Multiscale simulations of heterogeneous model membranes

This review will focus on computer modeling aimed at providing insights into the existence, structure, size, and thermodynamic stability of localized domains in membranes of heterogeneous composition. Modeling the lateral organization within a membrane is problematic due to the relatively slow lateral diffusion rate for lipid molecules so that microsecond or longer time scales are needed to fully model the formation and stability of a raft in a membrane. Although atomistic simulations currently are not able to reach this scale, they can provide data on the intermolecular forces and correlations that are involved in lateral organization. These data can be used to define coarse grained models that are capable of predictions of lateral organization in membranes. In this paper, we review modeling efforts that use interaction data from MD simulations to construct coarse grained models for heterogeneous bilayers. In this review we will discuss MD simulations done with the aim of gaining the information needed to build accurate coarse-grained models. We will then review some of the coarse-graining work, emphasizing modeling that has resulted from or has a basis in atomistic simulations.     

Detailed molecular dynamics simulations of model biological membranes containing cholesterol

Detailed molecular dynamics simulations performed to study the nature of lipid raft domains that appear in model membranes are reviewed in this paper. The described simulations were performed on hydrated bilayers containing binary mixtures of cholesterol with phospholipids and also on ternary mixtures containing cholesterol, a phospholipid with a high main transition temperature T_m, and a phospholipid with a low transition temperature T_m. These simulations provide qualitative and semi-quantitative information about cholesterol-lipid interactions and also a testing ground for major assumptions made to explain the nature of lipid rafts in model membranes.     

Lipid dependence of ABC transporter localization and function

Lipid rafts have been implicated in many cellular functions, including protein and lipid transport and signal transduction. ATP-binding cassette (ABC) transporters have also been localized in these membrane domains. In this review the evidence for this specific localization will be evaluated and discussed in terms of relevance to ABC transporter function. We will focus on three ABC transporters of the A, B and C subfamily, respectively. Two of these transporters are relevant to multidrug resistance in tumor cells (Pgp/ABCB1 and MRP1/ABCC1), while the third (ABCA1) is extensively studied in relation to the reverse cholesterol pathway and cellular cholesterol homeostasis. We will attempt to derive a generalized model of lipid rafts to which they associate based on the use of various different lipid raft isolation procedures. In the context of lipid rafts, modulation of ABC transporter localization and function by two relevant lipid classes, i.e. sphingolipids and cholesterol, will be discussed.     

Ordering effects of cholesterol and its analogues

Without any exaggeration, cholesterol is one of the most important lipid species in eukaryotic cells. Its effects on cellular membranes and functions range from purely mechanistic to complex metabolic ones, besides which it is also a precursor of the sex hormones (steroids) and several vitamins. In this review, we discuss the biophysical effects of cholesterol on the lipid bilayer, in particular the ordering and condensing effects, concentrating on the molecular level or inter-atomic interactions perspective, starting from two-component systems and proceeding to many-component ones e.g., modeling lipid rafts. Particular attention is paid to the roles of the methyl groups in the cholesterol ring system, and their possible biological function. Although our main research methodology is computer modeling, in this review we make extensive comparisons between experiments and different modeling approaches.     

Role of cholesterol in lipid raft formation: lessons from lipid model systems

Biochemical and cell-biological experiments have identified cholesterol as an important component of lipid ‘rafts’ and related structures (e.g., caveolae) in mammalian cell membranes, and membrane cholesterol levels as a key factor in determining raft stability and organization. Studies using cholesterol-containing bilayers as model systems have provided important insights into the roles that cholesterol plays in determining lipid raft behavior. This review will discuss recent progress in understanding two aspects of lipid-cholesterol interactions that are particularly relevant to understanding the formation and properties of lipid rafts. First, we will consider evidence that cholesterol interacts differentially with different membrane lipids, associating particularly strongly with saturated, high-melting phospho- and sphingolipids and particularly weakly with highly unsaturated lipid species. Second, we will review recent progress in reconstituting and directly observing segregated raft-like (liquid-ordered) domains in model membranes that mimic the lipid compositions of natural membranes incorporating raft domains.     

Pressure variation of the lateral diffusion in lipid bilayer membranes

A pressure-induced decrease of the lateral diffusion in pure and cholesterol containing phosphatidylcholine bilayer membranes has been determined by the excimer formation technique using pyrene as probe molecule. The experimental results at pressures up to 150 bars are described satisfactorily by the free volume theory of a molecular transport in liquids. A pressure increase of extrapolated 575 bars decreases the lateral diffusion of lipids by a factor of two in pure dipalmitoylphosphatidylcholine membranes. Higher pressures are necessary to induce the same effect in cholesterol containing membranes. This result is interpreted by the condensing effect of cholesterol in fluid bilayer membranes.     

Electroporation of heterogeneous lipid membranes

Electroporation is the basis for the transfection of genetic material and for drug delivery to cells, including electrochemotherapy for cancer. By means of molecular dynamics many aspects of membrane electroporation have been unveiled at the molecular detail in simple, homogeneous, lipid bilayers. However, the correspondence of these findings \with the process happening in cell membranes requires, at least, the consideration of laterally structured membranes. Here, I present a systematic molecular dynamics study of bilayers composed of different liquid-ordered and liquid-disordered lipid phases subjected to a transversal electric field. The simulations reveal two significant results. First, the electric field mainly affects the properties of the disordered phases, so that electroporation takes place in these membrane regions. Second, the smaller the disordered domains are, the faster they become electroporated. These findings may have a relevant significance in the experimental application of cell electroporation in vivo since it implies that electro-induced and pore-mediated transport processes occur in particularly small disordered domains of the plasma membrane, thus locally affecting only specific regions of the cell.     

GM1 ganglioside-mediated accumulation of amyloid beta-protein on cell membranes

The conversion of soluble, nontoxic amyloid beta-protein (Abeta) to aggregated, toxic Abeta is the key step in the development of Alzheimer's disease. Liposomal studies proposed that Abeta specifically recognizes a cholesterol-dependent cluster of monosialoganglioside GM1 and a conformationally altered form of Abeta promotes the aggregation of the protein. In this study, the accumulation of Abeta on living cells was investigated for the first time. The interaction of fluorescein-labeled Abeta (FL-Abeta) with rat pheochromocytoma PC12 cells was visualized using confocal laser microscopy. FL-Abeta was found to colocalize with GM1-rich domains on cell membranes and to accumulate in a concentration- and time-dependent manner, leading to cytotoxicity. Cholesterol depletion significantly reduced Abeta accumulation. These observations corroborate the GM1-mediated Abeta accumulation model.     

Functions of the nuclear lamins

Summary Raft membrane domains are envisioned as lateral assemblies of cholesterol and sphingolipids which adopt a liquid-ordered membrane phase. Our understanding of the raft architecture in cell membranes is developing rapidly. The current view describes raft domains as small and highly dynamic subdomains of cell membranes. The size and stability of raft domains are essential parameters which determine the function of raft domains in cells. Here we discuss how the architecture and stability of raft domains is regulated by oligomerisation of raft components and by modulation of their molecular composition.Abbreviations DIGs detergent-insoluble glycolipid-enriched membranes - GPI glycosylphosphatidylinositol - PDZ PSD95/discslarge/ZO-1 - PI phosphoinositide - PIP₂ phosphatidylinositol 4,5-bisphosphate - TCR T-cell receptor     

Phospholipid/cholesterol model membranes: formation of cholesterol crystallites

Experimental data that define conditions under which cholesterol crystallites form in cholesterol/phospholipid model membranes are reviewed. Structural features of the phospholipids that determine cholesterol crystallization include the length and degree of unsaturation of the acyl chains, the presence of charge on the headgroups and interheadgroup hydrogen bonds.     

Distinct localization of lipid rafts and externalized phosphatidylserine at the surface of apoptotic cells

Externalization of phosphatidylserine (PS) takes place in apoptotic cells as well as in viable cells under certain circumstances. Recent studies showed that externalized PS is localized at the lipid raft in viable activated immune cells. We found that lipid rafts and PS existed in a mutually exclusive manner in apoptotic cells. The number of PS-exposing apoptotic cells decreased when lipid rafts were disrupted. BCtheta;, which binds selectively to cholesterol in a cholesterol-rich region, did not effectively recognize lipid rafts of apoptotic cells. Lipid rafts rich in GM1 were successfully prepared from apoptotic cells, but the lipid raft protein LAT was not enriched in the preparation. Furthermore, the amount of PS and phosphatidylethanolamine but not of cholesterol in lipid rafts appeared to change after induction of apoptosis. These results suggest that lipid rafts are structurally modified during apoptosis and, despite being localized differently from PS, are involved in the externalization of PS.     

Amyloid beta-protein interactions with membranes and cholesterol: causes or casualties of Alzheimer's disease

Amyloid beta-protein (Aβ) is thought to be one of the primary factors causing neurodegeneration in Alzheimer's disease (AD). This protein is an amphipathic molecule that perturbs membranes, binds lipids and alters cell function. Several studies have reported that Aβ alters membrane fluidity but the direction of this effect has not been consistently observed and explanations for this lack of consistency are proposed. Cholesterol is a key component of membranes and cholesterol interacts with Aβ in a reciprocal manner. Aβ impacts on cholesterol homeostasis and modification of cholesterol levels alters Aβ expression. In addition, certain cholesterol lowering drugs (statins) appear to reduce the risk of AD in human subjects. However, the role of changes in the total amount of brain cholesterol in AD and the mechanisms of action of statins in lowering the risk of AD are unclear. Here we discuss data on membranes, cholesterol, Aβ and AD, and propose that modification of the transbilayer distribution of cholesterol in contrast to a change in the total amount of cholesterol provides a cooperative environment for Aβ synthesis and accumulation in membranes leading to cell dysfunction including disruption in cholesterol homeostasis.     

Molecular dynamics simulations of local anesthetic articaine in a lipid bilayer

In order to investigate structural and dynamical properties of local anesthetic articaine in a model lipid bilayer, a series of molecular dynamics simulations have been performed. Simulations were carried out for neutral and charged (protonated) forms of articaine inserted in fully hydrated dimyristoylphosphatidylcholine (DMPC) lipid bilayer. For comparison purpose, a fully hydrated DMPC bilayer without articaine was also simulated. The length of each simulation was 200 ns. Various properties of the lipid bilayer systems in the presence of both charged and uncharged forms of articaine taken at two different concentrations have been examined: membrane area per lipid, mass density distributions, order parameters, radial distribution functions, head group tilt, diffusion coefficients, electrostatic potential, etc, and compared with results of previous simulations of DMPC bilayer in the presence of lidocaine. It was shown that addition of both charged and neutral forms of articaine causes increase of the dipole electrostatic potential in the membrane interior.     

Interactions of amphotericin B derivatives with lipid membranes—A molecular dynamics study

Amphotericin B (AmB) is a well-known polyene macrolide antibiotic used to treat systemic fungal infections. AmB targets more efficiently fungal than animal membranes. However, there are only minor differences in the mode of action of AmB against both types of membranes, which is a source of AmB toxicity. In this work, we analyzed interactions of two low toxic derivatives of AmB (SAmE and PAmE), synthesized in our laboratory, with lipid membranes. Molecular dynamics simulations of the lipid bilayers containing ergosterol (fungal cells) or cholesterol (animal cells) and the studied antibiotic molecules were performed to compare the structural and dynamic properties of AmB derivatives and the parent drug inside the membrane. A number of differences was found for AmB and its derivatives' behavior in cholesterol- and ergosterol-containing membranes. We found that PAmE and SAmE can penetrate deeper into the hydrophobic region of the membrane compared to AmB. Modification of the amino and carboxyl group of AmB also resulted in the conformational transition within the antibiotic's polar head. Wobbling dynamics differentiation, depending on the sterol present, was discovered for the AmB derivatives. These differences may be interpreted as molecular factors responsible for the improved selectivity observed macroscopically for the studied AmB derivatives.     

Lateral diffusion of erythrocyte phospholipids in model membranes comparison between inner and outer leaflet components

The physical properties of lipid bilayers with a similar composition to the outer and inner leaflets of the human erythrocyte membrane have been examined in protein-free model systems. The outer leaflet (OL) was represented by a phospholipid mixture containing phosphatidylcholine and sphingomyelin extracted from human erythrocytes, while a mixture of phosphatidylcholine, phosphatidylserine and phosphatidylethanolamine represented the inner leaflet (IL). The ratio of cholesterol to phospholipid was varied in both mixtures. The lateral diffusion coefficient of fluorescent phospholipids diluted in such lipid mixtures was determined by the modulated fringe pattern photobleaching technique. Contrast curves with a single exponential decay, indicative of homogeneous samples, were obtained only for temperatures above 15 °C and for a cholesterol to phospholipid molar ratio below 0.8. The rate of lateral diffusion was approximately five times faster in IL than in OL multilayers, in agreement with former results obtained in human erythrocytes (Morrot et al. 1986). Varying the cholesterol to phospholipid ratio from 0 to 0.8 (mol/mol) enabled us to decrease the diffusion constant by only a factor of approximately 2 for both IL and OL mixtures. The order parameter of a spin-labeled phospholipid was determined in the different systems and found to be systematically smaller in IL mixtures than in OL mixtures. The present study indicates that the difference in lipid diffusivity of the two erythrocyte leaflets may be accounted for solely by a difference in phospholipid composition, and may be independent of cholesterol and protein asymmetry.Abbreviations OL outer leaflet - IL inner leaflet - RBC red blood cell - NBD-PC 1-acyl-2-[12-(7-nitrobenz-2-oxa-1,3-diazol-4-yl) amino] dodecanoyl phosphatidylcholine - NBD-PE 1-acyl-2-[12-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino] dodecanoyl phosphatidylethanolamine - NBD-PS 1-acyl-2-[12-(7-nitrobenz-2-oxy-1,3-diazol-4-yl)amino] dodecanoyl phosphatidylserine - DMPC 1,2 dimyristoyl-sn-glycero-3-phosphocholine - DMPS 1,2 dimyristoyl-snglycero-3-phosphoserine - PC phosphatidyleholine - C/P cholesterol over phospholipid molar ratio - D lateral diffusion coefficient - S order parameter - ESR electron spin resonance - NMR nuclear magnetic resonance - EDTA ethylene diamine tetraacetic acid - TRIS tris-(hydroxymethyl)amino ethane Offprint requests to: P. F Devaux     

Presenilin 1 modifies lipid raft composition of neuronal membranes

Protein-lipid interactions in the nervous system may provide insight into the causes of neurological disorders. In this study, we elucidated if expression of human presenilin 1 (PS1) in a mouse model changes the physico-chemical properties of brain membranes. PS1 is a multifunctional transmembrane protein and part of the γ-secretase complex. This complex is critical for the production of the Alzheimer related amyloid beta peptide. Brain membranes isolated from mice expressing a human wild-type PS1 transgene are less fluid and contain higher cholesterol and sphingomyelin levels. Moreover, our data reveal significant changes in membrane micro-domains and indicate that PS1 induces the formation of lipid rafts.     

Involvement of lipid rafts in macrophage apoptosis induced by cationic liposomes

We have demonstrated that protein kinase Cδ (PKCδ) could be involved in macrophage apoptosis induced by cationic liposomes composed of stearylamine (SA-liposomes), but the detailed mechanism of how SA-liposomes activate PKCδ has remained unclear. In this paper, we clarified whether lipid rafts are involved in the PKCδ activation induced by SA-liposomes. Co-localization of SA-liposomes and Cholera toxin B subunit (CBT), which specifically binds to ganglioside GM1 on lipid rafts, was found by microscopic observation. The incorporation of SA-liposomes into lipid rafts was clearly inhibited by the pretreatment of cells with an agent, 2,6-di-O-methyl-α-cyclodextrin (DM-α-CD) which disrupts lipid rafts. Activation of PKCδ and externalization of phosphatidylserine induced by SA-liposomes were also suppressed by DM-α-CD, which extracts sphingolipids and proteins from lipid rafts. Reactive oxygen species (ROS) generation, which could be involved in the macrophage apoptosis, was also inhibited by DM-α-CD. Furthermore, apoptosis induced by SA-liposomes was clearly inhibited when the cells were pre-treated with DM-α-CD, but not nystatin, a cholesterol-sequestering agent that disrupt lipid rafts. These findings suggest that sphingolipids in lipid rafts are involved in the activation of PKCδ which leads to apoptosis induced by cationic liposomes, SA-liposomes.     

The effect of hydroxylated fatty acid-containing phospholipids in the remodeling of lipid membranes

The synthetic fatty acid 2-hydroxyoleic acid (2OHOA) is an antitumor drug that regulates membrane lipid composition and structure. An important effect of this drug is the restoration of sphingomyelin (SM) levels in cancer cell membranes, where the SM concentration is lower than in non-tumor cells. It is well known that free fatty acid concentration in cell membranes is lower than 5%, and that fatty acid excess is rapidly incorporated into phospholipids. In a recent work, we have considered the effect of free 2OHOA in model membranes in liquid ordered (Lo) and liquid disordered (Ld) phases, by using all-atom molecular dynamics. This study concerns membranes that are modified upon incorporation of 2OHOA into different phospholipids. 2OHOA-containing phospholipids have a permanent effect on lipid membranes, making a Ld membrane surface more compact and less hydrated, whereas the opposite effect is observed in Lo domains. Moreover, the hydroxyl group of fatty acid chains increases the propensity of Ld model membranes to form hexagonal or other non-lamellar structures. This article is part of a Special Issue entitled: Membrane Structure and Function: Relevance in the Cell's Physiology, Pathology and Therapy.     

Susceptibility of plasma lipids to peroxidation

Lipid peroxidation is an old and yet novel subject. It induces membrane disturbance and damage and its products are known to induce the generation of various cytokines and cell signaling. In the present work, the susceptibility and specificity of human plasma lipids to oxidation were studied, aiming specifically at elucidating the effects of oxidation milieu and oxidants. Cholesteryl esters (CEs) and phosphatidylcholines (PCs) were more readily oxidized in plasma than in organic solution under similar conditions. The susceptibilities of PC and free cholesterol (FC) relative to CE to free radical-mediated lipid peroxidation induced by peroxyl radicals and peroxynitrite were smaller in plasma than in organic solution. The higher rate of CE oxidation by free radicals than PC may be accounted for by the physical effects as well as higher content of polyunsaturated lipids in CE than PC. On the contrary, PC was more readily oxidized than CE by lipoxygenases. The lipid hydroperoxides were stable in organic solution but reduced to the corresponding hydroxides in plasma, the rate being much faster for PC hydroperoxides than for CE and FC hydroperoxides. It was confirmed that free radical-mediated oxidation gave both cis,trans and trans,trans, racemic, random hydroperoxides, while that by lipoxygenase gave only regio- and stereo-specific cis,trans-hydroperoxide.     

P4 ATPases - The physiological relevance of lipid flipping transporters

P4 ATPases are integral transmembrane proteins implicated in phospholipid translocation from the exoplasmic to the cytosolic leaflet of biological membranes. Our present knowledge on the cellular physiology of P4 ATPases is mostly derived from studies in the yeast Saccharomyces cerevisiae, where P4 ATPases play a pivotal role in the biogenesis of intracellular transport vesicles, polarized protein transport and protein maturation. In contrast, the physiological and cellular functions of mammalian P4 ATPases are largely unexplored. P4 ATPases act in concert with members of the CDC50 protein family, which are putative β-subunits for P4 ATPases. This review highlights the current status of a slowly emerging research field and emphasizes the contribution of P4 ATPases to the vesicle-generating machinery.