Biology subjects

Roux, A.

Publications and source records attributed to Roux, A..

At least 19 recordsLinked to original sources

bioRxiv2026

A regulatory T cell signature provides a shared molecular basis for the therapeutic window of opportunity in rheumatic disease

Rheumatic diseases, including rheumatoid arthritis (RA), spondyloarthritis (SpA) and osteoarthritis (OA), show distinct phenotypes yet respond to overlapping therapies, implicating shared immune mechanisms. In the Transimmunom cohort, we profiled peripheral blood from 240 individuals (47 healthy, 44 OA, 91 RA, 58 SpA) across deep immunophenotyping, immunoproteomics and Treg-Teff transcriptomics. Single-layer analyses revealed broader Treg than Teff remodeling, along with a shared pattern of reduced activated Tregs and expanded Helios+ Tregs across all diseases, alongside a decrease in functional Treg subpopulations, including CTLA4+ and CD45RA- Tregs. In RA specifically, LAG3+ Tregs were also expanded. Combining omics layers outperformed single-layer approaches for disease classification. Among individual layers, Treg transcriptomes were most discriminative, and integration uncovered disease-specific programs. Unsupervised clustering identified a cross-disease cluster independent of activity, treatment and age, mapping to early disease (<= years) and dominated by a Treg dysfunction-associated program. These results provide a biological rationale for the therapeutic "window of opportunity" concept and duration-stratified Treg-directed trials.

immunology↗

bioRxiv2026

A switch in clathrin turnover controls endocytic coat size and organisation

Endocytosis internalises nutrients, regulates extracellular signals, and recycles membrane components. Clathrin polymerises into a coat that shapes the endocytic vesicle from the plasma membrane. However, the role of clathrins dynamic assembly in the endocytic process remains unclear. We show, using two-colour fluorescence recovery after photobleaching assays in yeast, that the clathrin coat turns over rapidly in the early phase of endocytosis, dependent on the auxilin Swa2 and its ATPase. In the late phase the turnover is stopped by the coat protein Sla1. Regulated clathrin turnover is critical for the timing of endocytic progression and for controlling coat size. In the absence of this dynamic regulation the endocytic coats become abnormally large, resulting in the failure of the final actin-driven vesicle budding. These findings reveal that, in addition to its classic structural function, the dynamic properties of the clathrin lattice are critical for both the temporal and mechanical aspects of endocytosis.

cell biology↗

bioRxiv2026

Direction of ESCRT-III-dependent membrane bending emerges from bilayer asymmetry

In cells, ESCRT-III is unique in mediating fission of membrane necks from inside, a process called reverse-topology fission. Yet, in vitro, the complex primarily assembles outside membrane necks and mediates fission with normal topology. Here, we show that the direction of ESCRT-mediated membrane deformation emerges from bilayer asymmetry rather than being intrinsically encoded by the ESCRT machinery alone. Using genetic perturbations in budding yeast, we find that disruption of phospholipid asymmetry and sphingolipid homeostasis does not abolish ESCRT-dependent trafficking but renders ILV formation highly sensitive to membrane physical state, leading to inefficient cargo sorting and accumulation of stalled endosomal intermediates. In vitro reconstitution experiments and synthetic in vivo cargo systems demonstrate that asymmetric protein distribution across the membrane is sufficient to bias curvature directionality, with luminal leaflet crowding promoting efficient ILV incorporation and cytosolic crowding inhibiting inward budding. Together, these results support a model in which ESCRT-mediated membrane bending directionality emerges from the intrinsic tension difference between the bilayer leaflets. This tension difference arises from both lipid and cargo crowding-encoded asymmetries within the bilayer, rather than being solely encoded by ESCRT polymer properties.

biophysics↗

bioRxiv2026

TIMS-Bench: Towards community standards for benchmarking untargeted trapped ion mobility metabolomics tools and datasets

Untargeted liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based metabolomics is an important technology for unbiased discovery of small molecules in biomedical (e.g., drug discovery to diagnostics), animal, plant, environmental, and microbial research. Over the past decade, ion mobility has added an additional dimension to the triplet of MS1, MS2, and retention time, helping resolve co-eluting or isomeric features in an LC-MS/MS that aid in compound identification. Here, we focused on evaluating the current trapped ion mobility spectrometry (TIMS)-amenable feature-finding tools (MZmine 4.9, MS-DIAL 5.5, and MetaboScape 2025 14.0.3) for pre-processing of metabolomics data generated using a popular ion mobility mass spectrometry (IM-MS) technique, TIMS. We leveraged ten public and three benchmark TIMS datasets to evaluate these tools for their strengths and weaknesses. Our results show that MZmine consistently identified the highest number of features and confidently annotated features; however, this performance was accompanied by an increased number of false positives, due to peak splitting, as well as reduced accuracy in collision cross section (CCS) measurements. In contrast, MetaboScape achieved the highest fraction of high-quality MS2 spectra, reflecting a more conservative feature detection strategy. MS-DIAL demonstrated balanced performance, identifying features that other tools missed. Finally, we publicly release the ground-truth datasets and code to support future developments in improving IMS data analysis.

bioinformatics↗

bioRxiv2026

Nematic order in cellular tissues: a standardized framework and anomalous defect dynamics

Cellular monolayers often exhibit orientational order, with nematic alignment of cell shape and cytoskeletal structures governing tissue-scale collective dynamics. Despite extensive studies, a unified analysis framework for characterizing active nematics in living systems remains partial, and key discrepancies with theory persist. Here, we present a systematic and comparative analysis of nematic order and tissue flow dynamics across twelve distinct cell types. We quantify the impact of analysis parameters and provide data-driven guidelines to improve reproducibility and cross-study comparability. Across all nematic systems, we uncover remarkably consistent static properties, supporting the universality of nematic behavior in living tissues. By combining orientation-field analysis with velocity-field measurements and numerical simulations, we show that all examined systems display contractile active nematic signatures, with characteristic flow structures around topological defects. However, direct tracking of individual defects reveals subdiffusive dynamics, in stark contrast with the superdiffusive, self-propelled motion predicted by the hydrodynamic theory of active nematics. Our results establish a standardized framework for nematic analysis in biological systems and highlight fundamental limitations of current active nematic models in describing defect dynamics in living tissues.

biophysics↗

bioRxiv2026

Analysis of 3D epithelial tissue packing reveals correlated patterns of cell height and skewing

Epithelia are tissues which envelop organs, exchange with and protect from their environment. The tight packing of cells within an epithelium guarantees its cohesiveness and impermeability, essential to its functions. Yet, how cells spatially arrange in three dimensions within an epithelium, and how the three-dimensional cell packing responds to geometrical constraints, is not fully understood. In a proliferating epithelium, cellular volumes vary from cell to cell, notably due to cell growth. At the same time, epithelia tend to display a smooth, continuous apical surface, indicating that cell shape determinants are spatially coupled. It is unclear how these and other factors modulate cell shape variability within the epithelium. Here, we segmented three-dimensional cell shapes of MDCK epithelia, grown in hollow spheres of alginate. With this assay, we could modulate the substrate adhesive properties, tissue curvature, and cell area density. We observed that as the tissue proliferates and cell density increases, average cell volume decreases. In contrast, cell height is relatively conserved over time, with an average value sensitive to the rigidity, curvature and adhesive strength of the substrate. We identified large and spatially correlated fluctuations in cell skewing, defined as the relative difference between apical and basal area that do not arise from curvature. Skewing is associated to spatial patterns of tilted cells whose apico-basal axis deviates from orthogonality to the substrate. Surprisingly, cell skewing and cell height are correlated, indicating internal rules for three-dimensional cell shapes. Altogether, our study identifies unexpected patterns of three-dimensional cell shape variation within a proliferative epithelium.

biophysics↗

bioRxiv2026

Multiscale curvature modulates epithelial architecture and nuclear organization.

Curved geometries are a defining feature of epithelial tissues, yet how cells integrate curvature cues across spatial scales remains unclear. Here, we combined wavy hydrogels with inducible self-rolling substrates to readily and independently impose local and large-scale curvatures on epithelial monolayers, recapitulating key geometric features of bronchiolar epithelia. By controlling the orientation of local curvature relative to the large-scale curvature set by the tube axis, we show that multiscale curvature induces scale-dependent and anisotropic remodeling of cell shape, nuclear organization, and tissue thickness. In contrast, nuclei maintain a robust and conserved three-dimensional geometry, with curvature primarily regulating nuclear orientation rather than shape. This hierarchical induction of specific changes to the tissue architecture shows that multiscale curvature sensing is a fundamental physical principle governing epithelial architecture.

biophysics↗

bioRxiv2025

Molecular basis for cellular compartmentalization by an ancient membrane fission mechanism

The emergence of cell compartmentalization depends on membrane fission to create the endomembrane compartments. In eukaryotes, membrane fission is commonly executed by ESCRT-III, a protein complex conserved in all domains of life. However, whether membrane fission was an ancestral ESCRT-III activity predating eukaryogenesis remains unknown. Here we show that ESCRT-IIIA from Asgard Heimdallarchaeota, the closest archaeal relatives of eukaryotes, performs membrane fission through an N-terminal amphipathic helix, which we term Hofund. In eukaryotes, Hofund is fragmented across ESCRT-IIIA paralogs, and disrupting these regions causes severe fission defect in yeast. Remarkably, Heimdallarchaeota Hofund restores fission when fused to defective eukaryotic paralogs. These findings suggest that ESCRT-III-mediated fission arose before eukaryogenesis and later diversified to support the regulatory complexity of eukaryotic compartmentalization.

cell biology↗

bioRxiv2025

Characterisation of novel bacteriophages against the cattle pathogen Moraxella bovis

BackgroundInfectious bovine keratoconjunctivitis is the most important cattle ocular disease worldwide. The infection is primarily caused by Moraxella bovis and is a highly contagious disease that significantly affects cattle welfare. Currently, antibiotic medication is the primary treatment for infectious bovine keratoconjunctivitis. However, with rising concerns over antibiotic resistance, we propose developing a more targeted therapeutic strategy using bacteriophages (phages). Materials and MethodsWe have isolated the first known Moraxella bovis phages, characterised them according to their genome sequence, local virulence index and with transmission electron microscopy. The host ranges were assessed using 41 clinical M. bovis strains isolated from infected cows. ResultsFour phages were isolated and characterised. Comparative analysis identified a high degree of genomic similarity between the phages MB15, MB16, MB26 and MB43. MB43 was the most distinct, with the smallest host range phenotype. ConclusionsThe isolated phages show therapeutic potential for further development against Moraxella infections.

microbiology↗

bioRxiv2025

Fluorescence lifetime estimation: a practical approach using Flipper-TR FLIM

Flipper-TR is a membrane dye sensitive to lipid packing widely used to probe membrane tension in live cells via fluorescence lifetime imaging microscopy (FLIM). However, no consensus currently exists on the optimal strategy for extracting lifetime values, particularly across varying experimental setups and biological systems. Here, we systematically compare multiple approaches to estimate Flipper-TR lifetime, including multi-exponential reconvolution fitting, tail fitting, mean photon arrival time (first moment), and phasor analysis. These estimators are tested against changes in photon budget, sample characteristics, microscope manufacturer, and laser frequency. This offers a comprehensive benchmark and decision-making framework for quantitative FLIM analysis of Flipper dyes in various contexts.

biophysics↗

bioRxiv2024

TORC2-regulated sterol redistribution mediates recovery from membrane perturbation by small amphipathic molecules.

To maintain plasma membrane (PM) integrity, cells need to acutely regulate PM lipid composition. The Target Of Rapamycin (TOR) complex 2 is a protein kinase that acts as a central regulator of PM homeostasis, but the mechanisms by which it monitors and reacts to membrane stresses are poorly understood. To address this knowledge gap, we characterized a family of amphiphilic molecules that physically perturb PM organization and in doing so inhibit TORC2 in yeast and mammalian cells. Using fluorescent lipid associated reporters in budding yeast, we show that these small molecules first cause a transient increase in the amount of biochemically accessible ergosterol at the PM. Contemporaneous TORC2 inhibition stimulates a rapid removal of accessible ergosterol from the PM by the PM-ER sterol transporters Lam2 and Lam4, necessary for TORC2 reactivation. Thus, we show that TORC2 acts in a feedback loop to control active sterol levels at the PM and introduce sterols as possible TORC2 signalling modulators.

cell biology↗

bioRxiv2024

Immunological Profiling in Knee Osteoarthritis: Treg Dysfunction as Key Driver of Pain

Pain is the hallmark symptom of osteoarthritis (OA) and its biological drivers remain poorly understood. While the role of innate immunity in OA has been extensively studied, the involvement of adaptive immunity, in particular regulatory T cells (Tregs), is not well understood. Using a comprehensive multi-omic approach on the peripheral blood from 46 knee OA patients with similar radiographic stage, including deep immunophenotyping, cytokine profiling, transcriptomic and T-cell receptor analysis on sorted CD4 Tregs and effector T cells (Teff), we identified an immunological signature associated with OA-related pain. Cytokines promoting Treg expansion and activation (with increases of sIL2-RA, sTNFR1, sTNFR2) were correlated with the Western Ontario and McMaster Universities Arthritis Index (WOMAC) pain subscore, suggesting a potential Treg dysfunction. Nineteen T cell subsets were correlated with WOMAC pain. Notably, we found a negative correlation of cell subsets associated with Treg expansion and activation (FoxP3+CTLA4+, CD4+CD57+, Treg CD95+, CD4 Treg CD45RA-). Differential gene expression analysis between patients with low and high WOMAC pain intensity (threshold [≥] 40/100) revealed an upregulation of inflammasome-related genes such as IL1RL1, IL31RA, IFITM3, NLRP3, IFNG in Tregs. Functional enrichment analysis highlighted an overrepresentation of innate immune response, IL-8, and interferon activation pathways suggesting a pro-inflammatory state in Tregs of patients with high pain intensity. Collectively, our systems immunology approach highlights multiple associations between Treg dysfunctionality and OA-related pain, providing new insights into the adaptive immune systems contribution to OA-related pain.

immunology↗

bioRxiv2024

Two-dimensional condensates of HRS drive the assembly of flat clathrin lattices on endosomes

In cells, the curved clathrin structures in vesicle budding are well characterized, while the flat ones remain poorly understood. We reconstituted the flat assembly of ESCRT-0 protein HRS and clathrin onto lipid membranes in vitro. HRS was found to form protein condensates. These condensates spread as a two-dimensional layer on negatively charged membranes and promoted the assembly of clathrin into a flat coat. Correlative cryo-tomography of HRS-labeled endosomes revealed a pure hexagonal lattice, consistent with flat clathrin structures. Cholesterol enhanced HRS recruitment to the membrane both in cells and in supported bilayers. Furthermore, cholesterol promoted the phase separation of HRS onto membranes, which in turn concentrated cholesterol underneath. This positive feedback promoted the formation of HRS-clathrin microdomains that sorted reconstituted ubiquitinated cargoes. Altogether, our results show that the unique architecture of ESCRT-0 is assembled by the two-dimensional phase-separation of HRS which drives the assembly of flat clathrin coats.

biochemistry↗

bioRxiv2024

Actin dynamics sustains spatial gradients of membrane tension in adherent cells

Tension propagates in lipid bilayers over hundreds of microns within milliseconds, precluding the formation of tension gradients. Nevertheless, plasma membrane tension gradients have been evidenced in migrating cells and along axons. Here, using a fluorescent membrane tension probe, we show that membrane tension gradients exist in all adherent cells, whether they migrate or not. Non-adhering cells do not display tension gradients. We further show that branched actin increases tension, while membrane-to-cortex attachments facilitate its propagation. Tension is the lowest at the edge of adhesion sites and highest at protrusions, setting the boundaries of the tension gradients. By providing a quantitative and mechanistic basis behind the organization of membrane tension gradients, our work explains how they are actively sustained in adherent cells.

cell biology↗

bioRxiv2024

Evolutionarily conserved principles of ESCRT-III-mediated membrane remodelling revealed by a two-subunit Asgard archaeal system

ESCRT-III proteins assemble into composite polymers that undergo stepwise changes in composition and structure to deform membranes across the tree of life. Here, using a phylogenetic analysis we demonstrate that the two ESCRT-III proteins present in our closest archaeal relatives are evolutionarily related to B-type and A-type eukaryotic paralogues, which initiate and execute membrane remodelling, respectively. This deep homology is reflected in ESCRT-III structure and function as demonstrated by the fact that ESCRT-IIIB assembles into parallel arrays on planar membranes to initiate membrane deformation, and is required to recruit ESCRT-IIIA to generate composite polymers. ESCRT-IIIA homopolymers can then remodel membranes into tubes, as a likely prelude to scission. Taken together, this analysis reveals a set of conserved principles governing ESCRT-III-dependent membrane remodelling that first emerged with the evolution of a two-component ESCRT-III system in the Asgard archaea, and which continue to underlie complex multi-component, ESCRT-III-dependent membrane remodelling in eukaryotes.

evolutionary biology↗

bioRxiv2024

Topology changes of the regenerating Hydra define actin nematic defects as mechanical organizers of morphogenesis.

AbstractHydra is named after the mythological animal for its regenerative capabilities, but contrary to its mythological counterpart, it only regenerates one head when cut. Here we show that soft compression of head regenerating tissues induces the regeneration of viable, two headed animals. Topological defects in the supracellular nematic organization of actin were previously correlated with the new head regeneration site1. Soft compression creates new topological defects associated with additional heads. To test the necessity of topological defects in head regeneration, we changed the topology of the tissue. By compressing the head regenerating tissues along their body axis, topological defects of the foot and of the regenerating head fused together, forming a toroid with no defects. Perfectly ordered toroids did not regenerate over eight days and eventually disintegrated. Spheroids made from excised body column tissue partially lose their actin order during regeneration. Compression of spheroids generated toroids with actin defects. These tissues regenerated into toroidal animals with functional head and foot, and a bifurcated body. Our results show that topological defects in the actin order are necessary to shape the head of the regenerating Hydra, supporting the notion that actin topological defects are mechanical organizers of morphogenesis.

developmental biology↗

bioRxiv2023

Mechanism for Vipp1 spiral formation, ring biogenesis and membrane repair

ESCRT-III family proteins build dynamic filaments that remodel membrane. Although the transition of planar filaments into 3D membrane budding structures is fundamental for their function, the geometric changes in polymer architecture driving the transition remain obscure. Here we show how bacterial Vipp1 polymerises into dynamic planar sheets and spirals on membrane. The spirals converge to form a central ring like those known to bud membrane. To probe how Vipp1 morphs between polymers, we determine the architecture of multiple helical filaments. As well as describing filament constriction and membrane tubulation, the geometric relationship between helical and planar lattices enables Vipp1 sheets and spirals to be modelled. Moreover, the helical structures show filaments twisting - a process needed for Vipp1 to transition between planar and 3D architectures. Given the structural conservation between Vipp1 and ESCRT-III, our results may represent the broad changes in geometry required for some ESCRT-III filaments to switch between 2D and 3D forms.

microbiology↗

bioRxiv2023

CryoEM architecture of a native stretch-sensitive membrane microdomain

Biological membranes are partitioned into functional zones containing specific lipids and proteins, termed membrane microdomains. Their composition and organization remain controversial owing to a paucity of techniques that can visualize lipids in situ without disrupting their native behavior1,2. The yeast eisosome, a membrane compartment scaffolded by the BAR-domain proteins Pil1 and Lsp1, senses and responds to mechanical stress by flattening and releasing sequestered factors3-7. Here, we isolated native eisosomes as helical filaments of Pil1/Lsp1 lattice bound to plasma membrane lipids and solved their structures by helical reconstruction. We observe remarkable organization within the lipid bilayer density from which we could assign headgroups of PI(4,5)P2 and phosphatidylserine bound to Pil1/Lsp1 and a pattern of membrane voids, signatures of sterols, beneath an amphipathic helix. We verified these assignments using in vitro reconstitutions and molecular dynamics simulations. 3D variability analysis of the native eisosomes revealed a dynamic stretching of the Pil1/Lsp1 lattice that affects functionally important lipid sequestration, supporting a mechanism in which membrane stretching liberates lipids otherwise anchored by the Pil1/Lsp1 coat. Our results provide mechanistic insight into how eisosome BAR-domain proteins create a mechanosensitive membrane microdomain and, more globally, resolve long-standing controversies about the architecture and nature of lipid microdomains.

biophysics↗