Mathieu Coppey

Magneto-fluorescent core-shell supernanoparticles

Magneto-fluorescent particles have been recognized as an emerging class of materials that exhibit great potential in advanced applications. However, synthesizing such magneto-fluorescent nanomaterials that simultaneously exhibit uniform and tunable sizes, high magnetic content loading, maximized fluorophore coverage at the surface, and a versatile surface functionality has proven challenging. Here we report a simple approach for co-assembling magnetic nanoparticles with fluorescent quantum dots to form colloidal magneto-fluorescent supernanoparticles. Importantly, these supernanoparticles exhibit a superstructure consisting of a close packed magnetic nanoparticle “core” which is fully surrounded by a “shell” of fluorescent quantum dots. A thin layer of silica-coating provides high colloidal stability and biocompatiblity and a versatile surface functionality. We demonstrate that after surface pegylation, these silica-coated magneto-fluorescent supernanoparticles can be magnetically manipulated inside living cells while being optically tracked. Moreover, our silica-coated magneto-fluorescent supernanoparticles can also serve as an in vivo multi-photon and magnetic resonance dual-modal imaging probe.

Actin Flows Mediate a Universal Coupling between Cell Speed and Cell Persistence

Cell movement has essential functions in development, immunity, and cancer. Various cell migration patterns have been reported, but no general rule has emerged so far. Here, we show on the basis of experimental data in vitro and in vivo that cell persistence, which quantifies the straightness of trajectories, is robustly coupled to cell migration speed. We suggest that this universal coupling constitutes a generic law of cell migration, which originates in the advection of polarity cues by an actin cytoskeleton undergoing flows at the cellular scale. Our analysis relies on a theoretical model that we validate by measuring the persistence of cells upon modulation of actin flow speeds and upon optogenetic manipulation of the binding of an actin regulator to actin filaments. Beyond the quantitative prediction of the coupling, the model yields a generic phase diagram of cellular trajectories, which recapitulates the full range of observed migration patterns.

MAPK substrate competition integrates patterning signals in the Drosophila embryo.

Terminal regions of the Drosophila embryo are patterned by the localized activation of the mitogen-activated protein kinase (MAPK) pathway. This depends on the MAPK-mediated downregulation of Capicua (Cic), a repressor of the terminal gap genes. We establish that downregulation of Cic is antagonized by the anterior patterning morphogen Bicoid (Bcd). We demonstrate that this effect does not depend on transcriptional activity of Bcd and provide evidence suggesting that Bcd, a direct substrate of MAPK, decreases the availability of MAPK for its other substrates, such as Cic. Based on the quantitative analysis of MAPK signaling in multiple mutants, we propose that MAPK substrate competition coordinates the actions of the anterior and terminal patterning systems. In addition, we identify Hunchback as a novel target of MAPK phosphorylation that can account for the previously described genetic interaction between the posterior and terminal systems. Thus, a common enzyme-substrate competition mechanism can integrate the actions of the anterior, posterior, and terminal patterning signals. Substrate competition can be a general signal integration strategy in networks where enzymes, such as MAPK, interact with their multiple regulators and targets.

Subcellular control of Rac-GTPase signalling by magnetogenetic manipulation inside living cells

Many cell functions rely on the coordinated activity of signalling pathways at a subcellular scale. However, there are few tools capable of probing and perturbing signalling networks with a spatial resolution matching the intracellular dimensions of their activity patterns. Here we present a generic magnetogenetic approach based on the self-assembly of signalling complexes on the surface of functionalized magnetic nanoparticles inside living cells. The nanoparticles act as nanoscopic hot spots that can be displaced by magnetic forces and trigger signal transduction pathways that bring about a cell response. We applied this strategy to Rho-GTPases, a set of molecular switches known to regulate cell morphology via complex spatiotemporal patterns of activity. We demonstrate that the nanoparticle-mediated activation of signalling pathways leads to local remodelling of the actin cytoskeleton and to morphological changes.

Live Imaging of Bicoid-Dependent Transcription in Drosophila Embryos

The early Drosophila embryo is an ideal model to understand the transcriptional regulation of well-defined patterns of gene expression in a developing organism. In this system, snapshots of transcription measurements obtained by RNA FISH on fixed samples cannot provide the temporal resolution needed to distinguish spatial heterogeneity from inherent noise. Here, we used the MS2-MCP system to visualize in living embryos nascent transcripts expressed from the canonical hunchback (hb) promoter under the control of Bicoid (Bcd). The hb-MS2 reporter is expressed as synchronously as endogenous hb in the anterior half of the embryo, but unlike hb it is also active in the posterior, though more heterogeneously and more transiently than in the anterior. The length and intensity of active transcription periods in the anterior are strongly reduced in absence of Bcd, whereas posterior ones are mostly Bcd independent. This posterior noisy signal decreases progressively through nuclear divisions, so that the MS2 reporter expression mimics the known anterior hb pattern at cellularization. We propose that the establishment of the hb pattern relies on Bcd-dependent lengthening of transcriptional activity periods in the anterior and may require two distinct repression mechanisms in the posterior.

Nuclear Trapping Shapes the Terminal Gradient in the Drosophila Embryo

Patterning of the terminal regions of the Drosophila embryo relies on the gradient of phosphorylated ERK/MAPK (dpERK), which is controlled by the localized activation of the Torso receptor tyrosine kinase [1-4]. This model is supported by a large amount of data, but the gradient itself has never been quantified. We present the first measurements of the dpERK gradient and establish a new intracellular layer of its regulation. Based on the quantitative analysis of the spatial pattern of dpERK in mutants with different levels of Torso as well as the dynamics of the wild-type dpERK pattern, we propose that the terminal-patterning gradient is controlled by a cascade of diffusion-trapping modules. A ligand-trapping mechanism establishes a sharply localized pattern of the Torso receptor occupancy on the surface of the embryo. Inside the syncytial embryo, nuclei play the role of traps that localize diffusible dpERK. We argue that the length scale of the terminal-patterning gradient is determined mainly by the intracellular module.

Magnetogenetic Control of Protein Gradients Inside Living Cells with High Spatial and Temporal Resolution

Tools for controlling the spatial organization of proteins are a major prerequisite for deciphering mechanisms governing the dynamic architecture of living cells. Here, we have developed a generic approach for inducing and maintaining protein gradients inside living cells by means of biofunctionalized magnetic nanoparticles (MNPs). For this purpose, we tailored the size and surface properties of MNPs in order to ensure unhindered mobility in the cytosol. These MNPs with a core diameter below 50 nm could be rapidly relocalized in living cells by exploiting biased diffusion at weak magnetic forces in the femto-Newton range. In combination with MNP surface functionalization for specific in situ capturing of target proteins as well as efficient delivery into the cytosplasm, we here present a comprehensive technology for controlling intracellular protein gradients with a temporal resolution of a few tens of seconds.

Averaged residence times of stochastic motions in bounded domains

Two years ago, Blanco and Fournier (Blanco S. and Fournier R., Europhys. Lett. 61 (2003) 168) calculated the mean first exit time of a domain of a particle undergoing a randomly reoriented ballistic motion which starts from the boundary. They showed that it is simply related to the ratio of the volumes domain over its surface. This work was extended by Mazzolo (Mazzolo A., Europhys. Lett. 68 (2004) 350), who studied the case of trajectories which start inside the volume. In this letter, we propose an alternative formulation of the problem which allows us to calculate not only the mean exit time, but also the mean residence time inside a sub-domain. The cases of any combinations of reflecting and absorbing boundary conditions are considered. Lastly, we generalize our results for a wide class of stochastic motions.

Transcriptional Memory in the Drosophila Embryo

Transmission of active transcriptional states from mother to daughter cells has the potential to foster precision in the gene expression programs underlying development. Such transcriptional memory has been specifically proposed to promote rapid reactivation of complex gene expression profiles after successive mitoses in Drosophila development [1]. By monitoring transcription in living Drosophila embryos, we provide the first evidence for transcriptional memory in animal development. We specifically monitored the activities of stochastically expressed transgenes in order to distinguish active and inactive mother cells and the behaviors of their daughter nuclei after mitosis. Quantitative analyses reveal that there is a 4-fold higher probability for rapid reactivation after mitosis when the mother experienced transcription. Moreover, memory nuclei activate transcription twice as fast as neighboring inactive mothers, thus leading to augmented levels of gene expression. We propose that transcriptional memory is a mechanism of precision, which helps coordinate gene activity during embryogenesis.

Modelling the early steps of transduction in insect olfactory receptor neurons

Olfactory transduction is a multistep process whose basic function is to convert a low energy reaction, the odorant-receptor interaction that may involve a single odorant molecule, into a whole cell electrical response, the receptor potential, which triggers the firing of one or more action potentials. Although much effort has been devoted to the experimental analysis of transduction in olfactory receptor neurons (ORNS), especially in the favorable moth sex-pheromone receptor neuron, its modelling is less advanced. The model we investigated, which takes into account the translocation of pheromone molecules from air to the extracellular space, their deactivation and their interaction with receptors, focuses on the membrane cascade. It involves the interaction of receptors, G-proteins and effector enzymes, whose reaction rates are limited by lateral diffusion in the membrane. The evolutions in time of these species in response to single pulse stimulation of various intensities were compared to one another and to the experimentally measured electrical response. The results obtained suggest that the receptor-to-effector conversion is fast with respect to the receptor response, that it presents a small amplification factor, contrary to the photoreceptor, and that most of the amplification is achieved in the post-effector processes involving the second messenger and ionic channels.