Science.gov

Sample records for regulate membrane surface

  1. Membrane Supply and Demand Regulates F-Actin in a Cell Surface Reservoir.

    PubMed

    Figard, Lauren; Wang, Mengyu; Zheng, Liuliu; Golding, Ido; Sokac, Anna Marie

    2016-05-01

    Cells store membrane in surface reservoirs of pits and protrusions. These membrane reservoirs facilitate cell shape change and buffer mechanical stress, but we do not know how reservoir dynamics are regulated. During cellularization, the first cytokinesis in Drosophila embryos, a reservoir of microvilli unfolds to fuel cleavage furrow ingression. We find that regulated exocytosis adds membrane to the reservoir before and during unfolding. Dynamic F-actin deforms exocytosed membrane into microvilli. Single microvilli extend and retract in ∼20 s, while the overall reservoir is depleted in sync with furrow ingression over 60-70 min. Using pharmacological and genetic perturbations, we show that exocytosis promotes microvillar F-actin assembly, while furrow ingression controls microvillar F-actin disassembly. Thus, reservoir F-actin and, consequently, reservoir dynamics are regulated by membrane supply from exocytosis and membrane demand from furrow ingression. PMID:27165556

  2. Surface area of lipid membranes regulates the DNA-binding capacity of cationic liposomes

    NASA Astrophysics Data System (ADS)

    Marchini, Cristina; Montani, Maura; Amici, Augusto; Pozzi, Daniela; Caminiti, Ruggero; Caracciolo, Giulio

    2009-01-01

    We have applied electrophoresis on agarose gels to investigate the DNA-binding capacity of cationic liposomes made of cationic DC-cholesterol and neutral dioleoylphosphatidylethanolamine as a function of membrane charge density and cationic lipid/DNA charge ratio. While each cationic liposome formulation exhibits a distinctive DNA-protection ability, here we show that such a capacity is universally regulated by surface area of lipid membranes available for binding in an aspecific manner. The relevance of DNA protection for gene transfection is also discussed.

  3. AMP-Activated Protein Kinase Regulates the Cell Surface Proteome and Integrin Membrane Traffic

    PubMed Central

    Thavarajah, Thanusi; Medvedev, Sergei; Bowden, Peter; Marshall, John G.; Antonescu, Costin N.

    2015-01-01

    The cell surface proteome controls numerous cellular functions including cell migration and adhesion, intercellular communication and nutrient uptake. Cell surface proteins are controlled by acute changes in protein abundance at the plasma membrane through regulation of endocytosis and recycling (endomembrane traffic). Many cellular signals regulate endomembrane traffic, including metabolic signaling; however, the extent to which the cell surface proteome is controlled by acute regulation of endomembrane traffic under various conditions remains incompletely understood. AMP-activated protein kinase (AMPK) is a key metabolic sensor that is activated upon reduced cellular energy availability. AMPK activation alters the endomembrane traffic of a few specific proteins, as part of an adaptive response to increase energy intake and reduce energy expenditure. How increased AMPK activity during energy stress may globally regulate the cell surface proteome is not well understood. To study how AMPK may regulate the cell surface proteome, we used cell-impermeable biotinylation to selectively purify cell surface proteins under various conditions. Using ESI-MS/MS, we found that acute (90 min) treatment with the AMPK activator A-769662 elicits broad control of the cell surface abundance of diverse proteins. In particular, A-769662 treatment depleted from the cell surface proteins with functions in cell migration and adhesion. To complement our mass spectrometry results, we used other methods to show that A-769662 treatment results in impaired cell migration. Further, A-769662 treatment reduced the cell surface abundance of β1-integrin, a key cell migration protein, and AMPK gene silencing prevented this effect. While the control of the cell surface abundance of various proteins by A-769662 treatment was broad, it was also selective, as this treatment did not change the cell surface abundance of the transferrin receptor. Hence, the cell surface proteome is subject to acute

  4. Alternative routes to the cell surface underpin insulin-regulated membrane trafficking of GLUT4

    PubMed Central

    Kioumourtzoglou, Dimitrios; Pryor, Paul R.; Gould, Gwyn W.; Bryant, Nia J.

    2015-01-01

    ABSTRACT Insulin-stimulated delivery of glucose transporters (GLUT4, also known as SLC2A4) from specialized intracellular GLUT4 storage vesicles (GSVs) to the surface of fat and muscle cells is central to whole-body glucose regulation. This translocation and subsequent internalization of GLUT4 back into intracellular stores transits through numerous small membrane-bound compartments (internal GLUT4-containing vesicles; IGVs) including GSVs, but the function of these different compartments is not clear. Cellugyrin (also known as synaptogyrin-2) and sortilin define distinct populations of IGV; sortilin-positive IGVs represent GSVs, but the function of cellugyrin-containing IGVs is unknown. Here, we demonstrate a role for cellugyrin in intracellular sequestration of GLUT4 in HeLa cells and have used a proximity ligation assay to follow changes in pairwise associations between cellugyrin, sortilin, GLUT4 and membrane trafficking machinery following insulin-stimulation of 3T3-L1 adipoctyes. Our data suggest that insulin stimulates traffic from cellugyrin-containing to sortilin-containing membranes, and that cellugyrin-containing IGVs provide an insulin-sensitive reservoir to replenish GSVs following insulin-stimulated exocytosis of GLUT4. Furthermore, our data support the existence of a pathway from cellugyrin-containing membranes to the surface of 3T3-L1 adipocytes that bypasses GSVs under basal conditions, and that insulin diverts traffic away from this into GSVs. PMID:26071524

  5. Nanocrack-regulated self-humidifying membranes.

    PubMed

    Park, Chi Hoon; Lee, So Young; Hwang, Doo Sung; Shin, Dong Won; Cho, Doo Hee; Lee, Kang Hyuck; Kim, Tae-Woo; Kim, Tae-Wuk; Lee, Mokwon; Kim, Deok-Soo; Doherty, Cara M; Thornton, Aaron W; Hill, Anita J; Guiver, Michael D; Lee, Young Moo

    2016-04-28

    The regulation of water content in polymeric membranes is important in a number of applications, such as reverse electrodialysis and proton-exchange fuel-cell membranes. External thermal and water management systems add both mass and size to systems, and so intrinsic mechanisms of retaining water and maintaining ionic transport in such membranes are particularly important for applications where small system size is important. For example, in proton-exchange membrane fuel cells, where water retention in the membrane is crucial for efficient transport of hydrated ions, by operating the cells at higher temperatures without external humidification, the membrane is self-humidified with water generated by electrochemical reactions. Here we report an alternative solution that does not rely on external regulation of water supply or high temperatures. Water content in hydrocarbon polymer membranes is regulated through nanometre-scale cracks ('nanocracks') in a hydrophobic surface coating. These cracks work as nanoscale valves to retard water desorption and to maintain ion conductivity in the membrane on dehumidification. Hydrocarbon fuel-cell membranes with surface nanocrack coatings operated at intermediate temperatures show improved electrochemical performance, and coated reverse-electrodialysis membranes show enhanced ionic selectivity with low bulk resistance. PMID:27121841

  6. Nanocrack-regulated self-humidifying membranes

    NASA Astrophysics Data System (ADS)

    Park, Chi Hoon; Lee, So Young; Hwang, Doo Sung; Shin, Dong Won; Cho, Doo Hee; Lee, Kang Hyuck; Kim, Tae-Woo; Kim, Tae-Wuk; Lee, Mokwon; Kim, Deok-Soo; Doherty, Cara M.; Thornton, Aaron W.; Hill, Anita J.; Guiver, Michael D.; Lee, Young Moo

    2016-04-01

    The regulation of water content in polymeric membranes is important in a number of applications, such as reverse electrodialysis and proton-exchange fuel-cell membranes. External thermal and water management systems add both mass and size to systems, and so intrinsic mechanisms of retaining water and maintaining ionic transport in such membranes are particularly important for applications where small system size is important. For example, in proton-exchange membrane fuel cells, where water retention in the membrane is crucial for efficient transport of hydrated ions, by operating the cells at higher temperatures without external humidification, the membrane is self-humidified with water generated by electrochemical reactions. Here we report an alternative solution that does not rely on external regulation of water supply or high temperatures. Water content in hydrocarbon polymer membranes is regulated through nanometre-scale cracks (‘nanocracks’) in a hydrophobic surface coating. These cracks work as nanoscale valves to retard water desorption and to maintain ion conductivity in the membrane on dehumidification. Hydrocarbon fuel-cell membranes with surface nanocrack coatings operated at intermediate temperatures show improved electrochemical performance, and coated reverse-electrodialysis membranes show enhanced ionic selectivity with low bulk resistance.

  7. Characterization of Surface Modification of Polyethersulfone Membrane

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Surface modification of polyethersulfone (PES) membrane surface using UV/ozone-treated grafting and interfacial polymerization on membrane surface was investigated in order to improve the resistance of membrane surface to protein adsorption. These methods of surface modification were compared in te...

  8. Lipid-modified azurin of Neisseria meningitidis is a copper protein localized on the outer membrane surface and not regulated by FNR.

    PubMed

    Deeudom, Manu; Huston, Wilhemina; Moir, James W B

    2015-04-01

    The laz gene of Neisseria meningitidis is predicted to encode a lipid-modified azurin (Laz). Laz is very similar to azurin, a periplasmic protein, which belongs to the copper-containing proteins in the cupredoxin superfamily. In other bacteria, azurin is an electron donor to nitrite reductase, an important enzyme in the denitrifying process. It is not known whether Laz could function as an electron transfer protein in this important pathogen. Laz protein was heterologously expressed in Escherichia coli and purified. Electrospray mass spectrometry indicated that the Laz protein contains one copper ion. Laz was shown to be redox-active in the presence of its redox center copper ion. When oxidized, Laz exhibits an intense blue colour and absorbs visible light around 626 nm. The absorption is lost when exposed to diethyldithiocarbamate, a copper chelating agent. Polyclonal antibodies were raised against purified Laz for detecting expression of Laz under different growth conditions and to determine the orientation of Laz on the outer membrane. The expression of Laz under microaerobic and microaerobic denitrifying conditions was slightly higher than that under aerobic conditions. However, the expression of Laz was similar between the wild type strain and an fnr mutant, suggesting that Fumarate/Nitrate reduction regulator (FNR) does not regulate the expression of Laz despite the presence of a partial FNR box upstream of the laz gene. We propose that some Laz protein is exposed on the outer membrane surface of N. meningitidis as the αLaz antibodies can increase killing by complement in a capsule deficient N. meningitidis strain, in a dose-dependent fashion. PMID:25666376

  9. Membrane potential shapes regulation of dopamine transporter trafficking at the plasma membrane

    PubMed Central

    Richardson, Ben D.; Saha, Kaustuv; Krout, Danielle; Cabrera, Elizabeth; Felts, Bruce; Henry, L. Keith; Swant, Jarod; Zou, Mu-Fa; Newman, Amy Hauck; Khoshbouei, Habibeh

    2016-01-01

    The dopaminergic system is essential for cognitive processes, including reward, attention and motor control. In addition to DA release and availability of synaptic DA receptors, timing and magnitude of DA neurotransmission depend on extracellular DA-level regulation by the dopamine transporter (DAT), the membrane expression and trafficking of which are highly dynamic. Data presented here from real-time TIRF (TIRFM) and confocal microscopy coupled with surface biotinylation and electrophysiology suggest that changes in the membrane potential alone, a universal yet dynamic cellular property, rapidly alter trafficking of DAT to and from the surface membrane. Broadly, these findings suggest that cell-surface DAT levels are sensitive to membrane potential changes, which can rapidly drive DAT internalization from and insertion into the cell membrane, thus having an impact on the capacity for DAT to regulate extracellular DA levels. PMID:26804245

  10. Fractal reconstruction of rough membrane surface related with membrane fouling in a membrane bioreactor.

    PubMed

    Zhang, Meijia; Chen, Jianrong; Ma, Yuanjun; Shen, Liguo; He, Yiming; Lin, Hongjun

    2016-09-01

    In this paper, fractal reconstruction of rough membrane surface with a modified Weierstrass-Mandelbrot (WM) function was conducted. The topography of rough membrane surface was measured by an atomic force microscopy (AFM), and the results showed that the membrane surface was isotropous. Accordingly, the fractal dimension and roughness of membrane surface were calculated by the power spectrum method. The rough membrane surface was reconstructed on the MATLAB platform with the parameter values acquired from raw AFM data. The reconstructed membrane was much similar to the real membrane morphology measured by AFM. The parameters (including average roughness and root mean square (RMS) roughness) associated with membrane morphology for the model and real membrane were calculated, and a good match of roughness parameters between the reconstructed surface and real membrane was found, indicating the feasibility of the new developed method. The reconstructed membrane surface can be potentially used for interaction energy evaluation. PMID:27318159

  11. Membrane tension regulates clathrin-coated pit dynamics

    NASA Astrophysics Data System (ADS)

    Liu, Allen

    2014-03-01

    Intracellular organization depends on close communication between the extracellular environment and a network of cytoskeleton filaments. The interactions between cytoskeletal filaments and the plasma membrane lead to changes in membrane tension that in turns help regulate biological processes. Endocytosis is thought to be stimulated by low membrane tension and the removal of membrane increases membrane tension. While it is appreciated that the opposing effects of exocytosis and endocytosis have on keeping plasma membrane tension to a set point, it is not clear how membrane tension affects the dynamics of clathrin-coated pits (CCPs), the individual functional units of clathrin-mediated endocytosis. Furthermore, although it was recently shown that actin dynamics counteracts membrane tension during CCP formation, it is not clear what roles plasma membrane tension plays during CCP initiation. Based on the notion that plasma membrane tension is increased when the membrane area increases during cell spreading, we designed micro-patterned surfaces of different sizes to control the cell spreading sizes. Total internal reflection fluorescence microscopy of living cells and high content image analysis were used to quantify the dynamics of CCPs. We found that there is an increased proportion of CCPs with short (<20s) lifetime for cells on larger patterns. Interestingly, cells on larger patterns have higher CCP initiation density, an effect unexpected based on the conventional view of decreasing endocytosis with increasing membrane tension. Furthermore, by analyzing the intensity profiles of CCPs that were longer-lived, we found CCP intensity decreases with increasing cell size, indicating that the CCPs are smaller with increasing membrane tension. Finally, disruption of actin dynamics significantly increased the number of short-lived CCPs, but also decreased CCP initiation rate. Together, our study reveals new mechanistic insights into how plasma membrane tension regulates

  12. Surface selective membranes for carbon dioxide separation

    SciTech Connect

    Luebke, D.R.; Pennline, H.W.; Myers, C.R.

    2005-09-01

    In this study, hybrid membranes have been developed for the selective separation of CO2 from mixtures containing H2. Beginning with commercially available Pall alumina membrane tubes with nominal pore diameter of 5 nm, hybrids were produced by silation with a variety of functionalities designed to facilitate the selective adsorption of CO2 onto the pore surface. The goal is to produce a membrane which can harness the power of surface diffusion to give the selectivity of polymer membranes with the permeance of inorganic membranes.

  13. Formation and Regulation of Mitochondrial Membranes

    PubMed Central

    Schenkel, Laila Cigana

    2014-01-01

    Mitochondrial membrane phospholipids are essential for the mitochondrial architecture, the activity of respiratory proteins, and the transport of proteins into the mitochondria. The accumulation of phospholipids within mitochondria depends on a coordinate synthesis, degradation, and trafficking of phospholipids between the endoplasmic reticulum (ER) and mitochondria as well as intramitochondrial lipid trafficking. Several studies highlight the contribution of dietary fatty acids to the remodeling of phospholipids and mitochondrial membrane homeostasis. Understanding the role of phospholipids in the mitochondrial membrane and their metabolism will shed light on the molecular mechanisms involved in the regulation of mitochondrial function and in the mitochondrial-related diseases. PMID:24578708

  14. Nuclear myosin I regulates cell membrane tension.

    PubMed

    Venit, Tomáš; Kalendová, Alžběta; Petr, Martin; Dzijak, Rastislav; Pastorek, Lukáš; Rohožková, Jana; Malohlava, Jakub; Hozák, Pavel

    2016-01-01

    Plasma membrane tension is an important feature that determines the cell shape and influences processes such as cell motility, spreading, endocytosis and exocytosis. Unconventional class 1 myosins are potent regulators of plasma membrane tension because they physically link the plasma membrane with adjacent cytoskeleton. We identified nuclear myosin 1 (NM1) - a putative nuclear isoform of myosin 1c (Myo1c) - as a new player in the field. Although having specific nuclear functions, NM1 localizes predominantly to the plasma membrane. Deletion of NM1 causes more than a 50% increase in the elasticity of the plasma membrane around the actin cytoskeleton as measured by atomic force microscopy. This higher elasticity of NM1 knock-out cells leads to 25% higher resistance to short-term hypotonic environment and rapid cell swelling. In contrast, overexpression of NM1 in wild type cells leads to an additional 30% reduction of their survival. We have shown that NM1 has a direct functional role in the cytoplasm as a dynamic linker between the cell membrane and the underlying cytoskeleton, regulating the degree of effective plasma membrane tension. PMID:27480647

  15. Nuclear myosin I regulates cell membrane tension

    PubMed Central

    Venit, Tomáš; Kalendová, Alžběta; Petr, Martin; Dzijak, Rastislav; Pastorek, Lukáš; Rohožková, Jana; Malohlava, Jakub; Hozák, Pavel

    2016-01-01

    Plasma membrane tension is an important feature that determines the cell shape and influences processes such as cell motility, spreading, endocytosis and exocytosis. Unconventional class 1 myosins are potent regulators of plasma membrane tension because they physically link the plasma membrane with adjacent cytoskeleton. We identified nuclear myosin 1 (NM1) - a putative nuclear isoform of myosin 1c (Myo1c) - as a new player in the field. Although having specific nuclear functions, NM1 localizes predominantly to the plasma membrane. Deletion of NM1 causes more than a 50% increase in the elasticity of the plasma membrane around the actin cytoskeleton as measured by atomic force microscopy. This higher elasticity of NM1 knock-out cells leads to 25% higher resistance to short-term hypotonic environment and rapid cell swelling. In contrast, overexpression of NM1 in wild type cells leads to an additional 30% reduction of their survival. We have shown that NM1 has a direct functional role in the cytoplasm as a dynamic linker between the cell membrane and the underlying cytoskeleton, regulating the degree of effective plasma membrane tension. PMID:27480647

  16. Regulation of Plasma Membrane Recycling by CFTR

    NASA Astrophysics Data System (ADS)

    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.

  17. Electrodiffusion of lipids on membrane surfaces

    NASA Astrophysics Data System (ADS)

    Zhou, Y. C.

    2012-05-01

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

  18. Functional dynamics of cell surface membrane proteins

    NASA Astrophysics Data System (ADS)

    Nishida, Noritaka; Osawa, Masanori; Takeuchi, Koh; Imai, Shunsuke; Stampoulis, Pavlos; Kofuku, Yutaka; Ueda, Takumi; Shimada, Ichio

    2014-04-01

    Cell surface receptors are integral membrane proteins that receive external stimuli, and transmit signals across plasma membranes. In the conventional view of receptor activation, ligand binding to the extracellular side of the receptor induces conformational changes, which convert the structure of the receptor into an active conformation. However, recent NMR studies of cell surface membrane proteins have revealed that their structures are more dynamic than previously envisioned, and they fluctuate between multiple conformations in an equilibrium on various timescales. In addition, NMR analyses, along with biochemical and cell biological experiments indicated that such dynamical properties are critical for the proper functions of the receptors. In this review, we will describe several NMR studies that revealed direct linkage between the structural dynamics and the functions of the cell surface membrane proteins, such as G-protein coupled receptors (GPCRs), ion channels, membrane transporters, and cell adhesion molecules.

  19. Diffusion mediated localization on membrane surfaces

    NASA Technical Reports Server (NTRS)

    Weaver, D. L.

    1982-01-01

    Using the model of a cell membrane of a spherical surface in which membrane components may diffuse, the rate of localization due to trapping under diffusion control has been estimated by computing an analytical expression for the mean trapping time including the possibilities of a trapping probability less than one and/or the establishment of an equilibrium at the trap boundary.

  20. Capillary origami and superhydrophobic membrane surfaces

    NASA Astrophysics Data System (ADS)

    Geraldi, N. R.; Ouali, F. F.; Morris, R. H.; McHale, G.; Newton, M. I.

    2013-05-01

    Capillary origami uses surface tension to fold and shape solid films and membranes into three-dimensional structures. It uses the fact that solid surfaces, no matter how hydrophobic, will tend to adhere to and wrap around the surface of a liquid. In this work, we report that a superhydrophobic coating can be created, which can completely suppress wrapping as a contacting water droplet evaporates. We also show that using a wetting azeotropic solution of allyl alcohol, which penetrates the surface features, can enhance liquid adhesion and create more powerful Capillary Origami. These findings create the possibility of selectively shaping membrane substrates.

  1. Vesicle trafficking and cell surface membrane patchiness.

    PubMed

    Tang, Q; Edidin, M

    2001-07-01

    Membrane proteins and lipids often appear to be distributed in patches on the cell surface. These patches are often assumed to be membrane domains, arising from specific molecular associations. However, a computer simulation (Gheber and Edidin, 1999) shows that membrane patchiness may result from a combination of vesicle trafficking and dynamic barriers to lateral mobility. The simulation predicts that the steady-state patches of proteins and lipids seen on the cell surface will decay if vesicle trafficking is inhibited. To test this prediction, we compared the apparent sizes and intensities of patches of class I HLA molecules, integral membrane proteins, before and after inhibiting endocytic vesicle traffic from the cell surface, either by incubation in hypertonic medium or by expression of a dominant-negative mutant dynamin. As predicted by the simulation, the apparent sizes of HLA patches increased, whereas their intensities decreased after endocytosis and vesicle trafficking were inhibited. PMID:11423406

  2. Key Amino Acid Residues within the Third Membrane Domains of NR1 and NR2 Subunits Contribute to the Regulation of the Surface Delivery of N-methyl-d-aspartate Receptors*

    PubMed Central

    Kaniakova, Martina; Krausova, Barbora; Vyklicky, Vojtech; Korinek, Miloslav; Lichnerova, Katarina; Vyklicky, Ladislav; Horak, Martin

    2012-01-01

    N-methyl-d-aspartate (NMDA) receptors are glutamate ionotropic receptors that play critical roles in synaptic transmission, plasticity, and excitotoxicity. The functional NMDA receptors, heterotetramers composed mainly of two NR1 and two NR2 subunits, likely pass endoplasmic reticulum quality control before they are released from the endoplasmic reticulum and trafficked to the cell surface. However, the mechanism underlying this process is not clear. Using truncated and mutated NMDA receptor subunits expressed in heterologous cells, we found that the M3 domains of both NR1 and NR2 subunits contain key amino acid residues that contribute to the regulation of the number of surface functional NMDA receptors. These key residues are critical neither for the interaction between the NR1 and NR2 subunits nor for the formation of the functional receptors, but rather they regulate the early trafficking of the receptors. We also found that the identified key amino acid residues within both NR1 and NR2 M3 domains contribute to the regulation of the surface expression of unassembled NR1 and NR2 subunits. Thus, our data identify the unique role of the membrane domains in the regulation of the number of surface NMDA receptors. PMID:22711533

  3. DMSO induces dehydration near lipid membrane surfaces.

    PubMed

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

    2015-07-21

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

  4. Surface recognition elements of membrane protein oligomerization.

    PubMed

    Rath, Arianna; Deber, Charles M

    2008-02-15

    Although certain membrane proteins are functional as monomeric polypeptides, others must assemble into oligomers to carry out their biological roles. High-resolution membrane protein structures provide a valuable resource for examining the sequence features that facilitate-or preclude-assembly of membrane protein monomers into multimeric structures. Here we have utilized a data set of 28 high-resolution alpha-helical membrane protein structures comprising 32 nonredundant polypeptides to address this issue. The lipid-exposed surfaces of membrane proteins that have reached their fully assembled and functional biological units have been compared with those of the individual subunits that build quaternary structures. Though the overall amino acid composition of each set of surfaces is similar, a key distinction-the distribution of small-xxx-small motifs-delineates subunits from membrane proteins that have reached a functioning oligomeric state. Quaternary structure formation may therefore be dictated by small-xxx-small motifs that are not satisfied by intrachain contacts. PMID:17729275

  5. Tetraspanins regulate the protrusive activities of cell membrane

    SciTech Connect

    Bari, Rafijul; Guo, Qiusha; Zhongnan Hospital, Wuhan University, Wuhan ; Xia, Bing; Zhang, Yanhui H.; Giesert, Eldon E.; Levy, Shoshana; Zheng, Jie J.; Zhang, Xin A.

    2011-12-02

    Highlights: Black-Right-Pointing-Pointer Tetraspanins regulate microvillus formation. Black-Right-Pointing-Pointer Tetraspanin CD81 promotes microvillus formation. Black-Right-Pointing-Pointer Tetraspanin CD82 inhibits microvillus formation. Black-Right-Pointing-Pointer Based on this study, we extrapolated a general cellular mechanism for tetraspanins. Black-Right-Pointing-Pointer Tetraspanins engage various functions by regulating membrane protrusion morphogenesis. -- Abstract: Tetraspanins have gained increased attention due to their functional versatility. But the universal cellular mechanism that governs such versatility remains unknown. Herein we present the evidence that tetraspanins CD81 and CD82 regulate the formation and/or development of cell membrane protrusions. We analyzed the ultrastructure of the cells in which a tetraspanin is either overexpressed or ablated using transmission electron microscopy. The numbers of microvilli on the cell surface were counted, and the radii of microvillar tips and the lengths of microvilli were measured. We found that tetraspanin CD81 promotes the microvillus formation and/or extension while tetraspanin CD82 inhibits these events. In addition, CD81 enhances the outward bending of the plasma membrane while CD82 inhibits it. We also found that CD81 and CD82 proteins are localized at microvilli using immunofluorescence. CD82 regulates microvillus morphogenesis likely by altering the plasma membrane curvature and/or the cortical actin cytoskeletal organization. We predict that membrane protrusions embody a common morphological phenotype and cellular mechanism for, at least some if not all, tetraspanins. The differential effects of tetraspanins on microvilli likely lead to the functional diversification of tetraspanins and appear to correlate with their functional propensity.

  6. Proton transport via the membrane surface.

    PubMed Central

    Georgievskii, Yuri; Medvedev, Emile S; Stuchebrukhov, Alexei A

    2002-01-01

    Some proton pumps, such as cytochrome c oxidase (C(c)O), translocate protons across biological membranes at a rate that considerably exceeds the rate of proton transport to the entrance of the proton-conducting channel via bulk diffusion. This effect is usually ascribed to a proton-collecting antenna surrounding the channel entrance. In this paper, we consider a realistic phenomenological model of such an antenna. In our model, a homogeneous membrane surface, which can mediate proton diffusion toward the channel entrance, is populated with protolytic groups that are in dynamic equilibrium with the solution. Equations that describe coupled surface-bulk proton diffusion are derived and analyzed. A general expression for the rate constant of proton transport via such a coupled surface-bulk diffusion mechanism is obtained. A rigorous criterion is formulated of when proton diffusion along the surface enhances the transport. The enhancement factor is found to depend on the ratio of the surface and bulk diffusional constants, pK(a) values of surface protolytic groups, and their concentration. A capture radius for a proton on the surface and an effective size of the antenna are found. The theory also predicts the effective distance that a proton can migrate on the membrane surface between a source (such as CcO) and a sink (such as ATP synthase) without fully equilibrating with the bulk. In pure aqueous solutions, protons can travel over long distances (microns). In buffered solutions, the travel distance is much shorter (nanometers); still the enhancement effect of the surface diffusion on the proton flow to a target on the surface can be tens to hundreds at physiological buffer concentrations. These results are discussed in a general context of chemiosmotic theory. PMID:12023208

  7. Surface modification of PVDF membrane via AGET ATRP directly from the membrane surface

    NASA Astrophysics Data System (ADS)

    Meng, Jian-Qiang; Chen, Chun-Lin; Huang, Li-Ping; Du, Qi-Yun; Zhang, Yu-Feng

    2011-05-01

    This contribution demonstrates a method for PVDF microporous membrane modification via surface-initiated activators generated by electron transfer atom transfer radical polymerization (AGET ATRP) directly from the membrane surface. Three hydrophilic polymers, poly(2-(N,N-dimethylamino) ethyl methacrylate) (PDMAEMA), poly(2-oligo (ethylene glycol) monomethyl ether methacrylate) (POEGMA), and poly(2-hydroxyethyl methacrylate) (PHEMA), were grafted from the PVDF membrane surface in aqueous solution at room temperature. Attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy and X-ray photoelectron spectroscopy (XPS) confirmed the successful covalent tethering of the polymer chains onto the PVDF membrane surface. The gravimetry results indicated an approximately linear increase of the graft yields, up to about 330 μg/cm 2 for DMAEMA and 470 μg/cm 2 for both HEMA and OEGMA, with the polymerization time. Block copolymer brushes were prepared by chain extension. Water contact angle decreased over 50% for high yields, indicating improved surface hydrophilicity. The effects of the graft polymerization on membrane surface morphology, pore structure and permeability were investigated. It was found that the surface roughness was decreased and the pore size distribution was narrowed. The membrane permeability increased at low graft yields due to the enhanced hydrophilicity and decreased at high graft yields due to the overall reduction of the pore diameters.

  8. DNA Inversion Regulates Outer Membrane Vesicle Production in Bacteroides fragilis

    PubMed Central

    Nakayama-Imaohji, Haruyuki; Hirota, Katsuhiko; Yamasaki, Hisashi; Yoneda, Saori; Nariya, Hirofumi; Suzuki, Motoo; Secher, Thomas; Miyake, Yoichiro; Oswald, Eric; Hayashi, Tetsuya; Kuwahara, Tomomi

    2016-01-01

    Phase changes in Bacteroides fragilis, a member of the human colonic microbiota, mediate variations in a vast array of cell surface molecules, such as capsular polysaccharides and outer membrane proteins through DNA inversion. The results of the present study show that outer membrane vesicle (OMV) formation in this anaerobe is also controlled by DNA inversions at two distantly localized promoters, IVp-I and IVp-II that are associated with extracellular polysaccharide biosynthesis and the expression of outer membrane proteins. These promoter inversions are mediated by a single tyrosine recombinase encoded by BF2766 (orthologous to tsr19 in strain NCTC9343) in B. fragilis YCH46, which is located near IVp-I. A series of BF2766 mutants were constructed in which the two promoters were locked in different configurations (IVp-I/IVp-II = ON/ON, OFF/OFF, ON/OFF or OFF/ON). ON/ON B. fragilis mutants exhibited hypervesiculating, whereas the other mutants formed only a trace amount of OMVs. The hypervesiculating ON/ON mutants showed higher resistance to treatment with bile, LL-37, and human β-defensin 2. Incubation of wild-type cells with 5% bile increased the population of cells with the ON/ON genotype. These results indicate that B. fragilis regulates the formation of OMVs through DNA inversions at two distantly related promoter regions in response to membrane stress, although the mechanism underlying the interplay between the two regions controlled by the invertible promoters remains unknown. PMID:26859882

  9. Small membranes under negative surface tension.

    PubMed

    Avital, Yotam Y; Farago, Oded

    2015-03-28

    We use computer simulations and a simple free energy model to study the response of a bilayer membrane to the application of a negative (compressive) mechanical tension. Such a tension destabilizes the long wavelength undulation modes of giant vesicles, but it can be sustained when small membranes and vesicles are considered. Our negative tension simulation results reveal two regimes-(i) a weak negative tension regime characterized by stretching-dominated elasticity and (ii) a strong negative tension regime featuring bending-dominated elastic behavior. This resembles the findings of the classic Evans and Rawicz micropipette aspiration experiment in giant unilamellar vesicles (GUVs) [E. Evans and W. Rawicz, Phys, Rev. Lett. 64, 2094 (1990)]. However, in GUVs the crossover between the two elasticity regimes occurs at a small positive surface tension, while in smaller membranes it takes place at a moderate negative tension. Another interesting observation concerning the response of a small membrane to negative surface tension is related to the relationship between the mechanical and fluctuation tensions, which are equal to each other for non-negative values. When the tension decreases to negative values, the fluctuation tension γ drops somewhat faster than the mechanical tension τ in the small negative tension regime, before it saturates (and becomes larger than τ) for large negative tensions. The bending modulus exhibits an "opposite" trend. It remains almost unchanged in the stretching-dominated elastic regime, and decreases in the bending-dominated regime. Both the amplitudes of the thermal height undulations and the projected area variations diverge at the onset of mechanical instability. PMID:25833604

  10. Small membranes under negative surface tension

    NASA Astrophysics Data System (ADS)

    Avital, Yotam Y.; Farago, Oded

    2015-03-01

    We use computer simulations and a simple free energy model to study the response of a bilayer membrane to the application of a negative (compressive) mechanical tension. Such a tension destabilizes the long wavelength undulation modes of giant vesicles, but it can be sustained when small membranes and vesicles are considered. Our negative tension simulation results reveal two regimes—(i) a weak negative tension regime characterized by stretching-dominated elasticity and (ii) a strong negative tension regime featuring bending-dominated elastic behavior. This resembles the findings of the classic Evans and Rawicz micropipette aspiration experiment in giant unilamellar vesicles (GUVs) [E. Evans and W. Rawicz, Phys, Rev. Lett. 64, 2094 (1990)]. However, in GUVs the crossover between the two elasticity regimes occurs at a small positive surface tension, while in smaller membranes it takes place at a moderate negative tension. Another interesting observation concerning the response of a small membrane to negative surface tension is related to the relationship between the mechanical and fluctuation tensions, which are equal to each other for non-negative values. When the tension decreases to negative values, the fluctuation tension γ drops somewhat faster than the mechanical tension τ in the small negative tension regime, before it saturates (and becomes larger than τ) for large negative tensions. The bending modulus exhibits an "opposite" trend. It remains almost unchanged in the stretching-dominated elastic regime, and decreases in the bending-dominated regime. Both the amplitudes of the thermal height undulations and the projected area variations diverge at the onset of mechanical instability.

  11. Ocular Surface Membrane-Associated Mucins.

    PubMed

    Ablamowicz, Anna F; Nichols, Jason J

    2016-07-01

    Ocular surface epithelial cells produce and secrete mucins that form a hydrophilic barrier for protection and lubrication of the eye. This barrier, the glycocalyx, is formed by high molecular weight heavily glycosylated membrane-associated mucins (MAMs) that include MUC1, MUC4, and MUC16. These mucins extend into the tear film from the anterior surfaces of the conjunctiva and cornea, and, through interactions with galectin-3, prevent penetrance of pathogens into the eye. Due primarily to the glycosylation of the mucins, the glycocalyx also creates less friction during blinking and enables the tear film to maintain wetting of the eye. The secretory mucins include soluble MUC7 and gel-forming MUC5AC. These mucins, particularly MUC5AC, assist with removal of debris from the tear film and contribute to the hydrophilicity of the tear film. While new methodologies and cell culture models have expanded our understanding of mucin structure and function on the ocular surface, there is still a paucity of studies characterizing the glycosylation of MAMs on a normal ocular surface and a diseased ocular surface. Although studies have shown alterations in mucin production and expression in dry eye diseases, the relationship between changes in mucins and functional consequences is unclear. This review focuses on comparing what is known about MAMs in wet-surfaced epithelia of the body to what has been studied on the eye. PMID:27154035

  12. Vaginal epithelial cells regulate membrane adhesiveness to co-ordinate bacterial adhesion.

    PubMed

    Younes, Jessica A; Klappe, Karin; Kok, Jan Willem; Busscher, Henk J; Reid, Gregor; van der Mei, Henny C

    2016-04-01

    Vaginal epithelium is colonized by different bacterial strains and species. The bacterial composition of vaginal biofilms controls the balance between health and disease. Little is known about the relative contribution of the epithelial and bacterial cell surfaces to bacterial adhesion and whether and how adhesion is regulated over cell membrane regions. Here, we show that bacterial adhesion forces with cell membrane regions not located above the nucleus are stronger than with regions above the nucleus both for vaginal pathogens and different commensal and probiotic lactobacillus strains involved in health. Importantly, adhesion force ratios over membrane regions away from and above the nucleus coincided with the ratios between numbers of adhering bacteria over both regions. Bacterial adhesion forces were dramatically decreased by depleting the epithelial cell membrane of cholesterol or sub-membrane cortical actin. Thus, epithelial cells can regulate membrane regions to which bacterial adhesion is discouraged, possibly to protect the nucleus. PMID:26477544

  13. Membrane surface modification via polymer grafting and interfacial polymerization

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Membrane separation is an important technology for separating food ingredients and fractionating high-value substances from food processing by-products. Long-term uses of polymeric membranes in food protein processing are impeded by formation of fouled layers on the membrane surface. Surface modif...

  14. Signaling reactions on membrane surfaces: breaking the law of averages

    NASA Astrophysics Data System (ADS)

    Groves, Jay T.

    Most intracellular signal transduction reactions take place on the membrane surface. The membrane provides much more than just a surface environment on which signaling molecules are concentrated. There is a growing realization that multiple physical and chemical mechanisms allow the membrane to actively participate in the signaling reactions. Using a combination of single molecule imaging and spectroscopic techniques, my research seeks to directly resolve the actual mechanics of signaling reactions on membrane surfaces both in reconstituted systems and in living cells. These observations are revealing new insights into cellular signaling processes as well as some unexpected functional behaviors of proteins on the membrane surface.

  15. Experimental Study of a Membrane Antenna Surface Adaptive Control System

    NASA Technical Reports Server (NTRS)

    Fang, H.; Quijano, U.; Bach, V.; Hill, J.; Wang, K. W.

    2011-01-01

    Due to their ultra lightweight and high packaging efficiency, membrane reflectors are getting more and more attentions for mission architectures that need extremely large inspace deployable antennas. However how to maintain the surface shape of a membrane reflector to the instrument precision requirements is a very challenging problem. This experimental study investigated using PVDF membrane piezoelectric material as actuators to control the surface figures of membrane reflectors. The feasibility of this approach is demonstrated by several sets of test results.

  16. BIN1 regulates dynamic t-tubule membrane.

    PubMed

    Fu, Ying; Hong, TingTing

    2016-07-01

    Cardiac transverse tubules (t-tubules) are specific membrane organelles critical in calcium signaling and excitation-contraction coupling required for beat-to-beat heart contraction. T-tubules are highly branched and form an interconnected network that penetrates the myocyte interior to form junctions with the sarcoplasmic reticulum. T-tubules are selectively enriched with specific ion channels and proteins crucial in calcium transient development necessary in excitation-contraction coupling, thus t-tubules are a key component of cardiac myocyte function. In this review, we focus primarily on two proteins concentrated within the t-tubular network, the L-type calcium channel (LTCC) and associated membrane anchor protein, bridging integrator 1 (BIN1). Here, we provide an overview of current knowledge in t-tubule morphology, composition, microdomains, as well as the dynamics of the t-tubule network. Secondly, we highlight multiple aspects of BIN1-dependent t-tubule function, which includes forward trafficking of LTCCs to t-tubules, LTCC clustering at t-tubule surface, microdomain organization and regulation at t-tubule membrane, and the formation of a slow diffusion barrier within t-tubules. Lastly, we describe progress in characterizing how acquired human heart failure can be attributed to abnormal BIN1 transcription and associated t-tubule remodeling. Understanding BIN1-regulated cardiac t-tubule biology in human heart failure management has the dual benefit of promoting progress in both biomarker development and therapeutic target identification. This article is part of a Special Issue entitled: Cardiomyocyte Biology: Integration of Developmental and Environmental Cues in the Heart edited by Marcus Schaub and Hughes Abriel. PMID:26578114

  17. Intravacuolar Membranes Regulate CD8 T Cell Recognition of Membrane-Bound Toxoplasma gondii Protective Antigen.

    PubMed

    Lopez, Jodie; Bittame, Amina; Massera, Céline; Vasseur, Virginie; Effantin, Grégory; Valat, Anne; Buaillon, Célia; Allart, Sophie; Fox, Barbara A; Rommereim, Leah M; Bzik, David J; Schoehn, Guy; Weissenhorn, Winfried; Dubremetz, Jean-François; Gagnon, Jean; Mercier, Corinne; Cesbron-Delauw, Marie-France; Blanchard, Nicolas

    2015-12-15

    Apicomplexa parasites such as Toxoplasma gondii target effectors to and across the boundary of their parasitophorous vacuole (PV), resulting in host cell subversion and potential presentation by MHC class I molecules for CD8 T cell recognition. The host-parasite interface comprises the PV limiting membrane and a highly curved, membranous intravacuolar network (IVN) of uncertain function. Here, using a cell-free minimal system, we dissect how membrane tubules are shaped by the parasite effectors GRA2 and GRA6. We show that membrane association regulates access of the GRA6 protective antigen to the MHC I pathway in infected cells. Although insertion of GRA6 in the PV membrane is key for immunogenicity, association of GRA6 with the IVN limits presentation and curtails GRA6-specific CD8 responses in mice. Thus, membrane deformations of the PV regulate access of antigens to the MHC class I pathway, and the IVN may play a role in immune modulation. PMID:26628378

  18. Biofouling behavior and performance of forward osmosis membranes with bioinspired surface modification in osmotic membrane bioreactor.

    PubMed

    Li, Fang; Cheng, Qianxun; Tian, Qing; Yang, Bo; Chen, Qianyuan

    2016-07-01

    Forward osmosis (FO) has received considerable interest for water and energy related applications in recent years. Biofouling behavior and performance of cellulose triacetate (CTA) forward osmosis membranes with bioinspired surface modification via polydopamine (PD) coating and poly (ethylene glycol) (PEG) grafting (PD-g-PEG) in a submerged osmotic membrane bioreactor (OMBR) were investigated in this work. The modified membranes exhibited lower flux decline than the pristine one in OMBR, confirming that the bioinspired surface modification improved the antifouling ability of the CTA FO membrane. The result showed that the decline of membrane flux related to the increase of the salinity and MLSS concentration of the mixed liquid. It was concluded that the antifouling ability of modified membranes ascribed to the change of surface morphology in addition to the improvement of membrane hydrophilicity. The bioinspired surface modifications might improve the anti-adhesion for the biopolymers and biocake. PMID:27089532

  19. Regulation of membrane trafficking by signalling on endosomal and lysosomal membranes

    PubMed Central

    Li, Xinran; Garrity, Abigail G; Xu, Haoxing

    2013-01-01

    Endosomal and lysosomal membrane trafficking requires the coordination of multiple signalling events to control cargo sorting and processing, and endosome maturation. The initiation and termination of signalling events in endosomes and lysosomes is not well understood, but several key regulators have been identified, which include small GTPases, phosphoinositides, and Ca2+. Small GTPases act as master regulators and molecular switches in a GTP-dependent manner, initiating signalling cascades to regulate the direction and specificity of endosomal trafficking. Phosphoinositides are membrane-bound lipids that indicate vesicular identities for recruiting specific cytoplasmic proteins to endosomal membranes, thus allowing specificity of membrane fusion, fission, and cargo sorting to occur within and between specific vesicle compartments. In addition, phosphoinositides regulate the function of membrane proteins such as ion channels and transporters in a compartment-specific manner to mediate transport and signalling. Finally, Ca2+, a locally acting second messenger released from intracellular ion channels, may provide precise spatiotemporal regulation of endosomal signalling and trafficking events. Small GTPase signalling can regulate phosphoinositide conversion during endosome maturation, and electrophysiological studies on isolated endosomes have shown that endosomal and lysosomal Ca2+ channels are directly modulated by endosomal lipids. Thus trafficking and maturation of endosomes and lysosomes can be precisely regulated by dynamic changes in GTPases and membrane lipids, as well as Ca2+ signalling. Importantly, impaired phosphoinositide and Ca2+ signalling can cause endosomal and lysosomal trafficking defects at the cellular level, and a spectrum of lysosome storage diseases. PMID:23878375

  20. Composite Membrane with Underwater-Oleophobic Surface for Anti-Oil-Fouling Membrane Distillation.

    PubMed

    Wang, Zhangxin; Hou, Deyin; Lin, Shihong

    2016-04-01

    In this study, we fabricated a composite membrane for membrane distillation (MD) by modifying a commercial hydrophobic polyvinylidene fluoride (PVDF) membrane with a nanocomposite coating comprising silica nanoparticles, chitosan hydrogel and fluoro-polymer. The composite membrane exhibits asymmetric wettability, with the modified surface being in-air hydrophilic and underwater oleophobic, and the unmodified surface remaining hydrophobic. By comparing the performance of the composite membrane and the pristine PVDF membrane in direct contact MD experiments using a saline emulsion with 1000 ppm crude oil (in water), we showed that the fabricated composite membrane was significantly more resistant to oil fouling compared to the pristine hydrophobic PVDF membrane. Force spectroscopy was conducted for the interaction between an oil droplet and the membrane surface using a force tensiometer. The difference between the composite membrane and the pristine PVDF membrane in their interaction with an oil droplet served to explain the difference in the fouling propensities between these two membranes observed in MD experiments. The results from this study suggest that underwater oleophobic coating can effectively mitigate oil fouling in MD operations, and that the fabricated composite membrane with asymmetric wettability can enable MD to desalinate hypersaline wastewater with high concentrations of hydrophobic contaminants. PMID:26958985

  1. Cell cycle regulation of Golgi membrane dynamics.

    PubMed

    Tang, Danming; Wang, Yanzhuang

    2013-06-01

    The Golgi apparatus is a membranous organelle in the cell that plays essential roles in protein and lipid trafficking, sorting, processing, and modification. Its basic structure is a stack of closely aligned flattened cisternae. In mammalian cells, dozens of Golgi stacks are often laterally linked into a ribbon-like structure. Biogenesis of the Golgi during cell division occurs through a sophisticated disassembly and reassembly process that can be divided into three distinct but cooperative steps, including the deformation and reformation of the Golgi cisternae, stacks, and ribbon. Here, we review our current understanding of the protein machineries that control these three steps in the cycle of mammalian cell division: GRASP65 and GRASP55 in Golgi stack and ribbon formation; ubiquitin and AAA ATPases in postmitotic Golgi membrane fusion; and golgins and cytoskeleton in Golgi ribbon formation. PMID:23453991

  2. MAL Is a Regulator of the Recruitment of Myelin Protein PLP to Membrane Microdomains

    PubMed Central

    Bijlard, Marjolein; de Jonge, Jenny C.; Klunder, Bert; Nomden, Anita; Hoekstra, Dick; Baron, Wia

    2016-01-01

    In oligodendrocytes (OLGs), an indirect, transcytotic pathway is mediating transport of de novo synthesized PLP, a major myelin specific protein, from the apical-like plasma membrane to the specialized basolateral-like myelin membrane to prevent its premature compaction. MAL is a well-known regulator of polarized trafficking in epithelial cells, and given its presence in OLGs it was therefore of interest to investigate whether MAL played a similar role in PLP transport in OLGs, taking into account its timely expression in these cells. Our data revealed that premature expression of mCherry-MAL in oligodendrocyte progenitor cells interfered with terminal OLG differentiation, although myelin membrane formation per se was not impaired. In fact, also PLP transport to myelin membranes via the cell body plasma membrane was unaffected. However, the typical shift of PLP from TX-100-insoluble membrane domains to CHAPS-resistant, but TX-100-soluble membrane domains, seen in the absence of MAL expression, is substantially reduced upon expression of the MAL protein. Interestingly, not only in vitro, but also in developing brain a strongly diminished shift from TX-100 resistant to TX-100 soluble domains was observed. Consistently, the MAL-expression mediated annihilation of the typical membrane microdomain shift of PLP is also reflected by a loss of the characteristic surface expression profile of conformation-sensitive anti-PLP antibodies. Hence, these findings suggest that MAL is not involved in vesicular PLP trafficking to either the plasma membrane and/or the myelin membrane as such. Rather, we propose that MAL may regulate PLP’s distribution into distinct membrane microdomains that allow for lateral diffusion of PLP, directly from the plasma membrane to the myelin membrane once the myelin sheath has been assembled. PMID:27171274

  3. Protein-Induced Surface Structuring in Myelin Membrane Monolayers

    PubMed Central

    Rosetti, Carla M.; Maggio, Bruno

    2007-01-01

    Monolayers prepared from myelin conserve all the compositional complexity of the natural membrane when spread at the air-water interface. They show a complex pressure-dependent surface pattern that, on compression, changes from the coexistence of two liquid phases to a viscous fractal phase embedded in a liquid phase. We dissected the role of major myelin protein components, myelin basic protein (MBP), and Folch-Lees proteolipid protein (PLP) as crucial factors determining the structural dynamics of the interface. By analyzing mixtures of a single protein with the myelin lipids we found that MBP and PLP have different surface pressure-dependent behaviors. MBP stabilizes the segregation of two liquid phases at low pressures and becomes excluded from the film under compression, remaining adjacent to the interface. PLP, on the contrary, organizes a fractal-like pattern at all surface pressures when included in a monolayer of the protein-free myelin lipids but it remains mixed in the MBP-induced liquid phase. The resultant surface topography and dynamics is regulated by combined near to equilibrium and out-of-equilibrium effects. PLP appears to act as a surface skeleton for the whole components whereas MBP couples the structuring to surface pressure-dependent extrusion and adsorption processes. PMID:17905850

  4. Influence of membrane surface roughness on interfacial interactions with sludge flocs in a submerged membrane bioreactor.

    PubMed

    Zhao, Leihong; Shen, Liguo; He, Yiming; Hong, Huachang; Lin, Hongjun

    2015-05-15

    In this study, the interfacial interactions between sludge flocs and a rough membrane surface in a submerged membrane bioreactor were investigated. Models describing these interfacial interactions were firstly proposed based on the surface element integration (SEI) method. Surface properties of sludge flocs and membrane were experimentally determined to simulate the models through composite Simpson's rule. It was found that, roughness on membrane surface significantly decreased interaction strength, which enabled the sludge flocs to more easily attach on and detach from the rough membrane surface. Further analysis showed that the value of total interaction energy increased with asperity radius, while the strength of total interaction energy decreased with asperity height. Results also demonstrated that increase in floc size would significantly decrease the attractive specific total interaction with rough membrane surface. It was revealed that there existed a critical asperity radius above which the total interaction energy in certain separation distance coverage was continuously repulsive, facilitating membrane fouling control in MBRs. This study demonstrated the possibility to mitigate membrane fouling by "tailoring" membrane surface roughness. PMID:25660708

  5. Proline transport across the intestinal microvillus membrane may be regulated by membrane physical properties.

    PubMed

    Sadowski, D C; Gibbs, D J; Meddings, J B

    1992-03-23

    There is now abundant evidence that integral membrane protein function may be modulated by the physical properties of membrane lipids. The intestinal brush border membrane represents a membrane system highly specialized for nutrient absorption and, thus, provides an opportunity to study the interaction between integral membrane transport proteins and their lipid environment. We have previously demonstrated that alterations in this environment may modulate the function of the sodium-dependent glucose transporter in terms of its affinity for glucose. In this communication we report that membrane lipid-protein interactions are distinctly different for the proline transport proteins. Maximal transport rates for L-proline by either the neutral brush border or imino transport systems are reduced 10-fold when the surrounding membrane environment is made more fluid over the physiological range that exists along the crypt-villus axis. Furthermore, in microvillus membrane vesicles prepared from enterocytes isolated from along the crypt-villus axis a similar gradient exists in the functional activity of these transport systems. This would imply that either the functional activity of these transporters are regulated by membrane physical properties or that the synthesis and insertion of these proteins is coordinated in concert with membrane physical properties as the enterocyte migrates up the crypt-villus axis. PMID:1567897

  6. Role of Phosphatidylinositol 4,5-Bisphosphate in Regulating EHD2 Plasma Membrane Localization

    PubMed Central

    Simone, Laura C.; Caplan, Steve; Naslavsky, Naava

    2013-01-01

    The four mammalian C-terminal Eps15 homology domain-containing proteins (EHD1-EHD4) play pivotal roles in endocytic membrane trafficking. While EHD1, EHD3 and EHD4 associate with intracellular tubular/vesicular membranes, EHD2 localizes to the inner leaflet of the plasma membrane. Currently, little is known about the regulation of EHD2. Thus, we sought to define the factors responsible for EHD2’s association with the plasma membrane. The subcellular localization of endogenous EHD2 was examined in HeLa cells using confocal microscopy. Although EHD partner proteins typically mediate EHD membrane recruitment, EHD2 was targeted to the plasma membrane independent of two well-characterized binding proteins, syndapin2 and EHBP1. Additionally, the EH domain of EHD2, which facilitates canonical EHD protein interactions, was not required to direct overexpressed EHD2 to the cell surface. On the other hand, several lines of evidence indicate that the plasma membrane phospholipid phosphatidylinositol 4,5-bisphosphate (PIP2) plays a crucial role in regulating EHD2 subcellular localization. Pharmacologic perturbation of PIP2 metabolism altered PIP2 plasma membrane distribution (as assessed by confocal microscopy), and caused EHD2 to redistribute away from the plasma membrane. Furthermore, overexpressed EHD2 localized to PIP2-enriched vacuoles generated by active Arf6. Finally, we show that although cytochalasin D caused actin microfilaments to collapse, EHD2 was nevertheless maintained at the plasma membrane. Intriguingly, cytochalasin D induced relocalization of both PIP2 and EHD2 to actin aggregates, supporting a role of PIP2 in controlling EHD2 subcellular localization. Altogether, these studies emphasize the significance of membrane lipid composition for EHD2 subcellular distribution and offer new insights into the regulation of this important endocytic protein. PMID:24040268

  7. Blood coagulation reactions on nanoscale membrane surfaces

    NASA Astrophysics Data System (ADS)

    Pureza, Vincent S.

    Blood coagulation requires the assembly of several membrane-bound protein complexes composed of regulatory and catalytic subunits. The biomembranes involved in these reactions not only provide a platform for these procoagulant proteins, but can also affect their function. Increased exposure of acidic phospholipids on the outer leaflet of the plasma membrane can dramatically modulate the catalytic efficiencies of such membrane-bound enzymes. Under physiologic conditions, however, these phospholipids spontaneously cluster into a patchwork of membrane microdomains upon which membrane binding proteins may preferentially assemble. As a result, the membrane composition surrounding these proteins is largely unknown. Through the development and use of a nanometer-scale bilayer system that provides rigorous control of the phospholipid membrane environment, I investigated the role of phosphatidylserine, an acidic phospholipid, in the direct vicinity (within nanometers) of two critical membrane-bound procoagulant protein complexes and their respective natural substrates. Here, I present how the assembly and function of the tissue factor˙factor VIIa and factor Va˙factor Xa complexes, the first and final cofactor˙enzyme complexes of the blood clotting cascade, respectively, are mediated by changes in their immediate phospholipid environments.

  8. EH domain proteins regulate cardiac membrane protein targeting

    PubMed Central

    Gudmundsson, Hjalti; Hund, Thomas J.; Wright, Patrick J.; Kline, Crystal F.; Snyder, Jedidiah S.; Qian, Lan; Koval, Olha M.; Cunha, Shane R.; George, Manju; Rainey, Mark A.; Kashef, Farshid E.; Dun, Wen; Boyden, Penelope A.; Anderson, Mark E.; Band, Hamid; Mohler, Peter J.

    2010-01-01

    Rationale Cardiac membrane excitability is tightly regulated by an integrated network of membrane-associated ion channels, transporters, receptors, and signaling molecules. Membrane protein dynamics in health and disease are maintained by a complex ensemble of intracellular targeting, scaffolding, recycling, and degradation pathways. Surprisingly, despite decades of research linking dysfunction in membrane protein trafficking with human cardiovascular disease, essentially nothing is known regarding the molecular identity or function of these intracellular targeting pathways in excitable cardiomyocytes. Objective We sought to discover novel pathways for membrane protein targeting in primary cardiomyocytes. Methods and Results We report the initial characterization of a large family of membrane trafficking proteins in human heart. We employed a tissue-wide screen for novel ankyrin-associated trafficking proteins and identified four members of a unique Eps15 homology (EH) domain-containing protein family (EHD1, EHD2, EHD3, EHD4) that serve critical roles in endosome-based membrane protein targeting in other cell types. We show that EHD1-4 directly associate with ankyrin, provide the first information on the expression and localization of these molecules in primary cardiomyocytes, and demonstrate that EHD1-4 are co-expressed with ankyrin-B in the myocyte perinuclear region. Notably, the expression of multiple EHD proteins is increased in animal models lacking ankyrin-B, and EHD3-deficient cardiomyocytes display aberrant ankyrin-B localization and selective loss of Na/Ca exchanger expression and function. Finally, we report significant modulation of EHD expression following myocardial infarction, suggesting that these proteins may play a key role in regulating membrane excitability in normal and diseased heart. Conclusions Our findings identify and characterize a new class of cardiac trafficking proteins, define the first group of proteins associated with the ankyrin

  9. Probing a Membrane Matrix Regulating Hormone Action

    PubMed Central

    Stowe, Bruce B.; Dotts, Mary Ann

    1971-01-01

    Auxin-induced pea (Pisum sativum) stem section elongation is enhanced at levels of 3 to 40 micromolar by six new classes of alkane derivatives additional to those described earlier, providing that length of their molecules exceeds 20 A. Increasingly longer homologous series of alkyl chlorides, bromides and iodides, alkyl benzenes, dialkyl ethers, and sulfides show a logarithmically linear increase in specific activity above this length, reaching an optimum near 28 to 30 Å. Longer dialkyl ethers and sulfides are less effective, while steroids, or alkanes with substituents at both ends, are ineffective. Neither common metabolism nor common physical properties, other than over-all length of active molecules, seem to explain these results. However, the dimensions of the most abundant phospholipid of etiolated peas, 1-palmitoyl-2-linoleoyl-3-sn-phosphatidyl choline are such that a monolayer of this lecithin would contain cavities 20 Å long. It is postulated that lipids of this length or longer are active in the pea assay by forcing apart lecithin molecules, changing the charge distribution or chelating properties of a regulatory membrane. Steroid dimensions also match the cavity in lecithin monolayers and steroids would be maintained within such a cavity by hydrogen bonds. The name lipometrin for lipids whose physiological activity varies with length is proposed, among these, substances active in the pea bioassay could be termed oleanimins. PMID:16657838

  10. Oriented adsorption of purple membrane to cationic surfaces.

    PubMed

    Fisher, K A; Yanagimoto, K; Stoeckenius, W

    1978-05-01

    We have investigated the orientation of isolated fragments of Halobacterium halobium purple membrane (PM) adsorbed to poly-L-lysine-treated glass (PL-glass), by quanitative electron microscopy. Three lines of evidence support the conclusion that the cytoplasmic side of the membrane is preferentially absorbed. First, monolayer freeze-fracture reveals nonrandom orientation; more fracture faces (89%) are particulate than smooth. Second, the amount of each membrane surface present can be assayed using polycationic ferritin; 90% of all adsorbed membrane fragments are labeled. Third, it is possible to distinguish two surfaces, "cracked" (the extracellular surface) and "pitted" (the cytoplasmic surface) , in slowly air-dried, platinum-carbon-shadowed membranes. When applied under standard conditions, more than 80% appear cracked. Selection for the cytoplasmic by the cationic substrate suggests that the isolated PM, buffered at pH 7.4 and in the light, has a higher negative charge on its cytoplasmic surface than on its extracellular surface. Nevertheless, cationic ferritin (CF) preferentially adsorbs to the extracellular surface. Orientation provides a striking example of biomembrane surface asymmetry as well as the means to examine the chemical reactivity and physical properties of surfaces of a purified, nonvesicular membrane fragment. PMID:649662

  11. Reduced fouling of ultrafiltration membranes via surface fluorination

    SciTech Connect

    Sedath, R.H.; Yates, S.F.; Li, N.N.

    1993-03-01

    Surface fluorination can affect significantly the performance of an ultrafiltration membrane used to concentrate a food-related stream. Membranes fluorinated and tested as flat sheets exhibit higher initial fluxes, and do not foul as rapidly as untreated membranes. This improvement is linked to increased surface hydrophilicity, as shown in decreased contact angle with water. This increased hydrophilicity, in turn, is linked to the addition of fluorine and oxygen to the surface. The pilot plant study did-not show the difference in membrane flux and fouling observed in the flat sheet study. Instead, fluorinated and unfluorinated modules behaved similarly. Fouling by potato waste feed was severe and resulted in formation of an extensive gel layer within the module on the membrane surface. XPS, SEM and FTIR indicate that buildup of organic material occurred on both fluorinated and unfluorinated membranes, but SEM indicates that a fibrous mat of material was observed only on the nonfluorinated membrane. We conclude that in the pilot study, membrane fouling and gel formation were so extensive that the surface interaction effect was overwhelmed.

  12. Intermembrane Docking Reactions Are Regulated by Membrane Curvature

    PubMed Central

    Kunding, Andreas H.; Mortensen, Michael W.; Christensen, Sune M.; Bhatia, Vikram K.; Makarov, Ivan; Metzler, Ralf; Stamou, Dimitrios

    2011-01-01

    The polymorphism of eukaryotic cellular membranes is a tightly regulated and well-conserved phenotype. Recent data have revealed important regulatory roles of membrane curvature on the spatio-temporal localization of proteins and in membrane fusion. Here we quantified the influence of membrane curvature on the efficiency of intermembrane docking reactions. Using fluorescence microscopy, we monitored the docking of single vesicle–vesicle pairs of different diameter (30–200 nm) and therefore curvature, as mediated by neuronal soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs) and streptavidin-biotin. Surprisingly, the intermembrane docking efficiency exhibited an ∼30–60 fold enhancement as a function of curvature. In comparison, synaptotagmin and calcium accelerate SNARE-mediated fusion in vitro by a factor of 2–10. To explain this finding, we formulated a biophysical model. On the basis of our findings, we propose that membrane curvature can regulate intermembrane tethering reactions and consequently any downstream process, including the fusion of vesicles and possibly viruses with their target membranes. PMID:22261058

  13. High Surface Area Inorganic Membrane for Water Removal

    SciTech Connect

    2008-12-01

    This factsheet describes a research project whose objective is to demonstrate the fabrication and performance advantages of minichannel planar membrane modules made of porous metallic supports of surface area packing density one order of magnitude higher than the conventional membrane tube. The new, transformational, ceramic/metallic, hybrid membrane technology will be used for water/ethanol separations and reduce energy consumption by >20% over distillation and adsorption.

  14. Probing a Membrane Matrix Regulating Hormone Action

    PubMed Central

    Iwata, Takashi; Stowe, Bruce B.

    1973-01-01

    heptadecylbenzene is added after 6 hours. Applied ethylene even at concentrations much above those produced by the tissue itself fails to stimulate or inhibit short term pea stem section growth. It is concluded that the effect of oleanmins on growth is not mediated by ethylene. The similarities in concentration and molecular dimensions of these structurally diverse lipids which simultaneously stimulate respiration, growth, and ethylene formation, suggest a single site of action located in a regulatory membrane. PMID:16658394

  15. Geometrical Membrane Curvature as an Allosteric Regulator of Membrane Protein Structure and Function

    PubMed Central

    Tonnesen, Asger; Christensen, Sune M.; Tkach, Vadym; Stamou, Dimitrios

    2014-01-01

    Transmembrane proteins are embedded in cellular membranes of varied lipid composition and geometrical curvature. Here, we studied for the first time the allosteric effect of geometrical membrane curvature on transmembrane protein structure and function. We used single-channel optical analysis of the prototypic transmembrane β-barrel α-hemolysin (α-HL) reconstituted on immobilized single small unilamellar liposomes of different diameter and therefore curvature. Our data demonstrate that physiologically abundant geometrical membrane curvatures can enforce a dramatic allosteric regulation (1000-fold inhibition) of α-HL permeability. High membrane curvatures (1/diameter ∼1/40 nm−1) compressed the effective pore diameter of α-HL from 14.2 ± 0.8 Å to 11.4 ± 0.6 Å. This reduction in effective pore area (∼40%) when combined with the area compressibility of α-HL revealed an effective membrane tension of ∼50 mN/m and a curvature-imposed protein deformation energy of ∼7 kBT. Such substantial energies have been shown to conformationally activate, or unfold, β-barrel and α-helical transmembrane proteins, suggesting that membrane curvature could likely regulate allosterically the structure and function of transmembrane proteins in general. PMID:24411252

  16. Dynamic DNA methylation regulates neuronal intrinsic membrane excitability.

    PubMed

    Meadows, Jarrod P; Guzman-Karlsson, Mikael C; Phillips, Scott; Brown, Jordan A; Strange, Sarah K; Sweatt, J David; Hablitz, John J

    2016-01-01

    Epigenetic modifications, such as DNA cytosine methylation, contribute to the mechanisms underlying learning and memory by coordinating adaptive gene expression and neuronal plasticity. Transcription-dependent plasticity regulated by DNA methylation includes synaptic plasticity and homeostatic synaptic scaling. Memory-related plasticity also includes alterations in intrinsic membrane excitability mediated by changes in the abundance or activity of ion channels in the plasma membrane, which sets the threshold for action potential generation. We found that prolonged inhibition of DNA methyltransferase (DNMT) activity increased intrinsic membrane excitability of cultured cortical pyramidal neurons. Knockdown of the cytosine demethylase TET1 or inhibition of RNA polymerase blocked the increased membrane excitability caused by DNMT inhibition, suggesting that this effect was mediated by subsequent cytosine demethylation and de novo transcription. Prolonged DNMT inhibition blunted the medium component of the after-hyperpolarization potential, an effect that would increase neuronal excitability, and was associated with reduced expression of the genes encoding small-conductance Ca(2+)-activated K(+) (SK) channels. Furthermore, the specific SK channel blocker apamin increased neuronal excitability but was ineffective after DNMT inhibition. Our results suggested that DNMT inhibition enables transcriptional changes that culminate in decreased expression of SK channel-encoding genes and decreased activity of SK channels, thus providing a mechanism for the regulation of neuronal intrinsic membrane excitability by dynamic DNA cytosine methylation. This study has implications for human neurological and psychiatric diseases associated with dysregulated intrinsic excitability. PMID:27555660

  17. Leukocyte Cell Surface Proteinases: Regulation of Expression, Functions, and Mechanisms of Surface Localization

    PubMed Central

    Owen, Caroline A.

    2008-01-01

    A number of proteinases are expressed on the surface of leukocytes including members of the serine, metallo-, and cysteine proteinase superfamilies. Some proteinases are anchored to the plasma membrane of leukocytes by a transmembrane domain or a glycosyl phosphatidyl inositol (GPI) anchor. Other proteinases bind with high affinity to classical receptors, or with lower affinity to integrins, proteoglycans, or other leukocyte surface molecules. Leukocyte surface levels of proteinases are regulated by: 1) cytokines, chemokines, bacterial products, and growth factors which stimulate synthesis and/or release of proteinase by cells; 2) the availability of surface binding sites for proteinases; and/or 3) internalization or shedding of surface-bound proteinases. The binding of proteinases to leukocyte surfaces serves many functions including: 1) concentrating the activity of proteinases to the immediate pericellular environment; 2) facilitating pro-enzyme activation; 3) increasing proteinase stability and retention in the extracellular space; 4) regulating leukocyte function by proteinases signaling through cell surface binding sites or other surface proteins; and 5) protecting proteinases from inhibition by extracellular proteinase inhibitors. There is strong evidence that membrane-associated proteinases on leukocytes play critical roles in wound healing, inflammation, extracellular matrix remodeling, fibrinolysis, and coagulation. This review will outline the biology of membrane-associated proteinases expressed by leukocytes and their roles in physiologic and pathologic processes. PMID:18329945

  18. Extracellular Protease Digestion to Evaluate Membrane Protein Cell Surface Localization

    PubMed Central

    Besingi, Richard N.; Clark, Patricia L.

    2016-01-01

    Membrane proteins play crucial roles in signaling and as anchors for cell surface display. Proper secretion of a membrane protein can be evaluated by its susceptibility to digestion by an extracellular protease, but this requires a crucial control to confirm membrane integrity during digestion. This protocol describes how to use this approach to determine how efficiently a protein is secreted to the outer surface of Gram-negative bacteria. Its success relies upon careful selection of an appropriate intracellular reporter protein that will remain undigested if the membrane barrier remains intact, but is rapidly digested when cells are lysed prior to evaluation. Reporter proteins that are resistant to proteases (e.g. maltose-binding protein) do not return accurate results; in contrast, proteins that are more readily digested (e.g. SurA) serve as more sensitive reporters of membrane integrity, yielding more accurate measurements of membrane protein localization. Similar considerations apply when evaluating membrane protein localization in other contexts, including eukaryotic cells and organelle membranes. Evaluating membrane protein localization using this approach requires only standard biochemistry laboratory equipment for cell lysis, gel electrophoresis and western blotting. After expression of the protein of interest, this procedure can be completed in 4 h. PMID:26584447

  19. Peptide-modified PELCL electrospun membranes for regulation of vascular endothelial cells.

    PubMed

    Zhou, Fang; Jia, Xiaoling; Yang, Yang; Yang, Qingmao; Gao, Chao; Zhao, Yunhui; Fan, Yubo; Yuan, Xiaoyan

    2016-11-01

    The efficiency of biomaterials used in small vascular repair depends greatly on their ability to interact with vascular endothelial cells (VECs). Rapid endothelialization of the vascular grafts is a promising way to prevent thrombosis and intimal hyperplasia. In this work, modification of electrospun membranes of poly(ethylene glycol)-b-poly(l-lactide-co-ε-caprolactone) (PELCL) by three different peptides for regulation of VECs were studied in order to obtain ideal bioactive biomaterials as small diameter vascular grafts. QK (a mimetic peptide to vascular endothelial growth factor), Arg-Glu-Asp-Val (REDV, a specific adhesive peptide to VECs) and Val-Ala-Pro-Gly (VAPG, a specific adhesive peptide to vascular smooth muscle cells) were investigated. Surface properties of the modified membranes and the response of VECs were verified. It was found that protein adsorption and platelet adhesion were effectively suppressed with the introduction of QK, REDV or VAPG peptides on the PELCL electrospun membranes. Both QK- and REDV-modified electrospun membranes could accelerate the proliferation of VECs in the first 9days, and the QK-modified electrospun membrane promoted cell proliferation more significantly than the REDV-modified one. The REDV-modified PELCL membrane was the most favorable for VECs adhesion than QK- and VAPG-modified membranes. It was suggested that QK- or REDV-modified PELCL electrospun membranes may have great potential applications in cardiovascular biomaterials for rapid endothelialization in situ. PMID:27524062

  20. Imaging membrane remodeling during regulated exocytosis in live mice.

    PubMed

    Shitara, Akiko; Weigert, Roberto

    2015-10-01

    In this mini-review we focus on the use of time-lapse light microscopy to study membrane remodeling during protein secretion in live animals. In particular, we highlight how subcellular intravital microscopy has enabled imaging the dynamics of both individual secretory vesicles and the plasma membrane, during different steps in the exocytic process. This powerful approach has provided us with the unique opportunity to unravel the role of the actin cytoskeleton in regulating this process under physiological conditions, and to overcome the shortcomings of more reductionist model systems. PMID:26160452

  1. Regulation of protein mobility via thermal membrane undulations.

    PubMed

    Brown, Frank L H

    2003-02-01

    The in-plane diffusivelike motion of membrane bound proteins on the surface of cells is considered. We suggest, on the basis of theoretical arguments and simulation, that thermally excited undulations of the lipid bilayer may serve as a mechanism for proteins to hop between adjacent regions on the cell surface separated by barriers composed of internal cellular structure (e.g., the cytoskeleton). We specifically investigate the mobility of band 3 dimer on the surface of red blood cells where the spectrin cytoskeletal meshwork defines a series of "corrals" on the cell surface known to hinder protein motion. Previous models of this system have postulated that the cytoskeleton must deform to allow passage of membrane bound proteins out of these corral regions and have ignored fluctuations of the bilayer. Our model provides a complementary mechanism and we posit that the mobility of real proteins in real cells is likely the result of several mechanisms acting in parallel. PMID:12547768

  2. Flavonoid-membrane Interactions: A Protective Role of Flavonoids at the Membrane Surface?

    PubMed Central

    Oteiza, Patricia I.; Erlejman, Alejandra G.; Verstraeten, Sandra V.; Keen, Carl L.; Fraga, César G.

    2005-01-01

    Flavonoids can exert beneficial health effects through multiple mechanisms. In this paper, we address the important, although not fully understood, capacity of flavonoids to interact with cell membranes. The interactions of polyphenols with bilayers include: (a) the partition of the more non-polar compounds in the hydrophobic interior of the membrane, and (b) the formation of hydrogen bonds between the polar head groups of lipids and the more hydrophilic flavonoids at the membrane interface. The consequences of these interactions are discussed. The induction of changes in membrane physical properties can affect the rates of membrane lipid and protein oxidation. The partition of certain flavonoids in the hydrophobic core can result in a chain breaking antioxidant activity. We suggest that interactions of polyphenols at the surface of bilayers through hydrogen bonding, can act to reduce the access of deleterious molecules (i.e. oxidants), thus protecting the structure and function of membranes. PMID:15712595

  3. Development of nanocomposite membranes with photocatalytic surfaces.

    PubMed

    Gehrke, I; Keuter, V; Gross, F

    2012-12-01

    In 2030, 47% of the world population will be living in areas of high water stress (UN World Water Development Report 3, 2009). The global water shortage results in an emerging need for novel, more efficient and cost saving water purification methods. Membrane technology with its very low use of chemical substances and secure retention of contaminants represents a promising water cleaning method. The approach presented focuses on the application of nanocoating procedures to membrane processes which aims at reducing the fouling potential of membranes and destroying water contaminants. Photocatalytic titanium dioxide nanoparticles are applied. Novel suitable microsieves including a homogeneous pore size distribution and high robustness were developed from metallic material. An appropriate multilayer coating system was developed and applied on a new nanocomposite filter resulting in high photocatalytic activities with maximum photon efficiencies of 0.0733. The coating layer do not have a negative influence on the permeate flux. The effectiveness of the nanocomposite filter was proven in a 12 days biofouling experiment with activated sludge suspensions. PMID:23447972

  4. Factors regulating the abundance and localization of synaptobrevin in the plasma membrane

    PubMed Central

    Dittman, Jeremy S.; Kaplan, Joshua M.

    2006-01-01

    After synaptic vesicle fusion, vesicle proteins must be segregated from plasma membrane proteins and recycled to maintain a functional vesicle pool. We monitored the distribution of synaptobrevin, a vesicle protein required for exocytosis, in Caenorhabditis elegans motor neurons by using a pH-sensitive synaptobrevin GFP fusion protein, synaptopHluorin. We estimated that 30% of synaptobrevin was present in the plasma membrane. By using a panel of endocytosis and exocytosis mutants, we found that the majority of surface synaptobrevin derives from fusion of synaptic vesicles and that, in steady state, synaptobrevin equilibrates throughout the axon. The surface synaptobrevin was enriched near active zones, and its spatial extent was regulated by the clathrin adaptin AP180. These results suggest that there is a plasma membrane reservoir of synaptobrevin that is supplied by the synaptic vesicle cycle and available for retrieval throughout the axon. The size of the reservoir is set by the relative rates of exo- and endocytosis. PMID:16844789

  5. An Ion Switch Regulates Fusion of Charged Membranes

    PubMed Central

    Siepi, Evgenios; Lutz, Silke; Meyer, Sylke; Panzner, Steffen

    2011-01-01

    Here we identify the recruitment of solvent ions to lipid membranes as the dominant regulator of lipid phase behavior. Our data demonstrate that binding of counterions to charged lipids promotes the formation of lamellar membranes, whereas their absence can induce fusion. The mechanism applies to anionic and cationic liposomes, as well as the recently introduced amphoteric liposomes. In the latter, an additional pH-dependent lipid salt formation between anionic and cationic lipids must occur, as indicated by the depletion of membrane-bound ions in a zone around pH 5. Amphoteric liposomes fuse under these conditions but form lamellar structures at both lower and higher pH values. The integration of these observations into the classic lipid shape theory yielded a quantitative link between lipid and solvent composition and the physical state of the lipid assembly. The key parameter of the new model, κ(pH), describes the membrane phase behavior of charged membranes in response to their ion loading in a quantitative way. PMID:21575575

  6. Mitochondrial membrane potential is regulated by vimentin intermediate filaments

    PubMed Central

    Chernoivanenko, Ivan S.; Matveeva, Elena A.; Gelfand, Vladimir I.; Goldman, Robert D.; Minin, Alexander A.

    2015-01-01

    This study demonstrates that the association of mitochondria with vimentin intermediate filaments (VIFs) measurably increases their membrane potential. This increase is detected by quantitatively comparing the fluorescence intensity of mitochondria stained with the membrane potential-sensitive dye tetramethylrhodamine-ethyl ester (TMRE) in murine vimentin-null fibroblasts with that in the same cells expressing human vimentin (∼35% rise). When vimentin expression is silenced by small hairpin RNA (shRNA) to reduce vimentin by 90%, the fluorescence intensity of mitochondria decreases by 20%. The increase in membrane potential is caused by specific interactions between a subdomain of the non-α-helical N terminus (residues 40 to 93) of vimentin and mitochondria. In rho 0 cells lacking mitochondrial DNA (mtDNA) and consequently missing several key proteins in the mitochondrial respiratory chain (ρ0 cells), the membrane potential generated by an alternative anaerobic process is insensitive to the interactions between mitochondria and VIF. The results of our studies show that the close association between mitochondria and VIF is important both for determining their position in cells and their physiologic activity.—Chernoivanenko, I. S., Matveeva, E. A., Gelfand, V. I., Goldman, R. D., Minin, A. A. Mitochondrial membrane potential is regulated by vimentin intermediate filaments. PMID:25404709

  7. Initial characterization of Trypanosoma lewisi surface membrane antigen.

    PubMed

    Viyanant, V; Jones, A W

    1978-03-01

    The purified T. lewisi were subjected to hypotonic lysis plus freezing and thawing in acetone dry ice bath. The trypanosome ghosts were obtained after repeated washing and centrifugation. The homogenized ghost suspension was assayed for enzyme Na++K+ ATPase activity to ratify the presence of the trypanosome surface membrane. Membrane solubilized in sodium dodecyl sulfate were fractionated by gel filtration on Sephadex columns equilibrated with the detergent and electrophoresis in polyacrylamide gel. Crude trypanosome surface membrane antigens were tested for their immunogenicity, administered to rats in Fraund's complete adjuvant. The results of these experiments indicated that the protective immunogen is tightly bound to the membrane since the use of strong anionic detergent is necessary in its extraction. PMID:212837

  8. Interchangeable adaptors regulate mitochondrial dynamin assembly for membrane scission

    PubMed Central

    Koirala, Sajjan; Guo, Qian; Kalia, Raghav; Bui, Huyen T.; Eckert, Debra M.; Frost, Adam; Shaw, Janet M.

    2013-01-01

    Mitochondrial fission is mediated by the dynamin-related GTPases Dnm1/Drp1 (yeast/mammals), which form spirals around constricted sites on mitochondria. Additional membrane-associated adaptor proteins (Fis1, Mdv1, Mff, and MiDs) are required to recruit these GTPases from the cytoplasm to the mitochondrial surface. Whether these adaptors participate in both GTPase recruitment and membrane scission is not known. Here we use a yeast strain lacking all fission proteins to identify the minimal combinations of GTPases and adaptors sufficient for mitochondrial fission. Although Fis1 is dispensable for fission, membrane-anchored Mdv1, Mff, or MiDs paired individually with their respective GTPases are sufficient to divide mitochondria. In addition to their role in Drp1 membrane recruitment, MiDs coassemble with Drp1 in vitro. The resulting heteropolymer adopts a dramatically different structure with a narrower diameter than Drp1 homopolymers assembled in isolation. This result demonstrates that an adaptor protein alters the architecture of a mitochondrial dynamin GTPase polymer in a manner that could facilitate membrane constriction and severing activity. PMID:23530241

  9. Membrane Vibration Tests Using Surface-Bonded Piezoelectric Patch Actuation

    NASA Technical Reports Server (NTRS)

    Gasper, James L.; Pappa, Richard S.

    2003-01-01

    This paper describes the status of on-going work at the NASA Langley Research Center to measure the dynamics of thin membranes. The test article is a one-meter square pre-tensioned Kapton membrane that incorporates small surface-bonded piezoelectric patches strategically positioned to excite many modes. It is shown that PVDF (polyvinylidene fluoride) and MFC (macro fiber composite) piezoelectric patch actuators provide adequate excitation energy to obtain modal frequencies and mode shapes. Results from modal tests performed on the membrane using piezoelectric patches of different sizes and positions are discussed.

  10. RACK1 interacts with filamin-A to regulate plasma membrane levels of the cystic fibrosis transmembrane conductance regulator.

    PubMed

    Smith, Laura; Litman, Paul; Kohli, Ekta; Amick, Joseph; Page, Richard C; Misra, Saurav; Liedtke, Carole M

    2013-07-01

    Mutations in cystic fibrosis transmembrane regulator (CFTR), a chloride channel in the apical membranes of secretory epithelial cells, underlie the fatal genetic disorder cystic fibrosis. Certain CFTR mutations, including the common mutation ΔF508-CFTR, result in greatly decreased levels of active CFTR at the apical membrane. Direct interactions between CFTR and the cytoskeletal adaptors filamin-A (FlnA) and Na(+)/H(+) exchanger regulatory factor 1 (NHERF1) stabilize the expression and localization of CFTR at the plasma membrane. The scaffold protein receptor for activated C kinase 1 (RACK1) also stabilizes CFTR surface expression; however, RACK1 does not interact directly with CFTR and its mechanism of action is unknown. In the present study, we report that RACK1 interacts directly with FlnA in vitro and in a Calu-3 airway epithelial cell line. We mapped the interaction between RACK1 and FlnA to the WD4 and WD6 repeats of RACK1 and to a segment of the large rod domain of FlnA, consisting of immunoglobulin-like repeats 8-15. Disruption of the RACK1-FlnA interaction causes a reduction in CFTR surface levels. Our results suggest that a novel RACK1-FlnA interaction is an important regulator of CFTR surface localization. PMID:23636454

  11. Surface modification of nanoporous alumina membranes by plasma polymerization

    NASA Astrophysics Data System (ADS)

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

    2008-06-01

    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.

  12. Spectral studies of Lanthanide interactions with membrane surfaces

    SciTech Connect

    Karukstis, K.K.; Kao, M.Y.; Savin, D.A.; Bittker, R.A.; Kaphengst, K.J.; Emetarom, C.M.; Naito, N.R.; Takamoto, D.Y.

    1995-03-23

    We have monitored the interactions of the series of trivalent lanthanide cations with the thylakoid membrane surface of spinach chloroplasts using two complementary spectral techniques. Measurements of the fluorescence emission of the extrinsic probe 2-p-toluidinonaphthalene-6-sulfonate (TNS) and the absorbance of the intrinsic chromophore chlorophyll provide two sensitive means of characterizing the dependence of the cation-membrane interaction on the nature of the cation. In these systems, added lanthanide cations adsorb onto the membrane surface to neutralize exposed segments of membrane-embedded protein complexes. The lanthanide-induced charge neutralization increases the proximity of added TNS anion to the membrane surface as evidenced by variations in the TNS fluorescence level and wavelength of maximum emission. Our results reveal a strong dependence of TNS fluorescence parameters on both lanthanide size and total orbital angular momentum L value. Lanthanides with greater charge density (small size and/or low L value) enhance the TNS fluorescence level to a greater extent. A possible origin for the lanthanide-dependent TNS fluorescence levels is suggested in terms of a heterogeneity in the number and type of TNS binding sites. The data are consistent with the proposal that larger lanthanides with smaller enthalpies of hydration induce more significant membrane appression. 59 refs., 9 figs., 2 tabs.

  13. The structure of ions and zwitterionic lipids regulates the charge of dipolar membranes.

    PubMed

    Szekely, Or; Steiner, Ariel; Szekely, Pablo; Amit, Einav; Asor, Roi; Tamburu, Carmen; Raviv, Uri

    2011-06-21

    In pure water, zwitterionic lipids form lamellar phases with an equilibrium water gap on the order of 2 to 3 nm as a result of the dominating van der Waals attraction between dipolar bilayers. Monovalent ions can swell those neutral lamellae by a small amount. Divalent ions can adsorb onto dipolar membranes and charge them. Using solution X-ray scattering, we studied how the structure of ions and zwitterionic lipids regulates the charge of dipolar membranes. We found that unlike monovalent ions that weakly interact with all of the examined dipolar membranes, divalent and trivalent ions adsorb onto membranes containing lipids with saturated tails, with an association constant on the order of ∼10 M(-1). One double bond in the lipid tail is sufficient to prevent divalent ion adsorption. We suggest that this behavior is due to the relatively loose packing of lipids with unsaturated tails that increases the area per lipid headgroup, enabling their free rotation. Divalent ion adsorption links two lipids and limits their free rotation. The ion-dipole interaction gained by the adsorption of the ions onto unsaturated membranes is insufficient to compensate for the loss of headgroup free-rotational entropy. The ion-dipole interaction is stronger for cations with a higher valence. Nevertheless, polyamines behave as monovalent ions near dipolar interfaces in the sense that they interact weakly with the membrane surface, whereas in the bulk their behavior is similar to that of multivalent cations. Advanced data analysis and comparison with theory provide insight into the structure and interactions between ion-induced regulated charged interfaces. This study models biologically relevant interactions between cell membranes and various ions and the manner in which the lipid structure governs those interactions. The ability to monitor these interactions creates a tool for probing systems that are more complex and forms the basis for controlling the interactions between dipolar

  14. Membrane organization and regulation of cellular Cholesterol homeostasis

    PubMed Central

    Jaureguiberry, María S.; Tricerri, M. Alejandra; Sanchez, Susana A; Garda, Horacio A; Finarelli, Gabriela S.; Gonzalez, Marina C.; Rimoldi, Omar J.

    2010-01-01

    An excess of intracellular free Cholesterol (Chol) is cytotoxic, and its homeostasis is crucial for cell viability. Apolipoprotein A–I (apoA-I) is a highly efficient Chol acceptor as it activates complex cellular pathways that tend to mobilize and export Chol from cellular depots. Here we hypothesize that membrane composition and/or organization is strongly involved in Chol homeostasis. To test this hypothesis, we constructed a cell line over expressing Stearoyl CoA desaturase (SCD-cells), which modifies plasma membrane (PM) composition by the enrichment of monounsaturated fatty,acids and determined this effect on membrane properties, cell viability and cholesterol homeostasis. PM in SCD-cells has a higher phospholipids/sphingomyelin ratio and is slightly enriched in Chol. These cells showed an increase in the cholesteryl esters/free Chol ratio, they were more resistant to Chol toxicity and in addition, they exported more caveolin than Control cells. The data suggest that cell functionality is preserved by regulating membrane fluidity and Chol exportation and storage. PMID:20336284

  15. Transcription Regulation and Membrane Stress Management in Enterobacterial Pathogens.

    PubMed

    Zhang, Nan; Jovanovic, Goran; McDonald, Christopher; Ces, Oscar; Zhang, Xiaodong; Buck, Martin

    2016-01-01

    Transcription regulation in a temporal and conditional manner underpins the lifecycle of enterobacterial pathogens. Upon exposure to a wide array of environmental cues, these pathogens modulate their gene expression via the RNA polymerase and associated sigma factors. Different sigma factors, either involved in general 'house-keeping' or specific responses, guide the RNA polymerase to their cognate promoter DNAs. The major alternative sigma54 factor when activated helps pathogens manage stresses and proliferate in their ecological niches. In this chapter, we review the function and regulation of the sigma54-dependent Phage shock protein (Psp) system-a major stress response when Gram-negative pathogens encounter damages to their inner membranes. We discuss the recent development on mechanisms of gene regulation, signal transduction and stress mitigation in light of different biophysical and biochemical approaches. PMID:27193545

  16. Surface functionalization of thin-film composite membranes with copper nanoparticles for antimicrobial surface properties.

    PubMed

    Ben-Sasson, Moshe; Zodrow, Katherine R; Genggeng, Qi; Kang, Yan; Giannelis, Emmanuel P; Elimelech, Menachem

    2014-01-01

    Biofouling is a major operational challenge in reverse osmosis (RO) desalination, motivating a search for improved biofouling control strategies. Copper, long known for its antibacterial activity and relatively low cost, is an attractive potential biocidal agent. In this paper, we present a method for loading copper nanoparticles (Cu-NPs) on the surface of a thin-film composite (TFC) polyamide RO membrane. Cu-NPs were synthesized using polyethyleneimine (PEI) as a capping agent, resulting in particles with an average radius of 34 nm and a copper content between 39 and 49 wt.%. The positive charge of the Cu-NPs imparted by the PEI allowed a simple electrostatic functionalization of the negatively charged RO membrane. We confirmed functionalization and irreversible binding of the Cu-NPs to the membrane surface with SEM and XPS after exposing the membrane to bath sonication. We also demonstrated that Cu-NP functionalization can be repeated after the Cu-NPs dissolve from the membrane surface. The Cu-NP functionalization had minimal impact on the intrinsic membrane transport parameters. Surface hydrophilicity and surface roughness were also maintained, and the membrane surface charge became positive after functionalization. The functionalized membrane exhibited significant antibacterial activity, leading to an 80-95% reduction in the number of attached live bacteria for three different model bacterial strains. Challenges associated with this functionalization method and its implementation in RO desalination are discussed. PMID:24308843

  17. Quantitative Analysis of Membrane Trafficking in Regulation of Cdc42 Polarity

    PubMed Central

    Watson, Leah J.; Rossi, Guendalina; Brennwald, Patrick

    2014-01-01

    Vesicle delivery of Cdc42 has been proposed as an important mechanism for generating and maintaining Cdc42 polarity at the plasma membrane. This mechanism requires the density of Cdc42 on secretory vesicles to be equal to or higher than the plasma membrane polarity cap. Using a novel method to estimate Cdc42 levels on post-Golgi secretory vesicles in intact yeast cells, we: 1) determined that endocytosis plays an important role in Cdc42’s association with secretory vesicles 2) found that a GFP-tag placed on the N-terminus of Cdc42 negatively impacts this vesicle association and 3) quantified the surface densities of Cdc42 on post-Golgi vesicles which revealed that the vesicle density of Cdc42 is three times more dilute than that at the polarity cap. This work suggests that the immediate consequence of secretory vesicle fusion with the plasma membrane polarity cap is to dilute the local Cdc42 surface density. This provides strong support for the model in which vesicle trafficking acts to negatively regulate Cdc42 polarity on the cell surface while also providing a means to recycle Cdc42 between the cell surface and internal membrane locations. PMID:25158298

  18. Detachment of surface membrane invagination systems by cationic amphiphilic drugs

    PubMed Central

    Osman, Sangar; Taylor, Kirk A.; Allcock, Natalie; Rainbow, Richard D.; Mahaut-Smith, Martyn P.

    2016-01-01

    Several cell types develop extensive plasma membrane invaginations to serve a specific physiological function. For example, the megakaryocyte demarcation membrane system (DMS) provides a membrane reserve for platelet production and muscle transverse (T) tubules facilitate excitation:contraction coupling. Using impermeant fluorescent indicators, capacitance measurements and electron microscopy, we show that multiple cationic amphiphilic drugs (CADs) cause complete separation of the DMS from the surface membrane in rat megakaryocytes. This includes the calmodulin inhibitor W-7, the phospholipase-C inhibitor U73122, and anti-psychotic phenothiazines. CADs also caused loss of T tubules in rat cardiac ventricular myocytes and the open canalicular system of human platelets. Anionic amphiphiles, U73343 (a less electrophilic U73122 analogue) and a range of kinase inhibitors were without effect on the DMS. CADs are known to accumulate in the inner leaflet of the cell membrane where they bind to anionic lipids, especially PI(4,5)P2. We therefore propose that surface detachment of membrane invaginations results from an ability of CADs to interfere with PI(4,5)P2 interactions with cytoskeletal or BAR domain proteins. This establishes a detubulating action of a large class of pharmaceutical compounds. PMID:26725955

  19. Rab43 regulates the sorting of a subset of membrane protein cargo through the medial Golgi.

    PubMed

    Cox, John V; Kansal, Rita; Whitt, Michael A

    2016-06-01

    To evaluate the role of cytoplasmic domains of membrane-spanning proteins in directing trafficking through the secretory pathway, we generated fluorescently tagged VSV G tsO45 with either the native G tail (G) or a cytoplasmic tail derived from the chicken AE1-4 anion exchanger (G(AE)). We previously showed that these two proteins progressed through the Golgi with distinct kinetics. To investigate the basis for the differential sorting of G and G(AE), we analyzed the role of several Golgi-associated small GTP-binding proteins and found that Rab43 differentially regulated their transport through the Golgi. We show that the expression of GFP-Rab43 arrested the anterograde transport of G(AE) in a Rab43-positive medial Golgi compartment. GFP-Rab43 expression also inhibited the acquisition of endoH-resistant sugars and the surface delivery of G(AE), as well as the surface delivery of the AE1-4 anion exchanger. In contrast, GFP-Rab43 expression did not affect the glycosylation or surface delivery of G. Unexpectedly, down-regulation of endogenous Rab43 using small interfering RNA resulted in an increase in the accumulation of G(AE) on the cell surface while having minimal effect on the surface levels of G. Our data demonstrate that Rab43 regulates the sorting of a subset of membrane-spanning cargo as they progress through the medial Golgi. PMID:27053659

  20. Regulation of the basement membrane by epithelia generated forces

    PubMed Central

    Tanner, Kandice

    2012-01-01

    Tumor metastasis involves a progressive loss of tissue architecture and dissolution of structural boundaries between the epithelium and connective tissue. The basement membrane (BM), a specialized network of extracellular matrix proteins forms a barrier that physically restricts pre- invasive lesions such that they remain as local insults. The BM is not a static structure, but one that is constantly regenerated and remodeled in the adult organism. Matrix organization also regulates cell function. Thus alterations in the balance of synthesis, remodeling and proteolytic degradation of the extracellular matrix proteins may contribute to a loss of structural integrity. However, the de novo assembly and maintenance of the complex structural properties of in vivo basement membranes remain elusive. Here, this paper highlights the current understanding on the structural properties and the establishment of the BM, and discusses the potential role of self-generated forces in adult tissue remodeling and the maintenance of the BM as a malignancy suppressor. PMID:23196920

  1. Homeostasis of the apical plasma membrane during regulated exocytosis in the salivary glands of live rodents

    PubMed Central

    Masedunskas, Andrius; Sramkova, Monika; Weigert, Roberto

    2011-01-01

    In exocrine organs such as the salivary glands, fluids and proteins are secreted into ductal structures by distinct mechanisms that are tightly coupled. In the acinar cells, the major secretory units of the salivary glands, fluids are secreted into the acinar canaliculi through paracellular and intracellular transport, whereas proteins are stored in large granules that undergo exocytosis and fuse with the apical plasma membranes releasing their content into the canaliculi. Both secretory processes elicit a remodeling of the apical plasma membrane that has not been fully addressed in in vitro or ex vivo models. Recently, we have studied regulated exocytosis in the salivary glands of live rodents, focusing on the role that actin and myosin plays in this process. We observed that during exocytosis both secretory granules and canaliculi are subjected to the hydrostatic pressure generated by fluid secretion. Furthermore, the absorption of the membranes of the secretory granules contributes to the expansion and deformation of the canaliculi. Here we suggest that the homeostasis of the apical plasma membranes during exocytosis is maintained by various strategies that include: (1) membrane retrieval via compensatory endocytosis, (2) increase of the surface area via membrane folds and (3) recruitment of a functional actomyosin complex. Our observations underscore the important relationship between tissue architecture and cellular response, and highlight the potential of investigating biological processes in vivo by using intravital microscopy. PMID:22754613

  2. Surface-Localized Spermidine Protects the Pseudomonas aeruginosa Outer Membrane from Antibiotic Treatment and Oxidative Stress

    PubMed Central

    Johnson, Lori; Mulcahy, Heidi; Kanevets, Uliana; Shi, Yan

    2012-01-01

    Extracellular DNA acts as a cation chelator and induces the expression of antibiotic resistance genes regulated by Mg2+ levels. Here we report the characterization of novel DNA-induced genes in Pseudomonas aeruginosa that are annotated as homologs of the spermidine synthesis genes speD (PA4773) and speE (PA4774). The addition of sublethal concentrations of DNA and membrane-damaging antibiotics induced expression of the genes PA4773 to PA4775, as shown using transcriptional lux fusions and quantitative RT-PCR. Exogenous polyamine addition prevented DNA- and peptide-mediated gene induction. Mutation of PA4774 resulted in an increased outer membrane (OM) susceptibility phenotype upon polymyxin B, CP10A, and gentamicin treatment. When the membrane-localized fluorescent probe C11-BODIPY581/591 was used as an indicator of peroxidation of membrane lipids, the PA4774::lux mutant demonstrated an increased susceptibility to oxidative membrane damage from H2O2 treatment. Addition of exogenous polyamines protected the membranes of the PA4774::lux mutant from polymyxin B and H2O2 treatment. Polyamines from the outer surface were isolated and shown to contain putrescine and spermidine by using high-performance liquid chromatography and mass spectrometry. The PA4774::lux mutant did not produce spermidine on the cell surface, but genetic complementation restored surface spermidine production as well as the antibiotic and oxidative stress resistance phenotypes of the membrane. We have identified new functions for spermidine on the cell surface and propose that polyamines are produced under Mg2+-limiting conditions as an organic polycation to bind lipopolysaccharide (LPS) and to stabilize and protect the outer membrane against antibiotic and oxidative damage. PMID:22155771

  3. Plasma Membrane Repair Is Regulated Extracellularly by Proteases Released from Lysosomes.

    PubMed

    Castro-Gomes, Thiago; Corrotte, Matthias; Tam, Christina; Andrews, Norma W

    2016-01-01

    Eukaryotic cells rapidly repair wounds on their plasma membrane. Resealing is Ca(2+)-dependent, and involves exocytosis of lysosomes followed by massive endocytosis. Extracellular activity of the lysosomal enzyme acid sphingomyelinase was previously shown to promote endocytosis and wound removal. However, whether lysosomal proteases released during cell injury participate in resealing is unknown. Here we show that lysosomal proteases regulate plasma membrane repair. Extracellular proteolysis is detected shortly after cell wounding, and inhibition of this process blocks repair. Conversely, surface protein degradation facilitates plasma membrane resealing. The abundant lysosomal cysteine proteases cathepsin B and L, known to proteolytically remodel the extracellular matrix, are rapidly released upon cell injury and are required for efficient plasma membrane repair. In contrast, inhibition of aspartyl proteases or RNAi-mediated silencing of the lysosomal aspartyl protease cathepsin D enhances resealing, an effect associated with the accumulation of active acid sphingomyelinase on the cell surface. Thus, secreted lysosomal cysteine proteases may promote repair by facilitating membrane access of lysosomal acid sphingomyelinase, which promotes wound removal and is subsequently downregulated extracellularly by a process involving cathepsin D. PMID:27028538

  4. Plasma Membrane Repair Is Regulated Extracellularly by Proteases Released from Lysosomes

    PubMed Central

    Castro-Gomes, Thiago; Corrotte, Matthias; Tam, Christina; Andrews, Norma W.

    2016-01-01

    Eukaryotic cells rapidly repair wounds on their plasma membrane. Resealing is Ca2+-dependent, and involves exocytosis of lysosomes followed by massive endocytosis. Extracellular activity of the lysosomal enzyme acid sphingomyelinase was previously shown to promote endocytosis and wound removal. However, whether lysosomal proteases released during cell injury participate in resealing is unknown. Here we show that lysosomal proteases regulate plasma membrane repair. Extracellular proteolysis is detected shortly after cell wounding, and inhibition of this process blocks repair. Conversely, surface protein degradation facilitates plasma membrane resealing. The abundant lysosomal cysteine proteases cathepsin B and L, known to proteolytically remodel the extracellular matrix, are rapidly released upon cell injury and are required for efficient plasma membrane repair. In contrast, inhibition of aspartyl proteases or RNAi-mediated silencing of the lysosomal aspartyl protease cathepsin D enhances resealing, an effect associated with the accumulation of active acid sphingomyelinase on the cell surface. Thus, secreted lysosomal cysteine proteases may promote repair by facilitating membrane access of lysosomal acid sphingomyelinase, which promotes wound removal and is subsequently downregulated extracellularly by a process involving cathepsin D. PMID:27028538

  5. Structural basis for the transcriptional regulation of membrane lipid homeostasis

    SciTech Connect

    Miller, Darcie J.; Zhang, Yong-Mei; Subramanian, Chitra; Rock, Charles O.; White, Stephen W.

    2010-11-09

    DesT is a transcriptional repressor that regulates the genes that control the unsaturated:saturated fatty acid ratio available for membrane lipid synthesis. DesT bound to unsaturated acyl-CoA has a high affinity for its cognate palindromic DNA-binding site, whereas DesT bound to saturated acyl-CoA does not bind this site. Structural analyses of the DesT-oleoyl-CoA-DNA and DesT-palmitoyl-CoA complexes reveal that acyl chain shape directly influences the packing of hydrophobic core residues within the DesT ligand-binding domain. These changes are propagated to the paired DNA-binding domains via conformational changes to modulate DNA binding. These structural interpretations are supported by the in vitro and in vivo characterization of site-directed mutants. The regulation of DesT by the unsaturated:saturated ratio of acyl chains rather than the concentration of a single ligand is a paradigm for understanding transcriptional regulation of membrane lipid homeostasis.

  6. Regulation of the divalent metal ion transporter via membrane budding

    PubMed Central

    Mackenzie, KimberlyD; Foot, Natalie J; Anand, Sushma; Dalton, Hazel E; Chaudhary, Natasha; Collins, Brett M; Mathivanan, Suresh; Kumar, Sharad

    2016-01-01

    The release of extracellular vesicles (EVs) is important for both normal physiology and disease. However, a basic understanding of the targeting of EV cargoes, composition and mechanism of release is lacking. Here we present evidence that the divalent metal ion transporter (DMT1) is unexpectedly regulated through release in EVs. This process involves the Nedd4-2 ubiquitin ligase, and the adaptor proteins Arrdc1 and Arrdc4 via different budding mechanisms. We show that mouse gut explants release endogenous DMT1 in EVs. Although we observed no change in the relative amount of DMT1 released in EVs from gut explants in Arrdc1 or Arrdc4 deficient mice, the extent of EVs released was significantly reduced indicating an adaptor role in biogenesis. Furthermore, using Arrdc1 or Arrdc4 knockout mouse embryonic fibroblasts, we show that both Arrdc1 and Arrdc4 are non-redundant positive regulators of EV release. Our results suggest that DMT1 release from the plasma membrane into EVs may represent a novel mechanism for the maintenance of iron homeostasis, which may also be important for the regulation of other membrane proteins. PMID:27462458

  7. Desalination by Membrane Distillation using Electrospun Polyamide Fiber Membranes with Surface Fluorination by Chemical Vapor Deposition.

    PubMed

    Guo, Fei; Servi, Amelia; Liu, Andong; Gleason, Karen K; Rutledge, Gregory C

    2015-04-22

    Fibrous membranes of poly(trimethyl hexamethylene terephthalamide) (PA6(3)T) were fabricated by electrospinning and rendered hydrophobic by applying a conformal coating of poly(1H,1H,2H,2H-perfluorodecyl acrylate) (PPFDA) using initiated chemical vapor deposition (iCVD). A set of iCVD-treated electrospun PA6(3)T fiber membranes with fiber diameters ranging from 0.25 to 1.8 μm were tested for desalination using the air gap membrane distillation configuration. Permeate fluxes of 2-11 kg/m2/h were observed for temperature differentials of 20-45 °C between the feed stream and condenser plate, with rejections in excess of 99.98%. The liquid entry pressure was observed to increase dramatically, from 15 to 373 kPa with reduction in fiber diameter. Contrary to expectation, for a given feed temperature the permeate flux was observed to increase for membranes of decreasing fiber diameter. The results for permeate flux and salt rejection show that it is possible to construct membranes for membrane distillation even from intrinsically hydrophilic materials after surface modification by iCVD and that the fiber diameter is shown to play an important role on the membrane distillation performance in terms of permeate flux, salt rejection, and liquid entry pressure. PMID:25835769

  8. Surface precision of optical membranes with curvature.

    PubMed

    Marker, D; Jenkins, C

    1997-11-24

    Space-based inflatable technology is of current interest to NASA and DOD, and in particular to the Air Force and Phillips Laboratory. Potentially large gains in lowering launch costs, through reductions in structure mass and volume, are driving this activity. Diverse groups are researching and developing this technology for radio and radar antennae, optical telescopes, and solar power and propulsion applications. Regardless of the use, one common requirement for successful application is the accuracy of the inflated surface shape. The work reported here concerns the shape control of an inflated thin circular disk through use of a nonlinear finite element analysis. First, a review of the important associated Hencky problem is given. Then we discuss a shape modification, achieved through enforced boundary displacements, which resulted in moving the inflated shape towards a desired parabolic profile. Minimization of the figure error is discussed and conclusions are drawn. PMID:19377552

  9. Late endosomal membranes rich in lysobisphosphatidic acid regulate cholesterol transport.

    PubMed

    Kobayashi, T; Beuchat, M H; Lindsay, M; Frias, S; Palmiter, R D; Sakuraba, H; Parton, R G; Gruenberg, J

    1999-06-01

    The fate of free cholesterol released after endocytosis of low-density lipoproteins remains obscure. Here we report that late endosomes have a pivotal role in intracellular cholesterol transport. We find that in the genetic disease Niemann-Pick type C (NPC), and in drug-treated cells that mimic NPC, cholesterol accumulates in late endosomes and sorting of the lysosomal enzyme receptor is impaired. Our results show that the characteristic network of lysobisphosphatidic acid-rich membranes contained within multivesicular late endosomes regulates cholesterol transport, presumably by acting as a collection and distribution device. The results also suggest that similar endosomal defects accompany the anti-phospholipid syndrome and NPC. PMID:10559883

  10. Regulation of membrane-type 1 matrix metalloproteinase activity by vacuolar H+-ATPases.

    PubMed Central

    Maquoi, Erik; Peyrollier, Karine; Noël, Agnès; Foidart, Jean-Michel; Frankenne, Francis

    2003-01-01

    Membrane-type 1 matrix metalloproteinase (MT1-MMP) is a key enzyme in normal development and malignant processes. The regulation of MT1-MMP activity on the cell surface is a complex process involving autocatalytic processing, tissue inhibitor of MMPs (TIMP) binding and constitutive internalization. However, the fate of internalized MT1-MMP is not known. Acidification of intracellular vacuolar compartments is essential for membrane trafficking, protein sorting and degradation. This acidification is controlled by vacuolar H(+)-ATPases, which can be selectively inhibited by bafilomycin-A(1). Here, we treated human tumour cell lines expressing MT1-MMP with bafilomycin-A(1), and analysed its effects on MT1-MMP activity, internalization and processing. We show that the activity of MT1-MMP on the cell surface is constitutively down-regulated through a vacuolar H(+)-ATPase-dependent degradation process. Blockade of this degradation caused the accumulation of TIMP-free active MT1-MMP molecules on the cell surface, although internalization was not affected. As a consequence of this impaired degradation, pro-MMP-2 activation was strongly enhanced. This study demonstrates that the catalytic activity of MT1-MMP on the cell surface is regulated through a vacuolar H(+)-ATPase-dependent degradation process. PMID:12667140

  11. Golgi sorting regulates organization and activity of GPI-proteins at apical membranes

    PubMed Central

    Tivodar, Simona; Formiggini, Fabio; Ossato, Giulia; Gratton, Enrico; Tramier, Marc; Coppey-Moisan, Maïté; Zurzolo, Chiara

    2014-01-01

    Here, we combined classical biochemistry with novel biophysical approaches to study with high spatial and temporal resolution the organization of GPI-anchored proteins (GPI-APs) at the plasma membrane of polarized epithelial cells. We show that in polarized MDCK cells, following sorting in the Golgi, each GPI-AP reaches the apical surface in homo-clusters. Golgi-derived homo-clusters are required for their subsequent plasma membrane organization into cholesterol-dependent hetero-clusters. By contrast, in non-polarized MDCK cells GPI-APs are delivered to the surface as monomers in an unpolarized manner and are not able to form hetero-clusters. We further demonstrate that this GPI-AP organization is regulated by the content of cholesterol in the Golgi apparatus and is required to maintain the functional state of the protein at the apical membrane. Thus, different from fibroblasts, in polarized epithelial cells a selective cholesterol-dependent sorting mechanism in the Golgi regulates both the organization and the function of GPI-APs at the apical surface. PMID:24681536

  12. Composite fluorocarbon membranes by surface-initiated polymerization from nanoporous gold-coated alumina.

    PubMed

    Escobar, Carlos A; Zulkifli, Ahmad R; Faulkner, Christopher J; Trzeciak, Alex; Jennings, G Kane

    2012-02-01

    This manuscript describes the versatile fabrication and characterization of a novel composite membrane that consists of a porous alumina support, a 100 nm thick nanoporous gold coating, and a selective poly(5-(perfluorohexyl)norbornene) (pNBF6) polymer that can be grown exclusively from the nanoporous gold or throughout the membrane. Integration of the three materials is achieved by means of silane and thiol chemistry, and the use of surface-initiated ring-opening metathesis polymerization (SI-ROMP) to grow the pNBF6. The use of SI-ROMP allows tailoring of the extent of polymerization of pNBF6 throughout the structure by varying polymerization time. Scanning electron microscopy (SEM) images indicate that the thin polymer films cover the structure entirely. Cross-sectional SEM images of the membrane not only corroborate growth of the pNBF6 polymer within both the porous alumina and the nanoporous gold coating but also show the growth of a pNBF6 layer between these porous substrates that lifts the nanoporous gold coating away from the alumina. Advancing contact angle (θ(A)) measurements show that the surfaces of these composite membranes exhibit both hydrophobic (θ(A) = 121-129)° and oleophobic (θ(A) = 69-74)° behavior due to the fluorocarbon side chains of the pNBF6 polymer that dominate the surface. Results from electrochemical impedance spectroscopy (EIS) confirm that the membranes provide effective barriers to aqueous ions, as evidenced by a resistive impedance on the order of 1 × 10(7) Ω cm(2). Sulfonation of the polymer backbone substantially enhances ion transport through the composite membrane, as indicated by a 40-60 fold reduction in resistive impedance. Ion transport and selectivity of the membrane change by regulating the polymerization time. The fluorinated nature of the sulfonated polymer renders the membrane selective toward molecules with similar chemical characteristics. PMID:22195729

  13. Progesterone receptor membrane component-1 regulates hepcidin biosynthesis

    PubMed Central

    Li, Xiang; Rhee, David K.; Malhotra, Rajeev; Mayeur, Claire; Hurst, Liam A.; Ager, Emily; Shelton, Georgia; Kramer, Yael; McCulloh, David; Keefe, David; Bloch, Kenneth D.; Bloch, Donald B.; Peterson, Randall T.

    2015-01-01

    Iron homeostasis is tightly regulated by the membrane iron exporter ferroportin and its regulatory peptide hormone hepcidin. The hepcidin/ferroportin axis is considered a promising therapeutic target for the treatment of diseases of iron overload or deficiency. Here, we conducted a chemical screen in zebrafish to identify small molecules that decrease ferroportin protein levels. The chemical screen led to the identification of 3 steroid molecules, epitiostanol, progesterone, and mifepristone, which decrease ferroportin levels by increasing the biosynthesis of hepcidin. These hepcidin-inducing steroids (HISs) did not activate known hepcidin-inducing pathways, including the BMP and JAK/STAT3 pathways. Progesterone receptor membrane component-1 (PGRMC1) was required for HIS-dependent increases in hepcidin biosynthesis, as PGRMC1 depletion in cultured hepatoma cells and zebrafish blocked the ability of HISs to increase hepcidin mRNA levels. Neutralizing antibodies directed against PGRMC1 attenuated the ability of HISs to induce hepcidin gene expression. Inhibiting the kinases of the SRC family, which are downstream of PGRMC1, blocked the ability of HISs to increase hepcidin mRNA levels. Furthermore, HIS treatment increased hepcidin biosynthesis in mice and humans. Together, these data indicate that PGRMC1 regulates hepcidin gene expression through an evolutionarily conserved mechanism. These studies have identified drug candidates and potential therapeutic targets for the treatment of diseases of abnormal iron metabolism. PMID:26657863

  14. Progesterone receptor membrane component-1 regulates hepcidin biosynthesis.

    PubMed

    Li, Xiang; Rhee, David K; Malhotra, Rajeev; Mayeur, Claire; Hurst, Liam A; Ager, Emily; Shelton, Georgia; Kramer, Yael; McCulloh, David; Keefe, David; Bloch, Kenneth D; Bloch, Donald B; Peterson, Randall T

    2016-01-01

    Iron homeostasis is tightly regulated by the membrane iron exporter ferroportin and its regulatory peptide hormone hepcidin. The hepcidin/ferroportin axis is considered a promising therapeutic target for the treatment of diseases of iron overload or deficiency. Here, we conducted a chemical screen in zebrafish to identify small molecules that decrease ferroportin protein levels. The chemical screen led to the identification of 3 steroid molecules, epitiostanol, progesterone, and mifepristone, which decrease ferroportin levels by increasing the biosynthesis of hepcidin. These hepcidin-inducing steroids (HISs) did not activate known hepcidin-inducing pathways, including the BMP and JAK/STAT3 pathways. Progesterone receptor membrane component-1 (PGRMC1) was required for HIS-dependent increases in hepcidin biosynthesis, as PGRMC1 depletion in cultured hepatoma cells and zebrafish blocked the ability of HISs to increase hepcidin mRNA levels. Neutralizing antibodies directed against PGRMC1 attenuated the ability of HISs to induce hepcidin gene expression. Inhibiting the kinases of the SRC family, which are downstream of PGRMC1, blocked the ability of HISs to increase hepcidin mRNA levels. Furthermore, HIS treatment increased hepcidin biosynthesis in mice and humans. Together, these data indicate that PGRMC1 regulates hepcidin gene expression through an evolutionarily conserved mechanism. These studies have identified drug candidates and potential therapeutic targets for the treatment of diseases of abnormal iron metabolism. PMID:26657863

  15. Interaction of Serum Proteins with Surface of Hemodialysis Fiber Membranes

    NASA Astrophysics Data System (ADS)

    Afrin, Rehana; Shirako, Yuji; Kishimoto, Kikuo; Ikai, Atsushi

    2012-08-01

    The poly(vinyl pyrrolidone)-covered hydrophilic surface of hollow-fiber membranes (fiber membrane, hereafter) for hemodialysis was mechanically probed using modified tips on an atomic force microscope (AFM) with covalent crosslinkers and several types of serum protein. The retraction part of many of the force extension (F-E) curves obtained with AFM tips coated with serum albumin had a long and smooth extension up to 200-300 nm indicating forced elongation of poly(vinyl pyrrolidone) chains. When fibrinogen-coated tips were used, long extension F-E curves up to 500 nm with multiple peaks were obtained in addition to smooth curves most likely reflecting the unfolding of fibrinogen molecules. The results indicated that individual polymer chains had a significant affinity toward serum proteins. The adhesion frequency of tips coated with serum proteins was lower on the poly(vinyl pyrrolidone) surface than on the uncoated hydrophobic polysulfone surface.

  16. Engineering a Highly Hydrophilic PVDF Membrane via Binding TiO₂Nanoparticles and a PVA Layer onto a Membrane Surface.

    PubMed

    Qin, Aiwen; Li, Xiang; Zhao, Xinzhen; Liu, Dapeng; He, Chunju

    2015-04-29

    A highly hydrophilic PVDF membrane was fabricated through chemically binding TiO2 nanoparticles and a poly(vinyl alcohol) (PVA) layer onto a membrane surface simultaneously. The chemical composition of the modified membrane surface was determined by X-ray photoelectron spectroscopy, and the binding performance of TiO2 nanoparticles and the PVA layer was investigated by a rinsing test. The results indicated that the TiO2 nanoparticles were uniformly and strongly tailored onto the membrane surface, while the PVA layer was firmly attached onto the surface of TiO2 nanoparticles and the membrane by adsorption-cross-linking. The possible mechanisms during the modification process and filtration performance, i.e., water permeability and bovine serum albumin (BSA) rejection, were investigated as well. Furthermore, antifouling property was discussed through multicycles of BSA solution filtration tests, where the flux recovery ratio was significantly increased from 20.0% for pristine PVDF membrane to 80.5% for PVDF/TiO2/PVA-modified membrane. This remarkable promotion is mainly ascribed to the improvement of surface hydrophilicity, where the water contact angle of the membrane surface was decreased from 84° for pristine membrane to 24° for PVDF/TiO2/PVA membrane. This study presents a novel and varied strategy for immobilization of nanoparticles and PVA layer on substrate surface, which could be easily adapted for a variety of materials for surface modification. PMID:25806418

  17. Lipid-linked cell wall precursors regulate membrane association of bacterial actin MreB

    PubMed Central

    Schirner, Kathrin; Eun, Ye-Jin; Dion, Mike; Luo, Yun; Helmann, John D.; Garner, Ethan C.; Walker, Suzanne

    2014-01-01

    Summary The bacterial actin homolog MreB, which is critical for rod shape determination, forms filaments that rotate around the cell width on the inner surface of the cytoplasmic membrane. What determines filament association with the membranes or with other cell wall elongation proteins is not known. Using specific chemical and genetic perturbations while following MreB filament motion, we find that MreB membrane association is an actively regulated process that depends on the presence of lipid-linked peptidoglycan precursors. When precursors are depleted, MreB filaments disassemble into the cytoplasm and peptidoglycan synthesis becomes disorganized. In cells that lack wall teichoic acids, but continue to make peptidoglycan, dynamic MreB filaments are observed, although their presence is not sufficient to establish a rod shape. We propose that the cell regulates MreB filament association with the membrane, allowing rapid and reversible inactivation of cell wall enzyme complexes in response to the inhibition of cell wall synthesis. PMID:25402772

  18. Effects of surface charge on interfacial interactions related to membrane fouling in a submerged membrane bioreactor based on thermodynamic analysis.

    PubMed

    Cai, Huihui; Fan, Hao; Zhao, Leihong; Hong, Huachang; Shen, Liguo; He, Yiming; Lin, Hongjun; Chen, Jianrong

    2016-03-01

    Effects of both membrane and sludge foulant surface zeta potentials on interfacial interactions between membrane and sludge foulant in different interaction scenarios were systematically investigated based on thermodynamic methods. Under conditions in this study, it was found that zeta potential had marginal effects on total interfacial interaction between two infinite planar surfaces, and the total interfacial interaction between foulant particles and membrane would be more repulsive with increase of absolute value of zeta potential. Adhesion of foulant particles on membrane surface should overcome an energy barrier. There exists a critical zeta potential below which energy barrier would disappear. Results also showed that rough surface membrane corresponded to significantly low strength of interfacial interactions. This study not only provided a series of methods to quantitatively assess the interfacial interactions between membrane and sludge foulants, but also reconciled the contradictory conclusions regarding effects of zeta potential in literature, giving important implications for membrane fouling mitigation. PMID:26641562

  19. Dynamic potential and surface morphology study of sertraline membrane sensors.

    PubMed

    Khater, M M; Issa, Y M; Hassib, H B; Mohammed, S H

    2015-05-01

    New rapid, sensitive and simple electrometric method was developed to determine sertraline hydrochloride (Ser-Cl) in its pure raw material and pharmaceutical formulations. Membrane sensors based on heteropolyacids as ion associating material were prepared. Silicomolybdic acid (SMA), silicotungstic acid (STA) and phosphomolybdic acid (PMA) were used. The slope and limit of detection are 50.00, 60.00 and 53.24 mV/decade and 2.51, 5.62 and 4.85 μmol L(-1) for Ser-ST, Ser-PM and Ser-SM membrane sensors, respectively. Linear range is 0.01-10.00 for the three sensors. These new sensors were used for the potentiometric titration of Ser-Cl using sodium tetraphenylborate as titrant. The surface morphologies of the prepared membranes with and without the modifier (ion-associate) were studied using scanning and atomic force microscopes. PMID:26257944

  20. Dynamic potential and surface morphology study of sertraline membrane sensors

    PubMed Central

    Khater, M.M.; Issa, Y.M.; Hassib, H.B.; Mohammed, S.H.

    2014-01-01

    New rapid, sensitive and simple electrometric method was developed to determine sertraline hydrochloride (Ser-Cl) in its pure raw material and pharmaceutical formulations. Membrane sensors based on heteropolyacids as ion associating material were prepared. Silicomolybdic acid (SMA), silicotungstic acid (STA) and phosphomolybdic acid (PMA) were used. The slope and limit of detection are 50.00, 60.00 and 53.24 mV/decade and 2.51, 5.62 and 4.85 μmol L−1 for Ser-ST, Ser-PM and Ser-SM membrane sensors, respectively. Linear range is 0.01–10.00 for the three sensors. These new sensors were used for the potentiometric titration of Ser-Cl using sodium tetraphenylborate as titrant. The surface morphologies of the prepared membranes with and without the modifier (ion-associate) were studied using scanning and atomic force microscopes. PMID:26257944

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

    NASA Astrophysics Data System (ADS)

    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.

  2. Membrane mimetic surface functionalization of nanoparticles: Methods and applications

    PubMed Central

    Weingart, Jacob; Vabbilisetty, Pratima; Sun, Xue-Long

    2013-01-01

    Nanoparticles (NPs), due to their size-dependent physical and chemical properties, have shown remarkable potential for a wide range of applications over the past decades. Particularly, the biological compatibilities and functions of NPs have been extensively studied for expanding their potential in areas of biomedical application such as bioimaging, biosensing, and drug delivery. In doing so, surface functionalization of NPs by introducing synthetic ligands and/or natural biomolecules has become a critical component in regards to the overall performance of the NP system for its intended use. Among known examples of surface functionalization, the construction of an artificial cell membrane structure, based on phospholipids, has proven effective in enhancing biocompatibility and has become a viable alternative to more traditional modifications, such as direct polymer conjugation. Furthermore, certain bioactive molecules can be immobilized onto the surface of phospholipid platforms to generate displays more reminiscent of cellular surface components. Thus, NPs with membrane-mimetic displays have found use in a range of bioimaging, biosensing, and drug delivery applications. This review herein describes recent advances in the preparations and characterization of integrated functional NPs covered by artificial cell membrane structures and their use in various biomedical applications. PMID:23688632

  3. Tuning the membrane surface potential for efficient toxin import

    PubMed Central

    Zakharov, Stanislav D.; Rokitskaya, Tatyana I.; Shapovalov, Vladimir L.; Antonenko, Yuri N.; Cramer, William A.

    2002-01-01

    Membrane surface electrostatic interactions impose structural constraints on imported proteins. An unprecedented sensitive dependence on these constraints was seen in the voltage-gated import and channel formation by the C-terminal pore-forming domain of the bacteriocin, colicin E1. At physiological ionic strengths, significant channel current was observed only in a narrow interval of anionic lipid content ([L−]), with the maximum current (Imax) at 25–30 mol% (dioleoyl)-phosphatidylglycerol ([L−]max) corresponding to a surface potential of the lipid bilayer in the absence of protein, ψ\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\setlength{\\oddsidemargin}{-69pt} \\begin{document} \\begin{equation*}{\\mathrm{_{o}^{max}}}\\end{equation*}\\end{document} = −60 ± 5 mV. Higher ionic strength shifted [L−]max to larger values, but ψ\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\setlength{\\oddsidemargin}{-69pt} \\begin{document} \\begin{equation*}{\\mathrm{_{o}^{max}}}\\end{equation*}\\end{document} remained approximately constant. It is proposed that the channel current (i) increases and (ii) decreases at |ψo| values <55 mV and >65 mV, because of (i) electrostatic interactions needed for effective insertion of the channel polypeptide and (ii) constraints due to electrostatic forces on the flexibility needed for cooperative insertion into the membrane. The loss of flexibility for |ψo| ≫ 65 mV was demonstrated by the absence of thermally induced intraprotein distance changes of the bound polypeptide. The anionic lipid content, 25–30 mol%, corresponding to the channel current maxima, is similar to that of the target Escherichia coli cytoplasmic membrane and membranes of mesophilic microorganisms. This suggests that one

  4. LDL Receptor-related Protein 1 Regulates the Abundance of Diverse Cell-signaling Proteins in the Plasma Membrane Proteome

    PubMed Central

    Gaultier, Alban; Simon, Gabriel; Niessen, Sherry; Dix, Melissa; Takimoto, Shinako; Cravatt, Benjamin F.; Gonias, Steven L.

    2010-01-01

    LDL receptor-related protein 1 (LRP1) is an endocytic receptor, reported to regulate the abundance of other receptors in the plasma membrane, including uPAR and tissue factor. The goal of this study was to identify novel plasma membrane proteins, involved in cell-signaling, which are regulated by LRP1. Membrane protein ectodomains were prepared from RAW 264.7 cells in which LRP1 was silenced and control cells using protease K. Peptides were identified by LC-MS/MS. By analysis of spectral counts, 31 transmembrane and secreted proteins were regulated in abundance at least 2-fold when LRP1 was silenced. Validation studies confirmed that semaphorin4D (Sema4D), plexin domain-containing protein-1 (Plxdc1), and neuropilin-1 were more abundant in the membranes of LRP1 gene-silenced cells. Regulation of Plxdc1 by LRP1 was confirmed in CHO cells, as a second model system. Plxdc1 co-immunoprecipitated with LRP1 from extracts of RAW 264.7 cells and mouse liver. Although Sema4D did not co-immunoprecipitate with LRP1, the cell-surface level of Sema4D was increased by RAP, which binds to LRP1 and inhibits binding of other ligands. These studies identify Plxdc1, Sema4D, and neuropilin-1 as novel LRP1-regulated cell-signaling proteins. Overall, LRP1 emerges as a generalized regulator of the plasma membrane proteome. PMID:20919742

  5. Engineering Surface Energy and Nanostructure of Microporous Films for Expanded Membrane Distillation Applications.

    PubMed

    Boo, Chanhee; Lee, Jongho; Elimelech, Menachem

    2016-08-01

    We investigated the factors that determine surface omniphobicity of microporous membranes and evaluated the potential application of these membranes in desalination of low surface tension wastewaters by membrane distillation (MD). Specifically, the effects of surface morphology and surface energy on membrane surface omniphobicity were systematically investigated by evaluating wetting resistance to low surface tension liquids. Single and multilevel re-entrant structures were achieved by using cylindrical glass fibers as a membrane substrate and grafting silica nanoparticles (SiNPs) on the fibers. Surface energy of the membrane was tuned by functionalizing the fiber substrate with fluoroalkylsilane (FAS) having two different lengths of fluoroalkyl chains. Results show that surface omniphobicity of the modified fibrous membrane increased with higher level of re-entrant structure and with lower surface energy. The secondary re-entrant structure achieved by SiNP coating on the cylindrical fibers was found to play a critical role in enhancing the surface omniphobicity. Membranes coated with SiNPs and chemically modified by the FAS with a longer fluoroalkyl chain (or lower surface energy) exhibited excellent surface omniphobicity and showed wetting resistance to low surface tension liquids such as ethanol (22.1 mN m(-1)). We further evaluated performance of the membranes in desalination of saline feed solutions with varying surface tensions by membrane distillation (MD). The engineered membranes exhibited stable MD performance with low surface tension feed waters, demonstrating the potential application omniphobic membranes in desalinating complex, high salinity industrial wastewaters. PMID:27391088

  6. Selective and programmed cleavage of GPI-anchored proteins from the surface membrane by phospholipase C.

    PubMed

    Müller, Alexandra; Klöppel, Christine; Smith-Valentine, Megan; Van Houten, Judith; Simon, Martin

    2012-01-01

    Many surface proteins of eukaryotic cells are tethered to the membrane by a GPI-anchor which is enzymatically cleavable. Here, we investigate cleavage and release of different GPI-proteins by phospholipase C from the outer membrane of the ciliate Paramecium tetraurelia. Our data indicate that different GPI-proteins are not equally cleaved as proteins of the surface antigen family are preferentially released in vitro compared to several smaller GPI-proteins. Likewise, the analysis of culture medium indicates exclusive in vivo release of surface antigens by two phospholipase C isoforms (PLC2 and PLC6). This suggests that phospholipase C shows affinity for select groups of GPI-anchored proteins. Our data also reveal an up-regulation of PLC isoforms in GPI-anchored protein cleavage during antigenic switching. As a consequence, silencing of these PLCs leads to a drastic decrease of antigen concentration in the medium. These results suggest a higher order of GPI-regulation by phospholipase C as cleavage occurs programmed and specific for single GPI-proteins instead of an unspecific shedding of the entire surface membrane GPI-content. PMID:22024023

  7. Organelle acidification negatively regulates vacuole membrane fusion in vivo.

    PubMed

    Desfougères, Yann; Vavassori, Stefano; Rompf, Maria; Gerasimaite, Ruta; Mayer, Andreas

    2016-01-01

    The V-ATPase is a proton pump consisting of a membrane-integral V0 sector and a peripheral V1 sector, which carries the ATPase activity. In vitro studies of yeast vacuole fusion and evidence from worms, flies, zebrafish and mice suggested that V0 interacts with the SNARE machinery for membrane fusion, that it promotes the induction of hemifusion and that this activity requires physical presence of V0 rather than its proton pump activity. A recent in vivo study in yeast has challenged these interpretations, concluding that fusion required solely lumenal acidification but not the V0 sector itself. Here, we identify the reasons for this discrepancy and reconcile it. We find that acute pharmacological or physiological inhibition of V-ATPase pump activity de-acidifies the vacuole lumen in living yeast cells within minutes. Time-lapse microscopy revealed that de-acidification induces vacuole fusion rather than inhibiting it. Cells expressing mutated V0 subunits that maintain vacuolar acidity were blocked in this fusion. Thus, proton pump activity of the V-ATPase negatively regulates vacuole fusion in vivo. Vacuole fusion in vivo does, however, require physical presence of a fusion-competent V0 sector. PMID:27363625

  8. Organelle acidification negatively regulates vacuole membrane fusion in vivo

    PubMed Central

    Desfougères, Yann; Vavassori, Stefano; Rompf, Maria; Gerasimaite, Ruta; Mayer, Andreas

    2016-01-01

    The V-ATPase is a proton pump consisting of a membrane-integral V0 sector and a peripheral V1 sector, which carries the ATPase activity. In vitro studies of yeast vacuole fusion and evidence from worms, flies, zebrafish and mice suggested that V0 interacts with the SNARE machinery for membrane fusion, that it promotes the induction of hemifusion and that this activity requires physical presence of V0 rather than its proton pump activity. A recent in vivo study in yeast has challenged these interpretations, concluding that fusion required solely lumenal acidification but not the V0 sector itself. Here, we identify the reasons for this discrepancy and reconcile it. We find that acute pharmacological or physiological inhibition of V-ATPase pump activity de-acidifies the vacuole lumen in living yeast cells within minutes. Time-lapse microscopy revealed that de-acidification induces vacuole fusion rather than inhibiting it. Cells expressing mutated V0 subunits that maintain vacuolar acidity were blocked in this fusion. Thus, proton pump activity of the V-ATPase negatively regulates vacuole fusion in vivo. Vacuole fusion in vivo does, however, require physical presence of a fusion-competent V0 sector. PMID:27363625

  9. Latent progenitor cells as potential regulators for tympanic membrane regeneration

    NASA Astrophysics Data System (ADS)

    Kim, Seung Won; Kim, Jangho; Seonwoo, Hoon; Jang, Kyung-Jin; Kim, Yeon Ju; Lim, Hye Jin; Lim, Ki-Taek; Tian, Chunjie; Chung, Jong Hoon; Choung, Yun-Hoon

    2015-06-01

    Tympanic membrane (TM) perforation, in particular chronic otitis media, is one of the most common clinical problems in the world and can present with sensorineural healing loss. Here, we explored an approach for TM regeneration where the latent progenitor or stem cells within TM epithelial layers may play an important regulatory role. We showed that potential TM stem cells present highly positive staining for epithelial stem cell markers in all areas of normal TM tissue. Additionally, they are present at high levels in perforated TMs, especially in proximity to the holes, regardless of acute or chronic status, suggesting that TM stem cells may be a potential factor for TM regeneration. Our study suggests that latent TM stem cells could be potential regulators of regeneration, which provides a new insight into this clinically important process and a potential target for new therapies for chronic otitis media and other eardrum injuries.

  10. Regulating the Transport of DNA through Biofriendly Nanochannels in a Thin Solid Membrane

    PubMed Central

    Wang, Deqiang; Harrer, Stefan; Luan, Binquan; Stolovitzky, Gustavo; Peng, Hongbo; Afzali-Ardakani, Ali

    2014-01-01

    Channels formed by membrane proteins regulate the transport of water, ions or nutrients that are essential to cells' metabolism. Recent advances in nanotechnology allow us to fabricate solid-state nanopores for transporting and analyzing biomolecules. However, uncontrollable surface properties of a fabricated nanopore cause irregular transport of biomolecules, limiting potential biomimetic applications. Here we show that a nanopore functionalized with a self-assembled monolayer (SAM) can potentially regulate the transport of a DNA molecule by changing functional groups of the SAM. We found that an enhanced interaction between DNA and a SAM-coated nanopore can slow down the translocation speed of DNA molecules and increase the DNA capture-rate. Our results demonstrate that the transport of DNA molecules inside nanopores could be modulated by coating a SAM on the pore surface. Our method to control the DNA motion inside a nanopore may find its applications in nanopore-based DNA sequencing devices. PMID:24496378

  11. Regulating the Transport of DNA through Biofriendly Nanochannels in a Thin Solid Membrane

    NASA Astrophysics Data System (ADS)

    Wang, Deqiang; Harrer, Stefan; Luan, Binquan; Stolovitzky, Gustavo; Peng, Hongbo; Afzali-Ardakani, Ali

    2014-02-01

    Channels formed by membrane proteins regulate the transport of water, ions or nutrients that are essential to cells' metabolism. Recent advances in nanotechnology allow us to fabricate solid-state nanopores for transporting and analyzing biomolecules. However, uncontrollable surface properties of a fabricated nanopore cause irregular transport of biomolecules, limiting potential biomimetic applications. Here we show that a nanopore functionalized with a self-assembled monolayer (SAM) can potentially regulate the transport of a DNA molecule by changing functional groups of the SAM. We found that an enhanced interaction between DNA and a SAM-coated nanopore can slow down the translocation speed of DNA molecules and increase the DNA capture-rate. Our results demonstrate that the transport of DNA molecules inside nanopores could be modulated by coating a SAM on the pore surface. Our method to control the DNA motion inside a nanopore may find its applications in nanopore-based DNA sequencing devices.

  12. Surface shape change during fusion of erythrocyte membranes is sensitive to membrane skeleton agents.

    PubMed Central

    Wu, Y; Rosenberg, J D; Sowers, A E

    1994-01-01

    We previously reported that the induction of membrane fusion between pairs of erythrocyte ghosts is accompanied by the formation of a multipore fusion zone that undergoes an area expansion with condition-dependent characteristics. These characteristics allowed us to hypothesize substantial, if not major, involvement of the spectrin-based membrane skeleton in controlling this expansion. It was also found that the fusion zone, which first appears in phase optics as a flat diaphragm, has a lifetime that is also highly condition-dependent. We report here that 2,3-diphosphoglycerate, wheat germ agglutinin, diamide, and N-ethylmaleimide, all known to have binding sites primarily on skeleton components (including spectrin), have condition-dependent effects on specific components of the fusion zone diameter versus time expansion curve and the flat diaphragm lifetime. We also report a pH/ionic strength condition that causes a dramatic stabilization of flat diaphragms in a manner consistent with the known pH/ionic strength dependence of the spectrin calorimetric transition, thus further supporting the hypothesis of spectrin involvement. Our data suggest that the influence of the membrane skeleton on cell fusion is to restrain the rounding up that takes place after membrane fusion and that it may have variable, rather than fixed, mechanical properties. Data show that WGA, a known ligand for sialic acid, and DPG, a known metabolite, influences the flat diaphragm stability and late period expansion rates, raising the possibility that some of these mechanical properties are biologically regulated. Images FIGURE 2 PMID:7858126

  13. Forming lipid bilayer membrane arrays on micropatterned polyelectrolyte film surfaces.

    PubMed

    Zhang, Ying; Wang, Lei; Wang, Xuejing; Qi, Guodong; Han, Xiaojun

    2013-07-01

    A novel method of forming lipid bilayer membrane arrays on micropatterned polyelectrolyte film surfaces is introduced. Polyelectrolyte films were fabricated by the layer-by-layer technique on a silicon oxide surface modified with a 3-aminopropyltriethoxysilane (APTES) monolayer. The surface pK(a) value of the APTES monolayer was determined by cyclic voltammetry to be approximately 5.61, on the basis of which a pH value of 2.0 was chosen for layer-by-layer assembly. Micropatterned polyelectrolyte films were obtained by deep-UV (254 nm) photolysis though a mask. Absorbed fluorescent latex beads were used to visualize the patterned surfaces. Lipid bilayer arrays were fabricated on the micropatterned surfaces by immersing the patterned substrates into a solution containing egg phosphatidylcholine vesicles. Fluorescence recovery after photobleaching studies yielded a lateral diffusion coefficient for probe molecules of 1.31±0.17 μm(2) s(-1) in the bilayer region, and migration of the lipid NBD PE in bilayer lipid membrane arrays was observed in an electric field. PMID:23695862

  14. Antimicrobial metallic copper surfaces kill Staphylococcus haemolyticus via membrane damage

    PubMed Central

    Santo, Christophe Espírito; Quaranta, Davide; Grass, Gregor

    2012-01-01

    Recently, copper (Cu) in its metallic form has regained interest for its antimicrobial properties. Use of metallic Cu surfaces in worldwide hospital trials resulted in remarkable reductions in surface contaminations. Yet, our understanding of why microbes are killed upon contact to the metal is still limited and different modes of action have been proposed. This knowledge, however, is crucial for sustained use of such surfaces in hospitals and other hygiene-sensitive areas. Here, we report on the molecular mechanisms by which the Gram-positive Staphylococcus haemolyticus is inactivated by metallic Cu. Staphylococcus haemolyticus was killed within minutes on Cu but not on stainless steel demonstrating the antimicrobial efficacy of metallic Cu. Inductively coupled plasma mass spectroscopy (ICP-MS) analysis and in vivo staining with Coppersensor-1 indicated that cells accumulated large amounts of Cu ions from metallic Cu surfaces contributing to lethal damage. Mutation rates of Cu- or steel-exposed cells were similarly low. Instead, live/dead staining indicated cell membrane damage in Cu- but not steel-exposed cells. These findings support a model of the cellular targets of metallic Cu toxicity in bacteria, which suggests that metallic Cu is not genotoxic and does not kill via DNA damage. In contrast, membranes constitute the likely Achilles’ heel of Cu surface-exposed cells. PMID:22950011

  15. Hypoxia regulates global membrane protein endocytosis through caveolin-1 in cancer cells

    PubMed Central

    Bourseau-Guilmain, E.; Menard, J. A.; Lindqvist, E.; Indira Chandran, V.; Christianson, H. C.; Cerezo Magaña, M.; Lidfeldt, J.; Marko-Varga, G.; Welinder, C.; Belting, M.

    2016-01-01

    Hypoxia promotes tumour aggressiveness and resistance of cancers to oncological treatment. The identification of cancer cell internalizing antigens for drug targeting to the hypoxic tumour niche remains a challenge of high clinical relevance. Here we show that hypoxia down-regulates the surface proteome at the global level and, more specifically, membrane proteome internalization. We find that hypoxic down-regulation of constitutive endocytosis is HIF-independent, and involves caveolin-1-mediated inhibition of dynamin-dependent, membrane raft endocytosis. Caveolin-1 overexpression inhibits protein internalization, suggesting a general negative regulatory role of caveolin-1 in endocytosis. In contrast to this global inhibitory effect, we identify several proteins that can override caveolin-1 negative regulation, exhibiting increased internalization at hypoxia. We demonstrate antibody-mediated cytotoxin delivery and killing specifically of hypoxic cells through one of these proteins, carbonic anhydrase IX. Our data reveal that caveolin-1 modulates cell-surface proteome turnover at hypoxia with potential implications for specific targeting of the hypoxic tumour microenvironment. PMID:27094744

  16. Hypoxia regulates global membrane protein endocytosis through caveolin-1 in cancer cells.

    PubMed

    Bourseau-Guilmain, E; Menard, J A; Lindqvist, E; Indira Chandran, V; Christianson, H C; Cerezo Magaña, M; Lidfeldt, J; Marko-Varga, G; Welinder, C; Belting, M

    2016-01-01

    Hypoxia promotes tumour aggressiveness and resistance of cancers to oncological treatment. The identification of cancer cell internalizing antigens for drug targeting to the hypoxic tumour niche remains a challenge of high clinical relevance. Here we show that hypoxia down-regulates the surface proteome at the global level and, more specifically, membrane proteome internalization. We find that hypoxic down-regulation of constitutive endocytosis is HIF-independent, and involves caveolin-1-mediated inhibition of dynamin-dependent, membrane raft endocytosis. Caveolin-1 overexpression inhibits protein internalization, suggesting a general negative regulatory role of caveolin-1 in endocytosis. In contrast to this global inhibitory effect, we identify several proteins that can override caveolin-1 negative regulation, exhibiting increased internalization at hypoxia. We demonstrate antibody-mediated cytotoxin delivery and killing specifically of hypoxic cells through one of these proteins, carbonic anhydrase IX. Our data reveal that caveolin-1 modulates cell-surface proteome turnover at hypoxia with potential implications for specific targeting of the hypoxic tumour microenvironment. PMID:27094744

  17. Antibacterial and hemocompatibility switchable polypropylene nonwoven fabric membrane surface.

    PubMed

    Zhao, Jie; Song, Lingjie; Shi, Qiang; Luan, Shifang; Yin, Jinghua

    2013-06-12

    In this article, a facile approach to fabricate a biofunctional polypropylene nonwoven fabric membrane (PP NWF) with a switchable surface from antibacterial property to hemocompatibility is presented. In the first step, a cationic carboxybetaine ester monomer, [(2-(methacryboxy) ethyl)]-N,N-dimethylamino-ethylammonium bromide, methyl ester (CABA-1-ester) was synthesized. Subsequently, this monomer was introduced on the PP NWF surface via plasma pretreatment and a UV-induced graft polymerization technique. Finally, a switchable surface from antibacterial property to hemocompatibility was easily realized by hydrolysis of poly(CABA-1-ester) moieties on the PP NWF surface under mild conditions. Surface hydrolysis behaviors under different pH conditions were investigated. These PP NWFs grafted with poly(CABA-1-ester) segments can cause significant suppression of S. aureus proliferation; after hydrolysis, these surfaces covered by poly[(2-(methacryloxy) ethyl)] carboxybetaine (poly(CABA)) chains exhibited obvious reduction in protein adsorption and platelet adhesion, and remarkably enhanced antithrombotic properties. This strategy demonstrated that a switchable PP NWF surface from antibacterial property to hemocompatibility was easily developed by plasma pretreatment and UV-induced surface graft polymerization and that this surface may become an attractive platform for a range of biomedical applications. PMID:23668294

  18. Roles of the reproductive tract in modifications of the sperm membrane surface

    PubMed Central

    KUO, Yu-Wen; LI, Sheng-Hsiang; MAEDA, Kei-Ichiro; GADELLA, Bart M.; TSAI, Pei Shiue J.

    2016-01-01

    Successful fertilization requires viable and functional spermatozoa to recognize and fuse with the oocyte. In most mammalian species, mature spermatozoa are not capable of fertilizing the oocytes immediately after ejaculation. However, unlike somatic cells, spermatozoa, after leaving the testis, are transcriptionally and translationally silent; therefore, upon completion of spermiogenesis, spermatozoa carry only a minimal amount of essential proteins on their membranes as well as within their restricted volume of cytoplasm. To develop into a fully functional and competent sperm that is capable of successful fertilization, modifications of the sperm membrane surface during its transit in the reproductive tracts is critical. These post-spermatogenesis modifications advance the maturation of epididymal spermatozoa. In addition, components secreted into the lumen of the reproductive tracts that are later added onto the sperm membrane surface also regulate (inhibit or activate) the functions of the spermatozoa. This acquisition of additional proteins from the reproductive tracts may compensate for the inactivity of morphologically mature spermatozoa. In this review, we discuss the contributions of the male and female genital tracts to modifications of the sperm membrane surface at different stages of fertilization. PMID:27009019

  19. Salmonella Rapidly Regulates Membrane Permeability To Survive Oxidative Stress

    PubMed Central

    van der Heijden, Joris; Reynolds, Lisa A.; Deng, Wanyin; Mills, Allan; Scholz, Roland; Imami, Koshi; Foster, Leonard J.; Duong, Franck

    2016-01-01

    ABSTRACT The outer membrane (OM) of Gram-negative bacteria provides protection against toxic molecules, including reactive oxygen species (ROS). Decreased OM permeability can promote bacterial survival under harsh circumstances and protects against antibiotics. To better understand the regulation of OM permeability, we studied the real-time influx of hydrogen peroxide in Salmonella bacteria and discovered two novel mechanisms by which they rapidly control OM permeability. We found that pores in two major OM proteins, OmpA and OmpC, could be rapidly opened or closed when oxidative stress is encountered and that the underlying mechanisms rely on the formation of disulfide bonds in the periplasmic domain of OmpA and TrxA, respectively. Additionally, we found that a Salmonella mutant showing increased OM permeability was killed more effectively by treatment with antibiotics. Together, these results demonstrate that Gram-negative bacteria regulate the influx of ROS for defense against oxidative stress and reveal novel targets that can be therapeutically targeted to increase bacterial killing by conventional antibiotics. PMID:27507830

  20. Regulation of peptidoglycan synthesis by outer membrane proteins

    PubMed Central

    Typas, Athanasios; Banzhaf, Manuel; van den Berg van Saparoea, Bart; Verheul, Jolanda; Biboy, Jacob; Nichols, Robert J.; Zietek, Matylda; Beilharz, Katrin; Kannenberg, Kai; von Rechenberg, Moritz; Breukink, Eefjan; den Blaauwen, Tanneke; Gross, Carol A.; Vollmer, Waldemar

    2011-01-01

    Summary Growth of the meshlike peptidoglycan (PG) sacculus located between the bacterial inner and outer membranes (OM) is tightly regulated to ensure cellular integrity, maintain cell shape and orchestrate division. Cytoskeletal elements direct placement and activity of PG synthases from inside the cell but precise spatiotemporal control over this process is poorly understood. We demonstrate that PG synthases are also controlled from outside the sacculus. Two OM lipoproteins, LpoA and LpoB, are essential for the function respectively of PBP1A and PBP1B, the major E. coli bifunctional PG synthases. Each Lpo protein binds specifically to its cognate PBP and stimulates its transpeptidase activity, thereby facilitating attachment of new PG to the sacculus. LpoB shows partial septal localization and our data suggest that the LpoB-PBP1B complex contributes to OM constriction during cell division. LpoA/ LpoB and their PBP docking regions are restricted to γ-proteobacteria, providing models for niche-specific regulation of sacculus growth. PMID:21183073

  1. Substrate-Supported Phospholipid Membranes Studied by Surface Plasmon Resonance and Surface Plasmon Fluorescence Spectroscopy

    PubMed Central

    Tawa, Keiko; Morigaki, Kenichi

    2005-01-01

    Substrate-supported planar lipid bilayer membranes are attractive model cellular membranes for biotechnological applications such as biochips and sensors. However, reliable fabrication of the lipid membranes on solid surfaces still poses significant technological challenges. In this study, simultaneous surface plasmon resonance (SPR) and surface plasmon fluorescence spectroscopy (SPFS) measurements were applied to the monitoring of adsorption and subsequent reorganization of phospholipid vesicles on solid substrates. The fluorescence intensity of SPFS depends very sensitively on the distance between the gold substrate and the fluorophore because of the excitation energy transfer to gold. By utilizing this distance dependency, we could obtain information about the topography of the adsorbed membranes: Adsorbed vesicles could be clearly distinguished from planar bilayers due to the high fluorescence intensity. SPSF can also incorporate various analytical techniques to evaluate the physicochemical properties of the adsorbed membranes. As an example, we demonstrated that the lateral mobility of lipid molecules could be estimated by observing the recovery of fluorescence after photobleaching. Combined with the film thickness information obtained by SPR, SPR-SPFS proved to be a highly informative technique to monitor the lipid membrane assembly processes on solid substrates. PMID:16040759

  2. Syndecan-4 Regulates Muscle Differentiation and Is Internalized from the Plasma Membrane during Myogenesis

    PubMed Central

    Rønning, Sissel B.; Carlson, Cathrine R.; Stang, Espen; Kolset, Svein O.; Hollung, Kristin; Pedersen, Mona E.

    2015-01-01

    The cell surface proteoglycan syndecan-4 has been reported to be crucial for muscle differentiation, but the molecular mechanisms still remain to be fully understood. During in vitro differentiation of bovine muscle cells immunocytochemical analyses showed strong labelling of syndecan-4 intracellularly, in close proximity with Golgi structures, in membranes of intracellular vesicles and finally, in the nuclear area including the nuclear envelope. Chase experiments showed that syndecan-4 was internalized from the plasma membrane during this process. Furthermore, when syndecan-4 was knocked down by siRNA more myotubes were formed, and the expression of myogenic transcription factors, β1-integrin and actin was influenced. However, when bovine muscle cells were treated with a cell-penetrating peptide containing the cytoplasmic region of syndecan-4, myoblast fusion and thus myotube formation was blocked, both in normal cells and in syndecan-4 knock down cells. Altogether this suggests that the cytoplasmic domain of syndecan-4 is important in regulation of myogenesis. The internalization of syndecan-4 from the plasma membrane during muscle differentiation and the nuclear localization of syndecan-4 in differentiated muscle cells may be part of this regulation, and is a novel aspect of syndecan biology which merits further studies. PMID:26068620

  3. Detection of molecular interactions at membrane surfaces through colloid phase transitions

    NASA Astrophysics Data System (ADS)

    Baksh, Michael M.; Jaros, Michal; Groves, Jay T.

    2004-01-01

    The molecular architecture of-and biochemical processes within-cell membranes play important roles in all living organisms, with many drugs and infectious disease agents targeting membranes. Experimental studies of biochemical reactions on membrane surfaces are challenging, as they require a membrane environment that is fluid (like cell membranes) but nevertheless allows for the efficient detection and characterization of molecular interactions. One approach uses lipid membranes supported on solid substrates such as silica or polymers: although the membrane is trapped near the solid interface, it retains natural fluidity and biological functionality and can be implanted with membrane proteins for functional studies. But the detection of molecular interactions involving membrane-bound species generally requires elaborate techniques, such as surface plasmon resonance or total internal reflection fluorescence microscopy. Here we demonstrate that colloidal phase transitions of membrane-coated silica beads provide a simple and label-free method for monitoring molecular interactions on lipid membrane surfaces. By adjusting the lipid membrane composition and hence the pair interaction potential between the membrane-supporting silica beads, we poise our system near a phase transition so that small perturbations on the membrane surface induce dramatic changes in the macroscopic organization of the colloid. We expect that this approach, used here to probe with high sensitivity protein binding events at membrane surfaces, can be applied to study a broad range of cell membrane processes.

  4. A new method for modeling rough membrane surface and calculation of interfacial interactions.

    PubMed

    Zhao, Leihong; Zhang, Meijia; He, Yiming; Chen, Jianrong; Hong, Huachang; Liao, Bao-Qiang; Lin, Hongjun

    2016-01-01

    Membrane fouling control necessitates the establishment of an effective method to assess interfacial interactions between foulants and rough surface membrane. This study proposed a new method which includes a rigorous mathematical equation for modeling membrane surface morphology, and combination of surface element integration (SEI) method and the composite Simpson's approach for assessment of interfacial interactions. The new method provides a complete solution to quantitatively calculate interfacial interactions between foulants and rough surface membrane. Application of this method in a membrane bioreactor (MBR) showed that, high calculation accuracy could be achieved by setting high segment number, and moreover, the strength of three energy components and energy barrier was remarkably impaired by the existence of roughness on the membrane surface, indicating that membrane surface morphology exerted profound effects on membrane fouling in the MBR. Good agreement between calculation prediction and fouling phenomena was found, suggesting the feasibility of this method. PMID:26519696

  5. Surface Tension, Pressure Difference and Laplace Formula for Membranes

    NASA Astrophysics Data System (ADS)

    Koibuchi, Hiroshi; Shobukhov, Andrey

    2015-01-01

    The surface tension γ and the pressure difference Δp for spherical membranes are calculated using Monte Carlo simulation technique. We study the so-called tethered and uid surface discrete models that are defined on the fixed-connectivity (tethered) and dynamically triangulated (uid) lattices respectively. Hamiltonians of the models include a self-avoiding potential, which makes the enclosed volume well defined. We find that there is reasonable accuracy in the technique for the calculation of γ using the real area A if the bending rigidity κ or A/N is sufficiently large. We also find that γ becomes constant in the limit of A/N → ∞ both in the tethered and uid surfaces. The property limA/N→∞ γ = const corresponds to certain experimental results in cell biology.

  6. Intermonolayer Friction and Surface Shear Viscosity of Lipid Bilayer Membranes

    PubMed Central

    den Otter, W. K.; Shkulipa, S. A.

    2007-01-01

    The flow behavior of lipid bilayer membranes is characterized by a surface viscosity for in-plane shear deformations, and an intermonolayer friction coefficient for slip between the two leaflets of the bilayer. Both properties have been studied for a variety of coarse-grained double-tailed model lipids, using equilibrium and nonequilibrium molecular dynamics simulations. For lipids with two identical tails, the surface shear viscosity rises rapidly with tail length, while the intermonolayer friction coefficient is less sensitive to the tail length. Interdigitation of lipid tails across the bilayer midsurface, as observed for lipids with two distinct tails, strongly enhances the intermonolayer friction coefficient, but hardly affects the surface shear viscosity. The simulation results are compared against the available experimental data. PMID:17468168

  7. Temporal changes in extracellular polymeric substances on hydrophobic and hydrophilic membrane surfaces in a submerged membrane bioreactor.

    PubMed

    Matar, Gerald; Gonzalez-Gil, Graciela; Maab, Husnul; Nunes, Suzana; Le-Clech, Pierre; Vrouwenvelder, Johannes; Saikaly, Pascal E

    2016-05-15

    Membrane surface hydrophilic modification has always been considered to mitigating biofouling in membrane bioreactors (MBRs). Four hollow-fiber ultrafiltration membranes (pore sizes ∼0.1 μm) differing only in hydrophobic or hydrophilic surface characteristics were operated at a permeate flux of 10 L/m(2) h in the same lab-scale MBR fed with synthetic wastewater. In addition, identical membrane modules without permeate production (0 L/m(2) h) were operated in the same lab-scale MBR. Membrane modules were autopsied after 1, 10, 20 and 30 days of MBR operation, and total extracellular polymeric substances (EPS) accumulated on the membranes were extracted and characterized in detail using several analytical tools, including conventional colorimetric tests (Lowry and Dubois), liquid chromatography with organic carbon detection (LC-OCD), fluorescence excitation - emission matrices (FEEM), fourier transform infrared (FTIR) and confocal laser scanning microscope (CLSM). The transmembrane pressure (TMP) quickly stabilized with higher values for the hydrophobic membranes than hydrophilic ones. The sulfonated polysulfone (SPSU) membrane had the highest negatively charged membrane surface, accumulated the least amount of foulants and displayed the lowest TMP. The same type of organic foulants developed with time on the four membranes and the composition of biopolymers shifted from protein dominance at early stages of filtration (day 1) towards polysaccharides dominance during later stages of MBR filtration. Nonmetric multidimensional scaling of LC-OCD data showed that biofilm samples clustered according to the sampling event (time) regardless of the membrane surface chemistry (hydrophobic or hydrophilic) or operating mode (with or without permeate flux). These results suggest that EPS composition may not be the dominant parameter for evaluating membrane performance and possibly other parameters such as biofilm thickness, porosity, compactness and structure should be

  8. Membrane and Integrative Nuclear Fibroblastic Growth Factor Receptor (FGFR) Regulation of FGF-23*

    PubMed Central

    Han, Xiaobin; Xiao, Zhousheng; Quarles, L. Darryl

    2015-01-01

    Fibroblastic growth factor receptor 1 (FGFR1) signaling pathways are implicated in the regulation of FGF-23 gene transcription, but the molecular pathways remain poorly defined. We used low molecular weight (LMW, 18 kDa) FGF-2 and high molecular weight (HMW) FGF-2 isoforms, which, respectively, activate cell surface FGF receptors and intranuclear FGFR1, to determine the roles of membrane FGFRs and integrative nuclear FGFR1 signaling (INFS) in the regulation of FGF-23 gene transcription in osteoblasts. We found that LMW-FGF-2 induced NFAT and Ets1 binding to conserved cis-elements in the proximal FGF-23 promoter and stimulated FGF-23 promoter activity through PLCγ/calcineurin/NFAT and MAPK pathways in SaOS-2 and MC3T3-E1 osteoblasts. In contrast, HMW-FGF-2 stimulated FGF-23 promoter activity in osteoblasts through a cAMP-dependent binding of FGFR1 and cAMP-response element-binding protein (CREB) to a conserved cAMP response element (CRE) contiguous with the NFAT binding site in the FGF-23 promoter. Mutagenesis of the NFAT and CRE binding sites, respectively, inhibited the effects of LMW-FGF-2 and HMW-FGF-23 to stimulate FGF-23 promoter activity. FGF-2 activation of both membrane FGFRs and INFS-dependent FGFR1 pathways may provide a means to integrate systemic and local regulation of FGF-23 transcription under diverse physiological and pathological conditions. PMID:25752607

  9. Regulation of human airway surface liquid.

    PubMed

    Widdicombe, J H; Widdicombe, J G

    1995-01-01

    Human airways are lined with a film of liquid from 5-100 microns in depth, consisting of a periciliary sol around and a mucous gel above the cilia. Microscopical studies have shown the sol to be invariably the same depth as the length of the cilia, and we discuss possible reasons for this. The composition and sources of the airway surface liquid are also described. In addition the forces regulating its volume are analyzed. Several airway diseases are characterised by dramatic changes in the volume and composition of airway liquid. We review recent research suggesting that the accumulation of airway mucous secretions in cystic fibrosis is caused by alterations in active transport of ions and water across both the surface and gland epithelia. PMID:7740210

  10. Generation, modulation and maintenance of the plasma membrane asymmetric phospholipid composition in yeast cells during growth: their relation to surface potential and membrane protein activity.

    PubMed

    Cerbón, J; Calderón, V

    1995-04-12

    During growth a cyclic exposure of anionic phospholipids to the external surface of the plasma membrane was found. The surface charge density (sigma) increased gradually reaching a maximum in the first 5 h of growth and returned gradually to their initial value at the end of the logarithmic phase of growth (10-12 h). Phosphatidylinositol, that determines to a large extent the magnitude of the sigma, increased 83% in the yeast cells during the first 4 h of growth and returned gradually to their initial level at 10-12 h. During the stationary phase (12-24 h), both sigma and the anionic/zwitterionic phospholipid ratio, remained without any significant variation. The high-affinity H-linked glutamate transport system that behaves as a sensor of the changes in the membrane surface potential (phi) increased its activity in the first 5 h and then decreased it, following with great accuracy the sigma variations and remained without changes during the stationary phase of growth. The phosphatidylserine (PS) relative concentration in the cells (9.0%) did not significantly change during the whole growth curve, but their asymmetric distribution varied, contributing to the changes in sigma. PS facing the outer membrane surface increased 2.45-times during the first 5 h of growth and then returned to their original value at the end of the log phase (12 h). Phosphatidylcholine (PC) remained constant during the whole growth curve (50%), while phosphatidylethanolamine (PE) decreased 3-fold in the first 4 h and then increased to its original value at 10 h. Interestingly, PE at the outer membrane surface remained constant (3% of the total phospholipids) during the whole growth curve. During growth yeast cells change their phospholipid composition originating altered patterns of the plasma membrane phospholipid composition and IN-OUT distribution. This dynamic asymmetry is involved in the regulation of the surface potential and membrane protein activity. PMID:7718598

  11. Theory and simulations of adhesion receptor dimerization on membrane surfaces.

    PubMed

    Wu, Yinghao; Honig, Barry; Ben-Shaul, Avinoam

    2013-03-19

    The equilibrium constants of trans and cis dimerization of membrane bound (2D) and freely moving (3D) adhesion receptors are expressed and compared using elementary statistical-thermodynamics. Both processes are mediated by the binding of extracellular subdomains whose range of motion in the 2D environment is reduced upon dimerization, defining a thin reaction shell where dimer formation and dissociation take place. We show that the ratio between the 2D and 3D equilibrium constants can be expressed as a product of individual factors describing, respectively, the spatial ranges of motions of the adhesive domains, and their rotational freedom within the reaction shell. The results predicted by the theory are compared to those obtained from a novel, to our knowledge, dynamical simulations methodology, whereby pairs of receptors perform realistic translational, internal, and rotational motions in 2D and 3D. We use cadherins as our model system. The theory and simulations explain how the strength of cis and trans interactions of adhesive receptors are affected both by their presence in the constrained intermembrane space and by the 2D environment of membrane surfaces. Our work provides fundamental insights as to the mechanism of lateral clustering of adhesion receptors after cell-cell contact and, more generally, to the formation of lateral microclusters of proteins on cell surfaces. PMID:23528081

  12. Influence of membrane galactolipids and surface pressure on plastoquinone behaviour.

    PubMed

    Hoyo, Javier; Guaus, Ester; Torrent-Burgués, Juan

    2016-10-01

    In this work biomimetic monolayers of a MGDG, monogalactosyldiacylglycerol, and DGDG, digalactosyldiacylglycerol mixture (MD), in a ratio close to that of the thylakoid membranes of oxygenic photosynthetic organisms, have been prepared. The lipid mixture incorporates plastoquinone-9 (PQ), that is the electron and proton shuttle of the photosynthetic reaction centres. The MD:PQ mixtures have been firstly studied using surface pressure-area isotherms. Langmuir-Blodgett (LB) films of those mixtures have been transferred onto a substrate forming a monolayer that mimics one of the bilayer sides of the thylakoid membranes. These monolayers have been characterized topographically and electrochemically. The results show the influence of PQ in the MD matrix and its partial expulsion when increasing the surface pressure, obtaining two main PQ positions in the MD matrix. The calculated apparent electron transfer rate constants indicate a different kinetic control for the reduction and the oxidation of the PQ/PQH2 couple, being kRapp(I)=0.7·10(-6)s(-1), kRapp(II)=2.2·10(-9)s(-1), kOapp(I)=7.4·10(-4)s(-1) and kOapp(II)=5.2·10(-5)s(-1), respectively. The comparison of the different galactolipid:PQ systems that our group has studied is also presented, concluding that the PQ position in the galactolipid matrix can be tuned according to several controlled variables. PMID:27317998

  13. Regulation of ubiquitin-dependent cargo sorting by multiple endocytic adaptors at the plasma membrane

    PubMed Central

    Mayers, Jonathan R.; Wang, Lei; Pramanik, Jhuma; Johnson, Adam; Sarkeshik, Ali; Wang, Yueju; Saengsawang, Witchuda; Yates, John R.; Audhya, Anjon

    2013-01-01

    Endocytic protein trafficking is directed by sorting signals on cargo molecules that are recognized by cytosolic adaptor proteins. However, the steps necessary to segregate the variety of cargoes during endocytosis remain poorly defined. Using Caenorhabditis elegans, we demonstrate that multiple plasma membrane endocytic adaptors function redundantly to regulate clathrin-mediated endocytosis and to recruit components of the endosomal sorting complex required for transport (ESCRT) machinery to the cell surface to direct the sorting of ubiquitin-modified substrates. Moreover, our data suggest that preassembly of cargoes with the ESCRT-0 complex at the plasma membrane enhances the efficiency of downstream sorting events in the endolysosomal system. In the absence of a heterooligomeric adaptor complex composed of FCHO, Eps15, and intersectin, ESCRT-0 accumulation at the cell surface is diminished, and the degradation of a ubiquitin-modified cargo slows significantly without affecting the rate of its clathrin-mediated internalization. Consistent with a role for the ESCRT machinery during cargo endocytosis, we further show that the ESCRT-0 complex accumulates at a subset of clathrin-coated pits on the surface of human cells. Our findings suggest a unique mechanism by which ubiquitin-modified cargoes are sequestered into the endolysosomal pathway. PMID:23818590

  14. Alternative 3'UTRs act as scaffolds to regulate membrane protein localization

    PubMed Central

    Berkovits, Binyamin D.; Mayr, Christine

    2015-01-01

    About half of human genes use alternative cleavage and polyadenylation (ApA) to generate mRNA transcripts that differ in the length of their 3' untranslated regions (3'UTRs) while producing the same protein 1–3. Here we show in human cell lines that alternative 3' UTRs differentially regulate the localization of membrane proteins. The long 3'UTR of CD47 enables efficient cell surface expression of CD47 protein, whereas the short 3'UTR primarily localizes CD47 protein to the endoplasmic reticulum. CD47 protein localization occurs post-translationally and independently of RNA localization. In our model of 3' UTR-dependent protein localization, the long 3' UTR of CD47 acts as a scaffold to recruit a protein complex containing the RNA-binding protein HuR (also known as ELAVL1) and SET4 to the site of translation. This facilitates interaction of SET with the newly translated cytoplasmic domains of CD47 and results in subsequent translocation of CD47 to the plasma membrane via activated RAC1 5. We also show that CD47 protein has different functions depending on whether it was generated by the short or long 3'UTR isoforms. Thus, ApA contributes to the functional diversity of the proteome without changing the amino acid sequence. 3' UTR-dependent protein localization has the potential to be a widespread trafficking mechanism for membrane proteins because HuR binds to thousands of mRNAs6–9, and we show that the long 3' UTRs of CD44, ITGA1 and TNFRSF13C, which are bound by HuR, increase surface protein expression compared to their corresponding short 3' UTRs. We propose that during translation the scaffold function of 3' UTRs facilitates binding of proteins to nascent proteins to direct their transport or function—and that this role of 3' UTRs can be regulated by ApA. PMID:25896326

  15. Effect of membrane polymeric materials on relationship between surface pore size and membrane fouling in membrane bioreactors

    NASA Astrophysics Data System (ADS)

    Miyoshi, Taro; Yuasa, Kotaku; Ishigami, Toru; Rajabzadeh, Saeid; Kamio, Eiji; Ohmukai, Yoshikage; Saeki, Daisuke; Ni, Jinren; Matsuyama, Hideto

    2015-03-01

    We investigated the effect of different membrane polymeric materials on the relationship between membrane pore size and development of membrane fouling in a membrane bioreactor (MBR). Membranes with different pore sizes were prepared using three different polymeric materials, cellulose acetate butyrate (CAB), polyvinyl butyral (PVB), and polyvinylidene fluoride (PVDF), and the development of membrane fouling in each membrane was evaluated by batch filtration tests using a mixed liquor suspension obtained from a laboratory-scale MBR. The results revealed that the optimal membrane pore size to mitigate membrane fouling differed depending on membrane polymeric material. For PVDF membranes, the degree of membrane fouling decreased as membrane pore size increased. In contrast, CAB membranes with smaller pores had less fouling propensity than those with larger ones. Such difference can be attributed to the difference in major membrane foulants in each membrane; in PVDF, they were small colloids or dissolved organics in which proteins are abundant, and in CAB, microbial flocs. The results obtained in this study strongly suggested that optimum operating conditions of MBRs differ depending on the characteristics of the used membrane.

  16. A RhoA and Rnd3 cycle regulates actin reassembly during membrane blebbing.

    PubMed

    Aoki, Kana; Maeda, Fumiyo; Nagasako, Tomoya; Mochizuki, Yuki; Uchida, Seiichi; Ikenouchi, Junichi

    2016-03-29

    The actin cytoskeleton usually lies beneath the plasma membrane. When the membrane-associated actin cytoskeleton is transiently disrupted or the intracellular pressure is increased, the plasma membrane detaches from the cortex and protrudes. Such protruded membrane regions are called blebs. However, the molecular mechanisms underlying membrane blebbing are poorly understood. This study revealed that epidermal growth factor receptor kinase substrate 8 (Eps8) and ezrin are important regulators of rapid actin reassembly for the initiation and retraction of protruded blebs. Live-cell imaging of membrane blebbing revealed that local reassembly of actin filaments occurred at Eps8- and activated ezrin-positive foci of membrane blebs. Furthermore, we found that a RhoA-ROCK-Rnd3 feedback loop determined the local reassembly sites of the actin cortex during membrane blebbing. PMID:26976596

  17. Membrane regulation of the stress response from prokaryotic models to mammalian cells.

    PubMed

    Vigh, Laszlo; Nakamoto, Hitoshi; Landry, Jacques; Gomez-Munoz, Antonio; Harwood, John L; Horvath, Ibolya

    2007-10-01

    "Membrane regulation" of stress responses in various systems is widely studied. In poikilotherms, membrane rigidification could be the first reaction to cold perception: reducing membrane fluidity of membranes at physiological temperatures is coupled with enhanced cold inducibility of a number of genes, including desaturases (see J.L. Harwood's article in this Proceedings volume). A similar role of changes in membrane physical state in heat (oxidative stress, etc.) sensing- and signaling gained support recently from prokaryotes to mammalian cells. Stress-induced remodeling of membrane lipids could influence generation, transduction, and deactivation of stress signals, either through global effects on the fluidity of the membrane matrix, or by specific interactions of boundary (or raft) lipids with receptor proteins, lipases, ion channels, etc. Our data point to membranes not only as targets of stress, but also as sensors in activating a stress response. PMID:17656573

  18. Sialidases as regulators of bioengineered cellular surfaces

    PubMed Central

    Zamora, Cristina Y; Ryan, Matthew J; d'Alarcao, Marc; Kumar, Krishna

    2015-01-01

    Human sialidases (NEUs) catalyze the removal of N-acetyl neuraminic acids from the glycome of the cell and regulate a diverse repertoire of nominal cellular functions, such as cell signaling and adhesion. A greater understanding of their substrate permissivity is of interest in order to discern their physiological functions in disease states and in the design of specific and effective small molecule inhibitors. Towards this, we have synthesized soluble fluorogenic reporters of mammalian sialidase activity bearing unnatural sialic acids commonly incorporated into the cellular glycocalyx via metabolic glycoengineering. We found cell-surface sialidases in Jurkat capable of cleaving unnatural sialic acids with differential activities toward a variety of R groups on neuraminic acid. In addition, we observed modulated structure–activity relationships when cell-surface sialidases were presented glycans with unnatural bulky, hydrophobic or fluorinated moieties incorporated directly via glycoengineering. Our results confirm the importance of cell-surface sialidases in glycoengineering incorporation data. We demonstrate the flexibility of human NEUs toward derivatized sugars and highlight the importance of native glycan presentation to sialidase binding and activity. These results stand to inform not only metabolic glycoengineering efforts but also inhibitor design. PMID:25795684

  19. Surface modification of titanium membrane by chemical vapor deposition and its electrochemical self-cleaning

    NASA Astrophysics Data System (ADS)

    Li, X. W.; Li, J. X.; Gao, C. Y.; Chang, M.

    2011-10-01

    Membrane separation is applied widely in many fields, while concentration polarization and membrane fouling, limiting its promotion and application greatly, are the bottlenecks in membrane application. Among which, membrane fouling is irreversible, membrane must be periodically cleaned or even replaced to restore permeability. Membrane cleaning has become one of the key issues in membrane separation areas. Considering incomparable electrochemical advantages of boron-doped diamond (BDD) film electrode over conventional electrode, a new composite membrane Ti/BDD, made by depositing CVD (chemical vapor deposition) boron-doped diamond film on titanium(Ti) membrane to modify porous titanium surface, that can be cleaned electrochemically is proposed. Feasibility of its preparation and application is discussed in this paper. Results shows that based on the unique electrochemical properties of diamond, cleaning level of this composite Ti/BDD membrane is significantly increased, making membrane life and efficiency improved prominently.

  20. An Unrecognized Function of Cholesterol: Regulating the Mechanism Controlling Membrane Phospholipid Asymmetry.

    PubMed

    Arashiki, Nobuto; Saito, Masaki; Koshino, Ichiro; Kamata, Kotoe; Hale, John; Mohandas, Narla; Manno, Sumie; Takakuwa, Yuichi

    2016-06-28

    An asymmetric distribution of phospholipids in the membrane bilayer is inseparable from physiological functions, including shape preservation and survival of erythrocytes, and by implication other cells. Aminophospholipids, notably phosphatidylserine (PS), are confined to the inner leaflet of the erythrocyte membrane lipid bilayer by the ATP-dependent flippase enzyme, ATP11C, counteracting the activity of an ATP-independent scramblase. Phospholipid scramblase 1 (PLSCR1), a single-transmembrane protein, was previously reported to possess scrambling activity in erythrocytes. However, its function was cast in doubt by the retention of scramblase activity in erythrocytes of knockout mice lacking this protein. We show that in the human erythrocyte PLSCR1 is the predominant scramblase and by reconstitution into liposomes that its activity resides in the transmembrane domain. At or below physiological intracellular calcium concentrations, total suppression of flippase activity nevertheless leaves the membrane asymmetry undisturbed. When liposomes or erythrocytes are depleted of cholesterol (a reversible process in the case of erythrocytes), PS quickly appears at the outer surface, implying that cholesterol acts in the cell as a powerful scramblase inhibitor. Thus, our results bring to light a previously unsuspected function of cholesterol in regulating phospholipid scrambling. PMID:27267274

  1. Decreased Membrane Complement Regulators in the RPE Contributes to AMD

    PubMed Central

    Ebrahimi, Katayoon B.; Fijalkowski, Natalia; Cano, Marisol; Handa, James T.

    2013-01-01

    Dysregulated complement is thought to play a central role in AMD pathogenesis, but the specific mechanisms have yet to be determined. In maculas of AMD specimens, we found that the complement regulatory protein, CD59, was increased in regions of uninvolved retinal pigmented epithelium (RPE) of early AMD, but decreased in the RPE overlying drusen and in geographic atrophy, an advanced form of AMD. While CD46 immunostaining was basolaterally distributed in the RPE of unaffected controls, it was decreased in diseased areas of early AMD samples. Since oxidized low density lipoproteins (oxLDL) collect in drusen of AMD and are a known complement trigger, we treated ARPE-19 cells with oxLDL and found that cellular CD46 and CD59 proteins were decreased by 2.9-fold and 9-fold (p<0.01), respectively. OxLDLs increased complement factor B mRNA and Bb protein, but not factor D, I, or H. OxLDLs increased C3b, but not C3a, C5 or C5b-9. C5b-9 was increased by 27% (p<0.01) when medium was supplemented with human serum, which was sufficient to induce poly (ADP-ribose) polymerase cleavage, a marker of apoptosis. The decreased levels of CD46 and CD59 were in part, explained by their release in exosomal and apoptotic membranous particles. In addition, CD59 was partially degraded through activation of IRE1α. Collectively, these results suggest that a combination of impaired complement regulators results in inadequately controlled complement by the RPE in AMD that induces RPE damage. PMID:23097248

  2. Effects of fluid flow on elution of hydrophilic modifier from dialysis membrane surfaces.

    PubMed

    Matsuda, Masato; Sato, Mika; Sakata, Hiroki; Ogawa, Takahisa; Yamamoto, Ken-ichiro; Yakushiji, Taiji; Fukuda, Makoto; Miyasaka, Takehiro; Sakai, Kiyotaka

    2008-01-01

    When uremic blood flows through dialyzers during hemodialysis, dialysis membrane surfaces are exposed to shear stress and internal filtration, which may affect the surface characteristics of the dialysis membranes. In the present study, we evaluated changes in the characteristics of membrane surfaces caused by shear stress and internal filtration using blood substitutes: water purified by reverse osmosis and 6.7 wt% dextran70 solution. We focused on the levels of a hydrophilic modifier, polyvinylpyrrolidone (PVP), on the membrane surface measured by attenuated total reflectance Fourier transform infrared spectroscopy. Experiments involving 4 h dialysis, 0-144 h shear-stress loading, and 4 h dead-end filtration were performed using polyester-polymer alloy (PEPA) and polysulfone (PS) membranes. After the dialysis experiments with accompanying internal filtration, average PVP retention on the PEPA membrane surface was 93.7% in all areas, whereas that on the PS membrane surface was 98.9% in all areas. After the shear-stress loading experiments, PVP retention on the PEPA membrane surface decreased as shear-stress loading time and the magnitude of shear stress increased. However, with the PS membrane, PVP retention scarcely changed. After the dead-end filtration experiments, PVP retention decreased in all areas for both PEPA and PS membranes, but PVP retention on the PEPA membrane surface was lower than that on the PS membrane surface. PVP on the PEPA membrane surface was eluted by both shear stress and internal filtration, while that on the PS membrane surface was eluted only by internal filtration. PMID:18836876

  3. Phosphate Ions Affect the Water Structure at Functionalized Membrane Surfaces.

    PubMed

    Barrett, Aliyah; Imbrogno, Joseph; Belfort, Georges; Petersen, Poul B

    2016-09-01

    Antifouling surfaces improve function, efficiency, and safety in products such as water filtration membranes, marine vehicle coatings, and medical implants by resisting protein and biofilm adhesion. Understanding the role of water structure at these materials in preventing protein adhesion and biofilm formation is critical to designing more effective coatings. Such fouling experiments are typically performed under biological conditions using isotonic aqueous buffers. Previous studies have explored the structure of pure water at a few different antifouling surfaces, but the effect of electrolytes and ionic strength (I) on the water structure at antifouling surfaces is not well studied. Here sum frequency generation (SFG) spectroscopy is used to characterize the interfacial water structure at poly(ether sulfone) (PES) and two surface-modified PES films in contact with 0.01 M phosphate buffer with high and low salt (Ionic strength, I= 0.166 and 0.025 M, respectively). Unmodified PES, commonly used as a filtration membrane, and modified PES with a hydrophobic alkane (C18) and with a poly(ethylene glycol) (PEG) were used. In the low ionic strength phosphate buffer, water was strongly ordered near the surface of the PEG-modified PES film due to exclusion of phosphate ions and the creation of a surface potential resulting from charge separation between phosphate anions and sodium cations. However, in the high ionic strength phosphate buffer, the sodium and potassium chloride (138 and 3 mM, respectively) in the phosphate buffered saline screened this charge and substantially reduced water ordering. A much smaller water ordering and subsequent reduction upon salt addition was observed for the C18-modified PES, and little water structure change was seen for the unmodified PES. The large difference in water structuring with increasing ionic strength between widely used phosphate buffer and phosphate buffered saline at the PEG interface demonstrates the importance of studying

  4. Quantitative analysis of cell surface membrane proteins using membrane-impermeable chemical probe coupled with 18O labeling

    SciTech Connect

    Zhang, Haizhen; Brown, Roslyn N.; Qian, Weijun; Monroe, Matthew E.; Purvine, Samuel O.; Moore, Ronald J.; Gritsenko, Marina A.; Shi, Liang; Romine, Margaret F.; Fredrickson, Jim K.; Pasa-Tolic, Ljiljana; Smith, Richard D.; Lipton, Mary S.

    2010-05-03

    We report a mass spectrometry-based strategy for quantitative analysis of cell surface membrane proteome changes. The strategy includes enrichment of surface membrane proteins using a membrane-impermeable chemical probe followed by stable isotope 18O labeling and LC-MS analysis. We applied this strategy for enriching membrane proteins expressed by Shewanella oneidensis MR-1, a gram-negative bacterium with known metal-reduction capability via extracellular electron transfer between outer membrane proteins and environmental electron receptors. LC/MS/MS analysis resulted in the identification of about 79% membrane proteins among all proteins identified from the enriched sample. To illustrate the quantification of membrane proteome changes, enriched membrane protein samples from wild-type and mutant cells (generated from deletion of a type II secretion protein, GspD) were further labeled with 16O and 18O at the peptide level prior to LC-MS analysis. A chemical-probe-labeled pure protein has also been used as an internal standard for normalization purpose. The quantitative data revealed reduced abundances of many outer membrane proteins such as OmcA and MtrC in ΔgspD mutant cells, which agreed well with previously published studies.

  5. Quantitative analysis of cell surface membrane proteins using membrane-impermeable chemical probe coupled with 18O labeling

    PubMed Central

    Zhang, Haizhen; Brown, Roslyn N.; Qian, Wei-Jun; Monroe, Matthew E.; Purvine, Samuel O.; Moore, Ronald J.; Gritsenko, Marina A.; Shi, Liang; Romine, Margaret F; Fredrickson, James K.; Paša-Tolić, Ljiljana; Smith, Richard D.; Lipton, Mary S.

    2010-01-01

    We report a mass spectrometry-based strategy for quantitative analysis of cell surface membrane proteome changes. The strategy includes enrichment of surface membrane proteins using a membrane-impermeable chemical probe followed by stable isotope 18O labeling and LC-MS analysis. We applied this strategy for enriching membrane proteins expressed by Shewanella oneidensis MR-1, a gram-negative bacterium with known metal-reduction capability via extracellular electron transfer between outer membrane proteins and extracellular electron receptors. LC/MS/MS analysis resulted in the identification of about 400 proteins with 79% of them being predicted to be membrane localized. Quantitative aspects of the membrane enrichment were shown by peptide level 16O and 18O labeling of proteins from wild-type and mutant cells (generated from deletion of a type II secretion protein, GspD) prior to LC-MS analysis. Using a chemical probe labeled pure protein as an internal standard for normalization, the quantitative data revealed reduced abundances in ΔgspD mutant cells of many outer membrane proteins including the outer membrane c-cype cytochromes OmcA and MtrC, in agreement with previously investigation demonstrating that these proteins are substrates of the type II secretion system. PMID:20380418

  6. Glial regulation of the axonal membrane at nodes of Ranvier.

    PubMed

    Schafer, Dorothy P; Rasband, Matthew N

    2006-10-01

    Action potential conduction in myelinated nerve fibers depends on a polarized axonal membrane. Voltage-gated Na(+) and K(+) channels are clustered at nodes of Ranvier and mediate the transmembrane currents necessary for rapid saltatory conduction. Paranodal junctions flank nodes and function as attachment sites for myelin and as paracellular and membrane protein diffusion barriers. Common molecular mechanisms, directed by myelinating glia, are used to establish these axonal membrane domains. Initially, heterophilic interactions between glial and axonal cell adhesion molecules define the locations where nodes or paranodes form. Subsequently, within each domain, axonal cell adhesion molecules are stabilized and retained through interactions with cytoskeletal and scaffolding proteins, including ankyrins and spectrins. PMID:16945520

  7. Quantitative assessment of interfacial interactions with rough membrane surface and its implications for membrane selection and fabrication in a MBR.

    PubMed

    Chen, Jianrong; Mei, Rongwu; Shen, Liguo; Ding, Linxian; He, Yiming; Lin, Hongjun; Hong, Huachang

    2015-03-01

    The interfacial interactions between a foulant particle and rough membrane surface in a submerged membrane bioreactor (MBR) were quantitatively assessed by using a new-developed method. It was found that the profile of total interaction versus separation distance was complicated. There were an energy barrier and two negative energy ranges in the profile. Further analysis showed that roughness scale significantly affected the strength and properties of interfacial interactions. It was revealed that there existed a critical range of roughness scale within which the total energy in the separation distance ranged from 0 to several nanometers was continually repulsive. Decrease in foulant size would increase the strength of specific interaction energy, but did not change the existence of a critical roughness scale range. These findings suggested the possibility to "tailor" membrane surface morphology for membrane fouling mitigation, and thus gave significant implications for membrane selection and fabrication in MBRs. PMID:25553567

  8. A Dry Membrane Protection Technique to Allow Surface Acoustic Wave Biosensor Measurements of Biological Model Membrane Approaches

    PubMed Central

    Reder-Christ, Katrin; Schmitz, Patrick; Bota, Marian; Gerber, Ursula; Falkenstein-Paul, Hildegard; Fuss, Christian; Enachescu, Marius; Bendas, Gerd

    2013-01-01

    Model membrane approaches have attracted much attention in biomedical sciences to investigate and simulate biological processes. The application of model membrane systems for biosensor measurements is partly restricted by the fact that the integrity of membranes critically depends on the maintenance of an aqueous surrounding, while various biosensors require a preconditioning of dry sensors. This is for example true for the well-established surface acoustic wave (SAW) biosensor SAM®5 blue. Here, a simple drying procedure of sensor-supported model membranes is introduced using the protective disaccharide trehalose. Highly reproducible model membranes were prepared by the Langmuir-Blodgett technique, transferred to SAW sensors and supplemented with a trehalose solution. Membrane rehydration after dry incorporation into the SAW device becomes immediately evident by phase changes. Reconstituted model membranes maintain their full functionality, as indicated by biotin/avidin binding experiments. Atomic force microscopy confirmed the morphological invariability of dried and rehydrated membranes. Approximating to more physiological recognition phenomena, the site-directed immobilization of the integrin VLA-4 into the reconstituted model membrane and subsequent VCAM-1 ligand binding with nanomolar affinity were illustrated. This simple drying procedure is a novel way to combine the model membrane generation by Langmuir-Blodgett technique with SAW biosensor measurements, which extends the applicability of SAM®5 blue in biomedical sciences. PMID:24064603

  9. Surface Membrane Glycopeptides Which Coincide with Virus Transformation and Tumorigenesis

    PubMed Central

    Glick, Mary Catherine; Rabinowitz, Zelig; Sachs, Leo

    1974-01-01

    Glycopeptides from the surface of clones of hamster embryo cells were examined at various intervals after infection with polyoma virus. Two types of transformed cells were examined: (i) clones that showed delayed transformation or an initially low tumorigenicity, and (ii) clones that were rapidly transformed showing an initially high tumorigenicity. The glycopeptides were removed from the cell surface by trypsin and, after Pronase digestion, were examined by filtration through Sephadex G-50. With delayed transformation, a specific group of glycopeptides was increasingly evident over an 85-day period as the cells showed phenotypic properties of transformation and the ability to form tumors. In the other series, all but one clone of hamster embryo cells showed rapid transformation after infection with polyoma virus. This clone was less tumorigenic and showed little of the specific glycopeptides. In all cases of delayed or rapid transformation examined, the specific group of glycopeptides increased proportionately to the ability of the cells to form tumors. All of the cells derived from progressively growing tumors formed by injection of these transformed hamster cells into adult animals showed an abundance of this group of glycopeptides. These results suggest that specific surface membrane glycopeptides accompany viral transformation and tumorigenesis. PMID:4363256

  10. Mechanisms regulating cell membrane localization of the chemokine receptor CXCR4 in human hepatocarcinoma cells.

    PubMed

    Cepeda, Edgar B; Dediulia, Tatjana; Fernando, Joan; Bertran, Esther; Egea, Gustavo; Navarro, Estanislao; Fabregat, Isabel

    2015-05-01

    Hepatocellular carcinoma (HCC) cells with a mesenchymal phenotype show an asymmetric subcellular distribution of the chemokine receptor CXCR4, which is required for cell migration and invasion. In this work we examine the mechanisms that regulate the intracellular trafficking of CXCR4 in HCC cells. Results indicate that HCC cells present CXCR4 at the cell surface, but most of this protein is in endomembranes colocalizing with markers of the Golgi apparatus and recycling endosomes. The presence of high protein levels of CXCR4 present at the cell surface correlates with a mesenchymal-like phenotype and a high autocrine activation of the Transforming Growth Factor-beta (TGF-β) pathway. CXCR4 traffics along the Golgi/exocyst/plasma membrane pathway and requires EXOC4 (Sec8) component of the exocyst complex. HCC cells use distinct mechanisms for the CXCR4 internalization such as dynamin-dependent endocytosis and macropinocytosis. Regardless of the endocytic mechanisms, colocalization of CXCR4 and Rab11 is observed, which could be involved not only in receptor recycling but also in its post-Golgi transport. In summary, this work highlights membrane trafficking pathways whose pharmacological targeting could subsequently result in the inactivation of one of the main guiding mechanisms used by metastatic cells to colonize secondary organs and tissues. PMID:25704914

  11. Optogenetic oligomerization of Rab GTPases regulates intracellular membrane trafficking.

    PubMed

    Nguyen, Mai Khanh; Kim, Cha Yeon; Kim, Jin Man; Park, Byung Ouk; Lee, Sangkyu; Park, Hyerim; Heo, Won Do

    2016-06-01

    Intracellular membrane trafficking, which is involved in diverse cellular processes, is dynamic and difficult to study in a spatiotemporal manner. Here we report an optogenetic strategy, termed light-activated reversible inhibition by assembled trap of intracellular membranes (IM-LARIAT), that uses various Rab GTPases combined with blue-light-induced hetero-interaction between cryptochrome 2 and CIB1. In this system, illumination induces a rapid and reversible intracellular membrane aggregation that disrupts the dynamics and functions of the targeted membrane. We applied IM-LARIAT to specifically perturb several Rab-mediated trafficking processes, including receptor transport, protein sorting and secretion, and signaling initiated from endosomes. We finally used this tool to reveal different functions of local Rab5-mediated and Rab11-mediated membrane trafficking in growth cones and soma of young hippocampal neurons. Our results show that IM-LARIAT is a versatile tool that can be used to dissect spatiotemporal functions of intracellular membranes in diverse systems. PMID:27065232

  12. Tuning the resistance of polycarbonate membranes by plasma-induced graft surface modification

    NASA Astrophysics Data System (ADS)

    Baumann, Lukas; Hegemann, Dirk; de Courten, Damien; Wolf, Martin; Rossi, René M.; Meier, Wolfgang P.; Scherer, Lukas J.

    2013-03-01

    To tune the permeability resistance of porous polycarbonate (PC) membranes for caffeine, their surfaces were plasma modified with different monomers in a grafting from process. These coatings provided characteristic surface hydrophilicities. It was found that membranes with more hydrophilic surfaces have lower resistances to let caffeine pass through than membranes with hydrophobic surfaces. Additionally, it was possible to post-modify a poly(2-aminoethyl methacrylate) (AEMA) coated PC membrane with octanoic acid (Oct) under mild conditions. This post modification allowed transforming a slightly hydrophilic PC-AEMA membrane with a moderate permeability resistance into a hydrophobic PC-AEMA-Oct membrane with a high permeability resistance. Overall, it was possible to tune the PC membrane resistance for caffeine in a range from 5100 up to 15,100 s/cm.

  13. Improved antifouling properties of polyamide nanofiltration membranes by reducing the density of surface carboxyl groups.

    PubMed

    Mo, Yinghui; Tiraferri, Alberto; Yip, Ngai Yin; Adout, Atar; Huang, Xia; Elimelech, Menachem

    2012-12-18

    Carboxyls are inherent functional groups of thin-film composite polyamide nanofiltration (NF) membranes, which may play a role in membrane performance and fouling. Their surface presence is attributed to incomplete reaction of acyl chloride monomers during the membrane active layer synthesis by interfacial polymerization. In order to unravel the effect of carboxyl group density on organic fouling, NF membranes were fabricated by reacting piperazine (PIP) with either isophthaloyl chloride (IPC) or the more commonly used trimesoyl chloride (TMC). Fouling experiments were conducted with alginate as a model hydrophilic organic foulant in a solution, simulating the composition of municipal secondary effluent. Improved antifouling properties were observed for the IPC membrane, which exhibited lower flux decline (40%) and significantly greater fouling reversibility or cleaning efficiency (74%) than the TMC membrane (51% flux decline and 40% cleaning efficiency). Surface characterization revealed that there was a substantial difference in the density of surface carboxyl groups between the IPC and TMC membranes, while other surface properties were comparable. The role of carboxyl groups was elucidated by measurements of foulant-surface intermolecular forces by atomic force microscopy, which showed lower adhesion forces and rupture distances for the IPC membrane compared to TMC membranes in the presence of calcium ions in solution. Our results demonstrated that a decrease in surface carboxyl group density of polyamide membranes fabricated with IPC monomers can prevent calcium bridging with alginate and, thus, improve membrane antifouling properties. PMID:23205860

  14. Mixing Water, Transducing Energy, and Shaping Membranes: Autonomously Self-Regulating Giant Vesicles.

    PubMed

    Ho, James C S; Rangamani, Padmini; Liedberg, Bo; Parikh, Atul N

    2016-03-01

    Giant lipid vesicles are topologically closed compartments bounded by semipermeable flexible shells, which isolate femto- to picoliter quantities of the aqueous core from the surrounding bulk. Although water equilibrates readily across vesicular walls (10(-2)-10(-3) cm(3) cm(-2) s(-1)), the passive permeation of solutes is strongly hindered. Furthermore, because of their large volume compressibility (∼10(9)-10(10) N m(-2)) and area expansion (10(2)-10(3) mN m(-1)) moduli, coupled with low bending rigidities (10(-19) N m), vesicular shells bend readily but resist volume compression and tolerate only a limited area expansion (∼5%). Consequently, vesicles experiencing solute concentration gradients dissipate the available chemical energy through the osmotic movement of water, producing dramatic shape transformations driven by surface-area-volume changes and sustained by the incompressibility of water and the flexible membrane interface. Upon immersion in a hypertonic bath, an increased surface-area-volume ratio promotes large-scale morphological remodeling, reducing symmetry and stabilizing unusual shapes determined, at equilibrium, by the minimal bending-energy configurations. By contrast, when subjected to a hypotonic bath, walls of giant vesicles lose their thermal undulation, accumulate mechanical tension, and, beyond a threshold swelling, exhibit remarkable oscillatory swell-burst cycles, with the latter characterized by damped, periodic oscillations in vesicle size, membrane tension, and phase behavior. This cyclical pattern of the osmotic influx of water, pressure, membrane tension, pore formation, and solute efflux suggests quasi-homeostatic self-regulatory behavior allowing vesicular compartments produced from simple molecular components, namely, water, osmolytes, and lipids, to sense and regulate their microenvironment in a negative feedback loop. PMID:26866787

  15. Investigation on surface molecular conformations and pervaporation performance of the poly(vinyl alcohol) (PVA) membrane.

    PubMed

    Zhang, Wei; Zhang, Zhennan; Wang, Xinping

    2009-05-01

    A simple method of changing pre-treatment temperature in the course of film formation was used to tune the surface structures of PVA membranes. Surface structure and property of the resulting membranes were characterized by X-ray photoelectron spectroscopy (XPS), sum frequency generation (SFG) vibrational spectroscopy, and contact angle measurements. The results show that PVA have different molecular conformations at the membrane surface while those membranes were prepared at different pre-treatment temperature. At higher pre-treatment temperatures, polar acetoxyl residues and hydroxyl groups of the PVA chains oriented in a more orderly fashion, as induced by the faster evaporation of water. When the membranes were in air, CH(3) groups adjacent to the acetoxyl groups covered the surface in order to minimize the surface free energy, while backbones of the PVA were rarely observed. These surfaces exhibited a hydrophilic nature upon contact with water due to rapid surface reconstruction. Conversely, at lower pre-treatment temperatures, the backbone CH(2) groups dominated the surface, forming a less hydrophilic surface. When the PVA membranes were employed to separate ethanol/water mixtures, it was found that the PVA membranes with more hydrophilic surface exhibited higher water selectivity. Our investigation indicates that molecular conformations on the membrane surface have considerable influence on pervaporation performance. PMID:19249794

  16. Activation of a heat-stable cytolytic protein associated with the surface membrane of Naegleria fowleri.

    PubMed Central

    Lowrey, D M; McLaughlin, J

    1985-01-01

    Surface membrane-enriched fractions of Naegleria fowleri obtained after isopycnic centrifugation experiments contain a potent cytolytic activity as determined by hemolysis and 51Cr release assays. This surface membrane cytolysin was unaffected by a treatment at 75 degrees C for 30 min and accounted for 70 to 90% of cytolysis by whole-cell lysates of amoebae. This heat resistance as well as intimate membrane association distinguished the surface membrane cytolytic activity from a second heat-labile cytolytic activity which appears to be latent within lysosomes. The surface membrane cytolysin was found to be specifically activated by diluted samples of lysosomal fractions. The possible role of this surface membrane cytotoxin in the pathogenicity of N. fowleri is discussed. PMID:4055029

  17. Surface Modification of Polypropylene Membrane by RF Methane/Oxygen Mixture Plasma Treatment

    NASA Astrophysics Data System (ADS)

    Tsai, Ching-Yuan; Juang, Ruey-Shin; Huang, Chun

    2011-08-01

    The hydrophilic surface modification of micro-porous polypropylene (PP) membranes is achieved by low-pressure 13.56 MHz RF methane (CH4)/oxygen (O2) gas mixture plasma treatment. The changes in surface wettability and surface free energy were examined by static contact angle analysis. The static water contact angle of the plasma modified membrane notably decreased with increases in treatment time and plasma power. The obvious increase in the surface energy of polypropylene membranes due to CH4/O2 mixture gas plasma treatments was also observed. Optical emission spectroscopy (OES) was used to analyze the chemical species of CH4/O2 mixture gas plasma treatment. The variations in the surface morphology and chemical structure of the micro-porous PP membranes were confirmed by confocal laser scanning microscopy (CLSM), Fourier transform infrared spectroscopy (FTIR), and X-ray photoelectron spectroscopy (XPS) measurements. XPS analysis showed significantly higher surface concentrations of oxygen functional groups for CH4/O2 mixture gas plasma-modified polypropylene membrane surfaces than for the originally unmodified polypropylene membrane surface. The experimental results show the important role of chemical species in the interaction between a CH4/O2 mixture gas plasma and a membrane surface, which can be controlled by surface modification to tailor the hydrophilicity of the membrane to the requirements of various applications.

  18. Surface roughness of cellulose hollow fiber dialysis membranes and platelet adhesion.

    PubMed

    Tsunoda, N; Kokubo, K; Sakai, K; Fukuda, M; Miyazaki, M; Hiyoshi, T

    1999-01-01

    A great deal of research has been conducted focusing on membrane materials with reference to their blood compatibility, but blood compatibility is influenced both by the material used in membranes and their structure, and by the flow conditions at the membrane surface. Accordingly, the relationship between membrane surface roughness and hemocompatibility has been evaluated using five types of membranes of differing surface roughness by evaluating the inner surfaces of the hollow fibers by atomic force microscopy (AFM) and by measuring platelet adhesion ratios using bovine blood. The yield stress, which equates to flow characteristics, was also evaluated using a glycerol suspension of polymethylmethacrylate (PMMA), a Bingham fluid. It was found that membranes having rough surfaces had high platelet adhesion ratios and poor hemocompatibility, whereas those with smoother surfaces had lower platelet adhesion ratios and better hemocompatibility. Measurement of the yield stresses for these membranes revealed higher values for those with rough surfaces, and lower values for those with smoother polyethylene glycol (PEG) grafted surfaces. This suggests that flow conditions at the membrane surface differ according to its surface roughness, and that this difference in flow conditions also influences hemocompatibility. PMID:10503618

  19. Regulation of membrane-shape transitions induced by I-BAR domains.

    PubMed

    Chen, Zhiming; Shi, Zheng; Baumgart, Tobias

    2015-07-21

    regulate membrane curvature. PMID:26200865

  20. A dimeric form of prothrombin on membrane surfaces.

    PubMed Central

    Anderson, P J

    1998-01-01

    Blood coagulation requires the conversion of zymogens to active enzymes. These reactions are facilitated by Ca2+-dependent protein binding to membrane surfaces containing anionic phospholipids. Here it is shown that only in the presence of both Ca2+ and phospholipid vesicles composed of phosphatidylcholine and phosphatidylserine can a prothrombin dimer be chemically cross-linked. A cross-linker containing evenly spaced reactive groups was prepared by activating the carboxy groups of a ten-residue glutamic acid peptide and allowed to react with physiological concentrations of prothrombin. When Ca2+ and anionic phospholipids were both present during exposure to the cross-linker, it was found that more than 50% of the prothrombin was trapped as a chemically defined dimer with reaction times of the order of 1 min. The dimer yield remained high even when reactions were performed at high phospholipid-to-protein ratios at protein concentrations an order of magnitude less than physiological. Amino acid sequencing of a CNBr peptide produced from the purified dimer localized the cross-link to residues Lys341 and Lys427 of prothrombin. The specificity and high yield under mild conditions of the cross-linking suggest that dimeric membrane bound prothrombin might be a physiologically relevant substrate for the formation of thrombin. Prothrombinase converts this modified protein to an enzyme that catalyses the hydrolysis of a thrombin chromogenic substrate as efficiently as thrombin and is inhibited by a thrombin active-site directed inhibitor, but is a thrombin dimer. The thrombin dimer has impaired activity compared with thrombin with respect to physiological functions requiring binding to exosite I. A model based on the known structure of thrombin is presented that can account for the prothrombin dimer and the properties of the dimeric thrombin formed from it. PMID:9841875

  1. Tunable Surface Hydrophobicity and Fluid Transport through Nanoporous Membranes

    NASA Astrophysics Data System (ADS)

    Ostrowski, Joseph H. J.

    There are more than three billion people across the globe that struggle to obtain clean drinkable water. One of the most promising avenues for generating potable water is through reverse osmosis and nanofiltration. Both solutions require a semipermeable membrane that prohibits passage of unwanted solute particles but allows passage of the solvent. Atomically thin two-dimensional membranes based on porous graphene show great promise as semipermeable materials, but modeling fluid flow on length scales between the microscopic (nanometer and smaller) and macroscopic (micron and larger) regimes presents formidable challenges. This thesis explores both equilibrium and nonequilibrium aspects of this problem and develops new methodology for simulating systems away from thermal equilibrium. First, we hypothesize that there is a wetting penalty for water as it tries to breach a sheet of graphene that should be naturally hydrophobic. By using equilibrium molecular dynamics simulations, we show that the hydrophobicity depends sensitively on the degree of electrical doping, offering an opportunity to tune the hydrophobic effect of graphene using small amounts of doping. The wetting contact angle, a measure of hydrophobicity, changes dramatically with the voltage applied to single layer graphene. We find that the sensitivity of the hydrophobic effect to voltage depends not on hydrogen bonding motifs at the interface between graphene and water, but instead on a phenomenon known as electrowetting. The theory of electrowetting predicts that the difference in surface tensions that defines the contact angle is quartic in the voltage, rather than quadratic, as it would be in bilayer graphene or in a two-dimensional metal. To explore the nonequilibrium aspects of fluid passage through atomically thin membranes, we developed a molecular dynamics methodology for simulating fluid flow at constant flux based on Gauss's principle of least constraint. This method develops microscopic

  2. Highly hydrophilic thin-film composite forward osmosis membranes functionalized with surface-tailored nanoparticles.

    PubMed

    Tiraferri, Alberto; Kang, Yan; Giannelis, Emmanuel P; Elimelech, Menachem

    2012-09-26

    Thin-film composite polyamide membranes are state-of-the-art materials for membrane-based water purification and desalination processes, which require both high rejection of contaminants and high water permeabilities. However, these membranes are prone to fouling when processing natural waters and wastewaters, because of the inherent surface physicochemical properties of polyamides. The present work demonstrates the fabrication of forward osmosis polyamide membranes with optimized surface properties via facile and scalable functionalization with fine-tuned nanoparticles. Silica nanoparticles are coated with superhydrophilic ligands possessing functional groups that impart stability to the nanoparticles and bind irreversibly to the native carboxyl moieties on the membrane selective layer. The tightly tethered layer of nanoparticles tailors the surface chemistry of the novel composite membrane without altering the morphology or water/solute permeabilities of the membrane selective layer. Surface characterization and interfacial energy analysis confirm that highly hydrophilic and wettable membrane surfaces are successfully attained. Lower intermolecular adhesion forces are measured between the new membrane materials and model organic foulants, indicating the presence of a bound hydration layer at the polyamide membrane surface that creates a barrier for foulant adhesion. PMID:22948042

  3. Lipid membrane domains in cell surface and vacuolar systems.

    PubMed

    Kobayashi, T; Hirabayashi, Y

    2000-01-01

    Detergent insoluble sphingolipid-cholesterol enriched 'raft'-like membrane microdomains have been implicated in a variety of biological processes including sorting, trafficking, and signaling. Mutant cells and knockout animals of sphingolipid biosynthesis are clearly useful to understand the biological roles of lipid components in raft-like domains. It is suggested that raft-like domains distribute in internal vacuolar membranes as well as plasma membranes. In addition to sphingolipid-cholesterol-rich membrane domains, recent studies suggest the existence of another lipid-membrane domain in the endocytic pathway. This domain is enriched with a unique phospholipid, lysobisphosphatidic acid (LBPA) and localized in the internal membrane of multivesicular endosome. LBPA-rich membrane domains are involved in lipid and protein sorting within the endosomal system. Possible interaction between sphingolipids and LBPA in sphingolipid-storage disease is discussed. PMID:11201787

  4. The surface charge density of plant cell membranes: an attempt to resolve conflicting values for intrinsic s.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The electrical potentials at membrane surfaces (Psi) may be computed with electrostatic models incorporating the intrinsic surface charge density of the membrane (Sigma), the ion composition of the bathing medium, and ion binding to the membrane. Ion activities at membrane surfaces may be computed ...

  5. The Ca2+-activated Cl- channel Ano1 controls microvilli length and membrane surface area in the oocyte.

    PubMed

    Courjaret, Raphael; Hodeify, Rawad; Hubrack, Satanay; Ibrahim, Awab; Dib, Maya; Daas, Sahar; Machaca, Khaled

    2016-07-01

    Ca(2+)-activated Cl(-) channels (CaCCs) play important physiological functions in epithelia and other tissues. In frog oocytes the CaCC Ano1 regulates resting membrane potential and the block to polyspermy. Here, we show that Ano1 expression increases the oocyte surface, revealing a novel function for Ano1 in regulating cell morphology. Confocal imaging shows that Ano1 increases microvilli length, which requires ERM-protein-dependent linkage to the cytoskeleton. A dominant-negative form of the ERM protein moesin precludes the Ano1-dependent increase in membrane area. Furthermore, both full-length and the truncated dominant-negative forms of moesin co-localize with Ano1 to the microvilli, and the two proteins co-immunoprecipitate. The Ano1-moesin interaction limits Ano1 lateral membrane mobility and contributes to microvilli scaffolding, therefore stabilizing larger membrane structures. Collectively, these results reveal a newly identified role for Ano1 in shaping the plasma membrane during oogenesis, with broad implications for the regulation of microvilli in epithelia. PMID:27173493

  6. Cdc42 controls the dilation of the exocytotic fusion pore by regulating membrane tension

    PubMed Central

    Bretou, Marine; Jouannot, Ouardane; Fanget, Isabelle; Pierobon, Paolo; Larochette, Nathanaël; Gestraud, Pierre; Guillon, Marc; Emiliani, Valentina; Gasman, Stéphane; Desnos, Claire; Lennon-Duménil, Ana-Maria; Darchen, François

    2014-01-01

    Membrane fusion underlies multiple processes, including exocytosis of hormones and neurotransmitters. Membrane fusion starts with the formation of a narrow fusion pore. Radial expansion of this pore completes the process and allows fast release of secretory compounds, but this step remains poorly understood. Here we show that inhibiting the expression of the small GTPase Cdc42 or preventing its activation with a dominant negative Cdc42 construct in human neuroendocrine cells impaired the release process by compromising fusion pore enlargement. Consequently the mode of vesicle exocytosis was shifted from full-collapse fusion to kiss-and-run. Remarkably, Cdc42-knockdown cells showed reduced membrane tension, and the artificial increase of membrane tension restored fusion pore enlargement. Moreover, inhibiting the motor protein myosin II by blebbistatin decreased membrane tension, as well as fusion pore dilation. We conclude that membrane tension is the driving force for fusion pore dilation and that Cdc42 is a key regulator of this force. PMID:25143404

  7. MYADM regulates Rac1 targeting to ordered membranes required for cell spreading and migration.

    PubMed

    Aranda, Juan F; Reglero-Real, Natalia; Kremer, Leonor; Marcos-Ramiro, Beatriz; Ruiz-Sáenz, Ana; Calvo, María; Enrich, Carlos; Correas, Isabel; Millán, Jaime; Alonso, Miguel A

    2011-04-15

    Membrane organization into condensed domains or rafts provides molecular platforms for selective recruitment of proteins. Cell migration is a general process that requires spatiotemporal targeting of Rac1 to membrane rafts. The protein machinery responsible for making rafts competent to recruit Rac1 remains elusive. Some members of the MAL family of proteins are involved in specialized processes dependent on this type of membrane. Because condensed membrane domains are a general feature of the plasma membrane of all mammalian cells, we hypothesized that MAL family members with ubiquitous expression and plasma membrane distribution could be involved in the organization of membranes for cell migration. We show that myeloid-associated differentiation marker (MYADM), a protein with unique features within the MAL family, colocalizes with Rac1 in membrane protrusions at the cell surface and distributes in condensed membranes. MYADM knockdown (KD) cells had altered membrane condensation and showed deficient incorporation of Rac1 to membrane raft fractions and, similar to Rac1 KD cells, exhibited reduced cell spreading and migration. Results of rescue-of-function experiments by expression of MYADM or active Rac1L61 in cells knocked down for Rac1 or MYADM, respectively, are consistent with the idea that MYADM and Rac1 act on parallel pathways that lead to similar functional outcomes. PMID:21325632

  8. Rab, Arf, and Arl-Regulated Membrane Traffic in Cortical Neuron Migration.

    PubMed

    Tang, Bor Luen

    2016-07-01

    The migration of projection neurons from its birthplace in the subventricular zone to their final destination in the cortical plate is a complex process that requires a series of highly coordinated cellular events. Amongst the key factors involved in the processes are modulators of cytoskeletal dynamics, as well as cellular membrane traffic. Members of the small GTPases family responsible for the latter process, the Rabs and Arfs, have been recently implicated in cortical neuron migration. Rab5 and Rab11, which are key modulators of endocytosis and endocytic recycling respectively, ensure proper surface expression and distribution of N-cadherin, a key adhesion protein that tethers migrating neurons to the radial glia fiber tracts during pia-directed migration. Rab7, which is associated with lysosomal biogenesis and function, is important for the final step of terminal translocation when N-cadherin is downregulated by lysosomal degradation. Arf6 activity, which is known to be important in neuronal processes outgrowth, may negatively impact the multipolar-bipolar transition of cortical neurons undergoing radial migration, but the downstream effector of Arf6 in this regard is not yet known. In addition to the above, members of the Arl family which have been recently shown to be important in radial glia scaffold formation, would also be important for cortical neuron migration. In this short review, we discuss recent advances in our understanding of the importance of membrane traffic regulated by the Rab, Arf, and Arl family members in cortical neuron migration. PMID:26587959

  9. Recycling endosome membrane incorporation into the leading edge regulates lamellipodia formation and macrophage migration.

    PubMed

    Veale, Kelly J; Offenhäuser, Carolin; Whittaker, Shane P; Estrella, Ruby P; Murray, Rachael Z

    2010-10-01

    In comparison to our knowledge of the recycling of adhesion receptors and actin assembly, exactly how the cell controls its surface membrane to form a lamellipodium during migration is poorly understood. Here, we show the recycling endosome membrane is incorporated into the leading edge of a migrating cell to expand lamellipodia membrane. We have identified the SNARE complex that is necessary for fusion of the recycling endosome with the cell surface, as consisting of the R-SNARE VAMP3 on the recycling endosome partnering with the surface Q-SNARE Stx4/SNAP23, which was found to translocate and accumulate on the leading edge of migrating cells. Increasing VAMP3-mediated fusion of the recycling endosome with the surface increased membrane ruffling, while inhibition of VAMP3-mediated fusion showed that incorporation of the recycling endosome is necessary for efficient lamellipodia formation. At the same time, insertion of this recycling endosome membrane also delivers its cargo integrin α5β1 to the cell surface. The loss of this extra membrane for lamellipodia expansion and delivery of cargo in cells resulted in macrophages with a diminished capacity to effectively migrate. Thus, the recycling endosome membrane is incorporated into the leading edge and this aids expansion of the lamellipodia and simultaneously delivers integrins necessary for efficient cell migration. PMID:20604897

  10. TspanC8 tetraspanins differentially regulate the cleavage of ADAM10 substrates, Notch activation and ADAM10 membrane compartmentalization.

    PubMed

    Jouannet, Stéphanie; Saint-Pol, Julien; Fernandez, Laurent; Nguyen, Viet; Charrin, Stéphanie; Boucheix, Claude; Brou, Christel; Milhiet, Pierre-Emmanuel; Rubinstein, Eric

    2016-05-01

    The metalloprotease ADAM10 mediates the shedding of the ectodomain of various cell membrane proteins, including APP, the precursor of the amyloid peptide Aβ, and Notch receptors following ligand binding. ADAM10 associates with the members of an evolutionary conserved subgroup of tetraspanins, referred to as TspanC8, which regulate its exit from the endoplasmic reticulum. Here we show that 4 of these TspanC8 (Tspan5, Tspan14, Tspan15 and Tspan33) which positively regulate ADAM10 surface expression levels differentially impact ADAM10-dependent Notch activation and the cleavage of several ADAM10 substrates, including APP, N-cadherin and CD44. Sucrose gradient fractionation, single molecule tracking and quantitative mass-spectrometry analysis of the repertoire of molecules co-immunoprecipitated with Tspan5, Tspan15 and ADAM10 show that these two tetraspanins differentially regulate ADAM10 membrane compartmentalization. These data represent a unique example where several tetraspanins differentially regulate the function of a common partner protein through a distinct membrane compartmentalization. PMID:26686862

  11. Regulation of a muralytic enzyme by dynamic membrane topology

    SciTech Connect

    Sun, Qingan; Kuty, Gabriel F.; Arockiasamy, Arulandu; Xu, Min; Young, Ry; Sacchettini, James C.

    2010-08-16

    R{sup 21}, the lysozyme of coliphage 21, has an N-terminal signal-anchor-release (SAR) domain that directs its secretion in a membrane-tethered, inactive form and then its release and activation in the periplasm. Both genetic and crystallographic studies show that the SAR domain, once extracted from the bilayer, refolds into the body of the enzyme and effects muralytic activation by repositioning one residue of the canonical lysozyme catalytic triad.

  12. Nonlinear conductance and heterogeneity of voltage-gated ion channels allow defining electrical surface domains in cell membranes

    NASA Astrophysics Data System (ADS)

    Cervera, Javier; Manzanares, José A.; Mafe, Salvador

    2015-07-01

    The membrane potential of a cell measured by typical electrophysiological methods is only an average magnitude and experimental techniques allowing a more detailed mapping of the cell surface have shown the existence of spatial domains with locally different electric potentials and currents. Electrical potentials in non-neural cells are regulated by the nonlinear conductance of membrane ion channels. Voltage-gated potassium channels participate in cell hyperpolarization/depolarization processes and control the electrical signals over the cell surface, constituting good candidates to study basic biological questions on a more simplified scale than the complex cell membrane. These channels show also a high heterogeneity, making it possible to analyze the effects of diversity in the electrical responses of channels localized on spatial domains. We use a phenomenological approach of voltage gating that reproduces the observed rectification characteristics of inward rectifying potassium channels and relate the threshold voltage heterogeneity of the channels to the establishment of spatial domains with different electrical sensitivities. Although our model is only a limited picture of the whole cell membrane, it shows that domains with different ion channels may permit or suppress steady state bioelectrical signals over the cell surface according to their particular voltage sensitivity. Also, the nonlinear electrical coupling of channels with different threshold potentials can lead to a rich variety of bioelectrical phenomena, including regions of membrane potential bi-stability.

  13. Membrane-Mediated Regulation of the Intrinsically Disordered CD3ϵ Cytoplasmic Tail of the TCR

    PubMed Central

    López, Cesar A.; Sethi, Anurag; Goldstein, Byron; Wilson, Bridget S.; Gnanakaran, S.

    2015-01-01

    The regulation of T-cell-mediated immune responses depends on the phosphorylation of immunoreceptor tyrosine-based activation motifs (ITAMs) on T-cell receptors. Although many details of the signaling cascades are well understood, the initial mechanism and regulation of ITAM phosphorylation remains unknown. We used molecular dynamics simulations to study the influence of different compositions of lipid bilayers on the membrane association of the CD3ϵ cytoplasmic tails of the T-cell receptors. Our results show that binding of CD3ϵ to membranes is modulated by both the presence of negatively charged lipids and the lipid order of the membrane. Free-energy calculations reveal that the protein-membrane interaction is favored by the presence of nearby basic residues and the ITAM tyrosines. Phosphorylation minimizes membrane association, rendering the ITAM motif more accessible to binding partners. In systems mimicking biological membranes, the CD3ϵ chain localization is modulated by different facilitator lipids (e.g., gangliosides or phosphoinositols), revealing a plausible regulatory effect on activation through the regulation of lipid composition in cell membranes. PMID:25992726

  14. Study of the Effect of Nanoparticles and Surface Morphology on Reverse Osmosis and Nanofiltration Membrane Productivity

    PubMed Central

    Fang, Yuming; Duranceau, Steven J.

    2013-01-01

    To evaluate the significance of reverse osmosis (RO) and nanofiltration (NF) surface morphology on membrane performance, productivity experiments were conducted using flat-sheet membranes and three different nanoparticles, which included SiO2, TiO2 and CeO2. In this study, the productivity rate was markedly influenced by membrane surface morphology. Atomic force microscopy (AFM) analysis of membrane surfaces revealed that the higher productivity decline rates associated with polyamide RO membranes as compared to that of a cellulose acetate NF membrane was due to the inherent ridge-and-valley morphology of the active layer. The unique polyamide active layer morphology was directly related to the surface roughness, and was found to contribute to particle accumulation in the valleys causing a higher flux decline than in smoother membranes. Extended RO productivity experiments using laboratory grade water and diluted pretreated seawater were conducted to compare the effect that different nanoparticles had on membrane active layers. Membrane flux decline was not affected by particle type when the feed water was laboratory grade water. On the other hand, membrane productivity was affected by particle type when pretreated diluted seawater served as feed water. It was found that CeO2 addition resulted in the least observable flux decline, followed by SiO2 and TiO2. A productivity simulation was conducted by fitting the monitored flux data into a cake growth rate model, where the model was modified using a finite difference method to incorporate surface thickness variation into the analysis. The ratio of cake growth term (k1) and particle back diffusion term (k2) was compared in between different RO and NF membranes. Results indicated that k2 was less significant for surfaces that exhibited a higher roughness. It was concluded that the valley areas of thin-film membrane surfaces have the ability to capture particles, limiting particle back diffusion. PMID:24956946

  15. Plasma Membrane H(+)-ATPase Regulation in the Center of Plant Physiology.

    PubMed

    Falhof, Janus; Pedersen, Jesper Torbøl; Fuglsang, Anja Thoe; Palmgren, Michael

    2016-03-01

    The plasma membrane (PM) H(+)-ATPase is an important ion pump in the plant cell membrane. By extruding protons from the cell and generating a membrane potential, this pump energizes the PM, which is a prerequisite for growth. Modification of the autoinhibitory terminal domains activates PM H(+)-ATPase activity, and on this basis it has been hypothesized that these regulatory termini are targets for physiological factors that activate or inhibit proton pumping. In this review, we focus on the posttranslational regulation of the PM H(+)-ATPase and place regulation of the pump in an evolutionary and physiological context. The emerging picture is that multiple signals regulating plant growth interfere with the posttranslational regulation of the PM H(+)-ATPase. PMID:26584714

  16. Surface modification of cellulose acetate membrane using thermal annealing to enhance produced water treatment

    NASA Astrophysics Data System (ADS)

    Kusworo, T. D.; Aryanti, N.; Firdaus, M. M. H.; Sukmawati, H.

    2015-12-01

    This study is performed primarily to investigate the effect of surface modification of cellulose acetate using thermal annealing on the enhancement of membrane performance for produced water treatment. In this study, Cellulose Acetate membranes were casted using dry/wet phase inversion technique. The effect of additive and post-treatment using thermal annealing on the membrane surface were examined for produced water treatment. Therma annealing was subjected to membrane surface at 60 and 70 °C for 5, 10 and 15 second, respectively. Membrane characterizations were done using membrane flux and rejection with produced water as a feed, Scanning Electron Microscopy (SEM) and Fourier Transform Infra Red (FTIR) analysis. Experimental results showed that asymmetric cellulose acetate membrane can be made by dry/wet phase inversion technique. The results from the Scanning Electron Microscopy (FESEM) analysis was also confirmed that polyethylene glycol as additivie in dope solution and thermal annealing was affected the morphology and membrane performance for produced water treatment, respectively. Scanning electron microscopy micrographs showed that the selective layer and the substructure of membrane became denser and more compact after the thermal annealing processes. Therefore, membrane rejection was significantly increased while the flux was slighty decreased, respectively. The best membrane performance is obtained on the composition of 18 wt % cellulose acetate, poly ethylene glycol 5 wt% with thermal annealing at 70° C for 15 second.

  17. Surface modification of cellulose acetate membrane using thermal annealing to enhance produced water treatment

    SciTech Connect

    Kusworo, T. D. Aryanti, N. Firdaus, M. M. H.; Sukmawati, H.

    2015-12-29

    This study is performed primarily to investigate the effect of surface modification of cellulose acetate using thermal annealing on the enhancement of membrane performance for produced water treatment. In this study, Cellulose Acetate membranes were casted using dry/wet phase inversion technique. The effect of additive and post-treatment using thermal annealing on the membrane surface were examined for produced water treatment. Therma annealing was subjected to membrane surface at 60 and 70 °C for 5, 10 and 15 second, respectively. Membrane characterizations were done using membrane flux and rejection with produced water as a feed, Scanning Electron Microscopy (SEM) and Fourier Transform Infra Red (FTIR) analysis. Experimental results showed that asymmetric cellulose acetate membrane can be made by dry/wet phase inversion technique. The results from the Scanning Electron Microscopy (FESEM) analysis was also confirmed that polyethylene glycol as additivie in dope solution and thermal annealing was affected the morphology and membrane performance for produced water treatment, respectively. Scanning electron microscopy micrographs showed that the selective layer and the substructure of membrane became denser and more compact after the thermal annealing processes. Therefore, membrane rejection was significantly increased while the flux was slighty decreased, respectively. The best membrane performance is obtained on the composition of 18 wt % cellulose acetate, poly ethylene glycol 5 wt% with thermal annealing at 70° C for 15 second.

  18. Factor Xa dimerization competes with prothrombinase complex formation on platelet-like membrane surfaces.

    PubMed

    Koklic, Tilen; Chattopadhyay, Rima; Majumder, Rinku; Lentz, Barry R

    2015-04-01

    Exposure of phosphatidylserine (PS) molecules on activated platelet membrane surface is a crucial event in blood coagulation. Binding of PS to specific sites on factor Xa (fXa) and factor Va (fVa) promotes their assembly into a complex that enhances proteolysis of prothrombin by approximately 10⁵. Recent studies demonstrate that both soluble PS and PS-containing model membranes promote formation of inactive fXa dimers at 5 mM Ca²⁺. In the present study, we show how competition between fXa dimerization and prothrombinase formation depends on Ca²⁺ and lipid membrane concentrations. We used homo-FRET measurements between fluorescein-E-G-R-chloromethylketone (CK)-Xa [fXa irreversibly inactivated by alkylation of the active site histidine residue with FEGR (FEGR-fXa)] and prothrombinase activity measurements to reveal the balance between fXa dimer formation and fXa-fVa complex formation. Changes in FEGR-fXa dimer homo-FRET with addition of fVa to model-membrane-bound FEGR-fXa unambiguously demonstrated that formation of the FEGR-fXa-fVa complex dissociated the dimer. Quantitative global analysis according to a model for protein interaction equilibria on a surface provided an estimate of a surface constant for fXa dimer dissociation (K(fXa×fXa)(d, σ)) approximately 10-fold lower than K(fXa×fVa)(d,σ) for fXa-fVa complex. Experiments performed using activated platelet-derived microparticles (MPs) showed that competition between fXa dimerization and fXa-fVa complex formation was even more prominent on MPs. In summary, at Ca²⁺ concentrations found in the maturing platelet plug (2-5 mM), fVa can compete fXa off of inactive fXa dimers to significantly amplify thrombin production, both because it releases dimer inhibition and because of its well-known cofactor activity. This suggests a hitherto unanticipated mechanism by which PS-exposing platelet membranes can regulate amplification and propagation of blood coagulation. PMID:25572019

  19. Surface patterning of polymeric separation membranes and its influence on the filtration performance

    NASA Astrophysics Data System (ADS)

    Maruf, Sajjad

    Polymeric membrane based separation technologies are crucial for addressing the global issues such as water purification. However, continuous operations of these processes are often hindered by fouling which increases mass transport resistance of the membrane to permeation and thus the energy cost, and eventually replacement of the membrane in the system. In comparison to other anti-fouling strategies, the use of controlled surface topography to mitigate fouling has not been realized mainly due to the lack of methods to create targeted topography on the porous membrane surface. This thesis aims to develop a new methodology to create surface-patterned polymeric separation membrane to improve their anti-fouling characteristics during filtration. First, successful fabrication of sub-micron surface patterns directly on a commercial ultrafiltration (UF) membrane surface using nanoimprint lithographic (NIL) technique was demonstrated. Comprehensive filtration studies revealed that the presence of these sub-micron surface patterns mitigates not only the onset of colloidal particle deposition, but also lowers the rate of growth of cake layer after initial deposition, in comparison with un-patterned membranes. The anti-fouling effects were also observed for model protein solutions. Staged filtration experiments, with backwash cleaning, revealed that the permeate flux of the patterned membrane after protein fouling was considerably higher than that of the pristine or un-patterned membrane. In addition to the surface-patterning of UF membranes, successful fabrication of a surface-patterned thin film composite (TFC) membrane was shown for the first time. A two-step fabrication process was carried out by (1) nanoimprinting a polyethersulfone (PES) support using NIL, and (2) forming a thin dense film atop the PES support via interfacial polymerization (IP). Fouling experiments suggest that the surface patterns alter the hydrodynamics at the membrane-feed interface, which is

  20. Design and simulation of the surface shape control system for membrane mirror

    NASA Astrophysics Data System (ADS)

    Zhang, Gengsheng; Tang, Minxue

    2009-11-01

    The surface shape control is one of the key technologies for the manufacture of membrane mirror. This paper presents a design of membrane mirror's surface shape control system on the basis of fuzzy logic control. The system contains such function modules as surface shape design, surface shape control, surface shape analysis, and etc. The system functions are realized by using hybrid programming technology of Visual C# and MATLAB. The finite element method is adopted to simulate the surface shape control of membrane mirror. The finite element analysis model is established through ANSYS Parametric Design Language (APDL). ANSYS software kernel is called by the system in background running mode when doing the simulation. The controller is designed by means of controlling the sag of the mirror's central crosssection. The surface shape of the membrane mirror and its optical aberration are obtained by applying Zernike polynomial fitting. The analysis of surface shape control and the simulation of disturbance response are performed for a membrane mirror with 300mm aperture and F/2.7. The result of the simulation shows that by using the designed control system, the RMS wavefront error of the mirror can reach to 142λ (λ=632.8nm), which is consistent to the surface accuracy of the membrane mirror obtained by the large deformation theory of membrane under the same condition.

  1. Temperature distribution on anodic surface of membrane electrode assembly in proton exchange membrane fuel cell with interdigitated flow bed

    NASA Astrophysics Data System (ADS)

    Guo, Hang; Wang, Mao Hai; Liu, Jia Xing; Nie, Zhi Hua; Ye, Fang; Ma, Chong Fang

    2015-01-01

    Temperature distribution on the surface of a membrane electrode assembly (MEA) significantly influences the performance, lifetime, and reliability of proton exchange membrane fuel cells (PEMFCs). Entire temperature fields on the surface of an MEA anode side under an interdigitated flow field are experimentally measured at non-humidification conditions with a self-designed PEMFC and infrared imaging technology. The highest temperature on the surface of the MEA anode side appears in the bottom bordered two side channels, and the lowest temperature exists in the area closed to the inlet of the middle channel. The hot region on the surface of the MEA anode side is easy to locate in the infrared temperature image. The reason for the temperature distribution under the interdigitated flow field is analyzed. The temperature of the MEA, the non-uniformity of temperature distribution on the surface of the MEA anode side, and the fuel cell temperature increase with the loaded current density.

  2. Highly hydrophilic polyvinylidene fluoride (PVDF) ultrafiltration membranes via postfabrication grafting of surface-tailored silica nanoparticles.

    PubMed

    Liang, Shuai; Kang, Yan; Tiraferri, Alberto; Giannelis, Emmanuel P; Huang, Xia; Elimelech, Menachem

    2013-07-24

    Polyvinylidene fluoride (PVDF) has drawn much attention as a predominant ultrafiltration (UF) membrane material due to its outstanding mechanical and physicochemical properties. However, current applications suffer from the low fouling resistance of the PVDF membrane due to the intrinsic hydrophobic property of the membrane. The present study demonstrates a novel approach for the fabrication of a highly hydrophilic PVDF UF membrane via postfabrication tethering of superhydrophilic silica nanoparticles (NPs) to the membrane surface. The pristine PVDF membrane was grafted with poly(methacrylic acid) (PMAA) by plasma induced graft copolymerization, providing sufficient carboxyl groups as anchor sites for the binding of silica NPs, which were surface-tailored with amine-terminated cationic ligands. The NP binding was achieved through a remarkably simple and effective dip-coating technique in the presence or absence of the N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (EDC)/N-hydroxysuccinimide (NHS) cross-linking process. The properties of the membrane prepared from the modification without EDC/NHS cross-linking were comparable to those for the membrane prepared with the EDC/NHS cross-linking. Both modifications almost doubled the surface energy of the functionalized membranes, which significantly improved the wettability of the membrane and converted the membrane surface from hydrophobic to highly hydrophilic. The irreversibly bound layer of superhydrophilic silica NPs endowed the membranes with strong antifouling performance as demonstrated by three sequential fouling filtration runs using bovine serum albumin (BSA) as a model organic foulant. The results suggest promising applications of the postfabrication surface modification technique in various membrane separation areas. PMID:23796125

  3. Cholinesterase activity per unit surface area of conducting membranes.

    PubMed

    Brzin, M; Dettbarn, W D; Rosenberg, P; Nachmansohn, D

    1965-08-01

    According to theory, the action of acetylcholine (ACh) and ACh-esterase is essential for the permeability changes of excitable membranes during activity. It is, therefore, pertinent to know the activity of ACh-esterase per unit axonal surface area instead of per gram nerve, as it has been measured in the past. Such information has now been obtained with the newly developed microgasometric technique using a magnetic diver. (1) The cholinesterase (Ch-esterase) activity per mm(2) surface of sensory axons of the walking leg of lobster is 1.2 x 10(-3) microM/hr. (sigma = +/- 0.3 x 10(-3); SE = 0.17 x 10(-3)); the corresponding value for the motor axons isslightly higher: 1.93 x 10(-3) microM/hr. (sigma = +/- 0.41 x 10(-3); SE = +/- 0.14 x 10(-3)). Referred to gram nerve, the Ch-esterase activity of the sensory axons is much higher than that of the motor axons: 741 microM/hr. (sigma = +/- 73.5; SE = +/- 32.6) versus 111.6 microM/hr. (sigma = +/- 28.3; SE = +/- 10). (2) The enzyme activity in the small fibers of the stellar nerve of squid is 3.2 x 10(-4) microM/mm(2)/hr. (sigma = +/- 0.96 x 10(-4); SE = +/- 0.4 x 10(-4)). (3) The Ch-esterase activity per mm(2) surface of squid giant axon is 9.5 x 10(-5) microM/hr. (sigma = +/- 1.55 x 10(-5); SE = +/- 0.38 x 10(-5)). The value was obtained with small pieces of carefully cleaned axons after removal of the axoplasm and exposure to sonic disintegration. Without the latter treatment the figurewas 3.85 x 10(-5) microM/mm(2)/hr. (sigma = +/- 3.24 x 10(-5); SE = +/- 0.93 x 10(-5)). The experiments indicate the existence of permeability barriers in the cell wall surrounding part of the enzyme, since the substrate cannot reach all the enzyme even when small fragments of the cell wall are used without disintegration. (4) On the basis of the data obtained, some tentative approximations are made of the ratio of ACh released to Na ions entering the squid giant axon per cm(2) per impulse. PMID:5865929

  4. Force Balance and Membrane Shedding at the Red-Blood-Cell Surface

    NASA Astrophysics Data System (ADS)

    Sens, Pierre; Gov, Nir

    2007-01-01

    During the aging of the red-blood cell, or under conditions of extreme echinocytosis, membrane is shed from the cell plasma membrane in the form of nanovesicles. We propose that this process is the result of the self-adaptation of the membrane surface area to the elastic stress imposed by the spectrin cytoskeleton, via the local buckling of membrane under increasing cytoskeleton stiffness. This model introduces the concept of force balance as a regulatory process at the cell membrane and quantitatively reproduces the rate of area loss in aging red-blood cells.

  5. Preparation of Light-responsive Membranes by a Combined Surface Grafting and Postmodification Process

    PubMed Central

    Schöller, Katrin; Baumann, Lukas; Hegemann, Dirk; De Courten, Damien; Wolf, Martin; Rossi, René M.; Scherer, Lukas J.

    2014-01-01

    In order to modify the surface tension of commercial available track-edged polymer membranes, a procedure of surface-initiated polymerization is presented. The polymerization from the membrane surface is induced by plasma treatment of the membrane, followed by reacting the membrane surface with a methanolic solution of 2-hydroxyethyl methacrylate (HEMA). Special attention is given to the process parameters for the plasma treatment prior to the polymerization on the surface. For example, the influence of the plasma-treatment on different types of membranes (e.g. polyester, polycarbonate, polyvinylidene fluoride) is studied. Furthermore, the time-dependent stability of the surface-grafted membranes is shown by contact angle measurements. When grafting poly(2-hydroxyethyl methacrylate) (PHEMA) in this way, the surface can be further modified by esterification of the alcohol moiety of the polymer with a carboxylic acid function of the desired substance. These reactions can therefore be used for the functionalization of the membrane surface. For example, the surface tension of the membrane can be changed or a desired functionality as the presented light-responsiveness can be inserted. This is demonstrated by reacting PHEMA with a carboxylic acid functionalized spirobenzopyran unit which leads to a light-responsive membrane. The choice of solvent plays a major role in the postmodification step and is discussed in more detail in this paper. The permeability measurements of such functionalized membranes are performed using a Franz cell with an external light source. By changing the wavelength of the light from the visible to the UV-range, a change of permeability of aqueous caffeine solutions is observed. PMID:24686898

  6. Preparation of light-responsive membranes by a combined surface grafting and postmodification process.

    PubMed

    Schöller, Katrin; Baumann, Lukas; Hegemann, Dirk; De Courten, Damien; Wolf, Martin; Rossi, René M; Scherer, Lukas J

    2014-01-01

    In order to modify the surface tension of commercial available track-edged polymer membranes, a procedure of surface-initiated polymerization is presented. The polymerization from the membrane surface is induced by plasma treatment of the membrane, followed by reacting the membrane surface with a methanolic solution of 2-hydroxyethyl methacrylate (HEMA). Special attention is given to the process parameters for the plasma treatment prior to the polymerization on the surface. For example, the influence of the plasma-treatment on different types of membranes (e.g. polyester, polycarbonate, polyvinylidene fluoride) is studied. Furthermore, the time-dependent stability of the surface-grafted membranes is shown by contact angle measurements. When grafting poly(2-hydroxyethyl methacrylate) (PHEMA) in this way, the surface can be further modified by esterification of the alcohol moiety of the polymer with a carboxylic acid function of the desired substance. These reactions can therefore be used for the functionalization of the membrane surface. For example, the surface tension of the membrane can be changed or a desired functionality as the presented light-responsiveness can be inserted. This is demonstrated by reacting PHEMA with a carboxylic acid functionalized spirobenzopyran unit which leads to a light-responsive membrane. The choice of solvent plays a major role in the postmodification step and is discussed in more detail in this paper. The permeability measurements of such functionalized membranes are performed using a Franz cell with an external light source. By changing the wavelength of the light from the visible to the UV-range, a change of permeability of aqueous caffeine solutions is observed. PMID:24686898

  7. Low-Fouling Antibacterial Reverse Osmosis Membranes via Surface Grafting of Graphene Oxide.

    PubMed

    Huang, Xinwei; Marsh, Kristofer L; McVerry, Brian T; Hoek, Eric M V; Kaner, Richard B

    2016-06-15

    Azide-functionalized graphene oxide (AGO) was covalently anchored onto commercial reverse osmosis (RO) membrane surfaces via azide photochemistry. Surface modification was carried out by coating the RO membrane with an aqueous dispersion of AGO followed by UV exposure under ambient conditions. This simple process produces a hydrophilic, smooth, antibacterial membrane with limited reduction in water permeability or salt selectivity. The GO-RO membrane exhibited a 17-fold reduction in biofouling after 24 h of Escherichia coli contact and almost 2 times reduced BSA fouling after a 1 week cross-flow test compared to its unmodified counterpart. PMID:27231843

  8. Regulation of ciliary motility by membrane potential in Paramecium: a role for cyclic AMP.

    PubMed

    Bonini, N M; Gustin, M C; Nelson, D L

    1986-01-01

    The membrane potential of Paramecium controls the frequency and direction of the ciliary beat, thus determining the cell's swimming behavior. Stimuli that hyperpolarize the membrane potential increase the ciliary beat frequency and therefore increase forward swimming speed. We have observed that 1) drugs that elevate intracellular cyclic AMP increased swimming speed 2-3-fold, 2) hyperpolarizing the membrane potential by manipulation of extracellular cations (e.g., K+) induced both a transient increase in, and a higher sustained level of cyclic AMP compared to the control, and 3) the swimming speed of detergent-permeabilized cells in MgATP was stimulated 2-fold by the addition of cyclic AMP. Our results suggest that the membrane potential can regulate intracellular cAMP in Paramecium and that control of swimming speed by membrane potential may in part be mediated by cAMP. PMID:2427226

  9. Fatty acid remodeling by LPCAT3 enriches arachidonate in phospholipid membranes and regulates triglyceride transport

    PubMed Central

    Hashidate-Yoshida, Tomomi; Harayama, Takeshi; Hishikawa, Daisuke; Morimoto, Ryo; Hamano, Fumie; Tokuoka, Suzumi M; Eto, Miki; Tamura-Nakano, Miwa; Yanobu-Takanashi, Rieko; Mukumoto, Yoshiko; Kiyonari, Hiroshi; Okamura, Tadashi; Kita, Yoshihiro; Shindou, Hideo; Shimizu, Takao

    2015-01-01

    Polyunsaturated fatty acids (PUFAs) in phospholipids affect the physical properties of membranes, but it is unclear which biological processes are influenced by their regulation. For example, the functions of membrane arachidonate that are independent of a precursor role for eicosanoid synthesis remain largely unknown. Here, we show that the lack of lysophosphatidylcholine acyltransferase 3 (LPCAT3) leads to drastic reductions in membrane arachidonate levels, and that LPCAT3-deficient mice are neonatally lethal due to an extensive triacylglycerol (TG) accumulation and dysfunction in enterocytes. We found that high levels of PUFAs in membranes enable TGs to locally cluster in high density, and that this clustering promotes efficient TG transfer. We propose a model of local arachidonate enrichment by LPCAT3 to generate a distinct pool of TG in membranes, which is required for normal directionality of TG transfer and lipoprotein assembly in the liver and enterocytes. DOI: http://dx.doi.org/10.7554/eLife.06328.001 PMID:25898003

  10. Fatty acid remodeling by LPCAT3 enriches arachidonate in phospholipid membranes and regulates triglyceride transport.

    PubMed

    Hashidate-Yoshida, Tomomi; Harayama, Takeshi; Hishikawa, Daisuke; Morimoto, Ryo; Hamano, Fumie; Tokuoka, Suzumi M; Eto, Miki; Tamura-Nakano, Miwa; Yanobu-Takanashi, Rieko; Mukumoto, Yoshiko; Kiyonari, Hiroshi; Okamura, Tadashi; Kita, Yoshihiro; Shindou, Hideo; Shimizu, Takao

    2015-01-01

    Polyunsaturated fatty acids (PUFAs) in phospholipids affect the physical properties of membranes, but it is unclear which biological processes are influenced by their regulation. For example, the functions of membrane arachidonate that are independent of a precursor role for eicosanoid synthesis remain largely unknown. Here, we show that the lack of lysophosphatidylcholine acyltransferase 3 (LPCAT3) leads to drastic reductions in membrane arachidonate levels, and that LPCAT3-deficient mice are neonatally lethal due to an extensive triacylglycerol (TG) accumulation and dysfunction in enterocytes. We found that high levels of PUFAs in membranes enable TGs to locally cluster in high density, and that this clustering promotes efficient TG transfer. We propose a model of local arachidonate enrichment by LPCAT3 to generate a distinct pool of TG in membranes, which is required for normal directionality of TG transfer and lipoprotein assembly in the liver and enterocytes. PMID:25898003

  11. Mapping of cystic fibrosis transmembrane conductance regulator membrane topology by glycosylation site insertion.

    PubMed

    Chang, X B; Hou, Y X; Jensen, T J; Riordan, J R

    1994-07-15

    Technical difficulties in obtaining three-dimensional structures of intrinsic membrane proteins continues to limit understanding of their function. However, considerable insight can be gained from their two-dimensional topological arrangement in the lipid bilayer. Efficient molecular genetic approaches are available to discern the topology of prokaryotic but not of eukaryotic membrane proteins. The absolute asymmetry of the sidedness of their N-glycosylation was employed here to develop such a method using the cystic fibrosis transmembrane conductance regulator (CFTR). Insertion by in vitro mutagenesis of N-glycosylation consensus sequences (NXS/T) in predicted cytoplasmic and extracytoplasmic loops between hydrophobic sequences capable of traversing the membrane established the membrane topology of CFTR. This provides the first experimental evaluation of the original topological model of CFTR based solely on hydropathy algorithms and a method which may be generally applicable for the in vivo evaluation of the topology of other mammalian membrane proteins. PMID:7518437

  12. Determination of surface density of nonporous membranes with air-coupled ultrasound

    NASA Astrophysics Data System (ADS)

    Lerch, T. P.

    2015-03-01

    The surface density or mass per unit area of a membrane is an important material property often used in acoustics and ultrasonics. In this paper, a new measurement and analysis technique for estimating the surface density as a function of frequency for a nonporous membrane or foil is introduced. This new, broadband technique is derived from the Thompson-Gray measurement model which can be simplified to the fluid layer transfer function commonly used in acoustics. The fluid layer transfer function can be further simplified to the limp-wall mass law for acoustically 'thin' membranes whose thickness is much less than a wavelength. The transfer function of the membrane can be efficiently measured with commercially available air-coupled ultrasonic transducers from which the surface density can be computed. Surface density estimates are presented for four membrane-like materials: aluminum foil, brass shim, polyester and polyethylene sheets.

  13. VANGL2 regulates membrane trafficking of MMP14 to control cell polarity and migration.

    PubMed

    Williams, B Blairanne; Cantrell, V Ashley; Mundell, Nathan A; Bennett, Andrea C; Quick, Rachel E; Jessen, Jason R

    2012-05-01

    Planar cell polarity (PCP) describes the polarized orientation of cells within the plane of a tissue. Unlike epithelial PCP, the mechanisms underlying PCP signaling in migrating cells remain undefined. Here, the establishment of PCP must be coordinated with dynamic changes in cell adhesion and extracellular matrix (ECM) organization. During gastrulation, the membrane type-1 matrix metalloproteinase (MT1-MMP or MMP14) is required for PCP and convergence and extension cell movements. We report that the PCP protein Vang-like 2 (VANGL2) regulates the endocytosis and cell-surface availability of MMP14 in manner that is dependent on focal adhesion kinase. We demonstrate that zebrafish trilobite/vangl2 mutant embryos exhibit increased Mmp14 activity and decreased ECM. Furthermore, in vivo knockdown of Mmp14 partially rescues the Vangl2 loss-of-function convergence and extension phenotype. This study identifies a mechanism linking VANGL2 with MMP14 trafficking and suggests that establishment of PCP in migrating gastrula cells requires regulated proteolytic degradation or remodeling of the ECM. Our findings implicate matrix metalloproteinases as downstream effectors of PCP and suggest a broadly applicable mechanism whereby VANGL2 affects diverse morphogenetic processes. PMID:22357946

  14. VANGL2 regulates membrane trafficking of MMP14 to control cell polarity and migration

    PubMed Central

    Williams, B. Blairanne; Cantrell, V. Ashley; Mundell, Nathan A.; Bennett, Andrea C.; Quick, Rachel E.; Jessen, Jason R.

    2012-01-01

    Planar cell polarity (PCP) describes the polarized orientation of cells within the plane of a tissue. Unlike epithelial PCP, the mechanisms underlying PCP signaling in migrating cells remain undefined. Here, the establishment of PCP must be coordinated with dynamic changes in cell adhesion and extracellular matrix (ECM) organization. During gastrulation, the membrane type-1 matrix metalloproteinase (MT1-MMP or MMP14) is required for PCP and convergence and extension cell movements. We report that the PCP protein Vang-like 2 (VANGL2) regulates the endocytosis and cell-surface availability of MMP14 in manner that is dependent on focal adhesion kinase. We demonstrate that zebrafish trilobite/vangl2 mutant embryos exhibit increased Mmp14 activity and decreased ECM. Furthermore, in vivo knockdown of Mmp14 partially rescues the Vangl2 loss-of-function convergence and extension phenotype. This study identifies a mechanism linking VANGL2 with MMP14 trafficking and suggests that establishment of PCP in migrating gastrula cells requires regulated proteolytic degradation or remodeling of the ECM. Our findings implicate matrix metalloproteinases as downstream effectors of PCP and suggest a broadly applicable mechanism whereby VANGL2 affects diverse morphogenetic processes. PMID:22357946

  15. Characterization of perfluorinated cation-exchange membranes MF-4SC surface modified with halloysite nanotubes

    NASA Astrophysics Data System (ADS)

    Filippov, A.; Afonin, D.; Kononenko, N.; Shkirskaya, S.

    2015-10-01

    The electrical conductivity and diffusion permeability through perfluorinated cation-exchange membranes MF-4SC (Russian analog of the Nafion-type membrane), whose surface is modified by nanotubes of halloysite using short exposures of low temperature microwave plasma, are theoretically investigated using the Nernst-Planck approach. The method of quantitative evaluation of physicochemical parameters (individual and averaged diffusion coefficients and averaged distribution coefficients of ion pairs in the membrane) of the systems `electrolyte solution - bi-layer ion-exchange membrane - water/electrolyte solution', which was proposed by us earlier, is further developed. The aforementioned parameters of modified MF-4SC/halloysite membranes were found using the least squares method. For this purpose we used electrical conductivity as well as diffusion permeability data experimentally obtained for NaCl and HCl solutions of different concentration. A new model of bi-layer membrane system can be used for refining the calculated results by taking into account the difference between co- and counter-ion diffusivities inside the membrane layers. We showed that grafting the layer of halloysite nanotubes onto the membrane surface noticeably affects the exchange capacity as well as the structural and transport characteristics of the original perfluorinated membrane. In particular, such a membrane may show weak asymmetry of diffusion permeability when its position inside a measuring cell is changed. Hybrid MF-4SC/halloysite membranes can thus be productively used in fuel cells and catalysis.

  16. Regulation of the V-ATPase along the Endocytic Pathway Occurs through Reversible Subunit Association and Membrane Localization

    PubMed Central

    Lafourcade, Céline; Sobo, Komla; Kieffer-Jaquinod, Sylvie; Garin, Jérome; van der Goot, F. Gisou

    2008-01-01

    The lumen of endosomal organelles becomes increasingly acidic when going from the cell surface to lysosomes. Luminal pH thereby regulates important processes such as the release of internalized ligands from their receptor or the activation of lysosomal enzymes. The main player in endosomal acidification is the vacuolar ATPase (V-ATPase), a multi-subunit transmembrane complex that pumps protons from the cytoplasm to the lumen of organelles, or to the outside of the cell. The active V-ATPase is composed of two multi-subunit domains, the transmembrane V0 and the cytoplasmic V1. Here we found that the ratio of membrane associated V1/Vo varies along the endocytic pathway, the relative abundance of V1 being higher on late endosomes than on early endosomes, providing an explanation for the higher acidity of late endosomes. We also found that all membrane-bound V-ATPase subunits were associated with detergent resistant membranes (DRM) isolated from late endosomes, raising the possibility that association with lipid-raft like domains also plays a role in regulating the activity of the proton pump. In support of this, we found that treatment of cells with U18666A, a drug that leads to the accumulation of cholesterol in late endosomes, affected acidification of late endosome. Altogether our findings indicate that the activity of the vATPase in the endocytic pathway is regulated both by reversible association/dissociation and the interaction with specific lipid environments. PMID:18648502

  17. Oxytocin regulates the plasma membrane Ca2+ transport in rat myometrium.

    PubMed Central

    Enyedi, A; Brandt, J; Minami, J; Penniston, J T

    1989-01-01

    Development of myometrium in young female rats was stimulated by administration of diethylstilboestrol. Plasma membrane and sarcoplasmic reticulum from rat myometrium were separated by a new and rapid method using a Percoll gradient. Calcium uptake was inhibited in plasma membrane vesicles isolated from oxytocin-treated myometrium, while no consistent effect of oxytocin was found on the Ca2+ uptake in the sarcoplasmic reticulum. Oxytocin regulated the plasma membrane Ca2+ pump by decreasing its apparent affinity for Ca2+ without affecting its maximal velocity. The K1/2 for Ca2+ in the absence of calmodulin was 0.41 +/- 0.04 microM in normal membranes; this was increased to 0.93 +/- 0.12 microM in oxytocin-treated membranes. Calmodulin decreased the K1/2 for Ca2+ to 0.27 +/- 0.027 microM and oxytocin also increased this, to 0.46 +/- 0.061 microM. The effect of oxytocin on the plasma membrane Ca2+ pump was highly dependent on the hormonal status of the animals. When the diethylstilboestrol was administered together with progesterone, the inhibitory action of oxytocin was totally suppressed, consistent with the expected action of this agent. The results suggest that regulation of the plasma membrane Ca2+ pump may be important in the prolonged elevation of intracellular Ca2+ caused by oxytocin. PMID:2775210

  18. Phosphatidylethanolamine Is a Key Regulator of Membrane Fluidity in Eukaryotic Cells.

    PubMed

    Dawaliby, Rosie; Trubbia, Cataldo; Delporte, Cédric; Noyon, Caroline; Ruysschaert, Jean-Marie; Van Antwerpen, Pierre; Govaerts, Cédric

    2016-02-12

    Adequate membrane fluidity is required for a variety of key cellular processes and in particular for proper function of membrane proteins. In most eukaryotic cells, membrane fluidity is known to be regulated by fatty acid desaturation and cholesterol, although some cells, such as insect cells, are almost devoid of sterol synthesis. We show here that insect and mammalian cells present similar microviscosity at their respective physiological temperature. To investigate how both sterols and phospholipids control fluidity homeostasis, we quantified the lipidic composition of insect SF9 and mammalian HEK 293T cells under normal or sterol-modified condition. As expected, insect cells show minimal sterols compared with mammalian cells. A major difference is also observed in phospholipid content as the ratio of phosphatidylethanolamine (PE) to phosphatidylcholine (PC) is inverted (4 times higher in SF9 cells). In vitro studies in liposomes confirm that both cholesterol and PE can increase rigidity of the bilayer, suggesting that both can be used by cells to maintain membrane fluidity. We then show that exogenously increasing the cholesterol amount in SF9 membranes leads to a significant decrease in PE:PC ratio whereas decreasing cholesterol in HEK 293T cells using statin treatment leads to an increase in the PE:PC ratio. In all cases, the membrane fluidity is maintained, indicating that both cell types combine regulation by sterols and phospholipids to control proper membrane fluidity. PMID:26663081

  19. βIV-Spectrin regulates TREK-1 membrane targeting in the heart

    PubMed Central

    Hund, Thomas J.; Snyder, Jedidiah S.; Wu, Xiangqiong; Glynn, Patric; Koval, Olha M.; Onal, Birce; Leymaster, Nicholas D.; Unudurthi, Sathya D.; Curran, Jerry; Camardo, Celia; Wright, Patrick J.; Binkley, Philip F.; Anderson, Mark E.; Mohler, Peter J.

    2014-01-01

    Aims Cardiac function depends on the highly regulated and co-ordinate activity of a large ensemble of potassium channels that control myocyte repolarization. While voltage-gated K+ channels have been well characterized in the heart, much less is known about regulation and/or targeting of two-pore K+ channel (K2P) family members, despite their potential importance in modulation of heart function. Methods and results Here, we report a novel molecular pathway for membrane targeting of TREK-1, a mechano-sensitive K2P channel regulated by environmental and physical factors including membrane stretch, pH, and polyunsaturated fatty acids (e.g. arachidonic acid). We demonstrate that βIV-spectrin, an actin-associated protein, is co-localized with TREK-1 at the myocyte intercalated disc, associates with TREK-1 in the heart, and is required for TREK-1 membrane targeting. Mice expressing βIV-spectrin lacking TREK-1 binding (qv4J) display aberrant TREK-1 membrane localization, decreased TREK-1 activity, delayed action potential repolarization, and arrhythmia without apparent defects in localization/function of other cardiac potassium channel subunits. Finally, we report abnormal βIV-spectrin levels in human heart failure. Conclusions These data provide new insight into membrane targeting of TREK-1 in the heart and establish a broader role for βIV-spectrin in organizing functional membrane domains critical for normal heart function. PMID:24445605

  20. Key diffusion mechanisms involved in regulating bidirectional water permeation across E. coli outer membrane lectin

    PubMed Central

    Sachdeva, Shivangi; Kolimi, Narendar; Nair, Sanjana Anilkumar; Rathinavelan, Thenmalarchelvi

    2016-01-01

    Capsular polysaccharides (CPSs) are major bacterial virulent determinants that facilitate host immune evasion. E. coli group1 K30CPS is noncovalently attached to bacterial surface by Wzi, a lectin. Intriguingly, structure based phylogenetic analysis indicates that Wzi falls into porin superfamily. Molecular dynamics (MD) simulations further shed light on dual role of Wzi as it also functions as a bidirectional passive water specific porin. Such a functional role of Wzi was not realized earlier, due to the occluded pore. While five water specific entry points distributed across extracellular & periplasmic faces regulate the water diffusion involving different mechanisms, a luminal hydrophobic plug governs water permeation across the channel. Coincidently, MD observed open state structure of “YQF” triad is seen in sugar-binding site of sodium-galactose cotransporters, implicating its involvement in K30CPS surface anchorage. Importance of Loop 5 (L5) in membrane insertion is yet another highlight. Change in water diffusion pattern of periplasmic substitution mutants suggests Wzi’s role in osmoregulation by aiding in K30CPS hydration, corroborating earlier functional studies. Water molecules located inside β-barrel of Wzi crystal structure further strengthens the role of Wzi in osmoregulation. Thus, interrupting water diffusion or L5 insertion may reduce bacterial virulence. PMID:27320406

  1. Membrane Type 1 Matrix Metalloproteinase Regulates Monocyte Migration and Collagen Destruction in Tuberculosis

    PubMed Central

    Sathyamoorthy, Tarangini; Tezera, Liku B.; Walker, Naomi F.; Brilha, Sara; Saraiva, Luisa; Mauri, Francesco A.; Wilkinson, Robert J.; Friedland, Jon S.

    2015-01-01

    Tuberculosis (TB) remains a global pandemic and drug resistance is rising. Multicellular granuloma formation is the pathological hallmark of Mycobacterium tuberculosis infection. The membrane type 1 matrix metalloproteinase (MT1-MMP or MMP-14) is a collagenase that is key in leukocyte migration and collagen destruction. In patients with TB, induced sputum MT1-MMP mRNA levels were increased 5.1-fold compared with matched controls and correlated positively with extent of lung infiltration on chest radiographs (r = 0.483; p < 0.05). M. tuberculosis infection of primary human monocytes increased MT1-MMP surface expression 31.7-fold and gene expression 24.5-fold. M. tuberculosis–infected monocytes degraded collagen matrix in an MT1-MMP–dependent manner, and MT1-MMP neutralization decreased collagen degradation by 73%. In human TB granulomas, MT1-MMP immunoreactivity was observed in macrophages throughout the granuloma. Monocyte–monocyte networks caused a 17.5-fold increase in MT1-MMP surface expression dependent on p38 MAPK and G protein–coupled receptor-dependent signaling. Monocytes migrating toward agarose beads impregnated with conditioned media from M. tuberculosis–infected monocytes expressed MT1-MMP. Neutralization of MT1-MMP activity decreased this M. tuberculosis network–dependent monocyte migration by 44%. Taken together, we demonstrate that MT1-MMP is central to two key elements of TB pathogenesis, causing collagen degradation and regulating monocyte migration. PMID:26091717

  2. Crystalline polymorphism induced by charge regulation in ionic membranes

    PubMed Central

    Leung, Cheuk-Yui; Palmer, Liam C.; Kewalramani, Sumit; Qiao, Baofu; Stupp, Samuel I.; Olvera de la Cruz, Monica; Bedzyk, Michael J.

    2013-01-01

    The crystallization of molecules with polar and hydrophobic groups, such as ionic amphiphiles and proteins, is of paramount importance in biology and biotechnology. By coassembling dilysine (+2) and carboxylate (–1) amphiphiles of various tail lengths into bilayer membranes at different pH values, we show that the 2D crystallization process in amphiphile membranes can be controlled by modifying the competition of long-range and short-range interactions among the polar and the hydrophobic groups. The pH and the hydrophobic tail length modify the intermolecular packing and the symmetry of their crystalline phase. For hydrophobic tail lengths of 14 carbons (C14), we observe the coassembly into crystalline bilayers with hexagonal molecular ordering via in situ small- and wide-angle X-ray scattering. As the tail length increases, the hexagonal lattice spacing decreases due to an increase in van der Waals interactions, as demonstrated by atomistic molecular dynamics simulations. For C16 and C18 we observe a reentrant crystalline phase transition sequence, hexagonal–rectangular-C–rectangular-P–rectangular-C–hexagonal, as the solution pH is increased from 3 to 10.5. The stability of the rectangular phases, which maximize tail packing, increases with increasing tail length. As a result, for very long tails (C22), the possibility of observing packing symmetries other than rectangular-C phases diminishes. Our work demonstrates that it is possible to systematically exchange chemical and mechanical energy by changing the solution pH value within a range of physiological conditions at room temperature in bilayers of molecules with ionizable groups. PMID:24065818

  3. Serotonin-induced down-regulation of cell surface serotonin transporter

    PubMed Central

    Jørgensen, Trine Nygaard; Christensen, Peter Møller; Gether, Ulrik

    2014-01-01

    The serotonin transporter (SERT) terminates serotonergic signaling and enables refilling of synaptic vesicles by mediating reuptake of serotonin (5-HT) released into the synaptic cleft. The molecular and cellular mechanisms controlling SERT activity and surface expression are not fully understood. Here we demonstrate that the substrate 5-HT itself causes acute down-regulation of SERT cell surface expression. To assess surface SERT expression by ELISA, we used a SERT variant (TacSERT) where the N-terminus of SERT was fused to the intracellular tail of the extracellularly FLAG-tagged single-membrane spanning protein Tac. In stably transfected HEK293 cells, 5-HT caused a dose-dependent reduction in TacSERT surface signal with an EC50 value equivalent to the Km value observed for 5-HT uptake. The 5-HT-induced reduction in surface signal reached maximum within 40-60 min and was blocked by the selective SERT inhibitor S-citalopram. 5-HT-induced reduction in SERT expression was further supported by surface biotinylation experiments showing 5-HT-induced reduction in wild type SERT plasma membrane levels. Moreover, preincubation with 5-HT lowered the Vmax for 5-HT uptake in cultured raphe serotonergic neurons, indicting that endogenous cell-surface resident SERT likewise is down-regulated in the presence of substrate. PMID:24462583

  4. PES Surface Modification Using Green Chemistry: New Generation of Antifouling Membranes.

    PubMed

    Nady, Norhan

    2016-01-01

    A major limitation in using membrane-based separation processes is the loss of performance due to membrane fouling. This drawback can be addressed thanks to surface modification treatments. A new and promising surface modification using green chemistry has been recently investigated. This modification is carried out at room temperature and in aqueous medium using green catalyst (enzyme) and nontoxic modifier, which can be safely labelled "green surface modification". This modification can be considered as a nucleus of new generation of antifouling membranes and surfaces. In the current research, ferulic acid modifier and laccase bio-catalyst were used to make poly(ethersulfone) (PES) membrane less vulnerable to protein adsorption. The blank and modified PES membranes are evaluated based on e.g., their flux and protein repellence. Both the blank and the modified PES membranes (or laminated PES on silicon dioxide surface) are characterized using many techniques e.g., SEM, EDX, XPS and SPM, etc. The pure water flux of the most modified membranes was reduced by 10% on average relative to the blank membrane, and around a 94% reduction in protein adsorption was determined. In the conclusions section, a comparison between three modifiers-ferulic acid, and two other previously used modifiers (4-hydroxybenzoic acid and gallic acid)-is presented. PMID:27096873

  5. PES Surface Modification Using Green Chemistry: New Generation of Antifouling Membranes

    PubMed Central

    Nady, Norhan

    2016-01-01

    A major limitation in using membrane-based separation processes is the loss of performance due to membrane fouling. This drawback can be addressed thanks to surface modification treatments. A new and promising surface modification using green chemistry has been recently investigated. This modification is carried out at room temperature and in aqueous medium using green catalyst (enzyme) and nontoxic modifier, which can be safely labelled “green surface modification”. This modification can be considered as a nucleus of new generation of antifouling membranes and surfaces. In the current research, ferulic acid modifier and laccase bio-catalyst were used to make poly(ethersulfone) (PES) membrane less vulnerable to protein adsorption. The blank and modified PES membranes are evaluated based on e.g., their flux and protein repellence. Both the blank and the modified PES membranes (or laminated PES on silicon dioxide surface) are characterized using many techniques e.g., SEM, EDX, XPS and SPM, etc. The pure water flux of the most modified membranes was reduced by 10% on average relative to the blank membrane, and around a 94% reduction in protein adsorption was determined. In the conclusions section, a comparison between three modifiers—ferulic acid, and two other previously used modifiers (4-hydroxybenzoic acid and gallic acid)—is presented. PMID:27096873

  6. Fabrication of dual-responsive cellulose-based membrane via simplified surface-initiated ATRP.

    PubMed

    Qiu, Xiaoyun; Ren, Xueqin; Hu, Shuwen

    2013-02-15

    An independently temperature- and pH-responsive membrane was developed by simultaneously grafting poly(N-isopropylacryamide) (PNIPAAm) and poly[(2-(diethylamino)ethyl methacrylate] (PDEAEMA) from different sides of a crosslinked cellulose membrane. The synthesis was simplified by using surface-initiated activators regenerated by electron transfer for atom-transfer radical polymerization in a diffusion device. The grafted membrane was heterostructured. The grafted polymer layer thickness was linearly related to reaction time. The wettabilities of the double-membrane sides responded individually and reversibly to temperature and pH. The surface grafted with PNIPAAm shifted from hydrophilicity to hydrophobicity above the lower critical solution temperature. The PDEAEMA side was hydrophilic in acidic aqueous solution and hydrophobic under basic conditions. This dual-response cellulose membrane has potential applications in water treatment, separations, and other membrane applications. PMID:23399233

  7. Snorkel: an epitope tagging system for measuring the surface expression of membrane proteins.

    PubMed

    Brown, Michael; Stafford, Lewis J; Onisk, Dale; Joaquim, Tony; Tobb, Alhagie; Goldman, Larissa; Fancy, David; Stave, James; Chambers, Ross

    2013-01-01

    Tags are widely used to monitor a protein's expression level, interactions, protein trafficking, and localization. Membrane proteins are often tagged in their extracellular domains to allow discrimination between protein in the plasma membrane from that in internal pools. Multipass membrane proteins offer special challenges for inserting a tag since the extracellular regions are often composed of small loops and thus inserting an epitope tag risks perturbing the structure, function, or location of the membrane protein. We have developed a novel tagging system called snorkel where a transmembrane domain followed by a tag is appended to the cytoplasmic C-terminus of the membrane protein. In this way the tag is displayed extracellularly, but structurally separate from the membrane protein. We have tested the snorkel tag system on a diverse panel of membrane proteins including GPCRs and ion channels and demonstrated that it reliably allows for monitoring of the surface expression. PMID:24023844

  8. Electroelastic coupling between membrane surface fluctuations and membrane-embedded charges: Continuum multidielectric treatment

    NASA Astrophysics Data System (ADS)

    Miloshevsky, Gennady V.; Hassanein, Ahmed; Partenskii, Michael B.; Jordan, Peter C.

    2010-06-01

    The coupling of electric fields and charges with membrane-water interfacial fluctuations affects membrane electroporation, ionic conductance, and voltage gating. A modified continuum model is introduced to study charge interaction with membrane-water interfacial fluctuations in multidielectric environments. By surrounding a point charge with a low dielectric sphere, the linear Poisson-Boltzmann equation is directly solved by calculating the reaction field potential via a method that eliminates singularity contributions. This allows treatment of charges located at dielectric boundaries. Two complementary mechanisms governing charge-fluctuation interactions are considered: (1) electroelastic deformation (EED), treating the membrane as an elastic slab (smectic bilayer model), and (2) electrohydrophobic solvation (EHS), accounting for water penetration into the membrane's hydrophobic core. EED often leads to large membrane thickness perturbations, far larger than those consistent with elastic model descriptions [M. B. Partenskii, G. V. Miloshevsky, and P. C. Jordan, Isr. J. Chem. 47, 385 (2007)]. We argue that a switch from EED to EHS can be energetically advantageous at intermediate perturbation amplitudes. Both perturbation mechanisms are simulated by introducing adjustable shapes optimized by the kinetic Monte Carlo reaction path following approach [G. V. Miloshevsky and P. C. Jordan, J. Chem. Phys. 122, 214901 (2005)]. The resulting energy profiles agree with those of recent atomistic molecular dynamics studies on translating a charged residue across a lipid bilayer [S. Dorairaj and T. W. Allen, Proc. Natl. Acad. Sci. U.S.A. 104, 4943 (2007)].

  9. Constructing a novel zwitterionic surface of PVDF membrane through the assembled chitosan and sodium alginate.

    PubMed

    Wang, Haiye; Zhao, Xinzhen; He, Chunju

    2016-06-01

    A novel zwitterionic surface of PVDF membrane with significantly improved antifouling properties was prepared though pressure-assisted layer by layer self-assembly method based on the electrostatic interactions of chitosan (CS), sodium alginate (SA) and polyfunctional lysine. For the modified C-S-C-S-L membrane, the contact angle decreased to 35°, the bovine serum albumin (BSA) adsorption mass of static fouling on the membrane surface decreased to 10μg/cm(2), and the secondary water flux recovery rate (FRR) of dynamic fouling of BSA and humic acid (HA) pollutants increased to 98% and 99%, respectively, exhibiting excellent antifouling performance. The results demonstrated that using charged bio-macromolecules and amino acids to build zwitterionic surface was effective and convenient to change the interface properties of the separation membrane through the pressure-assisted self-assembly modification method, and provided a new way for the industrial scale hydrophilic modification of hydrophobic porous membrane materials. PMID:26944663

  10. Cholesterol level affects surface charge of lipid membranes in saline solution

    PubMed Central

    Magarkar, Aniket; Dhawan, Vivek; Kallinteri, Paraskevi; Viitala, Tapani; Elmowafy, Mohammed; Róg, Tomasz; Bunker, Alex

    2014-01-01

    Cholesterol is an important component of all biological membranes as well as drug delivery liposomes. We show here that increasing the level of cholesterol in a phospholipid membrane decreases surface charge in the physiological environment. Through molecular dynamics simulation we have shown that increasing the level of cholesterol decreases Na+ ion binding. Complementary experimental ζ – potential measurements have shown a decreased ζ – potential with increasing cholesterol content, indicative of reduced surface charge. Both experiments and simulations have been carried out on both saturated 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC) and monounsaturated 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) membranes. This result is particularly important because membrane surface charge plays an important role in the interactions of biomembranes with peripheral membrane proteins and drug delivery liposomes with the immune system. PMID:24845659

  11. Computational redesign of the lipid-facing surface of the outer membrane protein OmpA

    PubMed Central

    Stapleton, James A.; Whitehead, Timothy A.; Nanda, Vikas

    2015-01-01

    Advances in computational design methods have made possible extensive engineering of soluble proteins, but designed β-barrel membrane proteins await improvements in our understanding of the sequence determinants of folding and stability. A subset of the amino acid residues of membrane proteins interact with the cell membrane, and the design rules that govern this lipid-facing surface are poorly understood. We applied a residue-level depth potential for β-barrel membrane proteins to the complete redesign of the lipid-facing surface of Escherichia coli OmpA. Initial designs failed to fold correctly, but reversion of a small number of mutations indicated by backcross experiments yielded designs with substitutions to up to 60% of the surface that did support folding and membrane insertion. PMID:26199411

  12. Plasma Membrane Calcium ATPase Activity Is Regulated by Actin Oligomers through Direct Interaction*

    PubMed Central

    Dalghi, Marianela G.; Fernández, Marisa M.; Ferreira-Gomes, Mariela; Mangialavori, Irene C.; Malchiodi, Emilio L.; Strehler, Emanuel E.; Rossi, Juan Pablo F. C.

    2013-01-01

    As recently described by our group, plasma membrane calcium ATPase (PMCA) activity can be regulated by the actin cytoskeleton. In this study, we characterize the interaction of purified G-actin with isolated PMCA and examine the effect of G-actin during the first polymerization steps. As measured by surface plasmon resonance, G-actin directly interacts with PMCA with an apparent 1:1 stoichiometry in the presence of Ca2+ with an apparent affinity in the micromolar range. As assessed by the photoactivatable probe 1-O-hexadecanoyl-2-O-[9-[[[2-[125I]iodo-4-(trifluoromethyl-3H-diazirin-3-yl)benzyl]oxy]carbonyl]nonanoyl]-sn-glycero-3-phosphocholine, the association of PMCA to actin produced a shift in the distribution of the conformers of the pump toward a calmodulin-activated conformation. G-actin stimulates Ca2+-ATPase activity of the enzyme when incubated under polymerizing conditions, displaying a cooperative behavior. The increase in the Ca2+-ATPase activity was related to an increase in the apparent affinity for Ca2+ and an increase in the phosphoenzyme levels at steady state. Although surface plasmon resonance experiments revealed only one binding site for G-actin, results clearly indicate that more than one molecule of G-actin was needed for a regulatory effect on the pump. Polymerization studies showed that the experimental conditions are compatible with the presence of actin in the first stages of assembly. Altogether, these observations suggest that the stimulatory effect is exerted by short oligomers of actin. The functional interaction between actin oligomers and PMCA represents a novel regulatory pathway by which the cortical actin cytoskeleton participates in the regulation of cytosolic Ca2+ homeostasis. PMID:23803603

  13. Extracellular Membrane Vesicles and Immune Regulation in the Brain

    PubMed Central

    Cossetti, Chiara; Smith, Jayden A.; Iraci, Nunzio; Leonardi, Tommaso; Alfaro-Cervello, Clara; Pluchino, Stefano

    2012-01-01

    The brain is characterized by a complex and integrated network of interacting cells in which cell-to-cell communication is critical for proper development and function. Initially considered as an immune privileged site, the brain is now regarded as an immune specialized system. Accumulating evidence reveals the presence of immune components in the brain, as well as extensive bidirectional communication that takes place between the nervous and the immune system both under homeostatic and pathological conditions. In recent years the secretion of extracellular membrane vesicles (EMVs) has been described as a new and evolutionary well-conserved mechanism of cell-to-cell communication, with EMVs influencing the microenvironment through the traffic of bioactive molecules that include proteins and nucleic acids, such as DNA, protein coding, and non-coding RNAs. Increasing evidence suggests that EMVs are a promising candidate to study cross-boundary cell-to-cell communication pathways. Herein we review the role of EMVs secreted by neural cells in modulating the immune response(s) within the brain under physiological and pathological circumstances. PMID:22557978

  14. Surface modification of PTMSP membranes by plasma treatment: Asymmetry of transport in organic solvent nanofiltration.

    PubMed

    Volkov, A V; Tsarkov, S E; Gilman, A B; Khotimsky, V S; Roldughin, V I; Volkov, V V

    2015-08-01

    For the first time, the effect of asymmetry of the membrane transport was studied for organic solvents and solutes upon their nanofiltration through the plasma-modified membranes based on poly(1-trimethylsilyl-1-propyne) (PTMSP). Plasma treatment is shown to provide a marked hydrophilization of the hydrophobic PTMSP surface (the contact angle of water decreases from 88 down to 20°) and leads to the development of a negative charge of -5.2 nC/cm(2). The XPS measurements prove the formation of the oxygen-containing groups (Si-O and C-O) due to the surface modification. The AFM images show that the small-scale surface roughness of the plasma-treated PTMSP sample is reduced but the large-scale surface heterogeneities become more pronounced. The modified membranes retain their hydrophilic surface properties even after the nanofiltration tests and 30-day storage under ambient conditions. The results of the filtration tests show that when the membrane is oriented so that its modified layer contacts the feed solution, the membrane permeability for linear alcohols (methanol-propanol) and acetone decreases nearly two times. When the modified membrane surface faces the permeate, the membrane is seen to regain its transport characteristics: the flux becomes equal to that of the unmodified PTMSP. The well-pronounced effect of the transport asymmetry is observed for the solution of the neutral dye Solvent Blue 35 in methanol, ethanol, and acetone. For example, the initial membrane shows the negative retention for the Solvent Blue 35 dye (-16%) upon its filtration from the ethanol solution whereas, for the modified PTMSP membrane, the retention increases up to 17%. Various effects contributing to the asymmetry of the membrane transport characteristics are discussed. PMID:25482845

  15. Airfoil-shaped micro-mixers for reducing fouling on membrane surfaces

    SciTech Connect

    Ho, Clifford K; Altman, Susan J; Clem, Paul G; Hibbs, Michael; Cook, Adam W

    2012-10-23

    An array of airfoil-shaped micro-mixers that enhances fluid mixing within permeable membrane channels, such as used in reverse-osmosis filtration units, while minimizing additional pressure drop. The enhanced mixing reduces fouling of the membrane surfaces. The airfoil-shaped micro-mixer can also be coated with or comprised of biofouling-resistant (biocidal/germicidal) ingredients.

  16. Performance improvement of passive direct methanol fuel cells with surface-patterned Nafion® membranes

    NASA Astrophysics Data System (ADS)

    Pu, Longjuan; Jiang, Jingjing; Yuan, Ting; Chai, Jieshi; Zhang, Haifeng; Zou, Zhiqing; Li, Xue-Mei; Yang, Hui

    2015-02-01

    Nafion® 115 membrane, patterned by thermal imprint lithography on the anode side, is used for passive direct methanol fuel cells (DMFCs). The membrane roughness factor, defined as the ratio between the actual and projected membrane surface area, was investigated for its effects on the performance of the DMFCs. When the anode Pt-Ru (1:1) catalyst loading is 1.0 mg cm-2, the maximum power density of the DMFC with a surface-patterned membrane (roughness factor: 5.4) using 3.0 M methanol as the fuel at 25 ± 1 °C reaches 27.2 ± 0.3 mW cm-2, an increase of ∼57.2% in comparison to DMFC using the pristine membrane (roughness factor: ∼1.0). Further, electrochemical characterization indicates that increased roughness factor of the membrane results in increased electrochemically active surface area and reduced charge transfer resistance in the cell. These performance improvements are ascribed to the increased surface roughness which enlarges the membrane/catalyst interface, possibly facilitating mass transport of the fuel and improving anode catalyst utilization. Thus, patterned membranes have great potential in improving the performance of fuel cells and reducing catalyst loading.

  17. SEPARATION PROPERTIES OF SURFACE MODIFIED SILICA SUPPORTED LIQUID MEMBRANES FOR DIVALENT METAL REMOVAL/RECOVERY

    EPA Science Inventory

    The synthesis and separation properties of a mesoporous silica supported liquid membrane (SLM) were studied. The membranes consisted of a silica layer, from dip-coated colloidal silica, on a a-alumina support, modified with DCDMS (dichlorodimethyl silane) to add surface methyl g...

  18. Surface pH controls purple-to-blue transition of bacteriorhodopsin. A theoretical model of purple membrane surface.

    PubMed Central

    Szundi, I; Stoeckenius, W

    1989-01-01

    We have developed a surface model of purple membrane and applied it in an analysis of the purple-to-blue color change of bacteriorhodopsin which is induced by acidification or deionization. The model is based on dissociation and double layer theory and the known membrane structure. We calculated surface pH, ion concentrations, charge density, and potential as a function of bulk pH and concentration of mono- and divalent cations. At low salt concentrations, the surface pH is significantly lower than the bulk pH and it becomes independent of bulk pH in the deionized membrane suspension. Using an experimental acid titration curve for neutral, lipid-depleted membrane, we converted surface pH into absorption values. The calculated bacteriohodopsin color changes for acidification of purple, and titrations of deionized blue membrane with cations or base agree well with experimental results. No chemical binding is required to reproduce the experimental curves. Surface charge and potential changes in acid, base and cation titrations are calculated and their relation to the color change is discussed. Consistent with structural data, 10 primary phosphate and two basic surface groups per bacteriorhodopsin are sufficient to obtain good agreement between all calculated and experimental curves. The results provide a theoretical basis for our earlier conclusion that the purple-to-blue transition must be attributed to surface phenomena and not to cation binding at specific sites in the protein. Images FIGURE 1 PMID:2775832

  19. Differential Regulation of Clathrin and Its Adaptor Proteins during Membrane Recruitment for Endocytosis.

    PubMed

    Wang, Chao; Hu, Tianwei; Yan, Xu; Meng, Tingting; Wang, Yutong; Wang, Qingmei; Zhang, Xiaoyue; Gu, Ying; Sánchez-Rodríguez, Clara; Gadeyne, Astrid; Lin, Jinxing; Persson, Staffan; Van Damme, Daniël; Li, Chuanyou; Bednarek, Sebastian Y; Pan, Jianwei

    2016-05-01

    In plants, clathrin-mediated endocytosis (CME) is dependent on the function of clathrin and its accessory heterooligomeric adaptor protein complexes, ADAPTOR PROTEIN2 (AP-2) and the TPLATE complex (TPC), and is negatively regulated by the hormones auxin and salicylic acid (SA). The details for how clathrin and its adaptor complexes are recruited to the plasma membrane (PM) to regulate CME, however, are poorly understood. We found that SA and the pharmacological CME inhibitor tyrphostin A23 reduce the membrane association of clathrin and AP-2, but not that of the TPC, whereas auxin solely affected clathrin membrane association, in Arabidopsis (Arabidopsis thaliana). Genetic and pharmacological experiments revealed that loss of AP2μ or AP2σ partially affected the membrane association of other AP-2 subunits and that the AP-2 subunit AP2σ, but not AP2μ, was required for SA- and tyrphostin A23-dependent inhibition of CME Furthermore, we show that although AP-2 and the TPC are both required for the PM recruitment of clathrin in wild-type cells, the TPC is necessary for clathrin PM association in AP-2-deficient cells. These results indicate that developmental signals may differentially modulate the membrane recruitment of clathrin and its core accessory complexes to regulate the process of CME in plant cells. PMID:26945051

  20. Differential Regulation of Clathrin and Its Adaptor Proteins during Membrane Recruitment for Endocytosis1[OPEN

    PubMed Central

    Wang, Chao; Hu, Tianwei; Yan, Xu; Meng, Tingting; Wang, Yutong; Wang, Qingmei; Zhang, Xiaoyue; Gu, Ying; Sánchez-Rodríguez, Clara; Gadeyne, Astrid; Lin, Jinxing

    2016-01-01

    In plants, clathrin-mediated endocytosis (CME) is dependent on the function of clathrin and its accessory heterooligomeric adaptor protein complexes, ADAPTOR PROTEIN2 (AP-2) and the TPLATE complex (TPC), and is negatively regulated by the hormones auxin and salicylic acid (SA). The details for how clathrin and its adaptor complexes are recruited to the plasma membrane (PM) to regulate CME, however, are poorly understood. We found that SA and the pharmacological CME inhibitor tyrphostin A23 reduce the membrane association of clathrin and AP-2, but not that of the TPC, whereas auxin solely affected clathrin membrane association, in Arabidopsis (Arabidopsis thaliana). Genetic and pharmacological experiments revealed that loss of AP2μ or AP2σ partially affected the membrane association of other AP-2 subunits and that the AP-2 subunit AP2σ, but not AP2μ, was required for SA- and tyrphostin A23-dependent inhibition of CME. Furthermore, we show that although AP-2 and the TPC are both required for the PM recruitment of clathrin in wild-type cells, the TPC is necessary for clathrin PM association in AP-2-deficient cells. These results indicate that developmental signals may differentially modulate the membrane recruitment of clathrin and its core accessory complexes to regulate the process of CME in plant cells. PMID:26945051

  1. Electroelastic coupling between membrane surface fluctuations and membrane-embedded charges: Continuum multidielectric treatment

    PubMed Central

    Miloshevsky, Gennady V.; Hassanein, Ahmed; Partenskii, Michael B.; Jordan, Peter C.

    2010-01-01

    The coupling of electric fields and charges with membrane-water interfacial fluctuations affects membrane electroporation, ionic conductance, and voltage gating. A modified continuum model is introduced to study charge interaction with membrane-water interfacial fluctuations in multidielectric environments. By surrounding a point charge with a low dielectric sphere, the linear Poisson–Boltzmann equation is directly solved by calculating the reaction field potential via a method that eliminates singularity contributions. This allows treatment of charges located at dielectric boundaries. Two complementary mechanisms governing charge-fluctuation interactions are considered: (1) electroelastic deformation (EED), treating the membrane as an elastic slab (smectic bilayer model), and (2) electrohydrophobic solvation (EHS), accounting for water penetration into the membrane’s hydrophobic core. EED often leads to large membrane thickness perturbations, far larger than those consistent with elastic model descriptions [M. B. Partenskii, G. V. Miloshevsky, and P. C. Jordan, Isr. J. Chem. 47, 385 (2007)]. We argue that a switch from EED to EHS can be energetically advantageous at intermediate perturbation amplitudes. Both perturbation mechanisms are simulated by introducing adjustable shapes optimized by the kinetic Monte Carlo reaction path following approach [G. V. Miloshevsky and P. C. Jordan, J. Chem. Phys. 122, 214901 (2005)]. The resulting energy profiles agree with those of recent atomistic molecular dynamics studies on translating a charged residue across a lipid bilayer [S. Dorairaj and T. W. Allen, Proc. Natl. Acad. Sci. U.S.A. 104, 4943 (2007)]. PMID:20572734

  2. Modification of polysulfone membranes via surface-initiated atom transfer radical polymerization

    NASA Astrophysics Data System (ADS)

    Dong, Han-Bang; Xu, You-Yi; Yi, Zhuan; Shi, Jun-Li

    2009-08-01

    Hydrophilic poly((poly(ethylene glycol) methyl ether methacrylate) (P(PEGMA)) and poly(glycidylmethacrylate) (PGMA) brushes were grafted from chloromethylated polysulfone (CMPSF) membrane surfaces via surface-initiated atom transfer radical polymerization (ATRP). Prior to ATRP, chloromethylation of PSF was performed beforehand and the obtained CMPSF was prepared into porous membranes by phase inversion process. It was demonstrated that the benzyl chloride groups on the CMPSF membrane surface afforded effective macroinitiators to graft the well-defined polymer brushes. 1H NMR was employed to confirm the structure of CMPSF. The grafting yield of P(PEGMA) and PGMA was determined by weight gain measurement. Attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy and X-ray photoelectron spectroscopy (XPS) confirmed the grafting of P(PEGMA) and PGMA chains. Water contact angle measurements indicated that the introduction of P(PEGMA) and PGMA graft chains promoted remarkably the surface hydrophilicity of PSF membranes. The effects of P(PEGMA) and PGMA immobilization on membrane morphology, permeability and fouling resistance were investigated. It was found that P(PEGMA) and PGMA grafts brought higher pure water flux, improved hydrophilic surface and better anti-protein absorption ability to PSF membranes after modification. And evidently, macromonomer P(PEGMA) brought much better properties to the PSF membranes than PGMA macromonomer.

  3. Regulation of developmental and environmental signaling by interaction between microtubules and membranes in plant cells.

    PubMed

    Zhang, Qun; Zhang, Wenhua

    2016-02-01

    Cell division and expansion require the ordered arrangement of microtubules, which are subject to spatial and temporal modifications by developmental and environmental factors. Understanding how signals translate to changes in cortical microtubule organization is of fundamental importance. A defining feature of the cortical microtubule array is its association with the plasma membrane; modules of the plasma membrane are thought to play important roles in the mediation of microtubule organization. In this review, we highlight advances in research on the regulation of cortical microtubule organization by membrane-associated and membrane-tethered proteins and lipids in response to phytohormones and stress. The transmembrane kinase receptor Rho-like guanosine triphosphatase, phospholipase D, phosphatidic acid, and phosphoinositides are discussed with a focus on their roles in microtubule organization. PMID:26687389

  4. The ubiquitin ligase HACE1 regulates Golgi membrane dynamics during the cell cycle

    PubMed Central

    Tang, Danming; Xiang, Yi; De Renzis, Stefano; Rink, Jochen; Zheng, Gen; Zerial, Marino; Wang, Yanzhuang

    2012-01-01

    Partitioning of the Golgi membrane into daughter cells during mammalian cell division occurs through a unique disassembly and reassembly process that is regulated by ubiquitination. However, the identity of the ubiquitin ligase is unknown. Here we show that the Homologous to the E6-AP Carboxyl Terminus (HECT) domain containing ubiquitin ligase HACE1 is targeted to the Golgi membrane through interactions with Rab proteins. The ubiquitin ligase activity of HACE1 in mitotic Golgi disassembly is required for subsequent postmitotic Golgi membrane fusion. Depletion of HACE1 using small interfering RNAs or expression of an inactive HACE1 mutant protein in cells impaired postmitotic Golgi membrane fusion. The identification of HACE1 as a Golgi-localized ubiquitin ligase provides evidence that ubiquitin has a critical role in Golgi biogenesis during the cell cycle. PMID:21988917

  5. Cell Surface and Membrane Engineering: Emerging Technologies and Applications

    PubMed Central

    Saeui, Christopher T.; Mathew, Mohit P.; Liu, Lingshui; Urias, Esteban; Yarema, Kevin J.

    2015-01-01

    Membranes constitute the interface between the basic unit of life—a single cell—and the outside environment and thus in many ways comprise the ultimate “functional biomaterial”. To perform the many and often conflicting functions required in this role, for example to partition intracellular contents from the outside environment while maintaining rapid intake of nutrients and efflux of waste products, biological membranes have evolved tremendous complexity and versatility. This article describes how membranes, mainly in the context of living cells, are increasingly being manipulated for practical purposes with drug discovery, biofuels, and biosensors providing specific, illustrative examples. Attention is also given to biology-inspired, but completely synthetic, membrane-based technologies that are being enabled by emerging methods such as bio-3D printers. The diverse set of applications covered in this article are intended to illustrate how these versatile technologies—as they rapidly mature—hold tremendous promise to benefit human health in numerous ways ranging from the development of new medicines to sensitive and cost-effective environmental monitoring for pathogens and pollutants to replacing hydrocarbon-based fossil fuels. PMID:26096148

  6. Peptides at Membrane Surfaces and their Role in Prebiotic Evolution

    NASA Technical Reports Server (NTRS)

    Pohorille, Andrew; Wilson, Michael A.; Chipot, Christophe; Fonda, Mark (Technical Monitor)

    2002-01-01

    Protocells had to transport ions and organic matter across membranes separating the interior of the cell from the environment, capture and utilize energy and transduce environmental signals. In a series of detailed, molecular-level computer simulations we show how these peptides in contact with membranes can acquire ordered structures and functions. We have investigated the stability of a simple alpha-helical peptide containing Leucine (L) and Serine (S) of the form (LSLLLSL)3 in a model membrane system. The parallel in-plane state is the most stable configuration. The transmembrane state is metastable, and about 15 kcal/mol is required to insert the peptide into the membrane. We investigated dimes of both (LSLLLSL)3 and glycophorin A, and show how the free energy of helix association can, at least partially, offset the free energy of insertion. We have also investigated the transmembrane pore of the influenza M2 protein. This aggregate of four identical alpha-helices, each built of 25 amino acids, forms an efficient and selective voltage-gated proton channel. Our simulations explain the gating mechanism, which can involve strands of hydrogen-bonded water through the pore or proton transfer through tautomerization of protein residues. The channel can be re-engineered to act as a simple proton pump.

  7. Plasma Membrane Abundance of Human Aquaporin 5 Is Dynamically Regulated by Multiple Pathways.

    PubMed

    Kitchen, Philip; Öberg, Fredrik; Sjöhamn, Jennie; Hedfalk, Kristina; Bill, Roslyn M; Conner, Alex C; Conner, Matthew T; Törnroth-Horsefield, Susanna

    2015-01-01

    Aquaporin membrane protein channels mediate cellular water flow. Human aquaporin 5 (AQP5) is highly expressed in the respiratory system and secretory glands where it facilitates the osmotically-driven generation of pulmonary secretions, saliva, sweat and tears. Dysfunctional trafficking of AQP5 has been implicated in several human disease states, including Sjögren's syndrome, bronchitis and cystic fibrosis. In order to investigate how the plasma membrane expression levels of AQP5 are regulated, we studied real-time translocation of GFP-tagged AQP5 in HEK293 cells. We show that AQP5 plasma membrane abundance in transfected HEK293 cells is rapidly and reversibly regulated by at least three independent mechanisms involving phosphorylation at Ser156, protein kinase A activity and extracellular tonicity. The crystal structure of a Ser156 phosphomimetic mutant indicates that its involvement in regulating AQP5 membrane abundance is not mediated by a conformational change of the carboxy-terminus. We suggest that together these pathways regulate cellular water flow. PMID:26569106

  8. Thermal Regulation of Membrane Lipid Fluidity by a Two-Component System in "Bacillus Subtilis"

    ERIC Educational Resources Information Center

    Bredeston, L. M.; Marciano, D.; Albanesi, D.; De Mendoza, D.; Delfino, J. M.

    2011-01-01

    This article describes a simple and robust laboratory exercise on the regulation of membrane unsaturated fatty acid composition in bacteria by a decrease in growth temperature. We take advantage of the well characterized Des pathway of "Bacillus subtilis", composed of a [delta]5-desaturase (encoded by the "des" gene) and the canonical…

  9. Plasma Membrane Abundance of Human Aquaporin 5 Is Dynamically Regulated by Multiple Pathways

    PubMed Central

    Kitchen, Philip; Öberg, Fredrik; Sjöhamn, Jennie; Hedfalk, Kristina; Bill, Roslyn M.; Conner, Alex C.; Conner, Matthew T.; Törnroth-Horsefield, Susanna

    2015-01-01

    Aquaporin membrane protein channels mediate cellular water flow. Human aquaporin 5 (AQP5) is highly expressed in the respiratory system and secretory glands where it facilitates the osmotically-driven generation of pulmonary secretions, saliva, sweat and tears. Dysfunctional trafficking of AQP5 has been implicated in several human disease states, including Sjögren’s syndrome, bronchitis and cystic fibrosis. In order to investigate how the plasma membrane expression levels of AQP5 are regulated, we studied real-time translocation of GFP-tagged AQP5 in HEK293 cells. We show that AQP5 plasma membrane abundance in transfected HEK293 cells is rapidly and reversibly regulated by at least three independent mechanisms involving phosphorylation at Ser156, protein kinase A activity and extracellular tonicity. The crystal structure of a Ser156 phosphomimetic mutant indicates that its involvement in regulating AQP5 membrane abundance is not mediated by a conformational change of the carboxy-terminus. We suggest that together these pathways regulate cellular water flow. PMID:26569106

  10. Human cell receptor CD46 is down regulated through recognition of a membrane-proximal region of the cytoplasmic domain in persistent measles virus infection.

    PubMed Central

    Hirano, A; Yant, S; Iwata, K; Korte-Sarfaty, J; Seya, T; Nagasawa, S; Wong, T C

    1996-01-01

    Monkey cells persistently infected by measles virus (MV) Biken strain (Biken-CV-1 cells) showed no cytopathic effects and lacked surface expression of a homolog of human cell receptor, membrane cofactor protein CD46. Transfection of a human CD46 gene into these cells induced extensive cell fusion, indicating that down regulation of the endogenous CD46 homolog was essential for the maintenance of a noncytopathic mode of infection. Surface expression of the exogenously introduced human CD46 was also drastically down regulated in the persistently infected cells compared with uninfected cells. The down regulation was specific for CD46 and did not affect surface expression of exogenously introduced CD4. Exogenous human CD46 was synthesized efficiently in the persistently infected cells, but it did not accumulate on the cell surface. Fusion of Biken-CV-1 cells required the extracellular hemagglutinin (H-protein)-binding domain but not the cytoplasmic domain. Replacing the transmembrane and cytoplasmic domains of CD46 with a glycosylphosphatidylinositol anchor did not prevent cell fusion but completely alleviated down regulation of the glycosylphosphatidylinositol-anchored CD46 in Biken-CV-1 cells. Deletion analyses revealed that the membrane-distal sequences of the CD46 cytoplasmic domain were not only unnecessary but also inhibitory for CD46 down regulation. By contrast, the six amino acid residues proximal to the membrane contained a sequence required for CD46 down regulation in the persistently infected cells. These results indicate that CD46 is down regulated in the persistently infected cells by a mechanism that recognizes a membrane-proximal sequence in the CD46 cytoplasmic domain. PMID:8794336

  11. Surface Segregation in a PdCu Alloy Hydrogen Separation Membrane

    SciTech Connect

    Miller, J.B.; Matranga, C.S.; Gellman, A.J.

    2007-06-01

    Separation of hydrogen from mixed gas streams is an important step for hydrogen generation technologies, including hydrocarbon reforming and coal/biomass gasification. Dense palladium-based membranes have received significant attention for this application because of palladium’s ability to dissociatively adsorb molecular hydrogen at its surface for subsequent transport of hydrogen atoms through its bulk. Alloying palladium with minor components, like copper, has been shown to improve both the membrane’s structural characteristics and resistance to poisoning of its catalytic surface [1]. Surface segregation—a composition difference between the bulk material and its surface—is common in alloys and can affect important surface processes. Rational design of alloy membranes requires that surface segregation be understood, and possibly controlled. In this work, we examine surface segregation in a polycrystalline Pd70Cu30 hydrogen separation membrane as a function of thermal treatment and adsorption of hydrogen sulfide.

  12. Probing the dynamic regulation of peripheral membrane proteins using hydrogen deuterium exchange-MS (HDX-MS).

    PubMed

    Vadas, Oscar; Burke, John E

    2015-10-01

    Many cellular signalling events are controlled by the selective recruitment of protein complexes to membranes. Determining the molecular basis for how lipid signalling complexes are recruited, assembled and regulated on specific membrane compartments has remained challenging due to the difficulty of working in conditions mimicking native biological membrane environments. Enzyme recruitment to membranes is controlled by a variety of regulatory mechanisms, including binding to specific lipid species, protein-protein interactions, membrane curvature, as well as post-translational modifications. A powerful tool to study the regulation of membrane signalling enzymes and complexes is hydrogen deuterium exchange-MS (HDX-MS), a technique that allows for the interrogation of protein dynamics upon membrane binding and recruitment. This review will highlight the theory and development of HDX-MS and its application to examine the molecular basis of lipid signalling enzymes, specifically the regulation and activation of phosphoinositide 3-kinases (PI3Ks). PMID:26517882

  13. A taste sensor based on surface imprinted TiO2 membrane

    NASA Astrophysics Data System (ADS)

    Xiao, Wenxiang; Chen, Zhencheng; Jiang, Xingguo; Zhao, Hongtian; Chu, Fugang; Hou, Hongbin

    2012-03-01

    Surface imprinted TiO2 membranes had been prepared and used as sensing membranes for basic tastes discrimination. Four basic taste molecules (citric acid, D-glucose, quinine hydrochloride and sodium L-glutamate for sour, sweet, bitter and umami respectively) were used as templates for imprinting. The sensor was fabricated in light-addressable potentiometric principle. Experimental results show that membranes imprinted by citric acid and quinine hydrochloride exhibit similar response behaviors towards four taste substances, that is citric acid > quinine hydrochloride > sodium L-glutamate > D-glucose. Membrane imprinted by sodium L-glutamate is sensitive towards quinine hydrochloride. Except for D-glucose imprinting membrane, other three membranes are inert to glucose. Combined with principal component analysis, four basic tastes can be well distinguished.

  14. A taste sensor based on surface imprinted TiO2 membrane

    NASA Astrophysics Data System (ADS)

    Xiao, Wenxiang; Chen, Zhencheng; Jiang, Xingguo; Zhao, Hongtian; Chu, Fugang; Hou, Hongbin

    2011-11-01

    Surface imprinted TiO2 membranes had been prepared and used as sensing membranes for basic tastes discrimination. Four basic taste molecules (citric acid, D-glucose, quinine hydrochloride and sodium L-glutamate for sour, sweet, bitter and umami respectively) were used as templates for imprinting. The sensor was fabricated in light-addressable potentiometric principle. Experimental results show that membranes imprinted by citric acid and quinine hydrochloride exhibit similar response behaviors towards four taste substances, that is citric acid > quinine hydrochloride > sodium L-glutamate > D-glucose. Membrane imprinted by sodium L-glutamate is sensitive towards quinine hydrochloride. Except for D-glucose imprinting membrane, other three membranes are inert to glucose. Combined with principal component analysis, four basic tastes can be well distinguished.

  15. Functionality in Electrospun Nanofibrous Membranes Based on Fiber's Size, Surface Area, and Molecular Orientation

    PubMed Central

    Matsumoto, Hidetoshi; Tanioka, Akihiko

    2011-01-01

    Electrospinning is a versatile method for forming continuous thin fibers based on an electrohydrodynamic process. This method has the following advantages: (i) the ability to produce thin fibers with diameters in the micrometer and nanometer ranges; (ii) one-step forming of the two- or three-dimensional nanofiber network assemblies (nanofibrous membranes); and (iii) applicability for a broad spectrum of molecules, such as synthetic and biological polymers and polymerless sol-gel systems. Electrospun nanofibrous membranes have received significant attention in terms of their practical applications. The major advantages of nanofibers or nanofibrous membranes are the functionalities based on their nanoscaled-size, highly specific surface area, and highly molecular orientation. These functionalities of the nanofibrous membranes can be controlled by their fiber diameter, surface chemistry and topology, and internal structure of the nanofibers. This report focuses on our studies and describes fundamental aspects and applications of electrospun nanofibrous membranes. PMID:24957735

  16. Cell Surface Area and Membrane Folding in Glioblastoma Cell Lines Differing in PTEN and p53 Status

    PubMed Central

    Höring, Marcus; Westerling, Katherine; Fiedler, Vanessa; Katzer, Astrid; Krohne, Georg; Flentje, Michael; Djuzenova, Cholpon S.

    2014-01-01

    Glioblastoma multiforme (GBM) is characterized by rapid growth, invasion and resistance to chemo−/radiotherapy. The complex cell surface morphology with abundant membrane folds, microvilli, filopodia and other membrane extensions is believed to contribute to the highly invasive behavior and therapy resistance of GBM cells. The present study addresses the mechanisms leading to the excessive cell membrane area in five GBM lines differing in mutational status for PTEN and p53. In addition to scanning electron microscopy (SEM), the membrane area and folding were quantified by dielectric measurements of membrane capacitance using the single-cell electrorotation (ROT) technique. The osmotic stability and volume regulation of GBM cells were analyzed by video microscopy. The expression of PTEN, p53, mTOR and several other marker proteins involved in cell growth and membrane synthesis were examined by Western blotting. The combined SEM, ROT and osmotic data provided independent lines of evidence for a large variability in membrane area and folding among tested GBM lines. Thus, DK-MG cells (wild type p53 and wild type PTEN) exhibited the lowest degree of membrane folding, probed by the area-specific capacitance Cm = 1.9 µF/cm2. In contrast, cell lines carrying mutations in both p53 and PTEN (U373-MG and SNB19) showed the highest Cm values of 3.7–4.0 µF/cm2, which corroborate well with their heavily villated cell surface revealed by SEM. Since PTEN and p53 are well-known inhibitors of mTOR, the increased membrane area/folding in mutant GBM lines may be related to the enhanced protein and lipid synthesis due to a deregulation of the mTOR-dependent downstream signaling pathway. Given that membrane folds and extensions are implicated in tumor cell motility and metastasis, the dielectric approach presented here provides a rapid and simple tool for screening the biophysical cell properties in studies on targeting chemo- or radiotherapeutically the migration and invasion

  17. Cholesterol Exposure at the Membrane Surface Is Necessary and Sufficient to Trigger Perfringolysin O Binding†

    PubMed Central

    Flanagan, John J.; Tweten, Rodney K.; Johnson, Arthur E.; Heuck, Alejandro P.

    2010-01-01

    Perfringolysin O (PFO) is the prototype for the cholesterol-dependent cytolysins, a family of bacterial pore-forming toxins that act on eukaryotic membranes. The pore-forming mechanism of PFO exhibits an absolute requirement for membrane cholesterol, but the complex interplay between the structural arrangement of the PFO C-terminal domain and the distribution of cholesterol in the target membrane is poorly understood. Herein we show that PFO binding to the bilayer and the initiation of the sequence of events that culminate in the formation of a transmembrane pore depend on the availability of free cholesterol at the membrane surface, while changes in the acyl chain packing of the phospholipids and cholesterol in the membrane core, or the presence or absence of detergent-resistant domains do not correlate with PFO binding. Moreover, PFO association with the membrane was inhibited by the addition of sphingomyelin, a typical component of membrane rafts in cell membranes. Finally, addition of molecules that do not interact with PFO, but intercalate into the membrane and displace cholesterol from its association with phospholipids (e.g., epicholesterol), reduced the amount of cholesterol required to trigger PFO binding. Taken together, our studies reveal that PFO binding to membranes is triggered when the concentration of cholesterol exceeds the association capacity of the phospholipids, and this cholesterol excess is then free to associate with the toxin. PMID:19292457

  18. Tuning surface hydrophilicity/hydrophobicity of hydrocarbon proton exchange membranes (PEMs).

    PubMed

    He, Chenfeng; Mighri, Frej; Guiver, Michael D; Kaliaguine, Serge

    2016-03-15

    The effect of annealing on the surface hydrophilicity of various representative classes of hydrocarbon-based proton exchange membranes (PEMs) is investigated. In all cases, a more hydrophilic membrane surface develops after annealing at elevated temperatures. The annealing time also had some influence, but in different ways depending on the class of PEM. Longer annealing times resulted in more hydrophilic membrane surfaces for copolymerized sulfonated poly(ether ether ketone) (SPEEK-HQ), while the opposite behavior occurred in sulfonated poly(aryl ether ether ketone) (Ph-SPEEK), sulfonated poly(aryl ether ether ketone ketone) (Ph-m-SPEEKK) and sulfonated poly (aryl ether ether nitrile) (SPAEEN-B). Increased surface hydrophilicity upon annealing results from ionic cluster decomposition, according to the "Eisenberg-Hird-Moore model" (EHM). The increased surface hydrophilicity is supported by contact angle (CA) measurements, and the cluster decomposition is auxiliarily supported by probing the level of atomic sulfur (sulfonic acid) within different surface depths using angle-dependent XPS as well as ATR-FTIR. Membrane acidification leads to more hydrophilic surfaces by elimination of the hydrogen bonding that occurs between strongly-bound residual solvent (dimethylacetamide, DMAc) and PEM sulfonic acid groups. The study of physicochemical tuning of surface hydrophilicity/hydrophobicity of PEMs by annealing and acidification provides insights for improving membrane electrode assembly (MEA) fabrication in fuel cell (FC). PMID:26722798

  19. Sutureless Fixation of Amniotic Membrane for Therapy of Ocular Surface Disorders

    PubMed Central

    Kotomin, Ilya; Valtink, Monika; Hofmann, Kai; Frenzel, Annika; Morawietz, Henning; Werner, Carsten; Funk, Richard H. W.; Engelmann, Katrin

    2015-01-01

    Amniotic membrane is applied to the diseased ocular surface to stimulate wound healing and tissue repair, because it releases supportive growth factors and cytokines. These effects fade within about a week after application, necessitating repeated application. Generally, amniotic membrane is fixed with sutures to the ocular surface, but surgical intervention at the inflamed or diseased site can be detrimental. Therefore, we have developed a system for the mounting of amniotic membrane between two rings for application to a diseased ocular surface without surgical intervention (sutureless amniotic membrane transplantation). With this system, AmnioClip, amniotic membrane can be applied like a large contact lens. First prototypes were tested in an experiment on oneself for wearing comfort. The final system was tested on 7 patients in a pilot study. A possible influence of the ring system on the biological effects of amniotic membrane was analyzed by histochemistry and by analyzing the expression of vascular endothelial growth factor-A (VEGF-A), hepatocyte growth factor (HGF), fibroblast growth factor 2 (FGF 2) and pigment epithelium-derived factor (PEDF) from amniotic membranes before and after therapeutic application. The final product, AmnioClip, showed good tolerance and did not impair the biological effects of amniotic membrane. VEGF-A and PEDF mRNA was expressed in amniotic membrane after storage and mounting before transplantation, but was undetectable after a 7-day application period. Consequently, transplantation of amniotic membranes with AmnioClip provides a sutureless and hence improved therapeutic strategy for corneal surface disorders. Trial Registration ClinicalTrials.gov NCT02168790 PMID:25955359

  20. Concise Review: Plasma and Nuclear Membranes Convey Mechanical Information to Regulate Mesenchymal Stem Cell Lineage.

    PubMed

    Uzer, Gunes; Fuchs, Robyn K; Rubin, Janet; Thompson, William R

    2016-06-01

    Numerous factors including chemical, hormonal, spatial, and physical cues determine stem cell fate. While the regulation of stem cell differentiation by soluble factors is well-characterized, the role of mechanical force in the determination of lineage fate is just beginning to be understood. Investigation of the role of force on cell function has largely focused on "outside-in" signaling, initiated at the plasma membrane. When interfaced with the extracellular matrix, the cell uses integral membrane proteins, such as those found in focal adhesion complexes to translate force into biochemical signals. Akin to these outside-in connections, the internal cytoskeleton is physically linked to the nucleus, via proteins that span the nuclear membrane. Although structurally and biochemically distinct, these two forms of mechanical coupling influence stem cell lineage fate and, when disrupted, often lead to disease. Here we provide an overview of how mechanical coupling occurs at the plasma and nuclear membranes. We also discuss the role of force on stem cell differentiation, with focus on the biochemical signals generated at the cell membrane and the nucleus, and how those signals influence various diseases. While the interaction of stem cells with their physical environment and how they respond to force is complex, an understanding of the mechanical regulation of these cells is critical in the design of novel therapeutics to combat diseases associated with aging, cancer, and osteoporosis. Stem Cells 2016;34:1455-1463. PMID:26891206

  1. Stem cell differentiation increases membrane-actin adhesion regulating cell blebability, migration and mechanics

    PubMed Central

    Sliogeryte, Kristina; Thorpe, Stephen D.; Lee, David A.; Botto, Lorenzo; Knight, Martin M.

    2014-01-01

    This study examines how differentiation of human mesenchymal stem cells regulates the interaction between the cell membrane and the actin cortex controlling cell behavior. Micropipette aspiration was used to measure the pressure required for membrane-cortex detachment which increased from 0.15 kPa in stem cells to 0.71 kPa following chondrogenic differentiation. This effect was associated with reduced susceptibility to mechanical and osmotic bleb formation, reduced migration and an increase in cell modulus. Theoretical modelling of bleb formation demonstrated that the increased stiffness of differentiated cells was due to the increased membrane-cortex adhesion. Differentiated cells exhibited greater F-actin density and slower actin remodelling. Differentiated cells also expressed greater levels of the membrane-cortex ezrin, radixin, moeisin (ERM) linker proteins which was responsible for the reduced blebability, as confirmed by transfection of stem cells with dominant active ezrin-T567D-GFP. This study demonstrates that stem cells have an inherently weak membrane-cortex adhesion which increases blebability thereby regulating cell migration and stiffness. PMID:25471686

  2. Eisosomes promote the ability of Sur7 to regulate plasma membrane organization in Candida albicans

    PubMed Central

    Wang, Hong X.; Douglas, Lois M.; Veselá, Petra; Rachel, Reinhard; Malinsky, Jan; Konopka, James B.

    2016-01-01

    The plasma membrane of the fungal pathogen Candida albicans forms a protective barrier that also mediates many processes needed for virulence, including cell wall synthesis, invasive hyphal morphogenesis, and nutrient uptake. Because compartmentalization of the plasma membrane is believed to coordinate these diverse activities, we examined plasma membrane microdomains termed eisosomes or membrane compartment of Can1 (MCC), which correspond to ∼200-nm-long furrows in the plasma membrane. A pil1∆ lsp1∆ mutant failed to form eisosomes and displayed strong defects in plasma membrane organization and morphogenesis, including extensive cell wall invaginations. Mutation of eisosome proteins Slm2, Pkh2, and Pkh3 did not cause similar cell wall defects, although pkh2∆ cells formed chains of furrows and pkh3∆ cells formed wider furrows, identifying novel roles for the Pkh protein kinases in regulating furrows. In contrast, the sur7∆ mutant formed cell wall invaginations similar to those for the pil1∆ lsp1∆ mutant even though it could form eisosomes and furrows. A PH-domain probe revealed that the regulatory lipid phosphatidylinositol 4,5-bisphosphate was enriched at sites of cell wall invaginations in both the sur7∆ and pil1∆ lsp1∆ cells, indicating that this contributes to the defects. The sur7∆ and pil1∆ lsp1∆ mutants displayed differential susceptibility to various types of stress, indicating that they affect overlapping but distinct functions. In support of this, many mutant phenotypes of the pil1∆ lsp1∆ cells were rescued by overexpressing SUR7. These results demonstrate that C. albicans eisosomes promote the ability of Sur7 to regulate plasma membrane organization. PMID:27009204

  3. Regulation of the transbilayer movement of diacylglycerol in the plasma membrane.

    PubMed

    Ueda, Yoshibumi; Ishitsuka, Reiko; Hullin-Matsuda, Françoise; Kobayashi, Toshihide

    2014-12-01

    This mini-review presents recent advances in the regulation of the membrane transbilayer movement (or flip-flop) of diacylglycerol (DAG), a key intermediate in lipid metabolism and a second messenger in lipid-mediated signaling. Despite progresses in lipid biophysics and imaging, little is known about the DAG dynamics across the two leaflets of the plasma membrane in living cells. Previous model membrane studies with DAG analogs demonstrated their fast flip-flop suggesting that DAG is evenly distributed between the two leaflets of the plasma membrane. However, recent molecular dynamics simulations indicate that DAG transbilayer movement depends on the lipid environment surrounding the lipid, i.e. DAG flips more slowly across a more ordered "lipid raft-like" bilayer (enriched in sphingomyelin/cholesterol) than across a more fluid bilayer (composed of unsaturated glycerophospholipids). Furthermore using the yellow fluorescent protein-tagged C1AB domain from protein kinase C-γ (EYFP-C1AB) that selectively binds DAG, we recently proved that the sphingomyelin (SM) content in the plasma membrane outer leaflet regulates DAG transbilayer movement in Madin-Darby canine kidney cells treated with bacterial phosphatidylcholine-specific phospholipase C. The dose-dependent inhibition of DAG flip-flop by SM could be reproduced in model membranes using fluorescent short chain DAG analog. Regulation of DAG transbilayer movement by the outer leaflet SM content is expected to modify the downstream recruitment of C1-domain containing effectors, thus bringing new insights on the role of DAG dynamics in cell pathophysiology. PMID:25241257

  4. VAMP7 regulates constitutive membrane incorporation of the cold-activated channel TRPM8

    PubMed Central

    Ghosh, Debapriya; Pinto, Silvia; Danglot, Lydia; Vandewauw, Ine; Segal, Andrei; Van Ranst, Nele; Benoit, Melissa; Janssens, Annelies; Vennekens, Rudi; Vanden Berghe, Pieter; Galli, Thierry; Vriens, Joris; Voets, Thomas

    2016-01-01

    The cation channel TRPM8 plays a central role in the somatosensory system, as a key sensor of innocuously cold temperatures and cooling agents. Although increased functional expression of TRPM8 has been implicated in various forms of pathological cold hypersensitivity, little is known about the cellular and molecular mechanisms that determine TRPM8 abundance at the plasma membrane. Here we demonstrate constitutive transport of TRPM8 towards the plasma membrane in atypical, non-acidic transport vesicles that contain lysosomal-associated membrane protein 1 (LAMP1), and provide evidence that vesicle-associated membrane protein 7 (VAMP7) mediates fusion of these vesicles with the plasma membrane. In line herewith, VAMP7-deficient mice exhibit reduced functional expression of TRPM8 in sensory neurons and concomitant deficits in cold avoidance and icilin-induced cold hypersensitivity. Our results uncover a cellular pathway that controls functional plasma membrane incorporation of a temperature-sensitive TRP channel, and thus regulates thermosensitivity in vivo. PMID:26843440

  5. VAMP7 regulates constitutive membrane incorporation of the cold-activated channel TRPM8.

    PubMed

    Ghosh, Debapriya; Pinto, Silvia; Danglot, Lydia; Vandewauw, Ine; Segal, Andrei; Van Ranst, Nele; Benoit, Melissa; Janssens, Annelies; Vennekens, Rudi; Vanden Berghe, Pieter; Galli, Thierry; Vriens, Joris; Voets, Thomas

    2016-01-01

    The cation channel TRPM8 plays a central role in the somatosensory system, as a key sensor of innocuously cold temperatures and cooling agents. Although increased functional expression of TRPM8 has been implicated in various forms of pathological cold hypersensitivity, little is known about the cellular and molecular mechanisms that determine TRPM8 abundance at the plasma membrane. Here we demonstrate constitutive transport of TRPM8 towards the plasma membrane in atypical, non-acidic transport vesicles that contain lysosomal-associated membrane protein 1 (LAMP1), and provide evidence that vesicle-associated membrane protein 7 (VAMP7) mediates fusion of these vesicles with the plasma membrane. In line herewith, VAMP7-deficient mice exhibit reduced functional expression of TRPM8 in sensory neurons and concomitant deficits in cold avoidance and icilin-induced cold hypersensitivity. Our results uncover a cellular pathway that controls functional plasma membrane incorporation of a temperature-sensitive TRP channel, and thus regulates thermosensitivity in vivo. PMID:26843440

  6. Sorting Nexin 11 Regulates Lysosomal Degradation of Plasma Membrane TRPV3.

    PubMed

    Li, Caiyue; Ma, Wenbo; Yin, Shikui; Liang, Xin; Shu, Xiaodong; Pei, Duanqing; Egan, Terrance M; Huang, Jufang; Pan, Aihua; Li, Zhiyuan

    2016-05-01

    The trafficking of ion channels to/from the plasma membrane is considered an important mechanism for cellular activity and an interesting approach for disease therapies. The transient receptor potential vanilloid 3 (TRPV3) ion channel is widely expressed in skin keratinocytes, and its trafficking mechanism to/from the plasma membrane is unknown. Here, we report that the vesicular trafficking protein sorting nexin 11 (SNX11) downregulates the level of the TRPV3 plasma membrane protein. Overexpression of SNX11 causes a decrease in the level of TRPV3 current and TRPV3 plasma membrane protein in TRPV3-transfected HEK293T cells. Subcellular localizations and western blots indicate that SNX11 interacts with TRPV3 and targets it to lysosomes for degradation, which is blocked by the lysosomal inhibitors chloroquine and leupeptin. Both TRPV3 and SNX11 are highly expressed in HaCaT cells. We show that TRPV3 agonists-activated Ca(2+) influxes and the level of native TRPV3 total protein in HaCaT cells are decreased by overexpression of SNX11 and increased by knockdown of SNX11. Our findings reveal that SNX11 promotes the trafficking of TRPV3 from the plasma membrane to lysosomes for degradation via protein-protein interactions, which demonstrates a previously unknown function of SNX11 as a regulator of TRPV3 trafficking from the plasma membrane to lysosomes. PMID:26818531

  7. New insights into the regulation of cholesterol efflux from the sperm membrane

    PubMed Central

    Leahy, Tamara; Gadella, Bart M

    2015-01-01

    Cholesterol is an essential component of the mammalian plasma membrane because it promotes membrane stability without comprising membrane fluidity. Given this important cellular role, cholesterol levels are tightly controlled at multiple levels. It has been clearly shown that cholesterol redistribution and depletion from the sperm membrane is a key part of the spermatozoon's preparation for fertilization. Some factors that regulate these events are described (e.g., bicarbonate, calcium) but the mechanisms underlying cholesterol export are poorly understood. How does a hydrophobic cholesterol molecule inserted in the sperm plasma membrane enter the energetically unfavorable aqueous surroundings? This review will provide an overview of knowledge in this area and highlight our gaps in understanding. The overall aim is to better understand cholesterol redistribution in the sperm plasma membrane, its relation to the possible activation of a cholesterol transporter and the role of cholesterol acceptors. Armed with such knowledge, sperm handling techniques can be adapted to better prepare spermatozoa for in vitro and in vivo fertilization. PMID:25926609

  8. The role of K+ conductances in regulating membrane excitability in human gastric corpus smooth muscle

    PubMed Central

    Lee, Ji Yeon; Ko, Eun-ju; Ahn, Ki Duck; Kim, Sung

    2015-01-01

    Changes in resting membrane potential (RMP) regulate membrane excitability. K+ conductance(s) are one of the main factors in regulating RMP. The functional role of K+ conductances has not been studied the in human gastric corpus smooth muscles (HGCS). To examine the role of K+ channels in regulation of RMP in HGCS we employed microelectrode recordings, patch-clamp, and molecular approaches. Tetraethylammonium and charybdotoxin did not affect the RMP, suggesting that BK channels are not involved in regulating RMP. Apamin, a selective small conductance Ca2+-activated K+ channel (SK) blocker, did not show a significant effect on the membrane excitability. 4-Aminopyridine, a Kv channel blocker, caused depolarization and increased the duration of slow wave potentials. 4-Aminopyridine also inhibited a delayed rectifying K+ current in isolated smooth muscle cells. End-product RT-PCR gel detected Kv1.2 and Kv1.5 in human gastric corpus muscles. Glibenclamide, an ATP-sensitive K+ channel (KATP) blocker, did not induce depolarization, but nicorandil, a KATP opener, hyperpolarized HGCS, suggesting that KATP are expressed but not basally activated. Kir6.2 transcript, a pore-forming subunit of KATP was expressed in HGCS. A low concentration of Ba2+, a Kir blocker, induced strong depolarization. Interestingly, Ba2+-sensitive currents were minimally expressed in isolated smooth muscle cells under whole-cell patch configuration. KCNJ2 (Kir2.1) transcript was expressed in HGCS. Unique K+ conductances regulate the RMP in HGCS. Delayed and inwardly rectifying K+ channels are the main candidates in regulating membrane excitability in HGCS. With the development of cell dispersion techniques of interstitial cells, the cell-specific functional significance will require further analysis. PMID:25591864

  9. Cell cycle-regulated membrane binding of NuMA contributes to efficient anaphase chromosome separation.

    PubMed

    Zheng, Zhen; Wan, Qingwen; Meixiong, Gerry; Du, Quansheng

    2014-03-01

    Accurate and efficient separation of sister chromatids during anaphase is critical for faithful cell division. It has been proposed that cortical dynein-generated pulling forces on astral microtubules contribute to anaphase spindle elongation and chromosome separation. In mammalian cells, however, definitive evidence for the involvement of cortical dynein in chromosome separation is missing. It is believed that dynein is recruited and anchored at the cell cortex during mitosis by the α subunit of heterotrimeric G protein (Gα)/mammalian homologue of Drosophila Partner of Inscuteable/nuclear mitotic apparatus (NuMA) ternary complex. Here we uncover a Gα/LGN-independent lipid- and membrane-binding domain at the C-terminus of NuMA. We show that the membrane binding of NuMA is cell cycle regulated-it is inhibited during prophase and metaphase by cyclin-dependent kinase 1 (CDK1)-mediated phosphorylation and only occurs after anaphase onset when CDK1 activity is down-regulated. Further studies indicate that cell cycle-regulated membrane association of NuMA underlies anaphase-specific enhancement of cortical NuMA and dynein. By replacing endogenous NuMA with membrane-binding-deficient NuMA, we can specifically reduce the cortical accumulation of NuMA and dynein during anaphase and demonstrate that cortical NuMA and dynein contribute to efficient chromosome separation in mammalian cells. PMID:24371089

  10. Calmodulin effects on steroids-regulated plasma membrane calcium pump activity.

    PubMed

    Zylinska, Ludmila; Kowalska, Iwona; Ferenc, Bozena

    2009-03-01

    It is now generally accepted that non-genomic steroids action precedes their genomic effects by modulation of intracellular signaling pathways within seconds after application. Ca(2+) is a very potent and ubiquitous ion in all cells, and its concentration is precisely regulated. The most sensitive on Ca(2+) increase is ATP-consuming plasma membrane calcium pump (PMCA). The enzyme is coded by four genes, but isoforms diversity was detected in excitable and non-excitable cells. It is the only ion pump stimulated directly by calmodulin (CaM). We examined the role of PMCA isoforms composition and CaM effect in regulation of Ca(2+) uptake by estradiol, dehydroepiandrosterone (DHEA), pregnenolone (PREG), and their sulfates in a concentration range from 10(-9) to 10(-6) M, using the membranes from rat cortical synaptosomes, differentiated PC12 cells, and human erythrocytes. In excitable membranes with full set of PMCAs steroids apparently increased Ca(2+) uptake, although to a variable extent. In most of the cases, CaM decreased transport by 30-40% below controls. Erythrocyte PMCA was regulated by the steroids somewhat differently than excitable cells. CaM strongly increased the potency for Ca(2+) extrusion in membranes incubated with 17-beta-estradiol and PREG. Our results indicated that steroids may sufficiently control cytoplasmic calcium concentration within physiological and therapeutic range. The response depended on the cell type, PMCA isoforms expression profile, CaM presence, and the steroids structure. PMID:19226536