Inactivating negative regulators of cortical branched actin enhances persistence of single cell migration.

The Rac1-WAVE-Arp2/3 pathway pushes the plasma membrane by polymerizing branched actin, thereby powering membrane protrusions that mediate cell migration. Here, using knockdown (KD) or knockout (KO), we combine the inactivation of the Arp2/3 inhibitory protein arpin, the Arp2/3 subunit ARPC1A and the WAVE complex subunit CYFIP2, all of which enhance the polymerization of cortical branched actin. Inactivation of the three negative regulators of cortical branched actin increases migration persistence of human breast MCF10A cells and of endodermal cells in the zebrafish embryo, significantly more than any single or double inactivation. In the triple KO cells, but not in triple KD cells, the 'super-migrator' phenotype was associated with a heterogenous downregulation of vimentin (VIM) expression and a lack of coordination in collective behaviors, such as wound healing and acinus morphogenesis. Re-expression of vimentin in triple KO cells largely restored normal persistence of single cell migration, suggesting that vimentin downregulation contributes to the maintenance of the super-migrator phenotype in triple KO cells. Constant excessive production of branched actin at the cell cortex thus commits cells into a motile state through changes in gene expression.

Live 3D imaging and mapping of shear stresses within tissues using incompressible elastic beads.

To investigate the role of mechanical constraints in morphogenesis and development, we have developed a pipeline of techniques based on incompressible elastic sensors. These techniques combine the advantages of incompressible liquid droplets, which have been used as precise in situ shear stress sensors, and of elastic compressible beads, which are easier to tune and to use. Droplets of a polydimethylsiloxane mix, made fluorescent through specific covalent binding to a rhodamin dye, are produced by a microfluidics device. The elastomer rigidity after polymerization is adjusted to the tissue rigidity. Its mechanical properties are carefully calibrated in situ, for a sensor embedded in a cell aggregate submitted to uniaxial compression. The local shear stress tensor is retrieved from the sensor shape, accurately reconstructed through an active contour method. In vitro, within cell aggregates, and in vivo, in the prechordal plate of the zebrafish embryo during gastrulation, our pipeline of techniques demonstrates its efficiency to directly measure the three dimensional shear stress repartition within a tissue.

Mid-infrared tunable filter based on a femtosecond-written silica volume Bragg grating.

A silica volume Bragg grating (VBG) is used to filter the light of a mid-infrared (mid-IR) supercontinuum laser. The VBG with a 7 µm period was inscribed with 800 nm pulses with a 100 fs duration and the phase-mask technique over a glass thickness of 3 mm. Despite silica's absorption, the VBG allows obtaining a narrowband light source tunable from 2.9 to 4.2 µm with a full width at half maximum (FWHM) of 29 nm. This demonstrates the great potential of using femtosecond-written VBGs as highly tunable, yet selective, spectral filters in the mid-IR.

Low temperature reaction kinetics inside an extended Laval nozzle: REMPI characterization and detection by broadband rotational spectroscopy.

Chirped-Pulse Fourier-Transform millimeter wave (CP-FTmmW) spectroscopy is a powerful method that enables detection of quantum state specific reactants and products in mixtures. We have successfully coupled this technique with a pulsed uniform Laval flow system to study photodissociation and reactions at low temperature, which we refer to as CPUF ("Chirped-Pulse/Uniform flow"). Detection by CPUF requires monitoring the free induction decay (FID) of the rotational coherence. However, the high collision frequency in high-density uniform supersonic flows can interfere with the FID and attenuate the signal. One way to overcome this is to sample the flow, but this can cause interference from shocks in the sampling region. This led us to develop an extended Laval nozzle that creates a uniform flow within the nozzle itself, after which the gas undergoes a shock-free secondary expansion to cold, low pressure conditions ideal for CP-FTmmW detection. Impact pressure measurements, commonly used to characterize Laval flows, cannot be used to monitor the flow within the nozzle. Therefore, we implemented a REMPI (resonance-enhanced multiphoton ionization) detection scheme that allows the interrogation of the conditions of the flow directly inside the extended nozzle, confirming the fluid dynamics simulations of the flow environment. We describe the development of the new 20 K extended flow, along with its characterization using REMPI and computational fluid dynamics. Finally, we demonstrate its application to the first low temperature measurement of the reaction kinetics of HCO with O2 and obtain a rate coefficient at 20 K of 6.66 ± 0.47 × 10-11 cm3 molec-1 s-1.

Synthesis of α-Chloroarylacetic Acid via Electrochemical Carboxylation of α,α-Dichloroarylmethane Derivatives.

The electrocarboxylation of α,α-dichloroarylmethane derivatives in the presence of CO2 was achieved, providing several α-chloroarylacetic acid derivatives with modest yields but high selectivity (chlorinated vs. non-chlorinated or dicarboxylic acid products). The obtained products were then involved in several chemical transformations, underlining their potential as versatile intermediates in synthetic chemistry. A mechanism was also proposed based upon a control experiment and cyclic voltammetry (CV) study.

The world around us and the picture(s) in our heads: The effects of news media use on belief organization.

Since Converse [1964. The nature of belief systems in mass publics. Critical Review, 18(1-3), 1 - 74 https://doi.org/10.1080/08913810608443650] asked "What goes with what?", research tries to answer this question. How individuals perceive the world around them depending on media use has been an endeavor of studying societal beliefs of societal issues separately. Building upon literature on cognitive architecture, we study how media use shapes the formation and stability of belief structures across issues in public opinion reflected in groups of individuals. Using a three-wave panel study, we found (1) that individuals' perceptions of different issues are interconnected, (2) translating into aggregate-stable, concurring groups in public opinion, and that (3) differential media use affects the formation and stability of these groups.

High-temperature hypersonic Laval nozzle for non-LTE cavity ringdown spectroscopy.

A small dimension Laval nozzle connected to a compact high enthalpy source equipped with cavity ringdown spectroscopy (CRDS) is used to produce vibrationally hot and rotationally cold high-resolution infrared spectra of polyatomic molecules in the 1.67 µm region. The Laval nozzle was machined in isostatic graphite, which is capable of withstanding high stagnation temperatures. It is characterized by a throat diameter of 2 mm and an exit diameter of 24 mm. It was designed to operate with argon heated up to 2000 K and to produce a quasi-unidirectional flow to reduce the Doppler effect responsible for line broadening. The hypersonic flow was characterized using computational fluid dynamics simulations, Pitot measurements, and CRDS. A Mach number evolving from 10 at the nozzle exit up to 18.3 before the occurrence of a first oblique shock wave was measured. Two different gases, carbon monoxide (CO) and methane (CH4), were used as test molecules. Vibrational (Tvib) and rotational (Trot) temperatures were extracted from the recorded infrared spectrum, leading to Tvib = 1346 ± 52 K and Trot = 12 ± 1 K for CO. A rotational temperature of 30 ± 3 K was measured for CH4, while two vibrational temperatures were necessary to reproduce the observed intensities. The population distribution between vibrational polyads was correctly described with Tvib I=894±47 K, while the population distribution within a given polyad (namely, the dyad or the pentad) was modeled correctly by Tvib II=54±4 K, testifying to a more rapid vibrational relaxation between the vibrational energy levels constituting a polyad.

Endodermal germ-layer formation through active actin-driven migration triggered by N-cadherin.

Germ-layer formation during gastrulation is both a fundamental step of development and a paradigm for tissue formation and remodeling. However, the cellular and molecular basis of germ-layer segregation is poorly understood, mostly because of the lack of direct in vivo observations. We used mosaic zebrafish embryos to investigate the formation of the endoderm. High-resolution live imaging and functional analyses revealed that endodermal cells reach their characteristic innermost position through an active, oriented, and actin-based migration dependent on Rac1, which contrasts with the previously proposed differential adhesion cell sorting. Rather than being attracted to their destination, the yolk syncytial layer, cells appear to migrate away from their neighbors. This migration depends on N-cadherin that, when imposed in ectodermal cells, is sufficient to trigger their internalization without affecting their fate. Overall, these results lead to a model of germ-layer formation in which, upon N-cadherin expression, endodermal cells actively migrate away from their epiblastic neighbors to reach their internal position, revealing cell-contact avoidance as an unexplored mechanism driving germ-layer formation.

Inhibitory signalling to the Arp2/3 complex steers cell migration.

Cell migration requires the generation of branched actin networks that power the protrusion of the plasma membrane in lamellipodia. The actin-related proteins 2 and 3 (Arp2/3) complex is the molecular machine that nucleates these branched actin networks. This machine is activated at the leading edge of migrating cells by Wiskott-Aldrich syndrome protein (WASP)-family verprolin-homologous protein (WAVE, also known as SCAR). The WAVE complex is itself directly activated by the small GTPase Rac, which induces lamellipodia. However, how cells regulate the directionality of migration is poorly understood. Here we identify a new protein, Arpin, that inhibits the Arp2/3 complex in vitro, and show that Rac signalling recruits and activates Arpin at the lamellipodial tip, like WAVE. Consistently, after depletion of the inhibitory Arpin, lamellipodia protrude faster and cells migrate faster. A major role of this inhibitory circuit, however, is to control directional persistence of migration. Indeed, Arpin depletion in both mammalian cells and Dictyostelium discoideum amoeba resulted in straighter trajectories, whereas Arpin microinjection in fish keratocytes, one of the most persistent systems of cell migration, induced these cells to turn. The coexistence of the Rac-Arpin-Arp2/3 inhibitory circuit with the Rac-WAVE-Arp2/3 activatory circuit can account for this conserved role of Arpin in steering cell migration.

Isomer-specific detection in the UV photodissociation of the propargyl radical by chirped-pulse mm-wave spectroscopy in a pulsed quasi-uniform flow.

Isomer-specific detection and product branching fractions in the UV photodissociation of the propargyl radical is achieved through the use of chirped-pulse Fourier-transform mm-wave spectroscopy in a pulsed quasi-uniform flow (CPUF). Propargyl radicals are produced in the 193 nm photodissociation of 1,2-butadiene. Absorption of a second photon leads to H atom elimination giving three possible C3H2 isomers: singlets cyclopropenylidene (c-C3H2) and propadienylidene (l-C3H2), and triplet propargylene (3HCCCH). The singlet products and their appearance kinetics in the flow are directly determined by rotational spectroscopy, but due to the negligible dipole moment of propargylene, it is not directly monitored. However, we exploit the time-dependent kinetics of H-atom catalyzed isomerization to infer the branching to propargylene as well. We obtain the overall branching among H loss channels to be 2.9% (+1.1/-0.5) l-C3H2 + H, 16.8% (+3.2/-1.3) c-C3H2 + H, and 80.2 (+1.8/-4.2) 3HCCCH + H. Our findings are qualitatively consistent with earlier RRKM calculations in that the major channel in the photodissociation of the propargyl radical at 193 nm is to 3HCCCH + H; however, a greater contribution to the energetically most favorable isomer, c-C3H2 + H is observed in this work. We do not detect the predicted HCCC + H2 channel, but place an upper bound on its yield of 1%.

Collective mesendoderm migration relies on an intrinsic directionality signal transmitted through cell contacts.

Collective cell migration is key to morphogenesis, wound healing, or cancer cell migration. However, its cellular bases are just starting to be unraveled. During vertebrate gastrulation, axial mesendoderm migrates in a group, the prechordal plate, from the embryonic organizer to the animal pole. How this collective migration is achieved remains unclear. Previous work has suggested that cells migrate as individuals, with collective movement resulting from the addition of similar individual cell behavior. Through extensive analyses of cell trajectories, morphologies, and polarization in zebrafish embryos, we reveal that all prechordal plate cells show the same behavior and rely on the same signaling pathway to migrate, as expected if they do so individually. However, by using cell transplants, we demonstrate that prechordal plate migration is a true collective process, as isolated cells do not migrate toward the animal pole. They are still polarized and motile but lose directionality. Directionality is restored upon contact with the endogenous prechordal plate. This contact dependent orientation relies on E-cadherin, Wnt-PCP signaling, and Rac1. Importantly, groups of cells also need contact with the endogenous plate to orient correctly, showing an instructive role of the plate in establishing directionality. Overall, our results lead to an original model of collective migration in which directional information is contained within the moving group rather than provided by extrinsic cues, and constantly maintained in cells by contacts with their neighbors. This self-organizing model could account for collective invasion of new territories, as observed in cancer strands, without requirement for any attractant in the colonized tissue.

[Collective cell migrations]. / Migrations cellulaires collectives.

Historically centered on the study of individual cell motility, the field of cell migration has recently moved up one level to look at cooperative behaviour within migratory cell populations. It is now well established that numerous physiological and pathological migration events involve collectively migrating cells rather than solitary cells or concomitantly migrating individual cells. In this review, we first discuss the criteria allowing a given migratory event to be classified as collective cell migration. We then summarize the main concepts that rule collective cell migration in epithelial and mesenchymal tissues with a main focus on mechanisms controlling polarity and directionality in cell collectives.

Reliability Investigation of Silicide-Based Thermoelectric Modules.

The reliability and failure mechanisms of silicide-based thermoelectric modules (n-type Mg2(Si,Sn)/p-type HMS) were investigated thanks to two types of thermal tests with either a fixed or a cycling thermal gradient, under different atmospheres. The hot interfaces of the thermoelectric modules were analyzed by scanning electron microscopy and X-ray diffraction after the reliability tests. The current thermoelectric modules do not exhibit any failure mechanism under ambient air for a hot side temperature of 250 °C for tests conducted either during 500 h at a fixed temperature gradient or after 1000 thermal cycles. However, when the temperature was increased to 350 °C, pesting phenomena were detected at the hot side of the n-type Mg2(Si,Sn) legs caused by the decomposition/oxidation of the material. These phenomena are strongly slowed down for thermoelectric modules tested under a primary vacuum, highlighting the predominant role of oxygen in the degradation mechanism. Interdiffusion phenomena are the most pronounced at the interface of the hot side of the n-type thermoelectric materials. The formation of a MgO layer, which is an electrical and thermal insulator, has decreased the thermoelectric modules' performances. For thermal cycling tests, cracks are observed on the hot side of the n-type legs. The presence of these cracks drastically increases the thermal and electrical resistances, leading to an overheating of the system and limiting its efficiency and failure by breaking electrical continuity. The interfaces at the hot side temperature of the p-type HMS legs remained intact whatever the test conditions were, indicating a high chemical stability and a good mechanical strength.

Necrosulfonamide causes oxidation of PCM1 and impairs ciliogenesis and autophagy.

Centriolar satellites are high-order assemblies, scaffolded by the protein PCM1, that gravitate as particles around the centrosome and play pivotal roles in fundamental cellular processes notably ciliogenesis and autophagy. Despite stringent control mechanisms involving phosphorylation and ubiquitination, the landscape of post-translational modifications shaping these structures remains elusive. Here, we report that necrosulfonamide (NSA), a small molecule known for binding and inactivating the pivotal effector of cell death by necroptosis MLKL, intersects with centriolar satellites, ciliogenesis, and autophagy independently of MLKL. NSA functions as a potent redox cycler and triggers the oxidation and aggregation of PCM1 alongside select partners, while minimally impacting the overall distribution of centriolar satellites. Additionally, NSA-mediated ROS production disrupts ciliogenesis and leads to the accumulation of autophagy markers, partially alleviated by PCM1 deletion. Together, these results identify PCM1 as a redox sensor protein and provide new insights into the interplay between centriolar satellites and autophagy.

Ultradifferentiable classes of entire functions.

We study classes of ultradifferentiable functions defined in terms of small weight sequences violating standard growth and regularity requirements. First, we show that such classes can be viewed as weighted spaces of entire functions for which the crucial weight is given by the associated weight function of the so-called conjugate weight sequence. Moreover, we generalize results from M. Markin from the so-called small Gevrey setting to arbitrary convenient families of (small) sequences and show how the corresponding ultradifferentiable function classes can be used to detect boundedness of normal linear operators on Hilbert spaces (associated with an evolution equation problem). Finally, we study the connection between small sequences and the recent notion of dual sequences introduced in the Ph.D. thesis of J. Jiménez-Garrido.

Thermal Stability of Mg2Si0.6Sn0.4 under Oxidation Conditions.

The use of Mg2Si0.6Sn0.4 under air in thermoelectric modules in the mid-temperature range of 400-600 °C is linked to its ability to resist oxidation. In this study, oxidation experiments performed at 400 °C under air evidenced the stability of the material, either under static conditions (up to 100 h) or under severe heating-cooling cyclic conditions (up to 400 cycles), showing its ability to be used in a reliable way at this temperature. By combining thermogravimetry, scanning electron microscopy, temperature X-ray diffraction analysis, and mechanical and thermodynamic considerations, a mechanism is proposed explaining how Mg2Si0.6Sn0.4 further undergoes decomposition with time under air when treated above 500 °C. The presence of Sn and the formation of various oxides are the key parameters of the material's degradation.

Chromatically Corrected Multicolor Multiphoton Microscopy.

Simultaneous imaging of multiple labels in tissues is key to studying complex biological processes. Although strategies for color multiphoton excitation have been established, chromatic aberration remains a major problem when multiple excitation wavelengths are used in a scanning microscope. Chromatic aberration introduces a spatial shift between the foci of beams of different wavelengths that varies across the field of view, severely degrading the performance of color imaging. In this work, we propose an adaptive correction strategy that solves this problem in two-beam microscopy techniques. Axial chromatic aberration is corrected by a refractive phase mask that introduces pure defocus into one beam, while lateral chromatic aberration is corrected by a piezoelectric mirror that dynamically compensates for lateral shifts during scanning. We show that this light-efficient approach allows seamless chromatic correction over the entire field of view of different multiphoton objectives without compromising spatial and temporal resolution and that the effective area for beam-mixing processes can be increased by more than 1 order of magnitude. We illustrate this approach with simultaneous three-color, two-photon imaging of developing zebrafish embryos and fixed Brainbow mouse brain slices over large areas. These results establish a robust and efficient method for chromatically corrected multiphoton imaging.

Label-free imaging of red blood cells and oxygenation with color third-order sum-frequency generation microscopy.

Mapping red blood cells (RBCs) flow and oxygenation is of key importance for analyzing brain and tissue physiology. Current microscopy methods are limited either in sensitivity or in spatio-temporal resolution. In this work, we introduce a novel approach based on label-free third-order sum-frequency generation (TSFG) and third-harmonic generation (THG) contrasts. First, we propose a novel experimental scheme for color TSFG microscopy, which provides simultaneous measurements at several wavelengths encompassing the Soret absorption band of hemoglobin. We show that there is a strong three-photon (3P) resonance related to the Soret band of hemoglobin in THG and TSFG signals from zebrafish and human RBCs, and that this resonance is sensitive to RBC oxygenation state. We demonstrate that our color TSFG implementation enables specific detection of flowing RBCs in zebrafish embryos and is sensitive to RBC oxygenation dynamics with single-cell resolution and microsecond pixel times. Moreover, it can be implemented on a 3P microscope and provides label-free RBC-specific contrast at depths exceeding 600 µm in live adult zebrafish brain. Our results establish a new multiphoton contrast extending the palette of deep-tissue microscopy.

MiniBAR/GARRE1 is a dual Rac and Rab effector required for ciliogenesis.

Cilia protrude from the cell surface and play critical roles in intracellular signaling, environmental sensing, and development. Reduced actin-dependent contractility and intracellular trafficking are both required for ciliogenesis, but little is known about how these processes are coordinated. Here, we identified a Rac1- and Rab35-binding protein with a truncated BAR (Bin/amphiphysin/Rvs) domain that we named MiniBAR (also known as KIAA0355/GARRE1), which plays a key role in ciliogenesis. MiniBAR colocalizes with Rac1 and Rab35 at the plasma membrane and on intracellular vesicles trafficking to the ciliary base and exhibits fast pulses at the ciliary membrane. MiniBAR depletion leads to short cilia, resulting from abnormal Rac-GTP/Rho-GTP levels and increased acto-myosin-II-dependent contractility together with defective trafficking of IFT88 and ARL13B into cilia. MiniBAR-depleted zebrafish embryos display dysfunctional short cilia and hallmarks of ciliopathies, including left-right asymmetry defects. Thus, MiniBAR is a dual Rac and Rab effector that controls both actin cytoskeleton and membrane trafficking for ciliogenesis.

Drosophila Ric-8 regulates Galphai cortical localization to promote Galphai-dependent planar orientation of the mitotic spindle during asymmetric cell division.

Localization and activation of heterotrimeric G proteins have a crucial role during asymmetric cell division. The asymmetric division of the Drosophila sensory precursor cell (pl) is polarized along the antero-posterior axis by Frizzled signalling and, during this division, activation of Galphai depends on Partner of Inscuteable (Pins). We establish here that Ric-8, which belongs to a family of guanine nucleotide-exchange factors for Galphai, regulates cortical localization of the subunits Galphai and Gbeta13F. Ric-8, Galphai and Pins are not necessary for the control of the anteroposterior orientation of the mitotic spindle during pl cell division downstream of Frizzled signalling, but they are required for maintainance of the spindle within the plane of the epithelium. On the contrary, Frizzled signalling orients the spindle along the antero-posterior axis but also tilts it along the apico-basal axis. Thus, Frizzled and heterotrimeric G-protein signalling act in opposition to ensure that the spindle aligns both in the plane of the epithelium and along the tissue polarity axis.