Etude multi échelle de trois effecteurs conservés des épithéliums ciliés

Nos organes sont recouverts d'une couche de cellules épithéliales créant des frontières biologiques. Dans cette étude, je me concentre sur l'épithélium cilié, composé de cellules multiciliées (CMC). Les CMCs possèdent à leur surface de nombreux cils motiles. Chaque cil se forme à partir d'un centriole modifié, le corps basal (CB). Les cils battent de manière synchronisée pour générer des flux directionnels de fluides, essentiels pour des mécanismes physiologiques comme : l'élimination de pathogènes des voies respiratoires et le transport des gamètes dans les voies génitales. Des mutations de gènes nécessaires à la formation et fonction des cils sont responsables de syndromes familiaux causant des infections des voies respiratoires et un risque élevé de stérilité. Ce travail aborde le rôle multi-échelle de trois protéines centriolaires conservées (Lrrcc1, Ccdc61 et Odf2) dans l'épiderme cilié du Xénope. Mes travaux montrent que Lrrcc1, Ccdc61 et Odf2 codent des composants centriolaires essentiels pour l’ancrage des CBs à la membrane apicale, leur espacement, leur polarisation coordonnée et l'organisation du cytosquelette apical. De plus, ces défauts entraînent un battement ciliaire lent et un flux inefficace, menant à une survie embryonnaire réduite en présence de pathogènes. En outre, j'aborde le rôle de Odf2 dans l’établissement du patron d’espacement des CMCs. Dans la peau des embryons de Xénope, les CMCs adoptent un espacement régulier, probablement important pour la production d'un flux homogène. Nous montrons que Odf2, vraisemblablement en tant que composant centrosomal important pour l’organisation des microtubules, est nécessaire à l’espacement régulier des CMCs.Our organs are covered with a layer of epithelial cells creating biological borders. In this study, I focus on the ciliated epithelium, composed of multiciliated cells (MCC). MCCs have many motile cilia on their surface. Each cilium is formed from a modified centriole, the basal body (BB). Cilia beat synchronously to generate directional fluid flows at their surface, which are essential for physiological mechanisms such as: the removal of pathogens from the respiratory tract and the transport of gametes along the genital tract. Mutations in genes necessary for the formation and function of cilia are responsible for familial syndromes causing respiratory tract infections and a high risk of infertility. This work addresses the multi-scale role of three conserved centriolar proteins (Lrrcc1, Ccdc61 and Odf2) in the ciliated epidermis of Xenopus. My work shows that Lrrcc1, Ccdc61 and Odf2 encode centriolar components essential for the anchoring of BBs to the apical membrane, their spacing, their coordinated polarization and the organization of the apical cytoskeleton. In addition, these defects cause a slower ciliary beating and an inefficient flow, leading to reduced embryonic survival in the presence of pathogens. In addition, I discuss the role of Odf2 in the establishment of MCCs spacing pattern. In the skin of Xenopus embryos, MCCs adopt a regular spacing pattern, probably important for the production of homogeneous flow. We show that Odf2, probably as an important centrosomal component for the organization of microtubules, is necessary for the regular spacing of MCCs

Copper and Cadmium administration induce toxicity and oxidative stress in the marine flatworm Macrostomum lignano

International audienceThe contamination of coastal regions with different toxicants, including heavy metal ions such as copper and cadmium jeopardizes health and survival of organisms exposed to this habitat. In order to study the effects of high copper and cadmium concentrations in these marine environments, we used the flatworm Macrostomum lignano as a model. This platyhelminth lives in shallow coastal waters and is exposed to high concentrations of all toxicants that accumulate in these shallow sea floors. We show that both, cadmium and copper induce toxicity at high concentrations, with copper being more toxic than cadmium. At concentrations below those inducing acute toxicity, long-term survival rates were reduced for both metal ions. The effects of sublethal doses comprise reduced physical activities and an increase in ROS levels within the worms

Salinity stress from the perspective of the energy-redox axis: Lessons from a marine intertidal flatworm

In the context of global change, there is an urgent need for researchers in conservation physiology to understand the physiological mechanisms leading to the acquisition of stress acclimation phenotypes. Intertidal organisms continuously cope with drastic changes in their environmental conditions, making them outstanding models for the study of physiological acclimation. As the implementation of such processes usually comes at a high bioenergetic cost, a mitochondrial/oxidative stress approach emerges as the most relevant approach when seeking to analyze whole-animal responses. Here we use the intertidal flatworm Macrostomum lignano to analyze the bioenergetics of salinity acclimation and its consequences in terms of reactive oxygen/nitrogen species formation and physiological response to counteract redox imbalance. Measures of water fluxes and body volume suggest that M. lignano is a hyper-/iso-regulator. Higher salinities were revealed to be the most energetically expensive conditions, with an increase in mitochondrial density accompanied by increased respiration rates. Such modifications came at the price of enhanced superoxide anion production, likely associated with a high caspase 3 upregulation. These animals nevertheless managed to live at high levels of environmental salinity through the upregulation of several mitochondrial antioxidant enzymes such as superoxide dismutase. Contrarily, animals at low salinities decreased their respiration rates, reduced their activity and increased nitric oxide formation, suggesting a certain degree of metabolic arrest. A contradictory increase in dichlorofluorescein fluorescence and an upregulation of gluthathione-S-transferase pi 1 (GSTP1) expression were observed in these individuals. If animals at low salinity are indeed facing metabolic depression, the return to seawater may result in an oxidative burst. We hypothesize that this increase in GSTP1 could be a “preparation for oxidative stress”, i.e. a mechanism to counteract the production of free radicals upon returning to seawater. The results of the present study shed new light on how tolerant organisms carry out subcellular adaptations to withstand environmental change

Lrrcc1 and Ccdc61 are conserved effectors of multiciliated cell function

International audienceCiliated epithelia perform essential functions across animal evolution, ranging from locomotion of marine organisms to mucociliary clearance of airways in mammals. These epithelia are composed of multiciliated cells (MCCs) harbouring myriads of motile cilia, which rest on modified centrioles called basal bodies (BBs), and beat coordinately to generate directed fluid flows. Thus, BB biogenesis and organization is central to MCC function. In basal eukaryotes, the coiled-coil domain proteins Lrrcc1 and Ccdc61 were shown to be required for proper BB construction and function. Here, we used the Xenopus embryonic ciliated epidermis to characterize Lrrcc1 and Ccdc61 in vertebrate MCCs. We found that they both encode BB components, localized proximally at the junction with striated rootlets. Knocking down either gene caused defects in BB docking, spacing, and polarization. Moreover, their depletion impaired the apical cytoskeleton, and altered ciliary beating. Consequently, cilia-powered fluid flow was greatly reduced in morphant tadpoles, which displayed enhanced mortality when exposed to pathogenic bacteria. This work illustrates how integration across organizational scales make elementary BB components essential for the emergence of the physiological function of ciliated epithelia

Active mucus–cilia hydrodynamic coupling drives self-organization of human bronchial epithelium

International audienceThe respiratory tract is protected by mucus, a complex fluid transported along the epithelial surface by the coordinated beating of millions of microscopic cilia, hence the name of mucociliary clearance. Its impairment is associated with all severe chronic respiratory diseases. Yet, the relationship between ciliary density and the spatial scale of mucus transport, as well as the mechanisms that drive ciliary-beat orientations are much debated. Here, we show on polarized human bronchial epithelia that mucus swirls and circular orientational order of the underlying ciliary beats emerge and grow during ciliogenesis, until a macroscopic mucus transport is achieved for physiological ciliary densities. By establishing that the macroscopic ciliary-beat order is lost and recovered by removing and adding mucus, respectively, we demonstrate that cilia–mucus hydrodynamic interactions govern the collective dynamics of ciliary-beat directions. We propose a two-dimensional model that predicts a phase diagram of mucus transport in accordance with the experiments. This paves the way to a predictive in silico modelling of bronchial mucus transport in health and disease