A functional partnership between a lipid scramblase and a lipid transfer protein in the regulation of phosphatidylserine homeostasis.

Phosphatidylserine is synthesised in the endoplasmic reticulum (ER), but this negatively charged lipid is highly concentrated in the plasma membrane (PM). There, it plays numerous roles in processes such as cell signalling and cell fusion, and it can also mediate apoptosis and synaptic pruning when exposed in the outer leaflet of the PM. In the yeast Saccharomyces cerevisiae, selective PS transport between the ER and the PM by the lipid transfer protein Osh6 is required for PS enrichment in the plasma membrane. Osh6 operates at membrane contact sites (MCS), where it interacts with the Ist2 protein embedded in the ER, tethering the ER to the PM via its long, disordered C-terminal tail.

In two studies conducted in collaboration with the CRBM (CNRS/University of Montpellier), the IPMC (CNRS/University of Nice Côte d'Azur), and the IBCP (CNRS/University of Lyon), we show that Ist2 is a membrane transporter that catalyses rapid lipid exchange across the two leaflets of the ER – acting as a lipid ‘scramblase’ – and that this scrambling activity sustains Osh6-mediated PS transfer between the ER and the PM.

First, we demonstrated that Ist2 catalyses the scrambling of different lipids in vitro after reconstituting purified Ist2 in proteoliposomes. Molecular dynamics simulations were then used to identify a cavity through which the lipid headgroup passes during transport from one leaflet to another. Further cellular studies revealed a close relationship between the COPII complex and the Ist2 protein. This was manifested by impaired yeast growth and vacuolar trafficking, as well as disruption of ER exit sites. We also found that Ist2 deletion stimulates the formation of ER-derived lipid droplets and changes their composition. Finally, using artificial, reconstituted ER-PM contact sites, we demonstrated that Ist2-mediated lipid scrambling sustains Osh6-mediated lipid transfer. 

Together, our studies identify Ist2 as a lipid scramblase and establish that lipid scrambling in the ER by Ist2 controls various cellular functions, such as vesicular transport and lipid droplet homeostasis. This highlights the importance of lipid dynamics for ER function. Furthermore, we reveal a functional partnership between Ist2-mediated lipid scrambling and Osh6-mediated lipid transfer at MCS.

More information :

https://doi-org.insb.bib.cnrs.fr/10.1083/jcb.202502112

https://www-science-org.insb.bib.cnrs.fr/doi/10.1126/sciadv.adz2217

Contact : Guillaume Lenoir guillaume.lenoir@i2bc.paris-saclay.fr

NMT in a new light: associated with the ribosome via the NAC complex, this enzyme not only adds lipid tags to the N-termini of nascent proteins but also acts as a chaperone, cooperating with MetAPs to ensure proper folding and delivery.

N-myristoylation is an essential cotranslational lipid modification catalyzed by N-myristoyltransferases (NMTs). Structural and cellular analyses reveal that NMT1 associates with the ribosomal tunnel exit via the nascent polypeptide–associated complex (NAC) and acts sequentially after MetAP-mediated initiator methionine removal, in contrast to previously described simultaneous cotranslational modification assemblies. Unexpectedly, NMT1 also exhibits chaperone-like activity, expanding its functional repertoire in cotranslational protein biogenesis.

More information : https://www-nature-com.insb.bib.cnrs.fr/articles/s41467-025-67962-4

Contact : Thierry Meinnel and Carmela Giglione thierry.meinnel@i2bc.paris-saclay.fr and carmela.giglione@i2bc.paris-saclay.fr 

Noncoding DNA can generate microproteins, some of which evolve into new genes. We show that de novo genes preferentially originate from GC-rich, foldable sequences, revealing how base composition channels the birth of new proteins.

Noncoding regions of eukaryotic genomes are widely transcribed and constitute a major source of novel microproteins, some of which eventually become fixed as de novo genes - a process known as de novo gene birth that plays a significant role in species adaptation. However, the structural properties of these nascent proteins and the factors governing their evolutionary fate remain poorly understood. In particular, the role of genome nucleotide composition (GC content) in shaping their biophysical properties has remained unclear. In this study, researchers of the I2BC, analyzed the foldability and sequence properties of millions of putative microproteins encoded by intergenic open reading frames (ORFs) from 3,379 eukaryotic species spanning a broad range of GC contents (18–79%). Results show that GC content strongly influences amino-acid composition and structural tendencies, suggesting distinct cellular impacts if non-genic regions are pervasively expressed. AT-rich species predominantly encode ORFs biased toward hydrophobic, aggregation-prone sequences, whereas GC-rich species tend to encode more hydrophilic, disorder-prone ORFs. ORFs from genomes with intermediate GC content display a more balanced composition and higher folding potential, with many expected to adopt proto-folds. To assess how these properties relate to gene emergence,the authors traced the evolutionary history of several hundred de novo proteins across 22 species using phylostratigraphy, targeted de novo gene searches, and ancestral sequence reconstruction. Researchers find that de novo genes preferentially originate from GC-rich ORFs with intrinsic folding potential. Together, our results reveal that the interplay between GC content and foldability - rooted in the structure of the genetic code - shapes the emergence of novel genes.

More information: https://www.nature.com/articles/s41467-025-68022-7

Contact: Anne Lopes anne.lopes@i2bc.paris-saclay.fr

We developed a flexizyme-based semi-synthetic strategy that provides access to stable 2′- and 3′-amide AA-tRNA analogs, offering robust tools for structural biology and for probing regiospecificity in AA-tRNA-dependent enzymes.

The study of the regiospecificity of aminoacyl-tRNA (AA-tRNA)-dependent enzymes and their structural characterization with AA-tRNAs are limited by rapid hydrolysis of the ester bond linking amino acid to tRNA. To overcome this limitation, stable AA-tRNA analogs bearing hydrolysis-resistant linkages, such as amide bonds or ester bioisosteres, have been developed. These analogs are valuable tools for investigating interactions between AA-tRNAs and various enzymes or ribonucleoproteins, including elongation factors, ribosomes, Fem-family transferases, and cyclodipeptide synthases. However, their synthesis remains technically challenging. Recently, flexizymes—engineered ribozymes capable of aminoacylating tRNAs with diverse amino acids or analogs—have enabled the synthesis of 3′-amide-linked AA-NH-tRNAs. Due to their inherent specificity for 3′-OH acylation, flexizymes have not been used to generate 2′-amide-linked analogs, and such regioisomers have remained unexplored. In this study, we demonstrate that while flexizymes cannot directly aminoacylate the 2′ position, they can nevertheless mediate the synthesis of 2′-aminoacyl-NH-tRNAs via a two-step regioisomerization mechanism with excellent yields. This finding provides new insights into the binding mode of AA-tRNAs to flexizymes and expands the chemical space of stable AA-tRNA analogs. Access to both 3′- and 2′-amide regioisomers will enable more precise studies of AA-tRNA recognition and catalysis by various AA-tRNA-dependent systems.

More information : https://doi.org/10.1002/chem.202503506

Contact person: Matthieu Fonvielle matthieu.fonvielle@i2bc.paris-saclay.fr

https://www.i2bc.paris-saclay.fr/enzymology-and-non-ribosomal-peptide-biosynthesis/

We identified a new target for antiparasitic compounds and it is in the mitochondria.

Chagas disease, caused by Trypanosoma cruzi, is a neglected parasitic infection. The very limited arsenal of anti-T. cruzi treatments calls for the development of new drugs. Recently, a library of 3-benzylmenadione derivatives was synthesized with cruzidione being the most efficient and specific compound against the parasite. To decipher its mode of action, we used the yeast Saccharomyces cerevisiae as model. Evidence pinpointed at the heme A synthase Cox15 as a primary target of cruzidione: 1) a mutation in Cox15 (i.e., S429F) renders the yeast cells highly sensitive to the drug, 2) treatment with cruzidione led to the loss of cytochrome c oxidase, an enzyme that relies on heme A as an essential cofactor and 3) replacement of the yeast Cox15 by T. cruzi enzyme resulted in a high sensitivity to cruzidione. We then investigated the effect of cruzidione in T. cruzi and observed a significant reduction of heme contents, most likely involving the inhibition of the heme A synthase. This, in turn, led to a decrease in O2 consumption by the parasite. Finally, using the yeast model, we showed that, similarly to what we previously found for the antimalarial benzylmenadione plasmodione, NADH-dehydrogenase plays a key role in cruzidione bioactivation. We proposed that the reduced benzoylmenadione metabolites produced by the reaction with NADH-dehydrogenase, act as Cox15 inhibitors. This study, through the identification of the mode of action of cruzidione, highlighted Cox15 as a novel target for antiparasitic drugs.

More information: https://journals.asm.org/doi/10.1128/aac.01161-25

Contact: Brigitte Meunier, brigitte.meunier@i2bc.paris-saclay.fr

Understanding how the essential complexes —the coregulator Mediator and the chromatin remodeling complex RSC/PBAF— cooperate in the nucleus

In eukaryotes, transcription and DNA repair occur in the crowded context of chromatin. Dysfunctions of these processes can lead to cancers. Mediator is an essential and conserved multisubunit coactivator complex, mutated in many cancers. However, it remains largely unknown how Mediator and chromatin regulators coordinate their functions. A recent publication of the team suggests the novel hypothesis that Mediator acts in conjunction with the chromatin remodeling complex RSC (Remodels the Structure of Chromatin) of SWI/SNF family, homologous to PBAF (Polybromo-associated BAF) in human, representing the most frequently mutated complexes in cancers. Building on this recent publication, a new project has been supported by Ligue Nationale contre le cancer. Using the yeast model, with a perspective to extend the study to human cells, the team intends to decipher the molecular mechanisms involved in functional cooperation between these essential coregulator complexes in transcription regulation, DNA repair and chromatin organization relevant for cancer biology.

Contact: Julie Soutourina julie.soutourina@i2bc.paris-saclay.fr

From structural modeling to natural transformation in bacteria: identification of a key protein complex for DNA transport across the membrane of Firmicutes.

Transformation is a key mechanism of horizontal gene transfer, central to bacterial adaptation. This evolutionarily conserved process allows bacteria to integrate exogenous genetic material into their genome, thereby facilitating, for example, the spread of antibiotic resistance.
In a study published in the journal PNAS, scientists from I2BC (CEA/CNRS/UPSaclay, Gif-sur-Yvette) and DRCM/IBFJ (CEA/UPSaclay/UP Cité, Fontenay-aux-Roses), in collaboration with a laboratory at CBI (CNRS/University of Toulouse), identified a protein complex involved in the transport of single-stranded DNA across the membrane during transformation. Using AlphaFold, they structurally modeled this three-protein complex, which is highly conserved in the phylum Firmicutes, in interaction with single-stranded DNA. The structural model allowed them to identify a possible path for DNA through a conserved channel in one of the three proteins, a transmembrane protein, and then along a groove formed by the other two proteins. This model was validated by a robust experimental strategy in the bacterium Streptococcus pneumoniae, by measuring the impact on transformation efficiency of disruptive mutations in several protein–protein and protein–DNA interfaces. The transmembrane channel was found to be conserved in a structural model in Helicobacter pylori, and its importance for transformation was also experimentally confirmed in this bacterium. This study sheds light on the molecular mechanisms of bacterial transformation and demonstrates the power of macromolecular structure prediction to generate molecular hypotheses and guide functional experiments.

More information: https://doi.org/10.1073/pnas.2511180122

Contact : Jessica Andreani jessica.andreani@i2bc.paris-saclay.fr

Study published in PLoS Pathogens of molecular features of phage receptor SlpA protein demonstrated the complexity of the interactions between human enteropathogen C. difficile and its phages

The emergence of Clostridioides difficile as a leading cause of healthcare-associated nosocomial intestinal infections calls for novel therapeutic strategies, particularly in the context of antibiotic resistance and recurrent disease. Phage therapy is a promising approach, but its clinical application against C. difficile remains hindered by our lack of understanding of the factors driving host specificity. Indeed, it is crucial to understand how phages specifically interact with their host to be able to select the best candidates for cocktail preparation. The main surface layer protein SlpA is a key phage receptor, but the molecular details governing phage-receptor interactions remain unclear. By dissecting the structural features of SlpA required for phage infection through engineered SlpA isoforms and domain modifications, in collaboration with our Canadian collegues from Sherbrooke University, we reveal how specific regions of SlpA mediate phage adsorption and infection. These new insights into phage–receptor interactions will be instrumental in guiding the future engineering of broad-host-range therapeutic phages.

More information : https://doi. org/10.1371/journal.ppat.1013724 

Contact : Olga Soutourina olga.soutourina@i2bc.paris-saclay.fr

https://www.i2bc.paris-saclay.fr/regulatory-rnas-in-clostridia/

Insects choose their bacterial gut symbionts by creating a multifactorial selective environment that promotes competition among symbiont candidates and colonization by the single, best-adapted strain

Funded by the European Research Council (ERC synergy 2025), the project “3Stops2Go” leaps over the “three red lights” of premature stop codons to re-express critical protein and correct genetic diseases.

The project, recently funded by an ERC Synergy Grant 2025, aims to target premature termination codon (PTC)mutations, which prematurely halt protein translation and are involved in about 11% of human genetic diseases.
By combining expertise in protist biology, RNA biology, and gene therapy, the consortium of four researchers (Leoš Valášek (Institute of Microbiology, Czech Academy of Sciences, Czech Republic), Julius Lukeš (Biology Centre, Czech Academy of Sciences, Czech Republic), Olivier Namy (Université Paris-Saclay, CEA, CNRS, France / I2BC, France), and Mark Osborn (University of Minnesota, USA)) aims to harness natural stop-codon bypass mechanisms to develop therapeutic tools (engineered tRNAs and readthrough inducers), test them in patient-derived cells and animal models, and ultimately pave the way toward clinical applications.

More information: https://erc.europa.eu/news-events/news/synergy-grants-2025-examples-projects

Contact: Olivier Namy olivier.namy@i2bc.paris-saclay.fr>

Cécile Courret, CNRS Researcher and recipient of an ATIP-Avenir grant, has joined the Department of Genome Biology to establish her team ‘Intragenomic Conflict and Evolution’.

Cécile's research focus on genetic conflicts and their impact on genome evolution. The basic principles of Mendelian inheritance state that in heterozygous individuals, two alleles have equal chances of being transmitted to the next generation. However, some genetic elements do not follow these rules. These so-called selfish genetic elements, such as transposons or meiotic drivers, bias inheritance in their own favor, often at the expense of the organism.

Because they disrupt essential processes like meiosis, heterochromatin regulation, and cell division, selfish elements create a persistent conflict with the host genome. This conflict triggers an evolutionary “arms race,” in which genomes evolve defense mechanisms to counterbalance the harmful effects of these elements. Far from being rare exceptions, such conflicts are now recognized as a major force shaping genome structure and function.

My research relies on the Drosophila model and combines genomic, molecular, and cytological approaches to investigate both how selfish elements perturb fundamental biological processes and how organisms respond to mitigate these disruptions. By studying systems where these conflicts are still active in natural populations, as well as the genomic signatures left by past events, Cécile's team aims to better understand how conflicts drive evolutionary innovation and shed light on the molecular basis of essential biological mechanisms.

Contact: Cécile Courret, cecile.courret@i2bc.paris-saclay.fr

Pushing Streptomyces engineering to new frontiers! The Samy phage tool targets the chromosome’s farthest terminal regions, rich in specialized metabolite genes, enabling precise integration across strains. A breakthrough for antibiotic production like albonoursin.

Phages are a valuable resource for the genetic engineering of Streptomyces antibiotic-producing bacteria. Indeed, a few integrative vectors based on phage integrase are available to insert transgene at specific genomic loci. Chromosome conformation captures previously demonstrated that the Streptomyces linear chromosome is organized in two spatial compartments: The central compartment encompassing most conserved and highly expressed genes in exponential phase, and the terminal compartments enriched in poorly conserved sequences including specialized metabolite biosynthetic gene clusters. This study introduces a new integrative tool based on a recently described phage, Samy, which specifically targets the terminal compartment of its native host chromosome. Samy is related to PhiC31 phage and, like the latter, encodes a serine integrase. Whereas PhiC31 targets a site generally located near the origin of replication, the Samy integration site is one of the farthest known attB sites from it. The authors demonstrated that the Samy integrase efficiently mediates the specific integration of a non-replicating plasmid in six Streptomyces strains from distinct clades. Bioinformatic analyses revealed that the Samy-attB site is rather conserved, and located in the terminal compartment of most Streptomyces chromosomes. Finally, heterologous expression of the albonoursin biosynthetic gene cluster from the Samy-, PhiC31-, and R4-attB sites yields quantitatively equivalent levels of production, though qualitative differences were observed. Altogether, these results demonstrate that the att-int Samy system expands Streptomyces genetic engineering tools by enabling targeted integration in the terminal chromosomal compartment.

More information: https://doi.org/10.1007/s00253-026-13707-2

Contact: Stéphanie BURY-MONE stephanie.bury-mone@i2bc.paris-saclay.fr

New discovery: a dynamic interplay between axis proteins and cohesins ensures chromosome stability during meiosis in Sordaria.

Faithful chromosome segregation during meiosis requires the coordinated action of cohesin complexes and chromosome axis proteins. How these factors interact and communicate along chromosome axes, especially during meiotic prophase I, remains however, only partially understood. Researchers of the Genome Biology Department of the I2BC investigated the functional interplay between the cohesin components and regulators (Rad21, Rec8, Wapl, Sororin, Spo76/Pds5) and two meiosis-specific axis proteins Red1 and Hop1. Analysis of multiple combinations of their corresponding null mutants and of their genetic-epistasis interactions in the fungus Sordaria macrospora revealed a hierarchical regulatory network for their recruitment and releasing. Their work uncovers an unexpected role of axis proteins Red1 and Hop1, that together with Sororin, provide stage-specific protection of Spo76/Pds5 against Wapl-mediated release. Furthermore, we identify that Spo76/Pds5 is the main target of Wapl and acts as a central guardian of kleisin stability against Slx8/STUbL-dependent proteasomal degradation.

Together, these findings show that a dynamic crosstalk between axis proteins and cohesins is crucial to preserve axis integrity and to ensure accurate meiotic progression.

More information: https://doi.org/10.1371/journal.pgen.1012001

Contact: Stéphanie BOISNARD stephanie.boisnard@i2bc.paris-saclay.fr

Using functional genomics in sorted Salmonella populations and high-resolution microscopy, the researchers of the I2BC show that the expression of Pathogenicity Island 1 is associated with chromatin remodeling and with the repositioning of this region toward the nucleoid periphery.

Chromatin provides a universal framework for organizing and regulating genomes across the three domains of life. In bacteria, it is composed of intrinsically supercoiled DNA associated with small DNA-binding proteins known as nucleoid-associated proteins (NAPs). Their binding can induce DNA bending, bridging, coating, and/or wrapping, giving rise to distinct modes of chromatin organization.
Bacterial chromatin can exist in a repressed state (silent chromatin) or in an actively transcribed state (active chromatin). Silent chromatin is largely associated with H-NS, a xenogeneic silencer that restricts the costly expression of genes acquired by horizontal transfer. In contrast, active chromatin is densely occupied by RNA polymerase and is characterized by different levels of DNA supercoiling. However, the changes in protein occupancy and chromatin organization that accompany transitions between these two states remain poorly understood.
Researchers of the Genome Biology Department of the I2BC in collaboration with the NGS and the Imaging facility of the I2BC and the Trinity College Dublin (Ireland), have unveiled the chromatin organization of horizontally acquired regions in Salmonella enterica serovar Typhimurium, which are essential for its pathogenicity. They show that expression of Pathogenicity Island 1 (SPI-1) is accompanied by local chromatin remodeling, marked by profound changes in three-dimensional organization and protein occupancy. This remodeling is also associated with the repositioning of SPI-1 toward the nucleoid periphery.
These findings provide new insights into the interplay between xenogeneic silencing, counter-silencing mechanisms, chromatin architecture, and the evolutionary integration of acquired DNA. They also reveal a finely tuned chromatin remodeling process that minimizes the cellular cost of activating pathogenicity islands, and they establish a direct link between the linear (1D) organization of the genome and its three-dimensional (3D) folding.

More information: https://www.nature.com/articles/s41467-025-67746-w

https://www.insb.cnrs.fr/fr/cnrsinfo/comment-les-genes-de-virulence-reorganisent-le-genome-de-salmonella-0

Contact: Vicky Lioy virginia.lioy@i2bc.paris-saclay.fr

Tight regulation of Fe-S clusters biosynthesis via a mutually antagonistic binding of frataxin and ferredoxin-2 to the assembly machinery, with several important implications for the Friedreich’s ataxia disease caused by frataxin deficiency.

Iron-sulfur (Fe-S) clusters are essential metallocofactors that perform a multitude of biological functions. They are synthesized de novo by multi-proteins machienries and any defect in their synthesis leads to severe diseases such as Friedreich’s ataxia (FRDA), caused by defective expression of frataxin (FXN). Here, we uncover that efficient [2Fe-2S] cluster assembly requires a fine-tuned balanced ratio of FXN and Ferredoxin-2 (FDX2), an essential enzyme of the assembly process. [2Fe-2S] clusters are assembled on the scaffold protein ISCU2 with sulfur provided as a persulfide by NFS1, which is cleaved into sulfide by FDX2. FXN stimulates the whole process by accelerating persulfide transfer to ISCU2. Using an in vitro reconstituted human system, we show that any deviation from a close-to-equal amount of FXN or FDX2 downregulates Fe-S cluster synthesis. We performed a structure-function investigation, which revealed that this is due to competition between FXN and FDX2 for the same binding site on the NFS1-ISCU2 complex. We found that higher levels of FXN impair the persulfide-reductase activity of FDX2 and higher levels of FDX2 slow FXN-accelerated persulfide transfer to ISCU2. We also discovered that FDX2 directly hinders persulfide generation and transfer to ISCU2 by interacting with the persulfide-carrying mobile loop of NFS1. We further found that knocking-down FDX2 expression in a FRDA drosophila model, increases fly lifespan. Altogether, this work highlights a direct regulation of Fe-S cluster biosynthesis through antagonistic binding of FXN and FDX2 and suggests that decreasing FDX2 in the context of FXN deficiency in FRDA might constitute a novel therapeutic axis.

More information: https://www.nature.com/articles/s41586-025-09822-1

Contact : Benoit D'Autréaux benoit.dautreaux@i2bc.paris-saclay.fr

Plants that hyperaccumulate nickel open the possibility to mine this metal from soils in an environmetally friendly manner. In this study, sequencing  of a nickel hyperaccumulating plant together with genomic and transcriptomic comparisons reveal the molecular mechanisms underlying this extreme trait.

Nickel hyperaccumulation is an extreme adaptation to ultramafic soils observed in more than 500 plant species. However, our understanding of the molecular mechanisms underlying the evolution of this trait remains limited. To shed light on these mechanisms, we have generated a high-quality genome assembly of the metal hyperaccumulator Noccaea caerulescens. We then used this genome as reference to conduct comparative intraspecific and interspecific transcriptomic analyses using various accessions of N. caerulescens and the non-accumulating relative Microthlaspi perfoliatum to identify genes associated with nickel hyperaccumulation.
Our results suggest a correlation between nickel hyperaccumulation and a decrease in the expression of genes involved in defense responses and the regulation of membrane trafficking. Surprisingly, these analyses did not reveal a significant enrichment of genes involved in the regulation of metal homeostasis. However, we found that the expression levels of selected metal transporter genes,
namely, NcHMA3, NcHMA4, and NcIREG2, are consistently elevated in N. caerulescens accessions hyperaccumulating nickel.
Furthermore, our analyses identified frameshift mutations in NcIRT1 associated with the loss of nickel hyperaccumulation in a few accessions. We further showed that the expression of a functional NcIRT1 in the roots of the La Calamine accession increases nickel accumulation in shoots. Our results demonstrate that NcIRT1 participates in nickel hyperaccumulation in N. caerulescens. They also
suggest that nickel hyperaccumulation is an ancient trait in N. caerulescens that has evolved from the high and constitutive expression of several metal transporters, including NcIREG2, and that the trait was subsequently lost in a few accessions due to mutations in NcIRT1.

More information: https://doi.org/10.1093/plcell/koaf281

Contacts:

Sylvain Merlot, sylvain.merlot@univ-tlse3.fr

Sébastien Thomine, sebastien.thomine@i2bc.paris-saclay.fr

Hijacking mechanism of membrane undecaprenyl phosphate recycling protein BacA in Enterococcus faecalis by a two-peptide bacteriocin revealed

Antibiotic-resistance is a critical global health concern stressing the urgent need for new therapeutic strategies beyond conventional drugs. In this context, peptide-based bacteriocins constitute potential medical use antibacterial alternatives. In the present study, we have highlighted the molecular mechanism of a two-peptide bacteriocin, enterocin C, with potent activity against Enterococcus faecalis, a major opportunistic pathogen notorious for multidrug resistance. In collaboration with a group from INRAE MICALIS, the team has uncovered how enterocin C specifically exploits the membrane-embedded protein BacA as a cell surface receptor. BacA, widely conserved across bacteria, plays a central role in bacterial cell-wall biogenesis through its dual phosphatase and flippase activities, which are essential for recycling the lipid carrier undecaprenyl phosphate.
Using a combination of biochemical, biophysical, and microbiological approaches supported by AlphaFold structural modelling, we dissected the cooperative action of the two enterocin C peptides. Acting at nanomolar concentrations, peptide EntC1 inserts deeply into BacA’s outward-open catalytic pocket, blocking its enzymatic function and facilitating the subsequent binding of peptide EntC2. This dual docking event anchors the bacteriocin deep within the membrane’s hydrophobic core, ultimately triggering membrane disruption and bacterial cell death.
The findings reveal the molecular determinants of this precision targeting and represent the first detailed mechanistic description of a two-peptide bacteriocin’s mode of action. This work identifies BacA as a valuable target for bacteriocin-mediated killing and open avenues for the rational design of peptide-based antimicrobials for tailor-made antimicrobials to help combat antibiotic-resistant infections.

More information :

Thierry Touzé :thierry.touze@i2bc.paris-saclay.fr

https://pubmed.ncbi.nlm.nih.gov/41232670/

A new ED-TA method which enabled genomics investigation of plasmid establishment during conjugative transfer was developed by I2BC researchers. They showed robust induction of a subset of plasmid genes at the early stages of conjugation through single-stranded DNA promoters.

Bacterial conjugation is a principal mode of horizontal gene transfer which has important life science implications including bacterial genome evolution, dissemination of genetic traits and bioengineering applications. Notably, the spread of multidrug resistance via conjugative plasmids is one of the biggest concerns in global public health. Although conjugation has been studied since 1940s and the overall procedure is widely known and well documented, molecular details of reactions that establishes a transferred plasmid in the new host cell remain elusive. In addition, there are specific regulatory mechanisms for temporal expression of plasmid genes, that are also crucial for successful conjugation as they allow timely expression and function of plasmid-encoded arsenal against host defense mechanisms.

Genomics-based studies of plasmid establishment were previously hampered by the nature of conjugation which takes place within a mixture of cell populations. Essentially, they require the separation of subpopulations before the DNA/RNA extraction to avoid contamination of indistinguishable plasmid DNA/RNA from the donor. Researchers of the Genome Biology Department of the I2BC, described the development of a new method, called ED-TA, which exploits a donor mutant hypersensitive to hypoosmotic shock. ED-TA allows unprecedently quick and efficient « Elimination of Donor population for Transconjugant Analysis». Using a clinically relevant model multidrug resistant conjugative plasmid, pESBL, they elucidated its transcription profile in successful and abortive conjugation. Researchers also showed single-stranded DNA promoters allow robust induction of a subset of genes at the early stages of conjugation. As the ED-TA method is straightforward and broadly applicable, further research taking advantage of the method will shed light on important molecular mechanisms of plasmid establishment after conjugation.

More information: doi.org/10.1093/nar/gkaf1299

https://www.insb.cnrs.fr/fr/cnrsinfo/une-nouvelle-methode-pour-suivre-de-pres-la-transmission-des-genes-chez-les-bacteries

Contact: Yoshiharu Yamaichi yoshiharu.yamaichi@i2bc.paris-saclay.fr

Despite having low homology with BRCA2 proteins from other organisms and no folded domain, the newly discovered BRCA2 protein from Physcomitrium patens (the green yeast) promotes homologous recombination by binding to recombinases and DNA.

BRCA2 is crucial for mediating homology-directed DNA repair (HDR) through its binding to single-stranded DNA (ssDNA) and the recombinases RAD51 and DMC1. Most BRCA2 orthologs have a canonical DNA-binding domain (DBD) with the exception of Drosophila melanogaster. It remains unclear whether such a noncanonical BRCA2 variant without DBD possesses a DNA-binding activity. Here, we identify a new noncanonical BRCA2 in the model plant Physcomitrium patens (PpBRCA2). We establish that PpBRCA2 is essential for genome integrity maintenance, somatic DNA double-strand break (DSB) repair, HDR-mediated gene targeting, and RAD51 foci recruitment at DNA break sites. PpBRCA2 is also critical for DSB repair during meiosis. Interestingly, PpBRCA2 interacts strongly with RAD51 but weakly with DMC1, suggesting a distinct meiotic function compared to other BRCA2 homologs. Despite lacking the canonical DBD, PpBRCA2 binds ssDNA through its disordered N-terminal region and efficiently promotes HDR. Our work highlights that the ssDNA binding capacity of BRCA2 homologs is conserved regardless of the presence of a canonical DBD and provides a deeper understanding of BRCA2’s functional diversity across species.

More information:

https://pmc.ncbi.nlm.nih.gov/articles/PMC12412785/

Contact: Sophie Zinn sophie.zinn@i2bc.paris-saclay.fr