Membrane Spontaneous Curvature Research Articles

Red Blood Cell Membrane Fluctuations and their Mechanisms: Passive Versus Active

Red blood cells (RBCs) are soft and flexible biconcave discs, which are able to pass through capillaries with diameters several times smaller than the RBC size. The RBC deformability also results in noticeable membrane fluctuations, which must be correlated with RBC membrane properties. However, it is still under debate whether RBC membrane fluctuations are simply passive thermal undulations or whether a red cell also experiences active fluctuations which are driven by a metabolic activity or other cell processes. We will present direct evidence that the RBC undulations are not solely passive thermal fluctuations, which has been obtained from a set of different experiments and simulations using a high spatiotemporal resolution: from 10 microseconds to several seconds in time and up to 20 nanometers in space. Experimental results show a violation of the fluctuation-dissipation theorem (FDT) for freshly prepared RBCs indicating the existence of active processes. However, the FDT is satisfied for starved cells demonstrating that the membrane fluctuations are passive when the energy supply is absent. Experiments also show a considerable change in the fluctuation amplitudes for fresh and starved cells. Subsequently, we perform simulations which fully mimic and quantify the experiments. We are able to quantitatively extract RBC membrane properties including shear elasticity, bending rigidity, and membrane viscosity. Furthermore, we test several models for active fluctuations, which mimic different possible mechanisms including spectrin network remodeling, ion pumps, and change in the spontaneous membrane curvature. Simulation results agree well with experimental data and suggest that several processes mentioned above may contribute to active RBC fluctuations. We will discuss which processes are more likely to take place.

How cholesterol is distributed between monolayers in asymmetric lipid membranes

The distribution of cholesterol in asymmetric lipid bilayers was studied by extensive coarse-grained molecular dynamics simulations. The effects of the lipid head group charge, acyl chain saturation, spontaneous membrane curvature and surface tension of the membrane were investigated. Four asymmetric bilayers containing DOPC, DOPS, DSPC or DSPS lipids were simulated on a time scale extended to tens of microseconds. We show that cholesterol strongly prefers anionic lipids to neutral and saturated lipid tails to unsaturated with a distribution ratio of ~0.7 in neutral/anionic bilayers and of ~0.4 in unsaturated/saturated bilayers. Multiple flip-flop transitions of cholesterol were observed directly, and their mean times ranged from 80 to 250 ns. It was shown that the distribution of cholesterol in the asymmetric membrane depends not only on the type of lipid, but also on the local membrane curvature and the surface tension. The membrane curvature enhances the influence of the lipid head groups on cholesterol distribution, while non-optimal surface tension caused by different areas per lipid in different monolayers increases the effect of the lipid tail saturation. It was clearly seen that the monolayers of asymmetric bilayers are interdependent. Mean distances from the bilayer center to cholesterol molecules depend not only on the type of the lipid in the considered monolayer but also on the composition of the opposite monolayer.

Munc18–1, exocytotic fusion pore regulation and local membrane anisotropy

The release of hormones and neurotransmitters from vesicles can be modified by the regulation of the fusion pore, an aqueous channel that forms upon the fusion of the vesicle membrane with the plasma membrane. However, the mechanisms are unclear. Munc18–1 protein interacts with Syntaxin1 (Synt 1), a member of the SNARE proteins, which plays an important role in exocytosis. It has been shown that Munc18–1 has multiple roles, both in pre- and post-fusion stages of exocytosis. It regulates the traffic of Synt1 to the plasma membrane. By inhibiting the tethering of the vesicle SNARE protein Synaptobrevin 2 (Syb2) solely to Synt1 at the plasma membrane, but favoring the vesicular tethering to the preformed binary cis SNARE complex of Synt1A-SNAP25B, Munc18–1 is tuning vesicle docking and the membrane merger process. Additionally, Munc18–1 affects exocytosis at the post-fusion stage by regulating the fusion pore properties (i.e., dwell-time and fusion pore diameter). Among many possible mechanisms that may regulate the fusion pore, but have never been considered previously, is the influence of Munc18–1 on the membrane anisotropy, which determines the local spontaneous membrane curvature and the architecture of the fusion pore. We here propose that Munc18–1 affects the fusion pore by modulating the dynamic local (re)arrangement of anisotropic membrane components within the highly curved fusion pore nanostructure, to which proteins, lipids or their complexes can participate.

Elastic, electrostatic and electrokinetic forces influencing membrane curvature

Many cellular and intracellular processes critically depend on membrane shape, but the shape generating mechanisms are still to be fully understood. In this study we evaluate how electrostatic/electrokinetic forces contribute to membrane curvature. Membrane bilayer had finite thickness and was either elastically anisotropic or anisotropic overall, but isotropic per sections (heads and tails). The physics of the situation was evaluated using a coupled system of elastic and electrostatic/electrokinetic (Poisson–Nernst–Planck) equations. The fixed charges present only on the upper membrane surface lead to the accumulation of counter-ions and depletion of co-ions that decay spatially very rapidly (Debye length<1nm), as does the potential and electric field. Spatially uneven electric field and the permittivity mismatch also induce charges at the membrane–solution interface, which are not fixed but influence the electrostatics nevertheless. Membrane bends due to — Coulomb force (caused by fixed membrane charges in the electric field) and the dielectric force (due to the non-uniform electric field and the permittivity mismatch between the membrane and the solution). Both act as membrane surface forces, and both depend supra-linearly on the fixed charge density. Regardless of sign of the fixed charges, the membrane bends toward the charged (upper) surface owing to the action of the Coulomb force, but this is opposed by the smaller dielectric force. The spontaneous membrane curvature becomes very pronounced at high fixed charge densities, leading to very small spontaneous radii (<50nm). In conclusion the electrostatic/electrokinetic forces contribute significantly to the membrane curvature.

Dynamical membrane curvature instability controlled by intermonolayer friction

We study a dynamical curvature instability caused by a local chemical modification of aphospholipid membrane. In our experiments, a basic solution is microinjected close to agiant unilamellar vesicle, which induces a local chemical modification of somelipids in the external monolayer of the membrane. This modification causes alocal deformation of the vesicle, which then relaxes. We present a theoreticaldescription of this instability, taking into account both the change of the equilibriumlipid density and the change of the spontaneous membrane curvature inducedby the chemical modification. We show that these two types of changes of themembrane properties yield different dynamics. In contrast, it is impossible todistinguish them when studying the equilibrium shape of a vesicle subjected to aglobal modification. In our model, the longest relaxation timescale is related to theintermonolayer friction, which plays an important part when there is a change in theequilibrium density in one monolayer. We compare our experimental results to thepredictions of our model by fitting the measured time evolution of the deformationheight to the solution of our dynamical equations. We obtain good agreementbetween theory and experiments. Our fits enable us to estimate the intermonolayerfriction coefficient, yielding values that are consistent with previous measurements.

The role of calcium in membrane condensation and spontaneous curvature variations in model lipidic systems

In this study, the dynamical behaviour of calcium-induced disordered to well-ordered structural transitions has been investigated by time-resolved synchrotron small-angle X-ray scattering (SAXS) in the milliseconds to seconds range. The in situ monitoring of the formed non-equilibrium self-assembled structures was achieved by the successful combination of synchrotron SAXS with stopped flow measurements. The effect of the rapid mixing of aqueous dispersions of dioleoylphosphatidyalglycerol (DOPG)/monoolein (MO) with low concentrations of Ca(2+) ions is reported. Under static conditions and in the absence of Ca(2+) ions, the evaluation of SAXS data for DOPG/MO aqueous dispersions prepared with three different DOPG/MO molar ratios indicates the formation of either a sponge-like L(3) phase or uncorrelated bilayers. Clearly, the lipid composition plays a vital role in modulating the structural behaviour of these aqueous dispersions in the absence and also in the presence of Ca(2+) ions. The rapid-mixing experiments revealed that the fast and strong interactions of Ca(2+) ions with the negatively charged DOPG/MO membranes triggers the transformation from the L(3) phase or the uncorrelated bilayers to the well-ordered dehydrated L(α) phase or to inverted type bicontinuous cubic phases, V(2), with either a symmetry of Pn3m or Im3m. Additionally, we recently reported (A. Yaghmur, P. Laggner, B. Sartori and M. Rappolt, PLoS ONE, 2008, 3, e2072) that low concentrations of Ca(2+) ions trigger the formation of the inverted type hexagonal (H(2)) phase in DOPG/MO aqueous dispersions with a molar DOPG/MO ratio of 30/70. These are also temperature-sensitive structural transitions. Intriguingly, the strong association of Ca(2+) ions with the negatively charged DOPG/MO membranes leads to fast re-organization of the two lipids and simultaneously induces fast tuning of the curvature.

Observing the Solubilization of Lipid Bilayers by Detergents with Optical Microscopy of GUVs

The solubilization of lipid bilayers by detergents was studied with optical microscopy of giant unilamellar vesicles (GUVs) composed of palmitoyl oleoyl phosphatidylcholine (POPC). A solution of the detergents Triton X-100 (TX-100) and sodium dodecyl sulfate (SDS) was injected with a micropipette close to single GUVs. The solubilization process was observed with phase contrast and fluorescence microscopy and found to be dependent on the detergent nature. In the presence of TX-100, GUVs initially showed an increase in their surface area, due to insertion of TX-100 with rapid equilibration between the two leaflets of the bilayer. Then, above a solubility threshold, several holes opened, rendering the bilayer a lace fabric appearance, and the bilayer gradually vanished. On the other hand, injection of SDS caused initially an increase in the membrane spontaneous curvature, which is mainly associated with incorporation of SDS in the outer layer only. This created a stress in the membrane, which caused either opening of transient macropores with substantial decrease in vesicle size or complete vesicle bursting. In another experimental setup, the extent of solubilization/destruction of a collection of GUVs was measured as a function of either TX-100 or SDS concentration.

Electrostatic Bending of Lipid Membranes: How Are Lipid and Electrostatic Properties Interrelated?

Electrostatic modification of lipid headgroups and its effect on membrane curvature are not only relevant in a variety of contexts such as cell shape transformation and membrane tubulation but also are intriguingly implicated in membrane functions. For instance, the gating (open vs closed) properties of mechanosensitive channels can be influenced by membrane curvature and ion valence. However, a full theoretical description of membrane electrostatics is still lacking; in the past, membrane bending has often been considered under a few assumptions about how bending modifies lipid arrangements and surface charges. Here, we present a unified theoretical approach to spontaneous membrane curvature, C(0), in which lipid properties (e.g., packing shape) and electrostatic effects are self-consistently integrated. For the description of electrostatic interactions, especially between a lipid charge and a divalent counterion, we implement the Poisson-Boltzmann (PB) approach by incorporation of finite ionic sizes, so as to capture both lateral and transverse charge correlations on the membrane surface. Our results show that C(0) is sensitive to the way lipid rearrangements and divalent counterions are modeled. Interestingly, it can change its sign in the presence of divalent counterions, thus stabilizing reverse hexagonal (H(II)) phases. Our results show how electrostatic modification of headgroups influences the preferred structure of lipid aggregates (inverted micelles vs bilayers).

Membrane Bending Energy and Fusion Pore Kinetics in Ca2+-Triggered Exocytosis

A fusion pore composed of lipid is an obligatory kinetic intermediate of membrane fusion, and its formation requires energy to bend membranes into highly curved shapes. The energetics of such deformations in viral fusion is well established, but the role of membrane bending in Ca2+-triggered exocytosis remains largely untested. Amperometry recording showed that during exocytosis in chromaffin and PC12 cells, fusion pores formed by smaller vesicles dilated more rapidly than fusion pores formed by larger vesicles. The logarithm of 1/(fusion pore lifetime) varied linearly with vesicle curvature. The vesicle size dependence of fusion pore lifetime quantitatively accounted for the nonexponential fusion pore lifetime distribution. Experimentally manipulating vesicle size failed to alter the size dependence of fusion pore lifetime. Manipulations of membrane spontaneous curvature altered this dependence, and applying the curvature perturbants to the opposite side of the membrane reversed their effects. These effects of curvature perturbants were opposite to those seen in viral fusion. These results indicate that during Ca2+-triggered exocytosis membrane bending opposes fusion pore dilation rather than fusion pore formation. Ca2+-triggered exocytosis begins with a proteinaceous fusion pore with less stressed membrane, and becomes lipidic as it dilates, bending membrane into a highly curved shape.

A Coarse Grained Molecular Dynamics Study of Self-Reproduction of Fatty Acid Vesicles

The formation of self-reproducing fatty acid vesicles is believed to be a quintessential building block required for (the origin of) life. In literature a number of vesicle reproduction experiments have been reported [1]. Of particular interest is the so called matrix effect where the final size distribution of vesicles formed upon addition of new surfactants is strongly biased toward the size distribution of pre-existing vesicles. However, the mechanism behind this is poorly understood. Here, we regard this mechanism using coarse grained molecular dynamics simulations with a parameter set based on a previously published force field [2]. These simulations show fission of a vesicle on continuous addition of new fatty acid molecules to the vesicle exterior. The new molecules are incorporated in the outer leaflet of the vesicle's membrane. As the rate of fatty acid redistribution among the leaflets by means of flip-flop is lower than the rate of fatty acid uptake in the outer leaflet, an excess of molecules in the outer leaflet compared to the inner leaflet is formed. This results in spontaneous membrane curvature, causing the vesicle to deform into twin-vesicles. Eventually, the built-up spontaneous curvature induces the formation of a narrow neck, that finally breaks, resulting in full fission. Importantly, no leakage from the interior of the vesicles was observed. The reproduction pathway shown in these simulations agrees with published experimental data, as well as with new data from our group (see abstract Nicole Pfleger). Our coarse grained molecular dynamics simulations offer further insight at the molecular level of the self-reproduction pathway of fatty acid vesicles.[1] Luisi et al. (2008) J. Phys. Chem. B 112, 14655-14664.[2] Markvoort et al. (2005) J. Phys. Chem. B 109, 22649-22654.

Modulation of Membrane Rigidity by Sar1, a Vesicle Trafficking Protein

Intracellular cargo trafficking involves dramatic changes in membrane shape, the mechanics of which remain poorly understood. We focus on Sar1, the key regulator of the coat protein complex II (COPII) family that ferries newly synthesized proteins from the endoplasmic reticulum (ER), and the only member of the COPII coat that interacts directly with the ER lipid bilayer membrane. To investigate whether Sar1 has a role beyond merely localizing the other COPII proteins, we directly measure the force involved in membrane deformation as a function of its concentration, using optically trapped microspheres to pull tethers from in vitro lipid membranes whose composition and large surface area mimic the composition and geometry of the ER. We find that Sar1 lowers the rigidity (bending modulus) of lipid membranes to nearly zero in a concentration-dependent manner. Moreover, Sar1 lacking its N-terminal amphipathic helix induces negative (concave) spontaneous membrane curvature. These results reveal a paradigm-altering insight into COPII trafficking: Sar1 actively alters the material properties of the membranes it binds to, lowering the energetic cost of curvature generation.View Large Image | View Hi-Res Image | Download PowerPoint Slide

Formation of Block Liposomes is a General Phenomenon of Charged Membranes

We have recently reported on the discovery of block liposomes (BLs), a new class of chain-melted (liquid) vesicles, formed in mixtures of curvature-stabilizing hexadecavalent cationic lipid MVLBG2, DOPC and water (Zidovska et al., Langmuir, 2008). BLs consist of distinctly shaped, yet connected spheres, pears, tubes, or rods. Unlike typical liposome systems, where spherical vesicles, tubular vesicles, and cylindrical micelles are separated on the macroscopic scale, within a BL, shapes are separated on the nanometer scale. We carried out structural studies of BLs with differential-contrast-microscopy (DIC) and cryo-transmission-electron-microscopy (cryo-TEM) and identified membrane charge and spontaneous membrane curvature as key parameters controlling the BL-formation. BL-formation was believed to be a special capacity of MVLBG2, a newly synthesized highly charged (16+) lipid (Ewert et al., JACS, 2006) with giant dendrimer-like headgroup leading to a conical molecular shape resulting into high spontaneous membrane curvature, when incorporated into lipid bilayer. In this work we report formation of BLs for other cationic lipids, demonstrating that BL-formation is a general phenomenon of all charged membranes. We carried out systematic study of binary lipid mixtures comprised of DOPC and a cationic lipid, varying the headgroup size and charge of cationic lipid from 1+, 3+ to 5+. We find that all cationic lipids form BLs on the micrometer and nanometer scale. We have also found that pure DOPC forms BLs in presence of monovalent salt, which is known to cause zwitterionic DOPC to become negatively charged. This latter finding confirms that BL-formation is a general capacity of charged membranes independent of the charge nature. Block liposomes may find a range of applications in chemical and nucleic acid delivery and as building blocks in the design of templates for hierarchical structures. Funding by DOE DE-FG-02-06ER46314, NIH GM-59288, NSF DMR-0503347.

Chemically Triggered Ejection of Membrane Tubules Controlled by Intermonolayer Friction

We report a chemically driven membrane shape instability that triggers the ejection of a tubule growing exponentially toward a chemical source. The instability is initiated by a dilation of the exposed monolayer, which is coupled to the membrane spontaneous curvature and slowed down by intermonolayer friction. Our experiments are performed by local delivery of a basic pH solution to a giant vesicle. Quantitative fits of the data give an intermonolayer friction coefficient b approximately 2x10;{9} J s/m;{4}. The exponential growth of the tubule may be explained by a Marangoni stress yielding a pulling force proportional to its length.

Curvature-driven lateral segregation of membrane constituents in Golgi cisternae

Lateral segregation of mobile membrane constituents (e.g. lipids, proteins or membrane domains) into the regions of their preferred curvature relaxes stresses in the membrane. The equilibrium distribution of the constituents in the membrane is thus a balance between the gains in the membrane elastic energy and the segregation-induced loss of entropy. The membrane in the Golgi cisternae is particularly susceptible to the curvature-driven segregation because it possesses two very different curvatures—the highly curved membrane in the cisternal rims and the flat membrane in the cisternal sides. In this work, we calculate the extent of lateral segregation in the Golgi cisternae in the case where the segregation is driven by the Helfrich bending energy. It is assumed that the membrane bending constant and spontaneous curvature depend on the local membrane composition. A simple analytical expression for the extent of the lateral segregation is derived. The results show that the segregation depends on the ratio between the bending constant and the thermal energy, the difference of the preferred curvatures of the constituents and the sizes of the constituents. Applying the model to a typical Golgi cisterna, it was found that entropy can effectively limit the extent of the curvature-driven lateral segregation.

Behavior of Giant Vesicles with Anchored DNA Molecules

We study changes in curvature and elastic properties of lipid membranes induced by anchoring of long hydrophilic polymers at low polymer surface concentrations (corresponding to the mushroom regime). The effect of anchored polymers on the membrane spontaneous curvature is characterized by monitoring the changes in the fluctuation spectra and the morphology of giant unilamellar vesicles. The polymers used in our study are fluorescently labeled and biotinylated λ-phage DNA molecules which bind to biotinylated giant unilamellar vesicles via a biotin-avidin-biotin linkage. By varying the amount of biotinylated lipid in the membrane, we control the surface concentration of anchors. At low anchor concentrations, the spontaneous curvature of the membrane increases linearly with the DNA concentration. The linear increase is consistent with theoretical predictions for polymer surface concentrations in the mushroom regime. At higher anchor concentrations, which should still belong to the mushroom regime, the vesicles undergo budding transitions. In this latter regime, the bud size is used to estimate the polymer-induced membrane curvature.

Theoretical model of reticulocyte to erythrocyte shape transformation

A theoretical model describing the kinetics of reticulocyte shape transformation was developed. The model considers the evolution of a simple cellular shape under transmembrane pressure difference, and proposes a four-parameter axisymmetric approximation of the cell surface. The mathematical analysis considers plasma membrane tension in the plane of bilayer leaflets, membrane spontaneous curvature and transmembrane transport of water. Cytoskeleton dilatational and shear rigidity, and the energetic barrier preventing the decrease of cell volume below a certain minimum are also incorporated. The set of adequate physical assumptions allowed for formulation of the equation for free energy of the investigated system. Computer simulations of cell shape changes, down to the state of free energy minimum, together with estimation of the time needed for the resulting transport of water, revealed a complex, three-phase picture of temporal alterations in cellular geometry with a wide spectrum of final results, and led to propose a standard model of reticulocyte–erythrocyte transformation. According to the model, both cell volume and surface undergo changes, and the work of the pressure, initially accumulated in the cytoskeleton, is consumed for local bending of the cell membrane. Further simulations with modified initial shape or parameters of the standard model show the trajectories of system evolution and help in better understanding the conditions for the erythro-, sphero-, ovalo-, stomato-, and leptoidal metamorphosis of maturing red blood cells. The stability of the final biconcave shape was also verified. Spherogenic modifications were discussed in the context of spherocytosis. Future development of the model was proposed.

Interaction of the antimicrobial peptide pheromone Plantaricin A with model membranes: Implications for a novel mechanism of action

Plantaricin A (plA) is a 26-residue bacteria-produced peptide pheromone with membrane-permeabilizing antimicrobial activity. In this study the interaction of plA with membranes is shown to be highly dependent on the membrane lipid composition. PlA bound readily to zwitterionic 1-stearoyl-2-oleoyl- sn-glycero-3-phosphocholine (SOPC) monolayers and liposomes, yet without significantly penetrating into these membranes. The presence of cholesterol attenuated the intercalation of plA into SOPC monolayers. The association of plA to phosphatidylcholine was, however, sufficient to induce membrane permeabilization, with nanomolar concentrations of the peptide triggering dye leakage from SOPC liposomes. The addition of the negatively charged phospholipid, 1-palmitoyl-2-oleoyl- sn-glycero-3-phospho- rac-glycerol POPG (SOPC/POPG; molar ratio 8:2) enhanced the membrane penetration of the peptide, as revealed by (i) peptide-induced increment in the surface pressure of lipid monolayers, (ii) increase in diphenylhexatriene (DPH) emission anisotropy measured for bilayers, and (iii) fluorescence characteristics of the two Trps of plA in the presence of liposomes, measured as such as well as in the presence of different quenchers. Despite deeper intercalation of plA into the SOPC/POPG lipid bilayer, much less peptide-induced dye leakage was observed for these liposomes than for the SOPC liposomes. Further changes in the mode of interaction of plA with lipids were evident when also the zwitterionic phospholipid, 1-palmitoyl-2-oleoyl- sn-glycerol-3-phosphoethanolaminne (POPE) was present (SOPC/POPG/POPE, molar ratio 3:2:5), thus suggesting increase in membrane spontaneous negative curvature to affect the mode of association of this peptide with lipid bilayer. PlA induced more efficient aggregation of the SOPC/POPG and SOPC/POPG/POPE liposomes than of the SOPC liposomes, which could explain the attenuated peptide-induced dye leakage from the former liposomes. At micromolar concentrations, plA killed human leukemic T-cells by both necrosis and apoptosis. Interestingly, plA formed supramolecular protein–lipid amyloid-like fibers upon binding to negatively charged phospholipid-containing membranes, suggesting a possible mechanistic connection between fibril formation and the cytotoxicity of plA.

SNARE-Mediated Lipid Mixing Depends on the Physical State of the Vesicles

Reconstitution experiments have suggested that N-ethylmaleimide sensitive factor attachment protein receptor (SNARE) proteins constitute a minimal membrane fusion machinery but have yielded contradictory results, and it is unclear whether the mechanism of membrane merger is related to the stalk mechanism that underlies physiological membrane fusion. Here we show that reconstitution of solubilized neuronal SNAREs into preformed 100 nm liposomes (direct method) yields proteoliposomes with more homogeneous sizes and protein densities than the standard reconstitution method involving detergent cosolubilization of proteins and lipids. Standard reconstitutions yield slow but efficient lipid mixing at high protein densities and variable amounts of lipid mixing at moderate protein densities. However, the larger, more homogenous proteoliposomes prepared by the direct method yield almost no lipid mixing at moderate protein densities. These results suggest that the lipid mixing observed for standard reconstitutions is dominated by the physical state of the membrane, perhaps due to populations of small vesicles (or micelles) with high protein densities and curvature stress created upon reconstitution. Accordingly, changing membrane spontaneous curvature by adding lysophospholipids inhibits the lipid mixing observed for standard reconstitutions. Our data indicate that the lipid mixing caused by high SNARE densities and/or curvature stress occurs by a stalk mechanism resembling the mechanism of fusion between biological membranes, but the neuronal SNAREs are largely unable to induce lipid mixing at physiological protein densities and limited curvature stress.

Dynamics of Membranes Driven by Actin Polymerization

A motile cell, when stimulated, shows a dramatic increase in the activity of its membrane, manifested by the appearance of dynamic membrane structures such as lamellipodia, filopodia, and membrane ruffles. The external stimulus turns on membrane bound activators, like Cdc42 and PIP2, which cause increased branching and polymerization of the actin cytoskeleton in their vicinity leading to a local protrusive force on the membrane. The emergence of the complex membrane structures is a result of the coupling between the dynamics of the membrane, the activators, and the protrusive forces. We present a simple model that treats the dynamics of a membrane under the action of actin polymerization forces that depend on the local density of freely diffusing activators on the membrane. We show that, depending on the spontaneous membrane curvature associated with the activators, the resulting membrane motion can be wavelike, corresponding to membrane ruffling and actin waves, or unstable, indicating the tendency of filopodia to form. Our model also quantitatively explains a variety of related experimental observations and makes several testable predictions.

Polymorphism of DNA–anionic liposome complexes reveals hierarchy of ion-mediated interactions

Self-assembled DNA delivery systems based on anionic lipids (ALs) complexed with DNA mediated by divalent cations have been recently introduced as an alternative to cationic lipid-DNA complexes because of their low cytotoxicity. We investigate AL-DNA complexes induced by different cations by using synchrotron small angle x-ray scattering and confocal microscopy to show how different ion-mediated interactions are expressed in the self-assembled structures and phase behavior of AL-DNA complexes. The governing interactions in AL-DNA systems are complex: divalent ions can mediate strong attractions between different combinations of the components (such as DNA-DNA and membrane-membrane). Moreover, divalent cations can coordinate non-electrostatically with lipids and modify the resultant membrane structure. We find that at low membrane charge densities AL-DNA complexes organize into a lamellar structure of alternating DNA and membrane layers crosslinked by ions. At high membrane charge densities, a new phase with no analog in cationic lipid-DNA systems is observed: DNA is expelled from the complex, and a lamellar stack of membranes and intercalated ions is formed. For a subset of the ionic species, high ion concentrations generate an inverted hexagonal phase comprised of DNA strands wrapped by ion-coated lipid tubes. A simple theoretical model that takes into account the electrostatic and membrane elastic contributions to the free energy shows that this transition is consistent with an ion-induced change in the membrane spontaneous curvature, c0. Moreover, the crossover between the lamellar and inverted hexagonal phases occurs at a critical c0 that agrees well with experimental values.