Léa Chazot-Franguiadakis

École Normale Supérieure de Lyon

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Bastien Molcrette

Université Claude Bernard Lyon 1

Fabien Montel

École Normale Supérieure de Lyon

Philippe Guégan

Centre National de la Recherche Scientifique

Anna Salvetti

Inserm

François Liénard

Laboratoire de Physique de l'ENS de Lyon

Pascal Bernard

École Normale Supérieure de Lyon

Marylène Mougel

Centre National de la Recherche Scientifique

Amélie Malapert

Université Claude Bernard Lyon 1

Véronique Bennevault

Centre National de la Recherche Scientifique

Jean Lacroix

Université Paris Cité

Cooperative Institutions

Centre National de la Recherche Scientifique

Université Claude Bernard Lyon 1

École Normale Supérieure de Lyon

Laboratoire de Physique de l'ENS de Lyon

Université Paris Cité

Inserm

Clinique Claude-Bernard

Sorbonne Université

Commissariat à l'Énergie Atomique et aux Énergies Alternatives

Institut Parisien de Chimie Moléculaire

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The sequence of the extruded non-template strand determines the architecture of R-loops

bioRxiv (Cold Spring Harbor Laboratory) (2019)

Yeraldinne Carrasco-Salas Amélie Malapert Shaheen Sulthana Bastien Molcrette Léa Chazot-Franguiadakis

ABSTRACT Three-stranded R-loop structures have been associated with genomic instability phenotypes. What underlies their wide-ranging effects on genome stability remains poorly understood. Here we combined biochemical and atomic force microscopy approaches with single molecule R-loop footprinting to demonstrate that R-loops formed at the model Airn locus in vitro adopt a defined set of three-dimensional conformations characterized by distinct shapes and volumes, which we call R-loop objects. Interestingly, we show that these R-loop objects impose specific physical constraints on the DNA, as revealed by the presence of stereotypical angles in the surrounding DNA. Biochemical probing and mutagenesis experiments revealed that the formation of R-loop objects at Airn is dictated by the sequence of the extruded non-template strand, suggesting that R-loops possess intrinsic sequence-driven properties. Consistent with this, we show that R-loops formed at the fission yeast gene sum3 do not form detectable R-loop objects. Our results reveal that R-loops differ by their architectures and that the organization of the non-template strand is a fundamental characteristic of R-loops, which could explain that only a subset of R-loops is associated with replication-dependent DNA breaks.

Sequence (biology)

10.1101/576561

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Citations (1)

Soft jamming of viral particles in nanopores

Nature Communications (2024)

Léa Chazot-Franguiadakis Joëlle Eid Gwendoline Delecourt Pauline J. Kolbeck Saskia Brugère

Abstract Viruses have remarkable physical properties and complex interactions with their environment. However, their aggregation in confined spaces remains unexplored, although this phenomenon is of paramount importance for understanding viral infectivity. Using hydrodynamical driving and optical detection, we developed a method to detect the transport of single virus in real time through synthetic nanopores. We unveiled a jamming phenomenon specifically associated with virus confinement under flow. We showed that the interactions of viral particles with themselves and with the pore surface were critical for clog formation. Based on the detailed screening of the physical and chemical determinants, we proposed a simple dynamical model that recapitulated all the experimental observations. Our results pave the way for the study of jamming phenomena in the presence of more complex interactions.

Nanopore

Infectivity

10.1038/s41467-024-50059-9

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The extruded non-template strand determines the architecture of R-loops

Nucleic Acids Research (2019)

Yeraldinne Carrasco-Salas Amélie Malapert Shaheen Sulthana Bastien Molcrette Léa Chazot-Franguiadakis

Abstract Three-stranded R-loop structures have been associated with genomic instability phenotypes. What underlies their wide-ranging effects on genome stability remains poorly understood. Here we combined biochemical and atomic force microscopy approaches with single molecule R-loop footprinting to demonstrate that R-loops formed at the model Airn locus in vitro adopt a defined set of three-dimensional conformations characterized by distinct shapes and volumes, which we call R-loop objects. Interestingly, we show that these R-loop objects impose specific physical constraints on the DNA, as revealed by the presence of stereotypical angles in the surrounding DNA. Biochemical probing and mutagenesis experiments revealed that the formation of R-loop objects at Airn is dictated by the extruded non-template strand, suggesting that R-loops possess intrinsic sequence-driven properties. Consistent with this, we show that R-loops formed at the fission yeast gene sum3 do not form detectable R-loop objects. Our results reveal that R-loops differ by their architectures and that the organization of the non-template strand is a fundamental characteristic of R-loops, which could explain that only a subset of R-loops is associated with replication-dependent DNA breaks.

10.1093/nar/gkz341

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Citations (37)

Optical Quantification by Nanopores of Viruses, Extracellular Vesicles, and Nanoparticles

Nano Letters (2022)

Léa Chazot-Franguiadakis Joëlle Eid Marius Socol Bastien Molcrette Philippe Guégan

Nanopores combined with optical approaches can be used to detect viral particles. In this work, we demonstrate the ability of hydrodynamical driving and optical sensing to identify and quantify viral particles in a biological sample. We have developed a simple and rapid method which requires only fluorescent labeling of the particles and can therefore be applied to a wide range of virus type. The system operates in real time and at the single particle level while providing a low error on concentration (4%) and a low limit of detection of 105 particles/mL for an acquisition time of 60 s with the ability to increase the acquisition time to achieve a lower limit.

Nanopore

Particle (ecology)

Extracellular Vesicles

10.1021/acs.nanolett.2c00253

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Citations (5)

Experimental study of a nanoscale translocation ratchet

Proceedings of the National Academy of Sciences (2022)

Bastien Molcrette Léa Chazot-Franguiadakis François Liénard Zsombor Balassy Céline Freton

Despite an extensive theoretical and numerical background, the translocation ratchet mechanism, which is fundamental for the transmembrane transport of biomolecules, has never been experimentally reproduced at the nanoscale. Only the Sec61 and bacterial type IV pilus pores were experimentally shown to exhibit a translocation ratchet mechanism. Here we designed a synthetic translocation ratchet and quantified its efficiency as a nanopump. We measured the translocation frequency of DNA molecules through nanoporous membranes and showed that polycations at the trans side accelerated the translocation in a ratchet-like fashion. We investigated the ratchet efficiency according to geometrical and kinetic parameters and observed the ratchet to be only dependent on the size of the DNA molecule with a power law [Formula: see text]. A threshold length of 3 kbp was observed, below which the ratchet did not operate. We interpreted this threshold in a DNA looping model, which quantitatively explained our results.

Ratchet effect

10.1073/pnas.2202527119

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Citations (7)

Thermally Switchable Nanogate Based on Polymer Phase Transition

Nano Letters (2023)

Pauline J. Kolbeck Dihia Benaoudia Léa Chazot-Franguiadakis Gwendoline Delecourt Jérôme Mathé

Mimicking and extending the gating properties of biological pores is of paramount interest for the fabrication of membranes that could be used in filtration or drug processing. Here, we build a selective and switchable nanopore for macromolecular cargo transport. Our approach exploits polymer graftings within artificial nanopores to control the translocation of biomolecules. To measure transport at the scale of individual biomolecules, we use fluorescence microscopy with a zero-mode waveguide set up. We show that grafting polymers that exhibit a lower critical solution temperature creates a toggle switch between an open and closed state of the nanopore depending on the temperature. We demonstrate tight control over the transport of DNA and viral capsids with a sharp transition (∼1 °C) and present a simple physical model that predicts key features of this transition. Our approach provides the potential for controllable and responsive nanopores in a range of applications.

Nanopore

Biomolecule

10.1021/acs.nanolett.3c00438

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Citations (3)

Catch and Thread: experimental study of a nanoscale translocation ratchet

Zenodo (CERN European Organization for Nuclear Research) (2022)

Bastien Molcrette Léa Chazot-Franguiadakis François Liénard Zsombor Balassy Céline Freton

10.5281/zenodo.6802447

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Quelques éléments de physique autour des nanopores biologiques

Reflets de la physique (2023)

Bastien Molcrette Léa Chazot-Franguiadakis Thomas Auger Fabien Montel

Les nanopores biologiques sont d’étonnantes machines moléculaires. Ils remplissent une grande variété de fonctions, allant du tri des biomolécules à la transmission des signaux dans nos neurones et au repliement des protéines nouvellement produites. Le membre le plus surprenant de ce club est le pore nucléaire. Il régule le flux de molécules entre le noyau et l’intérieur de la cellule. Ses performances, mesurées par son efficacité énergétique, sa directionnalité ou sa sélectivité, n’ont pas d’équivalent dans les systèmes artificiels. Nous verrons que la compréhension de son fonctionnement permet d’appréhender des phénomènes physiques nouveaux et d’imaginer des systèmes de filtration sélectifs, ainsi que des pompes moléculaires.

10.1051/refdp/202375018

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