-5509},
   pmid = {8366035},
   title = {Degradation of individual chromosomes in recA mutants of {{\textit{Escherichia coli}}}},
   volume = {175},
   url = {https://journals.asm.org/doi/10.1128/jb.175.17.5505-5509.1993},
   year = {1993}
}
@article{Cockram2015,
   abstract = {Understanding molecular mechanisms in the context of living cells requires the development of new methods of in vivo biochemical analysis to complement established in vitro biochemistry. A critically important molecular mechanism is genetic recombination, required for the beneficial reassortment of genetic information and for {DNA} double-strand break repair (DSBR). Central to recombination is the {RecA} (Rad51) protein that assembles into a spiral filament on {DNA} and mediates genetic exchange. Here we have developed a method that combines chromatin immunoprecipitation with next-generation sequencing (ChIP-Seq) and mathematical modeling to quantify {RecA} protein binding during the active repair of a single DSB in the chromosome of {{\textit{Escherichia coli}}}. We have used quantitative genomic analysis to infer the key in vivo molecular parameters governing {RecA} loading by the helicase/ nuclease {{RecB}CD} at recombination hot-spots, known as Chi. Our genomic analysis has also revealed that DSBR at the lacZ locus causes a second {{RecB}CD}-mediated DSBR event to occur in the terminus region of the chromosome, over 1 Mb away.},
   author = {Charlotte A. Cockram and Milana Filatenkova and Vincent Danos and Meriem El Karoui and David R.F. Leach},
   doi = {10.1073/pnas.1424269112},
   issn = {10916490},
   issue = {34},
   journal = {Proceedings of the National Academy of Sciences of the United States of America},
   keywords = {{DNA} repair,Homologous recombination,Mechanistic modelling,{RecA},{{RecB}CD}},
   month = {8},
   pages = {E4735-E4742},
   pmid = {26261330},
   publisher = {National Academy of Sciences},
   title = {Quantitative genomic analysis of {RecA} protein binding during {DNA} double-strand break repair reveals {{RecB}CD} action in vivo},
   volume = {112},
   url = {https://www.pnas.org/doi/abs/10.1073/pnas.1424269112},
   year = {2015}
}
@article{Thedie2017,
   abstract = {Green-to-red photoconvertible fluorescent proteins ({PCFP}s) such as {mEos2} and its derivatives are widely used in PhotoActivated Localization Microscopy (PALM). However, the complex photophysics of these genetically encoded markers complicates the quantitative analysis of PALM data. Here, we show that intense 561 nm light (∼1 kW/cm2) typically used to localize single red molecules considerably affects the green-state photophysics of {mEos2} by populating at least two reversible dark states. These dark states retard green-to-red photoconversion through a shelving effect, although one of them is rapidly depopulated by 405 nm light illumination. Multiple {mEos2} switching and irreversible photobleaching is thus induced by yellow/green and violet photons before green-to-red photoconversion occurs, contributing to explain the apparent limited signaling efficiency of this {PCFP}. Our data reveals that the photophysics of {PCFP}s of anthozoan origin is substantially more complex than previously thought, and suggests that intense 561 nm laser light should be used with care, notably for quantitative or fast PALM approaches.},
   author = {Daniel Thédié and Romain Berardozzi and Virgile Adam and Dominique Bourgeois},
   issn = {19487185},
   issue = {18},
   journal = {Journal of Physical Chemistry Letters},
   month = {9},
   pages = {4424-4430},
   pmid = {28850784},
   publisher = {American Chemical Society},
   title = {Photoswitching of Green {mEos2} by Intense 561 nm Light Perturbs Efficient Green-to-Red Photoconversion in Localization Microscopy},
   volume = {8},
   url = {https://pubs.acs.org/doi/full/10.1021/acs.jpclett.7b01701},
   year = {2017}
}
@article{Grimm2015,
   abstract = {A simple and general chemical structure change to a panel of cell-permeable small-molecule fluorophores increases their brightness and photostability, which will enable improved single-molecule studies and super-resolution imaging. Specific labeling of biomolecules with bright fluorophores is the keystone of fluorescence microscopy. Genetically encoded self-labeling tag proteins can be coupled to synthetic dyes inside living cells, resulting in brighter reporters than fluorescent proteins. Intracellular labeling using these techniques requires cell-permeable fluorescent ligands, however, limiting utility to a small number of classic fluorophores. Here we describe a simple structural modification that improves the brightness and photostability of dyes while preserving spectral properties and cell permeability. Inspired by molecular modeling, we replaced the N,N-dimethylamino substituents in tetramethylrhodamine with four-membered azetidine rings. This addition of two carbon atoms doubles the quantum efficiency and improves the photon yield of the dye in applications ranging from in vitro single-molecule measurements to super-resolution imaging. The novel substitution is generalizable, yielding a palette of chemical dyes with improved quantum efficiencies that spans the {UV} and visible range.},
   author = {Jonathan B. Grimm and Brian P. English and Jiji Chen and Joel P. Slaughter and Zhengjian Zhang and Andrey Revyakin and Ronak Patel and John J. Macklin and Davide Normanno and Robert H. Singer and Timothée Lionnet and Luke D. Lavis},
   doi = {10.1038/nmeth.3256},
   issn = {1548-7105},
   issue = {3},
   journal = {Nature Methods 2015 12:3},
   keywords = {Cellular imaging,Chemical synthesis,Chemical tools,Fluorescent dyes},
   month = {1},
   pages = {244-250},
   pmid = {25599551},
   publisher = {Nature Publishing Group},
   title = {A general method to improve fluorophores for live-cell and single-molecule microscopy},
   volume = {12},
   url = {https://www.nature.com/articles/nmeth.3256},
   year = {2015}
}
@article{Vickridge2017,
   abstract = {Aberrant {DNA} replication is a major source of the mutations and chromosomal rearrangements associated with pathological disorders. In bacteria, several different {DNA} lesions are repaired by homologous recombination, a process that involves sister chromatid pairing. Previous work in {{\textit{Escherichia coli}}} has demonstrated that sister chromatid interactions (SCIs) mediated by topological links termed precatenanes, are controlled by topoisomerase IV. In the present work, we demonstrate that during the repair of mitomycin C-induced lesions, topological links are rapidly substituted by an {SOS}-induced sister chromatid cohesion process involving the {RecN} protein. The loss of SCIs and viability defects observed in the absence of {RecN} were compensated by alterations in topoisomerase IV, suggesting that the main role of {RecN} during {DNA} repair is to promote contacts between sister chromatids. {RecN} also modulates whole chromosome organization and {RecA} dynamics suggesting that SCIs significantly contribute to the repair of {DNA} double-strand breaks (DSBs). Homologous recombination of {DNA} lesions in bacteria involves sister chromatid pairing. Here, the authors show that {RecN} promotes contacts between sister chromatids and facilitates repair.},
   author = {Elise Vickridge and Charlene Planchenault and Charlotte Cockram and Isabel Garcia Junceda and Olivier Espéli},
   doi = {10.1038/ncomms14618},
   issn = {2041-1723},
   issue = {1},
   journal = {Nature Communications 2017 8:1},
   keywords = {Bacterial genetics,Double,strand {DNA} breaks},
   month = {3},
   pages = {1-12},
   pmid = {28262707},
   publisher = {Nature Publishing Group},
   title = {Management of {{\textit{E. coli}}} sister chromatid cohesion in response to genotoxic stress},
   volume = {8},
   url = {https://www.nature.com/articles/ncomms14618},
   year = {2017}
}
@article{Keyamura2019,
   abstract = {Bacterial {RecN}, closely related to the structural maintenance of chromosomes (SMC) family of proteins, functions in the repair of {DNA} double-strand breaks (DSBs) by homologous recombination. Here we show that the purified {{\textit{Escherichia coli}}} {RecN} protein topologically loads onto both single-stranded {DNA} (ss{DNA}) and double-stranded {DNA} (ds{DNA}) that has a preference for ss{DNA}. {RecN} topologically bound to ds{DNA} slides off the end of linear ds{DNA}, but this is prevented by {RecA} nucleoprotein filaments on ss{DNA}, thereby allowing {RecN} to translocate to DSBs. Furthermore, we found that, once {RecN} is recruited onto ss{DNA}, it can topologically capture a second ds{DNA} substrate in an ATP-dependent manner, suggesting a role in synapsis. Indeed, {RecN} stimulates {RecA}-mediated D-loop formation and subsequent strand exchange activities. Our findings provide mechanistic insights into the recruitment of {RecN} to DSBs and sister chromatid interactions by {RecN}, both of which function in {RecA}-mediated DSB repair. Kenji Keyamura and Takashi Hishida demonstrate that {RecN} binds to single-stranded and double-stranded {DNA} through topological entrapment and stimulates {RecA}-mediated strand exchange activity in bacteria. The results provide mechanistic insights into how {RecN} is recruited to and repairs {DNA} double-strand breaks.},
   author = {Kenji Keyamura and Takashi Hishida},
   doi = {10.1038/s42003-019-0655-4},
   issn = {2399-3642},
   issue = {1},
   journal = {Communications Biology 2019 2:1},
   keywords = {{DNA},{DNA} recombination,Double,strand {DNA} breaks},
   month = {11},
   pages = {1-12},
   pmid = {31754643},
   publisher = {Nature Publishing Group},
   title = {Topological {DNA}-binding of structural maintenance of chromosomes-like {RecN} promotes {DNA} double-strand break repair in {{\textit{Escherichia coli}}}},
   volume = {2},
   url = {https://www.nature.com/articles/s42003-019-0655-4},
   year = {2019}
}
@article{Wyman2006,
   abstract = {Breaks in both {DNA} strands are a particularly dangerous threat to genome stability. At a {DNA} double-strand break (DSB), potentially lost sequence information cannot be recovered from the same {DNA} molecule. However, simple repair by joining two broken ends, though inherently error prone, is preferable to leaving ends broken and capable of causing genome rearrangements. To avoid DSB-induced genetic disinformation and disruption of vital processes, such as replication and transcription, cells possess robust mechanisms to repair DSBs. Because all breaks are not created equal, the particular repair mechanism used depends largely on what is possible and needed based on the structure of the broken {DNA}. We argue that although categorizing different DSB repair mechanisms along pathways and subpathways can be conceptually useful, in cells flexible and reversible interactions among DSB repair factors form a web from which a nonpredetermined path to repair for any number of different {DNA} breaks will emerge. Copyright © 2006 by Annual Reviews. All rights reserved.},
   author = {Claire Wyman and Roland Kanaar},
   doi = {10.1146/annurev.genet.40.110405.090451},
   isbn = {0824312406},
   issn = {00664197},
   issue = {Volume 40, 2006},
   journal = {Annual Review of Genetics},
   keywords = {{DNA} damage,Genome (in)stability,Homologous recombination,Nonhomologous {DNA} end-joining,Stalled {DNA} replication},
   month = {12},
   pages = {363-383},
   pmid = {16895466},
   publisher = {Annual Reviews},
   title = {{DNA} double-strand break repair: All's well that ends well},
   volume = {40},
   url = {https://www.annualreviews.org/content/journals/10.1146/annurev.genet.40.110405.090451},
   year = {2006}
}
@article{Wyman2004,
   abstract = {Exchange of strands between homologous {DNA} molecules is catalyzed by evolutionarily conserved recombinases. These proteins can occur in different quaternary arrangements: rings or helical filaments. Recent results reveal that recombinase function follows from the filamentous form.},
   author = {Claire Wyman and Roland Kanaar},
   doi = {10.1016/j.cub.2004.07.049},
   issn = {09609822},
   issue = {15},
   journal = {Current Biology},
   month = {8},
   pages = {R629-R631},
   pmid = {15296782},
   publisher = {Cell Press},
   title = {Homologous recombination: Down to the wire},
   volume = {14},
   url = {http://www.cell.com/article/S0960982204005536/fulltext http://www.cell.com/article/S0960982204005536/abstract https://www.cell.com/current-biology/abstract/S0960-9822(04)00553-6},
   year = {2004}
}
@article{Ojkic2020,
   abstract = {Fluoroquinolones, antibiotics that cause {DNA} damage by inhibiting {DNA} topoisomerases, are clinically important, but their mechanism of action is not yet fully understood. In particular, the dynamical response of bacterial cells to fluoroquinolone exposure has hardly been investigated, although the {SOS} response, triggered by {DNA} damage, is often thought to play a key role. Here, we investigated the growth inhibition of the bacterium {{\textit{Escherichia coli}}} by the fluoroquinolone ciprofloxacin at low concentrations. We measured the long-term and short-term dynamical response of the growth rate and {DNA} production rate to ciprofloxacin at both the population and single-cell levels. We show that, despite the molecular complexity of {DNA} metabolism, a simple roadblock-and-kill model focusing on replication fork blockage and {DNA} damage by ciprofloxacin-poisoned {DNA} topoisomerase II (gyrase) quantitatively reproduces long-term growth rates in the presence of ciprofloxacin. The model also predicts dynamical changes in the {DNA} production rate in wild-type {{\textit{E. coli}}} and in a recombination-deficient mutant following a step-up of ciprofloxacin. Our work highlights that bacterial cells show a delayed growth rate response following fluoroquinolone exposure. Most importantly, our model explains why the response is delayed: it takes many doubling times to fragment the {DNA} sufficiently to inhibit gene expression. We also show that the dynamical response is controlled by the timescale of {DNA} replication and gyrase binding/unbinding to the {DNA} rather than by the {SOS} response, challenging the accepted view. Our work highlights the importance of including detailed biophysical processes in biochemicalsystems models to quantitatively predict the bacterial response to antibiotics.},
   author = {Nikola Ojkic and Elin Lilja and Susana Direito and Angela Dawson and Rosalind J. Allen and Bartlomiej Waclaw},
   doi = {10.1128/AAC.02487-19/SUPPL_FILE/AAC.02487-19-S0001.PDF},
   issn = {10986596},
   issue = {9},
   journal = {Antimicrobial Agents and Chemotherapy},
   keywords = {Antibiotics,Computer modeling,Fluoroquinolones,Mechanisms of action},
   month = {9},
   pmid = {32601161},
   publisher = {American Society for Microbiology},
   title = {A roadblock-and-kill mechanism of action model for the {DNA}-targeting antibiotic ciprofloxacin},
   volume = {64},
   url = {https://journals.asm.org/doi/10.1128/aac.02487-19},
   year = {2020}
}
@article{Zahradka2009,
   abstract = {Exponentially growing recA mutant cells of {{\textit{Escherichia coli}}} display pronounced {DNA} degradation that starts at the sites of {DNA} damage and depends on {{RecB}CD} nuclease ({ExoV}) activity. As a consequence of this "reckless" {DNA} degradation, populations of recA mutants contain a large proportion of anucleate cells. We have found that both {DNA} degradation and anucleate-cell production are efficiently suppressed by mutations in the xonA (sbcB) and sbcD genes. The suppressive effects of these mutations were observed in normally grown, as well as in {UV}-irradiated, recA cells. The products of the xonA and sbcD genes are known to code for the Exol and {SbcCD} nucleases, respectively. Since both xonA and sbcD mutations are required for strong suppression of {DNA} degradation while individual mutations have only a weak suppressive effect, we infer that Exol and {SbcCD} play partially redundant roles in regulating {DNA} degradation in recA cells. We suggest that their roles might be in processing (blunting) {DNA} ends, thereby producing suitable substrates for {{RecB}CD} binding. Copyright © 2009, American Society for Microbiology. All Rights Reserved.},
   author = {Ksenija Zahradka and Maja Buljubašić and Mirjana Petranović and Davor Zahradka},
   doi = {10.1128/jb.01877-07},
   issn = {00219193},
   issue = {5},
   journal = {Journal of Bacteriology},
   month = {3},
   pages = {1677-1687},
   pmid = {19074388},
   publisher = {American Society for Microbiology},
   title = {Roles of {ExoI} and {SbcCD} nucleases in "Reckless" {DNA} degradation in recA mutants of {{\textit{Escherichia coli}}}},
   volume = {191},
   url = {https://journals.asm.org/doi/10.1128/jb.01877-07},
   year = {2009}
}
@article{Shee2013,
   abstract = {Spontaneous {DNA} breaks instigate genomic changes that fuel cancer and evolution, yet direct quantification of double-strand breaks (DSBs) has been limited. Predominant sources of spontaneous DSBs remain elusive. We report synthetic technology for quantifying DSBs using fluorescent-protein fusions of double-strand {DNA} end-binding protein, Gam of bacteriophage Mu. In {{\textit{Escherichia coli}}} Gam{GFP} forms foci at chromosomal DSBs and pinpoints their subgenomic locations. Spontaneous DSBs occur mostly one per cell, and correspond with generations, supporting replicative models for spontaneous breakage, and providing the first true breakage rates. In mammalian cells Gam{GFP}-labels laser-induced DSBs antagonized by end-binding protein Ku; co-localizes incompletely with DSB marker 53BP1 suggesting superior DSB-specificity; blocks resection; and demonstrates {DNA} breakage via APOBEC3A cytosine deaminase. We demonstrate directly that some spontaneous DSBs occur outside of S phase. The data illuminate spontaneous {DNA} breakage in {{\textit{E. coli}}} and human cells and illustrate the versatility of fluorescent-Gam for interrogation of DSBs in living cells. © Shee et al.},
   author = {Chandan Shee and Ben D. Cox and Franklin Gu and Elizabeth M. Luengas and Mohan C. Joshi and Li Ya Chiu and David Magnan and Jennifer A. Halliday and Ryan L. Frisch and Janet L. Gibson and Ralf Bernd Nehring and Huong G. Do and Marcos Hernandez and Lei Li and Christophe Herman and P. J. Hastings and David Bates and Reuben S. Harris and Kyle M. Miller and Susan M. Rosenberg},
   doi = {10.7554/ELIFE.01222.001},
   issn = {2050084X},
   issue = {2},
   journal = {eLife},
   month = {10},
   pmid = {24171103},
   publisher = {eLife Sciences Publications Ltd},
   title = {Engineered proteins detect spontaneous {DNA} breakage in human and bacterial cells},
   volume = {2013},
   year = {2013}
}
@article{Kalita2024,
   abstract = {All living organisms have developed strategies to respond to chromosomal damage and preserve genome integrity. One such response is the repair of {DNA} double-strand breaks (DSBs), one of the most toxic forms of {DNA} lesions. In {{\textit{Escherichia coli}}}, DSBs are repaired via {{RecB}CD}-dependent homologous recombination. {{RecB}CD} is essential for accurate chromosome maintenance, but its over-expression can lead to reduced {DNA} repair ability. This apparent paradox suggests that {{RecB}CD} copy numbers may need to be tightly controlled within an optimal range. Using single-molecule fluorescence mi-croscopy, we have established that {RecB} is present in very low abundance at mRNA and protein levels. {RecB} transcription shows high fluctuations, yet cell-to-cell protein variability remains remarkably low. Here, we show that the post-transcriptional regulator Hfq binds to recB mRNA and down-regulates {RecB} protein translation in vivo. Furthermore, specific disruption of the Hfq-binding site leads to more efficient translation of recB mRNAs. In addition, we observe a less effective reduction of {RecB} protein fluctuations in the absence of Hfq. This fine-tuning Hfq-mediated mechanism might have the underlying physiological function of maintaining {RecB} protein levels within an optimal range.},
   author = {Irina Kalita and Ira Alexandra Iosub and Lorna McLaren and Louise Goossens and Sander Granneman and Meriem El Karoui},
   doi = {10.7554/ELIFE.94918.1},
   journal = {eLife},
   month = {5},
   publisher = {eLife Sciences Publications Limited},
   title = {An Hfq-dependent post-transcriptional mechanism fine tunes {RecB} expression in {{\textit{Escherichia coli}}}},
   volume = {13},
   url = {https://elifesciences.org/reviewed-preprints/94918},
   year = {2024}
}
@article{Henrikus2020,
   abstract = {Several functions have been proposed for the {{\textit{Escherichia coli}}} {DNA} polymerase IV (pol IV). Although much research has focused on a potential role for pol IV in assisting pol III replisomes in the bypass of lesions, pol IV is rarely found at the replication fork in vivo. Pol IV is expressed at increased levels in {{\textit{E. coli}}} cells exposed to exogenous {DNA} damaging agents, including many commonly used antibiotics. Here we present live-cell single-molecule microscopy measurements indicating that double-strand breaks induced by antibiotics strongly stimulate pol IV activity. Exposure to the antibiotics ciprofloxacin and trimethoprim leads to the formation of double strand breaks in {{\textit{E. coli}}} cells. {RecA} and pol IV foci increase after treatment and exhibit strong colocalization. The induction of the {SOS} response, the appearance of {RecA} foci, the appearance of pol IV foci and {RecA}-pol IV colocalization are all dependent on {RecB} function. The positioning of pol IV foci likely reflects a physical interaction with the {RecA}∗ nucleoprotein filaments that has been detected previously in vitro. Our observations provide an in vivo substantiation of a direct role for pol IV in double strand break repair in cells treated with double strand break-inducing antibiotics.},
   author = {Sarah S. Henrikus and Camille Henry and Amy E. Mcgrath and Slobodan Jergic and John P. Mcdonald and Yvonne Hellmich and Steven T. Bruckbauer and Matthew L. Ritger and Megan E. Cherry and Elizabeth A. Wood and Phuong T. Pham and Myron F. Goodman and Roger Woodgate and Michael M. Cox and Antoine M. Van Oijen and Harshad Ghodke and Andrew Robinson},
   doi = {10.1093/NAR/GKAA597},
   issn = {0305-1048},
   issue = {15},
   journal = {Nucleic Acids Research},
   keywords = {antibiotics,binding (molecular function),ciprofloxacin,diagnostic imaging,dna,dna polymerase beta,double strand break repair,double-stranded dna breaks,molecule,sos response (genetics),trimethoprim},
   month = {9},
   pages = {8490-8508},
   pmid = {32687193},
   publisher = {Oxford Academic},
   title = {Single-molecule live-cell imaging reveals {RecB}-dependent function of {DNA} polymerase IV in double strand break repair},
   volume = {48},
   url = {https://dx.doi.org/10.1093/nar/gkaa597},
   year = {2020}
}
@article{Lepore2025,
   abstract = {Efficient {DNA} repair is essential for maintaining genome integrity and ensuring cell survival. In {{\textit{Escherichia coli}}}, {{RecB}CD} plays a crucial role in processing {DNA} ends, following a {DNA} double-strand break (DSB), to initiate repair. While {{RecB}CD} has been extensively studied in vitro, less is known about how it contributes to rapid and efficient repair in living bacteria. Here, we use single-molecule microscopy to investigate {DNA} repair in real time in {{\textit{E. coli}}}. We quantify {RecB} single-molecule mobility and monitor the induction of the {DNA} damage response ({SOS} response) in individual cells. We show that {RecB} binding to {DNA} ends caused by endogenous processes leads to efficient repair without {SOS} induction. In contrast, repair is less efficient in the presence of exogenous damage or in a mutant strain with modified {RecB} activities, leading to high {SOS} induction. Our findings reveal how subtle alterations in {RecB} activity profoundly impact the efficiency of {DNA} repair in {{\textit{E. coli}}}.},
   author = {Alessia Lepore and Daniel Thédié and Lorna Mclaren and Louise Goossens and Benura Azeroglu and Oliver J. Pambos and Achillefs N. Kapanidis and Meriem El Karoui},
   doi = {10.1093/NAR/GKAF454},
   issn = {13624962},
   issue = {10},
   journal = {Nucleic Acids Research},
   month = {5},
   pages = {454},
   pmid = {40464683},
   publisher = {Oxford Academic},
   title = {In vivo single-molecule imaging of {RecB} reveals efficient repair of {DNA} damage in {{\textit{Escherichia coli}}}},
   volume = {53},
   url = {https://dx.doi.org/10.1093/nar/gkaf454},
   year = {2025}
}
@article{delVal2021,
   abstract = {{RecA} plays a central role in {DNA} repair and is a main actor involved in recombination and activation of the {SOS} response. It is also used in the context of biotechnological applications in recombinase polymerase isothermal amplification (RPA). In this work, we studied the biological properties of seven {RecA} variants, in particular their recombinogenic activity and their ability to induce the {SOS} response, to better understand the structure–function relationship of {RecA} and the effect of combined mutations. We also investigated the biochemical properties of {RecA} variants that may be useful for the development of biotechnological applications. We showed that Dickeya dadantii {RecA} (Dd{RecA}) had an optimum strand exchange activity at 30&nbsp;°C and in the presence of a dNTP mixture that inhibited {{\textit{Escherichia coli}}} {RecA} (Ec{RecA}). The differences between the CTD and C-tail of the Ec{RecA} and Dd{RecA} domains could explain the altered behaviour of Dd{RecA}. D. radiodurans {RecA} (Dr{RecA}) was unable to perform recombination and activation of the {SOS} response in an {{\textit{E. coli}}} context, probably due to its inability to interact with {{\textit{E. coli}}} recombination accessory proteins and {SOS} LexA repressor. Dr{RecA} strand exchange activity was totally inhibited in the presence of chloride ions but worked well in acetate buffer. The overproduction of Pseudomonas aeruginosa {RecA} (Pa{RecA}) in an {{\textit{E. coli}}} context was responsible for a higher {SOS} response and defects in cellular growth. Pa{RecA} was less inhibited by the dNTP mixture than Ec{RecA}. Finally, the study of three variants, namely, EcPa, Ec{RecA}V1 and Ec{RecA}V2, that contained a combination of mutations that, taken independently, are described as improving recombination, led us to raise new hypotheses on the structure–function relationship and on the monomer–monomer interactions that perturb the activity of the protein as a whole.},
   author = {Elsa del Val and William Nasser and Hafid Abaibou and Sylvie Reverchon},
   doi = {10.1038/s41598-021-00589-9},
   isbn = {0123456789},
   issn = {2045-2322},
   issue = {1},
   journal = {Scientific Reports 2021 11:1},
   keywords = {Biochemistry,Microbiology},
   month = {10},
   pages = {1-17},
   pmid = {34702889},
   publisher = {Nature Publishing Group},
   title = {Design and comparative characterization of {RecA} variants},
   volume = {11},
   url = {https://www.nature.com/articles/s41598-021-00589-9},
   year = {2021}
}
@article{Pribis2019,
   abstract = {Antibiotics can induce mutations that cause antibiotic resistance. Yet, despite their importance, mechanisms of antibiotic-promoted mutagenesis remain elusive. We report that the fluoroquinolone antibiotic ciprofloxacin (cipro) induces mutations by triggering transient differentiation of a mutant-generating cell subpopulation, using reactive oxygen species (ROS). Cipro-induced {DNA} breaks activate the {{\textit{Escherichia coli}}} {SOS} {DNA}-damage response and error-prone {DNA} polymerases in all cells. However, mutagenesis is limited to a cell subpopulation in which electron transfer together with {SOS} induce ROS, which activate the sigma-S (σS) general-stress response, which allows mutagenic {DNA}-break repair. When sorted, this small σS-response-“on” subpopulation produces most antibiotic cross-resistant mutants. A U.S. Food and Drug Administration (FDA)-approved drug prevents σS induction, specifically inhibiting antibiotic-promoted mutagenesis. Further, {SOS}-inhibited cell division, which causes multi-chromosome cells, promotes mutagenesis. The data support a model in which within-cell chromosome cooperation together with development of a “gambler” cell subpopulation promote resistance evolution without risking most cells. Bacteria exposed to antibiotic acquire reactive oxygen in a transient “gambler” cell subpopulation that undertakes general stress response-induced mutagenic {DNA} break repair, evolves resistance to new antibiotics, and is inhibited by an FDA-approved drug that inhibits evolvability.},
   author = {John P. Pribis and Libertad García-Villada and Yin Zhai and Ohad Lewin-Epstein and Anthony Z. Wang and Jingjing Liu and Jun Xia and Qian Mei and Devon M. Fitzgerald and Julia Bos and Robert H. Austin and Christophe Herman and David Bates and Lilach Hadany and P. J. Hastings and Susan M. Rosenberg},
   doi = {10.1016/J.MOLCEL.2019.02.037},
   issn = {1097-4164},
   issue = {4},
   journal = {Molecular cell},
   keywords = {Anti-Bacterial Agents / adverse effects*,Bacterial / drug effects,Bacterial / genetics*,Cell Division / drug effects,Ciprofloxacin / adverse effects,{DNA} Damage / drug effects,{DNA}-Directed {DNA} Polymerase / genetics,Drug Resistance,{{\textit{Escherichia coli}}} / drug effects,{{\textit{Escherichia coli}}} / genetics*,{{\textit{Escherichia coli}}} / pathogenicity,Extramural,Gene Expression Regulation,Genetics / drug effects,John P Pribis,Libertad García-Villada,MEDLINE,Mutagenesis / drug effects,Mutagenesis / genetics*,Mutation,N.I.H.,NCBI,NIH,NLM,National Center for Biotechnology Information,National Institutes of Health,National Library of Medicine,Non-U.S. Gov't,PMC6553487,PubMed Abstract,Reactive Oxygen Species / metabolism,Research Support,{SOS} Response,Sigma Factor / genetics,Susan M Rosenberg,doi:10.1016/j.molcel.2019.02.037,pmid:30948267},
   month = {5},
   pages = {785-800.e7},
   pmid = {30948267},
   publisher = {Mol Cell},
   title = {Gamblers: An Antibiotic-Induced Evolvable Cell Subpopulation Differentiated by Reactive-Oxygen-Induced General Stress Response},
   volume = {74},
   url = {https://pubmed.ncbi.nlm.nih.gov/30948267/},
   year = {2019}
}