Pavel Bashkirov

GTPase cycle of dynamin is coupled to membrane squeeze and release, leading to spontaneous fission.

The GTPase dynamin is critically involved in membrane fission during endocytosis. How does dynamin use the energy of GTP hydrolysis for membrane remodeling? By monitoring the ionic permeability through lipid nanotubes (NT), we found that dynamin was capable of squeezing NT to extremely small radii, depending on the NT lipid composition. However, long dynamin scaffolds did not produce fission: instead, fission followed GTPase-dependent cycles of assembly and disassembly of short dynamin scaffolds and involved a stochastic process dependent on the curvature stress imposed by dynamin. Fission happened spontaneously upon NT release from the scaffold, without leakage. Our calculations revealed that local narrowing of NT could induce cooperative lipid tilting, leading to self-merger of the inner monolayer of NT (hemifission), consistent with the absence of leakage. We propose that dynamin transmits GTPs energy to periodic assembling of a limited curvature scaffold that brings lipids to an unstable intermediate.

Geometric Catalysis of Membrane Fission Driven by Flexible Dynamin Rings

Making the Cut Dynamin is the prototypical member of a large family of structurally related guanosine triphosphatases involved in membrane fission and fusion. A variety of models have been suggested to explain how dynamin works. Shnyrova et al. (p. 1433; see the Perspective by Holz) reconstituted dynamin-mediated membrane scission on lipid nanotubes and suggest a molecular model for dynamin activity that takes into consideration all known aspects of dynamin function. Guanosine triphosphate hydrolysis limits polymerization of the membrane protein dynamin on lipid nanotubes into short, metastable collars. [Also see Perspective by Holz] Biological membrane fission requires protein-driven stress. The guanosine triphosphatase (GTPase) dynamin builds up membrane stress by polymerizing into a helical collar that constricts the neck of budding vesicles. How this curvature stress mediates nonleaky membrane remodeling is actively debated. Using lipid nanotubes as substrates to directly measure geometric intermediates of the fission pathway, we found that GTP hydrolysis limits dynamin polymerization into short, metastable collars that are optimal for fission. Collars as short as two rungs translated radial constriction to reversible hemifission via membrane wedging of the pleckstrin homology domains (PHDs) of dynamin. Modeling revealed that tilting of the PHDs to conform with membrane deformations creates the low-energy pathway for hemifission. This local coordination of dynamin and lipids suggests how membranes can be remodeled in cells.

Ganglioside GM1 increases line tension at raft boundary in model membranes

Gangliosides are significant participants in suppression of immune system during tumor processes. It was shown that they can induce apoptosis of T-lymphocytes in a raft-dependent manner. Fluorescence confocal microscopy was used to study distribution and influence of ganglioside GM1 on raft properties in giant unilamellar vesicles. Both raft and non-raft phase markers were utilized. No visible phase separation was observed without GM1 unless lateral tension was applied to the membrane. At 2 mol % of GM1 large domains appeared indicating macroscopic phase separation. Increase of GM1 content to 5 mol % resulted in shape transformation of the domains consistent with growth of line tension at the domain boundary. At 10 mol % of GM1 almost all domains were pinched out from vesicles, forming their own homogeneous liposomes. Estimations showed that the change of the GM1 content from 2 to 5–10 mol % resulted in a several-fold increase of line tension. This finding provides a possible mechanism of apoptosis induction by GM1. Incorporation of GM1 into a membrane leads to an increase of the line tension. This results in a growth of the average size of rafts due to coalescence or merger of small domains. Thus, necessary proteins can find themselves in one common raft and start the corresponding cascade of reactions.

Variation of lipid membrane composition caused by strong bending

Dynamic coupling between the morphology and molecular composition of cellular membranes is crucial for formation of the intracellular organelles and transport vesicles. Most of the membrane proteins and lipids discriminate membrane curvatures. However, it remains unclear whether the curvature alone is sufficient to support heterogeneous distribution of lipids. Here we demonstrate that the curvature-driven redistribution of phospholipids, such as dioleoylphosphatidylethanolamine (DOPE), requires strong membrane bending. We used cylindrical lipid nanotubes (NTs) pulled from planar lipid membranes with lateral tension of ∼1 dyn/cm. Such high tensions forced extreme curvatures of the NT membrane, with luminal radius approaching the thickness of the lipid bilayer, 5nm. When the NT contained lipid species with high spontaneous curvature (SC), such as DOPE, we observed slow reduction of its radius. This reduction indicated the redistribution of DOPE between the inner and outer monolayers of the NT. Accordingly, the SC of DOPE was recovered from the measured changes in the radii: the SC value, calculated under the assumption that the DOPE content is coupled to the monolayer curvature, was ∼0.4 nm−1, consistent with the published data. Thus, redistribution of lipids should be taken into account in calculations of composition and material properties of strongly deformed membrane structures, such as intermediate structures arising in the processes of membrane fusion and fission.

Interaction of amphipathic peptides mediated by elastic membrane deformations

Amphipathic alpha-helical peptides are perspective antimicrobial drugs. These peptides are partially embedded into the membrane to a shallow depth so that the longitudinal axis of the helix is parallel to the plane of the membrane or deviates from it by a small angle. In the framework of theory of elasticity of liquid crystals, adapted to lipid membranes, we calculated the energy of deformations occurring near the peptides partially embedded into the membrane. The energy of deformations is minimal when two peptides are parallel to each other and stay at a distance of about 5 nm. This configuration is stable with respect to small parallel displacements of the peptides and with respect to small variation of the angle between their axes both in the plane of the membrane and in the perpendicular direction. As a result of deformation the average thickness of the membrane decreases. The distribution of the elastic energy density has a maximum in the middle between the peptides. This region is the most likely place for formation of the through pores in the membrane. Since the equilibrium distance between the peptides is relatively large, it is assumed that the originally appearing pore should be purely lipidic.

Mechanism of pore formation in stearoyl-oleoyl-phosphatidylcholine membranes subjected to lateral tension

Theoretical model of a through pore formation in lipid bilayer membrane under applied lateral tension was developed. In the framework of elastic theory of liquid crystals adapted to lipid membranes, we calculated a continuous trajectory from intact bilayer through a hydrophobic defect to a through pore. It was shown that the major energetic characteristic of membrane stability with respect to the pore formation, i. e., line tension, depends both on the pore radius and on the value of the applied lateral tension. This leads to a non-monotonous dependence of the average waiting time of the pore formation on the lateral tension: at low tensions the waiting time was large, then there was a local minimum, after which the average waiting time was increasing again. For membranes formed from stearoyl oleoyl phosphatidylcholine, the local minimum corresponded to the lateral tension of 7 mN/m; the calculated value of the edge line tension of a large pore was 16.5 pN. These results are consistent with available experimental data.

The 2018 biomembrane curvature and remodeling roadmap

The importance of curvature as a structural feature of biological membranes has been recognized for many years and has fascinated scientists from a wide range of different backgrounds. On the one hand, changes in membrane morphology are involved in a plethora of phenomena involving the plasma membrane of eukaryotic cells, including endo- and exocytosis, phagocytosis and filopodia formation. On the other hand, a multitude of intracellular processes at the level of organelles rely on generation, modulation, and maintenance of membrane curvature to maintain the organelle shape and functionality. The contribution of biophysicists and biologists is essential for shedding light on the mechanistic understanding and quantification of these processes. Given the vast complexity of phenomena and mechanisms involved in the coupling between membrane shape and function, it is not always clear in what direction to advance to eventually arrive at an exhaustive understanding of this important research area. The 2018 Biomembrane Curvature and Remodeling Roadmap of Journal of Physics D: Applied Physics addresses this need for clarity and is intended to provide guidance both for students who have just entered the field as well as established scientists who would like to improve their orientation within this fascinating area.

Detection of DNA molecules in a lipid nanotube channel in the low ion strength conditions

Investigation of the transport phenomena in the nanoscopic channels/pores with the diameter smaller than 100 nm is of utmost importance for various biological, medical, and technical applications. Presently, the main line of development of nanofluidics is creation of biosensors capable of detecting single molecules and manipulating them. Detection of molecules is based on the measurement of electric current through a channel of appropriate size: when the molecule enters the channel, which diameter is comparable with the molecule size, the ion current reduces. In order to improve transport properties of such channels, their walls are often coated with a lipid bilayer, which behaves as two-dimensional liquid and thus is capable of supporting transport phenomena. In the present work, we utilized this property of lipid membranes for the development of a method for detecting and controlling transport of single-stranded DNA through channels formed by membrane cylinders with the luminal radii of 5–7 nm. We have demonstrated that in the conditions of small ion strength, the appearance of a DNA molecule inside such channel is accompanied by an increase of its ion conductivity and can be controlled by the polarity of the applied voltage. The amplitude of the ion current increase allows evaluating the amount of DNA molecules inside the channels. It was also demonstrated that upon adsorption of DNA molecules on the lipid bilayer surface, the membrane cylinder behaves as a voltage-sensitive selective ion channel.