Explore the vast range of titles available.

RNA is a ubiquitous biomolecule that can serve as both catalyst and information carrier. Understanding how RNA bioactivity is controlled is crucial for elucidating its physiological roles and potential applications in synthetic biology. Here, we show that lipid membranes can act as RNA organization platforms, introducing a mechanism for riboregulation. The activity of R3C ribozyme can be modified by the presence of lipid membranes, with direct RNA–lipid interactions dependent on RNA nucleotide content, base pairing, and length. In particular, the presence of guanine in short RNAs is crucial for RNA–lipid interactions, and G-quadruplex formation further promotes lipid binding. Lastly, by artificially modifying the R3C substrate sequence to enhance membrane binding, we generated a lipid-sensitive ribozyme reaction with riboswitch-like behavior. These findings introduce RNA–lipid interactions as a tool for developing synthetic riboswitches and RNA-based lipid biosensors and bear significant implications for RNA world scenarios for the origin of life.

The lipid composition of cellular organelles is tailored to suit their specialized tasks. A fundamental transition in the lipid landscape divides the secretory pathway in early and late membrane territories, allowing an adaptation from biogenic to barrier functions. Defending the contrasting features of these territories against erosion by vesicular traffic poses a major logistical problem. To this end, cells evolved a network of lipid composition sensors and pipelines along which lipids are moved by non-vesicular mechanisms. We review recent insights into the molecular basis of this regulatory network and consider examples in which malfunction of its components leads to system failure and disease.

Giant vesicles are widely used as a model membrane system, both for basic biological systems and for their promising applications in the development of smart materials and cell mimetics, as well as in driving new technologies in synthetic biology and for the cosmetics and pharmaceutical industry. The reader is guided to use giant vesicles, from the formation of simple membrane platforms to advanced membrane and cell system models. It also includes fundamentals for understanding lipid or polymer membrane structure, properties and behavior. Every chapter includes ideas for further applications and discussions on the implications of the observed phenomena towards understanding membrane-related processes. The Giant Vesicle Book is meant to be a road companion, a trusted guide for those making their first steps in this field as well as a source of information required by experts. Key Features • A complete summary of the field, covering fundamental concepts, practical methods, core theory, and the most promising applications • A start-up package of theoretical and experimental information for newcomers in the field • Extensive protocols for establishing the required preparations and assays • Tips and instructions for carefully performing and interpreting measurements with giant vesicles or for observing them, including pitfalls • Approaches developed for investigating giant vesicles as well as brief overviews of previous studies implementing the described techniques • Handy tables with data and structures for ready reference Part I: The making of Chapter 1 Preparation methods for giant unilamellar vesicles - Rumiana Dimova, Pasquale Stano, Carlos M. Marques and Peter Walde Chapter 2 Preparation and properties of giant plasma membrane vesicles and giant unilamellar vesicles from natural membranes - Joseph H. Lorent and Ilya Levental Chapter 3 Protein reconstitution in giant vesicles - Matthias Garten, Daniel Lévy and Patricia Bassereau　 Chapter 4 GUVs with cytoskeleton - Tobias Härtel and Petra Schwille Part II: Giant vesicles theoretically and in silico Chapter 5 Understanding giant vesicles – a theoretical perspective - Reinhard Lipowsky Chapter 6 Simulating membranes, vesicles, and cells - Thorsten Auth, Dmitry A. Fedosov and Gerhard Gompper Chapter 7 Theory of vesicle dynamics in flow and electric fields - Petia M. Vlahovska and Chaouqi Misbah Chapter 8 Particle-membrane interactions - Jaime Agudo-Canalejo, Reinhard Lipowsky Chapter 9 Theory of polymer-membrane interactions - Fabrice Thalmann and Carlos M. Marques Part III: GUV-based techniques and what one can learn from them Chapter 10 Application of optical microscopy techniques on giant unilamellar vesicles - Luis A. Bagatolli Chapter 11 Mechanics assays of synthetic lipid membranes based on micropipette aspiration - Elisa Parra and David Needham Chapter 12 Atomic force microscopy of giant unilamellar vesicles - Andreas Janshoff Chapter 13 Manipulation and biophysical characterization of GUVs with an optical stretcher - Gheorghe Cojoc, Antoine Girot, Ulysse Delabre and Jochen Guck Chapter 14 Vesicle fluctuation analysis - John Hjort Ipsen, Allan Grønhøj Hansen and Tripta Bhatia Chapter 15 Using electric fields to assess membrane material properties in GUVs - Rumiana Dimova and　　Karin A. Riske Chapter 16 Creating membrane nanotubes from GUVs - Coline Prévost, Mijo Simunovic and Patricia Bassereau Chapter 17 Measuring GUV adhesion - Kheya Sengupta and Ana Smith Chapter 18 Phase diagrams and tie lines in GUVs - Matthew C. Blosser, Caitlin Cornell, Scott P. Rayermann and Sarah L. Keller Chapter 19 Vesicle dynamics in flow: an experimental approach - Victor Steinberg and Michael Levant Chapter 20 Membrane permeability measurements - Begoña Ugarte-Uribe, Ana J. García-Sáez and Mireille M. A. E. Claessens Part IV: GUVs as membrane interaction platforms Chapter 21 - Lipid and protein mobility in GUVs - Begoña Ugarte-Uribe, Kushal Kumar Das and Ana J. García-Sáez Chapter 22 Shining light on membranes - Rosangela Itri, Carlos M. Marques and Mauricio S. Baptista Chapter 23 Protein-membrane interactions - Eva M Schmid and Daniel A Fletcher Chapter 24 Effects of antimicrobial peptides and detergents on GUVs - Karin A. Riske Chapter 25 Lipid-polymer interactions: effect on GUVs shapes and behavior - Brigitte Pépin-Donat, François Quemeneur and Clément Campillo Part V: GUVs as complex membrane containers Chapter 26 Polymersomes - Praful Nair, David Christian and Dennis E. Discher Chapter 27 Giant hybrid polymer/lipid vesicles - Thi Phuong Tuyen Dao, Khalid Ferji, Fabio Fernandes, Manuel Prieto, Sébastien Lecommandoux, Emmanuel Ibarboure, Olivier Sandre and Jean-François Le Meins Chapter 28 Giant unilamellar vesicles: from protocell models to the construction of minimal cells - Masayuki Imai and Peter Walde Chapter 29 Encapsulation of aqueous two-phase systems and gels within giant lipid vesicles - Allyson M. Marianelli and Christine D. Keating Chapter 30 Droplet-supported giant lipid vesicles as compartments for synthetic biology - Johannes P. Frohnmayer, Marian Weiss, Lucia T. Benk, Jan-Willi Janiesch, Barbara Haller, Rafael B. Lira, Rumiana Dimova, Ilia Plazman and Joachim P. Spatz Appendices Appendix 1 List of lipids and physical constants of lipid bilayers Appendix 2 List of membrane dyes and fluorescent groups conjugated to lipids Appendix 3 List of detergents Appendix 4 List of water-soluble dyes or their fluorescent groups and their structures Rumiana Dimova leads an experimental lab in biophysics at the Max Planck Institute of Colloids and Interfaces in Potsdam, Germany. She has been working with giant vesicles already from the beginning of her scientific career. After being introduced into the magic of their preparation during her studies as a student in Bulgaria, she remained fascinated by their application and over the years pursued a variety of projects employing giant vesicles as a platform to develop new methods for the biophysical characterization of membranes and processes involving them. Until now, these studies have resulted in more than hundred peer-reviewed publications. Recently, she was also awarded the Emmy Noether distinction for women in physics of the European Physical Society. Carlos Marques , a CNRS senior scientist, founded the MCube group at the Charles Sadron Institute in Strasbourg, France, where he gears experimental and theoretical research towards the understanding of the physical properties of self-assembled lipid bilayers. Trained as a polymer theoretician, Carlos first got interested in membranes because they interact with polymers and published the first prediction for the membrane changes expected when polymers adsorb on lipid bilayers. He then expanded the scope of his group to include experiments and numerical simulations, and has now published many papers based on research with giant unilamellar vesicles, including the first study of lipid oxidation in GUVs and the discovery of the so-called PVA method for vesicle growth.

Lipid membranes, which are fundamental to cellular function, undergo various mechanical deformations. Accurate modeling of these processes necessitates a thorough understanding of membrane elasticity. The lateral shear modulus, a critical parameter describing membrane resistance to lateral stresses, remains elusive due to the membrane’s fluid nature. Two contrasting hypotheses, local fluidity and global fluidity, have been proposed. While the former suggests a zero local lateral shear modulus anywhere within lipid monolayers, the latter posits that only the integral of this modulus over the monolayer thickness vanishes. These differing models lead to distinct estimations of other elastic moduli and affect the modeling of biological processes, such as membrane fusion/fission and membrane-mediated interactions. Notably, they predict distinct local stress distributions in cylindrically curved membranes. The local fluidity model proposes isotropic local lateral stress, whereas the global fluidity model predicts anisotropy due to anisotropic local lateral stretching of lipid monolayers. Using molecular dynamics simulations, this study directly investigates these models by analyzing local stress in a cylindrically curved membrane. The results conclusively demonstrate the existence of static local lateral shear stress and anisotropy in local lateral stress within the monolayers of the cylindrical membrane, strongly supporting the global fluidity model. These findings have significant implications for the calculation of surface elastic moduli and offer novel insights into the fundamental principles governing lipid membrane elasticity.

Cholesterol is an integral component of eukaryotic cell membranes and a key molecule in controlling membrane fluidity, organization, and other physicochemical parameters. It also plays a regulatory function in antibiotic drug resistance and the immune response of cells against viruses, by stabilizing the membrane against structural damage. While it iswell understood that, structurally, cholesterol exhibits a densification effect on fluid lipid membranes, its effects on membrane bending rigidity are assumed to be nonuniversal; i.e., cholesterol stiffens saturated lipid membranes, but has no stiffening effect on membranes populated by unsaturated lipids, such as 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC). This observation presents a clear challenge to structure–property relationships and to our understanding of cholesterol-mediated biological functions. Here, using a comprehensive approach—combining neutron spin-echo (NSE) spectroscopy, solid-state deuterium NMR (²H NMR) spectroscopy, and molecular dynamics (MD) simulations—we report that cholesterol locally increases the bending rigidity of DOPC membranes, similar to saturated membranes, by increasing the bilayer’s packing density. All three techniques, inherently sensitive to mesoscale bending fluctuations, show up to a threefold increase in effective bending rigidity with increasing cholesterol content approaching a mole fraction of 50%. Our observations are in good agreement with the known effects of cholesterol on the area-compressibility modulus and membrane structure, reaffirming membrane structure–property relationships. The current findings point to a scale-dependent manifestation of membrane properties, highlighting the need to reassess cholesterol’s role in controlling membrane bending rigidity over mesoscopic length and time scales of important biological functions, such as viral budding and lipid–protein interactions.

Parkinson’s disease, the most common age-related movement disorder, is a progressive neurodegenerative disease with unclear etiology. Key neuropathological hallmarks are Lewy bodies and Lewy neurites: neuronal inclusions immunopositive for the protein α-synuclein. In-depth ultrastructural analysis of Lewy pathology is crucial to understanding pathogenesis of this disease. Using correlative light and electron microscopy and tomography on postmortem human brain tissue from Parkinson’s disease brain donors, we identified α-synuclein immunopositive Lewy pathology and show a crowded environment of membranes therein, including vesicular structures and dysmorphic organelles. Filaments interspersed between the membranes and organelles were identifiable in many but not all α-synuclein inclusions. Crowding of organellar components was confirmed by stimulated emission depletion (STED)-based super-resolution microscopy, and high lipid content within α-synuclein immunopositive inclusions was corroborated by confocal imaging, Fourier-transform coherent anti-Stokes Raman scattering infrared imaging and lipidomics. Applying such correlative high-resolution imaging and biophysical approaches, we discovered an aggregated protein–lipid compartmentalization not previously described in the Parkinsons’ disease brain.

A multitude of biological processes are enabled by complex interactions between lipid membranes and proteins. To understand such dynamic processes, it is crucial to differentiate the constituent biomolecular species and track their individual time evolution without invasive labels. Here, we present a label-free mid-infrared biosensor capable of distinguishing multiple analytes in heterogeneous biological samples with high sensitivity. Our technology leverages a multi-resonant metasurface to simultaneously enhance the different vibrational fingerprints of multiple biomolecules. By providing up to 1000-fold near-field intensity enhancement over both amide and methylene bands, our sensor resolves the interactions of lipid membranes with different polypeptides in real time. Significantly, we demonstrate that our label-free chemically specific sensor can analyze peptide-induced neurotransmitter cargo release from synaptic vesicle mimics. Our sensor opens up exciting possibilities for gaining new insights into biological processes such as signaling or transport in basic research as well as provides a valuable toolkit for bioanalytical and pharmaceutical applications. Complex protein-lipid interactions are difficult to study in real-time without labels. Here, Rodrigo et al . introduce a multimodal plasmonic infrared biosensor to simultaneously distinguish multiple analytes with high sensitivity.

Key Points Cellular membranes are laterally heterogeneous and consist of transient and dynamic domains with varying properties, which prominently include ordered lipid-driven domains that are referred to as lipid (or membrane) rafts. Membrane domains can be induced and regulated by a variety of interactions, which include specific lipid–lipid and lipid–protein interactions, bulk membrane properties, and interactions between membrane components and the underlying cytoskeleton. Advanced microscopy and biochemistry techniques facilitate the study of membrane domains; however, these domains still elude direct in vivo visualization. The multiplicity of possible organizational states and their context-dependent nature most likely account for experimental inconsistencies. Membrane rafts potentially have crucial physiological roles across cell types that range from immune cells to cancer cells. Membrane domains are conserved throughout the domains of life, which supports their functional importance in biological systems. Lipid rafts are relatively ordered membrane domains that are enriched in cholesterol and saturated lipids, and selectively recruit other lipids and proteins. They are dynamic and heterogeneous in composition and are thus challenging to visualize in vivo . New technologies are providing novel insights into the formation, organization and functions of these membrane domains. Cellular plasma membranes are laterally heterogeneous, featuring a variety of distinct subcompartments that differ in their biophysical properties and composition. A large number of studies have focused on understanding the basis for this heterogeneity and its physiological relevance. The membrane raft hypothesis formalized a physicochemical principle for a subtype of such lateral membrane heterogeneity, in which the preferential associations between cholesterol and saturated lipids drive the formation of relatively packed (or ordered) membrane domains that selectively recruit certain lipids and proteins. Recent studies have yielded new insights into this mechanism and its relevance in vivo , owing primarily to the development of improved biochemical and biophysical technologies.

This protocol from Lee et al . describes a method for the extraction of membrane proteins in their native lipid environment using styrene maleic anhydride (SMA) co-polymer. The method is applicable to both prokaryotic and eukaryotic expression systems. Despite the great importance of membrane proteins, structural and functional studies of these proteins present major challenges. A significant hurdle is the extraction of the functional protein from its natural lipid membrane. Traditionally achieved with detergents, purification procedures can be costly and time consuming. A critical flaw with detergent approaches is the removal of the protein from the native lipid environment required to maintain functionally stable protein. This protocol describes the preparation of styrene maleic acid (SMA) co-polymer to extract membrane proteins from prokaryotic and eukaryotic expression systems. Successful isolation of membrane proteins into SMA lipid particles (SMALPs) allows the proteins to remain with native lipid, surrounded by SMA. We detail procedures for obtaining 25 g of SMA (4 d); explain the preparation of protein-containing SMALPs using membranes isolated from Escherichia coli (2 d) and control protein-free SMALPS using E. coli polar lipid extract (1–2 h); investigate SMALP protein purity by SDS–PAGE analysis and estimate protein concentration (4 h); and detail biophysical methods such as circular dichroism (CD) spectroscopy and sedimentation velocity analytical ultracentrifugation (svAUC) to undertake initial structural studies to characterize SMALPs (∼2 d). Together, these methods provide a practical tool kit for those wanting to use SMALPs to study membrane proteins.

Background Lipid-protein interactions stabilize protein oligomers, shape their structure, and modulate their function. Whereas in vitro experiments already account for the functional importance of lipids by using natural lipid extracts, in silico methods lack behind by embedding proteins in single component lipid bilayers. However, to accurately complement in vitro experiments with molecular details at very high spatio-temporal resolution, molecular dynamics simulations have to be performed in natural(-like) lipid environments. Results To enable more accurate MD simulations, we have prepared four membrane models of E. coli polar lipid extract, a typical model organism, each at all-atom (CHARMM36) and coarse-grained (Martini3) representations. These models contain all main lipid headgroup types of the E. coli inner membrane, i.e., phosphatidylethanolamines, phosphatidylglycerols, and cardiolipins, symmetrically distributed between the membrane leaflets. The lipid tail (un)saturation and propanylation stereochemistry represent the bacterial lipid tail composition of E. coli grown at 37 ∘ C until 3/4 of the log growth phase. The comparison of the Simple three lipid component models to the complex 14-lipid component model Avanti over a broad range of physiologically relevant temperatures revealed that the balance of lipid tail unsaturation and propanylation in different positions and inclusion of lipid tails of various length maintain realistic values for lipid mobility, membrane area compressibility, lipid ordering, lipid volume and area, and the bilayer thickness. The only Simple model that was able to satisfactory reproduce most of the structural properties of the complex Avanti model showed worse agreement of the activation energy of basal water permeation with the here performed measurements. The Martini3 models reflect extremely well both experimental and atomistic behavior of the E. coli polar lipid extract membranes. Aquaporin-1 embedded in our native(-like) membranes causes partial lipid ordering and membrane thinning in its vicinity. Moreover, aquaporin-1 attracts and temporarily binds negatively charged lipids, mainly cardiolipins, with a distinct cardiolipin binding site in the crevice at the contact site between two monomers, most probably stabilizing the tetrameric protein assembly. Conclusions The here prepared and validated membrane models of E. coli polar lipids extract revealed that lipid tail complexity, in terms of double bond and cyclopropane location and varying lipid tail length, is key to stabilize membrane properties over a broad temperature range. In addition, they build a solid basis for manifold future simulation studies on more realistic lipid membranes bridging the gap between simulations and experiments.