Membrane Tension Research Articles

Native Cellular Membranes Facilitate Channel Activity of MscL by Enhancing Slow Collective Motions of Its Transmembrane Helices.

Mechanosensitive channels of large conductance (MscL) serve as a mechanoelectrical valve of cells in response to the membrane tension. The influence of membrane environments on the MscL channel activity and the underlying mechanism remains unclear. Herein, we developed a new sample preparation protocol that allows for the detection of high-quality 1H-detected solid-state NMR spectra of MscL in cellular membranes, enabling site-specific analysis of its dynamics. Dipolar order parameters and spin relaxation rates are measured for 51 residues of MscL in synthetic and native membranes. The dynamics data reveal that while MscL maintains a similar rigidity in both membrane environments, it exhibits enhanced slow collective motions in the native cellular membranes. Molecular dynamics simulations demonstrate the critical role of slow motions in the mechanosensitivity of MscL by promoting protein-membrane interactions. This study examines atomic-resolution dynamics of a membrane-protein in cellular membranes and provides novel insights into the functional significance of membrane-protein dynamics.

RhoA regulates oligodendrocyte differentiation and myelination by orchestrating cortical and membrane tension.

Timely differentiation and myelin formation by oligodendrocytes are essential for the physiological functioning of the central nervous system (CNS). While the Rho GTPase RhoA has been hinted as a negative regulator of myelin sheath formation, the precise in vivo mechanisms have remained elusive. Here we show that RhoA controls the timing and progression of myelination by oligodendrocytes through a fine-tuned balance between cortical tension, membrane tension and cell shape. Using a conditional mouse model, we observe that Rhoa ablation results in the acceleration of myelination driven by hastened differentiation and facilitated through membrane expansion induced by changes in MLCII activity and in F-actin redistribution and turnover within the cell. These findings reveal RhoA as a central molecular integrator of alterations in actin cytoskeleton, actomyosin contractility and membrane tension underlying precise morphogenesis of oligodendrocytes and normal myelination of the CNS.

Coordination of force-generating actin-based modules stabilizes and remodels membranes in vivo.

Membrane remodeling drives a broad spectrum of cellular functions, and it is regulated through mechanical forces exerted on the membrane by cytoplasmic complexes. Here, we investigate how actin filaments dynamically tune their structure to control the active transfer of membranes between cellular compartments with distinct compositions and biophysical properties. Using intravital subcellular microscopy in live rodents we show that a lattice composed of linear filaments stabilizes the granule membrane after fusion with the plasma membrane and a network of branched filaments linked to the membranes by Ezrin, a regulator of membrane tension, initiates and drives to completion the integration step. Our results highlight how the actin cytoskeleton tunes its structure to adapt to dynamic changes in the biophysical properties of membranes.

The Role of the Swollen State in Cell Proliferation.

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.

PREPRINT Machine Learning for the Sensitivity Analysis of a Model of the Cellular Uptake of Nanoparticles for the Treatment of Cancer.

Experimental studies on the cellular uptake of nanoparticles (NPs), useful for the investigation of NP-based drug delivery systems, are often difficult to interpret due to the large number of parameters that can contribute to the phenomenon. It is therefore of great interest to identify insignificant parameters to reduce the number of variables used for the design of experiments. In this work, a model of the wrapping of elliptical NPs by the cell membrane is used to compare the influence of the aspect ratio of the NP, the membrane tension, the NP-membrane adhesion, and its variation during the interaction with the NP on the equilibrium state of the wrapping process. Several surrogate models, such as Kriging, Polynomial Chaos Expansion (PCE), and artificial neural networks (ANN) have been built and compared to emulate the computationally expensive model. Only the ANN-based model outperformed the other approaches by providing much better predictivity metrics and could therefore be used to compute the sensitivity indices. Our results showed that the NP's aspect ratio, the initial NP-membrane adhesion, the membrane tension, and the delay for the increase of the NP-membrane adhesion after receptor dynamics are the main contributors to the cellular internalization of the NP, while the influence of other parameters is negligible.

Integrative analysis of gene expression, protein abundance, and metabolomic profiling elucidates complex relationships in chronic hyperglycemia-induced changes in human aortic smooth muscle cells

Type 2 diabetes mellitus (T2DM) is a major public health concern with significant cardiovascular complications (CVD). Despite extensive epidemiological data, the molecular mechanisms relating hyperglycemia to CVD remain incompletely understood. We here investigated the impact of chronic hyperglycemia on human aortic smooth muscle cells (HASMCs) cultured under varying glucose conditions in vitro, mimicking normal (5 mmol/L), pre-diabetic (10 mmol/L), and diabetic (20 mmol/L) conditions, respectively. Normal HASMC cultures served as baseline controls, and patient-derived T2DM-SMCs served as disease controls. Results showed significant increases in cellular proliferation, area, perimeter, and F-actin expression with increasing glucose concentration (p < 0.01), albeit not exceeding the levels in T2DM cells. Atomic force microscopy analysis revealed significant decreases in Young’s moduli, membrane tether forces, membrane tension, and surface adhesion in SMCs at higher glucose levels (p < 0.001), with T2DM-SMCs being the lowest among all the cases (p < 0.001). T2DM-SMCs exhibited elevated levels of selected pro-inflammatory markers (e.g., ILs-6, 8, 23; MCP-1; M-CSF; MMPs-1, 2, 3) compared to glucose-treated SMCs (p < 0.01). Conversely, growth factors (e.g., VEGF-A, PDGF-AA, TGF-β1) were higher in SMCs exposed to high glucose levels but lower in T2DM-SMCs (p < 0.01). Pathway enrichment analysis showed significant increases in the expression of inflammatory cytokine-associated pathways, especially involving IL-10, IL-4 and IL-13 signaling in genes that are up-regulated by elevated glucose levels. Differentially regulated gene analysis showed that compared to SMCs receiving normal glucose, 513 genes were upregulated and 590 genes were downregulated in T2DM-SMCs; fewer genes were differentially expressed in SMCs receiving higher glucose levels. Finally, the altered levels in genes involved in ECM organization, elastic fiber synthesis and formation, laminin interactions, and ECM proteoglycans were identified. Growing literature suggests that phenotypic switching in SMCs lead to arterial wall remodeling (e.g., change in stiffness, calcific deposits formation), with direct implications in the onset of CVD complications. Our results suggest that chronic hyperglycemia is one such factor that leads to morphological, biomechanical, and functional alterations in vascular SMCs, potentially contributing to the pathogenesis of T2DM-associated arterial remodeling. The observed differences in gene expression patterns between in vitro hyperglycemic models and patient-derived T2DM-SMCs highlight the complexity of T2DM pathophysiology and underline the need for further studies.

Arrangements of intramembranous muscles of wings are influenced by body mass across the radiation of phyllostomid bats.

Extensive research into bat flight mechanisms has highlighted the complex functional and evolutionary dynamics of their wing structures, yet the anatomical details of certain wing muscles remain elusive. In particular, the intramembranous plagiopatagiales proprii muscles, located within the plagiopatagium-an area of the wing lacking direct joint connections-exhibit remarkable variation across bat families. These muscles, which extend anteroposteriorly in macroscopic bundles, play a crucial role in wing stiffening, modulating membrane tension, and reducing wing curvature during flight. Since larger bats tend to have higher wing loading (WL; the ratio of body mass [BMa] to wing area) and may therefore experience increased patagial curvature and resultant drag, we hypothesized that body size significantly influences the evolutionary development of the plagiopatagiales proprii muscles. This study investigates the relationship between BMa and the morphology of the plagiopatagiales proprii in New World leaf-nosed bats (Phyllostomidae), employing bivariate allometry, multivariate analysis, and comparative phylogenetic methods across 24 species from eight phyllostomid subfamilies. Our findings reveal a significant phylogenetic signal in muscle architecture, along with positive evolutionary allometry in muscle area. This suggests an adaptive increase in muscle size in larger species, likely to counterbalance the increased WL, reduce wing curvature, and minimize drag. This research enhances our understanding of the functional and adaptive morphological evolution of intramembranous wing muscles in phyllostomid bats, underscoring their evolutionary significance.

Mechanical characterization of freestanding lipid bilayers with temperature-controlled phase.

Coexistence of lipid domains in cell membranes is associated with vital biological processes. Here, we investigate two such membranes: a multi-component membrane composed of DOPC and DPPC lipids with gel and fluid separated domains, and a single component membrane composed of PMPC lipids forming ripples. We characterize their mechanical properties below their melting point, where ordered and disordered regions coexist, and above their melting point, where they are in fluid phase. To conduct these inquiries, we create lipid bilayers in a microfluidic chip interfaced with a heating system and optical tweezers. The chip features a bubble trap and enables high-throughput formation of planar bilayers. Optical tweezers experiments reveal interfacial hydrodynamics (fluid-slip) and elastic properties (membrane tension and bending rigidity) at various temperatures. For PMPC bilayers, we demonstrate a higher fluid slip at the interface in the fluid-phase compared to the ripple phase, while for the DOPC:DPPC mixture, similar fluid slip is measured below and above the transition point. Membrane tension for both compositions increases after thermal fluidization. Bending rigidity is also measured using the forces required to extend a lipid nanotube pushed out of the freestanding membranes. This novel temperature-controlled microfluidic platform opens numerous possibilities for thermomechanical studies on freestanding planar membranes.

Single-Cell Hypertrophy Promotes Contractile Function of Cultured Human Airway Smooth Muscle Cells via Piezo1 and YAP Auto-Regulation.

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.

Biomolecular condensates mediate bending and scission of endosome membranes

Multivesicular bodies are key endosomal compartments implicated in cellular quality control through their degradation of membrane-bound cargo proteins1–3. The ATP-consuming ESCRT protein machinery mediates the capture and engulfment of membrane-bound cargo proteins through invagination and scission of multivesicular-body membranes to form intraluminal vesicles4,5. Here we report that the plant ESCRT component FREE16 forms liquid-like condensates that associate with membranes to drive intraluminal vesicle formation. We use a minimal physical model, reconstitution experiments and in silico simulations to identify the dynamics of this process and describe intermediate morphologies of nascent intraluminal vesicles. Furthermore, we find that condensate-wetting-induced line tension forces and membrane asymmetries are sufficient to mediate scission of the membrane neck without the ESCRT protein machinery or ATP consumption. Genetic manipulation of the ESCRT pathway in several eukaryotes provides additional evidence for condensate-mediated membrane scission in vivo. We find that the interplay between condensate and machinery-mediated scission mechanisms is indispensable for osmotic stress tolerance in plants. We propose that condensate-mediated scission represents a previously undescribed scission mechanism that depends on the physicomolecular properties of the condensate and is involved in a range of trafficking processes. More generally, FREE1 condensate-mediated membrane scission in multivesicular-body biogenesis highlights the fundamental role of wetting in intracellular dynamics and organization.

Membrane Tension Regulation is Required for Wound Repair.

Disruptions of the eukaryotic plasma membrane due to chemical and mechanical challenges are frequent and detrimental and thus need to be repaired to maintain proper cell function and avoid cell death. However, the cellular mechanisms involved in wound resealing and restoration of homeostasis are diverse and contended. Here, it is shown that clathrin-mediated endocytosis is induced at later stages of plasma membrane wound repair following the actual resealing of the wound. This compensatory endocytosis occurs near the wound, predominantly at sites of previous early endosome exocytosis which is required in the initial stage of membrane resealing, suggesting a spatio-temporal co-ordination of exo- and endocytosis during wound repair. Using cytoskeletal alterations and modulations of membrane tension and membrane area, membrane tension is identified as a major regulator of the wounding-associated exo- and endocytic events that mediate efficient wound repair. Thus, membrane tension changes are a universal trigger for plasma membrane wound repair modulating the exocytosis of early endosomes required for resealing and subsequent clathrin-mediated endocytosis acting at later stages to restore cell homeostasis and function.

Mechanosensitive fluorescence lifetime probes for investigating the dynamic mechanism of ferroptosis

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.

Clathrin-Mediated Endocytosis of Multiple Nanoparticles Tends to Be Less Cooperative: A Computational Study.

The internalization of nanoparticles is of great significance for their biological applications. Clathrin-mediated endocytosis (CME) is one of the main endocytic pathways. However, there is still a lack of a fundamental understanding regarding the internalization of multiple nanoparticles via CME. Therefore, in this study, we conducted computational investigations to uncover detailed molecular mechanisms and kinetic pathways for differently shaped nanoparticles in the presence of clathrin. Particular focus is given to understanding the CME of multiple-nanoparticle systems. We found that unlike receptor-mediated endocytosis, multiple nanoparticles did not get cooperatively wrapped by the membrane but tended to undergo independent endocytosis in the presence of clathrin. To further investigate the endocytosis mechanism, we studied the effects of clathrins, nanoparticle shape, nanoparticle size, nanoparticle arrangement, and membrane surface tension. The self-assembly of clathrin prefers independent endocytosis for multiple nanoparticles. Besides, the cooperative behavior is weak with increasing nanoparticle-shape anisotropy. However, when the membrane tension is reduced, the endocytosis pathway for multiple nanoparticles is cooperative endocytosis. Moreover, we found that the self-assembly of clathrins reduces the critical size of nanoparticles to undergo cooperative wrapping by the cell membrane. Our results provide valuable insights into the molecular mechanisms of multiple nanoparticles through CME and offer useful guidance for the design of nanoparticles as drug/gene delivery carriers.

High-frequency MHz-order vibration enables cell membrane remodeling and lipid microdomain manipulation

We elucidate the mechanism underpinning a recently discovered phenomenon in which cells respond to MHz-order mechanostimuli. Deformations induced along the plasma membrane under these external mechanical cues are observed to decrease the membrane tension, which, in turn, drives transient and reversible remodeling of its lipid structure. In particular, the increase and consequent coalescence of ordered lipid microdomains leads to closer proximity to mechanosensitive ion channels—Piezo1, in particular—that, due to crowding, results in their activation to mobilize influx of calcium (Ca2+) ions into the cell. It is the modulation of this second messenger that is responsible for the downstream signaling and cell fates that ensue. In addition, we show that such spatiotemporal control over the membrane microdomains in cells—without necessitating biochemical factors—facilitates aggregation and association of intrinsically disordered tau proteins in neuroblastoma cells, and their transformation to pathological conditions implicated in neurodegenerative diseases, thereby paving the way for the development of therapeutic intervention strategies.

Neutrophils exhibit flexible migration strategies and trail formation mechanisms on varying adhesive substrates

Substrate anchorage is essential for cell migration and actin polymerization at cell front and myosin contractility at cell rear are known to govern cell forward movement. Yet their differential driving strategies for neutrophil migration on distinct adhesiveness substrates and their contributions to the migration-induced trail formation remain unclear. Here we explore the morphological changes, migration dynamics, and trail formation of neutrophils on ICAM-1 and PLL substrates, with a focus on the relationships among adhesive forces, traction forces, and out-of-plane forces. Results indicate that, on ICAM-1, neutrophil migration and trail formation rely on the coordinated interactions of Arp2/3 and myosin, along with biochemical regulation (via Syk and calpain) of adhesion and de-adhesion. This pattern leads to traction forces being concentrated at relatively fewer adhesive sites, facilitating cell forward migration. On PLL, however, neutrophils primarily depend on Arp2/3-mediated actin polymerization, resulting in a broader distribution of traction forces and weaker adhesions, which allows for higher leading-edge migrating velocities. Elevated membrane tension and out-of-plane forces generated by bleb protrusions on PLL reduce the reliance on myosin-driven contraction at the trailing edge, enabling easier tail detachment through elastic recoil. This work highlights the differential impact of substrate adhesiveness on neutrophil migration and trail formation and dynamics, providing new insights into cell migration mechanisms and potential therapeutic targets for inflammatory and immune-related disorders.

Fission of double-membrane tubes under tension

The division of a cellular compartment culminates with the scission of a highly constricted membrane neck. Scission requires lipid rearrangements, topology changes, and transient formation of nonbilayer intermediate structures driven by curvature stress. Often, a side effect of this stress is pore formation, which may lead to content leakage and thus breaching of the membrane barrier function. In single-membrane systems, leakage is avoided through the formation of a hemifusion (HF) intermediate, whose structure is still a subject of debate. The consequences of curvature stress have not been explored in double-membrane systems, such as the mitochondrion. Here, we combine experimental and theoretical approaches to study neck constriction and scission driven by tension in biomimetic lipid systems, namely single- and double-membrane nanotubes (sNTs and dNTs), respectively. In sNTs, constriction by high tension gives rise to a metastable HF intermediate (seen as stalk or worm-like micelle), whereas poration is universally slower in a simple neck. In dNTs, high membrane tension causes sequential rupture of each membrane. In contrast, low tension leads to the HF of both membranes, which may lead to a leaky fusion pathway, or may progress to further fusion of the two membranes along a number of transformation pathways. These findings provide a new mechanistic basis for fundamental cellular processes.

Time-resolved proximity proteomics uncovers a membrane tension-sensitive caveolin-1 interactome at the rear of migrating cells.

Caveolae are small membrane pits with fundamental roles in mechanotransduction. Several studies have shown that caveolae flatten out in response to an increase in membrane tension, thereby acting as a mechanosensitive membrane reservoir that buffers acute mechanical stress. The dynamic assembly and disassembly of caveolae has 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 caveolae-mediated mechanotransduction is poorly understood. 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 disassembles the caveolar protein coat. Membrane tension also dislodges caveolin-1 from focal adhesion proteins and several mechanosensitive cortical actin regulators including filamins and cortactin. In addition, we present evidence that ROCK and the RhoGAP ARHGAP29 are associated with caveolin-1 in a membrane tension-dependent manner, and that ARHGAP29 regulates 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.

Structural Rearrangement of the AT1 Receptor Modulated by Membrane Thickness and Tension.

Membrane-embedded mechanosensitive (MS) proteins, including ion channels and G-protein coupled receptors (GPCRs), are essential for the transduction of external mechanical stimuli into biological signals. The angiotensin II type 1 (AT1) receptor plays many important roles in cardiovascular regulation and is associated with diseases such as hypertension and congestive heart failure. The membrane-mediated activation of the AT1 receptor is not well understood, despite this being one of the most widely studied GPCRs within the context of biased agonism. Here, we use extensive molecular dynamics (MD) simulations to characterize the effect of the local membrane environment on the activation of the AT1 receptor. We show that membrane thickness plays an important role in the stability of active and inactive states of the receptor, as well as the dynamic interchange between states. Furthermore, our simulation results show that membrane tension is effective in driving large-scale structural changes in the inactive state such as the outward movement of transmembrane helix 6 to stabilize intermediate active-like conformations. We conclude by comparing our simulation observations with AlphaFold 2 predictions, as a proxy to experimental structures, to provide a framework for how membrane mediated stimuli can facilitate activation of the AT1 receptor through the β-arrestin signaling pathway.

Effects of Normal and Lateral Electric Fields on Membrane Mechanical Properties.

As a core component of biological and synthetic membranes, lipid bilayers are key to compartmentalizing chemical processes. Bilayer morphology and mechanical properties are heavily influenced by electric fields, such as those caused by biological ion concentration gradients. We present atomistic simulations exploring the effects of electric fields applied normally and laterally to lipid bilayers. We find that normal fields decrease membrane tension, while lateral fields increase it. Free energy perturbation calculations indicate the importance of dipole-dipole interactions to these tension changes, especially for lateral fields. We additionally show that membrane area compressibilities can be related to their cohesive energies, allowing us to estimate changes in membrane bending rigidity under applied fields. We find that normal and lateral fields decrease and increase bending rigidity, respectively. These results point to the use of directed electric fields to locally control membrane stiffness, thereby modulating associated cellular processes.

Osmotic spawning vesicle.

We discovered a cascade vesicle division system driven by osmotic inflation. Binary giant unilamellar vesicles (GUVs) composed of sodium bis(2-ethylhexyl)sulfosuccinate (AOT) and cholesterol (Chol) were subjected to an osmotic pressure difference by encapsulating membrane-impermeable osmolytes (typically sucrose) in an external aqueous solution containing membrane-permeable osmolytes (typically fructose). This simple setup enabled the mother GUVs to repeatedly form small membrane buds and subsequently undergo divisions over several hundred seconds, resulting in the production of approximately 30-300 daughter GUVs from a single mother GUV. The observed morphological change of GUVs is well described by the mechanical balance between membrane bending, membrane tension, and osmotic pressure difference based on the spontaneous curvature model. This "osmotic spawning" behavior of GUVs does not rely on chemical reactions or functional macromolecules. Therefore, this cascade division system is compatible with various chemical systems and has the potential to implement proliferation ability in artificial cells, drug delivery systems, and protocells simply by modifying their membrane compartments and osmolytes.