Biology subjects

Cornet, F.

Publications and source records attributed to Cornet, F..

8 recordsLinked to original sources

bioRxiv2026

Chromosomal integration of blaCTX-M redefines the evolutionary trajectory of antibiotic resistance in E. coli through lineage-structured expansion

The global CTX-M pandemic is considered a plasmid-driven phenomenon, yet chromosomal integration of blaCTX-M has been reported in various settings. Screening 451,174 publicly available E. coli genomes and retaining 2,269 CTX-M-E. coli long-read or hybrid assemblies, we found that 33.5% carried at least one chromosomally integrated blaCTX-M. Chromosomal integration was strongly structured by E. coli lineage and CTX-M variant, reaching 68.5% in ST38 and 43.9% in ST131. In the latter, integration rates varied between subclades with modelling, suggesting recent acceleration within the ST131-C2 subclade. Additional resistance genes were frequently co-integrated with blaCTX-M (33%), with architectures largely inherited from plasmid-borne structures. Integrations were mainly ISEcp1- or IS26-mediated (77.3% and 18.9%, respectively) and occurred across the entire E. coli chromosome without spared zones. blaCTX-M chromosomal integration is a widespread, lineage-structured component of the CTX-M pandemic, providing stable vertical inheritance that complements or may supplant plasmid-mediated dissemination in successful ESBL-E. coli clones.

microbiology↗

bioRxiv2025

Fast assembly and in vivo coalescence of ParB biocondensates involved in bacterial DNA partition

Faithful DNA segregation in bacteria relies on ParABS systems, in which ParB assembles into condensates at centromere-like parS sites, while the ATPase ParA spatially organizes these complexes. How these ParB condensates maintain dynamic behavior without collapsing into a single structure has remained unclear. Here, we combine chromosome degradation with quantitative imaging to dissect the kinetics and physical principles governing ParB condensate dynamics in vivo. In the absence of the nucleoid, ParB condensates from the plasmid F diffuse freely and coalesce within seconds upon encounter. Strikingly, quantitative analyses indicate that condensates may operate near the fusion-separation boundary, such that minimal energy is sufficient to split them after replication, preventing irreversible coalescence. Using mutants, we demonstrate that condensate assembly is required for coalescence and uncover a dual role for ParAF: nucleoid tethering restricts condensate mobility and limits fusion, while ParAF also promotes a ParBF state competent for assembly and coalescence, likely by enhancing ParB-ParB interactions. Finally, condensates rapidly disassemble and reassemble upon 1,6-hexanediol treatment, underscoring their reversibility and the stabilizing contribution of ParB-DNA interactions. Together, our results establish ParBF complexes as bona-fide biocondensates tuned by ParAF to ensure robust DNA segregation. More broadly, these findings highlight regulated phase separation as a key organizing principle of bacterial replicons.

microbiology↗

bioRxiv2024

Interplay between the Xer recombination system and the dissemination of antibioresistance in Acinetobacter baumannii

Antibiotic-resistant infections pose a pressing challenge in clinical settings. Plasmids are widely recognized for hastening the emergence of resistance by facilitating horizontal gene transfer of antibiotic resistance genes among bacteria. We explore this inquiry in Acinetobacter baumannii, a globally emerging nosocomial pathogen responsible for a wide array of infections with worrying accumulation of resistances, notably involving plasmids. In this specie, plasmids of the Rep_3 family harbor adaptive genes within variable regions edged by potential site-specific recombination sites recognized by the XerCD recombinase. We first show that the Xer system of Acinetobacter baumannii functions as described in Escherichia coli, resolving chromosome dimers at the dif site as well as recombining plasmid-borne sites. The multiple Xer recombination sites found in Rep_3 plasmids do not, however, allow excising plasmid fragments. They rather recombine to co-integrate plasmids, which may then further evolve to exchange genes. Co-integrates represent a significative part of the plasmid population and their formation is controlled by the sequence of the recombination sites determining their compatibility between the recombining sites. We conclude that plasmids frequently exchange genes in Acinetobacter baumannii using Xer recombination, allowing a high level yet controlled plasticity involved in the acquisition and combination of resistance genes.

microbiology↗

bioRxiv2024

A mechanistic step in the genesis of secondary chromosomes from plasmids

Bacterial genomes contain a plethora of secondary replicons of divergent size. Circular replicons must carry a system for resolving dimeric forms, resulting from recombination between sister copies. These systems use site-specific recombinases. Among these, the XerCD recombinase resolves dimers of chromosomes and certain plasmids using different controls. We have analyzed the dimer resolution functions in enterobacterial secondary replicons and show that, in addition to the main chromosomes, XerCD is preferentially used by small plasmids and by the largest secondary replicons, megaplasmids and secondary chromosomes. Indeed, all replicons longer than 250 kb host an active XerCD recombination site. These sites, in contrast to those of small plasmids, use the same control as chromosomes, coupled to cell division by the FtsK protein. We conclude that a chromosome-like mode of dimer resolution is mandatory for the faithful inheritance of large plasmids and chromids, its acquisition being a prerequisite for the genesis of secondary chromosomes from plasmids.

microbiology↗

bioRxiv2024

In vivo Assembly of Bacterial Partition Condensates on Supercoiled and Linear DNA

In bacteria, faithful DNA segregation of chromosomes and plasmids is mainly mediated by ParABS systems. These systems, consisting of a ParA ATPase, a DNA binding ParB CTPase, and centromere sites parS, orchestrate the separation of newly replicated DNA copies and their intracellular positioning. Accurate segregation relies on the assembly of a high-molecular-weight complex, comprising a few hundreds of ParB dimers nucleated from parS sites. This complex assembles in a multi-step process and exhibits dynamic liquid-droplet properties. Despite various proposed models, the complete mechanism for partition complex assembly remains elusive. This study investigates the impact of DNA supercoiling on ParB DNA binding profiles in vivo, using the ParABS system of the plasmid F. We found that variations in DNA supercoiling does not significantly affect any steps in the assembly of the partition complex. Furthermore, physical modeling, leveraging ChIP-seq data from linear plasmids F, suggests that ParB sliding is restricted to approximately 2-Kbp from parS, highlighting the necessity for additional mechanisms beyond ParB sliding over DNA for concentrating ParB into condensates nucleated at parS. Lastly, explicit simulations of a polymer coated with bound ParB suggest a dominant role for ParB-ParB interactions in DNA compaction within ParB condensates.

molecular biology↗

bioRxiv2022

A subclass of the IS1202 family of bacterial insertion sequences targets XerCD recombination sites

IS1202, originally isolated from Streptococcus pneumonia in the mid-1990s had been previously tagged as an emerging IS family in ISfinder. While searching for plasmid-associated Xer recombinase recombination sites (xrs) in Acinetobacter baumannii, we observed that some insertion sequences related to IS1202 were repeatedly found abutting these sites in a number of plasmids. The plasmids often carried repeated xrs thought to form a new type of mobile genetic element (MGE) which uses the chromosomally-encoded XerCD recombinase for mobility. The MGE (xrs cassette) consist of xrs flanking one or a small number of genes often including different clinically important carbapenemase-encoding bla-OXA. The IS1202- related IS are inserted with their left, transposase proximal extremity, IRL, five base pairs from xrs and include a characteristic 5bp flanking target duplication. Further searches revealed that many different plasmid- and chromosome-borne xrs can be targeted and that IS1202-xrs combinations are not limited to Acinetobacter baumannii but occur in other bacteria. In addition to 28 IS1202 group ISs in ISfinder and a number which had been subsequently submitted, we undertook a survey of the NCBI (February 2020) and identified 138 additional IS1202-related IS. These could be divided into 3 principal subgroups based on their transposase sequences and on the length of the DR generated on insertion: subgroup IS1202 27-28bp DR); ISTde1 (15-17bp); and ISAba32 (5-6bp). Members of each group which lacked DR were also found. But since other examples of most of these were subsequently identified having DR, those lacking DR may have been generated by intra-replicon recombination. Only members of the group which generate 5bp DR were found to target xrs. These were not only identified in plasmids but also occurred at some individual xrs sites, dif, located at the chromosome replication terminus and involved in post-replication chromosome segregation. Further analysis showed the presence of subgroup-specific indels in their transposases which may be responsible for the differences in their behavior. We propose that this collection of IS be classed as a new insertion sequence family: the IS1202 family composed of at three subfamilies, only one of which specifically targets plasmid-borne xrs. We discuss the implications of xrs targeting for gene mobility.

genomics↗

bioRxiv2022

Characterization of the DNA binding domain of StbA, a key protein of a new type of DNA segregation system

Low-copy-number plasmids require sophisticated genetic devices to achieve efficient segregation of plasmid copies during cell division. Plasmid R388 uses a unique segregation mechanism, based on StbA, a small multifunctional protein. StbA is the key protein in a segregation system not involving a plasmid-encoded NTPase partner, it regulates the expression of several plasmid operons, and it is the main regulator of plasmid conjugation. The mechanisms by which StbA, together with the centromere-like sequence stbS, achieves segregation, is largely uncharacterized. To better understand the molecular basis of R388 segregation, we determined the crystal structure of the conserved N-terminal domain of StbA to 1.9 [A] resolution. It folds into an HTH DNA-binding motif, structurally related to that of the PadR subfamily II of transcriptional regulators. StbA is organized in two domains. Its N-terminal domain carries the specific stbS DNA binding activity. A truncated version of StbA, deleted of its C-terminal domain, displays only partial activities in vivo, indicating that the non-conserved C-terminal domain is required for efficient segregation and subcellular plasmid positioning. The structure of StbA DNA-binding domain also provides some insight into how StbA monomers cooperate to repress transcription by binding to the stbDR and to form the segregation complex with stbS.

microbiology↗

bioRxiv2019

Multiple activities of the MatP protein are involved in post-replicative pairing of sister chromosomes in Escherichia coli

The ter region of the bacterial chromosome, where replication terminates, is the last to be segregated before cell division in Escherichia coli. Delayed segregation is controlled by the MatP protein, which binds to specific sites (matS) within ter, and interacts with other proteins such as ZapB. Here, we investigate the role of MatP by combining short-time mobility analyses of the ter locus with biochemical approaches. We find that ter mobility is similar to that of a non-ter locus, except when sister ter loci are paired after replication. This effect depends on MatP, the persistence of catenanes, and ZapB. We characterise MatP/DNA complexes and conclude that MatP binds DNA as a tetramer, but bridging matS sites in a DNA-rich environment remains infrequent. We propose that tetramerisation of MatP links matS sites with ZapB and/or with non-specific DNA to promote optimal pairing of sister ter regions until cell division.Competing Interest StatementThe authors have declared no competing interest.View Full Text

microbiology↗