WorldWideScience

Sample records for plasma membrane lipids

  1. Lipid organization of the plasma membrane

    NARCIS (Netherlands)

    Ingólfsson, Helgi I; Melo, Manuel N; van Eerden, Floris J; Arnarez, Clément; Lopez, Cesar A; Wassenaar, Tsjerk A; Periole, Xavier; de Vries, Alex H; Tieleman, D Peter; Marrink, Siewert J

    2014-01-01

    The detailed organization of cellular membranes remains rather elusive. Based on large-scale molecular dynamics simulations, we provide a high-resolution view of the lipid organization of a plasma membrane at an unprecedented level of complexity. Our plasma membrane model consists of 63 different

  2. Lipid organization of the plasma membrane

    NARCIS (Netherlands)

    Ingólfsson, Helgi I; Melo, Manuel N; van Eerden, Floris J; Arnarez, Clément; Lopez, Cesar A; Wassenaar, Tsjerk A; Periole, Xavier; de Vries, Alex H; Tieleman, D Peter; Marrink, Siewert J

    2014-01-01

    The detailed organization of cellular membranes remains rather elusive. Based on large-scale molecular dynamics simulations, we provide a high-resolution view of the lipid organization of a plasma membrane at an unprecedented level of complexity. Our plasma membrane model consists of 63 different li

  3. Preparation of artificial plasma membrane mimicking vesicles with lipid asymmetry.

    Directory of Open Access Journals (Sweden)

    Qingqing Lin

    Full Text Available Lipid asymmetry, the difference in lipid distribution across the lipid bilayer, is one of the most important features of eukaryotic cellular membranes. However, commonly used model membrane vesicles cannot provide control of lipid distribution between inner and outer leaflets. We recently developed methods to prepare asymmetric model membrane vesicles, but facile incorporation of a highly controlled level of cholesterol was not possible. In this study, using hydroxypropyl-α-cyclodextrin based lipid exchange, a simple method was devised to prepare large unilamellar model membrane vesicles that closely resemble mammalian plasma membranes in terms of their lipid composition and asymmetry (sphingomyelin (SM and/or phosphatidylcholine (PC outside/phosphatidylethanolamine (PE and phosphatidylserine (PS inside, and in which cholesterol content can be readily varied between 0 and 50 mol%. We call these model membranes "artificial plasma membrane mimicking" ("PMm" vesicles. Asymmetry was confirmed by both chemical labeling and measurement of the amount of externally-exposed anionic lipid. These vesicles should be superior and more realistic model membranes for studies of lipid-lipid and lipid-protein interaction in a lipid environment that resembles that of mammalian plasma membranes.

  4. Preparation of artificial plasma membrane mimicking vesicles with lipid asymmetry.

    Science.gov (United States)

    Lin, Qingqing; London, Erwin

    2014-01-01

    Lipid asymmetry, the difference in lipid distribution across the lipid bilayer, is one of the most important features of eukaryotic cellular membranes. However, commonly used model membrane vesicles cannot provide control of lipid distribution between inner and outer leaflets. We recently developed methods to prepare asymmetric model membrane vesicles, but facile incorporation of a highly controlled level of cholesterol was not possible. In this study, using hydroxypropyl-α-cyclodextrin based lipid exchange, a simple method was devised to prepare large unilamellar model membrane vesicles that closely resemble mammalian plasma membranes in terms of their lipid composition and asymmetry (sphingomyelin (SM) and/or phosphatidylcholine (PC) outside/phosphatidylethanolamine (PE) and phosphatidylserine (PS) inside), and in which cholesterol content can be readily varied between 0 and 50 mol%. We call these model membranes "artificial plasma membrane mimicking" ("PMm") vesicles. Asymmetry was confirmed by both chemical labeling and measurement of the amount of externally-exposed anionic lipid. These vesicles should be superior and more realistic model membranes for studies of lipid-lipid and lipid-protein interaction in a lipid environment that resembles that of mammalian plasma membranes.

  5. Lateral mobility of plasma membrane lipids in dividing Xenopus eggs

    OpenAIRE

    de Laat, S W; Tetteroo, P.A.T.; Bluemink, J.G.; Dictus, W.J.A.G.; Zoelen, E.J.J. van

    1984-01-01

    The lateral mobility of plasma membrane lipids was analyzed during first cleavage of Xaopus Levis eggs by fluorescence photobleaching recovery (FPR) measurements, using the lipid analogs 5-(N-hexadecanoyl)aminofluorescein (“HEDAF”) and 5-(N-tetradecanoyl)aminofluorescein (“TEDAF”) as probes. The preexisting plasma membrane of the animal side showed an inhomogeneous, dotted fluorescence pattern after labeling and the lateral mobility of both probes used was below the detection limits of the FP...

  6. Plasma membrane organization and function: moving past lipid rafts.

    Science.gov (United States)

    Kraft, Mary L

    2013-09-01

    "Lipid raft" is the name given to the tiny, dynamic, and ordered domains of cholesterol and sphingolipids that are hypothesized to exist in the plasma membranes of eukaryotic cells. According to the lipid raft hypothesis, these cholesterol- and sphingolipid-enriched domains modulate the protein-protein interactions that are essential for cellular function. Indeed, many studies have shown that cellular levels of cholesterol and sphingolipids influence plasma membrane organization, cell signaling, and other important biological processes. Despite 15 years of research and the application of highly advanced imaging techniques, data that unambiguously demonstrate the existence of lipid rafts in mammalian cells are still lacking. This Perspective summarizes the results that challenge the lipid raft hypothesis and discusses alternative hypothetical models of plasma membrane organization and lipid-mediated cellular function.

  7. Revisiting transbilayer distribution of lipids in the plasma membrane.

    Science.gov (United States)

    Murate, Motohide; Kobayashi, Toshihide

    2016-01-01

    Whereas asymmetric transbilayer lipid distribution in the plasma membrane is well recognized, methods to examine the precise localization of lipids are limited. In this review, we critically evaluate the methods that are applied to study transbilayer asymmetry of lipids, summarizing the factors that influence the measurement. Although none of the present methods is perfect, the current application of immunoelectron microscopy-based technique provides a new picture of lipid asymmetry. Next, we summarize the transbilayer distribution of individual lipid in both erythrocytes and nucleated cells. Finally we discuss the concept of the interbilayer communication of lipids.

  8. Lateral mobility of plasma membrane lipids in dividing Xenopus eggs

    NARCIS (Netherlands)

    Laat, S.W. de; Tetteroo, P.A.T.; Bluemink, J.G.; Dictus, W.J.A.G.; Zoelen, E.J.J. van

    1984-01-01

    The lateral mobility of plasma membrane lipids was analyzed during first cleavage of Xaopus Levis eggs by fluorescence photobleaching recovery (FPR) measurements, using the lipid analogs 5-(N-hexadecanoyl)aminofluorescein (“HEDAF”) and 5-(N-tetradecanoyl)aminofluorescein (“TEDAF”) as probes. The pre

  9. Lateral mobility of plasma membrane lipids in dividing Xenopus eggs

    NARCIS (Netherlands)

    Laat, S.W. de; Tetteroo, P.A.T.; Bluemink, J.G.; Dictus, W.J.A.G.; Zoelen, E.J.J. van

    1984-01-01

    The lateral mobility of plasma membrane lipids was analyzed during first cleavage of Xaopus Levis eggs by fluorescence photobleaching recovery (FPR) measurements, using the lipid analogs 5-(N-hexadecanoyl)aminofluorescein (“HEDAF”) and 5-(N-tetradecanoyl)aminofluorescein (“TEDAF”) as probes. The

  10. Plasma membrane lipids and their role in fungal virulence.

    Science.gov (United States)

    Rella, Antonella; Farnoud, Amir M; Del Poeta, Maurizio

    2016-01-01

    There has been considerable evidence in recent years suggesting that plasma membrane lipids are important regulators of fungal pathogenicity. Various glycolipids have been shown to impart virulent properties in several fungal species, while others have been shown to play a role in host defense. In addition to their role as virulence factors, lipids also contribute to other virulence mechanisms such as drug resistance, biofilm formation, and release of extracellular vesicles. In addition, lipids also affect the mechanical properties of the plasma membrane through the formation of packed microdomains composed mainly of sphingolipids and sterols. Changes in the composition of lipid microdomains have been shown to disrupt the localization of virulence factors and affect fungal pathogenicity. This review gathers evidence on the various roles of plasma membrane lipids in fungal virulence and how lipids might contribute to the different processes that occur during infection and treatment. Insight into the role of lipids in fungal virulence can lead to an improved understanding of the process of fungal pathogenesis and the development of new lipid-mediated therapeutic strategies.

  11. Analysis of lipid-composition changes in plasma membrane microdomains.

    Science.gov (United States)

    Ogiso, Hideo; Taniguchi, Makoto; Okazaki, Toshiro

    2015-08-01

    Sphingolipids accumulate in plasma membrane microdomain sites, such as caveolae or lipid rafts. Such microdomains are considered to be important nexuses for signal transduction, although changes in the microdomain lipid components brought about by signaling are poorly understood. Here, we applied a cationic colloidal silica bead method to analyze plasma membrane lipids from monolayer cells cultured in a 10 cm dish. The detergent-resistant fraction from the silica bead-coated membrane was analyzed by LC-MS/MS to evaluate the microdomain lipids. This method revealed that glycosphingolipids composed the microdomains as a substitute for sphingomyelin (SM) in mouse embryonic fibroblasts (tMEFs) from an SM synthase 1/2 double KO (DKO) mouse. The rate of formation of the detergent-resistant region was unchanged compared with that of WT-tMEFs. C2-ceramide (Cer) stimulation caused greater elevations in diacylglycerol and phosphatidic acid levels than in Cer levels within the microdomains of WT-tMEFs. We also found that lipid changes in the microdomains of SM-deficient DKO-tMEFs caused by serum stimulation occurred in the same manner as that of WT-tMEFs. This practical method for analyzing membrane lipids will facilitate future comprehensive analyses of membrane microdomain-associated responses.

  12. Lipid signalling dynamics at the β-cell plasma membrane.

    Science.gov (United States)

    Wuttke, Anne

    2015-04-01

    Pancreatic β-cells are clustered in islets of Langerhans and secrete insulin in response to increased concentrations of circulating glucose. Insulin in turn acts on liver, muscle and fat tissue to store energy and normalize the blood glucose level. Inappropriate insulin release may lead to impaired glucose tolerance and diabetes. In addition to glucose, other nutrients, neural stimuli and hormonal stimuli control insulin secretion. Many of these signals are perceived at the plasma membrane, which is also the site where insulin granules undergo exocytosis. Therefore, it is not surprising that membrane lipids play an important role in the regulation of insulin secretion. β-cells release insulin in a pulsatile fashion. Signalling lipids integrate the nutrient and neurohormonal inputs to fine-tune, shape and co-ordinate the pulsatility. An important group of signalling lipids are phosphoinositides and their downstream messengers. This MiniReview will discuss new insights into lipid signalling dynamics in β-cells obtained from live-cell imaging experiments with fluorescent translocation biosensors. The plasma membrane concentration of several phosphoinositides and of their downstream messengers changes rapidly upon nutrient or neurohormonal stimulation. Glucose induces the most complex spatio-temporal patterns, typically involving oscillations of messenger concentrations, which sometimes are locally restricted. The tightly controlled levels of lipid messengers can mediate specific binding of downstream effectors to the plasma membrane, contributing to the appropriate regulation of insulin secretion.

  13. Super-resolution optical microscopy of lipid plasma membrane dynamics.

    Science.gov (United States)

    Eggeling, Christian

    2015-01-01

    Plasma membrane dynamics are an important ruler of cellular activity, particularly through the interaction and diffusion dynamics of membrane-embedded proteins and lipids. FCS (fluorescence correlation spectroscopy) on an optical (confocal) microscope is a popular tool for investigating such dynamics. Unfortunately, its full applicability is constrained by the limited spatial resolution of a conventional optical microscope. The present chapter depicts the combination of optical super-resolution STED (stimulated emission depletion) microscopy with FCS, and why it is an important tool for investigating molecular membrane dynamics in living cells. Compared with conventional FCS, the STED-FCS approach demonstrates an improved possibility to distinguish free from anomalous molecular diffusion, and thus to give new insights into lipid-protein interactions and the traditional lipid 'raft' theory.

  14. Membrane potential governs lateral segregation of plasma membrane proteins and lipids in yeast.

    Science.gov (United States)

    Grossmann, Guido; Opekarová, Miroslava; Malinsky, Jan; Weig-Meckl, Ina; Tanner, Widmar

    2007-01-10

    The plasma membrane potential is mainly considered as the driving force for ion and nutrient translocation. Using the yeast Saccharomyces cerevisiae as a model organism, we have discovered a novel role of the membrane potential in the organization of the plasma membrane. Within the yeast plasma membrane, two non-overlapping sub-compartments can be visualized. The first one, represented by a network-like structure, is occupied by the proton ATPase, Pma1, and the second one, forming 300-nm patches, houses a number of proton symporters (Can1, Fur4, Tat2 and HUP1) and Sur7, a component of the recently described eisosomes. Evidence is presented that sterols, the main lipid constituent of the plasma membrane, also accumulate within the patchy compartment. It is documented that this compartmentation is highly dependent on the energization of the membrane. Plasma membrane depolarization causes reversible dispersion of the H(+)-symporters, not however of the Sur7 protein. Mitochondrial mutants, affected in plasma membrane energization, show a significantly lower degree of membrane protein segregation. In accordance with these observations, depolarized membranes also considerably change their physical properties (detergent sensitivity).

  15. Plant lipid environment and membrane enzymes: the case of the plasma membrane H+-ATPase.

    Science.gov (United States)

    Morales-Cedillo, Francisco; González-Solís, Ariadna; Gutiérrez-Angoa, Lizbeth; Cano-Ramírez, Dora Luz; Gavilanes-Ruiz, Marina

    2015-04-01

    Several lipid classes constitute the universal matrix of the biological membranes. With their amphipathic nature, lipids not only build the continuous barrier that confers identity to every cell and organelle, but they are also active actors that modulate the activity of the proteins immersed in the lipid bilayer. The plasma membrane H(+)-ATPase, an enzyme from plant cells, is an excellent example of a transmembrane protein whose activity is influenced by the hydrophilic compartments at both sides of the membrane and by the hydrophobic domains of the lipid bilayer. As a result, an extensive documentation of the effect of numerous amphiphiles in the enzyme activity can be found. Detergents, membrane glycerolipids, and sterols can produce activation or inhibition of the enzyme activity. In some cases, these effects are associated with the lipids of the membrane bulk, but in others, a direct interaction of the lipid with the protein is involved. This review gives an account of reports related to the action of the membrane lipids on the H(+)-ATPase activity.

  16. Lipid-protein interactions in plasma membranes of fiber cells isolated from the human eye lens.

    Science.gov (United States)

    Raguz, Marija; Mainali, Laxman; O'Brien, William J; Subczynski, Witold K

    2014-03-01

    The protein content in human lens membranes is extremely high, increases with age, and is higher in the nucleus as compared with the cortex, which should strongly affect the organization and properties of the lipid bilayer portion of intact membranes. To assess these effects, the intact cortical and nuclear fiber cell plasma membranes isolated from human lenses from 41- to 60-year-old donors were studied using electron paramagnetic resonance spin-labeling methods. Results were compared with those obtained for lens lipid membranes prepared from total lipid extracts from human eyes of the same age group [Mainali, L., Raguz, M., O'Brien, W. J., and Subczynski, W. K. (2013) Biochim. Biophys. Acta]. Differences were considered to be mainly due to the effect of membrane proteins. The lipid-bilayer portions of intact membranes were significantly less fluid than lipid bilayers of lens lipid membranes, prepared without proteins. The intact membranes were found to contain three distinct lipid environments termed the bulk lipid domain, boundary lipid domain, and trapped lipid domain. However, the cholesterol bilayer domain, which was detected in cortical and nuclear lens lipid membranes, was not detected in intact membranes. The relative amounts of bulk and trapped lipids were evaluated. The amount of lipids in domains uniquely formed due to the presence of membrane proteins was greater in nuclear membranes than in cortical membranes. Thus, it is evident that the rigidity of nuclear membranes is greater than that of cortical membranes. Also the permeability coefficients for oxygen measured in domains of nuclear membranes were significantly lower than appropriate coefficients measured in cortical membranes. Relationships between the organization of lipids into lipid domains in fiber cells plasma membranes and the organization of membrane proteins are discussed.

  17. Reduction in lateral lipid mobility of lipid bilayer membrane by atmospheric pressure plasma irradiation

    Science.gov (United States)

    Suda, Yoshiyuki; Tero, Ryugo; Yamashita, Ryuma; Yusa, Kota; Takikawa, Hirofumi

    2016-03-01

    Plasma medicine is an emerging research field in which various applications of electrical discharge, especially in the form of nonequilibrium plasma at atmospheric pressure, are examined, for example, the application of plasma to biological targets for various purposes such as selective killing of tumor cells and blood stanching. We have focused on the behavior of an artificial cell membrane system at the solid-liquid interface. To evaluate the lateral lipid mobility, we measured the diffusion coefficient of the supported lipid bilayer (SLB) composed of dioleoylphosphatidylcholine with fluorescence recovery after photobleaching by confocal laser scanning microscopy. It was found that the diffusion coefficient was decreased by plasma irradiation and that the diffusion coefficient decreasing rate proceeded with increasing plasma power. We investigated the effects of stimulation with an equilibrium chemical, H2O2, on the SLB and confirmed that the diffusion coefficient did not change at least up to a H2O2 concentration of 5 mM. These results indicate that transient active species generated by plasma play critical roles in the reduction in SLB fluidity. The effects of the two generated major oxidized lipid species, hydroxyl- or hydroperoxy-phosphatidylcholine (PC) and acyl-chain-truncated PCs terminated with aldehyde or carboxyl group, on lateral lipid mobility are discussed.

  18. Efficient replacement of plasma membrane outer leaflet phospholipids and sphingolipids in cells with exogenous lipids.

    Science.gov (United States)

    Li, Guangtao; Kim, JiHyun; Huang, Zhen; St Clair, Johnna R; Brown, Deborah A; London, Erwin

    2016-12-06

    Our understanding of membranes and membrane lipid function has lagged far behind that of nucleic acids and proteins, largely because it is difficult to manipulate cellular membrane lipid composition. To help solve this problem, we show that methyl-α-cyclodextrin (MαCD)-catalyzed lipid exchange can be used to maximally replace the sphingolipids and phospholipids in the outer leaflet of the plasma membrane of living mammalian cells with exogenous lipids, including unnatural lipids. In addition, lipid exchange experiments revealed that 70-80% of cell sphingomyelin resided in the plasma membrane outer leaflet; the asymmetry of metabolically active cells was similar to that previously defined for erythrocytes, as judged by outer leaflet lipid composition; and plasma membrane outer leaflet phosphatidylcholine had a significantly lower level of unsaturation than phosphatidylcholine in the remainder of the cell. The data also provided a rough estimate for the total cellular lipids residing in the plasma membrane (about half). In addition to such lipidomics applications, the exchange method should have wide potential for investigations of lipid function and modification of cellular behavior by modification of lipids.

  19. Accumulation of raft lipids in T-cell plasma membrane domains engaged in TCR signalling

    DEFF Research Database (Denmark)

    Zech, Tobias; Ejsing, Christer S.; Gaus, Katharina;

    2009-01-01

    domains were also enriched in plasmenyl phosphatidylethanolamine and phosphatidylserine. Modulating the T-cell lipidome with polyunsaturated fatty acids impaired the plasma membrane condensation at TCR signalling foci and resulted in a perturbed molecular lipid composition. These results correlate...... and saturated phosphatidylcholine species as compared with control plasma membrane fragments. This provides, for the first time, direct evidence that TCR activation domains comprise a distinct molecular lipid composition reminiscent of liquid-ordered raft phases in model membranes. Interestingly, TCR activation...

  20. Influence of nonequilibrium lipid transport, membrane compartmentalization, and membrane proteins on the lateral organization of the plasma membrane

    Science.gov (United States)

    Fan, Jun; Sammalkorpi, Maria; Haataja, Mikko

    2010-01-01

    Compositional lipid domains (lipid rafts) in plasma membranes are believed to be important components of many cellular processes. The mechanisms by which cells regulate the sizes, lifetimes, and spatial localization of these domains are rather poorly understood at the moment. We propose a robust mechanism for the formation of finite-sized lipid raft domains in plasma membranes, the competition between phase separation in an immiscible lipid system and active cellular lipid transport processes naturally leads to the formation of such domains. Simulations of a continuum model reveal that the raft size distribution is broad and the average raft size is strongly dependent on the rates of cellular and interlayer lipid transport processes. We demonstrate that spatiotemporal variations in the recycling may enable the cell to localize larger raft aggregates at specific parts along the membrane. Moreover, we show that membrane compartmentalization may further facilitate spatial localization of the raft domains. Finally, we demonstrate that local interactions with immobile membrane proteins can spatially localize the rafts and lead to further clustering.

  1. Lipid domain structure of the plasma membrane revealed by patching of membrane components.

    Science.gov (United States)

    Harder, T; Scheiffele, P; Verkade, P; Simons, K

    1998-05-18

    Lateral assemblies of glycolipids and cholesterol, "rafts," have been implicated to play a role in cellular processes like membrane sorting, signal transduction, and cell adhesion. We studied the structure of raft domains in the plasma membrane of non-polarized cells. Overexpressed plasma membrane markers were evenly distributed in the plasma membrane. We compared the patching behavior of pairs of raft markers (defined by insolubility in Triton X-100) with pairs of raft/non-raft markers. For this purpose we cross-linked glycosyl-phosphatidylinositol (GPI)-anchored proteins placental alkaline phosphatase (PLAP), Thy-1, influenza virus hemagglutinin (HA), and the raft lipid ganglioside GM1 using antibodies and/or cholera toxin. The patches of these raft markers overlapped extensively in BHK cells as well as in Jurkat T-lymphoma cells. Importantly, patches of GPI-anchored PLAP accumulated src-like protein tyrosine kinase fyn, which is thought to be anchored in the cytoplasmic leaflet of raft domains. In contrast patched raft components and patches of transferrin receptor as a non-raft marker were sharply separated. Taken together, our data strongly suggest that coalescence of cross-linked raft elements is mediated by their common lipid environments, whereas separation of raft and non-raft patches is caused by the immiscibility of different lipid phases. This view is supported by the finding that cholesterol depletion abrogated segregation. Our results are consistent with the view that raft domains in the plasma membrane of non-polarized cells are normally small and highly dispersed but that raft size can be modulated by oligomerization of raft components.

  2. Ionic protein-lipid interaction at the plasma membrane: what can the charge do?

    Science.gov (United States)

    Li, Lunyi; Shi, Xiaoshan; Guo, Xingdong; Li, Hua; Xu, Chenqi

    2014-03-01

    Phospholipids are the major components of cell membranes, but they have functional roles beyond forming lipid bilayers. In particular, acidic phospholipids form microdomains in the plasma membrane and can ionically interact with proteins via polybasic sequences, which can have functional consequences for the protein. The list of proteins regulated by ionic protein-lipid interaction has been quickly expanding, and now includes membrane proteins, cytoplasmic soluble proteins, and viral proteins. Here we review how acidic phospholipids in the plasma membrane regulate protein structure and function via ionic interactions, and how Ca(2+) regulates ionic protein-lipid interactions via direct and indirect mechanisms.

  3. PLASMA-MEMBRANE LIPID ALTERATIONS INDUCED BY NACL IN WINTER-WHEAT ROOTS

    NARCIS (Netherlands)

    MANSOUR, MMF; VANHASSELT, PR; KUIPER, PJC

    1994-01-01

    A highly enriched plasma membrane fraction was isolated by two phase partitioning from wheat roots (Triticum aestivum L. cv. Vivant) grown with and without 100 mM NaCl. The lipids of the plasma membrane fraction were extracted and characterized. Phosphatidylcholine and phosphatidylethanolamine were

  4. LIPID RAFTS, FLUID/FLUID PHASE SEPARATION, AND THEIR RELEVANCE TO PLASMA MEMBRANE STRUCTURE AND FUNCTION

    OpenAIRE

    Sengupta, Prabuddha; Baird, Barbara; Holowka, David

    2007-01-01

    Novel biophysical approaches combined with modeling and new biochemical data have helped to recharge the lipid raft field and have contributed to the generation of a refined model of plasma membrane organization. In this review, we summarize new information in the context of previous literature to provide new insights into the spatial organization and dynamics of lipids and proteins in the plasma membrane of live cells. Recent findings of large-scale separation of liquid-ordered and liquid-di...

  5. Exclusive photorelease of signalling lipids at the plasma membrane.

    Science.gov (United States)

    Nadler, André; Yushchenko, Dmytro A; Müller, Rainer; Stein, Frank; Feng, Suihan; Mulle, Christophe; Carta, Mario; Schultz, Carsten

    2015-12-21

    Photoactivation of caged biomolecules has become a powerful approach to study cellular signalling events. Here we report a method for anchoring and uncaging biomolecules exclusively at the outer leaflet of the plasma membrane by employing a photocleavable, sulfonated coumarin derivative. The novel caging group allows quantifying the reaction progress and efficiency of uncaging reactions in a live-cell microscopy setup, thereby greatly improving the control of uncaging experiments. We synthesized arachidonic acid derivatives bearing the new negatively charged or a neutral, membrane-permeant coumarin caging group to locally induce signalling either at the plasma membrane or on internal membranes in β-cells and brain slices derived from C57B1/6 mice. Uncaging at the plasma membrane triggers a strong enhancement of calcium oscillations in β-cells and a pronounced potentiation of synaptic transmission while uncaging inside cells blocks calcium oscillations in β-cells and causes a more transient effect on neuronal transmission, respectively. The precise subcellular site of arachidonic acid release is therefore crucial for signalling outcome in two independent systems.

  6. The C-terminal Cytosolic Region of Rim21 Senses Alterations in Plasma Membrane Lipid Composition: INSIGHTS INTO SENSING MECHANISMS FOR PLASMA MEMBRANE LIPID ASYMMETRY.

    Science.gov (United States)

    Nishino, Kanako; Obara, Keisuke; Kihara, Akio

    2015-12-25

    Yeast responds to alterations in plasma membrane lipid asymmetry and external alkalization via the sensor protein Rim21 in the Rim101 pathway. However, the sensing mechanism used by Rim21 remains unclear. Here, we found that the C-terminal cytosolic domain of Rim21 (Rim21C) fused with GFP was associated with the plasma membrane under normal conditions but dissociated upon alterations in lipid asymmetry or external alkalization. This indicates that Rim21C contains a sensor motif. Rim21C contains multiple clusters of charged residues. Among them, three consecutive Glu residues (EEE motif) were essential for Rim21 function and dissociation of Rim21C from the plasma membrane in response to changes in lipid asymmetry. In contrast, positively charged residues adjacent to the EEE motif were required for Rim21C to associate with the membrane. We therefore propose an "antenna hypothesis," in which Rim21C moves to or from the plasma membrane and functions as the sensing mechanism of Rim21.

  7. Partitioning, diffusion, and ligand binding of raft lipid analogs in model and cellular plasma membranes.

    Science.gov (United States)

    Sezgin, Erdinc; Levental, Ilya; Grzybek, Michal; Schwarzmann, Günter; Mueller, Veronika; Honigmann, Alf; Belov, Vladimir N; Eggeling, Christian; Coskun, Unal; Simons, Kai; Schwille, Petra

    2012-07-01

    Several simplified membrane models featuring coexisting liquid disordered (Ld) and ordered (Lo) lipid phases have been developed to mimic the heterogeneous organization of cellular membranes, and thus, aid our understanding of the nature and functional role of ordered lipid-protein nanodomains, termed "rafts". In spite of their greatly reduced complexity, quantitative characterization of local lipid environments using model membranes is not trivial, and the parallels that can be drawn to cellular membranes are not always evident. Similarly, various fluorescently labeled lipid analogs have been used to study membrane organization and function in vitro, although the biological activity of these probes in relation to their native counterparts often remains uncharacterized. This is particularly true for raft-preferring lipids ("raft lipids", e.g. sphingolipids and sterols), whose domain preference is a strict function of their molecular architecture, and is thus susceptible to disruption by fluorescence labeling. Here, we analyze the phase partitioning of a multitude of fluorescent raft lipid analogs in synthetic Giant Unilamellar Vesicles (GUVs) and cell-derived Giant Plasma Membrane Vesicles (GPMVs). We observe complex partitioning behavior dependent on label size, polarity, charge and position, lipid headgroup, and membrane composition. Several of the raft lipid analogs partitioned into the ordered phase in GPMVs, in contrast to fully synthetic GUVs, in which most raft lipid analogs mis-partitioned to the disordered phase. This behavior correlates with the greatly enhanced order difference between coexisting phases in the synthetic system. In addition, not only partitioning, but also ligand binding of the lipids is perturbed upon labeling: while cholera toxin B binds unlabeled GM1 in the Lo phase, it binds fluorescently labeled GMI exclusively in the Ld phase. Fluorescence correlation spectroscopy (FCS) by stimulated emission depletion (STED) nanoscopy on intact

  8. Lateral diffusion of phospholipids in the plasma membrane of soybean protoplasts: Evidence for membrane lipid domains.

    Science.gov (United States)

    Metcalf, T N; Wang, J L; Schindler, M

    1986-01-01

    Fluorescent lipid and phospholipid probes were incorporated at 4 degrees C into soybean protoplasts prepared from cultured soybean (SB-1) cells. Fluorescence microscopy showed that the plasma membrane as well as the nucleus were labeled. Fluorescence redistribution after photobleaching (FRAP) analysis was performed on these cells at 18 degrees C to monitor the lateral mobility of the incorporated probes. After labeling at low concentrations (40 mug/ml) of phosphatidyl-N-(4-nitrobenzo-2-oxa-1,3-diazolyl)ethanolamine (NBD-PtdEtn), a single mobile component was observed with a diffusion coefficient (D) of approximately 3 x 10(-9) cm(2)/sec. After labeling at higher probe concentrations (>/=100 mug/ml), two diffusing species were observed, with diffusion coefficients of approximately 3 x 10(-9) cm(2)/sec ("fast") and approximately 5 x 10(-10) cm(2)/sec ("slow"). Similar results were observed with fluorescent derivatives of phosphatidylcholine and fatty acids. In contrast to these results, parallel analysis of 3T3 fibroblasts, using the same probes and conditions, yielded only a single diffusion component. These results suggest that the soybean plasma membrane may contain two distinct lipid domains in terms of lipid mobility. Consistent with this idea, experiments with soybean protoplasts yielded a single diffusion component under the following conditions: (i) labeling with NBD-PtdEtn (100 mug/ml), FRAP analysis at 37 degrees C (D = 1.1 x 10(-8) cm(2)/sec); (ii) labeling with NBD-PtdEtn (100 mug/ml), FRAP analysis at 18 degrees C in the presence of 2 mM EGTA (D = 4.2 x 10(-9) cm(2)/sec); (iii) labeling with 5-(N-dodecanoyl)aminofluorescein (a short-chain lipid probe), FRAP analysis at 18 degrees C or 37 degrees C (D = 2.5 x 10(-8) cm(2)/sec). These results suggest that the plasma membrane of soybean cells may contain stable immiscible domains of fluid and gel-like lipids.

  9. Influence of plasma-treatments on the structure, superstructure, and function of membrane lipids

    Science.gov (United States)

    Hammer, Malte U.; Forbrig, Enrico; Weltmann, Klaus-Dieter; Reuter, Stephan

    2012-10-01

    Every cell, eu- or prokaryotic, has a membrane as an interface to the environment. Every substance that is applied from outside the cell has to interact with it. This includes plasma-generated reactive species in the liquid cell environment created by plasma-treatment. By the Singer and Nicolson model, proteins are embedded in a lipid bilayer. Proteins are the functional elements, lipids are the structural elements. Due to the amphiphilic nature of the lipids, they form (super-) structures in an aqueous environment. The exact superstructure is determined by a structural parameter of the lipid, its shape. Here, we show experiments on lipids by fluorophore-based liposome assays and raman spectroscopy. The results show a membrane-activity of plasma-born reactive species against lipids and lipid structures. Based on this results and literature, we propose a model for a lesion-forming mechanism in membranes of some reactive species created by plasma-treatment. It is based on a hydrophobic-hydrophilic mismatch due to lipid peroxidization induced by reactive species generated in liquids by plasma-treatment.

  10. Independent mobility of proteins and lipids in the plasma membrane of Escherichia coli.

    Science.gov (United States)

    Nenninger, Anja; Mastroianni, Giulia; Robson, Alexander; Lenn, Tchern; Xue, Quan; Leake, Mark C; Mullineaux, Conrad W

    2014-06-01

    Fluidity is essential for many biological membrane functions. The basis for understanding membrane structure remains the classic Singer-Nicolson model, in which proteins are embedded within a fluid lipid bilayer and able to diffuse laterally within a sea of lipid. Here we report lipid and protein diffusion in the plasma membrane of live cells of the bacterium Escherichia coli, using Fluorescence Recovery after Photobleaching (FRAP) and Total Internal Reflection Fluorescence (TIRF) microscopy to measure lateral diffusion coefficients. Lipid and protein mobility within the membrane were probed by visualizing an artificial fluorescent lipid and a simple model membrane protein consisting of a single membrane-spanning alpha-helix with a Green Fluorescent Protein (GFP) tag on the cytoplasmic side. The effective viscosity of the lipid bilayer is strongly temperature-dependent, as indicated by changes in the lipid diffusion coefficient. Surprisingly, the mobility of the model protein was unaffected by changes in the effective viscosity of the bulk lipid, and TIRF microscopy indicates that it clusters in segregated, mobile domains. We suggest that this segregation profoundly influences the physical behaviour of the protein in the membrane, with strong implications for bacterial membrane function and bacterial physiology.

  11. Tetracyclines increase lipid phosphate phosphatase expression on plasma membranes and turnover of plasma lysophosphatidate.

    Science.gov (United States)

    Tang, Xiaoyun; Zhao, Yuan Y; Dewald, Jay; Curtis, Jonathan M; Brindley, David N

    2016-04-01

    Extracellular lysophosphatidate and sphingosine 1-phosphate (S1P) are important bioactive lipids, which signal through G-protein-coupled receptors to stimulate cell growth and survival. The lysophosphatidate and S1P signals are terminated partly by degradation through three broad-specificity lipid phosphate phosphatases (LPPs) on the cell surface. Significantly, the expression of LPP1 and LPP3 is decreased in many cancers, and this increases the impact of lysophosphatidate and S1P signaling. However, relatively little is known about the physiological or pharmacological regulation of the expression of the different LPPs. We now show that treating several malignant and nonmalignant cell lines with 1 μg/ml tetracycline, doxycycline, or minocycline significantly increased the extracellular degradation of lysophosphatidate. S1P degradation was also increased in cells that expressed high LPP3 activity. These results depended on an increase in the stabilities of the three LPPs and increased expression on the plasma membrane. We tested the physiological significance of these results and showed that treating rats with doxycycline accelerated the clearance of lysophosphatidate, but not S1P, from the circulation. However, administering 100 mg/kg/day doxycycline to mice decreased plasma concentrations of lysophosphatidate and S1P. This study demonstrates a completely new property of tetracyclines in increasing the plasma membrane expression of the LPPs.

  12. Membrane Compartmentalization Reducing the Mobility of Lipids and Proteins within a Model Plasma Membrane.

    Science.gov (United States)

    Koldsø, Heidi; Reddy, Tyler; Fowler, Philip W; Duncan, Anna L; Sansom, Mark S P

    2016-09-01

    The cytoskeleton underlying cell membranes may influence the dynamic organization of proteins and lipids within the bilayer by immobilizing certain transmembrane (TM) proteins and forming corrals within the membrane. Here, we present coarse-grained resolution simulations of a biologically realistic membrane model of asymmetrically organized lipids and TM proteins. We determine the effects of a model of cytoskeletal immobilization of selected membrane proteins using long time scale coarse-grained molecular dynamics simulations. By introducing compartments with varying degrees of restraints within the membrane models, we are able to reveal how compartmentalization caused by cytoskeletal immobilization leads to reduced and anomalous diffusional mobility of both proteins and lipids. This in turn results in a reduced rate of protein dimerization within the membrane and of hopping of membrane proteins between compartments. These simulations provide a molecular realization of hierarchical models often invoked to explain single-molecule imaging studies of membrane proteins.

  13. Plasma lipid pattern and red cell membrane structure in β-thalassemia patients in Jakarta

    Directory of Open Access Journals (Sweden)

    Seruni K.U. Freisleben

    2011-08-01

    Full Text Available Background: Over the last 10 years, we have investigated thalassemia patients in Jakarta to obtain a comprehensive picture of iron overload, oxidative stress, and cell damage.Methods: In blood samples from 15 transfusion-dependent patients (group T, 5 non-transfused patients (group N and 10 controls (group C, plasma lipids and lipoproteins, lipid-soluble vitamin E, malondialdehyde (MDA and thiol status were measured. Isolated eryhtrocyte membranes were investigated with electron paramagnetic resonance (EPR spectroscopy using doxyl-stearic acid and maleimido-proxyl spin lables. Data were analyzed statistically with ANOVA.Results: Plasma triglycerides were higher and cholesterol levels were lower in thalassemic patients compared to controls. Vitamin E, group C: 21.8 vs T: 6.2 μmol/L and reactive thiols (C: 144 vs. T: 61 μmol/L were considerably lower in transfused patients, who exert clear signs of oxidative stress (MDA, C: 1.96 vs T: 9.2 μmol/L and of tissue cell damage, i.e., high transaminases plasma levels. Non-transfused thalassemia patients have slight signs of oxidative stress, but no significant indication of cell damage. Erythrocyte membrane parameters from EPR spectroscopy differ considerably between all groups. In transfusion-dependent patients the structure of the erythrocyte membrane and the gradients of polarity and fluidity are destroyed in lipid domains; binding capacity of protein thiols in the membrane is lower and immobilized.Conclusion: In tranfusion-dependent thalassemic patients, plasma lipid pattern and oxidative stress are associated with structural damage of isolated erythrocyte membranes as measured by EPR spectroscopy with lipid and proteinthiol spin labels. (Med J Indones 2011; 20:178-84Keywords: electron paramagnetic resonance spectroscopy, erythrocyte membrane, lipoproteins, oxidative stress, thalassemia, plasma lipids.

  14. Drosophila Lipophorin Receptors Recruit the Lipoprotein LTP to the Plasma Membrane to Mediate Lipid Uptake.

    Directory of Open Access Journals (Sweden)

    Míriam Rodríguez-Vázquez

    2015-06-01

    Full Text Available Lipophorin, the main Drosophila lipoprotein, circulates in the hemolymph transporting lipids between organs following routes that must adapt to changing physiological requirements. Lipophorin receptors expressed in developmentally dynamic patterns in tissues such as imaginal discs, oenocytes and ovaries control the timing and tissular distribution of lipid uptake. Using an affinity purification strategy, we identified a novel ligand for the lipophorin receptors, the circulating lipoprotein Lipid Transfer Particle (LTP. We show that specific isoforms of the lipophorin receptors mediate the extracellular accumulation of LTP in imaginal discs and ovaries. The interaction requires the LA-1 module in the lipophorin receptors and is strengthened by a contiguous region of 16 conserved amino acids. Lipophorin receptor variants that do not interact with LTP cannot mediate lipid uptake, revealing an essential role of LTP in the process. In addition, we show that lipophorin associates with the lipophorin receptors and with the extracellular matrix through weak interactions. However, during lipophorin receptor-mediated lipid uptake, LTP is required for a transient stabilization of lipophorin in the basolateral plasma membrane of imaginal disc cells. Together, our data suggests a molecular mechanism by which the lipophorin receptors tether LTP to the plasma membrane in lipid acceptor tissues. LTP would interact with lipophorin particles adsorbed to the extracellular matrix and with the plasma membrane, catalyzing the exchange of lipids between them.

  15. The C2 domains of granuphilin are high-affinity sensors for plasma membrane lipids.

    Science.gov (United States)

    Lyakhova, Tatyana A; Knight, Jefferson D

    2014-09-01

    Membrane-targeting proteins are crucial components of many cell signaling pathways, including the secretion of insulin. Granuphilin, also known as synaptotagmin-like protein 4, functions in tethering secretory vesicles to the plasma membrane prior to exocytosis. Granuphilin docks to insulin secretory vesicles through interaction of its N-terminal domain with vesicular Rab proteins; however, the mechanisms of granuphilin plasma membrane targeting and release are less clear. Granuphilin contains two C2 domains, C2A and C2B, that interact with the plasma membrane lipid phosphatidylinositol-(4,5)-bisphosphate [PI(4,5)P2]. The goal of this study was to determine membrane-binding mechanisms, affinities, and kinetics of both granuphilin C2 domains using fluorescence spectroscopic techniques. Results indicate that both C2A and C2B bind anionic lipids in a Ca(2+)-independent manner. The C2A domain binds liposomes containing a physiological mixture of lipids including 2% PI(4,5)P2 or PI(3,4,5)P3 with high affinity (apparent K(d, PIPx) of 2-5 nM), and binds nonspecifically with moderate affinity to anionic liposomes lacking phosphatidylinositol phosphate (PIPx) lipids. The C2B domain binds with sub-micromolar affinity to liposomes containing PI(4,5)P2 but does not have a measurable affinity for background anionic lipids. Both domains can be competed away from their target lipids by the soluble PIPx analog inositol-(1,2,3,4,5,6)-hexakisphosphate (IP6), which is a positive regulator of insulin secretion. Potential roles of these interactions in the docking and release of granuphilin from the plasma membrane are discussed.

  16. Molecular dynamics study of lipid bilayers modeling the plasma membranes of mouse hepatocytes and hepatomas

    Science.gov (United States)

    Andoh, Yoshimichi; Aoki, Noriyuki; Okazaki, Susumu

    2016-02-01

    Molecular dynamics (MD) calculations of lipid bilayers modeling the plasma membranes of normal mouse hepatocytes and hepatomas in water have been performed under physiological isothermal-isobaric conditions (310.15 K and 1 atm). The changes in the membrane properties induced by hepatic canceration were investigated and were compared with previous MD calculations included in our previous study of the changes in membrane properties induced by murine thymic canceration. The calculated model membranes for normal hepatocytes and hepatomas comprised 23 and 24 kinds of lipids, respectively. These included phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, phosphatidylinositol, sphingomyelin, lysophospholipids, and cholesterol. We referred to previously published experimental values for the mole fraction of the lipids adopted in the present calculations. The calculated structural and dynamic properties of the membranes such as lateral structure, order parameters, lateral self-diffusion constants, and rotational correlation times all showed that hepatic canceration causes plasma membranes to become more ordered laterally and less fluid. Interestingly, this finding contrasts with the less ordered structure and increased fluidity of plasma membranes induced by thymic canceration observed in our previous MD study.

  17. Regional differences in the lateral mobility of plasma membrane lipids in a molluscan embryo

    NARCIS (Netherlands)

    Speksnijder, J.E.; Dohmen, M.R.; Tertoolen, L.G.J.; Laat, S.W. de

    1985-01-01

    Regional and temporal differences in plasma membrane lipid mobility have been analyzed during the first three cleavage cycles of the embryo of the polar-lobe-forming mollusc Nassarius reticulatus by the fluorescence photobleaching recovery (FPR) method, using 1,1′-ditetradecyl 3,3,3′,3′-tetramethyli

  18. Regional differences in the lateral mobility of plasma membrane lipids in a molluscan embryo

    NARCIS (Netherlands)

    Speksnijder, J.E.; Dohmen, M.R.; Tertoolen, L.G.J.; Laat, S.W. de

    1985-01-01

    Regional and temporal differences in plasma membrane lipid mobility have been analyzed during the first three cleavage cycles of the embryo of the polar-lobe-forming mollusc Nassarius reticulatus by the fluorescence photobleaching recovery (FPR) method, using 1,1′-ditetradecyl

  19. Lipid asymmetry in plant plasma membranes: phosphate deficiency-induced phospholipid replacement is restricted to the cytosolic leaflet

    DEFF Research Database (Denmark)

    Tjällström, H; Hellgren, Lars; Wieslander, Å;

    2010-01-01

    barrier) and rafts both contain only trace amounts of DGDG, we conclude that this lipid class is not compatible with membrane functions requiring a high degree of lipid order. By not replacing phospholipids site specifically with DGDG, negative functional effects of this lipid in the plasma membrane...... are avoided.-Tjellström, H., Hellgren, L. I., Wieslander, A., Sandelius, A. S. Lipid asymmetry in plant plasma membranes: phosphate deficiency-induced phospholipid replacement is restricted to the cytosolic leaflet.......As in other eukaryotes, plant plasma membranes contain sphingolipids, phospholipids, and free sterols. In addition, plant plasma membranes also contain sterol derivatives and usually 5 mol% DGDG was included. As both the apoplastic plasma membrane leaflet (probably the major water permeability...

  20. Detection of apoptosis through the lipid order of the outer plasma membrane leaflet.

    Science.gov (United States)

    Darwich, Zeinab; Klymchenko, Andrey S; Kucherak, Oleksandr A; Richert, Ludovic; Mély, Yves

    2012-12-01

    Cell plasma membranes of living cells maintain their asymmetry, so that the outer leaflet presents a large quantity of sphingomyelin, which is critical for formation of ordered lipid domains. Here, a recently developed probe based on Nile Red (NR12S) was applied to monitor changes in the lipid order specifically at the outer leaflet of cell membranes. Important key features of NR12S are its ratiometric response exclusively to lipid order (liquid ordered vs. liquid disordered phase) and not to surface charge, the possibility of using it at very low concentrations (10-20nM) and the very simple staining protocol. Cholesterol extraction, oxidation and sphingomyelin hydrolysis were found to red shift the emission spectrum of NR12S, indicating a decrease in the lipid order at the outer plasma membrane leaflet. Remarkably, apoptosis induced by three different agents (actinomycin D, camptothecin, staurosporine) produced very similar spectroscopic effects, suggesting that apoptosis also significantly decreases the lipid order at this leaflet. The applicability of NR12S to detect apoptosis was further validated by fluorescence microscopy and flow cytometry, using the ratio between the blue and red parts of its emission band. Thus, for the first time, an environment-sensitive probe, sensitive to lipid order, is shown to detect apoptosis, suggesting a new concept in apoptosis sensing.

  1. Lipids and glycosphingolipids in caveolae and surrounding plasma membrane of primary rat adipocytes.

    Science.gov (United States)

    Ortegren, Unn; Karlsson, Margareta; Blazic, Natascha; Blomqvist, Maria; Nystrom, Fredrik H; Gustavsson, Johanna; Fredman, Pam; Strålfors, Peter

    2004-05-01

    We have made a comprehensive and quantitative analysis of the lipid composition of caveolae from primary rat fat cells and compared the composition of plasma membrane inside and outside caveolae. We isolated caveolae from purified plasma membranes using ultrasonication in carbonate buffer to disrupt the membrane, or extraction with nonionic detergent, followed by density gradient ultracentrifugation. The carbonate-isolated caveolae fraction was further immunopurified using caveolin antibodies. Carbonate-isolated caveolae were enriched in cholesterol and sphingomyelin, and the concentration was three- and twofold higher, respectively, in caveolae compared to the surrounding plasma membrane. The concentration of glycerophospholipids was similar suggesting that glycerophospholipids constitute a constant core throughout the plasma membrane. The composition of detergent-insoluble fractions of the plasma membrane was very variable between preparations, but strongly enriched in sphingomyelin and depleted of glycerophospholipids compared to carbonate-isolated caveolae; indicating that detergent extraction is not a suitable technique for caveolae preparation. An average adipocyte caveola contained about 22 x 10(3) molecules of cholesterol, 7.5 x 10(3) of sphingomyelin and 23 x 10(3) of glycerophospholipid. The glycosphingolipid GD3 was highly enriched in caveolae, whereas GM3, GM1 and GD1a were present inside as well as outside the caveolae membrane. GD1b, GT1b, GM2, GQ1b, sulfatide and lactosylceramide sulfate were not detected in caveolae.

  2. Intrinsic stability of Brassicaceae plasma membrane in relation to changes in proteins and lipids as a response to salinity.

    Science.gov (United States)

    Chalbi, Najla; Martínez-Ballesta, Ma Carmen; Youssef, Nabil Ben; Carvajal, Micaela

    2015-03-01

    Changes in plasma membrane lipids, such as sterols and fatty acids, have been observed as a result of salt stress. These alterations, together with modification of the plasma membrane protein profile, confer changes in the physical properties of the membrane to be taken into account for biotechnological uses. In our experiments, the relationship between lipids and proteins in three different Brassicaceae species differing in salinity tolerance (Brassica oleracea, B. napus and Cakile maritima) and the final plasma membrane stability were studied. The observed changes in the sterol (mainly an increase in sitosterol) and fatty acid composition (increase in RUFA) in each species led to physical adaptation of the plasma membrane to salt stress. The in vitro vesicles stability was higher in the less tolerant (B. oleracea) plants together with low lipoxygenase activity. These results indicate that the proteins/lipids ratio and lipid composition is an important aspect to take into account for the use of natural vesicles in plant biotechnology.

  3. Lipid raft-dependent plasma membrane repair interferes with the activation of B lymphocytes.

    Science.gov (United States)

    Miller, Heather; Castro-Gomes, Thiago; Corrotte, Matthias; Tam, Christina; Maugel, Timothy K; Andrews, Norma W; Song, Wenxia

    2015-12-21

    Cells rapidly repair plasma membrane (PM) damage by a process requiring Ca(2+)-dependent lysosome exocytosis. Acid sphingomyelinase (ASM) released from lysosomes induces endocytosis of injured membrane through caveolae, membrane invaginations from lipid rafts. How B lymphocytes, lacking any known form of caveolin, repair membrane injury is unknown. Here we show that B lymphocytes repair PM wounds in a Ca(2+)-dependent manner. Wounding induces lysosome exocytosis and endocytosis of dextran and the raft-binding cholera toxin subunit B (CTB). Resealing is reduced by ASM inhibitors and ASM deficiency and enhanced or restored by extracellular exposure to sphingomyelinase. B cell activation via B cell receptors (BCRs), a process requiring lipid rafts, interferes with PM repair. Conversely, wounding inhibits BCR signaling and internalization by disrupting BCR-lipid raft coclustering and by inducing the endocytosis of raft-bound CTB separately from BCR into tubular invaginations. Thus, PM repair and B cell activation interfere with one another because of competition for lipid rafts, revealing how frequent membrane injury and repair can impair B lymphocyte-mediated immune responses.

  4. Inhibition of HIV-1 endocytosis allows lipid mixing at the plasma membrane, but not complete fusion

    Directory of Open Access Journals (Sweden)

    de la Vega Michelle

    2011-12-01

    Full Text Available Abstract Background We recently provided evidence that HIV-1 enters HeLa-derived TZM-bl and lymphoid CEMss cells by fusing with endosomes, whereas its fusion with the plasma membrane does not proceed beyond the lipid mixing step. The mechanism of restriction of HIV-1 fusion at the cell surface and/or the factors that aid the virus entry from endosomes remain unclear. Results We examined HIV-1 fusion with a panel of target cells lines and with primary CD4+ T cells. Kinetic measurements of fusion combined with time-resolved imaging of single viruses further reinforced the notion that HIV-1 enters the cells via endocytosis and fusion with endosomes. Furthermore, we attempted to deliberately redirect virus fusion to the plasma membrane, using two experimental strategies. First, the fusion reaction was synchronized by pre-incubating the viruses with cells at reduced temperature to allow CD4 and coreceptors engagement, but not the virus uptake or fusion. Subsequent shift to a physiological temperature triggered accelerated virus uptake followed by entry from endosomes, but did not permit fusion at the cell surface. Second, blocking HIV-1 endocytosis by a small-molecule dynamin inhibitor, dynasore, resulted in transfer of viral lipids to the plasma membrane without any detectable release of the viral content into the cytosol. We also found that a higher concentration of dynasore is required to block the HIV-endosome fusion compared to virus internalization. Conclusions Our results further support the notion that HIV-1 enters disparate cell types through fusion with endosomes. The block of HIV-1 fusion with the plasma membrane at a post-lipid mixing stage shows that this membrane is not conducive to fusion pore formation and/or enlargement. The ability of dynasore to interfere with the virus-endosome fusion suggests that dynamin could be involved in two distinct steps of HIV-1 entry - endocytosis and fusion within intracellular compartments.

  5. C2 domain of synaptotagminⅠassociates with lipid rafts of plasma membrane

    Institute of Scientific and Technical Information of China (English)

    L(U) JiHua; HE Li; SUI SenFang

    2008-01-01

    In this paper we report that the C2 domain of synaptotagmin I (syt I) could associate with lipid rafts of plasma membrane. We demonstrate that phosphatidylinositol 4,5-bisphosphate (PIP2) in the target membrane and Ca2+ are the key factors to enhance the raft association of the C2 domain. We also found that the raft association of the C2 domain could be fulfilled by either C2A or C2B alone, suggesting that their raft association might be complementary. Finally, we indicate that destroying lipid rafts or blocking syt I-raft association could significantly reduce the Ca2+-driven release of glutamates. Our data indicate that the raft association of the C2 domain might play an important role in the regulated exocytosis.

  6. Effects of Aluminum on ATPase Activity and Lipid Composition of Plasma Membranes from Wheat Roots

    Institute of Scientific and Technical Information of China (English)

    HE Long-fei; LIU You-liang; SHEN Zhen-guo; WANG Ai-qin

    2002-01-01

    The effects of aluminum on ATPase activity and lipid composition of the plasma membranes isolated from root tips of Al-tolerant (Altas 66) or Al-sensitive (Scout 66) cultivar of Triticum aestivum L.was assayed. The results showed that both cultivars had similar changes in H+ -ATPase and Ca2+ -ATPase activities after aluminum treatment. Exposure of both cultivars to 20 and 100 (mol/L aluminum for 5 d significantly decreased the activities of Ca2+ -ATPase of plasma membranes. The activities of H+-ATPasc in plasma membrane increased under 20 μmol/L aluminum and decreased at 100 μmol/L aluminum. With aluminum treatment, the PL content of plasma membrane decreased, but GL content increased. The ratio of PL to GL decreased more distinctly in Scout 66 than that in Altas 66. Treated with 20 and 100 μmol/L aluminum, linolenic acid content and the index of unsaturated fatty acids decreaced greatly in Scout 66, but the index of unsaturated fatty acids in Altas 66 increased slightly.

  7. Plasma membranes as heat stress sensors: from lipid-controlled molecular switches to therapeutic applications.

    Science.gov (United States)

    Török, Zsolt; Crul, Tim; Maresca, Bruno; Schütz, Gerhard J; Viana, Felix; Dindia, Laura; Piotto, Stefano; Brameshuber, Mario; Balogh, Gábor; Péter, Mária; Porta, Amalia; Trapani, Alfonso; Gombos, Imre; Glatz, Attila; Gungor, Burcin; Peksel, Begüm; Vigh, László; Csoboz, Bálint; Horváth, Ibolya; Vijayan, Mathilakath M; Hooper, Phillip L; Harwood, John L; Vigh, László

    2014-06-01

    The classic heat shock (stress) response (HSR) was originally attributed to protein denaturation. However, heat shock protein (Hsp) induction occurs in many circumstances where no protein denaturation is observed. Recently considerable evidence has been accumulated to the favor of the "Membrane Sensor Hypothesis" which predicts that the level of Hsps can be changed as a result of alterations to the plasma membrane. This is especially pertinent to mild heat shock, such as occurs in fever. In this condition the sensitivity of many transient receptor potential (TRP) channels is particularly notable. Small temperature stresses can modulate TRP gating significantly and this is influenced by lipids. In addition, stress hormones often modify plasma membrane structure and function and thus initiate a cascade of events, which may affect HSR. The major transactivator heat shock factor-1 integrates the signals originating from the plasma membrane and orchestrates the expression of individual heat shock genes. We describe how these observations can be tested at the molecular level, for example, with the use of membrane perturbers and through computational calculations. An important fact which now starts to be addressed is that membranes are not homogeneous nor do all cells react identically. Lipidomics and cell profiling are beginning to address the above two points. Finally, we observe that a deregulated HSR is found in a large number of important diseases where more detailed knowledge of the molecular mechanisms involved may offer timely opportunities for clinical interventions and new, innovative drug treatments. This article is part of a Special Issue entitled: Membrane Structure and Function: Relevance in the Cell's Physiology, Pathology and Therapy.

  8. Revisiting Plant Plasma Membrane Lipids in Tobacco: A Focus on Sphingolipids.

    Science.gov (United States)

    Cacas, Jean-Luc; Buré, Corinne; Grosjean, Kevin; Gerbeau-Pissot, Patricia; Lherminier, Jeannine; Rombouts, Yoann; Maes, Emmanuel; Bossard, Claire; Gronnier, Julien; Furt, Fabienne; Fouillen, Laetitia; Germain, Véronique; Bayer, Emmanuelle; Cluzet, Stéphanie; Robert, Franck; Schmitter, Jean-Marie; Deleu, Magali; Lins, Laurence; Simon-Plas, Françoise; Mongrand, Sébastien

    2016-01-01

    The lipid composition of plasma membrane (PM) and the corresponding detergent-insoluble membrane (DIM) fraction were analyzed with a specific focus on highly polar sphingolipids, so-called glycosyl inositol phosphorylceramides (GIPCs). Using tobacco (Nicotiana tabacum) 'Bright Yellow 2' cell suspension and leaves, evidence is provided that GIPCs represent up to 40 mol % of the PM lipids. Comparative analysis of DIMs with the PM showed an enrichment of 2-hydroxylated very-long-chain fatty acid-containing GIPCs and polyglycosylated GIPCs in the DIMs. Purified antibodies raised against these GIPCs were further used for immunogold-electron microscopy strategy, revealing the distribution of polyglycosylated GIPCs in domains of 35 ± 7 nm in the plane of the PM. Biophysical studies also showed strong interactions between GIPCs and sterols and suggested a role for very-long-chain fatty acids in the interdigitation between the two PM-composing monolayers. The ins and outs of lipid asymmetry, raft formation, and interdigitation in plant membrane biology are finally discussed.

  9. Low temperature alters plasma membrane lipid composition and ATPase activity of pineapple fruit during blackheart development.

    Science.gov (United States)

    Zhou, Yuchan; Pan, Xiaoping; Qu, Hongxia; Underhill, Steven J R

    2014-02-01

    Plasma membrane (PM) plays central role in triggering primary responses to chilling injury and sustaining cellular homeostasis. Characterising response of membrane lipids to low temperature can provide important information for identifying early causal factors contributing to chilling injury. To this end, PM lipid composition and ATPase activity were assessed in pineapple fruit (Ananas comosus) in relation to the effect of low temperature on the development of blackheart, a form of chilling injury. Chilling temperature at 10 °C induced blackheart development in concurrence with increase in electrolyte leakage. PM ATPase activity was decreased after 1 week at low temperature, followed by a further decrease after 2 weeks. The enzyme activity was not changed during 25 °C storage. Loss of total PM phospholipids was found during postharvest senescence, but more reduction was shown from storage at 10 °C. Phosphatidylcholine and phosphatidylethanolamine were the predominant PM phospholipid species. Low temperature increased the level of phosphatidic acid but decreased the level of phosphatidylinositol. Both phospholipid species were not changed during storage at 25 °C. Postharvest storage at both temperatures decreased the levels of C18:3 and C16:1, and increased level of C18:1. Low temperature decreased the level of C18:2 and increased the level of C14:0. Exogenous application of phosphatidic acid was found to inhibit the PM ATPase activity of pineapple fruit in vitro. Modification of membrane lipid composition and its effect on the functional property of plasma membrane at low temperature were discussed in correlation with their roles in blackheart development of pineapple fruit.

  10. Lipid rafts are essential for the regulation of SOCE by plasma membrane resident STIM1 in human platelets.

    Science.gov (United States)

    Dionisio, Natalia; Galán, Carmen; Jardín, Isaac; Salido, Ginés M; Rosado, Juan A

    2011-03-01

    STIM1 is a transmembrane protein essential for the activation of store-operated Ca²+ entry (SOCE), a major Ca²+ influx mechanism. STIM1 is either located in the endoplasmic reticulum, communicating the Ca²+ concentration in the stores to plasma membrane channels or in the plasma membrane, where it might sense the extracellular Ca²+ concentration. Plasma membrane-located STIM1 has been reported to mediate the SOCE sensitivity to extracellular Ca²+ through its interaction with Orai1. Here we show that plasma membrane lipid raft domains are essential for the regulation of SOCE by extracellular Ca²+. Treatment of platelets with the SERCA inhibitor thapsigargin (TG) induced Mn²+ entry, which was inhibited by increasing concentrations of extracellular Ca²+. Platelet treatment with methyl-β-cyclodextrin, which removes cholesterol and disrupts the lipid raft domains, impaired the inactivation of Ca²+ entry induced by extracellular Ca²+. Methyl-β-cyclodextrin also abolished translocation of STIM1 to the plasma membrane stimulated by treatment with TG and prevented TG-evoked co-immunoprecipitation between plasma membrane-located STIM1 and the Ca²+ permeable channel Orai1. These findings suggest that lipid raft domains are essential for the inactivation of SOCE by extracellular Ca²+ mediated by the interaction between plasma membrane-located STIM1 and Orai1. Copyright © 2011 Elsevier B.V. All rights reserved.

  11. Molecular dynamics study of lipid bilayers modeling the plasma membranes of normal murine thymocytes and leukemic GRSL cells.

    Science.gov (United States)

    Andoh, Yoshimichi; Okazaki, Susumu; Ueoka, Ryuichi

    2013-04-01

    Molecular dynamics (MD) calculations for the plasma membranes of normal murine thymocytes and thymus-derived leukemic GRSL cells in water have been performed under physiological isothermal-isobaric conditions (310.15K and 1 atm) to investigate changes in membrane properties induced by canceration. The model membranes used in our calculations for normal and leukemic thymocytes comprised 23 and 25 kinds of lipids, respectively, including phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, phosphatidylinositol, sphingomyelin, lysophospholipids, and cholesterol. The mole fractions of the lipids adopted here were based on previously published experimental values. Our calculations clearly showed that the membrane area was increased in leukemic cells, and that the isothermal area compressibility of the leukemic plasma membranes was double that of normal cells. The calculated membranes of leukemic cells were thus considerably bulkier and softer in the lateral direction compared with those of normal cells. The tilt angle of the cholesterol and the conformation of the phospholipid fatty acid tails both showed a lower level of order in leukemic cell membranes compared with normal cell membranes. The lateral radial distribution function of the lipids also showed a more disordered structure in leukemic cell membranes than in normal cell membranes. These observations all show that, for the present thymocytes, the lateral structure of the membrane is considerably disordered by canceration. Furthermore, the calculated lateral self-diffusion coefficient of the lipid molecules in leukemic cell membranes was almost double that in normal cell membranes. The calculated rotational and wobbling autocorrelation functions also indicated that the molecular motion of the lipids was enhanced in leukemic cell membranes. Thus, here we have demonstrated that the membranes of thymocyte leukemic cells are more disordered and more fluid than normal cell membranes.

  12. Effects of Calcium on ATPase Activity and Lipid Composition of Plasma Membranes from Wheat Roots Under Aluminum Stress

    Institute of Scientific and Technical Information of China (English)

    HE Long-fei; SHEN Zhen-guo; LIU You-liang

    2003-01-01

    Effects of calcium on ATPase activities, lipid contents, and fatty acid compositions of plasma membrane from wheat roots were assayed under aluminum stress. The results showed that the increase of calcium concentration in the nutrient solution increased the activity of H+-ATPase and the phospholipid content, decreased the activity of Ca2+-ATPase and the galactolipid of plasma membrane. Owing to the decrease of linolenic acid content, the index of unsaturated fatty acid (IUFA) and index of double bond (DBI) decreased in Altas66. The IUFA and DBI of plasma membrane from Scout66 roots increased because its linolenic acid content increased obviously and its palmitic acid content decreased apparently.

  13. Effects of cholesterol on plasma membrane lipid order in MCF-7 cells by two-photon microscopy

    Science.gov (United States)

    Zeng, Yixiu; Chen, Jianling; Yang, Hongqin; Wang, Yuhua; Li, Hui; Xie, Shusen

    2014-09-01

    Lipid rafts are cholesterol- and glycosphingolipids- enriched microdomains on plasma membrane surface of mammal cells, involved in a variety of cellular processes. Depleting cholesterol from the plasma membrane by drugs influences the trafficking of lipid raft markers. Optical imaging techniques are powerful tools to study lipid rafts in live cells due to its noninvasive feature. In this study, breast cancer cells MCF-7 were treated with different concentrations of MβCD to deplete cholesterol and an environmentally sensitive fluorescence probe, Laurdan was loaded to image lipid order by two-photon microscopy. The generalized polarization (GP) values were calculated to distinguish the lipid order and disorder phase. GP images and GP distributions of native and cholesterol-depleted MCF-7 cells were obtained. Our results suggest that even at low concentration (0.5 mM) of MβCD, the morphology of the MCF-7 cells changes. Small high GP areas (lipid order phase) decrease more rapidly than low GP areas (lipid disorder phase), indicating that lipid raft structure was altered more severely than nonraft domains. The data demonstrates that cholesterol dramatically affect raft coverage and plasma membrane fluidity in living cells.

  14. Biomechanics and thermodynamics of nanoparticle interactions with plasma and endosomal membrane lipids in cellular uptake and endosomal escape.

    Science.gov (United States)

    Peetla, Chiranjeevi; Jin, Shihua; Weimer, Jonathan; Elegbede, Adekunle; Labhasetwar, Vinod

    2014-07-01

    To be effective for cytoplasmic delivery of therapeutics, nanoparticles (NPs) taken up via endocytic pathways must efficiently transport across the cell membrane and subsequently escape from the secondary endosomes. We hypothesized that the biomechanical and thermodynamic interactions of NPs with plasma and endosomal membrane lipids are involved in these processes. Using model plasma and endosomal lipid membranes, we compared the interactions of cationic NPs composed of poly(D,L-lactide-co-glycolide) modified with the dichain surfactant didodecyldimethylammonium bromide (DMAB) or the single-chain surfactant cetyltrimethylammonium bromide (CTAB) vs anionic unmodified NPs of similar size. We validated our hypothesis in doxorubicin-sensitive (MCF-7, with relatively fluid membranes) and resistant breast cancer cells (MCF-7/ADR, with rigid membranes). Despite their cationic surface charges, DMAB- and CTAB-modified NPs showed different patterns of biophysical interaction: DMAB-modified NPs induced bending of the model plasma membrane, whereas CTAB-modified NPs condensed the membrane, thereby resisted bending. Unmodified NPs showed no effects on bending. DMAB-modified NPs also induced thermodynamic instability of the model endosomal membrane, whereas CTAB-modified and unmodified NPs had no effect. Since bending of the plasma membrane and destabilization of the endosomal membrane are critical biophysical processes in NP cellular uptake and endosomal escape, respectively, we tested these NPs for cellular uptake and drug efficacy. Confocal imaging showed that in both sensitive and resistant cells DMAB-modified NPs exhibited greater cellular uptake and escape from endosomes than CTAB-modified or unmodified NPs. Further, paclitaxel-loaded DMAB-modified NPs induced greater cytotoxicity even in resistant cells than CTAB-modified or unmodified NPs or drug in solution, demonstrating the potential of DMAB-modified NPs to overcome the transport barrier in resistant cells. In

  15. Lipid composition of pea (Pisum sativum L. and maize (Zea mays L. root plasma membrane and membrane-bound peroxidase and superoxide dismutase

    Directory of Open Access Journals (Sweden)

    Kukavica Biljana

    2007-01-01

    Full Text Available Plasma membrane was isolated from roots of pea and maize plants and used to analyze POD and SOD isoforms, as well as lipid composition. Among lipids, phospholipids were the main lipid class, with phosphatidylcho­line being the most abundant individual component in both pea and maize plasma membranes. Significant differences between the two plant species were found in the contents of cerebrosides, free sterols, and steryl glycosides. Most maize POD isoforms were with neutral and anionic pI values, but the opposite was observed in pea. While both anionic and cationic SOD isoforms were isolated from maize, only two anionic SOD isoforms were detected in pea.

  16. Ouabain Modulates the Lipid Composition of Hippocampal Plasma Membranes from Rats with LPS-induced Neuroinflammation.

    Science.gov (United States)

    Garcia, Israel José Pereira; Kinoshita, Paula Fernanda; Scavone, Cristoforo; Mignaco, Julio Alberto; Barbosa, Leandro Augusto de Oliveira; Santos, Hérica de Lima

    2015-12-01

    The effects of ouabain (OUA) and lipopolysaccharide (LPS) in vivo on hippocampal membranes (RHM) of Wistar male rats aged 3 months were analyzed. After intraperitoneal (i.p.) injection of OUA only, LPS only, OUA plus LPS, or saline, the content of proteins, phospholipids, cholesterol and gangliosides from RHM was analyzed. The total protein and cholesterol contents of RHM were not significantly affected by OUA or LPS for the experimentally paired groups. In contrast, total phospholipids and gangliosides were strongly modulated by either OUA or LPS treatments. LPS reduced the total phospholipids (roughly 23 %) and increased the total gangliosides (approximately 40 %). OUA alone increased the total phospholipids (around 23 %) and also the total gangliosides (nearly 34 %). OUA pretreatment compensated the LPS-induced changes, preserving the total phospholipids and gangliosides around the same levels of the control. Thus, an acute treatment with OUA not only modulated the composition of hippocampal membranes from 3-month-old rats, but also was apparently able to counteract membrane alterations resulting from LPS-induced neuroinflammation. This study demonstrates for the first time that the OUA capacity modulates the lipid composition of hippocampal plasma membranes from rats with LPS-induced neuroinflammation.

  17. The effect of Amaranth oil on monolayers of artificial lipids and hepatocyte plasma membranes with adrenalin-induced stress.

    Science.gov (United States)

    Yelisyeyeva, O P; Semen, K O; Ostrovska, G V; Kaminskyy, D V; Sirota, T V; Zarkovic, N; Mazur, D; Lutsyk, O D; Rybalchenko, K; Bast, A

    2014-03-15

    In this paper the oil from seeds of Amaranthus cruentus L. (AmO) was shown to be an efficient modulator of the physical chemical properties of artificial lipid and rat hepatocyte plasma membranes. AmO improved the membrane stability, their stress resistance and the adsorption of neurotensin to plasma membranes with the distinct biphasic interactions being observed even after adrenalin stress exposure. The analysis of pro-/antioxidant balance in rat blood revealed a mild prooxidant activity after AmO intake, which was accompanied by accumulation of oxidative destruction products in plasma membranes. This prooxidant action of AmO was corroborated in vitro in an adrenalin autooxidation model. On the other hand, the observed improved resistance to adrenalin stress in AmO supplemented rats was associated with an antioxidant response in blood and plasma membrane studies. The AmO effects can be attributed to the modulation of the metabolic pathways involved into oxygen and free radical homeostasis.

  18. Plasma membrane lipid alterations associated with cold acclimation of winter rye seedlings (Secale cereale L. cv Puma)

    Energy Technology Data Exchange (ETDEWEB)

    Lynch, D.V.; Steponkus, P.L. (Cornell Univ., Ithaca, NY (USA))

    1987-01-01

    Highly enriched plasma membrane fractions were isolated from leaves of nonacclimated (NA) and acclimated (ACC) rye (Secale cereale L. cv Puma) seedlings. Collectively, free sterols, steryl glucosides, and acylated steryl glucosides constituted > 50 mole % of the total lipid in both NA and ACC plasma membrane fractions. Glucocerebrosides containing hydroxy fatty acids constituted the major glycolipid class of the plasma membrane, accounting for 16 mole % of the total lipid. Phospholipids, primarily phosphatidylcholine and phosphatidylethanolamine with lesser amounts of phosphatidylglycerol, phosphatidic acid, phosphatidylserine, and phosphatidylinositol, comprised only 32 mole% of the total lipid in NA samples. Following cold acclimation, free sterols increased from 33 to 44 mole %, while steryl glucosides and acylated steryl glucosides decreased from 15 to 6 mole % and 4 to 1 mole %, respectively. Sterol analyses of these lipid classes demonstrated that free {beta}-sitosterol increased from 21 to 32 mole % (accounting for the increase in free sterols as a class) at the expense of sterol derivatives containing {beta}-sitosterol. Glucocerebrosides decreased from 16 to 7 mole % of the total lipid following cold acclimation. In addition, the relative proportions of associated hydroxy fatty acids, including 22:0 (h), 24:0 (h), 22:1 (h), and 24:1 (h) were altered. The phospholipid content of the plasma membrane fraction increased to 42 mole % of the total lipid following cold acclimation. Although the relative proportions of the individual phospholipids did not change appreciably after cold acclimation, there were substantial differences in the molecular species. Di-unsaturated molecular species of phosphatidylcholine and phosphatidylethanolamine increased following acclimation. These results demonstrate that cold acclimation results in substantial changes in the lipid composition of the plasma membrane.

  19. Effect of heavy metals on plasma membrane lipids and antioxidant enzymes of Zygophyllum species

    Directory of Open Access Journals (Sweden)

    Amal Ahmed Morsy

    2012-01-01

    Full Text Available Background: Heavy metals are major environmental pollutant when they present in high concentration in soil and have toxic effects on growth and development of plants. Industrial activities result in heavy metal pollution of large areas of land, which greatly affects natural vegetation. Understanding the mechanism of how plants combat heavy metals adverse effects is hence of great importance. Materials and Methods: Two different localities were chosen; one locality was in the vicinity of gypsum factory and the other one was 25 km away from the factory. Two Zygophyllum species (Z. album and Z. coccineum were naturally grown in the studied areas. The effects of soil heavy metal stress on shoot heavy metal concentrations, lipid peroxidation, antioxidant enzyme activities and the root plasma membrane (PM lipid composition were analyzed. Results: Heavy metal concentrations and Lipid peroxidation increased in the shoot of both species grown in the polluted area. The activities of ascorbate oxidase (ASO, guaiacal peroxidase (GPX, ascorbate peroxidase (APX and superoxide dismutase (SOD were increased whereas these of catalase (CAT were decreased in both species under the polluted conditions. PM total lipids, phospholipids, glycolipids and sterols were decreased in Z. album and Z. coccineum as a result of the polluted soil. Heavy metal stress increased phosphatidylethanolamine (PE and decreased phosphatidylinositol (PI and phophatidylglycerol (PG, with no significant change in phosphatidylcholine (PC in the root PM of both species. Phosphatidylserine (PS decreased in the PM of Z. album whereas it increased in the PM of Z. coccineum under the pollution conditions. Heavy metal stress changed the composition and concentration of fatty acids of the root PM, resulting in increased sat/unsat ratio of both species. Conclusion: the results suggest that efficient antioxidant machinery and favorable PM lipid homeostasis are important to enable Zygophyllum species

  20. Bright and photostable push-pull pyrene dye visualizes lipid order variation between plasma and intracellular membranes

    Science.gov (United States)

    Niko, Yosuke; Didier, Pascal; Mely, Yves; Konishi, Gen-Ichi; Klymchenko, Andrey S.

    2016-01-01

    Imaging lipid organization in cell membranes requires advanced fluorescent probes. Here, we show that a recently synthesized push-pull pyrene (PA), similarly to popular probe Laurdan, changes the emission maximum as a function of lipid order, but outperforms it by spectroscopic properties. In addition to red-shifted absorption compatible with common 405 nm diode laser, PA shows higher brightness and much higher photostability than Laurdan in apolar membrane environments. Moreover, PA is compatible with two-photon excitation at wavelengths >800 nm, which was successfully used for ratiometric imaging of coexisting liquid ordered and disordered phases in giant unilamellar vesicles. Fluorescence confocal microscopy in Hela cells revealed that PA efficiently stains the plasma membrane and the intracellular membranes at >20-fold lower concentrations, as compared to Laurdan. Finally, ratiometric imaging using PA reveals variation of lipid order within different cellular compartments: plasma membranes are close to liquid ordered phase of model membranes composed of sphingomyelin and cholesterol, while intracellular membranes are much less ordered, matching well membranes composed of unsaturated phospholipids without cholesterol. These differences in the lipid order were confirmed by fluorescence lifetime imaging (FLIM) at the blue edge of PA emission band. PA probe constitutes thus a new powerful tool for biomembrane research.

  1. Diffusion of lipids and GPI-anchored proteins in actin-free plasma membrane vesicles measured by STED-FCS

    DEFF Research Database (Denmark)

    Schneider, Falk; Waithe, Dominic; Clausen, Mathias P

    2017-01-01

    Diffusion and interaction dynamics of molecules at the plasma membrane play an important role in cellular signalling, and they are suggested to be strongly associated with the actin cytoskeleton. Here, we utilise super-resolution STED microscopy combined with fluorescence correlation spectroscopy...... (STED-FCS) to access and compare the diffusion characteristics of fluorescent lipid analogues and GPI-anchored proteins (GPI-APs) in the live cell plasma membrane and in actin cytoskeleton-free cell-derived giant plasma membrane vesicles (GPMVs). Hindered diffusion of phospholipids and sphingolipids...... forming immobile clusters, both of which disappear in GPMVs. Our data underline the crucial role of the actin cortex in maintaining hindered diffusion modes of many but not all of the membrane molecules, and highlight a powerful experimental approach to decipher specific influences on molecular plasma...

  2. Lateral mobility of plasma membrane lipids in a molluscan egg: Evidence for an animal/vegetal polarity

    NARCIS (Netherlands)

    Laat, S.W. de; Speksnijder, J.E.; Dohmen, M.R.; Zoelen, E. van; Tertoolen, L.G.J.; Bluemink, J.G.

    1984-01-01

    The lateral diffusion of the lipid analog C₁₄-diI (3', 3'-dihexadecylindocarbocyanine iodide) was measured in the plasma membrane of early embryos of the mollusc Nassarius reticulatus using the FPR-(Fluorescence Photobleaching Recovery) method. At almost all stages measured (from

  3. Lateral mobility of plasma membrane lipids in a molluscan egg: Evidence for an animal/vegetal polarity

    NARCIS (Netherlands)

    Laat, S.W. de; Speksnijder, J.E.; Dohmen, M.R.; Zoelen, E. van; Tertoolen, L.G.J.; Bluemink, J.G.

    1984-01-01

    The lateral diffusion of the lipid analog C₁₄-diI (3', 3'-dihexadecylindocarbocyanine iodide) was measured in the plasma membrane of early embryos of the mollusc Nassarius reticulatus using the FPR-(Fluorescence Photobleaching Recovery) method. At almost all stages measured (from fertilized egg up t

  4. Recent progress on lipid lateral heterogeneity in plasma membranes: From rafts to submicrometric domains.

    Science.gov (United States)

    Carquin, Mélanie; D'Auria, Ludovic; Pollet, Hélène; Bongarzone, Ernesto R; Tyteca, Donatienne

    2016-04-01

    The concept of transient nanometric domains known as lipid rafts has brought interest to reassess the validity of the Singer-Nicolson model of a fluid bilayer for cell membranes. However, this new view is still insufficient to explain the cellular control of surface lipid diversity or membrane deformability. During the past decades, the hypothesis that some lipids form large (submicrometric/mesoscale vs nanometric rafts) and stable (>min vs s) membrane domains has emerged, largely based on indirect methods. Morphological evidence for stable submicrometric lipid domains, well-accepted for artificial and highly specialized biological membranes, was further reported for a variety of living cells from prokaryot es to yeast and mammalian cells. However, results remained questioned based on limitations of available fluorescent tools, use of poor lipid fixatives, and imaging artifacts due to non-resolved membrane projections. In this review, we will discuss recent evidence generated using powerful and innovative approaches such as lipid-specific toxin fragments that support the existence of submicrometric domains. We will integrate documented mechanisms involved in the formation and maintenance of these domains, and provide a perspective on their relevance on membrane deformability and regulation of membrane protein distribution. Copyright © 2015 Elsevier Ltd. All rights reserved.

  5. Methods of staining and visualization of sphingolipid enriched and non-enriched plasma membrane regions of Arabidopsis thaliana with fluorescent dyes and lipid analogues

    Directory of Open Access Journals (Sweden)

    Blachutzik Jörg O

    2012-08-01

    Full Text Available Abstract Background Sterols and Sphingolipids form lipid clusters in the plasma membranes of cell types throughout the animal and plant kingdoms. These lipid domains provide a medium for protein signaling complexes at the plasma membrane and are also observed to be principal regions of membrane contact at the inception of infection. We visualized different specific fluorescent lipophilic stains of the both sphingolipid enriched and non-sphingolipid enriched regions in the plasma membranes of live protoplasts of Arabidopsis thaliana. Results Lipid staining protocols for several fluorescent lipid analogues in plants are presented. The most emphasis was placed on successful protocols for the single and dual staining of sphingolipid enriched regions and exclusion of sphingolipid enriched regions on the plasma membrane of Arabidopsis thaliana protoplasts. A secondary focus was placed to ensure that these staining protocols presented still maintain cell viability. Furthermore, the protocols were successfully tested with the spectrally sensitive dye Laurdan. Conclusion Almost all existing staining procedures of the plasma membrane with fluorescent lipid analogues are specified for animal cells and tissues. In order to develop lipid staining protocols for plants, procedures were established with critical steps for the plasma membrane staining of Arabidopsis leaf tissue and protoplasts. The success of the plasma membrane staining protocols was additionally verified by measurements of lipid dynamics by the fluorescence recovery after photobleaching technique and by the observation of new phenomena such as time dependent lipid polarization events in living protoplasts, for which a putative physiological relevance is suggested.

  6. Lipids of the Golgi membrane

    NARCIS (Netherlands)

    van Meer, G.

    1998-01-01

    The thin membrane of the endoplasmic reticulum matures into the thick plasma membrane in the Golgi apparatus. Along the way, the concentrations of cholesterol and sphingolipids increase. Here, Gerrit van Meer discusses how this phenomenon may reflect an intricate lipid-protein sorting machinery. Syn

  7. Lipid domains in intact fiber-cell plasma membranes isolated from cortical and nuclear regions of human eye lenses of donors from different age groups.

    Science.gov (United States)

    Raguz, Marija; Mainali, Laxman; O'Brien, William J; Subczynski, Witold K

    2015-03-01

    The results reported here clearly document changes in the properties and the organization of fiber-cell membrane lipids that occur with age, based on electron paramagnetic resonance (EPR) analysis of lens membranes of clear lenses from donors of age groups from 0 to 20, 21 to 40, and 61 to 80 years. The physical properties, including profiles of the alkyl chain order, fluidity, hydrophobicity, and oxygen transport parameter, were investigated using EPR spin-labeling methods, which also provide an opportunity to discriminate coexisting lipid domains and to evaluate the relative amounts of lipids in these domains. Fiber-cell membranes were found to contain three distinct lipid environments: bulk lipid domain, which appears minimally affected by membrane proteins, and two domains that appear due to the presence of membrane proteins, namely boundary and trapped lipid domains. In nuclear membranes the amount of boundary and trapped phospholipids as well as the amount of cholesterol in trapped lipid domains increased with the donors' age and was greater than that in cortical membranes. The difference between the amounts of lipids in domains uniquely formed due to the presence of membrane proteins in nuclear and cortical membranes increased with the donors' age. It was also shown that cholesterol was to a large degree excluded from trapped lipid domains in cortical membranes. It is evident that the rigidity of nuclear membranes was greater than that of cortical membranes for all age groups. The amount of lipids in domains of low oxygen permeability, mainly in trapped lipid domains, were greater in nuclear than cortical membranes and increased with the age of donors. These results indicate that the nuclear fiber cell plasma membranes were less permeable to oxygen than cortical membranes and become less permeable to oxygen with age. In clear lenses, age-related changes in the lens lipid and protein composition and organization appear to occur in ways that increase fiber

  8. Protein sorting by lipid phase-like domains supports emergent signaling function in B lymphocyte plasma membranes.

    Science.gov (United States)

    Stone, Matthew B; Shelby, Sarah A; Núñez, Marcos F; Wisser, Kathleen; Veatch, Sarah L

    2017-02-01

    Diverse cellular signaling events, including B cell receptor (BCR) activation, are hypothesized to be facilitated by domains enriched in specific plasma membrane lipids and proteins that resemble liquid-ordered phase-separated domains in model membranes. This concept remains controversial and lacks direct experimental support in intact cells. Here, we visualize ordered and disordered domains in mouse B lymphoma cell membranes using super-resolution fluorescence localization microscopy, demonstrate that clustered BCR resides within ordered phase-like domains capable of sorting key regulators of BCR activation, and present a minimal, predictive model where clustering receptors leads to their collective activation by stabilizing an extended ordered domain. These results provide evidence for the role of membrane domains in BCR signaling and a plausible mechanism of BCR activation via receptor clustering that could be generalized to other signaling pathways. Overall, these studies demonstrate that lipid mediated forces can bias biochemical networks in ways that broadly impact signal transduction.

  9. Proteomics of MUC1-containing lipid rafts from plasma membranes and exosomes of human breast carcinoma cells MCF-7.

    Science.gov (United States)

    Staubach, Simon; Razawi, Hanieh; Hanisch, Franz-Georg

    2009-05-01

    Apically expressed human MUC1 is known to become endocytosed and either to re-enter the secretory pathway for recycling to the plasma membrane or to be exported by the cells via the formation of multi-vesicular bodies and the release of exosomes. By using recombinant fusion-tagged MUC1 as a bait protein we followed an anti-myc affinity-based approach for isolating subpopulations of lipid rafts from the plasma membranes and exosomes of MCF-7 breast cancer cells. MUC1(+) lipid rafts were not only found to contain genuine raft proteins (flotillin-1, prohibitin, G protein, annexin A2), but also raft-associated proteins linking these to the cytoskeleton (ezrin/villin-2, profilin II, HSP27, gamma-actin, beta-actin) or proteins in complexes with raft proteins, including the bait protein (HSP60, HSP70). Major overlaps were revealed for the subproteomes of plasma membranous and exosomal lipid raft preparations, indicating that MUC1 is sorted into subpopulations of rafts for its trafficking via flotillin-dependent pathways and export via exosomes.

  10. Effects of moderately enhanced levels of ozone on the acyl lipid composition and dynamical properties of plasma membranes isolated from garden pea (Pisum sativum)

    DEFF Research Database (Denmark)

    Hellgren, Lars; Sellden, G.; Sandelius, A.S.

    2001-01-01

    Plasma membranes were isolated from leaves of 16-day-old garden pea, Pisum sativum L., that had been grown in the absence or presence of 65 nl l(-1) ozone for 4 days prior to membrane isolation, Plasma membranes from ozone-fumigated plants contained significantly more acyl lipids per protein than...... those from leaves of plants grown in filtered air on a molar/weight ratio, The ratio between the major acyl lipids, phosphatidylethanolamine (PE) and phosphatidylcholine (PC), also increased due to the ozone fumigation, while the fatty acid unsaturation level was unaltered in total plasma membrane acyl...... lipids, as well as in PC and PE, The amount of free sterols per protein was unaltered, but the percentage of campesterol increased, concomitant with a decrease in stigmasterol, The dynamical properties of the isolated plasma membranes were assessed using Laurdan fluorescence spectroscopy, which monitors...

  11. Barriers to the free diffusion of proteins and lipids in the plasma membrane.

    Science.gov (United States)

    Trimble, William S; Grinstein, Sergio

    2015-02-01

    Biological membranes segregate into specialized functional domains of distinct composition, which can persist for the entire life of the cell. How separation of their lipid and (glyco)protein components is generated and maintained is not well understood, but the existence of diffusional barriers has been proposed. Remarkably, the physical nature of such barriers and the manner whereby they impede the free diffusion of molecules in the plane of the membrane has rarely been studied in depth. Moreover, alternative mechanisms capable of generating membrane inhomogeneity are often disregarded. Here we describe prototypical biological systems where membrane segregation has been amply documented and discuss the role of diffusional barriers and other processes in the generation and maintenance of their structural and functional compartmentalization.

  12. Scanning STED-FCS reveals spatiotemporal heterogeneity of lipid interaction in the plasma membrane of living cells

    Science.gov (United States)

    Honigmann, Alf; Mueller, Veronika; Ta, Haisen; Schoenle, Andreas; Sezgin, Erdinc; Hell, Stefan W.; Eggeling, Christian

    2014-11-01

    The interaction of lipids and proteins plays an important role in plasma membrane bioactivity, and much can be learned from their diffusion characteristics. Here we present the combination of super-resolution STED microscopy with scanning fluorescence correlation spectroscopy (scanning STED-FCS, sSTED-FCS) to characterize the spatial and temporal heterogeneity of lipid interactions. sSTED-FCS reveals transient molecular interaction hotspots for a fluorescent sphingolipid analogue. The interaction sites are smaller than 80 nm in diameter and lipids are transiently trapped for several milliseconds in these areas. In comparison, newly developed fluorescent phospholipid and cholesterol analogues with improved phase-partitioning properties show more homogenous diffusion, independent of the preference for liquid-ordered or disordered membrane environments. Our results do not support the presence of nanodomains based on lipid-phase separation in the basal membrane of our cultured nonstimulated cells, and show that alternative interactions are responsible for the strong local trapping of our sphingolipid analogue.

  13. The α7 nicotinic acetylcholine receptor function in hippocampal neurons is regulated by the lipid composition of the plasma membrane.

    Science.gov (United States)

    Colón-Sáez, José O; Yakel, Jerrel L

    2011-07-01

    The α7 nicotinic acetylcholine receptors (nAChRs) play an important role in cellular events such as neurotransmitter release, second messenger cascades, cell survival and apoptosis. In addition, they are a therapeutic target for the treatment of neurological disorders such as Alzheimer's disease and schizophrenia, and drugs that potentiate α7 nAChRs through the regulation of desensitization are currently being developed. Recently, these channels were found to be localized into lipid rafts. Here we show that the disruption of lipid rafts in rat primary hippocampal neurons, through cholesterol-scavenging drugs (methyl-β-cyclodextrin) and the enzymatic breakdown of sphingomyelin (sphingomyelinase), results in significant changes in the desensitization kinetics of native and expressed α7 nAChRs. These effects can be prevented by cotreatment with cholesterol and sphingomyelin, and can be mimicked by treatment with cholesterol and sphingomyelin synthesis inhibitors (mevastatin and myriocin, respectively), suggesting that the effects on desensitization kinetics are indeed due to changes in the levels of cholesterol and sphingomyelin in the plasma membrane. These data provide new insights into themechanism of desensitization of α7 nAChRs by providing evidence that the lipid composition of the plasma membrane can modulate the activity of the α7 nAChRs.

  14. Lateral mobility of plasma membrane lipids in Xenopus eggs: Regional differences related to animal/vegetal polarity become extreme upon fertilization

    NARCIS (Netherlands)

    Bluemink, J.G.; Dictus, W.J.A.G.; Zoelen, E.J.J. van; Tetteroo, P.A.T.; Tertoolen, L.G.J.; Laat, S.W. de

    1984-01-01

    Regional differences in the lateral mobility properties of plasma membrane lipids have been studied in unfertilized and fertilizedxaqpus eggs by fluorescence photobleaching recovery (FPR) measurements. Out of a variety of commonly used lipid probes only the aminofluorescein-labeled fatty acids

  15. Lateral mobility of plasma membrane lipids in Xenopus eggs: Regional differences related to animal/vegetal polarity become extreme upon fertilization

    NARCIS (Netherlands)

    Bluemink, J.G.; Dictus, W.J.A.G.; Zoelen, E.J.J. van; Tetteroo, P.A.T.; Tertoolen, L.G.J.; Laat, S.W. de

    1984-01-01

    Regional differences in the lateral mobility properties of plasma membrane lipids have been studied in unfertilized and fertilizedxaqpus eggs by fluorescence photobleaching recovery (FPR) measurements. Out of a variety of commonly used lipid probes only the aminofluorescein-labeled fatty acids HEDAF

  16. Effect of Hypoxia on the Calcium and Magnesium Content, Lipid Peroxidation Level, and Ca2+-ATPase Activity of Syncytiotrophoblast Plasma Membranes from Placental Explants

    Directory of Open Access Journals (Sweden)

    Delia I. Chiarello

    2014-01-01

    Full Text Available In the current study the possible relationship between the Ca2+/Mg2+ ratio of human syncytiotrophoblast plasma membranes and their lipid peroxidation and Ca2+-ATPase activity was determined. Syncytiotrophoblast plasma membranes of placental explants cultured under hypoxia increased their lipid peroxidation and Ca2+ content, diminished their Ca2+-ATPase activity, and kept their Mg2+ content unchanged. Membranes preincubated with different concentrations of Ca2+ increased their Ca2+ content without changes in their Mg2+ content. There is a direct relationship between Ca2+ content and lipid peroxidation of the membranes, as well as an inverse relationship between their Ca2+ content and Ca2+-ATPase activity. On the contrary, preincubation of membranes with different concentrations of Mg2+ showed a higher Mg2+ content without changing their lipid peroxidation and Ca2+-ATPase activity. Explants cultured under hypoxia in the presence of 4 mM MgSO4 showed similar values of lipid peroxidation and Ca2+-ATPase activity of their membranes compared to those of explants cultured under normoxia. Increased Ca2+ content of the membranes by interacting with negatively charged phospholipids could result in destabilizing effects of the membrane structure, exposing hydrocarbon chains of fatty acids to the action of free radicals. Mg2+ might exert a stabilizing effect of the membranes, avoiding their exposure to free radicals.

  17. Membrane-lipid therapy in operation: the HSP co-inducer BGP-15 activates stress signal transduction pathways by remodeling plasma membrane rafts.

    Directory of Open Access Journals (Sweden)

    Imre Gombos

    Full Text Available Aging and pathophysiological conditions are linked to membrane changes which modulate membrane-controlled molecular switches, causing dysregulated heat shock protein (HSP expression. HSP co-inducer hydroxylamines such as BGP-15 provide advanced therapeutic candidates for many diseases since they preferentially affect stressed cells and are unlikely have major side effects. In the present study in vitro molecular dynamic simulation, experiments with lipid monolayers and in vivo ultrasensitive fluorescence microscopy showed that BGP-15 alters the organization of cholesterol-rich membrane domains. Imaging of nanoscopic long-lived platforms using the raft marker glycosylphosphatidylinositol-anchored monomeric green fluorescent protein diffusing in the live Chinese hamster ovary (CHO cell plasma membrane demonstrated that BGP-15 prevents the transient structural disintegration of rafts induced by fever-type heat stress. Moreover, BGP-15 was able to remodel cholesterol-enriched lipid platforms reminiscent of those observed earlier following non-lethal heat priming or membrane stress, and were shown to be obligate for the generation and transmission of stress signals. BGP-15 activation of HSP expression in B16-F10 mouse melanoma cells involves the Rac1 signaling cascade in accordance with the previous observation that cholesterol affects the targeting of Rac1 to membranes. Finally, in a human embryonic kidney cell line we demonstrate that BGP-15 is able to inhibit the rapid heat shock factor 1 (HSF1 acetylation monitored during the early phase of heat stress, thereby promoting a prolonged duration of HSF1 binding to heat shock elements. Taken together, our results indicate that BGP-15 has the potential to become a new class of pharmaceuticals for use in 'membrane-lipid therapy' to combat many various protein-misfolding diseases associated with aging.

  18. Impact of Lipid Composition and Receptor Conformation on the Spatio-temporal Organization of μ-Opioid Receptors in a Multi-component Plasma Membrane Model.

    Science.gov (United States)

    Marino, Kristen A; Prada-Gracia, Diego; Provasi, Davide; Filizola, Marta

    2016-12-01

    The lipid composition of cell membranes has increasingly been recognized as playing an important role in the function of various membrane proteins, including G Protein-Coupled Receptors (GPCRs). For instance, experimental and computational evidence has pointed to lipids influencing receptor oligomerization directly, by physically interacting with the receptor, and/or indirectly, by altering the bulk properties of the membrane. While the exact role of oligomerization in the function of class A GPCRs such as the μ-opioid receptor (MOR) is still unclear, insight as to how these receptors oligomerize and the relevance of the lipid environment to this phenomenon is crucial to our understanding of receptor function. To examine the effect of lipids and different MOR conformations on receptor oligomerization we carried out extensive coarse-grained molecular dynamics simulations of crystal structures of inactive and/or activated MOR embedded in an idealized mammalian plasma membrane composed of 63 lipid types asymmetrically distributed across the two leaflets. The results of these simulations point, for the first time, to specific direct and indirect effects of the lipids, as well as the receptor conformation, on the spatio-temporal organization of MOR in the plasma membrane. While sphingomyelin-rich, high-order lipid regions near certain transmembrane (TM) helices of MOR induce an effective long-range attractive force on individual protomers, both long-range lipid order and interface formation are found to be conformation dependent, with a larger number of different interfaces formed by inactive MOR compared to active MOR.

  19. [Isolation and purification of human blood plasma proteins able to form potassium channels in artificial bilayer lipid membrane].

    Science.gov (United States)

    Venediktova, N I; Kuznetsov, K V; Gritsenko, E N; Gulidova, G P; Mironova, G D

    2012-01-01

    Protein fraction able to induce K(+)-selective transport across bilayer lipid membrane was isolated from human blood plasma with the use of the detergent and proteolytic enzyme-free method developed at our laboratory. After addition of the studied sample to the artificial membrane in the presence of 100 mM KCl, a discrete current change was observed. No channel activity was recorded in the presence of calcium and sodium ions. Channel forming activity of fraction was observed only in the presence of K+. Using a threefold gradient of KCl in the presence of studied proteins the potassium-selective potential balanced by voltage of -29 mV was registered. This value is very close to the theoretical Nernst potential in this case. This means that the examined ion channel is cation-selective. According to data obtained with MS-MALDI-TOF/TOF and database NCBI three protein components were identified in isolated researched sample.

  20. Choroideremia Is a Systemic Disease With Lymphocyte Crystals and Plasma Lipid and RBC Membrane Abnormalities

    Science.gov (United States)

    Zhang, Alice Yang; Mysore, Naveen; Vali, Hojatollah; Koenekoop, Jamie; Cao, Sang Ni; Li, Shen; Ren, Huanan; Keser, Vafa; Lopez-Solache, Irma; Siddiqui, Sorath Noorani; Khan, Ayesha; Mui, Jeannie; Sears, Kelly; Dixon, Jim; Schwartzentruber, Jeremy; Majewski, Jacek; Braverman, Nancy; Koenekoop, Robert K.

    2015-01-01

    Purpose Photoreceptor neuronal degenerations are common, incurable causes of human blindness affecting 1 in 2000 patients worldwide. Only half of all patients are associated with known mutations in over 250 disease genes, prompting our research program to identify the remaining new genes. Most retinal degenerations are restricted to the retina, but photoreceptor degenerations can also be found in a wide variety of systemic diseases. We identified an X-linked family from Sri Lanka with a severe choroidal degeneration and postulated a new disease entity. Because of phenotypic overlaps with Bietti's crystalline dystrophy, which was recently found to have systemic features, we hypothesized that a systemic disease may be present in this new disease as well. Methods For phenotyping, we performed detailed eye exams with in vivo retinal imaging by optical coherence tomography. For genotyping, we performed whole exome sequencing, followed by Sanger sequencing confirmations and cosegregation. Systemic investigations included electron microscopy studies of peripheral blood cells in patients and in normal controls and detailed fatty acid profiles (both plasma and red blood cell [RBC] membranes). Fatty acid levels were compared to normal controls, and only values two standard deviations above or below normal controls were further evaluated. Results The family segregated a REP1 mutation, suggesting choroideremia (CHM). We then found crystals in peripheral blood lymphocytes and discovered significant plasma fatty acid abnormalities and RBC membrane abnormalities (i.e., elevated plasmalogens). To replicate our discoveries, we expanded the cohort to nine CHM patients, genotyped them for REP1 mutations, and found the same abnormalities (crystals and fatty acid abnormalities) in all patients. Conclusions Previously, CHM was thought to be restricted to the retina. We show, to our knowledge for the first time, that CHM is a systemic condition with prominent crystals in lymphocytes and

  1. Cell-penetrating compounds preferentially bind glycosaminoglycans over plasma membrane lipids in a charge density- and stereochemistry-dependent manner.

    Science.gov (United States)

    Prevette, Lisa E; Benish, Nicolas C; Schoenecker, Amber R; Braden, Kristin J

    2015-12-01

    Cell-penetrating compounds (CPCs) are often conjugated to drugs and genes to facilitate cellular uptake. We hypothesize that the electrostatic interaction between the positively charged amines of the cell-penetrating compounds and the negatively charged glycosaminoglycans (GAGs) extending from cell surfaces is the initiating step in the internalization process. The interactions of generation 5 PAMAM dendrimer, Tat peptide and 25 kDa linear PEI with four different GAGs have been studied using isothermal titration calorimetry to elucidate structure-function relationships that could lead to improved drug and gene delivery methods to a wide variety of cell types. Detailed thermodynamic analysis has determined that CPC-GAG binding constants range from 8.7×10(3) to 2.4×10(6)M(-1) and that affinity is dependent upon GAG charge density and stereochemistry and CPC molecular weight. The effect of GAG composition on affinity is likely due to hydrogen bonding between CPC amines and amides and GAG hydroxyl and amine groups. These results were compared to the association of CPCs with lipid vesicles of varying composition as model plasma membranes to finally clarify the relative importance of each cell surface component in initial cell recognition. CPC-lipid affinity increases with anionic lipid content, but GAG affinity is higher for all cell-penetrating compounds, confirming the role these heterogeneous polysaccharides play in cellular association and clustering.

  2. Terpenes increase the lipid dynamics in the Leishmania plasma membrane at concentrations similar to their IC50 values.

    Directory of Open Access Journals (Sweden)

    Heverton Silva Camargos

    Full Text Available Although many terpenes have shown antitumor, antibacterial, antifungal, and antiparasitic activity, the mechanism of action is not well established. Electron paramagnetic resonance (EPR spectroscopy of the spin-labeled 5-doxyl stearic acid revealed remarkable fluidity increases in the plasma membrane of terpene-treated Leishmania amazonensis promastigotes. For an antiproliferative activity assay using 5×10(6 parasites/mL, the sesquiterpene nerolidol and the monoterpenes (+-limonene, α-terpineol and 1,8-cineole inhibited the growth of the parasites with IC50 values of 0.008, 0.549, 0.678 and 4.697 mM, respectively. The IC50 values of these terpenes increased as the parasite concentration used in the cytotoxicity assay increased, and this behavior was examined using a theoretical treatment of the experimental data. Cytotoxicity tests with the same parasite concentration as in the EPR experiments revealed a correlation between the IC50 values of the terpenes and the concentrations at which they altered the membrane fluidity. In addition, the terpenes induced small amounts of cell lysis (4-9% at their respective IC50 values. For assays with high cell concentrations (2×10(9 parasites/mL, the incorporation of terpene into the cell membrane was very fast, and the IC50 values observed for 24 h and 5 min-incubation periods were not significantly different. Taken together, these results suggest that terpene cytotoxicity is associated with the attack on the plasma membrane of the parasite. The in vitro cytotoxicity of nerolidol was similar to that of miltefosine, and nerolidol has high hydrophobicity; thus, nerolidol might be used in drug delivery systems, such as lipid nanoparticles to treat leishmaniasis.

  3. Terpenes increase the lipid dynamics in the Leishmania plasma membrane at concentrations similar to their IC50 values.

    Science.gov (United States)

    Camargos, Heverton Silva; Moreira, Rodrigo Alves; Mendanha, Sebastião Antonio; Fernandes, Kelly Souza; Dorta, Miriam Leandro; Alonso, Antonio

    2014-01-01

    Although many terpenes have shown antitumor, antibacterial, antifungal, and antiparasitic activity, the mechanism of action is not well established. Electron paramagnetic resonance (EPR) spectroscopy of the spin-labeled 5-doxyl stearic acid revealed remarkable fluidity increases in the plasma membrane of terpene-treated Leishmania amazonensis promastigotes. For an antiproliferative activity assay using 5×10(6) parasites/mL, the sesquiterpene nerolidol and the monoterpenes (+)-limonene, α-terpineol and 1,8-cineole inhibited the growth of the parasites with IC50 values of 0.008, 0.549, 0.678 and 4.697 mM, respectively. The IC50 values of these terpenes increased as the parasite concentration used in the cytotoxicity assay increased, and this behavior was examined using a theoretical treatment of the experimental data. Cytotoxicity tests with the same parasite concentration as in the EPR experiments revealed a correlation between the IC50 values of the terpenes and the concentrations at which they altered the membrane fluidity. In addition, the terpenes induced small amounts of cell lysis (4-9%) at their respective IC50 values. For assays with high cell concentrations (2×10(9) parasites/mL), the incorporation of terpene into the cell membrane was very fast, and the IC50 values observed for 24 h and 5 min-incubation periods were not significantly different. Taken together, these results suggest that terpene cytotoxicity is associated with the attack on the plasma membrane of the parasite. The in vitro cytotoxicity of nerolidol was similar to that of miltefosine, and nerolidol has high hydrophobicity; thus, nerolidol might be used in drug delivery systems, such as lipid nanoparticles to treat leishmaniasis.

  4. Electroporation of heterogeneous lipid membranes.

    Science.gov (United States)

    Reigada, Ramon

    2014-03-01

    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.

  5. Criticality in Plasma Membranes

    Science.gov (United States)

    Machta, Benjamin; Papanikolaou, Stefanos; Sethna, James; Veatch, Sarah

    2011-03-01

    We are motivated by recent observations of micron-sized critical fluctuations in the 2d Ising Universality class in plasma membrane vesicles that are isolated from cortical cytoskeleton. We construct a minimal model of the plasma membrane's interaction with intact cytoskeleton which explains why large scale phase separation has not been observed in Vivo. In addition, we use analytical techniques from conformal field theory and numerical simulations to investigate the form of effective forces mediated by the membrane's proximity to criticality. We show that the range of this force is maximized near a critical point and we quantify its usefulness in mediating communication using techniques from information theory. Finally we use theoretical techniques from statistical physics in conjunction with Monte-Carlo simulations to understand how criticality can be used to increase the efficiency of membrane bound receptor mediated signaling. We expect that this sort of analysis will be broadly useful in understanding and quantifying the role of lipid ``rafts'' in a wide variety of membrane bound processes. Generally, we demonstrate that critical fluctuations provide a physical mechanism to organize and spatially segregate membrane components by providing channels for interaction over relatively large distances.

  6. Fusion of liposomes with the plasma membrane of epithelial cells: Fate of incorporated lipids as followed by freeze fracture and autoradiography of plastic sections

    NARCIS (Netherlands)

    Knoll, G.; Burger, K.N.J.; Bron, R.; van Meer, G.; Verkleij, A.J.

    1988-01-01

    The fusion of liposomes with the plasma membrane of influenza virus-infected monolayers of an epithelial cell line, Madin-Darby canine kidney cells (van Meer et al., 1985. Biochemistry, 24: 3593-3602), has been analyzed by morphological techniques. The distribution of liposomal lipids over the apica

  7. Comparison of the lipid composition of oat root and coleoptile plasma membranes: lack of short-term change in response to auxin

    Science.gov (United States)

    Sandstrom, R. P.; Cleland, R. E.

    1989-01-01

    The total lipid composition of plasma membranes (PM), isolated by the phase partitioning method from two different oat (Avena sativa L.) tissues, the root and coleoptile, was compared. In general, the PM lipid composition was not conserved between these two organs of the oat seedling. Oat roots contained 50 mole percent phospholipid, 25 mole percent glycolipid, and 25 mole percent free sterol, whereas comparable amounts in the coleoptile were 42, 39, and 19 mole percent, respectively. Individual lipid components within each lipid class also showed large variations between the two tissues. Maximum specific ATPase activity in the root PM was more than double the activity in the coleoptile. Treatment of coleoptile with auxin for 1 hour resulted in no detectable changes in PM lipids or extractable ATPase activity. Differences in the PM lipid composition between the two tissues that may define the limits of ATPase activity are discussed.

  8. Protein receptor-independent plasma membrane remodeling by HAMLET: a tumoricidal protein-lipid complex.

    Science.gov (United States)

    Nadeem, Aftab; Sanborn, Jeremy; Gettel, Douglas L; James, Ho C S; Rydström, Anna; Ngassam, Viviane N; Klausen, Thomas Kjær; Pedersen, Stine Falsig; Lam, Matti; Parikh, Atul N; Svanborg, Catharina

    2015-11-12

    A central tenet of signal transduction in eukaryotic cells is that extra-cellular ligands activate specific cell surface receptors, which orchestrate downstream responses. This ''protein-centric" view is increasingly challenged by evidence for the involvement of specialized membrane domains in signal transduction. Here, we propose that membrane perturbation may serve as an alternative mechanism to activate a conserved cell-death program in cancer cells. This view emerges from the extraordinary manner in which HAMLET (Human Alpha-lactalbumin Made LEthal to Tumor cells) kills a wide range of tumor cells in vitro and demonstrates therapeutic efficacy and selectivity in cancer models and clinical studies. We identify a ''receptor independent" transformation of vesicular motifs in model membranes, which is paralleled by gross remodeling of tumor cell membranes. Furthermore, we find that HAMLET accumulates within these de novo membrane conformations and define membrane blebs as cellular compartments for direct interactions of HAMLET with essential target proteins such as the Ras family of GTPases. Finally, we demonstrate lower sensitivity of healthy cell membranes to HAMLET challenge. These features suggest that HAMLET-induced curvature-dependent membrane conformations serve as surrogate receptors for initiating signal transduction cascades, ultimately leading to cell death.

  9. Therapeutic MSC exosomes are derived from lipid raft microdomains in the plasma membrane

    Directory of Open Access Journals (Sweden)

    Soon Sim Tan

    2013-12-01

    Full Text Available Background: Mesenchymal stem cell (MSC was previously shown to secrete lipid vesicles that when purified by high performance liquid chromatography as a population of homogenously sized particles with a hydrodynamic radius of 55–65 nm reduce infarct size in a mouse model of myocardial ischemia/reperfusion injury. As these vesicles exhibit many biophysical and biochemical properties of exosomes, they were identified as exosomes. Here we investigated if these lipid vesicles were indeed exosomes that have an endosomal biogenesis. Method: In most cells, endocytosis is thought to occur at specialized microdomains known as lipid rafts. To demonstrate an endosomal origin for MSC exosomes, MSCs were pulsed with ligands e.g. transferrin (Tfs and Cholera Toxin B (CTB that bind receptors in lipid rafts. The endocytosed ligands were then chased to determine if they were incorporated into the exosomes. Results: A fraction of exogenous Tfs was found to recycle into MSC exosomes. When MSCs were pulsed with labelled Tfs in the presence of chlorpromazine, an inhibitor of clathrin-mediated endocytosis, Tf incorporation in CD81-immunoprecipitate was reduced during the chase. CTB which binds GM1 gangliosides that are enriched in lipid rafts extracted exosome-associated proteins, CD81, CD9, Alix and Tsg101 from MSC-conditioned medium. Exogenous CTBs were pulse-chased into secreted vesicles. Extraction of Tf- or CTB-binding vesicles in an exosome preparation mutually depleted each other. Inhibition of sphingomyelinases reduced CTB-binding vesicles. Conclusion: Together, our data demonstrated that MSC exosomes are derived from endocytosed lipid rafts and that their protein cargo includes exosome-associated proteins CD81, CD9, Alix and Tsg101.

  10. Cationic Au Nanoparticle Binding with Plasma Membrane-like Lipid Bilayers: Potential Mechanism for Spontaneous Permeation to Cells Revealed by Atomistic Simulations

    DEFF Research Database (Denmark)

    Heikkila, E.; Martinez-Seara, H.; Gurtovenko, A. A.

    2014-01-01

    Despite being chemically inert as a bulk material, nanoscale gold can pose harmful side effects to living organisms. In particular, cationic Au nanoparticles (AuNP+) of 2 nm diameter or less permeate readily through plasma membranes and induce cell death. We report atomistic simulations of cationic...... Au nanoparticles interacting with realistic membranes and explicit solvent using a model system that comprises two cellular compartments, extracellular and cytosolic, divided by two asymmetric lipid bilayers. The membrane-AuNP+ binding and membrane reorganization processes are discovered...... to be governed by cooperative effects where AuNP+, counterions, water, and the two membrane leaflets all contribute. On the extracellular side, we find that the nanoparticle has to cross a free energy barrier of about 5 k(B)T prior forming a stable contact with the membrane. This results in a rearrangement...

  11. [Indices of total lipids in erythrocyte membranes and blood plasma in patients with gingivitis in the presence of concomitant diseases and after antioxidant therapy].

    Science.gov (United States)

    Zhirova, V G

    2001-01-01

    Enrolled in this study were those children of both sexes (n=82) from ecologically unfavourable zones who had been admitted to the Yevpatoriya sanatorium "Druzhba" (Frendship) for the rehabilitative period, their age ranging between 11 to 14 years. Our objective in this examination was to study total lipids in red cell membranes and blood plasma in using the antioxidant erbisol to treat inflammatory disease of the oral mucosa in the presence of concomitant illnesses. The performed analysis of the data obtained permitted reaching the conclusion that treatment of parodontal disorders having developed in the presence of concomitant troubles using in a complex therapy erbisol class antioxidants results in a decrease of activation of lipid peroxidation processes in red cell membranes and blood plasma, which fact has found confirmation in an apparent therapeutic effect.

  12. Plasma membrane ATPases

    DEFF Research Database (Denmark)

    Palmgren, Michael Broberg; Bækgaard, Lone; Lopez Marques, Rosa Laura

    2011-01-01

    The plasma membrane separates the cellular contents from the surrounding environment. Nutrients must enter through the plasma membrane in order to reach the cell interior, and toxic metabolites and several ions leave the cell by traveling across the same barrier. Biological pumps in the plasma me...

  13. Lipids and membrane lateral organization

    Directory of Open Access Journals (Sweden)

    Sandro eSonnino

    2010-11-01

    Full Text Available Shortly after the elucidation of the very basic structure and properties of cellular membranes, it became evident that cellular membranes are highly organized structures with multiple and multi-dimensional levels of order. Very early observations suggested that the lipid components of biological membranes might be active players in the creations of these levels of order. In the late 80’s, several different and diverse experimental pieces of evidence coalesced together giving rise to the lipid raft hypothesis. Lipid rafts became enormously (and, in the opinion of these authors, sometimes acritically popular, surprisingly not just within the lipidologist community (who is supposed to be naturally sensitive to the fascination of lipid rafts. Today, a PubMed search using the key word lipid rafts returned a list of 3767 papers, including 690 reviews (as a term of comparison, searching over the same time span for a very hot lipid-related key word, ceramide returned 6187 hits with 799 reviews, and a tremendous number of different cellular functions have been described as lipid raft-dependent. However, a clear consensus definition of lipid raft has been proposed only in recent times, and the basic properties, the ruling forces, and even the existence of lipid rafts in living cells have been recently matter of intense debate. The scenario that is gradually emerging from the controversies elicited by the lipid raft hypothesis emphasize multiple roles for membrane lipids in determining membrane order, that encompasses their tendency to phase separation but are clearly not limited to this. In this review, we would like to re-focus the attention of the readers on the importance of lipids in organizing the fine structure of cellular membranes.

  14. Dynamics of multicomponent lipid membranes

    Science.gov (United States)

    Camley, Brian Andrew

    We present theoretical and computational descriptions of the dynamics of multicomponent lipid bilayer membranes. These systems are both model systems for "lipid rafts" in cell membranes and interesting physical examples of quasi-two-dimensional fluids. Our chief tool is a continuum simulation that uses a phase field to track the composition of the membrane, and solves the hydrodynamic equations exactly using the appropriate Green's function (Oseen tensor) for the membrane. We apply this simulation to describe the diffusion of domains in phase-separated membranes, the dynamics of domain flickering, and the process of phase separation in lipid membranes. We then derive an analytical theory to describe domain flickering that is consistent with our simulation results, and use this to analyze experimental measurements of membrane domains. Through this method, we measure the membrane viscosity solely from fluorescence microscopy measurements. We study phase separation in quasi-two-dimensional membranes in depth with both simulations and scaling theory, and classify the different scaling regimes and morphologies, which differ from pure two-dimensional fluids. Our results may explain previous inconsistent measurements of the dynamical scaling exponent for phase separation in membranes. We also extend our theory beyond the simplest model, including the possibility that the membrane will be viscoelastic, as well as considering the inertia of the membrane and the fluid surrounding the membrane.

  15. Insulin sensitivity is independent of lipid binding protein trafficking at the plasma membrane in human skeletal muscle

    DEFF Research Database (Denmark)

    Jordy, Andreas Børsting; Serup, Annette Karen; Karstoft, Kristian

    2014-01-01

    The aim of the present study was to investigate lipid-induced regulation of lipid binding proteins in human skeletal muscle and the impact hereof on insulin sensitivity. Eleven healthy male subjects underwent a 3-day hyper-caloric and high-fat diet regime. Muscle biopsies were taken before......-regulated by increased fatty acid availability. This suggests a time dependency in the up-regulation of FAT/CD36 and FABPpm protein during high availability of plasma fatty acids. Furthermore, we did not detect FATP1 and FATP4 protein in giant sarcolemmal vesicles obtained from human skeletal muscle. In conclusion......, this study shows that a short-term lipid-load increases mRNA content of key lipid handling proteins in human muscle. However, decreased insulin sensitivity after high-fat diet is not accompanied with relocation of FAT/CD36 or FABPpm protein to the sarcolemma. Finally, FATP1 and FATP4 protein could...

  16. Electronic polymers in lipid membranes.

    Science.gov (United States)

    Johansson, Patrik K; Jullesson, David; Elfwing, Anders; Liin, Sara I; Musumeci, Chiara; Zeglio, Erica; Elinder, Fredrik; Solin, Niclas; Inganäs, Olle

    2015-06-10

    Electrical interfaces between biological cells and man-made electrical devices exist in many forms, but it remains a challenge to bridge the different mechanical and chemical environments of electronic conductors (metals, semiconductors) and biosystems. Here we demonstrate soft electrical interfaces, by integrating the metallic polymer PEDOT-S into lipid membranes. By preparing complexes between alkyl-ammonium salts and PEDOT-S we were able to integrate PEDOT-S into both liposomes and in lipid bilayers on solid surfaces. This is a step towards efficient electronic conduction within lipid membranes. We also demonstrate that the PEDOT-S@alkyl-ammonium:lipid hybrid structures created in this work affect ion channels in the membrane of Xenopus oocytes, which shows the possibility to access and control cell membrane structures with conductive polyelectrolytes.

  17. Gel Domains in the Plasma Membrane of Saccharomyces cerevisiae: HIGHLY ORDERED, ERGOSTEROL-FREE, AND SPHINGOLIPID-ENRICHED LIPID RAFTS*

    OpenAIRE

    Aresta-Branco, Francisco; Cordeiro, André M.; Marinho, H. Susana; Cyrne, Luísa; Antunes, Fernando; de Almeida, Rodrigo F. M.

    2010-01-01

    The plasma membrane of Saccharomyces cerevisiae was studied using the probes trans-parinaric acid and diphenylhexatriene. Diphenylhexatriene anisotropy is a good reporter of global membrane order. The fluorescence lifetimes of trans-parinaric acid are particularly sensitive to the presence and nature of ordered domains, but thus far they have not been measured in yeast cells. A long lifetime typical of the gel phase (>30 ns) was found in wild-type (WT) cells from two different genetic backgro...

  18. Exploration of lipid metabolism in relation with plasma membrane properties of Duchenne muscular dystrophy cells: influence of L-carnitine.

    Directory of Open Access Journals (Sweden)

    Françoise Le Borgne

    Full Text Available Duchenne muscular dystrophy (DMD arises as a consequence of mutations in the dystrophin gene. Dystrophin is a membrane-spanning protein that connects the cytoskeleton and the basal lamina. The most distinctive features of DMD are a progressive muscular dystrophy, a myofiber degeneration with fibrosis and metabolic alterations such as fatty infiltration, however, little is known on lipid metabolism changes arising in Duchenne patient cells. Our goal was to identify metabolic changes occurring in Duchenne patient cells especially in terms of L-carnitine homeostasis, fatty acid metabolism both at the mitochondrial and peroxisomal level and the consequences on the membrane structure and function. In this paper, we compared the structural and functional characteristics of DMD patient and control cells. Using radiolabeled L-carnitine, we found, in patient muscle cells, a marked decrease in the uptake and the intracellular level of L-carnitine. Associated with this change, a decrease in the mitochondrial metabolism can be seen from the analysis of mRNA encoding for mitochondrial proteins. Probably, associated with these changes in fatty acid metabolism, alterations in the lipid composition of the cells were identified: with an increase in poly unsaturated fatty acids and a decrease in medium chain fatty acids, mono unsaturated fatty acids and in cholesterol contents. Functionally, the membrane of cells lacking dystrophin appeared to be less fluid, as determined at 37°C by fluorescence anisotropy. These changes may, at least in part, be responsible for changes in the phospholipids and cholesterol profile in cell membranes and ultimately may reduce the fluidity of the membrane. A supplementation with L-carnitine partly restored the fatty acid profile by increasing saturated fatty acid content and decreasing the amounts of MUFA, PUFA, VLCFA. L-carnitine supplementation also restored muscle membrane fluidity. This suggests that regulating lipid metabolism

  19. Lipid membranes on nanostructured silicon.

    Energy Technology Data Exchange (ETDEWEB)

    Slade, Andrea Lynn; Lopez, Gabriel P. (University of New Mexico, Albuquerque, NM); Ista, Linnea K. (University of New Mexico, Albuquerque, NM); O' Brien, Michael J. (University of New Mexico, Albuquerque, NM); Sasaki, Darryl Yoshio; Bisong, Paul (University of New Mexico, Albuquerque, NM); Zeineldin, Reema R. (University of New Mexico, Albuquerque, NM); Last, Julie A.; Brueck, Stephen R. J. (University of New Mexico, Albuquerque, NM)

    2004-12-01

    A unique composite nanoscale architecture that combines the self-organization and molecular dynamics of lipid membranes with a corrugated nanotextured silicon wafer was prepared and characterized with fluorescence microscopy and scanning probe microscopy. The goal of this project was to understand how such structures can be assembled for supported membrane research and how the interfacial interactions between the solid substrate and the soft, self-assembled material create unique physical and mechanical behavior through the confinement of phases in the membrane. The nanometer scale structure of the silicon wafer was produced through interference lithography followed by anisotropic wet etching. For the present study, a line pattern with 100 nm line widths, 200 nm depth and a pitch of 360 nm pitch was fabricated. Lipid membranes were successfully adsorbed on the structured silicon surface via membrane fusion techniques. The surface topology of the bilayer-Si structure was imaged using in situ tapping mode atomic force microscopy (AFM). The membrane was observed to drape over the silicon structure producing an undulated topology with amplitude of 40 nm that matched the 360 nm pitch of the silicon structure. Fluorescence recovery after photobleaching (FRAP) experiments found that on the microscale those same structures exhibit anisotropic lipid mobility that was coincident with the silicon substructure. The results showed that while the lipid membrane maintains much of its self-assembled structure in the composite architecture, the silicon substructure indeed influences the dynamics of the molecular motion within the membrane.

  20. The physics of stratum corneum lipid membranes.

    Science.gov (United States)

    Das, Chinmay; Olmsted, Peter D

    2016-07-28

    The stratum corneum (SC), the outermost layer of skin, comprises rigid corneocytes (keratin-filled dead cells) in a specialized lipid matrix. The continuous lipid matrix provides the main barrier against uncontrolled water loss and invasion of external pathogens. Unlike all other biological lipid membranes (such as intracellular organelles and plasma membranes), molecules in the SC lipid matrix show small hydrophilic groups and large variability in the length of the alkyl tails and in the numbers and positions of groups that are capable of forming hydrogen bonds. Molecular simulations provide a route for systematically probing the effects of each of these differences separately. In this article, we present the results from atomistic molecular dynamics of selected lipid bilayers and multi-layers to probe the effect of these polydispersities. We address the nature of the tail packing in the gel-like phase, the hydrogen bond network among head groups, the bending moduli expected for leaflets comprising SC lipids and the conformation of very long ceramide lipids in multi-bilayer lipid assemblies.This article is part of the themed issue 'Soft interfacial materials: from fundamentals to formulation'.

  1. Polyunsaturated Lipids Regulate Membrane Domain Stability by Tuning Membrane Order.

    Science.gov (United States)

    Levental, Kandice R; Lorent, Joseph H; Lin, Xubo; Skinkle, Allison D; Surma, Michal A; Stockenbojer, Emily A; Gorfe, Alemayehu A; Levental, Ilya

    2016-04-26

    The plasma membrane (PM) serves as the functional interface between a cell and its environment, hosting extracellular signal transduction and nutrient transport among a variety of other processes. To support this extensive functionality, PMs are organized into lateral domains, including ordered, lipid-driven assemblies termed lipid rafts. Although the general requirements for ordered domain formation are well established, how these domains are regulated by cell-endogenous mechanisms or exogenous perturbations has not been widely addressed. In this context, an intriguing possibility is that dietary fats can incorporate into membrane lipids to regulate the properties and physiology of raft domains. Here, we investigate the effects of polyunsaturated fats on the organization of membrane domains across a spectrum of membrane models, including computer simulations, synthetic lipid membranes, and intact PMs isolated from mammalian cells. We observe that the ω-3 polyunsaturated fatty acid docosahexaenoic acid is robustly incorporated into membrane lipids, and this incorporation leads to significant remodeling of the PM lipidome. Across model systems, docosahexaenoic acid-containing lipids enhance the stability of ordered raft domains by increasing the order difference between them and coexisting nonraft domains. The relationship between interdomain order disparity and the stability of phase separation holds for a spectrum of different perturbations, including manipulation of cholesterol levels and high concentrations of exogenous amphiphiles, suggesting it as a general feature of the organization of biological membranes. These results demonstrate that polyunsaturated fats affect the composition and organization of biological membranes, suggesting a potential mechanism for the extensive effects of dietary fat on health and disease.

  2. Lipids in the Sperm Plasma Membrane and Their Role in Fertilization%精子胞膜脂质及其在受精中的作用

    Institute of Scientific and Technical Information of China (English)

    牛冬梅; 汪俊军

    2009-01-01

    Sexual reproduction is marked by the fusion of the sperm ceil with the oocyte during fertilization to produce the diploid zy-gote,in which the lipids in the sperm plasma membrane play an important role.Due to the loss of most cell organelles and DNA tran-seription,spermatozoa lack protein expression and vesicular transport.However,the lipids of the sperm plasma membrane undergo complicated dynamic changes,which may facilitate the capacitation,binding with zona pellucida,acrosome reaction and fusion of the sperm ceil with the oocyte.This paper summarizes the progress in the studies of the lipids in the sperm plasma membrane,their eompo-sition,structure,peroxidation,metabolism and role in fertilization.%通过受精来完成精子与卵子融合并产生二倍体合子(受精卵)是有性生殖的标志,在这一过程中精子胞膜脂质发挥了重要作用.精子在成熟过程中,由于细胞器的丢失和DNA转录的终止,精子最终停止合成膜脂质、蛋白质,并且囊泡转运也终止.然而精子胞膜脂质成分却仍发生着复杂变化,并籍此在精子获能、与卵子透明带结合、顶体反应以及精子与卵子膜融合等过程中均起着重要作用.本文就精子膜脂质的组成、结构特点、过氧化作用、代谢及其在受精中的作用进行综述.

  3. Effect of plasma etching on photoluminescence of SnO(x)/Sn nanoparticles deposited on DOPC lipid membrane.

    Science.gov (United States)

    An, Hyeun Hwan; Lee, Seung Jae; Baek, Seung Ha; Han, Won Bae; Kim, Young Ho; Yoon, Chong Seung; Suh, Sang Hee

    2012-02-15

    The photoluminescence characteristic of the SnO(x)/Sn nanoparticles deposited on a solid supported liquid-crystalline phospholipid (1,2-dioleoyl-sn-glycero-3-phosphocholine) membrane was probed after plasma etching the nanoparticle monolayer. It was shown that the plasma etching of the nanoparticle surface greatly altered the particle morphology and enhanced the PL effect, especially when the particle size was below 10 nm in spite of strong presence of surrounding carbon. The enhancement mainly stemmed from the growth of a new PL peak due to the additional defect states produced on the nanoparticle surface by the plasma etching. It was also shown that hydrating the SnO(x)/Sn nanoparticles similarly improved the PL response of the nanoparticles as the hydration produced an additional oxygen-rich oxide layer on the particle surface. Copyright © 2011 Elsevier Inc. All rights reserved.

  4. Localization of lipid raft proteins to the plasma membrane is a major function of the phospholipid transfer protein Sec14.

    Science.gov (United States)

    Curwin, Amy J; Leblanc, Marissa A; Fairn, Gregory D; McMaster, Christopher R

    2013-01-01

    The Sec14 protein domain is a conserved tertiary structure that binds hydrophobic ligands. The Sec14 protein from Saccharomyces cerevisiae is essential with studies of S. cerevisiae Sec14 cellular function facilitated by a sole temperature sensitive allele, sec14(ts). The sec14(ts) allele encodes a protein with a point mutation resulting in a single amino acid change, Sec14(G266D). In this study results from a genome-wide genetic screen, and pharmacological data, provide evidence that the Sec14(G266D) protein is present at a reduced level compared to wild type Sec14 due to its being targeted to the proteosome. Increased expression of the sec14(ts) allele ameliorated growth arrest, but did not restore the defects in membrane accumulation or vesicular transport known to be defective in sec14(ts) cells. We determined that trafficking and localization of two well characterized lipid raft resident proteins, Pma1 and Fus-Mid-GFP, were aberrant in sec14(ts) cells. Localization of both lipid raft proteins was restored upon increased expression of the sec14(ts) allele. We suggest that a major function provided by Sec14 is trafficking and localization of lipid raft proteins.

  5. Cellular membrane collapse by atmospheric-pressure plasma jet

    Science.gov (United States)

    Kim, Kangil; Jun Ahn, Hak; Lee, Jae-Hyeok; Kim, Jae-Ho; Sik Yang, Sang; Lee, Jong-Soo

    2014-01-01

    Cellular membrane dysfunction caused by air plasma in cancer cells has been studied to exploit atmospheric-pressure plasma jets for cancer therapy. Here, we report that plasma jet treatment of cervical cancer HeLa cells increased electrical conductivity across the cellular lipid membrane and caused simultaneous lipid oxidation and cellular membrane collapse. We made this finding by employing a self-manufactured microelectrode chip. Furthermore, increased roughness of the cellular lipid membrane and sequential collapse of the membrane were observed by atomic force microscopy following plasma jet treatment. These results suggest that the cellular membrane catastrophe occurs via coincident altered electrical conductivity, lipid oxidation, and membrane roughening caused by an atmospheric-pressure plasma jet, possibly resulting in cellular vulnerability to reactive species generated from the plasma as well as cytotoxicity to cancer cells.

  6. Cellular membrane collapse by atmospheric-pressure plasma jet

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Kangil; Sik Yang, Sang, E-mail: jsjlee@ajou.ac.kr, E-mail: ssyang@ajou.ac.kr [Department of Electrical and Computer Engineering, Ajou University, Suwon 443-749 (Korea, Republic of); Jun Ahn, Hak; Lee, Jong-Soo, E-mail: jsjlee@ajou.ac.kr, E-mail: ssyang@ajou.ac.kr [Department of Biological Sciences, Ajou University, Suwon 443-749 (Korea, Republic of); Lee, Jae-Hyeok; Kim, Jae-Ho [Department of Molecular Science and Technology, Ajou University, Suwon 443-749 (Korea, Republic of)

    2014-01-06

    Cellular membrane dysfunction caused by air plasma in cancer cells has been studied to exploit atmospheric-pressure plasma jets for cancer therapy. Here, we report that plasma jet treatment of cervical cancer HeLa cells increased electrical conductivity across the cellular lipid membrane and caused simultaneous lipid oxidation and cellular membrane collapse. We made this finding by employing a self-manufactured microelectrode chip. Furthermore, increased roughness of the cellular lipid membrane and sequential collapse of the membrane were observed by atomic force microscopy following plasma jet treatment. These results suggest that the cellular membrane catastrophe occurs via coincident altered electrical conductivity, lipid oxidation, and membrane roughening caused by an atmospheric-pressure plasma jet, possibly resulting in cellular vulnerability to reactive species generated from the plasma as well as cytotoxicity to cancer cells.

  7. Lipids: architects and regulators of membrane dynamics and trafficking.

    Science.gov (United States)

    Moreau, Patrick

    2007-05-01

    We have recently shown that an inhibition of sterol synthesis by fenpropimorph leads to an accumulation of sterol precursors, hydroxypalmitic acid-containing glucosylceramides and detergent resistant membranes in the Golgi bodies instead of the plasma membrane, suggesting that the individual molecules or the microdomains were blocked in the Golgi. These results and others from several eukaryotic models link lipid metabolism with membrane morphodynamics that are involved in membrane trafficking. Focus has been expanded to other lipid families, and numerous evidences are given showing lipids and lipid-modifying enzymes as key regulators of membrane homeostasis which can strongly regulate membrane morphodynamics and therefore trafficking. Beside protein-based machineries, lipid-based machineries are also shown as crucial regulatory forces involved in protein transport and sorting.

  8. Giant plasma membrane vesicles: models for understanding membrane organization.

    Science.gov (United States)

    Levental, Kandice R; Levental, Ilya

    2015-01-01

    The organization of eukaryotic membranes into functional domains continues to fascinate and puzzle cell biologists and biophysicists. The lipid raft hypothesis proposes that collective lipid interactions compartmentalize the membrane into coexisting liquid domains that are central to membrane physiology. This hypothesis has proven controversial because such structures cannot be directly visualized in live cells by light microscopy. The recent observations of liquid-liquid phase separation in biological membranes are an important validation of the raft hypothesis and enable application of the experimental toolbox of membrane physics to a biologically complex phase-separated membrane. This review addresses the role of giant plasma membrane vesicles (GPMVs) in refining the raft hypothesis and expands on the application of GPMVs as an experimental model to answer some of key outstanding problems in membrane biology. Copyright © 2015 Elsevier Inc. All rights reserved.

  9. Confined diffusion of transmembrane proteins and lipids induced by the same actin meshwork lining the plasma membrane.

    Science.gov (United States)

    Fujiwara, Takahiro K; Iwasawa, Kokoro; Kalay, Ziya; Tsunoyama, Taka A; Watanabe, Yusuke; Umemura, Yasuhiro M; Murakoshi, Hideji; Suzuki, Kenichi G N; Nemoto, Yuri L; Morone, Nobuhiro; Kusumi, Akihiro

    2016-04-01

    The mechanisms by which the diffusion rate in the plasma membrane (PM) is regulated remain unresolved, despite their importance in spatially regulating the reaction rates in the PM. Proposed models include entrapment in nanoscale noncontiguous domains found in PtK2 cells, slow diffusion due to crowding, and actin-induced compartmentalization. Here, by applying single-particle tracking at high time resolutions, mainly to the PtK2-cell PM, we found confined diffusion plus hop movements (termed "hop diffusion") for both a nonraft phospholipid and a transmembrane protein, transferrin receptor, and equal compartment sizes for these two molecules in all five of the cell lines used here (actual sizes were cell dependent), even after treatment with actin-modulating drugs. The cross-section size and the cytoplasmic domain size both affected the hop frequency. Electron tomography identified the actin-based membrane skeleton (MSK) located within 8.8 nm from the PM cytoplasmic surface of PtK2 cells and demonstrated that the MSK mesh size was the same as the compartment size for PM molecular diffusion. The extracellular matrix and extracellular domains of membrane proteins were not involved in hop diffusion. These results support a model of anchored TM-protein pickets lining actin-based MSK as a major mechanism for regulating diffusion.

  10. Organization and Dynamics of Receptor Proteins in a Plasma Membrane.

    Science.gov (United States)

    Koldsø, Heidi; Sansom, Mark S P

    2015-11-25

    The interactions of membrane proteins are influenced by their lipid environment, with key lipid species able to regulate membrane protein function. Advances in high-resolution microscopy can reveal the organization and dynamics of proteins and lipids within living cells at resolutions membranes of in vivo-like complexity. We explore the dynamics of proteins and lipids in crowded and complex plasma membrane models, thereby closing the gap in length and complexity between computations and experiments. Our simulations provide insights into the mutual interplay between lipids and proteins in determining mesoscale (20-100 nm) fluctuations of the bilayer, and in enabling oligomerization and clustering of membrane proteins.

  11. 95 Alteration of Plasma Lipid Profile and Atherogenic Indices of ...

    African Journals Online (AJOL)

    JTEkanem

    2009-07-15

    Jul 15, 2009 ... procumbens on the packed cell volume (PCV), daily weight gain, plasma lipid profiles and ... conditions, whether primary or secondary to diabetes mellitus, hypertension and obesity .... (1999) Abnormalities of membrane.

  12. Pore dynamics in lipid membranes

    Science.gov (United States)

    Gozen, I.; Dommersnes, P.

    2014-09-01

    Transient circular pores can open in plasma membrane of cells due to mechanical stress, and failure to repair such pores lead to cell death. Similar pores in the form of defects also exist among smectic membranes, such as in myelin sheaths or mitochondrial membranes. The formation and growth of membrane defects are associated with diseases, for example multiple sclerosis. A deeper understanding of membrane pore dynamics can provide a more refined picture of membrane integrity-related disease development, and possibly also treatment options and strategies. Pore dynamics is also of great importance regarding healthcare applications such as drug delivery, gene or as recently been implied, cancer therapy. The dynamics of pores significantly differ in stacks which are confined in 2D compared to those in cells or vesicles. In this short review, we will summarize the dynamics of different types of pores that can be observed in biological membranes, which include circular transient, fusion and hemi-fusion pores. We will dedicate a section to floral and fractal pores which were discovered a few years ago and have highly peculiar characteristics. Finally, we will discuss the repair mechanisms of large area pores in conjunction with the current cell membrane repair hypotheses.

  13. Role of Membrane Lipid Fatty Acids in Sperm Cryopreservation

    Directory of Open Access Journals (Sweden)

    Rajes Mandal

    2014-01-01

    Full Text Available Lipid is an important constituent of cell membrane. Membrane lipid composition of spermatozoa has been correlated to different function. Many researchers have related membrane lipid with survival success after cryopreservation or cold shock. Sperm maturation and acrosome reactions are natural phenomenon, but cryopreservation or cold shock is not. Therefore, sperm cells are not programmed for such change and undergo stress. So the change in membrane lipid composition due to cold shock or cryopreservation may be looked upon as response of spermatozoa to a certain stressed condition. A significant body of research worked on the relationship between membrane lipid and fatty acid composition and ability of cell to tolerate adverse change in temperature. However, as the approach of different research groups was different, it is very difficult to compare the changes. Studies have been done with different species, ejaculated/seminal or epididymal sperm. Lipid analyses have been done with whole cell membrane isolated by different methods. Fatty acids estimated were from whole cell, plasma membrane, head membrane, or phospholipids. The cryopreservation condition, media composition, and diluents/cryoprotectants were also different. At this onset a comprehensive review is needed to cover changes of sperm membrane lipid composition of different species under different cryopreservation conditions.

  14. Effects of salinity on activities of H+-ATPase, H+-PPase and membrane lipid composition in plasma membrane and tonoplast vesicles isolated from soybean (Glycine max L.) seedlings

    Institute of Scientific and Technical Information of China (English)

    YU Bing-jun; LAM Hon-ming; SHAO Gui-hua; LIU You-ling

    2005-01-01

    The effects of NaCl stress on the H+ -ATPase, H+ -PPase activity and lipid composition of plasma membrane(PM) and tonoplast(TP) vesicles isolated from roots and leaves of two soybean cultivars( Glycine max L. ) differing in salt tolerance(Wenfeng7,salt-tolerant; Union, salt-sensitive) were investigated. When Wenfeng7 was treated with 0.3% (W/V) NaCl for 3 d, the H+ -ATPase activities in PM and TP from roots and leaves exhibited a reduction and an enhancement, respectively. The H+ -PPase activity in TP from roots also increased. Similar effects were not observed in roots of Union. In addition, the increases of phospholipid content and ratios ofphospholipid to galactolipid in PM and TP from roots and leaves of Wenfeng7 may also change membrane permeability and hence affect salt tolerance.

  15. Lipid rafts and detergent-resistant membranes in epithelial keratinocytes.

    Science.gov (United States)

    McGuinn, Kathleen P; Mahoney, Mỹ G

    2014-01-01

    Our understanding of the plasma membrane has markedly increased since Singer and Nicolson proposed the fluid mosaic model in 1972. While their revolutionary theory of the lipid bilayer remains largely valid, it is now known that lipids and proteins are not randomly dispersed throughout the plasma membrane but instead may be organized within membrane microdomains, commonly referred to as lipid rafts. Lipid rafts are highly dynamic, detergent resistant, and enriched with both cholesterol and glycosphingolipids. The two main types are flotillin-rich planar lipid rafts and caveolin-rich caveolae. It is proposed that flotillin and caveolin proteins regulate cell communication by compartmentalizing and interacting with signal transduction proteins within their respective lipid microdomains. Consequently, membrane rafts play an important role in vital cellular functions including migration, invasion, and signaling; thus, alterations in their microenvironment can initiate signaling pathways that affect cellular function and behavior. Therefore, the identification of lipid rafts and their associated proteins is integral to the study of transmembrane signaling. Here, we review the current standard protocols and biochemical approaches used to isolate and define raft proteins from epithelial cells and tissues. Furthermore, in Section 3 of this chapter, detailed protocols are offered for isolating lipid rafts by subjection to detergent and sucrose density centrifugation, as well as an approach for selectively isolating caveolae. Methods to manipulate rafts with treatments such as methyl-β-cyclodextrin and flotillin III are also described.

  16. Glycans pattern the phase behaviour of lipid membranes

    Science.gov (United States)

    Subramaniam, Anand Bala; Guidotti, Guido; Manoharan, Vinothan N.; Stone, Howard A.

    2013-02-01

    Hydrated networks of glycans (polysaccharides)—in the form of cell walls, periplasms or gel-like matrices—are ubiquitously present adjacent to cellular plasma membranes. Yet, despite their abundance, the function of glycans in the extracellular milieu is largely unknown. Here we show that the spatial configuration of glycans controls the phase behaviour of multiphase model lipid membranes: inhomogeneous glycan networks stabilize large lipid domains at the characteristic length scale of the network, whereas homogeneous networks suppress macroscopic lipid phase separation. We also find that glycan-patterned phase separation is thermally reversible—thus indicating that the effect is thermodynamic rather than kinetic—and that phase patterning probably results from a preferential interaction of glycans with ordered lipid phases. These findings have implications for membrane-mediated transport processes, potentially rationalize long-standing observations that differentiate the behaviour of native and model membranes and may indicate an intimate coupling between cellular lipidomes and glycomes.

  17. Electrodiffusion of lipids on membrane surfaces

    Science.gov (United States)

    Zhou, Y. C.

    2012-05-01

    Lateral translocation of lipids and proteins is a universal process on membrane surfaces. Local aggregation or organization of lipids and proteins can be induced when the random lateral motion is mediated by the electrostatic interactions and membrane curvature. Although the lateral diffusion rates of lipids on membranes of various compositions are measured and the electrostatic free energies of predetermined protein-membrane-lipid systems can be computed, the process of the aggregation and the evolution to the electrostatically favorable states remain largely undetermined. Here we propose an electrodiffusion model, based on the variational principle of the free energy functional, for the self-consistent lateral drift-diffusion of multiple species of charged lipids on membrane surfaces. Finite sizes of lipids are modeled to enforce the geometrical constraint of the lipid concentration on membrane surfaces. A surface finite element method is developed to appropriate the Laplace-Beltrami operators in the partial differential equations of the model. Our model properly describes the saturation of lipids on membrane surfaces, and correctly predicts that the MARCKS peptide can consistently sequester three multivalent phosphatidylinositol 4,5-bisphosphate lipids through its basic amino acid residues, regardless of a wide range of the percentage of monovalent phosphatidylserine in the membrane.

  18. [Germ cell membrane lipids in spermatogenesis].

    Science.gov (United States)

    Wang, Ting; Shi, Xiao; Quan, Song

    2016-05-01

    Spermatogenesis is a complex developmental process in which a diploid progenitor germ cell transforms into highly specialized spermatozoa. During spermatogenesis, membrane remodeling takes place, and cell membrane permeability and liquidity undergo phase-specific changes, which are all associated with the alteration of membrane lipids. Lipids are important components of the germ cell membrane, whose volume and ratio fluctuate in different phases of spermatogenesis. Abnormal lipid metabolism can cause spermatogenic dysfunction and consequently male infertility. Germ cell membrane lipids are mainly composed of cholesterol, phospholipids and glycolipids, which play critical roles in cell adhesion and signal transduction during spermatogenesis. An insight into the correlation of membrane lipids with spermatogenesis helps us to better understand the mechanisms of spermatogenesis and provide new approaches to the diagnosis and treatment of male infertility.

  19. How Lipid Membranes Affect Pore Forming Toxin Activity.

    Science.gov (United States)

    Rojko, Nejc; Anderluh, Gregor

    2015-12-15

    Pore forming toxins (PFTs) evolved to permeate the plasma membrane of target cells. This is achieved in a multistep mechanism that usually involves binding of soluble protein monomer to the lipid membrane, oligomerization at the plane of the membrane, and insertion of part of the polypeptide chain across the lipid membrane to form a conductive channel. Introduced pores allow uncontrolled transport of solutes across the membrane, inflicting damage to the target cell. PFTs are usually studied from the perspective of structure-function relationships, often neglecting the important role of the bulk membrane properties on the PFT mechanism of action. In this Account, we discuss how membrane lateral heterogeneity, thickness, and fluidity influence the pore forming process of PFTs. In general, lipid molecules are more accessible for binding in fluid membranes due to steric reasons. When PFT specifically binds ordered domains, it usually recognizes a specific lipid distribution pattern, like sphingomyelin (SM) clusters or SM/cholesterol complexes, and not individual lipid species. Lipid domains were also suggested to act as an additional concentration platform facilitating PFT oligomerization, but this is yet to be shown. The last stage in PFT action is the insertion of the transmembrane segment across the membranes to build the transmembrane pore walls. Conformational changes are a spontaneous process, and sufficient free energy has to be available for efficient membrane penetration. Therefore, fluid bilayers are permeabilized more readily in comparison to highly ordered and thicker liquid ordered lipid phase (Lo). Energetically more costly insertion into the Lo phase can be driven by the hydrophobic mismatch between the thinner liquid disordered phase (Ld) and large protein complexes, which are unable to tilt like single transmembrane segments. In the case of proteolipid pores, membrane properties can directly modulate pore size, stability, and even selectivity. Finally

  20. DMSO Induces Dehydration near Lipid Membrane Surfaces

    Science.gov (United States)

    Cheng, Chi-Yuan; Song, Jinsuk; Pas, Jolien; Meijer, Lenny H.H.; Han, Songi

    2015-01-01

    Dimethyl sulfoxide (DMSO) has been broadly used in biology as a cosolvent, a cryoprotectant, and an enhancer of membrane permeability, leading to the general assumption that DMSO-induced structural changes in cell membranes and their hydration water play important functional roles. Although the effects of DMSO on the membrane structure and the headgroup dehydration have been extensively studied, the mechanism by which DMSO invokes its effect on lipid membranes and the direct role of water in this process are unresolved. By directly probing the translational water diffusivity near unconfined lipid vesicle surfaces, the lipid headgroup mobility, and the repeat distances in multilamellar vesicles, we found that DMSO exclusively weakens the surface water network near the lipid membrane at a bulk DMSO mole fraction (XDMSO) of DMSO was found to effectively destabilize the hydration water structure at the lipid membrane surface at XDMSO 0.1, DMSO enters the lipid interface and restricts the lipid headgroup motion. We postulate that DMSO acts as an efficient cryoprotectant even at low concentrations by exclusively disrupting the water network near the lipid membrane surface, weakening the cohesion between water and adhesion of water to the lipid headgroups, and so mitigating the stress induced by the volume change of water during freeze-thaw. PMID:26200868

  1. Transport and sorting of membrane lipids

    NARCIS (Netherlands)

    van Meer, G.|info:eu-repo/dai/nl/068570368

    1993-01-01

    The lipid composition of cellular membranes may seem unnecessarily complex. However, the lipid composition of each membrane is carefully regulated by local metabolism and specificity in transport, marking the functional significance for the cell. Recent research has revealed unexpected discoveries c

  2. Differential Effect of Plant Lipids on Membrane Organization

    Science.gov (United States)

    Grosjean, Kevin; Mongrand, Sébastien; Beney, Laurent; Simon-Plas, Françoise; Gerbeau-Pissot, Patricia

    2015-01-01

    The high diversity of the plant lipid mixture raises the question of their respective involvement in the definition of membrane organization. This is particularly the case for plant plasma membrane, which is enriched in specific lipids, such as free and conjugated forms of phytosterols and typical phytosphingolipids, such as glycosylinositolphosphoceramides. This question was here addressed extensively by characterizing the order level of membrane from vesicles prepared using various plant lipid mixtures and labeled with an environment-sensitive probe. Fluorescence spectroscopy experiments showed that among major phytosterols, campesterol exhibits a stronger ability than β-sitosterol and stigmasterol to order model membranes. Multispectral confocal microscopy, allowing spatial analysis of membrane organization, demonstrated accordingly the strong ability of campesterol to promote ordered domain formation and to organize their spatial distribution at the membrane surface. Conjugated sterol forms, alone and in synergy with free sterols, exhibit a striking ability to order membrane. Plant sphingolipids, particularly glycosylinositolphosphoceramides, enhanced the sterol-induced ordering effect, emphasizing the formation and increasing the size of sterol-dependent ordered domains. Altogether, our results support a differential involvement of free and conjugated phytosterols in the formation of ordered domains and suggest that the diversity of plant lipids, allowing various local combinations of lipid species, could be a major contributor to membrane organization in particular through the formation of sphingolipid-sterol interacting domains. PMID:25575593

  3. Organized living: formation mechanisms and functions of plasma membrane domains in yeast.

    Science.gov (United States)

    Ziółkowska, Natasza E; Christiano, Romain; Walther, Tobias C

    2012-03-01

    Plasma membrane proteins and lipids organize into lateral domains of specific composition. Domain formation is achieved by a combination of lipid-lipid and lipid-protein interactions, membrane-binding protein scaffolds and protein fences. The resulting domains function in membrane protein turnover and homeostasis, as well as in cell signaling. We review the mechanisms generating plasma membrane domains and the functional consequences of this organization, focusing on recent findings from research on the yeast model system.

  4. Effect of fish oil on lateral mobility of prostaglandin F2α (FP) receptors and spatial distribution of lipid microdomains in bovine luteal cell plasma membrane in vitro.

    Science.gov (United States)

    Plewes, M R; Burns, P D; Graham, P E; Hyslop, R M; Barisas, B G

    2017-01-01

    Lipid microdomains are ordered regions on the plasma membrane of cells, rich in cholesterol and sphingolipids, ranging in size from 10 to 200 nm in diameter. These lipid-ordered domains may serve as platforms to facilitate colocalization of intracellular signaling proteins during agonist-induced signal transduction. It is hypothesized that fish oil will disrupt the lipid microdomains, increasing spatial distribution of these lipid-ordered domains and lateral mobility of the prostaglandin (PG) F2α (FP) receptors in bovine luteal cells. The objectives of this study were to examine the effects of fish oil on (1) the spatial distribution of lipid microdomains, (2) lateral mobility of FP receptors, and (3) lateral mobility of FP receptors in the presence of PGF2α on the plasma membrane of bovine luteal cells in vitro. Bovine ovaries were obtained from a local abattoir and corpora lutea were digested using collagenase. In experiment 1, lipid microdomains were labeled using cholera toxin subunit B Alexa Fluor 555. Domains were detected as distinct patches on the plasma membrane of mixed luteal cells. Fish oil treatment decreased fluorescent intensity in a dose-dependent manner (P oil treatment on lateral mobility of FP receptors. Fish oil treatment increased microdiffusion and macrodiffusion coefficients of FP receptors as compared to control cells (P oil-treated cells (P oil treatment. Lateral mobility of receptors was decreased within 5 min following the addition of ligand for control cells (P oil-treated cells (P > 0.10). The data presented provide strong evidence that fish oil causes a disruption in lipid microdomains and affects lateral mobility of FP receptors in the absence and presence of PGF2α.

  5. Single Lipid Molecule Dynamics on Supported Lipid Bilayers with Membrane Curvature

    Directory of Open Access Journals (Sweden)

    Philip P. Cheney

    2017-03-01

    Full Text Available The plasma membrane is a highly compartmentalized, dynamic material and this organization is essential for a wide variety of cellular processes. Nanoscale domains allow proteins to organize for cell signaling, endo- and exocytosis, and other essential processes. Even in the absence of proteins, lipids have the ability to organize into domains as a result of a variety of chemical and physical interactions. One feature of membranes that affects lipid domain formation is membrane curvature. To directly test the role of curvature in lipid sorting, we measured the accumulation of two similar lipids, 1,2-Dihexadecanoyl-sn-glycero-3-phosphoethanolamine (DHPE and hexadecanoic acid (HDA, using a supported lipid bilayer that was assembled over a nanopatterned surface to obtain regions of membrane curvature. Both lipids studied contain 16 carbon, saturated tails and a head group tag for fluorescence microscopy measurements. The accumulation of lipids at curvatures ranging from 28 nm to 55 nm radii was measured and fluorescein labeled DHPE accumulated more than fluorescein labeled HDA at regions of membrane curvature. We then tested whether single biotinylated DHPE molecules sense curvature using single particle tracking methods. Similar to groups of fluorescein labeled DHPE accumulating at curvature, the dynamics of single molecules of biotinylated DHPE was also affected by membrane curvature and highly confined motion was observed.

  6. Pollen viability and membrane lipid composition.

    NARCIS (Netherlands)

    Bilsen, van D.G.J.L.

    1993-01-01

    In this thesis membrane lipid composition is studied in relation to pollen viability during storage. Chapter 1 reviews pollen viability, membranes in the dry state and membrane changes associated with cellular aging. This chapter is followed by a study of age-related changes in phospholipid composit

  7. Designing lipids for selective partitioning into liquid ordered membrane domains.

    Science.gov (United States)

    Momin, Noor; Lee, Stacey; Gadok, Avinash K; Busch, David J; Bachand, George D; Hayden, Carl C; Stachowiak, Jeanne C; Sasaki, Darryl Y

    2015-04-28

    Self-organization of lipid molecules into specific membrane phases is key to the development of hierarchical molecular assemblies that mimic cellular structures. While the packing interaction of the lipid tails should provide the major driving force to direct lipid partitioning to ordered or disordered membrane domains, numerous examples show that the headgroup and spacer play important but undefined roles. We report here the development of several new biotinylated lipids that examine the role of spacer chemistry and structure on membrane phase partitioning. The new lipids were prepared with varying lengths of low molecular weight polyethylene glycol (EGn) spacers to examine how spacer hydrophilicity and length influence their partitioning behavior following binding with FITC-labeled streptavidin in liquid ordered (Lo) and liquid disordered (Ld) phase coexisting membranes. Partitioning coefficients (Kp Lo/Ld) of the biotinylated lipids were determined using fluorescence measurements in studies with giant unilamellar vesicles (GUVs). Compared against DPPE-biotin, DPPE-cap-biotin, and DSPE-PEG2000-biotin lipids, the new dipalmityl-EGn-biotin lipids exhibited markedly enhanced partitioning into liquid ordered domains, achieving Kp of up to 7.3 with a decaethylene glycol spacer (DP-EG10-biotin). We further demonstrated biological relevance of the lipids with selective partitioning to lipid raft-like domains observed in giant plasma membrane vesicles (GPMVs) derived from mammalian cells. Our results found that the spacer group not only plays a pivotal role for designing lipids with phase selectivity but may also influence the structural order of the domain assemblies.

  8. The interaction between purple membrane and membrane lipid

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    Bacteriorhodopsin in purple membrane was reconstituted into different lipid vesicles. The effect of three different lipids on the structure and function of bacteriorhodopsin in lipid vesicles was studied by the observation on freeze-fracture eletron microscopy, the rotational diffusion of bacteriorhodopsin in lipid vesicles, the measurement of absorption spectrum, and the absorbance change with time. For these prepared samples, the results showed that DMPC was the stable lipid environment of bacteriorhodopsin; egg-pc causeed the loss of retinal chromophore of bacteriorhodopsin and it was not reversible change, cholesterol could stabilize the bacteriorhodopsin in lipid environment,but it caused the aggregation of bacteriorhodopsin.

  9. Sphingolipid symmetry governs membrane lipid raft structure.

    Science.gov (United States)

    Quinn, Peter J

    2014-07-01

    Lipid domain formation in membranes underlies the concept of rafts but their structure is controversial because the key role of cholesterol has been challenged. The configuration of glycosphingolipid receptors for agonists, bacterial toxins and enveloped viruses in plasma membrane rafts appears to be an important factor governing ligand binding and infectivity but the details are as yet unresolved. I have used X-ray diffraction methods to examine how cholesterol affects the distribution of glycosphingolipid in aqueous dispersions of an equimolar mixture of cholesterol and egg-sphingomyelin containing different proportions of glucosylceramide from human extracts. Three coexisting liquid-ordered bilayer structures are observed at 37°C in mixtures containing up to 20mol% glycosphingolipid. All the cholesterol was sequestered in one bilayer with the minimum amount of sphingomyelin (33mol%) to prevent formation of cholesterol crystals. The other two bilayers consisted of sphingomyelin and glucosylceramide. Asymmetric molecular species of glucosylceramide with N-acyl chains longer than 20 carbons form an equimolar complex with sphingomyelin in which the glycosidic residues are arranged in hexagonal array. Symmetric molecular species mix with sphingomyelin in proportions less than equimolar to form quasicrystalline bilayers. When the glycosphingolipid exceeds equimolar proportions with sphingomyelin cholesterol is incorporated into the structure and formation of a gel phase of glucosylceramide is prevented. The demonstration of particular structural features of ceramide molecular species combined with the diversity of sugar residues of glycosphingolipid classes paves the way for a rational approach to understanding the functional specificity of lipid rafts and how they are coupled across cell membranes.

  10. Cytoskeletal pinning controls phase separation in multicomponent lipid membranes.

    Science.gov (United States)

    Arumugam, Senthil; Petrov, Eugene P; Schwille, Petra

    2015-03-10

    We study the effect of a minimal cytoskeletal network formed on the surface of giant unilamellar vesicles by the prokaryotic tubulin homolog, FtsZ, on phase separation in freestanding lipid membranes. FtsZ has been modified to interact with the membrane through a membrane targeting sequence from the prokaryotic protein MinD. FtsZ with the attached membrane targeting sequence efficiently forms a highly interconnected network on membranes with a concentration-dependent mesh size, much similar to the eukaryotic cytoskeletal network underlying the plasma membrane. Using giant unilamellar vesicles formed from a quaternary lipid mixture, we demonstrate that the artificial membrane-associated cytoskeleton, on the one hand, suppresses large-scale phase separation below the phase transition temperature, and, on the other hand, preserves phase separation above the transition temperature. Our experimental observations support the ideas put forward in our previous simulation study: In particular, the picket fence effect on phase separation may explain why micrometer-scale membrane domains are observed in isolated, cytoskeleton-free giant plasma membrane vesicles, but not in intact cell membranes. The experimentally observed suppression of large-scale phase separation much below the transition temperatures also serves as an argument in favor of the cryoprotective role of the cytoskeleton. Copyright © 2015 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  11. Non-Brownian diffusion in lipid membranes: Experiments and simulations.

    Science.gov (United States)

    Metzler, R; Jeon, J-H; Cherstvy, A G

    2016-10-01

    The dynamics of constituents and the surface response of cellular membranes-also in connection to the binding of various particles and macromolecules to the membrane-are still a matter of controversy in the membrane biophysics community, particularly with respect to crowded membranes of living biological cells. We here put into perspective recent single particle tracking experiments in the plasma membranes of living cells and supercomputing studies of lipid bilayer model membranes with and without protein crowding. Special emphasis is put on the observation of anomalous, non-Brownian diffusion of both lipid molecules and proteins embedded in the lipid bilayer. While single component, pure lipid bilayers in simulations exhibit only transient anomalous diffusion of lipid molecules on nanosecond time scales, the persistence of anomalous diffusion becomes significantly longer ranged on the addition of disorder-through the addition of cholesterol or proteins-and on passing of the membrane lipids to the gel phase. Concurrently, experiments demonstrate the anomalous diffusion of membrane embedded proteins up to macroscopic time scales in the minute time range. Particular emphasis will be put on the physical character of the anomalous diffusion, in particular, the occurrence of ageing observed in the experiments-the effective diffusivity of the measured particles is a decreasing function of time. Moreover, we present results for the time dependent local scaling exponent of the mean squared displacement of the monitored particles. Recent results finding deviations from the commonly assumed Gaussian diffusion patterns in protein crowded membranes are reported. The properties of the displacement autocorrelation function of the lipid molecules are discussed in the light of their appropriate physical anomalous diffusion models, both for non-crowded and crowded membranes. In the last part of this review we address the upcoming field of membrane distortion by elongated membrane

  12. Vascular endothelial cell membranes differentiate between stretch and shear stress through transitions in their lipid phases.

    Science.gov (United States)

    Yamamoto, Kimiko; Ando, Joji

    2015-10-01

    Vascular endothelial cells (ECs) respond to the hemodynamic forces stretch and shear stress by altering their morphology, functions, and gene expression. However, how they sense and differentiate between these two forces has remained unknown. Here we report that the plasma membrane itself differentiates between stretch and shear stress by undergoing transitions in its lipid phases. Uniaxial stretching and hypotonic swelling increased the lipid order of human pulmonary artery EC plasma membranes, thereby causing a transition from the liquid-disordered phase to the liquid-ordered phase in some areas, along with a decrease in membrane fluidity. In contrast, shear stress decreased the membrane lipid order and increased membrane fluidity. A similar increase in lipid order occurred when the artificial lipid bilayer membranes of giant unilamellar vesicles were stretched by hypotonic swelling, indicating that this is a physical phenomenon. The cholesterol content of EC plasma membranes significantly increased in response to stretch but clearly decreased in response to shear stress. Blocking these changes in the membrane lipid order by depleting membrane cholesterol with methyl-β-cyclodextrin or by adding cholesterol resulted in a marked inhibition of the EC response specific to stretch and shear stress, i.e., phosphorylation of PDGF receptors and phosphorylation of VEGF receptors, respectively. These findings indicate that EC plasma membranes differently respond to stretch and shear stress by changing their lipid order, fluidity, and cholesterol content in opposite directions and that these changes in membrane physical properties are involved in the mechanotransduction that activates membrane receptors specific to each force.

  13. Cholecystokinin Elevates Mouse Plasma Lipids

    Science.gov (United States)

    Zhou, Lichun; Yang, Hong; Lin, Xinghua; Okoro, Emmanuel U.; Guo, Zhongmao

    2012-01-01

    Cholecystokinin (CCK) is a peptide hormone that induces bile release into the intestinal lumen which in turn aids in fat digestion and absorption in the intestine. While excretion of bile acids and cholesterol into the feces eliminates cholesterol from the body, this report examined the effect of CCK on increasing plasma cholesterol and triglycerides in mice. Our data demonstrated that intravenous injection of [Thr28, Nle31]-CCK at a dose of 50 ng/kg significantly increased plasma triglyceride and cholesterol levels by 22 and 31%, respectively, in fasting low-density lipoprotein receptor knockout (LDLR−/−) mice. The same dose of [Thr28, Nle31]-CCK induced 6 and 13% increases in plasma triglyceride and cholesterol, respectively, in wild-type mice. However, these particular before and after CCK treatment values did not achieve statistical significance. Oral feeding of olive oil further elevated plasma triglycerides, but did not alter plasma cholesterol levels in CCK-treated mice. The increased plasma cholesterol in CCK-treated mice was distributed in very-low, low and high density lipoproteins (VLDL, LDL and HDL) with less of an increase in HDL. Correspondingly, the plasma apolipoprotein (apo) B48, B100, apoE and apoAI levels were significantly higher in the CCK-treated mice than in untreated control mice. Ligation of the bile duct, blocking CCK receptors with proglumide or inhibition of Niemann-Pick C1 Like 1 transporter with ezetimibe reduced the hypercholesterolemic effect of [Thr28, Nle31]-CCK in LDLR−/− mice. These findings suggest that CCK-increased plasma cholesterol and triglycerides as a result of the reabsorption of biliary lipids from the intestine. PMID:23300532

  14. Cholecystokinin elevates mouse plasma lipids.

    Directory of Open Access Journals (Sweden)

    Lichun Zhou

    Full Text Available Cholecystokinin (CCK is a peptide hormone that induces bile release into the intestinal lumen which in turn aids in fat digestion and absorption in the intestine. While excretion of bile acids and cholesterol into the feces eliminates cholesterol from the body, this report examined the effect of CCK on increasing plasma cholesterol and triglycerides in mice. Our data demonstrated that intravenous injection of [Thr28, Nle31]-CCK at a dose of 50 ng/kg significantly increased plasma triglyceride and cholesterol levels by 22 and 31%, respectively, in fasting low-density lipoprotein receptor knockout (LDLR(-/- mice. The same dose of [Thr28, Nle31]-CCK induced 6 and 13% increases in plasma triglyceride and cholesterol, respectively, in wild-type mice. However, these particular before and after CCK treatment values did not achieve statistical significance. Oral feeding of olive oil further elevated plasma triglycerides, but did not alter plasma cholesterol levels in CCK-treated mice. The increased plasma cholesterol in CCK-treated mice was distributed in very-low, low and high density lipoproteins (VLDL, LDL and HDL with less of an increase in HDL. Correspondingly, the plasma apolipoprotein (apo B48, B100, apoE and apoAI levels were significantly higher in the CCK-treated mice than in untreated control mice. Ligation of the bile duct, blocking CCK receptors with proglumide or inhibition of Niemann-Pick C1 Like 1 transporter with ezetimibe reduced the hypercholesterolemic effect of [Thr28, Nle31]-CCK in LDLR(-/- mice. These findings suggest that CCK-increased plasma cholesterol and triglycerides as a result of the reabsorption of biliary lipids from the intestine.

  15. 质膜脂筏的研究进展%Research Progress of Lipid Raft on Plasma Membrane

    Institute of Scientific and Technical Information of China (English)

    姚茂平; 陈雄庭

    2007-01-01

    质膜具有一些被命名为"脂筏(lipid rafts)"和"质膜微囊(Caveolae)"的微区.脂筏由丰富的鞘糖脂、胆固醇和酰化的蛋白质动态聚集而成,形成相对稳定的具有一定功能的筹结构漂浮于二维流动的细胞质膜中,被人们形象地称之为"功能筏".具有参与胞吞胞饮、信号转导、运输胆固醇等重要功能,随着人们在分子水平上认识的不断深化,脂筏结构和功能的物理、化学基础研究方面也取得了初步的进展.

  16. Erythrocyte membrane, plasma and atherosclerotic plaque lipid pattern in coronary heart disease Perfil lipídico de membrana de eritrocito, plasma y placa ateromatosa en la enfermedad coronaria

    Directory of Open Access Journals (Sweden)

    Natalia R. Lausada

    2007-10-01

    Full Text Available The objective was to analyze the lipid composition of the atherosclerotic plaque (AP, plasma and erythrocyte membrane (EM in patients with advanced coronary heart disease (CHD. AP were obtained through endarterectomy in 18 patients. Ten normolipemic healthy subjects were selected to obtain the normal lipid pattern profile. Total lipids of AP and EM were determined by HPTLC, and the fatty acid profile from AP, EM and plasma using TLC-FID. The relative amount of the lipid species analyzed in AP was in line with the data in the literature [phospholipids: 23.5 mol% ± 3.5; total cholesterol 68.9 mol% ± 7.9; triglyceride 7.6 mol% ± 3.4]. Plasma and EM from CHD patients compared to controls, showed a decrease in polyunsaturated fatty acids and an increase in saturated fatty acids leading to a decrease in the unsaturation index (plasma: 1.67 ± 0.06 vs. 1.28 ± 0.03, PEl objetivo fue analizar la composición lipídica de las membranas de eritrocitos (ME, plasma y placas ateromatosas (PA en pacientes con enfermedad coronaria avanzada (ECV. Las PA fueron obtenidas de endarterectomías coronarias de 18 pacientes. Fueron seleccionados 10 sujetos sanos, normolipémicos, como grupo control. Los lípidos totales de PA y ME se determinaron utilizando HPTLC, y el perfil de ácidos grasos de las PA, ME y plasma mediante TLC-FID. La cantidad relativa de las especies lipídicas obtenidas de las PA coinciden con la literatura [fosfolípidos 23.5 mol% ± 3.5; colesterol total 68.9 mol% ± 7.9; triglicéridos 7.6 mol% ± 3.4]. En el plasma y en las ME de los pacientes con ECV se observó, comparando con los pacientes controles, una disminución de los ácidos grasos poli-no saturados acompañado de un aumento de los ácidos grasos saturados que provocó el descenso del índice de instauración (plasma: 1.67 ± 0.06 vs. 1.28 ± 0.03, P<0.05; ME: 2.28 ± 0.04 vs. 1.25 ± 0.010, P<0.05 y el incremento del cociente AG saturados/insaturados (plasma: 0.35 ± 0.02 vs. 0

  17. Chemically Stable Lipids for Membrane Protein Crystallization

    Energy Technology Data Exchange (ETDEWEB)

    Ishchenko, Andrii; Peng, Lingling; Zinovev, Egor; Vlasov, Alexey; Lee, Sung Chang; Kuklin, Alexander; Mishin, Alexey; Borshchevskiy, Valentin; Zhang, Qinghai; Cherezov, Vadim (MIPT); (USC); (Scripps)

    2017-05-01

    The lipidic cubic phase (LCP) has been widely recognized as a promising membrane-mimicking matrix for biophysical studies of membrane proteins and their crystallization in a lipidic environment. Application of this material to a wide variety of membrane proteins, however, is hindered due to a limited number of available host lipids, mostly monoacylglycerols (MAGs). Here, we designed, synthesized, and characterized a series of chemically stable lipids resistant to hydrolysis, with properties complementary to the widely used MAGs. In order to assess their potential to serve as host lipids for crystallization, we characterized the phase properties and lattice parameters of mesophases made of two most promising lipids at a variety of different conditions by polarized light microscopy and small-angle X-ray scattering. Both lipids showed remarkable chemical stability and an extended LCP region in the phase diagram covering a wide range of temperatures down to 4 °C. One of these lipids has been used for crystallization and structure determination of a prototypical membrane protein bacteriorhodopsin at 4 and 20 °C.

  18. Very long-chain fatty acid-containing lipids rather than sphingolipids per se are required for raft association and stable surface transport of newly synthesized plasma membrane ATPase in yeast.

    Science.gov (United States)

    Gaigg, Barbara; Toulmay, Alexandre; Schneiter, Roger

    2006-11-10

    The proton-pumping H+-ATPase, Pma1p, is an abundant and very long lived polytopic protein of the yeast plasma membrane. Pma1p constitutes a major cargo of the secretory pathway and thus serves as a model to study plasma membrane biogenesis. Pma1p associates with detergent-resistant membrane domains (lipid "rafts") already in the ER, and a lack of raft association correlates with mistargeting of the protein to the vacuole, where it is degraded. We are analyzing the role of specific lipids in membrane domain formation and have previously shown that surface transport of Pma1p is independent of newly synthesized sterols but that sphingolipids with C26 very long chain fatty acid are crucial for raft association and surface transport of Pma1p (Gaigg, B., Timischl, B., Corbino, L., and Schneiter, R. (2005) J. Biol. Chem. 280, 22515-22522). We now describe a more detailed analysis of the function that sphingolipids play in this process. Using a yeast strain in which the essential function of sphingolipids is substituted by glycerophospholipids containing C26 very long chain fatty acids, we find that sphingolipids per se are dispensable for raft association and surface delivery of Pma1p but that the C26 fatty acid is crucial. We thus conclude that the essential function of sphingolipids for membrane domain formation and stable surface delivery of Pma1p is provided by the C26 fatty acid that forms part of the yeast ceramide.

  19. Binding of Serotonin to Lipid Membranes

    DEFF Research Database (Denmark)

    Peters, Günther H.J.; Wang, Chunhua; Cruys-Bagger, Nicolaj;

    2013-01-01

    dynamics (MD) simulations. 5-HT was found to interact strongly with lipid bilayers (partitioning coefficient ∼1200 in mole fraction units), and this is highly unusual for a hydrophilic solute like 5-HT which has a bulk, oil–water partitioning coefficient well below unity. It follows that membrane affinity...... and a prevailing residence between the phosphate and the carbonyl groups of the lipid. The unprotonated form of 5-HT shows the opposite orientation, with the primary amine pointing toward the membrane core. Partitioning of 5-HT was found to decrease lipid chain order. These distinctive interactions of 5-HT...

  20. Imaging lipid domains in cell membranes: the advent of super-resolution fluorescence microscopy

    Directory of Open Access Journals (Sweden)

    Dylan Myers Owen

    2013-12-01

    Full Text Available The lipid bilayer of model membranes, liposomes reconstituted from cell lipids, and plasma membrane vesicles and spheres can separate into two distinct liquid phases to yield lipid domains with liquid-ordered and liquid-disordered properties. These observations are the basis of the lipid raft hypothesis that postulates the existence of cholesterol-enriched ordered-phase lipid domains in cell membranes that could regulate protein mobility, localization and interaction. Here we review the evidence that nano-scaled lipid complexes and meso-scaled lipid domains exist in cell membranes and how new fluorescence microscopy techniques that overcome the diffraction limit provide new insights into lipid organization in cell membranes.

  1. Atomistic Monte Carlo simulation of lipid membranes

    DEFF Research Database (Denmark)

    Wüstner, Daniel; Sklenar, Heinz

    2014-01-01

    Biological membranes are complex assemblies of many different molecules of which analysis demands a variety of experimental and computational approaches. In this article, we explain challenges and advantages of atomistic Monte Carlo (MC) simulation of lipid membranes. We provide an introduction......, as assessed by calculation of molecular energies and entropies. We also show transition from a crystalline-like to a fluid DPPC bilayer by the CBC local-move MC method, as indicated by the electron density profile, head group orientation, area per lipid, and whole-lipid displacements. We discuss the potential...... of local-move MC methods in combination with molecular dynamics simulations, for example, for studying multi-component lipid membranes containing cholesterol....

  2. Microcompartments within the yeast plasma membrane.

    Science.gov (United States)

    Merzendorfer, Hans; Heinisch, Jürgen J

    2013-02-01

    Recent research in cell biology makes it increasingly clear that the classical concept of compartmentation of eukaryotic cells into different organelles performing distinct functions has to be extended by microcompartmentation, i.e., the dynamic interaction of proteins, sugars, and lipids at a suborganellar level, which contributes significantly to a proper physiology. As different membrane compartments (MCs) have been described in the yeast plasma membrane, such as those defined by Can1 and Pma1 (MCCs and MCPs), Saccharomyces cerevisiae can serve as a model organism, which is amenable to genetic, biochemical, and microscopic studies. In this review, we compare the specialized microcompartment of the yeast bud neck with other plasma membrane substructures, focusing on eisosomes, cell wall integrity-sensing units, and chitin-synthesizing complexes. Together, they ensure a proper cell division at the end of mitosis, an intricately regulated process, which is essential for the survival and proliferation not only of fungal, but of all eukaryotic cells.

  3. How Membrane-Active Peptides Get into Lipid Membranes.

    Science.gov (United States)

    Sani, Marc-Antoine; Separovic, Frances

    2016-06-21

    The structure-function relationship for a family of antimicrobial peptides (AMPs) from the skin of Australian tree frogs is discussed and compared with that of peptide toxins from bee and Australian scorpion venoms. Although these membrane-active peptides induce a similar cellular fate by disrupting the lipid bilayer integrity, their lytic activity is achieved via different modes of action, which are investigated in relation to amino acid sequence, secondary structure, and membrane lipid composition. In order to better understand what structural features govern the interaction between peptides and lipid membranes, cell-penetrating peptides (CPPs), which translocate through the membrane without compromising its integrity, are also discussed. AMPs possess membrane lytic activities that are naturally designed to target the cellular membrane of pathogens or competitors. They are extremely diverse in amino acid composition and often show specificity against a particular strain of microbe. Since our antibiotic arsenal is declining precariously in the face of the rise in multiantibiotic resistance, AMPs increasingly are seen as a promising alternative. In an effort to understand their molecular mechanism, biophysical studies of a myriad of AMPs have been reported, yet no unifying mechanism has emerged, rendering difficult the rational design of drug leads. Similarly, a wide variety of cytotoxic peptides are found in venoms, the best known being melittin, yet again, predicting their activity based on a particular amino acid composition or secondary structure remains elusive. A common feature of these membrane-active peptides is their preference for the lipid environment. Indeed, they are mainly unstructured in solution and, in the presence of lipid membranes, quickly adsorb onto the surface, change their secondary structure, eventually insert into the hydrophobic core of the membrane bilayer, and finally disrupt the bilayer integrity. These steps define the molecular

  4. Assessing the nature of lipid raft membranes

    DEFF Research Database (Denmark)

    Niemelä, Perttu S; Ollila, Samuli; Hyvönen, Marja T

    2007-01-01

    to intriguing lateral pressure profiles that are distinctly different from corresponding profiles in nonraft-like membranes. The results propose that the functioning of certain classes of membrane proteins is regulated by changes in the lateral pressure profile, which can be altered by a change in lipid content....... of highly ordered lateral domains rich in sphingomyelin and cholesterol (CHOL). These domains, called functional lipid rafts, have been suggested to take part in a variety of dynamic cellular processes such as membrane trafficking, signal transduction, and regulation of the activity of membrane proteins....... However, despite the proposed importance of these domains, their properties, and even the precise nature of the lipid phases, have remained open issues mainly because the associated short time and length scales have posed a major challenge to experiments. In this work, we employ extensive atom...

  5. Menthol-induced action potentials in Conocephalum conicum as a result of unspecific interactions between menthol and the lipid phase of the plasma membrane.

    Science.gov (United States)

    Kupisz, Kamila; Trebacz, Kazimierz; Gruszecki, Wiesław I

    2015-07-01

    Our previous study has shown that the liverwort Conocephalum conicum generates action potentials (APs) in response to both temperature drop and menthol, which are also activators of the TRPM8 (transient receptor potential melastatin 8) receptor in animals. Not only similarities but also differences between electrical reactions to menthol and cooling observed in the liverwort aroused our interest in the action of menthol at the molecular level. Patch-clamp investigations have shown that menthol causes a reduction of current flowing through slow vacuolar (SV) channels to 29 ± 10% of the initial value (n = 9); simultaneously, it does not influence magnitudes of currents passing through a single SV channel. This may point to an unspecific interaction between menthol and the lipid phase of the membrane. An influence of menthol on lipid organization in membranes was investigated in two-component monomolecular layers formed with menthol and dipalmitoylphosphatidylcholine (DPPC) at the argon-water interface. Analyses of the mean molecular area parameters vs the molar fraction of the menthol component have shown over-additivity (approximately 20 Å(2) ) in the region of high molar fractions of menthol. Infrared absorption spectroscopy studies have shown that menthol, most probably, induces breaking of a hydrogen bond network formed by ester carbonyl groups and water bridges in the lipid membrane and binds to the polar head group region of DPPC. We conclude that the disruption in the lipid phase of the membrane influences ion channels and/or pumps and subsequently causes generation of APs in excitable plants such as C. conicum.

  6. Self-Segregation of Myelin Membrane Lipids in Model Membranes

    NARCIS (Netherlands)

    Yurlova, Larisa; Kahya, Nicoletta; Aggarwal, Shweta; Kaiser, Hermann-Josef; Chiantia, Salvatore; Bakhti, Mostafa; Pewzner-Jung, Yael; Ben-David, Oshrit; Futerman, Anthony H.; Bruegger, Britta; Simons, Mikael

    2011-01-01

    Rapid conduction of nerve impulses requires coating of axons by myelin sheaths, which are multilamellar, lipid-rich membranes produced by oligodendrocytes in the central nervous system. To act as an insulator, myelin has to form a stable and firm membrane structure. In this study, we have analyzed t

  7. Assessing the nature of lipid raft membranes.

    Directory of Open Access Journals (Sweden)

    Perttu S Niemelä

    2007-02-01

    Full Text Available The paradigm of biological membranes has recently gone through a major update. Instead of being fluid and homogeneous, recent studies suggest that membranes are characterized by transient domains with varying fluidity. In particular, a number of experimental studies have revealed the existence of highly ordered lateral domains rich in sphingomyelin and cholesterol (CHOL. These domains, called functional lipid rafts, have been suggested to take part in a variety of dynamic cellular processes such as membrane trafficking, signal transduction, and regulation of the activity of membrane proteins. However, despite the proposed importance of these domains, their properties, and even the precise nature of the lipid phases, have remained open issues mainly because the associated short time and length scales have posed a major challenge to experiments. In this work, we employ extensive atom-scale simulations to elucidate the properties of ternary raft mixtures with CHOL, palmitoylsphingomyelin (PSM, and palmitoyloleoylphosphatidylcholine. We simulate two bilayers of 1,024 lipids for 100 ns in the liquid-ordered phase and one system of the same size in the liquid-disordered phase. The studies provide evidence that the presence of PSM and CHOL in raft-like membranes leads to strongly packed and rigid bilayers. We also find that the simulated raft bilayers are characterized by nanoscale lateral heterogeneity, though the slow lateral diffusion renders the interpretation of the observed lateral heterogeneity more difficult. The findings reveal aspects of the role of favored (specific lipid-lipid interactions within rafts and clarify the prominent role of CHOL in altering the properties of the membrane locally in its neighborhood. Also, we show that the presence of PSM and CHOL in rafts leads to intriguing lateral pressure profiles that are distinctly different from corresponding profiles in nonraft-like membranes. The results propose that the functioning of

  8. The Plasma Membrane of Saccharomyces cerevisiae : Structure, Function, and Biogenesis

    NARCIS (Netherlands)

    VANDERREST, ME; KAMMINGA, AH; NAKANO, A; ANRAKU, Y; POOLMAN, B; KONINGS, WN

    1995-01-01

    The composition of phospholipids, sphingolipids, and sterols in the plasma membrane has a strong influence on the activity of the proteins associated or embedded in the lipid bilayer. Since most lipid-synthesizing enzymes in Saccharomyces cerevisiae are located in intracellular organelles, an extens

  9. The Plasma Membrane of Saccharomyces cerevisiae : Structure, Function, and Biogenesis

    NARCIS (Netherlands)

    VANDERREST, ME; KAMMINGA, AH; NAKANO, A; ANRAKU, Y; POOLMAN, B; KONINGS, WN

    The composition of phospholipids, sphingolipids, and sterols in the plasma membrane has a strong influence on the activity of the proteins associated or embedded in the lipid bilayer. Since most lipid-synthesizing enzymes in Saccharomyces cerevisiae are located in intracellular organelles, an

  10. Biotechnology Applications of Tethered Lipid Bilayer Membranes

    Directory of Open Access Journals (Sweden)

    Joshua A. Jackman

    2012-12-01

    Full Text Available The importance of cell membranes in biological systems has prompted the development of model membrane platforms that recapitulate fundamental aspects of membrane biology, especially the lipid bilayer environment. Tethered lipid bilayers represent one of the most promising classes of model membranes and are based on the immobilization of a planar lipid bilayer on a solid support that enables characterization by a wide range of surface-sensitive analytical techniques. Moreover, as the result of molecular engineering inspired by biology, tethered bilayers are increasingly able to mimic fundamental properties of natural cell membranes, including fluidity, electrical sealing and hosting transmembrane proteins. At the same time, new methods have been employed to improve the durability of tethered bilayers, with shelf-lives now reaching the order of weeks and months. Taken together, the capabilities of tethered lipid bilayers have opened the door to biotechnology applications in healthcare, environmental monitoring and energy storage. In this review, several examples of such applications are presented. Beyond the particulars of each example, the focus of this review is on the emerging design and characterization strategies that made these applications possible. By drawing connections between these strategies and promising research results, future opportunities for tethered lipid bilayers within the biotechnology field are discussed.

  11. Membrane lipid rafts and neurobiology: age-related changes in membrane lipids and loss of neuronal function.

    Science.gov (United States)

    Egawa, Junji; Pearn, Matthew L; Lemkuil, Brian P; Patel, Piyush M; Head, Brian P

    2016-08-15

    A better understanding of the cellular physiological role that plasma membrane lipids, fatty acids and sterols play in various cellular systems may yield more insight into how cellular and whole organ function is altered during the ageing process. Membrane lipid rafts (MLRs) within the plasma membrane of most cells serve as key organizers of intracellular signalling and tethering points of cytoskeletal components. MLRs are plasmalemmal microdomains enriched in sphingolipids, cholesterol and scaffolding proteins; they serve as a platform for signal transduction, cytoskeletal organization and vesicular trafficking. Within MLRs are the scaffolding and cholesterol binding proteins named caveolin (Cav). Cavs not only organize a multitude of receptors including neurotransmitter receptors (NMDA and AMPA receptors), signalling proteins that regulate the production of cAMP (G protein-coupled receptors, adenylyl cyclases, phosphodiesterases (PDEs)), and receptor tyrosine kinases involved in growth (Trk), but also interact with components that modulate actin and tubulin cytoskeletal dynamics (e.g. RhoGTPases and actin binding proteins). MLRs are essential for the regulation of the physiology of organs such as the brain, and age-related loss of cholesterol from the plasma membrane leads to loss of MLRs, decreased presynaptic vesicle fusion, and changes in neurotransmitter release, all of which contribute to different forms of neurodegeneration. Thus, MLRs provide an active membrane domain that tethers and reorganizes the cytoskeletal machinery necessary for membrane and cellular repair, and genetic interventions that restore MLRs to normal cellular levels may be exploited as potential therapeutic means to reverse the ageing and neurodegenerative processes.

  12. A mechanism of raft formation on both plasma membrane layers

    Science.gov (United States)

    Sornbundit, Kan; Modchang, Charin; Triampo, Wannapong; Triampo, Darapond; Nuttavut, Narin

    2013-10-01

    A double-layered membrane model is proposed to explain raft formation and induction on extracellular (outer) and cytoplasmic (inner) leaflets of plasma membranes in a situation where only the outer layer has a tendency to phase-separate. In the model, lipid exchange with the surrounding medium is allowed on both layers, but lipid exchange between layers is not allowed. Simulations display domain stabilization on both layers. The effect of the lipid recycling frequencies on stationary domain sizes is also investigated. It is found that stationary domain sizes decrease when lipid recycling frequencies are stronger. Linear stability analysis is used to verify the results.

  13. Atomistic Monte Carlo simulation of lipid membranes

    DEFF Research Database (Denmark)

    Wüstner, Daniel; Sklenar, Heinz

    2014-01-01

    , as assessed by calculation of molecular energies and entropies. We also show transition from a crystalline-like to a fluid DPPC bilayer by the CBC local-move MC method, as indicated by the electron density profile, head group orientation, area per lipid, and whole-lipid displacements. We discuss the potential......Biological membranes are complex assemblies of many different molecules of which analysis demands a variety of experimental and computational approaches. In this article, we explain challenges and advantages of atomistic Monte Carlo (MC) simulation of lipid membranes. We provide an introduction...... into the various move sets that are implemented in current MC methods for efficient conformational sampling of lipids and other molecules. In the second part, we demonstrate for a concrete example, how an atomistic local-move set can be implemented for MC simulations of phospholipid monomers and bilayer patches...

  14. Evidence for condensed complexes of cholesterol in lipid membranes

    Science.gov (United States)

    Ratajczak, Maria K.

    Although cholesterol is a predominant lipid in the eukaryotic plasma membrane, its interactions with other lipids are still not well understood. Insights into the nature of lipid assembly can be gained from examining lipid-cholesterol interaction using model systems. A key observation was the discovery of liquid-liquid phase diagrams with two critical points in the binary mixtures of cholesterol and lipids. The shape of the phase diagrams can be explained by a thermodynamic model of "condensed complexes". In our quest to characterize cholesterol-lipid interactions, we determined phase diagrams of cholesterol and phospholipids that point to the existence of condensed complexes. This complex formation hypothesis was further supported by experiments involving cholesterol removal by cyclodextrin, grazing x-ray diffraction and x-ray reflectivity studies and isothermal calorimetry. Our study aimed at establishing a correlation (or the lack of) between domain formation and complex formation, as well as determining the mode of cholesterol association with different lipids based on their structural and physical properties. We established a displacement assay by which we were able to probe cholesterol-lipid interactions by perturbing them in the presence of an intercalator that competes with cholesterol for association with lipids. Our data support the condensed complex model between cholesterol and lipids, and cholesterol when complexed with lipids shows low activity whereas free, uncomplexed cholesterol exhibits high activity. We were successful in modulating cholesterol activity by varying the level of intercalator while keeping the cholesterol content fixed. In this thesis, not only have we shown that cholesterol can be displaced by intercalators in model systems, we have further established that such displacement can take place in membranes of live cell.

  15. Regulation of the Plasma Membrane H+-ATPase

    DEFF Research Database (Denmark)

    Falhof, Janus

    The plasma membrane (PM) H+-ATPase is responsible for generating the electrochemical gradientthat drives the secondary transport of nutrients across the cellular membrane. It belongs to a familyof cation and lipid transporters that are vital to many organisms. PM H+-ATPases are Type P3AATPases...

  16. Nanodomain stabilization dynamics in plasma membranes of biological cells

    Science.gov (United States)

    Das, Tamal; Maiti, Tapas K.; Chakraborty, Suman

    2011-02-01

    We discover that a synergistically amplifying role of stabilizing membrane proteins and continuous lipid recycling can explain the physics governing the stability, polydispersity, and dynamics of lipid raft domains in plasma membranes of biological cells. We establish the conjecture using a generalized order parameter based on theoretical formalism, endorsed by detailed scaling arguments and domain mapping. Quantitative agreements with morphological distributions of raft complexes, as obtained from Förster resonance energy transfer based visualization, support the present theoretical conjecture.

  17. Microparticle Assembly Pathways on Lipid Membranes

    Science.gov (United States)

    van der Wel, Casper; Heinrich, Doris; Kraft, Daniela J.

    2017-09-01

    Understanding interactions between microparticles and lipid membranes is of increasing importance, especially for unraveling the influence of microplastics on our health and environment. Here, we study how a short-ranged adhesive force between microparticles and model lipid membranes causes membrane-mediated particle assembly. Using confocal microscopy, we observe the initial particle attachment to the membrane, then particle wrapping, and in rare cases spontaneous membrane tubulation. In the attached state, we measure that the particle mobility decreases by 26%. If multiple particles adhere to the same vesicle, their initial single-particle state determines their interactions and subsequent assembly pathways: 1) attached particles only aggregate when small adhesive vesicles are present in solution, 2) wrapped particles reversibly attract one another by membrane deformation, and 3) a combination of wrapped and attached particles form membrane-mediated dimers, which further assemble into a variety of complex structures. The experimental observation of distinct assembly pathways induced only by a short ranged membrane-particle adhesion, shows that a cellular cytoskeleton or other active components are not required for microparticle aggregation. We suggest that this membrane-mediated microparticle aggregation is a reason behind reported long retention times of polymer microparticles in organisms.

  18. Engineering Lipid Bilayer Membranes for Protein Studies

    Directory of Open Access Journals (Sweden)

    Muhammad Shuja Khan

    2013-10-01

    Full Text Available Lipid membranes regulate the flow of nutrients and communication signaling between cells and protect the sub-cellular structures. Recent attempts to fabricate artificial systems using nanostructures that mimic the physiological properties of natural lipid bilayer membranes (LBM fused with transmembrane proteins have helped demonstrate the importance of temperature, pH, ionic strength, adsorption behavior, conformational reorientation and surface density in cellular membranes which all affect the incorporation of proteins on solid surfaces. Much of this work is performed on artificial templates made of polymer sponges or porous materials based on alumina, mica, and porous silicon (PSi surfaces. For example, porous silicon materials have high biocompatibility, biodegradability, and photoluminescence, which allow them to be used both as a support structure for lipid bilayers or a template to measure the electrochemical functionality of living cells grown over the surface as in vivo. The variety of these media, coupled with the complex physiological conditions present in living systems, warrant a summary and prospectus detailing which artificial systems provide the most promise for different biological conditions. This study summarizes the use of electrochemical impedance spectroscopy (EIS data on artificial biological membranes that are closely matched with previously published biological systems using both black lipid membrane and patch clamp techniques.

  19. Insulin sensitivity is independent of lipid binding protein trafficking at the plasma membrane in human skeletal muscle: effect of a 3-day, high-fat diet.

    Science.gov (United States)

    Jordy, Andreas B; Serup, Annette K; Karstoft, Kristian; Pilegaard, Henriette; Kiens, Bente; Jeppesen, Jacob

    2014-11-01

    The aim of the present study was to investigate lipid-induced regulation of lipid binding proteins in human skeletal muscle and the impact hereof on insulin sensitivity. Eleven healthy male subjects underwent a 3-day hypercaloric and high-fat diet regime. Muscle biopsies were taken before and after the diet intervention, and giant sarcolemmal vesicles were prepared. The high-fat diet induced decreased insulin sensitivity, but this was not associated with a relocation of FAT/CD36 or FABPpm protein to the sarcolemma. However, FAT/CD36 and FABPpm mRNA, but not the proteins, were upregulated by increased fatty acid availability. This suggests a time dependency in the upregulation of FAT/CD36 and FABPpm protein during high availability of plasma fatty acids. Furthermore, we did not detect FATP1 and FATP4 protein in giant sarcolemmal vesicles obtained from human skeletal muscle. In conclusion, this study shows that a short-term lipid-load increases mRNA content of key lipid handling proteins in human muscle. However, decreased insulin sensitivity after a high-fat diet is not accompanied with relocation of FAT/CD36 or FABPpm protein to the sarcolemma. Finally, FATP1 and FATP4 protein was located intracellularly but not at the sarcolemma in humans. Copyright © 2014 the American Physiological Society.

  20. Nonlinear adhesion dynamics of confined lipid membranes

    Science.gov (United States)

    To, Tung; Le Goff, Thomas; Pierre-Louis, Olivier

    Lipid membranes, which are ubiquitous objects in biological environments are often confined. For example, they can be sandwiched between a substrate and the cytoskeleton between cell adhesion, or between other membranes in stacks, or in the Golgi apparatus. We present a study of the nonlinear dynamics of membranes in a model system, where the membrane is confined between two flat walls. The dynamics derived from the lubrication approximation is highly nonlinear and nonlocal. The solution of this model in one dimension exhibits frozen states due to oscillatory interactions between membranes caused by the bending rigidity. We develope a kink model for these phenomena based on the historical work of Kawasaki and Otha. In two dimensions, the dynamics is more complex, and depends strongly on the amount of excess area in the system. We discuss the relevance of our findings for experiments on model membranes, and for biological systems. Supported by the grand ANR Biolub.

  1. Adaptive lipid packing and bioactivity in membrane domains.

    Directory of Open Access Journals (Sweden)

    Erdinc Sezgin

    Full Text Available Lateral compositional and physicochemical heterogeneity is a ubiquitous feature of cellular membranes on various length scales, from molecular assemblies to micrometric domains. Segregated lipid domains of increased local order, referred to as rafts, are believed to be prominent features in eukaryotic plasma membranes; however, their exact nature (i.e. size, lifetime, composition, homogeneity in live cells remains difficult to define. Here we present evidence that both synthetic and natural plasma membranes assume a wide range of lipid packing states with varying levels of molecular order. These states may be adapted and specifically tuned by cells during active cellular processes, as we show for stimulated insulin secretion. Most importantly, these states regulate both the partitioning of molecules between coexisting domains and the bioactivity of their constituent molecules, which we demonstrate for the ligand binding activity of the glycosphingolipid receptor GM1. These results confirm the complexity and flexibility of lipid-mediated membrane organization and reveal mechanisms by which this flexibility could be functionalized by cells.

  2. Fatty acid profiles from the plasma membrane and detergent resistant membranes of two plant species.

    Science.gov (United States)

    Carmona-Salazar, Laura; El Hafidi, Mohammed; Gutiérrez-Nájera, Nora; Noyola-Martínez, Liliana; González-Solís, Ariadna; Gavilanes-Ruíz, Marina

    2015-01-01

    It is essential to establish the composition of the plant plasma membrane in order to understand its organization and behavior under continually changing environments. Knowledge of the lipid phase, in particular the fatty acid (FA) complex repertoire, is important since FAs determine many of the physical-chemical membrane properties. FAs are constituents of the membrane glycerolipid and sphingolipid backbones and can also be linked to some sterols. In addition, FAs are components of complex lipids that can constitute membrane micro-domains, and the use of detergent-resistant membranes is a common approach to study their composition. The diversity and cellular allocation of the membrane lipids containing FAs are very diverse and the approaches to analyze them provide only general information. In this work, a detailed FA analysis was performed using highly purified plasma membranes from bean leaves and germinating maize embryos and their respective detergent-resistant membrane preparations. The analyses showed the presence of a significant amount of very long chain FAs (containing 28C, 30C and 32C), in both plasma membrane preparations from bean and maize, that have not been previously reported. Herein is demonstrated that a significant enrichment of very long chain saturated FAs and saturated FAs can occur in detergent-resistant membrane preparations, as compared to the plasma membranes from both plant species. Considering that a thorough analysis of FAs is rarely performed in purified plasma membranes and detergent-resistant membranes, this work provides qualitative and quantitative evidence on the contributions of the length and saturation of FAs to the organization of the plant plasma membrane and detergent-resistant membranes.

  3. [Updated detection of the function of sperm plasma membrane].

    Science.gov (United States)

    Zhou, Xin; Xia, Xin-Yi; Huang, Yu-Feng

    2010-08-01

    The sperm plasma membrane is rich in polyunsaturated fatty acids and a variety of proteins, and its function is associated with sperm capacitation, acrosome reaction and sperm-egg fusion. Sperm fertilizability can be predicted by detecting the function of the sperm plasma membrane, which is performed mainly with the following five techniques: sperm hypoosmotic swelling test, Eosin gamma water test, sperm membrane lipid peroxidation determination, seminal plasma superoxide dismutase determination, and flow cytometry. The evaluation of the function of sperm plasma membrane can be applied in detecting semen quality, selecting semen centrifugation, assessing the quality and fertilizability of sex-sorted sperm, improving cryopreservation, and guiding the optimization of intracytoplasmic sperm injection. This review presents an update on the principles, methods and steps of the detection of sperm plasma membrane function, as well as an overview of its status quo and application.

  4. Atomistic Monte Carlo simulation of lipid membranes.

    Science.gov (United States)

    Wüstner, Daniel; Sklenar, Heinz

    2014-01-24

    Biological membranes are complex assemblies of many different molecules of which analysis demands a variety of experimental and computational approaches. In this article, we explain challenges and advantages of atomistic Monte Carlo (MC) simulation of lipid membranes. We provide an introduction into the various move sets that are implemented in current MC methods for efficient conformational sampling of lipids and other molecules. In the second part, we demonstrate for a concrete example, how an atomistic local-move set can be implemented for MC simulations of phospholipid monomers and bilayer patches. We use our recently devised chain breakage/closure (CBC) local move set in the bond-/torsion angle space with the constant-bond-length approximation (CBLA) for the phospholipid dipalmitoylphosphatidylcholine (DPPC). We demonstrate rapid conformational equilibration for a single DPPC molecule, as assessed by calculation of molecular energies and entropies. We also show transition from a crystalline-like to a fluid DPPC bilayer by the CBC local-move MC method, as indicated by the electron density profile, head group orientation, area per lipid, and whole-lipid displacements. We discuss the potential of local-move MC methods in combination with molecular dynamics simulations, for example, for studying multi-component lipid membranes containing cholesterol.

  5. Atomistic Monte Carlo Simulation of Lipid Membranes

    Directory of Open Access Journals (Sweden)

    Daniel Wüstner

    2014-01-01

    Full Text Available Biological membranes are complex assemblies of many different molecules of which analysis demands a variety of experimental and computational approaches. In this article, we explain challenges and advantages of atomistic Monte Carlo (MC simulation of lipid membranes. We provide an introduction into the various move sets that are implemented in current MC methods for efficient conformational sampling of lipids and other molecules. In the second part, we demonstrate for a concrete example, how an atomistic local-move set can be implemented for MC simulations of phospholipid monomers and bilayer patches. We use our recently devised chain breakage/closure (CBC local move set in the bond-/torsion angle space with the constant-bond-length approximation (CBLA for the phospholipid dipalmitoylphosphatidylcholine (DPPC. We demonstrate rapid conformational equilibration for a single DPPC molecule, as assessed by calculation of molecular energies and entropies. We also show transition from a crystalline-like to a fluid DPPC bilayer by the CBC local-move MC method, as indicated by the electron density profile, head group orientation, area per lipid, and whole-lipid displacements. We discuss the potential of local-move MC methods in combination with molecular dynamics simulations, for example, for studying multi-component lipid membranes containing cholesterol.

  6. Membrane Compartment Occupied by Can1 (MCC and Eisosome Subdomains of the Fungal Plasma Membrane

    Directory of Open Access Journals (Sweden)

    James B. Konopka

    2011-12-01

    Full Text Available Studies on the budding yeast Saccharomyces cerevisiae have revealed that fungal plasma membranes are organized into different subdomains. One new domain termed MCC/eisosomes consists of stable punctate patches that are distinct from lipid rafts. The MCC/eisosome domains correspond to furrows in the plasma membrane that are about 300 nm long and 50 nm deep. The MCC portion includes integral membrane proteins, such as the tetraspanners Sur7 and Nce102. The adjacent eisosome includes proteins that are peripherally associated with the membrane, including the BAR domains proteins Pil1 and Lsp1 that are thought to promote membrane curvature. Genetic analysis of the MCC/eisosome components indicates these domains broadly affect overall plasma membrane organization. The mechanisms regulating the formation of MCC/eisosomes in model organisms will be reviewed as well as the role of these plasma membrane domains in fungal pathogenesis and response to antifungal drugs.

  7. Targeting Membrane Lipid a Potential Cancer Cure?

    Science.gov (United States)

    Tan, Loh Teng-Hern; Chan, Kok-Gan; Pusparajah, Priyia; Lee, Wai-Leng; Chuah, Lay-Hong; Khan, Tahir Mehmood; Lee, Learn-Han; Goh, Bey-Hing

    2017-01-01

    Cancer mortality and morbidity is projected to increase significantly over the next few decades. Current chemotherapeutic strategies have significant limitations, and there is great interest in seeking novel therapies which are capable of specifically targeting cancer cells. Given that fundamental differences exist between the cellular membranes of healthy cells and tumor cells, novel therapies based on targeting membrane lipids in cancer cells is a promising approach that deserves attention in the field of anticancer drug development. Phosphatidylethanolamine (PE), a lipid membrane component which exists only in the inner leaflet of cell membrane under normal circumstances, has increased surface representation on the outer membrane of tumor cells with disrupted membrane asymmetry. PE thus represents a potential chemotherapeutic target as the higher exposure of PE on the membrane surface of cancer cells. This feature as well as a high degree of expression of PE on endothelial cells in tumor vasculature, makes PE an attractive molecular target for future cancer interventions. There have already been several small molecules and membrane-active peptides identified which bind specifically to the PE molecules on the cancer cell membrane, subsequently inducing membrane disruption leading to cell lysis. This approach opens up a new front in the battle against cancer, and is of particular interest as it may be a strategy that may be prove effective against tumors that respond poorly to current chemotherapeutic agents. We aim to highlight the evidence suggesting that PE is a strong candidate to be explored as a potential molecular target for membrane targeted novel anticancer therapy. PMID:28167913

  8. Tethered bimolecular lipid membranes - A novel model membrane platform

    Energy Technology Data Exchange (ETDEWEB)

    Knoll, Wolfgang; Koeper, Ingo; Naumann, Renate; Sinner, Eva-Kathrin [Max Planck Institute for Polymer Research, Ackermannweg 10, 55128 Mainz (Germany)

    2008-10-01

    This contribution summarizes some of our efforts in designing, synthesizing, assembling, and characterizing functional tethered bimolecular lipid membranes (tBLMs) as a novel platform for biophysical studies of and with artificial membranes or for sensor development employing, e.g., membrane integral receptor proteins. Chemical coupling schemes based on thiol groups for Au substrates or silanes used in the case of oxide surfaces allow for the covalent and, hence, chemically and mechanically robust attachment of anchor lipids to the solid support, stabilizing the proximal layer of a tethered membrane on the transducer surface. Surface plasmon optics, the quartz crystal microbalance, fluorescence- and IR spectroscopies, and electrochemical techniques are used to characterize the build-up of these complex supramolecular interfacial architectures. We demonstrate, in particular, that bilayers with a specific electrical resistance of better than 10 M{omega} cm{sup 2} can be achieved routinely with this approach. The functionalization of the lipid membranes by the incorporation of peptides is demonstrated for the carrier valinomycin which shows in our tBLMs the expected discrimination by four orders of magnitude between the translocation of K{sup +}- and Na{sup +}-ions across the hydrophobic barrier. For the synthetic channel-forming peptide M2 the high electrical resistance of the bilayer with the correspondingly low background current allows for the recording of even single channel current fluctuations. From the many membrane proteins that we reconstituted so far we describe results obtained with the redox-protein cytochrome c oxidase. Here, we also use a genetically modified mutant with a His-tag at either the C- or the N-terminus for the oriented attachment of the protein via the NTA/Ni{sup 2+} approach. With this strategy, we not only can control the density of the immobilized functional units, we introduce a completely new and alternative concept for the

  9. There Is No Simple Model of the Plasma Membrane Organization

    Science.gov (United States)

    Bernardino de la Serna, Jorge; Schütz, Gerhard J.; Eggeling, Christian; Cebecauer, Marek

    2016-01-01

    Ever since technologies enabled the characterization of eukaryotic plasma membranes, heterogeneities in the distributions of its constituents were observed. Over the years this led to the proposal of various models describing the plasma membrane organization such as lipid shells, picket-and-fences, lipid rafts, or protein islands, as addressed in numerous publications and reviews. Instead of emphasizing on one model we in this review give a brief overview over current models and highlight how current experimental work in one or the other way do not support the existence of a single overarching model. Instead, we highlight the vast variety of membrane properties and components, their influences and impacts. We believe that highlighting such controversial discoveries will stimulate unbiased research on plasma membrane organization and functionality, leading to a better understanding of this essential cellular structure. PMID:27747212

  10. There is no simple model of the plasma membrane organisation

    Directory of Open Access Journals (Sweden)

    Jorge Bernardino De La Serna

    2016-09-01

    Full Text Available Ever since technologies enabled the characterisation of eukaryotic plasma membranes, heterogeneities in the distributions of its constituents were observed. Over the years this led to the proposal of various models describing the plasma membrane organisation such as lipid shells, picket-and-fences, lipid rafts, or protein islands, as addressed in numerous publications and reviews. Instead of emphasising on one model we in this review give a brief overview over current models and highlight how current experimental work in one or the other way do not support the existence of a single overarching model. Instead, we highlight the vast variety of membrane properties and components, their influences and impacts. We believe that highlighting such controversial discoveries will stimulate unbiased research on plasma membrane organisation and functionality, leading to a better understanding of this essential cellular structure.

  11. Methyl-branched lipids promote the membrane adsorption of α-synuclein by enhancing shallow lipid-packing defects.

    Science.gov (United States)

    Garten, Matthias; Prévost, Coline; Cadart, Clotilde; Gautier, Romain; Bousset, Luc; Melki, Ronald; Bassereau, Patricia; Vanni, Stefano

    2015-06-28

    Alpha-synuclein (AS) is a synaptic protein that is directly involved in Parkinson's disease due to its tendency to form protein aggregates. Since AS aggregation can be dependent on the interactions between the protein and the cell plasma membrane, elucidating the membrane binding properties of AS is of crucial importance to establish the molecular basis of AS aggregation into toxic fibrils. Using a combination of in vitro reconstitution experiments based on Giant Unilamellar Vesicles (GUVs), confocal microscopy and all-atom molecular dynamics simulations, we have investigated the membrane binding properties of AS, with a focus on the relative contribution of hydrophobic versus electrostatic interactions. In contrast with previous observations, we did not observe any binding of AS to membranes containing the ganglioside GM1, even at relatively high GM1 content. AS, on the other hand, showed a stronger affinity for neutral flat membranes consisting of methyl-branched lipids. To rationalize these results, we used all-atom molecular dynamics simulations to investigate the influence of methyl-branched lipids on interfacial membrane properties. We found that methyl-branched lipids promote the membrane adsorption of AS by creating shallow lipid-packing defects to a larger extent than polyunsaturated and monounsaturated lipids. Our findings suggest that methyl-branched lipids may constitute a remarkably adhesive substrate for peripheral proteins that adsorb on membranes via hydrophobic insertions.

  12. Lipid Directed Intrinsic Membrane Protein Segregation

    DEFF Research Database (Denmark)

    Hansen, Jesper S.; Thompson, James R.; Helix Nielsen, Claus;

    2013-01-01

    We demonstrate a new approach for direct reconstitution of membrane proteins during giant vesicle formation. We show that it is straightforward to create a tissue-like giant vesicle film swelled with membrane protein using aquaporin SoPIP2;1 as an illustration. These vesicles can also be easily h...... harvested for individual study. By controlling the lipid composition we are able to direct the aquaporin into specific immiscible liquid domains in giant vesicles. The oligomeric α-helical protein cosegregates with the cholesterol-poor domains in phase separating ternary mixtures....

  13. Membrane lipids and the origin of life

    Science.gov (United States)

    Oro, J.; Holzer, G.; Rao, M.; Tornabene, T. G.

    1981-01-01

    The current state of knowledge regarding the development of biological systems is briefly reviewed. At a crucial stage concerning the evolution of such systems, the mechanisms leading to more complex structures must have evolved within the confines of a protected microenvironment, similar to those provided by the contemporary cell membranes. The major components found normally in biomembranes are phospholipids. The structure of the biomembrane is examined, and attention is given to questions concerning the availability of the structural components which are necessary in the formation of primitive lipid membranes. Two approaches regarding the study of protomembranes are discussed. The probability of obtaining ether lipids under prebiotic conditions is considered, taking into account the formation of cyclic and acyclic isoprenoids by the irradiation of isoprene with UV.

  14. Laurdan monitors different lipids content in eukaryotic membrane during embryonic neural development.

    Science.gov (United States)

    Bonaventura, Gabriele; Barcellona, Maria Luisa; Golfetto, Ottavia; Nourse, Jamison L; Flanagan, Lisa A; Gratton, Enrico

    2014-11-01

    We describe a method based on fluorescence-lifetime imaging microscopy (FLIM) to assess the fluidity of various membranes in neuronal cells at different stages of development [day 12 (E12) and day 16 (E16) of gestation]. For the FLIM measurements, we use the Laurdan probe which is commonly used to assess membrane water penetration in model and in biological membranes using spectral information. Using the FLIM approach, we build a fluidity scale based on calibration with model systems of different lipid compositions. In neuronal cells, we found a marked difference in fluidity between the internal membranes and the plasma membrane, being the plasma membrane the less fluid. However, we found no significant differences between the two cell groups, E12 and E16. Comparison with NIH3T3 cells shows that the plasma membranes of E12 and E16 cells are significantly more fluid than the plasma membrane of the cancer cells.

  15. Atomic force microscopy of model lipid membranes.

    Science.gov (United States)

    Morandat, Sandrine; Azouzi, Slim; Beauvais, Estelle; Mastouri, Amira; El Kirat, Karim

    2013-02-01

    Supported lipid bilayers (SLBs) are biomimetic model systems that are now widely used to address the biophysical and biochemical properties of biological membranes. Two main methods are usually employed to form SLBs: the transfer of two successive monolayers by Langmuir-Blodgett or Langmuir-Schaefer techniques, and the fusion of preformed lipid vesicles. The transfer of lipid films on flat solid substrates offers the possibility to apply a wide range of surface analytical techniques that are very sensitive. Among them, atomic force microscopy (AFM) has opened new opportunities for determining the nanoscale organization of SLBs under physiological conditions. In this review, we first focus on the different protocols generally employed to prepare SLBs. Then, we describe AFM studies on the nanoscale lateral organization and mechanical properties of SLBs. Lastly, we survey recent developments in the AFM monitoring of bilayer alteration, remodeling, or digestion, by incubation with exogenous agents such as drugs, proteins, peptides, and nanoparticles.

  16. Importance of the hexagonal lipid phase in biological membrane organisation

    Directory of Open Access Journals (Sweden)

    Juliette eJouhet

    2013-12-01

    Full Text Available Abstract:Domains are present in every natural membrane. They are characterised by a distinctive protein and/or lipid composition. Their size is highly variable from the nano- to the micrometer scale. The domains confer specific properties to the membrane leading to original structure and function. The determinants leading to domain organisation are therefore important but remain obscure. This review presents how the ability of lipids to organize into hexagonal II or lamellar phases can promote particular local structures within membranes. Since biological membranes are composed of a mixture of lipids, each with distinctive biophysical properties, lateral and transversal sorting of lipids can promote creation of domains inside the membrane through local modulation of the lipid phase. Lipid biophysical properties have been characterized for long based on in vitro analyses using non-natural lipid molecules; their re-examinations using natural lipids might open interesting perspectives on membrane architecture occurring in vivo in various cellular and physiological contexts.

  17. Topography and functional information of plasma membrane

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    By using atomic force microscope (AFM), the topography and function of the plasmalemma surface of the isolated protoplasts from winter wheat mesophyll cells were observed, and compared with dead protoplasts induced by dehydrating stress. The observational results revealed that the plasma membrane of living protoplasts was in a state of polarization. Lipid layers of different cells and membrane areas exhibited distinct active states. The surfaces of plasma membranes were unequal, and were characterized of regionalisation. In addition, lattice structures were visualized in some regions of the membrane surface. These typical structures were assumed to be lipid molecular complexes, which were measured to be 15.8±0.09 nm in diameter and 1.9±0.3 nm in height. Both two-dimensional and three-dimensional imaging showed that the plasmalemma surfaces of winter wheat protoplasts were covered with numerous protruding particles. In order to determine the chemical nature of the protruding particles, living protoplasts were treated by proteolytic enzyme. Under the effect of enzyme, large particles became relatively looser, resulting that their width was increased and their height decreased. The results demonstrated that these particles were likely to be of protein nature. These protein particles at plasmalemma surface were different in size and unequal in distribution. The diameter of large protein particles ranged from 200 to 440 nm, with a central micropore, and the apparent height of them was found to vary from 12 to 40 nm. The diameter of mid-sized protein particles was between 40―60 nm, and a range of 1.8―5 nm was given for the apparent height of them. As for small protein particles, obtained values were 12―40 nm for their diameter and 0.7―2.2 nm for height. Some invaginated pits were also observed at the plasma membrane. They were formed by the endocytosis of protoplast. Distribution density of them at plasmalemma was about 16 pits per 15 μm2. According to their

  18. Topography and functional information of plasma membrane

    Institute of Scientific and Technical Information of China (English)

    SUN DeLan; CHEN JianMin; SONG YanMei; ZHU ChuanFeng; PAN GeBo; WAN LiJun

    2008-01-01

    By using atomic force microscope (AFM), the topography and function of the plasmalemma surface of the isolated protoplasta from winter wheat mesophyll cells were observed, and compared with dead protoplssts induced by dehydrating stress. The observational results revealed that the plasma membrane of living protoplasta was in a state of polarization. Lipid layers of different cells and membrane areas exhibited distinct active states. The surfaces of plasma membranes were unequal, and were characterized of regionalisation. In addition, lattice structures were visualized in some regions of the membrane surface. These typical structures were assumed to be lipid molecular complexes, which were measured to be 15.8±0.09 nm in diameter and 1.9±0.3 nm in height. Both two-dimensional and three-dimensional imaging showed that the plasmalemma surfaces of winter wheat protoplasta were covered with numerous protruding particles. In order to determine the chemical nature of the protruding particles, living protoplasts were treated by proteolytic enzyme. Under the effect of enzyme, large particles became relatively looser, resulting that their width was increased and their height decreased.The results demonstrated that these particles were likely to be of protein nature. These protein particles at plasmalemma surface were different in size and unequal in distribution. The diameter of large protein particles ranged from 200 to 440 nm, with a central micropore, and the apparent height of them was found to vary from 12 to 40 nm. The diameter of mid-sized protein particles was between 40-60 nm,and a range of 1.8-5 nm was given for the apparent height of them. As for small protein particles, obtained values were 12-40 nm for their diameter and 0.7-2.2 nm for height. Some invaginated pits were also observed at the plasma membrane. They were formed by the endocytosis of protoplsst. Distributlon density of them at plasmalemma was about 16 pits per 15 μm2. According to their size, we

  19. Homeoviscous adaptation and the regulation of membrane lipids

    DEFF Research Database (Denmark)

    Ernst, Robert; Ejsing, Christer S; Antonny, Bruno

    2016-01-01

    Biological membranes are complex and dynamic assemblies of lipids and proteins. Poikilothermic organisms including bacteria, fungi, reptiles, and fish do not control their body temperature and must adapt their membrane lipid composition in order to maintain membrane fluidity in the cold. This ada......Biological membranes are complex and dynamic assemblies of lipids and proteins. Poikilothermic organisms including bacteria, fungi, reptiles, and fish do not control their body temperature and must adapt their membrane lipid composition in order to maintain membrane fluidity in the cold...

  20. Interaction measurement of particles bound to a lipid membrane

    Science.gov (United States)

    Sarfati, Raphael; Dufresne, Eric

    2015-03-01

    The local shape and dynamics of the plasma membrane play important roles in many cellular processes. Local membrane deformations are often mediated by the adsorption of proteins (notably from the BAR family), and their subsequent self-assembly. The emerging hypothesis is that self-assembly arises from long-range interactions of individual proteins through the membrane's deformation field. We study these interactions in a model system of micron-sized colloidal particles adsorbed onto a lipid bilayer. We use fluorescent microscopy, optical tweezers and particle tracking to measure dissipative and conservative forces as a function of the separation between the particles. We find that particles are driven together with forces of order 100 fN and remain bound in a potential well with a stiffness of order 100 fN/micron.

  1. Interaction of Hematoporphyrin with Lipid Membranes

    DEFF Research Database (Denmark)

    Stepniewski, M.; Kepczynski, M.; Jamroz, D.

    2012-01-01

    Natural or synthetic porphyrins are being used as photosensitizers in photodiagnosis (PD) and photodynamic therapy (PDT) of malignancies and some other diseases. Understanding the interactions between porphyrins and cell membranes is therefore important to rationalize the uptake of photosensitizers....... The dye molecules were found to reside in the phospholipid headgroup area close to the carbonyl groups of the POPC acyl chains. Their orientations were dependent on the protonation state of two propionic groups. Hp(2-) was found to have a lower affinity to enter the membrane than the neutral form....... The dianions, being in the aqueous phase, formed stable dimers with a strictly determined geometry. Our results fully supported the experimental data and provide a more detailed molecular-level description of the interactions of photosensitizers with lipid membranes....

  2. Lipid Gymnastics: Tethers and Fingers in membrane

    Science.gov (United States)

    Tayebi, Lobat; Miller, Gregory; Parikh, Atul

    2009-03-01

    A significant body of evidence now links local mesoscopic structure (e.g., shape and composition) of the cell membrane with its function; the mechanisms by which cellular membranes adopt the specific shapes remain poorly understood. Among all the different structures adopted by cellular membranes, the tubular shape is one of the most surprising one. While their formation is typically attributed to the reorganization of membrane cytoskeleton, many exceptions exist. We report the instantaneous formation of tubular membrane mesophases following the hydration under specific thermal conditions. The shapes emerge in a bimodal way where we have two distinct diameter ranges for tubes, ˜20μm and ˜1μm, namely fat fingers and narrow tethers. We study the roughening of hydrated drops of 3 lipids in 3 different spontaneous curvatures at various temp. and ionic strength to figure out the dominant effect in selection of tethers and fingers. Dynamics of the tubes are of particular interest where we observe four distinct steps of birth, coiling, uncoiling and retraction with different lifetime on different thermal condition. These dynamics appear to reflect interplay between membrane elasticity, surface adhesion, and thermal or hydrodynamic gradient.

  3. Ras diffusion is sensitive to plasma membrane viscosity.

    Science.gov (United States)

    Goodwin, J Shawn; Drake, Kimberly R; Remmert, Catha L; Kenworthy, Anne K

    2005-08-01

    The cell surface contains a variety of barriers and obstacles that slow the lateral diffusion of glycosylphosphatidylinositol (GPI)-anchored and transmembrane proteins below the theoretical limit imposed by membrane viscosity. How the diffusion of proteins residing exclusively on the inner leaflet of the plasma membrane is regulated has been largely unexplored. We show here that the diffusion of the small GTPase Ras is sensitive to the viscosity of the plasma membrane. Using confocal fluorescence recovery after photobleaching, we examined the diffusion of green fluorescent protein (GFP)-tagged HRas, NRas, and KRas in COS-7 cells loaded with or depleted of cholesterol, a well-known modulator of membrane bilayer viscosity. In cells loaded with excess cholesterol, the diffusional mobilities of GFP-HRas, GFP-NRas, and GFP-KRas were significantly reduced, paralleling the behavior of the viscosity-sensitive lipid probes DiIC(16) and DiIC(18). However, the effects of cholesterol depletion on protein and lipid diffusion in cell membranes were highly dependent on the depletion method used. Cholesterol depletion with methyl-beta-cyclodextrin slowed Ras diffusion by a viscosity-independent mechanism, whereas overnight cholesterol depletion slightly increased both protein and lipid diffusion. The ability of Ras to sense membrane viscosity may represent a general feature of proteins residing on the cytoplasmic face of the plasma membrane.

  4. Plasma lipid concentrations for some Brazilian lizards.

    Science.gov (United States)

    Gillett, M P; Lima, V L; Costa, J C; Sibrian, A M

    1979-01-01

    1. Plasma concentrations of cholesterol, cholesteryl esters, phospholipids and triglycerides were determined for ten species of Brazilian lizards, Iguana iguana, Tropidurus torquatos and T. semitaeniatus (Iguanidae), Tupinambis teguixin, Ameiva ameiva and Cnemidophorus ocellifer (Teiidae), Mabuya maculata (Scincidae), Hemidactylus mabouia (Gekkonidae), Amphisbaenia vermicularis and Leposternon polystegum (Amphisbaenidae). 2. Considerable inter- and intra-species variations in plasma lipid concentrations were observed. 3. The percentage of total cholesterol esterified and the individual phospholipid composition of plasma were relatively constant for each species. 4. Over 60% of the cholesteryl esters present in plasma from three species each of iguanid and teiid lizards were polyenoic.

  5. Lipid corralling and poloxamer squeeze-out in membranes

    DEFF Research Database (Denmark)

    Wu, G.H.; Majewski, J.; Ege, C.;

    2004-01-01

    Using x-ray scattering measurements we have quantitatively determined the effect of poloxamer 188 (P188), a polymer known to seal damaged membranes, on the structure of lipid monolayers. P188 selectively inserts into low lipid-density regions of the membrane and "corrals" lipid molecules to pack...... tightly, leading to unexpected Bragg peaks at low nominal lipid density and inducing lipid/poloxamer phase separation. At tighter lipid packing, the once inserted P188 is squeezed out, allowing the poloxamer to gracefully exit when the membrane integrity is restored....

  6. Interaction of antimicrobial peptides with lipid membranes

    Energy Technology Data Exchange (ETDEWEB)

    Hanulova, Maria

    2008-12-15

    This study aims to investigate the difference in the interaction of antimicrobial peptides with two classes of zwitterionic peptides, phosphatidylethanolamines (PE) and phosphatidylcholines (PC). Further experiments were performed on model membranes prepared from specific bacterial lipids, lipopolysaccharides (LPS) isolated from Salmonella minnesota. The structure of the lipid-peptide aqueous dispersions was studied by small-and wide-angle X-ray diffraction during heating and cooling from 5 to 85 C. The lipids and peptides were mixed at lipid-to-peptide ratios 10-10000 (POPE and POPC) or 2-50 (LPS). All experiments were performed at synchrotron soft condensed matter beamline A2 in Hasylab at Desy in Hamburg, Germany. The phases were identified and the lattice parameters were calculated. Alamethicin and melittin interact in similar ways with the lipids. Pure POPC forms only lamellar phases. POPE forms lamellar phases at low temperatures that upon heating transform into a highly curved inverse hexagonal phase. Insertion of the peptide induced inverse bicontinuous cubic phases which are an ideal compromise between the curvature stress and the packing frustration. Melittin usually induced a mixture of two cubic phases, Im3m and Pn3m, with a ratio of lattice parameters close to 1.279, related to the underlying minimal surfaces. They formed during the lamellar to hexagonal phase transition and persisted during cooling till the onset of the gel phase. The phases formed at different lipid-to-peptide ratios had very similar lattice parameters. Epitaxial relationships existed between coexisting cubic phases and hexagonal or lamellar phases due to confinement of all phases to an onion vesicle, a vesicle with several layers consisting of different lipid phases. Alamethicin induced the same cubic phases, although their formation and lattice parameters were dependent on the peptide concentration. The cubic phases formed during heating from the lamellar phase and their onset

  7. Randomly organized lipids and marginally stable proteins: a coupling of weak interactions to optimize membrane signaling.

    Science.gov (United States)

    Rice, Anne M; Mahling, Ryan; Fealey, Michael E; Rannikko, Anika; Dunleavy, Katie; Hendrickson, Troy; Lohese, K Jean; Kruggel, Spencer; Heiling, Hillary; Harren, Daniel; Sutton, R Bryan; Pastor, John; Hinderliter, Anne

    2014-09-01

    Eukaryotic lipids in a bilayer are dominated by weak cooperative interactions. These interactions impart highly dynamic and pliable properties to the membrane. C2 domain-containing proteins in the membrane also interact weakly and cooperatively giving rise to a high degree of conformational plasticity. We propose that this feature of weak energetics and plasticity shared by lipids and C2 domain-containing proteins enhance a cell's ability to transduce information across the membrane. We explored this hypothesis using information theory to assess the information storage capacity of model and mast cell membranes, as well as differential scanning calorimetry, carboxyfluorescein release assays, and tryptophan fluorescence to assess protein and membrane stability. The distribution of lipids in mast cell membranes encoded 5.6-5.8bits of information. More information resided in the acyl chains than the head groups and in the inner leaflet of the plasma membrane than the outer leaflet. When the lipid composition and information content of model membranes were varied, the associated C2 domains underwent large changes in stability and denaturation profile. The C2 domain-containing proteins are therefore acutely sensitive to the composition and information content of their associated lipids. Together, these findings suggest that the maximum flow of signaling information through the membrane and into the cell is optimized by the cooperation of near-random distributions of membrane lipids and proteins. This article is part of a Special Issue entitled: Interfacially Active Peptides and Proteins. Guest Editors: William C. Wimley and Kalina Hristova.

  8. Concise theory of chiral lipid membranes

    CERN Document Server

    Tu, Z C

    2007-01-01

    A theory of chiral lipid membranes is proposed on the basis of a concise free energy density which includes the contributions of the bending and the surface tension of membranes, as well as the chirality and orientational variation of tilting molecules. This theory is consistent with the previous experiments [J.M. Schnur \\textit{et al.}, Science \\textbf{264}, 945 (1994); M.S. Spector \\textit{et al.}, Langmuir \\textbf{14}, 3493 (1998); Y. Zhao, \\textit{et al.}, Proc. Natl. Acad. Sci. USA \\textbf{102}, 7438 (2005)] on self-assembled chiral lipid membranes of DC$_{8,9}$PC. A torus with the ratio between its two generated radii larger than $\\sqrt{2}$ is predicted from the Euler-Lagrange equations. It is found that tubules with helically modulated tilting state are not admitted by the Euler-Lagrange equations, and that they are less energetically favorable than helical ripples in tubules. The pitch angles of helical ripples are theoretically estimated to be about 0$^\\circ$ and 35$^\\circ$, which are close to the mo...

  9. The mystery of membrane organization: composition, regulation and roles of lipid rafts.

    Science.gov (United States)

    Sezgin, Erdinc; Levental, Ilya; Mayor, Satyajit; Eggeling, Christian

    2017-06-01

    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.

  10. Recycling from endosomes to the plasma membrane

    NARCIS (Netherlands)

    Dam, E.M. van

    2001-01-01

    Summary V Chapter?Summary Many membrane proteins are, after endocytic uptake, efficiently recycled back to the plasma membrane. The aim of the studies presented in this thesis was to determine pathways and molecular mechanisms that are involved in recycling. Plasma membrane-derived clathrin-coated v

  11. Recycling from endosomes to the plasma membrane

    NARCIS (Netherlands)

    Dam, E.M. van

    2001-01-01

    Summary V Chapter?Summary Many membrane proteins are, after endocytic uptake, efficiently recycled back to the plasma membrane. The aim of the studies presented in this thesis was to determine pathways and molecular mechanisms that are involved in recycling. Plasma membrane-derived clathrin-coated

  12. Lipid traffic: the ABC of transbilayer movement

    NARCIS (Netherlands)

    Raggers, R.J.; Pomorski, T.; Holthuis, J.C.M.; Kälin, N.; van Meer, G.

    2000-01-01

    Membrane lipids do not spontaneously exchange between the two leaflets of lipid bilayers because the polar headgroups cannot cross the hydrophobic membrane interior. Cellular membranes, notably eukaryotic plasma membranes, are equipped with special proteins that actively translocate lipids from one

  13. The molecular face of lipid rafts in model membranes

    NARCIS (Netherlands)

    Risselada, H. Jelger; Marrink, Siewert J.

    2008-01-01

    Cell membranes contain a large number of different lipid species. Such a multicomponent mixture exhibits a complex phase behavior with regions of structural and compositional heterogeneity. Especially domains formed in ternary mixtures, composed of saturated and unsaturated lipids together with

  14. The lipid organisation of the cell membrane

    Directory of Open Access Journals (Sweden)

    Ladha, S.

    2000-04-01

    Full Text Available Lipids and proteins in biological membranes are arranged in a mosaic of domains in the membrane. These domains represent small-scale heterogeneities in composition, shape and fluidity within the plane of the membrane, over the range of hundreds of nanometers to a few micrometers. They arise from the complex interactions of the heterogeneous mixtures of phospholipids, sterols, and proteins that make up all biological membranes.Los lípidos y las proteínas en las membranas biológicas están dispuestos en un mosaico de campos en la membrana. Estos campos representan heterogeneidades a pequeña escala en la composición, forma y fluidez dentro del plano de la membrana, en un rango que va de los cientos de nanómetros a los pocos micrómetros. Estos campos se originan de las complejas interacciones de las mezclas heterogéneas de fosfolípidos, esteroles y proteínas de las que están hechas todas y cada una de las membranas biológicas.

  15. Relation between plasma and brain lipids

    DEFF Research Database (Denmark)

    Wellington, Cheryl L; Frikke-Schmidt, Ruth

    2016-01-01

    : Plasma levels of traditional lipids and lipoproteins are not consistently associated with risk of dementia even though low plasma levels of apolipoprotein E, through unknown mechanisms, robustly predict future dementia. Experimental evidence suggests neuroprotective roles of several brain...... and cerebrospinal fluid apolipoproteins. Whether plasma levels of apolipoprotein E, or any other apolipoprotein with possible central nervous system and/or blood-brain barrier functions (apolipoproteins J, A-I, A-II, A-IV, D, C-I, and C-III) may become accessible biomarker components that improve risk prediction...

  16. Variations in plasma lipids and lipoproteins among cardiovascular ...

    African Journals Online (AJOL)

    Variations in plasma lipids and lipoproteins among cardiovascular disease patients in ... This study was designed to assess the changes in plasma lipids and lipoproteins, in particular highdensity lipoprotein (HDLC) in patients ... Article Metrics.

  17. Lipid-lipid and lipid-drug interactions in biological membranes

    Science.gov (United States)

    Martynowycz, Michael W.

    Interactions between lipids and drug molecules in biological membranes help govern proper biological function in organisms. The mechanisms responsible for hydrophobic drug permeation remain elusive. Many small molecule drugs are hydrophobic. These drugs inhibit proteins in the cellular interior. The rise of antibiotic resistance in bacteria is thought to be caused by mutations in protein structure, changing drug kinetics to favor growth. However, small molecule drugs have been shown to have different mechanisms depending in the structure of the lipid membrane of the target cell. Biological membranes are investigated using Langmuir monolayers at the air-liquid interface. These offer the highest level of control in the mimetic system and allow them to be investigated using complementary techniques. Langmuir isotherms and insertion assays are used to determine the area occupied by each lipid in the membrane and the change in area caused by the introduction of a drug molecule, respectively. Specular X-ray reflectivity is used to determine the electron density of the monolayer, and grazing incidence X-ray diffraction is used to determine the in-plane order of the monolayer. These methods determine the affinity of the drug and the mechanism of action. Studies are presented on hydrophobic drugs with mammalian membrane mimics using warfarin along with modified analogues, called superwarfarins. Data shows that toxicity of these modified drugs are modulated by the membrane cholesterol content in cells; explaining several previously unexplained effects of the drugs. Membrane mimics of bacteria are investigated along with their interactions with a hydrophobic antibiotic, novobiocin. Data suggests that permeation of the drug is mediated by modifications to the membrane lipids, and completely ceases translocation under certain circumstances. Circumventing deficiencies in small, hydrophobic drugs is approached by using biologically mimetic oligomers. Peptoids, mimetic of host

  18. Evidence for the presence of lipid-free monomolecular apolipoprotein A-1 in plasma[S

    Science.gov (United States)

    Miyazaki, Osamu; Ogihara, Jun; Fukamachi, Isamu; Kasumi, Takafumi

    2014-01-01

    The first step in reverse cholesterol transport is a process by which lipid-free or lipid-poor apoA-1 removes cholesterol from cells through the action of ATP binding cassette transporter A1 at the plasma membrane. However the structure and composition of lipid-free or -poor apoA-1 in plasma remains obscure. We previously obtained a monoclonal antibody (MAb) that specifically recognizes apoA-1 in preβ1-HDL, the smallest apoA-1-containing particle in plasma, which we used to establish a preβ1-HDL ELISA. Here, we purified preβ1-HDL from fresh normal plasma using said antibody, and analyzed the composition and structure. ApoA-1 was detected, but neither phospholipid nor cholesterol were detected in the purified preβ1-HDL. Only globular, not discoidal, particles were observed by electron microscopy. In nondenaturing PAGE, no difference in the mobility was observed between the purified preβ1-HDL and original plasma preβ1-HDL, or between the preβ1-HDL and lipid-free apoA-1 prepared by delipidating HDL. In sandwich ELISA using two anti-preβ1-HDL MAbs, reactivity with intact plasma preβ1-HDL was observed in ELISA using two MAbs with distinct epitopes but no reactivity was observed in ELISA using a single MAb, and the same phenomenon was observed with monomolecular lipid-free apoA-1. These results suggest that plasma preβ1-HDL is lipid-free monomolecular apoA-1. PMID:24304668

  19. Self assembly of HIV-1 Gag protein on lipid membranes generates PI(4,5)P2/Cholesterol nanoclusters

    Science.gov (United States)

    Yandrapalli, Naresh; Lubart, Quentin; Tanwar, Hanumant S.; Picart, Catherine; Mak, Johnson; Muriaux, Delphine; Favard, Cyril

    2016-01-01

    The self-assembly of HIV-1 Gag polyprotein at the inner leaflet of the cell host plasma membrane is the key orchestrator of virus assembly. The binding between Gag and the plasma membrane is mediated by specific interaction of the Gag matrix domain and the PI(4,5)P2 lipid (PIP2). It is unknown whether this interaction could lead to local reorganization of the plasma membrane lipids. In this study, using model membranes, we examined the ability of Gag to segregate specific lipids upon self-assembly. We show for the first time that Gag self-assembly is responsible for the formation of PIP2 lipid nanoclusters, enriched in cholesterol but not in sphingomyelin. We also show that Gag mainly partition into liquid-disordered domains of these lipid membranes. Our work strongly suggests that, instead of targeting pre-existing plasma membrane lipid domains, Gag is more prone to generate PIP2/Cholesterol lipid nanodomains at the inner leaflet of the plasma membrane during early events of virus assembly. PMID:28008947

  20. Membrane Binding of HIV-1 Matrix Protein: Dependence on Bilayer Composition and Protein Lipidation

    Science.gov (United States)

    Barros, Marilia; Nanda, Hirsh

    2016-01-01

    ABSTRACT By assembling in a protein lattice on the host's plasma membrane, the retroviral Gag polyprotein triggers formation of the viral protein/membrane shell. The MA domain of Gag employs multiple signals—electrostatic, hydrophobic, and lipid-specific—to bring the protein to the plasma membrane, thereby complementing protein-protein interactions, located in full-length Gag, in lattice formation. We report the interaction of myristoylated and unmyristoylated HIV-1 Gag MA domains with bilayers composed of purified lipid components to dissect these complex membrane signals and quantify their contributions to the overall interaction. Surface plasmon resonance on well-defined planar membrane models is used to quantify binding affinities and amounts of protein and yields free binding energy contributions, ΔG, of the various signals. Charge-charge interactions in the absence of the phosphatidylinositide PI(4,5)P2 attract the protein to acidic membrane surfaces, and myristoylation increases the affinity by a factor of 10; thus, our data do not provide evidence for a PI(4,5)P2 trigger of myristate exposure. Lipid-specific interactions with PI(4,5)P2, the major signal lipid in the inner plasma membrane, increase membrane attraction at a level similar to that of protein lipidation. While cholesterol does not directly engage in interactions, it augments protein affinity strongly by facilitating efficient myristate insertion and PI(4,5)P2 binding. We thus observe that the isolated MA protein, in the absence of protein-protein interaction conferred by the full-length Gag, binds the membrane with submicromolar affinities. IMPORTANCE Like other retroviral species, the Gag polyprotein of HIV-1 contains three major domains: the N-terminal, myristoylated MA domain that targets the protein to the plasma membrane of the host; a central capsid-forming domain; and the C-terminal, genome-binding nucleocapsid domain. These domains act in concert to condense Gag into a membrane

  1. Biophysical Changes of Lipid Membranes in the Presence of Ethanol at Varying Concentrations.

    Science.gov (United States)

    Konas, Ryan M; Daristotle, John L; Harbor, Ndubuisi B; Klauda, Jeffery B

    2015-10-15

    Ethanol is widely used as an additive to gasoline, and production of ethanol can come from single-celled organisms such as yeast. We systematically studied the influence of ethanol on common lipids found in yeast plasma membranes, specifically phosphatidylserine (PS), phosphatidylethanolamine (PE), and phosphatidylcholine (PC). Molecular dynamics simulations were used to probe changes to the biophysical properties of membranes with varying equilibrated bulk ethanol concentrations less than 25 mol %. The palmitoyl oleoyl (PO, 18:1/16:0) chain was used for all lipids, and a mixed bilayer of POPE/POPS (7:3 ratio) was also simulated. Ethanol was found to interact strongly with POPC, and thus its surface area per lipid, chain order, and electron density profiles differ the most from the neat bilayer. At 12 mol % ethanol in the bulk, ethanol penetrated into the hydrophobic core for all membranes studied, but POPC had the highest penetration. Although the anionic headgroup of POPS acted as a protectant for membrane structure compared to the zwitterionic lipids, this was not the case for the POPE/POPS mixture that showed more penetration of ethanol into the membrane than the single-component membranes. To fully characterize the impact of ethanol on yeast plasma membranes, our results suggest that experiments and simulations need to consider representative mixtures of lipids that exist in vivo.

  2. The Effect of Lidocaine · HCl on the Fluidity of Native and Model Membrane Lipid Bilayers.

    Science.gov (United States)

    Park, Jun-Seop; Jung, Tae-Sang; Noh, Yang-Ho; Kim, Woo-Sung; Park, Won-Ick; Kim, Young-Soo; Chung, In-Kyo; Sohn, Uy Dong; Bae, Soo-Kyung; Bae, Moon-Kyoung; Jang, Hye-Ock; Yun, Il

    2012-12-01

    The purpose of this study is to investigated the mechanism of pharmacological action of local anesthetic and provide the basic information about the development of new effective local anesthetics. Fluorescent probe techniques were used to evaluate the effect of lidocaine·HCl on the physical properties (transbilayer asymmetric lateral and rotational mobility, annular lipid fluidity and protein distribution) of synaptosomal plasma membrane vesicles (SPMV) isolated from bovine cerebral cortex, and liposomes of total lipids (SPMVTL) and phospholipids (SPMVPL) extracted from the SPMV. An experimental procedure was used based on selective quenching of 1,3-di(1-pyrenyl)propane (Py-3-Py) and 1,6-diphenyl-1,3,5-hexatriene (DPH) by trinitrophenyl groups, and radiationless energy transfer from the tryptophans of membrane proteins to Py-3-Py. Lidocaine·HCl increased the bulk lateral and rotational mobility of neuronal and model membrane lipid bilayes, and had a greater fluidizing effect on the inner monolayer than the outer monolayer. Lidocaine·HCl increased annular lipid fluidity in SPMV lipid bilayers. It also caused membrane proteins to cluster. The most important finding of this study is that there is far greater increase in annular lipid fluidity than that in lateral and rotational mobilities by lidocaine·HCl. Lidocaine·HCl alters the stereo or dynamics of the proteins in the lipid bilayers by combining with lipids, especially with the annular lipids. In conclusion, the present data suggest that lidocaine, in addition to its direct interaction with proteins, concurrently interacts with membrane lipids, fluidizing the membrane, and thus inducing conformational changes of proteins known to be intimately associated with membrane lipid.

  3. (poly)Phosphoinositide phosphorylation is a marker for plasma membrane in Friend erythroleukaemic cells

    NARCIS (Netherlands)

    Rawyler, A.J.; Roelofsen, B.; Wirtz, K.W.A.; Kamp, J.A.F. op den

    1982-01-01

    Upon subcellular fractionation of (murine) Friend erythroleukaemic cells (FELCs), purified plasma membranes were identified by their high enrichment in specific marker enzymes and typical plasma membrane lipids. When FELCs were incubated for short periods with 32Pi before cell fractionation, the

  4. (poly)Phosphoinositide phosphorylation is a marker for plasma membrane in Friend erythroleukaemic cells

    NARCIS (Netherlands)

    Rawyler, A.J.; Roelofsen, B.; Wirtz, K.W.A.; Kamp, J.A.F. op den

    1982-01-01

    Upon subcellular fractionation of (murine) Friend erythroleukaemic cells (FELCs), purified plasma membranes were identified by their high enrichment in specific marker enzymes and typical plasma membrane lipids. When FELCs were incubated for short periods with 32Pi before cell fractionation, the lip

  5. Modeling Yeast Organelle Membranes and How Lipid Diversity Influences Bilayer Properties.

    Science.gov (United States)

    Monje-Galvan, Viviana; Klauda, Jeffery B

    2015-11-17

    Membrane lipids are important for the health and proper function of cell membranes. We have improved computational membrane models for specific organelles in yeast Saccharomyces cerevisiae to study the effect of lipid diversity on membrane structure and dynamics. Previous molecular dynamics simulations were performed by Jo et al. [(2009) Biophys J. 97, 50-58] on yeast membrane models having six lipid types with compositions averaged between the endoplasmic reticulum (ER) and the plasma membrane (PM). We incorporated ergosterol, phosphatidic acid, phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, and phosphatidylinositol lipids in our models to better describe the unique composition of the PM, ER, and trans-Golgi network (TGN) bilayers of yeast. Our results describe membrane structure based on order parameters (SCD), electron density profiles (EDPs), and lipid packing. The average surface area per lipid decreased from 63.8 ± 0.4 Å(2) in the ER to 47.1 ± 0.3 Å(2) in the PM, while the compressibility modulus (KA) varied in the opposite direction. The high SCD values for the PM lipids indicated a more ordered bilayer core, while the corresponding lipids in the ER and TGN models had lower parameters by a factor of at least 0.7. The hydrophobic core thickness (2DC) as estimated from EDPs is the thickest for PM, which is in agreement with estimates of hydrophobic regions of transmembrane proteins from the Orientation of Proteins in Membranes database. Our results show the importance of lipid diversity and composition on a bilayer's structural and mechanical properties, which in turn influences interactions with the proteins and membrane-bound molecules.

  6. Electrophoretic separation method for membrane pore-forming proteins in multilayer lipid membranes.

    Science.gov (United States)

    Okamoto, Yukihiro; Tsujimoto, Yusuke; Umakoshi, Hiroshi

    2016-03-01

    In this paper, we report on a novel electrophoretic separation and analysis method for membrane pore-forming proteins in multilayer lipid membranes (MLMs) in order to overcome the problems related to current separation and analysis methods of membrane proteins, and to obtain a high-performance separation method on the basis of specific properties of the lipid membranes. We constructed MLMs, and subsequently characterized membrane pore-forming protein behavior in MLMs. Through the use of these MLMs, we were able to successfully separate and analyze membrane pore-forming proteins in MLMs. To the best of our knowledge, this research is the first example of membrane pore-forming protein separation in lipid membranes. Our method can be expected to be applied for the separation and analysis of other membrane proteins including intrinsic membrane proteins and to result in high-performance by utilizing the specific properties of lipid membranes.

  7. Impaired plasma lipid profiles in acute hepatitis

    Directory of Open Access Journals (Sweden)

    Wang Yongzhong

    2010-01-01

    Full Text Available Abstract The present study examined plasma lipid profiles in thirty patients suffered from acute viral hepatitis. Patients' blood samples were collected at both the debut and recovery of diseases. Thirty sex and age matched normal subjects were included as controls. Plasma total triglycerides (TG, total cholesterol, high density lipoprotein cholesterol (HDL-C, low density lipoprotein cholesterol (LDL-C, apolipoprotein AI (ApoAI, apolipoprotein B (ApoB, lipoprotein (a (Lp(a, blood coagulation status including prothrombin complex activity and activated partial tromboplastin time (APTT, and hepatic functions were determined by the automatic biochemical analytical instrument. It demonstrated that plasma levels of total cholesterol, HDL-C and apoAI were significantly lower in the patients at the acute phase of hepatitis than those in normal subjects, whereas plasma levels of TG and LDL-C were obviously higher in the patients than in normal subjects (P

  8. Critical point fluctuations in supported lipid membranes.

    Science.gov (United States)

    Connell, Simon D; Heath, George; Olmsted, Peter D; Kisil, Anastasia

    2013-01-01

    In this paper, we demonstrate that it is possible to observe many aspects of critical phenomena in supported lipid bilayers using atomic force microscopy (AFM) with the aid of stable and precise temperature control. The regions of criticality were determined by accurately measuring and calculating phase diagrams for the 2 phase L(d)-L(o) region, and tracking how it moves with temperature, then increasing the sampling density around the estimated critical regions. Compositional fluctuations were observed above the critical temperature (T(c)) and characterised using a spatial correlation function. From this analysis, the phase transition was found to be most closely described by the 2D Ising model, showing it is a critical transition. Below T(c) roughening of the domain boundaries occurred due to the reduction in line tension close to the critical point. Smaller scale density fluctuations were also detected just below T(c). At T(c), we believe we have observed fluctuations on length scales greater than 10 microm. The region of critically fluctuating 10-100 nm nanodomains has been found to extend a considerable distance above T(c) to temperatures within the biological range, and seem to be an ideal candidate for the actual structure of lipid rafts in cell membranes. Although evidence for this idea has recently emerged, this is the first direct evidence for nanoscale domains in the critical region.

  9. Linearly concatenated cyclobutane (ladderane) lipids form a dense bacterial membrane

    NARCIS (Netherlands)

    Sinninghe Damsté, J.S.; Strous, M.; Rijpstra, W.I.C.; Hopmans, E.C.; Geenevasen, J.A.J.; Duin, A.C.T. van; Niftrik, L.A.; Jetten, M.S.M.

    2002-01-01

    Lipid membranes are essential to the functioning of cells, enabling the existence of concentration gradients of ions and metabolites. Microbial membrane lipids can contain three-, five-, six- and even seven-membered aliphatic rings, but four-membered aliphatic cyclobutane rings have never been obser

  10. Differential effect of plant lipids on membrane organization: specificities of phytosphingolipids and phytosterols.

    Science.gov (United States)

    Grosjean, Kevin; Mongrand, Sébastien; Beney, Laurent; Simon-Plas, Françoise; Gerbeau-Pissot, Patricia

    2015-02-27

    The high diversity of the plant lipid mixture raises the question of their respective involvement in the definition of membrane organization. This is particularly the case for plant plasma membrane, which is enriched in specific lipids, such as free and conjugated forms of phytosterols and typical phytosphingolipids, such as glycosylinositolphosphoceramides. This question was here addressed extensively by characterizing the order level of membrane from vesicles prepared using various plant lipid mixtures and labeled with an environment-sensitive probe. Fluorescence spectroscopy experiments showed that among major phytosterols, campesterol exhibits a stronger ability than β-sitosterol and stigmasterol to order model membranes. Multispectral confocal microscopy, allowing spatial analysis of membrane organization, demonstrated accordingly the strong ability of campesterol to promote ordered domain formation and to organize their spatial distribution at the membrane surface. Conjugated sterol forms, alone and in synergy with free sterols, exhibit a striking ability to order membrane. Plant sphingolipids, particularly glycosylinositolphosphoceramides, enhanced the sterol-induced ordering effect, emphasizing the formation and increasing the size of sterol-dependent ordered domains. Altogether, our results support a differential involvement of free and conjugated phytosterols in the formation of ordered domains and suggest that the diversity of plant lipids, allowing various local combinations of lipid species, could be a major contributor to membrane organization in particular through the formation of sphingolipid-sterol interacting domains.

  11. Fluorescence interference contrast based approach to study real time interaction of melittin with plasma membranes

    Science.gov (United States)

    Gupta, Sharad; Gui, Dong; Zandi, Roya; Gill, Sarjeet; Mohideen, Umar

    2014-03-01

    Melittin is an anti-bacterial and hemolytic toxic peptide found in bee venom. Cell lysis behavior of peptides has been widely investigated, but the exact interaction mechanism of lytic peptides with lipid membranes and its constituents has not been understood completely. In this paper we study the melittin interaction with lipid plasma membranes in real time using non-invasive and non-contact fluorescence interference contrast microscopy (FLIC). Particularly the interaction of melittin with plasma membranes was studied in a controlled molecular environment, where these plasma membrane were composed of saturated lipid, 1,2-diphytanoyl-sn-glycero-3-phosphocholine (DPhPC) and unsaturated lipid, 1,2-dioleoyl-sn-glycero-3-phosphocholine(DOPC) with and without cholesterol. We found out that melittin starts to form nanometer size pores in the plasma membranes shortly after interacting with membranes. But the addition of cholesterol in plasma membrane slows down the pore formation process. Our results show that inclusion of cholesterol to the plasma membranes make them more resilient towards pore formation and lysis of membrane.

  12. Properties of Plasma Membrane from Pea Root Seedlings under Altered Gravity

    Science.gov (United States)

    Klymchuk, D.; Baranenko, V.; Vorobyova, T. V.; Kurylenko, I.; Chyzhykova, O.; Dubovoy, V.

    In this study, the properties of pea (Pisum sativum L.) plasma membrane were examined to determine how the membrane structure and functions are regulated in response to clinorotation (2 rev/min) conditions. Membrane preparations enriched by plasma membrane vesicles were obtained by aqueous two-phase partitioning from 6-day seedling roots. The specific characteristics of H^+-ATPase, lípid composition and peroxidation intensity as well as fluidity of lipid bilayer were analysed. ATP hydrolytic activity was inhibited by ortovanadate and was insensitive to aside and nitrate in sealed plasma membrane vesicles isolated from both clinorotated and control seedlings. Plasma membrane vesicles from clinorotated seedlings in comparison to controls were characterised by increase in the total lipid/protein ratio, ATP hydrolytic activity and intensifying of lipid peroxidation. Sitosterol and campesterol were the predominant free sterol species. Clinorotated seedlings contained a slightly higher level of unsaturated fatty acid than controls. Plasma membrane vesicles were labelled with pyrene and fluorescence originating from monomeric (I_M) molecules and excimeric (I_E) aggregates were measured. The calculated I_E/I_M values were higher in clinorotated seedlings compared with controls reflecting the reduction in membrane microviscosity. The involvement of the changes in plasma membrane lipid content and composition, fluidity and H^+-ATPase activity in response of pea seedlings to altered gravity is discussed.

  13. Properties of fiber cell plasma membranes isolated from the cortex and nucleus of the porcine eye lens.

    Science.gov (United States)

    Mainali, Laxman; Raguz, Marija; O'Brien, William J; Subczynski, Witold K

    2012-04-01

    The organization and physical properties of the lipid bilayer portion of intact cortical and nuclear fiber cell plasma membranes isolated from the eye lenses of two-year-old pigs were studied using electron paramagnetic resonance (EPR) spin-labeling. Membrane fluidity, hydrophobicity, and the oxygen transport parameter (OTP) were assessed from the EPR spectra of precisely positioned spin labels. Intact cortical and nuclear membranes, which include membrane proteins, were found to contain three distinct lipid environments. These lipid environments were termed the bulk lipid domain, boundary lipid domain, and trapped lipid domain (lipids in protein aggregates). The amount of boundary and trapped lipids was greater in intact nuclear membranes than in cortical membranes. The properties of intact membranes were compared with the organization and properties of lens lipid membranes made of the total lipid extracts from the lens cortex or nucleus. In cortical lens lipid membranes, only one homogenous environment was detected, which was designated as a bulk lipid domain (phospholipid bilayer saturated with cholesterol). Lens lipid membranes prepared from the lens nucleus possessed two domains, assigned as a bulk lipid domain and a cholesterol bilayer domain (CBD). In intact nuclear membranes, it was difficult to discriminate the CBD, which was clearly detected in nuclear lens lipid membranes, because the OTP measured in the CBD is the same as in the domain formed by trapped lipids. The two domains unique to intact membranes-namely, the domain formed by boundary lipids and the domain formed by trapped lipids-were most likely formed due to the presence of membrane proteins. It is concluded that formation of rigid and practically impermeable domains is enhanced in the lens nucleus, indicating changes in membrane composition that may help to maintain low oxygen concentration in this lens region.

  14. Counterion-mediated pattern formation in membranes containing anionic lipids.

    Science.gov (United States)

    Slochower, David R; Wang, Yu-Hsiu; Tourdot, Richard W; Radhakrishnan, Ravi; Janmey, Paul A

    2014-06-01

    Most lipid components of cell membranes are either neutral, like cholesterol, or zwitterionic, like phosphatidylcholine and sphingomyelin. Very few lipids, such as sphingosine, are cationic at physiological pH. These generally interact only transiently with the lipid bilayer, and their synthetic analogs are often designed to destabilize the membrane for drug or DNA delivery. However, anionic lipids are common in both eukaryotic and prokaryotic cell membranes. The net charge per anionic phospholipid ranges from -1 for the most abundant anionic lipids such as phosphatidylserine, to near -7 for phosphatidylinositol 3,4,5 trisphosphate, although the effective charge depends on many environmental factors. Anionic phospholipids and other negatively charged lipids such as lipopolysaccharides are not randomly distributed in the lipid bilayer, but are highly restricted to specific leaflets of the bilayer and to regions near transmembrane proteins or other organized structures within the plane of the membrane. This review highlights some recent evidence that counterions, in the form of monovalent or divalent metal ions, polyamines, or cationic protein domains, have a large influence on the lateral distribution of anionic lipids within the membrane, and that lateral demixing of anionic lipids has effects on membrane curvature and protein function that are important for biological control. Copyright © 2014. Published by Elsevier B.V.

  15. INTERACTION OF ALDEHYDES DERIVED FROM LIPID PEROXIDATION AND MEMBRANE PROTEINS.

    Directory of Open Access Journals (Sweden)

    Stefania ePizzimenti

    2013-09-01

    Full Text Available A great variety of compounds are formed during lipid peroxidation of polyunsaturated fatty acids of membrane phospholipids. Among them, bioactive aldehydes, such as 4-hydroxyalkenals, malondialdehyde (MDA and acrolein, have received particular attention since they have been considered as toxic messengers that can propagate and amplify oxidative injury. In the 4-hydroxyalkenal class, 4-hydroxy-2-nonenal (HNE is the most intensively studied aldehyde, in relation not only to its toxic function, but also to its physiological role. Indeed, HNE can be found at low concentrations in human tissues and plasma and participates in the control of biological processes, such as signal transduction, cell proliferation and differentiation. Moreover, at low doses, HNE exerts an anti-cancer effect, by inhibiting cell proliferation, angiogenesis, cell adhesion and by inducing differentiation and/or apoptosis in various tumor cell lines. It is very likely that a substantial fraction of the effects observed in cellular responses, induced by HNE and related aldehydes, be mediated by their interaction with proteins, resulting in the formation of covalent adducts or in the modulation of their expression and/or activity. In this review we focus on membrane proteins affected by lipid peroxidation-derived aldehydes, under physiological and pathological conditions.

  16. Computational studies of plasma lipoprotein lipids.

    Science.gov (United States)

    Pan, Lurong; Segrest, Jere P

    2016-10-01

    Plasma lipoproteins are macromolecular assemblies of proteins and lipids found in the blood. The lipid components of lipoproteins are amphipathic lipids such as phospholipids (PLs), and unesterified cholesterols (UCs) and hydrophobic lipids such as cholesteryl esters (CEs) and triglycerides (TGs). Since lipoproteins are soft matter supramolecular assemblies easily deformable by thermal fluctuations and they also exist in varying densities and protein/lipid components, a detailed understanding of their structure/function is experimentally difficult. Molecular dynamics (MD) simulation has emerged as a particularly promising way to explore the structure and dynamics of lipoproteins. The purpose of this review is to survey the current status of computational studies of the lipid components of the lipoproteins. Computational studies aim to explore three levels of complexity for the 3-dimensional structural dynamics of lipoproteins at various metabolic stages: (i) lipoprotein particles consist of protein with minimal lipid; (ii) lipoprotein particles consist of PL-rich discoidal bilayer-like lipid particles; (iii) mature circulating lipoprotein particles consist of CE-rich or TG-rich spheroidal lipid-droplet-like particles. Due to energy barriers involved in conversion between these species, other biomolecules also participate in lipoprotein biological assembly. For example: (i) lipid-poor apolipoprotein A-I (apoA-I) interacts with ATP-binding cassette transporter A1 (ABCA1) to produce nascent discoidal high density lipoprotein (dHDL) particles; (ii) lecithin-cholesterol acyltransferase (LCAT) mediates the conversion of UC to CE in dHDL, driving spheroidal HDL (sHDL) formation; (iii) transfer proteins, cholesterol ester transfer protein (CETP) and phospholipid transfer protein (PLTP), transfer both CE and TG and PL, respectively, between lipoprotein particles. Computational studies have the potential to explore different lipoprotein particles at each metabolic stage in

  17. Perforin rapidly induces plasma membrane phospholipid flip-flop.

    Directory of Open Access Journals (Sweden)

    Sunil S Metkar

    Full Text Available The cytotoxic cell granule secretory pathway is essential for host defense. This pathway is fundamentally a form of intracellular protein delivery where granule proteases (granzymes from cytotoxic lymphocytes are thought to diffuse through barrel stave pores generated in the plasma membrane of the target cell by the pore forming protein perforin (PFN and mediate apoptotic as well as additional biological effects. While recent electron microscopy and structural analyses indicate that recombinant PFN oligomerizes to form pores containing 20 monomers (20 nm when applied to liposomal membranes, these pores are not observed by propidium iodide uptake in target cells. Instead, concentrations of human PFN that encourage granzyme-mediated apoptosis are associated with pore structures that unexpectedly favor phosphatidylserine flip-flop measured by Annexin-V and Lactadherin. Efforts that reduce PFN mediated Ca influx in targets did not reduce Annexin-V reactivity. Antigen specific mouse CD8 cells initiate a similar rapid flip-flop in target cells. A lipid that augments plasma membrane curvature as well as cholesterol depletion in target cells enhance flip-flop. Annexin-V staining highly correlated with apoptosis after Granzyme B (GzmB treatment. We propose the structures that PFN oligomers form in the membrane bilayer may include arcs previously observed by electron microscopy and that these unusual structures represent an incomplete mixture of plasma membrane lipid and PFN oligomers that may act as a flexible gateway for GzmB to translocate across the bilayer to the cytosolic leaflet of target cells.

  18. A Coarse Grained Model for a Lipid Membrane with Physiological Composition and Leaflet Asymmetry.

    Directory of Open Access Journals (Sweden)

    Satyan Sharma

    Full Text Available The resemblance of lipid membrane models to physiological membranes determines how well molecular dynamics (MD simulations imitate the dynamic behavior of cell membranes and membrane proteins. Physiological lipid membranes are composed of multiple types of phospholipids, and the leaflet compositions are generally asymmetric. Here we describe an approach for self-assembly of a Coarse-Grained (CG membrane model with physiological composition and leaflet asymmetry using the MARTINI force field. An initial set-up of two boxes with different types of lipids according to the leaflet asymmetry of mammalian cell membranes stacked with 0.5 nm overlap, reliably resulted in the self-assembly of bilayer membranes with leaflet asymmetry resembling that of physiological mammalian cell membranes. Self-assembly in the presence of a fragment of the plasma membrane protein syntaxin 1A led to spontaneous specific positioning of phosphatidylionositol(4,5bisphosphate at a positively charged stretch of syntaxin consistent with experimental data. An analogous approach choosing an initial set-up with two concentric shells filled with different lipid types results in successful assembly of a spherical vesicle with asymmetric leaflet composition. Self-assembly of the vesicle in the presence of the synaptic vesicle protein synaptobrevin 2 revealed the correct position of the synaptobrevin transmembrane domain. This is the first CG MD method to form a membrane with physiological lipid composition as well as leaflet asymmetry by self-assembly and will enable unbiased studies of the incorporation and dynamics of membrane proteins in more realistic CG membrane models.

  19. Isolation and analysis of membrane lipids and lipid rafts in common carp (Cyprinus carpio L.).

    Science.gov (United States)

    Brogden, Graham; Propsting, Marcus; Adamek, Mikolaj; Naim, Hassan Y; Steinhagen, Dieter

    2014-03-01

    Cell membranes act as an interface between the interior of the cell and the exterior environment and facilitate a range of essential functions including cell signalling, cell structure, nutrient uptake and protection. It is composed of a lipid bilayer with integrated proteins, and the inner leaflet of the lipid bilayer comprises of liquid ordered (Lo) and liquid disordered (Ld) domains. Lo microdomains, also named as lipid rafts are enriched in cholesterol, sphingomyelin and certain types of proteins, which facilitate cell signalling and nutrient uptake. Lipid rafts have been extensively researched in mammals and the presence of functional lipid rafts was recently demonstrated in goldfish, but there is currently very little knowledge about their composition and function in fish. Therefore a protocol was established for the analysis of lipid rafts and membranous lipids in common carp (Cyprinus carpio L.) tissues. Twelve lipids were identified and analysed in the Ld domain of the membrane with the most predominant lipids found in all tissues being; triglycerides, cholesterol, phosphoethanolamine and phosphatidylcholine. Four lipids were identified in lipid rafts in all tissues analysed, triglycerides (33-62%) always found in the highest concentration followed by cholesterol (24-32%), phosphatidylcholine and sphingomyelin. Isolation of lipid rafts was confirmed by identifying the presence of the lipid raft associated protein flotillin, present at higher concentrations in the detergent resistant fraction. The data provided here build a lipid library of important carp tissues as a baseline for further studies into virus entry, protein trafficking or environmental stress analysis.

  20. Plant P4-ATPases: lipid translocators with a role in membrane traficking

    DEFF Research Database (Denmark)

    Lopez Marques, Rosa Laura

    recently completed the biochemical characterization of two ALA proteins: ALA2, a prevacuolar compartment-localized protein with an unusually tight specificity, and ALA10, a plasma membrane-localized protein with and unforeseen broad substrate specificity. Besides providing an insight into the mechanism...... of lipid translocation, our results suggest that the different transport features of these proteins might be related to their physiological function at the membrane where they are located....... a large family of membrane proteins involved in pumping different physiologically-relevant substrates across biological membranes [4]. The members of the P4 subfamily (also known as flippases) catalyze the energy-driven translocation of lipids necessary for establishing transbilayer lipid asymmetry [5...

  1. Anomalous diffusion of proteins in sheared lipid membranes

    CERN Document Server

    Khoshnood, Atefeh

    2013-01-01

    We use coarse grained molecular dynamics simulations to investigate diffusion properties of sheared lipid membranes with embedded transmembrane proteins. In membranes without proteins, we find normal in-plane diffusion of lipids in all flow conditions. Protein embedded membranes behave quite differently: by imposing a simple shear flow and sliding the monolayers of the membrane over each other, the motion of protein clusters becomes strongly superdiffusive in the shear direction. In such a circumstance, subdiffusion regime is predominant perpendicular to the flow. We show that superdiffusion is a result of accelerated chaotic motions of protein--lipid complexes within the membrane voids, which are generated by hydrophobic mismatch or the transport of lipids by proteins.

  2. The plant plasma membrane H+-ATPase

    DEFF Research Database (Denmark)

    Ekberg, Kira

      The very high mobility of protons in aqueous solutions demands special features of membrane proton transporters to sustain efficient yet regulated proton transport across biological membranes. By the use of the chemical energy of ATP, plasma-membrane-embedded H+-ATPases extrude protons from cells...... of plants and fungi to generate electrochemical proton gradients. A recently published crystal structure of a plasma membrane H(+)-ATPase contributes to our knowledge about the mechanism of these essential enzymes. Together with biochemical and structural data presented in this thesis we are now able...... to describe the basic molecular components that allow the plasma membrane proton H+-ATPase to carry out proton transport against large membrane potentials. Moreover, a completely new paradigm for post-translational activation of these proteins is presented. The talk will focus on the following themes...

  3. The plant plasma membrane H+-ATPase

    DEFF Research Database (Denmark)

    Ekberg, Kira

      The very high mobility of protons in aqueous solutions demands special features of membrane proton transporters to sustain efficient yet regulated proton transport across biological membranes. By the use of the chemical energy of ATP, plasma-membrane-embedded H+-ATPases extrude protons from cells...... of plants and fungi to generate electrochemical proton gradients. A recently published crystal structure of a plasma membrane H(+)-ATPase contributes to our knowledge about the mechanism of these essential enzymes. Together with biochemical and structural data presented in this thesis we are now able...... to describe the basic molecular components that allow the plasma membrane proton H+-ATPase to carry out proton transport against large membrane potentials. Moreover, a completely new paradigm for post-translational activation of these proteins is presented. The talk will focus on the following themes...

  4. G protein-membrane interactions II: Effect of G protein-linked lipids on membrane structure and G protein-membrane interactions.

    Science.gov (United States)

    Casas, Jesús; Ibarguren, Maitane; Álvarez, Rafael; Terés, Silvia; Lladó, Victoria; Piotto, Stefano P; Concilio, Simona; Busquets, Xavier; López, David J; Escribá, Pablo V

    2017-09-01

    G proteins often bear myristoyl, palmitoyl and isoprenyl moieties, which favor their association with the membrane and their accumulation in G Protein Coupled Receptor-rich microdomains. These lipids influence the biophysical properties of membranes and thereby modulate G protein binding to bilayers. In this context, we showed here that geranylgeraniol, but neither myristate nor palmitate, increased the inverted hexagonal (HII) phase propensity of phosphatidylethanolamine-containing membranes. While myristate and palmitate preferentially associated with phosphatidylcholine membranes, geranylgeraniol favored nonlamellar-prone membranes. In addition, Gαi1 monomers had a higher affinity for lamellar phases, while Gβγ and Gαβγ showed a marked preference for nonlamellar prone membranes. Moreover, geranylgeraniol enhanced the binding of G protein dimers and trimers to phosphatidylethanolamine-containing membranes, yet it decreased that of monomers. By contrast, both myristate and palmitate increased the Gαi1 preference for lamellar membranes. Palmitoylation reinforced the binding of the monomer to PC membranes and myristoylation decreased its binding to PE-enriched bilayer. Finally, binding of dimers and trimers to lamellar-prone membranes was decreased by palmitate and myristate, but it was increased in nonlamellar-prone bilayers. These results demonstrate that co/post-translational G protein lipid modifications regulate the membrane lipid structure and that they influence the physico-chemical properties of membranes, which in part explains why G protein subunits sort to different plasma membrane domains. This article is part of a Special Issue entitled: Membrane Lipid Therapy: Drugs Targeting Biomembranes edited by Pablo V. Escribá. Copyright © 2017 Elsevier B.V. All rights reserved.

  5. Challenges in the theoretical investigations of lipid membrane configurations

    Institute of Scientific and Technical Information of China (English)

    Tu Zhan-Chun

    2013-01-01

    We report some key results in the theoretical investigations of configurations of lipid membranes and present several challenges in this field,which involve (i) the exact solutions to the shape equation of lipid vesicles,(ii) the exact solutions to the governing equations of open lipid membranes,(iii) the neck condition of two-phase vesicles in the budding state,(iv)the nonlocal theory of membrane elasticity,and (v) the relationship between the symmetry and the magnitude of the free energy.

  6. Evaluation of plasma lipids and lipoproteins in nigerians suffering ...

    African Journals Online (AJOL)

    Evaluation of plasma lipids and lipoproteins in nigerians suffering from depressive illness. ... Very little is known about the lipid and lipoprotein status in Nigerian adults suffering from depression. One hundred subjects ... Article Metrics.

  7. Aspirin inhibits formation of cholesterol rafts in fluid lipid membranes.

    Science.gov (United States)

    Alsop, Richard J; Toppozini, Laura; Marquardt, Drew; Kučerka, Norbert; Harroun, Thad A; Rheinstädter, Maikel C

    2015-03-01

    Aspirin and other non-steroidal anti-inflammatory drugs have a high affinity for phospholipid membranes, altering their structure and biophysical properties. Aspirin has been shown to partition into the lipid head groups, thereby increasing membrane fluidity. Cholesterol is another well known mediator of membrane fluidity, in turn increasing membrane stiffness. As well, cholesterol is believed to distribute unevenly within lipid membranes leading to the formation of lipid rafts or plaques. In many studies, aspirin has increased positive outcomes for patients with high cholesterol. We are interested if these effects may be, at least partially, the result of a non-specific interaction between aspirin and cholesterol in lipid membranes. We have studied the effect of aspirin on the organization of 1,2-dipalmitoyl-sn-glycero-3-phosphatidylcholine (DPPC) membranes containing cholesterol. Through Langmuir-Blodgett experiments we show that aspirin increases the area per lipid and decreases compressibility at 32.5 mol% cholesterol, leading to a significant increase of fluidity of the membranes. Differential scanning calorimetry provides evidence for the formation of meta-stable structures in the presence of aspirin. The molecular organization of lipids, cholesterol and aspirin was studied using neutron diffraction. While the formation of rafts has been reported in binary DPPC/cholesterol membranes, aspirin was found to locally disrupt membrane organization and lead to the frustration of raft formation. Our results suggest that aspirin is able to directly oppose the formation of cholesterol structures through non-specific interactions with lipid membranes. Copyright © 2014 Elsevier B.V. All rights reserved.

  8. Endosomal recycling controls plasma membrane area during mitosis.

    Science.gov (United States)

    Boucrot, Emmanuel; Kirchhausen, Tomas

    2007-05-08

    The shape and total surface of a cell and its daughters change during mitosis. Many cells round up during prophase and metaphase and reacquire their extended and flattened shape during cytokinesis. How does the total area of plasma membrane change to accommodate these morphological changes and by what mechanism is control of total membrane area achieved? Using single-cell imaging methods, we have found that the amount of plasma membrane in attached cells in culture decreases at the beginning of mitosis and recovers rapidly by the end. Clathrin-based endocytosis is normal throughout all phases of cell division, whereas recycling of internalized membranes back to the cell surface slows considerably during the rounding up period and resumes at the time at which recovery of cell membrane begins. Interference with either one of these processes by genetic or chemical means impairs cell division. The total cell-membrane area recovers even in the absence of a functional Golgi apparatus, which would be needed for export of newly synthesized membrane lipids and proteins. We propose a mechanism by which modulation of endosomal recycling controls cell area and surface expression of membrane-bound proteins during cell division.

  9. Plasma membranes from insect midgut cells

    Directory of Open Access Journals (Sweden)

    Walter R. Terra

    2006-06-01

    Full Text Available Plasma membranes from insect midgut cells are separated into apical and basolateral domains. The apical domain is usually modified into microvilli with a molecular structure similar to other animals. Nevertheless, the microvillar structure should differ in some insects to permit the traffic inside them of secretory vesicles that may budd laterally or pinch-off from the tips of microvilli. Other microvillar modifications are associated with proton-pumping or with the interplay with an ensheathing lipid membrane (the perimicrovilllar membrane observed in the midgut cells of hemipterans (aphids and bugs. The perimicrovillar membranes are thought to be involved in amino acid absorption from diluted diets. The microvillar and perimicrovillar membranes have densities (and protein content that depend on the insect taxon. The role played by the microvillar and perimicrovillar proteins in insect midgut physiology is reviewed here trying to provide a coherent picture of data and highlighting further research areas.As membranas plasmáticas das células intestinais dos insetos apresentam um domínio apical e outro basal. O domínio apical é geralmente modificado em microvilosidades com organização molecular similar a de outros animais, embora possam diferir naqueles insetos que apresentam vesículas secretoras em trânsito que brotam lateralmente ou destacam-se das extremidades das microvilosidades. Outras modificações microvilares estão associadas a bombeamento de prótons ou a interrelações com uma membrana lipídica (a membrana perimicrovilar que reveste as microvilosidades de células intestinais de hemípteros (pulgões e percevejos. Admite-se que as membranas perimicrovilares estejam envolvidas na absorção de aminoácidos a partir de dietas diluídas. As membranas microvilares e perimicrovilares tem densidades distintas (e conteúdo protéico que dependem do táxon do inseto. O papel desempenhado pelas proteínas microvilares e

  10. Nucleic acid-lipid membrane interactions studied by DSC.

    Science.gov (United States)

    Giatrellis, Sarantis; Nounesis, George

    2011-01-01

    The interactions of nucleic acids with lipid membranes are of great importance for biological mechanisms as well as for biotechnological applications in gene delivery and drug carriers. The optimization of liposomal vectors for clinical use is absolutely dependent upon the formation mechanisms, the morphology, and the molecular organization of the lipoplexes, that is, the complexes of lipid membranes with DNA. Differential scanning calorimetry (DSC) has emerged as an efficient and relatively easy-to-operate experimental technique that can straightforwardly provide data related to the thermodynamics and the kinetics of the DNA-lipid complexation and especially to the lipid organization and phase transitions within the membrane. In this review, we summarize DSC studies considering nucleic acid-membrane systems, accentuating DSC capabilities, and data analysis. Published work involving cationic, anionic, and zwitterionic lipids as well as lipid mixtures interacting with RNA and DNA of different sizes and conformations are included. It is shown that despite limitations, issues such as DNA- or RNA-induced phase separation and microdomain lipid segregation, liposomal aggregation and fusion, alterations of the lipid long-range molecular order, as well as membrane-induced structural changes of the nucleic acids can be efficiently treated by systematic high-sensitivity DSC studies.

  11. Nucleic acid-lipid membrane interactions studied by DSC

    Directory of Open Access Journals (Sweden)

    Sarantis Giatrellis

    2011-01-01

    Full Text Available The interactions of nucleic acids with lipid membranes are of great importance for biological mechanisms as well as for biotechnological applications in gene delivery and drug carriers. The optimization of liposomal vectors for clinical use is absolutely dependent upon the formation mechanisms, the morphology, and the molecular organization of the lipoplexes, that is, the complexes of lipid membranes with DNA. Differential scanning calorimetry (DSC has emerged as an efficient and relatively easy-to-operate experimental technique that can straightforwardly provide data related to the thermodynamics and the kinetics of the DNA-lipid complexation and especially to the lipid organization and phase transitions within the membrane. In this review, we summarize DSC studies considering nucleic acid-membrane systems, accentuating DSC capabilities, and data analysis. Published work involving cationic, anionic, and zwitterionic lipids as well as lipid mixtures interacting with RNA and DNA of different sizes and conformations are included. It is shown that despite limitations, issues such as DNA- or RNA-induced phase separation and microdomain lipid segregation, liposomal aggregation and fusion, alterations of the lipid long-range molecular order, as well as membrane-induced structural changes of the nucleic acids can be efficiently treated by systematic high-sensitivity DSC studies.

  12. Membrane curvature enables N-Ras lipid anchor sorting to liquid-ordered membrane phases

    DEFF Research Database (Denmark)

    Larsen, Jannik Bruun; Jensen, Martin Borch; Bhatia, Vikram Kjøller;

    2015-01-01

    Trafficking and sorting of membrane-anchored Ras GTPases are regulated by partitioning between distinct membrane domains. Here, in vitro experiments and microscopic molecular theory reveal membrane curvature as a new modulator of N-Ras lipid anchor and palmitoyl chain partitioning. Membrane...... curvature was essential for enrichment in raft-like liquid-ordered phases; enrichment was driven by relief of lateral pressure upon anchor insertion and most likely affects the localization of lipidated proteins in general....

  13. Remodeling of the postsynaptic plasma membrane during neural development.

    Science.gov (United States)

    Tulodziecka, Karolina; Diaz-Rohrer, Barbara B; Farley, Madeline M; Chan, Robin B; Di Paolo, Gilbert; Levental, Kandice R; Waxham, M Neal; Levental, Ilya

    2016-11-07

    Neuronal synapses are the fundamental units of neural signal transduction and must maintain exquisite signal fidelity while also accommodating the plasticity that underlies learning and development. To achieve these goals, the molecular composition and spatial organization of synaptic terminals must be tightly regulated; however, little is known about the regulation of lipid composition and organization in synaptic membranes. Here we quantify the comprehensive lipidome of rat synaptic membranes during postnatal development and observe dramatic developmental lipidomic remodeling during the first 60 postnatal days, including progressive accumulation of cholesterol, plasmalogens, and sphingolipids. Further analysis of membranes associated with isolated postsynaptic densities (PSDs) suggests the PSD-associated postsynaptic plasma membrane (PSD-PM) as one specific location of synaptic remodeling. We analyze the biophysical consequences of developmental remodeling in reconstituted synaptic membranes and observe remarkably stable microdomains, with the stability of domains increasing with developmental age. We rationalize the developmental accumulation of microdomain-forming lipids in synapses by proposing a mechanism by which palmitoylation of the immobilized scaffold protein PSD-95 nucleates domains at the postsynaptic plasma membrane. These results reveal developmental changes in lipid composition and palmitoylation that facilitate the formation of postsynaptic membrane microdomains, which may serve key roles in the function of the neuronal synapse. © 2016 Tulodziecka et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

  14. Insights into thermophilic archaebacterial membrane stability from simplified models of lipid membranes

    Science.gov (United States)

    Davis, Charles H.; Nie, Huifen; Dokholyan, Nikolay V.

    2007-05-01

    Lipid aggregation into fluid bilayers is an essential process for sustaining life. Simplified models of lipid structure, which allow for long time scales or large length scales not obtainable with all-atom simulations, have recently been developed and show promise for describing lipid dynamics in biological systems. Here, we describe two simplified models, a reduced-lipid model and a bola-lipid model for thermophilic bacterial membranes, developed for use with the rapid discrete molecular dynamics simulation method. In the reduced-lipid model, we represent the lipid chain by a series of three beads interacting through pairwise discrete potentials that model hydrophobic attractions between hydrocarbon tails in implicit solvent. Our phase diagram recapitulates those produced by continuous potential models with similar coarse-grained lipid representations. We also find that phase transition temperatures for our reduced-lipid model are dependent upon the flexibility of the lipid chain, giving an insight into archaebacterial membrane stability and prompting development of a bola-lipid model specific for archaebacteria lipids. With both the reduced-lipid and bola-lipid model, we find that the reduced flexibility inherent in archaebacteria lipids yields more stable bilayers as manifested by increased phase transition temperatures. The results of these studies provide a simulation methodology for lipid molecules in biological systems and show that discrete molecular dynamics is applicable to lipid aggregation and dynamics.

  15. Reverse-osmosis membranes by plasma polymerization

    Science.gov (United States)

    Hollahan, J. R.; Wydeven, T.

    1972-01-01

    Thin allyl amine polymer films were developed using plasma polymerization. Resulting dry composite membranes effectively reject sodium chloride during reverse osmosis. Films are 98% sodium chloride rejective, and 46% urea rejective.

  16. Paracrine signaling through plasma membrane hemichannels

    National Research Council Canada - National Science Library

    Wang, Nan; De Bock, Marijke; Decrock, Elke; Bol, Mélissa; Gadicherla, Ashish; Vinken, Mathieu; Rogiers, Vera; Bukauskas, Feliksas F; Bultynck, Geert; Leybaert, Luc

    2013-01-01

    Plasma membrane hemichannels composed of connexin (Cx) proteins are essential components of gap junction channels but accumulating evidence suggests functions of hemichannels beyond the communication provided by junctional channels...

  17. Proteomics and the dynamic plasma membrane

    DEFF Research Database (Denmark)

    Sprenger, Richard R; Jensen, Ole Nørregaard

    2010-01-01

    plasma membrane is of particular interest, by not only serving as a barrier between the "cell interior" and the external environment, but moreover by organizing and clustering essential components to enable dynamic responses to internal and external stimuli. Defining and characterizing the dynamic plasma...... membrane proteome is crucial for understanding fundamental biological processes, disease mechanisms and for finding drug targets. Protein identification, characterization of dynamic PTMs and protein-ligand interactions, and determination of transient changes in protein expression and composition are among...... the challenges in functional proteomic studies of the plasma membrane. We review the recent progress in MS-based plasma membrane proteomics by presenting key examples from eukaryotic systems, including mammals, yeast and plants. We highlight the importance of enrichment and quantification technologies required...

  18. Role of zinc in plasma membrane function

    National Research Council Canada - National Science Library

    O'Dell, B L

    2000-01-01

    ... with a posttranslational change in plasma membrane proteins. Among the signs of zinc deficiency in rats is a bleeding tendency associated with failure of platelet aggregation, a phenomenon that correlates with impaired uptake of Ca(2+) when stimulated...

  19. Transport proteins of the plant plasma membrane

    Science.gov (United States)

    Assmann, S. M.; Haubrick, L. L.; Evans, M. L. (Principal Investigator)

    1996-01-01

    Recently developed molecular and genetic approaches have enabled the identification and functional characterization of novel genes encoding ion channels, ion carriers, and water channels of the plant plasma membrane.

  20. Electro-hydrodynamic effects on lipid membranes in giant vesicles

    Science.gov (United States)

    Staykova, Margarita; Yamamoto, Tetsuya; Lipowsky, Reinhard; Dimova, Rumiana

    2009-11-01

    Electric fields are widely applied for cell manipulation in numerous micron-scale systems. Here, we show for the first time that alternating electric fields may cause pronounced flows in the membrane of giant lipid vesicles as well as in the surrounding fluid media.^ The lipid vesicles are not only biomimetic model for the cell membrane but also have many potential biotechnological applications, e.g. as drug-delivery systems and micro-reactors. The reported effects should be considered in electric micro-manipulation procedures on cells and vesicles. They might be useful for applications in microfluidic technologies, for lipid mixing, trapping and displacement, as will be demonstrated. We also believe that our method for visualization of the lipid flows by fluorescently labeled intra-membrane domains will be helpful for studies on membrane behavior in vesicles subjected to shear or mechanical stresses.

  1. Chain ordering of hybrid lipids can stabilize domains in saturated/hybrid/cholesterol lipid membranes

    Science.gov (United States)

    Yamamoto, T.; Brewster, R.; Safran, S. A.

    2010-07-01

    We use a liquid-crystal model to predict that hybrid lipids (lipids that have one saturated and one unsaturated tail) can stabilize line interfaces between domains in mixed membranes of saturated lipids, hybrid lipids, and cholesterol (SHC membranes). The model predicts the phase separation of SHC membranes with both parabolic and loop binodals depending on the cholesterol concentration, modeled via an effective pressure. In some cases, the hybrid lipids can reduce the line tension to zero in SHC membranes at temperatures that approach the critical temperature as the pressure is increased. The differences in the hybrid saturated tail conformational order in bulk and at the interface are responsible for the reduction of the line tension.

  2. Sizes of lipid domains: What do we know from artificial lipid membranes? What are the possible shared features with membrane rafts in cells?

    Science.gov (United States)

    Rosetti, Carla M; Mangiarotti, Agustín; Wilke, Natalia

    2017-01-28

    In model lipid membranes with phase coexistence, domain sizes distribute in a very wide range, from the nanometer (reported in vesicles and supported films) to the micrometer (observed in many model membranes). Domain growth by coalescence and Ostwald ripening is slow (minutes to hours), the domain size being correlated with the size of the capture region. Domain sizes thus strongly depend on the number of domains which, in the case of a nucleation process, depends on the oversaturation of the system, on line tension and on the perturbation rate in relation to the membrane dynamics. Here, an overview is given of the factors that affect nucleation or spinodal decomposition and domain growth, and their influence on the distribution of domain sizes in different model membranes is discussed. The parameters analyzed respond to very general physical rules, and we therefore propose a similar behavior for the rafts in the plasma membrane of cells, but with obstructed mobility and with a continuously changing environment.

  3. The role of interfacial lipids in stabilizing membrane protein oligomers.

    Science.gov (United States)

    Gupta, Kallol; Donlan, Joseph A C; Hopper, Jonathan T S; Uzdavinys, Povilas; Landreh, Michael; Struwe, Weston B; Drew, David; Baldwin, Andrew J; Stansfeld, Phillip J; Robinson, Carol V

    2017-01-19

    Oligomerization of membrane proteins in response to lipid binding has a critical role in many cell-signalling pathways but is often difficult to define or predict. Here we report the development of a mass spectrometry platform to determine simultaneously the presence of interfacial lipids and oligomeric stability and to uncover how lipids act as key regulators of membrane-protein association. Evaluation of oligomeric strength for a dataset of 125 α-helical oligomeric membrane proteins reveals an absence of interfacial lipids in the mass spectra of 12 membrane proteins with high oligomeric stability. For the bacterial homologue of the eukaryotic biogenic transporters (LeuT, one of the proteins with the lowest oligomeric stability), we found a precise cohort of lipids within the dimer interface. Delipidation, mutation of lipid-binding sites or expression in cardiolipin-deficient Escherichia coli abrogated dimer formation. Molecular dynamics simulation revealed that cardiolipin acts as a bidentate ligand, bridging across subunits. Subsequently, we show that for the Vibrio splendidus sugar transporter SemiSWEET, another protein with low oligomeric stability, cardiolipin shifts the equilibrium from monomer to functional dimer. We hypothesized that lipids are essential for dimerization of the Na(+)/H(+) antiporter NhaA from E. coli, which has the lowest oligomeric strength, but not for the substantially more stable homologous Thermus thermophilus protein NapA. We found that lipid binding is obligatory for dimerization of NhaA, whereas NapA has adapted to form an interface that is stable without lipids. Overall, by correlating interfacial strength with the presence of interfacial lipids, we provide a rationale for understanding the role of lipids in both transient and stable interactions within a range of α-helical membrane proteins, including G-protein-coupled receptors.

  4. Atomistic study of lipid membranes containing chloroform: looking for a lipid-mediated mechanism of anesthesia.

    Directory of Open Access Journals (Sweden)

    Ramon Reigada

    Full Text Available The molecular mechanism of general anesthesia is still a controversial issue. Direct effect by linking of anesthetics to proteins and indirect action on the lipid membrane properties are the two hypotheses in conflict. Atomistic simulations of different lipid membranes subjected to the effect of small volatile organohalogen compounds are used to explore plausible lipid-mediated mechanisms. Simulations of homogeneous membranes reveal that electrostatic potential and lateral pressure transversal profiles are affected differently by chloroform (anesthetic and carbon tetrachloride (non-anesthetic. Simulations of structured membranes that combine ordered and disordered regions show that chloroform molecules accumulate preferentially in highly disordered lipid domains, suggesting that the combination of both lateral and transversal partitioning of chloroform in the cell membrane could be responsible of its anesthetic action.

  5. Bile acids modulate signaling by functional perturbation of plasma membrane domains.

    Science.gov (United States)

    Zhou, Yong; Maxwell, Kelsey N; Sezgin, Erdinc; Lu, Maryia; Liang, Hong; Hancock, John F; Dial, Elizabeth J; Lichtenberger, Lenard M; Levental, Ilya

    2013-12-13

    Eukaryotic cell membranes are organized into functional lipid and protein domains, the most widely studied being membrane rafts. Although rafts have been associated with numerous plasma membrane functions, the mechanisms by which these domains themselves are regulated remain undefined. Bile acids (BAs), whose primary function is the solubilization of dietary lipids for digestion and absorption, can affect cells by interacting directly with membranes. To investigate whether these interactions affected domain organization in biological membranes, we assayed the effects of BAs on biomimetic synthetic liposomes, isolated plasma membranes, and live cells. At cytotoxic concentrations, BAs dissolved synthetic and cell-derived membranes and disrupted live cell plasma membranes, implicating plasma membrane damage as the mechanism for BA cellular toxicity. At subtoxic concentrations, BAs dramatically stabilized domain separation in Giant Plasma Membrane Vesicles without affecting protein partitioning between coexisting domains. Domain stabilization was the result of BA binding to and disordering the nonraft domain, thus promoting separation by enhancing domain immiscibility. Consistent with the physical changes observed in synthetic and isolated biological membranes, BAs reorganized intact cell membranes, as evaluated by the spatial distribution of membrane-anchored Ras isoforms. Nanoclustering of K-Ras, related to nonraft membrane domains, was enhanced in intact plasma membranes, whereas the organization of H-Ras was unaffected. BA-induced changes in Ras lateral segregation potentiated EGF-induced signaling through MAPK, confirming the ability of BAs to influence cell signal transduction by altering the physical properties of the plasma membrane. These observations suggest general, membrane-mediated mechanisms by which biological amphiphiles can produce their cellular effects.

  6. Lipid partitioning at the nuclear envelope controls membrane biogenesis

    NARCIS (Netherlands)

    Barbosa, Antonio Daniel; Sembongi, Hiroshi; Su, Wen-Min; Abreu, Susana; Reggiori, Fulvio; Carman, George M.; Siniossoglou, Symeon

    2015-01-01

    Partitioning of lipid precursors between membranes and storage is crucial for cell growth, and its disruption underlies pathologies such as cancer, obesity, and type 2 diabetes. However, the mechanisms and signals that regulate this process are largely unknown. In yeast, lipid precursors are mainly

  7. Slow fusion of liposomes composed of membrane-spanning lipids

    NARCIS (Netherlands)

    Elferink, MGL; vanBreemen, J; Konings, WN; Driessen, AJM; Wilschut, J; Elferink, Marieke G.L.

    1997-01-01

    The fusion characteristics of large unilamellar liposomes composed of bipolar tetraether lipids extracted from the thermophilic archaeon Sulfolobus acidocaldarius, was investigated. These lipids span the entire membrane and form single monolayer liposomes in aqueous media [Elferink, M.G.L., de Wit,

  8. Aspirin Increases the Solubility of Cholesterol in Lipid Membranes

    Science.gov (United States)

    Alsop, Richard; Barrett, Matthew; Zheng, Sonbo; Dies, Hannah; Rheinstadter, Maikel

    2014-03-01

    Aspirin (ASA) is often prescribed for patients with high levels of cholesterol for the secondary prevention of myocardial events, a regimen known as the Low-Dose Aspirin Therapy. We have recently shown that Aspirin partitions in lipid bilayers. However, a direct interplay between ASA and cholesterol has not been investigated. Cholesterol is known to insert itself into the membrane in a dispersed state at moderate concentrations (under ~37.5%) and decrease fluidity of membranes. We prepared model lipid membranes containing varying amounts of both ASA and cholesterol molecules. The structure of the bilayers as a function of ASA and cholesterol concentration was determined using high-resolution X-ray diffraction. At cholesterol levels of more than 40mol%, immiscible cholesterol plaques formed. Adding ASA to the membranes was found to dissolve the cholesterol plaques, leading to a fluid lipid bilayer structure. We present first direct evidence for an interaction between ASA and cholesterol on the level of the cell membrane.

  9. The Deleterious Effects of Oxidative and Nitrosative Stress on Palmitoylation, Membrane Lipid Rafts and Lipid-Based Cellular Signalling: New Drug Targets in Neuroimmune Disorders.

    Science.gov (United States)

    Morris, Gerwyn; Walder, Ken; Puri, Basant K; Berk, Michael; Maes, Michael

    2016-09-01

    Oxidative and nitrosative stress (O&NS) is causatively implicated in the pathogenesis of Alzheimer's and Parkinson's disease, multiple sclerosis, chronic fatigue syndrome, schizophrenia and depression. Many of the consequences stemming from O&NS, including damage to proteins, lipids and DNA, are well known, whereas the effects of O&NS on lipoprotein-based cellular signalling involving palmitoylation and plasma membrane lipid rafts are less well documented. The aim of this narrative review is to discuss the mechanisms involved in lipid-based signalling, including palmitoylation, membrane/lipid raft (MLR) and n-3 polyunsaturated fatty acid (PUFA) functions, the effects of O&NS processes on these processes and their role in the abovementioned diseases. S-palmitoylation is a post-translational modification, which regulates protein trafficking and association with the plasma membrane, protein subcellular location and functions. Palmitoylation and MRLs play a key role in neuronal functions, including glutamatergic neurotransmission, and immune-inflammatory responses. Palmitoylation, MLRs and n-3 PUFAs are vulnerable to the corruptive effects of O&NS. Chronic O&NS inhibits palmitoylation and causes profound changes in lipid membrane composition, e.g. n-3 PUFA depletion, increased membrane permeability and reduced fluidity, which together lead to disorders in intracellular signal transduction, receptor dysfunction and increased neurotoxicity. Disruption of lipid-based signalling is a source of the neuroimmune disorders involved in the pathophysiology of the abovementioned diseases. n-3 PUFA supplementation is a rational therapeutic approach targeting disruptions in lipid-based signalling.

  10. The strength of side chain hydrogen bonds in the plasma membrane

    Science.gov (United States)

    Hristova, Kalina; Sarabipour, Sarvenaz

    2013-03-01

    There are no direct quantitative measurements of hydrogen bond strengths in membrane proteins residing in their native cellular environment. To address this knowledge gap, here we use fluorescence resonance energy transfer (FRET) to measure the impact of hydrogen bonds on the stability of a membrane protein dimer in vesicles derived from eukaryotic plasma membranes, and we compare these results to previous measurements of hydrogen bond strengths in model lipid bilayers. We demonstrate that FRET measurements of membrane protein interactions in plasma membrane vesicles have the requisite sensitivity to quantify the strength of hydrogen bonds. We find that the hydrogen bond-mediated stabilization in the plasma membrane is small, only -0.7 kcal/mole. It is the same as in model lipid bilayers, despite the different nature and dielectric properties of the two environments.

  11. The SNARE Sec22b has a non-fusogenic function in plasma membrane expansion.

    Science.gov (United States)

    Petkovic, Maja; Jemaiel, Aymen; Daste, Frédéric; Specht, Christian G; Izeddin, Ignacio; Vorkel, Daniela; Verbavatz, Jean-Marc; Darzacq, Xavier; Triller, Antoine; Pfenninger, Karl H; Tareste, David; Jackson, Catherine L; Galli, Thierry

    2014-05-01

    Development of the nervous system requires extensive axonal and dendritic growth during which neurons massively increase their surface area. Here we report that the endoplasmic reticulum (ER)-resident SNARE Sec22b has a conserved non-fusogenic function in plasma membrane expansion. Sec22b is closely apposed to the plasma membrane SNARE syntaxin1. Sec22b forms a trans-SNARE complex with syntaxin1 that does not include SNAP23/25/29, and does not mediate fusion. Insertion of a long rigid linker between the SNARE and transmembrane domains of Sec22b extends the distance between the ER and plasma membrane, and impairs neurite growth but not the secretion of VSV-G. In yeast, Sec22 interacts with lipid transfer proteins, and inhibition of Sec22 leads to defects in lipid metabolism at contact sites between the ER and plasma membrane. These results suggest that close apposition of the ER and plasma membrane mediated by Sec22 and plasma membrane syntaxins generates a non-fusogenic SNARE bridge contributing to plasma membrane expansion, probably through non-vesicular lipid transfer.

  12. Impact of ionic liquids in aqueous solution on bacterial plasma membranes studied with molecular dynamics simulations.

    Science.gov (United States)

    Lim, Geraldine S; Zidar, Jernej; Cheong, Daniel W; Jaenicke, Stephan; Klähn, Marco

    2014-09-04

    The impact of five different imidazolium-based ionic liquids (ILs) diluted in water on the properties of a bacterial plasma membrane is investigated using molecular dynamics (MD) simulations. Cations considered are 1-octyl-3-methylimidazolium (OMIM), 1-octyloxymethyl-3-methylimidazolium (OXMIM), and 1-tetradecyl-3-methylimidazolium (TDMIM), as well as the anions chloride and lactate. The atomistic model of the membrane bilayer is designed to reproduce the lipid composition of the plasma membrane of Gram-negative Escherichia coli. Spontaneous insertion of cations into the membrane is observed in all ILs. Substantially more insertions of OMIM than of OXMIM occur and the presence of chloride reduces cation insertions compared to lactate. In contrast, anions do not adsorb onto the membrane surface nor diffuse into the bilayer. Once inserted, cations are oriented in parallel to membrane lipids with cation alkyl tails embedded into the hydrophobic membrane core, while the imidazolium-ring remains mostly exposed to the solvent. Such inserted cations are strongly associated with one to two phospholipids in the membrane. The overall order of lipids decreased after OMIM and OXMIM insertions, while on the contrary the order of lipids in the vicinity of TDMIM increased. The short alkyl tails of OMIM and OXMIM generate voids in the bilayer that are filled by curling lipids. This cation induced lipid disorder also reduces the average membrane thickness. This effect is not observed after TDMIM insertions due to the similar length of cation alkyl chain and the fatty acids of the lipids. This lipid-mimicking behavior of inserted TDMIM indicates a high membrane affinity of this cation that could lead to an enhanced accumulation of cations in the membrane over time. Overall, the simulations reveal how cations are inserted into the bacterial membrane and how such insertions change its properties. Moreover, the different roles of cations and anions are highlighted and the fundamental

  13. DMSO induces dehydration near lipid membrane surfaces

    National Research Council Canada - National Science Library

    Cheng, Chi-Yuan; Song, Jinsuk; Pas, Jolien; Meijer, Lenny H H; Han, Songi

    2015-01-01

    Dimethyl sulfoxide (DMSO) has been broadly used in biology as a cosolvent, a cryoprotectant, and an enhancer of membrane permeability, leading to the general assumption that DMSO-induced structural changes in cell membranes...

  14. Undulation instability in a bilayer lipid membrane due to electric field interaction with lipid dipoles

    CERN Document Server

    Bingham, Richard J; Smye, Stephen W

    2010-01-01

    Bilayer lipid membranes [BLMs] are an essential component of all biological systems, forming a functional barrier for cells and organelles from the surrounding environment. The lipid molecules that form membranes contain both permanent and induced dipoles, and an electric field can induce the formation of pores when the transverse field is sufficiently strong (electroporation). Here, a phenomenological free energy is constructed to model the response of a BLM to a transverse static electric field. The model contains a continuum description of the membrane dipoles and a coupling between the headgroup dipoles and the membrane tilt. The membrane is found to become unstable through buckling modes, which are weakly coupled to thickness fluctuations in the membrane. The thickness fluctuations, along with the increase in interfacial area produced by membrane buckling, increase the probability of localized membrane breakdown, which may lead to pore formation. The instability is found to depend strongly on the strengt...

  15. Pushing the lipid envelope: using bio-inspired nanocomposites to understand and exploit lipid membrane limitations

    Science.gov (United States)

    Montano, Gabriel

    Lipids serve as the organizing matrix material for biological membranes, the site of interaction of cells with the external environment. . As such, lipids play a critical role in structure/function relationships of an extraordinary number of critical biological processes. In this talk, we will look at bio-inspired membrane assemblies to better understand the roles of lipids in biological systems as well as attempt to generate materials that can mimic and potentially advance upon biological membrane processes. First, we will investigate the response of lipids to adverse conditions. In particular, I will present data that demonstrates the response of lipids to harsh conditions and how such responses can be exploited to generate nanocomposite rearrangements. I will also show the effect of adding the endotoxin lipopolysaccharide (LPS) to lipid bilayer assemblies and describe implications on our understanding of LPS organization in biological systems as well as describe induced lipid modifications that can be exploited to organize membrane composites with precise, two-dimensional geometric control. Lastly, I will describe the use of amphiphilic block copolymers to create membrane nanocomposites capable of mimicking biological systems. In particular, I will describe the use of our polymer-based membranes in creating artificial photosynthetic assemblies that rival biological systems in function in a more flexible, dynamic matrix.

  16. Ionic channels and nerve membrane lipids. Cholesterol-tetrodotoxin interaction.

    Science.gov (United States)

    Villegas, R; Barnola, F V; Camejo, G

    1970-04-01

    Experiments were carried out to investigate possible interactions of tetrodotoxin (TTX) with lipid molecules isolated from nerve fiber plasma membranes of the squid Dosidicus gigas. TTX has a highly selective ability to block the channel normally used by Na(+) to cross the axolemma during nervous impulse conduction. In order to investigate the interaction each lipid sample was spread on 5 x 10(-7)M TTX and TTX-free 0.15 M NaCl solutions adjusted to pH 7.4 with 7 x 10(-3)M phosphate buffer. The surface pressure-area diagrams of the lipid monolayers revealed that TTX interacts only with cholesterol. The expansion of the cholesterol monolayers at 5 x 10(-7)M TTX was 2 A(2)/molecule at zero pressure for the experiments at 20 degrees C and 2.5 A(2)/molecule for those at 25 degrees C. Similar results were obtained in KCl subphases. The apparent dissociation constant of the cholesterol-TTX complex calculated from dose-response experiments is 2.6 x 10(-7)M. Experiments at pH 10.1 revealed that the zwitter ionic form of TTX is less active. Experiments with cholesterol derivatives (cholesteryl acetate, cholesterol methyl ether, cholestanol, and cholestanyl acetate) indicate that for the interaction with TTX a partial negatively charged group at C-3 and a double bond between C-5 and C-6 on the steroid nucleus are required. Tetrodonic acid, a biologically inactive derivative of TTX, does not interact with cholesterol. The results lead us to propose that cholesterol is part of the Na(+) channel.

  17. Clostridium Perfringens Epsilon Toxin Binds to Membrane Lipids and Its Cytotoxic Action Depends on Sulfatide.

    Directory of Open Access Journals (Sweden)

    Carles Gil

    Full Text Available Epsilon toxin (Etx is one of the major lethal toxins produced by Clostridium perfringens types B and D, being the causal agent of fatal enterotoxemia in animals, mainly sheep and goats. Etx is synthesized as a non-active prototoxin form (proEtx that becomes active upon proteolytic activation. Etx exhibits a cytotoxic effect through the formation of a pore in the plasma membrane of selected cell targets where Etx specifically binds due to the presence of specific receptors. However, the identity and nature of host receptors of Etx remain a matter of controversy. In the present study, the interactions between Etx and membrane lipids from the synaptosome-enriched fraction from rat brain (P2 fraction and MDCK cell plasma membrane preparations were analyzed. Our findings show that both Etx and proEtx bind to lipids extracted from lipid rafts from the two different models as assessed by protein-lipid overlay assay. Lipid rafts are membrane microdomains enriched in cholesterol and sphingolipids. Binding of proEtx to sulfatide, phosphatidylserine, phosphatidylinositol (3-phosphate and phosphatidylinositol (5-phosphate was detected. Removal of the sulphate groups via sulfatase treatment led to a dramatic decrease in Etx-induced cytotoxicity, but not in proEtx-GFP binding to MDCK cells or a significant shift in oligomer formation, pointing to a role of sulfatide in pore formation in rafts but not in toxin binding to the target cell membrane. These results show for the first time the interaction between Etx and membrane lipids from host tissue and point to a major role for sulfatides in C. perfringens epsilon toxin pathophysiology.

  18. Lipid protrusions membrane softness, and enzymatic activity

    DEFF Research Database (Denmark)

    Jensen, Morten Østergaard; Høyrup, P.; Callisen, T.H.;

    2004-01-01

    The activity of phospholipase A(2) on lipid bilayers displays a characteristic lag burst behavior that has previously been shown to reflect the physical properties of the substrate. It has remained unclear which underlying molecular mechanism is responsible for this phenomenon. We propose here...... that protrusions of single lipid molecules out of the bilayer plane could provide such a mechanism. The proposal is supported by a combination of atomic-scale molecular dynamics simulations, theory, and experiments that have been performed in order to investigate the relationship between on the one side lipid...... protrusion modes and mechanical softness of phospholipid bilayers and on the other side the activity of enzymes acting on lipid bilayers composed of different unsaturated lipids. Specifically, our experiments show a correlation between the bilayer bending rigidity and the apparent Arrhenius activation energy...

  19. Concerted diffusion of lipids in raft-like membranes

    NARCIS (Netherlands)

    Apajalahti, Touko; Niemela, Perttu; Govindan, Praveen Nedumpully; Miettinen, Markus S.; Salonen, Emppu; Marrink, Siewert-Jan; Vattulainen, Ilpo

    2010-01-01

    Currently, there is no comprehensive model for the dynamics of cellular membranes. The understanding of even the basic dynamic processes, such as lateral diffusion of lipids, is still quite limited. Recent studies of one-component membrane systems have shown that instead of single-particle motions,

  20. Anthrax toxin-induced rupture of artificial lipid bilayer membranes

    Science.gov (United States)

    Nablo, Brian J.; Panchal, Rekha G.; Bavari, Sina; Nguyen, Tam L.; Gussio, Rick; Ribot, Wil; Friedlander, Art; Chabot, Donald; Reiner, Joseph E.; Robertson, Joseph W. F.; Balijepalli, Arvind; Halverson, Kelly M.; Kasianowicz, John J.

    2013-08-01

    We demonstrate experimentally that anthrax toxin complexes rupture artificial lipid bilayer membranes when isolated from the blood of infected animals. When the solution pH is temporally acidified to mimic that process in endosomes, recombinant anthrax toxin forms an irreversibly bound complex, which also destabilizes membranes. The results suggest an alternative mechanism for the translocation of anthrax toxin into the cytoplasm.

  1. The lipid bilayer membrane and its interactions with additives

    NARCIS (Netherlands)

    Meijer, L.A.

    1994-01-01

    The aim of this study was to make accurate predictions on the interaction of biologically relevant molecules with lipid bilayer membranes. We emphasised on the partitioning of these molecules between the membrane phase, and the aqueous phase quantified by the partition coefficient. To make

  2. The lipid bilayer membrane and its interactions with additives.

    NARCIS (Netherlands)

    Meijer, L.A.

    1994-01-01

    The aim of this study was to make accurate predictions on the interaction of biologically relevant molecules with lipid bilayer membranes. We emphasised on the partitioning of these molecules between the membrane phase, and the aqueous phase quantified by the partition coefficient. To make detailed

  3. Lipids, membrane proteins and natural membranes studied by neutron scattering and diffraction: A review

    Science.gov (United States)

    Zaccai, Giuseppe

    1986-02-01

    Diffraction first observed from myelin 50 years ago was correctly attributed to a fluid crystal of lipids, because similar patterns were observed from extracted lipid preparations. Following on more recent X-ray work which characterized a variety of lipid-water structures, neutron diffraction experiments have provided detailed descriptions of the molecular conformations in lipid bilayers. For a long time, however, the molecular structure of membrane proteins remained elusive and the development of detergents for the extraction of active membrane proteins, and the discovery of naturally crystalline purple membrane were important breakthroughs in this field. Structural parameters of membrane proteins solubilised in detergent have been measured by neutron scattering with contrast variation techniques. Purple membrane has been studied extensively by neutron diffraction. It is an excellent illustration of the use of deuterium labeling by different approaches to address specific questions of molecular structure. These studies are reviewed with a special emphasis on aspects which are applicable to membranes in general.

  4. Plasma membrane microdomains regulate turnover of transport proteins in yeast

    Science.gov (United States)

    Grossmann, Guido; Malinsky, Jan; Stahlschmidt, Wiebke; Loibl, Martin; Weig-Meckl, Ina; Frommer, Wolf B.; Opekarová, Miroslava; Tanner, Widmar

    2008-01-01

    In this study, we investigate whether the stable segregation of proteins and lipids within the yeast plasma membrane serves a particular biological function. We show that 21 proteins cluster within or associate with the ergosterol-rich membrane compartment of Can1 (MCC). However, proteins of the endocytic machinery are excluded from MCC. In a screen, we identified 28 genes affecting MCC appearance and found that genes involved in lipid biosynthesis and vesicle transport are significantly overrepresented. Deletion of Pil1, a component of eisosomes, or of Nce102, an integral membrane protein of MCC, results in the dissipation of all MCC markers. These deletion mutants also show accelerated endocytosis of MCC-resident permeases Can1 and Fur4. Our data suggest that release from MCC makes these proteins accessible to the endocytic machinery. Addition of arginine to wild-type cells leads to a similar redistribution and increased turnover of Can1. Thus, MCC represents a protective area within the plasma membrane to control turnover of transport proteins. PMID:19064668

  5. Effect of Sound Stimulation on the Lipid Physical States and Metabolism of Plasma Membrane from Chrysanthemum Callus%声波刺激对菊花愈伤组织膜脂物理状态和膜脂代谢的影响

    Institute of Scientific and Technical Information of China (English)

    赵虎成; 王伯初; 蔡绍皙; 席葆树

    2002-01-01

    以蔗糖梯度法纯化的菊花(Dendranthema morifolium (Ramat.) Tzvel.)愈伤组织质膜微囊为材料,研究了声波刺激下质膜膜脂代谢和物理状态的变化.结果表明, 一定强度(100 dB)和频率(1 000 Hz)的声波刺激使质膜磷酸二脂含量和二苯己三烯(DPH)荧光偏振值降低,质膜光散射值、MC540荧光强度和磷酸单脂含量增加.表明一定强度和频率的声波刺激使质膜变疏松,膜的流动性增加,膜表面电荷密度和疏水性降低,膜脂合成代谢增加,分解代谢减弱.由此可见,膜脂物理状态和膜脂代谢对声波刺激极为敏感.%Plasma membrane vesicles of chrysanthemum (Dendranthema morifolium (Ramat.) Tzvel.) callus was purified by sucrose gradient centrifugation to investigate the influence of sound stimulation on the lipid physical states and metabolism of plasma membrane. The results showed that sound stimulation decreased the content of phosphodiesters and the fluorescent intensity of DPH, but increased the light scattering value of the membrane, the fluorescent intensity of MC540 and the content of phosphomonoesters, indicating that the vesicles got looser, the charge density and hydrophobicity of membrane surface decreased under sound stimulation of some strength and frequency. However, the membrane fluidity increased under the condition. Meanwhile, the anabolism of membrane lipid increased and the catabolism decreased. It can be seen that the physical state and metabolism of membrane lipid is sensitive to sound stimulation.

  6. Tethered bilayer lipid membranes (tBLMs): interest and applications for biological membrane investigations.

    Science.gov (United States)

    Rebaud, Samuel; Maniti, Ofelia; Girard-Egrot, Agnès P

    2014-12-01

    Biological membranes play a central role in the biology of the cell. They are not only the hydrophobic barrier allowing separation between two water soluble compartments but also a supra-molecular entity that has vital structural functions. Notably, they are involved in many exchange processes between the outside and inside cellular spaces. Accounting for the complexity of cell membranes, reliable models are needed to acquire current knowledge of the molecular processes occurring in membranes. To simplify the investigation of lipid/protein interactions, the use of biomimetic membranes is an approach that allows manipulation of the lipid composition of specific domains and/or the protein composition, and the evaluation of the reciprocal effects. Since the middle of the 80's, lipid bilayer membranes have been constantly developed as models of biological membranes with the ultimate goal to reincorporate membrane proteins for their functional investigation. In this review, after a brief description of the planar lipid bilayers as biomimetic membrane models, we will focus on the construction of the tethered Bilayer Lipid Membranes, the most promising model for efficient membrane protein reconstitution and investigation of molecular processes occurring in cell membranes. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

  7. The adrenal specific toxicant mitotane directly interacts with lipid membranes and alters membrane properties depending on lipid composition.

    Science.gov (United States)

    Scheidt, Holger A; Haralampiev, Ivan; Theisgen, Stephan; Schirbel, Andreas; Sbiera, Silviu; Huster, Daniel; Kroiss, Matthias; Müller, Peter

    2016-06-15

    Mitotane (o,p'.-DDD) is an orphan drug approved for the treatment of adrenocortical carcinoma. The mechanisms, which are responsible for this activity of the drug, are not completely understood. It can be hypothesized that an impact of mitotane is mediated by the interaction with cellular membranes. However, an interaction of mitotane with (lipid) membranes has not yet been investigated in detail. Here, we characterized the interaction of mitotane and its main metabolite o,p'-dichlorodiphenyldichloroacetic acid (o,p'-DDA) with lipid membranes by applying a variety of biophysical approaches of nuclear magnetic resonance, electron spin resonance, and fluorescence spectroscopy. We found that mitotane and o,p'-DDA bind to lipid membranes by inserting into the lipid-water interface of the bilayer. Mitotane but not o,p'-DDA directly causes a disturbance of bilayer structure leading to an increased permeability of the membrane for polar molecules. Mitotane induced alterations of the membrane integrity required the presence of phosphatidylethanolamine and/or cholesterol. Collectively, our data for the first time characterize the impact of mitotane on the lipid membrane structure and dynamics, which may contribute to a better understanding of specific mitotane effects and side effects.

  8. Stabilization of concentration fluctuations in mixed membranes by hybrid lipids

    Science.gov (United States)

    Palmieri, Benoit; Safran, Samuel

    2012-02-01

    Finite-size domains have been observed at the surface of cells. These lipids ``rafts'' are stable nanodomains enriched in saturated lipids and cholesterol. While line tension favors macrodomains, one explanation for raft stabilization suggests that the membrane composition is tuned close to a spinodal temperature. From this point of view, rafts are long-lived concentration fluctuations in the mixed phase. We propose a ternary mixture model for the cell membrane that includes hybrid lipids which have one saturated and one unsaturated hydrocarbon chain. Finite amount of hybrid lipids reduces the packing incompatibility at the saturated/unsaturated lipid interface and stabilizes the concentration fluctuations. Hybrid-Hybrid interactions are included in the model and further increase the life-time of the rafts and decrease their length-scales. Moreover, the hybrid has extra orientational degrees of freedom that may lead to modulated phases.

  9. Phase state dependent current fluctuations in pure lipid membranes

    CERN Document Server

    Wunderlich, B; Idzko, A-L; Keyser, U F; Wixforth, A; Myles, V M; Heimburg, T; Schneider, M F

    2009-01-01

    Current fluctuations in pure lipid membranes have been shown to occur under the influence of transmembrane electric fields (electroporation) as well as a result from structural rearrangements of the lipid bilayer during phase transition (soft perforation). We demonstrate that the ion permeability during lipid phase transition exhibits the same qualitative temperature dependence as the macroscopic heat capacity of a D15PC/DOPC vesicle suspension. Microscopic current fluctuations show distinct characteristics for each individual phase state. While current fluctuations in the fluid phase show spike-like behaviour of short time scales (~ 2ms) with a narrow amplitude distribution, the current fluctuations during lipid phase transition appear in distinct steps with time scales in the order of ~ 20ms. 1 We propose a theoretical explanation for the origin of time scales and permeability based on a linear relationship between lipid membrane susceptibilities and relaxation times in the vicinity of the phase transition.

  10. A reusable device for electrochemical applications of hydrogel supported black lipid membranes

    DEFF Research Database (Denmark)

    Mech-Dorosz, Agnieszka; Heiskanen, Arto; Bäckström, Sania

    2015-01-01

    Black lipid membranes (BLMs) are significant in studies of membrane transport, incorporated proteins/ion transporters, and hence in construction of biosensor devices. Although BLMs provide an accepted mimic of cellular membranes, they are inherently fragile. Techniques are developed to stabilize...... of the gold electrode microchips and plasma modification of the ETFE aperture arrays facilitating covalent "sandwiching" of the hydrogel. Both fluorescence microscopy and EIS were used to demonstrate the induced spontaneous thinning of a deposited lipid solution, leading to formation of stabilized hs......BLMs on average in 10 min. The determined specific membrane capacitance and resistance were shown to vary in the range 0.31-0.49 mu F/cm2 and 45-65 k Omega cm2, respectively, corresponding to partially solvent containing BLMs with an average life time of 60-80 min. The characterized hsBLM formation and devised...

  11. Edelfosine and miltefosine effects on lipid raft properties: membrane biophysics in cell death by antitumor lipids.

    Science.gov (United States)

    Castro, Bruno M; Fedorov, Aleksander; Hornillos, Valentin; Delgado, Javier; Acuña, A Ulises; Mollinedo, Faustino; Prieto, Manuel

    2013-07-03

    Edelfosine (1-O-octadecyl-2-O-methyl-sn-glycero-phosphocholine) and miltefosine (hexadecylphosphocholine) are synthetic alkylphospholipids (ALPs) that are reported to selectively accumulate in tumor cell membranes, inducing Fas clustering and activation on lipid rafts, triggering apoptosis. However, the exact mechanism by which these lipids elicit these events is still not fully understood. Recent studies propose that their mode of action might be related with alterations of lipid rafts biophysical properties caused by these lipid drugs. To achieve a clear understanding of this mechanism, we studied the effects of pharmacologically relevant amounts of edelfosine and miltefosine in the properties of model and cellular membranes. The influence of these molecules on membrane order, lateral organization, and lipid rafts molar fraction and size were studied by steady-state and time-resolved fluorescence methods, Förster resonance energy transfer (FRET), confocal and fluorescence lifetime imaging microscopy (FLIM). We found that the global membrane and lipid rafts biophysical properties of both model and cellular membranes were not significantly affected by both the ALPs. Nonetheless, in model membranes, a mild increase in membrane fluidity induced by both alkyl lipids was detected, although this effect was more noticeable for edelfosine than miltefosine. This absence of drastic alterations shows for the first time that ALPs mode of action is unlikely to be directly linked to alterations of lipid rafts biophysical properties caused by these drugs. The biological implications of this result are discussed in the context of ALPs effects on lipid metabolism, mitochondria homeostasis modulation, and their relationship with tumor cell death.

  12. Construction of higher-ordered monolayer membranes derived from archaeal membrane lipid-inspired cyclic lipids with longer alkyl chains.

    Science.gov (United States)

    Nakamura, Makoto; Goto, Rie; Tadokoro, Toshio; Shibakami, Motonari

    2007-06-15

    A series of artificial cyclic lipids that mimic archaeal membrane ones has been synthesized. The structural features of these molecules include a longer cyclic framework, in which the alkyl chain length ranges from 24 to 32 in carbon number, which is longer than our first analogous molecule with 20-carbon long alkyl chains [K. Miyawaki, T. Takagi, M. Shibakami, Synlett 8 (2002) 1326]. Microscopic observation reveals that these molecules have a self-assembling ability: hydration of the lipids yields multilamellar vesicles in aqueous solution and monolayer sheets on solid supports. High-sensitivity differential scanning calorimetry (24- and 28-carbon alkyl chain lipids) indicates that (i) the alkyl chain length affects their phase behavior and (ii) the enthalpies of endothermic peaks accompanied by phase transition were considerably lower than those of their monomeric phospholipid analogs. Fluorescence polarization measurements suggest that the membranes made from the 24-carbon alkyl chain lipid have a higher polarization factor than membranes composed of DMPC and DMPC plus cholesterol. These findings imply that the cyclic lipids containing 24- and 28-carbon alkyl chain construct well-organized monolayer membranes and, in particular, that the molecular order of the 24-carbon alkyl chain lipid is higher than that of bilayer membranes in the liquid-ordered phase.

  13. Towards structural and functional analysis of the plant plasma membrane proton pump

    DEFF Research Database (Denmark)

    Justesen, Bo Højen

    of plasma membrane H+-ATPases. Studies on the plasma membrane H+-ATPases have involved both in vivo and in vitro approaches, with the latter employing either solubilisation by detergent micelles, or reconstitution into lipid vesicles. Despite resulting in a large body of information on structure, function...... into soluble nanoscale lipid bilayers, also termed nanodiscs. Extensive analysis confirms the correct assembly and reconstitution of active proton pump into nanodiscs. The pump inserts as a monomer, which through activity analysis confirms this as the minimal functional unit of the plasma membrane H......+-ATPase. Reconstitution of the H+-ATPase into nanodiscs has the potential to enable structural and functional characterization using various techniques, exemplified by the specific immobilization of reconstituted proton pump using surface plasma resonance. The ability to efficiently separate empty from membrane protein...

  14. Adhesion and receptor clustering stabilizes lateral heterogeneity in cell plasma membranes

    Science.gov (United States)

    Veatch, Sarah

    2013-03-01

    The thermodynamic properties of plasma membrane lipids play a vital role in many functions that initiate at the mammalian cell surface. Some functions are thought to occur, at least in part, because plasma membrane lipids have a tendency to separate into two distinct liquid phases, called liquid-ordered and liquid-disordered. We find that isolated cell plasma membranes are poised near a miscibility critical point separating these two liquid phases, and postulate that critical composition fluctuations provide the physical basis of functional membrane heterogeneity in intact cells. In this talk I will describe several possible mechanisms through which dynamic fluctuations can be stabilized in super-critical membranes, and will present some preliminary evidence suggesting that these structures can be visualized in intact cells using quantitative super-resolution fluorescence localization imaging.

  15. Ceramides in the skin lipid membranes: length matters.

    Science.gov (United States)

    Skolová, Barbora; Janůšová, Barbora; Zbytovská, Jarmila; Gooris, Gert; Bouwstra, Joke; Slepička, Petr; Berka, Pavel; Roh, Jaroslav; Palát, Karel; Hrabálek, Alexandr; Vávrová, Kateřina

    2013-12-17

    Ceramides are essential constituents of the skin barrier that allow humans to live on dry land. Reduced levels of ceramides have been associated with skin diseases, e.g., atopic dermatitis. However, the structural requirements and mechanisms of action of ceramides are not fully understood. Here, we report the effects of ceramide acyl chain length on the permeabilities and biophysics of lipid membranes composed of ceramides (or free sphingosine), fatty acids, cholesterol, and cholesterol sulfate. Short-chain ceramides increased the permeability of the lipid membranes compared to a long-chain ceramide with maxima at 4-6 carbons in the acyl. By a combination of differential scanning calorimetry, Fourier transform infrared spectroscopy, X-ray diffraction, Langmuir monolayers, and atomic force microscopy, we found that the reason for this effect in short ceramides was a lower proportion of tight orthorhombic packing and phase separation of continuous short ceramide-enriched domains with shorter lamellar periodicity compared to native long ceramides. Thus, long acyl chains in ceramides are essential for the formation of tightly packed impermeable lipid lamellae. Moreover, the model skin lipid membranes are a valuable tool to study the relationships between the lipid structure and composition, lipid organization, and the membrane permeability.

  16. Simulation modeling of supported lipid membranes - a review.

    Science.gov (United States)

    Hirtz, Michael; Kumar, Naresh; Chi, Lifeng

    2014-03-01

    Lipid membranes are of great importance for many biological systems and biotechnological applications. One method to gain a profound understanding of the dynamics in lipid membranes and their interaction with other system components is by modeling these systems by computer simulations. Many different approaches have been undertaken in this endeavor that have led to molecular level insights into the underlying mechanisms of several experimental observations and biological processes with an extremely high temporal resolution. As compared to the free-standing lipid bilayers, there are fewer simulation studies addressing the systems of supported lipid membranes. Nevertheless, these have significantly enhanced our understanding of the behavior of lipid layers employed in applications spanning from biosensors to drug delivery and for biological processes such as the breathing cycle of lung surfactants. In this review, we give an account of the state of the art of methods and applications of the simulations of supported lipid bilayers, interfacial membranes at the air/water interface and on solid surfaces.

  17. Alterations in plasma lipid profile patterns in head and neck cancer and oral precancerous conditions

    Directory of Open Access Journals (Sweden)

    Patel Prabhudas

    2004-01-01

    Full Text Available BACKGROUND : The changes in lipid profile have long been associated with cancer because lipids play a key role in maintenance of cell integrity. AIMS : The present study evaluated alterations in plasma lipid profile in untreated head and neck cancer patients as well as patients with oral precancerous conditions (OPC and its association with habit of tobacco consumption. MATERIAL AND METHODS : This hospital-based case control study included 184 head and neck cancer patients, 153 patients with OPC and 52 controls. Plasma lipids including: (i Total cholesterol, (ii LDL cholesterol (LDLC, (iii HDL cholesterol (HDLC (iv VLDL cholesterol (VLDLC and (v triglycerides were analysed by spectrophotometric kits. STATISTICAL ANALYSIS USED : Student′s t-test was performed to compare mean values of the parameters. RESULTS : A significant decrease in plasma total cholesterol and HDLC was observed in cancer patients (P=0.008 and P=0.000 respectively as well as in patients with OPC (P=0.014 and P=0.000, respectively as compared to the controls. The plasma VLDL and triglycerides levels were significantly lower in cancer patients as compared to the patients with OPC (P=0.04 and controls (P=0.059. The tobacco habituates showed lower plasma lipid levels than the non-habituates. Our data strengthen the evidence of an inverse relationship between plasma lipid levels and head and neck malignancies as well as OPC. CONCLUSION :The lower levels of plasma cholesterol and other lipid constituents in patients might be due to their increased utilization by neoplastic cells for new membrane biogenesis. The findings strongly warrant an in-depth study of alterations in plasma lipid profile in head neck cancer patients.

  18. Biosynthesis of archaeal membrane ether lipids

    NARCIS (Netherlands)

    Jain, Samta; Caforio, Antonella; Driessen, Arnold J. M.

    2014-01-01

    A vital function of the cell membrane in all living organism is to maintain the membrane permeability barrier and fluidity. The composition of the phospholipid bilayer is distinct in archaea when compared to bacteria and eukarya. In archaea, isoprenoid hydrocarbon side chains are linked via an ether

  19. Interleaflet Coupling, Pinning, and Leaflet Asymmetry—Major Players in Plasma Membrane Nanodomain Formation

    Science.gov (United States)

    Fujimoto, Toyoshi; Parmryd, Ingela

    2017-01-01

    The plasma membrane has a highly asymmetric distribution of lipids and contains dynamic nanodomains many of which are liquid entities surrounded by a second, slightly different, liquid environment. Contributing to the dynamics is a continuous repartitioning of components between the two types of liquids and transient links between lipids and proteins, both to extracellular matrix and cytoplasmic components, that temporarily pin membrane constituents. This make plasma membrane nanodomains exceptionally challenging to study and much of what is known about membrane domains has been deduced from studies on model membranes at equilibrium. However, living cells are by definition not at equilibrium and lipids are distributed asymmetrically with inositol phospholipids, phosphatidylethanolamines and phosphatidylserines confined mostly to the inner leaflet and glyco- and sphingolipids to the outer leaflet. Moreover, each phospholipid group encompasses a wealth of species with different acyl chain combinations whose lateral distribution is heterogeneous. It is becoming increasingly clear that asymmetry and pinning play important roles in plasma membrane nanodomain formation and coupling between the two lipid monolayers. How asymmetry, pinning, and interdigitation contribute to the plasma membrane organization is only beginning to be unraveled and here we discuss their roles and interdependence. PMID:28119914

  20. Inducing morphological changes in lipid bilayer membranes with microfabricated substrates

    Science.gov (United States)

    Liu, Fangjie; Collins, Liam F.; Ashkar, Rana; Heberle, Frederick A.; Srijanto, Bernadeta R.; Collier, C. Patrick

    2016-11-01

    Lateral organization of lipids and proteins into distinct domains and anchoring to a cytoskeleton are two important strategies employed by biological membranes to carry out many cellular functions. However, these interactions are difficult to emulate with model systems. Here we use the physical architecture of substrates consisting of arrays of micropillars to systematically control the behavior of supported lipid bilayers - an important step in engineering model lipid membrane systems with well-defined functionalities. Competition between attractive interactions of supported lipid bilayers with the underlying substrate versus the energy cost associated with membrane bending at pillar edges can be systematically investigated as functions of pillar height and pitch, chemical functionalization of the microstructured substrate, and the type of unilamellar vesicles used for assembling the supported bilayer. Confocal fluorescent imaging and AFM measurements highlight correlations that exist between topological and mechanical properties of lipid bilayers and lateral lipid mobility in these confined environments. This study provides a baseline for future investigations into lipid domain reorganization on structured solid surfaces and scaffolds for cell growth.

  1. Laurdan fluorescence senses mechanical strain in the lipid bilayer membrane.

    Science.gov (United States)

    Zhang, Yan-Liang; Frangos, John A; Chachisvilis, Mirianas

    2006-09-01

    The precise molecular mechanisms by which cells transduce a mechanical stimulus into an intracellular biochemical response have not yet been established. Here, we show for the first time that the fluorescence emission of an environment-sensitive membrane probe Laurdan is modulated by mechanical strain of the lipid bilayer membrane. We have measured fluorescence emission of Laurdan in phospholipid vesicles of 30, 50, and 100 nm diameter to show that osmotically induced membrane tension leads to an increase in polarity (hydration depth) of the phospholipid bilayer interior. Our data indicate that the general polarization of Laurdan emission is linearly dependent on membrane tension. We also show that higher membrane curvature leads to higher hydration levels. We anticipate that the proposed method will facilitate future studies of mechanically induced changes in physical properties of lipid bilayer environment both in vitro and in vivo.

  2. Lipid nanotechnologies for structural studies of membrane-associated proteins.

    Science.gov (United States)

    Stoilova-McPhie, Svetla; Grushin, Kirill; Dalm, Daniela; Miller, Jaimy

    2014-11-01

    We present a methodology of lipid nanotubes (LNT) and nanodisks technologies optimized in our laboratory for structural studies of membrane-associated proteins at close to physiological conditions. The application of these lipid nanotechnologies for structure determination by cryo-electron microscopy (cryo-EM) is fundamental for understanding and modulating their function. The LNTs in our studies are single bilayer galactosylceramide based nanotubes of ∼20 nm inner diameter and a few microns in length, that self-assemble in aqueous solutions. The lipid nanodisks (NDs) are self-assembled discoid lipid bilayers of ∼10 nm diameter, which are stabilized in aqueous solutions by a belt of amphipathic helical scaffold proteins. By combining LNT and ND technologies, we can examine structurally how the membrane curvature and lipid composition modulates the function of the membrane-associated proteins. As proof of principle, we have engineered these lipid nanotechnologies to mimic the activated platelet's phosphtaidylserine rich membrane and have successfully assembled functional membrane-bound coagulation factor VIII in vitro for structure determination by cryo-EM. The macromolecular organization of the proteins bound to ND and LNT are further defined by fitting the known atomic structures within the calculated three-dimensional maps. The combination of LNT and ND technologies offers a means to control the design and assembly of a wide range of functional membrane-associated proteins and complexes for structural studies by cryo-EM. The presented results confirm the suitability of the developed methodology for studying the functional structure of membrane-associated proteins, such as the coagulation factors, at a close to physiological environment. © 2014 Wiley Periodicals, Inc.

  3. Sphingolipid Organization in the Plasma Membrane and the Mechanisms That Influence It.

    Science.gov (United States)

    Kraft, Mary L

    2016-01-01

    Sphingolipids are structural components in the plasma membranes of eukaryotic cells. Their metabolism produces bioactive signaling molecules that modulate fundamental cellular processes. The segregation of sphingolipids into distinct membrane domains is likely essential for cellular function. This review presents the early studies of sphingolipid distribution in the plasma membranes of mammalian cells that shaped the most popular current model of plasma membrane organization. The results of traditional imaging studies of sphingolipid distribution in stimulated and resting cells are described. These data are compared with recent results obtained with advanced imaging techniques, including super-resolution fluorescence detection and high-resolution secondary ion mass spectrometry (SIMS). Emphasis is placed on the new insight into the sphingolipid organization within the plasma membrane that has resulted from the direct imaging of stable isotope-labeled lipids in actual cell membranes with high-resolution SIMS. Super-resolution fluorescence techniques have recently revealed the biophysical behaviors of sphingolipids and the unhindered diffusion of cholesterol analogs in the membranes of living cells are ultimately in contrast to the prevailing hypothetical model of plasma membrane organization. High-resolution SIMS studies also conflicted with the prevailing hypothesis, showing sphingolipids are concentrated in micrometer-scale membrane domains, but cholesterol is evenly distributed within the plasma membrane. Reductions in cellular cholesterol decreased the number of sphingolipid domains in the plasma membrane, whereas disruption of the cytoskeleton eliminated them. In addition, hemagglutinin, a transmembrane protein that is thought to be a putative raft marker, did not cluster within sphingolipid-enriched regions in the plasma membrane. Thus, sphingolipid distribution in the plasma membrane is dependent on the cytoskeleton, but not on favorable interactions with

  4. Hybrid and nonhybrid lipids exert common effects on membrane raft size and morphology.

    Science.gov (United States)

    Heberle, Frederick A; Doktorova, Milka; Goh, Shih Lin; Standaert, Robert F; Katsaras, John; Feigenson, Gerald W

    2013-10-01

    Nanometer-scale domains in cholesterol-rich model membranes emulate lipid rafts in cell plasma membranes (PMs). The physicochemical mechanisms that maintain a finite, small domain size are, however, not well understood. A special role has been postulated for chain-asymmetric or hybrid lipids having a saturated sn-1 chain and an unsaturated sn-2 chain. Hybrid lipids generate nanodomains in some model membranes and are also abundant in the PM. It was proposed that they align in a preferred orientation at the boundary of ordered and disordered phases, lowering the interfacial energy and thus reducing domain size. We used small-angle neutron scattering and fluorescence techniques to detect nanoscopic and modulated liquid phase domains in a mixture composed entirely of nonhybrid lipids and cholesterol. Our results are indistinguishable from those obtained previously for mixtures containing hybrid lipids, conclusively showing that hybrid lipids are not required for the formation of nanoscopic liquid domains and strongly implying a common mechanism for the overall control of raft size and morphology. We discuss implications of these findings for theoretical descriptions of nanodomains.

  5. Structural determinants of protein partitioning into ordered membrane domains and lipid rafts.

    Science.gov (United States)

    Lorent, Joseph Helmuth; Levental, Ilya

    2015-11-01

    Increasing evidence supports the existence of lateral nanoscopic lipid domains in plasma membranes, known as lipid rafts. These domains preferentially recruit membrane proteins and lipids to facilitate their interactions and thereby regulate transmembrane signaling and cellular homeostasis. The functionality of raft domains is intrinsically dependent on their selectivity for specific membrane components; however, while the physicochemical determinants of raft association for lipids are known, very few systematic studies have focused on the structural aspects that guide raft partitioning of proteins. In this review, we describe biophysical and thermodynamic aspects of raft-mimetic liquid ordered phases, focusing on those most relevant for protein partitioning. Further, we detail the variety of experimental models used to study protein-raft interactions. Finally, we review the existing literature on mechanisms for raft targeting, including lipid post-translational modifications, lipid binding, and transmembrane domain features. We conclude that while protein palmitoylation is a clear raft-targeting signal, few other general structural determinants for raft partitioning have been revealed, suggesting that many discoveries lie ahead in this burgeoning field.

  6. Hybrid and Nonhybrid Lipids Exert Common Effects on Membrane Raft Size and Morphology

    Energy Technology Data Exchange (ETDEWEB)

    Heberle, Frederick A [ORNL; Doktorova, Milka [Cornell University; Goh, Shih Lin [Cornell University; Standaert, Robert F [ORNL; Katsaras, John [ORNL; Feigenson, Gerald [Cornell University

    2013-01-01

    Nanometer-scale domains in cholesterolrich model membranes emulate lipid rafts in cell plasma membranes (PMs). The physicochemical mechanisms that maintain a finite, small domain size are, however, not well understood. A special role has been postulated for chainasymmetric or hybrid lipids having a saturated sn-1 chain and an unsaturated sn-2 chain. Hybrid lipids generate nanodomains in some model membranes and are also abundant in the PM. It was proposed that they align in a preferred orientation at the boundary of ordered and disordered phases, lowering the interfacial energy and thus reducing domain size. We used small-angle neutron scattering and fluorescence techniques to detect nanoscopic and modulated liquid phase domains in a mixture composed entirely of nonhybrid lipids and cholesterol. Our results are indistinguishable from those obtained previously for mixtures containing hybrid lipids, conclusively showing that hybrid lipids are not required for the formation of nanoscopic liquid domains and strongly implying a common mechanism for the overall control of raft size and morphology. We discuss implications of these findings for theoretical descriptions of nanodomains.

  7. DNA release from lipoplexes by anionic lipids: correlation with lipid mesomorphism, interfacial curvature, and membrane fusion

    Energy Technology Data Exchange (ETDEWEB)

    Tarahovsky, Yury S.; Koynova, Rumiana; MacDonald, Robert C. (Northwestern)

    2010-01-18

    DNA release from lipoplexes is an essential step during lipofection and is probably a result of charge neutralization by cellular anionic lipids. As a model system to test this possibility, fluorescence resonance energy transfer between DNA and lipid covalently labeled with Cy3 and BODIPY, respectively, was used to monitor the release of DNA from lipid surfaces induced by anionic liposomes. The separation of DNA from lipid measured this way was considerably slower and less complete than that estimated with noncovalently labeled DNA, and depends on the lipid composition of both lipoplexes and anionic liposomes. This result was confirmed by centrifugal separation of released DNA and lipid. X-ray diffraction revealed a clear correlation of the DNA release capacity of the anionic lipids with the interfacial curvature of the mesomorphic structures developed when the anionic and cationic liposomes were mixed. DNA release also correlated with the rate of fusion of anionic liposomes with lipoplexes. It is concluded that the tendency to fuse and the phase preference of the mixed lipid membranes are key factors for the rate and extent of DNA release. The approach presented emphasizes the importance of the lipid composition of both lipoplexes and target membranes and suggests optimal transfection may be obtained by tailoring lipoplex composition to the lipid composition of target cells.

  8. Modeling the Elastic Properties of Lipid Bilayer Membranes

    Science.gov (United States)

    Barry, Edward; Gibaud, Thomas; Zakhary, Mark; Dogic, Zvonimir

    2011-03-01

    Model membranes such as lipid bilayers have been indispensable tools for our understanding of the elastic properties of biological membranes. In this talk, I will introduce a colloidal model for membranes and demonstrate that the physical properties of these colloidal membranes are identical to lipid bilayers. The model system is unique in that the constituent molecules are homogenous and non-amphiphilic, yet their self-assembly into membranes and other hierarchical assemblages, such as a lamellar type phases and chiral ribbons, proceeds spontaneously in solution. Owing to the large size of the constituent molecules, individual molecules can be directly visualized and simultaneous observations at the continuum and molecular lengthscales are used to characterize the behavior of model membranes with unprecedented detail. Moreover, once assembled in solution, molecular interactions can be controlled in situ. In particular, the strength of chiral interactions can be varied, leading to fascinating transitions in behavior that resembles the formation of starfish vesicles. These observations point towards the important role of line tension, and have potential implications for phase separated lipid mixtures or lipid rafts.

  9. EDTA-induced membrane fluidization and destabilization: biophysical studies on artificial lipid membranes.

    Science.gov (United States)

    Prachayasittikul, Virapong; Isarankura-Na-Ayudhya, Chartchalerm; Tantimongcolwat, Tanawut; Nantasenamat, Chanin; Galla, Hans-Joachim

    2007-11-01

    The molecular mechanism of ethylenediaminetetraacetic acid (EDTA)-induced membrane destabilization has been studied using a combination of four biophysical techniques on artificial lipid membranes. Data from Langmuir film balance and epifluorescence microscopy revealed the fluidization and expansion effect of EDTA on phase behavior of monolayers of either 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) or mixtures of DPPC and metal-chelating lipids, such as N(alpha),N(alpha)-Bis[carboxymethyl]-N(epsilon)-[(dioctadecylamino)succinyl]-L-lysine or 1,2-dioleoyl-sn-glycero-3-[N-(5-amino-1-carboxypentyl iminodiacetic acid) succinyl]. A plausible explanation could be drawn from the electrostatic interaction between negatively charged groups of EDTA and the positively charged choline head group of DPPC. Intercalation of EDTA into the lipid membrane induced membrane curvature as elucidated by atomic force microscopy. Growth in size and shape of the membrane protrusion was found to be time-dependent upon exposure to EDTA. Further loss of material from the lipid membrane surface was monitored in real time using a quartz crystal microbalance. This indicates membrane restabilization by exclusion of the protrusions from the surface. Loss of lipid components facilitates membrane instability, leading to membrane permeabilization and lysis.

  10. EDTA-induced Membrane Fluidization and Destabilization: Biophysical Studies on Artificial Lipid Membranes

    Institute of Scientific and Technical Information of China (English)

    Virapong PRACHAYASITTIKUL; Chartchalerm ISARANKURA-NA-AYUDHYA; Tanawut TANTIMONGCOLWAT; Chanin NANTASENAMAT; Hans-Joachim GALLA

    2007-01-01

    The molecular mechanism of ethylenediaminetetraacetic acid (EDTA)-induced membrane destabilization has been studied using a combination of four biophysical techniques on artificial lipid membranes.Data from Langmuir film balance and epifluorescence microscopy revealed the fluidization and expansion effect of EDTA on phase behavior of monolayers of either 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) or mixtures of DPPC and metal-chelating lipids, such as Nα,Nα-Bis[carboxymethyl]-Nε-[(dioctadecylamino)succinyl]-L-lysine or 1,2-dioleoyl-sn-glycero-3-[N-(5-amino- 1-carboxypentyl iminodiacetic acid) succinyl]. A plausible explanation could be drawn from the electrostatic interaction between negatively charged groups of EDTA and the positively charged choline head group of DPPC.Intercalation of EDTA into the lipid membrane induced membrane curvature as elucidated by atomic force microscopy. Growth in size and shape of the membrane protrusion was found to be time-dependent upon exposure to EDTA. Further loss of material from the lipid membrane surface was monitored in real time using a quartz crystal microbalance. This indicates membrane restabilization by exclusion of the protrusions from the surface. Loss of lipid components facilitates membrane instability, leading to membrane permeabilization and lysis.

  11. Acid sphingomyelinase activity is regulated by membrane lipids and facilitates cholesterol transfer by NPC2.

    Science.gov (United States)

    Oninla, Vincent O; Breiden, Bernadette; Babalola, Jonathan O; Sandhoff, Konrad

    2014-12-01

    During endocytosis, membrane components move to intraluminal vesicles of the endolysosomal compartment for digestion. At the late endosomes, cholesterol is sorted out mainly by two sterol-binding proteins, Niemann-Pick protein type C (NPC)1 and NPC2. To study the NPC2-mediated intervesicular cholesterol transfer, we developed a liposomal assay system. (Abdul-Hammed, M., B. Breiden, M. A. Adebayo, J. O. Babalola, G. Schwarzmann, and K. Sandhoff. 2010. Role of endosomal membrane lipids and NPC2 in cholesterol transfer and membrane fusion. J. Lipid Res. 51: 1747-1760.) Anionic lipids stimulate cholesterol transfer between liposomes while SM inhibits it, even in the presence of anionic bis(monoacylglycero)phosphate (BMP). Preincubation of vesicles containing SM with acid sphingomyelinase (ASM) (SM phosphodiesterase, EC 3.1.4.12) results in hydrolysis of SM to ceramide (Cer), which enhances cholesterol transfer. Besides SM, ASM also cleaves liposomal phosphatidylcholine. Anionic phospholipids derived from the plasma membrane (phosphatidylglycerol and phosphatidic acid) stimulate SM and phosphatidylcholine hydrolysis by ASM more effectively than BMP, which is generated during endocytosis. ASM-mediated hydrolysis of liposomal SM was also stimulated by incorporation of diacylglycerol (DAG), Cer, and free fatty acids into the liposomal membranes. Conversely, phosphatidylcholine hydrolysis was inhibited by incorporation of cholesterol, Cer, DAG, monoacylglycerol, and fatty acids. Our data suggest that SM degradation by ASM is required for physiological secretion of cholesterol from the late endosomal compartment, and is a key regulator of endolysosomal lipid digestion.

  12. The surface charge of a cell lipid membrane

    CERN Document Server

    Pekker, M

    2014-01-01

    In this paper the problem of surface charge of the lipid membrane is considered. It is shown that the membrane surface is negatively charged. Negative ions are in potential wells formed by the dipole heads of membrane phospholipids. The binding energy of the ion with the membrane surface is much greater than its thermal energy. A self-consistent model of the potential in solution is developed, and a stationary charge density on the membrane surface is found. The estimates given in the paper show that the potential difference across the membrane of the unexcited axon (resting potential) can be explained by the difference in surface densities of the bound charges on the inner and outer surfaces of the membrane.

  13. Pressure effects on lipids and bio-membrane assemblies

    Directory of Open Access Journals (Sweden)

    Nicholas J. Brooks

    2014-11-01

    Full Text Available Membranes are amongst the most important biological structures; they maintain the fundamental integrity of cells, compartmentalize regions within them and play an active role in a wide range of cellular processes. Pressure can play a key role in probing the structure and dynamics of membrane assemblies, and is also critical to the biology and adaptation of deep-sea organisms. This article presents an overview of the effect of pressure on the mesostructure of lipid membranes, bilayer organization and lipid–protein assemblies. It also summarizes recent developments in high-pressure structural instrumentation suitable for experiments on membranes.

  14. Organic and inorganic osmolytes at lipid membrane interfaces

    DEFF Research Database (Denmark)

    Westh, P.; Peters, Günther H.j.

    2008-01-01

    on membrane–osmolyte interactions. The physical properties of lipid membranes are strongly affected by the presence of “foreign molecules”—that is, components other than water and lipids—at the interface. In many cases, the solute-induced perturbations are rather complex and cannot be rationalized......This chapter discusses the interactions of organic osmolytes and membranous interfaces, and the effects of these interactions on the properties of the membrane. It also includes a treatment of inorganic ions at the membrane interface since osmolyte effects involve a balance between organic...

  15. HAMLET interacts with lipid membranes and perturbs their structure and integrity.

    Science.gov (United States)

    Mossberg, Ann-Kristin; Puchades, Maja; Halskau, Øyvind; Baumann, Anne; Lanekoff, Ingela; Chao, Yinxia; Martinez, Aurora; Svanborg, Catharina; Karlsson, Roger

    2010-02-23

    Cell membrane interactions rely on lipid bilayer constituents and molecules inserted within the membrane, including specific receptors. HAMLET (human alpha-lactalbumin made lethal to tumor cells) is a tumoricidal complex of partially unfolded alpha-lactalbumin (HLA) and oleic acid that is internalized by tumor cells, suggesting that interactions with the phospholipid bilayer and/or specific receptors may be essential for the tumoricidal effect. This study examined whether HAMLET interacts with artificial membranes and alters membrane structure. We show by surface plasmon resonance that HAMLET binds with high affinity to surface adherent, unilamellar vesicles of lipids with varying acyl chain composition and net charge. Fluorescence imaging revealed that HAMLET accumulates in membranes of vesicles and perturbs their structure, resulting in increased membrane fluidity. Furthermore, HAMLET disrupted membrane integrity at neutral pH and physiological conditions, as shown by fluorophore leakage experiments. These effects did not occur with either native HLA or a constitutively unfolded Cys-Ala HLA mutant (rHLA(all-Ala)). HAMLET also bound to plasma membrane vesicles formed from intact tumor cells, with accumulation in certain membrane areas, but the complex was not internalized by these vesicles or by the synthetic membrane vesicles. The results illustrate the difference in membrane affinity between the fatty acid bound and fatty acid free forms of partially unfolded HLA and suggest that HAMLET engages membranes by a mechanism requiring both the protein and the fatty acid. Furthermore, HAMLET binding alters the morphology of the membrane and compromises its integrity, suggesting that membrane perturbation could be an initial step in inducing cell death.

  16. Photon correlation spectroscopy of bilayer lipid membranes.

    Science.gov (United States)

    Crilly, J F; Earnshaw, J C

    1983-02-01

    Light scattering by thermal fluctuations on simple monoglyceride bilayer membranes has been used to investigate the viscoelastic properties of these structures. Spectroscopic analysis of these fluctuations (capillary waves) permits the nonperturbative measurement of the interfacial tension and a shear interfacial viscosity acting normal to the membrane plane. The methods were established by studies of solvent and nonsolvent bilayers of glycerol monooleate (GMO). Changes in the tension of GMO/n-decane membranes induced by altering the composition of the parent solution were detected and quantified. In a test of the reliability of the technique controlled variations of the viscosity of the aqueous bathing solution were accurately monitored. The technique was applied to solvent-free bilayers formed from dispersions of GMO in squalane. The lower tensions observed attested to the comparative absence of solvent in such bilayers. In contrast to the solvent case, the solvent-free membranes exhibited a significant transverse shear viscosity, indicative of the enhanced intermolecular interactions within the bilayer.

  17. Isolation and characterization of the plasma membrane from the yeast Pichia pastoris.

    Science.gov (United States)

    Grillitsch, Karlheinz; Tarazona, Pablo; Klug, Lisa; Wriessnegger, Tamara; Zellnig, Günther; Leitner, Erich; Feussner, Ivo; Daum, Günther

    2014-07-01

    Despite similarities of cellular membranes in all eukaryotes, every compartment displays characteristic and often unique features which are important for the functions of the specific organelles. In the present study, we biochemically characterized the plasma membrane of the methylotrophic yeast Pichia pastoris with emphasis on the lipids which form the matrix of this compartment. Prerequisite for this effort was the design of a standardized and reliable isolation protocol of the plasma membrane at high purity. Analysis of isolated plasma membrane samples from P. pastoris revealed an increase of phosphatidylserine and a decrease of phosphatidylcholine compared to bulk membranes. The amount of saturated fatty acids in the plasma membrane was higher than in total cell extracts. Ergosterol, the final product of the yeast sterol biosynthetic pathway, was found to be enriched in plasma membrane fractions, although markedly lower than in Saccharomyces cerevisiae. A further characteristic feature of the plasma membrane from P. pastoris was the enrichment of inositol phosphorylceramides over neutral sphingolipids, which accumulated in internal membranes. The detailed analysis of the P. pastoris plasma membrane is discussed in the light of cell biological features of this microorganism especially as a microbial cell factory for heterologous protein production.

  18. Affinity of four polar neurotransmitters for lipid bilayer membranes

    DEFF Research Database (Denmark)

    Wang, Chunhua; Ye, Fengbin; Valardez, Gustavo F.

    2011-01-01

    Weak interactions of neurotransmitters and the lipid matrix in the synaptic membrane have been hypothesized to play a role in synaptic transmission of nerve signals, particularly with respect to receptor desensitization (Cantor, R. S. Biochemistry 2003, 42, 11891). The strength of such interactions......, however, was not measured, and this is an obvious impediment for further evaluation and understanding of a possible role for desensitization. We have used dialysis equilibrium to directly measure the net affinity of selected neurotransmitters for lipid membranes and analyzed this affinity data...... with respect to calorimetric measurements and molecular dynamics simulations. We studied an anionic (glutamate), a cationic (acetylcholine), and two zwitterionic (-aminobutyric acid and glycine) neurotransmitters, and membranes of pure dimyristoyl phosphatidylcholine (DMPC), DMPC doped with 10% anionic lipid...

  19. Equilibrium Configurations of Lipid Bilayer Membranes and Carbon Nanostructures

    Institute of Scientific and Technical Information of China (English)

    Iva(i)lo M.Mladenov; Peter A.Djondjorov; Mariana Ts.Hadzhilazova; Vassil M.Vassilev

    2013-01-01

    The present article concerns the continuum modelling of the mechanical behaviour and equilibrium shapes of two types of nano-scale objects:fluid lipid bilayer membranes and carbon nanostructures.A unified continuum model is used to handle four different case studies.Two of them consist in representing in analytic form cylindrical and axisymmetric equilibrium configurations of single-wall carbon nanotubes and fluid lipid bilayer membranes subjected to uniform hydrostatic pressure.The third one is concerned with determination of possible shapes of junctions between a single-wall carbon nanotube and a fiat graphene sheet or another single-wall carbon nanotube.The last one deals with the mechanical behaviour of closed fluid lipid bilayer membranes (vesicles) adhering onto a fiat homogeneous rigid substrate subjected to micro-injection and uniform hydrostatic pressure.

  20. Tethered and Polymer Supported Bilayer Lipid Membranes: Structure and Function

    Directory of Open Access Journals (Sweden)

    Jakob Andersson

    2016-05-01

    Full Text Available Solid supported bilayer lipid membranes are model systems to mimic natural cell membranes in order to understand structural and functional properties of such systems. The use of a model system allows for the use of a wide variety of analytical tools including atomic force microscopy, impedance spectroscopy, neutron reflectometry, and surface plasmon resonance spectroscopy. Among the large number of different types of model membranes polymer-supported and tethered lipid bilayers have been shown to be versatile and useful systems. Both systems consist of a lipid bilayer, which is de-coupled from an underlying support by a spacer cushion. Both systems will be reviewed, with an emphasis on the effect that the spacer moiety has on the bilayer properties.

  1. Polymorphic Regulation of Outer Membrane Lipid A Composition

    Directory of Open Access Journals (Sweden)

    Russell E. Bishop

    2016-11-01

    Full Text Available The importance of the polymorphic-phase behavior of lipid A structural variations in determining their endotoxic activities has been recognized previously, but any potential role for lipid A polymorphism in controlling outer membrane structure and function has been largely ignored until now. In a recent article in mBio [7(5:e01532-16, https://doi.org/10.1128/mBio.01532-16], Katherine E. Bonnington and Meta J. Kuehn of Duke University’s Department of Biochemistry make a compelling case for considering how the molecular shapes of the various lipid A structural subtypes found in the outer membrane contribute to the process of outer membrane vesicle (OMV formation.

  2. Selective Interaction of a Cationic Polyfluorene with Model Lipid Membranes: Anionic versus Zwitterionic Lipids

    Directory of Open Access Journals (Sweden)

    Zehra Kahveci

    2014-03-01

    Full Text Available This paper explores the interaction mechanism between the conjugated polyelectrolyte {[9,9-bis(6'-N,N,N-trimethylammoniumhexyl]fluorene-phenylene}bromide (HTMA-PFP and model lipid membranes. The study was carried out using different biophysical techniques, mainly fluorescence spectroscopy and microscopy. Results show that despite the preferential interaction of HTMA-PFP with anionic lipids, HTMA-PFP shows affinity for zwitterionic lipids; although the interaction mechanism is different as well as HTMA-PFP’s final membrane location. Whilst the polyelectrolyte is embedded within the lipid bilayer in the anionic membrane, it remains close to the surface, forming aggregates that are sensitive to the physical state of the lipid bilayer in the zwitterionic system. The different interaction mechanism is reflected in the polyelectrolyte fluorescence spectrum, since the maximum shifts to longer wavelengths in the zwitterionic system. The intrinsic fluorescence of HTMA-PFP was used to visualize the interaction between polymer and vesicles via fluorescence microscopy, thanks to its high quantum yield and photostability. This technique allows the selectivity of the polyelectrolyte and higher affinity for anionic membranes to be observed. The results confirmed the appropriateness of using HTMA-PFP as a membrane fluorescent marker and suggest that, given its different behaviour towards anionic and zwitterionic membranes, HTMA-PFP could be used for selective recognition and imaging of bacteria over mammalian cells.

  3. Perfluorooctanoic acid rigidifies a model lipid membrane

    NARCIS (Netherlands)

    Brüning, B.A.; Farago, B.

    2014-01-01

    We report a combined dynamic light scattering and neutron spin-echo (NSE) study on vesicles composed of the phospholipid 1,2-dimyristoyl-sn-glycero-3-phosphatidylcholine under the influence of varying amounts of perfluorooctanoic acid. We study local lipid bilayer undulations using NSE on time

  4. Perfluorooctanoic acid rigidifies a model lipid membrane

    NARCIS (Netherlands)

    Brüning, B.A.; Farago, B.

    2014-01-01

    We report a combined dynamic light scattering and neutron spin-echo (NSE) study on vesicles composed of the phospholipid 1,2-dimyristoyl-sn-glycero-3-phosphatidylcholine under the influence of varying amounts of perfluorooctanoic acid. We study local lipid bilayer undulations using NSE on time scale

  5. Lipids and Membrane Microdomains in HIV-1 Replication

    OpenAIRE

    Waheed, Abdul A.; Freed, Eric O.

    2009-01-01

    Several critical steps in the replication cycle of human immunodeficiency virus type 1 (HIV-1) – entry, assembly and budding – are complex processes that take place at the plasma membrane of the host cell. A growing body of data indicates that these early and late steps in HIV-1 replication take place in specialized plasma membrane microdomains, and that many of the viral and cellular components required for entry, assembly, and budding are concentrated in these microdomains. In particular, a...

  6. Effect of oxidative stress on plasma membrane fluidity of THP-1 induced macrophages.

    Science.gov (United States)

    de la Haba, Carlos; Palacio, José R; Martínez, Paz; Morros, Antoni

    2013-02-01

    Plasma membrane is one of the preferential targets of reactive oxygen species which cause lipid peroxidation. This process modifies membrane properties such as membrane fluidity, a very important physical feature known to modulate membrane protein localization and function. The aim of this study is to evaluate the effect of oxidative stress on plasma membrane fluidity regionalization of single living THP-1 macrophages. These cells were oxidized with H(2)O(2) at different concentrations, and plasma membrane fluidity was analyzed by two-photon microscopy in combination with the environment-sensitive probe Laurdan. Results show a significant H(2)O(2) concentration dependent increase in the frequency of rigid lipid regions, mainly attributable to lipid rafts, at the expense of the intermediate fluidity regions. A novel statistical analysis evaluated changes in size and number of lipid raft domains under oxidative stress conditions, as lipid rafts are platforms aiding cell signaling and are thought to have relevant roles in macrophage functions. It is shown that H(2)O(2) causes an increase in the number, but not the size, of raft domains. As macrophages are highly resistant to H(2)O(2), these new raft domains might be involved in cell survival pathways.

  7. Plasma membrane regulates Ras signaling networks.

    Science.gov (United States)

    Chavan, Tanmay Sanjeev; Muratcioglu, Serena; Marszalek, Richard; Jang, Hyunbum; Keskin, Ozlem; Gursoy, Attila; Nussinov, Ruth; Gaponenko, Vadim

    2015-01-01

    Ras GTPases activate more than 20 signaling pathways, regulating such essential cellular functions as proliferation, survival, and migration. How Ras proteins control their signaling diversity is still a mystery. Several pieces of evidence suggest that the plasma membrane plays a critical role. Among these are: (1) selective recruitment of Ras and its effectors to particular localities allowing access to Ras regulators and effectors; (2) specific membrane-induced conformational changes promoting Ras functional diversity; and (3) oligomerization of membrane-anchored Ras to recruit and activate Raf. Taken together, the membrane does not only attract and retain Ras but also is a key regulator of Ras signaling. This can already be gleaned from the large variability in the sequences of Ras membrane targeting domains, suggesting that localization, environment and orientation are important factors in optimizing the function of Ras isoforms.

  8. The Effect of Lidocaine · HCl on the Fluidity of Native and Model Membrane Lipid Bilayers

    OpenAIRE

    Park, Jun-Seop; Jung, Tae-Sang; Noh, Yang-Ho; Kim, Woo-Sung; Park, Won-Ick; Kim, Young-Soo; Chung, In-Kyo; Sohn, Uy Dong; Bae, Soo-Kyung; Bae, Moon-Kyoung; Jang, Hye-Ock; Yun, Il

    2012-01-01

    The purpose of this study is to investigated the mechanism of pharmacological action of local anesthetic and provide the basic information about the development of new effective local anesthetics. Fluorescent probe techniques were used to evaluate the effect of lidocaine·HCl on the physical properties (transbilayer asymmetric lateral and rotational mobility, annular lipid fluidity and protein distribution) of synaptosomal plasma membrane vesicles (SPMV) isolated from bovine cerebral cortex, a...

  9. In vitro study of interaction of synaptic vesicles with lipid membranes

    Energy Technology Data Exchange (ETDEWEB)

    Ghosh, S K; Castorph, S; Salditt, T [Institute for X-ray Physics, University of Goettingen, 37077 Goettingen (Germany); Konovalov, O [European Synchrotron Radiation Facility, 38043 Grenoble Cedex (France); Jahn, R; Holt, M, E-mail: sghosh1@gwdg.d, E-mail: mholt@gwdg.d, E-mail: tsaldit@gwdg.d [Department of Neurobiology, Max Planck Institute for Biophysical Chemistry, 37077 Goettingen (Germany)

    2010-10-15

    The fusion of synaptic vesicles (SVs) with the plasma membrane in neurons is a crucial step in the release of neurotransmitters, which are responsible for carrying signals between nerve cells. While many of the molecular players involved in this fusion process have been identified, a precise molecular description of their roles in the process is still lacking. A case in point is the plasma membrane lipid phosphatidylinositol 4,5-bisphosphate (PIP{sub 2}). Although PIP{sub 2} is known to be essential for vesicle fusion, its precise role in the process remains unclear. We have re-investigated the role of this lipid in membrane structure and function using the complementary experimental techniques of x-ray reflectivity, both on lipid monolayers at an air-water interface and bilayers on a solid support, and grazing incidence x-ray diffraction on lipid monolayers. These techniques provide unprecedented access to structural information at the molecular level, and detail the profound structural changes that occur in a membrane following PIP{sub 2} incorporation. Further, we also confirm and extend previous findings that the association of SVs with membranes is enhanced by PIP{sub 2} incorporation, and reveal the structural changes that underpin this phenomenon. Further, the association is further intensified by a physiologically relevant amount of Ca{sup 2+} ions in the subphase of the monolayer, as revealed by the increase in interfacial pressure seen with the lipid monolayer system. Finally, a theoretical calculation concerning the products arising from the fusion of these SVs with proteoliposomes is presented, with which we aim to illustrate the potential future uses of this system.

  10. In vitro study of interaction of synaptic vesicles with lipid membranes

    Science.gov (United States)

    Ghosh, S. K.; Castorph, S.; Konovalov, O.; Jahn, R.; Holt, M.; Salditt, T.

    2010-10-01

    The fusion of synaptic vesicles (SVs) with the plasma membrane in neurons is a crucial step in the release of neurotransmitters, which are responsible for carrying signals between nerve cells. While many of the molecular players involved in this fusion process have been identified, a precise molecular description of their roles in the process is still lacking. A case in point is the plasma membrane lipid phosphatidylinositol 4,5-bisphosphate (PIP2). Although PIP2 is known to be essential for vesicle fusion, its precise role in the process remains unclear. We have re-investigated the role of this lipid in membrane structure and function using the complementary experimental techniques of x-ray reflectivity, both on lipid monolayers at an air-water interface and bilayers on a solid support, and grazing incidence x-ray diffraction on lipid monolayers. These techniques provide unprecedented access to structural information at the molecular level, and detail the profound structural changes that occur in a membrane following PIP2 incorporation. Further, we also confirm and extend previous findings that the association of SVs with membranes is enhanced by PIP2 incorporation, and reveal the structural changes that underpin this phenomenon. Further, the association is further intensified by a physiologically relevant amount of Ca2+ ions in the subphase of the monolayer, as revealed by the increase in interfacial pressure seen with the lipid monolayer system. Finally, a theoretical calculation concerning the products arising from the fusion of these SVs with proteoliposomes is presented, with which we aim to illustrate the potential future uses of this system.

  11. Hybrid lipids increase nanoscale fluctuation lifetimes in mixed membranes

    Science.gov (United States)

    Palmieri, Benoit; Safran, Samuel A.

    2013-09-01

    A recently proposed ternary mixture model is used to predict fluctuation domain lifetimes in the one phase region. The membrane is made of saturated, unsaturated, and hybrid lipids that have one saturated and one unsaturated hydrocarbon chain. The hybrid lipid is a natural linactant which can reduce the packing incompatibility between saturated and unsaturated lipids. The fluctuation lifetimes are predicted as a function of the hybrid lipid fraction and the fluctuation domain size. These lifetimes can be increased by up to three orders of magnitude compared to the case of no hybrids. With hybrid, small length scale fluctuations have sizable amplitudes even close to the critical temperature and, hence, benefit from enhanced critical slowing down. The increase in lifetime is particularly important for nanometer scale fluctuation domains where the hybrid orientation and the other lipids composition are highly coupled.

  12. Inhomogeneity Based Characterization of Distribution Patterns on the Plasma Membrane.

    Directory of Open Access Journals (Sweden)

    Laura Paparelli

    2016-09-01

    Full Text Available Cell surface protein and lipid molecules are organized in various patterns: randomly, along gradients, or clustered when segregated into discrete micro- and nano-domains. Their distribution is tightly coupled to events such as polarization, endocytosis, and intracellular signaling, but challenging to quantify using traditional techniques. Here we present a novel approach to quantify the distribution of plasma membrane proteins and lipids. This approach describes spatial patterns in degrees of inhomogeneity and incorporates an intensity-based correction to analyze images with a wide range of resolutions; we have termed it Quantitative Analysis of the Spatial distributions in Images using Mosaic segmentation and Dual parameter Optimization in Histograms (QuASIMoDOH. We tested its applicability using simulated microscopy images and images acquired by widefield microscopy, total internal reflection microscopy, structured illumination microscopy, and photoactivated localization microscopy. We validated QuASIMoDOH, successfully quantifying the distribution of protein and lipid molecules detected with several labeling techniques, in different cell model systems. We also used this method to characterize the reorganization of cell surface lipids in response to disrupted endosomal trafficking and to detect dynamic changes in the global and local organization of epidermal growth factor receptors across the cell surface. Our findings demonstrate that QuASIMoDOH can be used to assess protein and lipid patterns, quantifying distribution changes and spatial reorganization at the cell surface. An ImageJ/Fiji plugin of this analysis tool is provided.

  13. Inhomogeneity Based Characterization of Distribution Patterns on the Plasma Membrane.

    Science.gov (United States)

    Paparelli, Laura; Corthout, Nikky; Pavie, Benjamin; Wakefield, Devin L; Sannerud, Ragna; Jovanovic-Talisman, Tijana; Annaert, Wim; Munck, Sebastian

    2016-09-01

    Cell surface protein and lipid molecules are organized in various patterns: randomly, along gradients, or clustered when segregated into discrete micro- and nano-domains. Their distribution is tightly coupled to events such as polarization, endocytosis, and intracellular signaling, but challenging to quantify using traditional techniques. Here we present a novel approach to quantify the distribution of plasma membrane proteins and lipids. This approach describes spatial patterns in degrees of inhomogeneity and incorporates an intensity-based correction to analyze images with a wide range of resolutions; we have termed it Quantitative Analysis of the Spatial distributions in Images using Mosaic segmentation and Dual parameter Optimization in Histograms (QuASIMoDOH). We tested its applicability using simulated microscopy images and images acquired by widefield microscopy, total internal reflection microscopy, structured illumination microscopy, and photoactivated localization microscopy. We validated QuASIMoDOH, successfully quantifying the distribution of protein and lipid molecules detected with several labeling techniques, in different cell model systems. We also used this method to characterize the reorganization of cell surface lipids in response to disrupted endosomal trafficking and to detect dynamic changes in the global and local organization of epidermal growth factor receptors across the cell surface. Our findings demonstrate that QuASIMoDOH can be used to assess protein and lipid patterns, quantifying distribution changes and spatial reorganization at the cell surface. An ImageJ/Fiji plugin of this analysis tool is provided.

  14. The effect of electric fields on lipid membranes

    CERN Document Server

    Vasilkoski, Z

    2006-01-01

    Contrary to existing theoretical models, experimental evidence points out that electroporation (membrane defect formation under external electric fields) starts to occur within the range of transmembrane voltages that cells may routinely experience, curiously, just above the range of transmembrane voltages involved in neural signal transmission. Understanding the underlying principles of electric fields-lipid membrane interactions seems to carry a great biological importance. An argument is presented toward understanding the theoretical aspects of electroporation by using the DLVO theory, which has not been recognized previously in the context of electroporation. Further, the dispersion interactions (with its quantum nature), of the double layer counterions and membrane lipid molecules over the Stern layer are emphasized. The sign of these forces is such that they compress the membrane. A parallel is drawn to the theory of thin films. The argument is that the external electric field breaks the symmetry of the...

  15. Stabilization of Lipid Membranes With Dendritic Polymers

    Science.gov (United States)

    2004-12-01

    monolayer coverage was possible using the spin - coating procedure for concentrations of 10-5 w/w dendrimers in solution or lower. Higher coverages...The fluorescence intensity increased as coverage time before spin coating increased and as the dendrimer solution concentration increased (Fig...enhanced the adsorption of lipids to the substrate. Figure 7: Fluorescence intensity as a function of the PAMAM concentration used in the spin

  16. Sulfate transport in Penicillium chrysogenum plasma membranes

    NARCIS (Netherlands)

    Hillenga, Dirk J.; Versantvoort, Hanneke J.M.; Driessen, Arnold J.M.; Konings, Wil N.

    1996-01-01

    Transport studies with Penicillium chrysogenum plasma membranes fused with cytochrome c oxidase liposomes demonstrate that sulfate uptake is driven by the transmembrane pH gradient and not by the transmembrane electrical potential. Ca2+ and other divalent cations are not required. It is concluded th

  17. Sulfate transport in Penicillium chrysogenum plasma membranes.

    OpenAIRE

    Hillenga, Dirk J.; Versantvoort, Hanneke J.M.; Driessen, Arnold J. M.; Konings, Wil N.

    1996-01-01

    Transport studies with Penicillium chrysogenum plasma membranes fused with cytochrome c oxidase liposomes demonstrate that sulfate uptake is driven by the transmembrane pH gradient and not by the transmembrane electrical potential. Ca2+ and other divalent cations are not required. It is concluded that the sulfate transport system catalyzes the symport of two protons with one sulfate anion.

  18. Lipidomics reveals a remarkable diversity of lipids in human plasma.

    Science.gov (United States)

    Quehenberger, Oswald; Armando, Aaron M; Brown, Alex H; Milne, Stephen B; Myers, David S; Merrill, Alfred H; Bandyopadhyay, Sibali; Jones, Kristin N; Kelly, Samuel; Shaner, Rebecca L; Sullards, Cameron M; Wang, Elaine; Murphy, Robert C; Barkley, Robert M; Leiker, Thomas J; Raetz, Christian R H; Guan, Ziqiang; Laird, Gregory M; Six, David A; Russell, David W; McDonald, Jeffrey G; Subramaniam, Shankar; Fahy, Eoin; Dennis, Edward A

    2010-11-01

    The focus of the present study was to define the human plasma lipidome and to establish novel analytical methodologies to quantify the large spectrum of plasma lipids. Partial lipid analysis is now a regular part of every patient's blood test and physicians readily and regularly prescribe drugs that alter the levels of major plasma lipids such as cholesterol and triglycerides. Plasma contains many thousands of distinct lipid molecular species that fall into six main categories including fatty acyls, glycerolipids, glycerophospholipids, sphingolipids, sterols, and prenols. The physiological contributions of these diverse lipids and how their levels change in response to therapy remain largely unknown. As a first step toward answering these questions, we provide herein an in-depth lipidomics analysis of a pooled human plasma obtained from healthy individuals after overnight fasting and with a gender balance and an ethnic distribution that is representative of the US population. In total, we quantitatively assessed the levels of over 500 distinct molecular species distributed among the main lipid categories. As more information is obtained regarding the roles of individual lipids in health and disease, it seems likely that future blood tests will include an ever increasing number of these lipid molecules.

  19. Membrane protein crystallization in lipidic mesophases: detergent effects.

    OpenAIRE

    Ai, X.; Caffrey, M.

    2000-01-01

    The "cubic phase method" for growing crystals of membrane proteins uses a complex mixture of water, lipid, protein, and other components. The current view is that the cubic phase is integral to the process. Thus additives from whatever source introduce the possibility of destabilizing the phase, thereby compromising the crystallization process. Detergents are used to solubilize membrane proteins and are likely to be ported into the cubic medium with the target protein. Depending on the identi...

  20. A Variational Approach to Particles in Lipid Membranes

    Science.gov (United States)

    Elliott, Charles M.; Gräser, Carsten; Hobbs, Graham; Kornhuber, Ralf; Wolf, Maren-Wanda

    2016-11-01

    A variety of models for the membrane-mediated interaction of particles in lipid membranes, mostly well-established in theoretical physics, is reviewed from a mathematical perspective. We provide mathematically consistent formulations in a variational framework, relate apparently different modelling approaches in terms of successive approximation, and investigate existence and uniqueness. Numerical computations illustrate that the new variational formulations are directly accessible to effective numerical methods.

  1. Lipidic cubic phase injector facilitates membrane protein serial femtosecond crystallography.

    Science.gov (United States)

    Weierstall, Uwe; James, Daniel; Wang, Chong; White, Thomas A; Wang, Dingjie; Liu, Wei; Spence, John C H; Bruce Doak, R; Nelson, Garrett; Fromme, Petra; Fromme, Raimund; Grotjohann, Ingo; Kupitz, Christopher; Zatsepin, Nadia A; Liu, Haiguang; Basu, Shibom; Wacker, Daniel; Han, Gye Won; Katritch, Vsevolod; Boutet, Sébastien; Messerschmidt, Marc; Williams, Garth J; Koglin, Jason E; Marvin Seibert, M; Klinker, Markus; Gati, Cornelius; Shoeman, Robert L; Barty, Anton; Chapman, Henry N; Kirian, Richard A; Beyerlein, Kenneth R; Stevens, Raymond C; Li, Dianfan; Shah, Syed T A; Howe, Nicole; Caffrey, Martin; Cherezov, Vadim

    2014-01-01

    Lipidic cubic phase (LCP) crystallization has proven successful for high-resolution structure determination of challenging membrane proteins. Here we present a technique for extruding gel-like LCP with embedded membrane protein microcrystals, providing a continuously renewed source of material for serial femtosecond crystallography. Data collected from sub-10-μm-sized crystals produced with less than 0.5 mg of purified protein yield structural insights regarding cyclopamine binding to the Smoothened receptor.

  2. Interaction of pristine and functionalized carbon nanotubes with lipid membranes.

    Science.gov (United States)

    Baoukina, Svetlana; Monticelli, Luca; Tieleman, D Peter

    2013-10-10

    Carbon nanotubes are widely used in a growing number of applications. Their interactions with biological materials, cell membranes in particular, is of interest in applications including drug delivery and for understanding the toxicity of carbon nanotubes. We use extensive molecular dynamics simulations with the MARTINI model to study the interactions of model nanotubes of different thickness, length, and patterns of chemical modification with model membranes. In addition, we characterize the interactions of small bundles of carbon nanotubes with membrane models. Short pristine carbon nanotubes readily insert into membranes and adopt an orientation parallel to the plane of the membrane in the center of the membrane. Larger aggregates and functionalized nanotubes exhibit a range of possible interactions. The distribution and orientation of carbon nanotubes can be controlled by functionalizing the nanotubes. Free energy calculations provide thermodynamic insight into the preferred orientations of different nanotubes and quantify structural defects in the lipid matrix.

  3. Solid-Supported Lipid Membranes: Formation, Stability and Applications

    Science.gov (United States)

    Goh, Haw Zan

    This thesis presents a comprehensive investigation of the formation of supported lipid membranes with vesicle hemifusion, their stability under detergents and organic solvents and their applications in molecular biology. In Chapter 3, we describe how isolated patches of DOPC bilayers supported on glass surfaces are dissolved by various detergents (decyl maltoside, dodecyl maltoside, CHAPS, CTAB, SDS, TritonX-100 and Tween20) at their CMC, as investigated by fluorescence video microscopy. In general, detergents partition into distal leaflets of bilayers and lead to the expansion of the bilayers through a rolling motion of the distal over the proximal leaflets, in agreement with the first stage of the established 3-stage model of lipid vesicle solubilization by detergents. Subsequently, we study the partitioning of organic solvents (methanol, ethanol, isopropanol, propanol, acetone and chloroform) into isolated bilayer patches on glass in Chapter 4 with fluorescence microscopy. The area expansion of bilayers due to the partitioning of organic solvents is measured. From the titration of organic solvents, we measured the rate of area expansion as a function of the volume fraction of organic solvents, which is proposed to be a measure of strength of interactions between solvents and membranes. From the same experiments, we also measure the maximum expansion of bilayers (or the maximum binding stoichiometry between organic solvents and lipids) before structural breakdown, which depends on the depth of penetration of solvents to the membranes. In Chapter 5, we investigate the formation of sparsely-tethered bilayer lipid membranes (stBLMs) with vesicle hemifusion. In vesicle hemifusion, lipid vesicles in contact with a hydrophobic alkyl-terminated self-assembled monolayer (SAM) deposit a lipid monolayer to the SAM surface, thus completing the bilayer. Electrical Impedance Spectroscopy and Neutron Reflectivity are used to probe the integrity of stBLMs in terms of their

  4. Interaction of aspirin (acetylsalicylic acid) with lipid membranes.

    Science.gov (United States)

    Barrett, Matthew A; Zheng, Songbo; Roshankar, Golnaz; Alsop, Richard J; Belanger, Randy K R; Huynh, Chris; Kučerka, Norbert; Rheinstädter, Maikel C

    2012-01-01

    We studied the interaction of Aspirin (acetylsalicylic acid) with lipid membranes using x-ray diffraction for bilayers containing up to 50 mol% of aspirin. From 2D x-ray intensity maps that cover large areas of reciprocal space we determined the position of the ASA molecules in the phospholipid bilayers and the molecular arrangement of the molecules in the plane of the membranes. We present direct experimental evidence that ASA molecules participate in saturated lipid bilayers of DMPC (1,2-dimyristoyl-sn-glycero-3-phosphocholine) and preferably reside in the head group region of the membrane. Up to 50 mol% ASA molecules can be dissolved in this type of bilayer before the lateral membrane organization is disturbed and the membranes are found to form an ordered, 2D crystal-like structure. Furthermore, ASA and cholesterol were found to co-exist in saturated lipid bilayers, with the ASA molecules residing in the head group region and the cholesterol molecules participating in the hydrophobic membrane core.

  5. Interaction of aspirin (acetylsalicylic acid with lipid membranes.

    Directory of Open Access Journals (Sweden)

    Matthew A Barrett

    Full Text Available We studied the interaction of Aspirin (acetylsalicylic acid with lipid membranes using x-ray diffraction for bilayers containing up to 50 mol% of aspirin. From 2D x-ray intensity maps that cover large areas of reciprocal space we determined the position of the ASA molecules in the phospholipid bilayers and the molecular arrangement of the molecules in the plane of the membranes. We present direct experimental evidence that ASA molecules participate in saturated lipid bilayers of DMPC (1,2-dimyristoyl-sn-glycero-3-phosphocholine and preferably reside in the head group region of the membrane. Up to 50 mol% ASA molecules can be dissolved in this type of bilayer before the lateral membrane organization is disturbed and the membranes are found to form an ordered, 2D crystal-like structure. Furthermore, ASA and cholesterol were found to co-exist in saturated lipid bilayers, with the ASA molecules residing in the head group region and the cholesterol molecules participating in the hydrophobic membrane core.

  6. Effect of anesthetics on bending elasticity of lipid membranes

    Science.gov (United States)

    Yi, Zheng; Michihiro, Nagao; Bossev, Dobrin

    2008-03-01

    Change in physical and chemical properties of bio-membranes is of great interest for understanding the mechanism of anesthetic action on membranes. Hypothetically the anesthetic alters the lipid membrane structure (promoting pore formation across membranes or at least switching transmembrane channels) and therefore the biophysical properties of the membrane. We have used neutron spin echo (NSE) spectroscopy to study the effect of anesthetic molecule, lidocaine, on the bending elasticity (BE) of lipid membranes. BE of lipid bilayers made of (1,2-Dimyristoyl-sn-Glycero-3-Phosphocholine) DMPC and 1,2-Dipalmitoyl-sn-Glycero-3-Phosphocholine (DPPC) have been measured at different temperatures and different in the fluid (Lα) phase. Using Zilman-Granek theory the BE were obtained from the decay of the NSE intermediate scattering function. We have found that in the presence of lidocaine the BE of DMPC and DPPC bilayers increases. The results were correlated with those from differential scanning calorimetry. Increase in the lidocaine concentration leads to decrease in the liquid/crystalline transition temperature.

  7. Novel determinants of H-Ras plasma membrane localization and transformation

    DEFF Research Database (Denmark)

    Willumsen, B M; Cox, A D; Solski, P A;

    1996-01-01

    cysteine did not abolish palmitoylation. However, despite continued lipid modification the mutant proteins failed to bind to plasma membranes and instead accumulated on internal membranes and, importantly, were not transforming. Addition of an N-terminal myristoylation signal to these defective mutants......, or to proteins entirely lacking the C-terminal 25 residues restored both plasma membrane association and transforming activity. Thus, H-Ras does not absolutely require prenylation or palmitoylation nor indeed its hypervariable domain in order to interact with effectors that ultimately cause transformation....... However, in this native state, the C-terminus appears to provide a combination of lipids and a previously unrecognized signal for specific plasma membrane targeting that are essential for the correct localization and biological function of H-Ras....

  8. Plasma deposited fluorinated films on porous membranes

    Energy Technology Data Exchange (ETDEWEB)

    Gancarz, Irena [Department of Polymer and Carbon Materials, Wrocław University of Technology, 50-370 Wrocław (Poland); Bryjak, Marek, E-mail: marek.bryjak@pwr.edu.pl [Department of Polymer and Carbon Materials, Wrocław University of Technology, 50-370 Wrocław (Poland); Kujawski, Jan; Wolska, Joanna [Department of Polymer and Carbon Materials, Wrocław University of Technology, 50-370 Wrocław (Poland); Kujawa, Joanna; Kujawski, Wojciech [Nicolaus Copernicus University, Faculty of Chemistry, 7 Gagarina St., 87-100 Torun (Poland)

    2015-02-01

    75 KHz plasma was used to modify track etched poly(ethylene terephthalate) membranes and deposit on them flouropolymers. Two fluorine bearing monomers were used: perflourohexane and hexafluorobenzene. The modified surfaces were analyzed by means of attenuated total reflection infra-red spectroscopy, X-ray photoelectron spectroscopy, scanning electron microscopy, atomic force microscopy and wettability. It was detected that hexaflourobenxene deposited to the larger extent than perflourohaxane did. The roughness of surfaces decreased when more fluoropolymer was deposited. The hydrophobic character of surface slightly disappeared during 20-days storage of hexaflourobenzene modified membrane. Perfluorohexane modified membrane did not change its character within 120 days after modification. It was expected that this phenomenon resulted from post-reactions of oxygen with radicals in polymer deposits. The obtained membranes could be used for membrane distillation of juices. - Highlights: • Plasma deposited hydrophobic layer of flouropolymers. • Deposition degree affects the surface properties. • Hydrohilization of surface due to reaction of oxygen with entrapped radicals. • Possibility to use modified porous membrane for water distillation and apple juice concentration.

  9. Proteomic Profiling of Detergent Resistant Membranes (Lipid Rafts) of Prostasomes.

    Science.gov (United States)

    Dubois, Louise; Ronquist, Karl K Göran; Ek, Bo; Ronquist, Gunnar; Larsson, Anders

    2015-11-01

    Prostasomes are exosomes derived from prostate epithelial cells through exocytosis by multivesicular bodies. Prostasomes have a bilayered membrane and readily interact with sperm. The membrane lipid composition is unusual with a high contribution of sphingomyelin at the expense of phosphatidylcholine and saturated and monounsaturated fatty acids are dominant. Lipid rafts are liquid-ordered domains that are more tightly packed than the surrounding nonraft phase of the bilayer. Lipid rafts are proposed to be highly dynamic, submicroscopic assemblies that float freely within the liquid disordered membrane bilayer and some proteins preferentially partition into the ordered raft domains. We asked the question whether lipid rafts do exist in prostasomes and, if so, which proteins might be associated with them. Prostasomes of density range 1.13-1.19g/ml were subjected to density gradient ultracentrifugation in sucrose fabricated by phosphate buffered saline (PBS) containing 1% Triton X-100 with capacity for banding at 1.10 g/ml, i.e. the classical density of lipid rafts. Prepared prostasomal lipid rafts (by gradient ultracentrifugation) were analyzed by mass spectrometry. The clearly visible band on top of 1.10g/ml sucrose in the Triton X-100 containing gradient was subjected to liquid chromatography-tandem MS and more than 370 lipid raft associated proteins were identified. Several of them were involved in intraluminal vesicle formation, e.g. tetraspanins, ESCRTs, and Ras-related proteins. This is the first comprehensive liquid chromatography-tandem MS profiling of proteins in lipid rafts derived from exosomes. Data are available via ProteomeXchange with identifier PXD002163.

  10. α-Synuclein senses lipid packing defects and induces lateral expansion of lipids leading to membrane remodeling.

    Science.gov (United States)

    Ouberai, Myriam M; Wang, Juan; Swann, Marcus J; Galvagnion, Celine; Guilliams, Tim; Dobson, Christopher M; Welland, Mark E

    2013-07-19

    There is increasing evidence for the involvement of lipid membranes in both the functional and pathological properties of α-synuclein (α-Syn). Despite many investigations to characterize the binding of α-Syn to membranes, there is still a lack of understanding of the binding mode linking the properties of lipid membranes to α-Syn insertion into these dynamic structures. Using a combination of an optical biosensing technique and in situ atomic force microscopy, we show that the binding strength of α-Syn is related to the specificity of the lipid environment (the lipid chemistry and steric properties within a bilayer structure) and to the ability of the membranes to accommodate and remodel upon the interaction of α-Syn with lipid membranes. We show that this interaction results in the insertion of α-Syn into the region of the headgroups, inducing a lateral expansion of lipid molecules that can progress to further bilayer remodeling, such as membrane thinning and expansion of lipids out of the membrane plane. We provide new insights into the affinity of α-Syn for lipid packing defects found in vesicles of high curvature and in planar membranes with cone-shaped lipids and suggest a comprehensive model of the interaction between α-Syn and lipid bilayers. The ability of α-Syn to sense lipid packing defects and to remodel membrane structure supports its proposed role in vesicle trafficking.

  11. Lipid-Mediated Clusters of Guest Molecules in Model Membranes and Their Dissolving in the Presence of Lipid Rafts.

    Science.gov (United States)

    Kardash, Maria E; Dzuba, Sergei A

    2017-05-25

    The clustering of molecules is an important feature of plasma membrane organization. It is challenging to develop methods for quantifying membrane heterogeneities because of their transient nature and small size. Here, we obtained evidence that transient membrane heterogeneities can be frozen at cryogenic temperatures which allows the application of solid-state experimental techniques sensitive to the nanoscale distance range. We employed the pulsed version of electron paramagnetic resonance (EPR) spectroscopy, the electron spin echo (ESE) technique, for spin-labeled molecules in multilamellar lipid bilayers. ESE decays were refined for pure contribution of spin-spin magnetic dipole-dipolar interaction between the labels; these interactions manifest themselves at a nanometer distance range. The bilayers were prepared from different types of saturated and unsaturated lipids and cholesterol (Chol); in all cases, a small amount of guest spin-labeled substances 5-doxyl-stearic-acid (5-DSA) or 3β-doxyl-5α-cholestane (DChl) was added. The local concentration found of 5-DSA and DChl molecules was remarkably higher than the mean concentration in the bilayer, evidencing the formation of lipid-mediated clusters of these molecules. To our knowledge, formation of nanoscale clusters of guest amphiphilic molecules in biological membranes is a new phenomenon suggested only recently. Two-dimensional 5-DSA molecular clusters were found, whereas flat DChl molecules were found to be clustered into stacked one-dimensional structures. These clusters disappear when the Chol content is varied between the boundaries known for lipid raft formation at room temperatures. The room temperature EPR evidenced entrapping of DChl molecules in the rafts.

  12. Assessing plasma glucose and lipid levels, body weight and acute ...

    African Journals Online (AJOL)

    Assessing plasma glucose and lipid levels, body weight and acute toxicity following oral ... Diabetes was induced in male and female Wistar rats with alloxan ... had good hypoglycemic activity and good effects on cardiovascular risk factors.

  13. Stabilization of composition fluctuations in mixed membranes by hybrid lipids

    Science.gov (United States)

    Safran, Samuel; Palmieri, Benoit

    2013-03-01

    A ternary mixture model is proposed to describe composition fluctuations in mixed membranes composed of saturated, unsaturated and hybrid lipids. The asymmetric hybrid lipid has one saturated and one unsaturated hydrocarbon chain and it can reduce the packing incompatibility between saturated and unsaturated lipids. A methodology to recast the free-energy of the lattice in terms of a continuous isotropic field theory is proposed and used to analyze composition fluctuations above the critical temperature. The effect of hybrid lipids on fluctuations domains rich in saturated/unsaturated lipids is predicted. The correlation length of such fluctuations decreases significantly with increasing amounts of hybrids even if the temperature is maintained close to the critical temperature. This provides an upper bound for the domain sizes expected in rafts stabilized by hybrids, above the critical temperature. When the hybrid composition of the membrane is increased further, a crossover value is found above which ``stripe-like'' fluctuations are observed. The wavelength of these fluctuations decreases with increasing hybrid fraction and tends toward a molecular size in a membrane that contains only hybrids.

  14. Cholesterol:phospholipid ratio is elevated in platelet plasma membrane in patients with hypertension.

    Science.gov (United States)

    Benjamin, N; Robinson, B F; Graham, J G; Wilson, R B

    1990-06-01

    The cholesterol:phospholipid ratio was measured in platelet plasma membrane, red blood cell (RBC) membranes, low density lipoprotein (LDL) and whole plasma in patients with primary hypertension and in matched normal controls. The cholesterol:phospholipid ratio was raised in the platelet membrane from hypertensive patients compared with that from normal controls (0.65 +/- 0.03 vs 0.53 +/- 0.02: mean +/- SEM; P less than 0.01). The ratio observed in RBC membranes, LDL and whole blood was similar in the two groups. If this abnormality in the lipid composition of platelet plasma membrane is present in other cells it could account for some of the changes in cell membrane function that have been described in hypertension.

  15. A microfluidic platform for probing single cell plasma membranes using optically trapped Smart Droplet Microtools (SDMs).

    Science.gov (United States)

    Lanigan, Peter M P; Ninkovic, Tanja; Chan, Karen; de Mello, Andrew J; Willison, Keith R; Klug, David R; Templer, Richard H; Neil, Mark A A; Ces, Oscar

    2009-04-21

    We recently introduced a novel platform based upon optically trapped lipid coated oil droplets (Smart Droplet Microtools-SDMs) that were able to form membrane tethers upon fusion with the plasma membrane of single cells. Material transfer from the plasma membrane to the droplet via the tether was seen to occur. Here we present a customised version of the SDM approach based upon detergent coated droplets deployed within a microfluidic format. These droplets are able to differentially solubilise the plasma membrane of single cells with spatial selectivity and without forming membrane tethers. The microfluidic format facilitates separation of the target cells from the bulk SDM population and from downstream analysis modules. Material transfer from the cell to the SDM was monitored by tracking membrane localized EGFP.

  16. Effects of non-thermal plasma on the electrical properties of an erythrocyte membrane

    Science.gov (United States)

    Lee, Jin Young; Baik, Ku Youn; Kim, Tae Soo; Lim, Jaekwan; Uhm, Han S.; Choi, Eun Ha

    2015-09-01

    Non-thermal plasma is used here for membrane oxidation and permeabilization in which the electrical properties of an erythrocyte membrane are investigated after treatments. The zeta potential as measured by electrophoresis shows the increased negativity of the membrane surface potential (Ψs). The secondary electron emission coefficient ( γ) measured by a focused ion beam shows a decrease in the dipole potential (Ψd) of lipid molecules. The voltage-sensitive fluorescent intensity as measured by flow cytometry shows a decrease in the trans-membrane potential (ΔΨ) through the lipid bilayer membrane. These results allow us to take a step forward to unveil the complex events occurring in plasma-treated cells.

  17. Interaction of articaine hydrochloride with prokaryotic membrane lipids.

    Science.gov (United States)

    Lygre, Henning; Moe, Grete; Nerdal, Willy; Holmsen, Holm

    2009-01-01

    Local anesthetics are the most commonly used drugs in dentistry, with a wide range of effects, including antimicrobial activity. High antimicrobial effects have recently been reported on oral microbes from articaine hydrochloride, revealed by the minimum inhibitory concentration and minimal bactericidal concentration. Additionally, articaine has recently been used as an alkaline component in endodontic materials with a proposed antibacterial activity. However, the detailed mechanisms of action have not been discussed. We determined the Langmuir surface pressure/molecular area isotherms of prokaryotic lipid monolayers, as well as the phospholipid phase transitions, by employing differential scanning calorimetry on unilamellar prokaryotic liposomes (bilayers). Articaine hydrochloride was found to interact with the prokaryotic membrane lipids in both monolayers and bilayers. An increase of the phospholipid molecular area of acidic glycerophospholipids as well as a decrease in phase transition temperature and enthalpy were found with increasing articaine hydrochloride concentration. The thermodynamic changes by adding articaine hydrochloride to prokaryotic membrane lipids are potentially related to the effects observed from antimicrobial peptides resulting from membrane insertion, aggregate composition, pore formation, and lysis. Interaction of articaine hydrochloride with prokaryotic membrane lipids is indicated. Hence, further research is necessary to gain insight into where these compounds exert their effects at the molecular level.

  18. Partitioning of Lipids at Domain Boundaries in Model Membranes

    NARCIS (Netherlands)

    Schafer, Lars V.; Marrink, Siewert J.

    2010-01-01

    Line-active molecules ("linactants") that bind to the boundary interface between different fluid lipid domains in membranes have a strong potential as regulators of the lateral heterogeneity that is important for many biological processes. Here, we use molecular dynamics simulations in combination w

  19. Valinomycin acts as a channel in ultrathin lipid membranes.

    Science.gov (United States)

    Gliozzi, A; Robello, M; Fittabile, L; Relini, A; Gambacorta, A

    1996-08-14

    When the thickness of monolayer membranes formed by bolaform archaeal lipids is reduced to the approximate length of two valinomycin molecules, the zero-current conductance does not show any more a linear dependence on valinomycin concentration; instead, a quadratic behaviour is observed. This suggests that a dimer permeation pore is formed and therefore the conduction mechanism changes from carrier to channel.

  20. Computer simulation of cytoskeleton-induced blebbing in lipid membranes

    DEFF Research Database (Denmark)

    Spangler, E. J.; Harvey, C. W.; Revalee, J. D.

    2011-01-01

    Blebs are balloon-shaped membrane protrusions that form during many physiological processes. Using computer simulation of a particle-based model for self-assembled lipid bilayers coupled to an elastic meshwork, we investigated the phase behavior and kinetics of blebbing. We found that blebs form...

  1. Interactions between mycoplasma lipid-associated membrane proteins and the host cells

    Institute of Scientific and Technical Information of China (English)

    YOU Xiao-xing; ZENG Yan-hua; WU Yi-mou

    2006-01-01

    Mycoplamas are a group of wall-less prokaryotes widely distributed in nature, some of which are pathogenic for humans and animals. There are many lipoproteins anchored on the outer face of the plasma membrane, called lipid-associated membrane proteins (LAMPs). LAMPs are highly antigenic and could undergo phase and size variation, and are recognized by the innate immune system through Toll-like receptors (TLR) 2 and 6. LAMPs can modulate the immune system, and could induce immune cells apoptosis or death. In addition, they may associate with malignant transformation of host cells and are also considered to be cofactors in the progression of AIDS.

  2. Drug-induced hemolytic anemia and thrombocytopenia associated with alterations of cell membrane lipids and acanthocyte formation.

    Science.gov (United States)

    Poulet, Frederique M; Penraat, Kelley; Collins, Nathaniel; Evans, Ellen; Thackaberry, Evan; Manfra, Denise; Engstrom, Laura; Geissler, Richard; Geraci-Erck, Maria; Frugone, Carlos; Abutarif, Malaz; Fine, Jay S; Peterson, Brianna L; Cummings, Brian S; Johnson, Robert C

    2010-10-01

    CXCR3 is a chemokine receptor, upregulated upon activation of T cells and expressed on nearly 100% of T cells in sites of inflammation. SCH 900875 is a selective CXCR3 receptor antagonist. Thrombocytopenia and severe hemolytic anemia with acanthocytosis occurred in rats at doses of 75, 100, and 150 mg/kg/day. Massively enlarged spleens corresponded histologically to extramedullary hematopoiesis, macrophages, and hemosiderin pigment and sinus congestion. Phagocytosed erythrocytes and platelets were within splenic macrophages. IgG and/or IgM were not detected on erythrocyte and platelet membranes. Ex vivo increased osmotic fragility of RBCs was observed. Lipid analysis of the RBC membrane revealed modifications in phosphatidylcholine, overall cholesterol, and/or sphingomyelin. Platelets exhibited slender filiform processes on their plasma membranes, analogous to those of acanthocytes. The presence of similar morphological abnormalities in acanthocytes and platelets suggests that possibly similar alterations in the lipid composition of the plasma membrane have taken place in both cell types. This phenotype correlated with alterations in plasma lipids (hypercholesterolemia and low triglycerides) that occurred after SCH 900875 administration, although other factors cannot be excluded. The increased cell destruction was considered triggered by alterations in the lipid profile of the plasma membranes of erythrocytes and platelets, as reflected morphologically.

  3. An ER Protein Functionally Couples Neutral Lipid Metabolism on Lipid Droplets to Membrane Lipid Synthesis in the ER

    DEFF Research Database (Denmark)

    Markgraf, Daniel F; Klemm, Robin W; Junker, Mirco;

    2014-01-01

    Eukaryotic cells store neutral lipids such as triacylglycerol (TAG) in lipid droplets (LDs). Here, we have addressed how LDs are functionally linked to the endoplasmic reticulum (ER). We show that, in S. cerevisiae, LD growth is sustained by LD-localized enzymes. When LDs grow in early stationary...... phase, the diacylglycerol acyl-transferase Dga1p moves from the ER to LDs and is responsible for all TAG synthesis from diacylglycerol (DAG). During LD breakdown in early exponential phase, an ER membrane protein (Ice2p) facilitates TAG utilization for membrane-lipid synthesis. Ice2p has a cytosolic...... and explain how cells switch neutral lipid metabolism from storage to consumption....

  4. INTRACELLULAR TRANSPORT. PI4P/phosphatidylserine countertransport at ORP5- and ORP8-mediated ER-plasma membrane contacts.

    Science.gov (United States)

    Chung, Jeeyun; Torta, Federico; Masai, Kaori; Lucast, Louise; Czapla, Heather; Tanner, Lukas B; Narayanaswamy, Pradeep; Wenk, Markus R; Nakatsu, Fubito; De Camilli, Pietro

    2015-07-24

    Lipid transfer between cell membrane bilayers at contacts between the endoplasmic reticulum (ER) and other membranes help to maintain membrane lipid homeostasis. We found that two similar ER integral membrane proteins, oxysterol-binding protein (OSBP)-related protein 5 (ORP5) and ORP8, tethered the ER to the plasma membrane (PM) via the interaction of their pleckstrin homology domains with phosphatidylinositol 4-phosphate (PI4P) in this membrane. Their OSBP-related domains (ORDs) harbored either PI4P or phosphatidylserine (PS) and exchanged these lipids between bilayers. Gain- and loss-of-function experiments showed that ORP5 and ORP8 could mediate PI4P/PS countertransport between the ER and the PM, thus delivering PI4P to the ER-localized PI4P phosphatase Sac1 for degradation and PS from the ER to the PM. This exchange helps to control plasma membrane PI4P levels and selectively enrich PS in the PM.

  5. GPI-anchored proteins do not reside in ordered domains in the live cell plasma membrane

    Science.gov (United States)

    Sevcsik, Eva; Brameshuber, Mario; Fölser, Martin; Weghuber, Julian; Honigmann, Alf; Schütz, Gerhard J.

    2015-04-01

    The organization of proteins and lipids in the plasma membrane has been the subject of a long-lasting debate. Membrane rafts of higher lipid chain order were proposed to mediate protein interactions, but have thus far not been directly observed. Here we use protein micropatterning combined with single-molecule tracking to put current models to the test: we rearranged lipid-anchored raft proteins (glycosylphosphatidylinositol(GPI)-anchored-mGFP) directly in the live cell plasma membrane and measured the effect on the local membrane environment. Intriguingly, this treatment does neither nucleate the formation of an ordered membrane phase nor result in any enrichment of nanoscopic-ordered domains within the micropatterned regions. In contrast, we find that immobilized mGFP-GPIs behave as inert obstacles to the diffusion of other membrane constituents without influencing their membrane environment over distances beyond their physical size. Our results indicate that phase partitioning is not a fundamental element of protein organization in the plasma membrane.

  6. Spin-labelled vacuolar-ATPase inhibitors in lipid membranes.

    Science.gov (United States)

    Dixon, Neil; Páli, Tibor; Kee, Terence P; Marsh, Derek

    2004-10-11

    Two spin-labelled derivatives of the 5-(2-indolyl)-2,4-pentadienoyl class of inhibitors of the vacuolar ATPase have been synthesised and their EPR properties characterised in phospholipid membranes. One spin-labelled inhibitor is the amide derivative of pentadienic acid and 4-amino-TEMPO (INDOL6), and the other is the 3-hydroxymethyl-PROXYL ester (INDOL5). The response of the EPR spectra to the chain-melting transition of dimyristoyl phosphatidylcholine (DMPC) bilayers demonstrates that both derivatives incorporate in phospholipid membranes. The axially anisotropic EPR spectra of INDOL6 in fluid DMPC membranes indicate that the indolyl-pentadienoyl inhibitors intercalate between the lipid chains, in the membrane. INDOL5, designed to possess additional internal segmental mobility, exhibits more nearly isotropic motion of the spin-label moiety in fluid membranes than does INDOL6. The EPR characteristics of INDOL5 are therefore well suited to detecting specific ligand-protein interactions. Progressive saturation EPR experiments with polar and hydrophobic relaxation agents (aqueous Ni2+ and oxygen) show that the nitroxide group is buried in the membrane, with the indole moiety providing the anchor at the membrane polar-apolar interface. Rates of spin-label reduction by externally added ascorbate confirm this assignment. These two spin-labelled derivatives provide complementary EPR probes of the lipid environment (INDOL6), and of ligand-protein interactions (INDOL5), for this class of V-ATPase inhibitor.

  7. Lipid domains control myelin basic protein adsorption and membrane interactions between model myelin lipid bilayers.

    Science.gov (United States)

    Lee, Dong Woog; Banquy, Xavier; Kristiansen, Kai; Kaufman, Yair; Boggs, Joan M; Israelachvili, Jacob N

    2014-02-25

    The surface forces apparatus and atomic force microscope were used to study the effects of lipid composition and concentrations of myelin basic protein (MBP) on the structure of model lipid bilayers, as well as the interaction forces and adhesion between them. The lipid bilayers had a lipid composition characteristic of the cytoplasmic leaflets of myelin from "normal" (healthy) and "disease-like" [experimental allergic encephalomyelitis (EAE)] animals. They showed significant differences in the adsorption mechanism of MBP. MBP adsorbs on normal bilayers to form a compact film (3-4 nm) with strong intermembrane adhesion (∼0.36 mJ/m(2)), in contrast to its formation of thicker (7-8 nm) swelled films with weaker intermembrane adhesion (∼0.13 mJ/m(2)) on EAE bilayers. MBP preferentially adsorbs to liquid-disordered submicron domains within the lipid membranes, attributed to hydrophobic attractions. These results show a direct connection between the lipid composition of membranes and membrane-protein adsorption mechanisms that affects intermembrane spacing and adhesion and has direct implications for demyelinating diseases.

  8. Shear rheology of lipid monolayers and insights on membrane fluidity

    Science.gov (United States)

    Espinosa, Gabriel; López-Montero, Iván; Monroy, Francisco; Langevin, Dominique

    2011-01-01

    The concept of membrane fluidity usually refers to a high molecular mobility inside the lipid bilayer which enables lateral diffusion of embedded proteins. Fluids have the ability to flow under an applied shear stress whereas solids resist shear deformations. Biological membranes require both properties for their function: high lateral fluidity and structural rigidity. Consequently, an adequate account must include, in addition to viscosity, the possibility for a nonzero shear modulus. This knowledge is still lacking as measurements of membrane shear properties have remained incomplete so far. In the present contribution we report a surface shear rheology study of different lipid monolayers that model distinct biologically relevant situations. The results evidence a large variety of mechanical behavior under lateral shear flow. PMID:21444777

  9. DNA-Tile Structures Induce Ionic Currents through Lipid Membranes.

    Science.gov (United States)

    Göpfrich, Kerstin; Zettl, Thomas; Meijering, Anna E C; Hernández-Ainsa, Silvia; Kocabey, Samet; Liedl, Tim; Keyser, Ulrich F

    2015-05-13

    Self-assembled DNA nanostructures have been used to create man-made transmembrane channels in lipid bilayers. Here, we present a DNA-tile structure with a nominal subnanometer channel and cholesterol-tags for membrane anchoring. With an outer diameter of 5 nm and a molecular weight of 45 kDa, the dimensions of our synthetic nanostructure are comparable to biological ion channels. Because of its simple design, the structure self-assembles within a minute, making its creation scalable for applications in biology. Ionic current recordings demonstrate that the tile structures enable ion conduction through lipid bilayers and show gating and voltage-switching behavior. By demonstrating the design of DNA-based membrane channels with openings much smaller than that of the archetypical six-helix bundle, our work showcases their versatility inspired by the rich diversity of natural membrane components.

  10. Polyunsaturation in lipid membranes: dynamic properties and lateral pressure profiles.

    Science.gov (United States)

    Ollila, Samuli; Hyvönen, Marja T; Vattulainen, Ilpo

    2007-03-29

    We elucidate the influence of unsaturation on single-component membrane properties, focusing on their dynamical aspects and lateral pressure profiles across the membrane. To this end, we employ atomistic molecular dynamics simulations to study five different membrane systems with varying degrees of unsaturation, starting from saturated membranes and systematically increasing the level of unsaturation, ending up with a bilayer of phospholipids containing the docosahexaenoic acid. For an increasing level of unsaturation, we find considerable effects on dynamical properties, such as accelerated dynamics of the phosphocholine head groups and glycerol backbones and speeded up rotational dynamics of the lipid molecules. The lateral pressure profile is found to be altered by the degree of unsaturation. For an increasing number of double bonds, the peak in the middle of the bilayer decreases. This is compensated for by changes in the membrane-water interface region in terms of increasing peak heights of the lateral pressure profile. Implications of the findings are briefly discussed.

  11. Binding of Neurotransmitters to Lipid Membranes

    DEFF Research Database (Denmark)

    Peters, Günther H.J.; Werge, Mikkel; Elf-Lind, Maria Northved

    2014-01-01

    We have performed a series of thermodynamic measurements and molecular dynamics (MD) simulations to study the interactions between the neurotransmitters (NTs) 5-hydroxytryptamine (5-HT), g-aminobutyrate (GABA), glycine (GLY), acetylcholine (ACH) and glutamate (GLU) as well as the amidated...... as the most important interaction by which the NTs are anchored to the membrane. These distinctive interactions could be related to nonspecific effects of these neurotransmitters and could point to a bilayer-mediated modulation of nerve transmission. However, due to the strong variability in affinity observed...... for the different NTs, this attraction is not an inherent property of all neurotransmitters....

  12. Cyclohexane Rings Reduce Membrane Permeability to Small Ions in Archaea-Inspired Tetraether Lipids.

    Science.gov (United States)

    Koyanagi, Takaoki; Leriche, Geoffray; Onofrei, David; Holland, Gregory P; Mayer, Michael; Yang, Jerry

    2016-01-26

    Extremophile archaeal organisms overcome problems of membrane permeability by producing lipids with structural elements that putatively improve membrane integrity compared to lipids from other life forms. Herein, we describe a series of lipids that mimic some key structural features of archaeal lipids, such as: 1) single tethering of lipid tails to create fully transmembrane tetraether lipids and 2) the incorporation of small rings into these tethered segments. We found that membranes formed from pure tetraether lipids leaked small ions at a rate that was about two orders of magnitude slower than common bilayer-forming lipids. Incorporation of cyclopentane rings into the tetraether lipids did not affect membrane leakage, whereas a cyclohexane ring reduced leakage by an additional 40 %. These results show that mimicking certain structural features of natural archaeal lipids results in improved membrane integrity, which may help overcome limitations of many current lipid-based technologies.

  13. Direct visualization of lipid domains in human skin stratum corneum's lipid membranes

    DEFF Research Database (Denmark)

    Plasencia, I; Norlen, Lars; Bagatolli, Luis

    2007-01-01

    ; and iii), whether pH has a direct effect on the lipid matrix phase behavior. In this work the lateral structure of membranes composed of lipids extracted from human skin stratum corneum was studied in a broad temperature range (10 degrees C-90 degrees C) using different techniques such as differential......The main function of skin is to serve as a physical barrier between the body and the environment. This barrier capacity is in turn a function of the physical state and structural organization of the stratum corneum extracellular lipid matrix. This lipid matrix is essentially composed of very long...... scanning calorimetry, fluorescence spectroscopy, and two-photon excitation and laser scanning confocal fluorescence microscopy. Here we show that hydrated bilayers of human skin stratum corneum lipids express a giant sponge-like morphology with dimensions corresponding to the global three...

  14. Lipid engineering reveals regulatory roles for membrane fluidity in yeast flocculation and oxygen-limited growth

    DEFF Research Database (Denmark)

    Degreif, Daniel; de Rond, Tristan; Bertl, Adam

    2017-01-01

    Cells modulate lipid metabolism in order to maintain membrane homeostasis. Here we use a metabolic engineering approach to manipulate the stoichiometry of fatty acid unsaturation, a regulator of cell membrane fluidity, in Saccharomyces cerevisiae. Unexpectedly, reduced lipid unsaturation triggere...

  15. Long term effects on human plasma lipoproteins of a formulation enriched in butter milk polar lipid

    Directory of Open Access Journals (Sweden)

    Nilsson Åke

    2009-10-01

    Full Text Available Abstract Background Sphingolipids (SL, in particular sphingomyelin (SM are important components of milk fat polar lipids. Dietary SM inhibits cholesterol absorption in rats (Nyberg et al. J Nutr Biochem. 2000 and SLs decrease both cholesterol and TG concentrations in lipid- and cholesterol fed APOE*3Leiden mice (Duivenvoorden et al. Am J Clin Nutr. 2006. This human study examines effects of a butter milk formulation enriched in milk fat globule membrane material, and thereby in SLs, on blood lipids in healthy volunteers. In a four week parallel group study with 33 men and 15 women we examined the effects of an SL-enriched butter milk formulation (A and an equivalent control formulation (B on plasma lipid levels. Plasma concentrations of HDL and LDL cholesterol, triacylglycerols (TG, apolipoproteins AI and B, and lipoprotein (a were measured. The daily dose of SL in A was 975 mg of which 700 mg was SM. The participants registered food and drink intake four days before introducing the test formula and the last four days of the test period. Results A daily increase of SL intake did not significantly influence fasting plasma lipids or lipoproteins. In group B TG, cholesterol, LDL, HDL and apolipoprotein B concentrations increased, however, but not in group A after four weeks. The difference in LDL cholesterol was seen primarily in women and difference in TG primarily in men. No significant side effects were observed. Conclusion The study did not show any significant decrease on plasma lipids or lipoprotein levels of an SL-enriched formulation containing 2-3 times more SL than the normal dietary intake on cholesterol, other plasma lipids or on energy intake. The formulation A may, however, have counteracted the trend towards increased blood lipid concentrations caused by increased energy intake that was seen with the B formulation.

  16. Protein-lipid interactions in bilayer membranes: a lattice model.

    Science.gov (United States)

    Pink, D A; Chapman, D

    1979-04-01

    A lattice model has been developed to study the effects of intrinsic membrane proteins upon the thermodynamic properties of a lipid bilayer membrane. We assume that only nearest-neighbor van der Waals and steric interactions are important and that the polar group interactions can be represented by effective pressure-area terms. Phase diagrams, the temperature T(0), which locates the gel-fluid melting, the transition enthalpy, and correlations were calculated by mean field and cluster approximations. Average lipid chain areas and chain areas when the lipid is in a given protein environment were obtained. Proteins that have a "smooth" homogeneous surface ("cholesterol-like") and those that have inhomogeneous surfaces or that bind lipids specifically were considered. We find that T(0) can vary depending upon the interactions and that another peak can appear upon the shoulder of the main peak which reflects the melting of a eutectic mixture. The transition enthalpy decreases generally, as was found before, but when a second peak appears departures from this behavior reflect aspects of the eutectic mixture. We find that proteins have significant nonzero probabilities for being adjacent to one another so that no unbroken "annulus" of lipid necessarily exists around a protein. If T(0) does not increase much, or decreases, with increasing c, then lipids adjacent to a protein cannot all be all-trans on the time scale (10(-7) sec) of our system. Around a protein the lipid correlation depth is about one lipid layer, and this increases with c. Possible consequences of ignoring changes in polar group interactions due to clustering of proteins are discussed.

  17. Distribution of Fullerene Nanoparticles between Water and Solid Supported Lipid Membranes: Thermodynamics and Effects of Membrane Composition on Distribution.

    Science.gov (United States)

    Ha, Yeonjeong; Katz, Lynn E; Liljestrand, Howard M

    2015-12-15

    The distribution coefficient (Klipw) of fullerene between solid supported lipid membranes (SSLMs) and water was examined using different lipid membrane compositions. Klipw of fullerene was significantly higher with a cationic lipid membrane compared to that with a zwitterionic or anionic lipid membrane, potentially due to the strong interactions between negative fullerene dispersions and positive lipid head groups. The higher Klipw for fullerene distribution to ternary lipid mixture membranes was attributed to an increase in the interfacial surface area of the lipid membrane resulting from phase separation. These results imply that lipid composition can be a critical factor that affects bioconcentration of fullerene. Distribution of fullerene into zwitterionic unsaturated lipid membranes was dominated by the entropy contribution (ΔS) and the process was endothermic (ΔH > 0). This result contrasts the partitioning thermodynamics of highly and moderately hydrophobic chemicals indicating that the lipid-water distribution mechanism of fullerene may be different from that of molecular level chemicals. Potential mechanisms for the distribution of fullerene that may explain these differences include adsorption on the lipid membrane surfaces and partitioning into the center of lipid membranes (i.e., absorption).

  18. Triglyceride Blisters in Lipid Bilayers: Implications for Lipid Droplet Biogenesis and the Mobile Lipid Signal in Cancer Cell Membranes

    DEFF Research Database (Denmark)

    Khandelia, Himanshu; Duelund, Lars; Pakkanen, Kirsi Inkeri

    2010-01-01

    Triglycerides have a limited solubility, around 3%, in phosphatidylcholine lipid bilayers. Using millisecond-scale course grained molecular dynamics simulations, we show that the model lipid bilayer can accommodate a higher concentration of triolein (TO) than earlier anticipated, by sequestering...... aggregates of unknown function present in malignant cells, and to the early biogenesis of lipid droplets accommodated between the two leaflets of the endoplasmic reticulum membrane. The TO aggregates give the bilayer a blister-like appearance, and will hinder the formation of multi-lamellar phases in model...

  19. The plasma membrane: Penultimate regulator of ADAM sheddase function.

    Science.gov (United States)

    Reiss, Karina; Bhakdi, Sucharit

    2017-11-01

    ADAM10 and ADAM17 are the best characterized members of the ADAM (A Disintegrin and Metalloproteinase) - family of transmembrane proteases. Both are involved diverse physiological and pathophysiological processes. ADAMs are known to be regulated by posttranslational mechanisms. However, emerging evidence indicates that the plasma membrane with its unique dynamic properties may additionally play an important role in controlling sheddase function. Membrane events that could contribute to regulation of ADAM-function are summarized. Surface expression of peptidolytic activity should be differentiated from ADAM-sheddase function since the latter additionally requires that the protease finds its substrate in the lipid bilayer. We propose that this is achieved through horizontal and vertical reorganization of membrane nanoarchitecture coordinately occurring at the sites of sheddase activation. Reshuffling of nanodomains thereby guides traffic of enzyme and substrate to each other. For ADAM17 phosphatidylserine exposure is required to then induce its shedding function. The novel concept that physicochemical properties of the lipid bilayer govern the action of ADAM-proteases may be extendable to other functional proteins that act at the cell surface. This article is part of a Special Issue entitled: Proteolysis as a Regulatory Event in Pathophysiology edited by Stefan Rose-John. Copyright © 2017. Published by Elsevier B.V.

  20. Binding and Fusion of Extracellular Vesicles to the Plasma Membrane of Their Cell Targets.

    Science.gov (United States)

    Prada, Ilaria; Meldolesi, Jacopo

    2016-08-09

    Exosomes and ectosomes, extracellular vesicles of two types generated by all cells at multivesicular bodies and the plasma membrane, respectively, play critical roles in physiology and pathology. A key mechanism of their function, analogous for both types of vesicles, is the fusion of their membrane to the plasma membrane of specific target cells, followed by discharge to the cytoplasm of their luminal cargo containing proteins, RNAs, and DNA. Here we summarize the present knowledge about the interactions, binding and fusions of vesicles with the cell plasma membrane. The sequence initiates with dynamic interactions, during which vesicles roll over the plasma membrane, followed by the binding of specific membrane proteins to their cell receptors. Membrane binding is then converted rapidly into fusion by mechanisms analogous to those of retroviruses. Specifically, proteins of the extracellular vesicle membranes are structurally rearranged, and their hydrophobic sequences insert into the target cell plasma membrane which undergoes lipid reorganization, protein restructuring and membrane dimpling. Single fusions are not the only process of vesicle/cell interactions. Upon intracellular reassembly of their luminal cargoes, vesicles can be regenerated, released and fused horizontally to other target cells. Fusions of extracellular vesicles are relevant also for specific therapy processes, now intensely investigated.

  1. Effects of lipid membrane curvature on lipid packing state evaluated by isothermal titration calorimetry.

    Science.gov (United States)

    Yokoyama, Hirokazu; Ikeda, Keisuke; Wakabayashi, Masaki; Ishihama, Yasushi; Nakano, Minoru

    2013-01-22

    In this report, we present a novel approach for the elucidation of the physicochemical properties of lipid membranes by isothermal titration calorimetry (ITC) to quantify the heat absorbed during the solubilization of vesicles into TritonX-100 micelles. By using large and small unilamellar vesicles for comparison, this method provides calorimetric data on the gel-to-liquid-crystalline phase transition and its curvature effects and, in particular, the enthalpy change upon membrane deformation from a planar to a curved shape, which cannot be obtained by the conventional approach using differential scanning calorimetry. The results showed quantitatively that the increase in membrane curvature increases the enthalpy of 1,2-dimyristoyl-sn-glycero-3-phosphocholine membranes both below and above the phase-transition temperature, and that the effect is more significant for the former condition. The calorimetric data obtained are further discussed in relation to the elastic bending energy of the membranes and membrane-peptide interaction.

  2. Incorporation of large guest molecules into liposomes via chemical reactions in lipid membranes.

    Science.gov (United States)

    Tsuchiya, Yuki; Sugikawa, Kouta; Ueda, Masafumi; Ikeda, Atsushi

    2017-02-22

    The incorporation of hydrophobic guest molecules into lipid membranes by the exchange of the guest molecule from a cyclodextrin (CDx) complex to a liposome is limited to guest molecules that can be included in CDxs. To solve this problem, large guest molecules were incorporated into liposomes by chemical reactions of guest molecules in lipid membranes. Stable lipid-membrane-incorporated fullerene derivatives with large substituent(s) were prepared by Diels-Alder reactions in lipid membranes.

  3. Roles played by acidic lipids in HIV-1 Gag membrane binding.

    Science.gov (United States)

    Olety, Balaji; Ono, Akira

    2014-11-26

    The MA domain mediates plasma membrane (PM) targeting of HIV-1 Gag, leading to particle assembly at the PM. The interaction between MA and acidic phospholipids, in addition to N-terminal myristoyl moiety, promotes Gag binding to lipid membranes. Among acidic phospholipids, PI(4,5)P2, a PM-specific phosphoinositide, is essential for proper HIV-1 Gag localization to the PM and efficient virus particle production. Recent studies further revealed that MA-bound RNA negatively regulates HIV-1 Gag membrane binding and that PI(4,5)P2 is necessary to overcome this RNA-imposed block. In this review, we will summarize the current understanding of Gag-membrane interactions and discuss potential roles played by acidic phospholipids.

  4. Low Frequency Sound Propagation in Lipid Membranes

    CERN Document Server

    Mosgaard, Lars D; Heimburg, Thomas

    2012-01-01

    In the recent years we have shown that cylindrical biological membranes such as nerve axons under physiological conditions are able to support stable electromechanical pulses called solitons. These pulses share many similarities with the nervous impulse, e.g., the propagation velocity as well as the measured reversible heat production and changes in thickness and length that cannot be explained with traditional nerve models. A necessary condition for solitary pulse propagation is the simultaneous existence of nonlinearity and dispersion, i.e., the dependence of the speed of sound on density and frequency. A prerequisite for the nonlinearity is the presence of a chain melting transition close to physiological temperatures. The transition causes a density dependence of the elastic constants which can easily be determined by experiment. The frequency dependence is more difficult to determine. The typical time scale of a nerve pulse is 1 ms, corresponding to a characteristic frequency in the range up to one kHz. ...

  5. Uterine receptivity and the plasma membrane transformation

    Institute of Scientific and Technical Information of China (English)

    Christopher R MURPHY

    2004-01-01

    This review begins with a brief commentary on the diversity of placentation mechanisms, and then goes on to examine the extensive alterations which occur in the plasma membrane of uterine epithelial cells during early pregnancy across species. Ultrastructural, biochemical and more general morphological data reveal that strikingly common phenomena occur in this plasma membrane during early pregnancy despite the diversity of placental types-from epitheliochorial to hemochorial, which ultimately form in different species. To encapsulate the concept that common morphological and molecular alterations occur across species, that they are found basolaterally as well as apically, and that moreover they are an ongoing process during much of early pregnancy, not just an event at the time attachment,brane during early pregnancy are key to uterine receptivity.

  6. Single Molecule Kinetics of ENTH Binding to Lipid Membranes

    Energy Technology Data Exchange (ETDEWEB)

    Rozovsky, Sharon [Univ. of Delaware, Newark, DE (United States); Forstner, Martin B. [Syracuse Univ., NY (United States); Sondermann, Holger [Cornell Univ., Ithaca, NY (United States); Groves, Jay T. [Univ. of California, Berkeley, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States)

    2012-04-03

    Transient recruitment of proteins to membranes is a fundamental mechanism by which the cell exerts spatial and temporal control over proteins’ localization and interactions. Thus, the specificity and the kinetics of peripheral proteins’ membrane residence are an attribute of their function. In this article, we describe the membrane interactions of the interfacial epsin N-terminal homology (ENTH) domain with its target lipid phosphatidylinositol (4,5)-bisphosphate (PtdIns(4,5)P2). The direct visualization and quantification of interactions of single ENTH molecules with supported lipid bilayers is achieved using total internal reflection fluorescence microscopy (TIRFM) with a time resolution of 13 ms. This enables the recording of the kinetic behavior of ENTH interacting with membranes with physiologically relevant concentrations of PtdIns(4,5)P2 despite the low effective binding affinity. Subsequent single fluorophore tracking permits us to build up distributions of residence times and to measure ENTH dissociation rates as a function of membrane composition. In addition, due to the high time resolution, we are able to resolve details of the motion of ENTH associated with a simple, homogeneous membrane. In this case ENTH’s diffusive transport appears to be the result of at least three different diffusion processes.

  7. Evidence for the presence of lipid-free monomolecular apolipoprotein A-1 in plasma.

    Science.gov (United States)

    Miyazaki, Osamu; Ogihara, Jun; Fukamachi, Isamu; Kasumi, Takafumi

    2014-02-01

    The first step in reverse cholesterol transport is a process by which lipid-free or lipid-poor apoA-1 removes cholesterol from cells through the action of ATP binding cassette transporter A1 at the plasma membrane. However the structure and composition of lipid-free or -poor apoA-1 in plasma remains obscure. We previously obtained a monoclonal antibody (MAb) that specifically recognizes apoA-1 in preβ1-HDL, the smallest apoA-1-containing particle in plasma, which we used to establish a preβ1-HDL ELISA. Here, we purified preβ1-HDL from fresh normal plasma using said antibody, and analyzed the composition and structure. ApoA-1 was detected, but neither phospholipid nor cholesterol were detected in the purified preβ1-HDL. Only globular, not discoidal, particles were observed by electron microscopy. In nondenaturing PAGE, no difference in the mobility was observed between the purified preβ1-HDL and original plasma preβ1-HDL, or between the preβ1-HDL and lipid-free apoA-1 prepared by delipidating HDL. In sandwich ELISA using two anti-preβ1-HDL MAbs, reactivity with intact plasma preβ1-HDL was observed in ELISA using two MAbs with distinct epitopes but no reactivity was observed in ELISA using a single MAb, and the same phenomenon was observed with monomolecular lipid-free apoA-1. These results suggest that plasma preβ1-HDL is lipid-free monomolecular apoA-1.

  8. Vesicular and Plasma Membrane Transporters for Neurotransmitters

    OpenAIRE

    2012-01-01

    The regulated exocytosis that mediates chemical signaling at synapses requires mechanisms to coordinate the immediate response to stimulation with the recycling needed to sustain release. Two general classes of transporter contribute to release, one located on synaptic vesicles that loads them with transmitter, and a second at the plasma membrane that both terminates signaling and serves to recycle transmitter for subsequent rounds of release. Originally identified as the target of psychoacti...

  9. [Physical arrangement of membrane lipids susceptible to being used in the process of cell sorting of proteins].

    Science.gov (United States)

    Wolf, C; Quinn, P; Koumanov, K; Chachaty, C; Tenchov, B

    1999-01-01

    Detection of immiscible lipid domains in biological membranes offers an alternative support to protein sorting. Liquid ordered domains ("rafts") comprising cholesterol and saturated sphingolipids incorporate saturated glycosyl-phosphatidylinositol (GPI)-anchored or acylated (palmitoyl- and myristoyl-) proteins or particular transmembrane protein sequences. These lipid domains can be isolated in the form of Detergent resistant membranes (DRM) from biological plasma membrane preparations. Caveolae appear to be a differentiated fraction of plasma membranes comprising such numerous cross-linked microdomains associated with caveolin in different cell types. While the biological relevance of such membrane domains is evidenced in vivo by co-patching of proteins sharing the identical affinity for sphingolipids and by the disruption of co-patching following cell cholesterol depletion, only a few physical studies confort the principle of membrane heterogeneity. Results are now presented where cholesterol addition in a tertiary lipid mixture forces outphase-separation, as a realistic model where the lipid segregation can promote protein sorting to the segregated Lo phase. A lipid mixture comprising phosphatidylserine, phosphatidylethanolamine and sphingomyelin of natural origin in the ratio (1/4/3: mole/mole) has been rendered neatly heterogeneous after the addition of cholesterol (27 mole%). Xray diffraction (Small angle Xray scattering) showed the splitting of two neatly resolved lamellar diffractions in the presence of cholesterol. Above 37 degrees C the heterogeneity was traceable by a broadened diffraction spot up to the complete get-to-liquid transition of sphingomyelin at temperatures > 40 degrees C where the spot became again symmetrical and narrow. The large temperature range where the immiscible lamellar phases are detected, the specific requirement for cholesterol association with sphingomyelin, the positive influence of calcium and the reversibility of domain

  10. Targeting proteins to liquid-ordered domains in lipid membranes.

    Science.gov (United States)

    Stachowiak, Jeanne C; Hayden, Carl C; Sanchez, Mari Angelica A; Wang, Julia; Bunker, Bruce C; Voigt, James A; Sasaki, Darryl Y

    2011-02-15

    We demonstrate the construction of novel protein-lipid assemblies through the design of a lipid-like molecule, DPIDA, endowed with tail-driven affinity for specific lipid membrane phases and head-driven affinity for specific proteins. In studies performed on giant unilamellar vesicles (GUVs) with varying mole fractions of dipalymitoylphosphatidylcholine (DPPC), cholesterol, and diphytanoylphosphatidyl choline (DPhPC), DPIDA selectively partitioned into the more ordered phases, either solid or liquid-ordered (L(o)) depending on membrane composition. Fluorescence imaging established the phase behavior of the resulting quaternary lipid system. Fluorescence correlation spectroscopy confirmed the fluidity of the L(o) phase containing DPIDA. In the presence of CuCl(2), the iminodiacetic acid (IDA) headgroup of DPIDA forms the Cu(II)-IDA complex that exhibits a high affinity for histidine residues. His-tagged proteins were bound specifically to domains enriched in DPIDA, demonstrating the capacity to target protein binding selectively to both solid and L(o) phases. Steric pressure from the crowding of surface-bound proteins transformed the domains into tubules with persistence lengths that depended on the phase state of the lipid domains.

  11. The influence of solid scaffolds on flat and curved lipid membranes

    Science.gov (United States)

    de Jong, D. H.; Heuer, A.

    2017-07-01

    Solid-supported membranes have become a common tool to study lipid membrane properties in a controlled environment. One particular example is the study of membrane curvature and its effect on lipid sorting. Here we simulate solid-supported membranes using the coarse grain molecular dynamics Martini force field. We characterize basic properties of the solid surfaces and lipid membranes deposited on them. Subsequently we construct large, solid ridges and use them to induce curvature in DOPC membranes. We study membrane properties, such as lateral diffusion and tail order parameters, relative to the curved membrane. Finally, we study the effect of the induced curvature on lateral lipid sorting in a ternary lipid membrane. Thus, we obtain comprehensive and microscopic insight into the impact of curvature on a lipid membrane in terms of structure and dynamics.

  12. The influence of solid scaffolds on flat and curved lipid membranes

    Directory of Open Access Journals (Sweden)

    D. H. de Jong

    2017-07-01

    Full Text Available Solid-supported membranes have become a common tool to study lipid membrane properties in a controlled environment. One particular example is the study of membrane curvature and its effect on lipid sorting. Here we simulate solid-supported membranes using the coarse grain molecular dynamics Martini force field. We characterize basic properties of the solid surfaces and lipid membranes deposited on them. Subsequently we construct large, solid ridges and use them to induce curvature in DOPC membranes. We study membrane properties, such as lateral diffusion and tail order parameters, relative to the curved membrane. Finally, we study the effect of the induced curvature on lateral lipid sorting in a ternary lipid membrane. Thus, we obtain comprehensive and microscopic insight into the impact of curvature on a lipid membrane in terms of structure and dynamics.

  13. Important roles for membrane lipids in haloarchaeal bioenergetics.

    Science.gov (United States)

    Kellermann, Matthias Y; Yoshinaga, Marcos Y; Valentine, Raymond C; Wörmer, Lars; Valentine, David L

    2016-11-01

    Recent advances in lipidomic analysis in combination with various physiological experiments set the stage for deciphering the structure-function of haloarchaeal membrane lipids. Here we focused primarily on changes in lipid composition of Haloferax volcanii, but also performed a comparative analysis with four other haloarchaeal species (Halobacterium salinarum, Halorubrum lacusprofundi, Halorubrum sodomense and Haloplanus natans) all representing distinctive cell morphologies and behaviors (i.e., rod shape vs. pleomorphic behavior). Common to all five haloarchaea, our data reveal an extraordinary high level of menaquinone, reaching up to 72% of the total lipids. This ubiquity suggests that menaquinones may function beyond their ordinary role as electron and proton transporter, acting simultaneously as ion permeability barriers and as powerful shield against oxidative stress. In addition, we aimed at understanding the role of cations interacting with the characteristic negatively charged surface of haloarchaeal membranes. We propose for instance that by bridging the negative charges of adjacent anionic phospholipids, Mg(2+) acts as surrogate for cardiolipin, a molecule that is known to control curvature stress of membranes. This study further provides a bioenergetic perspective as to how haloarchaea evolved following oxygenation of Earth's atmosphere. The success of the aerobic lifestyle of haloarchaea includes multiple membrane-based strategies that successfully balance the need for a robust bilayer structure with the need for high rates of electron transport - collectively representing the molecular basis to inhabit hypersaline water bodies around the planet.

  14. Plasma membrane organization and dynamics is probe and cell line dependent.

    Science.gov (United States)

    Huang, Shuangru; Lim, Shi Ying; Gupta, Anjali; Bag, Nirmalya; Wohland, Thorsten

    2017-09-01

    The action and interaction of membrane receptor proteins take place within the plasma membrane. The plasma membrane, however, is not a passive matrix. It rather takes an active role and regulates receptor distribution and function by its composition and the interaction of its lipid components with embedded and surrounding proteins. Furthermore, it is not a homogenous fluid but contains lipid and protein domains of various sizes and characteristic lifetimes which are important in regulating receptor function and signaling. The precise lateral organization of the plasma membrane, the differences between the inner and outer leaflet, and the influence of the cytoskeleton are still debated. Furthermore, there is a lack of comparisons of the organization and dynamics of the plasma membrane of different cell types. Therefore, we used four different specific membrane markers to test the lateral organization, the differences between the inner and outer membrane leaflet, and the influence of the cytoskeleton of up to five different cell lines, including Chinese hamster ovary (CHO-K1), Human cervical carcinoma (HeLa), neuroblastoma (SH-SY5Y), fibroblast (WI-38) and rat basophilic leukemia (RBL-2H3) cells by Imaging Total Internal Reflection (ITIR)-Fluorescence Correlation Spectroscopy (FCS). We measure diffusion in the temperature range of 298-310K to measure the Arrhenius activation energy (EArr) of diffusion and apply the FCS diffusion law to obtain information on the spatial organization of the probe molecules on the various cell membranes. Our results show clear differences of the FCS diffusion law and EArr for the different probes in dependence of their localization. These differences are similar in the outer and inner leaflet of the membrane. However, these values can differ significantly between different cell lines raising the question how molecular plasma membrane events measured in different cell lines can be compared. This article is part of a Special Issue

  15. Buffers affect the bending rigidity of model lipid membranes.

    Science.gov (United States)

    Bouvrais, Hélène; Duelund, Lars; Ipsen, John H

    2014-01-14

    In biophysical and biochemical studies of lipid bilayers the influence of the used buffer is often ignored or assumed to be negligible on membrane structure, elasticity, or physical properties. However, we here present experimental evidence, through bending rigidity measurements performed on giant vesicles, of a more complex behavior, where the buffering molecules may considerably affect the bending rigidity of phosphatidylcholine bilayers. Furthermore, a synergistic effect on the bending modulus is observed in the presence of both salt and buffer molecules, which serves as a warning to experimentalists in the data interpretation of their studies, since typical lipid bilayer studies contain buffer and ion molecules.

  16. Membrane lipids regulate ganglioside GM2 catabolism and GM2 activator protein activity.

    Science.gov (United States)

    Anheuser, Susi; Breiden, Bernadette; Schwarzmann, Günter; Sandhoff, Konrad

    2015-09-01

    Ganglioside GM2 is the major lysosomal storage compound of Tay-Sachs disease. It also accumulates in Niemann-Pick disease types A and B with primary storage of SM and with cholesterol in type C. Reconstitution of GM2 catabolism with β-hexosaminidase A and GM2 activator protein (GM2AP) at uncharged liposomal surfaces carrying GM2 as substrate generated only a physiologically irrelevant catabolic rate, even at pH 4.2. However, incorporation of anionic phospholipids into the GM2 carrying liposomes stimulated GM2 hydrolysis more than 10-fold, while the incorporation of plasma membrane stabilizing lipids (SM and cholesterol) generated a strong inhibition of GM2 hydrolysis, even in the presence of anionic phospholipids. Mobilization of membrane lipids by GM2AP was also inhibited in the presence of cholesterol or SM, as revealed by surface plasmon resonance studies. These lipids also reduced the interliposomal transfer rate of 2-NBD-GM1 by GM2AP, as observed in assays using Förster resonance energy transfer. Our data raise major concerns about the usage of recombinant His-tagged GM2AP compared with untagged protein. The former binds more strongly to anionic GM2-carrying liposomal surfaces, increases GM2 hydrolysis, and accelerates intermembrane transfer of 2-NBD-GM1, but does not mobilize membrane lipids.

  17. Membrane lipids regulate ganglioside GM2 catabolism and GM2 activator protein activity[S

    Science.gov (United States)

    Anheuser, Susi; Breiden, Bernadette; Schwarzmann, Günter; Sandhoff, Konrad

    2015-01-01

    Ganglioside GM2 is the major lysosomal storage compound of Tay-Sachs disease. It also accumulates in Niemann-Pick disease types A and B with primary storage of SM and with cholesterol in type C. Reconstitution of GM2 catabolism with β-hexosaminidase A and GM2 activator protein (GM2AP) at uncharged liposomal surfaces carrying GM2 as substrate generated only a physiologically irrelevant catabolic rate, even at pH 4.2. However, incorporation of anionic phospholipids into the GM2 carrying liposomes stimulated GM2 hydrolysis more than 10-fold, while the incorporation of plasma membrane stabilizing lipids (SM and cholesterol) generated a strong inhibition of GM2 hydrolysis, even in the presence of anionic phospholipids. Mobilization of membrane lipids by GM2AP was also inhibited in the presence of cholesterol or SM, as revealed by surface plasmon resonance studies. These lipids also reduced the interliposomal transfer rate of 2-NBD-GM1 by GM2AP, as observed in assays using Förster resonance energy transfer. Our data raise major concerns about the usage of recombinant His-tagged GM2AP compared with untagged protein. The former binds more strongly to anionic GM2-carrying liposomal surfaces, increases GM2 hydrolysis, and accelerates intermembrane transfer of 2-NBD-GM1, but does not mobilize membrane lipids. PMID:26175473

  18. Complex roles of hybrid lipids in the composition, order, and size of lipid membrane domains.

    Science.gov (United States)

    Hassan-Zadeh, Ebrahim; Baykal-Caglar, Eda; Alwarawrah, Mohammad; Huang, Juyang

    2014-02-11

    Hybrid lipids (HL) are phospholipids with one saturated chain and one unsaturated chain. HL are hypothesized to act as linactants (i.e., 2D surfactants) in cell membranes, reducing line tension and creating nanoscopic lipid domains. Here we compare three hybrid lipids of different chain unsaturation (16:0-18:1PC (POPC), 16:0-18:2PC (PLPC), and 16:0-20:4PC (PAPC)) in their abilities to alter the composition, line tension, order, and compactness of lipid domains. We found that the liquid-ordered (Lo) and liquid-disordered (Ld) lipid domains in PAPC/di18:0PC(DSPC)/cholesterol and PLPC/DSPC/cholesterol mixtures are micrometer-sized, and only the POPC/DSPC/cholesterol system has nanoscopic domains. The results indicate that some HLs with polyunsaturated chains are not linactants, and the monounsaturated POPC displays both properties of weak linactants and "Ld-phase" lipids such as di18:1PC (DOPC). The obtained phase boundaries from giant unilamellar vesicles (GUV) show that both POPC and PLPC partition well in the Lo phases. Our MD simulations reveal that these hybrid lipids decrease the order and compactness of Lo domains. Thus, hybrid lipids distinguish themselves from other lipid groups in this combined "partitioning and loosening" ability, which could explain why the Lo domains of GUVs, which often do not contain HL, are more compact than the raft domains in cell membranes. Our line tension measurement and Monte Carlo simulation both show that even the monounsaturated POPC is a weak linactant with only modest ability to occupy domain boundaries and reduce line tension. A more important property of HLs is that they can reduce physical property differences of Lo and Ld bulk domains, which also reduces line tension at domain boundaries.

  19. Molecular dynamics simulations of the interactions of medicinal plant extracts and drugs with lipid bilayer membranes

    DEFF Research Database (Denmark)

    Kopec, Wojciech; Telenius, Jelena; Khandelia, Himanshu

    2013-01-01

    Several small drugs and medicinal plant extracts, such as the Indian spice extract curcumin, have a wide range of useful pharmacological properties that cannot be ascribed to binding to a single protein target alone. The lipid bilayer membrane is thought to mediate the effects of many...... studies of the interactions of drugs and plant extracts are therefore of interest. Molecular dynamics simulations, which can access time and length scales that are not simultaneously accessible by other experimental methods, are often used to obtain quantitative molecular and thermodynamic descriptions...... such molecules directly via perturbation of the plasma membrane structure and dynamics, or indirectly by modulating transmembrane protein conformational equilibria. Furthermore, for bioavailability, drugs must interact with and eventually permeate the lipid bilayer barrier on the surface of cells. Biophysical...

  20. Labeling the plasma membrane with TMA-DPH.

    Science.gov (United States)

    Chazotte, Brad

    2011-05-01

    INTRODUCTION TMA-DPH (trimethylamine-diphenylhexatriene) is a fluorescent membrane probe that has classically been used to label the outer leaflet of a membrane bilayer, to label the outer leaflet of the plasma membrane in cells, and to report on membrane dynamics using the techniques of fluorescence polarization and/or fluorescence lifetime. This probe has also been used to follow exocytosis and endocytosis of labeled plasma membranes. The interaction of the aqueous environment with mitochondrial inner membrane dynamics has also been studied following the fluorescence polarization and the lifetime of TMA-DPH. This protocol describes the use of TMA-DPH to label the plasma membrane.

  1. Triglyceride blisters in lipid bilayers: implications for lipid droplet biogenesis and the mobile lipid signal in cancer cell membranes.

    Directory of Open Access Journals (Sweden)

    Himanshu Khandelia

    Full Text Available Triglycerides have a limited solubility, around 3%, in phosphatidylcholine lipid bilayers. Using millisecond-scale course grained molecular dynamics simulations, we show that the model lipid bilayer can accommodate a higher concentration of triolein (TO than earlier anticipated, by sequestering triolein molecules to the bilayer center in the form of a disordered, isotropic, mobile neutral lipid aggregate, at least 17 nm in diameter, which forms spontaneously, and remains stable on at least the microsecond time scale. The results give credence to the hotly debated existence of mobile neutral lipid aggregates of unknown function present in malignant cells, and to the early biogenesis of lipid droplets accommodated between the two leaflets of the endoplasmic reticulum membrane. The TO aggregates give the bilayer a blister-like appearance, and will hinder the formation of multi-lamellar phases in model, and possibly living membranes. The blisters will result in anomalous membrane probe partitioning, which should be accounted for in the interpretation of probe-related measurements.

  2. Supramolecular organization of the sperm plasma membrane during maturation and capacitation

    Institute of Scientific and Technical Information of China (English)

    Roy Jones; Peter S. James; Liz Howes; Andreas Bruckbauer; David Klenerman

    2007-01-01

    Aim: In the present study, a variety of high resolution microscopy techniques were used to visualize the organization and motion of lipids and proteins in the sperm's plasma membrane. We have addressed questions such as the presence of diffusion barriers, confinement of molecules to specific surface domains, polarized diffusion and the role of cholesterol in regulating lipid rafts and signal transduction during capacitation. Methods: Atomic force microscopy identified a novel region (EqSS) within the equatorial segment of bovine, porcine and ovine spermatozoa that was enriched in constitutively phosphorylated proteins. The EqSS was assembled during epididymal maturation. Fluorescence imaging techniques were then used to follow molecular diffusion on the sperm head. Results: Single lipid molecules were freely exchangeable throughout the plasma membrane and showed no evidence for confinement within domains. Large lipid aggregates, however, did not cross over the boundary between the post-acrosome and equatorial segment suggesting the presence of a molecular filter between these two domains. Conclusion: A small reduction in membrane cholesterol enlarges or increases lipid rafts concomitant with phosphorylation of intracellular proteins. Excessive removal of cholesterol, however, disorganizes rafts with a cessation of phosphorylation. These techniques are forcing a revision of long-held views on how lipids and proteins in sperm membranes are assembled into larger complexes that mediate recognition and fusion with the egg.

  3. Fluorescence lifetime imaging of membrane lipid order with a ratiometric fluorescent probe.

    Science.gov (United States)

    Kilin, Vasyl; Glushonkov, Oleksandr; Herdly, Lucas; Klymchenko, Andrey; Richert, Ludovic; Mely, Yves

    2015-05-19

    To monitor the lateral segregation of lipids into liquid-ordered (Lo) and -disordered (Ld) phases in lipid membranes, environment-sensitive dyes that partition in both phases but stain them differently have been developed. Of particular interest is the dual-color F2N12S probe, which can discriminate the two phases through the ratio of its two emission bands. These bands are associated with the normal (N(∗)) and tautomer (T(∗)) excited-state species that result from an excited-state intramolecular proton transfer. In this work, we investigated the potency of the time-resolved fluorescence parameters of F2N12S to discriminate lipid phases in model and cell membranes. Both the long and mean lifetime values of the T(∗) form of F2N12S were found to differ by twofold between Ld and Lo phases as a result of the restriction in the relative motions of the two aromatic moieties of F2N12S imposed by the highly packed Lo phase. This differed from the changes in the ratio of the two emission bands between the two phases, which mainly resulted from the decreased hydration of the N(∗) form in the Lo phase. Importantly, the strong difference in lifetimes between the two phases was preserved when cholesterol was added to the Ld phase. The two phases could be imaged with high contrast by fluorescence lifetime imaging microscopy (FLIM) on giant unilamellar vesicles. FLIM images of F2N12S-labeled live HeLa cells confirmed that the plasma membrane was mainly in the Lo-like phase. Furthermore, the two phases were found to be homogeneously distributed all over the plasma membrane, indicating that they are highly mixed at the spatiotemporal resolution of the FLIM setup. Finally, FLIM could also be used to sensitively monitor the change in lipid phase upon cholesterol depletion and apoptosis.

  4. Topologically-Mediated Membrane Dynamics in Supported Lipid Bilayers

    Science.gov (United States)

    Gilmore, Sean Fitzpatrick

    2011-12-01

    This thesis is primarily design driven. It describes the development and application of dynamically tunable class of solid-fluid interfaces, which serves as a test-bed configuration for fundamental studies of soft condensed matter in reduced dimension. My specific focus is in developing these interfaces to recapitulate topology-mediated phenomenon in biological lipid membranes. The phenomena that the interfacial topology manifest in diffusional characteristics in model membranes are probed using wide-area epifluorescence microscopy and a semi-quantitative analysis of dynamic recovery following photobleaching. Furthermore, real-time remodeling of the membrane-substrate interface topology is shown to provide fundamental information regarding curvature-dependent molecular sorting and resorting. Specifically our experiments using putative raft composition mixtures confirm the conformation-dependent alignment of liquid-ordered domains and moreover reveal domain-domain interactions for the first time in model bilayers. Ongoing work aimed at delineating these inter-domain interactions in terms of membrane elastic properties is being performed. Future work that includes peptide-driven membrane deformation and sorting, as well large-scale, curvature-driven in vivo sorting of lipids is proposed and discussed.

  5. Structure and dynamics of water and lipid molecules in charged anionic DMPG lipid bilayer membranes

    DEFF Research Database (Denmark)

    Rønnest, A. K.; Peters, Günther H.J.; Hansen, Flemming Yssing;

    2016-01-01

    the sensitivity to confirm the diffusion of a small component of water bound to the lipids as found in the simulations. In addition, the orientation of the dipole moment of the water molecules has been determined as a function of their depth in the membrane. Previous indirect estimates of the electrostatic...... compared to experimental results and used to determine an average diffusion constant for all water molecules in the system. On extrapolating the diffusion constants inferred experimentally to a temperature of 310 K, reasonable agreement with the simulations is obtained. However, the experiments do not have...... potential within phospholipid membranes imply an enormous electric field of 108-109 V m-1, which is likely to have great significance in controlling the conformation of translocating membrane proteins and in the transfer of ions and molecules across the membrane. We have calculated the membrane potential...

  6. Tubular lipid membranes pulled from vesicles: Dependence of system equilibrium on lipid bilayer curvature

    Science.gov (United States)

    Golushko, I. Yu.; Rochal, S. B.

    2016-01-01

    Conditions of joint equilibrium and stability are derived for a spherical lipid vesicle and a tubular lipid membrane (TLM) pulled from this vesicle. The obtained equations establish relationships between the geometric and physical characteristics of the system and the external parameters, which have been found to be controllable in recent experiments. In particular, the proposed theory shows that, in addition to the pressure difference between internal and external regions of the system, the variable spontaneous average curvature of the lipid bilayer (forming the TLM) also influences the stability of the lipid tube. The conditions for stability of the cylindrical phase of TLMs after switching off the external force that initially formed the TLM from a vesicle are discussed. The loss of system stability under the action of a small axial force compressing the TLM is considered.

  7. Lipid membrane-mediated attraction between curvature inducing objects

    Science.gov (United States)

    van der Wel, Casper; Vahid, Afshin; Šarić, Anđela; Idema, Timon; Heinrich, Doris; Kraft, Daniela J.

    2016-09-01

    The interplay of membrane proteins is vital for many biological processes, such as cellular transport, cell division, and signal transduction between nerve cells. Theoretical considerations have led to the idea that the membrane itself mediates protein self-organization in these processes through minimization of membrane curvature energy. Here, we present a combined experimental and numerical study in which we quantify these interactions directly for the first time. In our experimental model system we control the deformation of a lipid membrane by adhering colloidal particles. Using confocal microscopy, we establish that these membrane deformations cause an attractive interaction force leading to reversible binding. The attraction extends over 2.5 times the particle diameter and has a strength of three times the thermal energy (‑3.3 kBT). Coarse-grained Monte-Carlo simulations of the system are in excellent agreement with the experimental results and prove that the measured interaction is independent of length scale. Our combined experimental and numerical results reveal membrane curvature as a common physical origin for interactions between any membrane-deforming objects, from nanometre-sized proteins to micrometre-sized particles.

  8. Forming lipid bilayer membrane arrays on micropatterned polyelectrolyte film surfaces.

    Science.gov (United States)

    Zhang, Ying; Wang, Lei; Wang, Xuejing; Qi, Guodong; Han, Xiaojun

    2013-07-01

    A novel method of forming lipid bilayer membrane arrays on micropatterned polyelectrolyte film surfaces is introduced. Polyelectrolyte films were fabricated by the layer-by-layer technique on a silicon oxide surface modified with a 3-aminopropyltriethoxysilane (APTES) monolayer. The surface pK(a) value of the APTES monolayer was determined by cyclic voltammetry to be approximately 5.61, on the basis of which a pH value of 2.0 was chosen for layer-by-layer assembly. Micropatterned polyelectrolyte films were obtained by deep-UV (254 nm) photolysis though a mask. Absorbed fluorescent latex beads were used to visualize the patterned surfaces. Lipid bilayer arrays were fabricated on the micropatterned surfaces by immersing the patterned substrates into a solution containing egg phosphatidylcholine vesicles. Fluorescence recovery after photobleaching studies yielded a lateral diffusion coefficient for probe molecules of 1.31±0.17 μm(2) s(-1) in the bilayer region, and migration of the lipid NBD PE in bilayer lipid membrane arrays was observed in an electric field.

  9. Estimation of plasma lipids and its significance on histopathological grades in oral cancer: Prognostic significance an original research

    Directory of Open Access Journals (Sweden)

    Eugenia J Sherubin

    2013-01-01

    Full Text Available Background Objectives: Alterations in the lipid profile have long been associated with various cancers because lipids play a key role in maintenance of cell integrity. This study was to estimate the plasma lipid levels in patients with oral cancer and to correlate the values with the histopathological grades. Materials and Methods: The study group included 50 patients with oral cancer aged between 20 and 60 years who had visited the Department of Oral Medicine and Radiology during the period of September 2005 to July 2007. After the histotopathological confirmation, their plasma lipid levels were estimated using auto analyzer and the data was statistically analyzed. Results: The study revealed a significant decrease in the total plasma lipid levels in patients with oral cancer in comparison with the standard values. Comparing the plasma lipid levels with the histopathological grades, we observed a significant variation in the levels of total cholesterol, very low density lipoprotein, low-density lipoprotein, high-density lipoprotein and triglycerides Conclusion: The variation in the levels of plasma cholesterol and other lipid constituents in patients with cancer might be due to their increased utilization by neoplastic cells for new membrane biosynthesis. This study was an attempt to estimate the plasma lipids in oral cancer patients and its significance on histopathological grades. We observed a relationship between lower plasma lipids and oral cancer. The result of our study strongly warrants an in-depth research with larger samples and a longer follow-up to consider the low plasma lipid status in oral cancer patients as a useful indicator to assess the course and prognosis of the disease.

  10. Analytical solution of lipid membrane morphology subjected to boundary forces on the edges of rectangular membranes

    Science.gov (United States)

    Belay, T.; Kim, C. I.; Schiavone, P.

    2016-03-01

    We develop a complete analytical solution predicting the deformation of rectangular lipid membranes resulting from boundary forces acting on the perimeter of the membrane. The shape equation describing the equilibrium state of a lipid membrane is taken from the classical Helfrich model. A linearized version of the shape equation describing membrane morphology (within the Monge representation) is obtained via a limit of superposed incremental deformations. We obtain a complete analytical solution by reducing the corresponding problem to a single partial differential equation and by using Fourier series representations for various types of boundary forces. The solution obtained predicts smooth morphological transition over the domain of interest. Finally, we note that the methods used in our analysis are not restricted to the particular type of boundary conditions considered here and can accommodate a wide class of practical and important edge conditions.

  11. Novel tilt-curvature coupling in lipid membranes

    Science.gov (United States)

    Terzi, M. Mert; Deserno, Markus

    2017-08-01

    On mesoscopic scales, lipid membranes are well described by continuum theories whose main ingredients are the curvature of a membrane's reference surface and the tilt of its lipid constituents. In particular, Hamm and Kozlov [Eur. Phys. J. E 3, 323 (2000)] have shown how to systematically derive such a tilt-curvature Hamiltonian based on the elementary assumption of a thin fluid elastic sheet experiencing internal lateral pre-stress. Performing a dimensional reduction, they not only derive the basic form of the effective surface Hamiltonian but also express its emergent elastic couplings as trans-membrane moments of lower-level material parameters. In the present paper, we argue, though, that their derivation unfortunately missed a coupling term between curvature and tilt. This term arises because, as one moves along the membrane, the curvature-induced change of transverse distances contributes to the area strain—an effect that was believed to be small but nevertheless ends up contributing at the same (quadratic) order as all other terms in their Hamiltonian. We illustrate the consequences of this amendment by deriving the monolayer and bilayer Euler-Lagrange equations for the tilt, as well as the power spectra of shape, tilt, and director fluctuations. A particularly curious aspect of our new term is that its associated coupling constant is the second moment of the lipid monolayer's lateral stress profile—which within this framework is equal to the monolayer Gaussian curvature modulus, κ¯ m. On the one hand, this implies that many theoretical predictions now contain a parameter that is poorly known (because the Gauss-Bonnet theorem limits access to the integrated Gaussian curvature); on the other hand, the appearance of κ¯ m outside of its Gaussian curvature provenance opens opportunities for measuring it by more conventional means, for instance by monitoring a membrane's undulation spectrum at short scales.

  12. Microdomains of SNARE proteins in the plasma membrane

    NARCIS (Netherlands)

    Bogaart, G. van den; Lang, T.; Jahn, R.

    2013-01-01

    Exocytosis is catalyzed by the engagement of SNARE proteins embedded in the plasma membrane with complementary SNAREs in the membrane of trafficking vesicles undergoing exocytosis. In most cells studied so far, SNAREs are not randomly distributed across the plasma membrane but are clustered and

  13. Nanoclustering as a dominant feature of plasma membrane organization

    NARCIS (Netherlands)

    Garcia-Parajo, M.F.; Cambi, A.; Torreno-Pina, J.A.; Thompson, N.; Jacobson, K.

    2014-01-01

    Early studies have revealed that some mammalian plasma membrane proteins exist in small nanoclusters. The advent of super-resolution microscopy has corroborated and extended this picture, and led to the suggestion that many, if not most, membrane proteins are clustered at the plasma membrane at

  14. Regulation of Plasma Membrane Recycling by CFTR

    Science.gov (United States)

    Bradbury, Neil A.; Jilling, Tamas; Berta, Gabor; Sorscher, Eric J.; Bridges, Robert J.; Kirk, Kevin L.

    1992-04-01

    The gene that encodes the cystic fibrosis transmembrane conductance regulator (CFTR) is defective in patients with cystic fibrosis. Although the protein product of the CFTR gene has been proposed to function as a chloride ion channel, certain aspects of its function remain unclear. The role of CFTR in the adenosine 3',5'-monophosphate (cAMP)-dependent regulation of plasma membrane recycling was examined. Adenosine 3',5'-monophosphate is known to regulate endocytosis and exocytosis in chloride-secreting epithelial cells that express CFTR. However, mutant epithelial cells derived from a patient with cystic fibrosis exhibited no cAMP-dependent regulation of endocytosis and exocytosis until they were transfected with complementary DNA encoding wild-type CFTR. Thus, CFTR is critical for cAMP-dependent regulation of membrane recycling in epithelial tissues, and this function of CFTR could explain in part the pleiotropic nature of cystic fibrosis.

  15. Elasto-plasticity in wrinkled polymerized lipid membranes

    KAUST Repository

    Chaieb, Sahraoui

    2014-01-15

    Biomembranes shown to behave like elastic sheets, can also suffer plastic deformations. Neutron scattering experiments on partially polymerised wrinkled membranes revealed that when a critical degree of polymerisation is crossed, the wrinkled membranes do not resume their spherical shapes. Instead they remain wrinkled and rigid while their non-polymerised counterparts resume their spherical floppy shapes. The yield stress of these membranes, measured for the first time via the fractal dimension, is intimately related to the degree of polymerisation probably through a 2D disorder that quenches the lateral diffusion of the lipid molecules. This work might shed light on the physical reason behind the irreversible deformation of echinocytes, acanthocytes and malaria infected red blood cells.

  16. Elasto-plasticity in wrinkled polymerized lipid membranes

    Science.gov (United States)

    Chaieb, Sahraoui

    2014-01-01

    Biomembranes shown to behave like elastic sheets, can also suffer plastic deformations. Neutron scattering experiments on partially polymerised wrinkled membranes revealed that when a critical degree of polymerisation is crossed, the wrinkled membranes do not resume their spherical shapes. Instead they remain wrinkled and rigid while their non-polymerised counterparts resume their spherical floppy shapes. The yield stress of these membranes, measured for the first time via the fractal dimension, is intimately related to the degree of polymerisation probably through a 2D disorder that quenches the lateral diffusion of the lipid molecules. This work might shed light on the physical reason behind the irreversible deformation of echinocytes, acanthocytes and malaria infected red blood cells.

  17. The mitochondria-plasma membrane contact site.

    Science.gov (United States)

    Westermann, Benedikt

    2015-08-01

    Mitochondria are dynamic organelles that are highly motile and frequently fuse and divide. It has recently become clear that their complex behavior is governed to a large extent by interactions with other cellular structures. This review will focus on a mitochondria-plasma membrane tethering complex that was recently discovered and molecularly analyzed in budding yeast, the Num1/Mdm36 complex. This complex attaches mitochondria to the cell cortex and ensures that a portion of the organelles is retained in mother cells during cell division. At the same time, it supports mitochondrial division and integrates mitochondrial dynamics into cellular architecture. Recent evidence suggests that similar mechanisms might exist also in mammalian cells.

  18. Fluid lipid membranes: from differential geometry to curvature stresses.

    Science.gov (United States)

    Deserno, Markus

    2015-01-01

    A fluid lipid membrane transmits stresses and torques that are fully determined by its geometry. They can be described by a stress- and torque-tensor, respectively, which yield the force or torque per length through any curve drawn on the membrane's surface. In the absence of external forces or torques the surface divergence of these tensors vanishes, revealing them as conserved quantities of the underlying Euler-Lagrange equation for the membrane's shape. This review provides a comprehensive introduction into these concepts without assuming the reader's familiarity with differential geometry, which instead will be developed as needed, relying on little more than vector calculus. The Helfrich Hamiltonian is then introduced and discussed in some depth. By expressing the quest for the energy-minimizing shape as a functional variation problem subject to geometric constraints, as proposed by Guven (2004), stress- and torque-tensors naturally emerge, and their connection to the shape equation becomes evident. How to reason with both tensors is then illustrated with a number of simple examples, after which this review concludes with four more sophisticated applications: boundary conditions for adhering membranes, corrections to the classical micropipette aspiration equation, membrane buckling, and membrane mediated interactions.

  19. Flow in a rotating membrane plasma separator.

    Science.gov (United States)

    Lueptow, R M; Hajiloo, A

    1995-01-01

    Rotating filter separators are very effective in the separation of plasma from whole blood, but details of the flow field in the device have not been investigated. The flow in a commercial device has been modeled computationally using the finite element code FIDAP. Taylor vortices appear in the upstream end of the annulus but disappear in the downstream end because of increasing blood viscosity as plasma is removed. Fluid transport at the upstream end of the annulus results from both translation of Taylor vortices and fluid winding around the vortices. If the inertial effects of the axial flow are reduced, less fluid winds around the vortices and more fluid is transported by the translation of the vortices. The pressure at the membrane is nonuniform in the region where vortices appear, although the relative magnitude of the fluctuations is small.

  20. Membrane potential modulates plasma membrane phospholipid dynamics and K-Ras signaling

    Science.gov (United States)

    Zhou, Yong; Wong, Ching-On; Cho, Kwang-jin; van der Hoeven, Dharini; Liang, Hong; Thakur, Dhananiay P.; Luo, Jialie; Babic, Milos; Zinsmaier, Konrad E.; Zhu, Michael X.; Hu, Hongzhen; Venkatachalam, Kartik; Hancock, John F.

    2015-01-01

    Plasma membrane depolarization can trigger cell proliferation, but how membrane potential influences mitogenic signaling is uncertain. Here, we show that plasma membrane depolarization induces nanoscale reorganization of phosphatidylserine and phosphatidylinositol 4,5-bisphosphate but not other anionic phospholipids. K-Ras, which is targeted to the plasma membrane by electrostatic interactions with phosphatidylserine, in turn undergoes enhanced nanoclustering. Depolarization-induced changes in phosphatidylserine and K-Ras plasma membrane organization occur in fibroblasts, excitable neuroblastoma cells, and Drosophila neurons in vivo and robustly amplify K-Ras–dependent mitogen-activated protein kinase (MAPK) signaling. Conversely, plasma membrane repolarization disrupts K-Ras nanoclustering and inhibits MAPK signaling. By responding to voltage-induced changes in phosphatidylserine spatiotemporal dynamics, K-Ras nanoclusters set up the plasma membrane as a biological field-effect transistor, allowing membrane potential to control the gain in mitogenic signaling circuits. PMID:26293964

  1. Hydrogen Production from Ammonia Using a Plasma Membrane Reactor

    Directory of Open Access Journals (Sweden)

    Shinji Kambara

    2016-06-01

    Full Text Available In this study, an efficient method for using pulsed plasma to produce hydrogen from ammonia was developed. An original pulsed plasma reactor with a hydrogen separation membrane was developed for efficient hydrogen production, and its hydrogen production performance was investigated. Hydrogen production in the plasma was affected by the applied voltage and flow rate of ammonia gas. The maximum hydrogen production flow rate of a typical plasma reactor was 8.7 L/h, whereas that of the plasma membrane reactor was 21.0 L/h. We found that ammonia recombination reactions in the plasma controlled hydrogen production in the plasma reactor. In the plasma membrane reactor, a significant increase in hydrogen production was obtained because ammonia recombination reactions were inhibited by the permeation of hydrogen radicals generated in the plasma through a palladium alloy membrane. The energy efficiency was 4.42 mol-H2/kWh depending on the discharge power.

  2. Amounts of phospholipids and cholesterol in lipid domains formed in intact lens membranes: Methodology development and its application to studies of porcine lens membranes.

    Science.gov (United States)

    Raguz, Marija; Mainali, Laxman; O'Brien, William J; Subczynski, Witold K

    2015-11-01

    An electron paramagnetic resonance spin-labeling method has been developed that allows quantitative evaluation of the amounts of phospholipids and cholesterol in lipid domains of intact fiber-cell plasma membranes isolated from cortical and nuclear regions of eye lenses. The long term goal of this research is the assessment of organizational changes in human lens fiber cell membranes that occur with age and during cataract development. The measurements needed to be performed on lens membranes prepared from eyes of single donors and from single eyes. For these types of studies it is necessary to separate the age/cataract related changes from preparation/technique related changes. Human lenses differ not only because of age, but also because of the varying health histories of the donors. To solve these problems the sample-to-sample preparation/technique related changes were evaluated for cortical and nuclear lens membranes prepared from single porcine eyes. It was assumed that the differences due to the age (animals were two year old) and environmental conditions for raising these animals were minimal. Mean values and standard deviations from preparation/technique changes for measured amounts of lipids in membrane domains were calculated. Statistical analysis (Student's t-test) of the data also allowed determining the differences of mean values which were statistically significant with P ≤ 0.05. These differences defined for porcine lenses will be used for comparison of amounts of lipids in domains in human lens membranes prepared from eyes of single donors and from single eyes. Greater separations will indicate that differences were statistically significant with (P ≤ 0.05) and that they came from different than preparation/technique sources. Results confirmed that in nuclear porcine membranes the amounts of lipids in domains created due to the presence of membrane proteins were greater than those in cortical membranes and the differences were larger than

  3. Transition metal ion FRET to measure short-range distances at the intracellular surface of the plasma membrane.

    Science.gov (United States)

    Gordon, Sharona E; Senning, Eric N; Aman, Teresa K; Zagotta, William N

    2016-02-01

    Biological membranes are complex assemblies of lipids and proteins that serve as platforms for cell signaling. We have developed a novel method for measuring the structure and dynamics of the membrane based on fluorescence resonance energy transfer (FRET). The method marries four technologies: (1) unroofing cells to isolate and access the cytoplasmic leaflet of the plasma membrane; (2) patch-clamp fluorometry (PCF) to measure currents and fluorescence simultaneously from a membrane patch; (3) a synthetic lipid with a metal-chelating head group to decorate the membrane with metal-binding sites; and (4) transition metal ion FRET (tmFRET) to measure short distances between a fluorescent probe and a transition metal ion on the membrane. We applied this method to measure the density and affinity of native and introduced metal-binding sites in the membrane. These experiments pave the way for measuring structural rearrangements of membrane proteins relative to the membrane.

  4. Carboxylic Acids Plasma Membrane Transporters in Saccharomyces cerevisiae.

    Science.gov (United States)

    Casal, Margarida; Queirós, Odília; Talaia, Gabriel; Ribas, David; Paiva, Sandra

    2016-01-01

    This chapter covers the functionally characterized plasma membrane carboxylic acids transporters Jen1, Ady2, Fps1 and Pdr12 in the yeast Saccharomyces cerevisiae, addressing also their homologues in other microorganisms, as filamentous fungi and bacteria. Carboxylic acids can either be transported into the cells, to be used as nutrients, or extruded in response to acid stress conditions. The secondary active transporters Jen1 and Ady2 can mediate the uptake of the anionic form of these substrates by a H(+)-symport mechanism. The undissociated form of carboxylic acids is lipid-soluble, crossing the plasma membrane by simple diffusion. Furthermore, acetic acid can also be transported by facilitated diffusion via Fps1 channel. At the cytoplasmic physiological pH, the anionic form of the acid prevails and it can be exported by the Pdr12 pump. This review will highlight the mechanisms involving carboxylic acids transporters, and the way they operate according to the yeast cell response to environmental changes, as carbon source availability, extracellular pH and acid stress conditions.

  5. Structure and function of thyroid hormone plasma membrane transporters.

    Science.gov (United States)

    Schweizer, Ulrich; Johannes, Jörg; Bayer, Dorothea; Braun, Doreen

    2014-09-01

    Thyroid hormones (TH) cross the plasma membrane with the help of transporter proteins. As charged amino acid derivatives, TH cannot simply diffuse across a lipid bilayer membrane, despite their notorious hydrophobicity. The identification of monocarboxylate transporter 8 (MCT8, SLC16A2) as a specific and very active TH transporter paved the way to the finding that mutations in the MCT8 gene cause a syndrome of psychomotor retardation in humans. The purpose of this review is to introduce the current model of transmembrane transport and highlight the diversity of TH transmembrane transporters. The interactions of TH with plasma transfer proteins, T3 receptors, and deiodinase are summarized. It is shown that proteins may bind TH owing to their hydrophobic character in hydrophobic cavities and/or by specific polar interaction with the phenolic hydroxyl, the aminopropionic acid moiety, and by weak polar interactions with the iodine atoms. These findings are compared with our understanding of how TH transporters interact with substrate. The presumed effects of mutations in MCT8 on protein folding and transport function are explained in light of the available homology model.

  6. Effects of Iron Overload on the Activity of Na,K-ATPase and Lipid Profile of the Human Erythrocyte Membrane

    Science.gov (United States)

    Sousa, Leilismara; Garcia, Israel J. P.; Costa, Tamara G. F.; Silva, Lilian N. D.; Renó, Cristiane O.; Oliveira, Eneida S.; Tilelli, Cristiane Q.; Santos, Luciana L.; Cortes, Vanessa F.; Santos, Herica L.; Barbosa, Leandro A.

    2015-01-01

    Iron is an essential chemical element for human life. However, in some pathological conditions, such as hereditary hemochromatosis type 1 (HH1), iron overload induces the production of reactive oxygen species that may lead to lipid peroxidation and a change in the plasma-membrane lipid profile. In this study, we investigated whether iron overload interferes with the Na,K-ATPase activity of the plasma membrane by studying erythrocytes that were obtained from the whole blood of patients suffering from iron overload. Additionally, we treated erythrocytes of normal subjects with 0.8 mM H2O2 and 1 μM FeCl3 for 24 h. We then analyzed the lipid profile, lipid peroxidation and Na,K-ATPase activity of plasma membranes derived from these cells. Iron overload was more frequent in men (87.5%) than in women and was associated with an increase (446%) in lipid peroxidation, as indicated by the amount of the thiobarbituric acid reactive substances (TBARS) and an increase (327%) in the Na,K-ATPase activity in the plasma membrane of erythrocytes. Erythrocytes treated with 1 μM FeCl3 for 24 h showed an increase (132%) in the Na,K-ATPase activity but no change in the TBARS levels. Iron treatment also decreased the cholesterol and phospholipid content of the erythrocyte membranes and similar decreases were observed in iron overload patients. In contrast, erythrocytes treated with 0.8 mM H2O2 for 24 h showed no change in the measured parameters. These results indicate that erythrocytes from patients with iron overload exhibit higher Na,K-ATPase activity compared with normal subjects and that this effect is specifically associated with altered iron levels. PMID:26197432

  7. Effects of Iron Overload on the Activity of Na,K-ATPase and Lipid Profile of the Human Erythrocyte Membrane.

    Directory of Open Access Journals (Sweden)

    Leilismara Sousa

    Full Text Available Iron is an essential chemical element for human life. However, in some pathological conditions, such as hereditary hemochromatosis type 1 (HH1, iron overload induces the production of reactive oxygen species that may lead to lipid peroxidation and a change in the plasma-membrane lipid profile. In this study, we investigated whether iron overload interferes with the Na,K-ATPase activity of the plasma membrane by studying erythrocytes that were obtained from the whole blood of patients suffering from iron overload. Additionally, we treated erythrocytes of normal subjects with 0.8 mM H2O2 and 1 μM FeCl3 for 24 h. We then analyzed the lipid profile, lipid peroxidation and Na,K-ATPase activity of plasma membranes derived from these cells. Iron overload was more frequent in men (87.5% than in women and was associated with an increase (446% in lipid peroxidation, as indicated by the amount of the thiobarbituric acid reactive substances (TBARS and an increase (327% in the Na,K-ATPase activity in the plasma membrane of erythrocytes. Erythrocytes treated with 1 μM FeCl3 for 24 h showed an increase (132% in the Na,K-ATPase activity but no change in the TBARS levels. Iron treatment also decreased the cholesterol and phospholipid content of the erythrocyte membranes and similar decreases were observed in iron overload patients. In contrast, erythrocytes treated with 0.8 mM H2O2 for 24 h showed no change in the measured parameters. These results indicate that erythrocytes from patients with iron overload exhibit higher Na,K-ATPase activity compared with normal subjects and that this effect is specifically associated with altered iron levels.

  8. Critical composition fluctuations in artificial and cell-derived lipid membranes

    Science.gov (United States)

    Honerkamp-Smith, Aurelia

    2014-03-01

    Cell plasma membranes contain a mixture of lipid types which can segregate into coexisting liquids, a thermodynamic phenomenon which may contribute to biological functions. Simplified, artificial three-component lipid vesicles can be prepared which display a critical miscibility transition near room temperature. We found that such vesicles exhibit concentration fluctuations whose size, composition, and timescales vary consistently with critical exponents for two-dimensional conserved order parameter systems. However, the critical miscibility transition is also observed in vesicles formed directly from the membranes of living cells, despite their more complex composition and the presence of membrane proteins. I will describe our critical fluctuation measurements and also review a variety of more recent work by other researchers. Proximity to a critical point alters the spatial distribution and aggregation tendencies of proteins, and makes lipid mixtures more susceptible to domain formation by protein-mediated interactions, such as adhesion zones. Recent work suggests that critical temperature depression may also be relevant to the mechanism of anaesthetic action.

  9. Genetic Analysis of Arabidopsis Mutants Impaired in Plastid Lipid Import Reveals a Role of Membrane Lipids in Chloroplast Division

    Energy Technology Data Exchange (ETDEWEB)

    Fan, J.; Xu, C.

    2011-03-01

    The biogenesis of photosynthetic membranes in plants relies largely on lipid import from the endoplasmic reticulum (ER) and this lipid transport process is mediated by TGD proteins in Arabidopsis. Such a dependency of chloroplast biogenesis on ER-to-plastid lipid transport was recently exemplified by analyzing double mutants between tgd1-1 or tgd4-3 and fad6 mutants. The fad6 mutants are defective in the desaturation of membrane lipids in chloroplasts and therefore dependent on import of polyunsaturated lipid precursors from the ER for constructing a competent thylakoid membrane system. In support of a critical role of TGD proteins in ER-to-plastid lipid trafficking, we showed that the introduction of the tgd mutations into fad6 mutant backgrounds led to drastic reductions in relative amounts of thylakoid lipids. Moreover, the tgd1-1 fad6 and tgd4-3 fad6 double mutants were deficient in polyunsaturated fatty acids in chloroplast membrane lipids, and severely compromised in the biogenesis of photosynthetic membrane systems. Here we report that these double mutants are severely impaired in chloroplast division. The possible role of membrane lipids in chloroplast division is discussed.

  10. Super-Resolution Microscopy: Shedding Light on the Cellular Plasma Membrane.

    Science.gov (United States)

    Stone, Matthew B; Shelby, Sarah A; Veatch, Sarah L

    2017-02-17

    Lipids and the membranes they form are fundamental building blocks of cellular life, and their geometry and chemical properties distinguish membranes from other cellular environments. Collective processes occurring within membranes strongly impact cellular behavior and biochemistry, and understanding these processes presents unique challenges due to the often complex and myriad interactions between membrane components. Super-resolution microscopy offers a significant gain in resolution over traditional optical microscopy, enabling the localization of individual molecules even in densely labeled samples and in cellular and tissue environments. These microscopy techniques have been used to examine the organization and dynamics of plasma membrane components, providing insight into the fundamental interactions that determine membrane functions. Here, we broadly introduce the structure and organization of the mammalian plasma membrane and review recent applications of super-resolution microscopy to the study of membranes. We then highlight some inherent challenges faced when using super-resolution microscopy to study membranes, and we discuss recent technical advancements that promise further improvements to super-resolution microscopy and its application to the plasma membrane.

  11. Carrier-mediated ion transport in lipid bilayer membranes.

    Science.gov (United States)

    Laprade, R; Grenier, F; Pagé-Dansereau, M; Dansereau, J

    1984-08-01

    The electrical properties predicted by a widely accepted model for carrier-mediated ion transport in lipid bilayers are described. The different steps leading to ion transport and their associated rate constants are reaction at the interface between an ion in the aqueous phase and a carrier in the membrane (kRi), followed by translocation of the ion-carrier complex across the membrane interior (kis) and its dissociation at the other interface (kDi) after which the free carrier crosses back the membrane interior (ks). Results on glyceryl monooleate (GMO) membranes for a family of homologue carriers, the macrotetralide actin antibiotics (nonactin, monactin, dinactin, trinactin, and tetranactin) and a variety of ions (Na+, Cs+, Rb+, K+, NH4+, and Tl+) are presented. Internally consistent data obtained from steady-state electrical measurements (zero-current potential and conductance, current-voltage relationship) allow us to obtain the equilibrium permeability ratios for the different ions and show that for a given carrier kRi is relatively invariant from one ion to the other, except for Tl+ (larger), which implies that the ionic selectivity is controlled by the dissociation of the complex. The values of the individual rate constants obtained from current relaxation experiments are also presented and confirm the findings from steady-state measurements, as well as the isostericity concept for complexes of different ions with the same carrier (kis invariant). These also allow us to determine the aqueous phase membrane and torus membrane partition coefficients. Finally, the observed increase in kis from nonactin to tetranactin and, for all homologues, from GMO-decane to solvent-free GMO membranes, together with the concomitant decrease in kDi, can be explained in terms of modifications of electrostatic energy profiles induced by variations in carrier size and membrane thickness.

  12. Dynamical and structural properties of lipid membranes in relation to liposomal drug delivery systems

    DEFF Research Database (Denmark)

    Jørgensen, Kent; Høyrup, Lise Pernille Kristine; Pedersen, Tina B.

    2001-01-01

    The structural and dynamical properties of DPPC liposomes containing lipopolymers (PEG-lipids) and charged DPPS lipids have been,studied in relation to the lipid membrane interaction of enzymes and peptides. The results suggest that both the lipid membrane structure and dynamics and in particular...... the appearance of small-scale lipid structures might be of importance for the activity of membrane associated and liposome degrading enzymes as well as for the membrane interaction of acylated peptides. The combined experimental and simulation results are of relevance for a rational development of peptide loaded...

  13. Plasma membrane ubiquinone controls ceramide production and prevents cell death induced by serum withdrawal.

    Science.gov (United States)

    Barroso, M P; Gómez-Díaz, C; Villalba, J M; Burón, M I; López-Lluch, G; Navas, P

    1997-06-01

    Serum provides cultured cells with survival factors required to maintain growth. Its withdrawal induces the development of programmed cell death. HL-60 cells were sensitive to serum removal, and an increase of lipid peroxidation and apoptosis was observed. Long-term treatment with ethidium bromide induced the mitochondria-deficient rho(o)HL-60 cell line. These cells were surprisingly more resistant to serum removal, displaying fewer apoptotic cells and lower lipid peroxidation. HL-60 cells contained less ubiquinone at the plasma membrane than rho(o)HL-60 cells. Both cell types increased plasma membrane ubiquinone in response to serum removal, although this increase was much higher in rho(o) cells. Addition of ubiquinone to both cell cultures in the absence of serum improved cell survival with decreasing lipid peroxidation and apoptosis. Ceramide was accumulated after serum removal in HL-60 but not in rho(o)HL-60 cells, and exogenous ubiquinone reduced this accumulation. These results demonstrate a relationship between ubiquinone levels in the plasma membrane and the induction of serum withdrawal-induced apoptosis, and ceramide accumulation. Thus, ubiquinone, which is a central component of the plasma membrane electron transport system, can represent a first level of protection against oxidative damage caused by serum withdrawal.

  14. SAXS investigations on lipid membranes under osmotic stress

    Energy Technology Data Exchange (ETDEWEB)

    Rubim, R.L.; Vieira, V.; Gerbelli, B.B.; Teixeira da Silva, E.R.; Oliveira, C.L.P.; Oliveira, E.A. [Universidade de Sao Paulo (USP), Sao Paulo, SP (Brazil)

    2012-07-01

    Full text: In this work we, experimentally, investigate the interactions between lipid bilayers. A structural characterization is performed by small angle x-ray scattering (SAXS) on multilamellar systems under known osmotic pressure. Changes in the composition of membranes can modify their mechanical properties and structural parameters, like the flexibility of these membranes, which plays a key role on the determination of the tridimensional organization of bilayers. The membranes are composed of soya lecithin, where the major component is DPPC (Dipalmitoylphosphatidylcholine), and fatty acids are incorporated to the membrane in different concentrations, in order to turn the membrane more fluid. The membranes are inserted in a solution of PVP [poly(vinyl-pyrrolidone) - 40000] and the polymer will apply an osmotic pressure on them. The osmotic pressure is controlled by preparing PVP solutions of desired composition and, as we know the concentration of polymer in solution, we can obtain the intensity of the osmotic pressure. SAXS experiments were done in order to determine the distance between the bilayer. From the position of the Bragg peaks, the lamellar periodicity (the thickness of the membranes plus their distance of separation) was determined. Using theoretical model for the form and structure factors we fitted those experimental data and determined the thickness of the membranes. The distance between the membranes was controlled by the osmotic pressure (P) applied to the membranes and, for a given pressure, we determine the distance between the bilayers (a) on equilibrium. The experimental curve P(a) is theoretically described by the different contributions from van der Waals, hydration and fluctuation forces. From the fitting of experimental curves, relevant parameters characterizing the strength of the different interactions are obtained, such as Hamaker and rigidity constant [2, 3]. We observe that the separation between the bilayers on equilibrium is

  15. Segregation of PIP2 and PIP3 into distinct nanoscale regions within the plasma membrane

    Directory of Open Access Journals (Sweden)

    Jie Wang

    2012-07-01

    PIP2 and PIP3 are implicated in a wide variety of cellular signaling pathways at the plasma membrane. We have used STORM imaging to localize clusters of PIP2 and PIP3 to distinct nanoscale regions within the plasma membrane of PC12 cells. With anti-phospholipid antibodies directly conjugated with AlexaFluor 647, we found that PIP2 clusters in membrane domains of 64.5±27.558 nm, while PIP3 clusters had a size of 125.6±22.408 nm. With two color direct STORM imaging we show that >99% of phospholipid clusters have only one or other phospholipid present. These results indicate that lipid nano-domains can be readily identified using super-resolution imaging techniques, and that the lipid composition and size of clusters is tightly regulated.

  16. Direct Capture of Functional Proteins from Mammalian Plasma Membranes into Nanodiscs.

    Science.gov (United States)

    Roy, Jahnabi; Pondenis, Holly; Fan, Timothy M; Das, Aditi

    2015-10-20

    Mammalian plasma membrane proteins make up the largest class of drug targets yet are difficult to study in a cell free system because of their intransigent nature. Herein, we perform direct encapsulation of plasma membrane proteins derived from mammalian cells into a functional nanodisc library. Peptide fingerprinting was used to analyze the proteome of the incorporated proteins in nanodiscs and to further demonstrate that the lipid composition of the nanodiscs directly affects the class of protein that is incorporated. Furthermore, the functionality of the incorporated membrane proteome was evaluated by measuring the activity of membrane proteins: Na(+)/K(+)-ATPase and receptor tyrosine kinases. This work is the first report of the successful establishment and characterization of a cell free functional library of mammalian membrane proteins into nanodiscs.

  17. The importance of plasma membrane coenzyme Q in aging and stress responses.

    Science.gov (United States)

    Navas, Plácido; Villalba, José Manuel; de Cabo, Rafael

    2007-06-01

    The plasma membrane of eukaryotic cells is the limit to interact with the environment. This position implies receiving stress signals that affects its components such as phospholipids. Inserted inside these components is coenzyme Q that is a redox compound acting as antioxidant. Coenzyme Q is reduced by diverse dehydrogenase enzymes mainly NADH-cytochrome b(5) reductase and NAD(P)H:quinone reductase 1. Reduced coenzyme Q can prevent lipid peroxidation chain reaction by itself or by reducing other antioxidants such as alpha-tocopherol and ascorbate. The group formed by antioxidants and the enzymes able to reduce coenzyme Q constitutes a plasma membrane redox system that is regulated by conditions that induce oxidative stress. Growth factor removal, ethidium bromide-induced rho degrees cells, and vitamin E deficiency are some of the conditions where both coenzyme Q and its reductases are increased in the plasma membrane. This antioxidant system in the plasma membrane has been observed to participate in the healthy aging induced by calorie restriction. Furthermore, coenzyme Q regulates the release of ceramide from sphingomyelin, which is concentrated in the plasma membrane. This results from the non-competitive inhibition of the neutral sphingomyelinase by coenzyme Q particularly by its reduced form. Coenzyme Q in the plasma membrane is then the center of a complex antioxidant system preventing the accumulation of oxidative damage and regulating the externally initiated ceramide signaling pathway.

  18. Visualization of lipid domains of lipid domains in giant unilamellar vesicles using an environment-sensitive membrane probe based on 3-hydroxyflavone

    DEFF Research Database (Denmark)

    Klymchenko, Andrey; Oncul, Sule; Didier, Pascal

    2009-01-01

    of the photoselection with the Ld phase indicated no predominant orientation of the probe in the Ld phase. Comparison of the present results with those reported previously for F2N12S in living cells suggests a high content of the Lo phase in the outer leaflet of the cell plasma membranes. Taking into account the high......We characterized the recently introduced environment-sensitive fluorescent membrane probe based on 3-hydroxyflavone, F2N12S, in model lipid membranes displaying liquid disordered (Ld) phase, liquid ordered (Lo) phase, or their coexistence. Steady-state fluorescence studies in large unilamellar...... vesicles show that the probe dual emission drastically changes with the lipid bilayer phase, which can be correlated with the difference in their hydration. Using two-photon excitation microscopy on giant unilamellar vesicles, the F2N12S probe was found to bind both Ld and Lo phases, allowing visualization...

  19. The insertion and transport of anandamide in synthetic lipid membranes are both cholesterol-dependent.

    Directory of Open Access Journals (Sweden)

    Eric Di Pasquale

    Full Text Available BACKGROUND: Anandamide is a lipid neurotransmitter which belongs to a class of molecules termed the endocannabinoids involved in multiple physiological functions. Anandamide is readily taken up into cells, but there is considerable controversy as to the nature of this transport process (passive diffusion through the lipid bilayer vs. involvement of putative proteic transporters. This issue is of major importance since anandamide transport through the plasma membrane is crucial for its biological activity and intracellular degradation. The aim of the present study was to evaluate the involvement of cholesterol in membrane uptake and transport of anandamide. METHODOLOGY/PRINCIPAL FINDINGS: Molecular modeling simulations suggested that anandamide can adopt a shape that is remarkably complementary to cholesterol. Physicochemical studies showed that in the nanomolar concentration range, anandamide strongly interacted with cholesterol monolayers at the air-water interface. The specificity of this interaction was assessed by: i the lack of activity of structurally related unsaturated fatty acids (oleic acid and arachidonic acid at 50 nM on cholesterol monolayers, and ii the weak insertion of anandamide into phosphatidylcholine or sphingomyelin monolayers. In agreement with these data, the presence of cholesterol in reconstituted planar lipid bilayers triggered the stable insertion of anandamide detected as an increase in bilayer capacitance. Kinetics transport studies showed that pure phosphatidylcholine bilayers were weakly permeable to anandamide. The incorporation of cholesterol in phosphatidylcholine bilayers dose-dependently stimulated the translocation of anandamide. CONCLUSIONS/SIGNIFICANCE: Our results demonstrate that cholesterol stimulates both the insertion of anandamide into synthetic lipid monolayers and bilayers, and its transport across bilayer membranes. In this respect, we suggest that besides putative anandamide protein

  20. Biological Membranes in Extreme Conditions: Simulations of Anionic Archaeal Tetraether Lipid Membranes.

    Directory of Open Access Journals (Sweden)

    Luis Felipe Pineda De Castro

    Full Text Available In contrast to the majority of organisms that have cells bound by di-ester phospholipids, archaeal membranes consist of di- and tetraether phospholipids. Originating from organisms that withstand harsh conditions (e.g., low pH and a wide range of temperatures such membranes have physical properties that make them attractive materials for biological research and biotechnological applications. We developed force-field parameters based on the widely used Generalized Amber Force Field (GAFF to enable the study of anionic tetraether membranes of the model archaean Sulfolobus acidocaldarius by computer simulations. The simulations reveal that the physical properties of these unique membranes depend on the number of cyclopentane rings included in each lipid unit, and on the size of cations that are used to ensure charge neutrality. This suggests that the biophysical properties of Sulfolobus acidocaldarius cells depend not only on the compositions of their membranes but also on the media in which they grow.

  1. Biological Membranes in Extreme Conditions: Simulations of Anionic Archaeal Tetraether Lipid Membranes

    Science.gov (United States)

    Pineda De Castro, Luis Felipe; Dopson, Mark

    2016-01-01

    In contrast to the majority of organisms that have cells bound by di-ester phospholipids, archaeal membranes consist of di- and tetraether phospholipids. Originating from organisms that withstand harsh conditions (e.g., low pH and a wide range of temperatures) such membranes have physical properties that make them attractive materials for biological research and biotechnological applications. We developed force-field parameters based on the widely used Generalized Amber Force Field (GAFF) to enable the study of anionic tetraether membranes of the model archaean Sulfolobus acidocaldarius by computer simulations. The simulations reveal that the physical properties of these unique membranes depend on the number of cyclopentane rings included in each lipid unit, and on the size of cations that are used to ensure charge neutrality. This suggests that the biophysical properties of Sulfolobus acidocaldarius cells depend not only on the compositions of their membranes but also on the media in which they grow. PMID:27167213

  2. Shear-Driven Circulation Patterns in Lipid Membrane Vesicles

    CERN Document Server

    Woodhouse, Francis G; 10.1017/jfm.2012.118

    2012-01-01

    Recent experiments have shown that when a near-hemispherical lipid vesicle attached to a solid surface is subjected to a simple shear flow it exhibits a pattern of membrane circulation much like a dipole vortex. This is in marked contrast to the toroidal circulation that would occur in the related problem of a drop of immiscible fluid attached to a surface and subjected to shear. This profound difference in flow patterns arises from the lateral incompressibility of the membrane, which restricts the observable flows to those in which the velocity field in the membrane is two-dimensionally divergence free. Here we study these circulation patterns within the simplest model of membrane fluid dynamics. A systematic expansion of the flow field based on Papkovich--Neuber potentials is developed for general viscosity ratios between the membrane and the surrounding fluids. Comparison with experimental results [C. V\\'ezy, G. Massiera, and A. Viallat, Soft Matter 3, 844 (2007)] is made, and it is shown how such studies ...

  3. Solubility and permeation of hydrogen sulfide in lipid membranes.

    Directory of Open Access Journals (Sweden)

    Ernesto Cuevasanta

    Full Text Available Hydrogen sulfide (H(2S is mainly known for its toxicity but has recently been shown to be produced endogenously in mammalian tissues and to be associated with physiological regulatory functions. To better understand the role of biomembranes in modulating its biological distribution and effects; we measured the partition coefficient of H(2S in models of biological membranes. The partition coefficients were found to be 2.1±0.2, 1.9±0.5 and 2.0±0.6 in n-octanol, hexane and dilauroylphosphatidylcholine liposome membranes relative to water, respectively (25°C. This two-fold higher concentration of H(2S in the membrane translates into a rapid membrane permeability, P(m = 3 cm s(-1. We used a mathematical model in three dimensions to gain insight into the diffusion of total sulfide in tissues. This model shows that the sphere of action of sulfide produced by a single cell expands to involve more than 200 neighboring cells, and that the resistance imposed by lipid membranes has a significant effect on the diffusional spread of sulfide at pH 7.4, increasing local concentrations. These results support the role of hydrogen sulfide as a paracrine signaling molecule and reveal advantageous pharmacokinetic properties for its therapeutic applications.

  4. Solubility and permeation of hydrogen sulfide in lipid membranes.

    Science.gov (United States)

    Cuevasanta, Ernesto; Denicola, Ana; Alvarez, Beatriz; Möller, Matías N

    2012-01-01

    Hydrogen sulfide (H(2)S) is mainly known for its toxicity but has recently been shown to be produced endogenously in mammalian tissues and to be associated with physiological regulatory functions. To better understand the role of biomembranes in modulating its biological distribution and effects; we measured the partition coefficient of H(2)S in models of biological membranes. The partition coefficients were found to be 2.1±0.2, 1.9±0.5 and 2.0±0.6 in n-octanol, hexane and dilauroylphosphatidylcholine liposome membranes relative to water, respectively (25°C). This two-fold higher concentration of H(2)S in the membrane translates into a rapid membrane permeability, P(m) = 3 cm s(-1). We used a mathematical model in three dimensions to gain insight into the diffusion of total sulfide in tissues. This model shows that the sphere of action of sulfide produced by a single cell expands to involve more than 200 neighboring cells, and that the resistance imposed by lipid membranes has a significant effect on the diffusional spread of sulfide at pH 7.4, increasing local concentrations. These results support the role of hydrogen sulfide as a paracrine signaling molecule and reveal advantageous pharmacokinetic properties for its therapeutic applications.

  5. Visualization of lipid domains in giant unilamellar vesicles using an environment-sensitive membrane probe based on 3-hydroxyflavone.

    Science.gov (United States)

    Klymchenko, Andrey S; Oncul, Sule; Didier, Pascal; Schaub, Emmanuel; Bagatolli, Luis; Duportail, Guy; Mély, Yves

    2009-02-01

    We characterized the recently introduced environment-sensitive fluorescent membrane probe based on 3-hydroxyflavone, F2N12S, in model lipid membranes displaying liquid disordered (Ld) phase, liquid ordered (Lo) phase, or their coexistence. Steady-state fluorescence studies in large unilamellar vesicles show that the probe dual emission drastically changes with the lipid bilayer phase, which can be correlated with the difference in their hydration. Using two-photon excitation microscopy on giant unilamellar vesicles, the F2N12S probe was found to bind both Ld and Lo phases, allowing visualization of the individual phases from the fluorescence intensity ratio of its two emission bands. By using a linearly polarized excitation light, a strong photoselection was observed for F2N12S in the Lo phase, indicating that its fluorophore is nearly parallel to the lipid chains of the bilayer. In contrast, the absence of the photoselection with the Ld phase indicated no predominant orientation of the probe in the Ld phase. Comparison of the present results with those reported previously for F2N12S in living cells suggests a high content of the Lo phase in the outer leaflet of the cell plasma membranes. Taking into account the high selectivity of F2N12S for the cell plasma membranes and its suitability for both single- and two-photon excitation, applications of this probe to study membrane lateral heterogeneity in biological membranes are foreseen.

  6. Influence of a hyperlipidic diet on the composition of the non-membrane lipid pool of red blood cells of male and female rats

    Directory of Open Access Journals (Sweden)

    Xavier Remesar

    2015-07-01

    Full Text Available Background and objectives. Red blood cells (RBC are continuously exposed to oxidative agents, affecting their membrane lipid function. However, the amount of lipid in RBCs is higher than the lipids of the cell membrane, and includes triacylglycerols, which are no membrane components. We assumed that the extra lipids originated from lipoproteins attached to the cell surface, and we intended to analyse whether the size and composition of this lipid pool were affected by sex or diet.Experimental design. Adult male and female Wistar rats were fed control or cafeteria diets. Packed blood cells and plasma lipids were extracted and analysed for fatty acids by methylation and GC-MS, taking care of not extracting membrane lipids.Results. The absence of ω3-PUFA in RBC extracts (but not in plasma suggest that the lipids extracted were essentially those in the postulated lipid surface pool and not those in cell membrane. In cells’ extracts, there was a marked depletion of PUFA (and, in general, of insaturation. Fatty acid patterns were similar for all groups studied, with limited effects of sex and no effects of diet in RBC (but not in plasma fatty acids. Presence of trans fatty acids was small but higher in RBC lipids, and could not be justified by dietary sources.Conclusions. The presence of a small layer of lipid on the RBC surface may limit oxidative damage to the cell outer structures, and help explain its role in the transport of lipophilic compounds. However, there may be other, so far uncovered, additional functions for this lipid pool.

  7. LDL uptake by Leishmania amazonensis: involvement of membrane lipid microdomains.

    Science.gov (United States)

    De Cicco, Nuccia N T; Pereira, Miria G; Corrêa, José R; Andrade-Neto, Valter V; Saraiva, Felipe B; Chagas-Lima, Alessandra C; Gondim, Katia C; Torres-Santos, Eduardo C; Folly, Evelize; Saraiva, Elvira M; Cunha-E-Silva, Narcisa L; Soares, Maurilio J; Atella, Georgia C

    2012-04-01

    Leishmania amazonensis lacks a de novo mechanism for cholesterol synthesis and therefore must scavenge this lipid from the host environment. In this study we show that the L. amazonensis takes up and metabolizes human LDL(1) particles in both a time and dose-dependent manner. This mechanism implies the presence of a true LDL receptor because the uptake is blocked by both low temperature and by the excess of non-labelled LDL. This receptor is probably associated with specific microdomains in the membrane of the parasite, such as rafts, because this process is blocked by methyl-β-cyclodextrin (MCBD). Cholesteryl ester fluorescently-labeled LDL (BODIPY-cholesteryl-LDL) was used to follow the intracellular distribution of this lipid. After uptake it was localized in large compartments along the parasite body. The accumulation of LDL was analyzed by flow cytometry using FITC-labeled LDL particles. Together these data show for the first time that L. amazonensis is able to compensate for its lack of lipid synthesis through the use of a lipid importing machinery largely based on the uptake of LDL particles from the host. Understanding the details of the molecular events involved in this mechanism may lead to the identification of novel targets to block Leishmania infection in human hosts.

  8. An ER protein functionally couples neutral lipid metabolism on lipid droplets to membrane lipid synthesis in the ER

    Science.gov (United States)

    Markgraf, Daniel F.; Klemm, Robin W.; Junker, Mirco; Hannibal-Bach, Hans K.; Ejsing, Christer S.; Rapoport, Tom A.

    2014-01-01

    Eukaryotic cells store neutral lipids, such as triacylglycerol (TAG), in lipid droplets (LDs). Here, we have addressed how LDs are functionally linked to the endoplasmic reticulum (ER). We show in S. cerevisiae that LD growth is sustained by LD-localized enzymes. When LDs grow in early stationary phase, the diacylglycerol acyl-transferase Dga1p moves from the ER to LDs and is responsible for all TAG synthesis from diacylglycerol (DAG). During LD breakdown in early exponential phase, an ER membrane protein, Ice2p, facilitates TAG utilization for membrane-lipid synthesis. Ice2p has a cytosolic domain with affinity for LDs and is required for the efficient utilization of LD-derived DAG in the ER. Ice2p breaks a futile cycle on LDs between TAG-degradation and -synthesis, promoting the rapid re-localization of Dga1p to the ER. Our results show that Ice2p functionally links LDs with the ER, and explain how cells switch neutral lipid metabolism from storage to consumption. PMID:24373967

  9. An ER protein functionally couples neutral lipid metabolism on lipid droplets to membrane lipid synthesis in the ER.

    Science.gov (United States)

    Markgraf, Daniel F; Klemm, Robin W; Junker, Mirco; Hannibal-Bach, Hans K; Ejsing, Christer S; Rapoport, Tom A

    2014-01-16

    Eukaryotic cells store neutral lipids such as triacylglycerol (TAG) in lipid droplets (LDs). Here, we have addressed how LDs are functionally linked to the endoplasmic reticulum (ER). We show that, in S. cerevisiae, LD growth is sustained by LD-localized enzymes. When LDs grow in early stationary phase, the diacylglycerol acyl-transferase Dga1p moves from the ER to LDs and is responsible for all TAG synthesis from diacylglycerol (DAG). During LD breakdown in early exponential phase, an ER membrane protein (Ice2p) facilitates TAG utilization for membrane-lipid synthesis. Ice2p has a cytosolic domain with affinity for LDs and is required for the efficient utilization of LD-derived DAG in the ER. Ice2p breaks a futile cycle on LDs between TAG degradation and synthesis, promoting the rapid relocalization of Dga1p to the ER. Our results show that Ice2p functionally links LDs with the ER and explain how cells switch neutral lipid metabolism from storage to consumption.

  10. An ER Protein Functionally Couples Neutral Lipid Metabolism on Lipid Droplets to Membrane Lipid Synthesis in the ER

    Directory of Open Access Journals (Sweden)

    Daniel F. Markgraf

    2014-01-01

    Full Text Available Eukaryotic cells store neutral lipids such as triacylglycerol (TAG in lipid droplets (LDs. Here, we have addressed how LDs are functionally linked to the endoplasmic reticulum (ER. We show that, in S. cerevisiae, LD growth is sustained by LD-localized enzymes. When LDs grow in early stationary phase, the diacylglycerol acyl-transferase Dga1p moves from the ER to LDs and is responsible for all TAG synthesis from diacylglycerol (DAG. During LD breakdown in early exponential phase, an ER membrane protein (Ice2p facilitates TAG utilization for membrane-lipid synthesis. Ice2p has a cytosolic domain with affinity for LDs and is required for the efficient utilization of LD-derived DAG in the ER. Ice2p breaks a futile cycle on LDs between TAG degradation and synthesis, promoting the rapid relocalization of Dga1p to the ER. Our results show that Ice2p functionally links LDs with the ER and explain how cells switch neutral lipid metabolism from storage to consumption.

  11. Hydrostatic Pressure Promotes Domain Formation in Model Lipid Raft Membranes.

    Science.gov (United States)

    Worcester, David L; Weinrich, Michael

    2015-11-01

    Neutron diffraction measurements demonstrate that hydrostatic pressure promotes liquid-ordered (Lo) domain formation in lipid membranes prepared as both oriented multilayers and unilamellar vesicles made of a canonical ternary lipid mixture for which demixing transitions have been extensively studied. The results demonstrate an unusually large dependence of the mixing transition on hydrostatic pressure. Additionally, data at 28 °C show that the magnitude of increase in Lo caused by 10 MPa pressure is much the same as the decrease in Lo produced by twice minimum alveolar concentrations (MAC) of general anesthetics such as halothane, nitrous oxide, and xenon. Therefore, the results may provide a plausible explanation for the reversal of general anesthesia by hydrostatic pressure.

  12. Small Intestine but Not Liver Lysophosphatidylcholine Acyltransferase 3 (Lpcat3) Deficiency Has a Dominant Effect on Plasma Lipid Metabolism.

    Science.gov (United States)

    Kabir, Inamul; Li, Zhiqiang; Bui, Hai H; Kuo, Ming-Shang; Gao, Guangping; Jiang, Xian-Cheng

    2016-04-01

    Lysophosphatidylcholine acyltransferase 3 (Lpcat3) is involved in phosphatidylcholine remodeling in the small intestine and liver. We investigated lipid metabolism in inducible intestine-specific and liver-specificLpcat3gene knock-out mice. We producedLpcat3-Flox/villin-Cre-ER(T2)mice, which were treated with tamoxifen (at days 1, 3, 5, and 7), to deleteLpcat3specifically in the small intestine. At day 9 after the treatment, we found that Lpcat3 deficiency in enterocytes significantly reduced polyunsaturated phosphatidylcholines in the enterocyte plasma membrane and reduced Niemann-Pick C1-like 1 (NPC1L1), CD36, ATP-binding cassette transporter 1 (ABCA1), and ABCG8 levels on the membrane, thus significantly reducing lipid absorption, cholesterol secretion through apoB-dependent and apoB-independent pathways, and plasma triglyceride, cholesterol, and phospholipid levels, as well as body weight. Moreover, Lpcat3 deficiency does not cause significant lipid accumulation in the small intestine. We also utilized adenovirus-associated virus-Cre to depleteLpcat3in the liver. We found that liver deficiency only reduces plasma triglyceride levels but not other lipid levels. Furthermore, there is no significant lipid accumulation in the liver. Importantly, small intestine Lpcat3 deficiency has a much bigger effect on plasma lipid levels than that of liver deficiency. Thus, inhibition of small intestine Lpcat3 might constitute a novel approach for treating hyperlipidemia.

  13. Association of plasma lipid levels with atherosclerosis prevalence in psittaciformes.

    Science.gov (United States)

    Beaufrère, Hugues; Vet, Dr Med; Cray, Carolyn; Ammersbach, Mélanie; Tully, Thomas N

    2014-09-01

    The prevalence of atherosclerosis is high in the captive psittacine population and increases with age and female sex. The genera Psittacus, Amazona, and Nymphicus are predisposed to atherosclerosis, whereas the genera Cacatua and Ara are less susceptible. Plasma cholesterol and lipoprotein abnormalities have been suggested as risk factors in the development of atherosclerosis as observed in mammals. To investigate whether the psittacine genera susceptibility to atherosclerosis and the known risk factors of age and sex could be associated with differences in the lipid profile, a retrospective analysis was conducted on blood lipid values from 5625 birds. Prevalence values were obtained from a previously published, large, case-control study and were compared with identified trends in plasma lipid profiles. Genus-specific differences were identified in plasma total cholesterol values that corresponded to observed trends in the prevalence of clinically important atherosclerotic lesions, which were also highly correlated. The effect of age was significant but was mild and may not account for the dramatic increase in atherosclerosis prevalence observed with age. In addition, Quaker parrots ( Myiopsitta monachus ), which were used as experimental models for psittacine atherosclerosis and dyslipidemia, were found to have the highest values in all lipid profile parameters. The results of this study suggest that the differences observed in prevalence among species of the psittacine genera may partly be explained by differences in plasma total cholesterol levels. Results also support the use of Quaker parrots as models for studying atherosclerosis and dyslipidemia.

  14. Electroporation of archaeal lipid membranes using MD simulations.

    Science.gov (United States)

    Polak, Andraž; Tarek, Mounir; Tomšič, Matija; Valant, Janez; Ulrih, Nataša Poklar; Jamnik, Andrej; Kramar, Peter; Miklavčič, Damijan

    2014-12-01

    Molecular dynamics (MD) simulations were used to investigate the electroporation of archaeal lipid bilayers when subjected to high transmembrane voltages induced by a charge imbalance, mimicking therefore millisecond electric pulse experiments. The structural characteristics of the bilayer, a 9:91 mol% 2,3-di-O-sesterterpanyl-sn-glicerol-1-phospho-myo-inositol (AI) and 2,3-di-O-sesterterpanyl-sn-glicerol-1-phospho-1'(2'-O-α-D-glucosyl)-myo-inositol (AGI) were compared to small angle X-ray scattering data. A rather good agreement of the electron density profiles at temperatures of 298 and 343 K was found assessing therefore the validity of the protocols and force fields used in simulations. Compared to dipalmitoyl-phosphatidylcholine (DPPC), the electroporation threshold for the bilayer was found to increase from ~2 V to 4.3 V at 323 K, and to 5.2 V at 298 K. Comparing the electroporation thresholds of the archaeal lipids to those of simple diphytanoyl-phosphatidylcholine (DPhPC) bilayers (2.5 V at 323 K) allowed one to trace back the stability of the membranes to the structure of their lipid head groups. Addition of DPPC in amounts of 50 mol% to the archaeal lipid bilayers decreases their stability and lowers the electroporation thresholds to 3.8 V and 4.1 V at respectively 323 and 298 K. The present study therefore shows how membrane compositions can be selected to cover a wide range of responses to electric stimuli. This provides new routes for the design of liposomes that can be efficiently used as drug delivery carriers, as the selection of their composition allows one to tune in their electroporation threshold for subsequent release of their load.

  15. Changes in Plasma Lipids during Exposure to Total Sleep Deprivation.

    Science.gov (United States)

    Chua, Eric Chern-Pin; Shui, Guanghou; Cazenave-Gassiot, Amaury; Wenk, Markus R; Gooley, Joshua J

    2015-11-01

    The effects of sleep loss on plasma lipids, which play an important role in energy homeostasis and signaling, have not been systematically examined. Our aim was to identify lipid species in plasma that increase or decrease reliably during exposure to total sleep deprivation. Twenty individuals underwent sleep deprivation in a laboratory setting. Blood was drawn every 4 h and mass spectrometry techniques were used to analyze concentrations of 263 lipid species in plasma, including glycerolipids, glycerophospholipids, sphingolipids, and sterols. Chronobiology and Sleep Laboratory, Duke-NUS Graduate Medical School. Healthy ethnic-Chinese males aged 21-28 y (n = 20). Subjects were kept awake for 40 consecutive hours. Each metabolite time series was modeled as a sum of sinusoidal (circadian) and linear components, and we assessed whether the slope of the linear component differed from zero. More than a third of all individually analyzed lipid profiles exhibited a circadian rhythm and/or a linear change in concentration during sleep deprivation. Twenty-five lipid species showed a linear and predominantly unidirectional trend in concentration levels that was consistent across participants. Choline plasmalogen levels decreased, whereas several phosphatidylcholine (PC) species and triacylglycerides (TAG) carrying polyunsaturated fatty acids increased. The decrease in choline plasmalogen levels during sleep deprivation is consistent with prior work demonstrating that these lipids are susceptible to degradation by oxidative stress. The increase in phosphatidylcholines and triacylglycerides suggests that sleep loss might modulate lipid metabolism, which has potential implications for metabolic health in individuals who do not achieve adequate sleep. © 2015 Associated Professional Sleep Societies, LLC.

  16. Computer Simulation of Cytoskeleton-Induced Blebbing in Lipid Membranes

    CERN Document Server

    Spangler, Eric J; Revalee, Joel D; Kumar, P B Sunil; Laradji, Mohamed

    2011-01-01

    Blebs are balloon-shaped membrane protrusions that form during many physiological processes. Using computer simulation of a particle-based model for self-assembled lipid bilayers coupled to an elastic meshwork, we investigated the phase behavior and kinetics of blebbing. We found that blebs form for large values of the ratio between the areas of the bilayer and the cytoskeleton. We also found that blebbing can be induced when the cytoskeleton is subject to a localized ablation or a uniform compression. The results obtained are qualitatively in agreement with the experimental evidence and the model opens up the possibility to study the kinetics of bleb formation in detail.

  17. Direct affinity of dopamine to lipid membranes investigated by Nuclear Magnetic Resonance spectroscopy.

    Science.gov (United States)

    Matam, Yashasvi; Ray, Bruce D; Petrache, Horia I

    2016-04-08

    Dopamine, a naturally occurring neurotransmitter, plays an important role in the brain's reward system and acts on sensory receptors in the brain. Neurotransmitters are contained in lipid membraned vesicles and are released by exocytosis. All neurotransmitters interact with transport and receptor proteins in glial cells, on neuronal dendrites, and at the axonal button, and also must interact with membrane lipids. However, the extent of direct interaction between lipid membranes in the absence of receptors and transport proteins has not been extensively investigated. In this report, we use UV and NMR spectroscopy to determine the affinity and the orientation of dopamine interacting with lipid vesicles made of either phosphatidylcholine (PC) or phosphatidylserine (PS) lipids which are primary lipid components of synaptic vesicles. We quantify the interaction of dopamine's aromatic ring with lipid membranes using our newly developed method that involves reference spectra in hydrophobic environments. Our measurements show that dopamine interacts with lipid membranes primarily through the aromatic side opposite to the hydroxyl groups, with this aromatic side penetrating deeper into the hydrophobic region of the membrane. Since dopamine's activity involves its release into extracellular space, we have used our method to also investigate dopamine's release from lipid vesicles. We find that dopamine trapped inside PC and PS vesicles is released into the external solution despite its affinity to membranes. This result suggests that dopamine's interaction with lipid membranes is complex and involves both binding as well as permeation through lipid bilayers, a combination that could be an effective trigger for apoptosis of dopamine-generating cells.

  18. Inverse lyotropic phases of lipids and membrane curvature

    Energy Technology Data Exchange (ETDEWEB)

    Shearman, G C; Ces, O; Templer, R H; Seddon, J M [Department of Chemistry, Imperial College London, SW7 2AZ (United Kingdom)

    2006-07-19

    In recent years it has become evident that many biological functions and processes are associated with the adoption by cellular membranes of complex geometries, at least locally. In this paper, we initially discuss the range of self-assembled structures that lipids, the building blocks of biological membranes, may form, focusing specifically on the inverse lyotropic phases of negative interfacial mean curvature. We describe the roles of curvature elasticity and packing frustration in controlling the stability of these inverse phases, and the experimental determination of the spontaneous curvature and the curvature elastic parameters. We discuss how the lyotropic phase behaviour can be tuned by the addition of compounds such as long-chain alkanes, which can relieve packing frustration. The latter section of the paper elaborates further on the structure, geometric properties, and stability of the inverse bicontinuous cubic phases.

  19. Quantitative studies of antimicrobial peptide-lipid membrane interactions

    DEFF Research Database (Denmark)

    Kristensen, Kasper

    into such novel therapeutics. However, limited understanding of the mechanisms underlying microbicidal activity of antimicrobial peptides has slowed down this development. A central step toward understanding the microbicidal mechanisms of action of antimicrobial peptides is to understand the mechanisms by which......The increasing occurrence of multi-drug-resistant bacteria poses a serious threat to modern society. Therefore, novel types of anti-infective therapeutics are highly warranted. Antimicrobial peptides are a class of naturally occurring host-defense molecules that potentially might be developed...... antimicrobial peptides interact with phospholipid membranes. Motivated by that fact, the scope of this thesis is to study these antimicrobial peptide-lipid membrane interactions. In particular, we attempt to study these interactions with a quantitative approach. For that purpose, we consider the three...

  20. Nonidet P-40 extraction of lymphocyte plasma membrane. Characterization of the insoluble residue.

    Science.gov (United States)

    Davies, A A; Wigglesworth, N M; Allan, D; Owens, R J; Crumpton, M J

    1984-04-01

    Purified preparations of lymphocyte plasma membrane were extracted exhaustively with Nonidet P-40 in Dulbecco's phosphate-buffered saline medium. The insoluble fraction, as defined by sedimentation at 10(6) g-min, contained about 10% of the membrane protein as well as cholesterol and phospholipid. The lipid/protein ratio, cholesterol/phospholipid ratio and sphingomyelin content were increased in the residue. Density-gradient centrifugation suggested that the lipid and protein form a common entity. As judged by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis, the Nonidet P-40-insoluble fractions of the plasma membranes of human B lymphoblastoid cells and pig mesenteric lymph-node lymphocytes possessed similar qualitative polypeptide compositions but differed quantitatively. Both residues comprised major polypeptides of Mr 28 000, 33 000, 45 000 and 68 000, together with a prominent band of Mr 120 000 in the human and of Mr 200 000 in the pig. The polypeptides of Mr 28 000, 33 000, 68 000 and 120 000 were probably located exclusively in the Nonidet P-40-insoluble residue, which also possessed a 4-fold increase in 5'-nucleotidase specific activity. The results indicate that a reproducible fraction of lymphocyte plasma membrane is insoluble in non-ionic detergents and that this fraction possesses a unique polypeptide composition. By analogy with similar studies with erythrocyte ghosts, it appears likely that the polypeptides are located on the plasma membrane's cytoplasmic face.

  1. Elucidating how bamboo salt interacts with supported lipid membranes: influence of alkalinity on membrane fluidity.

    Science.gov (United States)

    Jeong, Jong Hee; Choi, Jae-Hyeok; Kim, Min Chul; Park, Jae Hyeon; Herrin, Jason Scott; Kim, Seung Hyun; Lee, Haiwon; Cho, Nam-Joon

    2015-07-01

    Bamboo salt is a traditional medicine produced from sea salt. It is widely used in Oriental medicine and is an alkalizing agent with reported antiinflammatory, antimicrobial and chemotherapeutic properties. Notwithstanding, linking specific molecular mechanisms with these properties has been challenging to establish in biological systems. In part, this issue may be related to bamboo salt eliciting nonspecific effects on components found within these systems. Herein, we investigated the effects of bamboo salt solution on supported lipid bilayers as a model system to characterize the interaction between lipid membranes and bamboo salt. The atomic composition of unprocessed and processed bamboo salts was first analyzed by mass spectrometry, and we identified several elements that have not been previously reported in other bamboo salt preparations. The alkalinity of hydrated samples was also measured and determined to be between pH 10 and 11 for bamboo salts. The effect of processed bamboo salt solutions on the fluidic properties of a supported lipid bilayer on glass was next investigated by fluorescence recovery after photobleaching (FRAP) analysis. It was demonstrated that, with increasing ionic strength of the bamboo salt solution, the fluidity of a lipid bilayer increased. On the contrary, increasing the ionic strength of near-neutral buffer solutions with sodium chloride salt diminished fluidity. To reconcile these two observations, we identified that solution alkalinity is critical for the effects of bamboo salt on membrane fluidity, as confirmed using three additional commercial bamboo salt preparations. Extended-DLVO model calculations support that the effects of bamboo salt on lipid membranes are due to the alkalinity imparting a stronger hydration force. Collectively, the results of this work demonstrate that processing of bamboo salt strongly affects its atomic composition and that the alkalinity of bamboo salt solutions contributes to its effect on membrane

  2. Alteration of membrane lipid biophysical properties and resistance of human lung adenocarcinoma A549 cells to cisplatin

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Alterations of membrane lipid biophysical properties of sensitiveA549 and resistant A549/DDP cells to the Cis-dichlorodiammine platinum (Cisplatin) were performed by measurements of fluorescence and flow cytometry approaches using fluorescence dyes of DPH, N-AS and Merocyanine 540 (MC 540) respectively. Fatty acids of membrane lipid of the two cell lines were analyzed by gas chromatography. The results indicated clearly that fluorescence polarization (P) of the DPH probe is 0.169 for the sensitive A549 cell and 0.194 for the resistant A549/DDP cells. Statistical analysis showed significant difference between the two cell lines. The polarizations of 2-AS and 7-AS which reflect the fluidity of surface and middle of lipid bilayer are 0.134 and 0.144 for the sensitive A549 cells as well as 0.171 and 0.178 for the resistant A549/DDP cells respectively, but there is no significant difference of the polarization of 12-AS between the two cell lines. This shows that altera-tions of the membrane fluidity of both cells were mainly located on the surface and middle of the lipid bilayer. In addition, the packing density of phospholipid molecules in the membrane of the two cell lines detected by MC540 probe indicated that lipid packing of A549 cell membranes was looser than that of the A549/DDP cells. And unsaturation degree of plasma membrane fatty acids of the A549/DDP cells was also lower than that of A549 cells. Taken together, it was proposed that the al-teration of membrane lipid biophysical state may be involved in the resistance of A549/DDP cells to cisplatin.

  3. Shiga toxin induces membrane reorganization and formation of long range lipid order

    DEFF Research Database (Denmark)

    Solovyeva, Vita; Johannes, Ludger; Simonsen, Adam Cohen

    2015-01-01

    microscopy. A content of 1% of glycosphingolipid globotriaosylceramide (Gb3) receptor lipids in a bilayer was used to bind the Shiga toxin B-subunit to the surface of gel domains. Binding of the Shiga toxin B-subunit to lipids led to the modulation of orientational membrane texture in gel domains and induced...... membrane reordering. When Shiga toxin was added above the lipid chain melting temperature, the toxin interaction with the membrane induced rearrangement and clustering of Gb3 lipids that resulted in the long range order and alignment of lipids in gel domains. The toxin induced redistribution of Gb3 lipids...... inside gel domains is governed by the temperature at which Shiga toxin was added to the membrane: above or below the phase transition. The temperature is thus one of the critical factors controlling lipid organization and texture in the presence of Shiga toxin. Lipid chain ordering imposed by Shiga toxin...

  4. Detergent interaction with tethered bilayer lipid membranes for protein reconstitution

    Science.gov (United States)

    Broccio, Matteo; Zan Goh, Haw; Loesche, Mathias

    2009-03-01

    Tethered bilayer lipid membranes (tBLMs) are self-assembled biomimetic structures in which the membrane is separated from a solid substrate by a nm-thick hydrated submembrane space. These model systems are being used in binding studies of peripheral proteins and exotoxins. Here we aim at their application for the reconstitution of water-insoluble integral membrane proteins. As an alternative to fusion of preformed proteoliposomes we study the direct reconstitution of such proteins for applications in biosensing and pharmaceutical screening. For reconstitution, highly insulating tBLMs (R˜10^5-10^6 φ) were temporarily incubated with a detergent to screen for conditions that keep the detergent-saturated membranestable and ready to incorporate detergent-solubilized proteins. We assess the electrical characteristics, i.e. specific resistance and capacitance, by means of electrochemical impedance spectroscopy (EIS) under timed incubation with decylmaltoside and dodecylmaltoside detergents in a regime around their critical micelle concentration, 1.8 mM and 0.17 mM respectively and demonstrate the restoration of the tBLM upon detergent removal. Thereby a range of concentration and incubation times was identified, that represents optimal conditions for the subsequent membrane protein reconstitution.

  5. Effect of physical constraints on the mechanisms of membrane fusion: bolaform lipid vesicles as model systems.

    OpenAIRE

    1996-01-01

    Bolaform lipid vesicles were used to study the effect of physical constraints on membrane fusion. In these vesicles the membrane is organized in a single monolayer, because of the presence of covalent bonds in its middle plane. Therefore, the formation of fusion intermediates is subject to higher energy barriers and greater geometrical constraints than is usual in bilayer membranes. Bolaform lipids were extracted from the thermophilic archaeon Sulfolobus solfataricus. These lipids can be divi...

  6. Imaging of mobile long-lived nanoplatforms in the live cell plasma membrane.

    Science.gov (United States)

    Brameshuber, Mario; Weghuber, Julian; Ruprecht, Verena; Gombos, Imre; Horváth, Ibolya; Vigh, László; Eckerstorfer, Paul; Kiss, Endre; Stockinger, Hannes; Schütz, Gerhard J

    2010-12-31

    The plasma membrane has been hypothesized to contain nanoscopic lipid platforms, which are discussed in the context of "lipid rafts" or "membrane rafts." Based on biochemical and cell biological studies, rafts are believed to play a crucial role in many signaling processes. However, there is currently not much information on their size, shape, stability, surface density, composition, and heterogeneity. We present here a method that allows for the first time the direct imaging of nanoscopic long-lived platforms with raft-like properties diffusing in the live cell plasma membrane. Our method senses these platforms by their property to assemble a characteristic set of fluorescent marker proteins or lipids on a time scale of seconds. A special photobleaching protocol was used to reduce the surface density of labeled mobile platforms down to the level of well isolated diffraction-limited spots without altering the single spot brightness. The statistical distribution of probe molecules per platform was determined by single molecule brightness analysis. For demonstration, we used the consensus raft marker glycosylphosphatidylinositol-anchored monomeric GFP and the fluorescent lipid analog BODIPY-G(M1), which preferentially partitions into liquid-ordered phases. For both markers, we found cholesterol-dependent homo-association in the plasma membrane of living CHO and Jurkat T cells in the resting state, thereby demonstrating the existence of small, mobile, long-lived platforms containing these probes. We further applied the technology to address structural changes in the plasma membrane during fever-type heat shock: at elevated temperatures, the glycosylphosphatidylinositol-anchored monomeric GFP homo-association disappeared, accompanied by an increase in the expression of the small heat shock protein Hsp27.

  7. Proving lipid rafts exist: membrane domains in the prokaryote Borrelia burgdorferi have the same properties as eukaryotic lipid rafts.

    Directory of Open Access Journals (Sweden)

    Timothy J LaRocca

    Full Text Available Lipid rafts in eukaryotic cells are sphingolipid and cholesterol-rich, ordered membrane regions that have been postulated to play roles in many membrane functions, including infection. We previously demonstrated the existence of cholesterol-lipid-rich domains in membranes of the prokaryote, B. burgdorferi, the causative agent of Lyme disease [LaRocca et al. (2010 Cell Host & Microbe 8, 331-342]. Here, we show that these prokaryote membrane domains have the hallmarks of eukaryotic lipid rafts, despite lacking sphingolipids. Substitution experiments replacing cholesterol lipids with a set of sterols, ranging from strongly raft-promoting to raft-inhibiting when mixed with eukaryotic sphingolipids, showed that sterols that can support ordered domain formation are both necessary and sufficient for formation of B. burgdorferi membrane domains that can be detected by transmission electron microscopy or in living organisms by Förster resonance energy transfer (FRET. Raft-supporting sterols were also necessary and sufficient for formation of high amounts of detergent resistant membranes from B. burgdorferi. Furthermore, having saturated acyl chains was required for a biotinylated lipid to associate with the cholesterol-lipid-rich domains in B. burgdorferi, another characteristic identical to that of eukaryotic lipid rafts. Sterols supporting ordered domain formation were also necessary and sufficient to maintain B. burgdorferi membrane integrity, and thus critical to the life of the organism. These findings provide compelling evidence for the existence of lipid rafts and show that the same principles of lipid raft formation apply to prokaryotes and eukaryotes despite marked differences in their lipid compositions.

  8. Channelopathies linked to plasma membrane phosphoinositides.

    Science.gov (United States)

    Logothetis, Diomedes E; Petrou, Vasileios I; Adney, Scott K; Mahajan, Rahul

    2010-07-01

    The plasma membrane phosphoinositide phosphatidylinositol 4,5-bisphosphate (PIP2) controls the activity of most ion channels tested thus far through direct electrostatic interactions. Mutations in channel proteins that change their apparent affinity to PIP2 can lead to channelopathies. Given the fundamental role that membrane phosphoinositides play in regulating channel activity, it is surprising that only a small number of channelopathies have been linked to phosphoinositides. This review proposes that for channels whose activity is PIP2-dependent and for which mutations can lead to channelopathies, the possibility that the mutations alter channel-PIP2 interactions ought to be tested. Similarly, diseases that are linked to disorders of the phosphoinositide pathway result in altered PIP2 levels. In such cases, it is proposed that the possibility for a concomitant dysregulation of channel activity also ought to be tested. The ever-growing list of ion channels whose activity depends on interactions with PIP2 promises to provide a mechanism by which defects on either the channel protein or the phosphoinositide levels can lead to disease.

  9. Analysis of Membrane Lipids of Airborne Micro-Organisms

    Science.gov (United States)

    MacNaughton, Sarah

    2006-01-01

    A method of characterization of airborne micro-organisms in a given location involves (1) large-volume filtration of air onto glass-fiber filters; (2) accelerated extraction of membrane lipids of the collected micro-organisms by use of pressurized hot liquid; and (3) identification and quantitation of the lipids by use of gas chromatography and mass spectrometry. This method is suitable for use in both outdoor and indoor environments; for example, it can be used to measure airborne microbial contamination in buildings ("sick-building syndrome"). The classical approach to analysis of airborne micro-organisms is based on the growth of cultureable micro-organisms and does not provide an account of viable but noncultureable micro-organisms, which typically amount to more than 90 percent of the micro-organisms present. In contrast, the present method provides an account of all micro-organisms, including cultureable, noncultureable, aerobic, and anaerobic ones. The analysis of lipids according to this method makes it possible to estimate the number of viable airborne micro-organisms present in the sampled air and to obtain a quantitative profile of the general types of micro-organisms present along with some information about their physiological statuses.

  10. Towards structural and functional analysis of the plant plasma membrane proton pump

    DEFF Research Database (Denmark)

    Justesen, Bo Højen

    The plasma membrane H+-ATPase is a proton pump essential for several physiological important processes in plants. Through the extrusion of protons from the cell, the PM H+-ATPase establishes and maintains a proton gradient used by proton coupled transporters and secondary active transport......, and regulation of H+-ATPases, key questions, in particular concerning the detailed interaction of regulator proteins with the H+-ATPases, remains answering that may require the use of new approaches. In this work the proton pump Arabidopsis thaliana plasma membrane H+-ATPase isoform 2 has been reconstituted...... into soluble nanoscale lipid bilayers, also termed nanodiscs. Extensive analysis confirms the correct assembly and reconstitution of active proton pump into nanodiscs. The pump inserts as a monomer, which through activity analysis confirms this as the minimal functional unit of the plasma membrane H...

  11. TNAP, an Essential Player in Membrane Lipid Rafts of Neuronal Cells.

    Science.gov (United States)

    Ermonval, Myriam; Baychelier, Florence; Fonta, Caroline

    2015-01-01

    The tissue non-specific alkaline phosphatase (TNAP) is a glycosyl-phosphatidylinositol (GPI) anchored glycoprotein which exists under different forms and is expressed in different tissues. As the other members of the ecto-phosphatase family, TNAP is targeted to membrane lipid rafts. Such micro domains enriched in particular lipids, are involved in cell sorting, are in close contact with the cellular cytoskeleton and play the role of signaling platform. In addition to its location in functional domains, the extracellular orientation of TNAP and the fact this glycoprotein can be shed from plasma membranes, contribute to its different phosphatase activities by acting as a phosphomonoesterase on various soluble substrates (inorganic pyrophosphate -PPi-, pyridoxal phosphate -PLP-, phosphoethanolamine -PEA-), as an ectonucleotidase on nucleotide-phosphate and presumably as a phosphatase able to dephosphorylate phosphoproteins and phospholipids associated to cells or to extra cellular matrix. More and more data accumulate on an involvement of the brain TNAP both in physiological and pathological situations. This review will summarize what is known and expected from the TNAP localization in lipid rafts with a particular emphasis on the role of a neuronal microenvironment on its potential function in the central nervous system.

  12. Following the trail of lipids: Signals initiated by PI3K function at multiple cellular membranes.

    Science.gov (United States)

    Naguib, Adam

    2016-05-17

    Phosphatidylinositol 3,4,5-trisphosphate [PtdIns(3,4,5)P3] is the signaling currency of the phosphoinositide 3-kinase (PI3K)/AKT pathway; transduction through this axis depends on this signaling lipid. Formation of PtdIns(3,4,5)P3 is dictated not only by PI3K activation but also by the localization and access of PI3K to its substrate PtdIns(4,5)P2 (phosphatidylinositol 4,5-bisphosphate). PI3K/AKT-mediated signaling is antagonized by PtdIns(3,4,5)P3 dephosphorylation. Although previously typically considered an event associated with the plasma membrane, it is now appreciated that the formation and metabolism of PtdIns(3,4,5)P3 occur on multiple membranes with distinct kinetics. Modulated activity of phosphatidylinositol lipid kinases and phosphatases contributes to intricately orchestrated lipid gradients that define the signaling status of the pathway at multiple sites within the cell.

  13. Correlating anomalous diffusion with lipid bilayer membrane structure using single molecule tracking and atomic force microscopy

    Science.gov (United States)

    Skaug, Michael J.; Faller, Roland; Longo, Marjorie L.

    2011-06-01

    Anomalous diffusion has been observed abundantly in the plasma membrane of biological cells, but the underlying mechanisms are still unclear. In general, it has not been possible to directly image the obstacles to diffusion in membranes, which are thought to be skeleton bound proteins, protein aggregates, and lipid domains, so the dynamics of diffusing particles is used to deduce the obstacle characteristics. We present a supported lipid bilayer system in which we characterized the anomalous diffusion of lipid molecules using single molecule tracking, while at the same time imaging the obstacles to diffusion with atomic force microscopy. To explain our experimental results, we performed lattice Monte Carlo simulations of tracer diffusion in the presence of the experimentally determined obstacle configurations. We correlate the observed anomalous diffusion with obstacle area fraction, fractal dimension, and correlation length. To accurately measure an anomalous diffusion exponent, we derived an expression to account for the time-averaging inherent to all single molecule tracking experiments. We show that the length of the single molecule trajectories is critical to the determination of the anomalous diffusion exponent. We further discuss our results in the context of confinement models and the generating stochastic process.

  14. A reusable device for electrochemical applications of hydrogel supported black lipid membranes.

    Science.gov (United States)

    Mech-Dorosz, Agnieszka; Heiskanen, Arto; Bäckström, Sania; Perry, Mark; Muhammad, Haseena B; Hélix-Nielsen, Claus; Emnéus, Jenny

    2015-02-01

    Black lipid membranes (BLMs) are significant in studies of membrane transport, incorporated proteins/ion transporters, and hence in construction of biosensor devices. Although BLMs provide an accepted mimic of cellular membranes, they are inherently fragile. Techniques are developed to stabilize them, such as hydrogel supports. In this paper, we present a reusable device for studies on hydrogel supported (hs) BLMs. These are formed across an ethylene tetrafluoroethylene (ETFE) aperture array supported by the hydrogel, which is during in situ polymerization covalently "sandwiched" between the ETFE substrate and a gold electrode microchip, thus allowing direct electrochemical studies with the integrated working electrodes. Using electrochemical impedance spectroscopy (EIS), X-ray photoelectron spectroscopy and contact angle measurements, we demonstrate the optimized chemical modifications of the gold electrode microchips and plasma modification of the ETFE aperture arrays facilitating covalent "sandwiching" of the hydrogel. Both fluorescence microscopy and EIS were used to demonstrate the induced spontaneous thinning of a deposited lipid solution, leading to formation of stabilized hsBLMs on average in 10 min. The determined specific membrane capacitance and resistance were shown to vary in the range 0.31-0.49 μF/cm(2) and 45-65 kΩ cm(2), respectively, corresponding to partially solvent containing BLMs with an average life time of 60-80 min. The characterized hsBLM formation and devised equivalent circuit models lead to a schematic model to illustrate lipid molecule distribution in hydrogel-supported apertures. The functionality of stabilized hsBLMs and detection sensitivity of the platform were verified by monitoring the effect of the ion transporter valinomycin.

  15. Conformations of double-headed, triple-tailed phospholipid oxidation lipid products in model membranes

    DEFF Research Database (Denmark)

    Hermetter, Albin; Kopec, Wojciech; Khandelia, Himanshu

    2013-01-01

    lipid in a Schiff base reaction to form a conjugate lipid (SCH) with two head groups, and three acyl tails. We investigate the conformations and properties of this unique class of adduct lipids using molecular dynamics simulations, and show that their insertion into lipid bilayers of POPC increases...... between the two head groups of the SCH. Schiff base formation of lipids can alter the concentration, homeostasis and localizations of phosphatidylserine and phosphatidylethanol lipids in membranes, and can therefore influence several membrane-associated processes including fusion and budding. The current...

  16. Endoplasmic reticulum-plasma membrane junctions: structure, function and dynamics.

    Science.gov (United States)

    Okeke, Emmanuel; Dingsdale, Hayley; Parker, Tony; Voronina, Svetlana; Tepikin, Alexei V

    2016-06-01

    Endoplasmic reticulum (ER)-plasma membrane (PM) junctions are contact sites between the ER and the PM; the distance between the two organelles in the junctions is below 40 nm and the membranes are connected by protein tethers. A number of molecular tools and technical approaches have been recently developed to visualise, modify and characterise properties of ER-PM junctions. The junctions serve as the platforms for lipid exchange between the organelles and for cell signalling, notably Ca(2+) and cAMP signalling. Vice versa, signalling events regulate the development and properties of the junctions. Two Ca(2+) -dependent mechanisms of de novo formation of ER-PM junctions have been recently described and characterised. The junction-forming proteins and lipids are currently the focus of vigorous investigation. Junctions can be relatively short-lived and simple structures, forming and dissolving on the time scale of a few minutes. However, complex, sophisticated and multifunctional ER-PM junctions, capable of attracting numerous protein residents and other cellular organelles, have been described in some cell types. The road from simplicity to complexity, i.e. the transformation from simple 'nascent' ER-PM junctions to advanced stable multiorganellar complexes, is likely to become an attractive research avenue for current and future junctologists. Another area of considerable research interest is the downstream cellular processes that can be activated by specific local signalling events in the ER-PM junctions. Studies of the cell physiology and indeed pathophysiology of ER-PM junctions have already produced some surprising discoveries, likely to expand with advances in our understanding of these remarkable organellar contact sites. © 2016 The Authors. The Journal of Physiology © 2016 The Physiological Society.

  17. Remodeling of Membrane Lipids in Iron-starved Chlamydomonas*

    Science.gov (United States)

    Urzica, Eugen I.; Vieler, Astrid; Hong-Hermesdorf, Anne; Page, M. Dudley; Casero, David; Gallaher, Sean D.; Kropat, Janette; Pellegrini, Matteo; Benning, Christoph; Merchant, Sabeeha S.

    2013-01-01

    Chlamydomonas reinhardtii cells exposed to abiotic stresses (e.g. nitrogen, zinc, or phosphorus deficiency) accumulate triacylglycerols (TAG), which are stored in lipid droplets. Here, we report that iron starvation leads to formation of lipid droplets and accumulation of TAGs. This occurs between 12 and 24 h after the switch to iron-starvation medium. C. reinhardtii cells deprived of iron have more saturated fatty acid (FA), possibly due to the loss of function of FA desaturases, which are iron-requiring enzymes with diiron centers. The abundance of a plastid acyl-ACP desaturase (FAB2) is decreased to the same degree as ferredoxin. Ferredoxin is a substrate of the desaturases and has been previously shown to be a major target of the iron deficiency response. The increase in saturated FA (C16:0 and C18:0) is concomitant with the decrease in unsaturated FA (C16:4, C18:3, or C18:4). This change was gradual for diacylglyceryl-N,N,N-trimethylhomoserine (DGTS) and digalactosyldiacylglycerol (DGDG), whereas the monogalactosyldiacylglycerol (MGDG) FA profile remained stable during the first 12 h, whereas MGDG levels were decreasing over the same period of time. These changes were detectable after only 2 h of iron starvation. On the other hand, DGTS and DGDG contents gradually decreased until a minimum was reached after 24–48 h. RNA-Seq analysis of iron-starved C. reinhardtii cells revealed notable changes in many transcripts coding for enzymes involved in FA metabolism. The mRNA abundances of genes coding for components involved in TAG accumulation (diacylglycerol acyltransferases or major lipid droplet protein) were increased. A more dramatic increase at the transcript level has been observed for many lipases, suggesting that major remodeling of lipid membranes occurs during iron starvation in C. reinhardtii. PMID:23983122

  18. Remodeling of membrane lipids in iron-starved Chlamydomonas.

    Science.gov (United States)

    Urzica, Eugen I; Vieler, Astrid; Hong-Hermesdorf, Anne; Page, M Dudley; Casero, David; Gallaher, Sean D; Kropat, Janette; Pellegrini, Matteo; Benning, Christoph; Merchant, Sabeeha S

    2013-10-18

    Chlamydomonas reinhardtii cells exposed to abiotic stresses (e.g. nitrogen, zinc, or phosphorus deficiency) accumulate triacylglycerols (TAG), which are stored in lipid droplets. Here, we report that iron starvation leads to formation of lipid droplets and accumulation of TAGs. This occurs between 12 and 24 h after the switch to iron-starvation medium. C. reinhardtii cells deprived of iron have more saturated fatty acid (FA), possibly due to the loss of function of FA desaturases, which are iron-requiring enzymes with diiron centers. The abundance of a plastid acyl-ACP desaturase (FAB2) is decreased to the same degree as ferredoxin. Ferredoxin is a substrate of the desaturases and has been previously shown to be a major target of the iron deficiency response. The increase in saturated FA (C16:0 and C18:0) is concomitant with the decrease in unsaturated FA (C16:4, C18:3, or C18:4). This change was gradual for diacylglyceryl-N,N,N-trimethylhomoserine (DGTS) and digalactosyldiacylglycerol (DGDG), whereas the monogalactosyldiacylglycerol (MGDG) FA profile remained stable during the first 12 h, whereas MGDG levels were decreasing over the same period of time. These changes were detectable after only 2 h of iron starvation. On the other hand, DGTS and DGDG contents gradually decreased until a minimum was reached after 24-48 h. RNA-Seq analysis of iron-starved C. reinhardtii cells revealed notable changes in many transcripts coding for enzymes involved in FA metabolism. The mRNA abundances of genes coding for components involved in TAG accumulation (diacylglycerol acyltransferases or major lipid droplet protein) were increased. A more dramatic increase at the transcript level has been observed for many lipases, suggesting that major remodeling of lipid membranes occurs during iron starvation in C. reinhardtii.

  19. Yeast lipids can phase separate into micrometer-scale membrane domains

    DEFF Research Database (Denmark)

    Klose, Christian; Ejsing, Christer S; Garcia-Saez, Ana J;

    2010-01-01

    The lipid raft concept proposes that biological membranes have the potential to form functional domains based on a selective interaction between sphingolipids and sterols. These domains seem to be involved in signal transduction and vesicular sorting of proteins and lipids. Although...... there is biochemical evidence for lipid raft-dependent protein and lipid sorting in the yeast Saccharomyces cerevisiae, direct evidence for an interaction between yeast sphingolipids and the yeast sterol ergosterol, resulting in membrane domain formation, is lacking. Here we show that model membranes formed from yeast...... total lipid extracts possess an inherent self-organization potential resulting in Ld-Lo phase coexistence at physiologically relevant temperature. Analyses of lipid extracts from mutants defective in sphingolipid metabolism as well as reconstitution of purified yeast lipids in model membranes of defined...

  20. Macroscopic domain formation in the platelet plasma membrane

    DEFF Research Database (Denmark)

    Bali, Rachna; Savino, Laura; Ramirez, Diego A.;

    2009-01-01

    There has been ample debate on whether cell membranes can present macroscopic lipid domains as predicted by three-component phase diagrams obtained by fluorescence microscopy. Several groups have argued that membrane proteins and interactions with the cytoskeleton inhibit the formation of large d...

  1. Membrane lipid unsaturation as physiological adaptation to animal longevity.

    Science.gov (United States)

    Naudí, Alba; Jové, Mariona; Ayala, Victòria; Portero-Otín, Manuel; Barja, Gustavo; Pamplona, Reinald

    2013-12-17

    The appearance of oxygen in the terrestrial atmosphere represented an important selective pressure for ancestral living organisms and contributed toward setting up the pace of evolutionary changes in structural and functional systems. The evolution of using oxygen for efficient energy production served as a driving force for the evolution of complex organisms. The redox reactions associated with its use were, however, responsible for the production of reactive species (derived from oxygen and lipids) with damaging effects due to oxidative chemical modifications of essential cellular components. Consequently, aerobic life required the emergence and selection of antioxidant defense systems. As a result, a high diversity in molecular and structural antioxidant defenses evolved. In the following paragraphs, we analyze the adaptation of biological membranes as a dynamic structural defense against reactive species evolved by animals. In particular, our goal is to describe the physiological mechanisms underlying the structural adaptation of cellular membranes to oxidative stress and to explain the meaning of this adaptive mechanism, and to review the state of the art about the link between membrane composition and longevity of animal species.

  2. Pressure effects on the equilibrium configurations of bilayer lipid membranes

    Science.gov (United States)

    DeVita, Raffaella; Stewart, Iain W.; Leo, Donald J.

    2007-10-01

    Planar bilayer lipid membranes (BLMs) are currently employed to construct many bio-inspired material systems and structures. In order to characterize the pressure effects on the equilibrium configurations of these biological membranes, a novel continuum model is proposed. The BLM is assumed to be a two-layer smectic A liquid crystal. The mean orientation of the amphiphilic molecules comprising the membrane is postulated to be perpendicular to the layers and each layer is idealized as a two-dimensional liquid. Moreover, the BLM is modeled as a simply supported plate undergoing small deformations. It is subjected to a pressure load that acts perpendicularly to the layers. The equilibrium equations and boundary conditions are derived from the bulk elastic energy for smectic A liquid crystals as described by de Gennes and Prost (1993 The Physics of Liquid Crystals 2nd edn (Oxford Science Publications)) by using variational methods. The resulting fourth-order linear partial differential equation is solved by employing cylindrical functions and the series solution is proved to be convergent. The solution is numerically computed for values of the model parameters that are reported in the literature. This paper is dedicated to the memory of our colleagues, Professors Kevin P Granata and Liviu Librescv, who lost their lives during the sensless tragedy on 16 April, 2007 at Virginia Tech.

  3. Lipid hydroperoxides in human plasma after ethanol consumption.

    Science.gov (United States)

    Asano, Migiwa; Nushida, Hideyuki; Adachi, Junko; Nagasaki, Yasushi; Nakagawa, Kanako; Kuse, Azumi; Ueno, Yasuhiro

    2009-04-01

    Oxidative stress contributes to the pathogenesis of alcoholic liver disease. The purpose of this study is to estimate the amount of oxidative stress that is present when healthy humans consume moderate amounts of ethanol. Blood was collected from healthy volunteers before, 1 h, and 3 h after drinking 400 ml of Japanese rice wine at the rate of 100 ml per 5 min. The aldehyde dehydrogenase 2 genotype and the concentrations of blood ethanol, total lipid hydroperoxides (LOOH), and cholesterol hydroperoxides were determined. The plasma LOOH was found to have significantly increased 1h after drinking. Cholesterol hydroperoxides were not detected in plasma, either before or after drinking. There was no relationship between the LOOH and the ethanol concentration. We showed that one-shot of moderate ethanol consumption temporarily increases the plasma LOOH in healthy volunteers but excessive plasma LOOH compounds were eliminated within a short time.

  4. Bipolar tetraether lipids: chain flexibility and membrane polarity gradients from spin-label electron spin resonance.

    Science.gov (United States)

    Bartucci, R; Gambacorta, A; Gliozzi, A; Marsh, D; Sportelli, L

    2005-11-15

    Membranes of thermophilic Archaea are composed of unique tetraether lipids in which C40, saturated, methyl-branched biphytanyl chains are linked at both ends to polar groups. In this paper, membranes composed of bipolar lipids P2 extracted from the acidothermophile archaeon Sulfolobus solfataricus are studied. The biophysical basis for the membrane formation and thermal stability is investigated by using electron spin resonance (ESR) of spin-labeled lipids. Spectral anisotropy and isotropic hyperfine couplings are used to determine the chain flexibility and polarity gradients, respectively. For comparison, similar measurements have been carried out on aqueous dispersions of diacyl reference lipid dipalmitoyl phosphatidylcholine and also of diphytanoyl phosphatidylcholine, which has methyl-branched chains. At a given temperature, the bolaform lipid chains are more ordered and less flexible than in normal bilayer membranes. Only at elevated temperatures (80 degrees C) does the flexibility of the chain environment in tetraether lipid assemblies approach that of fluid bilayer membranes. The height of the hydrophobic barrier formed by a monolayer of archaebacterial lipids is similar to that in conventional fluid bilayer membranes, and the permeability barrier width is comparable to that formed by a bilayer of C16 lipid chains. At a mole ratio of 1:2, the tetraether P2 lipids mix well with dipalmitoyl phosphatidylcholine lipids and stabilize conventional bilayer membranes. The biological as well as the biotechnological relevance of the results is discussed.

  5. Lipid recognition propensities of amino acids in membrane proteins from atomic resolution data

    Directory of Open Access Journals (Sweden)

    Morita Mizuki

    2011-12-01

    Full Text Available Abstract Background Protein-lipid interactions play essential roles in the conformational stability and biological functions of membrane proteins. However, few of the previous computational studies have taken into account the atomic details of protein-lipid interactions explicitly. Results To gain an insight into the molecular mechanisms of the recognition of lipid molecules by membrane proteins, we investigated amino acid propensities in membrane proteins for interacting with the head and tail groups of lipid molecules. We observed a common pattern of lipid tail-amino acid interactions in two different data sources, crystal structures and molecular dynamics simulations. These interactions are largely explained by general lipophilicity, whereas the preferences for lipid head groups vary among individual proteins. We also found that membrane and water-soluble proteins utilize essentially an identical set of amino acids for interacting with lipid head and tail groups. Conclusions We showed that the lipophilicity of amino acid residues determines the amino acid preferences for lipid tail groups in both membrane and water-soluble proteins, suggesting that tightly-bound lipid molecules and lipids in the annular shell interact with membrane proteins in a similar manner. In contrast, interactions between lipid head groups and amino acids showed a more variable pattern, apparently constrained by each protein's specific molecular function.

  6. Hydrostatic pressure decreases membrane fluidity and lipid desaturase expression in chondrocyte progenitor cells.

    Science.gov (United States)

    Montagne, Kevin; Uchiyama, Hiroki; Furukawa, Katsuko S; Ushida, Takashi

    2014-01-22

    Membrane biomechanical properties are critical in modulating nutrient and metabolite exchange as well as signal transduction. Biological membranes are predominantly composed of lipids, cholesterol and proteins, and their fluidity is tightly regulated by cholesterol and lipid desaturases. To determine whether such membrane fluidity regulation occurred in mammalian cells under pressure, we investigated the effects of pressure on membrane lipid order of mouse chondrogenic ATDC5 cells and desaturase gene expression. Hydrostatic pressure linearly increased membrane lipid packing and simultaneously repressed lipid desaturase gene expression. We also showed that cholesterol mimicked and cholesterol depletion reversed those effects, suggesting that desaturase gene expression was controlled by the membrane physical state itself. This study demonstrates a new effect of hydrostatic pressure on mammalian cells and may help to identify the molecular mechanisms involved in hydrostatic pressure sensing in chondrocytes.

  7. Towards a membrane proteome in Drosophila: a method for the isolation of plasma membrane

    Directory of Open Access Journals (Sweden)

    Thomas Graham H

    2010-05-01

    Full Text Available Abstract Background The plasma membrane (PM is a compartment of significant interest because cell surface proteins influence the way in which a cell interacts with its neighbours and its extracellular environment. However, PM is hard to isolate because of its low abundance. Aqueous two-phase affinity purification (2PAP, based on PEG/Dextran two-phase fractionation and lectin affinity for PM-derived microsomes, is an emerging method for the isolation of high purity plasma membranes from several vertebrate sources. In contrast, PM isolation techniques in important invertebrate genetic model systems, such as Drosophila melanogaster, have relied upon enrichment by density gradient centrifugation. To facilitate genetic investigation of activities contributing to the content of the PM sub-proteome, we sought to adapt 2PAP to this invertebrate model to provide a robust PM isolation technique for Drosophila. Results We show that 2PAP alone does not completely remove contaminating endoplasmic reticulum and mitochondrial membrane. However, a novel combination of density gradient centrifugation plus 2PAP results in a robust PM preparation. To demonstrate the utility of this technique we isolated PM from fly heads and successfully identified 432 proteins using MudPIT, of which 37% are integral membrane proteins from all compartments. Of the 432 proteins, 22% have been previously assigned to the PM compartment, and a further 34% are currently unassigned to any compartment and represent candidates for assignment to the PM. The remainder have previous assignments to other compartments. Conclusion A combination of density gradient centrifugation and 2PAP results in a robust, high purity PM preparation from Drosophila, something neither technique can achieve on its own. This novel preparation should lay the groundwork for the proteomic investigation of the PM in different genetic backgrounds in Drosophila. Our results also identify two key steps in this

  8. Characterization of freeze-fractured epithelial plasma membranes on nanometer scale with ToF-SIMS.

    Science.gov (United States)

    Draude, Felix; Körsgen, Martin; Pelster, Andreas; Schwerdtle, Tanja; Müthing, Johannes; Arlinghaus, Heinrich F

    2015-03-01

    Time-of-flight secondary ion mass spectrometry (ToF-SIMS) was used to characterize the freeze-fracturing process of human epithelial PANC-1 and UROtsa cells. For this purpose, phosphatidylcholine, sphingomyelin, phosphatidylethanolamine, and phosphatidylserine standard samples were investigated to find specific signals with both high specificity and signal intensity. The results were used to investigate single cells of subconfluent cell layers prepared with a special silicon wafer sandwich preparation technique. This freeze-fracturing technique strips cell membranes off the cells, isolating them on opposing silicon wafer substrates. Criteria were found for defining regions with stripped off cell membranes and, on the opposing wafer, complementary regions with the remaining cells. Measured ethanolamine/choline and serine/choline ratios in these regions clearly showed that in the freeze-fracturing process, the lipid bilayer of the plasma membrane is split along its central zone. Accordingly, only the outer lipid monolayer is stripped off the cell, while the inner lipid monolayer remains attached to the cell on the opposing wafer, thus allowing detailed analysis of a single lipid monolayer. Furthermore, it could be shown that using different washing procedures did not influence the transmembrane lipid distribution. Under optimized preparation conditions, it became feasible to detect lipids with a lateral resolution of approximately 100 nm. The data indicate that ToF-SIMS would be a very useful technique to study with very high lateral resolution changes in lipid composition caused, for example, by lipid storage diseases or pharmaceuticals that interfere with the lipid metabolism.

  9. Activation of Raf as a result of recruitment to the plasma membrane.

    Science.gov (United States)

    Stokoe, D; Macdonald, S G; Cadwallader, K; Symons, M; Hancock, J F

    1994-06-01

    The small guanine nucleotide binding protein Ras participates in a growth promoting signal transduction pathway. The mechanism by which interaction of Ras with the protein kinase Raf leads to activation of Raf was studied. Raf was targeted to the plasma membrane by addition of the COOH-terminal localization signals of K-ras. This modified form of Raf (RafCAAX) was activated to the same extent as Raf coexpressed with oncogenic mutant Ras. Plasma membrane localization rather than farnesylation or the presence of the additional COOH-terminal sequence accounted for the activation of RafCAAX. The activation of RafCAAX was completely independent of Ras; it was neither potentiated by oncogenic mutant Ras nor abrogated by dominant negative Ras. Raf, once recruited to the plasma membrane, was not anchored there by Ras; most activated Raf in cells was associated with plasma membrane cytoskeletal elements, not the lipid bilayer. Thus, Ras functions in the activation of Raf by recruiting Raf to the plasma membrane where a separate, Ras-independent, activation of Raf occurs.

  10. The essence of being extremophilic: the role of the unique archaeal membrane lipids : the role of the unique archaeal membrane lipids

    NARCIS (Netherlands)

    van de Vossenberg, J.L C M; Driessen, A.J.M.; Konings, W.N

    1998-01-01

    In extreme environments, mainly Archaea are encountered. The archaeal cytoplasmic membrane contains unique ether lipids that cannot easily be degraded, are temperature- and mechanically resistant, and highly salt tolerant. Moreover, thermophilic and extreme acidophilic Archaea possess

  11. Lipid composition affects the rate of photosensitized dissipation of cross-membrane diffusion potential on liposomes

    Science.gov (United States)

    Ytzhak, Shany; Wuskell, Joseph P.; Loew, Leslie M.; Ehrenberg, Benjamin

    2010-01-01

    Hydrophobic or amphiphilic tetrapyrrole sensitizers are taken up by cells and are usually located in cellular lipid membranes. Singlet oxygen is photogenerated by the sensitizer and it diffuses in the membrane and causes oxidative damage to membrane components. This damage can occur to membrane lipids and to membrane-localized proteins. Depolarization of the Nernst electric potential on cells’ membranes has been observed in cellular photosensitization, but it was not established whether lipid oxidation is a relevant factor leading to abolishing the resting potential of cells’ membranes and to their death. In this work we studied the effect of liposomes’ lipid composition on the kinetics of hematoporphyrin-photosensitized dissipation of K+-diffusion electric potential that was generated across the membranes. We employed an electrochromic voltage-sensitive spectroscopic probe that possesses a high fluorescence signal response to the potential. We found a correlation between the structure and unsaturation of lipids and the leakage of the membrane, following photosensitization. As the extent of non-conjugated unsaturation of the lipids is increased from 1 to 6 double bonds, the kinetics of depolarization become faster. We also found that the kinetics of depolarization is affected by the percentage of the unsaturated lipids in the liposome: as the fraction of the unsaturated lipids increases the leakage trough the membrane is enhanced. When liposomes are composed of a lipid mixture similar to that of natural membranes and photosensitization is being carried out under usual photodynamic therapy (PDT) conditions, photodamage to the lipids is not likely to cause enhanced permeability of ions through the membrane, which would have been a mechanism that leads to cell death. PMID:20536150

  12. Membrane fusion

    DEFF Research Database (Denmark)

    Bendix, Pól Martin

    2015-01-01

    At Stanford University, Boxer lab, I worked on membrane fusion of small unilamellar lipid vesicles to flat membranes tethered to glass surfaces. This geometry closely resembles biological systems in which liposomes fuse to plasma membranes. The fusion mechanism was studied using DNA zippering...... between complementary strands linked to the two apposing membranes closely mimicking the zippering mechanism of SNARE fusion complexes....

  13. Molecular dynamics simulations of lipid membranes with lateral force: rupture and dynamic properties.

    Science.gov (United States)

    Xie, Jun Yu; Ding, Guang Hong; Karttunen, Mikko

    2014-03-01

    Membranes' response to lateral tension, and eventual rupture, remains poorly understood. In this study, pure dipalmitoylphosphatidylcholine (DPPC) lipid bilayers, under tension/pressure, were studied using molecular dynamics (MD) simulations. The irreversible membrane breakdown is demonstrated to depend on the amplitude of lateral tension, loading rate, and the