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Sample records for plasma membrane structure

  1. Crystal structure of the plasma membrane proton pump

    DEFF Research Database (Denmark)

    Pedersen, Bjørn Panyella; Buch-Pedersen, Morten J; Morth, Jens Preben

    2007-01-01

    define the functional unit of ATP-coupled proton transport across the plasma membrane, and the structure is locked in a functional state not previously observed in P-type ATPases. The transmembrane domain reveals a large cavity, which is likely to be filled with water, located near the middle......A prerequisite for life is the ability to maintain electrochemical imbalances across biomembranes. In all eukaryotes the plasma membrane potential and secondary transport systems are energized by the activity of P-type ATPase membrane proteins: H+-ATPase (the proton pump) in plants and fungi 1, 2......, 3 , and Na+,K+-ATPase (the sodium–potassium pump) in animals 4 . The name P-type derives from the fact that these proteins exploit a phosphorylated reaction cycle intermediate of ATP hydrolysis 5 . The plasma membrane proton pumps belong to the type III P-type ATPase subfamily, whereas Na...

  2. Crystal structure of the plasma membrane proton pump

    DEFF Research Database (Denmark)

    Pedersen, Bjørn Panyella; Buch-Pedersen, Morten J; Morth, Jens Preben;

    2007-01-01

    A prerequisite for life is the ability to maintain electrochemical imbalances across biomembranes. In all eukaryotes the plasma membrane potential and secondary transport systems are energized by the activity of P-type ATPase membrane proteins: H+-ATPase (the proton pump) in plants and fungi 1, 2......, 3 , and Na+,K+-ATPase (the sodium–potassium pump) in animals 4 . The name P-type derives from the fact that these proteins exploit a phosphorylated reaction cycle intermediate of ATP hydrolysis 5 . The plasma membrane proton pumps belong to the type III P-type ATPase subfamily, whereas Na...... define the functional unit of ATP-coupled proton transport across the plasma membrane, and the structure is locked in a functional state not previously observed in P-type ATPases. The transmembrane domain reveals a large cavity, which is likely to be filled with water, located near the middle...

  3. Magnetic apatite for structural insights on the plasma membrane.

    Science.gov (United States)

    Stanca, Sarmiza E; Müller, Robert; Dellith, Jan; Nietzsche, Sandor; Stöckel, Stephan; Biskup, Christoph; Deckert, Volker; Krafft, Christoph; Popp, Jürgen; Fritzsche, Wolfgang

    2015-01-21

    The iron oxide-hydroxyapatite (FeOxHA) nanoparticles reported here differ from those reported before by their advantage of homogeneity and simple preparation; moreover, the presence of carboxymethyldextran (CMD), together with hydroxyapatite (HA), allows access to the cellular membrane, which makes our magnetic apatite unique. These nanoparticles combine magnetic behavior, Raman label ability and the property of interaction with the cellular membrane; they therefore represent an interesting material for structural differentiation of the cell membrane. It was observed by Raman spectroscopy, scanning electron microscopy (SEM) and fluorescence microscopy that FeOxHA adheres to the plasma membrane and does not penetrate the membrane. These insights make the nanoparticles a promising material for magnetic cell sorting, e.g. in microfluidic device applications.

  4. Magnetic apatite for structural insights on the plasma membrane

    Science.gov (United States)

    Stanca, Sarmiza E.; Müller, Robert; Dellith, Jan; Nietzsche, Sandor; Stöckel, Stephan; Biskup, Christoph; Deckert, Volker; Krafft, Christoph; Popp, Jürgen; Fritzsche, Wolfgang

    2015-01-01

    The iron oxide-hydroxyapatite (FeOxHA) nanoparticles reported here differ from those reported before by their advantage of homogeneity and simple preparation; moreover, the presence of carboxymethyldextran (CMD), together with hydroxyapatite (HA), allows access to the cellular membrane, which makes our magnetic apatite unique. These nanoparticles combine magnetic behavior, Raman label ability and the property of interaction with the cellular membrane; they therefore represent an interesting material for structural differentiation of the cell membrane. It was observed by Raman spectroscopy, scanning electron microscopy (SEM) and fluorescence microscopy that FeOxHA adheres to the plasma membrane and does not penetrate the membrane. These insights make the nanoparticles a promising material for magnetic cell sorting, e.g. in microfluidic device applications.

  5. 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.

  6. 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

  7. 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

  8. 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.

  9. 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.

  10. 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...

  11. Structure-function relationships of ErbB RTKs in the plasma membrane of living cells.

    Science.gov (United States)

    Arndt-Jovin, Donna J; Botelho, Michelle G; Jovin, Thomas M

    2014-04-01

    We review the states of the ErbB family of receptor tyrosine kinases (RTKs), primarily the EGF receptor (EGFR, ErbB1, HER1) and the orphan receptor ErbB2 as they exist in living mammalian cells, focusing on four main aspects: (1) aggregation state and distribution in the plasma membrane; (2) conformational features of the receptors situated in the plasma membrane, compared to the crystallographic structures of the isolated extracellular domains; (3) coupling of receptor disposition on filopodia with the transduction of signaling ligand gradients; and (4) ligand-independent receptor activation by application of a magnetic field.

  12. 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.

  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. 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...

  15. Changes in the plasma membrane in metabolic disease: impact of the membrane environment on G protein-coupled receptor structure and function.

    Science.gov (United States)

    Desai, Aditya J; Miller, Laurence J

    2017-07-10

    Drug development targeting GPCRs often utilizes model heterologous cell expression systems, reflecting an implicit assumption that the membrane environment has little functional impact on these receptors or on their responsiveness to drugs. However, much recent data have illustrated that membrane components can have an important functional impact on intrinsic membrane proteins. This review is directed toward gaining a better understanding of the structure of the plasma membrane in health and disease, and how this organelle can influence GPCR structure, function and regulation. It is important to recognize that the membrane provides a potential mode of lateral allosteric regulation of GPCRs and can affect the effectiveness of drugs and their biological responses in various disease states, which can even vary among individuals across the population. The type 1 cholecystokinin receptor is reviewed as an exemplar of a class A GPCR that is affected in this way by changes in the plasma membrane. © 2017 The British Pharmacological Society.

  16. 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...

  17. 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.

  18. A structural overview of the plasma membrane Na+,K+-ATPase and H+-ATPase ion pumps

    DEFF Research Database (Denmark)

    Morth, Jens Preben; Pedersen, Bjørn Panella; Buch-Pedersen, Morten Jeppe

    2011-01-01

    Plasma membrane ATPases are primary active transporters of cations that maintain steep concentration gradients. The ion gradients and membrane potentials derived from them form the basis for a range of essential cellular processes, in particular Na(+)-dependent and proton-dependent secondary tran......(+),K(+)-ATPase maintains a Na(+) and K(+) gradient in animal cells. Structural information provides insight into the function of these two distinct but related P-type pumps....

  19. 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...

  20. [Structural modifications of the surface of Escherichia coli bacteria and copper-induced permeability of plasma membrane].

    Science.gov (United States)

    Lebedev, V S; Volodina, L A; Deĭnega, E Iu; Fedorov, Iu I

    2005-01-01

    The effect of Cu2+ on the structural organization of the cell surface of Escherichia coli bacteria during the induction of conductivity of a plasma membrane was studied. A fluorescent study did not reveal any substantial changes in the microviscosity of lipids by the action of copper ions. At the same time, a substantial reorganization of membrane proteins during plasmolysis was observed. A model of the copper-induced structural reorganization of membrane lipids was constructed, according to which the reorganization leads to the opening in the membrane of channels of nonspecific conductivity for cations. The opening of conductivity channels results from the break of disulfide bonds in critical membrane proteins during the interaction with Cu+, which form either due to the reduction of Cu2+ on specific sites of cell surface or by means of external reducing agents.

  1. 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...

  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 plasma membrane monoamine transporter (PMAT): Structure, function, and role in organic cation disposition.

    Science.gov (United States)

    Wang, J

    2016-11-01

    Plasma membrane monoamine transporter (PMAT) is a new polyspecific organic cation transporter that transports a variety of biogenic amines and xenobiotic cations. Highly expressed in the brain, PMAT represents a major uptake2 transporter for monoamine neurotransmitters. At the blood-cerebrospinal fluid (CSF) barrier, PMAT is the principal organic cation transporter for removing neurotoxins and drugs from the CSF. Here I summarize our latest understanding of PMAT and its roles in monoamine uptake and xenobiotic disposition. © 2016 American Society for Clinical Pharmacology and Therapeutics.

  4. 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.

  5. Multifaceted plasma membrane Ca(2+) pumps: From structure to intracellular Ca(2+) handling and cancer.

    Science.gov (United States)

    Padányi, Rita; Pászty, Katalin; Hegedűs, Luca; Varga, Karolina; Papp, Béla; Penniston, John T; Enyedi, Ágnes

    2016-06-01

    Plasma membrane Ca(2+) ATPases (PMCAs) are intimately involved in the control of intracellular Ca(2+) concentration. They reduce Ca(2+) in the cytosol not only by direct ejection, but also by controlling the formation of inositol-1,4,5-trisphosphate and decreasing Ca(2+) release from the endoplasmic reticulum Ca(2+) pool. In mammals four genes (PMCA1-4) are expressed, and alternative RNA splicing generates more than twenty variants. The variants differ in their regulatory characteristics. They localize into highly specialized membrane compartments and respond to the incoming Ca(2+) with distinct temporal resolution. The expression pattern of variants depends on cell type; a change in this pattern can result in perturbed Ca(2+) homeostasis and thus altered cell function. Indeed, PMCAs undergo remarkable changes in their expression pattern during tumorigenesis that might significantly contribute to the unbalanced Ca(2+) homeostasis of cancer cells. This article is part of a Special Issue entitled: Calcium and Cell Fate. Guest Editors: Jacques Haiech, Claus Heizmann, Joachim Krebs, Thierry Capiod and Olivier Mignen.

  6. Structure and dynamics of nano-sized raft-like domains on the plasma membrane

    Science.gov (United States)

    Herrera, Fernando E.; Pantano, Sergio

    2012-01-01

    Cell membranes are constitutively composed of thousands of different lipidic species, whose specific organization leads to functional heterogeneities. In particular, sphingolipids, cholesterol and some proteins associate among them to form stable nanoscale domains involved in recognition, signaling, membrane trafficking, etc. Atomic-detail information in the nanometer/second scale is still elusive to experimental techniques. In this context, molecular simulations on membrane systems have provided useful insights contributing to bridge this gap. Here we present the results of a series of simulations of biomembranes representing non-raft and raft-like nano-sized domains in order to analyze the particular structural and dynamical properties of these domains. Our results indicate that the smallest (5 nm) raft domains are able to preserve their distinctive structural and dynamical features, such as an increased thickness, higher ordering, lower lateral diffusion, and specific lipid-ion interactions. The insertion of a transmembrane protein helix into non-raft, extended raft-like, and raft-like nanodomain environments result in markedly different protein orientations, highlighting the interplay between the lipid-lipid and lipid-protein interactions.

  7. The plasma membrane Ca2+ ATPase of animal cells: structure, function and regulation.

    Science.gov (United States)

    Di Leva, Francesca; Domi, Teuta; Fedrizzi, Laura; Lim, Dmitry; Carafoli, Ernesto

    2008-08-01

    Most important processes in cell life are regulated by calcium (Ca2+). A number of mechanisms have thus been developed to maintain the concentration of free Ca2+ inside cells at the level (100-200nM) necessary for the optimal operation of the targets of its regulatory function. The systems that move Ca2+ back and forth across membranes are important actors in its control. The plasma membrane calcium ATPase (PMCA pump) which ejects Ca2+ from all eukaryotic cell types will be the topic of this contribution. The pump uses a molecule of ATP to transport one molecule of Ca2+ from the cytosol to the external environment. It is a P-type ATPase encoded by four genes (ATP2B1-4), the transcripts of which undergo different types of alternative splicing. Many pump variants thus exist. Their multiplicity is best explained by the specific Ca2+ demands in different cell types. In keeping with these demands, the isoforms are differently expressed in tissues and cell types and have differential Ca2+ extruding properties. At very low Ca2+ concentrations the PMCAs are nearly inactive. They must be activated by calmodulin, by acid phospholipids, by protein kinases, and by other means, e.g., a dimerization process. Other proteins interact with the PMCAs (i.e., MAGUK and NHERF at the PDZ domain and calcineurin A in the main intracellular domain) to sort them to specific regions of the cell membrane or to regulate their function. In some cases the interaction is isoform, or even splice variant specific. PMCAs knock out (KO) mice have been generated and have contributed information on the importance of PMCAs to cells and organisms. So far, only one human genetic disease, hearing loss, has been traced back to a PMCA defect.

  8. Physical association between a novel plasma-membrane structure and centrosome orients cell division

    Science.gov (United States)

    Negishi, Takefumi; Miyazaki, Naoyuki; Murata, Kazuyoshi; Yasuo, Hitoyoshi; Ueno, Naoto

    2016-01-01

    In the last mitotic division of the epidermal lineage in the ascidian embryo, the cells divide stereotypically along the anterior-posterior axis. During interphase, we found that a unique membrane structure invaginates from the posterior to the centre of the cell, in a microtubule-dependent manner. The invagination projects toward centrioles on the apical side of the nucleus and associates with one of them. Further, a cilium forms on the posterior side of the cell and its basal body remains associated with the invagination. A laser ablation experiment suggests that the invagination is under tensile force and promotes the posterior positioning of the centrosome. Finally, we showed that the orientation of the invaginations is coupled with the polarized dynamics of centrosome movements and the orientation of cell division. Based on these findings, we propose a model whereby this novel membrane structure orchestrates centrosome positioning and thus the orientation of cell division axis. DOI: http://dx.doi.org/10.7554/eLife.16550.001 PMID:27502556

  9. Complex formation between primycin and ergosterol: entropy-driven initiation of modification of the fungal plasma membrane structure

    National Research Council Canada - National Science Library

    Virág, Eszter; Pesti, Miklós; Kunsági-Máté, Sándor

    2012-01-01

    The interaction of the antibiotic primycin with the main fungal sterol, ergosterol, was investigated in vitro in order to monitor the effect of primycin on the fungal plasma membrane at the molecular level...

  10. A structural overview of the plasma membrane Na+,K+-ATPase and H+-ATPase ion pumps

    DEFF Research Database (Denmark)

    Morth, Jens Preben; Pedersen, Bjørn Panella; Buch-Pedersen, Morten Jeppe

    2011-01-01

    Plasma membrane ATPases are primary active transporters of cations that maintain steep concentration gradients. The ion gradients and membrane potentials derived from them form the basis for a range of essential cellular processes, in particular Na(+)-dependent and proton-dependent secondary...... transport systems that are responsible for uptake and extrusion of metabolites and other ions. The ion gradients are also both directly and indirectly used to control pH homeostasis and to regulate cell volume. The plasma membrane H(+)-ATPase maintains a proton gradient in plants and fungi and the Na...

  11. Disrupted plasma membrane localization and loss of function reveal regions of human equilibrative nucleoside transporter 1 involved in structural integrity and activity.

    Science.gov (United States)

    Nivillac, Nicole M I; Wasal, Karanvir; Villani, Daniela F; Naydenova, Zlatina; Hanna, W J Brad; Coe, Imogen R

    2009-10-01

    Human Equilibrative Nucleoside Transporter 1 (hENT1) is an integral membrane protein that transports nucleosides and analog drugs across cellular membranes. Very little is known about intracellular processing and localization of hENT1. Here we show that disruption of a highly conserved triplet (PWN) near the N-terminus, or the last eight C-terminal residues (two hydrophobic triplets separated by a positive arginine) result in loss of plasma membrane localization and/or transport function. To understand the role of specific residues within these regions, we studied the localization patterns of N- or C-terminal deletion and/or substitution mutants of GFP-hENT1 using confocal microscopy. Quantification of GFP-hENT1 (mutant and wildtype) protein at the plasma membrane was conducted using nitrobenzylthioinosine (NBTI) binding. Functionality of the GFP-hENT1 mutants was determined by heterologous expression in Xenopus laevis oocytes followed by measurement of uridine uptake. Mutation of the proline within the PWN motif disrupts plasma membrane localization. C-terminal mutations (primarily within the hydrophobic triplets) lead to hENT1 retention within the cell (e.g. in the ER). Some mutants still localize to the plasma membrane but show reduced transport activity. These data suggest that these two regions contribute to the structural integrity and thus correct processing and function of hENT1.

  12. The Structure of the Synaptic Vesicle-Plasma Membrane Interface Constrains SNARE Models of Rapid, Synchronous Exocytosis at Nerve Terminals

    Science.gov (United States)

    Gundersen, Cameron B.

    2017-01-01

    Contemporary models of neurotransmitter release invoke direct or indirect interactions between the Ca2+ sensor, synaptotagmin and the incompletely zippered soluble, N-ethyl-maleimide-sensitive factor attachment protein receptor (SNARE) complex. However, recent electron microscopic (EM) investigations have raised pragmatic issues concerning the mechanism by which SNAREs trigger membrane fusion at nerve terminals. The first issue is related to the finding that the area of contact between a “fully primed” synaptic vesicle and the plasma membrane can exceed 600 nm2. Approximately four-thousands lipid molecules can inhabit this contact zone. Thus, renewed efforts will be needed to explain how the zippering of as few as two SNARE complexes mobilizes these lipids to achieve membrane fusion. The second issue emerges from the finding that “docking filaments” are sandwiched within the area of vesicle-plasma membrane contact. It is challenging to reconcile the location of these filaments with SNARE models of exocytosis. Instead, this commentary outlines how these data are more compatible with a model in which a cluster of synaptotagmins catalyzes exocytotic membrane fusion. PMID:28280457

  13. 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

  14. 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

  15. 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

  16. 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

  17. 2D-ELDOR study of heterogeneity and domain structure changes in plasma membrane vesicles upon cross-linking of receptors.

    Science.gov (United States)

    Chiang, Yun-Wei; Costa-Filho, Antonio J; Baird, Barbara; Freed, Jack H

    2011-09-08

    2D electron-electron double resonance (2D-ELDOR) with the "full Sc-" method of analysis is applied to the study of plasma membrane vesicles. Membrane structural changes upon antigen cross-linking of IgE receptors (IgE-FcεRI) in plasma membrane vesicles (PMVs) isolated from RBL-2H3 mast cells are investigated, for the first time, by means of these 2D-ELDOR techniques. Spectra of 1-palmitoyl-2-(16-doxyl stearoyl) phosphatidylcholine (16-PC) from PMVs before and after this stimulation at several temperatures are reported. The results demonstrate a coexistence of liquid-ordered (L(o)) and liquid-disordered (L(d)) components. We find that upon cross-linking, the membrane environment is remodeled to become more disordered, as shown by a moderate increase in the population of the L(d) component. This change in the relative amount of the L(o) versus L(d) components upon cross-linking is consistent with a model wherein the IgE receptors, which when clustered by antigen to cause cell stimulation, lead to more disordered lipids, and their dynamic and structural properties are slightly altered. This study demonstrates that 2D-ELDOR, analyzed by the full Sc- method, is a powerful approach for capturing the molecular dynamics in biological membranes. This is a particular case showing how 2D-ELDOR can be applied to study physical processes in complex systems that yield subtle changes.

  18. 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).

  19. 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.

  20. 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

  1. 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

  2. Complex formation between primycin and ergosterol: entropy-driven initiation of modification of the fungal plasma membrane structure.

    Science.gov (United States)

    Virág, Eszter; Pesti, Miklós; Kunsági-Máté, Sándor

    2012-04-01

    The interaction of the antibiotic primycin with the main fungal sterol, ergosterol, was investigated in vitro in order to monitor the effect of primycin on the fungal plasma membrane at the molecular level. The thermodynamic parameters of complex formation were determined by measuring Rayleigh scattering as a signal sensitive to particle size. The Benesi-Hildebrand method validated the 1 : 1 stoichiometry of the primycin-ergosterol complexes. A very low enthalpy change (ΔH=-1.14 kJ mol(-1)) was measured during the complex formation, which itself cannot be responsible for the molecular association. However, the entropy production (ΔS=29.78 J mol K(-1)) observed during the complex formation can describe the molecular interaction. This effect is probably due to the partial destruction of the solvation shell of the interacting species before the interlinking of the molecules. The results highlight the importance of ergosterol as concerns the mode of effect of primycin in the treatment of fungal infections. As the entropy has a determinant role in the ergosterol-primycin interaction, this interaction exhibits a very high temperature dependence, with the important consequence that the effect exerted by primycin on the cell membranes increases with rising temperature, and the effect is therefore pronounced in fevered bodies.

  3. 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.

  4. 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...

  5. 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...

  6. 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...

  7. 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.

  8. Composite plasma polymerized sulfonated polystyrene membrane for PEMFC

    Energy Technology Data Exchange (ETDEWEB)

    Nath, Bhabesh Kumar; Khan, Aziz; Chutia, Joyanti, E-mail: jchutiaiasst@gmail.com

    2015-10-15

    Highlights: • Methyl methane sulfonate (MMS) is used as the sulfonating agent. • The proton conductivity of the membrane is found to be 0.141 S cm{sup −1}. • Power density of fuel cell with styrene/MMS membrane is 0.5 W cm{sup −2}. • The membrane exhibits thermal stability up to 140 °C. - Abstract: This work presents the introduction of an organic compound methyl methane sulfonate (MMS) for the first time in fabrication of polystyrene based proton exchange membrane (PEM) by plasma polymerization process. The membrane is fabricated by co-polymerizing styrene and MMS in capacitively coupled continuous RF plasma. The chemical composition of the plasma polymerized polymer membrane is investigated using Fourier Transform Infrared Spectroscopy which reveals the formation of composite structure of styrene and MMS. The surface morphology studied using AFM and SEM depicts the effect of higher partial pressure of MMS on surface topography of the membrane. The proton transport property of the membrane studied using electrochemical impedance spectroscopy shows the achievement of maximum proton conductivity of 0.141 S cm{sup −1} which is comparable to Nafion 117 membrane. Fuel cell performance test of the synthesized membrane shows a maximum power density of 500 mW cm{sup −2} and current density of 0.62 A cm{sup −2} at 0.6 V.

  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. 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

  12. Surface modification of nanoporous alumina membranes by plasma polymerization.

    Science.gov (United States)

    Losic, Dusan; Cole, Martin A; Dollmann, Björn; Vasilev, Krasimir; Griesser, Hans J

    2008-06-18

    The deposition of plasma polymer coatings onto porous alumina (PA) membranes was investigated with the aim of adjusting the surface chemistry and the pore size of the membranes. PA membranes from commercial sources with a range of pore diameters (20, 100 and 200 nm) were used and modified by plasma polymerization using n-heptylamine (HA) monomer, which resulted in a chemically reactive polymer surface with amino groups. Heptylamine plasma polymer (HAPP) layers with a thickness less than the pore diameter do not span the pores but reduce their diameter. Accordingly, by adjusting the deposition time and thus the thickness of the plasma polymer coating, it is feasible to produce any desired pore diameter. The structural and chemical properties of modified membranes were studied by scanning electron microscopy (SEM), atomic force microscopy (AFM) and x-ray electron spectroscopy (XPS). The resultant PA membranes with specific surface chemistry and controlled pore size are applicable for molecular separation, cell culture, bioreactors, biosensing, drug delivery, and engineering complex composite membranes.

  13. 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.

  14. Epithelial cell-cell junctions and plasma membrane domains

    NARCIS (Netherlands)

    Giepmans, Ben N. G.; van Ijzendoorn, Sven C. D.

    Epithelial cells form a barrier against the environment, but are also required for the regulated exchange of molecules between an organism and its surroundings. Epithelial cells are characterised by a remarkable polarization of their plasma membrane, evidenced by the appearance of structurally,

  15. Epithelial cell-cell junctions and plasma membrane domains

    NARCIS (Netherlands)

    Giepmans, Ben N. G.; van Ijzendoorn, Sven C. D.

    2009-01-01

    Epithelial cells form a barrier against the environment, but are also required for the regulated exchange of molecules between an organism and its surroundings. Epithelial cells are characterised by a remarkable polarization of their plasma membrane, evidenced by the appearance of structurally, comp

  16. 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.

  17. 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

  18. 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.

  19. 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.

  20. 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.

  1. [Effect of point substitutions of Asp-714 and Asp-720 residues on the structure and function of the H+ -ATPase of the yeast plasma membrane].

    Science.gov (United States)

    Petrov, V V; Ibragimov, R I

    2014-01-01

    Membrane-spanning M5 and M6 segments, which play a role in the formation of cation transport sites in H(+)-, Ca2(+)-, K(+)-, Na(+)-, and other P2-ATPases, are connected by a short extracytoplasmic loop. In the yeast plasma membrane H(+)-ATPase, which belongs to a family of P2-ATPases, the loop is connected to M5 and M6 through the Asp-714 and Asp-720 residues. In this work, the effect of point amino, acidreplacements of Asp-714 and Asp-720 by Ala, Val, Asn, and Glu residues on the function of the enzyme was studied. The Asp714Asn point mutant possessed activities similar to those of the wild-type enzyme, whereas the replacement of Asp-714 by other amino acid residues disrupted biogenesis and led to a loss of activity. All mutants with substitution of Asp-720 were expressed and possessed relatively high activity. The D720V mutant displayed significantly reduced expression levels, activity, H+ transport, and ATP hydrolyzing activity. Thus, substitutions of Asp-714, except for the D714N mutant, led to significant defects in biogenesis and/or function of the enzyme. The results indicate the important role for the Asp-714 residue in biogenesis, structure stability, and enzyme function.

  2. Effects of freezing and cold acclimation on the plasma membrane of isolated protoplasts, progress report

    Energy Technology Data Exchange (ETDEWEB)

    Steponkus, P L

    1993-01-01

    Our goal is to provide a mechanistic understanding of the cellular and molecular aspects of freezing injury and cold acclimation from a perspective of the structural and functional integrity of the plasma membrane -- the primary site of freezing injury in winter cereals. We have utilized protoplasts isolated from leaves of winter rye (Secale cereale L. cv Puma) to study the cryobehavior of the plasma membrane during a freeze/thaw cycle. The focus of our current studies is on lesions in the plasma membrane that result from severe freeze-induced dehydration and result in the alteration of the semipermeable characteristics of the plasma membrane so that the protoplasts are osmotically unresponsive. In protoplasts isolated from non-acclimated rye leaves (NA protoplasts), injury is associated with the formation of aparticulate domains in the plasma membrane, aparticulate lamellae subtending the plasma membrane, and lamellar-to-hexagonal II phase transitions in the plasma membrane and the subtending lamellae. However, lamellar-to-hexagonal II phase transitions are not observed following severe dehydration of protoplasts isolated from cold-acclimated rye leaves (ACC protoplasts). Rather, injury is associated with the fracture-jump lesion,'' which, in freeze-fracture electron microscopy studies, is manifested as localized deviations in the fracture face of the plasma membrane. The fracture plane jumps'' from the plasma membrane to either subtending aparticulate lamellae or aparticulate regions of various endomembranes (predominantly chloroplast envelopes) that are in close apposition with the plasma membrane.

  3. Structure of a 14-3-3 coordinated hexamer of the plant plasma membrane H+ -ATPase by combining X-ray crystallography and electron cryomicroscopy.

    Science.gov (United States)

    Ottmann, Christian; Marco, Sergio; Jaspert, Nina; Marcon, Caroline; Schauer, Nicolas; Weyand, Michael; Vandermeeren, Caroline; Duby, Geoffrey; Boutry, Marc; Wittinghofer, Alfred; Rigaud, Jean-Louis; Oecking, Claudia

    2007-02-09

    Regulatory 14-3-3 proteins activate the plant plasma membrane H(+)-ATPase by binding to its C-terminal autoinhibitory domain. This interaction requires phosphorylation of a C-terminal, mode III, recognition motif as well as an adjacent span of approximately 50 amino acids. Here we report the X-ray crystal structure of 14-3-3 in complex with the entire binding motif, revealing a previously unidentified mode of interaction. A 14-3-3 dimer simultaneously binds two H(+)-ATPase peptides, each of which forms a loop within the typical 14-3-3 binding groove and therefore exits from the center of the dimer. Several H(+)-ATPase mutants support this structure determination. Accordingly, 14-3-3 binding could result in H(+)-ATPase oligomerization. Indeed, by using single-particle electron cryomicroscopy, the 3D reconstruction of the purified H(+)-ATPase/14-3-3 complex demonstrates a hexameric arrangement. Fitting of 14-3-3 and H(+)-ATPase atomic structures into the 3D reconstruction map suggests the spatial arrangement of the holocomplex.

  4. 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.

  5. 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...

  6. 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.

  7. 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

  8. 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.

  9. 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.

  10. Self-Deployable Membrane Structures

    Science.gov (United States)

    Sokolowski, Witold M.; Willis, Paul B.; Tan, Seng C.

    2010-01-01

    Currently existing approaches for deployment of large, ultra-lightweight gossamer structures in space rely typically upon electromechanical mechanisms and mechanically expandable or inflatable booms for deployment and to maintain them in a fully deployed, operational configuration. These support structures, with the associated deployment mechanisms, launch restraints, inflation systems, and controls, can comprise more than 90 percent of the total mass budget. In addition, they significantly increase the stowage volume, cost, and complexity. A CHEM (cold hibernated elastic memory) membrane structure without any deployable mechanism and support booms/structure is deployed by using shape memory and elastic recovery. The use of CHEM micro-foams reinforced with carbon nanotubes is considered for thin-membrane structure applications. In this advanced structural concept, the CHEM membrane structure is warmed up to allow packaging and stowing prior to launch, and then cooled to induce hibernation of the internal restoring forces. In space, the membrane remembers its original shape and size when warmed up. After the internal restoring forces deploy the structure, it is then cooled to achieve rigidization. For this type of structure, the solar radiation could be utilized as the heat energy used for deployment and space ambient temperature for rigidization. The overall simplicity of the CHEM self-deployable membrane is one of its greatest assets. In present approaches to space-deployable structures, the stow age and deployment are difficult and challenging, and introduce a significant risk, heavy mass, and high cost. Simple procedures provided by CHEM membrane greatly simplify the overall end-to-end process for designing, fabricating, deploying, and rigidizing large structures. The CHEM membrane avoids the complexities associated with other methods for deploying and rigidizing structures by eliminating deployable booms, deployment mechanisms, and inflation and control systems

  11. 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

  12. 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

  13. Robust mixed conducting membrane structure

    DEFF Research Database (Denmark)

    2010-01-01

    The present invention provides a membrane structure, comprising in said order a first electronically conducting layer, an ionically conducting layer, and a second electronically conducting layer, characterized in that the first and second electronically conducting layers are internally short circ...

  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. Robust mixed conducting membrane structure

    DEFF Research Database (Denmark)

    2010-01-01

    The present invention provides a membrane structure, comprising in said order a first electronically conducting layer, an ionically conducting layer, and a second electronically conducting layer, characterized in that the first and second electronically conducting layers are internally short...... circuited. The present invention further provides a method of producing the above membrane structure, comprising the steps of : providing a ionically conducting layer; applying at least one layer of electronically conducting material on each side of said ionically conducting layer; sintering the multilayer...

  16. 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.

  17. 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

  18. 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.

  19. Interaction between La(III) and proteins on the plasma membrane of horseradish

    Science.gov (United States)

    Yang, Guang-Mei; Chu, Yun-Xia; Lv, Xiao-Fen; Zhou, Qing; Huang, Xiao-Hua

    2012-06-01

    Lanthanum (La) is an important rare earth element in the ecological environment of plant. The proteins on the plasma membrane control the transport of molecules into and out of cell. It is very important to investigate the effect of La(III) on the proteins on the plasma membrane in the plant cell. In the present work, the interaction between La(III) and proteins on the plasma membrane of horseradish was investigated using optimization of the fluorescence microscopy and fluorescence spectroscopy. It is found that the fluorescence of the complex system of protoplasts and 1-aniline Kenai-8-sulfonic acid in horseradish treated with the low concentration of La(III) is increased compared with that of the control horseradish. The opposite effect is observed in horseradish treated with the high concentration of La(III). These results indicated that the low concentration of La(III) can interact with the proteins on the plasma membrane of horseradish, causing the improvement in the structure of proteins on the plasma membrane. The high concentration of La(III) can also interact with the proteins on the plasma membrane of horseradish, leading to the destruction of the structure of proteins on the plasma membrane. We demonstrate that the proteins on the plasma membrane are the targets of La(III) action on plant cell.

  20. RNAi-mediated downregulation of poplar plasma membrane intrinsic proteins (PIPs) changes plasma membrane proteome composition and affects leaf physiology.

    Science.gov (United States)

    Bi, Zhen; Merl-Pham, Juliane; Uehlein, Norbert; Zimmer, Ina; Mühlhans, Stefanie; Aichler, Michaela; Walch, Axel Karl; Kaldenhoff, Ralf; Palme, Klaus; Schnitzler, Jörg-Peter; Block, Katja

    2015-10-14

    Plasma membrane intrinsic proteins (PIPs) are one subfamily of aquaporins that mediate the transmembrane transport of water. To reveal their function in poplar, we generated transgenic poplar plants in which the translation of PIP genes was downregulated by RNA interference investigated these plants with a comprehensive leaf plasma membrane proteome and physiome analysis. First, inhibition of PIP synthesis strongly altered the leaf plasma membrane protein composition. Strikingly, several signaling components and transporters involved in the regulation of stomatal movement were differentially regulated in transgenic poplars. Furthermore, hormonal crosstalk related to abscisic acid, auxin and brassinosteroids was altered, in addition to cell wall biosynthesis/cutinization, the organization of cellular structures and membrane trafficking. A physiological analysis confirmed the proteomic results. The leaves had wider opened stomata and higher net CO2 assimilation and transpiration rates as well as greater mesophyll conductance for CO2 (gm) and leaf hydraulic conductance (Kleaf). Based on these results, we conclude that PIP proteins not only play essential roles in whole leaf water and CO2 flux but have important roles in the regulation of stomatal movement.

  1. Plasma-based accelerator structures

    Energy Technology Data Exchange (ETDEWEB)

    Schroeder, Carl B. [Univ. of California, Berkeley, CA (United States)

    1999-12-01

    Plasma-based accelerators have the ability to sustain extremely large accelerating gradients, with possible high-energy physics applications. This dissertation further develops the theory of plasma-based accelerators by addressing three topics: the performance of a hollow plasma channel as an accelerating structure, the generation of ultrashort electron bunches, and the propagation of laser pulses is underdense plasmas.

  2. Structure Prediction of Membrane Proteins

    Institute of Scientific and Technical Information of China (English)

    Chunlong Zhou; Yao Zheng; Yan Zhou

    2004-01-01

    There is a large gap between the number of membrane protein (MP) sequences and that of their decoded 3D structures, especially high-resolution structures, due to difficulties in crystal preparation of MPs. However, detailed knowledge of the 3D structure is required for the fundamental understanding of the function of an MP and the interactions between the protein and its inhibitors or activators. In this paper, some computational approaches that have been used to predict MP structures are discussed and compared.

  3. Correlation Between the Mobility of Inner Plasma Membrane Structure and Agglutination by Concanavalin A in Two Cell Lines of MOPC 173 Plasmocytoma Cells

    Science.gov (United States)

    Guérin, Claudine; Zachowski, Alain; Prigent, Bernadette; Paraf, Alain; Dunia, Irène; Diawara, Marie-Aline; Benedetti, E. L.

    1974-01-01

    Both the distribution of the concanavalin A-binding sites and the rearrangement of the intramembranous particles revealed by the freeze-etching technique, have been studied by means of two variants of the same cell line issued from MOPC 173 murine plasmocytoma. One variant does not agglutinate even in presence of high lectin concentration. It has been shown that the number of binding sites and affinity are almost the same in the two variants. The clustered distribution of intramembranous particles is induced by the interaction of the concanavalin A and the cell surface only in the variant which is agglutinable. From these results it became apparent that the clustered distribution of the membrane particulate components is an acquired feature of the plasma membrane accompanying cell agglutination. Images PMID:4521044

  4. 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.

  5. 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.

  6. The Structural Basis of Cholesterol Activity in Membranes

    Energy Technology Data Exchange (ETDEWEB)

    Olsen, Brett N.; Bielska, Agata; Lee, Tiffany; Daily, Michael D.; Covey, Douglas F.; Schlesinger, Paul H.; Baker, Nathan A.; Ory, Daniel S.

    2013-10-15

    Although the majority of free cellular cholesterol is present in the plasma membrane, cholesterol homeostasis is principally regulated through sterol-sensing proteins that reside in the cholesterol-poor endoplasmic reticulum (ER). In response to acute cholesterol loading or depletion, there is rapid equilibration between the ER and plasma membrane cholesterol pools, suggesting a biophysical model in which the availability of plasma membrane cholesterol for trafficking to internal membranes modulates ER membrane behavior. Previous studies have predominantly examined cholesterol availability in terms of binding to extramembrane acceptors, but have provided limited insight into the structural changes underlying cholesterol activation. In this study, we use both molecular dynamics simulations and experimental membrane systems to examine the behavior of cholesterol in membrane bilayers. We find that cholesterol depth within the bilayer provides a reasonable structural metric for cholesterol availability and that this is correlated with cholesterol-acceptor binding. Further, the distribution of cholesterol availability in our simulations is continuous rather than divided into distinct available and unavailable pools. This data provide support for a revised cholesterol activation model in which activation is driven not by saturation of membrane-cholesterol interactions but rather by bulk membrane remodeling that reduces membrane-cholesterol affinity.

  7. [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.

  8. The role of abscisic acid and low temperature in chickpea (Cicer arietinum) cold tolerance. II. Effects on plasma membrane structure and function.

    Science.gov (United States)

    Bakht, Jehan; Bano, Asghari; Dominy, Peter

    2006-01-01

    The frost hardiness of many plants such as chickpea can be increased by exposure to low non-freezing temperatures and/or the application of abscisic acid (ABA), a process known as frost acclimation. Experiments were conducted to study the response over a 14 d period of enriched plasma membrane fractions isolated from chickpea plants exposed to low temperature and sprayed with exogenous ABA. Measurement of the temperatures inducing 50% foliar cell death (LT50), and subsequent statistical analysis suggest that, like many plants, exposure to low temperatures (5/-2 degrees C; day/night) induces a significant level (P chickpea when compared with control plants (20/7 degrees C; day/night). Spraying plants with exogenous ABA also increased frost tolerance (P chickpea plants to low temperatures increased the DBI by 15% at day 4 and 19% at day 14 when compared with untreated control plants. Application of ABA alone did not increase the DBI by more than 6% at any time; the effects of both treatments applied together was more than additive, inducing a DBI increase of 27% at day 14 when compared with controls. There was a good correlation (P properties of the plasma membrane other than fluidity are involved in frost acclimation in chickpea.

  9. 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.

  10. Corrugated Membrane Fuel Cell Structures

    Energy Technology Data Exchange (ETDEWEB)

    Grot, Stephen [President, Ion Power Inc.

    2013-09-30

    One of the most challenging aspects of traditional PEM fuel cell stacks is the difficulty achieving the platinum catalyst utilization target of 0.2 gPt/kWe set forth by the DOE. Good catalyst utilization can be achieved with state-of-the-art catalyst coated membranes (CCM) when low catalyst loadings (<0.3 mg/cm2) are used at a low current. However, when low platinum loadings are used, the peak power density is lower than conventional loadings, requiring a larger total active area and a larger bipolar plate. This results in a lower overall stack power density not meeting the DOE target. By corrugating the fuel cell membrane electrode structure, Ion Power?s goal is to realize both the Pt utilization targets as well as the power density targets of the DOE. This will be achieved by demonstrating a fuel cell single cell (50 cm2) with a twofold increase in the membrane active area over the geometric area of the cell by corrugating the MEA structure. The corrugating structure must be able to demonstrate the target properties of < 10 mOhm-cm2 electrical resistance at > 20 psi compressive strength over the active area, in combination with offering at least 80% of power density that can be achieved by using the same MEA in a flat plate structure. Corrugated membrane fuel cell structures also have the potential to meet DOE power density targets by essentially packaging more membrane area into the same fuel cell volume as compared to conventional stack constructions.

  11. 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.

  12. Surface modification of polyacrylonitrile co-polymer membranes using pulsed direct current nitrogen plasma

    Energy Technology Data Exchange (ETDEWEB)

    Pal, Dipankar; Neogi, Sudarsan; De, Sirshendu, E-mail: sde@che.iitkgp.ernet.in

    2015-12-31

    Low temperature plasma treatment using pulsed direct current discharge of nitrogen gas was employed to enhance hydrophilicity of the polyacrylonitrile co-polymer membranes. The membranes were characterized in terms of morphology, structure, hydrophilicity, and membrane performance. Properties and functional groups on the surface of polyacrylonitrile co-polymer membranes were investigated by contact angle, scanning electron microscopy, Fourier transform infrared and X-ray photoelectron spectroscopy. Effects of plasma conditions, namely, pulsed voltage, duty cycle and treatment time on increase in membrane hydrophilicity were studied. Permeability of treated membrane was increased by 47% and it was retained up to 70 days. Surface etching due to plasma treatment was confirmed by weight loss of the treated membranes. Due to surface etching, average pore size increased and rejection of 200 kDa polyethylene glycol decreased to about 70% for the treated membrane. Oxygen and nitrogen functional groups were responsible for surface hydrophilicity. - Highlights: • Surface modification of polyacrylonitrile co-polymer membranes by pulsed direct current nitrogen plasma • Hydrophilic functional groups incorporated on the membrane surface • Significant enhancement of the permeability and wettability of the membranes • Water contact angle increased with storage time and finally stabilized.

  13. Atmospheric-pressure plasma activation and surface characterization on polyethylene membrane separator

    Science.gov (United States)

    Tseng, Yu-Chien; Li, Hsiao-Ling; Huang, Chun

    2017-01-01

    The surface hydrophilic activation of a polyethylene membrane separator was achieved using an atmospheric-pressure plasma jet. The surface of the atmospheric-pressure-plasma-treated membrane separator was found to be highly hydrophilic realized by adjusting the plasma power input. The variations in membrane separator chemical structure were confirmed by Fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy. Chemical analysis showed newly formed carbonyl-containing groups and high surface concentrations of oxygen-containing species on the atmospheric-pressure-plasma-treated polymeric separator surface. It also showed that surface hydrophilicity primarily increased from the polar component after atmospheric-pressure plasma treatment. The surface and pore structures of the polyethylene membrane separator were examined by scanning electron microscopy, revealing a slight alteration in the pore structure. As a result of the incorporation of polar functionalities by atmospheric-pressure plasma activation, the electrolyte uptake and electrochemical impedance of the atmospheric-pressure-plasma-treated membrane separator improved. The investigational results show that the separator surface can be controlled by atmospheric-pressure plasma surface treatment to tailor the hydrophilicity and enhance the electrochemical performance of lithium ion batteries.

  14. 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.

  15. Boar seminal plasma exosomes maintain sperm function by infiltrating into the sperm membrane.

    Science.gov (United States)

    Du, Jian; Shen, Jian; Wang, Yuanxian; Pan, Chuanying; Pang, Weijun; Diao, Hua; Dong, Wuzi

    2016-09-13

    Seminal plasma ingredients are important for maintenance of sperm viability. This study focuses on the effect of boar seminal plasma exosomes on sperm function during long-term liquid storage. Boar seminal plasma exosomes had typical nano-structure morphology as measured by scanning electron microscopy (SEM) and molecular markers such as AWN, CD9 and CD63 by western blot analysis. The effect on sperm parameters of adding different ratio of boar seminal plasma exosomes to boar sperm preparations was analyzed. Compared to the diluent without exosomes, the diluent with four times or sixteen times exosomes compared to original semen had higher sperm motility, prolonged effective survival time, improved sperm plasma membrane integrity (p membrane of sperm head which could improve sperm plasma membrane integrity.

  16. The plasma membrane proteome of germinating barley embryos

    DEFF Research Database (Denmark)

    Hynek, Radovan; Svensson, Birte; Jensen, O.N.

    2009-01-01

    was used to reduce soluble protein contamination and enrich for hydrophobic proteins. Sixty-one proteins in 14 SDS-PAGE bands were identified by LC-MS/MS and database searches. The identifications provide new insight into the plasma membrane functions in seed germination.......Cereal seed germination involves a complex coordination between different seed tissues. Plasma membranes must play crucial roles in coordination and execution of germination; however, very little is known about seed plasma membrane proteomes due to limited tissue amounts combined...... with amphiphilicity and low abundance of membrane proteins. A fraction enriched in plasma membranes was prepared from embryos dissected from 18 h germinated barley seeds using aqueous two-phase partitioning. Reversed-phase chromatography on C-4 resin performed in micro-spin columns with stepwise elution by 2-propanol...

  17. Shuttling of G protein subunits between the plasma membrane and intracellular membranes.

    Science.gov (United States)

    Chisari, Mariangela; Saini, Deepak Kumar; Kalyanaraman, Vani; Gautam, Narasimhan

    2007-08-17

    Heterotrimeric G proteins (alphabetagamma) mediate the majority of signaling pathways in mammalian cells. It is long held that G protein function is localized to the plasma membrane. Here we examined the spatiotemporal dynamics of G protein localization using fluorescence recovery after photobleaching, fluorescence loss in photobleaching, and a photoswitchable fluorescent protein, Dronpa. Unexpectedly, G protein subunits shuttle rapidly (t1/2 bromopalmitate. Thus, contrary to present thought, G proteins do not reside permanently on the plasma membrane but are constantly testing the cytoplasmic surfaces of the plasma membrane and endomembranes to maintain G protein pools in intracellular membranes to establish direct communication between receptors and endomembranes.

  18. Molecular Structure of Membrane Tethers

    OpenAIRE

    Baoukina, Svetlana; Marrink, Siewert J.; Tieleman, D. Peter

    2012-01-01

    Membrane tethers are nanotubes formed by a lipid bilayer. They play important functional roles in cell biology and provide an experimental window on lipid properties. Tethers have been studied extensively in experiments and described by theoretical models, but their molecular structure remains unknown due to their small diameters and dynamic nature. We used molecular dynamics simulations to obtain molecular-level insight into tether formation. Tethers were pulled from single-component lipid b...

  19. 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.

  20. 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.

  1. Identification and role of plasma membrane aquaporin in maize root

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    Using antiserum against expressed aquaporin fusion protein, GST-RD28, the distribution of aquaporin in the plasma membrane of maize root protoplasts has been examined under confocal laser scanning microscopy by indirect fluorescence staining. Results indicate that there are abundant aquaporins in maize roots, which are distributed in plasma membrane unevenly. Western blotting analysis of total protein solubilized from maize root plasma membrane shows that antiserum against GST-RD28 can cross-react with one protein around 55 ku. Another 28 ku protein can also be detected when the concentration of SDS and DTT in SDS-PAGE sample buffer is increased. The 55 and 28 ku proteins may be dimeric and monomeric of aquaporin respectively. Functional experiments show that aquaporin blocker HgCl2 and aquaporin antiserum can suppress the swelling of maize root protoplasts in hypotonic solution, indicating that aquaporin in plasma membrane of protoplast facilitates rapid transmembrane water flow.

  2. Localization of plasma membrane t-SNAREs syntaxin 2 and 3 in intracellular compartments

    Directory of Open Access Journals (Sweden)

    Kuismanen Esa

    2005-05-01

    Full Text Available Abstract Background Membrane fusion requires the formation of a complex between a vesicle protein (v-SNARE and the target membrane proteins (t-SNAREs. Syntaxin 2 and 3 are t-SNAREs that, according to previous over-expression studies, are predominantly localized at the plasma membrane. In the present study we investigated localization of the endogenous syntaxin 2 and 3. Results Endogenous syntaxin 2 and 3 were found in NRK cells in intracellular vesicular structures in addition to regions of the plasma membrane. Treatment of these cells with N-ethylmaleimide (NEM, which is known to inactivate membrane fusion, caused syntaxin 3 to accumulate in the trans-Golgi network and syntaxin 2 in perinuclear membrane vesicles. Kinetic analysis in the presence of NEM indicated that this redistribution of syntaxin 2 and 3 takes place via actin containing structures. Conclusion Our data suggest that syntaxin 2 cycles between the plasma membrane and the perinuclear compartment whereas syntaxin 3 cycles between the plasma membrane and the trans-Golgi network. It is possible that this cycling has an important role in the regulation of t-SNARE function.

  3. 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...

  4. 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.

  5. 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...

  6. Autoinhibitory Regulation of Plasma Membrane H+-ATPases

    DEFF Research Database (Denmark)

    Pedersen, Jesper Torbøl

    Electrochemical gradients across cell membranes are essential for nutrient uptake. In plant and fungal cells the electrochemical gradient across the plasma membrane (PM) can build much higher than in mammalian cells. The protein responsible for this gradient is the essential PM H+-ATPase that uses...

  7. 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.

  8. The Role of the Plasma Membrane in the Response of Plant Roots to Aluminum Toxicity

    OpenAIRE

    Ahn, Sung-Ju; Matsumoto, Hideaki

    2006-01-01

    Al3+, the predominant form of solubilized aluminum at pH values below 5.0, has been shown to exert a profound inhibitory effect on root elongation. Al is known to accumulate at the root apex. The plasma membrane represents the first potential target for Al toxicity, due to its pronounced binding to phospholipids. Al appears to alter both the structure and functions of the plasma membrane, and a great deal of research has been conducted concerning the interactions between Al and the plasma mem...

  9. Sulfonated polystyrene-type plasma-polymerized membranes for miniature direct methanol fuel cells

    Science.gov (United States)

    Roualdes, Stéphanie; Topala, Ionut; Mahdjoub, Habiba; Rouessac, Vincent; Sistat, Philippe; Durand, Jean

    Sulfonated polystyrene-type membranes were synthesized by plasma polymerization of a mixture of styrene and trifluoromethane sulfonic acid monomers in a low-frequency after-glow discharge plasma reactor. Such a deposition process enables the preservation of the monomers structure, which was confirmed by mass spectrometry analysis. The synthesized plasma-polymerized membranes are dense and uniform with a few microns thickness. Their structure determined by Fourier-transform infra-red spectroscopy and X-ray photoelectron spectroscopy is very rich in sulfonic acid groups (up to 5%) and stable up to 120 °C. Even if their intrinsic proton conductivity is low (10 -1 mS cm -1), directly related to their disorganized and highly cross-linked structure, plasma-polymerized membranes present a proton conduction ability similar to Nafion ® because of their low thickness. Due to their highly cross-linked structure, these membranes enable a reduction of the methanol crossover in a factor 10 by comparison with Nafion ®. Thus, the integration of plasma-polymerized films in miniaturized direct methanol fuel cells as proton-exchange membranes seems promising.

  10. 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.

  11. 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.

  12. HIV-1 buds predominantly at the plasma membrane of primary human macrophages.

    Directory of Open Access Journals (Sweden)

    Sonja Welsch

    2007-03-01

    Full Text Available HIV-1 assembly and release are believed to occur at the plasma membrane in most host cells with the exception of primary macrophages, for which exclusive budding at late endosomes has been reported. Here, we applied a novel ultrastructural approach to assess HIV-1 budding in primary macrophages in an immunomarker-independent manner. Infected macrophages were fed with BSA-gold and stained with the membrane-impermeant dye ruthenium red to identify endosomes and the plasma membrane, respectively. Virus-filled vacuolar structures with a seemingly intracellular localization displayed intense staining with ruthenium red, but lacked endocytosed BSA-gold, defining them as plasma membrane. Moreover, HIV budding profiles were virtually excluded from gold-filled endosomes while frequently being detected on ruthenium red-positive membranes. The composition of cellular marker proteins incorporated into HIV-1 supported a plasma membrane-derived origin of the viral envelope. Thus, contrary to current opinion, the plasma membrane is the primary site of HIV-1 budding also in infected macrophages.

  13. Visualizing structural dynamics of thylakoid membranes

    Science.gov (United States)

    Iwai, Masakazu; Yokono, Makio; Nakano, Akihiko

    2014-01-01

    To optimize photosynthesis, light-harvesting antenna proteins regulate light energy dissipation and redistribution in chloroplast thylakoid membranes, which involve dynamic protein reorganization of photosystems I and II. However, direct evidence for such protein reorganization has not been visualized in live cells. Here we demonstrate structural dynamics of thylakoid membranes by live cell imaging in combination with deconvolution. We observed chlorophyll fluorescence in the antibiotics-induced macrochloroplast in the moss Physcomitrella patens. The three-dimensional reconstruction uncovered the fine thylakoid membrane structure in live cells. The time-lapse imaging shows that the entire thylakoid membrane network is structurally stable, but the individual thylakoid membrane structure is flexible in vivo. Our observation indicates that grana serve as a framework to maintain structural integrity of the entire thylakoid membrane network. Both the structural stability and flexibility of thylakoid membranes would be essential for dynamic protein reorganization under fluctuating light environments. PMID:24442007

  14. 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.

  15. Cryobehavior of the plasma membrane in protoplasts isolated from cold-acclimated Arabidopsis leaves is related to surface area regulation.

    Science.gov (United States)

    Yamazaki, Tomokazu; Kawamura, Yukio; Uemura, Matsuo

    2008-06-01

    Extracellular freezing in plants results in dehydration and mechanical stresses upon the plasma membrane. Plants that acquire enhanced freezing tolerance after cold acclimation can withstand these two physical stresses. To understand the tolerance to freeze-induced physical stresses, the cryobehavior of the plasma membrane was observed using protoplasts isolated from cold-acclimated Arabidopsis thaliana leaves with the combination of a lipophilic fluorescent dye FM 1-43 and cryomicroscopy. We found that many vesicular structures appeared in the cytoplasmic region near the plasma membrane just after extracellular freezing occurred. These structures, referred to as freeze-induced vesicular structures (FIVs), then developed horizontally near the plasma membrane during freezing. There was a strong correlation between the increase in individual FIV size and the decrease in the surface area of the protoplasts during freezing. Some FIVs fused with their neighbors as the temperature decreased. Occasionally, FIVs fused with the plasma membrane, which may be necessary to relax the stress upon the plasma membrane during freezing. Vesicular structures resembling FIVs were also induced when protoplasts were mechanically pressed between a coverslip and slide glass. Fewer FIVs formed when protoplasts were subjected to hyperosmotic solution, suggesting that FIV formation is associated with mechanical stress rather than dehydration. Collectively, these results suggest that cold-acclimated plant cells may balance membrane tension in the plasma membrane by regulating the surface area. This enables plant cells to withstand the direct mechanical stress imposed by extracellular freezing.

  16. Cell membrane fluid-mosaic structure and cancer metastasis.

    Science.gov (United States)

    Nicolson, Garth L

    2015-04-01

    Cancer cells are surrounded by a fluid-mosaic membrane that provides a highly dynamic structural barrier with the microenvironment, communication filter and transport, receptor and enzyme platform. This structure forms because of the physical properties of its constituents, which can move laterally and selectively within the membrane plane and associate with similar or different constituents, forming specific, functional domains. Over the years, data have accumulated on the amounts, structures, and mobilities of membrane constituents after transformation and during progression and metastasis. More recent information has shown the importance of specialized membrane domains, such as lipid rafts, protein-lipid complexes, receptor complexes, invadopodia, and other cellular structures in the malignant process. In describing the macrostructure and dynamics of plasma membranes, membrane-associated cytoskeletal structures and extracellular matrix are also important, constraining the motion of membrane components and acting as traction points for cell motility. These associations may be altered in malignant cells, and probably also in surrounding normal cells, promoting invasion and metastatic colonization. In addition, components can be released from cells as secretory molecules, enzymes, receptors, large macromolecular complexes, membrane vesicles, and exosomes that can modify the microenvironment, provide specific cross-talk, and facilitate invasion, survival, and growth of malignant cells.

  17. Development of Polysulfone Hollow Fiber Porous Supports for High Flux Composite Membranes: Air Plasma and Piranha Etching

    OpenAIRE

    Ilya Borisov; Anna Ovcharova; Danila Bakhtin; Stepan Bazhenov; Alexey Volkov; Rustem Ibragimov; Rustem Gallyamov; Galina Bondarenko; Rais Mozhchil; Alexandr Bildyukevich; Vladimir Volkov

    2017-01-01

    For the development of high efficiency porous supports for composite membrane preparation, polysulfone (PSf) hollow fiber membranes (outer diameter 1.57 mm, inner diameter 1.12 mm) were modified by air plasma using the low temperature plasma treatment pilot plant which is easily scalable to industrial level and the Piranha etch (H2O2 + H2SO4). Chemical and plasma modification affected only surface layers and did not cause PSf chemical structure change. The modifications led to surface roughne...

  18. 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.

  19. Elevated blood Hsp60, its structural similarities and cross-reactivity with thyroid molecules, and its presence on the plasma membrane of oncocytes point to the chaperonin as an immunopathogenic factor in Hashimoto's thyroiditis.

    Science.gov (United States)

    Marino Gammazza, Antonella; Rizzo, Manfredi; Citarrella, Roberto; Rappa, Francesca; Campanella, Claudia; Bucchieri, Fabio; Patti, Angelo; Nikolic, Dragana; Cabibi, Daniela; Amico, Giandomenico; Conaldi, Pier Giulio; San Biagio, Pier Luigi; Montalto, Giuseppe; Farina, Felicia; Zummo, Giovanni; Conway de Macario, Everly; Macario, Alberto J L; Cappello, Francesco

    2014-05-01

    The role Hsp60 might play in various inflammatory and autoimmune diseases is under investigation, but little information exists pertaining to Hashimoto's thyroiditis (HT). With the aim to fill this gap, in the present work, we directed our attention to Hsp60 participation in HT pathogenesis. We found Hsp60 levels increased in the blood of HT patients compared to controls. The chaperonin was immunolocalized in thyroid tissue specimens from patients with HT, both in thyrocytes and oncocytes (Hurthle cells) with higher levels compared to controls (goiter). In oncocytes, we found Hsp60 not only in the cytoplasm but also on the plasma membrane, as shown by double immunofluorescence performed on fine needle aspiration cytology. By bioinformatics, we found regions in the Hsp60 molecule with remarkable structural similarity with the thyroglobulin (TG) and thyroid peroxidase (TPO) molecules, which supports the notion that autoantibodies against TG and TPO are likely to recognize Hsp60 on the plasma membrane of oncocytes. This was also supported by data obtained by ELISA, showing that anti-TG and anti-TPO antibodies cross-react with human recombinant Hsp60. Antibody-antigen (Hsp60) reaction on the cell surface could very well mediate thyroid cell damage and destruction, perpetuating inflammation. Experiments with recombinant Hsp60 did not show stimulation of cytokine production by peripheral blood mononuclear cells from HT patients. All together, these results led us to hypothesize that Hsp60 may be an active player in HT pathogenesis via an antibody-mediated immune mechanism.

  20. Presence of membranous vesicles in cat seminal plasma: ultrastructural characteristics, protein profile and enzymatic activity.

    Science.gov (United States)

    Polisca, A; Troisi, A; Minelli, A; Bellezza, I; Fontbonne, A; Zelli, R

    2015-02-01

    This study sought to verify the presence of membranous vesicles in cat seminal plasma by means of transmission electron microscopy and to identify protein profile and some of the enzymatic activities associated with these particles. The transmission electron microscopy observations showed the existence of different sized vesicular membranous structures of more or less spherical shape. These vesicles were surrounded by single-, double- or multiple-layered laminar membranes. The vesicle diameters ranged from 16.3 to 387.4 nm, with a mean of 116.5 ± 70.7 nm. Enzyme activity determinations showed the presence of dipeptilpeptidase IV, aminopeptidase, alkaline and acid phosphatase. To our knowledge, this is the first report that identifies and characterizes the membranous vesicles in cat seminal plasma. However, further studies are necessary to identify the exact site of production of these membranous vesicles in the cat male genital tract and to determine their specific roles in the reproductive events of this species.

  1. The dynamics of plant plasma membrane proteins: PINs and beyond.

    Science.gov (United States)

    Luschnig, Christian; Vert, Grégory

    2014-08-01

    Plants are permanently situated in a fixed location and thus are well adapted to sense and respond to environmental stimuli and developmental cues. At the cellular level, several of these responses require delicate adjustments that affect the activity and steady-state levels of plasma membrane proteins. These adjustments involve both vesicular transport to the plasma membrane and protein internalization via endocytic sorting. A substantial part of our current knowledge of plant plasma membrane protein sorting is based on studies of PIN-FORMED (PIN) auxin transport proteins, which are found at distinct plasma membrane domains and have been implicated in directional efflux of the plant hormone auxin. Here, we discuss the mechanisms involved in establishing such polar protein distributions, focusing on PINs and other key plant plasma membrane proteins, and we highlight the pathways that allow for dynamic adjustments in protein distribution and turnover, which together constitute a versatile framework that underlies the remarkable capabilities of plants to adjust growth and development in their ever-changing environment.

  2. Facilitative plasma membrane transporters function during ER transit.

    Science.gov (United States)

    Takanaga, Hitomi; Frommer, Wolf B

    2010-08-01

    Although biochemical studies suggested a high permeability of the endoplasmic reticulum (ER) membrane for small molecules, proteomics identified few specialized ER transporters. To test functionality of transporters during ER passage, we tested whether glucose transporters (GLUTs, SGLTs) destined for the plasma membrane are active during ER transit. HepG2 cells were characterized by low-affinity ER transport activity, suggesting that ER uptake is protein mediated. The much-reduced capacity of HEK293T cells to take up glucose across the plasma membrane correlated with low ER transport. Ectopic expression of GLUT1, -2, -4, or -9 induced GLUT isoform-specific ER transport activity in HEK293T cells. In contrast, the Na(+)-glucose cotransporter SGLT1 mediated efficient plasma membrane glucose transport but no detectable ER uptake, probably because of lack of a sufficient sodium gradient across the ER membrane. In conclusion, we demonstrate that GLUTs are sufficient for mediating ER glucose transport en route to the plasma membrane. Because of the low volume of the ER, trace amounts of these uniporters contribute to ER solute import during ER transit, while uniporters and cation-coupled transporters carry out export from the ER, together potentially explaining the low selectivity of ER transport. Expression levels and residence time of transporters in the ER, as well as their coupling mechanisms, could be key determinants of ER permeability.

  3. Detergent-resistant membrane subfractions containing proteins of plasma membrane, mitochondrial, and internal membrane origins.

    Science.gov (United States)

    Mellgren, Ronald L

    2008-04-24

    HEK293 cell detergent-resistant membranes (DRMs) isolated by the standard homogenization protocol employing a Teflon pestle homogenizer yielded a prominent opaque band at approximately 16% sucrose upon density gradient ultracentrifugation. In contrast, cell disruption using a ground glass tissue homogenizer generated three distinct DRM populations migrating at approximately 10%, 14%, and 20% sucrose, named DRM subfractions A, B, and C, respectively. Separation of the DRM subfractions by mechanical disruption suggested that they are physically associated within the cellular environment, but can be dissociated by shear forces generated during vigorous homogenization. All three DRM subfractions possessed cholesterol and ganglioside GM1, but differed in protein composition. Subfraction A was enriched in flotillin-1 and contained little caveolin-1. In contrast, subfractions B and C were enriched in caveolin-1. Subfraction C contained several mitochondrial membrane proteins, including mitofilin and porins. Only subfraction B appeared to contain significant amounts of plasma membrane-associated proteins, as revealed by cell surface labeling studies. A similar distribution of DRM subfractions, as assessed by separation of flotillin-1 and caveolin-1 immunoreactivities, was observed in CHO cells, in 3T3-L1 adipocytes, and in HEK293 cells lysed in detergent-free carbonate. Teflon pestle homogenization of HEK293 cells in the presence of the actin-disrupting agent latrunculin B generated DRM subfractions A-C. The microtubule-disrupting agent vinblastine did not facilitate DRM subfraction separation, and DRMs prepared from fibroblasts of vimentin-null mice were present as a single major band on sucrose gradients, unless pre-treated with latrunculin B. These results suggest that the DRM subfractions are interconnected by the actin cytoskeleton, and not by microtubes or vimentin intermediate filaments. The subfractions described may prove useful in studying discrete protein

  4. 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.

  5. Detection of glycoproteins in the Acanthamoeba plasma membrane

    Energy Technology Data Exchange (ETDEWEB)

    Paatero, G.I.L. (Abo Akademi (Finland)); Gahmberg, C.G. (Univ. of Helsinki (Finland))

    1988-11-01

    In the present study the authors have shown that glycoproteins are present in the plasma membrane of Acanthamoeba castellanii by utilizing different radioactive labeling techniques. Plasma membrane proteins in the amoeba were iodinated by {sup 125}I-lactoperoxidase labeling and the solubilized radiolabeled glycoproteins were separated by lectin-Sepharose affinity chromatography followed by polyacrylamide gel electrophoresis. The periodate/NaB{sup 3}H{sub 4} and galactose oxidase/NaB{sup 3}H{sub 4} labeling techniques were used for labeling of surface carbohydrates in the amoeba. Several surface-labeled glycoproteins were observed in addition to a diffusely labeled region with M{sub r} of 55,000-75,000 seen on electrophoresis, which could represent glycolipids. The presence of glycoproteins in the plasma membrane of Acanthamoeba castellanii was confirmed by metabolic labeling with ({sup 35}S)methionine followed by lectin-Sepharose affinity chromatography and polyacrylamide gel electrophoresis.

  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. 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.

  8. Production of selective membranes using plasma deposited nanochanneled thin films

    Directory of Open Access Journals (Sweden)

    Rodrigo Amorim Motta Carvalho

    2006-12-01

    Full Text Available The hydrolization of thin films obtained by tetraethoxysilane plasma polymerization results in the formation of a nanochanneled silicone like structure that could be useful for the production of selective membranes. Therefore, the aim of this work is to test the permeation properties of hydrolyzed thin films. The films were tested for: 1 permeation of polar organic compounds and/or water in gaseous phase and 2 permeation of salt in liquid phase. The efficiency of permeation was tested using a quartz crystal microbalance (QCM technique in gas phase and conductimetric analysis (CA in liquid phase. The substrates used were: silicon for characterization of the deposited films, piezoelectric quartz crystals for tests of selective membranes and cellophane paper for tests of permeation. QCM analysis showed that the nanochannels allow the adsorption and/or permeation of polar organic compounds, such as acetone and 2-propanol, and water. CA showed that the films allow salt permeation after an inhibition time needed for hydrolysis of the organic radicals within the film. Due to their characteristics, the films can be used for grains protection against microorganism proliferation during storage without preventing germination.

  9. Surface Modification of Asymmetric Polysulfone/Polyethylene Glycol Membranes by DC Ar-Glow Discharge Plasma

    Directory of Open Access Journals (Sweden)

    Chalad Yuenyao

    2016-01-01

    Full Text Available Polysulfone/polyethylene glycol (PSF/PEG membranes were prepared by dry/wet phase inversion method. Effects of direct current glow discharge plasma using argon as working gas on morphological structures and gas separation properties of membranes were studied. Alteration of membrane characteristics were analyzed by various techniques like contact angle, scanning electron microscope, Fourier transform infrared spectroscopy, and dynamic mechanical thermal analysis. Gas separation properties were measured in terms of permeation and ideal O2/N2 selectivity. Results showed that hydrophilic and gas separation properties of PSF/PEG membranes increased by plasma surface modification. It was also shown that the dosage of PEG and plasma treatment affected the morphological structures and mechanical and gas separation properties. The macro voids and transmembrane structure disappeared with a little amount of PEG dosage. Pore size and mechanical strength tend to decrease with increasing PEG dosage up to 10 wt%. Glass transition temperature (Tg receded from 201.8 to 143.7°C for pure PSF and PSF/PEG with PEG dosage of 10 wt%. O2 and N2 gases permeation through the 10-minute plasma treated membranes tend to increase. However, the permeation strongly dispersed when treatment time was more extended.

  10. 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.

  11. Plasma membrane electron transport in frog blood vessels

    Indian Academy of Sciences (India)

    Rashmi P Rao; K Nalini; J Prakasa Rao

    2009-12-01

    In an attempt to see if frog blood vessels possess a plasma membrane electron transport system, the postcaval vein and aorta isolated from Rana tigrina were tested for their ability to reduce ferricyanide, methylene blue, and 2,6-dichloroindophenol. While the dyes remained unchanged, ferricyanide was reduced to ferrocyanide. This reduction was resistant to inhibition by cyanide and azide. Heptane extraction or formalin fixation of the tissues markedly reduced the capability to reduce ferricyanide. Denuded aortas retained only 30% of the activity of intact tissue. Our results indicate that the amphibian postcaval vein and aorta exhibit plasma membrane electron transport

  12. Therapeutic plasmapheresis using membrane plasma separation.

    Science.gov (United States)

    Sinha, Aditi; Tiwari, Anand Narain; Chanchlani, Rahul; Seetharamanjaneyulu, V; Hari, Pankaj; Bagga, Arvind

    2012-08-01

    The authors present their experience with therapeutic plasmapheresis (TPE) using membrane filters at the pediatric dialysis unit of a referral center. Between January 2006 and December 2010, 486 sessions of TPE were performed in 39 patients (range 6-17 y), chiefly for atypical hemolytic uremic syndrome (HUS, n = 22), crescentic glomerulonephritis (n = 8) and focal segmental glomerulosclerosis (n = 5). Satisfactory response was noted in 32 patients, particularly with HUS (n = 22) or crescentic glomerulonephritis (n = 6). Adverse effects included chills or urticaria (n = 8 sessions), hypocalcemia (n = 6) and hypotension (n = 5). The present findings highlight the safety, efficacy and feasibility of TPE using membrane filtration.

  13. Regulation of plant plasma membrane H+- and Ca2+-ATPases by terminal domains

    DEFF Research Database (Denmark)

    Bækgaard, Lone; Fuglsang, Anja Thoe; Palmgren, Michael Gjedde

    2005-01-01

    In the last few years, major progress has been made to elucidate the structure, function, and regulation of P-type plasma membrane H(+)-and Ca(2+)-ATPases. Even though a number of regulatory proteins have been identified, many pieces are still lacking in order to understand the complete regulator...... mechanisms of these pumps. In plant plasma membrane H(+)- and Ca(2+)-ATPases, autoinhibitory domains are situated in the C- and N-terminal domains, respectively. A model for a common mechanism of autoinhibition is discussed....

  14. Ternary structure reveals mechanism of a membrane diacylglycerol kinase

    Science.gov (United States)

    Li, Dianfan; Stansfeld, Phillip J.; Sansom, Mark S. P.; Keogh, Aaron; Vogeley, Lutz; Howe, Nicole; Lyons, Joseph A.; Aragao, David; Fromme, Petra; Fromme, Raimund; Basu, Shibom; Grotjohann, Ingo; Kupitz, Christopher; Rendek, Kimberley; Weierstall, Uwe; Zatsepin, Nadia A.; Cherezov, Vadim; Liu, Wei; Bandaru, Sateesh; English, Niall J.; Gati, Cornelius; Barty, Anton; Yefanov, Oleksandr; Chapman, Henry N.; Diederichs, Kay; Messerschmidt, Marc; Boutet, Sébastien; Williams, Garth J.; Marvin Seibert, M.; Caffrey, Martin

    2015-12-01

    Diacylglycerol kinase catalyses the ATP-dependent conversion of diacylglycerol to phosphatidic acid in the plasma membrane of Escherichia coli. The small size of this integral membrane trimer, which has 121 residues per subunit, means that available protein must be used economically to craft three catalytic and substrate-binding sites centred about the membrane/cytosol interface. How nature has accomplished this extraordinary feat is revealed here in a crystal structure of the kinase captured as a ternary complex with bound lipid substrate and an ATP analogue. Residues, identified as essential for activity by mutagenesis, decorate the active site and are rationalized by the ternary structure. The γ-phosphate of the ATP analogue is positioned for direct transfer to the primary hydroxyl of the lipid whose acyl chain is in the membrane. A catalytic mechanism for this unique enzyme is proposed. The active site architecture shows clear evidence of having arisen by convergent evolution.

  15. Membrane protein structure determination in membrana.

    Science.gov (United States)

    Ding, Yi; Yao, Yong; Marassi, Francesca M

    2013-09-17

    The two principal components of biological membranes, the lipid bilayer and the proteins integrated within it, have coevolved for specific functions that mediate the interactions of cells with their environment. Molecular structures can provide very significant insights about protein function. In the case of membrane proteins, the physical and chemical properties of lipids and proteins are highly interdependent; therefore structure determination should include the membrane environment. Considering the membrane alongside the protein eliminates the possibility that crystal contacts or detergent molecules could distort protein structure, dynamics, and function and enables ligand binding studies to be performed in a natural setting. Solid-state NMR spectroscopy is compatible with three-dimensional structure determination of membrane proteins in phospholipid bilayer membranes under physiological conditions and has played an important role in elucidating the physical and chemical properties of biological membranes, providing key information about the structure and dynamics of the phospholipid components. Recently, developments in the recombinant expression of membrane proteins, sample preparation, pulse sequences for high-resolution spectroscopy, radio frequency probes, high-field magnets, and computational methods have enabled a number of membrane protein structures to be determined in lipid bilayer membranes. In this Account, we illustrate solid-state NMR methods with examples from two bacterial outer membrane proteins (OmpX and Ail) that form integral membrane β-barrels. The ability to measure orientation-dependent frequencies in the solid-state NMR spectra of membrane-embedded proteins provides the foundation for a powerful approach to structure determination based primarily on orientation restraints. Orientation restraints are particularly useful for NMR structural studies of membrane proteins because they provide information about both three-dimensional structure

  16. A cell-free assay to determine the stoichiometry of plasma membrane proteins.

    Science.gov (United States)

    Trigo, Cesar; Vivar, Juan P; Gonzalez, Carlos B; Brauchi, Sebastian

    2013-04-01

    Plasma membrane receptors, transporters, and ion channel molecules are often found as oligomeric structures that participate in signaling cascades essential for cell survival. Different states of protein oligomerization may play a role in functional control and allosteric regulation. Stochastic GFP-photobleaching (SGP) has emerged as an affordable and simple method to determine the stoichiometry of proteins at the plasma membrane. This non-invasive optical approach can be useful for total internal reflection of fluorescence microscopy (TIRFM), where signal-to-noise ratio is very high at the plasma membrane. Here, we report an alternative methodology implemented on a standard laser scanning confocal microscope (LSCM). The simplicity of our method will allow for its implementation in any epifluorescence microscope of choice.

  17. The connection of cytoskeletal network with plasma membrane and the cell wall

    Institute of Scientific and Technical Information of China (English)

    Zengyu Liu; Staffan Persson; Yi Zhang

    2015-01-01

    The cell wall provides external support of the plant cells, while the cytoskeletons including the microtubules and the actin filaments constitute an internal framework. The cytoskeletons contribute to the cell wall biosynthesis by spatially and temporarily regulating the transportation and deposition of cell wall components. This tight control is achieved by the dynamic behavior of the cytoskeletons, but also through the tethering of these structures to the plasma membrane. This tethering may also extend beyond the plasma membrane and impact on the cell wall, possibly in the form of a feedback loop. In this review, we discuss the linking components between the cytoskeletons and the plasma membrane, and/or the cell wall. We also discuss the prospective roles of these components in cell wall biosyn-thesis and modifications, and aim to provide a platform for further studies in this field.

  18. Advanced Fluorescence Microscopy Approaches to Understand the Dynamic Organization of the Plasma Membrane in Eukaryotes

    DEFF Research Database (Denmark)

    Ziomkiewicz, Iwona

    The plasma membrane (PM) is a physical barrier that defines the boundaries of a cell. It not only isolates the cell interior from the environment, but also enables cell communication and a selective exchange of solutes. To serve those contrasting functions, the PM has a dynamic structure consisting...

  19. Identification of calcium-binding proteins associated with the human sperm plasma membrane

    National Research Council Canada - National Science Library

    Naaby-Hansen, Soren; Diekman, Alan; Shetty, Jagathpala; Flickinger, Charles J; Westbrook, Anne; Herr, John C

    2010-01-01

    The precise composition of the human sperm plasma membrane, the molecular interactions that define domain specific functions, and the regulation of membrane associated proteins during the capacitation...

  20. Kinetics of plasma membrane and mitochondrial alterations in cells undergoing apoptosis

    National Research Council Canada - National Science Library

    Lizard, G; Fournel, S; Genestier, L; Dhedin, N; Chaput, C; Flacher, M; Mutin, M; Panaye, G; Revillard, J P

    1995-01-01

    ... of the nucleus, whereas integrity of the plasma membrane and organelles is preserved. Conversely cells undergoing necrosis display an early desintegration of cytoplasmic membrane and swelling of mitochondria...

  1. Significance of the plasma membrane for the nerve cell function, development and plasticity.

    Science.gov (United States)

    Mourek, Jindrich; Langmeier, Milos; Pokorny, Jaroslav

    2009-01-01

    Lipoid character of plasma membrane namely the presence of polyenic fatty acids enables to interact with membrane proteins and in certain extent also to modulate their function. During the development, molecules of membrane fatty acids become more and more complex, and the ratio of polyenic fatty acids/saturated fatty acids in the brain rises, while the concentration of monoenic fatty acids remained relatively stable. This phenomenon is apparent also in the ratio of unsaturated fatty acids OMEGA-3 in plasma of newborns which correlates with the birth weight. Plasma membrane reflects local specializations of nerve cells. Its composition varies in functionally specialized regions called domains. Specialized domains of nerve cells determine the function of dendrites, soma, axon, axon hillock ect. Premature weaning of laboratory rats results in structural changes and in the increase of excitability of neuronal circuits in hypothalamus, septum and hippocampus which indicate the possibility of membrane composition changes. In synapses, transport proteins of synaptic vesicles, act together with the specific proteins of the presynaptic membrane. Membrane proteins determine the release of neurotransmitter at different conditions of synaptic activity, and they can contribute to the recovery of neurotransmitter content after the repeated hyperactivity. In the model of experimental kindling, repeated seizures bring about decreases and distribution changes of synaptic vesicles.

  2. Resolving mixed mechanisms of protein subdiffusion at the T cell plasma membrane

    Science.gov (United States)

    Golan, Yonatan; Sherman, Eilon

    2017-06-01

    The plasma membrane is a complex medium where transmembrane proteins diffuse and interact to facilitate cell function. Membrane protein mobility is affected by multiple mechanisms, including crowding, trapping, medium elasticity and structure, thus limiting our ability to distinguish them in intact cells. Here we characterize the mobility and organization of a short transmembrane protein at the plasma membrane of live T cells, using single particle tracking and photoactivated-localization microscopy. Protein mobility is highly heterogeneous, subdiffusive and ergodic-like. Using mobility characteristics, we segment individual trajectories into subpopulations with distinct Gaussian step-size distributions. Particles of low-to-medium mobility consist of clusters, diffusing in a viscoelastic and fractal-like medium and are enriched at the centre of the cell footprint. Particles of high mobility undergo weak confinement and are more evenly distributed. This study presents a methodological approach to resolve simultaneous mixed subdiffusion mechanisms acting on polydispersed samples and complex media such as cell membranes.

  3. Interaction of Mason-Pfizer monkey virus matrix protein with plasma membrane.

    Directory of Open Access Journals (Sweden)

    Jan ePrchal

    2014-01-01

    Full Text Available Budding is the final step of the late phase of retroviral life cycle. It begins with the interaction of Gag precursor with plasma membrane through its N-terminal domain, the matrix protein. However, single generas of Retroviridae family differ in the way how they interact with plasma membrane. While in case of lentiviruses (e.g. human immunodeficiency virus (HIV the structural polyprotein precursor Gag interacts with cellular membrane prior to the assembly, betaretroviruses (Mason-Pfizer monkey virus (M-PMV first assemble their virus-like particles in the pericentriolar region of the infected cell and therefore, already assembled particles interact with the membrane. Although both these types of retroviruses use similar mechanism of the interaction of Gag with the membrane, the difference in the site of assembly leads to some differences in the mechanism of the interaction. Here we describe the interaction of M-PMV matrix protein with plasma membrane with emphasis on the structural aspects of the interaction with single phospholipids.

  4. 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

  5. A plasma membrane association module in yeast amino acid transporters

    NARCIS (Netherlands)

    Popov-Čeleketić, Dušan; Bianchi, Frans; Ruiz, Stephanie J; Meutiawati, Febrina; Poolman, Bert

    2016-01-01

    Amino acid permeases (AAPs) in the plasma membrane (PM) of Saccharomyces cerevisiae are responsible for the uptake of amino acids and involved in regulation of their cellular levels. Here, we report on a strong and complex module for PM association found in the C-terminal tail of AAPs. Using in sili

  6. 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

  7. Mammalian gamete plasma membranes re-assessments and reproductive implications

    Science.gov (United States)

    Establishment of the diploid status occurs with the fusion of female and male gametes. Both the mammalian oocyte and spermatozoa are haploid cells surrounded with plasma membranes that are rich in various proteins playing a crucial role during fertilization. Fertilization is a complex and ordered st...

  8. A plasma membrane association module in yeast amino acid transporters

    NARCIS (Netherlands)

    Popov-Čeleketić, Dušan; Bianchi, Frans; Ruiz, Stephanie J; Meutiawati, Febrina; Poolman, Bert

    2016-01-01

    Amino acid permeases (AAPs) in the plasma membrane (PM) of Saccharomyces cerevisiae are responsible for the uptake of amino acids and involved in regulation of their cellular levels. Here, we report on a strong and complex module for PM association found in the C-terminal tail of AAPs. Using in

  9. 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.

  10. 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.

  11. Plant Phosphoproteomics: Analysis of Plasma Membrane Transporters by Mass Spectrometry

    DEFF Research Database (Denmark)

    Ye, Juanying; Rudashevskaya, Elena; Young, Clifford

    important physiological functions, such as stomata aperture, cell elongation, or cellular pH regulation. It is known that the activity of plant plasma membrane H+-ATPase is regulated by phosphorylation. Therefore, we first investigated the phosphorylation profile of plant H+-ATPase by enriching...... the phosphopeptides with optimized TiO2 and IMAC enrichment methods prior to MS analysis. We further investigated the global phosphorylation profile of the whole plant plasma membrane proteins using the combination of our recently established phosphopeptide enrichment method, Calcium phosphate precipitation......  Phosphorylation is a key regulatory factor in all aspects of eukaryotic biology including the regulation of plant membrane-bound transport proteins. To date, mass spectrometry (MS) has been introduced as powerful technology for study of post translational modifications (PTMs), including protein...

  12. Electrospun superhydrophobic membranes with unique structures for membrane distillation.

    Science.gov (United States)

    Liao, Yuan; Loh, Chun-Heng; Wang, Rong; Fane, Anthony G

    2014-09-24

    With modest temperature demand, low operating pressure, and high solute rejection, membrane distillation (MD) is an attractive option for desalination, waste treatment, and food and pharmaceutical processing. However, large-scale practical applications of MD are still hindered by the absence of effective membranes with high hydrophobicity, high porosity, and adequate mechanical strength, which are important properties for MD permeation fluxes, stable long-term performance, and effective packing in modules without damage. This study describes novel design strategies for highly robust superhydrophobic dual-layer membranes for MD via electrospinning. One of the newly developed membranes comprises a durable and ultrathin 3-dimensional (3D) superhydrophobic skin and porous nanofibrous support whereas another was fabricated by electrospinning 3D superhydrophobic layers on a nonwoven support. These membranes exhibit superhydrophobicity toward distilled water, salty water, oil-in-water emulsion, and beverages, which enables them to be used not only for desalination but also for other processes. The superhydrophobic dual-layer membrane #3S-N with nanofibrous support has a competitive permeation flux of 24.6 ± 1.2 kg m(-2) h(-1) in MD (feed and permeate temperate were set as 333 and 293 K, respectively) due to the higher porosity of the nanofibrous scaffold. Meanwhile, the membranes with the nonwoven support exhibit greater mechanical strength due to this support combined with better long-term performance because of the thicker 3D superhydrophobic layers. The morphology, pore size, porosity, mechanical properties, and liquid enter pressure of water of these superhydrophobic composite membranes with two different structures are reported and compared with commercial polyvinylidene fluoride membranes.

  13. Small unilamellar liposomes as a membrane model for cell inactivation by cold atmospheric plasma treatment

    Science.gov (United States)

    Maheux, S.; Frache, G.; Thomann, J. S.; Clément, F.; Penny, C.; Belmonte, T.; Duday, D.

    2016-09-01

    Cold atmospheric plasma is thought to be a promising tool for numerous biomedical applications due to its ability to generate a large diversity of reactive species in a controlled way. In some cases, it can also generate pulsed electric fields at the zone of treatment, which can induce processes such as electroporation in cell membranes. However, the interaction of these reactive species and the pulse electric field with cells in a physiological medium is very complex, and we still need a better understanding in order to be useful for future applications. A way to reach this goal is to work with model cell membranes such as liposomes, with the simplest physiological liquid and in a controlled atmosphere in order to limit the number of parallel reactions and processes. In this paper, where this approach has been chosen, 1,2-Dioleoyl-sn-glycero-3-phosphocholine (DOPC) small unilamellar vesicles (SUV) have been synthesized in a phosphate buffered aqueous solution, and this solution has been treated by a nanosecond pulsed plasma jet under a pure nitrogen atmosphere. It is only the composition of the plasma gas that has been changed in order to generate different cocktails of reactive species. After the quantification of the main plasma reactive species in the phosphate buffered saline (PBS) solution, structural, surface charge state, and chemical modifications generated on the plasma treated liposomes, due to the interaction with the plasma reactive species, have been carefully characterized. These results allow us to further understand the effect of plasma reactive species on model cell membranes in physiological liquids. The permeation through the liposomal membrane and the reaction of plasma reactive species with molecules encapsulated inside the liposomes have also been evaluated. New processes of degradation are finally presented and discussed, which come from the specific conditions of plasma treatment under the pure nitrogen atmosphere.

  14. Modeling branching pore structures in membrane filters

    Science.gov (United States)

    Sanaei, Pejman; Cummings, Linda J.

    2016-11-01

    Membrane filters are in widespread industrial use, and mathematical models to predict their efficacy are potentially very useful, as such models can suggest design modifications to improve filter performance and lifetime. Many models have been proposed to describe particle capture by membrane filters and the associated fluid dynamics, but most such models are based on a very simple structure in which the pores of the membrane are assumed to be simple circularly-cylindrical tubes spanning the depth of the membrane. Real membranes used in applications usually have much more complex geometry, with interconnected pores which may branch and bifurcate. Pores are also typically larger on the upstream side of the membrane than on the downstream side. We present an idealized mathematical model, in which a membrane consists of a series of bifurcating pores, which decrease in size as the membrane is traversed. Feed solution is forced through the membrane by applied pressure, and particles are removed from the feed either by sieving, or by particle adsorption within pores (which shrinks them). Thus the membrane's permeability decreases as the filtration progresses, ultimately falling to zero. We discuss how filtration efficiency depends on the characteristics of the branching structure. Partial support from NSF DMS 1261596 is gratefully acknowledged.

  15. Ssh4, Rcr2 and Rcr1 affect plasma membrane transporter activity in Saccharomyces cerevisiae.

    Science.gov (United States)

    Kota, Jhansi; Melin-Larsson, Monika; Ljungdahl, Per O; Forsberg, Hanna

    2007-04-01

    Nutrient uptake in the yeast Saccharomyces cerevisiae is a highly regulated process. Cells adjust levels of nutrient transporters within the plasma membrane at multiple stages of the secretory and endosomal pathways. In the absence of the ER-membrane-localized chaperone Shr3, amino acid permeases (AAP) inefficiently fold and are largely retained in the ER. Consequently, shr3 null mutants exhibit greatly reduced rates of amino acid uptake due to lower levels of AAPs in their plasma membranes. To further our understanding of mechanisms affecting AAP localization, we identified SSH4 and RCR2 as high-copy suppressors of shr3 null mutations. The overexpression of SSH4, RCR2, or the RCR2 homolog RCR1 increases steady-state AAP levels, whereas the genetic inactivation of these genes reduces steady-state AAP levels. Additionally, the overexpression of any of these suppressor genes exerts a positive effect on phosphate and uracil uptake systems. Ssh4 and Rcr2 primarily localize to structures associated with the vacuole; however, Rcr2 also localizes to endosome-like vesicles. Our findings are consistent with a model in which Ssh4, Rcr2, and presumably Rcr1, function within the endosome-vacuole trafficking pathway, where they affect events that determine whether plasma membrane proteins are degraded or routed to the plasma membrane.

  16. Hierarchically structured, nitrogen-doped carbon membranes

    KAUST Repository

    Wang, Hong

    2017-08-03

    The present invention is a structure, method of making and method of use for a novel macroscopic hierarchically structured, nitrogen-doped, nano-porous carbon membrane (HNDCMs) with asymmetric and hierarchical pore architecture that can be produced on a large-scale approach. The unique HNDCM holds great promise as components in separation and advanced carbon devices because they could offer unconventional fluidic transport phenomena on the nanoscale. Overall, the invention set forth herein covers a hierarchically structured, nitrogen-doped carbon membranes and methods of making and using such a membranes.

  17. Lectin receptor kinases participate in protein-protein interactions to mediate plasma membrane-cell wall adhesions in Arabidopsis

    NARCIS (Netherlands)

    Gouget, A.; Senchou, V.; Govers, F.; Sanson, A.; Barre, A.; Rougé, P.; Pont-Lezica, R.; Canut, H.

    2006-01-01

    Interactions between plant cell walls and plasma membranes are essential for cells to function properly, but the molecules that mediate the structural continuity between wall and membrane are unknown. Some of these interactions, which are visualized upon tissue plasmolysis in Arabidopsis (Arabidopsi

  18. Thermostabilisation of membrane proteins for structural studies

    Science.gov (United States)

    Magnani, Francesca; Serrano-Vega, Maria J.; Shibata, Yoko; Abdul-Hussein, Saba; Lebon, Guillaume; Miller-Gallacher, Jennifer; Singhal, Ankita; Strege, Annette; Thomas, Jennifer A.; Tate, Christopher G.

    2017-01-01

    The thermostability of an integral membrane protein in detergent solution is a key parameter that dictates the likelihood of obtaining well-diffracting crystals suitable for structure determination. However, many mammalian membrane proteins are too unstable for crystallisation. We developed a thermostabilisation strategy based on systematic mutagenesis coupled to a radioligand-binding thermostability assay that can be applied to receptors, ion channels and transporters. It takes approximately 6-12 months to thermostabilise a G protein-coupled receptor (GPCR) containing 300 amino acid residues. The resulting thermostabilised membrane proteins are more easily crystallised and result in high-quality structures. This methodology has facilitated structure-based drug design applied to GPCRs, because it is possible to determine multiple structures of the thermostabilised receptors bound to low affinity ligands. Protocols and advice are given on how to develop thermostability assays for membrane proteins and how to combine mutations to make an optimally stable mutant suitable for structural studies. PMID:27466713

  19. 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.

  20. Rapid Preparation of a Plasma Membrane Fraction: Western Blot Detection of Translocated Glucose Transporter 4 from Plasma Membrane of Muscle and Adipose Cells and Tissues.

    Science.gov (United States)

    Yamamoto, Norio; Yamashita, Yoko; Yoshioka, Yasukiyo; Nishiumi, Shin; Ashida, Hitoshi

    2016-08-01

    Membrane proteins account for 70% to 80% of all pharmaceutical targets, indicating their clinical relevance and underscoring the importance of identifying differentially expressed membrane proteins that reflect distinct disease properties. The translocation of proteins from the bulk of the cytosol to the plasma membrane is a critical step in the transfer of information from membrane-embedded receptors or transporters to the cell interior. To understand how membrane proteins work, it is important to separate the membrane fraction of cells. This unit provides a protocol for rapidly obtaining plasma membrane fractions for western blot analysis. © 2016 by John Wiley & Sons, Inc. Copyright © 2016 John Wiley & Sons, Inc.

  1. Development of Polysulfone Hollow Fiber Porous Supports for High Flux Composite Membranes: Air Plasma and Piranha Etching

    Directory of Open Access Journals (Sweden)

    Ilya Borisov

    2017-02-01

    Full Text Available For the development of high efficiency porous supports for composite membrane preparation, polysulfone (PSf hollow fiber membranes (outer diameter 1.57 mm, inner diameter 1.12 mm were modified by air plasma using the low temperature plasma treatment pilot plant which is easily scalable to industrial level and the Piranha etch (H2O2 + H2SO4. Chemical and plasma modification affected only surface layers and did not cause PSf chemical structure change. The modifications led to surface roughness decrease, which is of great importance for further thin film composite (TFC membranes fabrication by dense selective layer coating, and also reduced water and ethylene glycol contact angle values for modified hollow fibers surface. Furthermore, the membranes surface energy increased two-fold. The Piranha mixture chemical modification did not change the membranes average pore size and gas permeance values, while air plasma treatment increased pore size 1.5-fold and also 2 order enhanced membranes surface porosity. Since membranes surface porosity increased due to air plasma treatment the modified membranes were used as efficient supports for preparation of high permeance TFC membranes by using poly[1-(trimethylsilyl-1-propyne] as an example for selective layer fabrication.

  2. PLASMA POLYMERIZATION OF HYDROPHILIC AND HYDROPHOBIC MONOMERS FOR SURFACE MODIFICATION OF NUCLE-MICROPOROUS MEMBRANE

    Institute of Scientific and Technical Information of China (English)

    LI Xuefen; LI Zhifen; CHEN Chuanfu; WU Wenhui

    1990-01-01

    Surface modification of nucle-microporous membrane by plasma polymerization of HEMA, NVP and D4 has been studied. The hydrophilicity of membranes was increased with increasing of plasma polymerization time of hydrophilic monomers HEMA and NVP. The flow rate of water through the membrane was increased remarkably after plasma polymerization of HEMA on it.

  3. Effects of freezing and cold acclimation on the plasma membrane of isolated protoplasts. Progress report, May 16, 1992--January 9, 1993

    Energy Technology Data Exchange (ETDEWEB)

    Steponkus, P.L.

    1993-05-01

    Our goal is to provide a mechanistic understanding of the cellular and molecular aspects of freezing injury and cold acclimation from a perspective of the structural and functional integrity of the plasma membrane -- the primary site of freezing injury in winter cereals. We have utilized protoplasts isolated from leaves of winter rye (Secale cereale L. cv Puma) to study the cryobehavior of the plasma membrane during a freeze/thaw cycle. The focus of our current studies is on lesions in the plasma membrane that result from severe freeze-induced dehydration and result in the alteration of the semipermeable characteristics of the plasma membrane so that the protoplasts are osmotically unresponsive. In protoplasts isolated from non-acclimated rye leaves (NA protoplasts), injury is associated with the formation of aparticulate domains in the plasma membrane, aparticulate lamellae subtending the plasma membrane, and lamellar-to-hexagonal II phase transitions in the plasma membrane and the subtending lamellae. However, lamellar-to-hexagonal II phase transitions are not observed following severe dehydration of protoplasts isolated from cold-acclimated rye leaves (ACC protoplasts). Rather, injury is associated with the ``fracture-jump lesion,`` which, in freeze-fracture electron microscopy studies, is manifested as localized deviations in the fracture face of the plasma membrane. The fracture plane ``jumps`` from the plasma membrane to either subtending aparticulate lamellae or aparticulate regions of various endomembranes (predominantly chloroplast envelopes) that are in close apposition with the plasma membrane.

  4. 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.

  5. 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.

  6. 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.

  7. 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.

  8. Autoinhibitory Regulation of Plasma Membrane H+-ATPases

    DEFF Research Database (Denmark)

    Pedersen, Jesper Torbøl

    Electrochemical gradients across cell membranes are essential for nutrient uptake. In plant and fungal cells the electrochemical gradient across the plasma membrane (PM) can build much higher than in mammalian cells. The protein responsible for this gradient is the essential PM H+-ATPase that uses...... in mammalian cells and it has been speculated if they have a similar function in plants. In this thesis we show, that plant PM H+-ATPases are receptors for lysophospholipids and the autoinhibitory terminal inhibition is released upon lysophospholipid binding. Finally, we have used a group of stabilizing...

  9. Modulation of Erythrocyte Plasma Membrane Redox System Activity by Curcumin

    Directory of Open Access Journals (Sweden)

    Prabhakar Singh

    2016-01-01

    Full Text Available Plasma membrane redox system (PMRS is an electron transport chain system ubiquitously present throughout all cell types. It transfers electron from intracellular substrates to extracellular acceptors for regulation of redox status. Curcumin, isolated from Curcuma longa, has modulatory effects on cellular physiology due to its membrane interaction ability and antioxidant potential. The present study investigates the effect of curcumin on PMRS activity of erythrocytes isolated from Wistar rats in vitro and in vivo and validated through an in silico docking simulation study using Molegro Virtual Docker (MVD. Effects of curcumin were also evaluated on level of glutathione (GSH and the oxidant potential of plasma measured in terms of plasma ferric equivalent oxidative potentials (PFEOP. Results show that curcumin significantly (p<0.01 downregulated the PMRS activity in a dose-dependent manner. Molecular docking results suggest that curcumin interacts with amino acids at the active site cavity of cytochrome b5 reductase, a key constituent of PMRS. Curcumin also increased the GSH level in erythrocytes and plasma while simultaneously decreasing the oxidant potential (PFEOP of plasma. Altered PMRS activity and redox status are associated with the pathophysiology of several health complications including aging and diabetes; hence, the above finding may explain part of the role of curcumin in health beneficial effects.

  10. 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).

  11. Plant membranes a biophysical approach to structure, development and senescence

    CERN Document Server

    Leshem, Ya’Acov Y

    1992-01-01

    The plasma membrane is at once the window through which the cell senses the environment and the portal through which the environment influences the structure and activities of the cell. Its importance in cellular physiology can thus hardly be overestimated, since constant flow of materials between cell and environment is essential to the well-being of any biological system. The nature of the materials mov­ ing into the cell is also critical, since some substances are required for maintenance and growth, while others, because of their toxicity, must either be rigorously excluded or permitted to enter only after chemical alteration. Such alteration frequently permits the compounds to be sequestered in special cellular compartments having different types of membranes. This type of homogeneity, plus the fact that the wear and tear of transmembrane molecular traffic compels the system to be constantly monitored and repaired, means that the membrane system of any organism must be both structurally complex and dy­...

  12. A membrane-proximal, C-terminal α-helix is required for plasma membrane localization and function of the G Protein-coupled receptor (GPCR) TGR5.

    Science.gov (United States)

    Spomer, Lina; Gertzen, Christoph G W; Schmitz, Birte; Häussinger, Dieter; Gohlke, Holger; Keitel, Verena

    2014-02-07

    The C terminus of G protein-coupled receptors (GPCRs) is important for G protein-coupling and activation; in addition, sorting motifs have been identified in the C termini of several GPCRs that facilitate correct trafficking from the endoplasmic reticulum to the plasma membrane. The C terminus of the GPCR TGR5 lacks any known sorting motif such that other factors must determine its trafficking. Here, we investigate deletion and substitution variants of the membrane-proximal C terminus of TGR5 with respect to plasma membrane localization and function using immunofluorescence staining, flow cytometry, and luciferase assays. Peptides of the membrane-proximal C-terminal variants are subjected to molecular dynamics simulations and analyzed with respect to their secondary structure. Our results reveal that TGR5 plasma membrane localization and responsiveness to extracellular ligands is fostered by a long (≥ 9 residues) α-helical stretch at the C terminus, whereas the presence of β-strands or only a short α-helical stretch leads to retention in the endoplasmic reticulum and a loss of function. As a proof-of-principle, chimeras of TGR5 containing the membrane-proximal amino acids of the β2 adrenergic receptor (β2AR), the sphingosine 1-phosphate receptor-1 (S1P1), or the κ-type opioid receptor (κOR) were generated. These TGR5β2AR, TGR5S1P1, or TGR5κOR chimeras were correctly sorted to the plasma membrane. As the exchanged amino acids of the β2AR, the S1P1, or the κOR form α-helices in crystal structures but lack significant sequence identity to the respective TGR5 sequence, we conclude that the secondary structure of the TGR5 membrane-proximal C terminus is the determining factor for plasma membrane localization and responsiveness towards extracellular ligands.

  13. Measuring distances between TRPV1 and the plasma membrane using a noncanonical amino acid and transition metal ion FRET

    OpenAIRE

    Zagotta, William N.; Gordon, Moshe T.; Senning, Eric N.; Munari, Mika A.; Gordon, Sharona E.

    2016-01-01

    Despite recent advances, the structure and dynamics of membrane proteins in cell membranes remain elusive. We implemented transition metal ion fluorescence resonance energy transfer (tmFRET) to measure distances between sites on the N-terminal ankyrin repeat domains (ARDs) of the pain-transducing ion channel TRPV1 and the intracellular surface of the plasma membrane. To preserve the native context, we used unroofed cells, and to specifically label sites in TRPV1, we incorporated a fluorescent...

  14. Glycine transporter dimers: evidence for occurrence in the plasma membrane

    DEFF Research Database (Denmark)

    Bartholomäus, Ingo; Milan-Lobo, Laura; Nicke, Annette

    2008-01-01

    membrane based on hydrodynamic and native gel electrophoretic studies. Here, we used cysteine substitution and oxidative cross-linking to show that of GlyT1 and GlyT2 also form dimeric complexes within the plasma membrane. GlyT oligomerization at the cell surface was confirmed for both GlyT1 and GlyT2......Different Na(+)/Cl(-)-dependent neurotransmitter transporters of the SLC6a family have been shown to form dimers or oligomers in both intracellular compartments and at the cell surface. In contrast, the glycine transporters (GlyTs) GlyT1 and -2 have been reported to exist as monomers in the plasma...

  15. Structures of photosynthetic membrane complexes

    OpenAIRE

    Semchonok, Dmitry Alexandrovich

    2016-01-01

    Photosynthesis is essential to all life on Earth. It is the biological process that captures energy of sunlight and converts it into chemical compounds usable by any living organism. The processes occurring within photosynthesis can be divided into the light-dependent reactions and the light-independent reactions. In my PhD thesis several integral membrane protein complexes were investigated that catalyze the light dependent reactions. Primarily the photosystem I and II that are taking part i...

  16. Neobiosynthesis of glycosphingolipids by plasma membrane-associated glycosyltransferases.

    Science.gov (United States)

    Crespo, Pilar M; Demichelis, Vanina Torres; Daniotti, José L

    2010-09-17

    Gangliosides, complex glycosphingolipids containing sialic acids, are synthesized in the endoplasmic reticulum and in the Golgi complex. These neobiosynthesized gangliosides move via vesicular transport to the plasma membrane, becoming components of the external leaflet. Gangliosides can undergo endocytosis followed by recycling to the cell surface or sorting to the Golgi complex or lysosomes for remodeling and catabolism. Recently, glycosphingolipid catabolic enzymes (glycohydrolases) have been found to be associated with the plasma membrane, where they display activity on the membrane components. In this work, we demonstrated that ecto-ganglioside glycosyltransferases may catalyze ganglioside synthesis outside the Golgi compartment, particularly at the cell surface. Specifically, we report the first direct evidence of expression and activity of CMP-NeuAc:GM3 sialyltransferase (Sial-T2) at the cell surface of epithelial and melanoma cells, with membrane-integrated ecto-Sial-T2 being able to sialylate endogenously synthesized GM3 ganglioside as well as exogenously incorporated substrate. Interestingly, we also showed that ecto-Sial-T2 was able to synthesize GD3 ganglioside at the cell surface using the endogenously synthesized cytidine monophospho-N-acetylneuraminic acid (CMP-NeuAc) available at the extracellular milieu. In addition, the expression of UDP-GalNAc:LacCer/GM3/GD3 N-acetylgalactosaminyltransferase (GalNAc-T) was also detected at the cell surface of epithelial cells, whose catalytic activity was only observed after feeding the cells with exogenous GM3 substrate. Thus, the relative interplay between the plasma membrane-associated glycosyltransferase and glycohydrolase activities, even when acting on a common substrate, emerges as a potential level of regulation of the local glycosphingolipid composition in response to different external and internal stimuli.

  17. 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.

  18. The apical plasma membrane of chitin-synthesizing epithelia

    Institute of Scientific and Technical Information of China (English)

    Bernard Moussian

    2013-01-01

    Chitin is the second most abundant polysaccharide on earth.It is produced at the apical side of epidermal,tracheal,fore-,and hindgut epithelial cells in insects as a central component of the protective and supporting extracellular cuticle.Chitin is also an important constituent of the midgut peritrophic matrix that encases the food supporting its digestion and protects the epithelium against invasion by possibly ingested pathogens.The enzyme producing chitin is a glycosyltransferase that resides in the apical plasma membrane forming a pore to extrude the chains of chitin into the extracellular space.The apical plasma membrane is not only a platform for chitin synthases but,probably through its shape and equipment with distinct factors,also plays an important role in orienting and organizing chitin fibers.Here,I review findings on the cellular and molecular constitution of the apical plasma membrane of chitin-producing epithelia mainly focusing on work done in the fruit fly Drosophila melanogaster.

  19. 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.

  20. Inside job: ligand-receptor pharmacology beneath the plasma membrane.

    Science.gov (United States)

    Babcock, Joseph J; Li, Min

    2013-07-01

    Most drugs acting on the cell surface receptors are membrane permeable and thus able to engage their target proteins in different subcellular compartments. However, these drugs' effects on cell surface receptors have historically been studied on the plasma membrane alone. Increasing evidence suggests that small molecules may also modulate their targeted receptors through membrane trafficking or organelle-localized signaling inside the cell. These additional modes of interaction have been reported for functionally diverse ligands of GPCRs, ion channels, and transporters. Such intracellular drug-target engagements affect cell surface expression. Concurrent intracellular and cell surface signaling may also increase the complexity and therapeutic opportunities of small molecule modulation. Here we discuss examples of ligand-receptor interactions that are present in both intra- and extracellular sites, and the potential therapeutic opportunities presented by this phenomenon.

  1. Inside job: ligand-receptor pharmacology beneath the plasma membrane

    Institute of Scientific and Technical Information of China (English)

    Joseph J BABCOCK; Min LI

    2013-01-01

    Most drugs acting on the cell surface receptors are membrane permeable and thus able to engage their target proteins in different subcellular compartments.However,these drugs' effects on cell surface receptors have historically been studied on the plasma membrane alone.Increasing evidence suggests that small molecules may also modulate their targeted receptors through membrane trafficking or organelle-localized signaling inside the cell.These additional modes of interaction have been reported for functionally diverse ligands of GPCRs,ion channels,and transporters.Such intracellular drug-target engagements affect cell surface expression.Concurrent intracellular and cell surface signaling may also increase the complexity and therapeutic opportunities of small molecule modulation.Here we discuss examples of ligand-receptor interactions that are present in both intra- and extracellular sites,and the potential therapeutic opportunities presented by this phenomenon.

  2. Superhydrophilic poly(L-lactic acid) electrospun membranes for biomedical applications obtained by argon and oxygen plasma treatment

    Science.gov (United States)

    Correia, D. M.; Ribeiro, C.; Botelho, G.; Borges, J.; Lopes, C.; Vaz, F.; Carabineiro, S. A. C.; Machado, A. V.; Lanceros-Méndez, S.

    2016-05-01

    Poly(L-lactic acid), PLLA, electrospun membranes and films were plasma treated at different times and power with argon (Ar) and oxygen (O2), independently, in order to modify the hydrophobic nature of the PLLA membranes. Both Ar and O2 plasma treatments promote an increase in fiber average size of the electrospun membranes from 830 ± 282 nm to 866 ± 361 and 1179 ± 397 nm, respectively, for the maximum exposure time (970 s) and power (100 W). No influence of plasma treatment was detected in the physical-chemical characteristics of PLLA, such as chemical structure, polymer phase or degree of crystallinity. On the other hand, an increase in the roughness of the films was obtained both with argon and oxygen plasma treatments. Surface wettability studies revealed a decrease in the contact angle with increasing plasma treatment time for a given power and with increasing power for a given time in membranes and films and superhydrophilic electrospun fiber membranes were obtained. Results showed that the argon and oxygen plasma treatments can be used to tailor hydrophilicity of PLLA membranes for biomedical applications. MTT assay results indicated that plasma treatments under Ar and O2 do not influence the metabolic activity of MC3T3-E1 pre-osteoblast cells.

  3. 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

  4. Plasma membrane domains enriched in cortical endoplasmic reticulum function as membrane protein trafficking hubs.

    Science.gov (United States)

    Fox, Philip D; Haberkorn, Christopher J; Weigel, Aubrey V; Higgins, Jenny L; Akin, Elizabeth J; Kennedy, Matthew J; Krapf, Diego; Tamkun, Michael M

    2013-09-01

    In mammalian cells, the cortical endoplasmic reticulum (cER) is a network of tubules and cisterns that lie in close apposition to the plasma membrane (PM). We provide evidence that PM domains enriched in underlying cER function as trafficking hubs for insertion and removal of PM proteins in HEK 293 cells. By simultaneously visualizing cER and various transmembrane protein cargoes with total internal reflectance fluorescence microscopy, we demonstrate that the majority of exocytotic delivery events for a recycled membrane protein or for a membrane protein being delivered to the PM for the first time occur at regions enriched in cER. Likewise, we observed recurring clathrin clusters and functional endocytosis of PM proteins preferentially at the cER-enriched regions. Thus the cER network serves to organize the molecular machinery for both insertion and removal of cell surface proteins, highlighting a novel role for these unique cellular microdomains in membrane trafficking.

  5. 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.

  6. Elisidepsin Interacts Directly with Glycosylceramides in the Plasma Membrane of Tumor Cells to Induce Necrotic Cell Death

    Science.gov (United States)

    Molina-Guijarro, José Manuel; García, Carolina; Macías, Álvaro; García-Fernández, Luis Francisco; Moreno, Cristina; Reyes, Fernando; Martínez-Leal, Juan Fernando; Fernández, Rogelio; Martínez, Valentín; Valenzuela, Carmen; Lillo, M. Pilar; Galmarini, Carlos M.

    2015-01-01

    Plasma membrane integrity is essential for cell life. Any major break on it immediately induces the death of the affected cell. Different molecules were described as disrupting this cell structure and thus showing antitumor activity. We have previously defined that elisidepsin (Irvalec®, PM02734) inserts and self-organizes in the plasma membrane of tumor cells, inducing a rapid loss of membrane integrity, cell permeabilization and necrotic death. Here we show that, in sensitive HCT-116 colorectal cells, all these effects are consequence of the interaction of elisidepsin with glycosylceramides in the cell membrane. Of note, an elisidepsin-resistant subline (HCT-116-Irv) presented reduced levels of glycosylceramides and no accumulation of elisidepsin in the plasma membrane. Consequently, drug treatment did not induce the characteristic necrotic cell death. Furthermore, GM95, a mutant derivative from B16 mouse melanoma cells lacking ceramide glucosyltransferase (UGCG) activity and thus the synthesis of glycosylceramides, was also resistant to elisidepsin. Over-expression of UGCG gene in these deficient cells restored glycosylceramides synthesis, rendering them sensitive to elisidepsin, at a similar level than parental B16 cells. These results indicate that glycosylceramides act as membrane targets of elisidepsin, facilitating its insertion in the plasma membrane and the subsequent membrane permeabilization that leads to drug-induced cell death. They also indicate that cell membrane lipids are a plausible target for antineoplastic therapy. PMID:26474061

  7. Elisidepsin Interacts Directly with Glycosylceramides in the Plasma Membrane of Tumor Cells to Induce Necrotic Cell Death.

    Directory of Open Access Journals (Sweden)

    José Manuel Molina-Guijarro

    Full Text Available Plasma membrane integrity is essential for cell life. Any major break on it immediately induces the death of the affected cell. Different molecules were described as disrupting this cell structure and thus showing antitumor activity. We have previously defined that elisidepsin (Irvalec®, PM02734 inserts and self-organizes in the plasma membrane of tumor cells, inducing a rapid loss of membrane integrity, cell permeabilization and necrotic death. Here we show that, in sensitive HCT-116 colorectal cells, all these effects are consequence of the interaction of elisidepsin with glycosylceramides in the cell membrane. Of note, an elisidepsin-resistant subline (HCT-116-Irv presented reduced levels of glycosylceramides and no accumulation of elisidepsin in the plasma membrane. Consequently, drug treatment did not induce the characteristic necrotic cell death. Furthermore, GM95, a mutant derivative from B16 mouse melanoma cells lacking ceramide glucosyltransferase (UGCG activity and thus the synthesis of glycosylceramides, was also resistant to elisidepsin. Over-expression of UGCG gene in these deficient cells restored glycosylceramides synthesis, rendering them sensitive to elisidepsin, at a similar level than parental B16 cells. These results indicate that glycosylceramides act as membrane targets of elisidepsin, facilitating its insertion in the plasma membrane and the subsequent membrane permeabilization that leads to drug-induced cell death. They also indicate that cell membrane lipids are a plausible target for antineoplastic therapy.

  8. A complex of antioxidant vitamins effectively inhibits free-radical oxidation of LDL phospholipids in blood plasma and membrane structures of the liver and myocardium.

    Science.gov (United States)

    Konovalova, G G; Lisina, M O; Tikhaze, A K; Lankin, V Z

    2003-02-01

    Antioxidant effect of a complex preparation including antioxidant vitamins C, E, provitamin A and selenium was studied on the model of Cu(2+)-initiated free-radical oxidation of LDL isolated from human blood plasma. The antioxidant effect of combined administration of alpha-tocopherol+ascorbic acid and alpha-tocopherol+beta-carotene is far more pronounced that the antioxidant effect of individual components of these cocktails. Moreover, in the model system the combined action of all antioxidant components completely inhibited free-radical oxidation of LDL. A 30-day course of peroral administration of antioxidant vitamin cocktail and selenium to rats pronouncedly enhanced the antioxidant potential of liver and completely suppressed free-radical processes in the myocardium. It is suggested that preparations containing antioxidant vitamins and selenium can be perspective for prevention and complex therapy of atherosclerosis.

  9. A role for eisosomes in maintenance of plasma membrane phosphoinositide levels

    OpenAIRE

    Fröhlich, Florian; Christiano, Romain; Olson, Daniel K.; Alcazar-Roman, Abel; DeCamilli, Pietro; Walther, Tobias C

    2014-01-01

    The plasma membrane delineates the cell and mediates its communication and material exchange with the environment. Many processes of the plasma membrane occur through interactions of proteins with phosphatidylinositol(4,5)-bisphosphate (PI(4,5)P2), which is highly enriched in this membrane and is a key determinant of its identity. Eisosomes function in lateral organization of the plasma membrane, but the molecular function of their major protein subunits, the BAR domain–containing proteins Pi...

  10. Optimizing internal structure of membrane filters

    Science.gov (United States)

    Cummings, Linda; Sanaei, Pejman

    2016-11-01

    Membrane filters are in widespread use, and manufacturers have considerable interest in improving their performance, in terms of particle retention properties, and total throughput over the filter lifetime. In this regard, it has long been known that membrane properties should not be uniform over the membrane depth; rather, membrane permeability should decrease in the direction of flow. While much research effort has been focused on investigating favorable membrane permeability gradients, this work has been largely empirical in nature. We present a simple, first-principles model for flow through and fouling of a membrane filter, accounting for permeability gradients via variable pore size. Our model accounts for two fouling modes: sieving; and particle adsorption within pores. For filtration driven by a fixed pressure drop, flux through the membrane eventually goes to zero, as fouling occurs and pores close. We address issues of filter performance as the internal pore structure is varied, by comparing the total throughput obtained with equal-resistance membranes. Within certain classes of pore profiles we are able to find the optimum pore profile that maximizes total throughput over the filter lifetime, while maintaining acceptable particle removal from the feed. Partial support from NSF DMS 1261596 is gratefully acknowledged.

  11. 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.

  12. Water/O2-plasma-assisted treatment of PCL membranes for biosignal immobilization.

    Science.gov (United States)

    Saşmazel, Hilal Türkoğlu; Manolache, Sorin; Gümüşderelioğlu, Menemşe

    2009-01-01

    The main purpose of this study was to obtain COOH functionalities on the surface of poly-epsilon-caprolactone (PCL) membranes using low-pressure water/O(2)-plasma-assisted treatment. PCL membranes were prepared using the solvent-casting technique. Then, low-pressure water/O(2) plasma treatments were performed in a cylindrical, capacitively coupled RF-plasma-reactor in three steps: H(2)O/O(2)-plasma treatment; in situ (oxalyl chloride vapors) gas/solid reaction to convert -OH functionalities into -COCl groups; and hydrolysis for final -COOH functionalities. Optimization of plasma modification processes was done using the DoE software program. COOH and OH functionalities on modified surfaces were detected quantitatively using the fluorescent labeling technique and an UVX 300G sensor. Chemical structural information of untreated, plasma treated and oxalyl chloride functionalized PCL membranes were acquired using pyrolysis GC/MS and ESCA analysis. High-resolution AFM images revealed that nanopatterns were more affected than micropatterns by plasma treatments. AFM images recorded with amino-functionalized tips presented increased size of the features on the surface that suggests higher density of the carboxyls on the nanotopographical elements. Low-pressure water/O(2)-plasma-treated and oxalyl chloride functionalized samples were biologically activated with insulin and/or heparin biosignal molecules using a PEO (polyoxyethylene bis amine) spacer. The success of the immobilization process was checked qualitatively by ESCA analysis. In addition, fluorescent labeling techniques were used for the quantitative determination of immobilized biomolecules. Cell-culture experiments indicated that biomolecule immobilization onto PCL scaffolds was effective on L929 cell adhesion and proliferation, especially in the presence of heparin.

  13. From the endoplasmic reticulum to the plasma membrane: mechanisms of CFTR folding and trafficking.

    Science.gov (United States)

    Farinha, Carlos M; Canato, Sara

    2017-01-01

    CFTR biogenesis starts with its co-translational insertion into the membrane of endoplasmic reticulum and folding of the cytosolic domains, towards the acquisition of a fully folded compact native structure. Efficiency of this process is assessed by the ER quality control system that allows the exit of folded proteins but targets unfolded/misfolded CFTR to degradation. If allowed to leave the ER, CFTR is modified at the Golgi and reaches the post-Golgi compartments to be delivered to the plasma membrane where it functions as a cAMP- and phosphorylation-regulated chloride/bicarbonate channel. CFTR residence at the membrane is a balance of membrane delivery, endocytosis, and recycling. Several adaptors, motor, and scaffold proteins contribute to the regulation of CFTR stability and are involved in continuously assessing its structure through peripheral quality control systems. Regulation of CFTR biogenesis and traffic (and its dysregulation by mutations, such as the most common F508del) determine its overall activity and thus contribute to the fine modulation of chloride secretion and hydration of epithelial surfaces. This review covers old and recent knowledge on CFTR folding and trafficking from its synthesis to the regulation of its stability at the plasma membrane and highlights how several of these steps can be modulated to promote the rescue of mutant CFTR.

  14. 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.

  15. 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.

  16. Plasma membrane mechanical stress activates TRPC5 channels.

    Directory of Open Access Journals (Sweden)

    Bing Shen

    Full Text Available Mechanical forces exerted on cells impose stress on the plasma membrane. Cells sense this stress and elicit a mechanoelectric transduction cascade that initiates compensatory mechanisms. Mechanosensitive ion channels in the plasma membrane are responsible for transducing the mechanical signals to electrical signals. However, the mechanisms underlying channel activation in response to mechanical stress remain incompletely understood. Transient Receptor Potential (TRP channels serve essential functions in several sensory modalities. These channels can also participate in mechanotransduction by either being autonomously sensitive to mechanical perturbation or by coupling to other mechanosensory components of the cell. Here, we investigated the response of a TRP family member, TRPC5, to mechanical stress. Hypoosmolarity triggers Ca2+ influx and cationic conductance through TRPC5. Importantly, for the first time we were able to record the stretch-activated TRPC5 current at single-channel level. The activation threshold for TRPC5 was found to be 240 mOsm for hypoosmotic stress and between -20 and -40 mmHg for pressure applied to membrane patch. In addition, we found that disruption of actin filaments suppresses TRPC5 response to hypoosmotic stress and patch pipette pressure, but does not prevent the activation of TRPC5 by stretch-independent mechanisms, indicating that actin cytoskeleton is an essential transduction component that confers mechanosensitivity to TRPC5. In summary, our findings establish that TRPC5 can be activated at the single-channel level when mechanical stress on the cell reaches a certain threshold.

  17. 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.

  18. Surface monofunctionalized polymethyl pentene hollow fiber membranes by plasma treatment and hemocompatibility modification for membrane oxygenators

    Science.gov (United States)

    Huang, Xin; Wang, Weiping; Zheng, Zhi; Fan, Wenling; Mao, Chun; Shi, Jialiang; Li, Lei

    2016-01-01

    The hemocompatibility of polymethyl pentene (PMP) hollow fiber membranes (HFMs) was improved through surface modification for membrane oxygenator applications. The modification was performed stepwise with the following: (1) oxygen plasma treatment, (2) functionalization of monosort hydroxyl groups through NaBH4 reduction, and (3) grafting 2-methacryloyloxyethyl phosphorylcholine (MPC) or heparin. SEM, ATR-FTIR, and XPS analyses were conducted to confirm successful grafting during the modification. The hemocompatibility of PMP HFMs was analyzed and compared through protein adsorption, platelet adhesion, and coagulation tests. Pure CO2 and O2 permeation rates, as well as in vitro gas exchange rates, were determined to evaluate the mass transfer properties of PMP HFMs. SEM results showed that different nanofibril topographies were introduced on the HFM surface. ATR-FTIR and XPS spectra indicated the presence of functionalization of monosort hydroxyl group and the grafting of MPC and heparin. Hemocompatibility evaluation results showed that the modified PMP HFMs presented optimal hemocompatibility compared with pristine HFMs. Gas permeation results revealed that gas permeation flux increased in the modified HFMs because of dense surface etching during the plasma treatment. The results of in vitro gas exchange rates showed that all modified PMP HFMs presented decreased gas exchange rates because of potential surface fluid wetting. The proposed strategy exhibits a potential for fabricating membrane oxygenators for biomedical applications to prevent coagulation formation and alter plasma-induced surface topology and composition.

  19. Structure Biology of Membrane Bound Enzymes

    Energy Technology Data Exchange (ETDEWEB)

    Fu, Dax [Johns Hopkins Univ., Baltimore, MD (United States). School of Medicine. Dept. of Physiology

    2016-11-30

    The overall goal of the proposed research is to understand the membrane-associated active processes catalyzed by an alkane $\\square$-hydroxylase (AlkB) from eubacterium Pseudomonase oleovorans. AlkB performs oxygenation of unactivated hydrocarbons found in crude oils. The enzymatic reaction involves energy-demanding steps in the membrane with the uses of structurally unknown metal active sites featuring a diiron [FeFe] center. At present, a critical barrier to understanding the membrane-associated reaction mechanism is the lack of structural information. The structural biology efforts have been challenged by technical difficulties commonly encountered in crystallization and structural determination of membrane proteins. The specific aims of the current budget cycle are to crystalize AlkB and initiate X-ray analysis to set the stage for structural determination. The long-term goals of our structural biology efforts are to provide an atomic description of AlkB structure, and to uncover the mechanisms of selective modification of hydrocarbons. The structural information will help elucidating how the unactivated C-H bonds of saturated hydrocarbons are oxidized to initiate biodegradation and biotransformation processes. The knowledge gained will be fundamental to biotechnological applications to biofuel transformation of non-edible oil feedstock. Renewable biodiesel is a promising energy carry that can be used to reduce fossil fuel dependency. The proposed research capitalizes on prior BES-supported efforts on over-expression and purification of AlkB to explore the inner workings of a bioenergy-relevant membrane-bound enzyme.

  20. Solid polymer electrolyte composite membrane comprising plasma etched porous support

    Science.gov (United States)

    Liu, Han; LaConti, Anthony B.

    2010-10-05

    A solid polymer electrolyte composite membrane and method of manufacturing the same. According to one embodiment, the composite membrane comprises a rigid, non-electrically-conducting support, the support preferably being a sheet of polyimide having a thickness of about 7.5 to 15 microns. The support has a plurality of cylindrical pores extending perpendicularly between opposing top and bottom surfaces of the support. The pores, which preferably have a diameter of about 0.1 to 5 microns, are made by plasma etching and preferably are arranged in a defined pattern, for example, with fewer pores located in areas of high membrane stress and more pores located in areas of low membrane stress. The pores are filled with a first solid polymer electrolyte, such as a perfluorosulfonic acid (PFSA) polymer. A second solid polymer electrolyte, which may be the same as or different than the first solid polymer electrolyte, may be deposited over the top and/or bottom of the first solid polymer electrolyte.

  1. Plasma membrane repair: the adaptable cell life-insurance.

    Science.gov (United States)

    Jimenez, Ana Joaquina; Perez, Franck

    2017-08-01

    The plasma membrane is the most basic element necessary for the cell to exist and be distinguishable from its environment. Regulated mechanisms allow tightly controlled communication between intacellular and extracellular medium allowing the maintenance of a specific biochemical environment, optimized for cellular functions. The anarchic and uncontrolled opening of a hole in the PM induces a change in the concentration of ions and oxidizing agents perturbing homeostasis. Fortunately, the cell possesses mechanisms that are capable of reacting to sudden extracellular medium entry and to block the leakage locally. Here we summarize the known mechanisms of membrane repair and how the size of the wound and the resulting calcium entry activates preferentially one or another mechanism adapted to the magnitude of the injury. Copyright © 2017 Elsevier Ltd. All rights reserved.

  2. 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.

  3. STUDIES ON THE PERMEABILITY OF PVC /EBBA OVERLAPPED ULTRATHIN COMPOSITE MEMBRANES MODIFIED BY PLASMA- POLYMERIZATION WITH FLUOROCARBON MONOMERS

    Institute of Scientific and Technical Information of China (English)

    FU Xiucheng; JIN Xigao; Tisato KAJIYAMA

    1989-01-01

    The PVC/EBBA ultrathin composite membranes with thickness of about 100 nm were prepared by spreading the solution on water surface. The overlapped composite membrane showed a characteristic aggregation structure in which the polymer matrix exists as a three-dimensional spongy network and the liquid crystal domains were observedThe surface modification for the overlapped membranes was carried out by means of plasma-polymerization with the monomers of fluorocarbon compounds. Both Arrhenius plots of permeability coefficients for oxygen (-Po2) in the membrane samples before and after modification showed significant increase in the vicinity of the TKN of EBBA.

  4. [Radiation-induced changes in structural state of membranes of human blood cells].

    Science.gov (United States)

    Burlakova, E B; Atkarskaia, M V; Fatkullina, L D; Andreev, S G

    2014-01-01

    To evaluate radiation-induced changes in the structural state of the membranes, blood samples of healthy donors were subjected to gamma radiation in the range of small (1-10 cGy) and medium doses (50 cGy-2 Gy). After irradiation, the microviscosity of lipid membranes of red and white blood cells was measured by ESR spin probe method. At doses exceeding 1 cGy, statistically significant changes of the degree of spontaneous erythrocyte hemolysis and of the lymphocyte plasma membrane microviscosity were observed. Under identical irradiation conditions, the stability of lymphocyte membranes was less as compared to erythrocyte membranes.

  5. Characterization of Plasma Membrane Proteins from Ovarian Cancer Cells Using Mass Spectrometry

    Directory of Open Access Journals (Sweden)

    David L. Springer

    2004-01-01

    Full Text Available To determine how the repertoire of plasma membrane proteins change with disease state, specifically related to cancer, several methods for preparation of plasma membrane proteins were evaluated. Cultured cells derived from stage IV ovarian tumors were grown to 90% confluence and harvested in buffer containing CHAPS detergent. This preparation was centrifuged at low speed to remove insoluble cellular debris resulting in a crude homogenate. Glycosylated proteins in the crude homogenate were selectively enriched using lectin affinity chromatography. The crude homogenate and the lectin purified sample were prepared for mass spectrometric evaluation. The general procedure for protein identification began with trypsin digestion of protein fractions followed by separation by reversed phase liquid chromatography that was coupled directly to a conventional tandem mass spectrometer (i.e. LCQ ion trap. Mass and fragmentation data for the peptides were searched against a human proteome data base using the informatics program SEQUEST. Using this procedure 398 proteins were identified with high confidence, including receptors, membrane-associated ligands, proteases, phosphatases, as well as structural and adhesion proteins. Results indicate that lectin chromatography provides a select subset of proteins and that the number and quality of the identifications improve as does the confidence of the protein identifications for this subset. These results represent the first step in development of methods to separate and successfully identify plasma membrane proteins from advanced ovarian cancer cells. Further characterization of plasma membrane proteins will contribute to our understanding of the mechanisms underlying progression of this deadly disease and may lead to new targeted interventions as well as new biomarkers for diagnosis.

  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. Measuring distances between TRPV1 and the plasma membrane using a noncanonical amino acid and transition metal ion FRET.

    Science.gov (United States)

    Zagotta, William N; Gordon, Moshe T; Senning, Eric N; Munari, Mika A; Gordon, Sharona E

    2016-02-01

    Despite recent advances, the structure and dynamics of membrane proteins in cell membranes remain elusive. We implemented transition metal ion fluorescence resonance energy transfer (tmFRET) to measure distances between sites on the N-terminal ankyrin repeat domains (ARDs) of the pain-transducing ion channel TRPV1 and the intracellular surface of the plasma membrane. To preserve the native context, we used unroofed cells, and to specifically label sites in TRPV1, we incorporated a fluorescent, noncanonical amino acid, L-ANAP. A metal chelating lipid was used to decorate the plasma membrane with high-density/high-affinity metal-binding sites. The fluorescence resonance energy transfer (FRET) efficiencies between L-ANAP in TRPV1 and Co(2+) bound to the plasma membrane were consistent with the arrangement of the ARDs in recent cryoelectron microscopy structures of TRPV1. No change in tmFRET was observed with the TRPV1 agonist capsaicin. These results demonstrate the power of tmFRET for measuring structure and rearrangements of membrane proteins relative to the cell membrane.

  8. Effects of freezing and cold acclimation on the plasma membrane of isolated protoplasts

    Energy Technology Data Exchange (ETDEWEB)

    Steponkus, P.L.

    1991-01-01

    This project focuses on lesions in the plasma membrane of protoplasts that occur during freezing to temperatures below {minus}5{degrees} which result in changes in the semipermeablity of the plasma membrane. This injury, referred to as loss of osmotic responsiveness, is associated with the formation of large, aparticulate domains in the plasma membrane, aparticulate lamellae subtending the plasma membrane, and lamellar-to-hexagonal{sub II} phase transitions in the plasma membrane and subtending lamellar. The goals of this project are to provide a mechanistic understanding of the mechanism by which freeze-induced dehydration effects the formation of aparticulate domains and lamellar-to-hexagonal{sub II} phase transitions and to determine the mechanisms by which cold acclimation and cryoprotectants preclude or diminish these ultrastructural changes. Our working hypothesis is the formation of aparticulate domains and lamellar-to-hexagon{sub II} phase transitions in the plasma membrane and subtending lamellae are manifestations of hydration-dependent bilayer-bilayer interactions.

  9. Specific interaction of postsynaptic densities with membrane rafts isolated from synaptic plasma membranes.

    Science.gov (United States)

    Liu, Qian; Yao, Wei-Dong; Suzuki, Tatsuo

    2013-06-01

    Postsynaptic membrane rafts are believed to play important roles in synaptic signaling, plasticity, and maintenance. We recently demonstrated the presence, at the electron microscopic level, of complexes consisting of membrane rafts and postsynaptic densities (PSDs) in detergent-resistant membranes (DRMs) prepared from synaptic plasma membranes (SPMs) ( Suzuki et al., 2011 , J Neurochem, 119, 64-77). To further explore these complexes, here we investigated the nature of the binding between purified SPM-DRMs and PSDs in vitro. In binding experiments, we used SPM-DRMs prepared after treating SPMs with n-octyl-β-d-glucoside, because at concentrations of 1.0% or higher it completely separates SPM-DRMs and PSDs, providing substantially PSD-free unique SPM-DRMs as well as DRM-free PSDs. PSD binding to PSD-free DRMs was identified by mass spectrometry, Western blotting, and electron microscopy. PSD proteins were not incorporated into SPMs, and significantly less PSD proteins were incorporated into DRMs prepared from liver membranes, providing in vitro evidence that binding of PSDs to DRMs is specific and suggestion of the presence of specific interacting molecules. These specific interactions may have important roles in synaptic development, function, and plasticity in vivo. In addition, the binding system we developed may be a good tool to search for binding molecules and binding mechanisms between PSDs and rafts.

  10. Plasma-induced Styrene Grafting onto the Surface of Polytetrafluoroethylene Powder for Proton Exchange Membrane Application%Plasma-induced Styrene Grafting onto the Surface of Polytetrafluoroethylene Powder for Proton Exchange Membrane Application

    Institute of Scientific and Technical Information of China (English)

    兰彦; 程诚; 张素贞; 倪国华; 陈龙威; 杨光杰; M.NAGATSU; 孟月东

    2011-01-01

    Low-temperature plasma treatment was adopted to graft styrene onto polytetrafluo- roethylene (PTFE) powder, which is widely used in the fabrication of proton exchange membrane (PEM). The grafted PTFE powder was sulfonated in chlorosulfonic acid and fabricated into a membrane, which was used as inexpensive PEM material for a proton exchange membrane fuel cell (PEMFC). Fourier transform infrared spectroscopy attenuated total reflection spectroscopy (FTIR-ATR) and X-ray photoelectron spectroscopy (XPS) analysis were used to characterize the structure of the sulfonated PTFE powder. The results showed that all the PTFE powders were successfully grafted by nitrogen plasma and then sulfonated under such experimental conditions. A scanning electron microscopy (SEM) image indicated that the fabricated membrane exhibits flat morphology and homogenous structure. The ion exchange capacity (IEC) of this kind of PEM was also investigated.

  11. Plasma membrane localization is required for RasA-mediated polarized morphogenesis and virulence of Aspergillus fumigatus.

    Science.gov (United States)

    Fortwendel, Jarrod R; Juvvadi, Praveen R; Rogg, Luise E; Asfaw, Yohannes G; Burns, Kimberlie A; Randell, Scott H; Steinbach, William J

    2012-08-01

    Ras is a highly conserved GTPase protein that is essential for proper polarized morphogenesis of filamentous fungi. Localization of Ras proteins to the plasma membrane and endomembranes through posttranslational addition of farnesyl and palmitoyl residues is an important mechanism through which cells provide specificity to Ras signal output. Although the Aspergillus fumigatus RasA protein is known to be a major regulator of growth and development, the membrane distribution of RasA during polarized morphogenesis and the role of properly localized Ras signaling in virulence of a pathogenic mold remain unknown. Here we demonstrate that Aspergillus fumigatus RasA localizes primarily to the plasma membrane of actively growing hyphae. We show that treatment with the palmitoylation inhibitor 2-bromopalmitate disrupts normal RasA plasma membrane association and decreases hyphal growth. Targeted mutations of the highly conserved RasA palmitoylation motif also mislocalized RasA from the plasma membrane and led to severe hyphal abnormalities, cell wall structural changes, and reduced virulence in murine invasive aspergillosis. Finally, we provide evidence that proper RasA localization is independent of the Ras palmitoyltransferase homolog, encoded by erfB, but requires the palmitoyltransferase complex subunit, encoded by erfD. Our results demonstrate that plasma membrane-associated RasA is critical for polarized morphogenesis, cell wall stability, and virulence in A. fumigatus.

  12. New insights into the organization of plasma membrane and its role in signal transduction.

    Science.gov (United States)

    Suzuki, Kenichi G N

    2015-01-01

    Plasma membranes have heterogeneous structures for efficient signal transduction, required to perform cell functions. Recent evidence indicates that the heterogeneous structures are produced by (1) compartmentalization by actin-based membrane skeleton, (2) raft domains, (3) receptor-receptor interactions, and (4) the binding of receptors to cytoskeletal proteins. This chapter provides an overview of recent studies on diffusion, clustering, raft association, actin binding, and signal transduction of membrane receptors, especially glycosylphosphatidylinositol (GPI)-anchored receptors. Studies on diffusion of GPI-anchored receptors suggest that rafts may be small and/or short-lived in plasma membranes. In steady state conditions, GPI-anchored receptors form transient homodimers, which may represent the "standby state" for the stable homodimers and oligomers upon ligation. Furthermore, It is proposed that upon ligation, the binding of GPI-anchored receptor clusters to cytoskeletal actin filaments produces a platform for downstream signaling, and that the pulse-like signaling easily maintains the stability of the overall signaling activity.

  13. Regulation of Ras signaling and function by plasma membrane microdomains.

    Science.gov (United States)

    Goldfinger, Lawrence E; Michael, James V

    2017-02-07

    Together H-, N- and KRAS mutations are major contributors to ~30% of all human cancers. Thus, Ras inhibition remains an important anti-cancer strategy. The molecular mechanisms of isotypic Ras oncogenesis are still not completely understood. Monopharmacological therapeutics have not been successful in the clinic. These disappointing outcomes have led to attempts to target elements downstream of Ras, mainly targeting either the Phosphatidylinositol 3-Kinase (PI3K) or Mitogen-Activated Protein Kinase (MAPK) pathways. While several such approaches are moderately effective, recent efforts have focused on preclinical evaluation of combination therapies to improve efficacies. This review will detail current understanding of the contributions of plasma membrane microdomain targeting of Ras to mitogenic and tumorigenic signaling and tumor progression. Moreover, this review will outline novel approaches to target Ras in cancers, including targeting schemes for new drug development, as well as putative re-purposing of drugs in current use to take advantage of blunting Ras signaling by interfering with Ras plasma membrane microdomain targeting and retention.

  14. 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.

  15. MLKL Compromises Plasma Membrane Integrity by Binding to Phosphatidylinositol Phosphates

    Directory of Open Access Journals (Sweden)

    Yves Dondelinger

    2014-05-01

    Full Text Available Although mixed lineage kinase domain-like (MLKL protein has emerged as a specific and crucial protein for necroptosis induction, how MLKL transduces the death signal remains poorly understood. Here, we demonstrate that the full four-helical bundle domain (4HBD in the N-terminal region of MLKL is required and sufficient to induce its oligomerization and trigger cell death. Moreover, we found that a patch of positively charged amino acids on the surface of the 4HBD binds to phosphatidylinositol phosphates (PIPs and allows recruitment of MLKL to the plasma membrane. Importantly, we found that recombinant MLKL, but not a mutant lacking these positive charges, induces leakage of PIP-containing liposomes as potently as BAX, supporting a model in which MLKL induces necroptosis by directly permeabilizing the plasma membrane. Accordingly, we found that inhibiting the formation of PI(5P and PI(4,5P2 specifically inhibits tumor necrosis factor (TNF-mediated necroptosis but not apoptosis.

  16. 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.

  17. Plasma membrane is the site of productive HIV-1 particle assembly.

    Directory of Open Access Journals (Sweden)

    Nolwenn Jouvenet

    2006-12-01

    Full Text Available Recently proposed models that have gained wide acceptance posit that HIV-1 virion morphogenesis is initiated by targeting the major structural protein (Gag to late endosomal membranes. Thereafter, late endosome-based secretory pathways are thought to deliver Gag or assembled virions to the plasma membrane (PM and extracellular milieu. We present several findings that are inconsistent with this model. Specifically, we demonstrate that HIV-1 Gag is delivered to the PM, and virions are efficiently released into the extracellular medium, when late endosome motility is abolished. Furthermore, we show that HIV-1 virions are efficiently released when assembly is rationally targeted to the PM, but not when targeted to late endosomes. Recently synthesized Gag first accumulates and assembles at the PM, but a proportion is subsequently internalized via endocytosis or phagocytosis, thus accounting for observations of endosomal localization. We conclude that HIV-1 assembly is initiated and completed at the PM, and not at endosomal membranes.

  18. Plasma membrane calcium pump regulation by metabolic stress

    Institute of Scientific and Technical Information of China (English)

    Jason; IE; Bruce

    2010-01-01

    The plasma membrane Ca2+-ATPase(PMCA)is an ATPdriven pump that is critical for the maintenance of low resting[Ca2+]i in all eukaryotic cells.Metabolic stress, either due to inhibition of mitochondrial or glycolytic metabolism,has the capacity to cause ATP depletion and thus inhibit PMCA activity.This has potentially fatal consequences,particularly for non-excitable cells in which the PMCA is the major Ca2+efflux pathway.This is because inhibition of the PMCA inevitably leads to cytosolic Ca2+ overload and the consequent cell death.However,the relationship between metabolic stress,ATP depletion and inhibition of the PMCA is not as simple as one would have originally predicted.There is increasing evidence that metabolic stress can lead to the inhibition of PMCA activity independent of ATP or prior to substantial ATP depletion.In particular,there is evidence that the PMCA has its own glycolytic ATP supply that can fuel the PMCA in the face of impaired mitochondrial function.Moreover, membrane phospholipids,mitochondrial membrane potential,caspase/calpain cleavage and oxidative stress have all been implicated in metabolic stress-induced inhibition of the PMCA.The major focus of this review is to challenge the conventional view of ATP-dependent regulation of the PMCA and bring together some of the alternative or additional mechanisms by which metabolic stress impairs PMCA activity resulting in cytosolic Ca2+ overload and cytotoxicity.

  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. High-Throughput Microplate-Based Assay to Monitor Plasma Membrane Wounding and Repair

    Directory of Open Access Journals (Sweden)

    Sarika Pathak-Sharma

    2017-07-01

    Full Text Available The plasma membrane of mammalian cells is susceptible to disruption by mechanical and biochemical damages that frequently occur within tissues. Therefore, efficient and rapid repair of the plasma membrane is essential for maintaining cellular homeostasis and survival. Excessive damage of the plasma membrane and defects in its repair are associated with pathological conditions such as infections, muscular dystrophy, heart failure, diabetes, and lung and neurodegenerative diseases. The molecular events that remodel the plasma membrane during its repair remain poorly understood. In the present work, we report the development of a quantitative high-throughput assay that monitors the efficiency of the plasma membrane repair in real time using a sensitive microplate reader. In this assay, the plasma membrane of living cells is perforated by the bacterial pore-forming toxin listeriolysin O and the integrity and recovery of the membrane are monitored at 37°C by measuring the fluorescence intensity of the membrane impermeant dye propidium iodide. We demonstrate that listeriolysin O causes dose-dependent plasma membrane wounding and activation of the cell repair machinery. This assay was successfully applied to cell types from different origins including epithelial and muscle cells. In conclusion, this high-throughput assay provides a novel opportunity for the discovery of membrane repair effectors and the development of new therapeutic compounds that could target membrane repair in various pathological processes, from degenerative to infectious diseases.

  1. 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.

  2. Plasma membrane and cytoskeleton dynamics during single-cell wound healing.

    Science.gov (United States)

    Boucher, Eric; Mandato, Craig A

    2015-10-01

    Wounding leads not only to plasma membrane disruption, but also to compromised cytoskeleton structures. This results not only in unwarranted exchanges between the cytosol and extracellular milieu, but also in loss of tensegrity, which may further endanger the cell. Tensegrity can be described as the interplay between the tensile forces generated by the apparent membrane tension, actomyosin contraction, and the cytoskeletal structures resisting those changes (e.g., microtubules). It is responsible for the structural integrity of the cell and for its ability to sense mechanical signals. Recent reviews dealing with single-cell healing mostly focused on the molecular machineries controlling the traffic and fusion of specific vesicles, or their role in different pathologies. In this review, we aim to take a broader view of the different modes of single cell repair, while focussing on the different ways the changes in plasmalemma surface area and composition, plasmalemma tension, and cytoskeletal dynamics may influence and affect single-cell repair.

  3. Morphology-properties relationship of gas plasma treated hydrophobic meso-porous membranes and their improved performance for desalination by membrane distillation

    Energy Technology Data Exchange (ETDEWEB)

    Dumée, Ludovic F., E-mail: ludovic.dumee@deakin.edu.au [Deakin University, Geelong Victoria–Australia - Institute for Frontier Materials (Australia); Alglave, Hortense; Chaffraix, Thomas; Lin, Bao; Magniez, Kevin [Deakin University, Geelong Victoria–Australia - Institute for Frontier Materials (Australia); Schütz, Jürg [CSIRO, Manufacturing Flagship, 75 Pigdons Road, 3216 Waurn Ponds, Victoria (Australia)

    2016-02-15

    Graphical abstract: - Highlights: • Systematic surface modifications by gas plasma treatment of hydrophobic polymers. • Correlation between plasma parameters and materials surface energy and morphology. • Spectral analysis of the formation of functional groups across the membranes surface. • Relationship between wettability, roughness and performance. - Abstract: The impact on performance of the surface energy and roughness of membrane materials used for direct contact membrane distillation are critical but yet poorly investigated parameters. The capacity to alter the wettability of highly hydrophobic materials such as poly(tetra-fluoro-ethylene) (PTFE) by gas plasma treatments is reported in this paper. An equally important contribution from this investigation arises from illustrating how vaporized material from the treated sample participates after a short while in the composition of the plasma and fundamentally changes the result of surface chemistry processes. The water contact angle across the hydrophobic membranes is generally controlled by varying the plasma gas conditions, such as the plasma power, chamber pressure and irradiation duration. Changes to surface porosity and roughness of the bulk material as well as the surface chemistry, through specific and partial de-fluorination of the surface were detected and systematically studied by Fourier transform infra-red analysis and scanning electron microscopy. It was found that the rupture of fibrils, formed during membrane processing by thermal-stretching, led to the formation of a denser surface composed of nodules similar to these naturally acting as bridging points across the membrane material between fibrils. This structural change has a profound and impart a permanent effect on the permeation across the modified membranes, which was found to be enhanced by up to 10% for long plasma exposures while the selectivity of the membranes was found to remain unaffected by the treatment at a level higher

  4. [Effect of plasma membrane ion permeability modulators on respiration and heat output of wheat roots].

    Science.gov (United States)

    Alekseeva, V A; Gordon, L Kh; Loseva, N L; Rakhimova, G G; Tsentsevitskiĭ, A N

    2006-01-01

    A study was made of changes in the rates of respiration, heat production, and membrane characteristics in cells of excised roots of wheat seedlings under the modulation of plasma membrane ion permeability by two membrane active compounds: valinomycin (20 microM (V50)) and chlorpromazine (50 microM (CP50) and 100 microM (CP100)). Both compounds increased the loss of potassium ions, which correlated with the lowering of membrane potential, rate of respiration, and heat production after a 2 h exposure. The differences in alteration of these parameters were due to specific action of either compound on the membrane and to the extent of ion homeostasis disturbance. V20 had a weak effect on the studied parameters. V50 caused an increase of the rate of respiration and heat production, which enhanced following a prolonged action (5 h) and were associated with ion homeostatis restoration. The extent of alteration of membrane characteristics (an increase of potassium loss by roots, and lowering of cell membrane potential) as well as energy expense under the action of CP50 during the first period were more pronounced than in the presence of V50. During a prolonged action of CP50, the increase of respiration intensity and heat production correlated with partial recovery of ion homeostatis in cells. Essential lowering of membrane potential and substantial loss of potassium by cells, starting from the early stages of their response reaction, were followed by inhibition of respiration rate and heat production. Alterations of the structure and functional characteristics of excised root cells indicate the intensification of the membrane-tropic effect of a prolonged action of CP100, and the lack of cell energy resources.

  5. Spatiotemporal mapping of diffusion dynamics and organization in plasma membranes

    Science.gov (United States)

    Bag, Nirmalya; Ng, Xue Wen; Sankaran, Jagadish; Wohland, Thorsten

    2016-09-01

    Imaging fluorescence correlation spectroscopy (FCS) and the related FCS diffusion law have been applied in recent years to investigate the diffusion modes of lipids and proteins in membranes. These efforts have provided new insights into the membrane structure below the optical diffraction limit, new information on the existence of lipid domains, and on the influence of the cytoskeleton on membrane dynamics. However, there has been no systematic study to evaluate how domain size, domain density, and the probe partition coefficient affect the resulting imaging FCS diffusion law parameters. Here, we characterize the effects of these factors on the FCS diffusion law through simulations and experiments on lipid bilayers and live cells. By segmenting images into smaller 7  ×  7 pixel areas, we can evaluate the FCS diffusion law on areas smaller than 2 µm and thus provide detailed maps of information on the membrane structure and heterogeneity at this length scale. We support and extend this analysis by deriving a mathematical expression to calculate the mean squared displacement (MSDACF) from the autocorrelation function of imaging FCS, and demonstrate that the MSDACF plots depend on the existence of nanoscopic domains. Based on the results, we derive limits for the detection of domains depending on their size, density, and relative viscosity in comparison to the surroundings. Finally, we apply these measurements to bilayers and live cells using imaging total internal reflection FCS and single plane illumination microscopy FCS.

  6. Isolation of plasma and nuclear membranes of thymocytes. II. Biochemical composition.

    Science.gov (United States)

    Monneron, A; d'Alayer, J

    1978-04-01

    Thymocyte plasma and nuclear membranes obtained by the procedure described in the accompanying paper were analyzed for their biochemical composition. Plasma membranes were very rich in phospholipid, cholesterol, sialic aicd; they did not contain nucleic acids. In comparison, nuclear membranes had a lower phospholipid to protein ratio and contained much less sialic acid and cholesterol. 50% of the cellular cholesterol and of the membrane-bound sialic acid were found in the plasma membranes, 14% in the nuclear membranes. Live cells were labeled with 131I, and the acid-insoluble radioactivity was followed in the subfractions. A good correlation with the distribution and enrichment of plasma membrane market-enzymes was obtained. Label enrichment was about 50-fold in the two lightest of the three plasma membrane fractions. 60% of the label was contained in the plasma membranes, only 4% in the nuclear membranes. Cross-contamination of these two types of membranes was thus negligible. Sodium dodecyl sulfate-gel electrophoresis revealed three different patterns specific for, respectively, plasma membranes, the microsomal-mitochondrial fraction, and nuclear membranes. Each pattern was characterized by a set of proteins and glycoproteins, among which high molecular weight glycoproteins could be considered as marker-proteins of, respectively, 280,000, 260,000, and 230,000 daltons. 131I-labeling of live cells tagged with a very high specific activity three glycoproteins of mol wt 280,000, 200,000, and 135,000 daltons. Nuclear membranes prepared from labeled isolated nuclei had a set of labeled proteins completely different from plasma membranes.

  7. The Plasma Membrane Ca2+ ATPase and the Plasma Membrane Sodium Calcium Exchanger Cooperate in the Regulation of Cell Calcium

    Science.gov (United States)

    Brini, Marisa; Carafoli, Ernesto

    2011-01-01

    Calcium is an ambivalent signal: it is essential for the correct functioning of cell life, but may also become dangerous to it. The plasma membrane Ca2+ ATPase (PMCA) and the plasma membrane Na+/Ca2+ exchanger (NCX) are the two mechanisms responsible for Ca2+ extrusion. The NCX has low Ca2+ affinity but high capacity for Ca2+ transport, whereas the PMCA has a high Ca2+ affinity but low transport capacity for it. Thus, traditionally, the PMCA pump has been attributed a housekeeping role in maintaining cytosolic Ca2+, and the NCX the dynamic role of counteracting large cytosolic Ca2+ variations (especially in excitable cells). This view of the roles of the two Ca2+ extrusion systems has been recently revised, as the specific functional properties of the numerous PMCA isoforms and splicing variants suggests that they may have evolved to cover both the basal Ca2+ regulation (in the 100 nM range) and the Ca2+ transients generated by cell stimulation (in the μM range). PMID:21421919

  8. Phosphatidic acid phosphatase and phospholipdase A activities in plasma membranes from fusing muscle cells.

    Science.gov (United States)

    Kent, C; Vagelos, P R

    1976-06-17

    Plasma membrane from fusing embryonic muscle cells were assayed for phospholipase A activity to determine if this enzyme plays a role in cell fusion. The membranes were assayed under a variety of conditions with phosphatidylcholine as the substrate and no phospholipase A activity was found. The plasma membranes did contain a phosphatidic acid phosphatase which was optimally active in the presence of Triton X-100 and glycerol. The enzyme activity was constant from pH 5.2 to 7.0, and did not require divalent cations. Over 97% of the phosphatidic acid phosphatase activity was in the particulate fraction. The subcellular distribution of the phosphatidic acid phosphatase was the same as the distributions of the plasma membrane markers, (Na+ + k+)-ATPase and the acetylcholine receptor, which indicates that this phosphatase is located exclusively in the plasma membranes. There was no detectable difference in the phosphatidic acid phosphatase activities of plasma membranes from fusing and non-fusing cells.

  9. Single molecule studies of molecular diffusion in cellular membranes: determining membrane structure.

    Science.gov (United States)

    Ritchie, Ken; Spector, Jeff

    Since the advent of single particle/molecule microscopies, researchers have applied these techniques to understanding the fluid membranes of cells. By observing diffusion of membrane proteins and lipids in live cell membranes of eukaryotic cells, it has been found that membranes contain a mosaic of fluid compartments. Such structure may be instrumental in understanding key characteristics of the membrane. Recent single molecule observations on prokaryotic cell membranes will also be discussed.

  10. A Comparative Study of Hydrophilic Modification of Polypropylene Membranes by Remote and Direct Ar Plasma

    Institute of Scientific and Technical Information of China (English)

    ZHANG Suzhen; CHENG Cheng; LAN Yan; MENG Yuedong

    2009-01-01

    Surface modification of polypropylene membrane by argon (Ar) plasma-induced graft polymerization with hydrophilic monomer [acrylic acid (AA) in this work]was investigated.It was found that both the distance of the membrane from the Ar plasma center and the plasma power had a strong influence on the surface modification,hydrophilicity and graft yield (GY) of the treated membrane.Results suggest that remote plasma treatment with a proper sample position,plasma power and graft polymerization leads to a membrane surface with not only less damage,but also more permanent hydrophilicity,than direct plasma treatment does.By analyzing the morphology and the chemical composition of the membrane surface by scanning electron microscopy (SEM) and X-ray photoelectron spectroscopy (XPS),as well as Fourier transform infrared attenuated total reflection spectroscopy (FTIR-ATR) respectively,a possible mechanism was tentatively revealed.

  11. Antifouling enhancement of polysulfone/TiO2 nanocomposite separation membrane by plasma etching

    Science.gov (United States)

    Chen, Z.; Yin, C.; Wang, S.; Ito, K.; Fu, Q. M.; Deng, Q. R.; Fu, P.; Lin, Z. D.; Zhang, Y.

    2017-01-01

    A polysulfone/TiO2 nanocomposite membrane was prepared via casting method, followed by the plasma etching of the membrane surface. Doppler broadened energy spectra vs. positron incident energy were employed to elucidate depth profiles of the nanostructure for the as-prepared and treated membranes. The results confirmed that the near-surface of the membrane was modified by the plasma treatment. The antifouling characteristics for the membranes, evaluated using the degradation of Rhodamin B, indicated that the plasma treatment enhances the photo catalytic ability of the membrane, suggesting that more TiO2 nanoparticles are exposed at the membrane surface after the plasma treatment as supported by the positron result.

  12. 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.

  13. 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.

  14. One-step isolation of plasma membrane proteins using magnetic beads with immobilized concanavalin A1

    Science.gov (United States)

    Lee, Yu-Chen; Block, Gregory; Chen, Huiwen; Folch-Puy, Emma; Foronjy, Robert; Jalili, Roxana; Jendresen, Christian Bille; Kimura, Masashi; Kraft, Edward; Lindemose, Søren; Lu, Jin; McLain, Teri; Nutt, Leta; Ramon-Garcia, Santiago; Smith, Joseph; Spivak, Aaron; Wang, Michael L.; Zanic, Marija; Lin, Sue-Hwa

    2008-01-01

    We have developed a simple method for isolating and purifying plasma membrane proteins from various cell types. This one-step affinity-chromatography method uses the property of the lectin concanavalin A (ConA) and the technique of magnetic-bead separation to obtain highly purified plasma membrane proteins from crude membrane preparations or cell lines. ConA is immobilized onto magnetic beads by binding biotinylated ConA to streptavidin magnetic beads. When these ConA magnetic beads were used to enrich plasma membranes from a crude membrane preparation, this procedure resulted in 3.7-fold enrichment of plasma membrane marker 5′-nucleotidase activity with 70% recovery of the activity in the crude membrane fraction of rat liver. In agreement with the results of 5′-nucleotidase activity, immunoblotting with antibodies specific for a rat liver plasma membrane protein, CEACAM1, indicated that CEACAM1 was enriched about threefold relative to that of the original membranes. In similar experiments, this method produced 13-fold enrichment of 5′-nucleotidase activity with 45% recovery of the activity from a total cell lysate of PC-3 cells and 7.1-fold enrichment of 5′-nucleotidase activity with 33% recovery of the activity from a total cell lysate of HeLa cells. These results suggest that this one-step purification method can be used to isolate total plasma membrane proteins from tissue or cells for the identification of membrane biomarkers. PMID:18765283

  15. Identification of frog photoreceptor plasma and disk membrane proteins by radioiodination

    Energy Technology Data Exchange (ETDEWEB)

    Witt, P.L.; Bownds, M.D.

    1987-03-24

    Several functions have been identified for the plasma membrane of the rod outer segment, including control of light-dependent changes in sodium conductance and a sodium-calcium exchange mechanism. However, little is known about its constituent proteins. Intact rod outer segments substantially free of contaminants were prepared in the dark and purified on a density gradient of Percoll. Surface proteins were then labeled by lactoperoxidase-catalyzed radioiodination, and intact rod outer segments were reisolated. Membrane proteins were identified by polyacrylamide gel electrophoresis and autoradiography. The surface proteins labeled included rhodopsin, the major membrane protein, and 12 other proteins. To compare the protein composition of plasma membrane with that of the internal disk membrane, purified rod outer segments were lysed by hypotonic disruption or freeze-thawing, and plasma plus disk membranes were radioiodinated. In these membrane preparations, rhodopsin was the major iodinated constituent, with 12 other proteins also labeled. Autoradiographic evidence indicated some differences in protein composition between disk and plasma membranes. A quantitative comparison of the two samples showed that labeling of two proteins, 24 kilodaltons (kDa) and 13 kDa, was enriched in the plasma membrane, while labeling of a 220-kDa protein was enriched in the disk membrane. These plasma membrane proteins may be associated with important functions such as the light-sensitive conductance and the sodium-calcium exchanger.

  16. Irvalec inserts into the plasma membrane causing rapid loss of integrity and necrotic cell death in tumor cells.

    Directory of Open Access Journals (Sweden)

    José M Molina-Guijarro

    Full Text Available Irvalec is a marine-derived antitumor agent currently undergoing phase II clinical trials. In vitro, Irvalec induces a rapid loss of membrane integrity in tumor cells, accompanied of a significant Ca(2+ influx, perturbations of membrane conductivity, severe swelling and the formation of giant membranous vesicles. All these effects are not observed in Irvalec-resistant cells, or are significantly delayed by pretreating the cells with Zn(2+. Using fluorescent derivatives of Irvalec it was demonstrated that the compound rapidly interacts with the plasma membrane of tumor cells promoting lipid bilayer restructuration. Also, FRET experiments demonstrated that Irvalec molecules localize in the cell membrane close enough to each other as to suggest that the compound could self-organize, forming supramolecular structures that likely trigger cell death by necrosis through the disruption of membrane integrity.

  17. 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

  18. In silicio identification of glycosyl-phosphatidylinositol-anchored plasma-membrane and cell wall proteins of Saccharomyces cerevisiae.

    Science.gov (United States)

    Caro, L H; Tettelin, H; Vossen, J H; Ram, A F; van den Ende, H; Klis, F M

    1997-12-01

    Use of the Von Heijne algorithm allowed the identification of 686 open reading frames (ORFs) in the genome of Saccharomyces cerevisiae that encode proteins with a potential N-terminal signal sequence for entering the secretory pathway. On further analysis, 51 of these proteins contain a potential glycosyl-phosphatidylinositol (GPI)-attachment signal. Seven additional ORFs were found to belong to this group. Upon examination of the possible GPI-attachment sites, it was found that in yeast the most probable amino acids for GPI-attachment as asparagine and glycine. In yeast, GPI-proteins are found at the cell surface, either attached to the plasma-membrane or as an intrinsic part of the cell wall. It was noted that plasma-membrane GPI-proteins possess a dibasic residue motif just before their predicted GPI-attachment site. Based on this, and on homologies between proteins, families of plasma-membrane and cell wall proteins were assigned, revealing 20 potential plasma-membrane and 38 potential cell wall proteins. For members of three plasma-membrane protein families, a function has been described. On the other hand, most of the cell wall proteins seem to be structural components of the wall, responsive to different growth conditions. The GPI-attachment site of yeast slightly differs from mammalian cells. This might be of use in the development of anti-fungal drugs.

  19. Up against the wall: is yeast cell wall integrity ensured by mechanosensing in plasma membrane microdomains?

    Science.gov (United States)

    Kock, Christian; Dufrêne, Yves F; Heinisch, Jürgen J

    2015-02-01

    Yeast cell wall integrity (CWI) signaling serves as a model of the regulation of fungal cell wall synthesis and provides the basis for the development of antifungal drugs. A set of five membrane-spanning sensors (Wsc1 to Wsc3, Mid2, and Mtl1) detect cell surface stress and commence the signaling pathway upon perturbations of either the cell wall structure or the plasma membrane. We here summarize the latest advances in the structure/function relationship primarily of the Wsc1 sensor and critically review the evidence that it acts as a mechanosensor. The relevance and physiological significance of the information obtained for the function of the other CWI sensors, as well as expected future developments, are discussed.

  20. Glycosidases in the plasma membrane of Ceratitis capitata spermatozoa.

    Science.gov (United States)

    Intra, Jari; De Caro, Daniela; Perotti, Maria-Elisa; Pasini, Maria Enrica

    2011-02-01

    Fruit flies in the family Tephritidae are rated among the world's most destructive agricultural pests. The Mediterranean fruit fly Ceratitis capitata is emerging as a model organism to study the fertilization in Insects. Three integral proteins with glycosidase activity are present in the plasma membrane of spermatozoa. The glycosidases have been purified and characterized. We have demonstrated the presence of three enzymes, a β-N-acetylhexosaminidase, an α-mannosidase and an α-l-fucosidase. The molecular mass of the native enzymes estimated by gel filtration was 160 kDa for β-N-acetylhexosaminidase, 310 kDa for α-mannosidase and 140 kDa for α-l-fucosidase. SDS-PAGE showed that β-N-acetylhexosaminidase is a dimer of a single protein of 73 kDa, α-mannosidase consists of six subunits with different molecular weights and α-l-fucosidase is a dimer made up by two different monomers. Characterization of the purified enzymes included glycosylation pattern, pI, optimal pH, substrate preference, kinetic properties and thermal stability. Soluble forms similar to the sperm associated glycosidases are present. Polyclonal antibodies raised against synthetic peptides designed from the predicted products of the Drosophila melanogaster genes encoding β-N-acetylhexosaminidase and α-l-fucosidase were used. Immunofluorescence labelling of spermatozoa showed that the enzymes are present in the sperm plasma membrane overlying the acrosome and the tail. This work represents the first report on the characterization in C. capitata of sperm proteins that are potentially involved in primary gamete recognition. Copyright © 2010 Elsevier Ltd. All rights reserved.

  1. Large-Scale Structure of Magnetospheric Plasma

    Science.gov (United States)

    Moore, T. E.; Delcourt, D. C.

    1995-01-01

    Recent investigations of magnetospheric plasma structure are summarized under the broad categories of empirical models, transport across boundaries, formation, and dynamics of the plasma sheet. This report reviews work in these areas during the period 1991 to 1993. Fully three-dimensional empirical models and simulations have become important contributors to our understanding of the magnetospheric system. Some new structural concepts have appeared in the literature: the 'entry boundary' and 'geo-pause', the plasma sheet 'region 1 vortices', the 'low-energy layer', the 'adia-baticity boundary' or 'wall region', and a region in the tail to which we refer as the 'injection port'. Traditional structural concepts have also been the subject of recent study, notably the plasmapause, the magnetopause, and the plasma sheet. Significant progress has been made in understanding the nature of plasma sheet formation and dynamics, but the acceleration of electrons to high energy remains somewhat mysterious.

  2. Enquiry into the Topology of Plasma Membrane-Localized PIN Auxin Transport Components.

    Science.gov (United States)

    Nodzyński, Tomasz; Vanneste, Steffen; Zwiewka, Marta; Pernisová, Markéta; Hejátko, Jan; Friml, Jiří

    2016-11-07

    Auxin directs plant ontogenesis via differential accumulation within tissues depending largely on the activity of PIN proteins that mediate auxin efflux from cells and its directional cell-to-cell transport. Regardless of the developmental importance of PINs, the structure of these transporters is poorly characterized. Here, we present experimental data concerning protein topology of plasma membrane-localized PINs. Utilizing approaches based on pH-dependent quenching of fluorescent reporters combined with immunolocalization techniques, we mapped the membrane topology of PINs and further cross-validated our results using available topology modeling software. We delineated the topology of PIN1 with two transmembrane (TM) bundles of five α-helices linked by a large intracellular loop and a C-terminus positioned outside the cytoplasm. Using constraints derived from our experimental data, we also provide an updated position of helical regions generating a verisimilitude model of PIN1. Since the canonical long PINs show a high degree of conservation in TM domains and auxin transport capacity has been demonstrated for Arabidopsis representatives of this group, this empirically enhanced topological model of PIN1 will be an important starting point for further studies on PIN structure-function relationships. In addition, we have established protocols that can be used to probe the topology of other plasma membrane proteins in plants.

  3. Intrarenal localization of the plasma membrane ATP channel pannexin1.

    Science.gov (United States)

    Hanner, Fiona; Lam, Lisa; Nguyen, Mien T X; Yu, Alan; Peti-Peterdi, János

    2012-11-15

    In the renal tubules, ATP released from epithelial cells stimulates purinergic receptors, regulating salt and water reabsorption. However, the mechanisms by which ATP is released into the tubular lumen are multifaceted. Pannexin1 (Panx1) is a newly identified. ubiquitously expressed protein that forms connexin-like channels in the plasma membrane, which have been demonstrated to function as a mechanosensitive ATP conduit. Here, we report on the localization of Panx1 in the mouse kidney. Using immunofluorescence, strong Panx1 expression was observed in renal tubules, including proximal tubules, thin descending limbs, and collecting ducts, along their apical cell membranes. In the renal vasculature, Panx1 expression was localized to vascular smooth muscle cells in renal arteries, including the afferent and efferent arterioles. Additionally, we tested whether Panx1 channels expressed in renal epithelial cells facilitate luminal ATP release by measuring the ATP content of urine samples freshly collected from wild-type and Panx1(-/-) mice. Urinary ATP levels were reduced by 30% in Panx1(-/-) compared with wild-type mice. These results suggest that Panx1 channels in the kidney may regulate ATP release and via purinergic signaling may participate in the control of renal epithelial fluid and electrolyte transport and vascular functions.

  4. Detection of boar sperm plasma membrane protein using Rhodamine 640; implications for cryobiology and physiology

    Science.gov (United States)

    Rhodamine 640 (R640) was used to detect changes in boar sperm plasma membrane protein (PMP) during cryopreservation; a poorly understood phenomenon. The protocol was adapted for boar sperm so that semen samples (n = 17) could be analyzed for PMP (R640 positive) and plasma membrane integrity (PMI; Y...

  5. pH-induced proton permeability changes of plasma membrane vesicles

    NARCIS (Netherlands)

    Miedema, H; Prins, HBA; Staal, H.

    In vivo studies with leaf cells of aquatic plant species such as Elodea nuttallii revealed the proton permeability and conductance of the plasma membrane to be strongly pH dependent. The question was posed if similar pH dependent permeability changes also occur in isolated plasma membrane vesicles.

  6. (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

  7. (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

  8. Monitoring the native phosphorylation state of plasma membrane proteins from a single mouse cerebellum

    DEFF Research Database (Denmark)

    Schindler, J.; Ye, J. Y.; Jensen, Ole Nørregaard

    2013-01-01

    Neuronal processing in the cerebellum involves the phosphorylation and dephosphorylation of various plasma membrane proteins such as AMPA or NMDA receptors. Despite the importance of changes in phosphorylation pattern, no global phospho-proteome analysis has yet been performed. As plasma membrane...

  9. Enzymes of phosphoinositide synthesis in secretory vesicles destined for the plasma membrane in Saccharomyces cerevisiae.

    Science.gov (United States)

    Kinney, A J; Carman, G M

    1990-07-01

    CDP-diacylglycerol synthase, phosphatidylinositol synthase, and phosphatidylinositol kinase activities were associated with post-Golgi apparatus secretory vesicles destined for the plasma membrane of Saccharomyces cerevisiae. These results suggest that the plasma membrane is capable of synthesizing both CDP-diacylglycerol and phosphatidylinositol as well as phosphorylating phosphatidylinositol.

  10. 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

  11. One-step isolation of plasma membrane proteins using magnetic beads with immobilized concanavalin A

    DEFF Research Database (Denmark)

    Lee, Yu-Chen; Block, Gregory; Chen, Huiwen;

    2008-01-01

    We have developed a simple method for isolating and purifying plasma membrane proteins from various cell types. This one-step affinity-chromatography method uses the property of the lectin concanavalin A (ConA) and the technique of magnetic bead separation to obtain highly purified plasma membran...

  12. 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.

  13. Response of plasma membrane H+-ATPase in rice (Oryza sativa) seedlings to simulated acid rain.

    Science.gov (United States)

    Liang, Chanjuan; Ge, Yuqing; Su, Lei; Bu, Jinjin

    2015-01-01

    Understanding the adaptation of plants to acid rain is important to find feasible approaches to alleviate such damage to plants. We studied effects of acid rain on plasma membrane H(+)-ATPase activity and transcription, intracellular H(+), membrane permeability, photosynthetic efficiency, and relative growth rate during stress and recovery periods. Simulated acid rain at pH 5.5 did not affect plasma membrane H(+)-ATPase activity, intracellular H(+), membrane permeability, photosynthetic efficiency, and relative growth rate. Plasma membrane H(+)-ATPase activity and transcription in leaves treated with acid rain at pH 3.5 was increased to maintain ion homeostasis by transporting excessive H(+) out of cells. Then intracellular H(+) was close to the control after a 5-day recovery, alleviating damage on membrane and sustaining photosynthetic efficiency and growth. Simulated acid rain at pH 2.5 inhibited plasma membrane H(+)-ATPase activity by decreasing the expression of H(+)-ATPase at transcription level, resulting in membrane damage and abnormal intracellular H(+), and reduction in photosynthetic efficiency and relative growth rate. After a 5-day recovery, all parameters in leaves treated with pH 2.5 acid rain show alleviated damage, implying that the increased plasma membrane H(+)-ATPase activity and its high expression were involved in repairing process in acid rain-stressed plants. Our study suggests that plasma membrane H(+)-ATPase can play a role in adaptation to acid rain for rice seedlings.

  14. Red wine activates plasma membrane redox system in human erythrocytes.

    Science.gov (United States)

    Tedesco, Idolo; Moccia, Stefania; Volpe, Silvestro; Alfieri, Giovanna; Strollo, Daniela; Bilotto, Stefania; Spagnuolo, Carmela; Di Renzo, Massimo; Aquino, Rita P; Russo, Gian Luigi

    2016-01-01

    In the present study, we report that polyphenols present in red wine obtained by a controlled microvinification process are able to protect human erythrocytes from oxidative stress and to activate Plasma Membrane Redox System (PMRS). Human plasma obtained from healthy subjects was incubated in the presence of whole red wine at a concentration corresponding to 9.13-73 μg/ml gallic acid equivalents to verify the capacity to protect against hypochlorous acid (HOCl)-induced plasma oxidation and to minimize chloramine formation. Red wine reduced hemolysis and chloramine formation induced by HOCl of 40 and 35%, respectively. PMRS present on human erythrocytes transfers electrons from intracellular molecules to extracellular electron acceptors. We demonstrated that whole red wine activated PMRS activity in human erythrocytes isolated from donors in a dose-dependent manner with a maximum at about 70-100 μg/ml gallic acid equivalents. We also showed that red wine increased glutathione (GSH) levels and erythrocytic antioxidant capacity, measured by 2,2-diphenyl-1-picrylhydrazyl (DPPH) quenching assay. Furthermore, we reported that GSH played a crucial role in regulating PMRS activity in erythrocytes. In fact, the effect of iodoacetamide, an alkylating agent that induces depletion of intracellular GSH, was completely counteracted by red wine. Bioactive compounds present in red wine, such as gallic acid, resveratrol, catechin, and quercetin were unable to activate PMRS when tested at the concentrations normally present in aged red wines. On the contrary, the increase of PMRS activity was associated with the anthocyanin fraction, suggesting the capacity of this class of compounds to positively modulate PMRS enzymatic activity.

  15. Existence and characteristics of nitrate reductase in plasma membrane of maize roots

    Institute of Scientific and Technical Information of China (English)

    陈珈; 王学臣

    1995-01-01

    The existence and characteristics of nitrate reductase (NR) have been investigated with microsomes and purified plasma membrane vesicles (RV and IV) from the primary root tips of maize (Zea mays L.). An integral membrane protein capable of reducing nitrate is presented in the plasma membrane which is obviously different from the soluble cytoplasmic NR in respect of NO3- induction and Triton X-100 activation Plasma membrane-bound NR did not have direct coupling relationship with the transmembrane H-transport, however, it could inhibit the electron transmission from NADH to K3[Fe(CN)6]. The possible action mode of plasma membrane redox system that the membrane-bound NR is involved in is discussed.

  16. The isolation of plasma membrane from protoplasts of soybean suspension cultures.

    Science.gov (United States)

    Galbraith, D W; Northcote, D H

    1977-04-01

    A procedure for the isolation of plasma membranes from protoplasts of suspension-cultured soybean is described. Protoplasts were prepared by enzymic digestion of the cell wall and the plasma membrane was labelled with radioactive diazotized sulphanilic acid. The membrane systems from broken protoplasts were separated by continuous isopycnic sucrose gradient centrifugation. Radioactivity was localized in a band possessing a buoyant density of 1-14 g ml-1. The activities of NADPH- and NADH-cytochrome c reductase, fumarase, Mg2+-ATPase, IDPase and acid phosphodiesterase in the various regions of the density gradient were determined. A plasma membrane fraction was selected which was relatively uncontaminated with membranes derived from endoplasmic reticulum, tonoplasts and mitochondria. The results indicated that Mg2+-ATPase and possibly acid phosphodiesterase were associated with the plasma membrane.

  17. Charged anaesthetics alter LM-fibroblast plasma-membrane enzymes by selective fluidization of inner or outer membrane leaflets.

    Science.gov (United States)

    Sweet, W D; Schroeder, F

    1986-10-15

    The functional consequences of the differences in lipid composition and structure between the two leaflets of the plasma membrane were investigated. Fluorescence of 1,6-diphenylhexa-1,3,5-triene(DPH), quenching, and differential polarized phase fluorimetry demonstrated selective fluidization by local anaesthetics of individual leaflets in isolated LM-cell plasma membranes. As measured by decreased limiting anisotropy of DPH fluorescence, cationic (prilocaine) and anionic (phenobarbital and pentobarbital) amphipaths preferentially fluidized the cytofacial and exofacial leaflets respectively. Unlike prilocaine, procaine, also a cation, fluidized both leaflets of these membranes equally. Pentobarbital stimulated 5'-nucleotidase between 0.1 and 5 mM and inhibited at higher concentrations, whereas phenobarbital only inhibited, at higher concentrations. Cationic drugs were ineffective. Two maxima of (Na+ + K+)-ATPase activation were obtained with both anionic drugs. Only one activation maximum was obtained with both cationic drugs. The maximum in activity below 1 mM for all four drugs clustered about a single limiting anisotropy value in the cytofacial leaflet, whereas there was no correlation between activity and limiting anisotropy in the exofacial leaflets. Therefore, although phenobarbital and pentobarbital below 1 mM fluidized the exofacial leaflet more than the cytofacial leaflet, the smaller fluidization in the cytofacial leaflet was functionally significant for (Na+ + K+)-ATPase. Mg2+-ATPase was stimulated at 1 mM-phenobarbital, unaffected by pentobarbital and slightly stimulated by both cationic drugs at concentrations fluidizing both leaflets. Thus the activity of (Na+ + K+)-ATPase was highly sensitive to selective fluidization of the leaflet containing its active site, whereas the other enzymes examined were little affected by fluidization of either leaflet.

  18. Hemocompatibility of poly(vinylidene fluoride) membrane grafted with network-like and brush-like antifouling layer controlled via plasma-induced surface PEGylation.

    Science.gov (United States)

    Chang, Yung; Shih, Yu-Ju; Ko, Chao-Yin; Jhong, Jheng-Fong; Liu, Ying-Ling; Wei, Ta-Chin

    2011-05-03

    In this work, the hemocompatibility of PEGylated poly(vinylidene fluoride) (PVDF) microporous membranes with varying grafting coverage and structures via plasma-induced surface PEGylation was studied. Network-like and brush-like PEGylated layers on PVDF membrane surfaces were achieved by low-pressure and atmospheric plasma treatment. The chemical composition, physical morphology, grafting structure, surface hydrophilicity, and hydration capability of prepared membranes were determined to illustrate the correlations between grafting qualities and hemocompatibility of PEGylated PVDF membranes in contact with human blood. Plasma protein adsorption onto different PEGylated PVDF membranes from single-protein solutions and the complex medium of 100% human plasma were measured by enzyme-linked immunosorbent assay (ELISA) with monoclonal antibodies. Hemocompatibility of the PEGylated membranes was evaluated by the antifouling property of platelet adhesion observed by scanning electron microscopy (SEM) and the anticoagulant activity of the blood coagulant determined by testing plasma-clotting time. The control of grafting structures of PEGylated layers highly regulates the PVDF membrane to resist the adsorption of plasma proteins, the adhesion of platelets, and the coagulation of human plasma. It was found that PVDF membranes grafted with brush-like PEGylated layers presented higher hydration capability with binding water molecules than with network-like PEGylated layers to improve the hemocompatible character of plasma protein and blood platelet resistance in human blood. This work suggests that the hemocompatible nature of grafted PEGylated polymers by controlling grafting structures gives them great potential in the molecular design of antithrombogenic membranes for use in human blood.

  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. Changes of plasma membrane ATPase activity,membrane potential and transmembrane proton gradient in Kandelia candel and Avicennia marina seedlings with various salinities

    Institute of Scientific and Technical Information of China (English)

    ZHAO Zhong-qiu; ZHENG Hai-lei; ZHU Yong-guan

    2004-01-01

    The salt-secreting mangrove, Avicennia marina, and non-salt-secreting mangrove, Kandelia candel were cultivated in sand with various salinities(0 ‰, 10 ‰, 20 ‰, 30 ‰, 40 ‰) for 60 d. Plasma membrane vesicles of high-purity in leaves and roots of A.marina and K. candel seedlings were obtained by two-phase partitioning. The function of the plasma membranes, the activity of ATPase, membrane potential and transmembrane proton gradient, at various salinities were investigated. The results showed that within a certain range of salinity(A. marina and roots of K. candel: 0-30‰;leaves of K.candel: 0-20‰), the activity of ATPase increased with increasing salinity, while high salinity(above 30‰ or 20‰) inhibited ATPase activity. In comparison with A. marina, K. candel appeared to be more sensitive to salinity. The dynamics of membrane potential and transmembrane proton gradient in leaves and roots of A. marina and K. candel seedlings were similar to that of ATPase. When treated directly by NaCl all the indexes were inhibited markedly: there was a little increase within 0-10‰(K. candel) or 0-20‰(A. marina) followed by sharp declining. It indicated that the structure and function of plasma membrane was damaged severely.

  1. Graft polymerization and plasma treatment of polymer membranes for fouling reduction: a review.

    Science.gov (United States)

    Kochkodan, Victor M; Sharma, Virender K

    2012-01-01

    This article presents a review of recent developments in surface modification of polymer membranes via graft polymerization and plasma treatment for reduction of fouling with organic compounds and microorganisms in pressure driven membrane processes. The factors affecting membrane fouling, such as membrane hydrophilicity, charge and surface roughness are discussed. The recent studies in which the reduction of organic fouling and biofouling by the modification of the membrane surface via ultraviolet/redox initiated surface grafting of hydrophilic polymers and low temperature plasma treatment are reviewed.

  2. Characterization of auxin-binding proteins from zucchini plasma membrane

    Science.gov (United States)

    Hicks, G. R.; Rice, M. S.; Lomax, T. L.

    1993-01-01

    We have previously identified two auxin-binding polypeptides in plasma membrane (PM) preparations from zucchini (Cucurbita pepo L.) (Hicks et al. 1989, Proc. Natl. Acad. Sci. USA 86, 4948-4952). These polypeptides have molecular weights of 40 kDa and 42 kDa and label specifically with the photoaffinity auxin analog 5-N3-7-3H-IAA (azido-IAA). Azido-IAA permits both the covalent and radioactive tagging of auxin-binding proteins and has allowed us to characterize further the 40-kDa and 42-kDa polypeptides, including the nature of their attachment to the PM, their relationship to each other, and their potential function. The azido-IAA-labeled polypeptides remain in the pelleted membrane fraction following high-salt and detergent washes, which indicates a tight and possibly integral association with the PM. Two-dimensional electrophoresis of partially purified azido-IAA-labeled protein demonstrates that, in addition to the major isoforms of the 40-kDa and 42-kDa polypeptides, which possess isoelectric points (pIs) of 8.2 and 7.2, respectively, several less abundant isoforms that display unique pIs are apparent at both molecular masses. Tryptic and chymotryptic digestion of the auxin-binding proteins indicates that the 40-kDa and 42-kDa polypeptides are closely related or are modifications of the same polypeptide. Phase extraction with the nonionic detergent Triton X-114 results in partitioning of the azido-IAA-labeled polypeptides into the aqueous (hydrophilic) phase. This apparently paradoxical behavior is also exhibited by certain integral membrane proteins that aggregate to form channels. The results of gel filtration indicate that the auxin-binding proteins do indeed aggregate strongly and that the polypeptides associate to form a dimer or multimeric complex in vivo. These characteristics are consistent with the hypothesis that the 40-kDa and 42-kDa polypeptides are subunits of a multimeric integral membrane protein which has an auxin-binding site, and which may

  3. Plasma membranes modified by plasma treatment or deposition as solid electrolytes for potential application in solid alkaline fuel cells.

    Science.gov (United States)

    Reinholdt, Marc; Ilie, Alina; Roualdès, Stéphanie; Frugier, Jérémy; Schieda, Mauricio; Coutanceau, Christophe; Martemianov, Serguei; Flaud, Valérie; Beche, Eric; Durand, Jean

    2012-07-30

    In the highly competitive market of fuel cells, solid alkaline fuel cells using liquid fuel (such as cheap, non-toxic and non-valorized glycerol) and not requiring noble metal as catalyst seem quite promising. One of the main hurdles for emergence of such a technology is the development of a hydroxide-conducting membrane characterized by both high conductivity and low fuel permeability. Plasma treatments can enable to positively tune the main fuel cell membrane requirements. In this work, commercial ADP-Morgane® fluorinated polymer membranes and a new brand of cross-linked poly(aryl-ether) polymer membranes, named AMELI-32®, both containing quaternary ammonium functionalities, have been modified by argon plasma treatment or triallylamine-based plasma deposit. Under the concomitant etching/cross-linking/oxidation effects inherent to the plasma modification, transport properties (ionic exchange capacity, water uptake, ionic conductivity and fuel retention) of membranes have been improved. Consequently, using plasma modified ADP-Morgane® membrane as electrolyte in a solid alkaline fuel cell operating with glycerol as fuel has allowed increasing the maximum power density by a factor 3 when compared to the untreated membrane.

  4. Plasma Membranes Modified by Plasma Treatment or Deposition as Solid Electrolytes for Potential Application in Solid Alkaline Fuel Cells

    Directory of Open Access Journals (Sweden)

    Christophe Coutanceau

    2012-07-01

    Full Text Available In the highly competitive market of fuel cells, solid alkaline fuel cells using liquid fuel (such as cheap, non-toxic and non-valorized glycerol and not requiring noble metal as catalyst seem quite promising. One of the main hurdles for emergence of such a technology is the development of a hydroxide-conducting membrane characterized by both high conductivity and low fuel permeability. Plasma treatments can enable to positively tune the main fuel cell membrane requirements. In this work, commercial ADP-Morgane® fluorinated polymer membranes and a new brand of cross-linked poly(aryl-ether polymer membranes, named AMELI-32®, both containing quaternary ammonium functionalities, have been modified by argon plasma treatment or triallylamine-based plasma deposit. Under the concomitant etching/cross-linking/oxidation effects inherent to the plasma modification, transport properties (ionic exchange capacity, water uptake, ionic conductivity and fuel retention of membranes have been improved. Consequently, using plasma modified ADP-Morgane® membrane as electrolyte in a solid alkaline fuel cell operating with glycerol as fuel has allowed increasing the maximum power density by a factor 3 when compared to the untreated membrane.

  5. Preliminary study on plasma membrane fluidity of Psychrophilic Yeast Rhodotorula sp. NJ298 in low temperature

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    The ability of cell to modulate the fluidity of plasma membrane was crucial to the survival of microorganism at low temperature. Plasma membrane proteins, fatty acids and carotenoids profiles of Antarctic psychrophilc yeast Rhodotorula sp. NJ298 were investigated at -3 ℃, 0 ℃ and 8 ℃. The results showed that plasma membrane protein content was greater at -3 ℃ than that at 8 ℃, and a unique membrane polypeptide composition with an apparent molecular mass of 94.7 kDa was newly synthesized with SDS-PAGE analysis; GC analysis showed that the main changes of fatty acids were the percentage of unsaturated fatty acids (C18∶ 1 and C18∶ 2) and shorter chain saturated fatty acid (C10∶ 0) increased along with the decrease of the culture temperature from 8 ℃ to -3 ℃; HPLC analysis indicated that astaxanthin was the major functional carotenoids of the plasma membrane, percentage of which increased from 54.6±1.5% at 8 ℃ to 81.9±2.1% at -3 ℃. However the fluidity of plasma membrane which was determined by measuring fluorescence anisotropy was similar at -3 ℃, 0 ℃ and 8 ℃. Hence these changes in plasma membrane's characteristics were involved in the cellular cold-adaptation by which NJ298 could maintain normal plasma membrane fluidity at near-freezing temperature.

  6. 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.

  7. [Function of surface membrane structures in Thiobacillus thiooxidans].

    Science.gov (United States)

    Pivovarova, T A; Karavaĭko, G I

    1975-01-01

    The function of the surface membrane structures was studied with cytochemical techniques on ultrathin sections of Thiobacillus thiooxidans. The transport of elementary sulphur inside the cell involves the surface membrane structures, while oxidation of the sulphur to sulphuric acid takes place on the outer surface of the cytoplasmic membrane. The surface membrane structures are supposed also to participate in the primary dissolution of elementary sulphur at the site of contact of the cells with the mineral.

  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. Strong thin membrane structure. [solar sails

    Science.gov (United States)

    Frazer, R. E. (Inventor)

    1979-01-01

    A continuous process is described for producing strong lightweight structures for use as solar sails for spacecraft propulsion by radiation pressure. A thin reflective coating, such as aluminum, is applied to a rotating cylinder. A nylon mesh, applied over the aluminum coating, is then coated with a polymerizing material such as a para-xylylene monomer gas to polymerize as a film bound to the mesh and the aluminum. An emissivity increasing material such as chromium or silicon monoxide is applied to the polymer film to disperse such material colloidally into the growing polymer film, or to the final polymer film. The resulting membrane structure is then removed from the cylinder. Alternately, the membrane structure can be formed by etching a substrate in the form of an organic film such as a polymide, or a metal foil, to remove material from the substrate and reduce its thickness. A thin reflective coating (aluminum) is applied on one side of the substrate, and an emissivity increasing coating is applied on the reverse side of the substrate.

  10. An early nodulin-like protein accumulates in the sieve element plasma membrane of Arabidopsis

    DEFF Research Database (Denmark)

    Khan, Junaid A.; Wang, Qi; Sjölund, Richard D.

    2007-01-01

    that have a similar overall domain structure of an amino-terminal signal peptide, plastocyanin-like copper-binding domain, proline/serine-rich domain, and carboxy-terminal hydrophobic domain. The amino- and carboxy-terminal domains of the 21.5-kD sieve element-specific ENOD are posttranslationally cleaved...... from the precursor protein, resulting in a mature peptide of approximately 15 kD that is attached to the sieve element plasma membrane via a carboxy-terminal glycosylphosphatidylinositol membrane anchor. Many of the Arabidopsis ENOD-like proteins accumulate in gametophytic tissues, whereas in both...... floral and vegetative tissues, the sieve element-specific ENOD is expressed only within the phloem. Members of the ENOD subfamily of the cupredoxin superfamily do not appear to bind copper and have unknown functions. Phenotypic analysis of homozygous T-DNA insertion mutants for the gene At3g20570 shows...

  11. Differential cargo mobilisation within Weibel-Palade bodies after transient fusion with the plasma membrane.

    Directory of Open Access Journals (Sweden)

    Nikolai I Kiskin

    Full Text Available Inflammatory chemokines can be selectively released from Weibel-Palade bodies (WPBs during kiss-and-run exocytosis. Such selectivity may arise from molecular size filtering by the fusion pore, however differential intra-WPB cargo re-mobilisation following fusion-induced structural changes within the WPB may also contribute to this process. To determine whether WPB cargo molecules are differentially re-mobilised, we applied FRAP to residual post-fusion WPB structures formed after transient exocytosis in which some or all of the fluorescent cargo was retained. Transient fusion resulted in WPB collapse from a rod to a spheroid shape accompanied by substantial swelling (>2 times by surface area and membrane mixing between the WPB and plasma membranes. Post-fusion WPBs supported cumulative WPB exocytosis. To quantify diffusion inside rounded organelles we developed a method of FRAP analysis based on image moments. FRAP analysis showed that von Willebrand factor-EGFP (VWF-EGFP and the VWF-propolypeptide-EGFP (Pro-EGFP were immobile in post-fusion WPBs. Because Eotaxin-3-EGFP and ssEGFP (small soluble cargo proteins were largely depleted from post-fusion WPBs, we studied these molecules in cells preincubated in the weak base NH4Cl which caused WPB alkalinisation and rounding similar to that produced by plasma membrane fusion. In these cells we found a dramatic increase in mobilities of Eotaxin-3-EGFP and ssEGFP that exceeded the resolution of our method (∼ 2.4 µm2/s mean. In contrast, the membrane mobilities of EGFP-CD63 and EGFP-Rab27A in post-fusion WPBs were unchanged, while P-selectin-EGFP acquired mobility. Our data suggest that selective re-mobilisation of chemokines during transient fusion contributes to selective chemokine secretion during transient WPB exocytosis. Selective secretion provides a mechanism to regulate intravascular inflammatory processes with reduced risk of thrombosis.

  12. Differential cargo mobilisation within Weibel-Palade bodies after transient fusion with the plasma membrane.

    Science.gov (United States)

    Kiskin, Nikolai I; Babich, Victor; Knipe, Laura; Hannah, Matthew J; Carter, Tom

    2014-01-01

    Inflammatory chemokines can be selectively released from Weibel-Palade bodies (WPBs) during kiss-and-run exocytosis. Such selectivity may arise from molecular size filtering by the fusion pore, however differential intra-WPB cargo re-mobilisation following fusion-induced structural changes within the WPB may also contribute to this process. To determine whether WPB cargo molecules are differentially re-mobilised, we applied FRAP to residual post-fusion WPB structures formed after transient exocytosis in which some or all of the fluorescent cargo was retained. Transient fusion resulted in WPB collapse from a rod to a spheroid shape accompanied by substantial swelling (>2 times by surface area) and membrane mixing between the WPB and plasma membranes. Post-fusion WPBs supported cumulative WPB exocytosis. To quantify diffusion inside rounded organelles we developed a method of FRAP analysis based on image moments. FRAP analysis showed that von Willebrand factor-EGFP (VWF-EGFP) and the VWF-propolypeptide-EGFP (Pro-EGFP) were immobile in post-fusion WPBs. Because Eotaxin-3-EGFP and ssEGFP (small soluble cargo proteins) were largely depleted from post-fusion WPBs, we studied these molecules in cells preincubated in the weak base NH4Cl which caused WPB alkalinisation and rounding similar to that produced by plasma membrane fusion. In these cells we found a dramatic increase in mobilities of Eotaxin-3-EGFP and ssEGFP that exceeded the resolution of our method (∼ 2.4 µm2/s mean). In contrast, the membrane mobilities of EGFP-CD63 and EGFP-Rab27A in post-fusion WPBs were unchanged, while P-selectin-EGFP acquired mobility. Our data suggest that selective re-mobilisation of chemokines during transient fusion contributes to selective chemokine secretion during transient WPB exocytosis. Selective secretion provides a mechanism to regulate intravascular inflammatory processes with reduced risk of thrombosis.

  13. TiO2-Based Phosphoproteomic Analysis of the Plasma Membrane and the Effects of Phosphatase Inhibitor Treatment

    DEFF Research Database (Denmark)

    Thingholm, Tine; Larsen, Martin Røssel; Ingrell, Christian

    2008-01-01

    of the plasma membrane phosphoproteome are challenging. We present an optimized analytical strategy for plasma membrane phosphoproteomics that combines efficient plasma membrane protein preparation with TiO 2-based phosphopeptide enrichment and high-performance mass spectrometry for phosphopeptide sequencing...

  14. Sperm-Egg Interaction: Evidence for Boar Sperm Plasma Membrane Receptors for Porcine Zona Pellucida

    Science.gov (United States)

    Peterson, Rudolph N.; Russell, Lonnie; Bundman, Donna; Freund, Matthew

    1980-01-01

    Freshly ejaculated, noncapacitated boar sperm bind rapidly and in large numbers to pig egg zona pellucida in vitro. In the present study, the number of sperm bound decreased sharply when sperm motility was lowered by energy poisons or by reducing the temperature. Highly motile sperm from humans, guinea pigs, and rats, added at concentrations ten times higher than control sperm, did not bind to the porcine zona. At the same high concentration, a small number of hamster and bull sperm bound to the zona. Binding of boar sperm to the zona pellucida was blocked almost completely by diluted whole antiserum to sperm plasma membranes and by univalent (Fab) antibody to these membranes. When antibody to sperm plasma membrane was first absorbed with plasma membrane vesicles, sperm binding was not inhibited. These results provide direct evidence for the existence of sperm plasma membrane receptors for the zona pellucida of the pig.

  15. Plasma Membrane H+-ATPase in Maize Roots Induced for NO3- Uptake.

    Science.gov (United States)

    Santi, S.; Locci, G.; Pinton, R.; Cesco, S.; Varanini, Z.

    1995-12-01

    Plasma membrane H+-ATPase was studied in maize (Zea mays L.) roots induced for NO3- uptake. Membrane vesicles were isolated by means of Suc density gradient from roots exposed for 24 h either to 1.5 mM NO3- or 1.5 mM SO4-. The two populations of vesicles had similar composition as shown by diagnostic inhibitors of membrane-associated ATPases. However, both ATP-dependent intravesicular H+ accumulation and ATP hydrolysis were considerably enhanced (60-100%) in vesicles isolated from NO3--induced roots. Km for Mg:ATP and pH dependency were not influenced by NO3- treatment of the roots. ATP hydrolysis in plasma membrane vesicles for both control and NO3--induced roots was not affected by 10 to 150 mM NO3- or Cl-. On the other hand, kinetics of NO3-- or Cl--stimulated ATP-dependent intravesicular H+ accumulation were modified in plasma membrane vesicles isolated from NO3-- induced roots. Immunoassays carried out with polyclonal antibodies against plasma membrane H+-ATPase revealed an increased steady-state level of the enzyme in plasma membrane vesicles isolated from NO3--induced roots. Results are consistent with the idea of an involvement of plasma membrane H+-ATPase in the overall response of roots to NO3-.

  16. 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.

  17. Controlling Structure in Sulfonated Block Copolymer Membranes

    Science.gov (United States)

    Truong, Phuc; Stein, Gila; Strzalka, Joe

    2015-03-01

    In many ionic block copolymer systems, the strong incompatibility between ionic and non-ionic segments will trap non-equilibrium structures in the film, making it difficult to engineer the optimal domain sizes and transport pathways. The goal of this work is to establish a framework for controlling the solid-state structure of sulfonated pentablock copolymer membranes. They have ABCBA block sequence, where A is poly(t-butyl styrene), B is poly(hydrogenated isoprene), and C is poly(styrene sulfonate). To process into films, the polymer is dissolved in toluene/n-propanol solvent mixtures, where the solvent proportions and the polymer loading were both varied. Solution-state structure was measured with small angle X-ray scattering (SAXS). We detected micelles with radii that depend on the solvent composition and polymer loading. Film structure was measured with grazing-incidence SAXS, which shows (i) domain periodicity is constant throughout film thickness; (ii) domain periodicity depends on solvent composition and polymer loading, and approximately matches the micelle radii in solutions. The solid-state packing is consistent with a hard sphere structure factor. Results suggest that solid-state structure can be tuned by manipulating the solution-state self-assembly.

  18. 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.

  19. 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.

  20. 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.

  1. Free-cholesterol loading does not trigger phase separation of the fluorescent sterol dehydroergosterol in the plasma membrane of macrophages

    DEFF Research Database (Denmark)

    Wüstner, Daniel

    2008-01-01

    membrane distribution of the fluorescent cholesterol-mimicking sterol dehydroergosterol (DHE) was investigated in FC-loaded J774 macrophages. Wide field fluorescence and deconvolution microscopy were combined with quantitative assessment of sterol distribution in straightened plasma membrane image segments...... with increased membrane cholesterol content, sterols do not form a separate phase in the plasma membrane....

  2. Cryodamage to plasma membrane integrity in head and tail regions of human sperm

    Institute of Scientific and Technical Information of China (English)

    Wei-JieZHU; Xue-GaoLIU

    2000-01-01

    Aim: To investigate the effect of cryopreservation on the plasma membrane integrity in the head and tail regions of individual sperm, and the relationship between intact cryopreserved sperm and its motility and zona-free hamster oocyte penetration rate. Methods: The eosin Y exclusion and the hypoosmotic swelling tests were combined to form a single test (HOS-EY test) to identify the spermatozoa with four types of membrane integrity. Results: After cryopreservation, there was a marked decline in the percentage of spermatozoa with Type IV membrane integrity (head membrane intact/tail membrane intact), and a significant increase in those with Type Ⅰ (head membrane damaged/tail membrane damaged) and Type Ⅲ (head membrane damaged/tail membrane intact) membrane integrity (n = 50, P0.05). Conclusion: (1) The HOS-EY test has the advantage of showing four patterns of membrane integrity in individual spermatozoon; (2) Cryopreservation causes a significant membrane rupture in the head and tail regions of spermatozoa; Type IT[ is the main transitional state of membrane cryodamage; (3) Cryodamage to head and tail membrane may occur independently; the presence of an intact tail membrane does not necessarily indicate the intactness of head membrane. (4) Intact membranes am closely related to postthaw motility, but do not reflect the fertilizing potential.

  3. Plasma membrane calcium pumps and their emerging roles in cancer

    Institute of Scientific and Technical Information of China (English)

    Sarah; J; Roberts-Thomson; Merril; C; Curry; Gregory; R; Monteith

    2010-01-01

    Alterations in calcium signaling and/or the expression of calcium pumps and channels are an increasingly recognized property of some cancer cells.Alterations in the expression of plasma membrane calcium ATPase(PMCA) isoforms have been reported in a variety of cancer types,including those of breast and colon,with some studies of cancer cell line differentiation identifying specific PMCA isoforms,which may be altered in some cancers.Some studies have also begun to assess levels of PMCA isoforms in clinical tumor samples and to address mechanisms of altered PMCA expression in cancers.Both increases and decreases in PMCA expression have been reported in different cancer types and in many cases these alterations are isoform specific.In this review,we provide an overview of studies investigating the expression of PMCA in cancer and discuss how both the overexpression and reduced expression of a PMCA isoform in a cancer cell could bestow a growth advantage,through augmenting responses to proliferative stimuli or reducing sensitivity to apoptosis.

  4. Arrestin-mediated endocytosis of yeast plasma membrane transporters.

    Science.gov (United States)

    Nikko, Elina; Pelham, Hugh R B

    2009-12-01

    Many plasma membrane transporters in yeast are endocytosed in response to excess substrate or certain stresses and degraded in the vacuole. Endocytosis invariably requires ubiquitination by the HECT domain ligase Rsp5. In the cases of the manganese transporter Smf1 and the amino acid transporters Can1, Lyp1 and Mup1 it has been shown that ubiquitination is mediated by arrestin-like adaptor proteins that bind to Rsp5 and recognize specific transporters. As yeast contains a large family of arrestins, this has been suggested as a general model for transporter regulation; however, analysis is complicated by redundancy amongst the arrestins. We have tested this model by removing all the arrestins and examining the requirements for endocytosis of four more transporters, Itr1 (inositol), Hxt6 (glucose), Fur4 (uracil) and Tat2 (tryptophan). This reveals functions for the arrestins Art5/Ygr068c and Art4/Rod1, and additional roles for Art1/Ldb19, Art2/Ecm21 and Art8/Csr2. It also reveals functional redundancy between arrestins and the arrestin-like adaptors Bul1 and Bul2. In addition, we show that delivery to the vacuole often requires multiple additional ubiquitin ligases or adaptors, including the RING domain ligase Pib1, and the adaptors Bsd2, Ear1 and Ssh4, some acting redundantly. We discuss the similarities and differences in the requirements for regulation of different transporters.

  5. Liquid general anesthetics lower critical temperatures in plasma membrane vesicles

    CERN Document Server

    Gray, Ellyn; Machta, Benjamin B; Veatch, Sarah L

    2013-01-01

    A large and diverse array of small hydrophobic molecules induce general anesthesia. Their efficacy as anesthetics has been shown to correlate both with their affinity for a hydrophobic environment and with their potency in inhibiting certain ligand gated ion channels. Here we explore the effects that n-alcohols and other liquid anesthetics have on the two-dimensional miscibility critical point observed in cell derived giant plasma membrane vesicles (GPMVs). We show that anesthetics depress the critical temperature (Tc) of these GPMVs without strongly altering the ratio of the two liquid phases found below Tc. The magnitude of this affect is consistent across n-alcohols when their concentration is rescaled by the median anesthetic concentration (AC50) for tadpole anesthesia, but not when plotted against the overall concentration in solution. At AC50 we see a 4{\\deg}C downward shift in Tc, much larger than is typically seen in the main chain transition at these anesthetic concentrations. GPMV miscibility critic...

  6. The plasma membrane transport systems and adaptation to salinity.

    Science.gov (United States)

    Mansour, Mohamed Magdy F

    2014-11-15

    Salt stress represents one of the environmental challenges that drastically affect plant growth and yield. Evidence suggests that glycophytes and halophytes have a salt tolerance mechanisms working at the cellular level, and the plasma membrane (PM) is believed to be one facet of the cellular mechanisms. The responses of the PM transport proteins to salinity in contrasting species/cultivars were discussed. The review provides a comprehensive overview of the recent advances describing the crucial roles that the PM transport systems have in plant adaptation to salt. Several lines of evidence were presented to demonstrate the correlation between the PM transport proteins and adaptation of plants to high salinity. How alterations in these transport systems of the PM allow plants to cope with the salt stress was also addressed. Although inconsistencies exist in some of the information related to the responses of the PM transport proteins to salinity in different species/cultivars, their key roles in adaptation of plants to high salinity is obvious and evident, and cannot be precluded. Despite the promising results, detailed investigations at the cellular/molecular level are needed in some issues of the PM transport systems in response to salinity to further evaluate their implication in salt tolerance.

  7. Fractal structures in nonlinear plasma physics.

    Science.gov (United States)

    Viana, R L; da Silva, E C; Kroetz, T; Caldas, I L; Roberto, M; Sanjuán, M A F

    2011-01-28

    Fractal structures appear in many situations related to the dynamics of conservative as well as dissipative dynamical systems, being a manifestation of chaotic behaviour. In open area-preserving discrete dynamical systems we can find fractal structures in the form of fractal boundaries, associated to escape basins, and even possessing the more general property of Wada. Such systems appear in certain applications in plasma physics, like the magnetic field line behaviour in tokamaks with ergodic limiters. The main purpose of this paper is to show how such fractal structures have observable consequences in terms of the transport properties in the plasma edge of tokamaks, some of which have been experimentally verified. We emphasize the role of the fractal structures in the understanding of mesoscale phenomena in plasmas, such as electromagnetic turbulence.

  8. Surface Modification of Polypropylene Microporous Membrane by Atmospheric-pressure Plasma Induced N-vinyl-2-pyrrolidone Graft Polymerization

    Institute of Scientific and Technical Information of China (English)

    ZHONG Shaofeng

    2012-01-01

    Membrane surfaces modified with poly(N-vinyl-2-pyrrolidone) (PNVP) can be endowed with hydrophilicity,biocompatibility and functionality.In this work,atmospheric pressure dielectric barrier discharge plasma graft polymerization of N-vinyl-2-pyrrolidone (NVP) onto polypropylene (PP) microporous membrane surface was studied.The experimental results reveal that plasma treatment conditions,such as discharge power,treatment time and adsorbed NVP amount,have remarkable effects on the grafting degree of NVP.Structural and morphological changes on the membrane surfaces were characterized by attenuated total reflection-Fourier transform infrared spectroscopy (FT-IR/ATR),X-ray photoelectron spectroscope (XPS) and field emission scanning electron microscopy (FE-SEM).Water contact angles of the membrane surfaces were also measured by the sessile drop method.Water contact angles on the membrane surfaces decrease with the increase of NVP grafting degree,which indicates an enhanced hydrophilicity for the modified membranes.The effects of grafting degrees on pure water fluxes were also measured.It is shown that pure water fluxes increase with grafting degree firstly and then decrease adversely.Finally,filtration of bovine serum albumin (BSA) solution and platelets adhesion of the PNVP modified membranes show good protein resistance and potential biocompatibility due to the enhancement of surface hydrophilicity.

  9. Pinkbar is an epithelial-specific BAR domain protein that generates planar membrane structures

    Energy Technology Data Exchange (ETDEWEB)

    Pykäläinen, Anette; Boczkowska, Malgorzata; Zhao, Hongxia; Saarikangas, Juha; Rebowski, Grzegorz; Jansen, Maurice; Hakanen, Janne; Koskela, Essi V.; Peränen, Johan; Vihinen, Helena; Jokitalo, Eija; Salminen, Marjo; Ikonen, Elina; Dominguez, Roberto; Lappalainen, Pekka (Helsinki); (Penn)

    2013-05-29

    Bin/amphipysin/Rvs (BAR)-domain proteins sculpt cellular membranes and have key roles in processes such as endocytosis, cell motility and morphogenesis. BAR domains are divided into three subfamilies: BAR- and F-BAR-domain proteins generate positive membrane curvature and stabilize cellular invaginations, whereas I-BAR-domain proteins induce negative curvature and stabilize protrusions. We show that a previously uncharacterized member of the I-BAR subfamily, Pinkbar, is specifically expressed in intestinal epithelial cells, where it localizes to Rab13-positive vesicles and to the plasma membrane at intercellular junctions. Notably, the BAR domain of Pinkbar does not induce membrane tubulation but promotes the formation of planar membrane sheets. Structural and mutagenesis analyses reveal that the BAR domain of Pinkbar has a relatively flat lipid-binding interface and that it assembles into sheet-like oligomers in crystals and in solution, which may explain its unique membrane-deforming activity.

  10. Surface hydrophobic modification of cellulose membranes by plasma-assisted deposition of hydrocarbon films

    Directory of Open Access Journals (Sweden)

    Mudtorlep Nisoa

    2010-03-01

    Full Text Available Surface modification by plasma polymerization is an efficient method to change the surface properties of a membrane. Desirable functionality such as hydrophobicity or hydrophilicity can be obtained, depending on plasma chemistry of gas precursors and discharge conditions. In this work, RF magnetron plasma is produced using acetylene and nitrogen as precursor gases. Variations of RF power, particle flux, deposited time and pressure of the precursor gases have been made to observe coating effects on the cellulose membranes. When appropriated conditions are used, a thin brownish film of hydrocarbon was formed on the membrane, and the water contact angle increased from 35 to 130 degrees.

  11. Leucine-based receptor sorting motifs are dependent on the spacing relative to the plasma membrane

    DEFF Research Database (Denmark)

    Geisler, C; Dietrich, J; Nielsen, B L;

    1998-01-01

    amino acid, is constitutively active. In this study, we have investigated how the spacing relative to the plasma membrane affects the function of both types of leucine-based motifs. For phosphorylation-dependent leucine-based motifs, a minimal spacing of 7 residues between the plasma membrane...... and the phospho-acceptor was required for phosphorylation and thereby activation of the motifs. For constitutively active leucine-based motifs, a minimal spacing of 6 residues between the plasma membrane and the acidic residue was required for optimal activity of the motifs. In addition, we found that the acidic...

  12. 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.

  13. Direct in vivo interaction of the antibiotic primycin with the plasma membrane of Candida albicans: an EPR study.

    Science.gov (United States)

    Virág, Eszter; Belagyi, Joseph; Gazdag, Zoltán; Vágvölgyi, Csaba; Pesti, Miklós

    2012-01-01

    The direct interaction of the antibiotic primycin with the plasma membrane was investigated by employing the well-characterized ergosterol-producing, amphotericin B-sensitive parental Candida albicans strain 33erg(+) and its ergosterol-less amphotericin B-resistant plasma membrane mutant erg-2. The growth inhibition concentration in shaken liquid medium was 64 μgml(-1) for 33erg(+) and 128 μgml(-1) for erg-2, suggesting that the plasma membrane composition influences the mode of action of primycin. To determine the primycin-induced changes in the plasma membrane dynamic, electron paramagnetic resonance (EPR) spectroscopy methods were used, the spin-labeled fatty acid 5-(4,4-dimethyloxazolidine-N-oxyl)stearic acid) being applied for the in vivo measurements. The phase transition temperatures of untreated strain 33erg(+) and its mutant erg-2 were 12.5°C and 11°C, respectively. After 128 μgml(-1) primycin treatment, these values increased to 17.5°C and 16°C, revealing a significant reduction in the phospholipid flexibility. Saturation transfer EPR measurements demonstrated that, the rotational correlation times of the spin label molecule for the control samples of 33erg(+) and erg-2 were 60 ns and 100 ns. These correlation times gradually decreased on the addition of increasing primycin concentrations, reaching 8 μs and 1 μs. The results indicate the plasma membrane "rigidizing" effect of primycin, a feature that may stem from its ability to undergo complex formation with membrane constituent fatty acid molecules, causing alterations in the structures of phospholipids in the hydrophobic surface near the fatty acid chain region. Copyright © 2011 Elsevier B.V. All rights reserved.

  14. Plasma membrane Ca2+-ATPases:Targets of oxidative stress in brain aging and neurodegeneration

    Institute of Scientific and Technical Information of China (English)

    Asma; Zaidi

    2010-01-01

    The plasma membrane Ca2+-ATPase(PMCA)pumps play an important role in the maintenance of precise levels of intracellular Ca2+[Ca2+]i,essential to the functioning of neurons.In this article,we review evidence showing age-related changes of the PMCAs in synaptic plasma membranes(SPMs).PMCA activity and protein levels in SPMs diminish progressively with increasing age. The PMCAs are very sensitive to oxidative stress and undergo functional and structural changes when exposed to oxidants of physiological relevance.The major signatures of oxidative modification in the PMCAs are rapid inactivation,conformational changes,aggregation, internalization from the plasma membrane and proteolytic degradation.PMCA proteolysis appears to be mediated by both calpains and caspases.The predominance of one proteolytic pathway vs the other,the ensuing pattern of PMCA degradation and its consequence on pump activity depends largely on the type of insult,its intensity and duration.Experimental reduction of PMCA expression not only alters the dynamics of cellular Ca2+ handling but also has a myriad of downstream conse-quences on various aspects of cell function,indicating a broad role of these pumps.Age-and oxidation-related down-regulation of the PMCAs may play an important role in compromised neuronal function in the aging brain and its several-fold increased susceptibility to neurodegenerative disorders such as Alzheimer’s disease, Parkinson’s disease,and stroke.Therapeutic approaches that protect the PMCAs and stabilize[Ca2+]i homeostasis may be capable of slowing and/or preventing neuronal degeneration.The PMCAs are therefore emerging as a new class of drug targets for therapeutic interventions in various chronic degenerative disorders.

  15. Perspective on plasma membrane cholesterol efflux and spermatozoal function

    Directory of Open Access Journals (Sweden)

    Dhastagir Sultan Sheriff

    2010-01-01

    Full Text Available The process of sperm maturation, capacitation, and fertilization occur in different molecular milieu provided by epididymis and female reproductive tract including oviduct. The different tissue environment with different oxygen tension and temperature may still influence the process of sperm maturation and capacitation. Reactive oxygen species (ROS is reported to be an initial switch that may activate the molecular process of capacitation. Therefore, the generation of reactive oxygen species and its possible physiological role depends upon a balance between its formation and degradation in an open environment provided by female reproductive tract. The sensitivity of the spermatozoa to the action of ROS may be due to its exposure for the first time to an oxygen rich external milieu compared to its internal milieu in the male reproductive tract. Reduced temperature in testicular environment coupled with reduced oxygen tension may be the right molecular environment for the process of spermatogenesis and spermiogenesis. The morphologically mature spermatozoa then may attain its motility in an environment provided by the caput epididymis wherein, the dyenin motor can become active. This ability to move forward will make the spermatozoa physiologically fit to undertake its sojourn in the competitive race of fertilization in a new oxygen rich female reproductive tract. The first encounter may be oxygen trigger or preconditioning of the spermatozoa with reactive oxygen species that may alter the spermatozoal function. Infertility is still one of the major global health problems that need medical attention. Apart from the development of artificial methods of reproduction and development of newer techniques in the field of andrology focuses attention on spermatozoal structure and metabolism. Therefore, understanding the molecular mechanisms involved in fertilization in general and that of sperm capacitation in particular may help lead to new and better

  16. Modelling and Fabrication of Micro-SOFC Membrane Structure

    Directory of Open Access Journals (Sweden)

    Brigita ABAKEVIČIENĖ

    2014-06-01

    Full Text Available Fabrication process of micro-SOFC membrane structure using the bulk micromachining of silicon technique with SiO2 and Si3N4 sacrificial layers is presented in this study. The process involves back side photolithography, magnetron sputtering of platinum thin films, thermal evaporation of YSZ electrolyte, deep reactive ion etching of silicon, and, finally, release of free-standing membrane using CF4/O2 plasma etching.X-ray analysis shows the cubic phase of YSZ electrolyte and platinum electrodes. Modelling of normal stress distribution in the micro-SOFC structure with the Si3N4 sacrificial layer shows that at high temperatures the substrate expands less than the coating, causing tensile stresses in the substrate area next to the coating and compressive stresses in the coating, as the substrate material has a lower coefficient of thermal expansion than the layered Pt/YSZ/Pt coating. DOI: http://dx.doi.org/10.5755/j01.ms.20.2.5585

  17. Modelling and Fabrication of Micro-SOFC Membrane Structure

    Directory of Open Access Journals (Sweden)

    Brigita ABAKEVIČIENĖ

    2014-06-01

    Full Text Available Fabrication process of micro-SOFC membrane structure using the bulk micromachining of silicon technique with SiO2 and Si3N4 sacrificial layers is presented in this study. The process involves back side photolithography, magnetron sputtering of platinum thin films, thermal evaporation of YSZ electrolyte, deep reactive ion etching of silicon, and, finally, release of free-standing membrane using CF4/O2 plasma etching.X-ray analysis shows the cubic phase of YSZ electrolyte and platinum electrodes. Modelling of normal stress distribution in the micro-SOFC structure with the Si3N4 sacrificial layer shows that at high temperatures the substrate expands less than the coating, causing tensile stresses in the substrate area next to the coating and compressive stresses in the coating, as the substrate material has a lower coefficient of thermal expansion than the layered Pt/YSZ/Pt coating. DOI: http://dx.doi.org/10.5755/j01.ms.20.2.5585

  18. Plasma deposition of silver nanoparticles on ultrafiltration membranes: antibacterial and anti-biofouling properties.

    Science.gov (United States)

    Cruz, Mercedes Cecilia; Ruano, Gustavo; Wolf, Marcus; Hecker, Dominic; Vidaurre, Elza Castro; Schmittgens, Ralph; Rajal, Verónica Beatriz

    2015-02-01

    A novel and versatile plasma reactor was used to modify Polyethersulphone commercial membranes. The equipment was applied to: i) functionalize the membranes with low-temperature plasmas, ii) deposit a film of poly(methyl methacrylate) (PMMA) by Plasma Enhanced Chemical Vapor Deposition (PECVD) and, iii) deposit silver nanoparticles (SNP) by Gas Flow Sputtering. Each modification process was performed in the same reactor consecutively, without exposure of the membranes to atmospheric air. Scanning electron microscopy and transmission electron microscopy were used to characterize the particles and modified membranes. SNP are evenly distributed on the membrane surface. Particle fixation and transport inside membranes were assessed before- and after-washing assays by X-ray photoelectron spectroscopy depth profiling analysis. PMMA addition improved SNP fixation. Plasma-treated membranes showed higher hydrophilicity. Anti-biofouling activity was successfully achieved against Gram-positive (Enterococcus faecalis) and -negative (Salmonella Typhimurium) bacteria. Therefore, disinfection by ultrafiltration showed substantial resistance to biofouling. The post-synthesis functionalization process developed provides a more efficient fabrication route for anti-biofouling and anti-bacterial membranes used in the water treatment field. To the best of our knowledge, this is the first report of a gas phase condensation process combined with a PECVD procedure in order to deposit SNP on commercial membranes to inhibit biofouling formation.

  19. Plasma deposition of silver nanoparticles on ultrafiltration membranes: antibacterial and anti-biofouling properties

    Science.gov (United States)

    Cruz, Mercedes Cecilia; Ruano, Gustavo; Wolf, Marcus; Hecker, Dominic; Vidaurre, Elza Castro; Schmittgens, Ralph; Rajal, Verónica Beatriz

    2015-01-01

    A novel and versatile plasma reactor was used to modify Polyethersulphone commercial membranes. The equipment was applied to: i) functionalize the membranes with low-temperature plasmas, ii) deposit a film of poly(methyl methacrylate) (PMMA) by Plasma Enhanced Chemical Vapor Deposition (PECVD) and, iii) deposit silver nanoparticles (SNP) by Gas Flow Sputtering. Each modification process was performed in the same reactor consecutively, without exposure of the membranes to atmospheric air. Scanning electron microscopy and transmission electron microscopy were used to characterize the particles and modified membranes. SNP are evenly distributed on the membrane surface. Particle fixation and transport inside membranes were assessed before- and after-washing assays by X-ray photoelectron spectroscopy depth profiling analysis. PMMA addition improved SNP fixation. Plasma-treated membranes showed higher hydrophilicity. Anti-biofouling activity was successfully achieved against Gram-positive (Enterococcus faecalis) and -negative (Salmonella Typhimurium) bacteria. Therefore, disinfection by ultrafiltration showed substantial resistance to biofouling. The post-synthesis functionalization process developed provides a more efficient fabrication route for anti-biofouling and anti-bacterial membranes used in the water treatment field. To the best of our knowledge, this is the first report of a gas phase condensation process combined with a PECVD procedure in order to deposit SNP on commercial membranes to inhibit biofouling formation. PMID:26166926

  20. 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

  1. 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.

  2. Phosphosite mapping of P-type plasma membrane H+-ATPase in homologous and heterologous environments

    DEFF Research Database (Denmark)

    Rudashevskaya, Elena; Ye, Juanying; Jensen, Ole N;

    2012-01-01

    Phosphorylation is an important posttranslational modification of proteins in living cells and primarily serves regulatory purposes. Several methods were employed for isolating phosphopeptides from proteolytically digested plasma membranes of Arabidopsis thaliana. After a mass spectrometric...

  3. Laser induced wounding of the plasma membrane and methods to study the repair process.

    Science.gov (United States)

    Jimenez, Ana J; Maiuri, Paolo; Lafaurie-Janvore, Julie; Perez, Franck; Piel, Matthieu

    2015-01-01

    Cells are constantly exposed to agents that can trigger the perforation of their plasma membrane. This damage occurs naturally, and the frequency and intensity depends on how much cells are exposed to damaging threats. The following protocol is a simple and powerful method to damage the plasma membrane using laser ablation. It allows the induction of a single and localized wound at the plasma membrane of cultured cells, which can be followed with fast time-lapse imaging. The first part of the protocol describes simple cell culture techniques and the material ideal to make the experiments. A second part of the protocol gives advice about the procedures to make effective wounds in cells while ensuring a good survival rate. We also propose different ways to follow the opening and closure of the plasma membrane. Finally, we describe the procedure to efficiently analyze the data acquired after single cell photodamage to characterize the wounding process.

  4. Plasma membrane proteomics and its application in clinical cancer biomarker discovery

    DEFF Research Database (Denmark)

    Leth-Larsen, Rikke; Lund, Rikke; Ditzel, Henrik J

    2010-01-01

    Plasma membrane proteins that are exposed on the cell surface have important biological functions, such as signaling into and out of the cells, ion transport, and cell-cell and cell-matrix interactions. The expression level of many of the plasma membrane proteins involved in these key functions...... targeted by protein drugs, such as human antibodies, that have enhanced survival of several groups of cancer patients. The combination of novel analytical approaches and subcellular fractionation procedures has made it possible to study the plasma membrane proteome in more detail, which will elucidate...... cancer biology, particularly metastasis, and guide future development of novel drug targets. The technical advances in plasma membrane proteomics and the consequent biological revelations will be discussed herein. Many of the advances have been made using cancer cell lines, but because the main goal...

  5. Arabidopsis TWISTED DWARF1 functionally interacts with Auxin Exporter ABCB1 on the root plasma membrane

    DEFF Research Database (Denmark)

    Wang, Bangjun; Bailly, Aurélien; Zwiewka, Marta

    2013-01-01

    Plant architecture is influenced by the polar, cell-to-cell transport of auxin that is primarily provided and regulated by plasma membrane efflux catalysts of the PIN-FORMED and B family of ABC transporter (ABCB) classes. The latter were shown to require the functionality of the FK506 binding...... assays, we demonstrate a predominant lateral, mainly outward-facing, plasma membrane location for TWD1 in the root epidermis characterized by the lateral marker ABC transporter G36/PLEIOTROPIC DRUG-RESISTANCE8/PENETRATION3. At these epidermal plasma membrane domains, TWD1 colocalizes with nonpolar ABCB1....... In planta bioluminescence resonance energy transfer analysis was used to verify specific ABC transporter B1 (ABCB1)-TWD1 interaction. Our data support a model in which TWD1 promotes lateral ABCB-mediated auxin efflux via protein-protein interaction at the plasma membrane, minimizing reflux from the root...

  6. Recycling endosomes in apical plasma membrane domain formation and epithelial cell polarity

    NARCIS (Netherlands)

    Golachowska, Magdalena R.; Hoekstra, Dick; van IJzendoorn, Sven C. D.

    2010-01-01

    Recycling endosomes have taken central stage in the intracellular sorting and polarized trafficking of apical and basolateral plasma membrane components. Molecular players in the underlying mechanisms are now emerging, including small GTPases, class V myosins and adaptor proteins. In particular,

  7. Involvement of Plasma Membrane H+-ATPase in Adaption of Rice to Ammonium Nutrient

    Institute of Scientific and Technical Information of China (English)

    ZHU Yi-yong; LIAN Juan; ZENG Hou-qing; LIU GAN; DI Ting-jun; SHEN Qi-rong; XU Guo-hua

    2011-01-01

    The preference of paddy rice for NH4+ rather than NO3- is associated with its tolerance to low pH since a rhizosphere acidification occurs during NH4+ absorption.However,the adaptation of rice root to low pH has not been fully elucidated.The plasma membrane H+-ATPase is a universal electronic H+ pump,which uses ATP as energy source to pump H+ across the plasma membranes into the apoplast.The key function of this enzyme is to keep pH homeostasis of plant cells and generate a H+ electrochemical gradient,thereby providing the driving force for the active influx and efflux of ions and metabolites across the plasma membrane.This study investigated the acclimation of plasma membrane H+-ATPase of rice root to low pH.This mechanism might be partly responsible for the preference of rice plants to NH4+ nutrition.

  8. Expression patterns of genes encoding plasma membrane aquaporins during fruit development in cucumber (Cucumis sativus L.).

    Science.gov (United States)

    Shi, Jin; Wang, Jinfang; Li, Ren; Li, Dianbo; Xu, Fengfeng; Sun, Qianqian; Zhao, Bin; Mao, Ai-Jun; Guo, Yang-Dong

    2015-11-01

    Aquaporins are membrane channels precisely regulating water movement through cell membranes in most living organisms. Despite the advances in the physiology of fruit development, their participation during fruit development in cucumber still barely understood. In this paper, the expressions of 12 genes encoding plasma membrane intrinsic proteins (PIPs) were analyzed during cucumber fruit development in our work. Based on the homology search with known PIPs from rice, Arabidopsis and strawberry, 12 cucumber PIP genes subfamily members were identified. Cellular localization assays indicated that CsPIPs were localized in the plasma membrane. The qRT-PCR analysis of CsPIPs showed that 12 CsPIPs were differentially expressed during fruit development. These results suggest that 12 genes encoding plasma membrane intrinsic proteins (CsPIPs) play very important roles in cucumber life cycle and the data generated will be helpful in understanding their precise roles during fruit development in cucumber.

  9. Expression and structural analysis of membrane proteins

    OpenAIRE

    Eifler, Nora

    2006-01-01

    1.1 Membrane Proteins Between one quarter and one third of all genes in eukaryotic and prokaryotic organisms code for integral membrane proteins (IMPs) (Essen, 2002). These proteins are essential parts of biological membranes and confer various functions, such as energy conversion, transport, biosynthesis of lipids, signal transduction, or cell recognition. The enormous economical potential of membrane proteins is highlighted by the family of G-protein-coupled receptors (GPC...

  10. 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.

  11. Analysis of tensioned membrane structures considering cable sliding

    Institute of Scientific and Technical Information of China (English)

    宋昌永

    2003-01-01

    In routine design of tensioned membrane structures, the membrane is generally modeled using space membrane elements and the cables by space cable elements, with no sliding allowed between the membrane and the cables. On the other hand, large deflections are expected and sliding between the membrane and the cables is inevitable. In the present paper, the general finite element code ABAQUS was employed to investigate the influence of cable sliding on membrane surface on the structural behavior. Three analysis models were devised to fulfill this purpose: (1) The membrane element shares nodes with the cable element; (2) The cable can slide on the membrane surface freely (without friction) and (3) The cable can slide on the membrane surface, but with friction between the cable and the membrane. The sliding problem is modeled using a surface - based contact algorithm. The results from three analysis models are compared, showing that cable sliding has only little influence on the structure shape and on the stress distributions in the membrane. The main influence of cable sliding may be its effect on the dynamic behavior of tensioned membrane structures.

  12. The plasma membrane redox system: a candidate source of aging-related oxidative stress

    OpenAIRE

    de Grey, Aubrey D. N. J

    2005-01-01

    The plasma membrane redox system (PMRS) is an electron transport chain in the plasma membrane that transfers electrons from either intra- or extracellular donors to extracellular acceptors. Unlike the superoxide-generating NADPH oxidase of phagocytes and the homologous (but much less active) enzymes found in some other cells, the PMRS is still incompletely characterised at the molecular level. Much is known, however, concerning its function and affinity for both physiological and non-physiolo...

  13. 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.

  14. Yeast cell wall integrity sensors form specific plasma membrane microdomains important for signalling.

    Science.gov (United States)

    Kock, Christian; Arlt, Henning; Ungermann, Christian; Heinisch, Jürgen J

    2016-09-01

    The cell wall integrity (CWI) pathway of the yeast Saccharomyces cerevisiae relies on the detection of cell surface stress by five sensors (Wsc1, Wsc2, Wsc3, Mid2, Mtl1). Each sensor contains a single transmembrane domain and a highly mannosylated extracellular region, and probably detects mechanical stress in the cell wall or the plasma membrane. We here studied the distribution of the five sensors at the cell surface by using fluorescently tagged variants in conjunction with marker proteins for established membrane compartments. We find that each of the sensors occupies a specific microdomain at the plasma membrane. The novel punctate 'membrane compartment occupied by Wsc1' (MCW) shows moderate overlap with other Wsc-type sensors, but not with those of the Mid-type sensors or other established plasma membrane domains. We further observed that sensor density and formation of the MCW compartment depends on the cysteine-rich head group near the N-terminus of Wsc1. Yet, signalling capacity depends more on the sensor density in the plasma membrane than on clustering within its microcompartment. We propose that the MCW microcompartment provides a quality control mechanism for retaining functional sensors at the plasma membrane to prevent them from endocytosis.

  15. 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.

  16. Dusty plasma liquid: structure and transfer phenomena

    Energy Technology Data Exchange (ETDEWEB)

    Fortov, Vladimir E; Vaulina, Olga S; Petrov, Oleg F [Institute for High Energy Densities, Russian Academy of Sciences, Izhorskaya 13/19, Moscow (Russian Federation)

    2005-12-15

    Results are given of the experimental investigation of three-particle correlation for liquid plasma-dust structures formed in the electrode layer of a capacitive rf discharge. The obtained three-particle correlation functions for experimental and numerical data are analysed and compared with the superposition approximation. The forming of clusters of macroparticles in plasma-dust systems being analysed is revealed. The experiments in heat transfer were performed in plasma of a capacitive radio-frequency (rf) discharge in argon (P {approx} 20 Pa) with particles 4 {mu}m in mean diameter. The results are given of an experimental investigation of processes of heat transfer for fluid dust structures in rf-discharge. The analysis of steady-state, and unsteady-state heat transfer are used to obtain the thermal conductivity and diffusivity constants.

  17. Effects of n-3 PUFAs on breast cancer cells through their incorporation in plasma membrane

    Directory of Open Access Journals (Sweden)

    Berra Bruno

    2011-05-01

    Full Text Available Abstract Background PUFAs are important molecules for membrane order and function; they can modify inflammation-inducible cytokines production, eicosanoid production, plasma triacylglycerol synthesis and gene expression. Recent studies suggest that n-3 PUFAs can be cancer chemopreventive, chemosuppressive and auxiliary agents for cancer therapy. N-3 PUFAs could alter cancer growth influencing cell replication, cell cycle, and cell death. The question that remains to be answered is how n-3 PUFAs can affect so many physiological processes. We hypothesize that n-3 PUFAs alter membrane stability, modifying cellular signalling in breast cancer cells. Methods Two lines of human breast cancer cells characterized by different expression of ER and EGFR receptors were treated with AA, EPA or DHA. We have used the MTT viability test and expression of apoptotic markers to evaluate the effect of PUFAs on cancer growth. Phospholipids were analysed by HPLC/GC, to assess n-3 incorporation into the cell membrane. Results We have observed that EPA and DHA induce cell apoptosis, a reduction of cell viability and the expression of Bcl2 and procaspase-8. Moreover, DHA slightly reduces the concentration of EGFR but EPA has no effect. Both EPA and DHA reduce the activation of EGFR. N-3 fatty acids are partially metabolized in both cell lines; AA is integrated without being further metabolized. We have analysed the fatty acid pattern in membrane phospholipids where they are incorporated with different degrees of specificity. N-3 PUFAs influence the n-6 content and vice versa. Conclusions Our results indicate that n-3 PUFA feeding might induce modifications of breast cancer membrane structure that increases the degree of fatty acid unsaturation. This paper underlines the importance of nutritional factors on health maintenance and on disease prevention.

  18. Enhancement of proton conductivity of sulfonated polystyrene membrane prepared by plasma polymerization process

    Indian Academy of Sciences (India)

    Bhabesh Kumar Nath; Aziz Khan; Joyanti Chutia; Arup Ratan Pal; Heremba Bailung; Neelotpal Sen Sarma; Devasish Chowdhury; Nirab Chandra Adhikary

    2014-12-01

    This work reports the achievement of higher proton conductivity of polystyrene based proton exchange membrane synthesized in a continuous RF plasma polymerization process using two precursors, styrene (C8H8) and trifluoromethane sulfonic acid (CF3SO3H). The chemical composition of the developed membranes is investigated using Fourier transform infrared spectroscopy and energy dispersive spectroscopy. Scanning electron microscopy has been used for the study of surface morphology and thickness measurement of the membrane. The membranes deposited in the power range from 0.114 to 0.318 Wcm-2 exhibit a lot of variation in the properties like proton transport, water uptake, sulfonation rate, ion exchange capacity and thermal behaviour. The proton conductivity of the membranes is achieved up to 0.6 Scm-1, measured with the help of potentiostat/galvanostat. The thermogravimetric study of the plasma polymerized membrane shows the thermal stability up to 140 °C temperature.

  19. Normal chemotaxis in Dictyostelium discoideum cells with a depolarized plasma membrane potential

    NARCIS (Netherlands)

    Duijn, Bert van; Vogelzang, Sake A.; Ypey, Dirk L.; Molen, Loek G. van der; Haastert, Peter J.M. van

    1990-01-01

    We examined a possible role for the plasma membrane potential in signal transduction during cyclic AMP-induced chemotaxis in the cellular slime mold Dictyostelium discoideum. Chemotaxis, cyclic GMP and cyclic AMP responses in cells with a depolarized membrane potential were measured. Cells can be

  20. Why Do Some T Cell Receptor Cytoplasmic Domains Associate with the Plasma Membrane?

    OpenAIRE

    Philip Anton evan der Merwe; Hao eZhang; Shaun-Paul eCordoba

    2012-01-01

    Based on studies in model systems it has been proposed that the cytoplasmic domains of T cell receptor signaling subunits that have polybasic motifs associate with the plasma membrane, and that this regulates their phosphorylation. Recent experiments in more physiological systems have confirmed membrane association but raised questions as to its function.

  1. Basic Amino Acid Transport in Plasma Membrane Vesicles of Penicillium chrysogenum

    NARCIS (Netherlands)

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

    1996-01-01

    The characteristics of the basic amino acid permease (system VI) of the filamentous fungus Penicillium chrysogenum were studied in plasma membranes fused with liposomes containing the beef heart mitochondrial cytochrome c oxidase. In the presence of reduced cytochrome c, the hybrid membranes accumul

  2. Integrity of the plasma membrane, the acrosomal membrane, and the mitochondrial membrane potential of sperm in Nelore bulls from puberty to sexual maturity

    Directory of Open Access Journals (Sweden)

    L.S.L.S. Reis

    2016-06-01

    Full Text Available ABSTRACT This study evaluated the plasma membrane integrity, acrosomal membrane integrity, and mitochondrial membrane potential of Nelore bull sperm from early puberty to early sexual maturity and their associations with sperm motility and vigor, the mass motility of the spermatozoa (wave motion, scrotal circumference, and testosterone. Sixty Nelore bulls aged 18 to 19 months were divided into four lots (n=15 bulls/lot and evaluated over 280 days. Semen samples, collected every 56 days by electroejaculation, were evaluated soon after collection for motility, vigor and wave motion under an optical microscope. Sperm membrane integrity, acrosomal integrity, and mitochondrial activity were evaluated under a fluorescent microscope using probe association (FITC-PSA, PI, JC-1, H342. The sperm were classified into eight integrity categories depending on whether they exhibited intact or damaged membranes, an intact or damaged acrosomal membrane, and high or low mitochondrial potential. The results show that bulls have a low amount of sperm with intact membranes at puberty, and the sperm show low motility, vigor, and wave motion; however, in bulls at early sexual maturity, the integrity of the sperm membrane increased significantly. The rate of sperm membrane damage was negatively correlated with motility, vigor, wave motion, and testosterone in the bulls, and a positive correlation existed between sperm plasma membrane integrity and scrotal circumference. The integrity of the acrosomal membrane was not influenced by puberty. During puberty and into early sexual maturity, bulls show low sperm mitochondrial potential, but when bulls reached sexual maturity, high membrane integrity with high mitochondrial potential was evident.

  3. Plasma Membranes Modified by Plasma Treatment or Deposition as Solid Electrolytes for Potential Application in Solid Alkaline Fuel Cells

    OpenAIRE

    Christophe Coutanceau; Marc Reinholdt; Jean Durand; Valérie Flaud; Serguei Martemianov; Alina Ilie; Eric Beche; Stéphanie Roualdès; Mauricio Schieda; Jérémy Frugier

    2012-01-01

    In the highly competitive market of fuel cells, solid alkaline fuel cells using liquid fuel (such as cheap, non-toxic and non-valorized glycerol) and not requiring noble metal as catalyst seem quite promising. One of the main hurdles for emergence of such a technology is the development of a hydroxide-conducting membrane characterized by both high conductivity and low fuel permeability. Plasma treatments can enable to positively tune the main fuel cell membrane requirements. In this work, com...

  4. Analysis of tensioned membrane structures considering cable sliding

    Institute of Scientific and Technical Information of China (English)

    宋昌永

    2003-01-01

    In routine design of tensioned membrane st ructures, the membrane is gen erally modeled using space membrane elements and the cables by space cable eleme nts, with no sliding allowed between the membrane and the cables. On the other h and, large deflections are expected and sliding between the membrane and the cab les is inevitable. In the present paper, the general finite element code ABAQUS was employed to investigate the influence of cable sliding on membrane surface o n the structural behavior. Three analysis models were devised to fulfill this pu rpose: (1) The membrane element shares nodes with the cable element; (2) The cab le can slide on the membrane surface freely (without friction) and (3) The cable can slide on the membrane surface, but with friction between the cable and the membrane. The sliding problem is modeled using a surface-based contact algorithm . The results from three analysis models are compared, showing that cable slidin g has only little influence on the structure shape and on the stress distributio ns in the membrane. The main influence of cable sliding may be its effect on the dynamic behavior of tensioned membrane structures.

  5. Towards Enhanced Performance Thin-film Composite Membranes via Surface Plasma Modification

    OpenAIRE

    Rackel Reis; Dumée, Ludovic F.; Tardy, Blaise L.; Raymond Dagastine; John D. Orbell; Jürg A. Schutz; Duke, Mikel C.

    2016-01-01

    Advancing the design of thin-film composite membrane surfaces is one of the most promising pathways to deal with treating varying water qualities and increase their long-term stability and permeability. Although plasma technologies have been explored for surface modification of bulk micro and ultrafiltration membrane materials, the modification of thin film composite membranes is yet to be systematically investigated. Here, the performance of commercial thin-film composite desalination membra...

  6. Protein receptor-independent plasma membrane remodeling by HAMLET

    DEFF Research Database (Denmark)

    Nadeem, Aftab; Sanborn, Jeremy; Gettel, Douglas L.

    2015-01-01

    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...

  7. Parallel artificial liquid membrane extraction of acidic drugs from human plasma

    DEFF Research Database (Denmark)

    Roldan-Pijuan, Mercedes; Pedersen-Bjergaard, Stig; Gjelstad, Astrid

    2015-01-01

    The new sample preparation concept “Parallel artificial liquid membrane extraction (PALME)” was evaluated for extraction of the acidic drugs ketoprofen, fenoprofen, diclofenac, flurbiprofen, ibuprofen, and gemfibrozil from human plasma samples. Plasma samples (250 μL) were loaded into individual...

  8. Tailoring the properties of asymmetric cellulose acetate membranes by gas plasma etching

    NARCIS (Netherlands)

    Olde riekerink, M.B.; Engbers, G.H.M.; Wessling, Matthias; Feijen, Jan

    2002-01-01

    Cellulose triacetate (CTA) ultrafilters and cellulose acetate blend (CAB) desalination membranes were treated with a radiofrequency gas plasma (tetrafluoromethane (CF4) or carbon dioxide (CO2), 47¿49 W, 0.04¿0.08 mbar). Treatment times were varied between 15 s and 120 min. The plasma-treated top

  9. Structural basis for the membrane association of ankyrinG via palmitoylation

    Science.gov (United States)

    Fujiwara, Yuichiro; Kondo, Hiroko X.; Shirota, Matsuyuki; Kobayashi, Megumi; Takeshita, Kohei; Nakagawa, Atsushi; Okamura, Yasushi; Kinoshita, Kengo

    2016-04-01

    By clustering various ion channels and transporters, ankyrin-G (AnkG) configures the membrane-excitation platforms in neurons and cardiomyocytes. AnkG itself localizes to specific areas on the plasma membrane via s-palmitoylation of Cys. However, the structural mechanism by which AnkG anchors to the membrane is not understood. In this study, we solved the crystal structures of the reduced and oxidized forms of the AnkG s-palmitoylation domain and used multiple long-term coarse-grained molecular dynamics simulations to analyze their membrane association. Here we report that the membrane anchoring of AnkG was facilitated by s-palmitoylation, defining a stable binding interface on the lipid membrane, and that AnkG without s-palmitoylation also preferred to stay near the membrane but did not have a unique binding interface. This suggests that AnkG in the juxtamembrane region is primed to accept lipid modification at Cys, and once that happens AnkG constitutes a rigid structural base upon which a membrane-excitation platform can be assembled.

  10. Antidiabetogenic Effects of Chromium Mitigate Hyperinsulinemia-Induced Cellular Insulin Resistance via Correction of Plasma Membrane Cholesterol Imbalance

    OpenAIRE

    Horvath, Emily M.; Tackett, Lixuan; McCarthy, Alicia M.; Raman, Priya; Brozinick, Joseph T.; Elmendorf, Jeffrey S.

    2007-01-01

    Previously, we found that a loss of plasma membrane (PM) phosphatidylinositol 4,5-bisphosphate (PIP2)-regulated filamentous actin (F-actin) structure contributes to insulin-induced insulin resistance. Interestingly, we also demonstrated that chromium picolinate (CrPic), a dietary supplement thought to improve glycemic status in insulin-resistant individuals, augments insulin-regulated glucose transport in insulin-sensitive 3T3-L1 adipocytes by lowering PM cholesterol. Here, to gain mechanisti...

  11. Effects of clary sage oil and its main components, linalool and linalyl acetate, on the plasma membrane of Candida albicans: an in vivo EPR study.

    Science.gov (United States)

    Blaskó, Ágnes; Gazdag, Zoltán; Gróf, Pál; Máté, Gábor; Sárosi, Szilvia; Krisch, Judit; Vágvölgyi, Csaba; Makszin, Lilla; Pesti, Miklós

    2017-02-01

    The effects of clary sage (Salvia sclarea L.) oil (CS-oil), and its two main components, linalool (Lol) and linalyl acetate (LA), on cells of the eukaryotic human pathogen yeast Candida albicans were studied. Dynamic and thermodynamic properties of the plasma membrane were investigated by electron paramagnetic resonance (EPR) spectroscopy, with 5-doxylstearic acid (5-SASL) and 16-SASL as spin labels. The monitoring of the head group regions with 5-SASL revealed break-point frequency decrease in a temperature dependent manner of the plasma membrane between 9.55 and 13.15 °C in untreated, in CS-oil-, Lol- and LA-treated membranes. The results suggest a significant increase in fluidity of the treated plasma membranes close to the head groups. Comparison of the results observed with the two spin labels demonstrated that CS-oil and LA induced an increased level of fluidization at both depths of the plasma membrane. Whereas Lol treatment induced a less (1 %) ordered bilayer organization in the superficial regions and an increased (10 %) order of the membrane leaflet in deeper layers. Acute toxicity tests and EPR results indicated that both the apoptotic and the effects exerted on the plasma membrane fluidity depended on the composition and chemical structure of the examined materials. In comparison with the control, treatment with CS-oil, Lol or LA induced 13.0, 12.3 and 26.4 % loss respectively, of the metabolites absorbing at 260 nm, as a biological consequence of the plasma membrane fluidizing effects. Our results confirmed that clary sage oil causes plasma membrane perturbations which leads to cell apoptosis process.

  12. Lectin receptor kinases participate in protein-protein interactions to mediate plasma membrane-cell wall adhesions in Arabidopsis.

    Science.gov (United States)

    Gouget, Anne; Senchou, Virginie; Govers, Francine; Sanson, Arnaud; Barre, Annick; Rougé, Pierre; Pont-Lezica, Rafael; Canut, Hervé

    2006-01-01

    Interactions between plant cell walls and plasma membranes are essential for cells to function properly, but the molecules that mediate the structural continuity between wall and membrane are unknown. Some of these interactions, which are visualized upon tissue plasmolysis in Arabidopsis (Arabidopsis thaliana), are disrupted by the RGD (arginine-glycine-aspartic acid) tripeptide sequence, a characteristic cell adhesion motif in mammals. In planta induced-O (IPI-O) is an RGD-containing protein from the plant pathogen Phytophthora infestans that can disrupt cell wall-plasma membrane adhesions through its RGD motif. To identify peptide sequences that specifically bind the RGD motif of the IPI-O protein and potentially play a role in receptor recognition, we screened a heptamer peptide library displayed in a filamentous phage and selected two peptides acting as inhibitors of the plasma membrane RGD-binding activity of Arabidopsis. Moreover, the two peptides also disrupted cell wall-plasma membrane adhesions. Sequence comparison of the RGD-binding peptides with the Arabidopsis proteome revealed 12 proteins containing amino acid sequences in their extracellular domains common with the two RGD-binding peptides. Eight belong to the receptor-like kinase family, four of which have a lectin-like extracellular domain. The lectin domain of one of these, At5g60300, recognized the RGD motif both in peptides and proteins. These results imply that lectin receptor kinases are involved in protein-protein interactions with RGD-containing proteins as potential ligands, and play a structural and signaling role at the plant cell surfaces.

  13. Lectin Receptor Kinases Participate in Protein-Protein Interactions to Mediate Plasma Membrane-Cell Wall Adhesions in Arabidopsis1

    Science.gov (United States)

    Gouget, Anne; Senchou, Virginie; Govers, Francine; Sanson, Arnaud; Barre, Annick; Rougé, Pierre; Pont-Lezica, Rafael; Canut, Hervé

    2006-01-01

    Interactions between plant cell walls and plasma membranes are essential for cells to function properly, but the molecules that mediate the structural continuity between wall and membrane are unknown. Some of these interactions, which are visualized upon tissue plasmolysis in Arabidopsis (Arabidopsis thaliana), are disrupted by the RGD (arginine-glycine-aspartic acid) tripeptide sequence, a characteristic cell adhesion motif in mammals. In planta induced-O (IPI-O) is an RGD-containing protein from the plant pathogen Phytophthora infestans that can disrupt cell wall-plasma membrane adhesions through its RGD motif. To identify peptide sequences that specifically bind the RGD motif of the IPI-O protein and potentially play a role in receptor recognition, we screened a heptamer peptide library displayed in a filamentous phage and selected two peptides acting as inhibitors of the plasma membrane RGD-binding activity of Arabidopsis. Moreover, the two peptides also disrupted cell wall-plasma membrane adhesions. Sequence comparison of the RGD-binding peptides with the Arabidopsis proteome revealed 12 proteins containing amino acid sequences in their extracellular domains common with the two RGD-binding peptides. Eight belong to the receptor-like kinase family, four of which have a lectin-like extracellular domain. The lectin domain of one of these, At5g60300, recognized the RGD motif both in peptides and proteins. These results imply that lectin receptor kinases are involved in protein-protein interactions with RGD-containing proteins as potential ligands, and play a structural and signaling role at the plant cell surfaces. PMID:16361528

  14. Identification of Novel Membrane Structures in Plasmodium falciparum Infected Erythrocytes

    Directory of Open Access Journals (Sweden)

    Clavijo Carlos A

    1998-01-01

    Full Text Available Little is known about the molecular mechanisms underlying the release of merozoites from malaria infected erythrocytes. In this study membranous structures present in the culture medium at the time of merozoite release have been characterized. Biochemical and ultrastructural evidence indicate that membranous structures consist of the infected erythrocyte membrane, the parasitophorous vacuolar membrane and a residual body containing electron dense material. These are subcellular compartments expected in a structure that arises as a consequence of merozoite release from the infected cell. Ultrastructural studies show that a novel structure extends from the former parasite compartment to the surface membrane. Since these membrane modifications are detected only after merozoites have been released from the infected erythrocyte, it is proposed that they might play a role in the release of merozoites from the host cell

  15. Sheath Structures of Strongly Electronegative Plasmas

    Institute of Scientific and Technical Information of China (English)

    段萍; 王正汹; 王文春; 刘金远; 刘悦; 王晓钢

    2005-01-01

    The sheath structures of strongly electronegative plasmas are investigated on basis of the accurate Bohm criterion obtained by Sagdeev potential. It is found that the presheath transition between the bulk plasma and the sheath almost does not exist there, and that distributions of electrons, negative and positive ions in the sheath form a pure positive ion sheath near the boundary of the electrode. Furthermore, the density distribution of space net charge has a peak near the sheath edge, the spatial potential within the sheath falls faster, and the sheath thickness becomes thinner.

  16. Theoretical Studies of Long Lived Plasma Structures

    CERN Document Server

    Dvornikov, Maxim

    2010-01-01

    We construct the model of a long lived plasma structure based on spherically symmetric oscillations of electrons in plasma. Oscillations of electrons are studied in frames of both classical and quantum approaches. We obtain the density profile of electrons and the dispersion relations for these oscillations. The differences between classical and quantum approaches are discussed. Then we study the interaction between electrons participating in spherically symmetric oscillations. We find that this interaction can be attractive and electrons can form bound states. The applications of the obtained results to the theory of natural plasmoids are considered.

  17. Surface hydrophobic modification of cellulose membranes by plasma-assisted deposition of hydrocarbon films

    OpenAIRE

    Mudtorlep Nisoa; Pikul Wanichapichart

    2010-01-01

    Surface modification by plasma polymerization is an efficient method to change the surface properties of a membrane. Desirable functionality such as hydrophobicity or hydrophilicity can be obtained, depending on plasma chemistry of gas precursors and discharge conditions. In this work, RF magnetron plasma is produced using acetylene and nitrogen as precursor gases. Variations of RF power, particle flux, deposited time and pressure of the precursor gases have been made to observe coating effec...

  18. 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...

  19. Prostasomes of canine seminal plasma - zinc-binding ability and effects on motility characteristics and plasma membrane integrity of spermatozoa.

    Science.gov (United States)

    Mogielnicka-Brzozowska, M; Strzeżek, R; Wasilewska, K; Kordan, W

    2015-06-01

    Prostasomes are small lipid membrane-confined vesicles that are involved in various fertilization-related processes. The aim of this study was to demonstrate canine seminal plasma prostasomes' ability to bind zinc ions, as well as examining their effects on sperm motility characteristics and plasma membrane integrity during cold storage. Ejaculates, collected from five cross-bred dogs (n = 50), were subjected to ultracentrifugation followed by gel filtration (GF) on a Superose 6 column. Prostasomes appeared as a single fraction in the elution profile. Transmission electron microscopy (TEM) analysis of canine prostasomes revealed the presence of membrane vesicles with diameters ranging from 20.3 to 301 nm. The zinc-affinity chromatography on a Chelating Sepharose Fast Flow - Zn(2 +) showed that from 93 to 100% of the prostasome proteins bind zinc ions (P(+) Zn). SDS-PAGE revealed that canine P(+) Zn comprised four protein bands, with low molecular weights (10.2-12 kDa). We have also shown a positive effect of prostasomes (p spermatozoa with intact plasma membrane (SYBR/PI dual staining) and intact acrosome (Giemsa stained), after 2 h storage at 5°C, was showed, in variant A (1.5% of total seminal plasma protein) and B, when compared with Control and variant C (2.5% of total seminal plasma protein). The prostasomes' effect on motility and plasma membrane integrity of canine cold-stored spermatozoa may be related to their ability to bind zinc ions and regulate their availability to the sperm.

  20. 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....

  1. Investigation of pig sperm plasma membrane reorganization using progesterone-albumin-fluorescein probes

    Institute of Scientific and Technical Information of China (English)

    Alfredo Medrano; Paul F Watson; William V Holt

    2012-01-01

    Objective:To relate semen susceptibility in cooling protocols to sperm plasma membrane properties.Methods:A series of experiments was performed using the fluorescent markers, progesterone-BSA-FITC andBSA-FITC.Results:These experiments indicated that both progesterone-BSA-FITC andBSA-FITC bound to specific sperm plasma membrane domains, thus producing four different binding patterns, revealing probable changes in membrane organization during capacitation and during cooling.Those patterns seem to make a sequence progressing from non-capacitated status to capacitated status.The proportion of each pattern was different during incubation than during cooling, showing the latter had a higher proportion of dead sperm than the former.Conclusions:At this stage, the association of sperm plasma membrane alterations was revealed byBSA-FITC probes and cryosensitivity remains unclear.

  2. Analysis of the induction of the myelin basic protein binding to the plasma membrane phospholipid monolayer

    Science.gov (United States)

    Zhang, Lei; Hao, Changchun; Feng, Ying; Gao, Feng; Lu, Xiaolong; Li, Junhua; Sun, Runguang

    2016-09-01

    Myelin basic protein (MBP) is an essential structure involved in the generation of central nervous system (CNS) myelin. Myelin shape has been described as liquid crystal structure of biological membrane. The interactions of MBP with monolayers of different lipid compositions are responsible for the multi-lamellar structure and stability of myelin. In this paper, we have designed MBP-incorporated model lipid monolayers and studied the phase behavior of MBP adsorbed on the plasma membrane at the air/water interface by thermodynamic method and atomic force microscopy (AFM). By analyzing the pressure-area (π-A) and pressure-time (π-T) isotherms, univariate linear regression equation was obtained. In addition, the elastic modulus, surface pressure increase, maximal insertion pressure, and synergy factor of monolayers were detected. These parameters can be used to modulate the monolayers binding of protein, and the results show that MBP has the strongest affinity for 1,2-dipalmitoyl-sn-glycero-3- phosphoserine (DPPS) monolayer, followed by DPPC/DPPS mixed and 1,2-dipalmitoyl-sn-glycero-3-phospho-choline (DPPC) monolayers via electrostatic and hydrophobic interactions. AFM images of DPPS and DPPC/DPPS mixed monolayers in the presence of MBP (5 nM) show a phase separation texture at the surface pressure of 20 mN/m and the incorporation of MBP put into the DPPC monolayers has exerted a significant effect on the domain structure. MBP is not an integral membrane protein but, due to its positive charge, interacts with the lipid head groups and stabilizes the membranes. The interaction between MBP and phospholipid membrane to determine the nervous system of the disease has a good biophysical significance and medical value. Project supported by the National Natural Science Foundation of China (Grant Nos. 21402114 and 11544009), the Natural Science Basic Research Plan in Shaanxi Province of China (Grant No. 2016JM2010), the Fundamental Research Funds for the Central

  3. 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

  4. 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.

  5. 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.

  6. Molecular structure, function, and dynamics of clathrin-mediated membrane traffic.

    Science.gov (United States)

    Kirchhausen, Tom; Owen, David; Harrison, Stephen C

    2014-05-01

    Clathrin is a molecular scaffold for vesicular uptake of cargo at the plasma membrane, where its assembly into cage-like lattices underlies the clathrin-coated pits of classical endocytosis. This review describes the structures of clathrin, major cargo adaptors, and other proteins that participate in forming a clathrin-coated pit, loading its contents, pinching off the membrane as a lattice-enclosed vesicle, and recycling the components. It integrates as much of the structural information as possible at the time of writing into a sketch of the principal steps in coated-pit and coated-vesicle formation.

  7. Studying Membrane Protein Structure and Function Using Nanodiscs

    DEFF Research Database (Denmark)

    Huda, Pie

    The structure and dynamic of membrane proteins can provide valuable information about general functions, diseases and effects of various drugs. Studying membrane proteins are a challenge as an amphiphilic environment is necessary to stabilise the protein in a functionally and structurally relevan...

  8. Characterization of plasma-induced cell membrane permeabilization: focus on OH radical distribution

    Science.gov (United States)

    Sasaki, Shota; Honda, Ryosuke; Hokari, Yutaro; Takashima, Keisuke; Kanzaki, Makoto; Kaneko, Toshiro

    2016-08-01

    Non-equilibrium atmospheric-pressure plasma (APP) is used medically for plasma-induced cell permeabilization. However, how plasma irradiation specifically triggers permeabilization remains unclear. In an attempt to identify the dominant factor(s), the distribution of plasma-produced reactive species was investigated, primarily focusing on OH radicals. A stronger plasma discharge, which produced more OH radicals in the gas phase, also produced more OH radicals in the liquid phase (OHaq), enhancing the cell membrane permeability. In addition, plasma irradiation-induced enhancement of cell membrane permeability decreased markedly with increased solution thickness (<1 mm), and the plasma-produced OHaq decayed in solution (diffusion length on the order of several hundred micrometers). Furthermore, the horizontally center-localized distribution of OHaq corresponded with the distribution of the permeabilized cells by plasma irradiation, while the overall plasma-produced oxidizing species in solution (detected by iodine-starch reaction) exhibited a doughnut-shaped horizontal distribution. These results suggest that OHaq, among the plasma-produced oxidizing species, represents the dominant factor in plasma-induced cell permeabilization. These results enhance the current understanding of the mechanism of APP as a cell-permeabilization tool.

  9. Gas-liquid interfacial plasmas producing reactive species for cell membrane permeabilization

    Science.gov (United States)

    Kaneko, Toshiro; Sasaki, Shota; Takashima, Keisuke; Kanzaki, Makoto

    2017-01-01

    Gas-liquid interfacial atmospheric-pressure plasma jets (GLI-APPJ) are used medically for plasma-induced cell-membrane permeabilization. In an attempt to identify the dominant factors induced by GLI-APPJ responsible for enhancing cell-membrane permeability, the concentration and distribution of plasma-produced reactive species in the gas and liquid phase regions are measured. These reactive species are classified in terms of their life-span: long-lived (e.g., H2O2), short-lived (e.g., O2•−), and extremely-short-lived (e.g., •OH). The concentration of plasma-produced •OHaq in the liquid phase region decreases with an increase in solution thickness (plasma-induced cell-membrane permeabilization is found to decay markedly as the thickness of the solution increases. Furthermore, the horizontally center-localized distribution of •OHaq, resulting from the center-peaked distribution of •OH in the gas phase region, corresponds with the distribution of the permeabilized cells upon APPJ irradiation, whereas the overall plasma-produced oxidizing species such as H2O2aq in solution exhibit a doughnut-shaped horizontal distribution. These results suggest that •OHaq is likely one of the dominant factors responsible for plasma-induced cell-membrane permeabilization. PMID:28163376

  10. 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.

  11. 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.

  12. 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.

  13. Phosphorylation of plant plasma membrane H+-ATPase by the heterologous host S.cerevisiae

    DEFF Research Database (Denmark)

    L. Rudashevskaya, Elena; Ye, Juanying; Jensen, Ole Nørregaard

     It is known, that phosphorylation of both plant and yeast plasma membrane H+-ATPase results in enzyme activation or inhibition. Several sites at the regulatory C-terminus of the enzyme have been found to undergo phosphorylation in vivo in both plant and yeast. The C-termini of plant H......+-ATPases are app. 60 amino acid residues longer than their yeast homologous. Yeast is found to phosphorylate at least one residue within the plant C-terminus. At the same time a wide range of investigations on structure, function, regulation and interaction of H+-ATPase is carried out with implication......-mutated to alanine residues (to prevent possible phosphorylation) or aspartate residues (to mimic phosphorylation of residue) and the mutated aha2 enzyme expressed in the yeast strain RS-72. Most of the mutations show positive or negative effect on yeast growth in functional complementation assays. It shows in vivo...

  14. Phosphorylation of plant plasma membrane H+-ATPase by the heterologous host S.cerevisiae

    DEFF Research Database (Denmark)

    L. Rudashevskaya, Elena; Ye, Juanying; Jensen, Ole Nørregaard

    +-ATPases are app. 60 amino acid residues longer than their yeast homologous. Yeast is found to phosphorylate at least one residue within the plant C-terminus. At the same time a wide range of investigations on structure, function, regulation and interaction of H+-ATPase is carried out with implication...... It is known, that phosphorylation of both plant and yeast plasma membrane H+-ATPase results in enzyme activation or inhibition. Several sites at the regulatory C-terminus of the enzyme have been found to undergo phosphorylation in vivo in both plant and yeast. The C-termini of plant H...... of heterologous system of yeast cells, expressing plant proton pump. Therefore identification of possible regulatory effects by phosphorylation events in plant H+-ATPase in the system is significant. A number of putative phosphorylation sites at regulatory C-domain of H+-ATPase (AHA2) have been point...

  15. Isothermal titration calorimetric studies on the interaction of the major bovine seminal plasma protein, PDC-109 with phospholipid membranes.

    Directory of Open Access Journals (Sweden)

    V Anbazhagan

    Full Text Available The interaction of the major bovine seminal plasma protein, PDC-109 with lipid membranes was investigated by isothermal titration calorimetry. Binding of the protein to model membranes made up of diacyl phospholipids was found to be endothermic, with positive values of binding enthalpy and entropy, and could be analyzed in terms of a single type of binding sites on the protein. Enthalpies and entropies for binding to diacylphosphatidylcholine membranes increased with increase in temperature, although a clear-cut linear dependence was not observed. The entropically driven binding process indicates that hydrophobic interactions play a major role in the overall binding process. Binding of PDC-109 with dimyristoylphosphatidylcholine membranes containing 25 mol% cholesterol showed an initial increase in the association constant as well as enthalpy and entropy of binding with increase in temperature, whereas the values decreased with further increase in temperature. The affinity of PDC-109 for phosphatidylcholine increased at higher pH, which is physiologically relevant in view of the basic nature of the seminal plasma. Binding of PDC-109 to Lyso-PC could be best analysed in terms of two types of binding interactions, a high affinity interaction with Lyso-PC micelles and a low-affinity interaction with the monomeric lipid. Enthalpy-entropy compensation was observed for the interaction of PDC-109 with phospholipid membranes, suggesting that water structure plays an important role in the binding process.

  16. Structure and structure-preserving algorithms for plasma physics

    Science.gov (United States)

    Morrison, P. J.

    2016-10-01

    Conventional simulation studies of plasma physics are based on numerically solving the underpinning differential (or integro-differential) equations. Usual algorithms in general do not preserve known geometric structure of the physical systems, such as the local energy-momentum conservation law, Casimir invariants, and the symplectic structure (Poincaré invariants). As a consequence, numerical errors may accumulate coherently with time and long-term simulation results may be unreliable. Recently, a series of geometric algorithms that preserve the geometric structures resulting from the Hamiltonian and action principle (HAP) form of theoretical models in plasma physics have been developed by several authors. The superiority of these geometric algorithms has been demonstrated with many test cases. For example, symplectic integrators for guiding-center dynamics have been constructed to preserve the noncanonical symplectic structures and bound the energy-momentum errors for all simulation time-steps; variational and symplectic algorithms have been discovered and successfully applied to the Vlasov-Maxwell system, MHD, and other magnetofluid equations as well. Hamiltonian truncations of the full Vlasov-Maxwell system have opened the field of discrete gyrokinetics and led to the GEMPIC algorithm. The vision that future numerical capabilities in plasma physics should be based on structure-preserving geometric algorithms will be presented. It will be argued that the geometric consequences of HAP form and resulting geometric algorithms suitable for plasma physics studies cannot be adapted from existing mathematical literature but, rather, need to be discovered and worked out by theoretical plasma physicists. The talk will review existing HAP structures of plasma physics for a variety of models, and how they have been adapted for numerical implementation. Supported by DOE DE-FG02-04ER-54742.

  17. Structural Requirements for Membrane Assembly of Proteins Spanning the Membrane Several Times

    OpenAIRE

    Lipp, Joachim; Flint, Nicholas; Haeuptle, Marie-Theres; Dobberstein, Bernhard

    1989-01-01

    We have investigated the structural requirements for the biogenesis of proteins spanning the membrane several times. Proteins containing various combinations of topological signals (signal anchor and stop transfer sequences) were synthesized in a cell-free translation system and their membrane topology was determined. Proteins spanning the membrane twice were obtained when a signal anchor sequence was followed by either a stop transfer sequence or a second signal anchor sequence. Thus, a sig...

  18. Capillary high-performance liquid chromatography/mass spectrometric analysis of proteins from affinity-purified plasma membrane.

    Science.gov (United States)

    Zhao, Yingxin; Zhang, Wei; White, Michael A; Zhao, Yingming

    2003-08-01

    Proteomics analysis of plasma membranes is a potentially powerful strategy for the discovery of proteins involved in membrane remodeling under diverse cellular environments and identification of disease-specific membrane markers. A key factor for successful analysis is the preparation of plasma membrane fractions with low contamination from subcellular organelles. Here we report the characterization of plasma membrane prepared by an affinity-purification method, which involves biotinylation of cell-surface proteins and subsequent affinity enrichment with strepavidin beads. Western blotting analysis showed this method was able to achieve a 1600-fold relative enrichment of plasma membrane versus mitochondria and a 400-fold relative enrichment versus endoplasmic reticulum, two major contaminants in plasma membrane fractions prepared by conventional ultracentrifugation methods. Capillary-HPLC/MS analysis of 30 microg of affinity-purified plasma membrane proteins led to the identification of 918 unique proteins, which include 16.4% integral plasma membrane proteins and 45.5% cytosol proteins (including 8.6% membrane-associated proteins). Notable among the identified membrane proteins include 30 members of ras superfamily, receptors (e.g., EGF receptor, integrins), and signaling molecules. The low number of endoplasmic reticulum and mitochondria proteins (approximately 3.3% of the total) suggests the plasma membrane preparation has minimum contamination from these organelles. Given the importance of integral membrane proteins for drug design and membrane-associated proteins in the regulation cellular behaviors, the described approach will help expedite the characterization of plasma membrane subproteomes, identify signaling molecules, and discover therapeutic membrane-protein targets in diseases.

  19. Chromium(VI)—induces Production of Reactive Oxygen Species,Change of Plasma Membrane Potential and Dissipation of Mitochondria Membrane otential in Chinese Hamster Lung Cell Cultures

    Institute of Scientific and Technical Information of China (English)

    XIEYI; ZHUANGZHI-XIONG

    2001-01-01

    Objective:To examine whether Reactive Oxygen Species(ROS) is generated,and whether plasma membrane potential and mitochnodrial membrane potential are depolarized in Chinese Hamster Lung(CHL)cell lines exposed to Cr(VI),Methods:CHL Cells were incubated with Cr(VI) at 10 umol/L,2.5umol/L,0.65umol/L for 3 and 6 hours,respectively.The rpoduction of ROS was performed by using 2,7-dichlorofluorescin discetate;The changes in plasma membrane potential were performed by using 2,7-dichlorofluorescin discetate;The changes in plasma membrane potential were performed by using 2,7-dichlorofluorescin diacetate;The changes in plasma membrane potential were estimated using fluorescent cationic dye DiBAC4;And the changes in mitochondria membrane potential were estimated using fluorescent dye Rhodamine 123,Results:The ROS levels in CHL cells increased in all treated groups compared with the control group(P<0.01);The plasma membrane potential and mitochondrial membrane potential in CHL cells dissipated after incubated with Cr(VI) at 10umol/L for 3 hours and 6 hours(P<0.01),at 2.5umol/L for 6 hours(P<0.01 or 0.05),Conclusion:Cr(VI) causes the dissipation of plasma membrane potential and mitochnodrial membrane otential in CHL cell cultrues,and Cr(VI)-induced ROS may play a role in the injuries.

  20. Elevated cAMP increases aquaporin-3 plasma membrane diffusion

    DEFF Research Database (Denmark)

    Marlar, Saw; Christensen, Eva Arnspang; Koffman, Jennifer Skaarup

    2014-01-01

    Regulated urine concentration takes place in the renal collecting duct upon arginine vasopressin (AVP) stimulation, where subapical vesicles containing aquaporin-2 (AQP2) are inserted into the apical membrane instantly increasing water reabsorption and urine concentration. The reabsorped water ex...

  1. Protein receptor-independent plasma membrane remodeling by HAMLET

    DEFF Research Database (Denmark)

    Nadeem, Aftab; Sanborn, Jeremy; Gettel, Douglas L.;

    2015-01-01

    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...

  2. Characterization of ion-exchange membrane materials: properties vs structure.

    Science.gov (United States)

    Berezina, N P; Kononenko, N A; Dyomina, O A; Gnusin, N P

    2008-06-22

    This review focuses on the preparation, structure and applications of ion-exchange membranes formed from various materials and exhibiting various functions (electrodialytic, perfluorinated sulphocation-exchange and novel laboratory-tested membranes). A number of experimental techniques for measuring electrotransport properties as well as the general procedure for membrane testing are also described. The review emphasizes the relationships between membrane structures, physical and chemical properties and mechanisms of electrochemical processes that occur in charged membrane materials. The water content in membranes is considered to be a key factor in the ion and water transfer and in polarization processes in electromembrane systems. We suggest the theoretical approach, which makes it possible to model and characterize the electrochemical properties of heterogeneous membranes using several transport-structural parameters. These parameters are extracted from the experimental dependences of specific electroconductivity and diffusion permeability on concentration. The review covers the most significant experimental and theoretical research on ion-exchange membranes that have been carried out in the Membrane Materials Laboratory of the Kuban State University. These results have been discussed at the conferences "Membrane Electrochemistry", Krasnodar, Russia for many years and were published mainly in Russian scientific sources.

  3. New membrane structures with proton conducting properties

    DEFF Research Database (Denmark)

    Nørgaard, Casper Frydendal

    Perfluorosulfonic acid membranes (e.g. Nafion®) are the most widely applied electrolytes in Polymer Electrolyte Membrane Fuel Cells (PEMFCs) because of their good chemical stability, mechanical properties and high proton conductivity, when well hydrated. The upper limit of operating temperature...... [1, 2, 3]. Improved fuel cell performance from incorporation of hygroscopic oxides or solid proton conductors (e.g. zirconium phosphates) has been reported. The poster exhibits upcoming work in the field of composite electrolyte membranes at the University of Southern Denmark, combining radiation...

  4. The plasma membrane as a capacitor for energy and metabolism.

    Science.gov (United States)

    Ray, Supriyo; Kassan, Adam; Busija, Anna R; Rangamani, Padmini; Patel, Hemal H

    2016-02-01

    When considering which components of the cell are the most critical to function and physiology, we naturally focus on the nucleus, the mitochondria that regulate energy and apoptotic signaling, or other organelles such as the endoplasmic reticulum, Golgi, ribosomes, etc. Few people will suggest that the membrane is the most critical element of a cell in terms of function and physiology. Those that consider the membrane critical will point to its obvious barrier function regulated by the lipid bilayer and numerous ion channels that regulate homeostatic gradients. What becomes evident upon closer inspection is that not all membranes are created equal and that there are lipid-rich microdomains that serve as platforms of signaling and a means of communication with the intracellular environment. In this review, we explore the evolution of membranes, focus on lipid-rich microdomains, and advance the novel concept that membranes serve as "capacitors for energy and metabolism." Within this framework, the membrane then is the primary and critical regulator of stress and disease adaptation of the cell.

  5. Sheath Structure of an Electronegative Plasma

    Institute of Scientific and Technical Information of China (English)

    王正汹; 刘金远; 邹秀; 刘悦; 王晓钢

    2003-01-01

    We investigate the sheath structure of an electronegative plasma at steady state with the assumptions of cold positive ions and hot negative ions. The modified Bohm criterion is obtained with the Sagdeev potential by introducing a modified ion sound velocity. At the same time the electric potential, net space charge and particles densities in the sheath are analysed in several cases of different temperature ratios of electrons to negative ions and different density ratios of negative ions to positive ions.

  6. 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.

  7. Towards Enhanced Performance Thin-film Composite Membranes via Surface Plasma Modification

    Science.gov (United States)

    Reis, Rackel; Dumée, Ludovic F.; Tardy, Blaise L.; Dagastine, Raymond; Orbell, John D.; Schutz, Jürg A.; Duke, Mikel C.

    2016-07-01

    Advancing the design of thin-film composite membrane surfaces is one of the most promising pathways to deal with treating varying water qualities and increase their long-term stability and permeability. Although plasma technologies have been explored for surface modification of bulk micro and ultrafiltration membrane materials, the modification of thin film composite membranes is yet to be systematically investigated. Here, the performance of commercial thin-film composite desalination membranes has been significantly enhanced by rapid and facile, low pressure, argon plasma activation. Pressure driven water desalination tests showed that at low power density, flux was improved by 22% without compromising salt rejection. Various plasma durations and excitation powers have been systematically evaluated to assess the impact of plasma glow reactions on the physico-chemical properties of these materials associated with permeability. With increasing power density, plasma treatment enhanced the hydrophilicity of the surfaces, where water contact angles decreasing by 70% were strongly correlated with increased negative charge and smooth uniform surface morphology. These results highlight a versatile chemical modification technique for post-treatment of commercial membrane products that provides uniform morphology and chemically altered surface properties.

  8. Grafting of molecularly imprinted polymer to porous polyethylene filtration membranes by plasma polymerization.

    Science.gov (United States)

    Cowieson, D; Piletska, E; Moczko, E; Piletsky, S

    2013-08-01

    An application of plasma-induced grafting of polyethylene membranes with a thin layer of molecularly imprinted polymer (MIP) was presented. High-density polyethylene (HDPE) membranes, "Vyon," were used as a substrate for plasma grafting modification. The herbicide atrazine, one of the most popular targets of the molecular imprinting, was chosen as a template. The parameters of the plasma treatment were optimized in order to achieve a good balance between polymerization and ablation processes. Modified HDPE membranes were characterized, and the presence of the grafted polymeric layer was confirmed based on the observed weight gain, pore size measurements, and infrared spectrometry. Since there was no significant change in the porosity of the modified membranes, it was assumed that only a thin layer of the polymer was introduced on the surface. The experiments on the re-binding of the template atrazine to the membranes modified with MIP and blank polymers were performed. HDPE membranes which were grafted with polymer using continuous plasma polymerization demonstrated the best result which was expressed in an imprinted factor equal to 3, suggesting that molecular imprinting was successfully achieved.

  9. Electron crystallography for structural and functional studies of membrane proteins.

    Science.gov (United States)

    Fujiyoshi, Yoshinori

    2011-01-01

    Membrane proteins are important research targets for basic biological sciences and drug design, but studies of their structure and function are considered difficult to perform. Studies of membrane structures have been greatly facilitated by technological and instrumental advancements in electron microscopy together with methodological advancements in biology. Electron crystallography is especially useful in studying the structure and function of membrane proteins. Electron crystallography is now an established method of analyzing the structures of membrane proteins in lipid bilayers, which resembles their natural biological environment. To better understand the neural system function from a structural point of view, we developed the cryo-electron microscope with a helium-cooled specimen stage, which allows for analysis of the structures of membrane proteins at a resolution higher than 3 Å. This review introduces recent instrumental advances in cryo-electron microscopy and presents some examples of structure analyses of membrane proteins, such as bacteriorhodopsin, water channels and gap junction channels. This review has two objectives: first, to provide a personal historical background to describe how we came to develop the cryo-electron microscope and second, to discuss some of the technology required for the structural analysis of membrane proteins based on cryo-electron microscopy.

  10. Improvement of alcoholic fermentation by calcium ions under enological conditions involves the increment of plasma membrane H(+)-ATPase activity.

    Science.gov (United States)

    Li, Jingyuan; Huang, Weidong; Wang, Xiuqin; Tang, Tian; Hua, Zhaozhe; Yan, Guoliang

    2010-07-01

    The effect of Ca(2+) on alcoholic fermentation and plasma membrane H(+)-ATPase activity of wine yeast under enological conditions were investigated in this study. The results showed that fermentation rate, cell growth and ethanol production were improved by 0.5 and 1.5 mM Ca(2+) supplementation, which correlated well with the increment of ATPase activity and protein levels. Considering the important role of ATPase in the tolerance of yeast to ethanol, the improvement could be, at least partially, attributed to the increment of ATPase activity. No activation of ATPase by Ca(2+) was observed in the early phase of fermentation and the increment of activity was only observed when ethanol concentration exceeded 6.5%. Therefore, the enhancement of ATPase activity by Ca(2+) was ascribed to alleviating the inhibition of ATPase activity by ethanol through protection of membrane structure. Our results suggest that, besides maintenance of cell membrane structure, the increment of plasma membrane ATPase activity was also responsible for the improvement of alcoholic fermentation by Ca(2+) supplementation.

  11. A role for eisosomes in maintenance of plasma membrane phosphoinositide levels.

    Science.gov (United States)

    Fröhlich, Florian; Christiano, Romain; Olson, Daniel K; Alcazar-Roman, Abel; DeCamilli, Pietro; Walther, Tobias C

    2014-09-15

    The plasma membrane delineates the cell and mediates its communication and material exchange with the environment. Many processes of the plasma membrane occur through interactions of proteins with phosphatidylinositol(4,5)-bisphosphate (PI(4,5)P2), which is highly enriched in this membrane and is a key determinant of its identity. Eisosomes function in lateral organization of the plasma membrane, but the molecular function of their major protein subunits, the BAR domain-containing proteins Pil1 and Lsp1, is poorly understood. Here we show that eisosomes interact with the PI(4,5)P2 phosphatase Inp51/Sjl1, thereby recruiting it to the plasma membrane. Pil1 is essential for plasma membrane localization and function of Inp51 but not for the homologous phosphatidylinositol bisphosphate phosphatases Inp52/Sjl2 and Inp53/Sjl3. Consistent with this, absence of Pil1 increases total and available PI(4,5)P2 levels at the plasma membrane. On the basis of these findings, we propose a model in which the eisosomes function in maintaining PI(4,5)P2 levels by Inp51/Sjl1 recruitment. © 2014 Fröhlich 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).

  12. Aqueous two-phase partition applied to the isolation of plasma membranes and Golgi apparatus from cultured mammalian cells

    Science.gov (United States)

    Morre, D. M.; Morre, D. J.

    2000-01-01

    Partitioning in dextran-poly(ethylene)glycol (PEG) aqueous-aqueous phase systems represents a mature technology with many applications to separations of cells and to the preparation of membranes from mammalian cells. Most applications to membrane isolation and purification have focused on plasma membranes, plasma membrane domains and separation of right side-out and inside-out plasma membrane vesicles. The method exploits a combination of membrane properties, including charge and hydrophobicity. Purification is based upon differential distributions of the constituents in a sample between the two principal compartments of the two phases (upper and lower) and at the interface. The order of affinity of animal cell membranes for the upper phase is: endoplasmic reticulum Golgi apparatusGolgi apparatus from transformed mammalian cells that combines aqueous two-phase partition and centrifugation. Also described is a periodic NADH oxidase, a new enzyme marker for right side-out plasma membrane vesicles not requiring detergent disruptions for measurement of activity.

  13. STRUCTURE AND PROPERTIES OF COMPOSITE POLYURETHANE HOLLOW FIBER MEMBRANES

    Institute of Scientific and Technical Information of China (English)

    Xian-feng Li; Chang-fa Xiao

    2005-01-01

    Composite polyurethane (PU)-SiO2 hollow fiber membranes were successfully prepared via optimizing the technique of dry-jet wet spinning, and their pressure-responsibilities were confirmed by the relationships of pure water fluxtransmembrane pressure (PWF-TP) for the first time. The origin for this phenomenon was analyzed on the basis of membrane structure and material characteristics. The effects of SiO2 content on the structure and properties of membrane were investigated. The experimental results indicated that SiO2 in membrane created a great many interfacial micro-voids and played an important role in pressure-responsibility, PWF and rejection of membrane: with the increase of SiO2 content, the ability of membrane recovery weakened, PWF increased, and rejection decreased slightly.

  14. 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

  15. 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.

  16. Denitrification by plant roots? New aspects of plant plasma membrane-bound nitrate reductase.

    Science.gov (United States)

    Eick, Manuela; Stöhr, Christine

    2012-10-01

    A specific form of plasma membrane-bound nitrate reductase in plants is restricted to roots. Two peptides originated from plasma membrane integral proteins isolated from Hordeum vulgare have been assigned as homologues to the subunit NarH of respiratory nitrate reductase of Escherichia coli. Corresponding sequences have been detected for predicted proteins of Populus trichocarpa with high degree of identities for the subunits NarH (75%) and NarG (65%), however, with less accordance for the subunit NarI. These findings coincide with biochemical properties, particularly in regard to the electron donors menadione and succinate. Together with the root-specific and plasma membrane-bound nitrite/NO reductase, nitric oxide is produced under hypoxic conditions in the presence of nitrate. In this context, a possible function in nitrate respiration of plant roots and an involvement of plants in denitrification processes are discussed.

  17. Ubiquitination regulates the plasma membrane expression of renal UT-A urea transporters.

    Science.gov (United States)

    Stewart, Gavin S; O'Brien, Jennifer H; Smith, Craig P

    2008-07-01

    The renal UT-A urea transporters UT-A1, UT-A2, and UT-A3 are known to play an important role in the urinary concentrating mechanism. The control of the cellular localization of UT-A transporters is therefore vital to overall renal function. In the present study, we have investigated the effect of ubiquitination on UT-A plasma membrane expression in Madin-Darby canine kidney (MDCK) cell lines expressing each of the three renal UT-A transporters. Inhibition of the ubiquitin-proteasome pathway caused an increase in basal transepithelial urea flux across MDCK-rat (r)UT-A1 and MDCK-mouse (m)UT-A2 monolayers (P UT-A transporter expression in the plasma membrane (P UT-A3 expression in the plasma membrane (P UT-A urea transporters, but that this is not the mechanism primarily used by vasopressin to produce its physiological effects.

  18. Preferential transfer of certain plasma membrane proteins onto T and B cells by trogocytosis.

    Directory of Open Access Journals (Sweden)

    Sandrine Daubeuf

    Full Text Available T and B cells capture antigens via membrane fragments of antigen presenting cells (APC in a process termed trogocytosis. Whether (and how a preferential transfer of some APC components occurs during trogocytosis is still largely unknown. We analyzed the transfer onto murine T and B cells of a large panel of fluorescent proteins with different intra-cellular localizations in the APC or various types of anchors in the plasma membrane (PM. Only the latter were transferred by trogocytosis, albeit with different efficiencies. Unexpectedly, proteins anchored to the PM's cytoplasmic face, or recruited to it via interaction with phosphinositides, were more efficiently transferred than those facing the outside of the cell. For proteins spanning the PM's whole width, transfer efficiency was found to vary quite substantially, with tetraspanins, CD4 and FcRgamma found among the most efficiently transferred proteins. We exploited our findings to set immunodiagnostic assays based on the capture of preferentially transferred components onto T or B cells. The preferential transfer documented here should prove useful in deciphering the cellular structures involved in trogocytosis.

  19. An early nodulin-like protein accumulates in the sieve element plasma membrane of Arabidopsis

    DEFF Research Database (Denmark)

    Khan, Junaid A.; Wang, Qi; Sjölund, Richard D.

    2007-01-01

    Membrane proteins within the sieve element-companion cell complex have essential roles in the physiological functioning of the phloem. The monoclonal antibody line RS6, selected from hybridomas raised against sieve elements isolated from California shield leaf (Streptanthus tortuosus; Brassicaceae......) tissue cultures, recognizes an antigen in the Arabidopsis (Arabidopsis thaliana) ecotype Columbia that is associated specifically with the plasma membrane of sieve elements, but not companion cells, and accumulates at the earliest stages of sieve element differentiation. The identity of the RS6 antigen...... from the precursor protein, resulting in a mature peptide of approximately 15 kD that is attached to the sieve element plasma membrane via a carboxy-terminal glycosylphosphatidylinositol membrane anchor. Many of the Arabidopsis ENOD-like proteins accumulate in gametophytic tissues, whereas in both...

  20. Ultrastructural and immunohistochemical localization of plasma membrane Ca2+-ATPase 4 in Ca2+-transporting epithelia

    DEFF Research Database (Denmark)

    Alexander, R Todd; Beggs, Megan R; Zamani, Reza

    2015-01-01

    Plasma Membrane Ca(2+)-ATPase's (PMCA) participate in epithelial Ca(2+) transport and intracellular Ca(2+) signaling. The Pmca4 isoform is enriched in distal nephron isolates and decreased in mice lacking the epithelial Ca(2+) channel, Trpv5. We therefore hypothesized that Pmca4 plays a significant...... in distal nephron cells at both the basolateral membrane and intracellular perinuclear compartments, but not submembranous vesicles, suggesting rapid trafficking to the plasma membrane is unlikely to occur in vivo. Pmca4 expression was not altered by perturbations in Ca(2+) balance, pointing...... detected Pmca1 in lateral membranes of enterocytes. In kidney, Pmca4 showed broad localization to the distal nephron. In mouse, expression was most abundant in segments coexpressing the epithelial Ca(2+) channel, Trpv5. Significant, albeit lower expression, was also evident in the region encompassing...

  1. Study and prediction of secondary structure for membrane proteins

    NARCIS (Netherlands)

    Amirova, Svetlana R.; Milchevsky, Juri V.; Filatov, Ivan V.; Esipova, Natalia G.; Tumanyan, Vladimir G.

    2007-01-01

    In this paper we present a novel approach to membrane protein secondary structure prediction based on the statistical stepwise discriminant analysis method. A new aspect of our approach is the possibility to derive physical -chemical properties that may affect the formation of membrane protein secon

  2. Nidovirus replication structures : hijacking membranes to support viral RNA synthesis

    NARCIS (Netherlands)

    Knoops, Kèvin

    2011-01-01

    Positive-stranded RNA viruses replicate in the cytoplasm of host cells and their replication complexes are associated with modified cell membranes. We investigated the structure of the nidovirus-induced membrane modifications and found that nidoviruses transform the endoplasmic reticulum into a reti

  3. Structural basis for lipopolysaccharide insertion in the bacterial outer membrane.

    Science.gov (United States)

    Qiao, Shuai; Luo, Qingshan; Zhao, Yan; Zhang, Xuejun Cai; Huang, Yihua

    2014-07-03

    One of the fundamental properties of biological membranes is the asymmetric distribution of membrane lipids. In Gram-negative bacteria, the outer leaflet of the outer membrane is composed predominantly of lipopolysaccharides (LPS). The export of LPS requires seven essential lipopolysaccharide transport (Lpt) proteins to move LPS from the inner membrane, through the periplasm to the surface. Of the seven Lpt proteins, the LptD-LptE complex is responsible for inserting LPS into the external leaflet of the outer membrane. Here we report the crystal structure of the ∼110-kilodalton membrane protein complex LptD-LptE from Shigella flexneri at 2.4 Å resolution. The structure reveals an unprecedented two-protein plug-and-barrel architecture with LptE embedded into a 26-stranded β-barrel formed by LptD. Importantly, the secondary structures of the first two β-strands are distorted by two proline residues, weakening their interactions with neighbouring β-strands and creating a potential portal on the barrel wall that could allow lateral diffusion of LPS into the outer membrane. The crystal structure of the LptD-LptE complex opens the door to new antibiotic strategies targeting the bacterial outer membrane.

  4. Structure and Water Transport in Nafion Nanocomposite Membranes

    Science.gov (United States)

    Davis, Eric; Page, Kirt

    2014-03-01

    Perfluorinated ionomers, specifically Nafion, are the most widely used ion exchange membranes for vanadium redox flow battery applications, where an understanding of the relationship between membrane structure and transport of water/ions is critical to battery performance. In this study, the structure of Nafion/SiO2 nanocomposite membranes, synthesized using sol-gel chemistry, as well as cast directly from Nafion/SiO2 nanoparticle dispersions, was measured using both small-angle neutron scattering (SANS) and ultra-small-angle neutron scattering (USANS). Through contrast match studies of the SiO2 nanoparticles, direct information on the change in the structure of the Nafion membranes and the ion-transport channels within was obtained, where differences in membrane structure was observed between the solution-cast membranes and the membranes synthesized using sol-gel chemistry. Additionally, water sorption and diffusion in these Nafion/SiO2 nanocomposite membranes were measured using in situ time-resolved Fourier transform infrared-attenuated total reflectance (FTIR-ATR) spectroscopy and dynamic vapor sorption (DVS).

  5. A Polybasic Plasma Membrane Binding Motif in the I-II Linker Stabilizes Voltage-gated CaV1.2 Calcium Channel Function.

    Science.gov (United States)

    Kaur, Gurjot; Pinggera, Alexandra; Ortner, Nadine J; Lieb, Andreas; Sinnegger-Brauns, Martina J; Yarov-Yarovoy, Vladimir; Obermair, Gerald J; Flucher, Bernhard E; Striessnig, Jörg

    2015-08-21

    L-type voltage-gated Ca(2+) channels (LTCCs) regulate many physiological functions like muscle contraction, hormone secretion, gene expression, and neuronal excitability. Their activity is strictly controlled by various molecular mechanisms. The pore-forming α1-subunit comprises four repeated domains (I-IV), each connected via an intracellular linker. Here we identified a polybasic plasma membrane binding motif, consisting of four arginines, within the I-II linker of all LTCCs. The primary structure of this motif is similar to polybasic clusters known to interact with polyphosphoinositides identified in other ion channels. We used de novo molecular modeling to predict the conformation of this polybasic motif, immunofluorescence microscopy and live cell imaging to investigate the interaction with the plasma membrane, and electrophysiology to study its role for Cav1.2 channel function. According to our models, this polybasic motif of the I-II linker forms a straight α-helix, with the positive charges facing the lipid phosphates of the inner leaflet of the plasma membrane. Membrane binding of the I-II linker could be reversed after phospholipase C activation, causing polyphosphoinositide breakdown, and was accelerated by elevated intracellular Ca(2+) levels. This indicates the involvement of negatively charged phospholipids in the plasma membrane targeting of the linker. Neutralization of four arginine residues eliminated plasma membrane binding. Patch clamp recordings revealed facilitated opening of Cav1.2 channels containing these mutations, weaker inhibition by phospholipase C activation, and reduced expression of channels (as quantified by ON-gating charge) at the plasma membrane. Our data provide new evidence for a membrane binding motif within the I-II linker of LTCC α1-subunits essential for stabilizing normal Ca(2+) channel function. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  6. Heterogeneity of Arabinogalactan-Proteins on the Plasma Membrane of Rose Cells.

    Science.gov (United States)

    Serpe, M. D.; Nothnagel, E. A.

    1996-11-01

    Arabinogalactan-proteins (AGPs) have been purified from the plasma membrane of suspension-cultured Paul's Scarlet rose (Rosa sp.) cells. The two most abundant and homogeneous plasma membrane AGP fractions were named plasma membrane AGP1 (PM-AGP1) and plasma membrane AGP2 (PM-AGP2) and had apparent molecular masses of 140 and 217 kD, respectively. Both PM-AGP1 and PM-AGP2 had [beta]-(1-3)-, [beta]-(1,6)-, and [beta]-(1,3,6)-galactopyranosyl residues, predominantly terminal [alpha]-arabinofuranosyl residues, and (1,4)- and terminal glucuronopyranosyl residues. The protein moieties of PM-AGP1 and PM-AGP2 were both rich in hydroxyproline, alanine, and serine, but differed in the abundance of hydroxyproline, which was 1.6 times higher in PM-AGP2 than in PM-AGP1. Another difference was the overall protein content, which was 3.7% (w/w) in PM-AGP1 and 15% in PM-AGP2. As judged by their behavior on reverse-phase chromatography, PM-AGP1 and PM-AGP2 were not more hydrophobic than AGPs from the cell wall or culture medium. In contrast, a minor plasma membrane AGP fraction eluted later on reverse-phase chromatography and was more negatively charged at pH 5 than either PM-AGP1 or PM-AGP2. The more negatively charged fraction contained molecules with a glycosyl composition characteristic of AGPs and included at least two different macromolecules. The results of this investigation indicate that Rosa plasma membrane contains at least four distinct AGPs or AGP-like molecules. These molecules differed from each other in size, charge, hydrophobicity, amino-acyl composition, and/or protein content.

  7. Identification of type-2 phosphatidic acid phosphohydrolase (PAPH-2) in neutrophil plasma membranes.

    Science.gov (United States)

    Boder, E; Taylor, G; Akard, L; Jansen, J; English, D

    1994-11-01

    Plasma membrane phosphatidic acid phosphohydrolase (PAPH) plays an important role in signal transduction by converting phosphatidic acid to diacylglycerol. PAPH-2, a Mg(2+)-independent, detergent-dependent enzyme involved in cellular signal transduction, is reportedly absent from the plasma membranes of neutrophilic leukocytes, a cell that responds to metabolic stimulation with abundant phospholipase D-dependent diacylglycerol generation. The present study was designed to resolve this discrepancy, focusing on the influence of cellular disruption techniques, detergent availability and cation sensitivity on the apparent distribution of PAPH in neutrophil subcellular fractions. The results clearly indicate the presence of two distinct types of PAPH within the particulate and cytosolic fractions of disrupted cells. Unlike the cytosolic enzyme, the particulate enzymes was not potentiated by magnesium and was strongly detergent-dependent. The soluble and particulate enzymes displayed dissimilar pH profiles. Separation of neutrophil particulate material into fractions rich in plasma membranes, specific granules and azurophilic granules by high speed discontinuous density gradient centrifugation revealed that the majority of the particulate activity was confined to plasma membranes. This activity was not inhibited by pretreatment with n-ethyl-maleimide in concentrations as high as 25 mM. PAPH activity recovered in the cytosolic fraction of disrupted neutrophils was almost completely inhibited by 5.0 mM n-ethylmaleimide. We conclude that resting neutrophils possess n-ethylmaleimide-resistant PAPH (type 2) within their plasma membranes. This enzyme may markedly influence the kinetics of cell activation by metabolizing second messengers generated as a result of activation of plasma membrane phospholipase D.

  8. Duration of ultrasound-mediated enhanced plasma membrane permeability

    NARCIS (Netherlands)

    Lammertink, Bart; Deckers, Roel; Storm, Gert; Moonen, Chrit; Bos, Clemens

    2015-01-01

    Ultrasound (US) induced cavitation can be used to enhance the intracellular delivery of drugs by transiently increasing the cell membrane permeability. The duration of this increased permeability, termed temporal window, has not been fully elucidated. In this study, the temporal window was investiga

  9. Duration of ultrasound-mediated enhanced plasma membrane permeability

    NARCIS (Netherlands)

    Lammertink, Bart; Deckers, Roel; Storm, Gerrit; Moonen, Chrit; Bos, Clemens

    2015-01-01

    Ultrasound (US) induced cavitation can be used to enhance the intracellular delivery of drugs by transiently increasing the cell membrane permeability. The duration of this increased permeability, termed temporal window, has not been fully elucidated. In this study, the temporal window was

  10. New membrane structures with proton conducting properties

    DEFF Research Database (Denmark)

    Nørgaard, Casper Frydendal

    operating temperatures (>100 °C) are desired as they improve reaction kinetics and reduce the problem of CO poisoning the catalyst, thus allowing reduced noble metal loading in the catalyst layers of the membrane electrode assembly of the fuel cell. Moreover water and heat management can be simplified...... [1, 2, 3]. Improved fuel cell performance from incorporation of hygroscopic oxides or solid proton conductors (e.g. zirconium phosphates) has been reported. The poster exhibits upcoming work in the field of composite electrolyte membranes at the University of Southern Denmark, combining radiation...

  11. DCCD inhibits protein translocation into plasma membrane vesicles from Escherichia coli at two different steps.

    OpenAIRE

    1987-01-01

    In vitro translocation of periplasmic and outer membrane proteins into inverted plasma membrane vesicles from Escherichia coli was completely prevented by the H+-ATPase inhibitor N,N'-dicyclohexylcarbodiimide (DCCD). DCCD was inhibitory to both co- and post-translational translocations, suggesting an involvement of the H+-translocating F1F0-ATPase in either mode of transport. This was verified by (i) the dependence of efficient co-translational translocation upon a low salt, i.e. F1-containin...

  12. Why cholesterol should be found predominantly in the cytoplasmic leaf of the plasma membrane

    CERN Document Server

    Giang, Ha

    2014-01-01

    In the mammalian plasma membrane, cholesterol can translocate rapidly between the exoplasmic and cytoplasmic leaves, and is found predominantly in the latter. We hypothesize that it is drawn to the inner leaf to reduce the bending free energy of the membrane caused by the presence there of phosphatidylethanolamine. Incorporating this mechanism into a model free energy for the bilayer, we calculate that approximately two thirds of the total cholesterol should be in the inner leaf.

  13. 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...

  14. Selective regulation of maize plasma membrane aquaporin trafficking and activity by the SNARE SYP121.

    Science.gov (United States)

    Besserer, Arnaud; Burnotte, Emeline; Bienert, Gerd Patrick; Chevalier, Adrien S; Errachid, Abdelmounaim; Grefen, Christopher; Blatt, Michael R; Chaumont, François

    2012-08-01

    Plasma membrane intrinsic proteins (PIPs) are aquaporins facilitating the diffusion of water through the cell membrane. We previously showed that the traffic of the maize (Zea mays) PIP2;5 to the plasma membrane is dependent on the endoplasmic reticulum diacidic export motif. Here, we report that the post-Golgi traffic and water channel activity of PIP2;5 are regulated by the SNARE (for soluble N-ethylmaleimide-sensitive factor protein attachment protein receptor) SYP121, a plasma membrane resident syntaxin involved in vesicle traffic, signaling, and regulation of K(+) channels. We demonstrate that the expression of the dominant-negative SYP121-Sp2 fragment in maize mesophyll protoplasts or epidermal cells leads to a decrease in the delivery of PIP2;5 to the plasma membrane. Protoplast and oocyte swelling assays showed that PIP2;5 water channel activity is negatively affected by SYP121-Sp2. A combination of in vitro (copurification assays) and in vivo (bimolecular fluorescence complementation, Förster resonance energy transfer, and yeast split-ubiquitin) approaches allowed us to demonstrate that SYP121 and PIP2;5 physically interact. Together with previous data demonstrating the role of SYP121 in regulating K(+) channel trafficking and activity, these results suggest that SYP121 SNARE contributes to the regulation of the cell osmotic homeostasis.

  15. Selective Regulation of Maize Plasma Membrane Aquaporin Trafficking and Activity by the SNARE SYP121[W

    Science.gov (United States)

    Besserer, Arnaud; Burnotte, Emeline; Bienert, Gerd Patrick; Chevalier, Adrien S.; Errachid, Abdelmounaim; Grefen, Christopher; Blatt, Michael R.; Chaumont, François

    2012-01-01

    Plasma membrane intrinsic proteins (PIPs) are aquaporins facilitating the diffusion of water through the cell membrane. We previously showed that the traffic of the maize (Zea mays) PIP2;5 to the plasma membrane is dependent on the endoplasmic reticulum diacidic export motif. Here, we report that the post-Golgi traffic and water channel activity of PIP2;5 are regulated by the SNARE (for soluble N-ethylmaleimide-sensitive factor protein attachment protein receptor) SYP121, a plasma membrane resident syntaxin involved in vesicle traffic, signaling, and regulation of K+ channels. We demonstrate that the expression of the dominant-negative SYP121-Sp2 fragment in maize mesophyll protoplasts or epidermal cells leads to a decrease in the delivery of PIP2;5 to the plasma membrane. Protoplast and oocyte swelling assays showed that PIP2;5 water channel activity is negatively affected by SYP121-Sp2. A combination of in vitro (copurification assays) and in vivo (bimolecular fluorescence complementation, Förster resonance energy transfer, and yeast split-ubiquitin) approaches allowed us to demonstrate that SYP121 and PIP2;5 physically interact. Together with previous data demonstrating the role of SYP121 in regulating K+ channel trafficking and activity, these results suggest that SYP121 SNARE contributes to the regulation of the cell osmotic homeostasis. PMID:22942383

  16. Comparative studies on the soluble and plasma membrane associated nitrate reductase from Cucumis sativus L.

    Directory of Open Access Journals (Sweden)

    Grażyna Kłobus

    2014-02-01

    Full Text Available The biochemical comparison between two forms of nitrate reductase from cucumber roots: the soluble enzyme and the plasma membrane-associated one was made. Soluble nitrate reductase was purified on the blue-Sepharose 4B. The nitrate reductase bound with plasma membranes was isolated from cucumber roots by partition of microsomes in the 6.5% dextran-PEG two phase system. The molecular weight of native enzyme estimated with HPLC was 240 kDa and 114 kDa for the soluble and membrane bounded enzyme, respectively. Temperature induced phase separation in Triton X-114 indicated a huge difference in hydrophobicity of the plasma membrane associated nitrate reductase and soluble form of enzyme. Small differences were observed in partial activities of plasma membrane nitrate reductase and soluble nitrate reductase. Also experiments with polyclonal antiserum raised against the native nitrate reductase showed some differences in the immunological properties of both forms of the nitrate reductase. The above results indicated that in cucumber roots two different forms of the nitrate reductase are present.

  17. 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.

  18. Function of plasma membrane microdomain-associated proteins during legume nodulation.

    Science.gov (United States)

    Qiao, Zhenzhen; Libault, Marc

    2017-08-17

    Plasma membrane microdomains are plasma membrane sub-compartments enriched in sphingolipids and sterols, and composed by a specific set of proteins. They are involved in recognizing signal molecules, transducing these signals, and controlling endocytosis and exocytosis processes. In a recent study, applying biochemical and microscopic methods, we characterized the soybean GmFWL1 protein, a major regulator of soybean nodulation, as a new membrane microdomain-associated protein. Interestingly, upon rhizobia inoculation of the soybean root system, GmFWL1 and one of its interacting partners, GmFLOT2/4, both translocate to the root hair cell tip, the primary site of interaction and infection between soybean and Rhizobium. The role of GmFWL1 as a plasma membrane microdomain-associated protein is also supported by immunoprecipitation assays performed on soybean nodules, which revealed 178 GmFWL1 protein partners including a large number of microdomain-associated proteins such as GmFLOT2/4. In this addendum, we provide additional information about the identity of the soybean proteins repetitively identified as GmFWL1 protein partners. Their function is discussed especially in regard to plant-microbe interactions and microbial symbiosis. This addendum will provide new insights in the role of plasma membrane microdomains in regulating legume nodulation.

  19. Amine Enrichment of Thin-Film Composite Membranes via Low Pressure Plasma Polymerization for Antimicrobial Adhesion.

    Science.gov (United States)

    Reis, Rackel; Dumée, Ludovic F; He, Li; She, Fenghua; Orbell, John D; Winther-Jensen, Bjorn; Duke, Mikel C

    2015-07-15

    Thin-film composite membranes, primarily based on poly(amide) (PA) semipermeable materials, are nowadays the dominant technology used in pressure driven water desalination systems. Despite offering superior water permeation and salt selectivity, their surface properties, such as their charge and roughness, cannot be extensively tuned due to the intrinsic fabrication process of the membranes by interfacial polymerization. The alteration of these properties would lead to a better control of the materials surface zeta potential, which is critical to finely tune selectivity and enhance the membrane materials stability when exposed to complex industrial waste streams. Low pressure plasma was employed to introduce amine functionalities onto the PA surface of commercially available thin-film composite (TFC) membranes. Morphological changes after plasma polymerization were analyzed by SEM and AFM, and average surface roughness decreased by 29%. Amine enrichment provided isoelectric point changes from pH 3.7 to 5.2 for 5 to 15 min of plasma polymerization time. Synchrotron FTIR mappings of the amine-modified surface indicated the addition of a discrete 60 nm film to the PA layer. Furthermore, metal affinity was confirmed by the enhanced binding of silver to the modified surface, supported by an increased antimicrobial functionality with demonstrable elimination of E. coli growth. Essential salt rejection was shown minimally compromised for faster polymerization processes. Plasma polymerization is therefore a viable route to producing functional amine enriched thin-film composite PA membrane surfaces.

  20. Ca2+-Transport through Plasma Membrane as a Test of Auxin Sensitivity

    Directory of Open Access Journals (Sweden)

    Anastasia A. Kirpichnikova

    2014-03-01

    Full Text Available Auxin is one of the crucial regulators of plant growth and development. The discovered auxin cytosolic receptor (TIR1 is not involved in the perception of the hormone signal at the plasma membrane. Instead, another receptor, related to the ABP1, auxin binding protein1, is supposed to be responsible for the perception at the plasma membrane. One of the fast and sensitive auxin-induced reactions is an increase of Ca2+ cytosolic concentration, which is suggested to be dependent on the activation of Ca2+ influx through the plasma membrane. This investigation was carried out with a plasmalemma enriched vesicle fraction, obtained from etiolated maize coleoptiles. The magnitude of Ca2+ efflux through the membrane vesicles was estimated according to the shift of potential dependent fluorescent dye diS-C3-(5. The obtained results showed that during coleoptiles ageing (3rd, 4th and 5th days of seedling etiolated growth the magnitude of Ca2+ efflux from inside-out vesicles was decreased. Addition of ABP1 led to a recovery of Ca2+ efflux to the level of the youngest and most sensitive cells. Moreover, the efflux was more sensitive, responding from 10−8 to 10−6 M 1-NAA, in vesicles containing ABP1, whereas native vesicles showed the highest efflux at 10−6 M 1-NAA. We suggest that auxin increases plasma membrane permeability to Ca2+ and that ABP1 is involved in modulation of this reaction.

  1. Plasma-deposited hybrid silica membranes with a controlled retention of organic bridges

    Energy Technology Data Exchange (ETDEWEB)

    Ngamou, P.H.T.; Creatore, M. [Department of Applied Physics, Eindhoven University of Technology, 5600 MB Eindhoven (Netherlands); Overbeek, J.P.; Kreiter, R.; Van Veen, H.M.; Vente, J.F. [ECN, Energy research Centre of the Netherlands, Petten (Netherlands); Wienk, I.M.; Cuperus, P.F. [SolSep BV, Apeldoorn (Netherlands)

    2013-03-05

    Hybrid organically bridged silica membranes are suitable for energy-efficient molecular separations under harsh industrial conditions. Such membranes can be useful in organic solvent nanofiltration if they can be deposited on flexible, porous and large area supports. Here, we report the proof of concept for applying an expanding thermal plasma to the synthesis of perm-selective hybrid silica films from an organically bridged monomer, 1,2-bis(triethoxysilyl)ethane. This membrane is the first in its class to be produced by plasma enhanced chemical vapor deposition. By tuning the plasma and process parameters, the organic bridging groups could be retained in the separating layer. This way, a defect free film could be made with pervaporation performances of an n-butanol-water mixture comparable with those of conventional ceramic supported membranes made by sol-gel technology (i.e. a water flux of [similar]1.8 kg m'-{sup 2} h{sup -1}, a water concentration in the permeate higher than 98% and a separation factor of >1100). The obtained results show the suitability of expanding thermal plasma as a technology for the deposition of hybrid silica membranes for molecular separations.

  2. 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.

  3. Roles of charged particles and reactive species on cell membrane permeabilization induced by atmospheric-pressure plasma irradiation

    Science.gov (United States)

    Sasaki, Shota; Kanzaki, Makoto; Hokari, Yutaro; Tominami, Kanako; Mokudai, Takayuki; Kanetaka, Hiroyasu; Kaneko, Toshiro

    2016-07-01

    As factors that influence cell membrane permeabilization during direct and indirect atmospheric-pressure plasma irradiation, charged particle influx, superoxide anion radicals (O2 -•), and hydrogen peroxide (H2O2) in plasma-irradiated solution were evaluated. These are the three strong candidate factors and might multiply contribute to cell membrane permeabilization. In particular, a shorter plasma diffusion distance leads to the enhancement of the direct effects such as charged particle influx and further increase cell membrane permeability. In addition, O2 -• dissipates over time (a life span of the order of minutes) in plasma-irradiated water, and the deactivation of a plasma-irradiated solution in term of cell membrane permeabilization occurs in a life span of the same order. These results could promote the understanding of the mechanism of plasma-induced cell membrane permeabilization.

  4. 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.

  5. Plant polyphenols as electron donors for erythrocyte plasma membrane redox system: validation through in silico approach

    OpenAIRE

    Kesharwani, Rajesh Kumar; Singh, Durg Vijay; Misra, Krishna; Rizvi, Syed Ibrahim

    2012-01-01

    Background The plasma membrane redox system (PMRS) has extensively been studied in erythrocytes. The PMRS plays an important role in maintaining plasma redox balance and provides a protective mechanism against oxidative stress. Earlier it was proposed that only NADH or NADPH provided reducing equivalents to PMRS; however, now it is acknowledged that some polyphenols also have the ability to donate reducing equivalents to PMRS. Methods Two different docking simulation softwares, Molegro Virtua...

  6. Sandwich-structured hollow fiber membranes for osmotic power generation

    KAUST Repository

    Fu, Feng Jiang

    2015-11-01

    In this work, a novel sandwich-structured hollow fiber membrane has been developed via a specially designed spinneret and optimized spinning conditions. With this specially designed spinneret, the outer layer, which is the most crucial part of the sandwich-structured membrane, is maintained the same as the traditional dual-layer membrane. The inner substrate layer is separated into two layers: (1) an ultra-thin middle layer comprising a high molecular weight polyvinylpyrrolidone (PVP) additive to enhance integration with the outer polybenzimidazole (PBI) selective layer, and (2) an inner-layer to provide strong mechanical strength for the membrane. Experimental results show that a high water permeability and good mechanical strength could be achieved without the expensive post treatment process to remove PVP which was necessary for the dual-layer pressure retarded osmosis (PRO) membranes. By optimizing the composition, the membrane shows a maximum power density of 6.23W/m2 at a hydraulic pressure of 22.0bar when 1M NaCl and 10mM NaCl are used as the draw and feed solutions, respectively. To our best knowledge, this is the best phase inversion hollow fiber membrane with an outer selective PBI layer for osmotic power generation. In addition, this is the first work that shows how to fabricate sandwich-structured hollow fiber membranes for various applications. © 2015 Elsevier B.V.

  7. Nanodevices based on Membrane-Carbon Nanotube Hybrid Structures

    Science.gov (United States)

    Jin, Hye Jun; Kim, Tae Hyun; Namgung, Seon; Hong, Seunghun; Lee, Sang Hun; Park, Tai Hyun

    2010-03-01

    Proteins in cell membrane have been drawing attention due to their versatile functionalities such as ion transfer for neuronal activity and selective binding for sensory systems. However, it is still very difficult to manipulate and study those proteins because they easily lose their functionalities without lipid membranes. We developed a method to coat lipid membranes containing various functional membrane proteins on single-walled carbon nanotube (swCNT)-based field effect transistors (FETs). In this hybrid structure, the activity of membrane proteins can be monitored by underlying swCNT-FETs, allowing us to easily study the functionalities of membrane proteins. Furthermore, we built advanced devices based on these hybrid structures. For an example, we coated lipid membrane containing `olfactory receptors' on swCNT-FETs, resulting in `bioelectric nose' systems. The bioelectric nose system had high sensitivity and human nose-like selectivity to odorant molecules. This talk will also discuss about the future prospect of these membrane-CNT hybrid structures.

  8. Influence of Low-Energy Ion Irradiation on Plasma MembranePermeability of Cells

    Institute of Scientific and Technical Information of China (English)

    ZHANG Dong-Mei; CUI Fu-Zhai; SUN Su-Qin; LIN You-Bo; TIAN Min-Bo; CHEN Guo-Qiang

    2000-01-01

    Effect of low-energy ion irradiation on plasma membrane permeability has been investigated by using electron spin resonance (ESR) spectroscopy of spin probe technique. The investigated system is plumule cells of wheat (Triticum aestivum L.) seeds implanted by 30keV N+ ions. ESR spectra indicated that plasmalemma permeability is sensitive to low-energyion irradiation. Ion irradiations with increasing fluences up to semi-lethal dose lead to gradual increase in plasmalemma permeability of the plumule cells. The possible factors relevant to the changes in membrane permeability are discussed in relation to the changes in the physical state and chemical nature of membranes.

  9. Resistive interchange modes and plasma flow structures

    Science.gov (United States)

    Paccagnella, Roberto

    2011-10-01

    Interchange modes are ubiquitous in magnetic confinement systems and are likely to determine or influence their transport properties. For example a good agreement between theory predictions for linear interchange modes and experimental results has been found recently in a Reverse Field Pinch device. In this work a set of magneto-hydro-dynamic (MHD) equations that describe the dynamical evolution for the pressure driven interchange modes in a magnetic confinement system are studied. Global and local solutions relevant for tokamaks and Reversed Field Pinches (RFPs) configurations are considered. The emphasis is especially in the characterization of the plasma flow structures associated with the dominant modes.

  10. 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.

  11. Fine-Structured Plasma Flows in Prominences

    Science.gov (United States)

    Panasenco, O.; Velli, M.; Landi, S.

    2008-12-01

    Plasmas in prominences (filaments against the disk) exhibit a very wide spectrum of different kind of motions. Here we analyze the plasma motions inside prominences observed by Hinode/SOT during 2006-2007 with focus on two spectacular examples from 25 April 2007 in Halpha line and 30 November 2006 in CaH line and then carry out some simulations of the possible dynamics. Most filaments are composed of fine threads of similar dimensions rooted in the chromosphere/photosphere. Recent observations of counter-streaming motions together with oscillations along the threads provide strong evidence that the threads are field aligned. To more correctly interpret the nature of observed downward flows of dense and cool plasma as well as the upward dark flows of less dense plasma, we take into account the geometry of the prominence structures and the viewing angle. The dark upflows exhibit turbulent patterns such as vortex formation and shedding that are consistent with the motions predicted by instabilities of the interchange type. Sometimes an appearance of dark motions is generated by dark voids opened in the prominence sheet after initiation of nearby downflow streams, implying mass drainage in the downflows. Based on 304 A observations, there is more filament mass in prominences than is visible in either the Halpha or CaH lines. The source of the downward moving plasma may be located either higher above the visible upper edge of the prominence or on the far end of the prominence spine. The bright downward motions of the more cool and dense plasma may be partly due to the counter-streaming motion along the magnetic fields lines and also to the presence of Rayleigh-Taylor type or ballooning/interchange instabilities in the upper regions of the prominence. Transverse motions of filament threads caused by magnetic instabilities constantly provide the conditions for reconnection in the low part of the corona and the chromosphere. We suggest that the combination of flows along

  12. Membrane Protein Mobility and Orientation Preserved in Supported Bilayers Created Directly from Cell Plasma Membrane Blebs.

    Science.gov (United States)

    Richards, Mark J; Hsia, Chih-Yun; Singh, Rohit R; Haider, Huma; Kumpf, Julia; Kawate, Toshimitsu; Daniel, Susan

    2016-03-29

    Membrane protein interactions with lipids are crucial for their native biological behavior, yet traditional characterization methods are often carried out on purified protein in the absence of lipids. We present a simple method to transfer membrane proteins expressed in mammalian cells to an assay-friendly, cushioned, supported lipid bilayer platform using cell blebs as an intermediate. Cell blebs, expressing either GPI-linked yellow fluorescent proteins or neon-green fused transmembrane P2X2 receptors, were induced to rupture on glass surfaces using PEGylated lipid vesicles, which resulted in planar supported membranes with over 50% mobility for multipass transmembrane proteins and over 90% for GPI-linked proteins. Fluorescent proteins were tracked, and their diffusion in supported bilayers characterized, using single molecule tracking and moment scaling spectrum (MSS) analysis. Diffusion was characterized for individual proteins as either free or confined, revealing details of the local lipid membrane heterogeneity surrounding the protein. A particularly useful result of our bilayer formation process is the protein orientation in the supported planar bilayer. For both the GPI-linked and transmembrane proteins used here, an enzymatic assay revealed that protein orientation in the planar bilayer results in the extracellular domains facing toward the bulk, and that the dominant mode of bleb rupture is via the "parachute" mechanism. Mobility, orientation, and preservation of the native lipid environment of the proteins using cell blebs offers advantages over proteoliposome reconstitution or disrupted cell membrane preparations, which necessarily result in significant scrambling of protein orientation and typically immobilized membrane proteins in SLBs. The bleb-based bilayer platform presented here is an important step toward integrating membrane proteomic studies on chip, especially for future studies aimed at understanding fundamental effects of lipid interactions

  13. Coherent structures in two-dimensional plasma turbulence

    DEFF Research Database (Denmark)

    Huld, T.; Nielsen, A.H.; Pécseli, H.L.;

    1991-01-01

    -band turbulent fluctuations is demonstrated by a conditional sampling technique. Depending on plasma parameters, the dominant structures can appear as monopole or multipole vortices, dipole vortices in particular. The importance of large structures for the turbulent plasma diffusion is discussed. A statistical...... analysis of the randomly varying plasma flux is presented....

  14. A conserved signaling network monitors delivery of sphingolipids to the plasma membrane in budding yeast.

    Science.gov (United States)

    Clarke, Jesse; Dephoure, Noah; Horecka, Ira; Gygi, Steven; Kellogg, Douglas

    2017-08-09

    In budding yeast, cell cycle progression and ribosome biogenesis are dependent upon plasma membrane growth, which ensures that events of cell growth are coordinated with each other and with the cell cycle. However, the signals that link the cell cycle and ribosome biogenesis to membrane growth are poorly understood. Here, we used proteome-wide mass spectrometry to systematically discover signals associated with membrane growth. The results suggest that membrane trafficking events required for membrane growth generate sphingolipid-dependent signals. A conserved signaling network appears to play an essential role in signaling by responding to delivery of sphingolipids to the plasma membrane. In addition, sphingolipid-dependent signals control phosphorylation of protein kinase C (Pkc1), which plays an essential role in the pathways that link the cell cycle and ribosome biogenesis to membrane growth. Together, these discoveries provide new clues to how growth-dependent signals control cell growth and the cell cycle. © 2017 by The American Society for Cell Biology.

  15. Trans-activity of plasma membrane-associated ganglioside sialyltransferase in mammalian cells.

    Science.gov (United States)

    Vilcaes, Aldo A; Demichelis, Vanina Torres; Daniotti, Jose L

    2011-09-09

    Gangliosides are acidic glycosphingolipids that contain sialic acid residues and are expressed in nearly all vertebrate cells. They are synthesized at the Golgi complex by a combination of glycosyltransferase activities followed by vesicular delivery to the plasma membrane, where they participate in a variety of physiological as well as pathological processes. Recently, a number of enzymes of ganglioside anabolism and catabolism have been shown to be associated with the plasma membrane. In particular, it was observed that CMP-NeuAc:GM3 sialyltransferase (Sial-T2) is able to sialylate GM3 at the plasma membrane (cis-catalytic activity). In this work, we demonstrated that plasma membrane-integrated ecto-Sial-T2 also displays a trans-catalytic activity at the cell surface of epithelial and melanoma cells. By using a highly sensitive enzyme-linked immunosorbent assay combined with confocal fluorescence microscopy, we observed that ecto-Sial-T2 was able to sialylate hydrophobically or covalently immobilized GM3 onto a solid surface. More interestingly, we observed that ecto-Sial-T2 was able to sialylate GM3 exposed on the membrane of neighboring cells by using both the exogenous and endogenous donor substrate (CMP-N-acetylneuraminic acid) available at the extracellular milieu. In addition, the trans-activity of ecto-Sial-T2 was considerably reduced when the expression of the acceptor substrate was inhibited by using a specific inhibitor of biosynthesis of glycolipids, indicating the lipidic nature of the acceptor. Our findings provide the first direct evidence that an ecto-sialyltransferase is able to trans-sialylate substrates exposed in the plasma membrane from mammalian cells, which represents a novel insight into the molecular events that regulate the local glycosphingolipid composition.

  16. Integral membrane protein structure determination using pseudocontact shifts

    Energy Technology Data Exchange (ETDEWEB)

    Crick, Duncan J.; Wang, Jue X. [University of Cambridge, Department of Biochemistry (United Kingdom); Graham, Bim; Swarbrick, James D. [Monash University, Monash Institute of Pharmaceutical Sciences (Australia); Mott, Helen R.; Nietlispach, Daniel, E-mail: dn206@cam.ac.uk [University of Cambridge, Department of Biochemistry (United Kingdom)

    2015-04-15

    Obtaining enough experimental restraints can be a limiting factor in the NMR structure determination of larger proteins. This is particularly the case for large assemblies such as membrane proteins that have been solubilized in a membrane-mimicking environment. Whilst in such cases extensive deuteration strategies are regularly utilised with the aim to improve the spectral quality, these schemes often limit the number of NOEs obtainable, making complementary strategies highly beneficial for successful structure elucidation. Recently, lanthanide-induced pseudocontact shifts (PCSs) have been established as a structural tool for globular proteins. Here, we demonstrate that a PCS-based approach can be successfully applied for the structure determination of integral membrane proteins. Using the 7TM α-helical microbial receptor pSRII, we show that PCS-derived restraints from lanthanide binding tags attached to four different positions of the protein facilitate the backbone structure determination when combined with a limited set of NOEs. In contrast, the same set of NOEs fails to determine the correct 3D fold. The latter situation is frequently encountered in polytopical α-helical membrane proteins and a PCS approach is thus suitable even for this particularly challenging class of membrane proteins. The ease of measuring PCSs makes this an attractive route for structure determination of large membrane proteins in general.

  17. Proteomic analysis of plasma membrane and secretory vesicles from human neutrophils

    Directory of Open Access Journals (Sweden)

    Campbell Kevin P

    2007-08-01

    Full Text Available Abstract Background Polymorphonuclear neutrophils (PMN constitute an essential cellular component of innate host defense against microbial invasion and exhibit a wide array of responses both to particulate and soluble stimuli. As the cells recruited earliest during acute inflammation, PMN respond rapidly and release a variety of potent cytotoxic agents within minutes of exposure to microbes or their products. PMN rely on the redistribution of functionally important proteins, from intracellular compartments to the plasma membrane and phagosome, as the means by which to respond quickly. To determine the range of membrane proteins available for rapid recruitment during PMN activation, we analyzed the proteins in subcellular fractions enriched for plasma membrane and secretory vesicles recovered from the light membrane fraction of resting PMN after Percoll gradient centrifugation and free-flow electrophoresis purification using mass spectrometry-based proteomics methods. Results To identify the proteins light membrane fractions enriched for plasma membrane vesicles and secretory vesicles, we employed a proteomic approach, first using MALDI-TOF (peptide mass fingerprinting and then by HPLC-MS/MS using a 3D ion trap mass spectrometer to analyze the two vesicle populations from resting PMN. We identified several proteins that are functionally important but had not previously been recovered in PMN secretory vesicles. Two such proteins, 5-lipoxygenase-activating protein (FLAP and dysferlin were further validated by immunoblot analysis. Conclusion Our data demonstrate the broad array of proteins present in secretory vesicles that provides the PMN with the capacity for remarkable and rapid reorganization of its plasma membrane after exposure to proinflammatory agents or stimuli.

  18. Solid-state NMR structures of integral membrane proteins.

    Science.gov (United States)

    Patching, Simon G

    2015-01-01

    Solid-state NMR is unique for its ability to obtain three-dimensional structures and to measure atomic-resolution structural and dynamic information for membrane proteins in native lipid bilayers. An increasing number and complexity of integral membrane protein structures have been determined by solid-state NMR using two main methods. Oriented sample solid-state NMR uses macroscopically aligned lipid bilayers to obtain orientational restraints that define secondary structure and global fold of embedded peptides and proteins and their orientation and topology in lipid bilayers. Magic angle spinning (MAS) solid-state NMR uses unoriented rapidly spinning samples to obtain distance and torsion angle restraints that define tertiary structure and helix packing arrangements. Details of all current protein structures are described, highlighting developments in experimental strategy and other technological advancements. Some structures originate from combining solid- and solution-state NMR information and some have used solid-state NMR to refine X-ray crystal structures. Solid-state NMR has also validated the structures of proteins determined in different membrane mimetics by solution-state NMR and X-ray crystallography and is therefore complementary to other structural biology techniques. By continuing efforts in identifying membrane protein targets and developing expression, isotope labelling and sample preparation strategies, probe technology, NMR experiments, calculation and modelling methods and combination with other techniques, it should be feasible to determine the structures of many more membrane proteins of biological and biomedical importance using solid-state NMR. This will provide three-dimensional structures and atomic-resolution structural information for characterising ligand and drug interactions, dynamics and molecular mechanisms of membrane proteins under physiological lipid bilayer conditions.

  19. Androgen Receptor Localizes to Plasma Membrane by Binding to Caveolin-1 in Mouse Sertoli Cells.

    Science.gov (United States)

    Deng, Qiong; Wu, Yong; Zhang, Zeng; Wang, Yue; Li, Minghua; Liang, Hui; Gui, Yaoting

    2017-01-01

    The nonclassical androgen signaling pathway translates signals into alterations in cellular function within minutes, and this action is proposed to be mediated by an androgen receptor (AR) localized to the plasma membrane. This study was designed to determine the mechanism underlying the membrane association of androgen receptor in TM4 cells, a mouse Sertoli cell line. Western blot analysis indicated testosterone-induced AR translocation to the cell membrane. Data from coimmunoprecipitation indicated that AR is associated with caveolin-1, and testosterone enhanced this association. Knockdown of caveolin-1 by shRNA decreased the amount of AR localized to membrane fraction and prevented AR membrane trafficking after being exposed to testosterone at physiological concentration. The palmitoylation inhibitor 2-bromopalmitate decreased AR membrane localization in basal condition and completely blocked testosterone-induced AR translocation to membrane fraction. These data suggested that AR localized to membrane fraction by binding with caveolin-1 through palmitoylation of the cysteine residue. This study provided a new evidence for AR membrane localization and its application for clarifying the nonclassical signaling pathway of androgens.

  20. Androgen Receptor Localizes to Plasma Membrane by Binding to Caveolin-1 in Mouse Sertoli Cells

    Directory of Open Access Journals (Sweden)

    Qiong Deng

    2017-01-01

    Full Text Available The nonclassical androgen signaling pathway translates signals into alterations in cellular function within minutes, and this action is proposed to be mediated by an androgen receptor (AR localized to the plasma membrane. This study was designed to determine the mechanism underlying the membrane association of androgen receptor in TM4 cells, a mouse Sertoli cell line. Western blot analysis indicated testosterone-induced AR translocation to the cell membrane. Data from coimmunoprecipitation indicated that AR is associated with caveolin-1, and testosterone enhanced this association. Knockdown of caveolin-1 by shRNA decreased the amount of AR localized to membrane fraction and prevented AR membrane trafficking after being exposed to testosterone at physiological concentration. The palmitoylation inhibitor 2-bromopalmitate decreased AR membrane localization in basal condition and completely blocked testosterone-induced AR translocation to membrane fraction. These data suggested that AR localized to membrane fraction by binding with caveolin-1 through palmitoylation of the cysteine residue. This study provided a new evidence for AR membrane localization and its application for clarifying the nonclassical signaling pathway of androgens.

  1. Fine velocity structures collisional dissipation in plasmas

    Science.gov (United States)

    Pezzi, Oreste; Valentini, Francesco; Veltri, Pierluigi

    2016-04-01

    In a weakly collisional plasma, such as the solar wind, collisions are usually considered far too weak to produce any significant effect on the plasma dynamics [1]. However, the estimation of collisionality is often based on the restrictive assumption that the particle velocity distribution function (VDF) shape is close to Maxwellian [2]. On the other hand, in situ spacecraft measurements in the solar wind [3], as well as kinetic numerical experiments [4], indicate that marked non-Maxwellian features develop in the three-dimensional VDFs, (temperature anisotropies, generation of particle beams, ring-like modulations etc.) as a result of the kinetic turbulent cascade of energy towards short spatial scales. Therefore, since collisional effects are proportional to the velocity gradients of the VDF, the collisionless hypothesis may fail locally in velocity space. Here, the existence of several characteristic times during the collisional relaxation of fine velocity structures is investigated by means of Eulerian numerical simulations of a spatially homogeneous force-free weakly collisional plasma. The effect of smoothing out velocity gradients on the evolution of global quantities, such as temperature and entropy, is discussed, suggesting that plasma collisionality can increase locally due to the velocity space deformation of the particle velocity distribution. In particular, by means of Eulerian simulations of collisional relaxation of a spatially homogeneous force-free plasma, in which collisions among particles of the same species are modeled through the complete Landau operator, we show that the system entropy growth occurs over several time scales, inversely proportional to the steepness of the velocity gradients in the VDF. We report clear evidences that fine velocity structures are dissipated by collisions in a time much shorter than global non-Maxwellian features, like, for example, temperature anisotropies. Moreover we indicate that, if small-scale structures

  2. High-resolution Structures of Protein-Membrane Complexes by Neutron Reflection and MD Simulation: Membrane Association of the PTEN Tumor Suppressor

    Science.gov (United States)

    Lösche, Matthias

    2012-02-01

    The lipid matrix of biomembranes is an in-plane fluid, thermally and compositionally disordered leaflet of 5 nm thickness and notoriously difficult to characterize in structural terms. Yet, biomembranes are ubiquitous in the cell, and membrane-bound proteins are implicated in a variety of signaling pathways and intra-cellular transport. We developed methodology to study proteins associated with model membranes using neutron reflection measurements and showed recently that this approach can resolve the penetration depth and orientation of membrane proteins with ångstrom resolution if their crystal or NMR structure is known. Here we apply this technology to determine the membrane bindung and unravel functional details of the PTEN phosphatase, a key player in the PI3K apoptosis pathway. PTEN is an important regulatory protein and tumor suppressor that performs its phosphatase activity as an interfacial enzyme at the plasma membrane-cytoplasm boundary. Acting as an antagonist to phosphoinositide-3-kinase (PI3K) in cell signaling, it is deleted in many human cancers. Despite its importance in regulating the levels of the phosphoinositoltriphosphate PI(3,4,5)P3, there is little understanding of how PTEN binds to membranes, is activated and then acts as a phosphatase. We investigated the structure and function of PTEN by studying its membrane affinity and localization on in-plane fluid, thermally disordered synthetic membrane models. The membrane association of the protein depends strongly on membrane composition, where phosphatidylserine (PS) and phosphatidylinositol diphosphate (PI(4,5)P2) act synergetically in attracting the enzyme to the membrane surface. Membrane affinities depend strongly on membrane fluidity, which suggests multiple binding sites on the protein for PI(4,5)P2. Neutron reflection measurements show that the PTEN phosphatase ``scoots'' along the membrane surface (penetration protein, ˜ 60 å away from the bilayer surface, in a rather compact

  3. Graphene-based structure, method of suspending graphene membrane, and method of depositing material onto graphene membrane

    Science.gov (United States)

    Zettl, Alexander K.; Meyer, Jannik Christian

    2013-04-02

    An embodiment of a method of suspending a graphene membrane across a gap in a support structure includes attaching graphene to a substrate. A pre-fabricated support structure having the gap is attached to the graphene. The graphene and the pre-fabricated support structure are then separated from the substrate which leaves the graphene membrane suspended across the gap in the pre-fabricated support structure. An embodiment of a method of depositing material includes placing a support structure having a graphene membrane suspended across a gap under vacuum. A precursor is adsorbed to a surface of the graphene membrane. A portion of the graphene membrane is exposed to a focused electron beam which deposits a material from the precursor onto the graphene membrane. An embodiment of a graphene-based structure includes a support structure having a gap, a graphene membrane suspended across the gap, and a material deposited in a pattern on the graphene membrane.

  4. High affinity RGD-binding sites at the plasma membrane of Arabidopsis thaliana links the cell wall.

    Science.gov (United States)

    Canut, H; Carrasco, A; Galaud, J P; Cassan, C; Bouyssou, H; Vita, N; Ferrara, P; Pont-Lezica, R

    1998-10-01

    The heptapeptide Tyr-Gly-Arg-Gly-Asp-Ser-Pro containing the sequence Arg-Gly-Asp (RGD--the essential structure recognised by animal cells in substrate adhesion molecules) was tested on epidermal cells of onion and cultured cells of Arabidopsis upon plasmolysis. Dramatic changes were observed on both types of cells following treatment: on onion cells, Hechtian strands linking the cell wall to the membrane were lost, while Arabidopsis cells changed from concave to convex plasmolysis. A control heptapeptide Tyr-Gly-Asp-Gly-Arg-Ser-Pro had no effect on the shape of plasmolysed cells. Protoplasts isolated from Arabidopsis cells agglutinate in the presence of ProNectinF, a genetically engineered protein of 72 kDa containing 13 RGD sequences: several protoplasts may adhere to a single molecule of ProNectinF. The addition of the RGD-heptapeptide disrupted the adhesion between the protoplasts. Purified plasma membrane from Arabidopsis cells exhibits specific binding sites for the iodinated RGD-heptapeptide. The binding is saturable, reversible, and two types of high affinity sites (Kd1 approximately 1 nM, and Kd2 approximately 40 nM) can be discerned. Competitive inhibition by several structurally related peptides and proteins noted the specific requirement for the RGD sequence. Thus, the RGD-binding activity of Arabidopsis fulfils the adhesion features of integrins, i.e. peptide specificity, subcellular location, and involvement in plasma membrane-cell wall attachments.

  5. Soulamarin isolated from Calophyllum brasiliense (Clusiaceae induces plasma membrane permeabilization of Trypanosoma cruzi and mytochondrial dysfunction.

    Directory of Open Access Journals (Sweden)

    Alexandre Rea

    Full Text Available Chagas disease is caused by the parasitic protozoan Trypanosoma cruzi. It has high mortality as well as morbidity rates and usually affects the poorer sections of the population. The development of new, less harmful and more effective drugs is a promising research target, since current standard treatments are highly toxic and administered for long periods. Fractioning of methanol (MeOH extract of the stem bark of Calophyllum brasiliense (Clusiaceae resulted in the isolation of the coumarin soulamarin, which was characterized by one- and two-dimensional (1H- and (13C NMR spectroscopy as well as ESI mass spectrometry. All data obtained were consistent with a structure of 6-hydroxy-4-propyl-5-(3-hydroxy-2-methyl-1-oxobutyl-6″,6″-dimethylpyrane-[2″,3″:8,7]-benzopyran-2-one for soulamarin. Colorimetric MTT assays showed that soulamarin induces trypanocidal effects, and is also active against trypomastigotes. Hemolytic activity tests showed that soulamarin is unable to induce any observable damage to erythrocytes (cmax. = 1,300 µM. The lethal action of soulamarin against T. cruzi was investigated by using amino(4-(6-(amino(iminiomethyl-1H-indol-2-ylphenylmethaniminium chloride (SYTOX Green and 1H,5H,11H,15H-Xantheno[2,3,4-ij:5,6,7-i'j']diquinolizin-18-ium, 9-[4-(chloromethylphenyl]-2,3,6,7,12,13,16,17-octahydro-chloride (MitoTracker Red as fluorimetric probes. With the former, soulamarin showed dose-dependent permeability of the plasma membrane, relative to fully permeable Triton X-100-treated parasites. Spectrofluorimetric and fluorescence microscopy with the latter revealed that soulamarin also induced a strong depolarization (ca. 97% of the mitochondrial membrane potential. These data demonstrate that the lethal action of soulamarin towards T. cruzi involves damages to the plasma membrane of the parasite and mitochondrial dysfunction without the additional generation of reactive oxygen species, which may have also contributed to the

  6. Soulamarin isolated from Calophyllum brasiliense (Clusiaceae) induces plasma membrane permeabilization of Trypanosoma cruzi and mytochondrial dysfunction.

    Science.gov (United States)

    Rea, Alexandre; Tempone, Andre G; Pinto, Erika G; Mesquita, Juliana T; Rodrigues, Eliana; Silva, Luciana Grus M; Sartorelli, Patricia; Lago, João Henrique G

    2013-01-01

    Chagas disease is caused by the parasitic protozoan Trypanosoma cruzi. It has high mortality as well as morbidity rates and usually affects the poorer sections of the population. The development of new, less harmful and more effective drugs is a promising research target, since current standard treatments are highly toxic and administered for long periods. Fractioning of methanol (MeOH) extract of the stem bark of Calophyllum brasiliense (Clusiaceae) resulted in the isolation of the coumarin soulamarin, which was characterized by one- and two-dimensional (1)H- and (13)C NMR spectroscopy as well as ESI mass spectrometry. All data obtained were consistent with a structure of 6-hydroxy-4-propyl-5-(3-hydroxy-2-methyl-1-oxobutyl)-6″,6″-dimethylpyrane-[2″,3″:8,7]-benzopyran-2-one for soulamarin. Colorimetric MTT assays showed that soulamarin induces trypanocidal effects, and is also active against trypomastigotes. Hemolytic activity tests showed that soulamarin is unable to induce any observable damage to erythrocytes (cmax. = 1,300 µM). The lethal action of soulamarin against T. cruzi was investigated by using amino(4-(6-(amino(iminio)methyl)-1H-indol-2-yl)phenyl)methaniminium chloride (SYTOX Green and 1H,5H,11H,15H-Xantheno[2,3,4-ij:5,6,7-i'j']diquinolizin-18-ium, 9-[4-(chloromethyl)phenyl]-2,3,6,7,12,13,16,17-octahydro-chloride (MitoTracker Red) as fluorimetric probes. With the former, soulamarin showed dose-dependent permeability of the plasma membrane, relative to fully permeable Triton X-100-treated parasites. Spectrofluorimetric and fluorescence microscopy with the latter revealed that soulamarin also induced a strong depolarization (ca. 97%) of the mitochondrial membrane potential. These data demonstrate that the lethal action of soulamarin towards T. cruzi involves damages to the plasma membrane of the parasite and mitochondrial dysfunction without the additional generation of reactive oxygen species, which may have also contributed to the death of

  7. Easy measurement of diffusion coefficients of EGFP-tagged plasma membrane proteins using k-space Image Correlation Spectroscopy

    DEFF Research Database (Denmark)

    Christensen, Eva Arnspang; Koffman, Jennifer Skaarup; Marlar, Saw

    2014-01-01

    Lateral diffusion and compartmentalization of plasma membrane proteins are tightly regulated in cells and thus, studying these processes will reveal new insights to plasma membrane protein function and regulation. Recently, k-Space Image Correlation Spectroscopy (kICS)1 was developed to enable ro...

  8. Specific photoaffinity labeling of two plasma membrane polypeptides with an azido auxin

    Science.gov (United States)

    Hicks, G. R.; Rayle, D. L.; Jones, A. M.; Lomax, T. L.

    1989-01-01

    Plasma membrane vesicles were isolated from zucchini (Cucurbita pepo) hypocotyl tissue by aqueous phase partitioning and assessed for homogeneity by the use of membrane-specific enzyme assays. The highly pure (ca. 95%) plasma membrane vesicles maintained a pH differential across the membrane and accumulated a tritiated azido analogue of 3-indoleacetic acid (IAA), 5-azido-[7-3H]IAA ([3H]N3IAA), in a manner similar to the accumulation of [3H]IAA. The association of the [3H]N3IAA with membrane vesicles was saturable and subject to competition by IAA and auxin analogues. Auxin-binding proteins were photoaffinity labeled by addition of [3H]N3IAA to plasma membrane vesicles prior to exposure to UV light (15 sec; 300 nm) and detected by subsequent NaDodSO4/PAGE and fluorography. When the reaction temperature was lowered to -196 degrees C, high-specific-activity labeling of a 40-kDa and a 42-kDa polypeptide was observed. Triton X-100 (0.1%) increased the specific activity of labeling and reduced the background, which suggests that the labeled polypeptides are intrinsic membrane proteins. The labeled polypeptides are of low abundance, as expected for auxin receptors. Further, the addition of IAA and auxin analogues to the photoaffinity reaction mixture resulted in reduced labeling that was qualitatively similar to their effects on the accumulation of radiolabeled IAA in membrane vesicles. Collectively, these results suggest that the radiolabeled polypeptides are auxin receptors. The covalent nature of the label should facilitate purification and further characterization of the receptors.

  9. Evaluation of a new microporous filtration membrane system for therapeutic plasma exchange.

    Science.gov (United States)

    Kurtz, S R; Carey, P M; McGill, M; Pineda, A A; Zaroulis, C G; Case, M T

    1987-01-01

    A new therapeutic plasma exchange device developed by Sarns Inc./3M was evaluated in plasmapheresis of 20 healthy volunteers and in a multicenter clinical study of therapeutic plasma exchange that included 49 patients. Safety and efficacy of plasma separation from whole blood were assessed for a module that contains Durapore microporous surfactant-free polyvinylidene fluoride membrane (Millipore Corp., Bedford, Mass., USA). The extra-corporeal volume was 80 ml. Citrate and heparin anticoagulants were utilized. Mean plasma separation efficiency was 62% with unhindered passage of plasma proteins through the membrane pores and no hemolysis or activation of complement as measured by total hemolytic complement (CH50) and C3 conversion. Mean decrease in platelet count after procedures was 10%. No severe reactions occurred, and citrate effects (13%) were comparable to values reported with centrifugal instruments. The Sarns Inc./3M Therapore device is a rapid, safe and efficient system for plasma exchange and potentially for source plasma collection. The principal benefits are small extracorporeal volume and cell-free filtrate.

  10. Proteomic analysis of liver plasma membrane from hepatitis B surface antigen transgenic mice

    Institute of Scientific and Technical Information of China (English)

    贾小芳

    2012-01-01

    Objective To explore the differential liver plasma membrane( PM) proteins that may be related to the occurrence,development and reversal process of hepatitis and to understand the pathogenesis of hepatitis and the new drug targets by performing a comparative proteomics research of liver PM between

  11. Gateway to understanding microparticles: standardized isolation and identification of plasma membrane-derived vesicles

    NARCIS (Netherlands)

    Dinkla, S.; Brock, R.; Joosten, I.; Bosman, G.J.C.G.M.

    2013-01-01

    Microparticles (MPs) are small plasma membrane-derived vesicles that can expose molecules originating from their parental cells. As vectors of biological information they are likely to play an active role in both homeostasis and pathogenesis, making them promising biomarkers and nanomedicine tools.

  12. Visualization of plasma membrane compartmentalization by high-speed quantum dot tracking

    DEFF Research Database (Denmark)

    Clausen, M. P.; Lagerholm, B. C.

    2013-01-01

    In this study, we have imaged plasma membrane molecules labeled with quantum dots in live cells using a conventional wide-field microscope with high spatial precision at sampling frequencies of 1.75 kHz. Many of the resulting single molecule trajectories are sufficiently long (up to several thous...

  13. Redox enzymes in the plant plasma membrane and their possible roles

    DEFF Research Database (Denmark)

    Berczi, A.; Møller, I.M.

    2000-01-01

    Purified plasma membrane (PM) vesicles from higher plants contain redox proteins with low-molecular-mass prosthetic groups such as flavins (both FMN and FAD), hemes, metals (Cu, Fe and Mn), thiol groups and possibly naphthoquinone (vitamin K-1), all of which are likely to participate in redox...

  14. Double Potential Pulse Chronocoulometry for Detection of Plasma Membrane Cholesterol Efflux at Disk Platinum Microelectrodes

    Science.gov (United States)

    West, Richard H.; Lu, Hui; Shaw, Kendrick; Chiel, Hillel J.; Kelley, Thomas J.; Burgess, James D.

    2016-01-01

    A double potential pulse scheme is reported for observation of cholesterol efflux from the plasma membrane of a single neuron cell. Capillary Pt disk microelectrodes having a thin glass insulator allow the 10 μm diameter electrode and cell to be viewed under optical magnification. The electrode, covalently functionalized with cholesterol oxidase, is positioned in contact with the cell surface resulting in enzyme catalyzed cholesterol oxidation and efflux of cholesterol from the plasma membrane at the electrode contact site. Enzymatically generated hydrogen peroxide accumulates at the electrode/cell interface during a 5 s hold-time and is oxidized during application of a potential pulse. A second, replicate potential pulse is applied 0.5 s after the first potential pulse to gauge background charge prior to significant accumulation of hydrogen peroxide. The difference in charge passed between the first and second potential pulse provides a measure of hydrogen peroxide generated by the enzyme and is an indication of the cholesterol efflux. Control experiments for bare Pt microelectrodes in contact with the cell plasma membrane show difference charge signals in the range of about 7–10 pC. Enzyme-modified electrodes in contact with the plasma membrane show signals in the range of 16–26 pC. PMID:27330196

  15. Potassium as an intrinsic uncoupler of the plasma membrane H+-ATPase

    DEFF Research Database (Denmark)

    Palmgren, Michael Gjedde; Buch-Pedersen, Morten Jeppe

    The plant plasma membrane proton pump (H(+)-ATPase) is stimulated by potassium, but it has remained unclear whether potassium is actually transported by the pump or whether it serves other roles. We now show that K(+) is bound to the proton pump at a site involving Asp(617) in the cytoplasmic...

  16. Method of preparing water purification membranes. [polymerization of allyl amine as thin films in plasma discharge

    Science.gov (United States)

    Hollahan, J. R.; Wydeven, T. J., Jr. (Inventor)

    1974-01-01

    Allyl amine and chemically related compounds are polymerized as thin films in the presence of a plasma discharge. The monomer compound can be polymerized by itself or in the presence of an additive gas to promote polymerization and act as a carrier. The polymerized films thus produced show outstanding advantages when used as reverse osmosis membranes.

  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. Video Views and Reviews: Golgi Export, Targeting, and Plasma Membrane Caveolae

    Science.gov (United States)

    Watters, Christopher

    2004-01-01

    In this article, the author reviews videos from "Molecular Biology of the Cell (MBC)" depicting various aspects of plasma membrane (PM) dynamics, including the targeting of newly synthesized components and the organization of those PM invaginations called caveolae. The papers accompanying these videos describe, respectively, the constitutive…

  19. ADENOSINE-TRIPHOSPHATE DEPENDENT TAUROCHOLATE TRANSPORT IN HUMAN LIVER PLASMA-MEMBRANES

    NARCIS (Netherlands)

    WOLTERS, H; KUIPERS, F; SLOOFF, MJH; VONK, RJ

    1992-01-01

    Transport systems involved in uptake and biliary secretion of bile salts have been extensively studied in rat liver; however, little is known about these systems in the human liver. In this study, we investigated taurocholate (TC) transport in canalicular and basolateral plasma membrane vesicles iso

  20. 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

  1. 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.

  2. 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.

  3. Calcium ion transport across plasma membranes isolated from rat kidney cortex.

    Science.gov (United States)

    Gmaj, P; Murer, H; Kinne, R

    1979-03-15

    Basal-lateral-plasma-membrane vesicles and brush-border-membrane vesicles were isolated from rat kidney cortex by differential centrifugation followed by free-flow-electrophoresis. Ca2+ uptake into these vesicles was investigated by a rapid filtration method. Both membranes show a considerable binding of Ca2+ to the vesicle interior, making the analysis of passive fluxes in uptake experiments difficult. Only the basal-lateral-plasma-membrane vesicles exhibit an ATP-dependent pump activity which can be distinguished from the activity in mitochondrial and endoplasmic reticulum by virtue of the different distribution during free-flow electrophoresis and its lack of sensitivity to oligomycin. The basal-lateral plasma membranes contain in addition a Na+/Ca2+-exchange system which mediates a probably rheogenic counter-transport of Ca2+ and Na+ across the basal cell border. The latter system is probably involved in the secondary active Na+-dependent and ouabain-inhibitable Ca2+ reabsorption in the proximal tubule, the ATP-driven system is probably more important for the maintenance of a low concentration of intracellular Ca2+.

  4. Plasma membrane cholesterol level and agonist-induced internalization of δ-opioid receptors; colocalization study with intracellular membrane markers of Rab family.

    Science.gov (United States)

    Brejchova, Jana; Vosahlikova, Miroslava; Roubalova, Lenka; Parenti, Marco; Mauri, Mario; Chernyavskiy, Oleksandr; Svoboda, Petr

    2016-08-01

    Decrease of cholesterol level in plasma membrane of living HEK293 cells transiently expressing FLAG-δ-OR by β-cyclodextrin (β-CDX) resulted in a slight internalization of δ-OR. Massive internalization of δ-OR induced by specific agonist DADLE was diminished in cholesterol-depleted cells. These results suggest that agonist-induced internalization of δ-OR, which has been traditionally attributed exclusively to clathrin-mediated pathway, proceeds at least partially via membrane domains. Identification of internalized pools of FLAG-δ-OR by colocalization studies with proteins of Rab family indicated the decreased presence of receptors in early endosomes (Rab5), late endosomes and lysosomes (Rab7) and fast recycling vesicles (Rab4). Slow type of recycling (Rab11) was unchanged by cholesterol depletion. As expected, agonist-induced internalization of oxytocin receptors was totally suppressed in β-CDX-treated cells. Determination of average fluorescence lifetime of TMA-DPH, the polar derivative of hydrophobic membrane probe diphenylhexatriene, in live cells by FLIM indicated a significant alteration of the overall PM structure which may be interpreted as an increased "water-accessible space" within PM area. Data obtained by studies of HEK293 cells transiently expressing FLAG-δ-OR by "antibody feeding" method were extended by analysis of the effect of cholesterol depletion on distribution of FLAG-δ-OR in sucrose density gradients prepared from HEK293 cells stably expressing FLAG-δ-OR. Major part of FLAG-δ-OR was co-localized with plasma membrane marker Na,K-ATPase and β-CDX treatment resulted in shift of PM fragments containing both FLAG-δ-OR and Na,K-ATPase to higher density. Thus, the decrease in content of the major lipid constituent of PM resulted in increased density of resulting PM fragments.

  5. GLUT-4 content in plasma membrane of muscle from patients with non-insulin-dependent diabetes mellitus

    DEFF Research Database (Denmark)

    Lund, S; Vestergaard, H; Andersen, P H

    1993-01-01

    The abundance of GLUT-4 protein in both total crude membrane and plasma membrane fractions of vastus lateralis muscle from 13 obese non-insulin-dependent diabetes mellitus (NIDDM) patients and 14 healthy subjects were examined in the fasting state and after supraphysiological hyperinsulinemia....... In the basal state the immunoreactive mass of GLUT-4 protein both in the crude membrane preparation and in the plasma membrane fraction was similar in NIDDM patients and control subjects. Moreover, in vivo insulin exposure neither for 30 min nor for 4 h had any impact on the content of GLUT-4 protein in plasma...... membranes. With the use of the same methodology, antibody, and achieving the same degree of plasma membrane purification and recovery, we found, however, that intraperitoneal administration of insulin to 7-wk-old rats within 30 min increased the content of GLUT-4 protein more than twofold (P

  6. Carbon nanotube embedded PVDF membranes: Effect of solvent composition on the structural morphology for membrane distillation

    Science.gov (United States)

    Mapunda, Edgar C.; Mamba, Bhekie B.; Msagati, Titus A. M.

    2017-08-01

    Rapid population increase, growth in industrial and agricultural sectors and global climate change have added significant pressure on conventional freshwater resources. Tapping freshwater from non-conventional water sources such as desalination and wastewater recycling is considered as sustainable alternative to the fundamental challenges of water scarcity. However, affordable and sustainable technologies need to be applied for the communities to benefit from the treatment of non-conventional water source. Membrane distillation is a potential desalination technology which can be used sustainably for this purpose. In this work multi-walled carbon nanotube embedded polyvinylidene fluoride membranes for application in membrane distillation desalination were prepared via non-solvent induced phase separation method. The casting solution was prepared using mixed solvents (N, N-dimethylacetamide and triethyl phosphate) at varying ratios to study the effect of solvent composition on membrane morphological structures. Membrane morphological features were studied using a number of techniques including scanning electron microscope, atomic force microscope, SAXSpace tensile strength analysis, membrane thickness, porosity and contact angle measurements. It was revealed that membrane hydrophobicity, thickness, tensile strength and surface roughness were increasing as the composition of N, N-dimethylacetamide in the solvent was increasing with maximum values obtained between 40 and 60% N, N-dimethylacetamide. Internal morphological structures were changing from cellular structures to short finger-like and sponge-like pores and finally to large macro void type of pores when the amount of N, N-dimethylacetamide in the solvent was changed from low to high respectively. Multi-walled carbon nanotube embedded polyvinylidene fluoride membranes of desired morphological structures and physical properties can be synthesized by regulating the composition of solvents used to prepare the

  7. Auxiliary Subunits: Shepherding AMPA Receptors to the Plasma Membrane

    Directory of Open Access Journals (Sweden)

    Simon C. Haering

    2014-08-01

    Full Text Available Ionotropic glutamate receptors (iGluRs are tetrameric ligand-gated cation channels that mediate excitatory signal transmission in the central nervous system (CNS of vertebrates. The members of the iGluR subfamily of α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA receptors (AMPARs mediate most of the fast excitatory signal transmission, and their abundance in the postsynaptic membrane is a major determinant of the strength of excitatory synapses. Therefore, regulation of AMPAR trafficking to the postsynaptic membrane is an important constituent of mechanisms involved in learning and memory formation, such as long-term potentiation (LTP and long-term depression (LTD. Auxiliary subunits play a critical role in the facilitation and regulation of AMPAR trafficking and function. The currently identified auxiliary subunits of AMPARs are transmembrane AMPA receptor regulatory proteins (TARPs, suppressor of lurcher (SOL, cornichon homologues (CNIHs, synapse differentiation-induced gene I (SynDIG I, cysteine-knot AMPAR modulating proteins 44 (CKAMP44, and germ cell-specific gene 1-like (GSG1L protein. In this review we summarize our current knowledge of the modulatory influence exerted by these important but still underappreciated proteins.

  8. Structure and membrane interactions of the homodimeric antibiotic peptide homotarsinin

    Science.gov (United States)

    Verly, Rodrigo M.; Resende, Jarbas M.; Junior, Eduardo F. C.; de Magalhães, Mariana T. Q.; Guimarães, Carlos F. C. R.; Munhoz, Victor H. O.; Bemquerer, Marcelo Porto; Almeida, Fábio C. L.; Santoro, Marcelo M.; Piló-Veloso, Dorila; Bechinger, Burkhard

    2017-01-01

    Antimicrobial peptides (AMPs) from amphibian skin are valuable template structures to find new treatments against bacterial infections. This work describes for the first time the structure and membrane interactions of a homodimeric AMP. Homotarsinin, which was found in Phyllomedusa tarsius anurans, consists of two identical cystine-linked polypeptide chains each of 24 amino acid residues. The high-resolution structures of the monomeric and dimeric peptides were determined in aqueous buffers. The dimer exhibits a tightly packed coiled coil three-dimensional structure, keeping the hydrophobic residues screened from the aqueous environment. An overall cationic surface of the dimer assures enhanced interactions with negatively charged membranes. An extensive set of biophysical data allowed us to establish structure-function correlations with antimicrobial assays against Gram-positive and Gram-negative bacteria. Although both peptides present considerable antimicrobial activity, the dimer is significantly more effective in both antibacterial and membrane biophysical assays. PMID:28102305

  9. Structure and membrane interactions of the homodimeric antibiotic peptide homotarsinin

    Science.gov (United States)

    Verly, Rodrigo M.; Resende, Jarbas M.; Junior, Eduardo F. C.; de Magalhães, Mariana T. Q.; Guimarães, Carlos F. C. R.; Munhoz, Victor H. O.; Bemquerer, Marcelo Porto; Almeida, Fábio C. L.; Santoro, Marcelo M.; Piló-Veloso, Dorila; Bechinger, Burkhard

    2017-01-01

    Antimicrobial peptides (AMPs) from amphibian skin are valuable template structures to find new treatments against bacterial infections. This work describes for the first time the structure and membrane interactions of a homodimeric AMP. Homotarsinin, which was found in Phyllomedusa tarsius anurans, consists of two identical cystine-linked polypeptide chains each of 24 amino acid residues. The high-resolution structures of the monomeric and dimeric peptides were determined in aqueous buffers. The dimer exhibits a tightly packed coiled coil three-dimensional structure, keeping the hydrophobic residues screened from the aqueous environment. An overall cationic surface of the dimer assures enhanced interactions with negatively charged membranes. An extensive set of biophysical data allowed us to establish structure-function correlations with antimicrobial assays against Gram-positive and Gram-negative bacteria. Although both peptides present considerable antimicrobial activity, the dimer is significantly more effective in both antibacterial and membrane biophysical assays.

  10. 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.

  11. Inhibition of cell adhesion by xARVCF indicates a regulatory function at the plasma membrane.

    Science.gov (United States)

    Reintsch, Wolfgang E; Mandato, Craig A; McCrea, Pierre D; Fagotto, François

    2008-09-01

    The cytoplasmic tail of cadherins is thought to regulate the strength and dynamics of cell-cell adhesion. Part of its regulatory activity has been attributed to a membrane-proximal region, the juxtamembrane domain (JMD), and its interaction with members of the p120 catenin subfamily. We show that titration of xARVCF, a member of this family, to the plasma membrane disrupts adhesion in the early embryo. Adhesion can be restored by coexpression of constitutively active Rac, suggesting that intracellular signaling is the primary cause in the loss of adhesion phenotype. Our observations suggest that the recruitment of p120 type catenins to the plasma membrane by the cadherin cytoplasmic tail may create protein complexes, which actively modulate the adhesion "status" of embryonic cells.

  12. Low energy plasma treatment of a proton exchange membrane used for low temperature fuel cells

    Energy Technology Data Exchange (ETDEWEB)

    Charles, C [Space Plasma, Power, and Propulsion group, Research School of Physical Sciences and Engineering, Australian National University, Canberra, ACT 0200 (Australia); Ramdutt, D [Space Plasma, Power, and Propulsion group, Research School of Physical Sciences and Engineering, Australian National University, Canberra, ACT 0200 (Australia); Brault, P [GREMI-CNRS Laboratory, University of Orleans, BP 6744, F-45067, Orleans (France); Caillard, A [Space Plasma, Power, and Propulsion group, Research School of Physical Sciences and Engineering, Australian National University, Canberra, ACT 0200 (Australia); Bulla, D [Space Plasma, Power, and Propulsion group, Research School of Physical Sciences and Engineering, Australian National University, Canberra, ACT 0200 (Australia); Boswell, R [Space Plasma, Power, and Propulsion group, Research School of Physical Sciences and Engineering, Australian National University, Canberra, ACT 0200 (Australia); Rabat, H [GREMI-CNRS Laboratory, University of Orleans, BP 6744, F-45067, Orleans (France); Dicks, A [School of Engineering, University of Queensland, Brisbane, QLD 4072 (Australia)

    2007-05-15

    A low energy ({approx}30 V) plasma treatment of Nafion, a commercial proton exchange membrane used for low temperature fuel cells, is performed in a helicon radiofrequency (13.56 MHz) plasma system. For argon densities in the 10{sup 9}-10{sup 10} cm{sup -3} range, the water contact angle (hydrophobicity) of the membrane surface linearly decreases with an increase in the plasma energy dose, which is maintained below 5.1 J cm{sup -2}, and which results from the combination of an ion energy dose (up to 3.8 J cm{sup -2}) and a photon (mostly UV) energy dose (up to 1.3 J cm{sup -2}). The decrease in water contact angle is essentially a result of the energy brought to the surface by ion bombardment. The measured effect of the energy brought to the surface by UV light is found to be negligible.

  13. Low energy plasma treatment of a proton exchange membrane used for low temperature fuel cells

    Science.gov (United States)

    Charles, C.; Ramdutt, D.; Brault, P.; Caillard, A.; Bulla, D.; Boswell, R.; Rabat, H.; Dicks, A.

    2007-05-01

    A low energy (~30 V) plasma treatment of Nafion, a commercial proton exchange membrane used for low temperature fuel cells, is performed in a helicon radiofrequency (13.56 MHz) plasma system. For argon densities in the 109-1010 cm-3 range, the water contact angle (hydrophobicity) of the membrane surface linearly decreases with an increase in the plasma energy dose, which is maintained below 5.1 J cm-2, and which results from the combination of an ion energy dose (up to 3.8 J cm-2) and a photon (mostly UV) energy dose (up to 1.3 J cm-2). The decrease in water contact angle is essentially a result of the energy brought to the surface by ion bombardment. The measured effect of the energy brought to the surface by UV light is found to be negligible.

  14. 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.

  15. Polyethylene glycol acrylate-grafted polysulphone membrane for artificial lungs: plasma modification and haemocompatibility improvement.

    Science.gov (United States)

    Wang, Weiping; Huang, Xin; Yin, Haiyan; Fan, Wenling; Zhang, Tao; Li, Lei; Mao, Chun

    2015-12-14

    In this study, polyethylene glycol acrylate (PEGA) was introduced onto the surface of polysulphone (PSF) membrane to prepare PSF-PEGA membranes through low-temperature plasma technology for haemocompatibility improvement of artificial lungs. The effects of plasma power, PEGA solution concentration and dipcoating temperature on surface modification were systematically investigated. Results of Fourier transform infrared spectroscopy, x-ray photoelectron spectroscopy and PEGA grafting degree confirmed that PEGA was successfully grafted onto the PSF membranes. Contact angle values showed that the hydrophilicity of the PSF-PEGA membrane surface increased by 21.5%. The results of the protein adsorption, platelet adhesion and coagulation tests further showed the excellent haemocompatibility of the modified membrane. Gas exchange tests also revealed that at a porcine blood flow rate of 5 l min(-1), O2 and CO2 exchange rates through the PSF-PEGA membrane were 198.6 and 170.9 ml min(-1), respectively; approximately this is the gas exchange capacity of commercial respiratory assistance devices.

  16. Forward transport of proteins in the plasma membrane of migrating cerebellar granule cells.

    Science.gov (United States)

    Wang, Dong; She, Liang; Sui, Ya-nan; Yuan, Xiao-bing; Wen, Yunqing; Poo, Mu-ming

    2012-12-18

    Directional flow of membrane components has been detected at the leading front of fibroblasts and the growth cone of neuronal processes, but whether there exists global directional flow of plasma membrane components over the entire migrating neuron remains largely unknown. By analyzing the trajectories of antibody-coated single quantum dots (QDs) bound to two membrane proteins, overexpressed myc-tagged synaptic vesicle-associated membrane protein VAMP2 and endogenous neurotrophin receptor TrkB, we found that these two proteins exhibited net forward transport, which is superimposed upon Brownian motion, in both leading and trailing processes of migrating cerebellar granule cells in culture. Furthermore, no net directional transport of membrane proteins was observed in nonmigrating cells with either growing or stalling leading processes. Analysis of the correlation of motion direction between two QDs on the same process in migrating neurons also showed a higher frequency of correlated forward than rearward movements. Such correlated QD movements were markedly reduced in the presence of myosin II inhibitor blebbistatin,suggesting the involvement of myosin II-dependent active transport processes. Thus, a net forward transport of plasma membrane proteins exists in the leading and trailing processes of migrating neurons, in line with the translocation of the soma.

  17. A method to modify PVDF microfiltration membrane via ATRP with low-temperature plasma pretreatment

    Science.gov (United States)

    Han, Yu; Song, Shuijun; Lu, Yin; Zhu, Dongfa

    2016-08-01

    The hydrophilic modification of a polyvinylidene fluoride (PVDF) microfiltration membrane via pretreatment with argon plasma and direct surface-initiated atom transfer radical polymerization (ATRP) was studied. Both modified and unmodified PVDF membranes were characterized by Fourier transform infrared spectroscopy (FTIR), water contact angle, scanning electron microscopy (SEM), X-ray photoelectron spectroscopy (XPS), and pore size distribution measurements. FTIR and XPS spectra confirmed that sulfobetaine methacrylate (SBMA) had been grafted onto the membrane surface. The initial contact angle decreased from 87.0° to 29.8° and a water drop penetrated into the modified membrane completely in 8 s. The pore size distribution of the modified membrane exhibited a smaller mean value than that of the original membrane. The antifouling properties of the modified PVDF membrane were evaluated by a filtration test using bovine serum albumin (BSA) solution. The results showed that the initial flux of the modified membrane increased from 2140.1 L/m2 h to 2812.7 L/m2 h and the equilibrium flux of BSA solution increased from 31 L/m2 h to 53 L/m2 h.

  18. 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.

  19. Glycosylphosphatidyl Inositol-anchored Proteins and fyn Kinase Assemble in Noncaveolar Plasma Membrane Microdomains Defined by Reggie-1 and -2

    OpenAIRE

    Stürmer, Claudia; Lang, Dirk M.; Kirsch, Friederike; Wiechers, Marianne F.; Deininger, Sören-Oliver; Plattner, Helmut

    2001-01-01

    Using confocal laser scanning and double immunogold electron microscopy, we demonstrate that reggie-1 and -2 are colocalized in ≤0.1-μm plasma membrane microdomains of neurons and astrocytes. In astrocytes, reggie-1 and -2 do not occur in caveolae but clearly outside these structures. Microscopy and coimmunoprecipitation show that reggie-1 and -2 are associated with fyn kinase and with the glycosylphosphatidyl inositol-anchored proteins Thy-1 and F3 that, when activated by antibody cross-link...

  20. Ion induced changes in the structure of bordered pit membranes.

    Science.gov (United States)

    Lee, Jinkee; Holbrook, N Michele; Zwieniecki, Maciej A

    2012-01-01

    Ion-mediated changes in xylem hydraulic resistance are hypothesized to result from hydrogel like properties of pectins located in the bordered pit membranes separating adjacent xylem vessels. Although the kinetics of the ion-mediated changes in hydraulic resistance are consistent with the swelling/deswelling behavior of pectins, there is no direct evidence of this activity. In this report we use atomic force microscopy (AFM) to investigate structural changes in bordered pit membranes associated with changes in the ionic concentration of the surrounding solution. When submerged in de-ionized water, AFM revealed bordered pit membranes as relatively smooth, soft, and lacking any sharp edges surface, in contrast to pictures from scanning electron microscope (SEM) or AFM performed on air-dry material. Exposure of the bordered pit membranes to 50 mM KCl solution resulted in significant changes in both surface physical properties and elevation features. Specifically, bordered pit membranes became harder and the fiber edges were clearly visible. In addition, the membrane contracted and appeared much rougher due to exposed microfibers. In neither solution was there any evidence of discrete pores through the membrane whose dimensions were altered in response to the ionic composition of the surrounding solution. Instead the variable hydraulic resistance appears to involve changes in the both the permeability and the thickness of the pit membrane.

  1. Effects of vitamin E supplementation on plasma membrane permeabilization and fluidization induced by chlorpromazine in the rat brain.

    Science.gov (United States)

    Maruoka, Nobuyuki; Murata, Tetsuhito; Omata, Naoto; Takashima, Yasuhiro; Fujibayashi, Yasuhisa; Wada, Yuji

    2008-03-01

    Neurotransmitter receptors play a key role in most research on antipsychotic drugs, but little is known about the effects of these drugs on the plasma membrane in the central nervous system. Therefore, we investigated whether chlorpromazine (CPZ), a typical phenothiazine antipsychotic drug, affects the plasma membrane integrity in the rat brain, and if so, whether these membrane alterations can be prevented by dietary supplementation with vitamin E, which has been shown to be an antioxidant and also a membrane-stabilizer. Leakage of [(18)F]2-fluoro-2-deoxy-D-glucose ([(18)F]FDG)-6-phosphate from rat striatal slices and decrease in 1,6-diphenyl-1,3,5-hexatriene fluorescence anisotropy were used as indexes for plasma membrane permeabilization and fluidization, respectively. CPZ induced leakage of [(18)F]FDG-6-phosphate from striatal slices, and the leakage was delayed in the vitamin E-supplemented group compared to that in the normal diet group. The decrease in plasma membrane anisotropy induced by CPZ was significantly attenuated by vitamin E supplementation. Chronic treatment with alpha-phenyl-N-tert-butyl nitrone, a free radical scavenger, had no effect on CPZ-induced plasma membrane permeabilization, and the treatment with CPZ did not induce lipid peroxidation. CPZ can reduce plasma membrane integrity in the brain, and this reduction can be prevented by vitamin E via its membrane-stabilizing properties, not via its antioxidant activity.

  2. Membrane transport mechanism 3D structure and beyond

    CERN Document Server

    Ziegler, Christine

    2014-01-01

    This book provides a molecular view of membrane transport by means of numerous biochemical and biophysical techniques. The rapidly growing number of atomic structures of transporters in different conformations and the constant progress in bioinformatics have recently added deeper insights.   The unifying mechanism of energized solute transport across membranes is assumed to consist of the conformational cycling of a carrier protein to provide access to substrate binding sites from either side of a cellular membrane. Due to the central role of active membrane transport there is considerable interest in deciphering the principles of one of the most fundamental processes in nature: the alternating access mechanism.   This book brings together particularly significant structure-function studies on a variety of carrier systems from different transporter families: Glutamate symporters, LeuT-like fold transporters, MFS transporters and SMR (RND) exporters, as well as ABC-type importers.   The selected examples im...

  3. Development of topologically structured membranes of aluminum oxide

    Science.gov (United States)

    Bankova, A.; Videkov, V.; Tzaneva, B.

    2014-05-01

    In recent years, nanomembranes have become one of the most widely used construction material for ultrasensitive and ultrathin applications in micro-electromechanical systems (MEMS) and other sensor structures due to their remarkable mechanical properties. Among these, the mechanical stability is of particular importance. We present an approach to the analysis of the stability of nanostructured anodic aluminum oxide free membranes subjected to mechanical bending. The membranes tested were with a thickness of 500 nm to 15 urn in various topological shapes; we describe the technological schemes of their preparation. Bends were applied to membranes prepared by using a selective process of etching and anodizing. The results of the preparation of the membranes are discussed, together with the influence of the angle of deflection, and the number of bendings. The results obtained can be used in designing MEMS structures and sensors which use nanostructured anodic aluminum oxide.

  4. Tumor-specific Hsp70 plasma membrane localization is enabled by the glycosphingolipid Gb3.

    Directory of Open Access Journals (Sweden)

    Mathias Gehrmann

    Full Text Available BACKGROUND: Human tumors differ from normal tissues in their capacity to present Hsp70, the major stress-inducible member of the HSP70 family, on their plasma membrane. Membrane Hsp70 has been found to serve as a prognostic indicator of overall patient survival in leukemia, lower rectal and non small cell lung carcinomas. Why tumors, but not normal cells, present Hsp70 on their cell surface and the impact of membrane Hsp70 on cancer progression remains to be elucidated. METHODOLOGY/PRINCIPAL FINDINGS: Although Hsp70 has been reported to be associated with cholesterol rich microdomains (CRMs, the partner in the plasma membrane with which Hsp70 interacts has yet to be identified. Herein, global lipid profiling demonstrates that Hsp70 membrane-positive tumors differ from their membrane-negative counterparts by containing significantly higher amounts of globotriaoslyceramide (Gb3, but not of other lipids such as lactosylceramide (LacCer, dodecasaccharideceramide (DoCer, galactosylceramide (GalCer, ceramide (Cer, or the ganglioside GM1. Apart from germinal center B cells, normal tissues are Gb3 membrane-negative. Co-localization of Hsp70 and Gb3 was selectively determined in Gb3 membrane-positive tumor cells, and these cells were also shown to bind soluble Hsp70-FITC protein from outside in a concentration-dependent manner. Given that the latter interaction can be blocked by a Gb3-specific antibody, and that the depletion of globotriaosides from tumors reduces the amount of membrane-bound Hsp70, we propose that Gb3 is a binding partner for Hsp70. The in vitro finding that Hsp70 predominantly binds to artificial liposomes containing Gb3 (PC/SM/Chol/Gb3, 17/45/33/5 confirms that Gb3 is an interaction partner for Hsp70. CONCLUSIONS/SIGNIFICANCE: These data indicate that the presence of Gb3 enables anchorage of Hsp70 in the plasma membrane of tumors and thus they might explain tumor-specific membrane localization of Hsp70.

  5. Structure formation of surfactant membranes under shear flow

    Science.gov (United States)

    Shiba, Hayato; Noguchi, Hiroshi; Gompper, Gerhard

    2013-07-01

    Shear-flow-induced structure formation in surfactant-water mixtures is investigated numerically using a meshless-membrane model in combination with a particle-based hydrodynamics simulation approach for the solvent. At low shear rates, uni-lamellar vesicles and planar lamellae structures are formed at small and large membrane volume fractions, respectively. At high shear rates, lamellar states exhibit an undulation instability, leading to rolled or cylindrical membrane shapes oriented in the flow direction. The spatial symmetry and structure factor of this rolled state agree with those of intermediate states during lamellar-to-onion transition measured by time-resolved scatting experiments. Structural evolution in time exhibits a moderate dependence on the initial condition.

  6. Class I Cytokine Receptors: Structure and function in the Membrane

    DEFF Research Database (Denmark)

    Bugge, Katrine Østergaard

    Class I cytokine receptors are involved in important biological functions of both physiological and pathological nature in mammals. However, the molecular details of the cross-membrane signal transduction through these receptors remain obscure. One of the major reasons for this is the lack...... of structural knowledge on their membrane-embedded transmembrane domains (TMDs), which connect the extracellular ligand binding domains to the intracellular signaling platforms. The overall aim of this thesis work was to improve our understanding of the class I cytokine receptor signaling across the membrane...... ample material of high quality for structural studies with NMR spectroscopy of several class I cytokine receptor TMDs. Furthermore, the structure of a class I cytokine receptor TMD in DHPC micelles was solved with solution-state NMR spectroscopy. Additionally, since structural studies of intact proteins...

  7. A beam-membrane structure micromachined differential pressure flow sensor.

    Science.gov (United States)

    Chen, P; Zhao, Y L; Tian, B; Li, C; Li, Y Y

    2015-04-01

    A beam-membrane structure micromachined flow sensor is designed, depending on the principle of differential pressure caused by the mass flow, which is directly proportional to the square flow rate. The FSI (fluid structure interaction) characteristics of the differential pressure flow sensor are investigated via numerical analysis and analog simulation. The working mechanism of the flow sensor is analyzed depending on the FSI results. Then, the flow sensor is fabricated and calibrated. The calibration results show that the beam-membrane structure differential pressure flow sensor achieves ideal static characteristics and works well in the practical applications.

  8. Correct use of Membrane Elements in Structural Analysis

    Directory of Open Access Journals (Sweden)

    Rothman Timothy

    2016-01-01

    Full Text Available Structural analysis of consumer electronic devices such as phones and tablets involves Finite Element Analysis (FEA. Dynamic loading conditions such as device dropping and bending dictate accurate FEA models to reduce design risk in many areas. The solid elements typically used in structural analysis do not have integration points on the surface. The outer surface is of most interest because that is where the cracks start. Analysts employ a post processing trick through using membranes to bring accurate stress/strain results to the surface. This paper explains numerical issues with implementation of membranes and recommends a methodology for accurate structural analysis.

  9. The biological response of cells to nanosecond pulsed electric fields is dependent on plasma membrane cholesterol.

    Science.gov (United States)

    Cantu, Jody C; Tarango, Melissa; Beier, Hope T; Ibey, Bennett L

    2016-11-01

    Previous work from our laboratory demonstrated nanopore formation in cell membranes following exposure to nanosecond pulsed electric fields (nsPEF). We observed differences in sensitivity to nsPEF in both acute membrane injury and 24h lethality across multiple cells lines. Based on these data, we hypothesize that the biological response of cells to nsPEF is dependent on the physical properties of the plasma membrane (PM), including regional cholesterol content. Results presented in this paper show that depletion of membrane cholesterol disrupts the PM and increases the permeability of cells to small molecules, including propidium iodide and calcium occurring after fewer nsPEF. Additionally, cholesterol depletion concurrently decreases the "dose" of nsPEF required to induce lethality. In summary, the results of the current study suggest that the PM cholesterol composition is an important determinant in the cellular response to nsPEF. Copyright © 2016 Elsevier B.V. All rights reserved.

  10. Lagrangian coherent structures and plasma transport processes

    CERN Document Server

    Falessi, M V; Schep, T J

    2015-01-01

    A dynamical system framework is used to describe transport processes in plasmas embedded in a magnetic field. For periodic systems with one degree of freedom the Poincar\\'e map provides a splitting of the phase space into regions where particles have different kinds of motion: periodic, quasi-periodic or chaotic. The boundaries of these regions are transport barriers; i.e., a trajectory cannot cross such boundaries during the whole evolution of the system. Lagrangian Coherent Structure (LCS) generalize this method to systems with the most general time dependence, splitting the phase space into regions with different qualitative behaviours. This leads to the definition of finite-time transport barriers, i.e. trajectories cannot cross the barrier for a finite amount of time. This methodology can be used to identify fast recirculating regions in the dynamical system and to characterize the transport between them.

  11. Protein amino acid composition of plasma membranes affects membrane fluidity and thereby ethanol tolerance in a self-flocculating fusant of Schizosaccharomyces pombe and Saccharomyces cerevisiae.

    Science.gov (United States)

    Hu, Chun-Keng; Bai, Feng-Wu; An, Li-Jia

    2005-09-01

    A combination of three amino acids including 1.0 g/L isoleucine, 0.5 g/L methionine and 2.0 g/L phenylalanine was found to enhance ethanol tolerance of a self-flocculating fusant of Schizosaccharomyces pombe and Saccharomyces cerevisiae. When subjected to 20% (V/V) ethanol for 9 h at 30 degrees C, all cells died whereas 57% remained viable for the cells grown in the presence of the three amino acids. Based on the analysis of protein amino acid composition of plasma membranes and the determination of plasma membrane fluidity by measuring fluorescence anisotropy using diphenylhexatriene as a probe, it was found that the significantly increased ethanol tolerance of cells grown with the three amino acids was due to the incorporation of the supplementary amino acids into the plasma membranes, thus resulting in enhanced ability of the plasma membranes to efficiently counteract the fluidizing effect of ethanol when subjected to ethanol stress. This is the first time to report that plasma membrane fluidity can be influenced by protein amino acid composition of plasma membranes.

  12. Membrane proteins structure and dynamics by nuclear magnetic resonance.

    Science.gov (United States)

    Maltsev, Sergey; Lorigan, Gary A

    2011-10-01

    Membrane proteins represent a challenging class of biological systems to study. They are extremely difficult to crystallize and in most cases they retain their structure and functions only in membrane environments. Therefore, commonly used diffraction methods fail to give detailed molecular structure and other approaches have to be utilized to obtain biologically relevant information. Nuclear magnetic resonance (NMR) spectroscopy, however, can provide powerful structural and dynamical constraints on these complicated systems. Solution- and solid-state NMR are powerful methods for investigating membrane proteins studies. In this work, we briefly review both solution and solid-state NMR techniques for membrane protein studies and illustrate the applications of these methods to elucidate proteins structure, conformation, topology, dynamics, and function. Recent advances in electronics, biological sample preparation, and spectral processing provided opportunities for complex biological systems, such as membrane proteins inside lipid vesicles, to be studied faster and with outstanding quality. New analysis methods therefore have emerged, that benefit from the combination of sample preparation and corresponding specific high-end NMR techniques, which give access to more structural and dynamic information.

  13. Finite Element Analysis of Wrinkled Membrane Structures for Sunshield Applications

    Science.gov (United States)

    Johnston, John D.; Brodeur, Stephen J. (Technical Monitor)

    2002-01-01

    The deployable sunshield is an example of a gossamer structure envisioned for use on future space telescopes. The basic structure consists of multiple layers of pretensioned, thin-film membranes supported by deployable booms. The prediction and verification of sunshield dynamics has been identified as an area in need of technology development due to the difficulties inherent in predicting nonlinear structural behavior of the membranes and because of the challenges involved. in ground testing of the full-scale structure. This paper describes a finite element analysis of a subscale sunshield that has been subjected to ground testing in support of the Next Generation Space Telescope (NGST) program. The analysis utilizes a nonlinear material model that accounts for wrinkling of the membranes. Results are presented from a nonlinear static preloading analysis and subsequent dynamics analyses to illustrate baseline sunshield structural characteristics. Studies are then described which provide further insight into the effect of membrane. preload on sunshield dynamics and the performance of different membrane modeling techniques. Lastly, a comparison of analytical predictions and ground test results is presented.

  14. Research on EM pulse protection property of plasma-microwave absorptive material-plasma sandwich structure

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    A plasma-microwave absorptive material (MAM)-plasma sandwich structure is presented to protect the electronic device against high power electromagnetic pulse. The model of electromagnetic wave reflected by and transmitting through the structure is established. Based on the characteristic parameters of plasma generated by discharge and usual MAM, the electromagnetic transmissive properties of the sandwich structure are investigated by the method of finite difference in time domain. The results indicate that in a rather broad frequency range, the electromagnetic attenuations by the structure are obviously better than the sum of attenuations resulted from plasma and MAM respectively. The models and results presented are instructive for electromagnetic pulse protection.

  15. 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

  16. Plant plasma membrane-bound staphylococcal-like DNases as a novel class of eukaryotic nucleases

    Directory of Open Access Journals (Sweden)

    Leśniewicz Krzysztof

    2012-10-01

    Full Text Available Abstract Background The activity of degradative nucleases responsible for genomic DNA digestion has been observed in all kingdoms of life. It is believed that the main function of DNA degradation occurring during plant programmed cell death is redistribution of nucleic acid derived products such as nitrogen, phosphorus and nucleotide bases. Plant degradative nucleases that have been studied so far belong mainly to the S1-type family and were identified in cellular compartments containing nucleic acids or in the organelles where they are stored before final application. However, the explanation of how degraded DNA components are exported from the dying cells for further reutilization remains open. Results Bioinformatic and experimental data presented in this paper indicate that two Arabidopsis staphylococcal-like nucleases, named CAN1 and CAN2, are anchored to the cell membrane via N-terminal myristoylation and palmitoylation modifications. Both proteins possess a unique hybrid structure in their catalytic domain consisting of staphylococcal nuclease-like and tRNA synthetase anticodon binding-like motifs. They are neutral, Ca2+-dependent nucleaces showing a different specificity toward the ssDNA, dsDNA and RNA substrates. A study of microarray experiments and endogenous nuclease activity revealed that expression of CAN1 gene correlates with different forms of programmed cell death, while the CAN2 gene is constitutively expressed. Conclusions In this paper we present evidence showing that two plant staphylococcal-like nucleases belong to a new, as yet unidentified class of eukaryotic nucleases, characterized by unique plasma membrane localization. The identification of this class of nucleases indicates that plant cells possess additional, so far uncharacterized, mechanisms responsible for DNA and RNA degradation. The potential functions of these nucleases in relation to their unique intracellular location are discussed.

  17. Co-overexpressing a plasma membrane and a vacuolar membrane sodium/proton antiporter significantly improves salt tolerance in transgenic Arabidopsis plants.

    Science.gov (United States)

    The Arabidopsis gene AtNHX1 encodes a vacuolar membrane bound sodium/proton (Sodium/Hydrogen) antiporter that transports sodium into the vacuole and exports hydrogen into the cytoplasm. The Arabidopsis gene SOS1 encodes a plasma membrane bound sodium/hydrogen antiporter that exports sodium to the ex...

  18. Chromium(VI)-induced Production of Reactive Oxygen Species, Change of Plasma Membrane Potential and Dissipation of Mitochondria Membrane Potential in Chinese Hamster Lung Cell Cultures

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Objective To examine whether Reactive Oxygen Species (ROS) is generated, and whether plasma membrane potential and mitochondrial membrane potential are depolarized in Chinese Hamster Lung (CHL) cell lines exposed to Cr (VI). Methods CHL cells were incubated with Cr(VI) at 10 μmol/L, 2.5 μmol/L, 0.65 μmol/L for 3 and 6 hours, respectively. The production of ROS was performed by using 2,7_dichlorofluorescin diacetate; The changes in plasma membrane potential were estimated using fluorescent cationic dye DiBAC4; And the changes in mitochondria membrane potential were estimated using fluorescent dye Rhodamine 123. Results The ROS levels in CHL cells increased in all treated groups compared with the control group (P<0.01); The plasma membrane potential and mitochondrial membrane potential in CHL cells dissipated after incubated with Cr(VI) at 10 μmol/L for 3 hours and 6 hours (P<0.01), at 2.5 μmol/L for 6 hours (P<0.01 or 0.05). Conclusion Cr(VI) causes the dissipation of plasma membrane potential and mitochondrial membrane potential in CHL cell cultures, and Cr(VI)_induced ROS may play a role in the injuries.

  19. Dissipative nonlinear structures in tokamak plasmas

    Directory of Open Access Journals (Sweden)

    K. A. Razumova

    2001-01-01

    Full Text Available A lot of different kinds of instabilities may be developed in high temperature plasma located in a strong toroidal magnetic field (tokamak plasma. Nonlinear effects in the instability development result in plasma self-organization. Such plasma has a geometrically complicated configuration, consisting of the magnetic surfaces imbedded into each other and split into islands with various characteristic numbers of helical twisting. The self-consistency of the processes means that the transport coefficients in plasma do not depend just on the local parameters, being a function of the whole plasma configuration and of the forces affecting it. By disrupting the bonds between separate magnetic surfaces filled with islands, one can produce zones of reduced transport in the plasma, i.e. “internal thermal barriers”, allowing one essentially to increase the plasma temperature and density.

  20. Course of organized structures in thermal plasma inside and outside argon plasma torch

    Science.gov (United States)

    Gruber, Jan; Sonsky, Jiri; Hlina, Jan

    2016-09-01

    Arc chamber of direct-current (dc) argon plasma torch and area just above the nozzle outside of this dc plasma torch were observed by hi-speed camera. System of reflecting mirrors and transparent silica arc chamber walls were used to obtain simultaneous records of both i) cathode area with electric arc inside the plasma torch and ii) nozzle exit with resulting plasma jet outside the plasma torch. Such experimental arrangement allowed us to track localized repeating patterns (organized structures) in the arc chamber and in the plasma flow. Identification of various organized structures - for different experimental conditions - according to their origin and typical development is presented in this paper. Impact of 300 Hz ripple in arc current was compared between different areas of the plasma. Additional simultaneous observation of plasma flow in the same system by series of photodiodes was used for verification of the results. The work was possible with institutional support RVO:61388998.

  1. Citrinin-induced fluidization of the plasma membrane of the fission yeast Schizosaccharomyces pombe.

    Science.gov (United States)

    Blaskó, Ágnes; Mike, Nóra; Gróf, Pál; Gazdag, Zoltán; Czibulya, Zsuzsanna; Nagy, Lívia; Kunsági-Máté, Sándor; Pesti, Miklós

    2013-09-01

    Citrinin (CTN) is a toxic fungal metabolite that is a hazardous contaminant of foods and feeds. In the present study, its acute toxicity and effects on the plasma membrane of Schizosaccharomyces pombe were investigated. The minimum inhibitory concentration of CTN against the yeast cells proved to be 500 μM. Treatment with 0, 250, 500 or 1000 μM CTN for 60 min resulted in a 0%, 2%, 21% or 100% decrease, respectively, in the survival rate of the cell population. Treatment of cells with 0, 100, 500 or 1000 μM CTN for 20 min induced decrease in the phase-transition temperature of the 5-doxylstearic acid-labeled plasma membrane to 16.51, 16.04, 14.18 or 13.98°C, respectively as measured by electron paramagnetic resonance spectroscopy. This perturbation was accompanied by the efflux of essential K⁺ from the cells. The existence of an interaction between CTN and glutathione was detected for the first time by spectrofluorometry. Our observations may suggest a direct interaction of CTN with the free sulfhydryl groups of the integral proteins of the plasma membrane, leading to dose-dependent membrane fluidization. The change in fluidity disturbed the ionic homeostasis, contributing to the death of the cells, which is a novel aspect of CTN cytotoxicity.

  2. Receptor dimer stabilization by hierarchical plasma membrane microcompartments regulates cytokine signaling.

    Science.gov (United States)

    You, Changjiang; Marquez-Lago, Tatiana T; Richter, Christian Paolo; Wilmes, Stephan; Moraga, Ignacio; Garcia, K Christopher; Leier, André; Piehler, Jacob

    2016-12-01

    The interaction dynamics of signaling complexes is emerging as a key determinant that regulates the specificity of cellular responses. We present a combined experimental and computational study that quantifies the consequences of plasma membrane microcompartmentalization for the dynamics of type I interferon receptor complexes. By using long-term dual-color quantum dot (QD) tracking, we found that the lifetime of individual ligand-induced receptor heterodimers depends on the integrity of the membrane skeleton (MSK), which also proved important for efficient downstream signaling. By pair correlation tracking and localization microscopy as well as by fast QD tracking, we identified a secondary confinement within ~300-nm-sized zones. A quantitative spatial stochastic diffusion-reaction model, entirely parameterized on the basis of experimental data, predicts that transient receptor confinement by the MSK meshwork allows for rapid reassociation of dissociated receptor dimers. Moreover, the experimentally observed apparent stabilization of receptor dimers in the plasma membrane was reproduced by simulations of a refined, hierarchical compartment model. Our simulations further revealed that the two-dimensional association rate constant is a key parameter for controlling the extent of MSK-mediated stabilization of protein complexes, thus ensuring the specificity of this effect. Together, experimental evidence and simulations support the hypothesis that passive receptor confinement by MSK-based microcompartmentalization promotes maintenance of signaling complexes in the plasma membrane.

  3. Discovery of novel membrane binding structures and functions

    Science.gov (United States)

    Kufareva, Irina; Lenoir, Marc; Dancea, Felician; Sridhar, Pooja; Raush, Eugene; Bissig, Christin; Gruenberg, Jean; Abagyan, Ruben; Overduin, Michael

    2014-01-01

    The function of a protein is determined by its intrinsic activity in the context of its subcellular distribution. Membranes localize proteins within cellular compartments and govern their specific activities. Discovering such membrane-protein interactions is important for understanding biological mechanisms, and could uncover novel sites for therapeutic intervention. Here we present a method for detecting membrane interactive proteins and their exposed residues that insert into lipid bilayers. Although the development process involved analysis of how C1b, C2, ENTH, FYVE, Gla, pleckstrin homology (PH) and PX domains bind membranes, the resulting Membrane Optimal Docking Area (MODA) method yields predictions for a given protein of known three dimensional structures without referring to canonical membrane-targeting modules. This approach was tested on the Arf1 GTPase, ATF2 acetyltransferase, von Willebrand factor A3 domain and Neisseria gonorrhoeae MsrB protein, and further refined with membrane interactive and non-interactive FAPP1 and PKD1 pleckstrin homology domains, respectively. Furthermore we demonstrate how this tool can be used to discover unprecedented membrane binding functions as illustrated by the Bro1 domain of Alix, which was revealed to recognize lysobisphosphatidic acid (LBPA). Validation of novel membrane-protein interactions relies on other techniques such as nuclear magnetic resonance spectroscopy (NMR) which was used here to map the sites of micelle interaction. Together this indicates that genome-wide identification of known and novel membrane interactive proteins and sites is now feasible, and provides a new tool for functional annotation of the proteome. PMID:25394204

  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. Molecular recognition based iron removal from human plasma with imprinted membranes.

    Science.gov (United States)

    Yavuz, H; Andaç, M; Uzun, L; Say, R; Denizli, A

    2006-09-01

    The aim of this study is to prepare ion-imprinted poly(2-hydroxyethyl methacrylate) (HEMA) based membranes which can be used for the selective removal of Fe3+ ions from Fe3+-overdosed human plasma. N-methacryloyl-(L)-glutamic acid (MAGA) was chosen as the ion-complexing monomer. In the first step, Fe3+ was complexed with MAGA and then, the Fe3+-imprinted poly(HEMA-MAGA) membranes were prepared by UV-initiated photo-polymerization of HEMA and MAGA-Fe3+ complex in the presence of an initiator (benzoyl peroxide). After that, the template (i.e., Fe3+ ions) was removed by using 0.1 M EDTA solution at room temperature. The specific surface area of the Fe3+-imprinted poly(HEMA-MAGA) membranes was found to be 49.2 m2/g and the swelling ratio was 92%. According to the elemental analysis results, the polymeric membranes contained 145.7 micromol MAGA/g polymer. The maximum adsorption capacity was 164.2 micromol Fe3+/g membrane. The relative selectivity coefficients of ion-imprinted membranes for Fe3+/Zn2+ and Fe3+/Cr3+ were 12.6 and 62.5 times greater than the non-imprinted matrix, respectively. The Fe3+-imprinted poly(HEMA-MAGA) membranes could be used many times without decreasing their Fe3+ adsorption capacities significantly.

  6. Regulation of glycolytic oscillations by mitochondrial and plasma membrane H+-ATPases

    DEFF Research Database (Denmark)

    Olsen, Lars Folke; Andersen, Ann Zahle; Lunding, Anita

    2009-01-01

    We investigated the coupling between glycolytic and mitochondrial membrane potential oscillations in Saccharomyces cerevisiae under semianaerobic conditions. Glycolysis was measured as NADH autofluorescence, and mitochondrial membrane potential was measured using the fluorescent dye 3,3'-diethylo......We investigated the coupling between glycolytic and mitochondrial membrane potential oscillations in Saccharomyces cerevisiae under semianaerobic conditions. Glycolysis was measured as NADH autofluorescence, and mitochondrial membrane potential was measured using the fluorescent dye 3......,3'-diethyloxacarbocyanine iodide. The responses of glycolytic and membrane potential oscillations to a number of inhibitors of glycolysis, mitochondrial electron flow, and mitochondrial and plasma membrane H(+)-ATPase were investigated. Furthermore, the glycolytic flux was determined as the rate of production of ethanol...... in a number of different situations (changing pH or the presence and absence of inhibitors). Finally, the intracellular pH was determined and shown to oscillate. The results support earlier work suggesting that the coupling between glycolysis and mitochondrial membrane potential is mediated by the ADP...

  7. Parallel artificial liquid membrane extraction of acidic drugs from human plasma.

    Science.gov (United States)

    Roldán-Pijuán, Mercedes; Pedersen-Bjergaard, Stig; Gjelstad, Astrid

    2015-04-01

    The new sample preparation concept "Parallel artificial liquid membrane extraction (PALME)" was evaluated for extraction of the acidic drugs ketoprofen, fenoprofen, diclofenac, flurbiprofen, ibuprofen, and gemfibrozil from human plasma samples. Plasma samples (250 μL) were loaded into individual wells in a 96-well donor plate and diluted with HCl to protonate the acidic drugs. The acidic drugs were extracted as protonated species from the individual plasma samples, through corresponding artificial liquid membranes each comprising 2 μL of dihexyl ether, and into corresponding acceptor solutions each comprising 50 μL of 25 mM ammonia solution (pH 10). The liquid membranes and the acceptor solutions were located in a 96-well filter plate, which was sandwiched with the 96-well donor plate during extraction. Parallel extraction of several samples was performed for 15 to 60 min, followed by high-performance liquid chromatography-ultraviolet detection of the individual acceptor solutions. Important PALME parameters including the chemical composition of the liquid membrane, extraction time, and sample pH were optimized, and the extraction performance was evaluated. Except for flurbiprofen, exhaustive extraction was accomplished from plasma. Linearity was obtained for all six drugs in the range 0.025-10 μg/mL, with r (2) values ranging between 0.998 and 1.000. Precision data were in the range 3-22% RSD, and accuracy data were within 72-130% with spiked plasma samples. Based on the current experiences, PALME showed substantial potential for future high-throughput bioanalysis of non-polar acidic drugs.

  8. 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.

  9. 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.

  10. Temperature-Induced Protein Conformational Changes in Barley Root Plasma Membrane-Enriched Microsomes

    Science.gov (United States)

    Caldwell, Charles R.

    1987-01-01

    The membrane-bound proteins of barley (Hordeum vulgare L. cv Conquest) root plasma membrane-enriched microsomes displayed fluorescence typical of protein-associated trytophan residues. The protein fluorescence intensity was sensitive to variations in sample temperature. The temperature-induced decline in protein fluorescence intensity was nonlinear with slope discontinuities at about 12 and 32°C. Detergents at levels above their critical micelle concentration enhanced protein fluorescence. Glutaraldehyde reduced protein fluorescence. Protein fluorescence polarization increased at temperatures above 30°C. Both the rate of tryptophan photoionization and the fluorescence intensity of the photoionization products suggested alterations in membrane protein conformation between 12 and 32°C. The quenching of the intrinsic protein fluorescence by acrylamide and potassium iodide indicated changes in accessibility of the extrinsic agents to the protein tryptophan residues beginning at about 14°C. The results indicate thermally induced changes in the dynamics of the membrane proteins over the temperature range of 12 to 32°C which could account for the complex temperature dependence of the barley root plasma membrane ATPase. PMID:16665545

  11. Triggered Ca2+ influx is required for extended synaptotagmin 1-induced ER-plasma membrane tethering.

    Science.gov (United States)

    Idevall-Hagren, Olof; Lü, Alice; Xie, Beichen; De Camilli, Pietro

    2015-09-01

    The extended synaptotagmins (E-Syts) are ER proteins that act as Ca(2+)-regulated tethers between the ER and the plasma membrane (PM) and have a putative role in lipid transport between the two membranes. Ca(2+) regulation of their tethering function, as well as the interplay of their different domains in such function, remains poorly understood. By exposing semi-intact cells to buffers of variable Ca(2+) concentrations, we found that binding of E-Syt1 to the PI(4,5)P2-rich PM critically requires its C2C and C2E domains and that the EC50 of such binding is in the low micromolar Ca(2+) range. Accordingly, E-Syt1 accumulation at ER-PM contact sites occurred only upon experimental manipulations known to achieve these levels of Ca(2+) via its influx from the extracellular medium, such as store-operated Ca(2+) entry in fibroblasts and membrane depolarization in β-cells. We also show that in spite of their very different physiological functions, membrane tethering by E-Syt1 (ER to PM) and by synaptotagmin (secretory vesicles to PM) undergo a similar regulation by plasma membrane lipids and cytosolic Ca(2+).

  12. Elevation of plasma membrane permeability by laser irradiation of selectively bound nanoparticles.

    Science.gov (United States)

    Yao, Cuiping; Rahmanzadeh, Ramtin; Endl, Elmar; Zhang, Zhenxi; Gerdes, Johannes; Hüttmann, Gereon

    2005-01-01

    Irradiation of nanoabsorbers with pico- and nanosecond laser pulses could result in thermal effects with a spatial confinement of less than 50 nm. Therefore absorbing nanoparticles could be used to create controlled cellular effects. We describe a combination of laser irradiation with nanoparticles, which changes the plasma membrane permeability. We demonstrate that the system enables molecules to penetrate impermeable cell membranes. Laser light at 532 nm is used to irradiate conjugates of colloidal gold, which are delivered by antibodies to the plasma membrane of the Hodgkin's disease cell line L428 and/or the human large-cell anaplastic lymphoma cell line Karpas 299. After irradiation, membrane permeability is evaluated by fluorescence microscopy and flow cytometry using propidium iodide (PI) and fluorescein isothiocyanate (FITC) dextran. The fraction of transiently permeabilized and then resealed cells is affected by the laser parameter, the gold concentration, and the membrane protein of the different cell lines to which the nanoparticles are bound. Furthermore, a dependence on particle size is found for these interactions in the different cell lines. The results suggest that after optimization, this method could be used for gene transfection and gene therapy.

  13. 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.

  14. 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.

  15. Proteomic analysis identifies interleukin 11 regulated plasma membrane proteins in human endometrial epithelial cells in vitro

    Directory of Open Access Journals (Sweden)

    Stanton Peter G

    2011-05-01

    Full Text Available Abstract Background During the peri-implantation period, the embryo adheres to an adequately prepared or receptive endometrial surface epithelium. Abnormal embryo adhesion to the endometrium results in embryo implantation failure and infertility. Endometrial epithelial cell plasma membrane proteins critical in regulating adhesion may potentially be infertility biomarkers or targets for treating infertility. Interleukin (IL 11 regulates human endometrial epithelial cells (hEEC adhesion. Its production is abnormal in women with infertility. The objective of the study was to identify IL11 regulated plasma membrane proteins in hEEC in vitro using a proteomic approach. Methods Using a 2D-differential in-gel electrophoresis (DIGE electrophoresis combined with LCMS/MS mass spectrometry approach, we identified 20 unique plasma membrane proteins differentially regulated by IL11 in ECC-1 cells, a hEEC derived cell line. Two IL11 regulated proteins with known roles in cell adhesion, annexin A2 (ANXA2 and flotillin-1 (FLOT1, were validated by Western blot and immunocytochemistry in hEEC lines (ECC-1 and an additional cell line, Ishikawa and primary hEEC. Flotilin-1 was further validated by immunohistochemistry in human endometrium throughout the menstrual cycle (n = 6-8/cycle. Results 2D-DIGE analysis identified 4 spots that were significantly different between control and IL11 treated group. Of these 4 spots, there were 20 proteins that were identified with LCMS/MS. Two proteins; ANXA2 and FLOT1 were chosen for further analyses and have found to be significantly up-regulated following IL11 treatment. Western blot analysis showed a 2-fold and a 2.5-fold increase of ANXA2 in hEEC membrane fraction of ECC-1 and Ishikawa cells respectively. Similarly, a 1.8-fold and a 2.3/2.4-fold increase was also observed for FLOT1 in hEEC membrane fraction of ECC-1 and Ishikawa cells respectively. In vitro, IL11 induced stronger ANXA2 expression on cell surface of primary h

  16. Structure and formation of egg membranes in Aedes aegypti. (L. ) (Diptera:Culicidae)