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1.
Methods Enzymol ; 701: 175-236, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-39025572

RESUMO

Biomembranes and vesicles cover a wide range of length scales. Indeed, small nanovesicles have a diameter of a few tens of nanometers whereas giant vesicles can have diameters up to hundreds of micrometers. The remodeling of giant vesicles on the micron scale can be observed by light microscopy and understood by the theory of curvature elasticity, which represents a top-down approach. The theory predicts the formation of multispherical shapes as recently observed experimentally. On the nanometer scale, much insight has been obtained via coarse-grained molecular dynamics simulations of nanovesicles, which provides a bottom-up approach based on the lipid numbers assembled in the two bilayer leaflets and the resulting leaflet tensions. The remodeling processes discussed here include the shape transformations of vesicles, their morphological responses to the adhesion of condensate droplets, the instabilities of lipid bilayers and nanovesicles, as well as the topological transformations of vesicles by membrane fission and fusion. The latter processes determine the complex topology of the endoplasmic reticulum.


Assuntos
Bicamadas Lipídicas , Simulação de Dinâmica Molecular , Bicamadas Lipídicas/química , Membrana Celular/química , Membrana Celular/metabolismo , Fusão de Membrana/fisiologia , Retículo Endoplasmático/metabolismo , Retículo Endoplasmático/ultraestrutura , Animais , Humanos
2.
Proc Natl Acad Sci U S A ; 121(26): e2321579121, 2024 Jun 25.
Artigo em Inglês | MEDLINE | ID: mdl-38900795

RESUMO

Cellular membranes exhibit a multitude of highly curved morphologies such as buds, nanotubes, cisterna-like sheets defining the outlines of organelles. Here, we mimic cell compartmentation using an aqueous two-phase system of dextran and poly(ethylene glycol) encapsulated in giant vesicles. Upon osmotic deflation, the vesicle membrane forms nanotubes, which undergo surprising morphological transformations at the liquid-liquid interfaces inside the vesicles. At these interfaces, the nanotubes transform into cisterna-like double-membrane sheets (DMS) connected to the mother vesicle via short membrane necks. Using super-resolution (stimulated emission depletion) microscopy and theoretical considerations, we construct a morphology diagram predicting the tube-to-sheet transformation, which is driven by a decrease in the free energy. Nanotube knots can prohibit the tube-to-sheet transformation by blocking water influx into the tubes. Because both nanotubes and DMSs are frequently formed by cellular membranes, understanding the formation and transformation between these membrane morphologies provides insight into the origin and evolution of cellular organelles.


Assuntos
Nanotubos , Polietilenoglicóis , Nanotubos/química , Polietilenoglicóis/química , Membrana Celular/metabolismo , Dextranos/química , Dextranos/metabolismo
3.
Adv Sci (Weinh) ; 11(23): e2309864, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38582523

RESUMO

Interactions between membranes and biomolecular condensates can give rise to complex phenomena such as wetting transitions, mutual remodeling, and endocytosis. In this study, light-triggered manipulation of condensate engulfment is demonstrated using giant vesicles containing photoswitchable lipids. UV irradiation increases the membrane area, which can be stored in nanotubes. When in contact with a condensate droplet, the UV light triggers rapid condensate endocytosis, which can be reverted by blue light. The affinity of the protein-rich condensates to the membrane and the reversibility of the engulfment processes is quantified from confocal microscopy images. The degree of photo-induced engulfment, whether partial or complete, depends on the vesicle excess area and the relative sizes of vesicles and condensates. Theoretical estimates suggest that utilizing the light-induced excess area to increase the vesicle-condensate adhesion interface is energetically more favorable than the energy gain from folding the membrane into invaginations and tubes. The overall findings demonstrate that membrane-condensate interactions can be easily and quickly modulated via light, providing a versatile system for building platforms to control cellular events and design intelligent drug delivery systems for cell repair.


Assuntos
Condensados Biomoleculares , Endocitose , Endocitose/fisiologia , Condensados Biomoleculares/metabolismo , Condensados Biomoleculares/química , Microscopia Confocal/métodos , Raios Ultravioleta
4.
Eur Phys J E Soft Matter ; 47(1): 4, 2024 Jan 11.
Artigo em Inglês | MEDLINE | ID: mdl-38206459

RESUMO

Phase separation of biomembranes into two fluid phases, a and b, leads to the formation of vesicles with intramembrane a- and b-domains. These vesicles can attain multispherical shapes consisting of several spheres connected by closed membrane necks. Here, we study the morphological complexity of these multispheres using the theory of curvature elasticity. Vesicles with two domains form two-sphere shapes, consisting of one a- and one b-sphere, connected by a closed ab-neck. The necks' effective mean curvature is used to distinguish positive from negative necks. Two-sphere shapes of two-domain vesicles can attain four different morphologies that are governed by two different stability conditions. The closed ab-necks are compressed by constriction forces which induce neck fission and vesicle division for large line tensions and/or large spontaneous curvatures. Multispherical shapes with one ab-neck and additional aa- and bb-necks involve several stability conditions, which act to reduce the stability regimes of the multispheres. Furthermore, vesicles with more than two domains form multispheres with more than one ab-neck. The multispherical shapes described here represent generalized constant-mean-curvature surfaces with up to four constant mean curvatures. These shapes are accessible to experimental studies using available methods for giant vesicles prepared from ternary lipid mixtures.

5.
Microb Cell Fact ; 22(1): 244, 2023 Nov 30.
Artigo em Inglês | MEDLINE | ID: mdl-38037098

RESUMO

Protein complex assembly facilitates the combination of individual protein subunits into functional entities, and thus plays a crucial role in biology and biotechnology. Recently, we developed quasi-twodimensional, silicon-based compartmental biochips that are designed to study and administer the synthesis and assembly of protein complexes. At these biochips, individual protein subunits are synthesized from locally confined high-density DNA brushes and are captured on the chip surface by molecular traps. Here, we investigate single-gene versions of our quasi-twodimensional synthesis systems and introduce the trap-binding efficiency to characterize their performance. We show by mathematical and computational modeling how a finite trap density determines the dynamics of protein-trap binding and identify three distinct regimes of the trap-binding efficiency. We systematically study how protein-trap binding is governed by the system's three key parameters, which are the synthesis rate, the diffusion constant and the trap-binding affinity of the expressed protein. In addition, we describe how spatially differential patterns of traps modulate the protein-trap binding dynamics. In this way, we extend the theoretical knowledge base for synthesis, diffusion, and binding in compartmental systems, which helps to achieve better control of directed molecular self-assembly required for the fabrication of nanomachines for synthetic biology applications or nanotechnological purposes.


Assuntos
Nanotecnologia , Biossíntese de Proteínas , Subunidades Proteicas , Nanotecnologia/métodos , Simulação por Computador , Silício
6.
Soft Matter ; 19(36): 6929-6944, 2023 Sep 20.
Artigo em Inglês | MEDLINE | ID: mdl-37664906

RESUMO

Biological and biomimetic membranes are based on lipid bilayers, consisting of two monolayers or leaflets. One important but challenging physical parameter of these membranes is their tension. For a long time, this tension was explicitly or implicitly taken to be the bilayer tension, acting on the whole bilayer membrane. More recently, it has been realized that it is useful to decompose the bilayer tension into two leaflet tensions and that these tensions are accessible to molecular dynamics simulations. To divide the bilayer up into two leaflets, it is necessary to introduce a midsurface that defines the spatial extent of the two leaflets. In previous studies, this midsurface was obtained from the density profiles across the bilayer and was then used to compute the molecular area per lipid. Here, we develop an alternative approach based on three-dimensional Voronoi tessellation and molecular volume per lipid. Using this volume-based approach, we determine the reference states with tensionless leaflets as well as the optimal volumes and areas per lipid. The optimal lipid volumes have practically the same value in both leaflets, irrespective of the size and curvature of the nanovesicles, whereas the optimal lipid areas are different for the two leaflets and depend on the vesicle size. In addition, we introduce lateral volume compressibilities to describe the elastic response of the lipid volume to the leaflet tensions. We show that the outer leaflet of a nanovesicle is more densely packed and less compressible than the inner leaflet and that this difference becomes more pronounced for smaller vesicles.


Assuntos
Biomimética , Bicamadas Lipídicas , Simulação de Dinâmica Molecular
7.
ACS Nano ; 17(13): 11957-11968, 2023 07 11.
Artigo em Inglês | MEDLINE | ID: mdl-37377213

RESUMO

Artificial or synthetic organelles are a key challenge for bottom-up synthetic biology. So far, synthetic organelles have typically been based on spherical membrane compartments, used to spatially confine selected chemical reactions. In vivo, these compartments are often far from being spherical and can exhibit rather complex architectures. A particularly fascinating example is provided by the endoplasmic reticulum (ER), which extends throughout the whole cell by forming a continuous network of membrane nanotubes connected by three-way junctions. The nanotubes have a typical diameter of between 50 and 100 nm. In spite of much experimental progress, several fundamental aspects of the ER morphology remain elusive. A long-standing puzzle is the straight appearance of the tubules in the light microscope, which form irregular polygons with contact angles close to 120°. Another puzzling aspect is the nanoscopic shapes of the tubules and junctions, for which very different images have been obtained by electron microcopy and structured illumination microscopy. Furthermore, both the formation and maintenance of the reticular networks require GTP and GTP-hydrolyzing membrane proteins. In fact, the networks are destroyed by the fragmentation of nanotubes when the supply of GTP is interrupted. Here, it is argued that all of these puzzling observations are intimately related to each other and to the dimerization of two membrane proteins anchored to the same membrane. So far, the functional significance of this dimerization process remained elusive and, thus, seemed to waste a lot of GTP. However, this process can generate an effective membrane tension that stabilizes the irregular polygonal geometry of the reticular networks and prevents the fragmentation of their tubules, thereby maintaining the integrity of the ER. By incorporating the GTP-hydrolyzing membrane proteins into giant unilamellar vesicles, the effective membrane tension will become accessible to systematic experimental studies.


Assuntos
Retículo Endoplasmático , Proteínas de Membrana , Proteínas de Membrana/metabolismo , Microscopia , Guanosina Trifosfato/metabolismo
8.
Biomolecules ; 13(6)2023 05 31.
Artigo em Inglês | MEDLINE | ID: mdl-37371505

RESUMO

Biological and biomimetic membranes are based on lipid bilayers, which consist of two monolayers or leaflets. To avoid bilayer edges, which form when the hydrophobic core of such a bilayer is exposed to the surrounding aqueous solution, a single bilayer closes up into a unilamellar vesicle, thereby separating an interior from an exterior aqueous compartment. Synthetic nanovesicles with a size below 100 nanometers, traditionally called small unilamellar vesicles, have emerged as potent platforms for the delivery of drugs and vaccines. Cellular nanovesicles of a similar size are released from almost every type of living cell. The nanovesicle morphology has been studied by electron microscopy methods but these methods are limited to a single snapshot of each vesicle. Here, we review recent results of molecular dynamics simulations, by which one can monitor and elucidate the spatio-temporal remodeling of individual bilayers and nanovesicles. We emphasize the new concept of leaflet tensions, which control the bilayers' stability and instability, the transition rates of lipid flip-flops between the two leaflets, the shape transformations of nanovesicles, the engulfment and endocytosis of condensate droplets and rigid nanoparticles, as well as nanovesicle adhesion and fusion. To actually compute the leaflet tensions, one has to determine the bilayer's midsurface, which represents the average position of the interface between the two leaflets. Two particularly useful methods to determine this midsurface are based on the density profile of the hydrophobic lipid chains and on the molecular volumes.


Assuntos
Bicamadas Lipídicas , Simulação de Dinâmica Molecular , Bicamadas Lipídicas/química , Membrana Celular/metabolismo , Endocitose
9.
Nat Commun ; 14(1): 2809, 2023 05 22.
Artigo em Inglês | MEDLINE | ID: mdl-37217523

RESUMO

Cells compartmentalize parts of their interiors into liquid-like condensates, which can be reconstituted in vitro. Although these condensates interact with membrane-bound organelles, their potential for membrane remodeling and the underlying mechanisms of such interactions are not well-understood. Here, we demonstrate that interactions between protein condensates - including hollow ones, and membranes can lead to remarkable morphological transformations and provide a theoretical framework to describe them. Modulation of solution salinity or membrane composition drives the condensate-membrane system through two wetting transitions, from dewetting, through a broad regime of partial wetting, to complete wetting. When sufficient membrane area is available, fingering or ruffling of the condensate-membrane interface is observed, an intriguing phenomenon producing intricately curved structures. The observed morphologies are governed by the interplay of adhesion, membrane elasticity, and interfacial tension. Our results highlight the relevance of wetting in cell biology, and pave the way for the design of synthetic membrane-droplet based biomaterials and compartments with tunable properties.


Assuntos
Condensados Biomoleculares , Proteínas , Molhabilidade , Proteínas/química
10.
Membranes (Basel) ; 13(2)2023 Feb 10.
Artigo em Inglês | MEDLINE | ID: mdl-36837726

RESUMO

Condensate droplets are formed in aqueous solutions of macromolecules that undergo phase separation into two liquid phases. A well-studied example are solutions of the two polymers PEG and dextran which have been used for a long time in biochemical analysis and biotechnology. More recently, phase separation has also been observed in living cells where it leads to membrane-less or droplet-like organelles. In the latter case, the condensate droplets are enriched in certain types of proteins. Generic features of condensate droplets can be studied in simple binary mixtures, using molecular dynamics simulations. In this review, I address the interactions of condensate droplets with biomimetic and biological membranes. When a condensate droplet adheres to such a membrane, the membrane forms a contact line with the droplet and acquires a very high curvature close to this line. The contact angles along the contact line can be observed via light microscopy, lead to a classification of the possible adhesion morphologies, and determine the affinity contrast between the two coexisting liquid phases and the membrane. The remodeling processes generated by condensate droplets include wetting transitions, formation of membrane nanotubes as well as complete engulfment and endocytosis of the droplets by the membranes.

11.
Nat Commun ; 14(1): 615, 2023 02 04.
Artigo em Inglês | MEDLINE | ID: mdl-36739277

RESUMO

During endocytosis of nanoparticles by cells, the cellular membranes engulf the particles, thereby forming a closed membrane neck that subsequently undergoes fission. For solid nanoparticles, these endocytic processes have been studied in some detail. Recently, such processes have also been found for liquid and condensate droplets, both in vitro and in vivo. These processes start with the spreading of the droplet onto the membrane followed by partial or complete engulfment of the droplet. Here, we use molecular dynamics simulations to study these processes at the nanoscale, for nano-sized droplets and vesicles. For both partial and complete engulfment, we observe two different endocytic pathways. Complete engulfment leads to a closed membrane neck which may be formed in a circular or strongly non-circular manner. A closed circular neck undergoes fission, thereby generating two nested daughter vesicles whereas a non-circular neck hinders the fission process. Likewise, partial engulfment of larger droplets leads to open membrane necks which can again have a circular or non-circular shape. Two key parameters identified here for these endocytic pathways are the transbilayer stress asymmetry of the vesicle membrane and the positive or negative line tension of the membrane-droplet contact line.


Assuntos
Nanopartículas , Membrana Celular/metabolismo , Endocitose
12.
iScience ; 26(1): 105765, 2023 Jan 20.
Artigo em Inglês | MEDLINE | ID: mdl-36590172

RESUMO

The endosomal sorting complex required for transport (ESCRT) is a multi-protein machinery involved in several membrane remodeling processes. Different approaches have been used to resolve how ESCRT proteins scission membranes. However, the underlying mechanisms generating membrane deformations are still a matter of debate. Here, giant unilamellar vesicles, microfluidic technology, and micropipette aspiration are combined to continuously follow the ESCRT-III-mediated membrane remodeling on the single-vesicle level for the first time. With this approach, we identify different mechanisms by which a minimal set of three ESCRT-III proteins from Entamoeba histolytica reshape the membrane. These proteins modulate the membrane stiffness and spontaneous curvature to regulate bud size and generate intraluminal vesicles even in the absence of ATP. We demonstrate that the bud stability depends on the protein concentration and membrane tension. The approaches introduced here should open the road to diverse applications in synthetic biology for establishing artificial cells with several membrane compartments.

13.
Glycobiology ; 32(11): 933-948, 2022 10 31.
Artigo em Inglês | MEDLINE | ID: mdl-36197124

RESUMO

Many proteins are anchored to the cell surface of eukaryotes using a unique family of glycolipids called glycosylphosphatidylinositol (GPI) anchors. These glycolipids also exist without a covalently bound protein, in particular on the cell surfaces of protozoan parasites where they are densely populated. GPIs and GPI-anchored proteins participate in multiple cellular processes such as signal transduction, cell adhesion, protein trafficking and pathogenesis of Malaria, Toxoplasmosis, Trypanosomiasis and prion diseases, among others. All GPIs share a common conserved glycan core modified in a cell-dependent manner with additional side glycans or phosphoethanolamine residues. Here, we use atomistic molecular dynamic simulations and perform a systematic study to evaluate the structural properties of GPIs with different side chains inserted in lipid bilayers. Our results show a flop-down orientation of GPIs with respect to the membrane surface and the presentation of the side chain residues to the solvent. This finding agrees well with experiments showing the role of the side residues as active epitopes for recognition of GPIs by macrophages and induction of GPI-glycan-specific immune responses. Protein-GPI interactions were investigated by attaching parasitic GPIs to Green Fluorescent Protein. GPIs are observed to recline on the membrane surface and pull down the attached protein close to the membrane facilitating mutual contacts between protein, GPI and the lipid bilayer. This model is efficient in evaluating the interaction of GPIs and GPI-anchored proteins with membranes and can be extended to study other parasitic GPIs and proteins and develop GPI-based immunoprophylaxis to treat infectious diseases.


Assuntos
Glicosilfosfatidilinositóis , Simulação de Dinâmica Molecular , Glicosilfosfatidilinositóis/química , Glicosilfosfatidilinositóis/metabolismo , Glicolipídeos , Polissacarídeos , Proteínas Ligadas por GPI
14.
Soft Matter ; 18(34): 6372-6383, 2022 Aug 31.
Artigo em Inglês | MEDLINE | ID: mdl-35975692

RESUMO

His-tagged molecules can be attached to lipid bilayers via certain anchor lipids, a method that has been widely used for the biofunctionalization of membranes and vesicles. To observe the membrane-bound molecules, it is useful to consider His-tagged molecules that are fluorescent as well. Here, we study two such molecules, green fluorescence protein (GFP) and green-fluorescent fluorescein isothiocyanate (FITC), both of which are tagged with a chain of six histidines (6H) that bind to the anchor lipids within the bilayers. The His-tag 6H is much smaller than the GFP molecule but somewhat larger than the FITC dye. The lipid bilayers form giant unilamellar vesicles (GUVs), the behavior of which can be directly observed in the optical microscope. We apply and compare three well-established preparation methods for GUVs: electroformation on platinum wire, polyvinyl alcohol (PVA) hydrogel swelling, and electroformation on indium tin oxide (ITO) glass. Microfluidics is used to expose the GUVs to a constant fluorophore concentration in the exterior solution. The brightness of membrane-bound 6H-GFP exceeds the brightness of membrane-bound 6H-FITC, in contrast to the quantum yields of the two fluorophores in solution. In fact, 6H-FITC is observed to be strongly quenched by the anchor lipids which bind the fluorophores via Ni2+ ions. For both 6H-GFP and 6H-FITC, the membrane fluorescence is measured as a function of the fluorophores' molar concentration. The theoretical analysis of these data leads to the equilibrium dissociation constants Kd = 37.5 nM for 6H-GFP and Kd = 18.5 nM for 6H-FITC. We also observe a strong pH-dependence of the membrane fluorescence.


Assuntos
Bicamadas Lipídicas , Lipossomas Unilamelares , Fluoresceína-5-Isotiocianato , Corantes Fluorescentes , Proteínas de Fluorescência Verde , Bicamadas Lipídicas/química , Microscopia de Fluorescência , Lipossomas Unilamelares/química
15.
Soft Matter ; 18(32): 6066-6078, 2022 Aug 17.
Artigo em Inglês | MEDLINE | ID: mdl-35929498

RESUMO

Much effort has been devoted to lipid bilayers and nanovesicles with a compositional asymmetry between the two leaflets of the bilayer membranes. Here, we address another fundamental asymmetry related to lipid densities and membrane tensions. To avoid membrane rupture, the osmotic conditions must be adjusted in such a way that the bilayer membranes are subject to a relatively low bilayer tension. However, even for vanishing bilayer tension, the individual leaflets can still experience significant leaflet tensions if one leaflet is stretched whereas the other leaflet is compressed. Such a stress asymmetry between the two leaflets can be directly controlled in molecular dynamics simulations by the initial assembly of the lipid bilayers. This stress asymmetry is varied here over a wide range to determine the stability and instability regimes of the asymmetric bilayers. The stability regime shrinks with decreasing size and increasing membrane curvature of the nanovesicle. In the instability regimes, the lipids undergo stress-induced flip-flops with a flip-flop rate that increases with increasing stress asymmetry. The onset of flip-flops can be characterized by a cumulative distribution function that is well-fitted by an exponential function for planar bilayers but has a sigmoidal shape for nanovesicles. In addition, the bilayer membranes form transient non-bilayer structures that relax back towards ordered bilayers with a reduced stress asymmetry. Our study reveals intrinsic limits for the possible magnitude of the transbilayer stress asymmetry and shows that the leaflet tensions represent key parameters for the flip-flop rates.


Assuntos
Bicamadas Lipídicas , Simulação de Dinâmica Molecular , Bicamadas Lipídicas/química
16.
Adv Colloid Interface Sci ; 301: 102613, 2022 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-35228127

RESUMO

Giant lipid vesicles form unusual multispherical or "multi-balloon" shapes consisting of several spheres that are connected by membrane necks. Such multispherical shapes have been recently observed when the two sides of the membranes were exposed to different sugar solutions. This sugar asymmetry induced a spontaneous curvature, the sign of which could be reversed by swapping the interior with the exterior solution. Here, previous studies of multispherical shapes are reviewed and extended to develop a comprehensive theory for these shapes. Each multisphere consists of large and small spheres, characterized by two radii, the large-sphere radius, Rl, and the small-sphere radius, Rs. For positive spontaneous curvature, the multisphere can be built up from variable numbers Nl and Ns of large and small spheres. In addition, multispheres consisting of N*=Nl+Ns equally sized spheres are also possible and provide examples for constant-mean-curvature surfaces. For negative spontaneous curvature, all multispheres consist of one large sphere that encloses a variable number Ns of small spheres. These general features of multispheres arise from two basic properties of curvature elasticity: the local shape equation for spherical membrane segments and the stability conditions for closed membrane necks. In addition, the (Nl+Ns)-multispheres can form several (Nl+Ns)-patterns that differ in the way, in which the spheres are mutually connected. These patterns may involve multispherical junctions consisting of individual spheres that are connected to more than two neighboring spheres. The geometry of the multispheres is governed by two polynomial equations which imply that (Nl+Ns)-multispheres can only be formed within a certain restricted range of vesicle volumes. Each (Nl+Ns)-pattern can be characterized by a certain stability regime that depends both on the stability of the closed necks and on the multispherical geometry. Interesting and challenging topics for future studies include the response of multispheres to locally applied external forces, membrane fusion between spheres to create multispherical shapes of higher-genus topology, and the enlarged morphological complexity of multispheres arising from lipid phase separation and intramembrane domains.


Assuntos
Bicamadas Lipídicas , Fusão de Membrana , Elasticidade
17.
Adv Biol (Weinh) ; 6(1): e2101020, 2022 01.
Artigo em Inglês | MEDLINE | ID: mdl-34859961

RESUMO

Cellular membranes exhibit a fascinating variety of different morphologies, which are continuously remodeled by transformations of membrane shape and topology. This remodeling is essential for important biological processes (cell division, intracellular vesicle trafficking, endocytosis) and can be elucidated in a systematic and quantitative manner using synthetic membrane systems. Here, recent insights obtained from such synthetic systems are reviewed, integrating experimental observations and molecular dynamics simulations with the theory of membrane elasticity. The study starts from the polymorphism of biomembranes as observed for giant vesicles by optical microscopy and small nanovesicles in simulations. This polymorphism reflects the unusual elasticity of fluid membranes and includes the formation of membrane necks or fluid 'worm holes'. The proliferation of membrane necks generates stable multi-spherical shapes, which can form tubules and tubular junctions. Membrane necks are also essential for the remodeling of membrane topology via membrane fission and fusion. Neck fission can be induced by fine-tuning of membrane curvature, which leads to the controlled division of giant vesicles, and by adhesion-induced membrane tension as observed for small nanovesicles. Challenges for future research include the interplay of curvature elasticity and membrane tension during membrane fusion and the localization of fission and fusion processes within intramembrane domains.


Assuntos
Fusão de Membrana , Divisão Celular , Membrana Celular , Elasticidade , Membranas
18.
Adv Mater ; 34(4): e2106633, 2022 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-34710248

RESUMO

Molecular crowding is an inherent feature of cell interiors. Synthetic cells as provided by giant unilamellar vesicles (GUVs) encapsulating macromolecules (poly(ethylene glycol) and dextran) represent an excellent mimetic system to study membrane transformations associated with molecular crowding and protein condensation. Similarly to cells, such GUVs exhibit highly curved structures like nanotubes. Upon liquid-liquid phase separation their membrane deforms into apparent kinks at the contact line of the interface between the two aqueous phases. These structures, nanotubes, and kinks, have dimensions below optical resolution. Here, these are studied with super-resolution stimulated emission depletion (STED) microscopy facilitated by immobilization in a microfluidic device. The cylindrical nature of the nanotubes based on the superior resolution of STED and automated data analysis is demonstrated. The deduced membrane spontaneous curvature is in excellent agreement with theoretical predictions. Furthermore, the membrane kink-like structure is resolved as a smoothly curved membrane demonstrating the existence of the intrinsic contact angle, which describes the wettability contrast of the encapsulated phases to the membrane. Resolving these highly curved membrane structures with STED imaging provides important insights in the membrane properties and interactions underlying cellular activities.


Assuntos
Nanotubos , Lipossomas Unilamelares , Microscopia , Nanotubos/química , Proteínas , Lipossomas Unilamelares/química
19.
Adv Sci (Weinh) ; 8(21): e2102109, 2021 11.
Artigo em Inglês | MEDLINE | ID: mdl-34569194

RESUMO

Biological cells are contained by a fluid lipid bilayer (plasma membrane, PM) that allows for large deformations, often exceeding 50% of the apparent initial PM area. Isolated lipids self-organize into membranes, but are prone to rupture at small (<2-4%) area strains, which limits progress for synthetic reconstitution of cellular features. Here, it is shown that by preserving PM structure and composition during isolation from cells, vesicles with cell-like elasticity can be obtained. It is found that these plasma membrane vesicles store significant area in the form of nanotubes in their lumen. These act as lipid reservoirs and are recruited by mechanical tension applied to the outer vesicle membrane. Both in experiment and theory, it is shown that a "superelastic" response emerges from the interplay of lipid domains and membrane curvature. This finding allows for bottom-up engineering of synthetic biomaterials that appear one magnitude softer and with threefold larger deformability than conventional lipid vesicles. These results open a path toward designing superelastic synthetic cells possessing the inherent mechanics of biological cells.


Assuntos
Membrana Celular/química , Lipossomas Unilamelares/química , Linhagem Celular Tumoral , Colesterol/química , Elasticidade , Humanos , Fosfatidilgliceróis/química , Esfingomielinas/química , Tensão Superficial
20.
Elife ; 102021 09 07.
Artigo em Inglês | MEDLINE | ID: mdl-34490842

RESUMO

We investigate the structural and orientational variability of the membrane-embedded T cell receptor (TCR) - CD3 complex in extensive atomistic molecular dynamics simulations based on the recent cryo-EM structure determined by Dong et al., 2019. We find that the TCR extracellular (EC) domain is highly variable in its orientation by attaining tilt angles relative to the membrane normal that range from 15° to 55°. The tilt angle of the TCR EC domain is both coupled to a rotation of the domain and to characteristic changes throughout the TCR - CD3 complex, in particular in the EC interactions of the Cß FG loop of the TCR, as well as in the orientation of transmembrane helices. The concerted motions of the membrane-embedded TCR - CD3 complex revealed in our simulations provide atomistic insights on conformational changes of the complex in response to tilt-inducing forces on antigen-bound TCRs.


Assuntos
Complexo CD3/metabolismo , Membrana Celular/metabolismo , Complexo Receptor-CD3 de Antígeno de Linfócitos T/metabolismo , Receptores de Antígenos de Linfócitos T alfa-beta/metabolismo , Linfócitos T/metabolismo , Complexo CD3/ultraestrutura , Membrana Celular/ultraestrutura , Microscopia Crioeletrônica , Humanos , Simulação de Dinâmica Molecular , Ligação Proteica , Conformação Proteica em alfa-Hélice , Complexo Receptor-CD3 de Antígeno de Linfócitos T/ultraestrutura , Receptores de Antígenos de Linfócitos T alfa-beta/ultraestrutura , Relação Estrutura-Atividade , Linfócitos T/imunologia , Linfócitos T/ultraestrutura
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