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2023

RSL24D1 sustains steady-state ribosome biogenesis and pluripotency translational programs in embryonic stem cells.

Author(s) : Durand S, Bruelle M, Bourdelais F, Bennychen B, Blin-Gonthier J, Isaac C, Huyghe A, Martel S, Seyve A, Vanbelle C, Adrait A, Couté Y, Meyronet D, Catez F, Diaz J, Lavial F, Ricci E, Ducray F, Gabut M,
Journal : Nat Commun
2023
Embryonic stem cell (ESC) fate decisions are regulated by a complex circuitrythat coordinates gene expression at multiple levels from chromatin to mRNAprocessing. Recently, ribosome biogenesis and translation have emerged as keypathways that efficiently control stem cell homeostasis, yet the underlyingmolecular mechanisms remain largely unknown. Here, we identified RSL24D1 ashighly expressed in both mouse and human pluripotent stem cells. RSL24D1 isassociated with nuclear pre-ribosomes and is required for the biogenesis of 60Ssubunits in mouse ESCs. Interestingly, RSL24D1 depletion significantly impairsglobal translation, particularly of key pluripotency factors and of componentsfrom the Polycomb Repressive Complex 2 (PRC2). While having a moderate impact ondifferentiation, RSL24D1 depletion significantly alters ESC self-renewal andlineage commitment choices. Altogether, these results demonstrate thatRSL24D1-dependant ribosome biogenesis is both required to sustain the expressionof pluripotent transcriptional programs and to silence PRC2-regulateddevelopmental programs, which concertedly dictate ESC homeostasis.

Severe COVID-19 patients have impaired plasmacytoid dendritic cell-mediated control of SARS-CoV-2.

Author(s) : Venet M, Ribeiro M, Décembre E, Bellomo A, Joshi G, Nuovo C, Villard M, Cluet D, Perret M, Pescamona R, Paidassi H, Walzer T, Allatif O, Belot A, Trouillet-Assant S, Ricci E, Dreux M,
Journal : Nat Commun
2023
Type I and III interferons (IFN-I/λ) are important antiviral mediators againstSARS-CoV-2 infection. Here, we demonstrate that plasmacytoid dendritic cells(pDC) are the predominant IFN-I/λ source following their sensing ofSARS-CoV-2-infected cells. Mechanistically, this short-range sensing by pDCsrequires sustained integrin-mediated cell adhesion with infected cells. In turn,pDCs restrict viral spread by an IFN-I/λ response directed towardSARS-CoV-2-infected cells. This specialized function enables pDCs to efficientlyturn-off viral replication, likely via a local response at the contact site withinfected cells. By exploring the pDC response in SARS-CoV-2 patients, we furtherdemonstrate that pDC responsiveness inversely correlates with the severity of thedisease. The pDC response is particularly impaired in severe COVID-19 patients.Overall, we propose that pDC activation is essential to controlSARS-CoV-2-infection. Failure to develop this response could be important tounderstand severe cases of COVID-19.

Discriminating physiological from non-physiological interfaces in structures of protein complexes: a community-wide study

Author(s) : Schweke H, Xu Q, Tauriello G, Pantolini L, Schwede T, Cazals F, Lhéritier A, Fernandez-Recio J, Rodríguez-Lumbreras L, Schueler-Furman O, others,
Journal : Proteomics
2023

Expulsion mechanism of the substrate-translocating subunit in ECF transporters

Author(s) : Thangaratnarajah C, Nijland M, Borges-Araújo L, Jeucken A, Rheinberger J, Marrink S, Souza P, Paulino C, Slotboom D,
Journal : Nature Communications
2023

Transmembrane dimers of type 1 receptors sample alternate configurations: MD simulations using coarse grain Martini 3 versus AlphaFold2 Multimer

Author(s) : Sahoo A, Souza P, Meng Z, Buck M,
Journal : Structure
2023

An implementation of the Martini coarse-grained force field in OpenMM

Author(s) : MacCallum J, Hu S, Lenz S, Souza P, Corradi V, Tieleman D,
Journal : Biophysical Journal
2023

Coevolution-Guided Mapping of the Type VI Secretion Membrane Complex-Baseplate Interface.

Author(s) : Vanlioğlu E, Santin Y, Filella-Merce I, Pellarin R, Cascales E,
Journal : J Mol Biol
2023
The type VI secretion system (T6SS) is a multiprotein weapon evolved byGram-negative bacteria to deliver effectors into eukaryotic cells or bacterialrivals. The T6SS uses a contractile mechanism to propel an effector-loaded needleinto its target. The contractile tail is built on an assembly platform, thebaseplate, which is anchored to a membrane complex. Baseplate-membrane complexinteractions are mainly mediated by contacts between the C-terminal domain of theTssK baseplate component and the cytoplasmic domain of the TssL inner membraneprotein. Currently, the structural details of this interaction are unknown due tothe marginal stability of the TssK-TssL complex. Here we conducted a mutagenesisstudy based on putative TssK-TssL contact pairs identified by co-evolutionanalyses. We then evaluated the impact of these mutations on T6SS activity,TssK-TssL interaction and sheath assembly and dynamics in enteroaggregativeEscherichia coli. Finally, we probed the TssK-TssL interface by disulfidecross-linking, allowing to propose a model for the baseplate-membrane complexinterface.

Acinetobacter type VI secretion system comprises a non-canonical membrane complex.

Author(s) : Kandolo O, Cherrak Y, Filella-Merce I, Le Guenno H, Kosta A, Espinosa L, Santucci P, Verthuy C, Lebrun R, Nilges M, Pellarin R, Durand E,
Journal : PLoS Pathog
2023
A. baumannii can rapidly acquire new resistance mechanisms and persist on abioticsurface, enabling the colonization of asymptomatic human host. In Acinetobacterthe type VI secretion system (T6SS) is involved in twitching, surface motilityand is used for interbacterial competition allowing the bacteria to uptake DNA.A. baumannii possesses a T6SS that has been well studied for its regulation andspecific activity, but little is known concerning its assembly and architecture.The T6SS nanomachine is built from three architectural sub-complexes. Unlike thebaseplate (BP) and the tail-tube complex (TTC), which are inherited frombacteriophages, the membrane complex (MC) originates from bacteria. The MC is themost external part of the T6SS and, as such, is subjected to evolution andadaptation. One unanswered question on the MC is how such a gigantesque molecularedifice is inserted and crosses the bacterial cell envelope. The A. baumannii MClacks an essential component, the TssJ lipoprotein, which anchors the MC to theouter membrane. In this work, we studied how A. baumannii compensates the absenceof a TssJ. We have characterized for the first time the A. baumannii's specificT6SS MC, its unique characteristic, its membrane localization, and assemblydynamics. We also defined its composition, demonstrating that its biogenesisemploys three Acinetobacter-specific envelope-associated proteins that define anintricate network leading to the assembly of a five-proteins membranesuper-complex. Our data suggest that A. baumannii has divided the function ofTssJ by (1) co-opting a new protein TsmK that stabilizes the MC and by (2)evolving a new domain in TssM for homo-oligomerization, a prerequisite to buildthe T6SS channel. We believe that the atypical species-specific features wereport in this study will have profound implication in our understanding of theassembly and evolutionary diversity of different T6SSs, that warrants futureinvestigation.

Carm1-arginine methylation of the transcription factor C/EBPα regulates transdifferentiation velocity

Author(s) : Torcal Garcia G, Kowenz-Leutz E, Tian T, Klonizakis A, Lerner J, De Andres-Aguayo L, Sapozhnikova V, Berenguer C, Carmona M, Casadesus M, Bulteau R, Francesconi M, Peiro S, Mertins P, Zaret K, Leutz A, Graf T,
Journal : eLife
2023
Here, we describe how the speed of C/EBPα-induced B cell to macrophage transdifferentiation (BMT) can be regulated, using both mouse and human models. The identification of a mutant of C/EBPα (C/EBPαR35A) that greatly accelerates BMT helped to illuminate the mechanism. Thus, incoming C/EBPα binds to PU.1, an obligate partner expressed in B cells, leading to the release of PU.1 from B cell enhancers, chromatin closing and silencing of the B cell program. Released PU.1 redistributes to macrophage enhancers newly occupied by C/EBPα, causing chromatin opening and activation of macrophage genes. All these steps are accelerated by C/EBPαR35A, initiated by its increased affinity for PU.1. Wild-type C/EBPα is methylated by Carm1 at arginine 35 and the enzyme’s perturbations modulate BMT velocity as predicted from the observations with the mutant. Increasing the proportion of unmethylated C/EBPα in granulocyte/macrophage progenitors by inhibiting Carm1 biases the cell’s differentiation toward macrophages, suggesting that cell fate decision velocity and lineage directionality are closely linked processes.

Mechanism of homology search expansion during recombinational DNA break repair

Author(s) : Dumont A, Mendiboure N, Savocco J, Anani L, Moreau P, Thierry A, Modolo L, Jost D, Piazza A,
Journal : BioRxiv
2023
Homology search catalyzed by a RecA/Rad51 nucleoprotein filament (NPF) is a central step of DNA double-strand break (DSB) repair by homologous recombination. How it operates in cells remains elusive. Here we developed a Hi-C-based methodology to map single-stranded DNA (ssDNA) contacts genome-wide in S. cerevisiae, which revealed two main homology search phases. Initial search conducted by short NPFs is confined in cis by cohesin-mediated chromatin loop folding. Progressive growth of stiff NPFs enables exploration of distant genomic sites. Long-range resection by Exo1 drives this transition from local to genome-wide search by providing ssDNA substrates for assembly of extensive NPFs. DSB end-tethering promotes coordinated homology search by NPFs formed on the two DSB ends. Finally, an autonomous genetic element on chromosome III engages the NPF and stimulates homology search in its vicinity. This work reveals the mechanism of the progressive and uneven expansion of homology search orchestrated by chromatin organizers, long-range resection, end-tethering, specialized genetic elements, and that exploits the stiff NPF structure conferred by Rad51 oligomerization.

Topological Constraints and Finite-Size Effects in Quantitative Polymer Models of Chromatin Organization

Author(s) : Abdulla A, Tortora M, Vaillant C, Jost D,
Journal : Macromolecules
2023

Transcription induces context-dependent remodeling of chromatin architecture during differentiation.

Author(s) : Chahar S, Ben Zouari Y, Salari H, Kobi D, Maroquenne M, Erb C, Molitor A, Mossler A, Karasu N, Jost D, Sexton T,
Journal : PLoS Biol
2023
Metazoan chromosomes are organized into discrete spatial domains (TADs), believedto contribute to the regulation of transcriptional programs. Despite extensivecorrelation between domain organization and gene activity, a direct mechanisticlink is unclear, with perturbation studies often showing little effect. To followchromatin architecture changes during development, we used Capture Hi-C tointerrogate the domains around key differentially expressed genes during mousethymocyte maturation, uncovering specific remodeling events. Notably, one TADboundary was broadened to accommodate RNA polymerase elongation past the border,and subdomains were formed around some activated genes without changes in CTCFbinding. The ectopic induction of some genes was sufficient to recapitulatedomain formation in embryonic stem cells, providing strong evidence thattranscription can directly remodel chromatin structure. These results suggestthat transcriptional processes drive complex chromosome folding patterns that canbe important in certain genomic contexts.

CGCompiler: Automated Coarse-Grained Molecule Parametrization via Noise-Resistant Mixed-Variable Optimization

Author(s) : Stroh K, Souza P, Monticelli L, Risselada H,
Journal : Journal of Chemical Theory and Computation
2023
Coarse-grained force fields (CG FFs) such as the Martini model entail a predefined, fixed set of Lennard-Jones parameters (building blocks) to model virtually all possible nonbonded interactions between chemically relevant molecules. Owing to its universality and transferability, the building-block coarse-grained approach has gained tremendous popularity over the past decade. The parametrization of molecules can be highly complex and often involves the selection and fine-tuning of a large number of parameters (e.g., bead types and bond lengths) to optimally match multiple relevant targets simultaneously. The parametrization of a molecule within the building-block CG approach is a mixed-variable optimization problem: the nonbonded interactions are discrete variables, whereas the bonded interactions are continuous variables. Here, we pioneer the utility of mixed-variable particle swarm optimization in automatically parametrizing molecules within the Martini 3 coarse-grained force field by matching both structural (e.g., RDFs) as well as thermodynamic data (phase-transition temperatures). For the sake of demonstration, we parametrize the linker of the lipid sphingomyelin. The important advantage of our approach is that both bonded and nonbonded interactions are simultaneously optimized while conserving the search efficiency of vector guided particle swarm optimization (PSO) methods over other metaheuristic search methods such as genetic algorithms. In addition, we explore noise-mitigation strategies in matching the phase-transition temperatures of lipid membranes, where nucleation and concomitant hysteresis introduce a dominant noise term within the objective function. We propose that noise-resistant mixed-variable PSO methods can both improve and automate parametrization of molecules within building-block CG FFs, such as Martini.

Histone removal in sperm protects paternal chromosomes from premature division at fertilization

Author(s) : Dubruille R, Herbette M, Revel M, Horard B, Chang C, Loppin B,
Journal : Science
2023
The global replacement of histones with protamines in sperm chromatin is widespread in animals, including insects, but its actual function remains enigmatic. We show that in the Drosophila paternal effect mutant paternal loss ( pal ), sperm chromatin retains germline histones H3 and H4 genome wide without impairing sperm viability. However, after fertilization, pal sperm chromosomes are targeted by the egg chromosomal passenger complex and engage into a catastrophic premature division in synchrony with female meiosis II. We show that pal encodes a rapidly evolving transition protein specifically required for the eviction of (H3-H4) 2 tetramers from spermatid DNA after the removal of H2A-H2B dimers. Our study thus reveals an unsuspected role of histone eviction from insect sperm chromatin: safeguarding the integrity of the male pronucleus during female meiosis. , Editor’s summary Ultracompaction of sperm DNA in many species is often associated with genome-wide replacement of histones with protamines, but the actual role of this radical change in chromatin organization remains largely enigmatic. Looking at a multidecade-old Drosophila mutant, Dubruille et al . found that histones were massively retained in sperm without affecting its ability to fertilize (see the Perspective by Levine). However, at fertilization, male chromosomes were aberrantly recognized by maternal factors that control female meiotic progression, leading to a deleterious premature division and early loss of male chromosomes. This work highlights the role of sperm chromatin in protecting male chromosomes in the egg. —Di Jiang , Histone retention in Drosophila sperm chromatin jeopardizes paternal chromosome integrity during female meiosis.

Condensin positioning at telomeres by shelterin proteins drives sister-telomere disjunction in anaphase

Author(s) : Colin L, Reyes C, Berthezene J, Maestroni L, Modolo L, Toselli E, Chanard N, Schaak S, Cuvier O, Gachet Y, Coulon S, Bernard P, Tournier S,
Journal : eLife
2023
The localization of condensin along chromosomes is crucial for their accurate segregation in anaphase. Condensin is enriched at telomeres but how and for what purpose had remained elusive. Here we show that fission yeast condensin accumulates at telomere repeats through the balancing acts of Taz1, a core component of the shelterin complex that ensures telomeric functions, and Mit1, a nucleosome-remodeler associated with shelterin. We further show that condensin takes part in sister-telomere separation in anaphase, and that this event can be uncoupled from the prior separation of chromosome arms, implying a telomere-specific separation mechanism. Consistent with a cis-acting process, increasing or decreasing condensin occupancy specifically at telomeres modifies accordingly the efficiency of their separation in anaphase. Genetic evidence suggests that condensin promotes sister-telomere separation by counteracting cohesin. Thus, our results reveal a shelterin-based mechanism that enriches condensin at telomeres to drive in cis their separation during mitosis.

SIN3 acts in distinct complexes to regulate the germline transcriptional program in C. elegans.

Author(s) : Robert V, Caron M, Gely L, Adrait A, Pakulska V, Couté Y, Chevalier M, Riedel C, Bedet C, Palladino F,
Journal : Development
2023
The SIN3 transcriptional coregulator influences gene expression through multipleinteractions that include histone deacetylases (HDACs). Haploinsufficiency andmutations in SIN3 are the underlying cause of Witteveen-Kolk syndrome and relatedintellectual disability (ID)/autism syndromes, emphasizing its key role indevelopment. However, little is known about the diversity of its interactions andfunctions in developmental processes. Here we show that loss of SIN-3, the singleSIN3 homologue in Caenorhabditis elegans, results in maternal effect sterilityassociated with deregulation of the germline transcriptome, including desilencingof X-linked genes. We identify at least two distinct SIN3 complexes containingspecific HDACs, and show that they differentially contribute to fertility. Singlecell smFISH reveals that in sin-3 mutants, the X chromosome becomes re-expressedprematurely and in a stochastic manner in individual germ cells, suggesting arole for SIN-3 in its silencing. Furthermore, we identify histone residues whoseacetylation increases in the absence of SIN3. Together, this work provides apowerful framework for the in vivo study of SIN3 and associated proteins.

Programmed DNA elimination in Mesorhabditis nematodes

Author(s) : Rey C, Launay C, Wenger E, Delattre M,
Journal : Curr Biol
2023

Cosegregation of recombinant chromatids maintains genome-wide heterozygosity in an asexual nematode

Author(s) : Blanc C, Saclier N, Le Faou E, Marie-Orleach L, Wenger E, Diblasi C, Glemin S, Galtier N, Delattre M,
Journal : Sci Adv
2023

APOLLO, a testis-specific Drosophila ortholog of importin-4, mediates the loading of protamine-like protein Mst77F into sperm chromatin

Author(s) : Emelyanov A, Barcenilla-Merino D, Loppin B, Fyodorov D,
Journal : Journal of Biological Chemistry
2023

Neural network and kinetic modelling of human genome replication reveal replication origin locations and strengths.

Author(s) : Arbona J, Kabalane H, Barbier J, Goldar A, Hyrien O, Audit B,
Journal : PLoS Comput Biol
2023
In human and other metazoans, the determinants of replication origin location andstrength are still elusive. Origins are licensed in G1 phase and fired in S phaseof the cell cycle, respectively. It is debated which of these two temporallyseparate steps determines origin efficiency. Experiments can independentlyprofile mean replication timing (MRT) and replication fork directionality (RFD)genome-wide. Such profiles contain information on multiple origins' propertiesand on fork speed. Due to possible origin inactivation by passive replication,however, observed and intrinsic origin efficiencies can markedly differ. Thus,there is a need for methods to infer intrinsic from observed origin efficiency,which is context-dependent. Here, we show that MRT and RFD data are highlyconsistent with each other but contain information at different spatial scales.Using neural networks, we infer an origin licensing landscape that, when insertedin an appropriate simulation framework, jointly predicts MRT and RFD data withunprecedented precision and underlies the importance of dispersive origin firing.We furthermore uncover an analytical formula that predicts intrinsic fromobserved origin efficiency combined with MRT data. Comparison of inferredintrinsic origin efficiencies with experimental profiles of licensed origins(ORC, MCM) and actual initiation events (Bubble-seq, SNS-seq, OK-seq, ORM) showthat intrinsic origin efficiency is not solely determined by licensingefficiency. Thus, human replication origin efficiency is set at both the originlicensing and firing steps.

HP1-driven phase separation recapitulates the thermodynamics and kinetics of heterochromatin condensate formation.

Author(s) : Tortora M, Brennan L, Karpen G, Jost D,
Journal : Proc Natl Acad Sci U S A
2023
The spatial segregation of pericentromeric heterochromatin (PCH) into distinct,membrane-less nuclear compartments involves the binding of HeterochromatinProtein 1 (HP1) to H3K9me2/3-rich genomic regions. While HP1 exhibitsliquid-liquid phase separation properties in vitro, its mechanistic impact on thestructure and dynamics of PCH condensate formation in vivo remains largelyunresolved. Here, using a minimal theoretical framework, we systematicallyinvestigate the mutual coupling between self-interacting HP1-like molecules andthe chromatin polymer. We reveal that the specific affinity of HP1 for H3K9me2/3loci facilitates coacervation in nucleo and promotes the formation of stable PCHcondensates at HP1 levels far below the concentration required to observe phaseseparation in purified protein assays in vitro. These heterotypic HP1-chromatininteractions give rise to a strong dependence of the nucleoplasmic HP1 density onHP1-H3K9me2/3 stoichiometry, consistent with the thermodynamics of multicomponentphase separation. The dynamical cross talk between HP1 and the viscoelasticchromatin scaffold also leads to anomalously slow equilibration kinetics, whichstrongly depend on the genomic distribution of H3K9me2/3 domains and result inthe coexistence of multiple long-lived, microphase-separated PCH compartments.The morphology of these complex coacervates is further found to be governed bythe dynamic establishment of the underlying H3K9me2/3 landscape, which may drivetheir increasingly abnormal, aspherical shapes during cell development. Thesefindings compare favorably to 4D microscopy measurements of HP1 condensateformation in live Drosophila embryos and suggest a general quantitative model ofPCH formation based on the interplay between HP1-based phase separation andchromatin polymer mechanics.

Biophysical ordering transitions underlie genome 3D re-organization during cricket spermiogenesis.

Author(s) : Orsi G, Tortora M, Horard B, Baas D, Kleman J, Bucevičius J, Lukinavičius G, Jost D, Loppin B,
Journal : Nat Commun
2023
Spermiogenesis is a radical process of differentiation whereby sperm cellsacquire a compact and specialized morphology to cope with the constraints ofsexual reproduction while preserving their main cargo, an intact copy of thepaternal genome. In animals, this often involves the replacement of most histonesby sperm-specific nuclear basic proteins (SNBPs). Yet, how the SNBP-structuredgenome achieves compaction and accommodates shaping remain largely unknown. Here,we exploit confocal, electron and super-resolution microscopy, coupled withpolymer modeling to identify the higher-order architecture of sperm chromatin inthe needle-shaped nucleus of the emerging model cricket Gryllus bimaculatus.Accompanying spermatid differentiation, the SNBP-based genome is strikinglyreorganized as ~25nm-thick fibers orderly coiled along the elongated nucleusaxis. This chromatin spool is further found to achieve large-scale helicaltwisting in the final stages of spermiogenesis, favoring its ultracompaction. Wereveal that these dramatic transitions may be recapitulated by a surprisinglysimple biophysical principle based on a nucleated rigidification of chromatinlinked to the histone-to-SNBP transition within a confined nuclear space. Ourwork highlights a unique, liquid crystal-like mode of higher-order genomeorganization in ultracompact cricket sperm, and establishes a multidisciplinarymethodological framework to explore the diversity of non-canonical modes of DNAorganization.

A burning question from the first international BPAN symposium: is restoration of autophagy a promising therapeutic strategy for BPAN?

Author(s) : Mollereau B, Hayflick S, Escalante R, Mauthe M, Papandreou A, Iuso A, Celle M, Aniorte S, Issa A, Lasserre J, Lesca G, Thobois S, Burger P, Walter L,
Journal : Autophagy
2023
Beta-propeller protein-associated neurodegeneration (BPAN) is a rareneurodegenerative disease associated with severe cognitive and motor deficits.BPAN pathophysiology and phenotypic spectrum are still emerging due to the factthat mutations in the WDR45 (WD repeat domain 45) gene, a regulator ofmacroautophagy/autophagy, were only identified a decade ago. In the firstinternational symposium dedicated to BPAN, which was held in Lyon, France, apanel of international speakers, including several researchers from the autophagycommunity, presented their work on human patients, cellular and animal models,carrying WDR45 mutations and their homologs. Autophagy researchers found anopportunity to explore the defective function of autophagy mechanisms associatedwith WDR45 mutations, which underlie neuronal dysfunction and early death.Importantly, BPAN is one of the few human monogenic neurological diseasestargeting a regulator of autophagy, which raises the possibility that it is arelevant model to directly assess the roles of autophagy in neurodegeneration andto develop autophagy restorative therapeutic strategies for more commondisorders.

Delineation of two multi-invasion-induced rearrangement pathways that differently affect genome stability

Author(s) : Reitz D, Djeghmoum Y, Piazza A,
Journal : Genes and Development
2023
Punctuated bursts of structural genomic variations (SVs) have been described in various organisms, but their etiology remains incompletely understood. Homologous recombination (HR) is a template-guided mechanism of repair of DNA double-strand breaks and stalled or collapsed replication forks. We recently identified a DNA break amplification and genome rearrangement pathway originating from the endonucleolytic processing of a multi-invasion (MI) DNA joint molecule formed during HR. Genome-wide approaches confirmed that multi-invasion-induced rearrangement (MIR) frequently leads to several repeat-mediated SVs and aneuploidies. Using molecular and genetic analysis and a novel, highly sensitive proximity ligation-based assay for chromosomal rearrangement quantification, we further delineate two MIR subpathways. MIR1 is a universal pathway occurring in any sequence context, which generates secondary breaks and frequently leads to additional SVs. MIR2 occurs only if recombining donors exhibit substantial homology and results in sequence insertion without additional breaks or SVs. The most detrimental MIR1 pathway occurs late on a subset of persisting DNA joint molecules in a PCNA/Polδ-independent manner, unlike recombinational DNA synthesis. This work provides a refined mechanistic understanding of these HR-based SV formation pathways and shows that complex repeat-mediated SVs can occur without displacement DNA synthesis. Sequence signatures for inferring MIR1 from long-read data are proposed.

Essential and recurrent roles for hairpin RNAs in silencing de novo sex chromosome conflict in Drosophila simulans

Author(s) : Vedanayagam J, Herbette M, Mudgett H, Lin C, Lai C, McDonough-Goldstein C, Dorus S, Loppin B, Meiklejohn C, Dubruille R, Lai E,
Journal : PLOS Biology
2023
AMUeio: tPicledarsievceolnofcirimditshtaotratllthheeandoinrgmleavlelylseaqreuraelpsresgernetgedactioornreocftlayl:leles, which benefits their own transmission even in the face of severe fitness costs to their host organism. However, relatively little is known about the molecular identity of meiotic drivers, their strategies of action, and mechanisms that can suppress their activity. Here, we present data from the fruitfly Drosophila simulans that address these questions. We show that a family of de novo, protamine-derived X-linked selfish genes (the Dox gene family) is silenced by a pair of newly emerged hairpin RNA (hpRNA) small interfering RNA (siRNA)-class loci, Nmy and Tmy. In the w[XD1] genetic background, knockout of nmy derepresses Dox and MDox in testes and depletes male progeny, whereas knockout of tmy causes misexpression of PDox genes and renders males sterile. Importantly, genetic interactions between nmy and tmy mutant alleles reveal that Tmy also specifically maintains male progeny for normal sex ratio. We show the Dox loci are functionally polymorphic within D. simulans, such that both nmy-associated sex ratio bias and tmy-associated sterility can be rescued by wild-type X chromosomes bearing natural deletions in different Dox family genes. Finally, using tagged transgenes of Dox and PDox2, we provide the first experimental evidence Dox family genes encode proteins that are strongly derepressed in cognate hpRNA mutants. Altogether, these studies support a model in which protamine-derived drivers and hpRNA suppressors drive repeated cycles of sex chromosome conflict and resolution that shape genome evolution and the genetic control of male gametogenesis.

Two decades of Martini: Better beads, broader scope

Author(s) : Marrink S, Monticelli L, Melo M, Alessandri R, Tieleman D, Souza P,
Journal : WIREs Computational Molecular Science
2023
Abstract The Martini model, a coarse-grained force field for molecular dynamics simulations, has been around for nearly two decades. Originally developed for lipid-based systems by the groups of Marrink and Tieleman, the Martini model has over the years been extended as a community effort to the current level of a general-purpose force field. Apart from the obvious benefit of a reduction in computational cost, the popularity of the model is largely due to the systematic yet intuitive building-block approach that underlies the model, as well as the open nature of the development and its continuous validation. The easy implementation in the widely used Gromacs software suite has also been instrumental. Since its conception in 2002, the Martini model underwent a gradual refinement of the bead interactions and a widening scope of applications. In this review, we look back at this development, culminating with the release of the Martini 3 version in 2021. The power of the model is illustrated with key examples of recent important findings in biological and material sciences enabled with Martini, as well as examples from areas where coarse-grained resolution is essential, namely high-throughput applications, systems with large complexity, and simulations approaching the scale of whole cells. This article is categorized under: Software > Molecular Modeling Molecular and Statistical Mechanics > Molecular Dynamics and Monte-Carlo Methods Structure and Mechanism > Computational Materials Science Structure and Mechanism > Computational Biochemistry and Biophysics

Automatic Optimization of Lipid Models in the Martini Force Field Using SwarmCG

Author(s) : Empereur-mot C, Pedersen K, Capelli R, Crippa M, Caruso C, Perrone M, Souza P, Marrink S, Pavan G,
Journal : Journal of Chemical Information and Modeling
2023
After two decades of continued development of the Martini coarse-grained force field (CG FF), further refinment of the already rather accurate Martini lipid models has become a demanding task that could benefit from integrative data-driven methods. Automatic approaches are increasingly used in the development of accurate molecular models, but they typically make use of specifically designed interaction potentials that transfer poorly to molecular systems or conditions different than those used for model calibration. As a proof of concept, here, we employ SwarmCG, an automatic multiobjective optimization approach facilitating the development of lipid force fields, to refine specifically the bonded interaction parameters in building blocks of lipid models within the framework of the general Martini CG FF. As targets of the optimization procedure, we employ both experimental observables (top-down references: area per lipid and bilayer thickness) and all-atom molecular dynamics simulations (bottom-up reference), which respectively inform on the supra-molecular structure of the lipid bilayer systems and on their submolecular dynamics. In our training sets, we simulate at different temperatures in the liquid and gel phases up to 11 homogeneous lamellar bilayers composed of phosphatidylcholine lipids spanning various tail lengths and degrees of (un)saturation. We explore different CG representations of the molecules and evaluate improvements a posteriori using additional simulation temperatures and a portion of the phase diagram of a DOPC/DPPC mixture. Successfully optimizing up to ∼80 model parameters within still limited computational budgets, we show that this protocol allows the obtainment of improved transferable Martini lipid models. In particular, the results of this study demonstrate how a fine-tuning of the representation and parameters of the models may improve their accuracy and how automatic approaches, such as SwarmCG, may be very useful to this end.

Protocol to measure protein-RNA binding using double filter-binding assays followed by phosphorimaging or high-throughput sequencing

Author(s) : Vega-Badillo J, Zamore P, Jouravleva K,
Journal : STAR Protoc
2023
Binding affinity quantitatively describes the strength of a molecular interaction and is reported by the equilibrium dissociation constant (KD). Here, we present a protocol to measure KD of mammalian microRNA-loaded Argonaute2 protein by double filter binding. We describe steps for radiolabeling target RNA, measuring concentration of binding-competent protein, setting up binding reactions, separating protein-bound RNA from protein-unbound RNA, preparing library for Illumina sequencing, and performing data analysis. Our protocol is easily applied to other RNA- or DNA-binding proteins. For complete details on the use and execution of this protocol, please refer to Jouravleva et al.

Orientational Wetting and Topological Transitions in Confined Solutions of Semiflexible Polymers

Author(s) : Tortora M, Jost D,
Journal : Macromolecules
2023