UMR168 – Laboratoire Physico-Chimie Curie

Publications de l’UMR 168

Année de publication : 2016

Benjamin Brunel, Grégory Beaune, Usharani Nagarajan, Sylvie Dufour, Françoise Brochard-Wyart, Françoise M Winnik (2016 Oct 8)

Nanostickers for cells: a model study using cell-nanoparticle hybrid aggregates.

Soft matter : 7902-7907 En savoir plus
Résumé

We present direct evidence that nanoparticles (NPs) can stick together cells that are inherently non-adhesive. Using cadherin-depleted S180 murine cells lines, which exhibit very low cell-cell adhesion, we show that NPs can assemble dispersed single cells into large cohesive aggregates. The dynamics of aggregation, which is controlled by diffusion and collision, can be described as a second-order kinetic law characterized by a rate of collision that depends on the size, concentration, and surface chemistry of the NPs. We model the cell-cell adhesion induced by the « nanostickers » using a three-state dynamical model, where the NPs are free, adsorbed on the cell membrane or internalized by the cells. We define a « sticking efficiency parameter » to compare NPs and look for the most efficient type of NP. We find that 20 nm carboxylated polystyrene NPs are more efficient nanostickers than 20 nm silica NPs which were reported to induce fast wound healing and to glue soft tissues. Nanostickers, by increasing the cohesion of tissues and tumors, may have important applications for tissue engineering and cancer treatment.

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Année de publication : 2017

Duclos G., Erlenkämper C., Joanny J.-F., Silberzan P. (2016 Sep 12)

Topological defects in confined populations of spindle-shaped cells

Nature Physics : 13 : 58-62 : DOI : 10.1038/nphys3876 En savoir plus
Résumé

Most spindle-shaped cells (including smooth muscles and sarcomas) organize in vivo into well-aligned ‘nematic’ domains, creating intrinsic topological defects that may be used to probe the behaviour of these active nematic systems. Active non-cellular nematics have been shown to be dominated by activity, yielding complex chaotic flows. However, the regime in which live spindle-shaped cells operate, and the importance of cell–substrate friction in particular, remains largely unexplored. Using in vitro experiments, we show that these active cellular nematics operate in a regime in which activity is effectively damped by friction, and that the interaction between defects is controlled by the system’s elastic nematic energy. Due to the activity of the cells, these defects behave as self-propelled particles and pairwise annihilate until all displacements freeze as cell crowding increases. When confined in mesoscopic circular domains, the system evolves towards two identical +1/2 disclinations facing each other. The most likely reduced positions of these defects are independent of the size of the disk,the cells’ activity or even the cell type, but are well described by equilibrium liquid crystal theory. These cell-based systems thus operate in a regime more stable than other active nematics, which may be necessary for their biological function.

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Année de publication : 2016

Marina Casiraghi, Marjorie Damian, Ewen Lescop, Elodie Point, Karine Moncoq, Nelly Morellet, Daniel Levy, Jacky Marie, Eric Guittet, Jean-Louis Banères, Laurent J Catoire (2016 Aug 5)

Functional Modulation of a G Protein-Coupled Receptor Conformational Landscape in a Lipid Bilayer.

Journal of the American Chemical Society : 11170-5 : DOI : 10.1021/jacs.6b04432 En savoir plus
Résumé

Mapping the conformational landscape of G protein-coupled receptors (GPCRs), and in particular how this landscape is modulated by the membrane environment, is required to gain a clear picture of how signaling proceeds. To this end, we have developed an original strategy based on solution-state nuclear magnetic resonance combined with an efficient isotope labeling scheme. This strategy was applied to a typical GPCR, the leukotriene B4 receptor BLT2, reconstituted in a lipid bilayer. Because of this, we are able to provide direct evidence that BLT2 explores a complex landscape that includes four different conformational states for the unliganded receptor. The relative distribution of the different states is modulated by ligands and the sterol content of the membrane, in parallel with the changes in the ability of the receptor to activate its cognate G protein. This demonstrates a conformational coupling between the agonist and the membrane environment that is likely to be fundamental for GPCR signaling.

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A Bertin, E Nogales (2016 Jul 31)

Preparing recombinant yeast septins and their analysis by electron microscopy.

Methods in cell biology : 21-34 : DOI : 10.1016/bs.mcb.2016.03.010 En savoir plus
Résumé

Septins are highly conserved and essential eukaryotic cytoskeletal proteins that interact with the inner plasma membrane. They are involved in essential functions requiring cell membrane remodeling and compartmentalization, such as cell division and dendrite morphogenesis, and have been implicated in numerous diseases. Depending on the organisms and on the type of tissue, a specific set of septins genes are expressed, ranging from 2 to 13. Septins self-assemble into linear, symmetric rods that can further organize into linear filaments several microns in length. Only a subset of human septins has been described at high resolution by X-ray crystallography (Sirajuddin et al., 2007). Electron microscopy (EM) has proven to be a method of choice for analyzing the molecular organization of septins. It is possible to localize each septin subunit within the rod complex using genetic tags, such as maltose-binding protein or green fluorescent protein, to generate a visible label of a specific septin subunit in EM images that are processed using single-particle EM methodology. In this chapter we present, in detail, the methods that we have used to analyze the molecular organization of budding yeast septins (Bertin et al., 2008). These methods include purification of septin complexes, sample preparation for EM, and image processing procedures. Such methods can be generalized to analyze the organization of septins from any organism.

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Caorsi V, Lemière J, Campillo C, Bussonnier M, Manzi J, Betz T, Plastino J, Carvalho K, Sykes C (2016 Jul 20)

Cell-sized liposome doublets reveal active tension build-up driven by acto-myosin dynamics

Soft Matter : 12 : 6223-31 : DOI : 10.1039/c6sm00856a En savoir plus
Résumé

Cells modulate their shape to fulfill specific functions, mediated by the cell cortex, a thin actin shell bound to the plasma membrane. Myosin motor activity, together with actin dynamics, contributes to cortical tension. Here, we examine the individual contributions of actin polymerization and myosin activity to tension increase with a non-invasive method. Cell-sized liposome doublets are covered with either a stabilized actin cortex of preformed actin filaments, or a dynamic branched actin network polymerizing at the membrane. The addition of myosin II minifilaments in both cases triggers a change in doublet shape that is unambiguously related to a tension increase. Preformed actin filaments allow us to evaluate the effect of myosin alone while, with dynamic actin cortices, we examine the synergy of actin polymerization and myosin motors in driving shape changes. Our assay paves the way for a quantification of tension changes triggered by various actin-associated proteins in a cell-sized system.

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Leïla Perié, Ken R Duffy (2016 Jul 13)

Retracing the in vivo haematopoietic tree using single-cell methods.

FEBS letters : DOI : 10.1002/1873-3468.12299 En savoir plus
Résumé

The dynamic process by which self-renewing stem cells and their offspring proliferate and differentiate to create the erythroid, myeloid and lymphoid lineages of the blood system has long since been an important topic of study. A range of recent single cell and family tracing methodologies such as massively parallel single-cell RNA-sequencing, mass cytometry, integration site barcoding, cellular barcoding and transposon barcoding are enabling unprecedented analysis, dissection and re-evaluation of the haematopoietic tree. In addition to the substantial experimental advances, these new techniques have required significant theoretical development in order to make biological deductions from their data. Here, we review these approaches from both an experimental and inferential point of view, considering their discoveries to date, their capabilities, limitations and opportunities for further development.

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Mijo Simunovic, Coline Prévost, Andrew Callan-Jones, Patricia Bassereau (2016 Jun 15)

Physical basis of some membrane shaping mechanisms.

Philosophical transactions. Series A, Mathematical, physical, and engineering sciences : DOI : 10.1098/rsta.2016.0034 En savoir plus
Résumé

In vesicular transportation pathways, membrane lipids and proteins are internalized, externalized or transported Within cells, not by bulk diffusion of single molecules, goal embedded in the membrane of small vesicles or thin tubules. The formation of These ‘transportation carriers’ Follows sequential events: bending membrane fission from the donor compartment, and transportation Eventually fusion with the acceptor membrane. A similar sequence is Involved During the internalization of drug or gene carriers inside cells. These membrane-shaping events are mediated by proteins Generally binding to membranes. The thesis Mechanisms behind biological processes are Actively Studied Both in the context of cell biology and biophysics. Bin / Amphiphysin / Rvs (BAR) domain proteins are Ideally suited for single Illustrating how soft matter principles can account for deformation by membrane proteins. We review here Some experimental methods and theoretical models to measure Corresponding thesis how proteins affect the mechanics and the shape of membranes. In more detail, we show how an experimental method Employing optical tweezers to pull a tube from a giant vesicle May give significant quantitative insights into the mechanism by which proteins sense and generate membrane curvature and the mechanism of membrane scission.This article is share of the themed issue ‘Soft interfacial materials: from fundamentals to formulation’.

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Renaud Renault, Jean-Baptiste Durand, Jean-Louis Viovy, Catherine Villard (2016 May 27)

Asymmetric axonal edge guidance: a new paradigm for building oriented neuronal networks.

Lab on a chip : 2188-91 : DOI : 10.1039/c6lc00479b En savoir plus
Résumé

We present a novel kind of directional axon guides for brain-on-a-chip applications. Contrarily to previous works, the directionality in our design is created by rerouting axons growing in the unwanted direction back to their original compartment while leaving the other growth direction unaffected. This design yields state-of-the-art levels of directionality without the disadvantages of previously reported technologies.

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Evangelos Gogolides, Angeliki Tserepi, Gerhard Jobst, Jean-Michel Friedt, David Rabus, Bruno Dupuy, Zuzana Bilkova, Stephanie Descroix, Jean-Louis Viovy, George Papadakis, Electra Gizeli (2016 May 27)

Micro-Nano-Bio Diagnostic System for Food Pathogen Detection Revolutionizes Food Safety Management & Protects Consumers Health.

Studies in health technology and informatics : 67-72 En savoir plus
Résumé

The development of integrated, fast and affordable platforms for pathogen detection is an emerging area where a multidisciplinary approach is necessary for designing microsystems employing miniaturized devices; these new technologies promise a significant advancement of the current state of analytical testing leading to improved healthcare. In this work, the development of a lab-on-chip microsystem platform for the genetic analysis of Salmonella in milk samples is presented. The heart of the platform is an acoustic detection biochip, integrated with a microfluidic module. This detection platform is combined with a micro-processor, which, alongside with magnetic beads technology and a DNA micro-amplification module, are responsible for performing sample pre-treatment, bacteria lysis, nucleic acid purification and amplification. Automated, multiscale manipulation of fluids in complex microchannel networks is combined with novel sensing principles developed by some of the partners. This system is expected to have a significant impact in food-pathogen detection by providing for the first time an integrated detection test for Salmonella screening in a very short time. Finally, thanks to the low cost and compact technologies involved, the proposed set-up is expected to provide a competitive analytical platform for direct application in field settings.

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Ayako Yamada, Maéva Vignes, Cécile Bureau, Alexandre Mamane, Bastien Venzac, Stéphanie Descroix, Jean-Louis Viovy, Catherine Villard, Jean-Michel Peyrin, Laurent Malaquin (2016 May 13)

In-mold patterning and actionable axo-somatic compartmentalization for on-chip neuron culture.

Lab on a chip : 2059-68 : DOI : 10.1039/c6lc00414h En savoir plus
Résumé

Oriented neuronal networks with controlled connectivity are required for many applications ranging from studies of neurodegeneration to neuronal computation. To build such networks in vitro, an efficient, directed and long lasting guidance of axons toward their target is a pre-requisite. The best guidance achieved so far, however, relies on confining axons in enclosed microchannels, making them poorly accessible for further investigation. Here we describe a method providing accessible and highly regular arrays of axons, emanating from somas positioned in distinct compartments. This method combines the use of a novel removable partition, allowing soma positioning outside of the axon guidance patterns, and in-mold patterning (iMP), a hybrid method combining chemical and mechanical cell positioning clues applied here for the first time to neurons. The axon guidance efficiency of iMP is compared to that of conventional patterning methods, e.g. micro-contact printing (chemical constraints by a poly-l-lysine motif) and micro-grooves (physical constraints by homogeneously coated microstructures), using guiding tracks of different widths and spacing. We show that iMP provides a gain of 10 to 100 in axon confinement efficiency on the tracks, yielding mm-long, highly regular, and fully accessible on-chip axon arrays. iMP also allows well-defined axon guidance from small populations of several neurons confined at predefined positions in μm-sized wells. iMP will thus open new routes for the construction of complex and accurately controlled neuronal networks.

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Davide Ferraro, Jérôme Champ, Bruno Teste, Marco Serra, Laurent Malaquin, Jean-Louis Viovy, Patricia de Cremoux, Stephanie Descroix (2016 May 10)

Microfluidic platform combining droplets and magnetic tweezers: application to HER2 expression in cancer diagnosis.

Scientific reports : 25540 : DOI : 10.1038/srep25540 En savoir plus
Résumé

The development of precision medicine, together with the multiplication of targeted therapies and associated molecular biomarkers, call for major progress in genetic analysis methods, allowing increased multiplexing and the implementation of more complex decision trees, without cost increase or loss of robustness. We present a platform combining droplet microfluidics and magnetic tweezers, performing RNA purification, reverse transcription and amplification in a fully automated and programmable way, in droplets of 250nL directly sampled from a microtiter-plate. This platform decreases sample consumption about 100 fold as compared to current robotized platforms and it reduces human manipulations and contamination risk. The platform’s performance was first evaluated on cell lines, showing robust operation on RNA quantities corresponding to less than one cell, and then clinically validated with a cohort of 21 breast cancer samples, for the determination of their HER2 expression status, in a blind comparison with an established routine clinical analysis.

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Jean-Francois Rupprecht, Jacques Prost (2016 Apr 30)

PHYSICAL BIOLOGY. A fresh eye on nonequilibrium systems.

Science (New York, N.Y.) : 514-5 : DOI : 10.1126/science.aaf4611 En savoir plus
Résumé

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Alexander Y Grosberg, Jean-François Joanny, Watee Srinin, Yitzhak Rabin (2016 Apr 28)

Scale-Dependent Viscosity in Polymer Fluids.

The journal of physical chemistry. B En savoir plus
Résumé

In this communication, we use simple physical arguments to construct a « phase diagram » of various frequency and wave vector-dependent regimes of effective viscosity for polymer fluids, including nonentangled and entangled melts, semidilute solutions without and with hydrodynamic interactions, as well as the more exotic case of a melt of unconcatenated ring polymers.

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Laura Wagstaff, Maja Goschorska, Kasia Kozyrska, Guillaume Duclos, Iwo Kucinski, Anatole Chessel, Lea Hampton-O'Neil, Charles R Bradshaw, George E Allen, Emma L Rawlins, Pascal Silberzan, Rafael E Carazo Salas, Eugenia Piddini (2016 Apr 26)

Mechanical cell competition kills cells via induction of lethal p53 levels.

Nature communications : 11373 : DOI : 10.1038/ncomms11373 En savoir plus
Résumé

Cell competition is a quality control mechanism that eliminates unfit cells. How cells compete is poorly understood, but it is generally accepted that molecular exchange between cells signals elimination of unfit cells. Here we report an orthogonal mechanism of cell competition, whereby cells compete through mechanical insults. We show that MDCK cells silenced for the polarity gene scribble (scrib(KD)) are hypersensitive to compaction, that interaction with wild-type cells causes their compaction and that crowding is sufficient for scrib(KD) cell elimination. Importantly, we show that elevation of the tumour suppressor p53 is necessary and sufficient for crowding hypersensitivity. Compaction, via activation of Rho-associated kinase (ROCK) and the stress kinase p38, leads to further p53 elevation, causing cell death. Thus, in addition to molecules, cells use mechanical means to compete. Given the involvement of p53, compaction hypersensitivity may be widespread among damaged cells and offers an additional route to eliminate unfit cells.

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Vincent Nier, Shreyansh Jain, Chwee Teck Lim, Shuji Ishihara, Benoit Ladoux, Philippe Marcq (2016 Apr 14)

Inference of Internal Stress in a Cell Monolayer.

Biophysical journal : 1625-35 : DOI : 10.1016/j.bpj.2016.03.002 En savoir plus
Résumé

We combine traction force data with Bayesian inversion to obtain an absolute estimate of the internal stress field of a cell monolayer. The method, Bayesian inversion stress microscopy, is validated using numerical simulations performed in a wide range of conditions. It is robust to changes in each ingredient of the underlying statistical model. Importantly, its accuracy does not depend on the rheology of the tissue. We apply Bayesian inversion stress microscopy to experimental traction force data measured in a narrow ring of cohesive epithelial cells, and check that the inferred stress field coincides with that obtained by direct spatial integration of the traction force data in this quasi one-dimensional geometry.

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