Biology subjects

Juillot, D.

Publications and source records attributed to Juillot, D..

6 recordsLinked to original sources

bioRxiv2026

Two temperature-dependent membrane fluidity regimes in Gram-positive bacteria

It is widely accepted that bacterial cells maintain a constant membrane fluidity in response to temperature changes. This process, known as membrane fluidity homeostasis, occurs through remodeling of plasma membrane composition. We tested this using an assay based on total internal reflection-fluorescence correlation spectroscopy (TIR-FCS) that directly quantifies membrane fluidity as the diffusion speed of a membrane marker in Bacillus subtilis and two other Gram-positive bacteria, Streptococcus pneumoniae and Staphylococcus aureus, across a temperature range of 20{degrees}C to 37{degrees}C. Instead of the expected constant membrane fluidity, we identified a two-component regime: membrane fluidity is maintained at low temperatures (<26{degrees}C), but freely increases at higher temperatures. Significance statementTemperature changes affect the physical properties of the bacterial plasma membrane. Typically, with a reduction in temperature comes a loss of membrane fluidity. It is known that bacteria like Bacillus subtilis adapt their membrane composition when this happens, and it was thought to maintain membrane fluidity constant. We found that this is not true: fluidity is maintained only in a low (<26{degrees}C) temperature regime, while at higher temperatures fluidity is not maintained. We found this to be true for several Gram+ bacteria.

biophysics↗

bioRxiv2025

A toxin/antitoxin system targeting the replication sliding-clamp induces competence in Streptococcus pneumoniae.

Streptococcus pneumoniae is a pathogenic bacterium capable of entering a cellular differentiation state, called competence, which enables it to acquire new genetic functions by natural transformation, as well as physiological functions such as tolerance to a number of antibiotics. The transition to this state is regulated by various environmental or intracellular signals that converge on the comCDE operon, which groups together the competence initiation genes. A fraction of activated cells is sufficient to propagate competence to the whole population via the product of the comC gene, the competence stimulating peptide (CSP). Remarkably, depletion of the essential ClpX/ ClpP AAA+ protease has been shown to induce the comCDE operon. Here we demonstrate that the ClpX-dependent induction of competence relies on the Spr1630 toxin (RipA), part of a Rosmer toxin-antitoxin system. We show that this toxin generates replicative stress by acting on the sliding clamp of replication, inducing transcription of the comCDE operon. Bacteria that produce RipA appear to lose their viability but remain metabolically active and able to produce CSP, thereby transferring competence to viable neighbouring cells. Authors summaryThe environment in which bacteria live puts them under a great deal of stress, forcing them to adapt constantly, either temporarily or permanently. Streptococcus pneumoniae, a pathogenic bacterium implicated in various pathologies such as otitis, meningitis and pneumonia, is also subject to stress, whether from its host, antibiotic treatments or the microbiota in which it lives. In response to this, S. pneumoniae is able to switch to a differentiated state called competence. This allows it to acquire new genetic characteristics through natural transformation, but also to better tolerate stresses such as antibiotics pressure. The underlying signals and signaling pathways of this phenotypic switch remain poorly characterized. In this study, we identified a novel toxin-antitoxin system that, when activated, causes a subset of the population to self-sacrifice by disrupting its own DNA replication. This self-induced arrest serves as a signal that promotes the transition to competence in neighboring cells, thereby improving the capacity for adaptation at the populational level.

microbiology↗

bioRxiv2024

Real-Time Nanoscale Investigation of Spore Coat Assembly in Bacillus subtilis

Spores of Bacillaceae, ubiquitous in soil, can withstand extreme conditions virtually indefinitely. The spore coat, one of the most sophisticated multiprotein complexes built by bacteria, protects the spore from environmental stresses and predators. In Bacillus subtilis, assembly of the multilayered coat initiates at the forespore pole and is perceived as a stable process that progresses continuously until covering the entire forespore surface. In contrast, in Bacillus cereus, coat formation initiates in the midspore region and extends outward to the spore poles. Using Structured Illumination Microscopy, we monitored B. subtilis coat development in real-time at the single-sporangium level with lateral resolution of 70 nm. We found that late-synthesized proteins from the innermost coat layers first assemble in the midspore region and are subsequently displaced towards the poles. This process is coupled with a unique redistribution of pre-assembled coat material across the forespore surface and influenced by outer coat development, highlighting a dynamic interplay in coat layers co-construction.

microbiology↗

bioRxiv2024

DivIVA controls the dynamics of septum splitting and cell elongation in Streptococcus pneumoniae

Bacterial shape and division rely on the dynamics of cell wall assembly, which involves regulated synthesis and cleavage of the peptidoglycan. In ovococci, these processes are coordinated in an annular mid-cell region with nanometric dimensions. More precisely, the cross-wall that is synthesized by the divisome is split to generate lateral wall, whose expansion is insured by insertion of so-called peripheral peptidoglycan by the elongasome. Septum cleavage and peripheral peptidoglycan synthesis are thus crucial remodeling events for ovococcal cell division and elongation. The structural DivIVA protein has long been known as a major regulator of these processes but its mode of action remains unknown. Here, we integrate click chemistry-based peptidoglycan labeling, direct stochastic optical reconstruction microscopy and in silico modeling, as well as epifluorescence and stimulated emission depletion microscopy to investigate the role of DivIVA in Streptococcus pneumoniae cell morphogenesis. Our work reveals two distinct phases of peptidoglycan remodeling along the cell cycle, that are differentially controlled by DivIVA. In particular, we show that DivIVA ensures homogeneous septum cleavage and peripheral peptidoglycan synthesis around the division site, and their maintenance throughout the cell cycle. Our data additionally suggest that DivIVA impacts the contribution of the elongasome and class A PBPs to cell elongation. We also report the position of DivIVA on either side of the septum, consistent with its known affinity for negatively curved membranes. Finally, we take the opportunity provided by these new observations to propose hypotheses for the mechanism of action of this key morphogenetic protein. IO_SCPLOWMPORTANCEC_SCPLOWThis study sheds light on fundamental processes governing bacterial cell growth and division, using integrated click chemistry, advanced microscopy and computational modeling approaches. More precisely, it addresses mechanisms involved in the regulation of cell wall synthesis and remodeling in Streptococcus pneumoniae. This bacterium belongs to the morphological group of ovococci, which includes many human pathogens, such as streptococci and enterococci. In this study, we have dissected the function of DivIVA, which is a structural protein involved in cell division, cell morphogenesis and chromosome partitioning in Gram-positive bacteria. This work unveils the role of DivIVA in the orchestration of cell division and elongation along the pneumococcal cell cycle. It not only helps understanding how ovoid bacteria proliferate, but also offers an opportunity to consider how DivIVA might serve as a scaffold and sensor for particular membrane regions, and thus be involved in various processes associated with the cell cycle.

microbiology↗

bioRxiv2024

Transient inhibition of cell division in competent pneumococcal cells results from deceleration of the septal peptidoglycan complex

Bacterial cells are known to produce inhibitors of cell division in response to stress responses and developmental programs. Knowledge of the underlying molecular mechanisms remains however largely limited. In this study, we investigated the mechanism of transient cell division inhibition observed during the development of competence for transformation in the human pathogen Streptococcus pneumoniae. In this species, ComM, a membrane protein specifically produced during competence, transiently inhibits cell division to preserve genomic integrity during transformation. We show that ComM reduces specifically the dynamics of the septal peptidoglycan synthetic complex FtsW:PBP2x. We also present evidence that ComM interacts with the peptidoglycan precursor synthetic enzyme MurA, and show that overproduction of MurA suppresses FtsW:PBP2x deceleration along the cell division delay in competent cells. Collectively, our data support a model in which ComM interferes with MurA activity to reduce septal peptidoglycan synthesis during competence in S. pneumoniae.

microbiology↗

bioRxiv2021

A high content microscopy screening identifies genes involved in cell width control in Bacillus subtilis

How cells control their shape and size is a fundamental question of biology. In most bacteria, cell shape is imposed by the peptidoglycan (PG) polymeric meshwork that surrounds the cell. Thus, bacterial cell morphogenesis results from the coordinated action of the proteins assembling and degrading the PG shell. Remarkably, during steady-state growth, most bacteria maintain a defined shape along generations, suggesting that error-proof mechanisms tightly control the process. In the rod-shaped model for Gram-positive bacteria Bacillus subtilis, the average cell length varies as a function of the growth rate but the cell diameter remains constant throughout the cell cycle and across growth conditions. Here, in an attempt to shed light on the cellular circuits controlling bacterial cell width, we developed a screen to identify genetic determinants of cell width in B. subtilis. Using high-content screening (HCS) fluorescence microscopy and semi-automated measurement of single-cell dimensions, we screened a library of ~ 4000 single knockout mutants. We identified 13 mutations significantly altering cell diameter, in genes that belong to several functional groups. In particular, our results indicate that metabolism plays a major role in cell width control in B. subtilis.

microbiology↗