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Cell swelling is known to be involved in various stages of the growth of plant cells and microorganisms but in mammalian cells how crucial a swollen state is for determining the fate of the cellular proliferation remains unclear. Recent evidence has increased our understanding of how the loss of the cell surface interactions with the extracellular matrix at early mitosis decreases the membrane tension triggering curvature changes in the plasma membrane and the activation of the sodium/hydrogen (Na +/H +) exchanger (NHE1) that drives osmotic swelling. Such a swollen state is temporary, but it is critical to alter essential membrane biophysical parameters that are required to activate Ca2 + channels and modulate the opening of K + channels involved in setting the membrane potential. A decreased membrane potential across the mitotic cell membrane enhances the clustering of Ras proteins involved in the Ca2 + and cytoskeleton-driven events that lead to cell rounding. Changes in the external mechanical and osmotic forces also have an impact on the lipid composition of the plasma membrane during mitosis. [ABSTRACT FROM AUTHOR]

Adhesion between a solid sphere and a thin film is a common but crucial issue in the study of biological membranes and two-dimensional materials. To supplement quantitative knowledge of membrane adhesion, this work addresses the axisymmetric adhesive contact between a rigid sphere and a circular freestanding elastic membrane clamped at the perimeter. For the membranes following linear stretching elasticity with initial radial tension, both the Johnson-Kendall-Roberts (JKR)- and Derjaguin-Muller-Toporov (DMT)-type adhesion as well as the transition regime in-between are considered. The dependency of contact radius and displacement on external force is studied analytically. In essence, the general solution is governed by three dimensionless parameters, reflecting the effects of membrane stretching elasticity, the range of adhesion force, and the membrane size. It is interestingly found that the membrane size does not affect the contact radius and displacement at zero external force at all and has minor influence on the value of pull-off force. The presented closed form solutions might be useful for the understanding of adhesion behaviors of sphere-membrane systems. [ABSTRACT FROM AUTHOR]

Severe asthma is characterized by increased cell volume (hypertrophy) and enhanced contractile function (hyperresponsiveness) of the airway smooth muscle cells (ASMCs). The causative relationship and underlying regulatory mechanisms between them, however, have remained unclear. Here, we manipulated the single-cell volume of in vitro cultured human ASMCs to increase from 2.7 to 5.2 and 8.2 × 103 μm3 as a simulated ASMC hypertrophy by culturing the cells on micropatterned rectangular substrates with a width of 25 μm and length from 50 to 100 and 200 μm, respectively. We found that as the cell volume increased, ASMCs exhibited a pro-contractile function with increased mRNA expression of contractile proteins, increased cell stiffness and traction force, and enhanced response to contractile stimulation. We also uncovered a concomitant increase in membrane tension and Piezo1 mRNA expression with increasing cell volume. Perhaps more importantly, we found that the enhanced contractile function due to cell volume increase was largely attenuated when membrane tension and Piezo1 mRNA expression were downregulated, and an auto-regulatory loop between Piezo1 and YAP mRNA expression was also involved in perpetuating the contractile function. These findings, thus, provide convincing evidence of a direct link between hypertrophy and enhanced contractile function of ASMCs that was mediated via Piezo1 mRNA expression, which may be specifically targeted as a novel therapeutic strategy to treat pulmonary diseases associated with ASMC hypertrophy such as severe asthma. [ABSTRACT FROM AUTHOR]

Deciphering the dynamic mechanism of ferroptosis can provide insights into pathogenesis, which is valuable for disease diagnosis and treatment. However, due to the lack of suitable time-resolved mechanosensitive tools, researchers have been unable to determine the membrane tension and morphology of the plasma membrane and the nuclear envelope during ferroptosis. With this research, we propose a rational strategy to develop robust mechanosensitive fluorescence lifetime probes which can facilitate simultaneous fluorescence lifetime imaging of the plasma membrane and nuclear envelope. Fluorescence lifetime imaging microscopy using the unique mechanosensitive probes reveal a dynamic mechanism for ferroptosis: The membrane tension of both the plasma membrane and the nuclear envelope decreases during ferroptosis, and the nuclear envelope exhibits budding during the advanced stage of ferroptosis. Significantly, the membrane tension of the plasma membrane is always larger than that of the nuclear envelope, and the membrane tension of the nuclear envelope is slightly larger than that of the nuclear membrane bubble. Meanwhile, the membrane lesions are repaired in the low-tension regions through exocytosis. [ABSTRACT FROM AUTHOR]

Controlling cell-substrate interactions via the microstructural characteristics of biomaterials offers an advantageous path for modulating cell dynamics, mechanosensing, and migration, as well as for designing immune-modulating implants, all without the drawbacks of chemical-based triggers. Specifically, recent in vivo studies have suggested that a porous implants microscale curvature landscape can significantly impact cell behavior and ultimately the immune response. To investigate such cell-substrate interactions, we utilized a 3D computational model incorporating the minimum necessary physics of cell migration and cell-substrate interactions needed to replicate known in vitro behaviors. This model specifically incorporates the effect of membrane tension, which was found to be necessary to replicate in vitro cell behavior on curved surfaces. Our simulated substrates represent two classes of porous materials recently used in implant studies, which have markedly different microscale curvature distributions and pore geometries. We found distinct differences between the overall migration behaviors, shapes, and actin polymerization dynamics of cells interacting with the two substrates. These differences were correlated to the shape energy of the cells as they interacted with the porous substrates, in effect interpreting substrate topography as an energetic landscape interrogated by cells. Our results demonstrate that microscale curvature directly influences cell shape and migration and, therefore, is likely to influence cell behavior. This supports further investigation of the relationship between the surface topography of implanted materials and the characteristic immune response, a complete understanding of which would broadly advance principles of biomaterial design.

Membrane tension is thought to be a long-range integrator of cell physiology. Membrane tension has been proposed to enable cell polarity during migration through front-back coordination and long-range protrusion competition. These roles necessitate effective tension transmission across the cell. However, conflicting observations have left the field divided as to whether cell membranes support or resist tension propagation. This discrepancy likely originates from the use of exogenous forces that may not accurately mimic endogenous forces. We overcome this complication by leveraging optogenetics to directly control localized actin-based protrusions or actomyosin contractions while simultaneously monitoring the propagation of membrane tension using dual-trap optical tweezers. Surprisingly, actin-driven protrusions and actomyosin contractions both elicit rapid global membrane tension propagation, whereas forces applied to cell membranes alone do not. We present a simple unifying mechanical model in which mechanical forces that engage the actin cortex drive rapid, robust membrane tension propagation through long-range membrane flows.

Abstract Background Among mechanoporation techniques for intracellular delivery, microfluidic approaches succeed in high delivery efficiency and throughput. However, especially the entry of large cargoes (e.g. DNA origami, mRNAs, organic/inorganic nanoparticles) is currently impaired since it requires large cell membrane pores with the need to apply multi-step processes and high forces, dramatically reducing cell viability. Results Here, HiViPore presents as a microfluidic viscoelastic contactless compression for one-step cell mechanoporation to produce large pores while preserving high cell viability. Inducing an increase of curvature at the equatorial region of cells, formation of a pore with a size of ~ 1 μm is obtained. The poration is coupled to an increase of membrane tension, measured as a raised fluorescence lifetime of 12% of a planarizable push-pull fluorescent probe (Flipper-TR) labelling the cell plasma membrane. Importantly, the local disruptions of cell membrane are transient and non-invasive, with a complete recovery of cell integrity and functions in ~ 10 min. As result, HiViPore guarantees cell viability as high as ~ 90%. In such conditions, an endocytic-free diffusion of large nanoparticles is obtained with typical size up to 500 nm and with a delivery efficiency up to 12 times higher than not-treated cells. Conclusions The proposed one-step contactless mechanoporation results in an efficient and safe approach for advancing intracellular delivery strategies. In detail, HiViPore solves the issues of low cell viability when multiple steps of poration are required to obtain large pores across the cell plasma membrane. Moreover, the compression uses a versatile, low-cost, biocompatible viscoelastic fluid, thus also optimizing the operational costs. With HiViPore, we aim to propose an easy-to-use microfluidic device to a wide range of users, involved in biomedical research, imaging techniques and nanotechnology for intracellular delivery applications in cell engineering.

The transient cellular organelles known as migrasomes, which form during cell migration along retraction fibers, have emerged as a crutial factor in various fundamental cellular processes and pathologies. These membrane vesicles originate from local membrane swellings, encapsulate specific cytoplasmic content, and are eventually released to the extracellular environment or taken up by recipient cells. Migrasome biogenesis entails a sequential membrane remodeling process involving a complex interplay between various molecular factors such as tetraspanin proteins, and mechanical properties like membrane tension and bending rigidity. In this review, we summarize recent studies exploring the mechanism of migrasome formation. We emphasize how physical forces, together with molecular factors, shape migrasome biogenesis, and detail the involvement of migrasomes in various cellular processes and pathologies. A comprehensive understanding of the exact mechanism underlying migrasome formation and the identification of key molecules involved hold promise for advancing their therapeutic and diagnostic applications. [ABSTRACT FROM AUTHOR]

Background: Among mechanoporation techniques for intracellular delivery, microfluidic approaches succeed in high delivery efficiency and throughput. However, especially the entry of large cargoes (e.g. DNA origami, mRNAs, organic/inorganic nanoparticles) is currently impaired since it requires large cell membrane pores with the need to apply multi-step processes and high forces, dramatically reducing cell viability. Results: Here, HiViPore presents as a microfluidic viscoelastic contactless compression for one-step cell mechanoporation to produce large pores while preserving high cell viability. Inducing an increase of curvature at the equatorial region of cells, formation of a pore with a size of ~ 1 μm is obtained. The poration is coupled to an increase of membrane tension, measured as a raised fluorescence lifetime of 12% of a planarizable push-pull fluorescent probe (Flipper-TR) labelling the cell plasma membrane. Importantly, the local disruptions of cell membrane are transient and non-invasive, with a complete recovery of cell integrity and functions in ~ 10 min. As result, HiViPore guarantees cell viability as high as ~ 90%. In such conditions, an endocytic-free diffusion of large nanoparticles is obtained with typical size up to 500 nm and with a delivery efficiency up to 12 times higher than not-treated cells. Conclusions: The proposed one-step contactless mechanoporation results in an efficient and safe approach for advancing intracellular delivery strategies. In detail, HiViPore solves the issues of low cell viability when multiple steps of poration are required to obtain large pores across the cell plasma membrane. Moreover, the compression uses a versatile, low-cost, biocompatible viscoelastic fluid, thus also optimizing the operational costs. With HiViPore, we aim to propose an easy-to-use microfluidic device to a wide range of users, involved in biomedical research, imaging techniques and nanotechnology for intracellular delivery applications in cell engineering. [ABSTRACT FROM AUTHOR]

Cytokinesis is the process where the mother cell's cytoplasm separates into daughter cells. This is driven by an actomyosin contractile ring that produces cortical contractility and drives cleavage furrow ingression, resulting in the formation of a thin intercellular bridge. While cytoskeletal reorganization during cytokinesis has been extensively studied, less is known about the spatiotemporal dynamics of the plasma membrane. Here, we image and model plasma membrane lipid and protein dynamics on the cell surface during leukemia cell cytokinesis. We reveal an extensive accumulation and folding of the plasma membrane at the cleavage furrow and the intercellular bridge, accompanied by a depletion and unfolding of the plasma membrane at the cell poles. These membrane dynamics are caused by two actomyosin-driven biophysical mechanisms: the radial constriction of the cleavage furrow causes local compression of the apparent cell surface area and accumulation of the plasma membrane at the furrow, while actomyosin cortical flows drag the plasma membrane toward the cell division plane as the furrow ingresses. The magnitude of these effects depends on the plasma membrane fluidity, cortex adhesion, and cortical contractility. Overall, our work reveals cell-intrinsic mechanical regulation of plasma membrane accumulation at the cleavage furrow that is likely to generate localized differences in membrane tension across the cytokinetic cell. This may locally alter endocytosis, exocytosis, and mechanotransduction, while also serving as a self-protecting mechanism against cytokinesis failures that arise from high membrane tension at the intercellular bridge. [ABSTRACT FROM AUTHOR]

Caveolae are small membrane pits with fundamental roles in mechanotransduction. Several studies have shown that caveolae flatten out in response to increased membrane tension, thereby acting as a mechanosensitive membrane reservoir that buffers acute mechanical stress. Caveolae have also been implicated in the control of RhoA/ROCK-mediated actomyosin contractility at the rear of migrating cells. However, how membrane tension controls the organisation of caveolae and their role in mechanotransduction remains unclear. To address this, we systematically quantified protein–protein interactions of caveolin-1 in migrating RPE1 cells at steady state and in response to an acute increase in membrane tension using biotin-based proximity labelling and quantitative mass spectrometry. Our data show that caveolae are highly enriched at the rear of migrating RPE1 cells and that membrane tension rapidly and reversibly disrupts the caveolar protein coat. Membrane tension also detaches caveolin-1 from focal adhesion proteins and several mechanosensitive regulators of cortical actin including filamins and cortactin. In addition, we present evidence that ROCK and the RhoGAP ARHGAP29 associate with caveolin-1 in a manner dependent on membrane tension, with ARHGAP29 influencing caveolin-1 Y14 phosphorylation, caveolae rear localisation, and RPE1 cell migration. Taken together, our work uncovers a membrane tension-sensitive coupling between caveolae and the rear-localised F-actin cytoskeleton. This provides a framework for dissecting the molecular mechanisms underlying caveolae-regulated mechanotransduction pathways.

Drought is one of the most serious abiotic stresses to land plants. Plants sense and respond to drought stress to survive under water deficiency. Scientists have studied how plants sense drought stress, or osmotic stress caused by drought, ever since Charles Darwin, and gradually obtained clues about osmotic stress sensing and signaling in plants. Osmotic stress is a physical stimulus that triggers many physiological changes at the cellular level, including changes in turgor, cell wall stiffness and integrity, membrane tension, and cell fluid volume, and plants may sense some of these stimuli and trigger downstream responses. In this review, we emphasized water potential and movements in organisms, compared putative signal inputs in cell wall‐containing and cell wall‐free organisms, prospected how plants sense changes in turgor, membrane tension, and cell fluid volume under osmotic stress according to advances in plants, animals, yeasts, and bacteria, summarized multilevel biochemical and physiological signal outputs, such as plasma membrane nanodomain formation, membrane water permeability, root hydrotropism, root halotropism, Casparian strip and suberin lamellae, and finally proposed a hypothesis that osmotic stress responses are likely to be a cocktail of signaling mediated by multiple osmosensors. We also discussed the core scientific questions, provided perspective about the future directions in this field, and highlighted the importance of robust and smart root systems and efficient source‐sink allocations for generating future high‐yield stress‐resistant crops and plants. [ABSTRACT FROM AUTHOR]

The budding of membranes and curvature generation is common to many forms of trafficking in cells. Clathrin-mediated endocytosis, as a prototypical example of trafficking, has been studied in great detail using a variety of experimental systems and methods. Recently, advances in experimental methods have led to great strides in insights on the molecular mechanisms and the spatiotemporal dynamics of the protein machinery associated with membrane curvature generation. These advances have been ably supported by computational models, which have given us insights into the underlying mechanical principles of clathrin-mediated endocytosis. On the other hand, targeted experimental perturbation of membranes has lagged behind that of proteins in cells. In this area, modeling is especially critical to interpret experimental measurements in a mechanistic context. Here, we discuss the contributions made by these models to our understanding of endocytosis and identify opportunities to strengthen the connections between models and experiments.

Drought causes osmotic stress and is the most serious natural disaster leading to crop failure. Ever since Darwin studied how plants sense and respond to drought, we have understood the mechanism of ABA（abscisic acid）signaling and gained some clues about drought and osmotic stress sensing and signaling in plants. In this review, we summarized recent advances in plant osmotic stress sensing and signaling. We proposed the putative manners of signal inputs during drought and osmotic stresses and discussed how plants sense and transduce these signals. We also discussed the core scientific questions and made perspective about the future directions in this field, aiming to provide clues for crop genetic improvement with drought resistance. [ABSTRACT FROM AUTHOR]

The inherent membrane tension of biological materials could vitally affect their responses to contact loading but is generally ignored in existing indentation analysis. In this paper, the authors theoretically investigate the contact stiffness of axisymmetric indentations of elastic solids covered with thin tensed membranes. When the indentation size decreases to the same order as the ratio of membrane tension to elastic modulus, the contact stiffness accounting for the effect of membrane tension becomes much higher than the prediction of conventional contact theory. An explicit expression is derived for the contact stiffness, which is universal for axisymmetric indentations using indenters of arbitrary convex profiles. On this basis, a simple method of analysis is proposed to estimate the membrane tension and elastic modulus of biological materials from the indentation load-depth data, which is successfully applied to analyze the indentation experiments of cells and lungs. This study might be helpful for the comprehensive assessment of the mechanical properties of soft biological systems. This paper highlights the crucial effect of the inherent membrane tension on the indentation response of soft biomaterials, which has been generally ignored in existing analysis of experiments. For typical indentation tests on cells and organs, the contact stiffness can be twice or higher than the prediction of conventional contact model. A universal expression of the contact stiffness accounting for the membrane tension effect is derived. On this basis, a simple method of analysis is proposed to abstract the membrane tension of biomaterials from the experimentally recorded indentation load-depth data. With this method, the elasticity of soft biomaterials can be characterized more comprehensively. [Display omitted] [ABSTRACT FROM AUTHOR]

Abstract Background Prostate cancer (PCa) is usually considered as cold tumor. Malignancy is associated with cell mechanic changes that contribute to extensive cell deformation required for metastatic dissemination. Thus, we established stiff and soft tumor subtypes for PCa patients from perspective of membrane tension. Methods Nonnegative matrix factorization algorithm was used to identify molecular subtypes. We completed analyses using software R 3.6.3 and its suitable packages. Results We constructed stiff and soft tumor subtypes using eight membrane tension-related genes through lasso regression and nonnegative matrix factorization analyses. We found that patients in stiff subtype were more prone to biochemical recurrence than those in soft subtype (HR 16.18; p

Metabolism and mechanics are two key facets of structural and functional processes in cells, such as growth, proliferation, homeostasis and regeneration. Their reciprocal regulation has been increasingly acknowledged in recent years: external physical and mechanical cues entail metabolic changes, which in return regulate cell mechanosensing and mechanotransduction. Since mitochondria are pivotal regulators of metabolism, we review here the reciprocal links between mitochondrial morphodynamics, mechanics and metabolism. Mitochondria are highly dynamic organelles which sense and integrate mechanical, physical and metabolic cues to adapt their morphology, the organization of their network and their metabolic functions. While some of the links between mitochondrial morphodynamics, mechanics and metabolism are already well established, others are still poorly documented and open new fields of research. First, cell metabolism is known to correlate with mitochondrial morphodynamics. For instance, mitochondrial fission, fusion and cristae remodeling allow the cell to fine‐tune its energy production through the contribution of mitochondrial oxidative phosphorylation and cytosolic glycolysis. Second, mechanical cues and alterations in mitochondrial mechanical properties reshape and reorganize the mitochondrial network. Mitochondrial membrane tension emerges as a decisive physical property which regulates mitochondrial morphodynamics. However, the converse link hypothesizing a contribution of morphodynamics to mitochondria mechanics and/or mechanosensitivity has not yet been demonstrated. Third, we highlight that mitochondrial mechanics and metabolism are reciprocally regulated, although little is known about the mechanical adaptation of mitochondria in response to metabolic cues. Deciphering the links between mitochondrial morphodynamics, mechanics and metabolism still presents significant technical and conceptual challenges but is crucial both for a better understanding of mechanobiology and for potential novel therapeutic approaches in diseases such as cancer. [ABSTRACT FROM AUTHOR]

Membrane curvature reflects physical forces operating on the lipid membrane, which plays important roles in cellular processes. Here, we design a mechanosensitive DNA (MSD) nanomachine that mimics natural mechanosensitive PIEZO channels to convert the membrane tension changes of lipid vesicles with different sizes into fluorescence signals in real time. The MSD nanomachine consists of an archetypical six‐helix‐bundle DNA nanopore, cholesterol‐based membrane anchors, and a solvatochromic fluorophore, spiropyran (SP). We find that the DNA nanopore effectively amplifies subtle variations of the membrane tension, which effectively induces the isomerization of weakly emissive SP into highly emissive merocyanine isomers for visualizing membrane tension changes. By measuring the membrane tension via the fluorescence of MSD nanomachine, we establish the correlation between the membrane tension and the curvature that follows the Young‐Laplace equation. This DNA nanotechnology‐enabled strategy opens new routes to studying membrane mechanics in physiological and pathological settings. [ABSTRACT FROM AUTHOR]

Vincristine, a widely used chemotherapeutic agent for treating different cancer, often induces severe peripheral neuropathic pain. A common symptom of vincristine-induced peripheral neuropathic pain is mechanical allodynia and hyperalgesia. However, mechanisms underlying vincristine-induced mechanical allodynia and hyperalgesia are not well understood. In the present study, we show with behavioral assessment in rats that vincristine induces mechanical allodynia and hyperalgesia in a PIEZO2 channel-dependent manner since gene knockdown or pharmacological inhibition of PIEZO2 channels alleviates vincristine-induced mechanical hypersensitivity. Electrophysiological results show that vincristine potentiates PIEZO2 rapidly adapting (RA) mechanically-activated (MA) currents in rat dorsal root ganglion (DRG) neurons. We have found that vincristine-induced potentiation of PIEZO2 MA currents is due to the enhancement of static plasma membrane tension (SPMT) of these cells following vincristine treatment. Reducing SPMT of DRG neurons by cytochalasin D (CD), a disruptor of the actin filament, abolishes vincristine-induced potentiation of PIEZO2 MA currents, and suppresses vincristine-induced mechanical hypersensitivity in rats. Collectively, enhancing SPMT and subsequently potentiating PIEZO2 MA currents in primary afferent neurons may be an underlying mechanism responsible for vincristine-induced mechanical allodynia and hyperalgesia in rats. Targeting to inhibit PIEZO2 channels may be an effective analgesic method to attenuate vincristine-induced mechanical hypersensitivity. Vincristine leads to the swelling of DRG neurons, increases their membrane tension and promotes PIEZO2 channel membrane trafficking, which results in the potentiation of PIEZO2 mechanically-activated currents, thereby mediates vincristine-induced mechanical hypersensitivity. [Display omitted] [ABSTRACT FROM AUTHOR]

Endothelial cells (ECs) elongate in the direction of blood flow, are stiffer, and are considered atheroprotective in areas of the vasculature where flow‐induced shear stress is high and unidirectional and are softer, atherogenic, and polygonal in areas experiencing oscillatory multidirectional flow. To understand the precise roles of EC mechanics and morphology in the uptake of therapeutic nanoparticles (NPs) by atherogenic endothelium, human aortic ECs are induced to adopt prescribed shapes and areas imposed by microcontact patterned adhesive islands. NP uptake per cell increases with increasing spreading area and decreases with increasing cell aspect ratio at constant cell spreading area. Biomechanical analysis shows that elongated cells exhibit higher cellular stress and stiffer membranes than cells with low aspect ratios, indicating a strong correlation between morphology, mechanical phenotype, and NP uptake. Further, ECs elongated by high laminar shear endocytosed NPs to a far lesser extent than those that are nonelongated in the chaotic, lower shear areas when cocultured in the same chamber. Results indicate that conditions leading to atherogenesis, such as low, chaotic shear‐induction of EC polygonal morphology may be used to increase the uptake of therapeutic NPs as a preventative measure against atherosclerosis. [ABSTRACT FROM AUTHOR]

Background: Prostate cancer (PCa) is usually considered as cold tumor. Malignancy is associated with cell mechanic changes that contribute to extensive cell deformation required for metastatic dissemination. Thus, we established stiff and soft tumor subtypes for PCa patients from perspective of membrane tension. Methods: Nonnegative matrix factorization algorithm was used to identify molecular subtypes. We completed analyses using software R 3.6.3 and its suitable packages. Results: We constructed stiff and soft tumor subtypes using eight membrane tension-related genes through lasso regression and nonnegative matrix factorization analyses. We found that patients in stiff subtype were more prone to biochemical recurrence than those in soft subtype (HR 16.18; p < 0.001), which was externally validated in other three cohorts. The top ten mutation genes between stiff and soft subtypes were DNAH, NYNRIN, PTCHD4, WNK1, ARFGEF1, HRAS, ARHGEF2, MYOM1, ITGB6 and CPS1. E2F targets, base excision repair and notch signaling pathway were highly enriched in stiff subtype. Stiff subtype had significantly higher TMB and T cells follicular helper levels than soft subtype, as well as CTLA4, CD276, CD47 and TNFRSF25. Conclusions: From the perspective of cell membrane tension, we found that stiff and soft tumor subtypes were closely associated with BCR-free survival for PCa patients, which might be important for the future research in the field of PCa. [ABSTRACT FROM AUTHOR]

Flipper probes have been introduced as small molecule fluorophores to image physical forces, that is, membrane tension in living systems. Their emergence over one decade is described, from evolution in design and synthesis to spectroscopic properties. Responsiveness to physical compression in equilibrium at the ground state is identified as the ideal origin of mechanosensitivity to image membrane tension in living cells. A rich collection of flippers is described to deliver and release in any subcellular membrane of interest in a leaflet‐specific manner. Chalcogen‐bonding cascade switching and dynamic covalent flippers are developed for super‐resolution imaging and dual‐sensing of membrane compression and hydration. Availability and broad use in the community validate flipper probes as a fine example of the power of translational supramolecular chemistry, moving from fundamental principles to success on the market. [ABSTRACT FROM AUTHOR]

Lateral phase separation within lipid bilayer membranes has attracted considerable attention in the fields of biophysics and cell biology. Living cells organize laterally segregated compartments, such as raft domains in an ordered phase, and regulate their dynamic structures under isothermal conditions to promote cellular functions. Model membrane systems with minimum components are powerful tools for investigating the basic phenomena of membrane phase separation. With the use of such model systems, several physicochemical characteristics of phase separation have been revealed. This review focuses on the isothermal triggering of membrane phase separation from a physical point of view. We consider the free energy of the membrane that describes lateral phase separation and explain the experimental results of model membranes to regulate domain formation under isothermal conditions. Three possible regulation factors are discussed: electrostatic interactions, chemical reactions and membrane tension. These findings may contribute to a better understanding of membrane lateral organization within living cells that function under isothermal conditions and could be useful for the development of artificial cell engineering. [ABSTRACT FROM AUTHOR]

Abstract Endothelial cells (ECs) elongate in the direction of blood flow, are stiffer, and are considered atheroprotective in areas of the vasculature where flow‐induced shear stress is high and unidirectional and are softer, atherogenic, and polygonal in areas experiencing oscillatory multidirectional flow. To understand the precise roles of EC mechanics and morphology in the uptake of therapeutic nanoparticles (NPs) by atherogenic endothelium, human aortic ECs are induced to adopt prescribed shapes and areas imposed by microcontact patterned adhesive islands. NP uptake per cell increases with increasing spreading area and decreases with increasing cell aspect ratio at constant cell spreading area. Biomechanical analysis shows that elongated cells exhibit higher cellular stress and stiffer membranes than cells with low aspect ratios, indicating a strong correlation between morphology, mechanical phenotype, and NP uptake. Further, ECs elongated by high laminar shear endocytosed NPs to a far lesser extent than those that are nonelongated in the chaotic, lower shear areas when cocultured in the same chamber. Results indicate that conditions leading to atherogenesis, such as low, chaotic shear‐induction of EC polygonal morphology may be used to increase the uptake of therapeutic NPs as a preventative measure against atherosclerosis.

Micromechanics techniques are playing an increasing role in characterization of biomembranes. The mechanical properties of membranes play an important role for a whole range of cellular processes. Lipid-protein biomembranes display lateral heterogeneity, domain formation, and morphological changes at mesoscopic and nanoscopic length scales. An attempt is made to introduce how membrane's material properties can be measured. Both fluctuation analysis and micro-pipette aspiration experiments have been used to quantify the micromechanics of membranes. The relationship between the structure and function of biomembranes is a critical concern in modern biology. This overview calls for a deeper understanding of how the cell complexity might be related to the mechanical properties of the lipid-protein membrane. Mechanical properties can influence cellular response to processes like adhesion, transport, differentiation, proliferation and migration. [ABSTRACT FROM AUTHOR]

Mechanical forces are inseparable from most cellular functions. Cell division, contraction, and adhesion generate intrinsic forces in the cells, whereas perturbations in the environment such as osmotic shifts, mechanical pressure, shear, or sound represent the external forces that the cells gauge and respond to. Mechanosensitive (MS) ion channels, which are the fastest mechanotransducers, represent a polyphyletic group with vastly diverse structural designs. In this review we briefly outline the history of the field by presenting major findings in a nearly chronological order, describe structural features of different groups, and attempt to illustrate some common physical principles of their gating mechanisms. [ABSTRACT FROM AUTHOR]

Here, we study the time-dependent regulation of fluctuation-tension during myogenesis and the role of the fusogen, myomerger. We measure nanometric height fluctuations of the basal membrane of C2C12 cells after triggering differentiation. Fusion of cells increases fluctuation-tension but prefers a transient lowering of tension (at ~2-24 h). Cells fail to fuse if early tension is continuously enhanced by methyl-β-cyclodextrin (MβCD). Perturbing tension regulation also reduces fusion. During this pre-fusion window, cells that finally differentiate usually display lower tension than other non-fusing cells, validating early tension states to be linked to fate decision. Early tension reduction is accompanied by low but gradually increasing level of the surface myomerger. Locally too, regions with higher myomerger intensity display lower tension. However, this negative correlation is lost in the early phase by MβCD-based cholesterol depletion or later as differentiation progresses. We find that with tension and surface-myomerger's enrichment under these conditions, myomerger clusters become pronouncedly diffused. We, therefore, propose that low tension aided by clustered surface-myomerger at the early phase is crucial for fusion and can be disrupted by cholesterol-reducing molecules, implying the potential to affect muscle health. [ABSTRACT FROM AUTHOR]

Here, we study the time-dependent regulation of fluctuation–tension during myogenesis and the role of the fusogen, myomerger. We measure nanometric height fluctuations of the basal membrane of C2C12 cells after triggering differentiation. Fusion of cells increases fluctuation–tension but prefers a transient lowering of tension (at ∼2–24 h). Cells fail to fuse if early tension is continuously enhanced by methyl-β-cyclodextrin (MβCD). Perturbing tension regulation also reduces fusion. During this pre-fusion window, cells that finally differentiate usually display lower tension than other non-fusing cells, validating early tension states to be linked to fate decision. Early tension reduction is accompanied by low but gradually increasing level of the surface myomerger. Locally too, regions with higher myomerger intensity display lower tension. However, this negative correlation is lost in the early phase by MβCD-based cholesterol depletion or later as differentiation progresses. We find that with tension and surface-myomerger’s enrichment under these conditions, myomerger clusters become pronouncedly diffused. We, therefore, propose that low tension aided by clustered surface-myomerger at the early phase is crucial for fusion and can be disrupted by cholesterol-reducing molecules, implying the potential to affect muscle health.

Osmotic pressure can break the fluid balance between intracellular and extracellular solutions. In hypo-osmotic solution, water molecules, which transfer into the cell and burst, are driven by the concentration difference of solute across the semi-permeable membrane. The complicated dynamic processes of intermittent bursts have been previously observed. However, the underlying physical mechanism has yet to be thoroughly explored and analyzed. Here, the intermittent release of inclusion in giant unilamellar vesicles was investigated quantitatively, applying the combination of experimental and theoretical methods in the hypo-osmotic medium. Experimentally, we adopted a highly sensitive electron multiplying charge-coupled device to acquire intermittent dynamic images. Notably, the component of the vesicle phospholipids affected the stretch velocity, and the prepared solution of vesicles adjusted the release time. Theoretically, we chose equations and numerical simulations to quantify the dynamic process in phases and explored the influences of physical parameters such as bilayer permeability and solution viscosity on the process. It was concluded that the time taken to achieve the balance of giant unilamellar vesicles was highly dependent on the molecular structure of the lipid. The pore lifetime was strongly related to the internal solution environment of giant unilamellar vesicles. The vesicles prepared in viscous solution were able to visualize long-lived pores. Furthermore, the line tension was measured quantitatively by the release velocity of inclusion, which was of the same order of magnitude as the theoretical simulation. In all, the experimental values well matched the theoretical values. Our investigation clarified the physical regulatory mechanism of intermittent pore formation and inclusion release, which provides an important reference for the development of novel technologies such as gene therapy based on transmembrane transport as well as controlled drug delivery based on liposomes. [ABSTRACT FROM AUTHOR]

Background: The plasma membrane provides a highly dynamic barrier for cancer cells to interact with their surrounding microenvironment. Membrane tension, a pivotal physical property of the plasma membrane, has attracted widespread attention since it plays a role in the progression of various cancers. This study aimed to identify a prognostic signature in colon cancer from membrane tension-related genes (MTRGs) and explore its implications for the disease. Methods: Bulk RNA-seq data were obtained from The Cancer Genome Atlas (TCGA) database, and then applied to the differentially expressed gene analysis. By implementing a univariate Cox regression and a LASSO-Cox regression, we developed a prognostic model based on four MTRGs. The prognostic efficacy of this model was evaluated in combination with a Kaplan–Meier analysis and receiver operating characteristic (ROC) curve analysis. Moreover, the relationships between the signature and immune cell infiltration, immune status, and somatic mutation were further explored. Lastly, by utilizing single-cell RNA-seq data, cell type annotation, pseudo-time analysis, drug sensitivity, and molecular docking were implemented. Results: We constructed a 4-MTRG signature. The risk score derived from the model was further validated as an independent variable for survival prediction. Two risk groups were divided based on the risk score calculated by the 4-MTRG signature. In addition, we observed a significant difference in immune cell infiltration, such as subsets of CD4 T cells and macrophages, between the high- and low-risk groups. Moreover, in the pseudo-time analysis, TIMP1 was found to be more highly expressed with the progression of time. Finally, three small molecule drugs, elesclomol, shikonin, and bryostatin-1, exhibited a binding potential to TIMP-1. Conclusions: The novel 4-MTRG signature is a promising biomarker in predicting clinical outcomes for colon cancer patients, and TIMP1, a member of the signature, may be a sensitive regulator of the progression of colon cancer. [ABSTRACT FROM AUTHOR]

Cells need to rapidly and precisely react to multiple mechanical and chemical stimuli in order to ensure precise context‐dependent responses. This requires dynamic cellular signalling events that ensure homeostasis and plasticity when needed. A less well‐understood process is cellular response to elevated interstitial fluid pressure, where the cell senses and responds to changes in extracellular hydrostatic pressure. Here, using quantitative label‐free digital holographic imaging, combined with genome editing, biochemical assays and confocal imaging, we analyse the temporal cellular response to hydrostatic pressure. Upon elevated cyclic hydrostatic pressure, the cell responds by rapid, dramatic and reversible changes in cellular volume. We show that YAP and TAZ, the co‐transcriptional regulators of the Hippo signalling pathway, control cell volume and that cells without YAP and TAZ have lower plasma membrane tension. We present direct evidence that YAP/TAZ drive the cellular response to hydrostatic pressure, a process that is at least partly mediated via clathrin‐dependent endocytosis. Additionally, upon elevated oscillating hydrostatic pressure, YAP/TAZ are activated and induce TEAD‐mediated transcription and expression of cellular components involved in dynamic regulation of cell volume and extracellular matrix. This cellular response confers a feedback loop that allows the cell to robustly respond to changes in interstitial fluid pressure. Synopsis: YAP/TAZ transcriptional co‐activators are central regulators of cellular response to changes in the extracellular and intracellular mechanical environment. This study identifies oscillating hydrostatic pressure as a new regulator of cell volume via YAP/TAZ activation. YAP/TAZ regulate cell volume and plasma membrane tensionOscillating hydrostatic pressure induces rapid and reversible changes in cellular volumeYAP/TAZ activation are essential for the cellular response to hydrostatic pressureCellular response to oscillating hydrostatic pressure involves clathrin‐dependent endocytosis and the actin cytoskeleton [ABSTRACT FROM AUTHOR]

Simple Summary: Here, we review the known characteristics and functions of proteins called myosin-I. These mechanoenzymes belong to an ancient family of actin-dependent motors, found throughout eukaryotic cells, which are characterized by intracellular membrane-bound compartments. We elaborate on the surprising fact that many different functions have been attributed to these proteins, and highlight that we now need to understand how their enzymatic activity supports these functions. We propose to focus on the remodeling of the actin cytoskeleton, a higher-order dynamic scaffold typical for eukaryotic cells. The actin cytoskeleton plays crucial roles in cell morphogenesis and functions. The main partners of cortical actin are molecular motors of the myosin superfamily. Although our understanding of myosin functions is heavily based on myosin-II and its ability to dimerize, the largest and most ancient class is represented by myosin-I. Class 1 myosins are monomeric, actin-based motors that regulate a wide spectrum of functions, and whose dysregulation mediates multiple human diseases. We highlight the current challenges in identifying the "pantograph" for myosin-I motors: we need to reveal how conformational changes of myosin-I motors lead to diverse cellular as well as multicellular phenotypes. We review several mechanisms for scaling, and focus on the (re-) emerging function of class 1 myosins to remodel the actin network architecture, a higher-order dynamic scaffold that has potential to leverage molecular myosin-I functions. Undoubtfully, understanding the molecular functions of myosin-I motors will reveal unexpected stories about its big partner, the dynamic actin cytoskeleton. [ABSTRACT FROM AUTHOR]

A critical step in cellular-trafficking pathways is the budding of membranes by protein coats, which recent experiments have demonstrated can be inhibited by elevated membrane tension. The robustness of processes like clathrin-mediated endocytosis (CME) across a diverse range of organisms and mechanical environments suggests that the protein machinery in this process has evolved to take advantage of some set of physical design principles to ensure robust vesiculation against opposing forces like membrane tension. Using a theoretical model for membrane mechanics and membrane protein interaction, we have systematically investigated the influence of membrane rigidity, curvature induced by the protein coat, area covered by the protein coat, membrane tension, and force from actin polymerization on bud formation. Under low tension, the membrane smoothly evolves from a flat to budded morphology as the coat area or spontaneous curvature increases, whereas the membrane remains essentially flat at high tensions. At intermediate, physiologically relevant, tensions, the membrane undergoes a "snap-through instability" in which small changes in the coat area, spontaneous curvature or membrane tension cause the membrane to "snap" from an open, U-shape to a closed bud. This instability can be smoothed out by increasing the bending rigidity of the coat, allowing for successful budding at higher membrane tensions. Additionally, applied force from actin polymerization can bypass the instability by inducing a smooth transition from an open to a closed bud. Finally, a combination of increased coat rigidity and force from actin polymerization enables robust vesiculation even at high membrane tensions.

Mechanics has been a central focus of physical biology in the past decade. In comparison, how cells manage their size is less understood. Here, we show that a parameter central to both the physics and the physiology of the cell, its volume, depends on a mechano-osmotic coupling. We found that cells change their volume depending on the rate at which they change shape, when they spontaneously spread or when they are externally deformed. Cells undergo slow deformation at constant volume, while fast deformation leads to volume loss. We propose a mechanosensitive pump and leak model to explain this phenomenon. Our model and experiments suggest that volume modulation depends on the state of the actin cortex and the coupling of ion fluxes to membrane tension. This mechano-osmotic coupling defines a membrane tension homeostasis module constantly at work in cells, causing volume fluctuations associated with fast cell shape changes, with potential consequences on cellular physiology.

Contrary to sulfides, sulfoxides or sulfones, sulfoximines have been mostly neglected in the design of fluorescent probes until recently. In this study, we elaborate systematically on sulfoximine acceptors in fluorescent flipper probes. Fluorescent flippers have been introduced as mechanosensitive probes to image membrane order and tension. They consist of twisted dithienothiophene dimers with sulfide and sulfone bridges to produce the essential primary dipole of the coupled push‐pull system. The objective of this study was to replace the sulfone acceptor by a series of sulfoximines. This is intriguing as a synthetic challenge and worthwhile because the extra nitrogen substituent offers a variability that is attractive to understand and control the performance of the probes. The new sulfoximine flippers provide corroborative evidence for the importance of the primary dipole of the planarizable push‐pull probe. Partitioning into differently ordered membranes and positioning within these different membranes is shown to correlate directly and dramatically with fluorescence lifetimes and mechanosensitivity. Sufficient partitioning into ordered membranes is confirmed as particularly important to image membrane tension by probe compression in the ground state. Compared to the conventional sulfone homolog, the best sulfoximine flipper has more red‐shifted absorption and emission maxima, longer fluorescence lifetime in cell membranes, and larger difference in lifetime upon application of membrane tension. [ABSTRACT FROM AUTHOR]

Many signal transductions resulting from ligand–receptor interactions occur at the cell surface. These signaling pathways play essential roles in cell polarization, membrane morphogenesis, and the modulation of membrane tension at the cell surface. However, due to the large number of membrane-binding proteins, including actin-membrane linkers, and transmembrane proteins present at the cell surface, the molecular mechanisms underlying the regulation at the cell surface are yet unclear. Here, we describe the molecular functions of one of the key players at the cell surface, ezrin/radixin/moesin (ERM) proteins from a biophysical point of view. We focus our discussion on biophysical properties of ERM proteins revealed by using biophysical tools in live cells and in vitro reconstitution systems. We first describe the structural properties of ERM proteins and then discuss the interactions of ERM proteins with PI(4,5)P2 and the actin cytoskeleton. These properties of ERM proteins revealed by using biophysical approaches have led to a better understanding of their physiological functions in cells and tissues. [ABSTRACT FROM AUTHOR]

Plasma membrane tension is known to regulate many cell functions, such as motility and membrane trafficking. Membrane tether pulling is an effective method for measuring the apparent membrane tension of cells and exploring membrane-cytoskeleton interactions. In this article, the mechanical properties of HP1 α -depleted MCF7 breast cancer cells are explored in comparison to controls, by pulling membrane tethers using optical tweezers. These studies were inspired by previous findings that a loss of HP1 α correlates with an increase in the invasive potential of malignant cancer cells. Specifically, the membrane tension and force relaxation curves for tethers pulled from MCF7 breast cancer cells with HP1 α knockdown and their matched controls were measured, and shown to be significantly different. • The mechanical properties of MCF7 breast cancer cells have been explored. • Membrane tethers were pulled from the cells using optical tweezers. • Results for membrane tension were compared when HP1 α was depleted. [ABSTRACT FROM AUTHOR]

Mechanosensitive flipper probes are attracting interest as fluorescent reporters of membrane order and tension in biological systems. We introduce PhotoFlippers, which contain a photocleavable linker and an ultralong tether between mechanophore and various targeting motifs. Upon irradiation, the original probe is released and labels the most ordered membrane that is accessible by intermembrane transfer. Spatiotemporal control from photocleavable flippers is essential to access open, dynamic or elusive membrane motifs without chemical or physical interference. For instance, fast release with light is shown to place the original small‐molecule probes into the innermost leaflet of the nuclear envelope to image changes in membrane tension, at specific points in time of membrane trafficking along the secretory pathway, or in the inner leaflet of the plasma membrane to explore membrane asymmetry. These results identify PhotoFlippers as useful chemistry tools to enable research in biology. [ABSTRACT FROM AUTHOR]

Sterols are a class of lipids critical for fundamental biological processes and membrane dynamics. These molecules are synthesized in the endoplasmic reticulum (ER) and are transported bi-directionally between the ER and plasma membrane (PM). However, the trafficking mechanism of sterols and their relationship with macroautophagy/autophagy are still poorly understood in the rice blast fungus Magnaporthe oryzae. Here, we identified the VAD1 Analog of StAR-related lipid transfer (VASt) domain-containing protein MoVast1 via co-immunoprecipitation in M. oryzae. Loss of MoVAST1 resulted in conidial defects, impaired appressorium development, and reduced pathogenicity. The MoTor (target of rapamycin in M. oryzae) activity is inhibited because MoVast1 deletion leads to high levels of sterol accumulation in the PM. Site-directed mutagenesis showed that the 902 T site is essential for localization and function of MoVast1. Through filipin or Flipper-TR staining, autophagic flux detection, MoAtg8 lipidation, and drug sensitivity assays, we uncovered that MoVast1 acts as a novel autophagy inhibition factor that monitors tension in the PM by regulating the sterol content, which in turn modulates the activity of MoTor. Lipidomics and transcriptomics analyses further confirmed that MoVast1 is an important regulator of lipid metabolism and the autophagy pathway. Our results revealed and characterized a novel sterol transfer protein important for M. oryzae pathogenicity. Abbreviations: AmB: amphotericin B; ATMT: Agrobacterium tumefaciens-mediated transformation; CM: complete medium; dpi: days post-inoculation; ER: endoplasmic reticulum; Flipper-TR: fluorescent lipid tension reporter; GO: Gene ontology; hpi: hours post-inoculation; IH: invasive hyphae; KEGG: kyoto encyclopedia of genes and genomes; MoTor: target of rapamycin in Magnaporthe oryzae; PalmC: palmitoylcarnitine; PM: plasma membrane; SD-N: synthetic defined medium without amino acids and ammonium sulfate; TOR: target of rapamycin; VASt: VAD1 Analog of StAR-related lipid transfer; YFP, yellow fluorescent protein. [ABSTRACT FROM AUTHOR]

Erythrocytes swell owing to the osmotic shock caused by hypotonic liquids, and when the membrane tension exceeds a certain limit, hemolysis occurs. The base tension in the membrane of a spherically shaped erythrocyte is usually ignored in the mechanical evaluation of hemolysis. However, the base tension cannot be ignored when the rigidity of the erythrocyte membrane increases owing to lesions, oxidative stress, and other phenomena. Therefore, it is necessary to re-evaluate the tension level at which hemolysis occurs by considering the increased base tension, which is caused by a combined increase in the bending and shear rigidity of the membrane. To achieve this, we calculated the effect of increases in the combined rigidity on the increases in the internal pressure and membrane tension of the erythrocyte. In this study, assuming the surface area to be constant, the swelling process of erythrocytes was evaluated under the condition that hemolysis does not occur. Evaluation was performed by minimizing strain energy, which is the sum of bending strain and shear strain. When the erythrocyte was spherical, the membrane base tension increased linearly with combined rigidity. Even when the bending rigidity was increased to 100 times that of normal erythrocytes, the effect of the base tension on the hemolysis tension level (15 mN/m) was negligible. However, when shear rigidity was increased to 50-100 times that of normal erythrocytes, it became necessary to decrease the hemolysis tension level by 10% and 20%, respectively, because the base tensions were approximately 1.5 and 3.0 mN/m, respectively.

Endocytic traffic is a complex and elegant operation involving cargo sorting, membrane budding and tubulation, generation of force, and the formation of organellar contacts. The role of specific proteins and lipids in these processes has been studied extensively. By comparison, precious little is understood about the contribution of the endocytic fluid to these events, despite much evidence that alteration of the contents can severely affect membrane traffic along the endocytic pathway. In particular, it has long been appreciated that dissipation of ionic gradients arrests endosome-to-lysosome maturation. How cells sense inorganic ions and transmit this information have remained largely enigmatic. Herein, we review the experimental findings that reveal an intimate association between luminal ions, their transport, and endocytic traffic. We then discuss the ionic sensors and the mechanisms proposed to convert ion concentrations into protein-based trafficking events, highlighting the current paucity of convincing explanations. [ABSTRACT FROM AUTHOR]