A mechanosensing mechanism controls plasma membrane shape homeostasis at the nanoscale.

As cells migrate and experience forces from their surroundings, they constantly undergo mechanical deformations which reshape their plasma membrane (PM). To maintain homeostasis, cells need to detect and restore such changes, not only in terms of overall PM area and tension as previously described, but also in terms of local, nanoscale topography. Here, we describe a novel phenomenon, by which cells sense and restore mechanically induced PM nanoscale deformations. We show that cell stretch and subsequent compression reshape the PM in a way that generates local membrane evaginations in the 100 nm scale. These evaginations are recognized by I-BAR proteins, which triggers a burst of actin polymerization mediated by Rac1 and Arp2/3. The actin polymerization burst subsequently re-flattens the evagination, completing the mechanochemical feedback loop. Our results demonstrate a new mechanosensing mechanism for PM shape homeostasis, with potential applicability in different physiological scenarios.

Estimation of anisotropic bending rigidities and spontaneous curvatures of crescent curvature-inducing proteins from tethered-vesicle experimental data.

The Bin/amphiphysin/Rvs (BAR) superfamily proteins have a crescent binding domain and bend biomembranes along the domain axis. However, their anisotropic bending rigidities and spontaneous curvatures have not been experimentally determined. Here, we estimated these values from the bound protein densities on tethered vesicles using a mean-field theory of anisotropic bending energy and orientation-dependent excluded volume. The dependence curves of the protein density on the membrane curvature are fitted to the experimental data for the I-BAR and N-BAR domains reported by C. Prévost et al. Nat. Commun., 2015, 6, 8529 and F.-C. Tsai et al. Soft Matter, 2021, 17, 4254-4265, respectively. For the I-BAR domain, all three density curves of different chemical potentials exhibit excellent fits with a single parameter set of anisotropic bending energy. When the classical isotropic bending energy is used instead, one of the curves can be fitted well, but the others exhibit large deviations. In contrast, for the N-BAR domain, two curves are not well fitted simultaneously the anisotropic model, although it is significantly improved compared to the isotropic model. This deviation likely suggests a cluster formation of the N-BAR domains.

Caveolin-1 dolines form a distinct and rapid caveolae-independent mechanoadaptation system.

In response to different types and intensities of mechanical force, cells modulate their physical properties and adapt their plasma membrane (PM). Caveolae are PM nano-invaginations that contribute to mechanoadaptation, buffering tension changes. However, whether core caveolar proteins contribute to PM tension accommodation independently from the caveolar assembly is unknown. Here we provide experimental and computational evidence supporting that caveolin-1 confers deformability and mechanoprotection independently from caveolae, through modulation of PM curvature. Freeze-fracture electron microscopy reveals that caveolin-1 stabilizes non-caveolar invaginations-dolines-capable of responding to low-medium mechanical forces, impacting downstream mechanotransduction and conferring mechanoprotection to cells devoid of caveolae. Upon cavin-1/PTRF binding, doline size is restricted and membrane buffering is limited to relatively high forces, capable of flattening caveolae. Thus, caveolae and dolines constitute two distinct albeit complementary components of a buffering system that allows cells to adapt efficiently to a broad range of mechanical stimuli.

Dynamic mechanochemical feedback between curved membranes and BAR protein self-organization.

In many physiological situations, BAR proteins reshape membranes with pre-existing curvature (templates), contributing to essential cellular processes. However, the mechanism and the biological implications of this reshaping process remain unclear. Here we show, both experimentally and through modelling, that BAR proteins reshape low curvature membrane templates through a mechanochemical phase transition. This phenomenon depends on initial template shape and involves the co-existence and progressive transition between distinct local states in terms of molecular organization (protein arrangement and density) and membrane shape (template size and spherical versus cylindrical curvature). Further, we demonstrate in cells that this phenomenon enables a mechanotransduction mode, in which cellular stretch leads to the mechanical formation of membrane templates, which are then reshaped into tubules by BAR proteins. Our results demonstrate the interplay between membrane mechanics and BAR protein molecular organization, integrating curvature sensing and generation in a comprehensive framework with implications for cell mechanical responses.

A theory of ordering of elongated and curved proteins on membranes driven by density and curvature.

Cell membranes interact with a myriad of curvature-active proteins that control membrane morphology and are responsible for mechanosensation and mechanotransduction. Some of these proteins, such as those containing BAR domains, are curved and elongated, and hence may adopt different states of orientational order, from isotropic to maximize entropy to nematic as a result of crowding or to adapt to the curvature of the underlying membrane. Here, extending the classical work of Onsager for ordering in hard particle systems and that of [E. S. Nascimento et al., Phys. Rev. E, 2017, 96, 022704], we develop a mean-field density functional theory to predict the orientational order and evaluate the free energy of ensembles of elongated and curved objects on curved membranes. This theory depends on the microscopic properties of the particles and explains how a density-dependent isotropic-to-nematic transition is modified by anisotropic curvature. We also examine the coexistence of isotropic and nematic phases. This theory predicts how ordering depends on geometry but we assume here that the geometry is fixed. It also lays the ground to understand the interplay between membrane reshaping by BAR proteins and molecular order, examined by [Le Roux et al., submitted, 2020].

A physical mechanism of TANGO1-mediated bulky cargo export.

The endoplasmic reticulum (ER)-resident protein TANGO1 assembles into a ring around ER exit sites (ERES), and links procollagens in the ER lumen to COPII machinery, tethers, and ER-Golgi intermediate compartment (ERGIC) in the cytoplasm (Raote et al., 2018). Here, we present a theoretical approach to investigate the physical mechanisms of TANGO1 ring assembly and how COPII polymerization, membrane tension, and force facilitate the formation of a transport intermediate for procollagen export. Our results indicate that a TANGO1 ring, by acting as a linactant, stabilizes the open neck of a nascent COPII bud. Elongation of such a bud into a transport intermediate commensurate with bulky procollagens is then facilitated by two complementary mechanisms: (i) by relieving membrane tension, possibly by TANGO1-mediated fusion of retrograde ERGIC membranes and (ii) by force application. Altogether, our theoretical approach identifies key biophysical events in TANGO1-driven procollagen export.

The plasma membrane as a mechanochemical transducer.

Cells are constantly submitted to external mechanical stresses, which they must withstand and respond to. By forming a physical boundary between cells and their environment that is also a biochemical platform, the plasma membrane (PM) is a key interface mediating both cellular response to mechanical stimuli, and subsequent biochemical responses. Here, we review the role of the PM as a mechanosensing structure. We first analyse how the PM responds to mechanical stresses, and then discuss how this mechanical response triggers downstream biochemical responses. The molecular players involved in PM mechanochemical transduction include sensors of membrane unfolding, membrane tension, membrane curvature or membrane domain rearrangement. These sensors trigger signalling cascades fundamental both in healthy scenarios and in diseases such as cancer, which cells harness to maintain integrity, keep or restore homeostasis and adapt to their external environment. This article is part of a discussion meeting issue 'Forces in cancer: interdisciplinary approaches in tumour mechanobiology'.

Clathrin polymerization exhibits high mechano-geometric sensitivity.

How tension modulates cellular transport has become a topic of interest in the recent past. However, the effect of tension on clathrin assembly and vesicle growth remains less understood. Here, we use the classical Helfrich theory to predict the energetic cost that clathrin is required to pay to remodel the membrane at different stages of vesicle formation. Our study reveals that this energetic cost is highly sensitive to not only the tension in the membrane but also to the instantaneous geometry of the membrane during shape evolution. Our study predicts a sharp reduction in clathrin coat size in the intermediate tension regime (0.01-0.1 mN m-1). Remarkably, the natural propensity of the membrane to undergo bending beyond the Ω shape causes a significant decrease in the energy needed from clathrin to drive vesicle growth. Our studies in mammalian cells confirm a reduction in clathrin coat size in an increased tension environment. In addition, our findings suggest that the two apparently distinct clathrin assembly modes, namely coated pits and coated plaques, observed in experimental investigations might be a consequence of varied tensions in the plasma membrane. Overall, the mechano-geometric sensitivity revealed in this study might also be at play during the polymerization of other membrane remodeling proteins.

Endocytic proteins drive vesicle growth via instability in high membrane tension environment.

Clathrin-mediated endocytosis (CME) is a key pathway for transporting cargo into cells via membrane vesicles; it plays an integral role in nutrient import, signal transduction, neurotransmission, and cellular entry of pathogens and drug-carrying nanoparticles. Because CME entails substantial local remodeling of the plasma membrane, the presence of membrane tension offers resistance to bending and hence, vesicle formation. Experiments show that in such high-tension conditions, actin dynamics is required to carry out CME successfully. In this study, we build on these pioneering experimental studies to provide fundamental mechanistic insights into the roles of two key endocytic proteins-namely, actin and BAR proteins-in driving vesicle formation in high membrane tension environment. Our study reveals an actin force-induced "snap-through instability" that triggers a rapid shape transition from a shallow invagination to a highly invaginated tubular structure. We show that the association of BAR proteins stabilizes vesicles and induces a milder instability. In addition, we present a rather counterintuitive role of BAR depolymerization in regulating the shape evolution of vesicles. We show that the dissociation of BAR proteins, supported by actin-BAR synergy, leads to considerable elongation and squeezing of vesicles. Going beyond the membrane geometry, we put forth a stress-based perspective for the onset of vesicle scission and predict the shapes and composition of detached vesicles. We present the snap-through transition and the high in-plane stress as possible explanations for the intriguing direct transformation of broad and shallow invaginations into detached vesicles in BAR mutant yeast cells.

Anisotropic spontaneous curvatures in lipid membranes.

Symmetry restrictions due to fluidity require the strain energy in the Helfrich theory of lipid membranes to be locally isotropic in nature. Although this framework is suitable for modeling the interaction of membranes with proteins that generate spherical curvature such as clathrin, there are other important membrane-bending proteins such as BIN-amphiphysin-Rvs proteins that form a cylindrical coat with different curvatures in the longitudinal and the circumferential directions. In this work, we present a detailed mathematical treatment of the theory of lipid membranes incorporating anisotropic spontaneous curvatures. We derive the associated Euler-Lagrange equations and the edge conditions in a generalized setting that allows spatial heterogeneities in the properties of the membrane-protein system. We employ this theory to model the constriction of a membrane tubule by a cylindrical scaffold. In particular, we highlight the role of the equilibrium equation in the tangential plane in regulating the spatial variation of the surface tension field.