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Matile Stefan - One of the best experts on this subject based on the ideXlab platform.

  • HydroFlipper Membrane Tension probes: imaging Membrane hydration and mechanical compression simultaneously in living cells
    'Royal Society of Chemistry (RSC)', 2022
    Co-Authors: Garcia José, Maillard, Jimmy Stéphane, Mercier Vincent, Roux Aurélien, Sakai Naomi, Fuerstenberg Alexandre, Roffay Chloé, López-andarias Javier, Matile Stefan
    Abstract:

    HydroFlippers are introduced as the first fluorescent Membrane Tension probes that report simultaneously on Membrane compression and hydration. The probe design is centered around a sensing cycle that couples the mechanical planarization of twisted push–pull fluorophores with the dynamic covalent hydration of their exocyclic acceptor. In FLIM images of living cells, Tension-induced deplanarization is reported as a decrease in fluorescence lifetime of the dehydrated mechanophore. Membrane hydration is reported as the ratio of the photon counts associated to the hydrated and dehydrated mechanophores in reconvoluted lifetime frequency histograms. Trends for Tension-induced decompression and hydration of cellular Membranes of interest (MOIs) covering plasma Membrane, lysosomes, mitochondria, ER, and Golgi are found not to be the same. Tension-induced changes in mechanical compression are rather independent of the nature of the MOI, while the responsiveness to changes in hydration are highly dependent on the intrinsic order of the MOI. These results confirm the mechanical planarization of push–pull probes in the ground state as most robust mechanism to routinely image Membrane Tension in living cells, while the availability of simultaneous information on Membrane hydration will open new perspectives in mechanobiology

  • Fluorescent Membrane Tension Probes for Early Endosomes
    'Wiley', 2022
    Co-Authors: Piazzolla Francesca, Mercier Vincent, Roux Aurélien, Sakai Naomi, Assies Léa, Matile Stefan
    Abstract:

    Fluorescent flipper probes have been introduced recently to image Membrane Tension in live cells, and strategies to target these probes to specific Membranes are emerging. In this context, early endosomes (EE) targeting without use of protein engineering is especially appealing because it translates into a fascinating transport problem. Weakly basic probes, commonly used to track inside acidic late endosomes and lysosomes, are poorly retained in EE because they are sufficiently neutralized in weakly acidic EE, thus able to diffuse out. Here, we disclose a rational strategy to target EE using a substituted benzylamine with higher pK a as a head group of the flipper probe. The resulting EE flippers are validated for preserved mechanosensitivity, ready for use in biology, particularly to elucidate the mechanics of endocytosis

  • Photocleavable Fluorescent Membrane Tension Probes: Fast Release with Spatiotemporal Control in Inner Leaflets of Plasma Membrane, Nuclear Envelope, and Secretory Pathway
    'Wiley', 2022
    Co-Authors: López-andarias Javier, Sakai Naomi, Assies Léa, Eblighatian, Krikor Bared, Pasquer, Quentin Titouan Loïc, Hoogendoorn Sascha, Matile Stefan
    Abstract:

    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

  • Mitochondrial Membrane Tension governs fission
    'Elsevier BV', 2021
    Co-Authors: Mahecic Dora, Colom Diego Adai, Roux Aurélien, Matile Stefan, Carlini Lina, Kleele Tatjana, Goujon Antoine, Manley Suliana
    Abstract:

    During mitochondrial fission, key molecular and cellular factors assemble on the outer mitochondrial Membrane, where they coordinate to generate constriction. Constriction sites can eventually divide or reverse upon disassembly of the machinery. However, a role for Membrane Tension in mitochondrial fission, although speculated, has remained undefined. We capture the dynamics of constricting mitochondria in mammalian cells using live-cell structured illumination microscopy (SIM). By analyzing the diameters of tubules that emerge from mitochondria and implementing a fluorescence lifetime-based mitochondrial Membrane Tension sensor, we discover that mitochondria are indeed under Tension. Under perturbations that reduce mitochondrial Tension, constrictions initiate at the same rate, but are less likely to divide. We propose a model based on our estimates of mitochondrial Membrane Tension and bending energy in living cells which accounts for the observed probability distribution for mitochondrial constrictions to divide

  • Mitochondrial Membrane Tension Governs Fission
    'Elsevier BV', 2021
    Co-Authors: Mahecic Dora, Colom Diego Adai, Roux Aurélien, Matile Stefan, Carlini Lina, Kleele Tatjana, Goujon Antoine, Manley Suliana
    Abstract:

    During mitochondrial fission, key molecular and cellular factors assemble on the outer mitochondrial Membrane, where they coordinate to generate constriction. Constriction sites can eventually divide or reverse upon disassembly of the machinery. However, a role for Membrane Tension in mitochondrial fission, although speculated, has remained undefined. We capture the dynamics of constricting mitochondria in mammalian cells using live-cell structured illumination microscopy (SIM). By analyzing the diameters of tubules that emerge from mitochondria and implementing a fluorescence lifetime-based mitochondrial Membrane Tension sensor, we discover that mitochondria are indeed under Tension. Under perturbations that reduce mitochondrial Tension, constrictions initiate at the same rate, but are less likely to divide. We propose a model based on our estimates of mitochondrial Membrane Tension and bending energy in living cells which accounts for the observed probability distribution for mitochondrial constrictions to divide.This work was supported in part by the National Centre of Competence in Research Chemical Biology (S. Manley, S. Matile, and A.R.). S. Manley also acknowledges SNSF Project Grant 31003A_182429 (to T.K. and D.M). T.K. received funding from the European Molecular Biology Organization (ALTF-739-2016) and the Munich Cluster for Systems Neurology (SyNergy). A.C. received funding from MCIU, MINECO G19/P66, RYC-18/02, and T1270-1

Charles A Coomer - One of the best experts on this subject based on the ideXlab platform.

  • single cell glycolytic activity regulates Membrane Tension and hiv 1 fusion
    PLOS Pathogens, 2020
    Co-Authors: Charles A Coomer, Irene Carlonandres, Maro Iliopoulou, Michael L Dustin, Ewoud B Compeer, Alex A Compton, Sergi Padillaparra
    Abstract:

    There has been resurgence in determining the role of host metabolism in viral infection yet deciphering how the metabolic state of single cells affects viral entry and fusion remains unknown. Here, we have developed a novel assay multiplexing genetically-encoded biosensors with single virus tracking (SVT) to evaluate the influence of global metabolic processes on the success rate of virus entry in single cells. We found that cells with a lower ATP:ADP ratio prior to virus addition were less permissive to virus fusion and infection. These results indicated a relationship between host metabolic state and the likelihood for virus-cell fusion to occur. SVT revealed that HIV-1 virions were arrested at hemifusion in glycolytically-inactive cells. Interestingly, cells acutely treated with glycolysis inhibitor 2-deoxyglucose (2-DG) become resistant to virus infection and also display less surface Membrane cholesterol. Addition of cholesterol in these in glycolytically-inactive cells rescued the virus entry block at hemifusion and enabled completion of HIV-1 fusion. Further investigation with FRET-based Membrane Tension and Membrane order reporters revealed a link between host cell glycolytic activity and host Membrane order and Tension. Indeed, cells treated with 2-DG possessed lower plasma Membrane lipid order and higher Tension values, respectively. Our novel imaging approach that combines lifetime imaging (FLIM) and SVT revealed not only changes in plasma Membrane Tension at the point of viral fusion, but also that HIV is less likely to enter cells at areas of higher Membrane Tension. We therefore have identified a connection between host cell glycolytic activity and Membrane Tension that influences HIV-1 fusion in real-time at the single-virus fusion level in live cells.

  • single cell glycolytic activity regulates Membrane Tension and hiv 1 fusion
    bioRxiv, 2019
    Co-Authors: Charles A Coomer, Irene Carlonandres, Maro Iliopoulou, Michael L Dustin, Ewoud B Compeer, Alex A Compton, Sergi Padillaparra
    Abstract:

    ABSTRACT There has been resurgence in determining the role of host metabolism in viral infection yet deciphering how the metabolic state of single cells affects viral entry and fusion remains unknown. Here, we have developed a novel assay multiplexing genetically encoded biosensors with single virus tracking (SVT) to evaluate the influence of global metabolic processes on the success rate of virus entry in single cells. We found that cells with a lower ATP:ADP ratio prior to virus addition were less permissive to virus fusion and infection. These results indicated a relationship between host metabolic state and the likelihood for virus-cell fusion to occur. SVT revealed that HIV-1 viruses were arrested at hemifusion in glycolytically-inactive cells. Interestingly, cells acutely treated with glycolysis inhibitor 2-deoxyglucose (2-DG) become resistant to virus infection and also display less surface Membrane cholesterol. Addition of cholesterol in these in glycolytically-inactive cells rescued the virus entry block at hemifusion and enabled completion of HIV-1 fusion. Further investigation with FRET-based Membrane Tension and Membrane-order reporters revealed a link between host cell glycolytic activity and host Membrane order and Tension. Indeed, cells treated with 2-DG possessed lower plasma Membrane lipid order and higher Tension values, respectively. Our novel imaging approach that combines lifetime imaging (FLIM) and SVT revealed not only changes in plasma Membrane Tension at the point of viral fusion, but also that HIV is less likely to enter cells at areas of higher Membrane Tension. We therefore have identified a connection between host cell glycolytic activity and Membrane Tension that influences HIV-1 fusion in real-time at the single-virus fusion level in live cells. As glycolytic activity sets Membrane Tension levels by altering cellular cholesterol surface levels, our results suggest additional previously unknown benefits of cholesterol-lowering medication in HIV-1 infection.

Michael P Sheetz - One of the best experts on this subject based on the ideXlab platform.

  • mechanical feedback between Membrane Tension and dynamics
    Trends in Cell Biology, 2012
    Co-Authors: Nils C Gauthier, Michael P Sheetz, Thomas A Masters
    Abstract:

    The plasma Membrane represents a physical inelastic barrier with a given area that adheres to the underlying cytoskeleton. The Tension in the Membrane physically affects cell functions and recent studies have highlighted that this physical signal orchestrates complex aspects of trafficking and motility. Despite its undeniable importance, little is known about the mechanisms by which Membrane Tension regulates cell functions or stimulates signals. The maintenance of Membrane Tension is also a matter of debate, particularly the nature of the Membrane reservoir and trafficking pathways that buffer Tension. In this review we discuss the importance of Membrane area and of Tension as a master integrator of cell functions, particularly for Membrane traffic.

  • temporary increase in plasma Membrane Tension coordinates the activation of exocytosis and contraction during cell spreading
    Proceedings of the National Academy of Sciences of the United States of America, 2011
    Co-Authors: Nils C Gauthier, Michael P Sheetz, Marcantoine Fardin, Pere Rocacusachs
    Abstract:

    Cell migration and spreading involve the coordination of Membrane trafficking, actomyosin contraction, and modifications to plasma Membrane Tension and area. The biochemical or biophysical basis for this coordination is however unknown. In this study, we show that during cell spreading, lamellipodia protrusion flattens plasma Membrane folds and blebs and, once the plasma Membrane area is depleted, there is a temporary increase in Membrane Tension by over twofold that is followed by activation of exocytosis and myosin contraction. Further, an artificial increase in plasma Membrane Tension stopped lamellipodia protrusion and activated an exocytotic burst. Subsequent decrease in Tension restored spreading with activation of contraction. Conversely, blebbistatin inhibition of actomyosin contraction resulted in an even greater increase in plasma Membrane Tension and exocytosis activation. This spatiotemporal synchronization indicates that Membrane Tension is the signal that coordinates Membrane trafficking, actomyosin contraction, and plasma Membrane area change. We suggest that cells use plasma Membrane Tension as a global physical parameter to control cell motility.

  • cell spreading and lamellipodial exTension rate is regulated by Membrane Tension
    Journal of Cell Biology, 2000
    Co-Authors: Drazen Raucher, Michael P Sheetz
    Abstract:

    Cell spreading and motility require the exTension of the plasma Membrane in association with the assembly of actin. In vitro, exTension must overcome resistance from Tension within the plasma Membrane. We report here that the addition of either amphiphilic compounds or fluorescent lipids that expanded the plasma Membrane increased the rate of cell spreading and lamellipodial exTension, stimulated new lamellipodial exTensions, and caused a decrease in the apparent Membrane Tension. Further, in PDGF-stimulated motility, the increase in the lamellipodial exTension rate was associated with a decrease in the apparent Membrane Tension and decreased Membrane–cytoskeleton adhesion through phosphatidylinositol diphosphate hydrolysis. Conversely, when Membrane Tension was increased by osmotically swelling cells, the exTension rate decreased. Therefore, we suggest that the lamellipodial exTension process can be activated by a physical signal (perhaps secondarily), and the rate of exTension is directly dependent upon the Tension in the plasma Membrane. Quantitative analysis shows that the lamellipodial exTension rate is inversely correlated with the apparent Membrane Tension. These studies describe a physical chemical mechanism involving changes in Membrane–cytoskeleton adhesion through phosphatidylinositol 4,5-biphosphate–protein interactions for modulating and stimulating the biochemical processes that power lamellipodial exTension.

  • Membrane tether formation from blebbing cells
    Biophysical Journal, 1999
    Co-Authors: Jianwu Dai, Michael P Sheetz
    Abstract:

    Membrane Tension has been proposed to be important in regulating cell functions such as endocytosis and cell motility. The apparent Membrane Tension has been calculated from tether forces measured with laser tweezers. Both Membrane-cytoskeleton adhesion and Membrane Tension contribute to the tether force. Separation of the plasma Membrane from the cytoskeleton occurs in Membrane blebs, which could remove the Membrane-cytoskeleton adhesion term. In renal epithelial cells, tether forces are significantly lower on blebs than on Membranes that are supported by cytoskeleton. Furthermore, the tether forces are equal on apical and basolateral blebs. In contrast, tether forces from Membranes supported by the cytoskeleton are greater in apical than in basolateral regions, which is consistent with the greater apparent cytoskeletal density in the apical region. We suggest that the tether force on blebs primarily contains only the Membrane Tension term and that the Membrane Tension may be uniform over the cell surface. Additional support for this hypothesis comes from observations of melanoma cells that spontaneously bleb. In melanoma cells, tether forces on blebs are proportional to the radius of the bleb, and as large blebs form, there are spikes in the tether force in other cell regions. We suggest that an internal osmotic pressure inflates the blebs, and the pressure calculated from the Law of Laplace is similar to independent measurements of intracellular pressures. When the Membrane Tension term is subtracted from the apparent Membrane Tension over the cytoskeleton, the Membrane-cytoskeleton adhesion term can be estimated. In both cell systems, Membrane-cytoskeleton adhesion was the major factor in generating the tether force.

  • Membrane expansion increases endocytosis rate during mitosis
    Journal of Cell Biology, 1999
    Co-Authors: Drazen Raucher, Michael P Sheetz
    Abstract:

    Mitosis in mammalian cells is accompanied by a dramatic inhibition of endocytosis. We have found that the addition of amphyphilic compounds to metaphase cells increases the endocytosis rate even to interphase levels. Detergents and solvents all increased endocytosis rate, and the extent of increase was in direct proportion to the concentration added. Although the compounds could produce a variety of different effects, we have found a strong correlation with a physical alteration in the Membrane Tension as measured by the laser tweezers. Plasma Membrane tethers formed by latex beads pull back on the beads with a force that was related to the in-plane bilayer Tension and Membrane– cytoskeletal adhesion. We found that as cells enter mitosis, the Membrane Tension rises as the endocytosis rate decreases; and as cells exited mitosis, the endocytosis rate increased as the Membrane Tension decreased. The addition of amphyphilic compounds decreased Membrane Tension and increased the endocytosis rate. With the detergent, deoxycholate, the endocytosis rate was restored to interphase levels when the Membrane Tension was restored to interphase levels. Although biochemical factors are clearly involved in the alterations in mitosis, we suggest that endocytosis is blocked primarily by the increase in apparent plasma Membrane Tension. Higher Tensions inhibit both the binding of the endocytic complex to the Membrane and mechanical deformation of the Membrane during invagination. We suggest that Membrane Tension is an important regulator of the endocytosis rate and alteration of Tension is sufficient to modify endocytosis rates during mitosis. Further, we postulate that the rise in Membrane Tension causes cell rounding and the inhibition of motility, characteristic of mitosis.

Stefan Matile - One of the best experts on this subject based on the ideXlab platform.

  • mechanosensitive fluorescent probes to image Membrane Tension in mitochondria endoplasmic reticulum and lysosomes
    Journal of the American Chemical Society, 2019
    Co-Authors: Antoine Goujon, Suliana Manley, Aurelien Roux, Adai Colom, Karolina Strakova, Vincent Mercier, Dora Mahecic, Naomi Sakai, Stefan Matile
    Abstract:

    Measuring forces inside cells is particularly challenging. With the development of quantitative microscopy, fluorophores which allow the measurement of forces became highly desirable. We have previously introduced a mechanosensitive flipper probe, which responds to the change of plasma Membrane Tension by changing its fluorescence lifetime and thus allows Tension imaging by FLIM. Herein, we describe the design, synthesis, and evaluation of flipper probes that selectively label intracellular organelles, i.e., lysosomes, mitochondria, and the endoplasmic reticulum. The probes respond uniformly to osmotic shocks applied extracellularly, thus confirming sensitivity toward changes in Membrane Tension. At rest, different lifetimes found for different organelles relate to known differences in Membrane organization rather than Membrane Tension and allow colabeling in the same cells. At the organelle scale, lifetime heterogeneity provides unprecedented insights on ER tubules and sheets, and nuclear Membranes. Exam...

  • mechanosensitive fluorescent probes to image Membrane Tension in mitochondria endoplasmic reticulum and lysosomes
    Journal of the American Chemical Society, 2019
    Co-Authors: Antoine Goujon, Suliana Manley, Aurelien Roux, Adai Colom, Karolina Strakova, Vincent Mercier, Dora Mahecic, Naomi Sakai, Stefan Matile
    Abstract:

    Measuring forces inside cells is particularly challenging. With the development of quantitative microscopy, fluorophores which allow the measurement of forces became highly desirable. We have previously introduced a mechanosensitive flipper probe, which responds to the change of plasma Membrane Tension by changing its fluorescence lifetime and thus allows Tension imaging by FLIM. Herein, we describe the design, synthesis, and evaluation of flipper probes that selectively label intracellular organelles, i.e., lysosomes, mitochondria, and the endoplasmic reticulum. The probes respond uniformly to osmotic shocks applied extracellularly, thus confirming sensitivity toward changes in Membrane Tension. At rest, different lifetimes found for different organelles relate to known differences in Membrane organization rather than Membrane Tension and allow colabeling in the same cells. At the organelle scale, lifetime heterogeneity provides unprecedented insights on ER tubules and sheets, and nuclear Membranes. Examples on endosomal trafficking or increase of Tension at mitochondrial constriction sites outline the potential of intracellularly targeted fluorescent Tension probes to address essential questions that were previously beyond reach.

Andreas Janshoff - One of the best experts on this subject based on the ideXlab platform.

  • Membrane Tension increases fusion efficiency of model Membranes in the presence of SNAREs
    Scientific Reports, 2017
    Co-Authors: Torben-tobias Kliesch, Jörn Dietz, Laura Turco, Partho Halder, Elena Polo, Marco Tarantola, Andreas Janshoff
    Abstract:

    The large gap in time scales between Membrane fusion occurring in biological systems during neurotransmitter release and fusion observed between model Membranes has provoked speculations over a large number of possible factors that might explain this discrepancy. One possible reason is an elevated lateral Membrane Tension present in the presynaptic Membrane. We investigated the Tension-dependency of fusion using model Membranes equipped with a minimal fusion machinery consisting of syntaxin 1, synaptobrevin and SNAP 25. Two different strategies were realized; one based on supported bilayers and the other one employing sessile giant liposomes. In the first approach, isolated patches of planar bilayers derived from giant unilamellar vesicles containing syntaxin 1 and preassembled SNAP 25 (ΔN-complex) were deposited on a dilatable PDMS sheet. In a second approach, lateral Membrane Tension was controlled through the adhesion of intact giant unilamellar vesicles on a functionalized surface. In both approaches fusion efficiency increases considerably with lateral Tension and we identified a threshold Tension of 3.4 mN m^−1, at which the number of fusion events is increased substantially.

  • epsin n terminal homology domain enth activity as a function of Membrane Tension
    Journal of Biological Chemistry, 2016
    Co-Authors: Martin Gleisner, Andreas Janshoff, Torben-tobias Kliesch, Benjamin Kroppen, Christian Fricke, Nelli Teske, Michael Meinecke, Claudia Steinem
    Abstract:

    The epsin N-terminal homology domain (ENTH) is a major player in clathrin-mediated endocytosis. To investigate the influence of initial Membrane Tension on ENTH binding and activity, we established a bilayer system based on adhered giant unilamellar vesicles (GUVs) to be able to control and adjust the Membrane Tension σ covering a broad regime. The shape of each individual adhered GUV as well as its adhesion area was monitored by spinning disc confocal laser microscopy. Control of σ in a range of 0.08-1.02 mN/m was achieved by altering the Mg(2+) concentration in solution, which changes the surface adhesion energy per unit area of the GUVs. Specific binding of ENTH to phosphatidylinositol 4,5-bisphosphate leads to a substantial increase in adhesion area of the sessile GUV. At low Tension (<0.1 mN/m) binding of ENTH can induce tubular structures, whereas at higher Membrane Tension the ENTH interaction deflates the sessile GUV and thereby increases the adhesion area. The increase in adhesion area is mainly attributed to a decrease in the area compressibility modulus KA We propose that the insertion of the ENTH helix-0 into the Membrane is largely responsible for the observed decrease in KA, which is supported by the observation that the mutant ENTH L6E shows a reduced increase in adhesion area. These results demonstrate that even in the absence of tubule formation, the area compressibility modulus and, as such, the bending rigidity of the Membrane is considerably reduced upon ENTH binding. This renders Membrane bending and tubule formation energetically less costly.

  • ezrin is a major regulator of Membrane Tension in epithelial cells
    Scientific Reports, 2015
    Co-Authors: Bastian Rouven Bruckner, Anna Pietuch, Stefan Nehls, Jan Rother, Andreas Janshoff
    Abstract:

    Plasma Membrane Tension is responsible for a variety of cellular functions such as motility, cell division, and endocytosis. Since Membrane Tension is dominated by the attachment of the actin cortex to the inner leaflet of the plasma Membrane, we investigated the importance of ezrin, a major cross-linker of the Membrane-cytoskeleton interface, for cellular mechanics of confluent MDCK II cells. For this purpose, we carried out ezrin depletion experiments and also enhanced the number of active ezrin molecules at the interface. Mechanical properties were assessed by force indentation experiments followed by Membrane tether extraction. PIP2 micelles were injected into individual living cells to reinforce the linkage between plasma Membrane and actin-cortex, while weakening of this connection was reached by ezrin siRNA and administration of the inhibitors neomycin and NSC 668394, respectively. We observed substantial stiffening of cells and an increase in Membrane Tension after addition of PIP2 micelles. In contrast, reduction of active ezrin led to a decrease of Membrane Tension accompanied by loss of excess surface area, increase in cortical Tension, remodelling of actin cytoskeleton, and reduction of cell height. The data confirm the importance of the ezrin-mediated connection between plasma Membrane and cortex for cellular mechanics and cell morphology.

  • Membrane Tension homeostasis of epithelial cells through surface area regulation in response to osmotic stress
    Biochimica et Biophysica Acta, 2013
    Co-Authors: Anna Pietuch, Bastian Rouven Bruckner, Andreas Janshoff
    Abstract:

    Abstract Osmotic stress poses one of the most fundamental challenges to living cells. Particularly, the largely inextensible plasma Membrane of eukaryotic cells easily ruptures under in-plane Tension calling for sophisticated strategies to readily respond to osmotic stress. We describe how epithelial cells react and adapt mechanically to the exposure to hypotonic and hypertonic solutions in the context of a confluent monolayer. Site-specific indentation experiments in conjunction with tether pulling on individual cells have been carried out with an atomic force microscope to reveal spatio-temporal changes in Membrane Tension and surface area. We found that cells compensate for an increase in lateral Tension due to hypoosmotic stress by sacrificing excess of Membrane area stored in protrusions and invaginations such as microvilli and caveolae. At mild hypotonic conditions lateral Tension increases partly compensated by surface are regulation, i.e. the cell sacrifices some of its Membrane reservoirs. A loss of Membrane–actin contacts occurs upon exposure to stronger hypotonic solutions giving rise to a drop in lateral Tension. Tension release recovers on longer time scales by an increasing endocytosis, which efficiently removes excess Membrane from the apical side to restore the initial pre-stress. Hypertonic solutions lead to shrinkage of cells and collapse of the apical Membrane onto the cortex. Exposure to distilled water leads to stiffening of cells due to removal of excess surface area and Tension increase due to elevated osmotic pressure across the plasma Membrane.