High-Resolution Whole-Genome Analysis of Sister-Chromatid Contacts

Espinosa, Elena; Paly, Evelyne; Barre, François Xavier

doi:10.1016/j.molcel.2020.06.033

Cited by 25 publications

(26 citation statements)

References 73 publications

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“…After 12 h (~25 generations) of incubation, cells were plated on LB agar containing Sm, Km and Amp and ~890,000 colonies were scraped to collect as the ‘output’ library. NGS library preparation, including gDNA preparation, MmeI digestion, adapter ligation and PCR amplification, was carried out as described in [ 22 ]. Three independent libraries were prepared with distinct ‘barcodes’ and subjected to independent Illumina sequencing twice, which was carried out at the I2BC NGS facility (Gif-sur-Yvette, France).…”

Section: Methodsmentioning

confidence: 99%

Vertical and Horizontal Transmission of ESBL Plasmid from Escherichia coli O104:H4

Daniel

Goldlust

Quèbre

et al. 2020

Genes

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Multidrug resistance (MDR) often results from the acquisition of mobile genetic elements (MGEs) that encode MDR gene(s), such as conjugative plasmids. The spread of MDR plasmids is founded on their ability of horizontal transference, as well as their faithful inheritance in progeny cells. Here, we investigated the genetic factors involved in the prevalence of the IncI conjugative plasmid pESBL, which was isolated from the Escherichia coli O104:H4 outbreak strain in Germany in 2011. Using transposon-insertion sequencing, we identified the pESBL partitioning locus (par). Genetic, biochemical and microscopic approaches allowed pESBL to be characterized as a new member of the Type Ib partitioning system. Inactivation of par caused mis-segregation of pESBL followed by post-segregational killing (PSK), resulting in a great fitness disadvantage but apparent plasmid stability in the population of viable cells. We constructed a variety of pESBL derivatives with different combinations of mutations in par, conjugational transfer (oriT) and pnd toxin-antitoxin (TA) genes. Only the triple mutant exhibited plasmid-free cells in viable cell populations. Time-lapse tracking of plasmid dynamics in microfluidics indicated that inactivation of pnd improved the survival of plasmid-free cells and allowed oriT-dependent re-acquisition of the plasmid. Altogether, the three factors—active partitioning, toxin-antitoxin and conjugational transfer—are all involved in the prevalence of pESBL in the E. coli population.

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Section: Methodsmentioning

confidence: 99%

Vertical and Horizontal Transmission of ESBL Plasmid from Escherichia coli O104:H4

Daniel

Goldlust

Quèbre

et al. 2020

Genes

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“…Samples for Tn-seq analysis were prepared as described ( Espinosa et al 2020 ). Libraries were paired-end sequenced in a Next-Seq (Illumina).…”

Section: Methodsmentioning

confidence: 99%

Plesiomonas shigelloides, an Atypical Enterobacterales with aVibrio-Related Secondary Chromosome

Adam

Brézellec

Espinosa

et al. 2022

Genome Biology and Evolution

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About 10% of bacteria have a multi-chromosome genome with a primary replicon of bacterial origin, called the chromosome, and other replicons of plasmid origin, the chromids. Studies on multi-chromosome bacteria revealed potential points of coordination between the replication/segregation of chromids and the progression of the cell cycle. For example, replication of the chromid of Vibrionales (called Chr2) is initiated upon duplication of a sequence carried by the primary chromosome (called Chr1), in such a way that replication of both replicons is completed synchronously. Also, Chr2 uses the Chr1 as a scaffold for its partition in the daughter cells. How many of the features detected so far are required for the proper integration of a secondary chromosome in the cell cycle? How many more features remain to be discovered? We hypothesized that critical features for the integration of the replication/segregation of a given chromid within the cell cycle program would be conserved independently of the species in which the chromid has settled. Hence, we searched for a chromid related to that found in Vibrionales outside of this order. We identified one in Plesiomonas shigelloides, an aquatic and pathogenic enterobacterium that diverged early within the clade of Enterobacterales. Our results suggest that the chromids present in P. shigelloides and Vibrionales derive from a common ancestor. We initiated in sillico genomic and proteomic comparative analyses of P. shigelloides, Vibrionales and Enterobacterales that enabled us to establish a list of features likely involved in the maintenance of the chromid within the host cell cycle.

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“…Previous works showed that the choreography of chromosome segregation restricts the activity of FtsK to the terminus region of the two V. cholerae chromosomes, suggesting that FtsK could not drive integration at the lacZ locus (Figure S2, (25)(26)(27)(28)). Correspondingly, no dif1specific integration events were observed at the lacZ locus with XafTmini-TLCΦ libraries.…”

Section: Xaft Drives the Integration Of Any Attp Site Harbouring The Same 5 Th Xerd-arm Bp Than Dif1mentioning

confidence: 95%

The Xer activation factor of TLCΦ expands the possibilities for Xer recombination

Miele

Vergnes

Possoz

et al. 2021

Preprint

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Many mobile elements take advantage of the highly-conserved chromosome dimer resolution system of bacteria, Xer. They participate in the transmission of antibiotic resistance and pathogenicity determinants. In particular, the toxin-linked cryptic satellite phage (TLCΦ) plays an essential role in the continuous emergence of new toxigenic clones of the Vibrio cholerae strain at the origin of the ongoing 7th cholera pandemic. The Xer machinery is composed of two chromosomally-encoded tyrosine recombinases, XerC and XerD. They resolve chromosome dimers by adding a crossover between sister copies of a specific 28 base pair site of bacterial chromosomes, dif. The activity of XerD depends on a direct contact with a cell division protein, FtsK, which spatially and temporally constrains the process. TLCΦ encodes for a XerD-activation factor (XafT), which drives the integration of the phage into the dif site of the primary chromosome of V. cholerae independently of FtsK. However, XerD does not bind to the attachment site (attP) of TLCΦ, which raised questions on the integration process. Here, we compared the integration efficiency of thousands of synthetic mini-TLCΦ plasmids harbouring different attP sites and assessed their stability in vivo. In addition, we compared the efficiency with which XafT and the XerD activation domain of FtsK drive recombination reactions in vitro. Taken together, our results suggest that XafT promotes the formation of synaptic complexes between canonical Xer recombination sites and imperfect sites.

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High-Resolution Whole-Genome Analysis of Sister-Chromatid Contacts

Cited by 25 publications

References 73 publications

Vertical and Horizontal Transmission of ESBL Plasmid from Escherichia coli O104:H4

Vertical and Horizontal Transmission of ESBL Plasmid from Escherichia coli O104:H4

Plesiomonas shigelloides, an Atypical Enterobacterales with aVibrio-Related Secondary Chromosome

The Xer activation factor of TLCΦ expands the possibilities for Xer recombination

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