Biology subjects

Pellequer, J.-L.

Publications and source records attributed to Pellequer, J.-L..

6 recordsLinked to original sources

bioRxiv2026

Elongasome Dysfunction Triggers Dependence on MepM-Mediated Peptidoglycan Recycling

The bacterial peptidoglycan (PG) layer is responsible for maintaining cell shape and protecting the cell against both external stress and internal turgor pressure. In rod-shaped bacteria, PG synthesis is orchestrated by two major multi-protein complexes: the elongasome and the divisome, which drive lateral and septal cell wall synthesis, respectively. The membrane protein MreD plays a central role in regulating the elongasome activity. Here, we leveraged a viable Pseudomonas aeruginosa transposon mutant, with reduced mreD expression, to dissect cellular adaptations to impaired elongasome function. A genome-wide synthetic lethality screen revealed that cell division and PG recycling pathways become essential in the context of an impaired elongasome machinery. Moreover, Tn-seq analysis identified the endopeptidase MepM as essential in the mreD downregulated background, whereas its overexpression promoted cell elongation in the parental strain, supporting a conserved role in lateral wall biogenesis. We demonstrate that MepM sustains viability during elongasome impairment by maintaining an active MurU-dependent PG recycling pathway. Together, our findings uncover a functional coupling between PG hydrolysis, recycling, and synthesis. We also identify MepM and PG recycling as critical determinants of bacterial survival when elongasome activity is compromised, positioning these pathways as attractive targets for the development of next-generation or combinatory antibacterial therapies.

microbiology↗

bioRxiv2025

Lipopolysaccharides nanodiscs, a biomimetic platform to study bacterial surface.

Lipopolysaccharides (LPS) are essential components of the outer membranes of Gram-negative bacteria, playing a crucial role in antimicrobial resistance, virulence, and the hosts immune response. Self-assembled particles displaying LPS are essential for biophysical studies addressing the behavior of bacterial surfaces under specific biomimetic conditions. Styrene-maleic acid (SMA) copolymers were employed to form LPS nanodiscs, either from extracted LPS or directly from purified outer membranes. These nanodiscs, derived from pathogenic O157:H7 or laboratory E. coli strains, are well-defined in size and yield high-resolution nuclear magnetic resonance (NMR) spectra. They have been successfully used to investigate molecular recognition by a human C-type Lectin Receptor (CLR) of the immune system and interaction with polymyxin antibiotics using various biophysical methods. This study high-lights the significance of LPS nanodiscs as bacterial surface mimetics and opens promising avenues for further research into LPS structure and interactions. The ability to generate well-defined LPS nanodiscs offers a powerful tool for studying the molecular mechanisms underlying bacterial pathogenesis and immune response.

biophysics↗

bioRxiv2024

AFMfit : Deciphering conformational dynamics in AFM data using fast nonlinear NMA and FFT-based search

Atomic Force Microscopy (AFM) offers a unique opportunity to study the conformational dynamics of proteins in near-physiological conditions at the single-molecule level. However, interpreting the two-dimensional molecular surfaces of multiple molecules measured in AFM experiments as three-dimensional conformational dynamics of a single molecule poses a significant challenge. Here, we present AFMfit, a flexible fitting procedure that deforms an input atomic model to match multiple AFM observations. The fitted models form a conformational ensemble that unambiguously describes the AFM experiment. Our method uses a new fast fitting algorithm based on the nonlinear Normal Mode Analysis (NMA) method NOLB to associate each molecule with its conformational state. AFMfit processes conformations of hundreds of AFM images of a single molecule in a few minutes on a single workstation, enabling analysis of larger datasets, including high-speed (HS)-AFM. We demonstrate the applications of our methods to synthetic and experimental AFM/HS-AFM data that include activated factor V and a membrane-embedded transient receptor potential channel TRPV3. AFMfit is an open-source Python package available at https://gricad-gitlab.univ-grenoble-alpes.fr/GruLab/AFMfit/.

bioinformatics↗

bioRxiv2023

Correlation between plant cell wall stiffening and root extension arrest phenotype in the combined abiotic stress of Fe and Al

The plasticity and growth of plant cell walls (CWs) remain poorly understood at the molecular level. In this work, we used atomic force microscopy (AFM) to observe elastic responses of the root transition zone of 4-day-old Arabidopsis thaliana wild type and almt1 mutant seedlings grown under Fe or Al stresses. The elastic parameters were deduced from force-distance measurements by AFM using the trimechanic-3PCS framework. In all metal stresses tested, the presence of single metal species Fe2+ or Al3+ at 10 {micro}M exerts no noticeable effect on the root growth compared with the control conditions. On the contrary, a mix of both the metal ions produced a strong root extension arrest concomitant with significant increase of CW stiffness. This was not found for the almt1 mutant which substantially abolishes the ability to exude malate. By raising the concentration of either Fe2+ or Al3+ to 20 {micro}M, no root extension arrest was observed; nevertheless, a rise of root stiffness occurred. Our results indicate that the combination of Fe2+ and Al3+ with exuded malate is crucial for both CW stiffening and root extension arrest. However, stiffness increase induced by single Fe or Al metal is not sufficient for arresting root growth. Summary statementWe record the change in stiffness of the external primary cell wall of living Arabidopsis thaliana seedlings in presence of metallic stress using atomic force microscopy. Results reveals for the first time the uncoupling between mechanical response (CW stiffening) and root extension arrest.

plant biology↗

bioRxiv2021

Structural and functional characterization of DdrC, a novel DNA damage-induced nucleoid associated protein involved in DNA compaction

Deinococcus radiodurans is a spherical bacterium well-known for its outstanding resistance to DNA-damaging agents. Exposure to such agents leads to drastic changes in the transcriptome of D. radiodurans. In particular, four Deinococcus-specific genes, known as DNA Damage Response genes, are strongly up-regulated and have been shown to contribute to the resistance phenotype of D. radiodurans. One of these, DdrC, is expressed shortly after exposure to {gamma}-radiation and is rapidly recruited to the nucleoid. In vitro, DdrC has been shown to compact circular DNA, circularize linear DNA, anneal complementary DNA strands and protect DNA from nucleases. To shed light on the possible functions of DdrC in D. radiodurans, we determined the crystal structure of the domain-swapped DdrC dimer at a resolution of 2.2 [A] and further characterized its DNA binding and compaction properties. Notably, we show that DdrC bears two asymmetric DNA binding sites located on either side of the dimer and can modulate the topology and level of compaction of circular DNA. These findings suggest that DdrC may be a DNA damage-induced nucleoid-associated protein that enhances nucleoid compaction to limit the dispersion of the fragmented genome and facilitate DNA repair after exposure to severe DNA damaging conditions.

microbiology↗

bioRxiv2019

The plasma Factor XIII heterotetrameric complex structure: unexpected unequal pairing within a symmetric complex

Factor XIII (FXIII) is a predominant determinant of clot stability, strength, and composition. Plasma FXIII circulates as a pro-transglutaminase with 2 catalytic A subunits and 2 carrier-protective B subunits in a heterotetramer (FXIII-A2B2). FXIII-A2 and -B2 subunits are synthesized separately and then assembled in plasma. Following proteolytic activation by thrombin and calcium-mediated dissociation of the B-subunits, activated FXIII (FXIIIa) covalently cross-links fibrin, promoting clot stability. The zymogen and active states of the FXIII-A subunits have been structurally characterized; however, the structure of FXIII-B subunits and the FXIII-A2B2 complex have remained elusive. Using integrative hybrid approaches including atomic force microscopy, cross-linking mass spectrometry, and computational approaches, we have constructed the first all-atom model of the FXIII-A2B2 complex. We also used molecular dynamic simulations in combination with isothermal titration calorimetry to characterize FXIII-A2B2 assembly, activation, and dissociation. Our data reveal unequal pairing of individual subunit monomers in an otherwise symmetric complex, and suggest this unusual structure is critical for both assembly and activation of this complex. Our findings enhance understanding of mechanisms associating FXIII-A2B2 mutations with disease and have important implications for the rational design of molecules to alter FXIII assembly and/or activity to reduce bleeding and thrombotic complications.

molecular biology↗