Annuaire
Publications
Les 20 dernières publications
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The SARS-CoV-2 nucleocapsid protein interferes with the full enzymatic activation of UPF1 and its interaction with UPF2.
- Journal : Nucleic Acids Res
- 2025
- The nonsense-mediated mRNA decay (NMD) pathway triggers the degradation ofdefective mRNAs and governs the expression of mRNAs with specificcharacteristics. Current understanding indicates that NMD is often significantlysuppressed during viral infections to protect the viral genome. In numerousviruses, this inhibition is achieved through direct or indirect interference withthe RNA helicase UPF1, thereby promoting viral replication and enhancingpathogenesis. In this study, we employed biochemical, biophysical assays andcellular investigations to explore the interplay between UPF1 and thenucleocapsid (Np) protein of SARS-CoV-2. We evaluated their direct interactionand its impact on inhibiting cellular NMD. Furthermore, we characterized how thisinteraction affects UPF1's enzymatic function. Our findings demonstrate that Npinhibits the unwinding activity of UPF1 by physically obstructing its access tostructured nucleic acid substrates. Additionally, we showed that Np bindsdirectly to UPF2, disrupting the formation of the UPF1/UPF2 complex essential forNMD progression. Intriguingly, our research also uncovered a surprising pro-viralrole of UPF1 and an antiviral function of UPF2. These results unveil a novel,multi-faceted mechanism by which SARS-CoV-2 evades the host's defenses andmanipulates cellular components. This underscores the potential therapeuticstrategy of targeting Np-UPF1/UPF2 interactions to treat COVID-19.
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Hijacking a real time detection thermocycler for enzymology: Improvement of a fluorescent bulk assay monitoring helicase activity.
- Journal : Biochimie
- 2025
- Helicases are enzymes involved in all aspects of nucleic acid synthesis,regulation and degradation. As a consequence, several methods were developed tomonitor their enzymatic activity. In this report, we described an improvement ofbulk fluorescent helicase assays to overcome their specific limitations (cost,health and safety regulations, etc.). Using a real time detection thermocycler tomonitor the fluorescence in real-time, we managed to precisely control theinitiation of the helicase reaction through temperature tuning. Therefore, wewere able to demonstrate that this setup could provide a qualitative and aquantitative evaluation of the helicase domain of the UPF1 helicase (UPF1-HD) andthat several fluorophores could be used in parallel during the same run.
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NAL1 forms a molecular cage to regulate FZP phase separation.
- Journal : Proc Natl Acad Sci U S A
- 2025
- NARROW LEAF 1 (NAL1), originally identified for its role in shaping leafmorphology, plant architecture, and various agronomic traits in rice, hasremained enigmatic in terms of the molecular mechanisms governing itsmultifaceted functions. In this study, we present a comprehensive structuralanalysis of NAL1 proteins, shedding light on how NAL1 regulates the phaseseparation of its physiological substrate, FRIZZY PANICLE (FZP), a transcriptionfactor. We determined that NAL1 assembles as a hexamer and forms a molecular cagewith a wide central channel and three narrower lateral channels, which coulddiscriminate its different substrates into the catalytic sites. Most notably, ourinvestigation unveils that FZP readily forms molecular condensates via phaseseparation both in vitro and in vivo. NAL1 fine-tunes FZP condensation,maintaining optimal concentrations to enhance transcriptional activity. Whilephase separation roles include sequestration and suppression of transcriptionalor enzymatic activity, our study highlights its context-dependent contribution totranscriptional regulation. NAL1 assumes a pivotal role in regulating the statesof these molecular condensates through its proteolytic activity, subsequentlyenhancing transcriptional cascades. Our findings offer insights intocomprehending the molecular mechanisms underpinning NAL1's diverse functions,with far-reaching implications for the field of plant biology. Additionally,these insights provide valuable guidance for the development of rational breedingstrategies aimed at enhancing crop productivity.
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A selfish supergene causes meiotic drive through both sexes in Drosophila
- Journal : Proceedings of the National Academy of Sciences
- 2025
- Meiotic drivers are selfish genetic elements that bias their own transmission during meiosis or gamete formation. Due to the fundamental differences between male and female meiosis in animals and plants, meiotic drivers operate through distinct mechanisms in the two sexes: In females, they exploit the asymmetry of meiosis to ensure their inclusion in the egg, whereas in males, they eliminate competing gametes after symmetric meiosis. Meiotic drive is commonly reported in males, where it strongly influences the evolution of spermatogenesis, while the few known cases in females have highlighted its crucial role in centromere evolution. Despite a growing number of examples in a wide range of organisms, meiotic drive has so far only been observed in one sex or the other since its discovery nearly 100 y ago. Here, we show that a selfish X chromosome known to cause meiotic drive in male Drosophila testacea flies also causes meiotic drive in females. We find that this X chromosome has supergene architecture, harboring extensive structural rearrangements that suppress recombination between the two X chromosomes. This has contributed to a substantial expansion of its size compared to the wild-type chromosome, partly due to the accumulation of species-specific repetitive elements. Our findings suggest that female meiotic drive may play an important role in the evolutionary dynamics of polymorphic structural variants that suppress recombination, including inversions, translocations, and supergenes.
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delfies: a Python package for the detection of DNA breakpoints with neo-telomere addition
- Journal : Journal of Open Source Software
- 2025
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Rhodamine6G and Hœchst33342 narrow BmrA conformational spectrum for a more efficient use of ATP.
- Journal : Nat Commun
- 2025
- Multidrug ABC transporters harness the energy of ATP binding and hydrolysis totranslocate substrates out of the cell and detoxify them. While this involves awell-accepted alternating access mechanism, molecular details of this interplayare still elusive. Rhodamine6G binding on a catalytic inactive mutant of thehomodimeric multidrug ABC transporter BmrA triggers a cooperative binding of ATPon the two identical nucleotide-binding-sites, otherwise michaelian. Here, weinvestigate this asymmetric behavior via a structural-enzymology approach,solving cryoEM structures of BmrA at defined ATP ratios, highlighting theplasticity of BmrA as it undergoes the transition from inward to outward facingconformations. Analysis of continuous heterogeneity within cryoEM data andstructural dynamics, reveals that Rhodamine6G narrows the conformational spectrumexplored by the nucleotide-binding domains. We observe the same behavior for theother drug Hœchst33342. Following on these findings, the effect of drug-bindingshowed an ATPase stimulation and a maximal transport activity of the wild-typeprotein at the concentration-range where the cooperative transition occurs.Altogether, these findings provide a description of the influence of drug bindingon the ATP-binding sites through a change in conformational dynamics.
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Donor transcription suppresses D-loops in cis and promotes genome stability
- Journal : BioRxiv
- 2025
- D-loops are DNA joint molecule intermediates central to DNA break repair by homologous recombination (HR). Priority rules between recombination and transcription at the donor locus have not been investigated. Here, using a controlled break induction system and physical detection of D-loops in S. cerevisiae, we show that donor transcription by RNA polymerase II acutely suppresses D-loops in cis, in an orientation-dependent manner. This inhibition does not rely on endogenous transcription factors, the RNA product, RNA:DNA hybrids, or previously characterized D-loop disruption factors. Transcription can be the major D-loop suppression pathway and inhibits the formation of repeat-mediated genome rearrangements. Transcription is therefore a negative regulator of HR at the D-loop level that promotes genome stability. These findings reveal the functional prioritization between two universal DNA-dependent processes.
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A guide to the biogenesis and functions of endogenous small non-coding RNAs in animals
- Journal : Nat Rev Mol Cell Biol
- 2025
- Small non-coding RNAs can be categorized into two main classes: structural RNAs and regulatory RNAs. Structural RNAs, which are abundant and ubiquitously expressed, have essential roles in the maturation of pre-mRNAs, modification of rRNAs and the translation of coding transcripts. By contrast, regulatory RNAs are often expressed in a developmental-specific, tissue-specific or cell-type-specific manner and exert precise control over gene expression. Reductions in cost and improvements in the accuracy of high-throughput RNA sequencing have led to the identification of many new small RNA species. In this Review, we provide a broad discussion of the genomic origins, biogenesis and functions of structural small RNAs, including tRNAs, small nuclear RNAs (snRNAs), small nucleolar RNAs (snoRNAs), vault RNAs (vtRNAs) and Y RNAs as well as their derived RNA fragments, and of regulatory small RNAs, such as microRNAs (miRNAs), endogenous small interfering RNAs (siRNAs) and PIWI-interacting RNAs (piRNAs), in animals.
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Translation-dependent and -independent mRNA decay occur through mutually exclusive pathways defined by ribosome density during T cell activation
- Journal : Genome Res
- 2024
- T cells. Our results indicate that most cellular transcripts are decayed to some extent in a translation-dependent manner. Our analysis further identifies the length of untranslated regions, the density of ribosomes, and GC3 content as important determinants of TDD magnitude. Consistently, all transcripts that undergo changes in ribosome density within their coding sequence upon T cell activation display a corresponding change in their TDD level. Moreover, we reveal a dynamic modulation in the relationship between GC3 content and TDD upon T cell activation, with a reversal in the impact of GC3- and AU3-rich codons. Altogether, our data show a strong and dynamic interconnection between mRNA translation and decay in mammalian primary cells.
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The white gene as a transgenesis marker for the cricket Gryllus bimaculatus
- Journal : G3: Genes, Genomes, Genetics
- 2024
- Abstract The cricket Gryllus bimaculatus is an emerging model insect of the order Orthoptera that is used in a wide variety of biological research themes. This hemimetabolous species appears highly complementary to Drosophila and other well-established holometabolous models. To improve transgenesis applications in G. bimaculatus, we have designed a transformation marker gene inspired from the widespread Drosophila mini-white+. Using CRISPR/Cas9, we first generated a loss-of-function mutant allele of the Gb-white gene (Gb-w), which exhibits a white eye coloration at all developmental stages. We then demonstrate that transgenic insertions of a piggyBac vector containing a 3xP3-Gb-w+ cassette rescue eye pigmentation. As an application, we used this vector to generate G. bimaculatus lines expressing a centromeric histone H3 variant (CenH3.1) fused to EGFP and validated EGFP-CenH3.1 detection at cricket centromeres. Finally, we demonstrate that Minos-based germline transformation and site-specific plasmid insertion with the ΦC31 integrase system function in G. bimaculatus.
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Deciphering Molecular Mechanisms Involved in the Modulation of Human Aquaporins' Water Permeability by Zinc Cations: A Molecular Dynamics Approach.
- Journal : Int J Mol Sci
- 2024
- Aquaporins (AQPs) constitute a wide family of water channels implicated in allkind of physiological processes. Zinc is the second most abundant trace elementin the human body and a few studies have highlighted regulation of AQP0 and AQP4by zinc. In the present work, we addressed the putative regulation of AQPs byzinc cations in silico through molecular dynamics simulations of human AQP0,AQP2, AQP4, and AQP5. Our results align with other scales of study and several invitro techniques, hence strengthening the reliability of this regulation by zinc.We also described two distinct putative molecular mechanisms associated with theincrease or decrease in AQPs' water permeability after zinc binding. Inassociation with other studies, our work will help deciphering the interactionnetworks existing between zinc and channel proteins.
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Retinal atrophy, inflammation, phagocytic and metabolic disruptions develop in the MerTK-cleavage-resistant mouse model.
- Journal : Front Neurosci
- 2024
- In the eye, cells from the retinal pigment epithelium (RPE) facing theneurosensory retina exert several functions that are all crucial for long-termsurvival of photoreceptors (PRs) and vision. Among those, RPE cells phagocytoseunder a circadian rhythm photoreceptor outer segment (POS) tips that areconstantly subjected to light rays and oxidative attacks. The MerTK tyrosinekinase receptor is a key element of this phagocytic machinery required for POSinternalization. Recently, we showed that MerTK is subjected to the cleavage ofits extracellular domain to finely control its function. In addition, monocytesin retinal blood vessels can migrate inside the inner retina and differentiateinto macrophages expressing MerTK, but their role in this context has not beenstudied yet. We thus investigated the ocular phenotype of MerTKcleavage-resistant (MerTK(CR)) mice to understand the relevance of thischaracteristic on retinal homeostasis at the RPE and macrophage levels. MerTK(CR)retinae appear to develop and function normally, as observed in retinal sections,by electroretinogram recordings and optokinetic behavioral tests. Monitoring ofMerTK(CR) and control mice between the ages of 3 and 18 months showed thedevelopment of large degenerative areas in the central retina as early as 4months when followed monthly by optical coherence tomography (OCT) plus fundusphotography (FP)/autofluorescence (AF) detection but not by OCT alone. Thedegenerative areas were associated with AF, which seems to be due to infiltratedmacrophages, as observed by OCT and histology. MerTK(CR) RPE primary culturesphagocytosed less POS in vitro, while in vivo, the circadian rhythm of POSphagocytosis was deregulated. Mitochondrial function and energy production werereduced in freshly dissected RPE/choroid tissues at all ages, thus showing ametabolic impairment not present in macrophages. RPE anomalies were detected byelectron microscopy, including phagosomes retained in the apical area andvacuoles. Altogether, this new mouse model displays a novel phenotype that couldprove useful to understanding the interplay between RPE and PRs in inflammatoryretinal degenerations and highlights new roles for MerTK in the regulation of theenergetic metabolism and the maintenance of the immune privilege in the retina.
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Structural insights into the N-terminal APHB domain of HrpA: mediating canonical and i-motif recognition.
- Journal : Nucleic Acids Res
- 2024
- RNA helicases function as versatile enzymes primarily responsible for remodelingRNA secondary structures and organizing ribonucleoprotein complexes. In ourstudy, we conducted a systematic analysis of the helicase-related activities ofEscherichia coli HrpA and presented the structures of both its apo form and itscomplex bound with both conventional and non-canonical DNAs. Our findings revealthat HrpA exhibits NTP hydrolysis activity and binds to ssDNA and ssRNA indistinct sequence-dependent manners. While the helicase core plays an essentialrole in unwinding RNA/RNA and RNA/DNA duplexes, the N-terminal extension in HrpA,consisting of three helices referred to as the APHB domain, is crucial for ssDNAbinding and RNA/DNA duplex unwinding. Importantly, the APHB domain is implicatedin binding to non-canonical DNA structures such as G-quadruplex and i-motif, andthis report presents the first solved i-motif-helicase complex. This research notonly provides comprehensive insights into the multifaceted roles of HrpA as anRNA helicase but also establishes a foundation for further investigations intothe recognition and functional implications of i-motif DNA structures in variousbiological processes.
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The catalytic triad of rice NARROW LEAF1 involves H234.
- Journal : Nat Plants
- 2024
- NARROW LEAF1 (NAL1) exerts a multifaceted influence on leaf morphology and cropyield. Recent crystal study proposed that histidine 233 (H233) is part of thecatalytic triad. Here we report that unlike suggested previously, H234 instead ofH233 is a component of the catalytic triad alongside residues D291 and S385 inNAL1. Remarkably, residue 233 unexpectedly plays a pivotal role in regulatingNAL1's proteolytic activity. These findings establish a strong foundation forutilizing NAL1 in breeding programs aimed at improving crop yield.
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Omecamtiv mecarbil and Mavacamten target the same myosin pocket despite opposite effects in heart contraction.
- Journal : Nat Commun
- 2024
- Inherited cardiomyopathies are common cardiac diseases worldwide, leading in thelate stage to heart failure and death. The most promising treatments againstthese diseases are small molecules directly modulating the force produced byβ-cardiac myosin, the molecular motor driving heart contraction. Omecamtivmecarbil and Mavacamten are two such molecules that completed phase 3 clinicaltrials, and the inhibitor Mavacamten is now approved by the FDA. In contrast toMavacamten, Omecamtiv mecarbil acts as an activator of cardiac contractility.Here, we reveal by X-ray crystallography that both drugs target the same pocketand stabilize a pre-stroke structural state, with only few local differences.All-atom molecular dynamics simulations reveal how these molecules producedistinct effects in motor allostery thus impacting force production in oppositeway. Altogether, our results provide the framework for rational drug developmentfor the purpose of personalized medicine.
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Stimulation of ATP Hydrolysis by ssDNA Provides the Necessary Mechanochemical Energy for G4 Unfolding.
- Journal : J Mol Biol
- 2024
- The G-quadruplex (G4) is a distinct geometric and electrophysical structurecompared to classical double-stranded DNA, and its stability can impede essentialcellular processes such as replication, transcription, and translation. Thisstudy focuses on the BsPif1 helicase, revealing its ability to bind independentlyto both single-stranded DNA (ssDNA) and G4 structures. The unfolding activity ofBsPif1 on G4 relies on the presence of a single tail chain, and the covalentcontinuity between the single tail chain and the G4's main chain is necessary forefficient G4 unwinding. This suggests that ATP hydrolysis-driven ssDNAtranslocation exerts a pull force on G4 unwinding. Molecular dynamics simulationsidentified a specific region within BsPif1 that contains five crucial amino acidsites responsible for G4 binding and unwinding. A "molecular wire stripper" modelis proposed to explain BsPif1's mechanism of G4 unwinding. These findings providea new theoretical foundation for further exploration of the G4 developmentmechanism in Pif1 family helicases.
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Aquaporin Modulation by Cations, a Review.
- Journal : Curr Issues Mol Biol
- 2024
- Aquaporins (AQPs) are transmembrane channels initially discovered for their rolein water flux facilitation through biological membranes. Over the years, a muchmore complex and subtle picture of these channels appeared, highlighting manyother solutes accommodated by AQPs and a dense regulatory network finely tuningcell membranes' water permeability. At the intersection between severaltransduction pathways (e.g., cell volume regulation, calcium signaling, potassiumcycling, etc.), this wide and ancient protein family is considered an importanttherapeutic target for cancer treatment and many other pathophysiologies.However, a precise and isoform-specific modulation of these channels function isstill challenging. Among the modulators of AQPs functions, cations have beenshown to play a significant contribution, starting with mercury beinghistorically associated with the inhibition of AQPs since their discovery. Whilethe comprehension of AQPs modulation by cations has improved, a unifyingmolecular mechanism integrating all current knowledge is still lacking. In aneffort to extract general trends, we reviewed all known modulations of AQPs bycations to capture a first glimpse of this regulatory network. We paid particularattention to the associated molecular mechanisms and pinpointed the residuesinvolved in cation binding and in conformational changes tied up to themodulation of the channel function.
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Plasticity and environment-specific relationships between gene expression and fitness in Saccharomyces cerevisiae.
- Journal : Nat Ecol Evol
- 2024
- The environment influences how an organism's genotype determines its phenotype and how this phenotype affects its fitness. Here, to better understand this dual role of environment in the production and selection of phenotypic variation, we determined genotype-phenotype-fitness relationships for mutant strains of Saccharomyces cerevisiae in four environments. Specifically, we measured how promoter mutations of the metabolic gene TDH3 modified expression level and affected growth for four different carbon sources. In each environment, we observed a clear relationship between TDH3 expression level and fitness, but this relationship differed among environments. Mutations with similar effects on expression in different environments often had different effects on fitness and vice versa. Such environment-specific relationships between phenotype and fitness can shape the evolution of phenotypic plasticity. We also found that mutations disrupting binding sites for transcription factors had more variable effects on expression among environments than those disrupting the TATA box, which is part of the core promoter. However, mutations with the most environmentally variable effects on fitness were located in the TATA box, because of both the lack of plasticity of TATA box mutations and environment-specific fitness functions. This observation suggests that mutations affecting different molecular mechanisms contribute unequally to regulatory sequence evolution in changing environments.
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Condensin loop extrusion properties, roadblocks, and role in homology search in S. cerevisiae
- Journal : BioRxiv
- 2024
- The in vivo mechanism, regulations by cis-acting roadblocks, and biological functions of loop extrusion by eukaryotic SMC complexes are incompletely defined. Here, using Hi-C, we identified two condensin-dependent contact stripes at the Recombination Enhancer (RE) and the rDNA in S. cerevisiae. We show that oriented, unidirectional loop extrusion proceeds from these sites with an estimated processivity ∼170 kb and a density ∼0.04-0.18 that varies across the cell cycle. Centromeres and highly-transcribed RNA PolII-dependent genes are permeable condensin roadblocks. Other positionally labile elements such as replication forks and Smc5/6 complexes bound to substrates generated in the absence of Top2 also hinder loop extrusion by condensin. Cohesin is not an obstacle for condensin. Finally, a DNA double-strand break at MAT blocks condensin, which results in the rapid establishment of a long-range RE-MAT loop that juxtaposes the recombination machinery with its HMLα donor target. Hence, all budding yeast SMCs are involved in recombinational DNA repair. We propose a revised model for donor selection during MAT switching that exploits specific properties of loop extrusion by condensin. It can serve as a paradigm for the establishment of other types of selective interactions along chromosomes.
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ProA and ProB repeat sequences shape genome organization, and enhancers open domains
- Journal : bioRxiv
- 2023
- There is a growing awareness that repeat sequences (RepSeq) - the main constituents of the human genome - are also prime players in its organization. Here we propose that the genome should be envisioned as a supersystem with three main subsystems, each composed of functionally redundant, cooperating elements. We define herein ProA and ProB RepSeqs as sequences that promote either the A/euchromatin or the B/heterochromatin compartment. ProA and ProB RepSeqs shape A/B partitioning, such that the relative proportions of ProA and ProB RepSeqs determine the propensity of a chromosome segment to adopt either an A or a B configuration. In human, core ProA RepSeqs are essentially made of Alu elements, whereas core ProB RepSeqs consist of young L1 and some Endogenous Retroviruses (ERVs) as well as a panel of AT-rich microsatellites and pericentromeric and telomeric satellites. Additionally, RepSeqs with more indefinite character and, importantly, their derivatives known as “transcriptional enhancers”, can shift between ProA and ProB functions and thus act to open or close specific chromatin domains depending on the cellular context. In this framework, genes and their promoters appear as a special class of RepSeqs that, in their active, transcribed state, reinforce the openness of their surroundings. Molecular mechanisms involve cooperativity between ProB elements, presumably underpinned by the condensate-like properties of heterochromatin, which ProA elements oppose in several ways. We provide strong arguments that altered CpG methylation patterns in cancer including a marked loss in the B compartment, result primarily from a global imbalance in the process of CpG methylation and its erasure. Our results suggest that the resulting altered methylation and impaired function of ProB RepSeqs globally weaken the B compartment, rendering it more plastic, which in turn may confer fate plasticity to the cancer cell.Competing Interest StatementThe authors have declared no competing interest.
Link to PubMed entry
