Biology subjects

Delevoye, C.

Publications and source records attributed to Delevoye, C..

14 recordsLinked to original sources

bioRxiv2025

Vimentin intermediate filaments support autophagosome biogenesis at ER-endosomes contact sites in response to starvation

Macroautophagy (autophagy) is a fundamental catabolic process requiring the biogenesis of the autophagosome to support cell survival during stress. While the roles of F-actin and microtubule cytoskeleton in autophagy are well established, the contribution of intermediate filaments (IFs) remains poorly understood. Here, we investigated the role of the type III IF vimentin in supporting the early steps of starvation-induced autophagy. We demonstrate that starvation triggers a rapid, perinuclear compaction of vimentin IFs, correlating with transient phosphorylation at serine 56 and enhanced overlap with the endoplasmic reticulum (ER). We reveal that autophagic proteins accumulate at the vimentin/ER interface, physically connecting the autophagosome biogenesis machinery to the vimentin IF network. Knock-out or pharmacological perturbation of vimentin-IFs dynamics using Withaferin-A significantly impairs starvation-induced autophagic flux. Mechanistically, we reveal that vimentin IFs are essential coordinators for the mobilization of endosome-ER-membrane contact sites (EERCS), a critical hub for autophagosome nucleation. Together, our findings uncover a novel role for vimentin IFs as a dynamic cytoskeletal coordinator that spatially organizes membrane contact sites to promote the efficient initiation of autophagosome biogenesis in response to nutrient stress. Summary statementThis study reveals that vimentin intermediate filaments rapidly reorganize to mobilize ER-endosome contact sites, establishing a critical spatial platform for starvation-induced autophagy initiation.

cell biology↗

bioRxiv2025

KIF20A cooperates with Myosin II to drive endosomal carriers fission and protein trafficking

Formation of membrane transport carrier relies on cytoskeleton coordination, however how microtubules and actin dynamics cooperate is still poorly understood. In this study, we reveal a novel mechanism by which a kinesin, KIF20A, regulates actomyosin and branched actin dynamics in coordination with Myosin II, facilitating the fission of transport intermediates at early endosomes. We demonstrate that KIF20A is required for maintaining endosomal homeostasis as its inhibition results in the enlargement of both early and late endosomes. Moreover, our findings demonstrate that KIF20A is required for the proper trafficking of Transferrin (Tf), and for ensuring the correct plasma membrane localization of {beta}1 Integrin, a key protein that is critical for cell adhesion and migration. Collectively, these results highlight a new pivotal role of KIF20A as a key regulator of endosomal dynamics and function, contributing to metastatic potential.

cell biology↗

bioRxiv2025

Keratin intermediate filaments mechanically position melanin pigments for genome photoprotection

Melanin pigments block genotoxic agents by positioning on the sun-exposed side of human skin keratinocytes nucleus. How this position is regulated and its role in genome photoprotection remains unknown. By developing a model of human keratinocytes internalizing extracellular melanin into pigment organelles, we show that keratin 5/14 intermediate filaments mechanically control the 3D perinuclear position of pigments, shielding DNA from photodamage. Imaging and microrheology in human disease-related model identify structural keratin cages surrounding pigment organelles to stiffen their microenvironment and maintain their 3D position. Optimum pigment spatialization is required for DNA photoprotection and rely on the interplay between intermediate filaments and microtubules bridged by plectin cytolinkers. Thus, the mechanically-driven proximity of pigment organelles to the nucleus is a key photoprotective parameter. Uncovering how human skin counteracts solar radiation by positioning the melanin microparasol next to the genome anticipates that dynamic spatialization of organelles is a physiological UV stress response. Short summaryMelanin pigments shield DNA from photodamage by positioning atop nuclei in skin keratinocytes. We show keratin 5/14 intermediate filaments control this 3D spatialization, forming protective cages around pigments. This positioning, together with microtubule function, optimizes genome protection, revealing cytoskeletons and organelle dynamics as a UV stress response.

cell biology↗

bioRxiv2024

Influenza A virus-induced production of PI4P at the endoplasmic reticulum involves ATG16L1 and promotes the egress of viral ribonucleoproteins

The genomic RNAs of influenza A viruses (IAVs) are replicated in the nucleus of infected cells in the form of viral ribonucleoproteins (vRNP) before being exported to the cytoplasm. The small GTPase RAB11A is involved in the transport of vRNPs to the sites of viral assembly at the plasma membrane, but the molecular mechanisms involved remain largely unknown. Here we show that IAV infection remodels the architecture of the endoplasmic reticulum (ER) sheets, where vRNPs tend to accumulate in the absence of RAB11A. To decipher the interplay between RAB11A, vRNPs and the ER, we investigated viral-induced perturbations of RAB11A proximity interactome. To this end, we generated cells stably expressing a TurboID-RAB11A fusion protein and performed biotin-based proximity labeling upon viral infection. We found that cellular regulators of phophatidylinositol-4-phosphate (PI4P) homeostasis, including the autophagic and stress response protein ATG16L1, are significantly enriched at the vicinity of RAB11A in infected cells. Infection induces an increase in cellular PI4P levels in an ATG16L1-dependent manner, while ATG16L1 relocalizes to ER membranes upon infection. Depletion of ATG16L1 decreases the co-distribution of vRNPs with PI4P punctae on ER membranes, and reduces the accumulation of vRNPs at the plasma membrane as well as the production of IAV infectious particles. Our data extend to IAVs the notion that viruses can modulate the metabolism and localization of phosphoinositides to control host membrane dynamics and point to the ER as an essential platform for vRNP transport. They provide evidence for a pivotal role of ATG16L1 in regulating the identity of endomembranes and coordinating RAB11A and PI4P-enriched membranes to ensure delivery of vRNPs to the plasma membrane.

microbiology↗

bioRxiv2023

How Myosin VI Traps its Off-State, is Activated and Dimerizes

Myosin VI (Myo6) is the only minus-end directed nanomotor on actin, allowing it to uniquely contribute to numerous cellular functions. As for other nanomotors, proper functioning of Myo6 relies on precise spatio-temporal control of motor activity via a poorly defined off-state and interactions with partners. Our structural, functional, and cellular studies reveal key features of myosin regulation and indicate that not all partners can activate Myo6. TOM1 and Dab2 cannot bind the off-state while, GIPC1 binds Myo6, releases its auto-inhibition and triggers proximal dimerization. Myo6 partners thus differentially recruit Myo6. We solved a crystal structure of the proximal dimerization domain, and show that its disruption compromises endocytosis in HeLa cells, emphasizing the importance of Myo6 dimerization. Finally, we show that the L926Q deafness mutation disrupts Myo6 auto-inhibition and indirectly impairs proximal dimerization. Our study thus demonstrates the importance of partners in the control of Myo6 auto-inhibition, localization, and activation.

biochemistry↗

bioRxiv2023

The recycling endosome biogenesis machinery coordinates BACE1 endosomal sorting and amyloid-Beta production

Alzheimers disease (AD) is the most common form of dementia worldwide. One of ADs main pathological hallmarks is the cerebral plaque deposits of {beta}-amyloid (A{beta}). A{beta} is generated through sequential enzymatic cleavage of the amyloid precursor protein (APP). The {beta}-secretase or {beta}-site APP-cleaving enzyme 1 (BACE1) initiates this cleavage and is thus key to regulate A{beta} formation. Both APP and BACE1 transit through the endolysosomal system but the exact nature of the compartment(s) where APP cleavage occurs as well as the molecular mechanisms that govern their endosomal sorting remain poorly known. Here we show that RAB11 not only regulates BACE1 transport from early/sorting endosomes (EEs/SEs) and drives the exocytosis of BACE1-containing recycling carriers. Moreover, recycling endosome-associated KIF13A, as well as its closely related homolog KIF13B, which are known RAB11 effectors involved in the biogenesis of recycling endosome (RE) from EEs/SEs, also participate in BACE1 endosomal sorting. Importantly, depletion of KIF13A or KIF13B leads to an increase in A{beta} generation. Depletion of the BLOC-1 complex, previously described as an essential partner for KIF13A-dependent RE biogenesis, also induces increased amount of A{beta}. Altogether, our findings support a model where the EEs/SEs represent a major organelle for A{beta} formation and identify the recycling endosome biogenesis machinery as a master coordinator of BACE1 endosomal sorting and transport.

cell biology↗

bioRxiv2022

Dual blockade of misfolded alpha-sarcoglycan degradation by bortezomib and givinostat combination

Limb-girdle muscular dystrophy type R3 (LGMD R3) is a rare genetic disorder characterized by a progressive proximal muscle weakness and caused by mutations in the SGCA gene encoding alpha-sarcoglycan (-SG). Here, we report the results of a mechanistic screening ascertaining the molecular mechanisms involved in the degradation of the most prevalent misfolded R77C--SG protein. We performed a combinatorial study to identify drugs potentializing the effect of a low dose of the proteasome inhibitor bortezomib on the R77C--SG degradation inhibition. Analysis of the screening associated to artificial intelligence-based predictive ADMET characterization of the hits led to identification of the HDAC inhibitor givinostat as potential therapeutical candidate. Functional characterization revealed that givinostat effect was related to autophagic pathway inhibition, unveiling new theories concerning degradation pathways of misfolded SG proteins. Beyond the identification of a new therapeutic option for LGMD R3 patients, our results shed light on the potential repurposing of givinostat for the treatment of other genetic diseases sharing similar protein degradation defects such as LGMD R5 and cystic fibrosis.

pharmacology and toxicology↗

bioRxiv2022

Caveolae promote successful abscission by controlling intercellular bridge tension during cytokinesis

During cytokinesis, the intercellular bridge (ICB) connecting the daughter cells experiences pulling forces, which delay abscission by preventing the assembly of the ESCRT scission machinery. Abscission is thus triggered by tension release, but how ICB tension is controlled is unknown. Here, we report that caveolae, which are known to control membrane tension upon mechanical stress in interphase cells, are located at the midbody, at the abscission site and at the ICB/cell interface in dividing cells. Functionally, the loss of caveolae delays ESCRT-III recruitment during cytokinesis and impairs abscission. This is the consequence of a 2-fold increase of ICB tension measured by laser ablation, associated with a local increase in myosin II activity at the ICB/cell interface. We thus propose that caveolae buffer membrane tension and limit contractibility at the ICB to promote ESCRT-III assembly and cytokinetic abscission. Altogether, this work reveals an unexpected connection between caveolae and the ESCRT machinery and the first role of caveolae in cell division. TEASERCaveolae limit the tension in the intercellular bridge during cytokinesis to enable ESCRT-III assembly and successful abscission.

cell biology↗

bioRxiv2021

PI4P and BLOC-1 remodel endosomal membranes into tubules

Intracellular trafficking is mediated by transport carriers that originate by membrane remodeling from donor organelles. Tubular carriers play major roles in the flux of membrane lipids and proteins to acceptor organelles. However, how lipids and proteins impose a tubular geometry on the carriers is incompletely understood. By exploiting imaging approaches at different scales on cells and in vitro membrane systems, we show that phosphatidylinositol-4-phosphate (PI4P) and biogenesis of lysosome-related organelles complex 1 (BLOC-1) govern the formation, stability and functions of recycling endosomal tubules. Endosomal PI4P production by type II PI4-kinases is needed to form nascent curved tubules through binding of BLOC-1 that stabilize and elongate them. Membrane remodeling by the PI4P/ BLOC-1 module functions not only in the recycling of endosomal cargoes, but also in the lifecycles of intracellular pathogens such as Chlamydia bacteria and influenza virus. This study demonstrates how a phospholipid and a protein complex coordinate as a minimal machinery to remodel cellular membranes into functional tubes.

cell biology↗

bioRxiv2021

Two type II phosphatidylinositol 4-kinases function sequentially in tubule-mediated cargo delivery from early endosomes to melanosomes

Melanosomes are pigment cell-specific lysosome-related organelles in which melanin pigments are synthesized and stored. Melanosome maturation requires delivery of melanogenic cargoes via tubular transport carriers that emanate from early endosomes and that require BLOC-1 for their formation. Here we show that phosphatidylinositol-4-phosphate (PtdIns4P) and the type II PtdIns-4-kinases (PI4KII and PI4KII{beta}) support BLOC-1-dependent tubule formation to regulate melanosome biogenesis. Depletion of either PI4KII or PI4KII{beta} with shRNAs in melanocytes reduced melanin content and misrouted BLOC-1-dependent cargoes to late endosomes/lysosomes. Genetic epistasis, cell fractionation, and quantitative live-cell imaging analyses show that PI4KII and PI4KII{beta} function sequentially and non-redundantly downstream of BLOC-1 during tubule elongation towards melanosomes by generating local pools of PtdIns4P. The data show that both type II PtdIns-4-kinases are necessary for efficient BLOC-1-dependent tubule elongation and subsequent melanosome contact and content delivery during melanosome biogenesis. The independent functions of PtdIns-4-kinases in tubule extension are downstream of likely redundant functions in BLOC-1-dependent tubule initiation. SUMMARYContents are delivered to maturing melanosomes from early endosomal intermediates through tubular transport carriers. Zhu et al show that two type II phosphatidylinositol kinases, PI4KII and PI4KII{beta}, sequentially generate phosphatidylinositol-4-phosphate during tubule initiation and elongation for ultimate melanosome content delivery.

cell biology↗

bioRxiv2021

Catabolism of lysosome-related organelles in color-changing spiders supports intracellular turnover of pigments

Pigment organelles of vertebrates belong to the lysosome-related organelle (LRO) family, of which melanin-producing melanosomes are the prototypes. While their anabolism has been extensively unraveled through the study of melanosomes in skin melanocytes, their catabolism remains poorly known. Here, we tap into the unique ability of crab spiders to reversibly change body coloration to examine the catabolism of their pigment organelles. By combining ultrastructural and metal analyses on high-pressure frozen integuments, we first assess whether pigment organelles of crab spiders belong to the LRO family and, second, how their catabolism is intracellularly processed. Using scanning-transmission electron microscopy, electron tomography and nanoscale Synchrotron-based scanning X-ray fluorescence, we show that pigment organelles possess ultrastructural and chemical hallmarks of LROs, including intraluminal vesicles and metal deposits, similar to melanosomes. Monitoring ultrastructural changes during bleaching suggests that the catabolism of pigment organelles involves the degradation and removal of their intraluminal content, possibly through lysosomal mechanisms. In contrast to skin melanosomes, anabolism and catabolism of pigments proceed within the same cell without requiring either cell death or secretion/phagocytosis. Our work hence provides support for the hypothesis that the endolysosomal system is fully functionalized for within-cell turnover of pigments, leading to functional maintenance under adverse conditions and phenotypic plasticity. First formulated for eye melanosomes in the context of human vision, the hypothesis of intracellular turnover of pigments gets unprecedented strong support from pigment organelles of spiders.

cell biology↗

bioRxiv2021

Dynlt3, a novel fundamental regulator of melanosome movement, distribution, maturation and transfer

Skin pigmentation is dependent on cellular processes including melanosome biogenesis, transport, maturation and transfer to keratinocytes. However, how the cells finely control these processes in space and time to ensure proper pigmentation remains unclear. Here, we show that a component of the cytoplasmic dynein complex, Dynlt3, is required for efficient melanosome transport, maturation and transfer. In melanocytes with decreased levels of Dynlt3, pigmented melanosomes undergo a more directional (convective) motion leading to their peripheral location in the cell, and are not fully matured. Stage IV melanosomes are more acidic, but still heavily pigmented, resulting in a less efficient melanosome transfer. Finally, the level of Dynlt3 is dependent on {beta}-catenin activity, revealing a novel function of the Wnt/{beta}-catenin signalling pathway during melanocyte and skin pigmentation, by coupling the transport, position and maturation of melanosomes required for their transfer.

cell biology↗

bioRxiv2020

Dopachrome tautomerase variants in patients with oculocutaneous albinism.

PurposeAlbinism is a clinically and genetically heterogeneous condition. Despite analysis of the nineteen known genes, [~]30% patients remain unsolved. We aimed to identify new genes involved in albinism. MethodsWe sequenced a panel of genes with known or predicted involvement in melanogenesis in 230 unsolved albinism patients. ResultsWe identified variants in the Dopachrome tautomerase (DCT) gene in two patients. One was compound heterozygous for a 14 bp deletion in exon 9 and c.118T>A p.(Cys40Ser). The second was homozygous for c.183C>G p.(Cys61Trp). Both patients had mild hair and skin hypopigmentation, and classical ocular features. CRISPR/Cas9 was used in C57BL/6J mice to create mutations identical to the missense mutations carried by the patients, along with one loss-of-function indel mutation. When bred to homozygosity the three mutations revealed hypopigmentation of the coat, milder for Cys40Ser compared to Cys61Trp or the frameshift mutation. Histological analysis identified significant hypopigmentation of the retinal pigmented epithelium (RPE) indicating that defective RPE melanogenesis could be associated with eye and vision defects. DCT loss of function in zebrafish embryos elicited hypopigmentation both in melanocytes and RPE cells. ConclusionsDCT is the gene for a new type of oculocutaneous albinism that we propose to name OCA8.

genetics↗

bioRxiv2019

Caveolae coupling of melanocytes signaling and mechanics is required for human skin pigmentation

Tissue homeostasis requires regulation of cell-cell communication, which relies on signaling molecules and cell contacts. In skin epidermis, keratinocytes secrete specific factors transduced by melanocytes into signaling cues to promote their pigmentation and dendrite outgrowth, while melanocytes transfer melanin pigments to keratinocytes to convey skin photoprotection. How epidermal cells integrate these functions remains poorly characterized. Here, we found that caveolae polarize in melanocytes and are particularly abundant at melanocyte-keratinocyte interface. Caveolae in melanocytes are sensitive to ultra-violet radiations and miRNAs released by keratinocytes. Preventing caveolae formation in melanocytes results in increased production of intracellular cAMP and melanin pigments, but decreases cell protrusions, cell-cell contacts, pigment transfer and epidermis pigmentation. Altogether, our data establish that, in melanocytes, caveolae serve as key molecular hubs that couple signaling outputs from keratinocytes to mechanical plasticity. This process is crucial to maintain cell-cell contacts and intercellular communication, skin pigmentation and tissue homeostasis.

cell biology↗