Biology subjects

Moriscot, C.

Publications and source records attributed to Moriscot, C..

9 recordsLinked to original sources

bioRxiv2026

Thylakoid plasticity primes photoprotection in diatoms

For many millions of years, diatoms have been major primary producers in marine ecosystems, supporting food webs and contributing to carbon sequestration through the export of photosynthetically fixed carbon to the deep ocean, mitigating atmospheric carbon dioxide levels and global climate. A key factor in their ecological success is their ability to rapidly adapt to fluctuating light, balancing efficient light harvesting under dim conditions with energy dissipation under excess light. However, the mechanisms behind this regulation have not been fully elucidated. This study shows that several processes contribute to dissipating excess excitation energy in diatoms exposed to light stress: (i) the detachment of the oxygen-evolving complex proteins from the PSII core, leading to energy dissipation via charge recombination, (ii) Non-Photochemical Quenching (NPQ) in the antenna, i.e., thermal dissipation of excited states and (iii) excitation energy transfer from PSII to PSI via energy spillover. The latter process is observed in parallel with high light-induced topological changes of the thylakoids that may favor transient interactions between the photosystems. Along with the accumulation of light-stress-protecting carotenoids on longer time-scales, this ensemble of photoprotective mechanisms provides a dynamic and flexible response to the light challenges expected in coastal marine habitats.

biochemistry↗

bioRxiv2024

Role of the clathrin adaptor AP-1 in cell wall integrity and virulence factor secretion in the plant pathogen Botrytis cinerea

In filamentous fungi, the secretory pathway plays a key role in polarized growth through the dlivery of vesicles carrying enzymes and components necessary to cell wall synthesis and fungal nutrition. In pathogenic species, it moreover supports the delivery of virulence factors. In metazoa and yeast, the formation of several secretory vesicles occurs at the Golgi apparatus and endosomes through a complex and partially characterized machinery involving clathrin and adaptor proteins such as AP-1. Data on the molecular mechanisms controlling this process are still scarce in filamentous fungi. Using a mutant under-expressing the {beta}-subunit of the heterotetrameric AP-1 clathrin adaptor complex, we investigated the importance of intracellular vesicle trafficking in the phytopathogenic fungus Botrytis cinerea. The mutant exhibits pleiotropic developmental defects, an abnormality in cell wall integrity associated with a mislocalization of the plasma-membrane chitine synthase BcCHSIIIa. Moreover, the AP-1 mutant is affected in the secretion of hydrolytic enzymes and displays a severe defect in pathogenicity. This study confirms the importance of the AP1/clathrin machinery in the secretory pathway of B. cinerea.

microbiology↗

bioRxiv2024

New assessment of teichoic acids in the cell envelope of Streptococcus pneumoniae

Teichoic acids (TA) are linear phospho-saccharidic polymers and important constituents of the cell envelope of Gram-positive bacteria, either bound to the peptidoglycan as wall teichoic acids (WTA) or to the membrane as lipoteichoic acids (LTA). The chemical composition of TA varies greatly but the presence of both WTA and LTA is highly conserved, hinting at an underlying fundamental function that is distinct from their numerous specific roles in diverse organisms. We report here the observation of a periplasmic space in the Gram-positive Streptococcus pneumoniae by cryo-electron microscopy of vitreous sections. The thickness and appearance of this region change upon deletion of genes involved in the attachment of teichoic acids, supporting the role of TA in the maintenance of a periplasmic space in Gram-positive bacteria as a possible universal function. Consequences of these mutations were further examined by super-resolved microscopy (dSTORM), following metabolic and fluorophore coupling by click-chemistry in pulse and pulse-chase experiments. This novel labeling method also enabled in-gel analysis of cell fractions, revealing that LTA-containing membranes sediment at low centrifugal forces. Owing to this easy separation approach, we were able to titrate the actual amount of TA per cell and to determine the ratio of WTA to LTA. In addition, we followed the change of TA length during growth phases, and discovered that a mutant devoid of LTA accumulates the membrane-bound polymerized TA precursor. SignificanceThe existence of a periplasmic space in Gram-positive bacteria has long been debated. The finding that compromising the attachment of teichoic acids changes the appearance and thickness of the periplasm in the pneumococcus indicates a role of these polymers in the maintenance of this space between the membrane and the cell wall. Metabolic labeling and electrophoresis showed that LTA-containing membranes are easily sedimented. This finding indicates that the LTA/WTA ratios reported in previous studies were likely underestimated, since most LTA were probably unknowingly discarded in these studies. Our method of TA analysis opens a new era in the investigation of these important and poorly known bacterial polymers and their role in the periplasmic space of Gram-positive organisms.

microbiology↗

bioRxiv2023

An inducible ESCRT-III inhibition tool to control HIV-1 budding

HIV-1 budding as well as many other cellular processes require the Endosomal Sorting Complex Required for Transport (ESCRT) machinery. Understanding the architecture of the native ESCRT-III complex at HIV-1 budding sites is limited due to spatial resolution and transient ESCRT-III recruitment. Here, we developed a drug-inducible transient HIV-1 budding inhibitory tool to enhance the ESCRT-III lifetime at budding sites. We generated auto-cleavable CHMP2A, CHMP3, and CHMP4B fusion proteins with the hepatitis C virus NS3 protease. We characterized the CHMP-NS3 fusion proteins in the absence and presence of protease inhibitor Glecaprevir with regard to expression, stability, localization and HIV-1 Gag VLP budding. Immunoblotting experiments revealed rapid and stable accumulation of CHMP-NS3 fusion proteins with variable modification of Gag VLP budding upon drug administration. Notably, CHMP2A-NS3 and CHMP4B-NS3 fusion proteins substantially decrease VLP release while CHMP3-NS3 exerted a minor effect and synergized with CHMP2A-NS3. Localization studies demonstrated the re-localization of CHMP-NS3 fusion proteins to the plasma membrane, endosomes, and Gag VLP budding sites. Through the combined use of transmission electron microscopy and video-microscopy, we unveiled drug-dependent accumulation of CHMP2A-NS3 and CHMP4B-NS3, causing a delay in HIV-1 Gag-VLP release. Our findings provide novel insight into the functional consequences of inhibiting ESCRT-III during HIV-1 budding and establish new tools to decipher the role of ESCRT-III at HIV-1 budding sites and other ESCRT-catalyzed cellular processes.

microbiology↗

bioRxiv2023

Ultrastructural details of resistance factors of the bacterial spore revealed by in situ cryo-electron tomography

The bacterial spore owes its incredible resistance capacities to various molecular structures that protect the cell content from external aggressions. Among the determinants of resistance are the quaternary structure of the chromosome and an extracellular shell made of proteinaceous layers (the coat), the assembly of which remains poorly understood. Here, in situ cryo-electron tomography (cryo-ET) on bacteria lamellae generated by cryo-focused ion beam micromachining (cryo-FIBM) provides insights into the ultrastructural organization of Bacillus subtilis sporangia, including that of the DNA and nascent coat layers. Analysis of the reconstructed tomograms reveal that rather early during sporulation, the chromosome in the developing spore (the forespore) adopts a toroidal structure harboring 5.5-nm thick fibers. At the same stage, coat proteins at the surface of the forespore form a complex stack of amorphous or structured layers with distinct electron density, dimensions and organization. We investigated the nature of the nascent coat layers in various mutant strains using cryo-FIBM/ET and transmission electron microscopy on resin sections of freeze-substituted bacteria. Combining these two cellular electron microscopy approaches, we distinguish seven nascent coat regions with different molecular properties, and propose a model for the contribution of the morphogenetic proteins SpoIVA, SpoVID, SafA and/or CotE. Significance statementBacterial spores are dormant cells that can resist to multiple stresses, including antibiotics, detergents, irradiation and high temperatures. Such resilience is an asset when spores are used for the benefit of humans, as in the case of probiotics, or a major problem for public health, food safety or biowarfare when it comes to spores of pathogenic bacteria. In this study, we combined state-of-the-art cryo-electron tomography and conventional cellular electron microscopy to provide insights into intermediate stages of spore development. Our data reveal the intracellular reorganization of the chromosome into a toroidal fibrillar structure and the complex assembly of the multi-protein, multilayered extracellular coat, shedding light on the mechanisms by which the spore acquires its incredible resistance capacities.

microbiology↗

bioRxiv2022

The AAA+ ATPase RavA and its binding partner ViaA modulate E. coli aminoglycoside sensitivity through interaction with the inner membrane

Enteric bacteria have to adapt to environmental stresses in the human gastrointestinal tract such as acid and nutrient stress, oxygen limitation and exposure to antibiotics. Membrane lipid composition has recently emerged as a key factor for stress adaptation. The E. coli ravA-viaA operon is essential for aminoglycoside bactericidal activity under anaerobiosis but its mechanism of action is unclear. Here we characterise the VWA domain-protein ViaA and its interaction with the AAA+ ATPase RavA, and find that both proteins localise at the inner cell membrane. We demonstrate that RavA and ViaA target specific phospholipids and subsequently identify their lipid-binding sites. We further show that mutations abolishing interaction with lipids restore induced changes in cell membrane morphology and lipid composition. Finally we reveal that these mutations render E. coli gentamicin-resistant under fumarate respiration conditions. Our work thus uncovers a ravA-viaA-based pathway which is mobilised in response to antibiotics under anaerobiosis and has a major impact on cell membrane regulation.

microbiology↗

bioRxiv2021

Molecular features underlying Pseudomonas aeruginosa persistence in human plasma

Pseudomonas aeruginosa, an opportunistic Gram-negative pathogen, is a leading cause of bacteremia with a high mortality rate. We recently reported that P. aeruginosa forms a persister-like sub-population of evaders in human plasma and blood. However, the molecular mechanisms underlying the formation of evaders remained unknown. Here, using a gain-of-function genetic screen, we examined the molecular determinants of P. aeruginosa persistence in plasma. We found that, among other factors, ATP and biotin availability greatly influence bacterial survival in plasma; mutants in pur and bio genes display higher tolerance and persistence, respectively. Electron microscopy combined with energy-dispersive X-ray spectroscopy (EDX) revealed the formation of polyphosphate granules upon incubation in plasma in several clinical strains, implying the bacterial response to a low-energy stress signal. Indeed, mutants with transposon insertions in ppk genes were eliminated in the plasma. Analysis of several steps of the complement cascade and exposure to an outer-membrane-impermeable drug, nisin, suggested that the mutants impede membrane attack complex (MAC) activity per se. Through this study, we shed light on P. aeruginosa response to the plasma conditions and discovered the multifactorial origin of bacterial resilience to MAC that provides a comprehensive picture of the complex interplay between P. aeruginosa and the human complement system. Author summaryPersistence of bacterial pathogens is a main cause of treatment failure and establishment of chronic bacterial infection. Despite innate immune responses, some bacteria may persist in human blood and plasma. Here we used a genome-wide screen to investigate the molecular determinants influencing Pseudomonas aeruginosa persistence in human plasma facing the complement system. Alongside a multifactorial strategy, we found intracellular levels of ATP and biotin to significantly influence bacterial capacity to deal with membrane attack complex (MAC)-dependent killing. These results underline the need to understand the complex interplay between bacterial pathogens and the human immune system when seeking to develop efficient antibacterial strategies.

microbiology↗

bioRxiv2020

In-cell quantitative structural imaging of phytoplankton using 3D electron microscopy

Phytoplankton is a minor fraction of the global biomass playing a major role in primary production and climate. Despite improved understanding of phytoplankton diversity and genomics, we lack nanoscale subcellular imaging approaches to understand their physiology and cell biology. Here, we present a complete Focused Ion Beam - Scanning Electron Microscopy (FIB-SEM) workflow (from sample preparation to image processing) to generate nanometric 3D phytoplankton models. Tomograms of entire cells, representatives of six ecologically-successful phytoplankton unicellular eukaryotes, were used for quantitative morphometric analysis. Besides lineage-specific cellular architectures, we observed common features related to cellular energy management: i) conserved cell-volume fractions occupied by the different organelles; ii) consistent plastid-mitochondria interactions, iii) constant volumetric ratios in these energy-producing organelles. We revealed detailed subcellular features related to chromatin organization and to biomineralization. Overall, this approach opens new perspectives to study phytoplankton acclimation responses to abiotic and biotic factors at a relevant biological scale.

plant biology↗

bioRxiv2019

Visualization of silver nanoparticle intracellular trafficking revealed nuclear translocation of silver ions leading to nuclear receptor impairment

The impact on human health of the increasing use of silver nanoparticles (AgNPs) in medical devices remains understudied, even though AgNP-containing dressings are known to release silver in the bloodstream leading to accumulation and slow clearance in the liver. Cellular studies have shown the intracellular dissolution of AgNPs within endo-lysosomes followed by Ag(I) binding to biomolecular thiolate-containing molecules. However, the precise subcellular distribution of Ag(I) and the nature of the disrupted physiological pathways remained unknown. Novel imaging approaches enabled us to visualize the trafficking of AgNP-containing lysosomes towards a perinuclear location and a direct nuclear transfer of Ag(I) species with accumulation in the nucleoli. These Ag(I) species impaired nuclear receptor activity, disrupting critical mechanisms of liver physiology in very low dose exposure scenarios, thus justifying further research into defining a framework for the safe use of AgNPs.

pharmacology and toxicology↗