Giant Unilamellar Vesicles Research Articles

Benefits of Combined Fluorescence Lifetime Imaging Microscopy and Fluorescence Correlation Spectroscopy for Biomedical Studies Demonstrated by Using a Liposome Model System

Drug delivery systems play a pivotal role in targeted pharmaceutical transport and controlled release at specific sites. Liposomes, commonly used as drug carriers, constitute a fundamental part of these systems. Moreover, the drug–liposome model serves as a robust platform for investigating interaction processes at both cellular and molecular levels. To advance our understanding of drug carrier uptake mechanisms, we employed fluorescence lifetime imaging microscopy (FLIM) and fluorescence correlation spectroscopy (FCS), leveraging the unique benefits of two-photon (2P) excitation. Our approach utilized giant unilamellar vesicles (GUVs) as a simplified model system for cell membranes, labelled with the amphiphilic fluorescent dye 3,3′-dioctadecyloxa-carbocyanine (DiOC18(3)). Additionally, large unilamellar vesicles (LUVs) functioned as a drug carrier system, incorporating the spectrally distinct fluorescent sulforhodamine 101 (SRh101) as a surrogate drug. The investigation emphasized the diverse interactions between GUVs and LUVs based on the charged lipids employed. We examined the exchange kinetics and structural alterations of liposome carriers during the uptake process. Our study underscores the significance of employing 2P excitation in conjunction with FLIM and FCS. This powerful combination offers a valuable methodological approach for studying liposome interactions, positioning them as an exceptionally versatile model system with a distinct technical advantage.

Influence of pH and lipid membrane on the liquid–liquid phase separation of wheat γ-gliadin in aqueous conditions

Protein body (PB) formation in wheat seeds is a critical process influencing seed content and nutritional quality. In this study, we investigate the potential mechanisms governing PB formation through an in vitro approach, focusing on γ-gliadin, a key wheat storage protein. We used a microfluidic technique to encapsulate γ-gliadin within giant unilamellar vesicles (GUVs) and tune the physicochemical conditions in a controlled and rapid way. We examined the influence of pH and protein concentration on LLPS and protein-membrane interactions using various microscopy and spectroscopy techniques. We showed that γ-gliadin encapsulated in GUVs can undergo a pH-triggered liquid–liquid phase separation (LLPS) by two distinct mechanisms depending on the γ-gliadin concentration. At low protein concentrations, γ-gliadins phase separate by a nucleation and growth-like process, while, at higher protein concentration and pH above 6.0, γ-gliadin formed a bi-continuous phase suggesting a spinodal decomposition-like mechanism. Fluorescence and microscopy data suggested that γ-gliadin dense phase exhibited affinity for the GUV membrane, forming a layer at the interface and affecting the reversibility of the phase separation.

Glycerophospholipid polyunsaturation modulates resveratrol action on biomimetic membranes

The phytoalexin resveratrol has received increasing attention for its potential to prevent oxidative damages in human organism. To shed further light on molecular mechanisms of its interaction with lipid membranes we study resveratrol influence on the organisation and mechanical properties of biomimetic lipid systems composed of synthetic phosphatidylcholines with mixed aliphatic chains and different degree of unsaturation at sn-2 position (1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine, POPC, and 1-palmitoyl-2-docosahexaenoyl-sn-glycero-3-phosphocholine, PDPC). High-sensitivity isothermal titration calorimetric measurements reveal stronger spontaneous resveratrol association to polyunsaturated phosphatidylcholine bilayers compared to the monounsaturated ones resulting from hydrophobic interactions, conformational changes of the interacting species and desolvation of molecular surfaces. The latter is supported by the results from Laurdan spectroscopy of large unilamellar vesicles providing data on hydration at the glycerol backbones of glycerophospholipides. Higher degree of lipid order is reported for POPC membranes compared to PDPC. While resveratrol mostly enhances the hydration of PDPC membranes, increasing POPC dehydration is reported upon treatment with the polyphenol. Dehydration of the polyunsaturated lipid bilayers is measured only at the highest phytoalexin content studied (resveratrol/lipid 0.5 mol/mol) and is less pronounced than the effect reported for POPC membranes. The polyphenol effect on membrane mechanics is probed by thermal shape fluctuation analysis of quasispherical giant unilamellar vesicles. Markedly different trend of the bending elasticity with increasing resveratrol concentration is reported for the two types of phospholipid bilayers studied. POPC membranes become more rigid in the presence of resveratrol, whereas PDPC-containing bilayers exhibit softening at lower concentrations of the polyphenol followed by a slight growth without bilayer stiffening even at the highest resveratrol content explored. The new data on the structural organization and membrane properties of resveratrol-treated phosphatidylcholine membranes may underpin the development of future liposomal applications of the polyphenol in medicinal chemistry.

Biomimetic Vesicles with Designer Phospholipids Can Sense Environmental Redox Cues.

Cell-like materials that sense environmental cues can serve as next-generation biosensors and help advance the understanding of intercellular communication. Currently, bottom-up engineering of protocell models from molecular building blocks remains a grand challenge chemists face. Herein, we describe giant unilamellar vesicles (GUVs) with biomimetic lipid membranes capable of sensing environmental redox cues. The GUVs employ activity-based sensing through designer phospholipids that are fluorescently activated in response to specific reductive (hydrogen sulfide) or oxidative (hydrogen peroxide) conditions. These synthetic phospholipids are derived from 1,2-dipalmitoyl-rac-glycero-3-phosphocholine and they possess a headgroup with heterocyclic aromatic motifs. Despite their structural deviation from the phosphocholine headgroup, the designer phospholipids (0.5-1.0 mol %) mixed with natural lipids can vesiculate, and the resulting GUVs (7-20 μm in diameter) remain intact over the course of redox sensing. All-atom molecular dynamics simulations gave insight into how these lipids are positioned within the hydrophobic core of the membrane bilayer and at the membrane-water interface. This work provides a purely chemical method to investigate potential redox signaling and opens up new design opportunities for soft materials that mimic protocells.

Tuning the double lipidation of salmon calcitonin to introduce a pore-like membrane translocation mechanism

A widespread strategy to increase the transport of therapeutic peptides across cellular membranes has been to attach lipid moieties to the peptide backbone (lipidation) to enhance their intrinsic membrane interaction. Efforts in vitro and in vivo investigating the correlation between lipidation characteristics and peptide membrane translocation efficiency have traditionally relied on end-point read-out assays and trial-and-error-based optimization strategies. Consequently, the molecular details of how therapeutic peptide lipidation affects it’s membrane permeation and translocation mechanisms remain unresolved. Here we employed salmon calcitonin as a model therapeutic peptide and synthesized nine double lipidated analogs with varying lipid chain lengths. We used single giant unilamellar vesicle (GUV) calcein influx time-lapse fluorescence microscopy to determine how tuning the lipidation length can lead to an All-or-None GUV filling mechanism, indicative of a peptide mediated pore formation. Finally, we used a GUVs-containing-inner-GUVs assay to demonstrate that only peptide analogs capable of inducing pore formation show efficient membrane translocation. Our data provided the first mechanistic details on how therapeutic peptide lipidation affects their membrane perturbation mechanism and demonstrated that fine-tuning lipidation parameters could induce an intrinsic pore-forming capability. These insights and the microscopy based workflow introduced for investigating structure–function relations could be pivotal for optimizing future peptide design strategies.

Construction of Out-of-Equilibrium Metabolic Networks in Nano- and Micrometer-Sized Vesicles.

We present a method to incorporate into vesicles complex protein networks, involving integral membrane proteins, enzymes, and fluorescence-based sensors, using purified components. This method is relevant for the design and construction of bioreactors and the study of complex out-of-equilibrium metabolic reaction networks. We start by reconstituting (multiple) membrane proteins into large unilamellar vesicles (LUVs) according to a previously developed protocol. We then encapsulate a mixture of purified enzymes, metabolites, and fluorescence-based sensors (fluorescent proteins or dyes) via freeze-thaw-extrusion and remove non-incorporated components by centrifugation and/or size-exclusion chromatography. The performance of the metabolic networks is measured in real time by monitoring the ATP/ADP ratio, metabolite concentration, internal pH, or other parameters by fluorescence readout. Our membrane protein-containing vesicles of 100-400 nm diameter can be converted into giant-unilamellar vesicles (GUVs), using existing but optimized procedures. The approach enables the inclusion of soluble components (enzymes, metabolites, sensors) into micrometer-size vesicles, thus upscaling the volume of the bioreactors by orders of magnitude. The metabolic network containing GUVs aretrapped in microfluidic devices for analysis by optical microscopy.

Molecular Interactions Between Egg White Peptides and Giant Unilamellar Vesicle Membranes: Effect of Peptide Localization on Membrane Fluidity.

Bioactive peptides have been shown to affect cell membrane fluidity, which is an important indicator of the cell membrane structure and function. However, the underlying mechanism of egg white-derived bioactive peptide regulation of cell membrane fluidity has not been elucidated yet. The cell membrane fluidity was investigated by giant unilamellar vesicles in the present study. The results showed that peptides TCNW, ADWAK, ESIINF, VPIEGII, LVEEY, and WKLC connect to membranes through intermolecular interactions, such as hydrogen bonding and regulated membrane fluidity, in a concentration-dependent way. In addition, peptides prefer to localize in the hydrophobic core of the bilayers. This study provides a theoretical basis for analyzing the localization of egg white bioactive peptides in specific cell membrane regions and their influence on the cell membrane fluidity.

Assessing the negative impact of chlorantraniliprole, isoxaflutole, and simazine pesticides on phospholipid membrane models and tilapia gill tissues

The indiscriminate and, very often, incorrect use of pesticides in Brazil, as well as in other countries, results in severe levels of environmental pollution and intoxication of human life. Herein, we studied plasma membrane models (monolayer and bilayer) of the phospholipid Dioleoyl-sn-glycerol-3-phosphocholine (DOPC) using Langmuir films, and large (LUVs) and giant (GUVs) unilamellar vesicles, to determine the effect of the pesticides chlorantraniliprole (CLTP), isoxaflutole (ISF), and simazine (SMZ), used in sugarcane. CLTP affects the lipid organization of the bioinspired models of DOPC π-A isotherms, while ISF and SMZ pesticides significantly affect the LUVs and GUVs. Furthermore, the in vivo study of the gill tissue in fish in the presence of pesticides (2.0 × 10−10 mol/L for CLTP, 8.3 × 10−9 mol/L for ISF, and SMZ at 9.9 × 10−9 mol/L) was performed using optical and fluorescence images. This investigation was motivated by the gill lipid membranes, which are vital for regulating transporter activity through transmembrane proteins, crucial for maintaining ionic balance in fish gills. In this way, the presence of phospholipids in gills offers a model for understanding their effects on fish health. Histological results show that exposure to CLTP, ISF, and SMZ may interfere with vital gill functions, leading to respiratory disorders and osmoregulation dysfunction. The results indicate that exposure to pesticides caused severe morphological alterations in fish, which could be correlated with their impact on the bioinspired membrane models. Moreover, the effect does not depend on the exposure period (24h and 96h), showing that animals exposed to pesticides for a short period suffer irreparable damage to gill tissue. In summary, we can conclude that the harm caused by pesticides, both in membrane models and in fish gills, occurs due to contamination of the aquatic system with pesticides. Therefore, water quality is vital for the preservation of ecosystems.

Melittin-Phospholipase A2 Synergism Is Mediated by Liquid-Liquid Miscibility Phase Transition in Giant Unilamellar Vesicles.

The primary constituents of honeybee venom, melittin and phospholipase A2 (PLA2), display toxin synergism in which the PLA2 activity is significantly enhanced by the presence of melittin. It has been shown previously that this is accomplished by the disruption in lipid packing, which allows PLA2 to become processive on the membrane surface. In this work, we show that melittin is capable of driving miscibility phase transition in giant unilamellar vesicles (GUVs) and that it raises the miscibility transition temperature (Tmisc) in a concentration-dependent manner. The induced phase separation enhances the processivity of PLA2, particularly at its boundaries, where a substantial difference in domain thickness creates a membrane discontinuity. The catalytic action of PLA2, in response, induces changes in the membrane, rendering it more conducive to melittin binding. This, in turn, facilitates further lipid phase separation and eventual vesicle lysis. Overall, our results show that melittin has powerful membrane-altering capabilities that activate PLA2 in various membrane contexts. More broadly, they exemplify how this biochemical system actively modulates and capitalizes on the spatial distribution of membrane lipids to efficiently achieve its objectives.

Electroformation of Giant Unilamellar Vesicles from Damp Lipid Films with a Focus on Vesicles with High Cholesterol Content.

Giant unilamellar vesicles (GUVs) are membrane models used to study membrane properties. Electroformation is one of the methods used to produce GUVs. During electroformation protocol, dry lipid film is formed. The drying of the lipid film induces the cholesterol (Chol) demixing artifact, in which Chol forms anhydrous crystals which do not participate in the formation of vesicles. This leads to a lower Chol concentration in the vesicle bilayers compared to the Chol concentration in the initial lipid solution. To address this problem, we propose a novel electroformation protocol that includes rapid solvent exchange (RSE), plasma cleaning, and spin-coating methods to produce GUVs. We tested the protocol, focusing on vesicles with a high Chol content using different spin-coating durations and vesicle type deposition. Additionally, we compared the novel protocol using completely dry lipid film. The optimal spin-coating duration for vesicles created from the phosphatidylcholine/Chol mixture was 30 s. Multilamellar vesicles (MLVs), large unilamellar vesicles (LUVs) obtained by the extrusion of MLVs through 100 nm membrane pores and LUVs obtained by extrusion of previously obtained LUVs through 50 nm membrane pores, were deposited on an electrode for 1.5/1 Chol/phosphatidylcholine (POPC) lipid mixture, and the results were compared. Electroformation using all three deposited vesicle types resulted in a high GUV yield, but the deposition of LUVs obtained by the extrusion of MLVs through 100 nm membrane pores provided the most reproducible results. Using the deposition of these LUVs, we produced high yield GUVs for six different Chol concentrations (from 0% to 71.4%). Using a protocol that included dry lipid film GUVs resulted in lower yields and induced the Chol demixing artifact, proving that the lipid film should never be subjected to drying when the Chol content is high.

Role of Shape in Particle-Lipid Membrane Interactions: From Surfing to Full Engulfment.

Understanding and manipulating the interactions between foreign bodies and cell membranes during endo- and phagocytosis is of paramount importance, not only for the fate of living cells but also for numerous biomedical applications. This study aims to elucidate the role of variables such as anisotropic particle shape, curvature, orientation, membrane tension, and adhesive strength in this essential process using a minimal experimental biomimetic system comprising giant unilamellar vesicles and rod-like particles with different curvatures and aspect ratios. We find that the particle wrapping process is dictated by the balance between the elastic free energy penalty and adhesion free energy gain, leading to two distinct engulfment pathways, tip-first and side-first, emphasizing the significance of the particle orientation in determining the pathway. Moreover, our experimental results are consistent with theoretical predictions in a state diagram, showcasing how to control the wrapping pathway from surfing to partial to complete wrapping by the interplay between membrane tension and adhesive strength. At moderate particle concentrations, we observed the formation of rod clusters, which exhibited cooperative and sequential wrapping. Our study contributes to a comprehensive understanding of the mechanistic intricacies of endocytosis by highlighting how the interplay between the anisotropic particle shape, curvature, orientation, membrane tension, and adhesive strength can influence the engulfment pathway.

Reconstitution of the Bacterial Glutamate Receptor Channel by Encapsulation of a Cell-Free Expression System.

Cell-free expression (CFE) systems are powerful tools in synthetic biology that allow biomimicry of cellular functions like biosensing and energy regeneration in synthetic cells. Reconstruction of a wide range of cellular processes, however, requires successful reconstitution of membrane proteins into the membrane of synthetic cells. While the expression of soluble proteins is usually successful in common CFE systems, the reconstitution of membrane proteins in lipid bilayers of synthetic cells has proven to be challenging. Here, a method for reconstitution of a model membrane protein, bacterial glutamate receptor (GluR0), in giant unilamellar vesicles (GUVs) as model synthetic cells based on encapsulation and incubation of the CFE reaction inside synthetic cells is demonstrated. Utilizing this platform, the effect of substituting the N-terminal signal peptide of GluR0 with proteorhodopsin signal peptide on successful cotranslational translocation of GluR0 into membranes of hybrid GUVs is demonstrated. This method provides a robust procedure that will allow cell-free reconstitution of various membrane proteins in synthetic cells.

Effect of Phosphatidylethanolamine on Pore Formation Induced by the Antimicrobial Peptide PGLa.

Most antimicrobial peptides (AMPs) induce pore formation and a burst of lipid bilayers and plasma membranes. This causes severe leakage of the internal contents and cell death. The AMP PGLa forms nanopores in giant unilamellar vesicles (GUVs) comprising dioleoylphosphatidylcholine (DOPC) and dioleoylphosphatidylglycerol (DOPG). We here elucidated the effect of the line tension of a prepore rim on PGLa-induced nanopore formation by investigating the interaction of PGLa with single GUVs comprising dioleoylphosphatidylethanolamine (DOPE)/DOPG (6:4) in buffer using the single GUV method. We found that PGLa forms nanopores in the GUV membrane, which evolved into a local burst and burst of GUVs. The rate of pore formation in DOPE/DOPG-GUVs was smaller than that in DOPC/DOPG-GUVs. PGLa is located only in the outer leaflet of a GUV bilayer just before a fluorescent probe AF647 leakage from the inside, indicating that this asymmetric distribution induces nanopore formation. PGLa-induced local burst and burst of GUVs were observed at 10 ms-time resolution. After nanopore formation started, dense particles and small vesicles appeared in the GUVs, followed by a decrease in the GUV diameter. The GUV was finally converted into smaller GUV or lipid membrane aggregates. We discuss the mechanisms of PGLa-induced nanopore formation and its direct evolution to a GUV burst.

Negative lipid membranes enhance the adsorption of TAT-decorated elastin-like polypeptide micelles

A cell-penetrating peptide (CPP) is a short amino-acid sequence capable of efficiently translocating across the cellular membrane of mammalian cells. However, the potential of CPPs as a delivery vector is hampered by the strong reduction of its translocation efficiency when it bears an attached molecular cargo. To overcome this problem, we used previously developed diblock copolymers of elastin-like polypeptides (ELPBCs), which we end functionalized with TAT (transactivator of transcription), an archetypal CPP built from a positively charged amino acid sequence of the HIV-1 virus. These ELPBCs self-assemble into micelles at a specific temperature and present the TAT peptide on their corona. These micelles can recover the lost membrane affinity of TAT and can trigger interactions with the membrane despite the presence of a molecular cargo. Herein, we study the influence of membrane surface charge on the adsorption of TAT-functionalized ELP micelles onto giant unilamellar vesicles (GUVs). We show that the TAT-ELPBC micelles show an increased binding constant toward negatively charged membranes compared to neutral membranes, but no translocation is observed. The affinity of the TAT-ELPBC micelles for the GUVs displays a stepwise dependence on the lipid charge of the GUV, which, to our knowledge, has not been reported previously for interactions between peptides and lipid membranes. By unveiling the key steps controlling the interaction of an archetypal CPP with lipid membranes, through regulation of the charge of the lipid bilayer, our results pave the way for a better design of delivery vectors based on CPPs.

Dynamic Light-Induced Protein Patterns at Model Membranes.

The precise localization and activation of proteins at the cell membrane at a certain time gives rise to many cellular processes, including cell polarization, migration, and division. Thus, methods to recruit proteins to model membranes with subcellular resolution and high temporal control are essential when reproducing and controlling such processes in synthetic cells. Here, a method is described for fabricating light-regulated reversible protein patterns at lipid membranes with high spatiotemporal precision. For this purpose, we immobilize the photoswitchable protein iLID (improved light-inducible dimer) on supported lipid bilayers (SLBs) and on the outer membrane of giant unilamellar vesicles (GUVs). Upon local blue light illumination, iLID binds to its partner Nano (wild-type SspB) and allows the recruitment of any protein of interest (POI) fused to Nano from the solution to the illuminated area on the membrane. This binding is reversible in the dark, which provides dynamic binding and release of the POI. Overall, this is a flexible and versatile method for regulating the localization of proteins with high precision in space and time using blue light.

Sedimentation‐Based Confinement of Individual Giant Unilamellar Vesicles in Microchamber Arrays with a Dynamically Exchangeable Outer Medium

AbstractGiant unilamellar vesicles (GUVs) are an ideal model to study cellular membrane functions in vitro, yet difficult to manipulate due to their fragile nature, especially when subjected to dynamic change of their external microenvironment. Here, an original microfluidic concept is introduced for constraint‐free confinement of individual GUVs in microchambers with a dynamically exchangeable outer medium. GUVs self‐confine in an array of laterally separated microchambers by sedimentation, avoiding any mechanical constraint while allowing diffusion‐based exchange of the outer medium via a microfluidic channel and time‐resolved microscopy observation. The geometric and flow parameters optimizing medium exchange while preventing GUV from lifting out are numerically established. Different aqueous solutions separated by air plugs can be flowed into the channel by taking advantage of a polydimethylsiloxane‐based hydrophilic channel wall. The possibility to manipulate microliter sample volumes for in situ observation of protein cell‐free expression is also exploited. It is found that the membrane‐targeting sequence of Bacillus subtilis MinD binds to GUVs and induces extensive membrane tubulation. This technically simple method offers a robust way to confine GUVs and dynamically control their outer medium, thus constituting an ideal platform to study the spatio‐temporal response of reconstituted membranes and/or synthetic cell studies subjected to dynamic micro‐environments.

Bilayer Charge Asymmetry and Oil Residues Destabilize Membranes upon Poration.

Transmembrane asymmetry is ubiquitous in cells, particularly with respect to lipids, where charged lipids are mainly restricted to one monolayer. We investigate the influence of anionic lipid asymmetry on the stability of giant unilamellar vesicles (GUVs), minimal plasma membrane models. To quantify asymmetry, we apply the fluorescence quenching assay, which is often difficult to reproduce, and caution in handling the quencher is generally underestimated. We first optimize this assay and then apply it to GUVs prepared with the inverted emulsion transfer protocol by using increasing fractions of anionic lipids restricted to one leaflet. This protocol is found to produce highly asymmetric bilayers but with ∼20% interleaflet mixing. To probe the stability of asymmetric versus symmetric membranes, we expose the GUVs to porating electric pulses and monitor the fraction of destabilized vesicles. The pulses open macropores, and the GUVs either completely recover or exhibit leakage or bursting/collapse. Residual oil destabilizes porated membranes, and destabilization is even more pronounced in asymmetrically charged membranes. This is corroborated by the measured pore edge tension, which is also found to decrease with increasing charge asymmetry. Using GUVs with imposed transmembrane pH asymmetry, we confirm that poration-triggered destabilization does not depend on the approach used to generate membrane asymmetry.

Electroformation of giant unilamellar vesicles from large liposomes

AbstractGiant unilamellar vesicles (GUVs) are widely used as model systems for biological membranes to study membrane-related processes in a precisely controlled in vitro environment, owing to their biophysical properties. The classical technique for the formation of giant liposomes starts with the dissolution of lipids or lipid mixtures in an organic solvent, which is then deposited as a thin lipid solution film on a support substrate. In this study, we present a comprehensive analysis investigating the effect of different lipid compositions on the generation of GUVs from preformed liposomes under non-ionic and ionic conditions. For all liposome types tested, the electroformation process, whether performed on indium tin oxide-coated glass slides or platinum wires, consistently produced GUVs that typically ranged in size from 5 to 20 μm. However, the yield of GUVs varied depending on the specific non-ionic or ionic conditions and the lipid composition of the preformed liposomes used. In general, the resulting population of giant vesicles was predominantly characterised by the presence of unilamellar and multivesicular vesicles. These findings have the potential to improve the refinement of protocol parameters for the formation of GUVs containing membrane proteins and for the study of the effects of lipid composition on membrane protein activity.

Rearrangement of GUV-confined actin networks in response to micropipette aspiration.

Although diverse actin network architectures found inside the cell have been individually reconstituted outside of the cell, how different types of actin architectures reorganize under applied forces is not entirely understood. Recently, bottom-up reconstitution has enabled studies where dynamic and phenotypic characteristics of various actin networks can be recreated in an isolated cell-like environment. Here, by creating a giant unilamellar vesicle (GUV)-based cell model encapsulating actin networks, we investigate how actin networks rearrange in response to localized stresses applied by micropipette aspiration. We reconstitute actin bundles and branched bundles in GUVs separately and mechanically perturb them. Interestingly, we find that, when aspirated, protrusive actin bundles that are otherwise randomly oriented in the GUV lumen collapse and align along the axis of the micropipette. However, when branched bundles are aspirated, the network remains intact and outside of the pipette while the GUV membrane is aspirated into the micropipette. These results reveal distinct responses in the rearrangement of actin networks in a network architecture-dependent manner when subjected to physical forces.

Role of Interaction Range and Buoyancy on the Adhesion of Vesicles.

Vesicles on substrates play a fundamental role in many biological processes, ranging from neurotransmitter release at the synapse on small scales to the nutrient intake of trees by large vesicles. For these processes, the adsorption or desorption of vesicles to biological substrates is crucial. Consequently, it is important to understand the factors determining whether and for how long a vesicle adsorbs to a substrate and what shape it will adopt. Here, we systematically study the adsorption of a vesicle to planar substrates with short- and long-range interactions, with and without buoyancy. We assume an axially symmetric system throughout our simulations. Previous studies often considered a contact potential of zero range and neutral buoyancy. The interaction range alters the location and order of the adsorption transition and is particularly important for small vesicles, e.g., in the synapse. Whereas even small density differences between the inside and the outside of the vesicle give rise to strong buoyancy effects for large vesicles, e.g., giant unilamellar vesicles, as buoyancy effects scale with the fourth power of the vesicle size. We find that (i) an attractive membrane-substrate potential with nonzero spatial extension leads to a pinned state, where the vesicle benefits from the attractive membrane-substrate interaction without significant deformation. The adsorption transition is of first order and occurs when the substrate switches from repulsive to attractive. (ii) Buoyancy shifts the transversality condition, which relates the maximal curvature in the contact zone to the adhesion strength and bending rigidity, up/downward, depending on the direction of the buoyancy force. The magnitude of the shift is influenced by the range of the potential. For upward buoyancy, adsorbed vesicles are at most metastable. We determine the stability limit and the desorption mechanisms and compile the thermodynamic data into an adsorption diagram. Our findings reveal that buoyancy, as well as spatially extended interactions, are essential when quantitatively comparing experiments to theory.