Biology subjects

Le Borgne, R.

Publications and source records attributed to Le Borgne, R..

At least 19 recordsLinked to original sources

bioRxiv2026

The Endoplasmic Reticulum as a Cytoplasmic Anchor for Microtubule-Driven Aster Centration

The proper positioning of microtubule (MT) asters is essential to regulate division positioning and multi-cellular morphogenesis, yet the mechanisms regulating this process remain poorly understood. Using sea urchin zygotes as a model, we combined high-resolution imaging, perturbation assays, and 3D modeling to investigate the nature of forces driving sperm aster centration after fertilization. We show that centering asters are highly anisotropic, featuring long microtubules extending at the aster front that may engage with organelles in bulk cytoplasm to generate pulling forces. While vesicular compartments, like acidic organelles, mitochondria or yolk, contributed minimally, we found that the endoplasmic reticulum (ER) undergoes a massive compaction around the centering aster and drags cytosol to produce a reactive centering force on the aster. Perturbing ER organization by cytoplasmic stratification or Atlastin-mediated fragmentation disrupted aster centration. A two-fluid model suggests that the frictional interaction between the compacting ER and cytosol fluid is sufficient to recapitulate key features of aster centration. These findings identify the ER as an important cytoplasmic anchor for MT force exertion in bulk cytoplasm, to guide aster centering motion and influence early embryonic division geometries. More broadly, our results highlight the role of cytoplasmic architecture in shaping the mechanics of organelle positioning during development.

cell biology↗

bioRxiv2026

Mechanosensation Promotes Local Cell Wall Repair

Walled cells, such as those of fungi, plants and bacteria, must continuously survey the integrity of their protective cell wall (CW), as they grow, divide, or face mechanical challenges from their high internal turgor and environment. To date, however, how cells detect defects in their CW to activate local reinforcement or repair machineries remains unclear. We implemented a laser irradiation assay to locally wound the CW of model rod-shaped fission yeast cells. We found that laser irradiation causes a near instantaneous local thinning of the CW accompanied by the formation of a local bulge, a drop in turgor and growth arrest, followed by a progressive recovery of CW thickness over tens of minutes with cells eventually resuming growth. Remarkably, wounding the CW away from cell tips, caused the re-localization of canonical polarity regulators including the Rho GTPases Cdc42 and Rho1, that recruited actin assembly and vesicular trafficking of CW synthases to repair the CW. A candidate screen approach suggests that the transmembrane surface mechanosensors Wsc1 and Mlt2 detect local CW defects and promote the recruitment of the Rho GEFs Gef1 and Rgf1 that activate Cdc42 and Rho1. Therefore, these findings delineate a mechanochemical pathway, coupling surface mechanosensing to polarity regulation and CW synthesis, that defines how walled cells may detect and repair local defects in their CWs to safeguard surface integrity.

cell biology↗

bioRxiv2026

A Cell Size-Dependent Competition Between Geometry and Polarity Governs Nuclear and Spindle positioning in Early Embryos

Nuclei and mitotic spindles are actively positioned at defined locations within cells to regulate cell polarity, division and multicellular morphogenesis1-4. Forces generated by cytoskeleton networks regulate the positioning of these organelles and are commonly influenced by extrinsic cues such as cell geometry or polarity5-12. To date, however, most studies have investigated this problem in one given cell type, hampering our understanding for how mechanical systems that position nuclei and spindles may scale during multicellular development. We tracked the spatiotemporal behaviour of centrosomes, nuclei and spindles in early sea urchin embryos from the 1-cell to the [~]1000 cells blastula stage. We found that they are initially located at cell centers, but that they undergo a progressive decentration towards the embryo apical surface, as cells become smaller during development. This apical shift is mediated by microtubule (MTs) pulling forces which are influenced by both cell shapes and apical polarity domains. Using 3D mathematical models and embryo dissections, we propose that MT centering forces that derive from cell geometry decay in strength during development as a consequence of cell size reduction, allowing apical polarity decentering forces to take over. Our results support a self-organized scenario in which polarity cues progressively outcompete cell geometry, to modulate the overall balance of MT forces and pattern nuclear and spindle positioning throughout early embryo development.

cell biology↗

bioRxiv2024

An interkinetic envelope surrounds chromosomes between meiosis I and II in C. elegans oocytes

At the end of cell division, the nuclear envelope reassembles around the decondensing chromosomes. Female meiosis culminates in two consecutive cell divisions of the oocyte, meiosis I and II, which are separated by a brief transition phase known as interkinesis. Due to the absence of chromosome decondensation and the suppression of genome replication during interkinesis, it has been widely assumed that the nuclear envelope does not reassemble between meiosis I and II. By analyzing interkinesis in C. elegans oocytes, we instead show that an atypical structure made of two lipid bilayers, which we termed the interkinetic envelope, surrounds the surface of the segregating chromosomes. The interkinetic envelope shares common features with the nuclear envelope but also exhibits specific characteristics that distinguish it, including its lack of continuity with the endoplasmic reticulum, unique protein composition, assembly mechanism, and function in chromosome segregation. These distinct attributes collectively define the interkinetic envelope as a unique and specialized structure that has been previously overlooked.

cell biology↗

bioRxiv2024

The directionality of collective cell delamination is governed by tissue architecture and cell adhesion in a Drosophila carcinoma model

Carcinomas originate in epithelia and are the most frequent type of cancers. Tumor progression starts with the collective delamination of live tumors out of the epithelium, and requires modulation of cell adhesion. Yet, it remains elusive exactly how remodeling of epithelial cell-cell and cell-extracellular matrix contacts contribute to metastasis onset and delamination directionality. We used Drosophila melanogaster larval eye disc to study cooperative oncogenesis and determine the contribution of bi- and tricellular septate junction (SJ) components to tumor progression in flat and pseudostratified epithelia. We reveal that loss-of-function of septate junction components alone can promote cell death whereas synergic interaction with oncogenic Ras triggers the collective delamination of living tumorigenic cells. Spatiotemporal analyses reveal apical and basal delamination processes differ in terms of cell identity, cell polarity and remodeling of cell-cell and cell-ECM contacts. Using a combination of in vivo and in silico approaches, we report that SJ-depleted RasV12 tumors in pseudostratified epithelia trigger tissue folding or basal collective delamination regardless of the order in which cell contacts are remodeled. In striking contrast, SJ-depleted RasV12 tumors formed in flat, squamous epithelia, first undergo apical constriction and acquire a dome-like shape. Tumors are enriched in adhesion molecules and form an apical neck at the interface of contact between mutant cells and wild-type neighbors. Concomitant to cytoskeleton remodeling, tumors emit apical protrusions in the lumen and progressively delaminate through the apical neck while remaining cohesive. Our study reveals that tissue architecture and changes in cell adhesion drive the directionality of collective delamination of neoplastic tumors out of an epithelium.

developmental biology↗

bioRxiv2024

Myocardial disarray drives metabolic inefficiency in human cardiomyocytes

Adult cardiomyocytes are embedded within a highly organized myocardial microenvironment that imposes critical geometric cues essential for the alignment and distribution of organelles and the shaping of their unique, rectangular cellular morphology. Despite the association of cardiomyocyte disarray with human heart disease, the functional consequences of this cellular disorganization remain poorly understood. Here, we leveraged micropatterned substrates to promote structural alignment in human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs), contrasting the effects of mechanical alignment on mitochondrial form and function with hiPSC-CMs cultured under standard unconstrained conditions. Cardiomyocytes cultured under unconstrained conditions exhibited misaligned sarcomeres and a perinuclear mitochondrial distribution while micropatterned hiPSC-CMs developed linear myofibrils and reconfigured sarcomere and mitochondrial organization, which increased mitochondrial respiration without augmenting mitochondrial mass. Notably, micropatterned hiPSC-CMs exhibited an increased number of mitochondrial-associated membranes, as determined by proximity ligation assays and transmission electron microscopy, suggesting enhanced interactions between the sarcoplasmic reticulum and mitochondria. Together, these findings demonstrate that mitochondrial-sarcoplasmic architecture and geometry are critical spatial features that ensure bioenergetic efficiency of cardiomyocytes. This work underscores the importance of cellular organization in cardiomyocyte metabolism and function, providing insights into the pathophysiology of cardiac diseases marked by cellular disarray.

cell biology↗

bioRxiv2024

Oncogenic Ras-driven Dorsal/NF-κB signaling contributes to tumorigenesis in a Drosophila carcinoma model

Cancer-driving mutations synergize with inflammatory stress signaling pathways during carcinogenesis. Drosophila melanogaster tumour models are increasingly recognized as models to inform conserved molecular mechanisms of tumorigenesis with both local and systemic effects of cancer. Although initial discoveries of the Toll-NF{kappa}B signaling pathway in development and immunity was pioneered in Drosophila, limited information is available for its role in cancer progression. Using a well-studied cooperative RasV12 -driven epithelial-derived tumour model, we here describe functions of Toll-NF-{kappa}B signaling in malignant RasV12, scrib- tumors. The extracellular Toll pathway components ModSP and PGRP-SA and intracellular signaling Kinase, Pelle/IRAK, are rate-limiting for tumor growth. The Toll pathway NF{kappa}B protein Dorsal, as well as cactus/I{kappa}B show elevated expression in tumors with highest expression in invasive cell populations. Oncogenic RasV12, and not loss of scribble, confers increased expression and heterogenous distribution of two Dorsal isoforms, DorsalA and DorsalB in different tumour cell populations. Mechanistic analyses demonstrates that Dorsal drives growth and malignancy by suppressing differentiation, counteracting apoptosis and promoting invasion of RasV12, scrib- tumors genetically dependent on twist and snail.

cancer biology↗

bioRxiv2023

ESCRT-III-dependent adhesive and mechanical changes are triggered by a mechanism sensing paracellular diffusion barrier alteration in Drosophila epithelial cells

Barrier functions of proliferative epithelia are constantly challenged by mechanical and chemical constraints. How epithelia respond to and cope with disturbances of the paracellular diffusion barrier to allow tissue integrity maintenance has been poorly characterized. Cellular junctions play an important role in this process and intracellular traffic contribute to their homeostasis. Here, we reveal that, in Drosophila pupal notum, alteration of the bi- or tricellular septate junctions (SJs) triggers a mechanism with two prominent outcomes. On one hand, there is an increase in the levels of E-cadherin, F- Actin and non-muscle myosin II in the plane of adherens junctions. On the other hand, {beta}-integrin/Vinculin-positive cell contacts are reinforced along the lateral and basal membranes. We report that the weakening of SJ integrity, caused by the depletion of bi- or tricellular SJ components, reduces ESCRT-III/Vps32/Shrub-dependent degradation and promotes instead Retromer-dependent recycling of SJ components. The consequence of the reduction in Shrub-dependent degradation extends to other transmembrane protein cargoes. Consequently, this trigger increased levels of {beta}- integrin, Crumbs and the Crumbs effectors {beta}-Heavy Spectrin Karst. We propose a mechanism by which epithelial cells, upon sensing alterations in the paracellular diffusion barrier, target Shrub to adjust the degradation/recycling balance and thereby compensate for barrier defects while maintaining epithelial integrity.

developmental biology↗

bioRxiv2023

Fascin-induced bundling protects actin filaments from disassembly by cofilin

Actin filament turnover plays a central role in shaping actin networks, yet the feedback mechanism between network architecture and filament assembly dynamics remains unclear. The activity of ADF/cofilin, the main protein family responsible for filament disassembly, has been mainly studied at the single filament level. Here, we report that fascin, by crosslinking filaments into bundles, strongly slows down filament disassembly by cofilin. We show that this is mainly due to a slower nucleation of the first cofilin clusters, which occurs up to 100-fold slower on large bundles compared to single filaments. In contrast, severing at cofilin cluster boundaries is unaffected by fascin bundling. After the nucleation of an initial cofilin cluster on a filament of a bundle, we observe the local removal of fascin. Surprisingly, the nucleation of cofilin clusters on adjacent filaments is highly enhanced, locally. We propose that this inter-filament cooperativity in cofilin binding arises from the local propagation of the cofilin-induced change in helicity from one filament to the other filaments of the bundle. Taken together, these observations reveal the molecular events explaining why, despite inter-filament cooperativity, fascin crosslinking protects actin filaments from cofilin-induced disassembly. These findings highlight the important role played by crosslinkers in organizing actin networks and modulating the activity of other regulatory proteins.

biochemistry↗

bioRxiv2022

Cell-intrinsic and -extrinsic functions of the ESCRT-III component Shrub in cytokinetic abscission of Drosophila Sensory Organ precursor

While the molecular mechanisms underlying the abscission of isolated cells are largely decrypted, those of fast-cycling, epithelial progenitors surrounded by epidermal cells (ECs) connected by junctions remain largely unexplored. Here, we investigated the remodeling of the permeability barrier ensured by septate junctions (SJs) during cytokinesis of Drosophila sensory organ precursor (SOP). We report that SOP cytokinesis involves the coordinated polarized assembly and remodeling of SJs in the dividing cell and its neighbors, which remained connected via membrane protrusions pointing toward the SOP midbody. SJs assembly and midbody basal displacement occur more rapidly in SOP than in ECs, leading to a faster disentanglement of the protrusions that precedes midbody release. As reported in isolated cells, the endosomal sorting complex required for transport-III component Shrub/CHMP4B is recruited at the midbody and cell-autonomously regulates abscission. In addition, we found that Shrub is recruited to membrane protrusions, is required for SJ integrity, and that alteration of SJ integrity leads to premature abscission. Our study uncovers cell-intrinsic and -extrinsic functions of Shrub in epithelial abscission to support the coordination of permeability barrier maintenance and abscission in SOPs.

developmental biology↗

bioRxiv2022

The ER tether VAPA is required for proper cell motility and for anchoring ER-plasma membrane contact sites to focal adhesions

Cell motility processes highly depend on the membrane distribution of Phosphoinositides (PInst), giving rise to cytoskeleton reshaping and membrane trafficking events. Membrane contact sites serve as platforms for lipid exchange and calcium fluxes between two organelles. Here, we show that VAPA, an ER membrane-resident contact site tether, plays a crucial role during cell motility. CaCo2 adenocarcinoma epithelial cells depleted for VAPA exhibit several collective and individual motility defects, disorganized actin cytoskeleton and altered protrusive activity. During migration, VAPA is required for the maintenance of PI(4,5)P2 levels at the plasma membrane, but not for PI(4)P homeostasis in the Golgi and endosomal compartments. Importantly, we show that VAPA regulates the dynamics of focal adhesions (FA) through its MSP domain, and is essential to stabilize and anchor ventral ER-PM contact sites to FA, thus mediating microtubule-dependent FA disassembly. To conclude, our results reveal unprecedented functions for VAPA-mediated membrane contact sites during cell motility and provides a dynamic picture of ER-PM contact sites connection with FA mediated by VAPA.

cell biology↗

bioRxiv2022

Cell Wall Dynamics in a Filamentous Fungus

Hyphal tip growth allows filamentous fungi to colonize space, reproduce or infect. It features remarkable morphogenetic plasticity including unusually fast elongation rates, tip turning, branching or bulging. These shape changes are all driven from the expansion of a protective cell wall (CW) secreted from apical pools of exocytic vesicles. How CW secretion, remodeling and deformation are modulated in concert to support rapid tip growth and morphogenesis while ensuring surface integrity remains poorly understood. We implemented sub-resolution imaging to map the dynamics of CW thickness and secretory vesicles in Aspergillus nidulans. We found that tip growth is associated with balanced rates of CW secretion and expansion, which limit temporal fluctuations in CW thickness, elongation speed and vesicle amount, to less than 10-15%. Affecting this balance through modulations of growth or trafficking yield to near-immediate changes in CW thickness, mechanics and shape. We developed a model with mechanical feedback which accounts for steady states of hyphal growth as well as rapid adaptation of CW mechanics and vesicle recruitment to different perturbations. These data provide unprecedented details on how CW dynamics emerges from material secretion and expansion, to stabilize fungal tip growth as well as promote its morphogenetic plasticity.

cell biology↗

bioRxiv2022

The AMA1-RON complex drives Plasmodium sporozoite invasion in the mosquito and mammalian hosts

Plasmodium sporozoites that are transmitted by blood-feeding female Anopheles mosquitoes invade hepatocytes for an initial round of intracellular replication, leading to the release of merozoites that invade and multiply within red blood cells. Sporozoites and merozoites share a number of proteins that are expressed by both stages, including the Apical Membrane Antigen 1 (AMA1) and the Rhoptry Neck Proteins (RONs). Although AMA1 and RONs are essential for merozoite invasion of erythrocytes during asexual blood stage replication of the parasite, their function in sporozoites is still unclear. Here we show that AMA1 interacts with RONs in mature sporozoites. By using DiCre-mediated conditional gene deletion in P. berghei, we demonstrate that loss of AMA1, RON2 or RON4 in sporozoites impairs colonization of the mosquito salivary glands and invasion of mammalian hepatocytes, without affecting transcellular parasite migration. Our data establish that AMA1 and RONs facilitate host cell invasion across Plasmodium invasive stages, and suggest that sporozoites use the AMA1-RON complex to safely enter the mosquito salivary glands without causing cell damage, to ensure successful parasite transmission. These results open up the possibility of targeting the AMA1-RON complex for transmission-blocking antimalarial strategies.

microbiology↗

bioRxiv2021

Contribution of cytoplasm viscoelastic properties to mitotic spindle positioning

Cells are filled with macromolecules and polymer networks that set scale-dependent viscous and elastic properties to the cytoplasm. Although the role of these parameters in molecular diffusion, reaction kinetics and cellular biochemistry is being increasingly recognized, their contributions to the motion and positioning of larger organelles, such as mitotic spindles for cell division remain unknown. Here, using magnetic tweezers to displace and rotate mitotic spindles in living embryos, we uncovered that the cytoplasm can impart viscoelastic reactive forces that move spindles, or passive objects with similar size, back to their original position. These forces are independent of cytoskeletal force generators, yet reach hundreds of piconewtons and scale with cytoplasm crowding. Spindle motion shears and fluidizes the cytoplasm, dissipating elastic energy and limiting spindle recoils with functional implications for asymmetric and oriented divisions. These findings suggest that bulk cytoplasm material properties may constitute important control elements for the regulation of division positioning and cellular organization. Significance StatementThe regulation of mitotic spindle positioning is a key process for tissue architecture, embryo development and stem cells. To date, most models have assumed that spindles are positioned by forces exerted by polar cytoskeleton networks, like microtubule asters or acto-myosin bundles. Here, using in situ magnetic tweezers to apply calibrated forces and torques to mitotic spindles in live dividing sea urchin cells, we found that the viscoelastic properties of the cytoplasm medium in which spindles are embedded can hold spindles in place, and move them back if their original position is perturbed. These viscoelastic forces are large and may significantly participate in the force balance that position and orient mitotic spindles in many cell types.

cell biology↗

bioRxiv2021

Trans-cellular tunnels induced by the fungal pathogen Candida albicans facilitate invasion through successive epithelial cells without host damage

The opportunistic fungal pathogen Candida albicans is normally commensal, residing in the mucosa of most healthy individuals. In susceptible hosts, its filamentous hyphal form can invade epithelial layers leading to superficial or severe systemic infection. Invasion is mainly intracellular, though it causes no apparent damage to host cells. We investigated the invasive lifestyle of C. albicans in vitro using live-cell imaging and the damage-sensitive reporter galectin-3. Quantitative single cell analysis showed that invasion can result in host membrane breaching at different stages of invasion and cell death, or in traversal of host cells without membrane breaching. Membrane labelling and three-dimensional "volume" electron microscopy revealed that hyphae can traverse several host cells within trans-cellular tunnels that are progressively remodelled and may undergo inflations linked to host glycogen stores, possibly during nutrient uptake. Thus, C. albicans invade epithelial tissues by either inflicting or avoiding host damage, the latter facilitated by trans-cellular tunnelling.

microbiology↗

bioRxiv2020

Interplay between Anakonda, Gliotactin and M6 for tricellular junction assembly and anchoring of septate junctions in Drosophila epithelium

In epithelia, Tricellular junctions (TCJs) serve as pivotal sites for barrier function and integration of both biochemical and mechanical signals. While essential for tissue homeostasis, TCJ assembly, composition and links to adjacent bicellular junctions (BCJs) remain poorly understood. Here we have characterized the assembly of TCJs within the plane of adherens junctions (tAJ) and the plane of septate junctions (tSJ) in Drosophila and report that their formation is spatiotemporally decoupled. The assembly and stabilization of previously described tSJ components Anakonda (Aka) and Gliotactin (Gli) as well as the newly reported tSJ proteolipid protein M6, is shown to be a complex process. Aka and M6, whose localization is interdependent, act upstream to locate Gli. In turn, Gli stabilizes Aka at tSJ. Those results unravel a previous unknown role of M6 at tSJ and a tight interplay between tSJ components to assemble and maintain tSJs. In addition, tSJ components are not only essential at vertex as we found that loss of tSJ integrity also induces micron-length bicellular SJs deformations that are free of tensile forces. This phenotype is associated with the disappearance of SJ components at tricellular contacts, indicating that bSJ are no longer connected to tSJs. Reciprocally, SJ components are in turn required to restrict the localization of Aka and Gli at vertex. We propose that tSJs function as pillars to anchor bSJs to ensure the maintenance of tissue integrity in Drosophila proliferative epithelia.

developmental biology↗

bioRxiv2020

Mitochondrial Aurora kinase A induces mitophagy by interacting with MAP1LC3 and Prohibitin 2

Epithelial and haematologic tumours often show the overexpression of the serine/threonine kinase AURKA. Recently, AURKA was shown to localise at mitochondria, where it regulates mitochondrial dynamics and ATP production. Here we define the molecular mechanisms of AURKA in regulating mitochondrial turnover by mitophagy. When overexpressed, AURKA induces the disappearance of Outer Mitochondrial Membrane proteins by the proteasome. Then, AURKA triggers the degradation of Inner Mitochondrial Membrane (IMM)/matrix proteins by interacting with core components of the autophagy pathway. On the IMM, the kinase forms a tripartite complex with MAP1LC3 and the mitophagy receptor PHB2. This complex is necessary to trigger mitophagy in a PARK2/Parkin-independent manner. The formation of the tripartite complex is induced by the phosphorylation of PHB2 on Ser39, which is required for MAP1LC3 to interact with PHB2. Last, treatment with the PHB2 ligand Xanthohumol blocks AURKA-induced mitophagy by destabilising the tripartite complex. This treatment also restores normal ATP production levels. Altogether, these data provide evidence for a previously undetected role of AURKA in promoting mitophagy through the interaction with PHB2 and MAP1LC3. This work paves the way to the use of function-specific pharmacological inhibitors to counteract the effects of the overexpression of AURKA in cancer.

cell biology↗

bioRxiv2020

Super-resolved live-cell imaging using Random Illumination Microscopy

Super-resolution fluorescence microscopy has been instrumental to progress in biology. Yet, the photo-induced toxicity, the loss of resolution into scattering samples or the complexity of the experimental setups curtail its general use for functional cell imaging. Here, we describe a new technology for tissue imaging reaching a 114nm/8Hz resolution at 30 {micro}m depth. Random Illumination Microscopy (RIM) consists in shining the sample with uncontrolled speckles and extracting a high-fidelity super-resolved image from the variance of the data using a reconstruction scheme accounting for the spatial correlation of the illuminations. Super-resolution unaffected by optical aberrations, undetectable phototoxicity, fast image acquisition rate and ease of use, altogether, make RIM ideally suited for functional live cell imaging in situ. RIM ability to image molecular and cellular processes in three dimensions and at high resolution is demonstrated in a wide range of biological situations such as the motion of Myosin II minifilaments in Drosophila.

bioengineering↗