Friction Mediates Scission of Tubular Membranes Scaffolded by BAR Proteins.

Membrane scission is essential for intracellular trafficking. While BAR domain proteins such as endophilin have been reported in dynamin-independent scission of tubular membrane necks, the cutting mechanism has yet to be deciphered. Here, we combine a theoretical model, in vitro, and in vivo experiments revealing how protein scaffolds may cut tubular membranes. We demonstrate that the protein scaffold bound to the underlying tube creates a frictional barrier for lipid diffusion; tube elongation thus builds local membrane tension until the membrane undergoes scission through lysis. We call this mechanism friction-driven scission (FDS). In cells, motors pull tubes, particularly during endocytosis. Through reconstitution, we show that motors not only can pull out and extend protein-scaffolded tubes but also can cut them by FDS. FDS is generic, operating even in the absence of amphipathic helices in the BAR domain, and could in principle apply to any high-friction protein and membrane assembly.

Chromatin Remodeling Due to Transient-Link-and-Pass Activity Enhances Subnuclear Dynamics.

Spatiotemporal coordination of chromatin and subnuclear compartments is crucial for cells. Numerous enzymes act inside nucleus-some of those transiently link and pass two chromatin segments. Here, we study how such an active perturbation affects fluctuating dynamics of an inclusion in the chromatic medium. Using numerical simulations and a versatile effective model, we categorize inclusion dynamics into three distinct modes. The transient-link-and-pass activity speeds up inclusion dynamics by affecting a slow mode related to chromatin remodeling, viz., size and shape of the chromatin meshes.

Membrane Tilt Drives Phase Separation of Adhesion Receptors.

Cell adhesion receptors are transmembrane proteins that bind cells to their environment. These proteins typically cluster into disk-shaped or linear structures. Here, we show that such clustering patterns spontaneously emerge when the receptor senses the membrane deformation gradient, for example, by reaching a lower-energy conformation when the membrane is tilted relative to the underlying binding substrate. Increasing the strength of the membrane gradient-sensing mechanism first yields isolated disk-shaped clusters and then long linear structures. Our theory is coherent with experimental estimates of the parameters, suggesting that a tilt-induced clustering mechanism is relevant in the context of cell adhesion.

Quantitative comparison of cell-cell detachment force in different experimental setups.

We compare three different setups for measuring cell-cell adhesion. We show that the measured strength depends on the type of setup that is used. For identical cells different assays measure different detachment forces. This can be understood from the fact that cell-cell detachment is a global property of the system. We also analyse the role of external force and line tension on contact angle and cell-cell detachment. Comparison with the experiments suggest that viscous forces play an important role in the process. We dedicate this article to Fyl Pincus who for many of us is an example to be followed not only for outstanding science but also for a marvelous human behavior.

Hydraulic and electric control of cell spheroids.

We use a theoretical approach to examine the effect of a radial fluid flow or electric current on the growth and homeostasis of a cell spheroid. Such conditions may be generated by a drain of micrometric diameter. To perform this analysis, we describe the tissue as a continuum. We include active mechanical, electric, and hydraulic components in the tissue material properties. We consider a spherical geometry and study the effect of the drain on the dynamics of the cell aggregate. We show that a steady fluid flow or electric current imposed by the drain could be able to significantly change the spheroid long-time state. In particular, our work suggests that a growing spheroid can systematically be driven to a shrinking state if an appropriate external field is applied. Order-of-magnitude estimates suggest that such fields are of the order of the indigenous ones. Similarities and differences with the case of tumors and embryo development are briefly discussed.

Active Nematic Flows over Curved Surfaces.

Cell monolayers are a central model system in the study of tissue biophysics. In vivo, epithelial tissues are curved on the scale of microns, and the curvature's role in the onset of spontaneous tissue flows is still not well understood. Here, we present a hydrodynamic theory for an apical-basal asymmetric active nematic gel on a curved strip. We show that surface curvature qualitatively changes monolayer motion compared with flat space: the resulting flows can be thresholdless, and the transition to motion may change from continuous to discontinuous. Surface curvature, friction, and active tractions are all shown to control the flow pattern selected, from simple shear to vortex chains.

Fluid pumping and active flexoelectricity can promote lumen nucleation in cell assemblies.

We discuss the physical mechanisms that promote or suppress the nucleation of a fluid-filled lumen inside a cell assembly or a tissue. We discuss lumen formation in a continuum theory of tissue material properties in which the tissue is described as a 2-fluid system to account for its permeation by the interstitial fluid, and we include fluid pumping as well as active electric effects. Considering a spherical geometry and a polarized tissue, our work shows that fluid pumping and tissue flexoelectricity play a crucial role in lumen formation. We furthermore explore the large variety of long-time states that are accessible for the cell aggregate and its lumen. Our work reveals a role of the coupling of mechanical, electrical, and hydraulic phenomena in tissue lumen formation.

Physics of lumen growth.

We model the dynamics of formation of intercellular secretory lumens. Using conservation laws, we quantitatively study the balance between paracellular leaks and the build-up of osmotic pressure in the lumen. Our model predicts a critical pumping threshold to expand stable lumens. Consistently with experimental observations in bile canaliculi, the model also describes a transition between a monotonous and oscillatory regime during luminogenesis as a function of ion and water transport parameters. We finally discuss the possible importance of regulation of paracellular leaks in intercellular tubulogenesis.

Liquid Phase Separation Controlled by pH.

We present a minimal model to study the effects of pH on liquid phase separation of macromolecules. Our model describes a mixture composed of water and macromolecules that exist in three different charge states and have a tendency to phase separate. This phase separation is affected by pH via a set of chemical reactions describing protonation and deprotonation of macromolecules, as well as self-ionization of water. We consider the simple case in which interactions are captured by Flory-Huggins interaction parameters corresponding to Debye screening lengths shorter than a nanometer, which is relevant to proteins inside biological cells under physiological conditions. We identify the conjugate thermodynamic variables at chemical equilibrium and discuss the effective free energy at fixed pH. First, we study phase diagrams as a function of macromolecule concentration and temperature at the isoelectric point of the macromolecules. We find a rich variety of phase diagram topologies, including multiple critical points, triple points, and first-order transition points. Second, we change the pH relative to the isoelectric point of the macromolecules and study how phase diagrams depend on pH. We find that these phase diagrams as a function of pH strongly depend on whether oppositely charged macromolecules or neutral macromolecules have a stronger tendency to phase separate. One key finding is that we predict the existence of a reentrant behavior as a function of pH. In addition, our model predicts that the region of phase separation is typically broader at the isoelectric point. This model could account for both in vitro phase separation of proteins as a function of pH and protein phase separation in yeast cells for pH values close to the isoelectric point of many cytosolic proteins.

The Roles of Microtubules and Membrane Tension in Axonal Beading, Retraction, and Atrophy.

Axonal beading, or the formation of a series of swellings along the axon, and retraction are commonly observed shape transformations that precede axonal atrophy in Alzheimer's disease, Parkinson's disease, and other neurodegenerative conditions. The mechanisms driving these morphological transformations are poorly understood. Here, we report controlled experiments that can induce either beading or retraction and follow the time evolution of these responses. By making quantitative analysis of the shape modes under different conditions, measurement of membrane tension, and using theoretical considerations, we argue that membrane tension is the main driving force that pushes cytosol out of the axon when microtubules are degraded, causing axonal thinning. Under pharmacological perturbation, atrophy is always retrograde, and this is set by a gradient in the microtubule stability. The nature of microtubule depolymerization dictates the type of shape transformation, vis-à-vis beading or retraction. Elucidating the mechanisms of these shape transformations may facilitate development of strategies to prevent or arrest axonal atrophy due to neurodegenerative conditions.

Theory of epithelial sheet morphology in three dimensions.

Morphogenesis during embryo development requires the coordination of mechanical forces to generate the macroscopic shapes of organs. We propose a minimal theoretical model, based on cell adhesion and actomyosin contractility, which describes the various shapes of epithelial cells and the bending and buckling of epithelial sheets, as well as the relative stability of cellular tubes and spheres. We show that, to understand these processes, a full 3D description of the cells is needed, but that simple scaling laws can still be derived. The morphologies observed in vivo can be understood as stable points of mechanical equations and the transitions between them are either continuous or discontinuous. We then focus on epithelial sheet bending, a ubiquitous morphogenetic process. We calculate the curvature of an epithelium as a function of actin belt tension as well as of cell-cell and and cell-substrate tension. The model allows for a comparison of the relative stabilities of spherical or cylindrical cellular structures (acini or tubes). Finally, we propose a unique type of buckling instability of epithelia, driven by a flattening of individual cell shapes, and discuss experimental tests to verify our predictions.

Dynamics of membrane tethers reveal novel aspects of cytoskeleton-membrane interactions in axons.

Mechanical properties of cell membranes are known to be significantly influenced by the underlying cortical cytoskeleton. The technique of pulling membrane tethers from cells is one of the most effective ways of studying the membrane mechanics and the membrane-cortex interaction. In this article, we show that axon membranes make an interesting system to explore as they exhibit both free membrane-like behavior where the tether-membrane junction is movable on the surface of the axons (unlike many other cell membranes) as well as cell-like behavior where there are transient and spontaneous eruptions in the tether force that vanish when F-actin is depolymerized. We analyze the passive and spontaneous responses of axonal membrane tethers and propose theoretical models to explain the observed behavior.

Homeostatic Fluctuations of a Tissue Surface.

We study the surface fluctuations of a tissue with a dynamics dictated by cell-rearrangement, cell-division, and cell-death processes. Surface fluctuations are calculated in the homeostatic state, where cell division and cell death equilibrate on average. The obtained fluctuation spectrum can be mapped onto several other spectra such as those characterizing incompressible fluids, compressible Maxwell elastomers, or permeable membranes in appropriate asymptotic regimes. Since cell division and cell death are out-of-equilibrium processes, detailed balance is broken, but a generalized fluctuation-response relation is satisfied in terms of appropriate observables. Our work is a first step toward the description of the out-of-equilibrium fluctuations of the surface of a thick epithelium and its dynamical response to external perturbations.

Theoretical study of actin layers attachment and separation.

We use the theory of active gels to study theoretically the merging and separation of two actin dense layers akin to cortical layers of animal cells. The layers bind at a distance equal to twice the thickness of a free layer, thus forming a single dense layer, similar in this sense to a lamellipodium. When that unique layer is stretched apart, it is resilient to break apart up to a critical length larger than twice the thickness of a free layer. We show that this behavior can result from the high contractile properties of the actomyosin gel due to the activity of myosin molecular motors. Furthermore, we establish that the stability of the stretched single layer is highly dependent on the properties of the gel. Indeed, the nematic order of the actin filaments along the polymerizing membranes is a destabilizing factor.

Nature of curvature coupling of amphiphysin with membranes depends on its bound density.

Cells are populated by a vast array of membrane-binding proteins that execute critical functions. Functions, like signaling and intracellular transport, require the abilities to bind to highly curved membranes and to trigger membrane deformation. Among these proteins is amphiphysin 1, implicated in clathrin-mediated endocytosis. It contains a Bin-Amphiphysin-Rvs membrane-binding domain with an N-terminal amphipathic helix that senses and generates membrane curvature. However, an understanding of the parameters distinguishing these two functions is missing. By pulling a highly curved nanotube of controlled radius from a giant vesicle in a solution containing amphiphysin, we observed that the action of the protein depends directly on its density on the membrane. At low densities of protein on the nearly flat vesicle, the distribution of proteins and the mechanical effects induced are described by a model based on spontaneous curvature induction. The tube radius and force are modified by protein binding but still depend on membrane tension. In the dilute limit, when practically no proteins were present on the vesicle, no mechanical effects were detected, but strong protein enrichment proportional to curvature was seen on the tube. At high densities, the radius is independent of tension and vesicle protein density, resulting from the formation of a scaffold around the tube. As a consequence, the scaling of the force with tension is modified. For the entire density range, protein was enriched on the tube as compared to the vesicle. Our approach shows that the strength of curvature sensing and mechanical effects on the tube depends on the protein density.

Furrow constriction in animal cell cytokinesis.

Cytokinesis is the process of physical cleavage at the end of cell division; it proceeds by ingression of an acto-myosin furrow at the equator of the cell. Its failure leads to multinucleated cells and is a possible cause of tumorigenesis. Here, we calculate the full dynamics of furrow ingression and predict cytokinesis completion above a well-defined threshold of equatorial contractility. The cortical acto-myosin is identified as the main source of mechanical dissipation and active forces. Thereupon, we propose a viscous active nonlinear membrane theory of the cortex that explicitly includes actin turnover and where the active RhoA signal leads to an equatorial band of myosin overactivity. The resulting cortex deformation is calculated numerically, and reproduces well the features of cytokinesis such as cell shape and cortical flows toward the equator. Our theory gives a physical explanation of the independence of cytokinesis duration on cell size in embryos. It also predicts a critical role of turnover on the rate and success of furrow constriction. Scaling arguments allow for a simple interpretation of the numerical results and unveil the key mechanism that generates the threshold for cytokinesis completion: cytoplasmic incompressibility results in a competition between the furrow line tension and the cell poles' surface tension.

Compressive stress inhibits proliferation in tumor spheroids through a volume limitation.

In most instances, the growth of solid tumors occurs in constrained environments and requires a competition for space. A mechanical crosstalk can arise from this competition. In this article, we dissect the biomechanical sequence caused by a controlled compressive stress on multicellular spheroids (MCSs) used as a tumor model system. On timescales of minutes, we show that a compressive stress causes a reduction of the MCS volume, linked to a reduction of the cell volume in the core of the MCS. On timescales of hours, we observe a reversible induction of the proliferation inhibitor, p27Kip1, from the center to the periphery of the spheroid. On timescales of days, we observe that cells are blocked in the cell cycle at the late G1 checkpoint, the restriction point. We show that the effect of pressure on the proliferation can be antagonized by silencing p27Kip1. Finally, we quantify a clear correlation between the pressure-induced volume change and the growth rate of the spheroid. The compression-induced proliferation arrest that we studied is conserved for five cell lines, and is completely reversible. It demonstrates a generic crosstalk between mechanical stresses and the key players of cell cycle regulation. Our results suggest a role of volume change in the sensitivity to pressure, and that p27Kip1 is strongly influenced by this change.

Curvature-induced clustering of cell adhesion proteins.

Cell adhesion proteins typically form stable clusters that anchor the cell membrane to its environment. Several works have suggested that cell membrane protein clusters can emerge from a local feedback between the membrane curvature and the density of proteins. Here, we investigate the effect of such a curvature-sensing mechanism in the context of cell adhesion proteins. We show how clustering emerges in an intermediate range of adhesion and curvature-sensing strengths. We identify key differences with the tilt-induced gradient sensing mechanism we previously proposed (Lin et al., arXiv:2307.03670).

Mechanical control of cell flow in multicellular spheroids.

Collective cell motion is observed in a wide range of biological processes. In tumors, physiological gradients of nutrients, growth factors, or even oxygen give rise to gradients of proliferation. We show using fluorescently labeled particles that these gradients drive a velocity field resulting in a cellular flow in multicellular spheroids. Under mechanical stress, the cellular flow is drastically reduced. We describe the results with a hydrodynamic model that considers only convection of the particles by the cellular flow.

Fluidization of tissues by cell division and apoptosis.

During the formation of tissues, cells organize collectively by cell division and apoptosis. The multicellular dynamics of such systems is influenced by mechanical conditions and can give rise to cell rearrangements and movements. We develop a continuum description of tissue dynamics, which describes the stress distribution and the cell flow field on large scales. In the absence of division and apoptosis, we consider the tissue to behave as an elastic solid. Cell division and apoptosis introduce stress sources that, in general, are anisotropic. By combining cell number balance with dynamic equations for the stress source, we show that the tissue effectively behaves as a viscoelastic fluid with a relaxation time set by the rates of division and apoptosis. If the system is confined in a fixed volume, it reaches a homeostatic state in which division and apoptosis balance. In this state, cells undergo a diffusive random motion driven by the stochasticity of division and apoptosis. We calculate the expression for the effective diffusion coefficient as a function of the tissue parameters and compare our results concerning both diffusion and viscosity to simulations of multicellular systems using dissipative particle dynamics.