scholarly journals Dissecting the Control Mechanisms for DNA Replication and Cell Division in E. coli

Cell Reports ◽  
2018 ◽  
Vol 25 (3) ◽  
pp. 761-771.e4 ◽  
Author(s):  
Gabriele Micali ◽  
Jacopo Grilli ◽  
Jacopo Marchi ◽  
Matteo Osella ◽  
Marco Cosentino Lagomarsino
Keyword(s):  
Cell Division ◽  
Dna Replication ◽  
Control Mechanisms ◽  
E Coli

scholarly journals Dissecting the control mechanisms for DNA replication and cell division in E. coli

2018 ◽  
Author(s):  
Gabriele Micali ◽  
Jacopo Grilli ◽  
Jacopo Marchi ◽  
Matteo Osella ◽  
Marco Cosentino Lagomarsino
Keyword(s):  
Cell Division ◽  
Dna Replication ◽  
Cell Cycle Progression ◽  
Genome Replication ◽  
Control Mechanisms ◽  
Cycle Progression ◽  
E Coli ◽  
Classic Problem ◽  
Initiation Of Replication ◽  
Initiation Of Dna Replication

Understanding how single E. coli cells coordinate the timing of cell division with genome replication would unlock a classic problem of biology, and open the way to address cell-cycle progression at the single-cell level. Several recent studies produced new data and proposed different models, based on the hypothesis that replication-segregation is the bottleneck process for cell division. However, due to the apparent contrast in both experimental results and proposed mechanisms, the emerging picture is fragmented and unclear. In this work, we re-evaluate jointly available data and models, and we show that, while each model contains useful insights, none of the proposed models, as well as generalizations based on the same assumptions, correctly describes all the correlation patterns observed in data. This analysis leads us to conclude that the assumption that replication is the bottleneck process for cell division is too restrictive. Instead, we propose that two concurrent cycles responsible for division and initiation of DNA replication together set the time of cell division. This framework correctly captures available data and allows us to select a nearly constant added size per origin between subsequent initiations as the most likely mechanism setting initiation of replication.

scholarly journals DNA Double Strand Break Repair in E. coli Perturbs Cell Division and Chromosome Dynamics

2019 ◽  
Author(s):  
M.A. White ◽  
E. Darmon ◽  
M.A. Lopez-Vernaza ◽  
D.R.F. Leach
Keyword(s):  
Cell Cycle ◽  
Cell Division ◽  
Dna Replication ◽  
Cell Size ◽  
Strand Break ◽  
Dsb Repair ◽  
Average Cell Size ◽  
Average Cell ◽  
E Coli ◽  
Dna Double Strand Break

AbstractTo prevent the transmission of damaged genomic material between generations, cells require a system for accommodating DNA repair within their cell cycles. We have previously shown that Escherichia coli cells subject to a single, repairable site-specific DNA double-strand break (DSB) per DNA replication cycle reach a new average cell length, with a negligible effect on population growth rate. We show here that this new cell size distribution is caused by a DSB repair-dependent delay in completion of cell division. This delay occurs despite unperturbed cell size regulated initiation of both chromosomal DNA replication and cell division. Furthermore, despite DSB repair altering the profile of DNA replication across the genome, the time required to complete chromosomal duplication is invariant. The delay in completion of cell division is accompanied by a DSB repair-dependent delay in individualization of sister nucleoids. We suggest that DSB repair events create inter-sister connections that persist until those chromosomes are separated by a closing septum.Author SummaryThe bacterium Escherichia coli has a remarkable cell cycle where overlapping rounds of DNA replication can occur in a single generation between cell birth and division. This implies a complex coordination network between growth, genome duplication and cell division to ensure that the right number of genomes are created and distributed to daughter cells at all growth rates. This network must be robust to a number of unpredictable challenges. One such challenge is broken DNA, something that in E. coli is estimated to occur in ~20% of cell division cycles. In this work we perturb the E. coli cell cycle by elevating the frequency of repairable DNA double-strand breaks to determine which parameters of the cell cycle are conserved and which are changed. Our results demonstrate that this perturbation does not alter the average cell size at initiation of DNA replication or initiation of cell division. Furthermore, it does not alter the time taken to replicate the genome or the generation time. However, it does delay the segregation of the DNA to daughter cells and the completion of cell division explaining the increase in average cell size observed previously.

An inducible DNA replication–cell division coupling mechanism in E. coli

Nature ◽  
1981 ◽  
Vol 290 (5809) ◽  
pp. 797-799 ◽  
Author(s):  
Olivier Huisman ◽  
Richard D'Ari
Keyword(s):  
Cell Division ◽  
Dna Replication ◽  
Coupling Mechanism ◽  
E Coli

Replication of Bacteriophage 186 DNA

1972 ◽  
Vol 30 ◽  
pp. 194-195
Author(s):  
Dhruba K. Chattoraj ◽  
Ross B. Inman
Keyword(s):  
Electron Microscopy ◽  
Dna Replication ◽  
Frame Of Reference ◽  
Dna Molecules ◽  
E Coli ◽  
Phage Dna ◽  
Partial Denaturation

Electron microscopy of replicating intermediates has been quite useful in understanding the mechanism of DNA replication in DNA molecules of bacteriophage, mitochondria and plasmids. The use of partial denaturation mapping has made the tool more powerful by providing a frame of reference by which the position of the replicating forks in bacteriophage DNA can be determined on the circular replicating molecules. This provided an easy means to find the origin and direction of replication in λ and P2 phage DNA molecules. DNA of temperate E. coli phage 186 was found to have an unique denaturation map and encouraged us to look into its mode of replication.

scholarly journals E. coli primase and DNA polymerase III holoenzyme are able to bind concurrently to a primed template during DNA replication

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Andrea Bogutzki ◽  
Natalie Naue ◽  
Lidia Litz ◽  
Andreas Pich ◽  
Ute Curth
Keyword(s):  
Dna Replication ◽  
Dna Polymerase ◽  
Protein Interactions ◽  
Dna Polymerase Iii ◽  
Protein Protein Interactions ◽  
Single Stranded Dna ◽  
E Coli ◽  
Polymerase Iii ◽  
C Terminus ◽  
Pol Iii

Abstract During DNA replication in E. coli, a switch between DnaG primase and DNA polymerase III holoenzyme (pol III) activities has to occur every time when the synthesis of a new Okazaki fragment starts. As both primase and the χ subunit of pol III interact with the highly conserved C-terminus of single-stranded DNA-binding protein (SSB), it had been proposed that the binding of both proteins to SSB is mutually exclusive. Using a replication system containing the origin of replication of the single-stranded DNA phage G4 (G4ori) saturated with SSB, we tested whether DnaG and pol III can bind concurrently to the primed template. We found that the addition of pol III does not lead to a displacement of primase, but to the formation of higher complexes. Even pol III-mediated primer elongation by one or several DNA nucleotides does not result in the dissociation of DnaG. About 10 nucleotides have to be added in order to displace one of the two primase molecules bound to SSB-saturated G4ori. The concurrent binding of primase and pol III is highly plausible, since even the SSB tetramer situated directly next to the 3′-terminus of the primer provides four C-termini for protein-protein interactions.

scholarly journals Regulation of Cell Division in Bacteria by Monitoring Genome Integrity and DNA Replication Status

2019 ◽  
Vol 202 (2) ◽  
Author(s):  
Peter E. Burby ◽  
Lyle A. Simmons
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Cell Division ◽  
Dna Replication ◽  
Cell Cycle Progression ◽  
Genome Instability ◽  
Sos Response ◽  
Bacterial Cells ◽  
Cycle Progression ◽  
Content Type

ABSTRACT All organisms regulate cell cycle progression by coordinating cell division with DNA replication status. In eukaryotes, DNA damage or problems with replication fork progression induce the DNA damage response (DDR), causing cyclin-dependent kinases to remain active, preventing further cell cycle progression until replication and repair are complete. In bacteria, cell division is coordinated with chromosome segregation, preventing cell division ring formation over the nucleoid in a process termed nucleoid occlusion. In addition to nucleoid occlusion, bacteria induce the SOS response after replication forks encounter DNA damage or impediments that slow or block their progression. During SOS induction, Escherichia coli expresses a cytoplasmic protein, SulA, that inhibits cell division by directly binding FtsZ. After the SOS response is turned off, SulA is degraded by Lon protease, allowing for cell division to resume. Recently, it has become clear that SulA is restricted to bacteria closely related to E. coli and that most bacteria enforce the DNA damage checkpoint by expressing a small integral membrane protein. Resumption of cell division is then mediated by membrane-bound proteases that cleave the cell division inhibitor. Further, many bacterial cells have mechanisms to inhibit cell division that are regulated independently from the canonical LexA-mediated SOS response. In this review, we discuss several pathways used by bacteria to prevent cell division from occurring when genome instability is detected or before the chromosome has been fully replicated and segregated.

scholarly journals Artificial Septal Targeting of Bacillus subtilis Cell Division Proteins in Escherichia coli: an Interspecies Approach to the Study of Protein-Protein Interactions in Multiprotein Complexes

2008 ◽  
Vol 190 (18) ◽  
pp. 6048-6059 ◽  
Author(s):  
Carine Robichon ◽  
Glenn F. King ◽  
Nathan W. Goehring ◽  
Jon Beckwith
Keyword(s):  
Escherichia Coli ◽  
Bacillus Subtilis ◽  
Cell Division ◽  
Protein Interactions ◽  
Fluorescent Protein ◽  
Bacterial Cell Division ◽  
Protein Protein Interactions ◽  
Positive Interaction ◽  
E Coli ◽  
Multiprotein Complexes

ABSTRACT Bacterial cell division is mediated by a set of proteins that assemble to form a large multiprotein complex called the divisome. Recent studies in Bacillus subtilis and Escherichia coli indicate that cell division proteins are involved in multiple cooperative binding interactions, thus presenting a technical challenge to the analysis of these interactions. We report here the use of an E. coli artificial septal targeting system for examining the interactions between the B. subtilis cell division proteins DivIB, FtsL, DivIC, and PBP 2B. This technique involves the fusion of one of the proteins (the “bait”) to ZapA, an E. coli protein targeted to mid-cell, and the fusion of a second potentially interacting partner (the “prey”) to green fluorescent protein (GFP). A positive interaction between two test proteins in E. coli leads to septal localization of the GFP fusion construct, which can be detected by fluorescence microscopy. Using this system, we present evidence for two sets of strong protein-protein interactions between B. subtilis divisomal proteins in E. coli, namely, DivIC with FtsL and DivIB with PBP 2B, that are independent of other B. subtilis cell division proteins and that do not disturb the cytokinesis process in the host cell. Our studies based on the coexpression of three or four of these B. subtilis cell division proteins suggest that interactions among these four proteins are not strong enough to allow the formation of a stable four-protein complex in E. coli in contrast to previous suggestions. Finally, our results demonstrate that E. coli artificial septal targeting is an efficient and alternative approach for detecting and characterizing stable protein-protein interactions within multiprotein complexes from other microorganisms. A salient feature of our approach is that it probably only detects the strongest interactions, thus giving an indication of whether some interactions suggested by other techniques may either be considerably weaker or due to false positives.

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