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Students from CentraleSupélec visit I2BC groups of the Cellular Biology department
80 students of the Bachelor of Global Engineering, a joint novel program of CentraleSupéléc and McGilles University visited the research groups Kühl, Sala and Siudeja at the Cellular Biology department at I2BC in March. During two afternoons in March three groups from the Cellular Biology department, Kühl, Sala and Siudeja, explained their research questions and model organisms to a total of 80 students from the Bachelor of Global Engineering (BoGe), a joint novel program of CentraleSupéléc, Paris-Saclay and McGilles University, Canada (https://www.centralesupelec.fr/en/bachelor-global-engineering) . The visit was part of the BoGe Biology course taught by Sébastien Bloyer (Genome dep.) and Inge Kühl (BioCell dep.). We thank all the participants for their help, and the I2BC for the support.
Plant nucleoside N-ribohydrolases: riboside binding and role in nitrogen storage mobilization
Plants can benefit from fast nucleoside breakdown Cells save their energy during nitrogen starvation by selective autophagy of ribosomes and degradation of RNA to ribonucleotides and nucleosides. Nucleosides are hydrolyzed by nucleoside N-ribohydrolases (nucleosidases, NRHs). Subclass I of NRHs preferentially hydrolyzes the purine ribosides while subclass II is more active towards uridine and xanthosine. Here, we performed a crystallographic and kinetic study to shed light on nucleoside preferences among plant NRHs followed by in vivo metabolomic and phenotyping analyses to reveal the consequences of enhanced nucleoside break-down. We report the crystal structure of Zea mays NRH2b (subclass II) and NRH3 (subclass I) in complexes with the substrate analog forodesine. Purine and pyrimidine catabolism are inseparable because nucleobase binding in the active site of ZmNRH is mediated via a water network and is thus unspecific. Dexamethasone-inducible ZmNRH overexpressor lines of Arabidopsis thaliana, as well as double nrh knockout lines of moss Physcomitrium patents, reveal a fine control of adenosine in contrast to other ribosides. ZmNRH overexpressor lines display an accelerated early vegetative phase including faster root and rosette growth upon nitrogen starvation or osmotic stress. Moreover, the lines enter the bolting and flowering phase much earlier. We observe changes in the pathways related to nitrogen-containing compounds such as β-alanine and several polyamines, which allow plants to reprogram their metabolism to escape stress. Taken together, crop plant breeding targetting enhanced NRH-mediated nitrogen recycling could therefore be a strategy to enhance plant growth tolerance and productivity under adverse growth conditions. more information: https://onlinelibrary.wiley.com/doi/10.1111/tpj.16572 contact: Solange Morera solange.morera@i2bc.paris-saclay.fr
Male manipulation impinges on social-dependent tumor suppression in Drosophila melanogaster females
This study shows that tumorous Drosophila females perceive a cognitive-like signal from their social environment that acts on tumor growth, while this effect is lost in mated females because of peptides transmitted through the male ejaculate. Physiological status can influence social behavior, which reciprocally can impinge on physiology and health. Previously, we reported that tumor growth in Drosophila virgin females depends on the social context, but did not investigate the underlying physiological mechanisms. Here, we sought to characterize the signal perceived between tumorous flies, ultimately discovering that the tumor suppressive effect varies depending on reproductive status. Firstly, we show that the tumor suppressive effect is neither dependent on remnant pheromone-like products nor on the microbiota. Transcriptome analysis of the heads of these tumorous flies reveals social-dependent gene-expression changes related to nervous-system activity, suggesting that a cognitive-like relay might mediate the tumor suppressive effect. The transcriptome also reveals changes in the expression of genes related to mating behavior. Surprisingly, we observed that this social-dependent tumor-suppressive effect is lost in fertilized females. After mating, Drosophila females change their behavior ─favoring offspring survival─ in response to peptides transferred via the male ejaculate, a pehnomenon called “male manipulation”. Remarkably, the social-dependent tumor suppressive effect is restored in females mated by sex-peptide deficient males. Since male manipulation has likely been selected to favor male gene transmission, our findings indicate that this evolutionary trait impedes social-dependent tumor growth slowdown. more information: https://doi.org/10.1038/s41598-024-57003-3 contact: Jacques MONTAGNE <jacques.montagne@i2bc.paris-saclay.fr>
Catalytic Mechanism of Fatty Acid Photodecarboxylase: on the Detection and Stability of the Initial Carbonyloxy Radical Intermediate
Quality trumps quantity: photoenzymatic decarboxylation of fatty acids does indeed occur in 300 ps. Despite the dismissal by our "slightly overproductive" competitors, citing it as 30x slower, our original findings stand strong! In fatty acid photodecarboxylase (FAP), light-induced formation of the primary radical product RCOO● from fatty acid RCOO– occurs in 300 ps, upon which CO2 is released quasi-immediately. Based on the hypothesis that aliphatic RCOO●(spectroscopically uncharacterized because unstable) absorbs in the red similarly to aromatic carbonyloxy radicals such as 2,6-dichlorobenzoyloxy radical (DCB●), much longer-lived linear RCOO● has been suggested recently. We performed quantum chemical reaction pathway and spectral calculations. Thesecalculations are in line with the experimental DCB● decarboxylation dynamics and spectral properties and show that in contrast to DCB●, aliphatic RCOO● radicals a) decarboxylate with a very low energetic barrier and on the timescale of a few ps and b) exhibit little red absorption. A time-resolved infrared spectroscopy experiment confirms very rapid, <<300 ps RCOO● decarboxylation in FAP. We argue that this property is required for the observed high quantum yield of hydrocarbons formation by FAP. more information: https://onlinelibrary.wiley.com/doi/10.1002/anie.202401376 contact: Pavel MULLER <pavel.muller@i2bc.paris-saclay.fr>
Transcription-induced DNA supercoiling clears RNA polymerase’s path in bacterial chromatin
Positive DNA supercoiling generated ahead of RNA polymerase during transcription elongation dislodges nucleoid-structuring protein H-NS from the DNA. Bacterial chromosomal DNA is condensed more than 1000 times inside the cell. This condensation results from a slight underwinding of the double helix, leading to the formation of loops of negatively supercoiled DNA (plectonemes) and from interactions with structural proteins that help compact these loops. However, these chromatin-associated proteins obstruct transcription and need to be transiently removed to allow gene expression. This is the case for H-NS, an abundant protein that blocks the expression of numerous genes. In Gram-negative pathogenic bacteria such as Salmonella, H-NS represses the expression of the majority of virulence genes. Displacement of H-NS becomes necessary during the infection process, which relies on the function of virulence gene products.
In a study published in the journal Nature Communications, researchers from the Genome Biology department of the Institute for Integrative Biology of the Cell (I2BC) present evidence that binding of H-NS to DNA is destabilized, from a distance, by the positive supercoils forming ahead of approaching RNA polymerase. It is known that the process of transcription induces DNA supercoiling by forcing the rotation of the DNA axis during elongation. To explain the destabilization of H-NS:DNA complexes by positive supercoiling, the researchers propose a model in which H-NS oligomers play a scaffolding role in negatively supercoiled DNA by bridging opposite arms of the plectonemic structure. Accumulation of positive supercoils and concomitant DNA axis rotation ahead of the approaching RNA polymerase cause the H-NS-bound, negatively supercoiled plectoneme to unroll, leading to the collapse of the scaffold and allowing RNA polymerase to continue on its path. More information: https://www.nature.com/articles/s41467-024-47114-w Contact: Lionelo Bossi lionello.bossi@i2bc.paris-saclay.fr
A highly conserved ligand-binding site for AccA transporters of antibiotic and quorum-sensing regulator in Agrobacterium leads to a different specificity
The highly conserved ligand binding site of the AccA transporters of antibiotic and quorum-sensing regulator in Agrobacterium is not linked to a conserved specificity. Plants genetically modified by the pathogenic Agrobacterium strain C58 synthesize agrocinopines A and B, whereas those modified by the pathogenic strain Bo542 produce agrocinopines C and D. The four agrocinopines (A, B, C and D) serve as nutrients by agrobacteria and signaling molecule for the dissemination of virulence genes. They share the uncommon pyranose-2-phosphate motif, represented by the L-arabinopyranose moiety in agrocinopines A/B and the D-glucopyranose moiety in agrocinopines C/D, also found in the antibiotic agrocin 84. They are imported into agrobacterial cytoplasm via the Acc transport system, including the solute-binding protein AccA coupled to an ABC transporter. We have previously shown that unexpectedly, AccA from strain C58 (AccAC58) recognizes the pyranose-2-phosphate motif present in all four agrocinopines and agrocin 84, meaning that strain C58 is able to import agrocinopines C/D, originating from the competitor strain Bo542. Here, using agrocinopine derivatives and combining crystallography, affinity and stability measurements, modeling, molecular dynamics, in vitro and vivo assays, we show that AccABo542 and AccAC58 behave differently despite 75% sequence identity and a nearly identical ligand binding site. Indeed, strain Bo542 imports only compounds containing the D-glucopyranose-2-phosphate moiety, and with a lower affinity compared to strain C58. This difference in import efficiency makes C58 more competitive than Bo542 in culture media. We can now explain why Agrobacterium/Allorhizobium vitis strain S4 is insensitive to agrocin 84, although its genome contains a conserved Acc transport system. Overall, our work highlights AccA proteins as a case study, for which stability and dynamics drive specificity. More information: https://portlandpress.com/biochemj/article-abstract/481/2/93/233802/A-highly-conserved-ligand-binding-site-for-AccA?redirectedFrom=fulltext Contact: Solange Morera solange.morera@i2bc.paris-saclay.fr
Members of gut microbiota prime insect immunity
Burkholderia bacteria from the gut microbiota in the bean bug insect can cross the epithelium of the gut and trigger a protective systemic immune response that has no adverse effects on the insect fitness but confers a general protection against pathogens. Insects lack acquired immunity and were thought to have no immune memory, but recent studies reported a phenomenon called immune priming, wherein sub-lethal dose of pathogens or non-pathogenic microbes stimulate immunity and prevent subsequential pathogen infection. Although the evidence for insect immune priming is accumulating, the underlying mechanisms are still unclear. The bean bug Riptortus pedestris acquires its gut microbiota from ambient soil and spatially structures them into a multispecies and variable community in the anterior midgut and a specific, mono-species Caballeronia symbiont population in the posterior region. We demonstrate that some particular Burkholderia strains colonizing the anterior midgut, stimulate systemic immunity by penetrating gut epithelia and migrating into the hemolymph. This hemolymph colonization and activated immunity, consisting of a humoral and a cellular response, has no negative effect on the host fitness, but on the contrary protected the insect from subsequent infection by pathogenic bacteria. Interruption of contact between the Burkholderia and epithelia of the gut weakened the host immunity back to pre-infection levels and made the insects again vulnerable to microbial infection, demonstrating that persistent acquisition of environmental bacteria is important to maintain an efficient immunity. This suggests that priming is only activated when it might be most helpful, e.g. when pathogen encounters are most likely in a microbially rich environment. Together, these findings not only highlight a role of environmental microbes in shaping insect immunity but also put a new, symbiotic perspective on bacterial intestinal barrier breaching and hemolymph colonization, which has been generally viewed only as a pathogenic phenomenon. More information: https://doi.org/10.1073/pnas.2315540121 Contact : Peter MERGAERT <peter.mergaert@i2bc.paris-saclay.fr>
JAK-STAT-dependent contact between follicle cells and the oocyte controls Drosophila anterior-posterior polarity and germline development
The authors identified a population of somatic cells in Drosophila follicles that elaborate filopodia penetrating the oocyte, thereby maintaining the contact between the two tissues. This is essential to control polarity and germline development of the future embryo. The number of embryonic primordial germ cells is determined, in Drosophila, by the quantity of germ plasm, whose assembly starts in the posterior region of the oocyte during oogenesis. Here, we report that extending JAK-STAT activity in the posterior somatic follicular epithelium leads to an excess of primordial germ cells in the future embryo. We show that JAK-STAT signaling is necessary for the differentiation of approximately 20 specialized follicle cells maintaining tight contact with the oocyte. These cells define, in the underlying posterior oocyte cortex, the anchoring of the germ cell determinant oskar mRNA. We reveal that the apical surface of these posterior anchoring cells extends long filopodia penetrating the oocyte. We identify two JAK-STAT targets in these cells that are each sufficient to extend the zone of contact with the oocyte, thereby leading to production of extra primordial germ cells. JAK-STAT signaling thus determines a fixed number of posterior anchoring cells required for anterior-posterior oocyte polarity and for the development of the future germline. More information: https://doi-org.insb.bib.cnrs.fr/10.1038/s41467-024-45963-z Contact: Marianne MALARTRE <marianne.malartre@i2bc.paris-saclay.fr>
Activity of MukBEF for chromosome management in E. coli and its inhibition by MatP
The MukBEF condensin of E. coli loads at the replication fork and is unloaded at the Ter region by MatP. Structural maintenance of chromosomes (SMC) complexes share conserved structures and serve a common role in maintaining chromosome architecture. In the bacterium Escherichia coli, the SMC complex MukBEF is necessary for rapid growth and the accurate segregation and positioning of the chromosome, although the specific molecular mechanisms involved are still unknown. Here, we used a number of in vivo assays to reveal how MukBEF controls chromosome conformation and how the MatP/matS system prevents MukBEF activity. Our results indicate that the loading of MukBEF occurs preferentially on newly replicated DNA, at multiple loci on the chromosome where it can promote long-range contacts in cis even though MukBEF can promote long-range contacts in the absence of replication. Using Hi-C and ChIP-seq analyses in strains with rearranged chromosomes, the prevention of MukBEF activity increases with the number of matS sites and this effect likely results from the unloading of MukBEF by MatP. Altogether, our results reveal how MukBEF operates to control chromosome folding and segregation in E. coli. More information: https://elifesciences.org/articles/91185 Contact: Stéphane DUIGOU <stephane.duigou@i2bc.paris-saclay.fr>
"Jeudis de la Recherche" at I2BC
As part of the "Jeudis de la Recherche" organized by the COMPAS (Paris-Saclay University), the CNRS and the town of Gif sur Yvette, Sylvie Lautru's team (I2BC, Microbiology Department) welcomed around twenty people to take part in 5 workshops on the theme of "Antibiotics and the microbial response: a never-ending battle". Thanks to the very interactive format, participants who were very interested in this topical issue, asked many questions and exchanged a lot with the research team. https://www.ville-gif.fr/215/que-faire-a-gif/culture/conferences/jeudis-de-la-recherche.htm
EMBO conference "Molecular Biology of Archaea" 23-27th of june 2024 in Palaiseau (near Paris)!
The registration in now open for the next EMBO conference "Molecular Biology of Archaea" that will take place near Paris (Palaiseau) on 23-27th of june 2024! Archaea are fascinating organisms with a bacteria-like morphology, but information-processing systems that are homologous to eukaryotic counterparts, for instance replication, transcription and translation. Recently, novel groups of archaea (e.g. Asgard archaea) have been discovered that represent the closest living relatives to eukaryotes and shed new light on their evolution. Archaeal research has led to seminal change-of-concept mechanistic discoveries in the area of molecular biology, and many studies are currently aiming to unravel the global environmental impact of archaea. Their viruses have a remarkable diversity of morphotypes, exceeding that of bacterial or eukaryotic viruses.The EMBO Workshop on Molecular biology of Archaea will cover cutting-edge research in various fields with a focus on archaeal molecular biology, evolution, and ecology and their interconnections. The proposed talks will present data at different scales ranging from single molecule and structural studies to archaeal physiology, metabolism and ecological impact. This is of high interest as molecular studies of archaea are a very active field, reflecting the wealth of metagenomics information revealing novel biology that can now be addressed by the latest experimental and computational techniques. all information: https://meetings.embo.org/event/24-archaea contact: Tamara BASTA-LE BERRE <tamara.basta@i2bc.paris-saclay.fr>
Transposon sequencing reveals genes enabling insect gut colonization by the symbiont Caballeronia insecticola
High-througput genetic screens with Tn-seq in an insect gut bacterium reveals gut-derived nutrients consumed by the symbiont. Caballeronia insecticola is a bacterium belonging to the Burkholderia sensu lato, able to colonize multiple environments like soils and the gut of the bean bug Riptortus pedestris. We constructed a saturated Himar1 mariner transposon library and revealed by transposon-sequencing (Tn-seq) that 498 protein-coding genes constitute the essential genome of C. insecticola for growth in free-living conditions. By comparing essential gene sets of C. insecticola and seven related Burkholderia s.l. strains, only 120 common genes were identified indicating that a large part of the essential genome is strain-specific. In order to reproduce specific nutritional conditions that are present in the gut of R. pedestris, we grew the mutant library in minimal media supplemented with candidate gut nutrients and identified several condition-dependent fitness-defect genes by Tn-seq. To validate the robustness of the approach, insertion mutants in six fitness genes were constructed and their growth-deficiency in media supplemented with the corresponding nutrient was confirmed. The mutants were further tested for their efficiency in R. pedestris gut colonization, confirming that gluconeogenic carbon sources, taurine and inositol, are nutrients consumed by the symbiont in the gut. Thus, our study provides insights about specific contributions provided by the insect host to the bacterial symbiont. More information: https://doi.org/10.1093/ismeco/ycad001 Contact: Peter MERGAERT <peter.mergaert@i2bc.paris-saclay.fr>
The stringent response is strongly activated in the high antibiotic producer, Streptomyces coelicolor
The stringent response controls positively antibiotic biosynthesis. Antibiotics are thus part of the stringent response. In most bacteria, the stringent response (SR) was initially characterized as a response to nitrogen (N) limitation resulting into the depletion of aminoacylated tRNAs leading to the stalling of ribosomes on mRNA. The recruitment of the ppGpp synthetase, RelA, at the stalled ribosomes activates (p)ppGpp synthesis from GTP. ppGpp that is the mediator of the SR controls negatively, at the transcriptional and translational levels, the expression of most ribosomal proteins leading to the down regulation of the translational process and thus of growth.
The model strains Streptomyces coelicolor (SC) and Streptomyces lividans (SL), strong and weak producers of the same antibiotics, respectively, were grown in condition of phosphate (Pi) limitation or proficiency and the abundance of proteins of their translational apparatus was compared. This study revealed that the expression of RelA was induced in Pi limitation suggesting that, besides N limitation, Pi limitation also contributes to the triggering of the SR. Interestingly, most proteins of the translational apparatus had a similar or slightly higher abundance in SL than in SC, in Pi limitation whereas most of these proteins were far more abundant in SL than in SC, in Pi proficiency. This indicated an alleviation of the SR in Pi proficiency in SL, but not in SC. This suggested an alteration of Pi up-take and/or Pi-mediated regulation in SC whose molecular basis remain to be elucidated.
Interestingly, the production of specialized metabolites in SC (CDA, RED and ACT) is usually concomitant of phases of growth slow down and it is known that ppGpp controls positively the expression of their biosynthetic pathways. Their production could thus be considered as part of the SR. Indeed, these metabolites were proposed to regulate negatively, through different processes, the energetic metabolism and thus the generation of ATP, in SC, a process that might contribute to the slower growth rate of SC compared to SL. More information: https://www.sciencedirect.com/science/article/pii/S0923250823001547?via%3Dihub Contact: Marie-Joëlle VIROLLE <marie-joelle.virolle@i2bc.paris-saclay.fr>
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Physics and Biological Systems 2024
Mark your calendars for Jun. 26-28, 2024! Join us at Ecole Polytechnique in Palaiseau for the 7th International Conference on Physics and Biological Systems, where top-notch scientists converge to explore the dynamic interplay between physics and life sciences. The 7th International Conference on Physics and Biological Systems will be held on Jun. 26-28 2024 at Ecole Polytechnique in Palaiseau, in the south of Paris. It aims to bring together a broad range of physical and life scientists working at the interface between the two disciplines around in-depth talks by first-rate international speakers. Attendance will be limited to 200 participants. We look forward to welcoming you in Palaiseau! more information: https://www.lptms.universite-paris-saclay.fr/physbio2024/ contact: Pavel MULLER <pavel.muller@i2bc.paris-saclay.fr>
How does DNA methylation, an epigenetic modification involved in cancer and disease, change the folding of our chromosomes?
In this literature review in Nature Structural and Molecular Biology, scientists from the Institute Jacques Monod and the I2BC discuss how the methylation of DNA changes the folding of our chromosomes and how this can cause defects and disease. DNA methylation is used as an epigenetic mark on human chromosomes to regulate the transcriptional activity of genes and parasitic transposable elements. Within the nucleus of human cells, chromosomes are non-randomly organized, which strongly influences transcriptional activity as well. DNA methylation and 3D chromosome organization are strongly reorganized during the earliest stages of embryonic development and in diseases like cancer. In this literature review, published in Nature Structural and Molecular Biology, scientists from the Institute Jacques Monod and the I2BC discuss the links between the two levels of chromosome organization. The authors particularly focus on the CTCF insulator protein, whose binding to the DNA directly translates 3D chromosome organization to transcriptional activity. CTCF binding to the DNA has been reported as DNA methylation sensitive in certain contexts, which for certain genes explains the link between transcriptional activity, 3D chromosome organization and DNA methylation. Yet, changes in CTCF binding near other genes appears less obviously coupled to this epigenetic mark, indicating that other mechanisms must play a role as well. more information: https://www.nature.com/articles/s41594-024-01241-6 contact: Daan Noordermeer <daan.noordermeer@i2bc.paris-saclay.fr>
A high-end Glacios2 cryo-electron microscope at I2BC
A Glacios2 200 kV has been delivered to I2BC, heralding, together with a Titan Krios 300 kV at synchrotron SOLEIL, a new era for cryo-electron microscopy in the Paris-Saclay area Cryo-electron microscopy (cryo-EM) has become an indispensable tool in structural biology, enabling researchers to unravel the complexities of biological macromolecules and understand their roles in various cellular processes and diseases. These last ten years the field has undergone a revolution, with technical advances that have made atomic resolution reconstructions of large macromolecular complexes possible (the so-called 'resolution revolution'), I2BC received the latest generation 200kV microscope, Glacios2, on March 27, 2024. Joint operation is planned with the 300kV Titan Krios also just delivered at synchrotron SOLEIL. This will allow researchers to study structures of living machines at work in fine detail, both for purified and/or reconstituted complexes and in cells. These two cryo-electron microscopes are gathering scientists from all over the Paris-Saclay area, sparking further interest in future developments at the interface beween structural and cellular biology. contact: Stéphane Bressanelli (plt-cryoem@i2bc.paris-saclay.fr)
Uncoupling programmed DNA cleavage and repair scrambles the Paramecium somatic genome
No Ku – no cut: how the ciliate Paramecium tetraurelia protects its somatic genome from translocations and aberrant chromosome fragmentation during developmentally programmed rearrangements. DNA double-strand breaks (DSBs) are a major threat to genome integrity. When incorrectly repaired, they can lead to genome rearrangements, chromosome instability, diseases, or cell death. Yet, key physiological processes, such as antibody gene assembly or meiotic recombination, rely on DSB induction. How living organisms cope with programmed DSBs is a major question in genome biology. The “Programmed genome rearrangements” team studies Paramecium to dissect developmentally programmed DNA elimination mechanisms. During each sexual cycle, tens of thousands of germline Internal Eliminated Sequences (IESs), scattered all along chromosomes, undergo precise excision. This process requires DSB introduction at IES boundaries, followed by error-free stitching of flanking DNA fragments by the non-homologous end joining (NHEJ) pathway. The team previously reported that the presence of a specialized variant of the NHEJ factor Ku is a pre-requisite for the activation of DNA cleavage, indicating that the two steps of the reaction are coupled. In this study, they engineered a DNA binding-deficient Ku mutant, which they characterized in collaboration with the IntGen team from the B3S department. They showed that the mutant Ku activates DNA cleavage, but is defective in repair. Unrepaired broken ends at IES boundaries are then trimmed and healed by telomere addition. By co-expressing wild-type Ku with the mutant, they were able to uncouple the cleavage and repair steps during IES excision. Using high throughput DNA sequencing and novel dedicated bioinformatic tools, they showed that chromosome translocations were largely favored under these conditions. This work demonstrates that coupling DNA cleavage and DSB repair ensures faithful genome assembly during programmed rearrangements. More information: https://www.cell.com/cell-reports/fulltext/S2211-1247(24)00329-2 Contact: Mireille Bétermier mireille.betermier@i2bc.paris-saclay.fr – Julien Bischerour – julien.bischerour@i2bc.paris-saclay.fr
A Fatty Acid Anabolic Pathway in Specialized-Cells Sustains a Remote Signal that Controls Egg Activation in Drosophila
While egg activation, i.e. oocyte to embryo transition, was not known to depend on physiological non-genital signal, this study shows that in Drosophila it depends on a very-long-chain-fatty-acid produced in the oenocytes (lipid metabolism specialized cells). Egg activation, representing the critical oocyte-to-embryo transition, provokes meiosis completion, modification of the vitelline membrane to prevent polyspermy, and translation of maternally provided mRNAs. This transition is triggered by a calcium signal induced by spermatozoon fertilization in most animal species, but not in insects. In Drosophila melanogaster, mature oocytes remain arrested at metaphase-I of meiosis and the calcium-dependent activation occurs while the oocyte moves through the genital tract. Here, we discovered that the oenocytes of fruitfly females are required for egg activation. Oenocytes, cells specialized in lipid-metabolism, are located beneath the abdominal cuticle. In adult flies, they synthesize the fatty acids (FAs) that are the precursors of cuticular hydrocarbons (CHCs), including pheromones. The oenocyte-targeted knockdown of a set of FA-anabolic enzymes, involved in very-long-chain fatty acid (VLCFA) synthesis, leads to a defect in egg activation. Given that some but not all of the identified enzymes are required for CHC/pheromone biogenesis, this putative VLCFA-dependent remote control may rely on an as-yet unidentified CHC or may function in parallel to CHC biogenesis. Additionally, we discovered that the most posterior ventral oenocyte cluster is in close proximity to the uterus. Since oocytes dissected from females deficient in this FA-anabolic pathway can be activated in vitro, this regulatory loop likely operates upstream of the calcium trigger. To our knowledge, our findings provide the first evidence that a physiological extra-genital signal remotely controls egg activation. Moreover, our study highlights a potential metabolic link between pheromone-mediated partner recognition and egg activation. More information: https://doi.org/10.1371/journal.pgen.1011186 Contact: Jacques Montagne jacques.montagne@i2bc.paris-saclay.fr
Epigenetics: combining flexible and rigid regions into a single structure to ensure genome replication and stability
The AMIG team at I2BC, in collaboration with teams from the Institut Curie and the Synchrotron Soleil, shows that the CAF-1 protein combines in its spatial organization flexible regions and rigid modules to deposit histones on DNA and effectively couple this process to DNA synthesis. Genome and epigenome integrity in eukaryotes depends on the proper coupling of histone deposition with DNA synthesis. This process relies on the evolutionary conserved histone chaperone CAF-1 for which the links between structure and functions are still a puzzle. While studies of the Saccharomyces cerevisiae CAF-1 complex enabled to propose a model for the histone deposition mechanism, we still lack a framework to demonstrate its generality and in particular, how its interaction with the polymerase accessory factor PCNA is operating. Here, we reconstituted a complete SpCAF-1 from fission yeast. We characterized its dynamic structure using NMR, SAXS and molecular modeling together with in vitro and in vivo functional studies on rationally designed interaction mutants. Importantly, we identify the unfolded nature of the acidic domain which folds up when binding to histones. We also show how the long KER helix mediates DNA binding and stimulates SpCAF-1 association with PCNA. Our study highlights how the organization of CAF-1 comprising both disordered regions and folded modules enables the dynamics of multiple interactions to promote synthesis-coupled histone deposition essential for its DNA replication, heterochromatin maintenance, and genome stability functions. More information: https://elifesciences.org/articles/91461 Contact: Françoise Ochsenbein francoise.ochsenbein@i2bc.paris-saclay.fr
Bettencourt-Schueller Foundation seminar on Art & Science: How does creativity emerge?
Chloé @Chl0e_Girard and Daan @DaanNoordermeer were invited to an interdisciplinary workshop organized by the Bettencourt-Schueller Foundation @Fondation_BS about creativity as motor for scientific research. A 3-days seminar was organized by the Bettencourt-Schueller Foundation at the Domaine de Chaumont-sur-Loire for former laureates (Prix Jeune Chercheur, Prix Impulsience, Prix Coups d’Elan pour la Recherche Française). This interdisciplinary seminar centered around creativity combined Arts & Science, the two main components of the foundation's patronage. The I2BC scientists participated in thematic workshops around Music, Mathematics, Visual Arts and Biology, presented by world-reknown artists and scientists. Daan Nordermeer was a recipient of the "Coup d'élan pour la recherche française”, while Chloé Girard received the "Prix jeune Chercheur", both in 2015.
Cooperation between two modes for DNA replication initiation in the archaeon Thermococcus barophilus
Demonstration that diverse physiological states influence the mode of DNA replication initiation in the archaeon Thermococcus. The mechanisms underpinning the replication of genomic DNA have recently been challenged in Archaea. Indeed, the lack of origin of replication has no deleterious effect on growth, suggesting that replication initiation relies on homologous recombination. Recombination-dependent replication (RDR) appears to be based on the recombinase RadA, which is of absolute requirement when no initiation origins are detected. The origin of this flexibility in the initiation of replication and the extent to which it is used in nature are yet to be understood. We combined deep sequencing and genetics to elucidate the dynamics of oriC utilization according to growth phases. We discovered that in Thermococcus barophilus, the use of oriC diminishes from the lag to the middle of the log phase, and subsequently increases gradually upon entering the stationary phase. Although oriC demonstrates no indispensability, RadA does exhibit essentiality. Notably, a knockdown mutant strain provides confirmation of the pivotal role of RadA in RDR for the first time. Thus, we demonstrate the existence of a tight combination between oriC utilization and homologous recombination to initiate DNA replication along the growth phases. Overall, this study demonstrates how diverse physiological states can influence the initiation of DNA replication, offering insights into how environmental sensing might impact this fundamental mechanism of life. More information: https://pubmed.ncbi.nlm.nih.gov/38421162/ Contact: Jacques OBERTO <jacques.oberto@i2bc.paris-saclay.fr>
A genome-wide comprehensive analysis of nucleosome positioning in yeast
We show that the regulation and compartmentalisation of nucleosomal organisation require the concomitant actions of local and global processes that are maintained actively by energy consuming factors. In eukaryotic cells, the one-dimensional DNA molecules need to be tightly packaged into the spatially constraining nucleus. Folding is achieved on its lowest level by wrapping the DNA around nucleosomes. Their arrangement regulates other nuclear processes, such as tran- scription and DNA repair. Despite strong efforts to study nucleosome positioning using Next Generation Sequencing (NGS) data, the mechanism of their collective arrangement along the gene body remains poorly understood. Here, we classify nucleosome distributions of protein-coding genes in Saccharomyces cerevisiae according to their profile similarity and analyse their differences using functional Principal Component Analysis. By decomposing the NGS signals into their main descriptive functions, we compared wild type and chromatin remodeler-deficient strains, keeping position-specific details preserved whilst considering the nucleosome arrangement as a whole. A correlation analysis with other genomic proper- ties, such as gene size and length of the upstream Nucleosome Depleted Region (NDR), identified key factors that influence the nucleosome distribution. We reveal that the RSC chromatin remodeler—which is responsible for NDR maintenance—is indispensable for decoupling nucleosome arrangement within the gene from positioning outside, which inter- fere in rsc8-depleted conditions. Moreover, nucleosome profiles in chd1Δ strains displayed a clear correlation with RNA polymerase II presence, whereas wild type cells did not indicate a noticeable interdependence. We propose that RSC is pivotal for global nucleosome orga- nisation, whilst Chd1 plays a key role for maintaining local arrangement. More information: https://journals.plos.org/ploscompbiol/article?id=10.1371/journal.pcbi.1011799 Contact: Arach GOLDAR <arach.goldar@i2bc.paris-saclay.fr>
Portrait Jeune Chercheuse - Cécile Sauvanet, Professeure junior en Biologie Structurale
Cécile Sauvanet est titulaire d’une Chaire de Professeur Junior du CNRS et est affectée à l’Institut de Biologie Intégrative de la Cellule - I2BC (CNRS/CEA/UPSaclay) à Gif-sur-Yvette. Ses recherches portent sur les mécanismes régulant le transport intracellulaire des organites en particulier les mitochondries. Elle utilise des approches de biologie intégrative alliant la biologie cellulaire, la biochimie et la cryo-microscopie électronique (Cryo-EM). Elle a effectué sa thèse de biochimie à l’Université de Bordeaux où son travail cherchait à caractériser la machinerie de fusion mitochondriale. Elle a pu montrer in vivo dans la levure Saccharomyces cerevisiae que des défauts bioénergétiques entrainaient l’inhibition spécifique de la fusion de la membrane mitochondriale interne. Elle a également caractérisé le rôle des mitofusines humaines dans la fusion mitochondriale grâce à des approches in vitro de biochimie et biophysique. Elle a ensuite poursuivi sa formation post-doctorale tout d’abord à l’université de Cornell (Ithaca, NY, USA) dans le laboratoire d’Anthony Bretscher. Ses recherches portaient sur les protéines responsables de la formation et de la régulation des microvillosités au niveau du site apical des cellules épithéliales. Elle a ensuite rejoint l'équipe de Francesca Giordano à l'I2BC, où elle a participé à l’étude des sites de contact membranaire entre le réticulum endoplasmique, les mitochondries et les gouttelettes lipidiques. Cécile a finalement rejoint le groupe de Dorit Hanein à l'Institut Pasteur où elle a concentré ses travaux sur le développement de méthodes de cryo-EM et cryo-tomographie qu’elle a utilisées pour étudier l'entrée du SARS-CoV-2 dans les cellules, les jonctions cellulaires ou encore la structure des protéines membranaires. Depuis Septembre 2023, elle a rejoint l’I2BC et l’équipe « Biochimie structurale des microtubules, des kinésines et leurs cargos » co-dirigée par Julie Ménétrey et Benoît Gigant. Elle y développe sa propre thématique de recherche sur les mécanismes régulant le transport des mitochondries dans les neurones. L’objectif est de comprendre comment les mitochondries sont positionnées au bon moment, au bon endroit et dans la bonne quantité pour répondre aux besoins de la cellule. « Je n'accepte plus les choses que je ne peux pas changer. Je change les choses que je ne peux pas accepter. » - Angela Davis Contact : cecile.sauvanet@i2bc.paris-saclay.fr
Via Life Sciences UPSaclay
Prize "Relève de l'étoile" for the excellence of the research work carried out by Alexia Royer
Alexia Royer received the prize "Relève de l'étoile" for her article “Clostridioides difficile S-Layer Protein A (SlpA) Serves as a General Phage Receptor” published in Microbiology Spectrum in February 2023. Antibiotics have been used for more than 80 years to treat bacterial infections, but bacteria have developed resistance to these treatments, making them difficult to eradicate. One of the side effects of antibiotics is that they tend to kill a broad spectrum of bacteria, including “good bacteria” important for the proper functioning of the human body. The resulting bacterial imbalance, called dysbiosis, will open the door to opportunistic bacteria such as Clostridioides difficile. This bacterium is the main cause of severe diarrhea in industrialized countries. As the infection is now treated with antibiotics, which are themselves responsible for the infection, many relapses are reported. In the laboratories of Professor Louis-Charles Fortier and Professor Olga Soutourina, we are interested in phage therapy, which is a treatment based on the use of bacteria-killing viruses called bacteriophages. Phages have the advantage of being specific and targeting a species or a few bacterial strains unlike antibiotics. Thus, they could be used as an alternative or complementary therapy to antibiotics to treat patients in therapeutic failure. The key step to set up a phage therapy is to understand the mechanism of recognition of the bacteria by phages. For a phage to attack and kill a bacterium, it must first attach to a receptor on its surface. In the article, we identified the SlpA protein as the receptor on the surface of the bacteria that is targeted by phages. We have shown that the infection specificity of phages is linked to the presence of different forms of SlpA, called isoforms, each strain expressing one isoform. We also identified a domain of SlpA, named D2, playing a role in receptor recognition by certain phages. These data are essential because they improve our understanding of phage/bacteria interactions and make phage therapy possible one day against C. difficile infections. https://frq.gouv.qc.ca/histoire-et-rapport/releve-etoile-jacques-genest-decembre-2023/ Alexia ROYER <alexia.royer@i2bc.paris-saclay.fr>
Protein-protein interactions: how to push forward the limits of the revolutionary AlphaFold2 programme?
AlphaFold2 is revolutionising protein structure prediction and structural biology practices. However, it may prove less effective for certain protein assemblies, particularly when they depend on intrinsically disordered regions. In an article published in Nature Communications, researchers from the I2BC show that applying a fragmentation strategy to the protein partners of such assemblies very significantly improves AlphaFold2’s prediction capacity. Mapping protein-protein interaction networks is essential for understanding the dynamics of cellular functions and their cross-regulation. Precise knowledge of interaction sites makes it possible to specifically perturb the proteins in these networks and understand the synergies and competitions that ensure cell function. Unfortunately, a great amount of structural information is still lacking to provide a detailed understanding of the organisation of interaction networks. The AlphaFold2 artificial intelligence programme has demonstrated a remarkable ability to predict the structures of protein assemblies that have co-evolved over long time scales. Its performance remained poorly characterised for assemblies involving intrinsically disordered regions, which often mediate transient and dynamic interactions during evolution. In a study published in the journal Nature Communications, researchers from the AMIG team at the Institut de Biologie Intégrative de la Cellule – I2BC (CNRS/CEA/UPSaclay, Gif-sur-Yvette) have shown that AlphaFold2 performs poorly if large disordered regions are used directly for prediction (40% success rate). A protein fragmentation strategy was found to be particularly well adapted to predicting the interfaces between folded domains and small protein motifs that fold on contact with the partner. Applied on a large scale using the Jean Zay HPC infrastructure on more than 900 complexes, this strategy achieved a success rate of almost 90%, a very encouraging result for the systematic screening of protein interaction networks. Nevertheless, the study calls for vigilance with regard to the risks of detecting false positives, which will be at the heart of future developments in artificial intelligence strategies such as AlphaFold2. More information: https://www.nature.com/articles/s41467-023-44288-7 Contact: Jessica ANDREANI and Raphaël GUEROIS <jessica.andreani@i2bc.paris-saclay.fr> <raphael.guerois@i2bc.paris-saclay.fr>
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