Biology subjects

Valotteau, C.

Publications and source records attributed to Valotteau, C..

7 recordsLinked to original sources

bioRxiv2026

Scaling the Dynamics of Coiled Coils

Coiled coils are structural motifs in proteins that play diverse functions. In MRE11-RAD50 (MR) complexes, ATP-driven changes in coiled coils are essential for DNA break sensing. However, coiled coil dynamics and its modulation by protein conformational changes remain unclear, partly due to the lack of quantitative tools. Here, we used high-speed atomic force microscopy (HS-AFM) for real-time visualization of the coiled coil conformational dynamics of individual MR complexes from bacteria and human homologs, and a biomedically relevant variant. The mean square deviation of the end-to-end distance of the coiled coils revealed a power-law scaling with time, conserved across conformational states, homologs, and variants, suggesting a universal dynamic scaling. Coiled coils behave as semi-flexible filaments with strong internal friction, leading to relaxation times that were seconds-long and varied among conformational states and variants. Molecular dynamics simulations indicated that strong friction arose from long-lifetime contacts between coils. Our results suggest that MR complexes modulate the coiled coil dynamics to mediate long-range allosteric and allodynamic communication during DNA repair.

biophysics↗

bioRxiv2026

A non-invasive approach for understanding localized force generation in 3D tissues

The development, maintenance and repair of epithelial tissues critically rely on adhesion complexes that ensure structural integrity while enabling dynamic remodeling. Such tissue remodeling underpins both physiological morphogenesis and pathological transformation. Central to these processes are mechanical forces, which tightly couple cytoskeletal organization to adhesion dynamics. Despite extensive investigations in two-dimensional (2D) systems, how these interactions are orchestrated within polarized three-dimensional (3D) epithelia remains largely unresolved. Here, we introduce a new, non-invasive strategy to probe localized force generation within 3D epithelial tissues. We engineered elastic polyacrylamide (PAAm) microbeads with cell-mimetic size and mechanical properties, enabling their seamless integration. In contrast to conventional bead injection approaches, these PAAm microbeads were spontaneously engulfed by the tissue, thereby establishing an intrinsic interface through which bead deformation can be directly correlated with local cytoskeletal architecture and adhesion organization, as visualized through high-resolution imaging combined with quantitative 3D computational reconstruction. Using this approach, we demonstrated that localized mechanical perturbations trigger pronounced cytoskeletal remodelling while preserving global tissue polarity. We further identified the extracellular matrix composition as key determinant of bead-tissue interactions, with collagen-I coating promoting robust adhesion and efficient incorporation. At the bead-cell interface, cells assembled tension-bearing focal adhesions and organized actin stress fibers, revealing the emergence of active cortical stress. Strikingly, quantitative analysis of bead deformation revealed a previously unrecognized mechanical duality: spatially segregated regions of pulling and pushing forces coexisted at the microscale, directly correlated with local cytoskeleton dynamics. This finding challenges the prevailing view of homogenous force application and instead supports a model in which cells deploy highly coordinated and spatially patterned force-generating strategies. Altogether, this integrative and non-invasive strategy offers a comprehensive pipeline for dissecting the dynamic interplay between cellular processes and tissue mechanics during morphogenesis in 3D model systems.

cell biology↗

bioRxiv2026

Pre-cuticle DPY 6 acts as a blueprint for aECM periodic organization in C. elegans

Apical extracellular matrices (aECMs) are essential for tissue integrity and function in multicellular organisms, but there is limited understanding of how such matrices are assembled and organized in the extracellular environment. The Caenorhabditis elegans cuticle, a model aECM that undergoes morphogenesis during each of the worms four larval molts, requires periodic circumferential furrows for structural integrity and immune regulation. Here, we show that furrow collagens must be cleaved from their N-terminal transmembrane domain for secretion and depend on the mucin-like pre-cuticle protein DPY-6 for their periodic assembly. While DPY-6 is dispensable for initial embryonic furrow formation, it acts as a mold during subsequent molts, ensuring pattern replication via its C-terminal cysteine cradle domain. These results reveal a central role for a transient matrix factor in organizing a complex periodically structured aECM. Author SummaryIn multicellular organisms, the extracellular matrix (ECM) provides structural support and regulates tissue function. Using the free-living worm Caenorhabditis elegans, we investigated how its apical ECM, the cuticle, forms a precise, repeating pattern of ridges called furrows. The cuticle is rebuilt at each of the worms four larval stages, providing a unique opportunity to study matrix morphogenesis in real time. We discovered that a transient protein, DPY-6, acts as a molecular mold to guide the self-organization of the matrix outside the epidermal cells. DPY-6 ensures that newly secreted proteins assemble into the correct periodic pattern during each rebuilding phase. Without DPY-6, the furrows lose their organization, leading to structural defects and immune system activation. Our findings reveal how a temporary scaffold can template the assembly of a complex, self-organizing structure. This work provides new insights into how biological matrices are built and maintained, with broader implications for understanding ECM assembly in health and disease.

developmental biology↗

bioRxiv2025

Near equilibrium unbinding of streptavidin/biotin using single molecule acoustic force spectroscopy.

The dissociation of the streptavidin-biotin (SA-b) bond has been widely characterized using bulk and single molecule force spectroscopy (SMFS) techniques. However, the dissociation rates (koff) from SMFS ([~]10-1 s-1) typically do not align with those from bulk approaches ([~]10-6 - 10-5 s-1), likely because SMFS measurements are conducted far from equilibrium. Near equilibrium SMFS requires high throughput measurements to obtain large enough statistics, and high stability over long time measurements at ultraslow loading force rates, impractical in most SMFS techniques. Here, we developed in-situ force calibration strategies for acoustic force spectroscopy (AFS) to probe the unbinding forces of SA-b in the near equilibrium regime, from 10 pN/s down to 10- 3 pN/s. The resulting koff was in excellent agreement with that from bulk measurements on the very same system. Combined with our previous SA-b data, we covered 15 orders of magnitude in loading rate, setting the ground for SMFS over the widest dynamic range, essential to fully describe the energy landscape of biomolecular processes.

biophysics↗

bioRxiv2024

Modulation of SARS-CoV-2 spike binding to ACE2 throughconformational selection

The first step of SARS-CoV-2 infection involves the interaction between the trimeric viral spike protein (S) and the host angiotensin-converting enzyme 2 (ACE2). The receptor binding domain (RBD) of S adopts two conformations: open and closed, respectively, accessible and inaccessible to ACE2. Therefore, RBD motions are suspected to affect ACE2 binding; yet a quantitative description of the underlying mechanism has been elusive. Here, using single-molecule approaches, we visualize RBD opening and closing and probe the S/ACE2 interaction. Our results show that RBD dynamics affect ACE2 binding but not unbinding. The resulting modulation is quantitatively predicted by a conformational selection model in which each protomer behaves independently. Our work reveals a general molecular mechanism affecting binding affinity without altering binding strength, helping to understand coronavirus infection and immune evasion.

biophysics↗

bioRxiv2022

Coupled mechanical mapping and interference contrast microscopy reveal viscoelastic and adhesion hallmarks of monocytes differentiation into macrophages

Monocytes in the blood torrent, when activated by pro-inflammatory signals, adhere to the vascular endothelium and migrate into the tissue for ultimately differentiate into macrophages. Mechanics and adhesion play a crucial role in macrophage functions, such as migration and phagocytosis. However, how monocytes change their adhesion and mechanical properties upon differentiation into macrophages is still not well understood. In this work, we combined atomic force microscopy (AFM) viscoelastic mapping with interference contrast microscopy (ICM) to simultaneously probe, at the single-cell level, viscoelasticity and adhesion during monocyte differentiation. THP-1 monocytic cells were differentiated into macrophages through phorbol 12-myristate 13-acetate (PMA). Morphological quantification was achieved using holographic tomography imaging and the expression of integrin subunit CD11b was tracked as a marker of differentiation. Holographic tomography proved to be a quantitative in vivo technique, revealing a dramatic increase in macrophage volume and surface area and two subpopulations, spread and round macrophages. AFM viscoelastic mapping revealed an increased stiffness and more solid-like behavior of differentiated macrophages, especially in the lamellipodia and microvilli regions. Differentiated cells revealed an important increase of the apparent Youngs modulus (E0) and a decrease of cell fluidity ({beta}) on differentiated cells, which correlated with an increase in adhesion area. Macrophages with a spreading phenotype enhanced these changes. Remarkably, when adhesion was eliminated, differentiated macrophages remained stiffer and more solid-like than monocytes, suggesting a permanent reorganization of the cytoskeleton. We speculate that the more solid-like microvilli and lamellipodia might help macrophages to minimize energy dissipation during mechanosensitive activity, such as phagocytosis, making it more efficient. Our proposed approach revealed viscoelastic and adhesion hallmarks of monocyte differentiation that may be important for biological function.

biophysics↗

bioRxiv2022

Combining DNA scaffolds and acoustic force spectroscopy to characterize individual protein bonds

Single-molecule data are of great significance in biology, chemistry, and medicine. However, experimental tools to characterize, in a multiplexed manner, protein bond rupture under force are needed. Acoustic force spectroscopy (AFS) is an emerging manipulation technique which generates acoustic waves to apply force in parallel on a large population of microbeads tethered to a surface. We have exploited this configuration on a recently developed modular Junctured-DNA (J-DNA) scaffold designed to study protein-protein interactions at the single-molecule level. By applying repetitive constant force steps on the FKBP12-rapamycin-FRB complex, we measured its unbinding kinetics under force at the single-bond level. Special effort was made in analyzing the data in order to identify potential pitfalls. We established a calibration method allowing in situ force determination during the course of the unbinding measurement. We compare our results with well established techniques, such as magnetic tweezers, to ensure their accuracy. We also apply our strategy for measuring the force dependent rupture of a single domain antibody with its antigen. We get a good agreement with standard measurement at zero force. Our technique offers single molecule precision for multiplexed measurements of interactions of biotechnological and medical interest.

biophysics↗