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Sample records for membrane fusion

  1. Membrane fusion

    DEFF Research Database (Denmark)

    Bendix, Pól Martin

    2015-01-01

    At Stanford University, Boxer lab, I worked on membrane fusion of small unilamellar lipid vesicles to flat membranes tethered to glass surfaces. This geometry closely resembles biological systems in which liposomes fuse to plasma membranes. The fusion mechanism was studied using DNA zippering...... between complementary strands linked to the two apposing membranes closely mimicking the zippering mechanism of SNARE fusion complexes....

  2. Viral membrane fusion

    International Nuclear Information System (INIS)

    Harrison, Stephen C.

    2015-01-01

    Membrane fusion is an essential step when enveloped viruses enter cells. Lipid bilayer fusion requires catalysis to overcome a high kinetic barrier; viral fusion proteins are the agents that fulfill this catalytic function. Despite a variety of molecular architectures, these proteins facilitate fusion by essentially the same generic mechanism. Stimulated by a signal associated with arrival at the cell to be infected (e.g., receptor or co-receptor binding, proton binding in an endosome), they undergo a series of conformational changes. A hydrophobic segment (a “fusion loop” or “fusion peptide”) engages the target-cell membrane and collapse of the bridging intermediate thus formed draws the two membranes (virus and cell) together. We know of three structural classes for viral fusion proteins. Structures for both pre- and postfusion conformations of illustrate the beginning and end points of a process that can be probed by single-virion measurements of fusion kinetics. - Highlights: • Viral fusion proteins overcome the high energy barrier to lipid bilayer merger. • Different molecular structures but the same catalytic mechanism. • Review describes properties of three known fusion-protein structural classes. • Single-virion fusion experiments elucidate mechanism

  3. Fusion of biological membranes

    Indian Academy of Sciences (India)

    small hemifusion diaphragm. To obtain a direct view of the fusion process, we have carried out extensive simulations of two bilayers, composed of block copolymers, which are immersed in a solvent which favors one of the blocks. As in the biological case, the membranes are placed under tension. This is essential as fusion ...

  4. Fusion of biological membranes

    Indian Academy of Sciences (India)

    Logo of the Indian Academy of Sciences. Indian Academy of ... Home; Journals; Pramana – Journal of Physics; Volume 64; Issue 6. Fusion of biological ... The process of membrane fusion has been examined by Monte Carlo simulation, and is found to be very different than the conventional picture. The differences in ...

  5. Fusion of biological membranes

    Indian Academy of Sciences (India)

    Abstract. The process of membrane fusion has been examined by Monte Carlo simu- lation, and is found to be very different than the conventional picture. The differences in mechanism lead to several predictions, in particular that fusion is accompanied by tran- sient leakage. This prediction has recently been verified.

  6. Fusion of biological membranes

    Indian Academy of Sciences (India)

    The process of membrane fusion has been examined by Monte Carlo simulation, and is found to be very different than the conventional picture. The differences in mechanism lead to several predictions, in particular that fusion is accompanied by transient leakage. This prediction has recently been verified. Self-consistent ...

  7. A Model for Membrane Fusion

    Science.gov (United States)

    Ngatchou, Annita

    2010-01-01

    Pheochromocytoma is a tumor of the adrenal gland which originates from chromaffin cells and is characterized by the secretion of excessive amounts of neurotransmitter which lead to high blood pressure and palpitations. Pheochromocytoma contain membrane bound granules that store neurotransmitter. The release of these stored molecules into the extracellular space occurs by fusion of the granule membrane with the cell plasma membrane, a process called exocytosis. The molecular mechanism of this membrane fusion is not well understood. It is proposed that the so called SNARE proteins [1] are the pillar of vesicle fusion as their cleavage by clostridial toxin notably, Botulinum neurotoxin and Tetanus toxin abrogate the secretion of neurotransmitter [2]. Here, I describe how physical principles are applied to a biological cell to explore the role of the vesicle SNARE protein synaptobrevin-2 in easing granule fusion. The data presented here suggest a paradigm according to which the movement of the C-terminal of synaptobrevin-2 disrupts the lipid bilayer to form a fusion pore through which molecules can exit.

  8. Membrane Trafficking and Vesicle Fusion

    Indian Academy of Sciences (India)

    Home; Journals; Resonance – Journal of Science Education; Volume 19; Issue 5. Membrane Trafficking and Vesicle Fusion: Post-Palade Era Researchers Win the Nobel Prize. Riddhi Atul Jani Subba Rao Gangi Setty. General Article Volume 19 Issue 5 May 2014 pp 421-445 ...

  9. Lipid Acrobatics in the Membrane Fusion Arena

    NARCIS (Netherlands)

    Markvoort, Albert J.; Marrink, Siewert J.; Chernomordik, Leonid V.; Kozlov, Michael M.

    2011-01-01

    In this review, we describe the recent contribution of computer simulation approaches to unravel the molecular details of membrane fusion. Over the past decade, fusion between apposed membranes and vesicles has been studied using a large variety of simulation methods and systems. Despite the variety

  10. Coiled coil driven membrane fusion between cyclodextrin vesicles and liposomes

    NARCIS (Netherlands)

    Versluis, Frank; Voskuhl, Jens; Vos, Jan; Friedrich, Heiner; Ravoo, Bart Jan; Bomans, Paul H H; Stuart, Marc C A; Sommerdijk, Nico A J M; Kros, Alexander

    2014-01-01

    Controlled fusion events between natural membranes composed of phospholipids with synthetic unnatural membranes will yield valuable fundamental information on the mechanism of membrane fusion. Here, fusion between vastly different phospholipid liposomes and cyclodextrin amphiphile based vesicles

  11. Assembly of intermediates for rapid membrane fusion.

    Science.gov (United States)

    Harner, Max; Wickner, William

    2018-01-26

    Membrane fusion is essential for intracellular protein sorting, cell growth, hormone secretion, and neurotransmission. Rapid membrane fusion requires tethering and Sec1-Munc18 (SM) function to catalyze R-, Qa-, Qb-, and Qc-SNARE complex assembly in trans , as well as SNARE engagement by the SNARE-binding chaperone Sec17/αSNAP. The hexameric vacuolar HOPS ( ho motypic fusion and vacuole p rotein s orting) complex in the yeast Saccharomyces cerevisiae tethers membranes through its affinities for the membrane Rab GTPase Ypt7. HOPS also has specific affinities for the vacuolar SNAREs and catalyzes SNARE complex assembly, but the order of their assembly into a 4-SNARE complex is unclear. We now report defined assembly intermediates on the path to membrane fusion. We found that a prefusion intermediate will assemble with HOPS and the R, Qa, and Qc SNAREs, and that this assembly undergoes rapid fusion upon addition of Qb and Sec17. HOPS-tethered membranes and all four vacuolar SNAREs formed a complex that underwent an even more dramatic burst of fusion upon Sec17p addition. These findings provide initial insights into an ordered fusion pathway consisting of the following intermediates and events: 1) Rab- and HOPS-tethered membranes, 2) a HOPS:R:Qa:Qc trans -complex, 3) a HOPS:4-SNARE trans -complex, 4) an engagement with Sec17, and 5) the rapid lipid rearrangements during fusion. In conclusion, our results indicate that the R:Qa:Qc complex forms in the context of membrane, Ypt7, HOPS, and trans -SNARE assembly and serves as a functional intermediate for rapid fusion after addition of the Qb-SNARE and Sec17 proteins. © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

  12. Membrane Trafficking and Vesicle Fusion

    Indian Academy of Sciences (India)

    IAS Admin

    hemophagocytic syndrome) and metabolic (diabe- tes) disorders [2, 23, 33]. Mutations in the genes of the basic secretory protein machinery lead to a number of membrane trafficking diseases such as Charcot–Marie–Tooth disease, Cohen.

  13. Membrane Trafficking and Vesicle Fusion

    Indian Academy of Sciences (India)

    IAS Admin

    investigators were awarded the Nobel Prize in Physiology or. Medicine in 2013. Introduction. Membrane Transport: In the eukaryotic cell, a majority of proteins are made in the cytosol. But the transmembrane and secretory proteins are synthesized in an organelle called the rough endoplasmic reticulum (ER). These proteins ...

  14. Peptides and membrane fusion : Towards an understanding of the molecular mechanism of protein-induced fusion

    NARCIS (Netherlands)

    Pecheur, EI; Sainte-Marie, J; Bienvenue, A; Hoekstra, D

    1999-01-01

    Processes such as endo- or exocytosis, membrane recycling, fertilization and enveloped viruses infection require one or more critical membrane fusion reactions. A key feature in viral and cellular fusion phenomena is the involvement of specific fusion proteins. Among the few well-characterized

  15. Dissipative Particle Dynamics of tension-induced membrane fusion

    DEFF Research Database (Denmark)

    Shillcock, Julian C.

    2009-01-01

    Recent studies of tension-induced membrane fusion using dissipative particle dynamics (DPD) simulations are briefly reviewed. The stochastic nature of the fusion process makes it necessary to simulate a large number of fusion attempts in order to obtain reliable fusion statistics and to extract...... meaningful values for the fusion probability and the average fusion times. All successful fusion events follow the same pathway. In this fusion pathway, configurations of individual lipids play an important role. Fusion starts with individual lipids assuming a splayed tail configuration with one tail......, three sub-processes have been identified in the fusion pathway. Their energy barriers are estimated to lie in the range 8-15kBT. The fusion probability is found to possess a maximum at intermediate tension values. As one decreases the tension, the fusion probability seems to vanish before...

  16. Fusion Pore Diameter Regulation by Cations Modulating Local Membrane Anisotropy

    Directory of Open Access Journals (Sweden)

    Doron Kabaso

    2012-01-01

    Full Text Available The fusion pore is an aqueous channel that is formed upon the fusion of the vesicle membrane with the plasma membrane. Once the pore is open, it may close again (transient fusion or widen completely (full fusion to permit vesicle cargo discharge. While repetitive transient fusion pore openings of the vesicle with the plasma membrane have been observed in the absence of stimulation, their frequency can be further increased using a cAMP-increasing agent that drives the opening of nonspecific cation channels. Our model hypothesis is that the openings and closings of the fusion pore are driven by changes in the local concentration of cations in the connected vesicle. The proposed mechanism of fusion pore dynamics is considered as follows: when the fusion pore is closed or is extremely narrow, the accumulation of cations in the vesicle (increased cation concentration likely leads to lipid demixing at the fusion pore. This process may affect local membrane anisotropy, which reduces the spontaneous curvature and thus leads to the opening of the fusion pore. Based on the theory of membrane elasticity, we used a continuum model to explain the rhythmic opening and closing of the fusion pore.

  17. On the mechanism of intracellular membrane fusion : In search of the genuine fusion factor

    NARCIS (Netherlands)

    Pecheur, EI; Maier, O; Hoekstra, D

    2000-01-01

    Intracellular membrane Fusion events require a general protein machinery that functions in vesicular traffic and in assembly and maintenance of organelles. An array of cytosolic and integral membrane proteins are currently identified, and in conjunction with ongoing detailed structural studies,

  18. Direct observation of intermediate states in model membrane fusion

    Science.gov (United States)

    Keidel, Andrea; Bartsch, Tobias F.; Florin, Ernst-Ludwig

    2016-01-01

    We introduce a novel assay for membrane fusion of solid supported membranes on silica beads and on coverslips. Fusion of the lipid bilayers is induced by bringing an optically trapped bead in contact with the coverslip surface while observing the bead’s thermal motion with microsecond temporal and nanometer spatial resolution using a three-dimensional position detector. The probability of fusion is controlled by the membrane tension on the particle. We show that the progression of fusion can be monitored by changes in the three-dimensional position histograms of the bead and in its rate of diffusion. We were able to observe all fusion intermediates including transient fusion, formation of a stalk, hemifusion and the completion of a fusion pore. Fusion intermediates are characterized by axial but not lateral confinement of the motion of the bead and independently by the change of its rate of diffusion due to the additional drag from the stalk-like connection between the two membranes. The detailed information provided by this assay makes it ideally suited for studies of early events in pure lipid bilayer fusion or fusion assisted by fusogenic molecules. PMID:27029285

  19. Tension-induced fusion of bilayer membranes and vesicles

    Science.gov (United States)

    Shillcock, Julian C.; Lipowsky, Reinhard

    2005-03-01

    Maintaining the integrity of their protective plasma membrane is a primary requirement of cells. Accordingly, cellular events that breach the membrane are tightly regulated. Artificial vesicles used in drug delivery must also stay intact until they have reached the desired target. In both cases, the intrinsic resistance of the membrane to rupture must be overcome to allow the efflux of the vesicle's contents. Here, we use mesoscopic simulations to study the fusion of 28-nm-diameter vesicles to 50 × 50 nm2 planar membrane patches over 2 μs. We monitor the time evolution of 93 different fusion attempts. This allows us to construct a global morphology diagram, using the initial tensions of the vesicle and the planar membrane patch as control parameters, and to determine the corresponding fusion statistics. All successful fusion events are observed to occur within 350 ns, which reflects the presence of alternative pathways for the tension relaxation.

  20. Herpesvirus glycoproteins undergo multiple antigenic changes before membrane fusion.

    Directory of Open Access Journals (Sweden)

    Daniel L Glauser

    Full Text Available Herpesvirus entry is a complicated process involving multiple virion glycoproteins and culminating in membrane fusion. Glycoprotein conformation changes are likely to play key roles. Studies of recombinant glycoproteins have revealed some structural features of the virion fusion machinery. However, how the virion glycoproteins change during infection remains unclear. Here using conformation-specific monoclonal antibodies we show in situ that each component of the Murid Herpesvirus-4 (MuHV-4 entry machinery--gB, gH/gL and gp150--changes in antigenicity before tegument protein release begins. Further changes then occurred upon actual membrane fusion. Thus virions revealed their final fusogenic form only in late endosomes. The substantial antigenic differences between this form and that of extracellular virions suggested that antibodies have only a limited opportunity to block virion membrane fusion.

  1. The dengue virus type 2 envelope protein fusion peptide is essential for membrane fusion

    International Nuclear Information System (INIS)

    Huang, Claire Y.-H.; Butrapet, Siritorn; Moss, Kelly J.; Childers, Thomas; Erb, Steven M.; Calvert, Amanda E.; Silengo, Shawn J.; Kinney, Richard M.; Blair, Carol D.; Roehrig, John T.

    2010-01-01

    The flaviviral envelope (E) protein directs virus-mediated membrane fusion. To investigate membrane fusion as a requirement for virus growth, we introduced 27 unique mutations into the fusion peptide of an infectious cDNA clone of dengue 2 virus and recovered seven stable mutant viruses. The fusion efficiency of the mutants was impaired, demonstrating for the first time the requirement for specific FP AAs in optimal fusion. Mutant viruses exhibited different growth kinetics and/or genetic stabilities in different cell types and adult mosquitoes. Virus particles could be recovered following RNA transfection of cells with four lethal mutants; however, recovered viruses could not re-infect cells. These viruses could enter cells, but internalized virus appeared to be retained in endosomal compartments of infected cells, thus suggesting a fusion blockade. Mutations of the FP also resulted in reduced virus reactivity with flavivirus group-reactive antibodies, confirming earlier reports using virus-like particles.

  2. Membrane fusion triggers rapid degradation of two gamete-specific, fusion-essential proteins in a membrane block to polygamy in Chlamydomonas

    OpenAIRE

    Liu, Yanjie; Misamore, Michael J.; Snell, William J.

    2010-01-01

    The plasma membranes of gametes are specialized for fusion, yet, once fusion occurs, in many organisms the new zygote becomes incapable of further membrane fusion reactions. The molecular mechanisms that underlie this loss of fusion capacity (block to polygamy) remain unknown. During fertilization in the green alga Chlamydomonas, the plus gamete-specific membrane protein FUS1 is required for adhesion between the apically localized sites on the plasma membranes of plus and minus gametes that a...

  3. MEMBRANE-FUSION OF SEMLIKI FOREST VIRUS INVOLVES HOMOTRIMERS OF THE FUSION PROTEIN

    NARCIS (Netherlands)

    WAHLBERG, JM; WILSCHUT, J; GAROFF, H

    1992-01-01

    Infection of cells with enveloped viruses is accomplished through membrane fusion. The binding and fusion Processes are mediated by the spike proteins in the envelope of the virus particle and usually involve a series of conformational changes in these proteins. We have studied the low-pH-mediated

  4. Molecular View of the Role of Fusion Peptides in Promoting Positive Membrane Curvature

    NARCIS (Netherlands)

    Fuhrmans, Marc; Marrink, Siewert J.

    2012-01-01

    Fusion peptides are moderately hydrophobic segments of viral and nonviral membrane fusion proteins that enable these proteins to fuse two closely apposed biological membranes. In vitro assays furthermore show that even isolated fusion peptides alone can support membrane fusion in model systems. In

  5. Inner membrane fusion mediates spatial distribution of axonal mitochondria

    Science.gov (United States)

    Yu, Yiyi; Lee, Hao-Chih; Chen, Kuan-Chieh; Suhan, Joseph; Qiu, Minhua; Ba, Qinle; Yang, Ge

    2016-01-01

    In eukaryotic cells, mitochondria form a dynamic interconnected network to respond to changing needs at different subcellular locations. A fundamental yet unanswered question regarding this network is whether, and if so how, local fusion and fission of individual mitochondria affect their global distribution. To address this question, we developed high-resolution computational image analysis techniques to examine the relations between mitochondrial fusion/fission and spatial distribution within the axon of Drosophila larval neurons. We found that stationary and moving mitochondria underwent fusion and fission regularly but followed different spatial distribution patterns and exhibited different morphology. Disruption of inner membrane fusion by knockdown of dOpa1, Drosophila Optic Atrophy 1, not only increased the spatial density of stationary and moving mitochondria but also changed their spatial distributions and morphology differentially. Knockdown of dOpa1 also impaired axonal transport of mitochondria. But the changed spatial distributions of mitochondria resulted primarily from disruption of inner membrane fusion because knockdown of Milton, a mitochondrial kinesin-1 adapter, caused similar transport velocity impairment but different spatial distributions. Together, our data reveals that stationary mitochondria within the axon interconnect with moving mitochondria through fusion and fission and that local inner membrane fusion between individual mitochondria mediates their global distribution. PMID:26742817

  6. Amphiphilic gold nanoparticles as modulators of lipid membrane fusion

    Science.gov (United States)

    Tahir, Mukarram; Alexander-Katz, Alfredo

    The fusion of lipid membranes is central to biological functions like inter-cellular transport and signaling and is coordinated by proteins of the soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs) superfamily. We utilize molecular dynamics simulations to demonstrate that gold nanoparticles functionalized with a mixed-monolayer of hydrophobic and hydrophilic alkanethiol ligands can act as synthetic analogues of these fusion proteins and mediate lipid membrane fusion by catalyzing the formation of a toroidal stalk between adjacent membranes and enabling the formation of a fusion pore upon influx of Ca2+ into the exterior solvent. The fusion pathway enabled by these synthetic nanostructures is analogous to the regulated fast fusion pathway observed during synaptic vesicle fusion; it therefore provides novel physical insights into this important biological process while also being relevant in a number of single-cell therapeutic applications. Computational resources from NSF XSEDE contract TG-DMR130042. Financial support from DOE CSGF fellowship DE-FG02-97ER25308.

  7. Vesicle fusion with bilayer lipid membrane controlled by electrostatic interaction

    Directory of Open Access Journals (Sweden)

    Azusa Oshima

    2017-09-01

    Full Text Available The fusion of proteoliposomes is a promising approach for incorporating membrane proteins in artificial lipid membranes. In this study, we employed an electrostatic interaction between vesicles and supported bilayer lipid membranes (s-BLMs to control the fusion process. We combined large unilamellar vesicles (LUVs containing anionic lipids, which we used instead of proteoliposomes, and s-BLMs containing cationic lipids to control electrostatic interaction. Anionic LUVs were never adsorbed or ruptured on the SiO2 substrate with a slight negative charge, and selectively fused with cationic s-BLMs. The LUVs can be fused effectively to the target position. Furthermore, as the vesicle fusion proceeds and some of the positive charges are neutralized, the attractive interaction weakens and finally the vesicle fusion saturates. In other words, we can control the number of LUVs fused with s-BLMs by controlling the concentration of the cationic lipids in the s-BLMs. The fluidity of the s-BLMs after vesicle fusion was confirmed to be sufficiently high. This indicates that the LUVs attached to the s-BLMs were almost completely fused, and there were few intermediate state vesicles in the fusion process. We could control the position and amount of vesicle fusion with the s-BLMs by employing an electrostatic interaction.

  8. Conflicting views on the membrane fusion machinery and the fusion pore

    DEFF Research Database (Denmark)

    Sørensen, Jakob B

    2009-01-01

    of the assembly of the fusogenic SNARE-complex. Here, I review conflicting views on the function of the core fusion machinery consisting of the SNAREs, Munc18, complexin, and synaptotagmin. Munc18 controls docking of vesicles to the plasma membrane and initial SNARE-complex assembly, whereas complexin...... of the water-soluble vesicle content. The fusion pore remains the most elusive part of the exocytosis pathway, owing to its short lifetime....

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

    NARCIS (Netherlands)

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

    1997-01-01

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

  10. A Cell-Cell Fusion Assay to Assess Arenavirus Envelope Glycoprotein Membrane-Fusion Activity.

    Science.gov (United States)

    York, Joanne; Nunberg, Jack H

    2018-01-01

    For many viruses that enter their target cells through pH-dependent fusion of the viral and endosomal membranes, cell-cell fusion assays can provide an experimental platform for investigating the structure-function relationships that promote envelope glycoprotein membrane-fusion activity. Typically, these assays employ effector cells expressing the recombinant envelope glycoprotein on the cell surface and target cells engineered to quantitatively report fusion with the effector cell. In the protocol described here, Vero cells are transfected with a plasmid encoding the arenavirus envelope glycoprotein complex GPC and infected with the vTF7-3 vaccinia virus expressing the bacteriophage T7 RNA polymerase. These effector cells are mixed with target cells infected with the vCB21R-lacZ vaccinia virus encoding a β-galactosidase reporter under the control of the T7 promoter. Cell-cell fusion is induced upon exposure to low-pH medium (pH 5.0), and the resultant expression of the β-galactosidase reporter is quantitated using a chemiluminescent substrate. We have utilized this robust microplate cell-cell fusion assay extensively to study arenavirus entry and its inhibition by small-molecule fusion inhibitors.

  11. Fusion of Sendai virus with vesicles of oligomerizable lipids: a microcalorimetric analysis of membrane fusion.

    Science.gov (United States)

    Ravoo, B J; Weringa, W D; Engberts, J B

    2000-01-01

    Sendai virus fuses efficiently with small and large unilamellar vesicles of the lipid 1,2-di-n-hexadecyloxypropyl-4- (beta-nitrostyryl) phosphate (DHPBNS) at pH 7.4 and 37 degrees C, as shown by lipid mixing assays and electron microscopy. However, fusion is strongly inhibited by oligomerization of the head groups of DHPBNS in the bilayer vesicles. The enthalpy associated with fusion of Sendai virus with DHPBNS vesicles was measured by isothermal titration microcalorimetry, comparing titrations of Sendai virus into (i) solutions of DHPBNS vesicles (which fuse with the virus) and (ii) oligomerized DHPBNS vesicles (which do not fuse with the virus), respectively. The observed heat effect of fusion of Sendai virus with DHPBNS vesicles is strongly dependent on the buffer medium, reflecting a partial charge neutralization of the Sendai F and HN proteins upon insertion into the negatively-charged vesicle membrane. No buffer effect was observed for the titration of Sendai virus into oligomerized DHPBNS vesicles, indicating that inhibition of fusion is a result of inhibition of insertion of the fusion protein into the target membrane. Fusion of Sendai virus with DHPBNS vesicles is endothermic and entropy-driven. The positive enthalpy term is dominated by heat effects resulting from merging of the protein-rich viral envelope with the lipid vesicle bilayers rather than by the fusion of the viral with the vesicle bilayers per se. Copyright 2000 Academic Press.

  12. Shear-Induced Membrane Fusion in Viscous Solutions

    KAUST Repository

    Kogan, Maxim

    2014-05-06

    Large unilamellar lipid vesicles do not normally fuse under fluid shear stress. They might deform and open pores to relax the tension to which they are exposed, but membrane fusion occurring solely due to shear stress has not yet been reported. We present evidence that shear forces in a viscous solution can induce lipid bilayer fusion. The fusion of 1,2-dioleoyl-sn-glycero-3- phosphocholine (DOPC) liposomes is observed in Couette flow with shear rates above 3000 s-1 provided that the medium is viscous enough. Liposome samples, prepared at different viscosities using a 0-50 wt % range of sucrose concentration, were studied by dynamic light scattering, lipid fusion assays using Förster resonance energy transfer (FRET), and linear dichroism (LD) spectroscopy. Liposomes in solutions with 40 wt % (or more) sucrose showed lipid fusion under shear forces. These results support the hypothesis that under suitable conditions lipid membranes may fuse in response to mechanical-force- induced stress. © 2014 American Chemical Society.

  13. The coronavirus spike protein : mechanisms of membrane fusion and virion incorporation

    NARCIS (Netherlands)

    Bosch, B.J.

    2004-01-01

    The coronavirus spike protein is a membrane-anchored glycoprotein responsible for virus-cell attachment and membrane fusion, prerequisites for a successful virus infection. In this thesis, two aspects are described regarding the molecular biology of the coronavirus spike protein: its membrane fusion

  14. A compensatory mutation provides resistance to disparate HIV fusion inhibitor peptides and enhances membrane fusion.

    Directory of Open Access Journals (Sweden)

    Matthew P Wood

    Full Text Available Fusion inhibitors are a class of antiretroviral drugs used to prevent entry of HIV into host cells. Many of the fusion inhibitors being developed, including the drug enfuvirtide, are peptides designed to competitively inhibit the viral fusion protein gp41. With the emergence of drug resistance, there is an increased need for effective and unique alternatives within this class of antivirals. One such alternative is a class of cyclic, cationic, antimicrobial peptides known as θ-defensins, which are produced by many non-human primates and exhibit broad-spectrum antiviral and antibacterial activity. Currently, the θ-defensin analog RC-101 is being developed as a microbicide due to its specific antiviral activity, lack of toxicity to cells and tissues, and safety in animals. Understanding potential RC-101 resistance, and how resistance to other fusion inhibitors affects RC-101 susceptibility, is critical for future development. In previous studies, we identified a mutant, R5-tropic virus that had evolved partial resistance to RC-101 during in vitro selection. Here, we report that a secondary mutation in gp41 was found to restore replicative fitness, membrane fusion, and the rate of viral entry, which were compromised by an initial mutation providing partial RC-101 resistance. Interestingly, we show that RC-101 is effective against two enfuvirtide-resistant mutants, demonstrating the clinical importance of RC-101 as a unique fusion inhibitor. These findings both expand our understanding of HIV drug-resistance to diverse peptide fusion inhibitors and emphasize the significance of compensatory gp41 mutations.

  15. In vitro assay using engineered yeast vacuoles for neuronal SNARE-mediated membrane fusion

    Science.gov (United States)

    Ko, Young-Joon; Lee, Miriam; Kang, KyeongJin; Song, Woo Keun; Jun, Youngsoo

    2014-01-01

    Intracellular membrane fusion requires not only SNARE proteins but also other regulatory proteins such as the Rab and Sec1/Munc18 (SM) family proteins. Although neuronal SNARE proteins alone can drive the fusion between synthetic liposomes, it remains unclear whether they are also sufficient to induce the fusion of biological membranes. Here, through the use of engineered yeast vacuoles bearing neuronal SNARE proteins, we show that neuronal SNAREs can induce membrane fusion between yeast vacuoles and that this fusion does not require the function of the Rab protein Ypt7p or the SM family protein Vps33p, both of which are essential for normal yeast vacuole fusion. Although excess vacuolar SNARE proteins were also shown to mediate Rab-bypass fusion, this fusion required homotypic fusion and vacuole protein sorting complex, which bears Vps33p and was accompanied by extensive membrane lysis. We also show that this neuronal SNARE-driven vacuole fusion can be stimulated by the neuronal SM protein Munc18 and blocked by botulinum neurotoxin serotype E, a well-known inhibitor of synaptic vesicle fusion. Taken together, our results suggest that neuronal SNARE proteins are sufficient to induce biological membrane fusion, and that this new assay can be used as a simple and complementary method for investigating synaptic vesicle fusion mechanisms. PMID:24821814

  16. Visualization and Sequencing of Membrane Remodeling Leading to Influenza Virus Fusion

    Science.gov (United States)

    Gui, Long; Ebner, Jamie L.; Mileant, Alexander; Williams, James A.

    2016-01-01

    ABSTRACT Protein-mediated membrane fusion is an essential step in many fundamental biological events, including enveloped virus infection. The nature of protein and membrane intermediates and the sequence of membrane remodeling during these essential processes remain poorly understood. Here we used cryo-electron tomography (cryo-ET) to image the interplay between influenza virus and vesicles with a range of lipid compositions. By following the population kinetics of membrane fusion intermediates imaged by cryo-ET, we found that membrane remodeling commenced with the hemagglutinin fusion protein spikes grappling onto the target membrane, followed by localized target membrane dimpling as local clusters of hemagglutinin started to undergo conformational refolding. The local dimples then transitioned to extended, tightly apposed contact zones where the two proximal membrane leaflets were in most cases indistinguishable from each other, suggesting significant dehydration and possible intermingling of the lipid head groups. Increasing the content of fusion-enhancing cholesterol or bis-monoacylglycerophosphate in the target membrane led to an increase in extended contact zone formation. Interestingly, hemifused intermediates were found to be extremely rare in the influenza virus fusion system studied here, most likely reflecting the instability of this state and its rapid conversion to postfusion complexes, which increased in population over time. By tracking the populations of fusion complexes over time, the architecture and sequence of membrane reorganization leading to efficient enveloped virus fusion were thus resolved. IMPORTANCE Enveloped viruses employ specialized surface proteins to mediate fusion of cellular and viral membranes that results in the formation of pores through which the viral genetic material is delivered to the cell. For influenza virus, the trimeric hemagglutinin (HA) glycoprotein spike mediates host cell attachment and membrane fusion. While

  17. Henipavirus Mediated Membrane Fusion, Virus Entry and Targeted Therapeutics

    Directory of Open Access Journals (Sweden)

    Dimitar B. Nikolov

    2012-02-01

    Full Text Available The Paramyxoviridae genus Henipavirus is presently represented by the type species Hendra and Nipah viruses which are both recently emerged zoonotic viral pathogens responsible for repeated outbreaks associated with high morbidity and mortality in Australia, Southeast Asia, India and Bangladesh. These enveloped viruses bind and enter host target cells through the coordinated activities of their attachment (G and class I fusion (F envelope glycoproteins. The henipavirus G glycoprotein interacts with host cellular B class ephrins, triggering conformational alterations in G that lead to the activation of the F glycoprotein, which facilitates the membrane fusion process. Using the recently published structures of HeV-G and NiV-G and other paramyxovirus glycoproteins, we review the features of the henipavirus envelope glycoproteins that appear essential for mediating the viral fusion process, including receptor binding, G-F interaction, F activation, with an emphasis on G and the mutations that disrupt viral infectivity. Finally, recent candidate therapeutics for henipavirus-mediated disease are summarized in light of their ability to inhibit HeV and NiV entry by targeting their G and F glycoproteins.

  18. FUSION OF ARTIFICIAL MEMBRANES WITH MAMMALIAN SPERMATOZOA - SPECIFIC INVOLVEMENT OF THE EQUATORIAL SEGMENT AFTER ACROSOME REACTION

    NARCIS (Netherlands)

    ARTS, EGJM; KUIKEN, J; JAGER, S; HOEKSTRA, D

    1993-01-01

    The fusogenic properties of bovine and human spermatozoa membranes were investigated, using phospholipid bilayers (liposomes) as target membranes. Fusion was monitored by following lipid mixing, as revealed by an assay based on resonance-energy transfer. In addition, fusion was visualized by

  19. Paramyxovirus membrane fusion: Lessons from the F and HN atomic structures

    International Nuclear Information System (INIS)

    Lamb, Robert A.; Paterson, Reay G.; Jardetzky, Theodore S.

    2006-01-01

    Paramyxoviruses enter cells by fusion of their lipid envelope with the target cell plasma membrane. Fusion of the viral membrane with the plasma membrane allows entry of the viral genome into the cytoplasm. For paramyxoviruses, membrane fusion occurs at neutral pH, but the trigger mechanism that controls the viral entry machinery such that it occurs at the right time and in the right place remains to be elucidated. Two viral glycoproteins are key to the infection process-an attachment protein that varies among different paramyxoviruses and the fusion (F) protein, which is found in all paramyxoviruses. For many of the paramyxoviruses (parainfluenza viruses 1-5, mumps virus, Newcastle disease virus and others), the attachment protein is the hemagglutinin/neuraminidase (HN) protein. In the last 5 years, atomic structures of paramyxovirus F and HN proteins have been reported. The knowledge gained from these structures towards understanding the mechanism of viral membrane fusion is described

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

    Directory of Open Access Journals (Sweden)

    de la Vega Michelle

    2011-12-01

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

  1. Coiled-coil driven membrane fusion: zipper-like vs. non-zipper-like peptide orientation.

    Science.gov (United States)

    Versluis, Frank; Dominguez, Juan; Voskuhl, Jens; Kros, Alexander

    2013-01-01

    Membrane fusion plays a central role in biological processes such as neurotransmission and exocytosis. An important class of proteins that induce membrane fusion are called SNARE (soluble N-ethyl malemeide sensitive factor attachment protein receptors) proteins. To induce membrane fusion, two SNARE proteins embedded in opposing membranes form a four-helix coiled-coil motif together with a third, cytoplasmic, SNARE protein. Coiled-coil formation brings the two membranes into close proximity allowing fusion to occur. Importantly, structural investigations have demonstrated that native membrane fusion only occurs when the orientation of the coiled-coil motif resembles that of a zipper. The zipper orientation arises when parallel coiled-coil formation takes place between peptides that are anchored into apposing membranes at identical termini, thereby forcing the membranes into close contact. Recently, we have designed a synthetic model for membrane fusion, which is based on a set of lipidated coiled-coil forming peptide pairs which are denoted E-K. When incorporated into liposomal membranes, coiled-coil formation between these lipidated peptides induces targeted and efficient membrane fusion of liposomes. Our model system mimics SNARE-driven membrane fusion, as it contains a coiled-coil motif which has a zipper-like orientation, similar to that of the SNARE proteins. Here we investigate whether the zipper-like orientation of the coiled-coil motifs is a prerequisite for membrane fusion in our model system. Our strategy is based on conjugation of the transmembrane anchor to either the N- or the C-terminus of peptides E and K. Whereas the use of a set of complementary peptides with the membrane anchor on identical peptide termini yields the zipper-like orientation of the coiled-coil complex, membrane anchors on opposite peptide termini results in a non-zipper-like coiled-coil orientation. Surprisingly, it was observed that efficient and targeted membrane fusion was

  2. Sphingolipids activate membrane fusion of Semliki Forest virus in a stereospecific manner

    DEFF Research Database (Denmark)

    Moesby, Lise; Corver, J; Erukulla, R K

    1995-01-01

    on degradation of the viral capsid protein by trypsin encapsulated in the target liposomes. Fusion mediated by D-erythro-ceramide was not affected by the additional presence in the target liposomes of ceramide stereoisomers incapable of fusion activation. Binding of the virus to the liposomes, as assessed...... by flotation on sucrose density gradients, was not dependent on the presence of fusion-competent or fusion-incompetent sphingolipids in the liposomes. The results of this study support the notion that a stereospecific interaction of the viral fusion protein with D-erythro sphingolipids in the target membrane......The alphavirus Semliki Forest virus (SFV) enters cells through receptor-mediated endocytosis. Subsequently, triggered by the acid pH in endosomes, the viral envelope fuses with the endosomal membrane. Membrane fusion of SFV has been shown previously to be dependent on the presence of cholesterol...

  3. A tethering complex drives the terminal stage of SNARE-dependent membrane fusion

    Science.gov (United States)

    D'Agostino, Massimo; Risselada, Herre Jelger; Lürick, Anna; Ungermann, Christian; Mayer, Andreas

    2017-11-01

    Membrane fusion in eukaryotic cells mediates the biogenesis of organelles, vesicular traffic between them, and exo- and endocytosis of important signalling molecules, such as hormones and neurotransmitters. Distinct tasks in intracellular membrane fusion have been assigned to conserved protein systems. Tethering proteins mediate the initial recognition and attachment of membranes, whereas SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) protein complexes are considered as the core fusion engine. SNARE complexes provide mechanical energy to distort membranes and drive them through a hemifusion intermediate towards the formation of a fusion pore. This last step is highly energy-demanding. Here we combine the in vivo and in vitro fusion of yeast vacuoles with molecular simulations to show that tethering proteins are critical for overcoming the final energy barrier to fusion pore formation. SNAREs alone drive vacuoles only into the hemifused state. Tethering proteins greatly increase the volume of SNARE complexes and deform the site of hemifusion, which lowers the energy barrier for pore opening and provides the driving force. Thereby, tethering proteins assume a crucial mechanical role in the terminal stage of membrane fusion that is likely to be conserved at multiple steps of vesicular traffic. We therefore propose that SNAREs and tethering proteins should be considered as a single, non-dissociable device that drives fusion. The core fusion machinery may then be larger and more complex than previously thought.

  4. Protein-induced fusion can be modulated by target membrane lipids through a structural switch at the level of the fusion peptide

    NARCIS (Netherlands)

    Pecheur, EI; Martin, [No Value; Bienvenue, A; Ruysschaert, JM; Hoekstra, D

    2000-01-01

    Regulatory features of protein-induced membrane fusion are largely unclear, particularly at the level of the fusion peptide. Fusion peptides being part of larger protein complexes, such investigations are met with technical limitations. Here, we show that the fusion activity of influenza virus or

  5. The TIP30 protein complex, arachidonic acid and coenzyme A are required for vesicle membrane fusion.

    Directory of Open Access Journals (Sweden)

    Chengliang Zhang

    Full Text Available Efficient membrane fusion has been successfully mimicked in vitro using artificial membranes and a number of cellular proteins that are currently known to participate in membrane fusion. However, these proteins are not sufficient to promote efficient fusion between biological membranes, indicating that critical fusogenic factors remain unidentified. We have recently identified a TIP30 protein complex containing TIP30, acyl-CoA synthetase long-chain family member 4 (ACSL4 and Endophilin B1 (Endo B1 that promotes the fusion of endocytic vesicles with Rab5a vesicles, which transport endosomal acidification enzymes vacuolar (H⁺-ATPases (V-ATPases to the early endosomes in vivo. Here, we demonstrate that the TIP30 protein complex facilitates the fusion of endocytic vesicles with Rab5a vesicles in vitro. Fusion of the two vesicles also depends on arachidonic acid, coenzyme A and the synthesis of arachidonyl-CoA by ACSL4. Moreover, the TIP30 complex is able to transfer arachidonyl groups onto phosphatidic acid (PA, producing a new lipid species that is capable of inducing close contact between membranes. Together, our data suggest that the TIP30 complex facilitates biological membrane fusion through modification of PA on membranes.

  6. Binding and Fusion of Extracellular Vesicles to the Plasma Membrane of Their Cell Targets.

    Science.gov (United States)

    Prada, Ilaria; Meldolesi, Jacopo

    2016-08-09

    Exosomes and ectosomes, extracellular vesicles of two types generated by all cells at multivesicular bodies and the plasma membrane, respectively, play critical roles in physiology and pathology. A key mechanism of their function, analogous for both types of vesicles, is the fusion of their membrane to the plasma membrane of specific target cells, followed by discharge to the cytoplasm of their luminal cargo containing proteins, RNAs, and DNA. Here we summarize the present knowledge about the interactions, binding and fusions of vesicles with the cell plasma membrane. The sequence initiates with dynamic interactions, during which vesicles roll over the plasma membrane, followed by the binding of specific membrane proteins to their cell receptors. Membrane binding is then converted rapidly into fusion by mechanisms analogous to those of retroviruses. Specifically, proteins of the extracellular vesicle membranes are structurally rearranged, and their hydrophobic sequences insert into the target cell plasma membrane which undergoes lipid reorganization, protein restructuring and membrane dimpling. Single fusions are not the only process of vesicle/cell interactions. Upon intracellular reassembly of their luminal cargoes, vesicles can be regenerated, released and fused horizontally to other target cells. Fusions of extracellular vesicles are relevant also for specific therapy processes, now intensely investigated.

  7. Multi-layered nanoparticles for penetrating the endosome and nuclear membrane via a step-wise membrane fusion process.

    Science.gov (United States)

    Akita, Hidetaka; Kudo, Asako; Minoura, Arisa; Yamaguti, Masaya; Khalil, Ikramy A; Moriguchi, Rumiko; Masuda, Tomoya; Danev, Radostin; Nagayama, Kuniaki; Kogure, Kentaro; Harashima, Hideyoshi

    2009-05-01

    Efficient targeting of DNA to the nucleus is a prerequisite for effective gene therapy. The gene-delivery vehicle must penetrate through the plasma membrane, and the DNA-impermeable double-membraned nuclear envelope, and deposit its DNA cargo in a form ready for transcription. Here we introduce a concept for overcoming intracellular membrane barriers that involves step-wise membrane fusion. To achieve this, a nanotechnology was developed that creates a multi-layered nanoparticle, which we refer to as a Tetra-lamellar Multi-functional Envelope-type Nano Device (T-MEND). The critical structural elements of the T-MEND are a DNA-polycation condensed core coated with two nuclear membrane-fusogenic inner envelopes and two endosome-fusogenic outer envelopes, which are shed in stepwise fashion. A double-lamellar membrane structure is required for nuclear delivery via the stepwise fusion of double layered nuclear membrane structure. Intracellular membrane fusions to endosomes and nuclear membranes were verified by spectral imaging of fluorescence resonance energy transfer (FRET) between donor and acceptor fluorophores that had been dually labeled on the liposome surface. Coating the core with the minimum number of nucleus-fusogenic lipid envelopes (i.e., 2) is essential to facilitate transcription. As a result, the T-MEND achieves dramatic levels of transgene expression in non-dividing cells.

  8. Mitochondrial DNA mutations provoke dominant inhibition of mitochondrial inner membrane fusion.

    Directory of Open Access Journals (Sweden)

    Cécile Sauvanet

    Full Text Available Mitochondria are highly dynamic organelles that continuously move, fuse and divide. Mitochondrial dynamics modulate overall mitochondrial morphology and are essential for the proper function, maintenance and transmission of mitochondria and mitochondrial DNA (mtDNA. We have investigated mitochondrial fusion in yeast cells with severe defects in oxidative phosphorylation (OXPHOS due to removal or various specific mutations of mtDNA. We find that, under fermentative conditions, OXPHOS deficient cells maintain normal levels of cellular ATP and ADP but display a reduced mitochondrial inner membrane potential. We demonstrate that, despite metabolic compensation by glycolysis, OXPHOS defects are associated to a selective inhibition of inner but not outer membrane fusion. Fusion inhibition was dominant and hampered the fusion of mutant mitochondria with wild-type mitochondria. Inhibition of inner membrane fusion was not systematically associated to changes of mitochondrial distribution and morphology, nor to changes in the isoform pattern of Mgm1, the major fusion factor of the inner membrane. However, inhibition of inner membrane fusion correlated with specific alterations of mitochondrial ultrastructure, notably with the presence of aligned and unfused inner membranes that are connected to two mitochondrial boundaries. The fusion inhibition observed upon deletion of OXPHOS related genes or upon removal of the entire mtDNA was similar to that observed upon introduction of point mutations in the mitochondrial ATP6 gene that are associated to neurogenic ataxia and retinitis pigmentosa (NARP or to maternally inherited Leigh Syndrome (MILS in humans. Our findings indicate that the consequences of mtDNA mutations may not be limited to OXPHOS defects but may also include alterations in mitochondrial fusion. Our results further imply that, in healthy cells, the dominant inhibition of fusion could mediate the exclusion of OXPHOS-deficient mitochondria from

  9. Effect of lipid molecule headgroup mismatch on non steroidal anti-inflammatory drugs induced membrane fusion.

    Science.gov (United States)

    Mondal Roy, Sutapa; Sarkar, Munna

    2011-12-20

    Membrane fusion is an essential process guiding many important biological events, which most commonly requires the aid of proteins and peptides as fusogenic agents. Small drug induced fusion at low drug concentration is a rare event. Only three drugs, namely, meloxicam (Mx), piroxicam (Px), and tenoxicam (Tx), belonging to the oxicam group of non steroidal anti-inflammatory drugs (NSAIDs) have been shown by us to induce membrane fusion successfully at low drug concentration. A better elucidation of the mechanism and the effect of different parameters in modulating the fusion process will allow the use of these common drugs to induce and control membrane fusion in various biochemical processes. In this study, we monitor the effect of lipid headgroup size mismatch in the bilayer on oxicam NSAIDs induced membrane fusion, by introducing dimyristoylphosphatidylethanolamine (DMPE) in dimyristoylphosphatidylcholine (DMPC) small unilamellar vesicles (SUVs). Such headgroup mismatch affects various lipid parameters which includes inhibition of trans-bilayer motion, domain formation, decrease in curvature, etc. Changes in various lipidic parameters introduce defects in the membrane bilayer and thereby modulate membrane fusion. SUVs formed by DMPC with increasing DMPE content (10, 20, and 30 mol %) were used as simple model membranes. Transmission electron microscopy (TEM) and differential scanning calorimetry (DSC) were used to characterize the DMPC-DMPE mixed vesicles. Fluorescence assays were used to probe the time dependence of lipid mixing, content mixing, and leakage and also used to determine the partitioning of the drugs in the membrane bilayer. How the inhibition of trans-bilayer motion, heterogeneous distribution of lipids, decrease in vesicle curvature, etc., arising due to headgroup mismatch affect the fusion process has been isolated and identified here. Mx amplifies these effects maximally followed by Px and Tx. This has been correlated to the enhanced

  10. The fusion of membranes and vesicles: pathway and energy barriers from Dissipative Particle Dynamics

    DEFF Research Database (Denmark)

    Shillcock, Julian C.

    2009-01-01

    are simulated using the optimized parameter set. In the observed fusion pathway, configurations of individual lipids play an important role. Fusion starts with individual lipids assuming a splayed tail configuration with one tail inserted in each membrane. To determine the corresponding energy barrier, we...

  11. Influenza-virus membrane fusion by cooperative fold-back of stochastically induced hemagglutinin intermediates

    NARCIS (Netherlands)

    Ivanovic, Tijana; Choi, Jason L.; Whelan, Sean P.; Oijen, Antoine M. van; Harrison, Stephen C.

    2013-01-01

    Influenza virus penetrates cells by fusion of viral and endosomal membranes catalyzed by the viral hemagglutinin (HA). Structures of the initial and final states of the HA trimer define the fusion endpoints, but do not specify intermediates. We have characterized these transitions by analyzing

  12. Switching between Successful and Dead-End Intermediates in Membrane Fusion.

    Science.gov (United States)

    Molotkovsky, Rodion J; Galimzyanov, Timur R; Jiménez-Munguía, Irene; Pavlov, Konstantin V; Batishchev, Oleg V; Akimov, Sergey A

    2017-12-02

    Fusion of cellular membranes during normal biological processes, including proliferation, or synaptic transmission, is mediated and controlled by sophisticated protein machinery ensuring the preservation of the vital barrier function of the membrane throughout the process. Fusion of virus particles with host cell membranes is more sparingly arranged and often mediated by a single fusion protein, and the virus can afford to be less discriminative towards the possible different outcomes of fusion attempts. Formation of leaky intermediates was recently observed in some fusion processes, and an alternative trajectory of the process involving formation of π-shaped structures was suggested. In this study, we apply the methods of elasticity theory and Lagrangian formalism augmented by phenomenological and molecular geometry constraints and boundary conditions to investigate the traits of this trajectory and the drivers behind the choice of one of the possible scenarios depending on the properties of the system. The alternative pathway proved to be a dead end, and, depending on the parameters of the participating membranes and fusion proteins, the system can either reversibly enter the corresponding "leaky" configuration or be trapped in it. A parametric study in the biologically relevant range of variables emphasized the fusion protein properties crucial for the choice of the fusion scenario.

  13. Switching between Successful and Dead-End Intermediates in Membrane Fusion

    Directory of Open Access Journals (Sweden)

    Rodion J. Molotkovsky

    2017-12-01

    Full Text Available Fusion of cellular membranes during normal biological processes, including proliferation, or synaptic transmission, is mediated and controlled by sophisticated protein machinery ensuring the preservation of the vital barrier function of the membrane throughout the process. Fusion of virus particles with host cell membranes is more sparingly arranged and often mediated by a single fusion protein, and the virus can afford to be less discriminative towards the possible different outcomes of fusion attempts. Formation of leaky intermediates was recently observed in some fusion processes, and an alternative trajectory of the process involving formation of π-shaped structures was suggested. In this study, we apply the methods of elasticity theory and Lagrangian formalism augmented by phenomenological and molecular geometry constraints and boundary conditions to investigate the traits of this trajectory and the drivers behind the choice of one of the possible scenarios depending on the properties of the system. The alternative pathway proved to be a dead end, and, depending on the parameters of the participating membranes and fusion proteins, the system can either reversibly enter the corresponding “leaky” configuration or be trapped in it. A parametric study in the biologically relevant range of variables emphasized the fusion protein properties crucial for the choice of the fusion scenario.

  14. Mitotic phosphorylation of VCIP135 blocks p97ATPase-mediated Golgi membrane fusion

    International Nuclear Information System (INIS)

    Totsukawa, Go; Matsuo, Ayaka; Kubota, Ayano; Taguchi, Yuya; Kondo, Hisao

    2013-01-01

    Highlights: •VCIP135 is mitotically phosphorylated on Threonine-760 and Serine-767 by Cdc2. •Phosphorylated VCIP135 does not bind to p97ATPase. •The phosphorylation of VCIP135 inhibits p97ATPase-mediated Golgi membrane fusion. -- Abstract: In mammals, the Golgi apparatus is disassembled early mitosis and reassembled at the end of mitosis. For Golgi disassembly, membrane fusion needs to be blocked. Golgi biogenesis requires two distinct p97ATPase-mediated membrane fusion, the p97/p47 and p97/p37 pathways. We previously reported that p47 phosphorylation on Serine-140 and p37 phosphorylation on Serine-56 and Threonine-59 result in mitotic inhibition of the p97/p47 and the p97/p37 pathways, respectively [11,14]. In this study, we show another mechanism of mitotic inhibition of p97-mediated Golgi membrane fusion. We clarified that VCIP135, an essential factor in both p97 membrane fusion pathways, is phosphorylated on Threonine-760 and Serine-767 by Cdc2 at mitosis and that this phosphorylated VCIP135 does not bind to p97. An in vitro Golgi reassembly assay revealed that VCIP135(T760E, S767E), which mimics mitotic phosphorylation, caused no cisternal regrowth. Our results indicate that the phosphorylation of VCIP135 on Threonine-760 and Serine-767 inhibits p97-mediated Golgi membrane fusion at mitosis

  15. Mitotic phosphorylation of VCIP135 blocks p97ATPase-mediated Golgi membrane fusion

    Energy Technology Data Exchange (ETDEWEB)

    Totsukawa, Go; Matsuo, Ayaka; Kubota, Ayano; Taguchi, Yuya; Kondo, Hisao, E-mail: hk228@med.kyushu-u.ac.jp

    2013-04-05

    Highlights: •VCIP135 is mitotically phosphorylated on Threonine-760 and Serine-767 by Cdc2. •Phosphorylated VCIP135 does not bind to p97ATPase. •The phosphorylation of VCIP135 inhibits p97ATPase-mediated Golgi membrane fusion. -- Abstract: In mammals, the Golgi apparatus is disassembled early mitosis and reassembled at the end of mitosis. For Golgi disassembly, membrane fusion needs to be blocked. Golgi biogenesis requires two distinct p97ATPase-mediated membrane fusion, the p97/p47 and p97/p37 pathways. We previously reported that p47 phosphorylation on Serine-140 and p37 phosphorylation on Serine-56 and Threonine-59 result in mitotic inhibition of the p97/p47 and the p97/p37 pathways, respectively [11,14]. In this study, we show another mechanism of mitotic inhibition of p97-mediated Golgi membrane fusion. We clarified that VCIP135, an essential factor in both p97 membrane fusion pathways, is phosphorylated on Threonine-760 and Serine-767 by Cdc2 at mitosis and that this phosphorylated VCIP135 does not bind to p97. An in vitro Golgi reassembly assay revealed that VCIP135(T760E, S767E), which mimics mitotic phosphorylation, caused no cisternal regrowth. Our results indicate that the phosphorylation of VCIP135 on Threonine-760 and Serine-767 inhibits p97-mediated Golgi membrane fusion at mitosis.

  16. SNARE motif: A common motif used by pathogens to manipulate membrane fusion

    Science.gov (United States)

    Wesolowski, Jordan

    2010-01-01

    To penetrate host cells through their membranes, pathogens use a variety of molecular components in which the presence of heptad repeat motifs seems to be a prevailing element. Heptad repeats are characterized by a pattern of seven, generally hydrophobic, residues. In order to initiate membrane fusion, viruses use glycoproteins-containing heptad repeats. These proteins are structurally and functionally similar to the SNARE proteins known to be involved in eukaryotic membrane fusion. SNAREs also display a heptad repeat motif called the “SNARE motif”. As bacterial genomes are being sequenced, microorganisms also appear to be carrying membrane proteins resembling eukaryotic SNAREs. This category of SNARE-like proteins might share similar functions and could be used by microorganisms to either promote or block membrane fusion. Such a recurrence across pathogenic organisms suggests that this architectural motif was evolutionarily selected because it most effectively ensures the survival of pathogens within the eukaryotic environment. PMID:21178463

  17. Measuring the strength of interaction between the Ebola fusion peptide and lipid rafts: implications for membrane fusion and virus infection.

    Directory of Open Access Journals (Sweden)

    Mônica S Freitas

    Full Text Available The Ebola fusion peptide (EBO₁₆ is a hydrophobic domain that belongs to the GP2 membrane fusion protein of the Ebola virus. It adopts a helical structure in the presence of mimetic membranes that is stabilized by the presence of an aromatic-aromatic interaction established by Trp8 and Phe12. In spite of its infectious cycle becoming better understood recently, several steps still remain unclear, a lacuna that makes it difficult to develop strategies to block infection. In order to gain insight into the mechanism of membrane fusion, we probed the structure, function and energetics of EBO₁₆ and its mutant W8A, in the absence or presence of different lipid membranes, including isolated domain-resistant membranes (DRM, a good experimental model for lipid rafts. The depletion of cholesterol from living mammalian cells reduced the ability of EBO₁₆ to induce lipid mixing. On the other hand, EBO₁₆ was structurally sensitive to interaction with lipid rafts (DRMs, but the same was not observed for W8A mutant. In agreement with these data, W8A showed a poor ability to promote membrane aggregation in comparison to EBO₁₆. Single molecule AFM experiments showed a high affinity force pattern for the interaction of EBO₁₆ and DRM, which seems to be a complex energetic event as observed by the calorimetric profile. Our study is the first to show a strong correlation between the initial step of Ebola virus infection and cholesterol, thus providing a rationale for Ebola virus proteins being co-localized with lipid-raft domains. In all, the results show how small fusion peptide sequences have evolved to adopt highly specific and strong interactions with membrane domains. Such features suggest these processes are excellent targets for therapeutic and vaccine approaches to viral diseases.

  18. HIV gp41 fusion peptide increases membrane ordering in a cholesterol-dependent fashion.

    Science.gov (United States)

    Lai, Alex L; Freed, Jack H

    2014-01-07

    Fusion between viral envelopes and host cell membranes, which is mediated by special glycoproteins anchored on the viral membrane, is required for HIV viral entry and infection. The HIV gp41 fusion peptide (FP), which initiates membrane fusion, adopts either an α-helical or β-sheeted structure depending on the cholesterol concentration. We used phosphocholine spin labels on the lipid headgroup and different positions on the acyl chain to detect its perturbation on lipid bilayers containing different cholesterol concentrations by electron-spin resonance. Our findings were as follows. 1), gp41 FP affects the lipid order in the same manner as previously shown for influenza hemagglutinin FP, i.e., it has a cooperative effect versus the peptide/lipid ratio, supporting our hypothesis that membrane ordering is a common prerequisite for viral membrane fusion. 2), gp41 FP induces membrane ordering in all lipid compositions studied, whereas a nonfusion mutant FP perturbs lipid order to a significantly smaller extent. 3), In high-cholesterol-containing lipid bilayers, where gp41 FP is in the β-aggregation conformation, its effect on the lipid ordering reaches deeper into the bilayer. The different extent to which the two conformers perturb is correlated with their fusogenicity. The possible role of the two conformers in membrane fusion is discussed. Copyright © 2014 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  19. The Structural Dynamics of the Flavivirus Fusion Peptide–Membrane Interaction

    Science.gov (United States)

    Souza, Theo L. F.; Sousa, Ivanildo P.; Bianconi, M. Lucia; Bernardi, Rafael C.; Pascutti, Pedro G.; Silva, Jerson L.; Gomes, Andre M. O.; Oliveira, Andréa C.

    2012-01-01

    Membrane fusion is a crucial step in flavivirus infections and a potential target for antiviral strategies. Lipids and proteins play cooperative roles in the fusion process, which is triggered by the acidic pH inside the endosome. This acidic environment induces many changes in glycoprotein conformation and allows the action of a highly conserved hydrophobic sequence, the fusion peptide (FP). Despite the large volume of information available on the virus-triggered fusion process, little is known regarding the mechanisms behind flavivirus–cell membrane fusion. Here, we evaluated the contribution of a natural single amino acid difference on two flavivirus FPs, FLAG (98DRGWGNGCGLFGK110) and FLAH (98DRGWGNHCGLFGK110), and investigated the role of the charge of the target membrane on the fusion process. We used an in silico approach to simulate the interaction of the FPs with a lipid bilayer in a complementary way and used spectroscopic approaches to collect conformation information. We found that both peptides interact with neutral and anionic micelles, and molecular dynamics (MD) simulations showed the interaction of the FPs with the lipid bilayer. The participation of the indole ring of Trp appeared to be important for the anchoring of both peptides in the membrane model, as indicated by MD simulations and spectroscopic analyses. Mild differences between FLAG and FLAH were observed according to the pH and the charge of the target membrane model. The MD simulations of the membrane showed that both peptides adopted a bend structure, and an interaction between the aromatic residues was strongly suggested, which was also observed by circular dichroism in the presence of micelles. As the FPs of viral fusion proteins play a key role in the mechanism of viral fusion, understanding the interactions between peptides and membranes is crucial for medical science and biology and may contribute to the design of new antiviral drugs. PMID:23094066

  20. Membrane fusion-competent virus-like proteoliposomes and proteinaceous supported bilayers made directly from cell plasma membranes.

    Science.gov (United States)

    Costello, Deirdre A; Hsia, Chih-Yun; Millet, Jean K; Porri, Teresa; Daniel, Susan

    2013-05-28

    Virus-like particles are useful materials for studying virus-host interactions in a safe manner. However, the standard production of pseudovirus based on the vesicular stomatitis virus (VSV) backbone is an intricate procedure that requires trained laboratory personnel. In this work, a new strategy for creating virus-like proteoliposomes (VLPLs) and virus-like supported bilayers (VLSBs) is presented. This strategy uses a cell blebbing technique to induce the formation of nanoscale vesicles from the plasma membrane of BHK cells expressing the hemagglutinin (HA) fusion protein of influenza X-31. These vesicles and supported bilayers contain HA and are used to carry out single particle membrane fusion events, monitored using total internal reflection fluorescence microscopy. The results of these studies show that the VLPLs and VLSBs contain HA proteins that are fully competent to carry out membrane fusion, including the formation of a fusion pore and the release of fluorophores loaded into vesicles. This new strategy for creating spherical and planar geometry virus-like membranes has many potential applications. VLPLs could be used to study fusion proteins of virulent viruses in a safe manner, or they could be used as therapeutic delivery particles to transport beneficial proteins coexpressed in the cells to a target cell. VLSBs could facilitate high throughput screening of antiviral drugs or pathogen-host cell interactions.

  1. Influenza-virus membrane fusion by cooperative fold-back of stochastically induced hemagglutinin intermediates.

    Science.gov (United States)

    Ivanovic, Tijana; Choi, Jason L; Whelan, Sean P; van Oijen, Antoine M; Harrison, Stephen C

    2013-02-19

    Influenza virus penetrates cells by fusion of viral and endosomal membranes catalyzed by the viral hemagglutinin (HA). Structures of the initial and final states of the HA trimer define the fusion endpoints, but do not specify intermediates. We have characterized these transitions by analyzing low-pH-induced fusion kinetics of individual virions and validated the analysis by computer simulation. We detect initial engagement with the target membrane of fusion peptides from independently triggered HAs within the larger virus-target contact patch; fusion then requires engagement of three or four neighboring HA trimers. Effects of mutations in HA indicate that withdrawal of the fusion peptide from a pocket in the pre-fusion trimer is rate-limiting for both events, but the requirement for cooperative action of several HAs to bring the fusing membranes together leads to a long-lived intermediate state for single, extended HA trimers. This intermediate is thus a fundamental aspect of the fusion mechanism. DOI:http://dx.doi.org/10.7554/eLife.00333.001.

  2. Sphingolipids activate membrane fusion of Semliki Forest virus in a stereospecific manner

    DEFF Research Database (Denmark)

    Moesby, Lise; Corver, J; Erukulla, R K

    1995-01-01

    The alphavirus Semliki Forest virus (SFV) enters cells through receptor-mediated endocytosis. Subsequently, triggered by the acid pH in endosomes, the viral envelope fuses with the endosomal membrane. Membrane fusion of SFV has been shown previously to be dependent on the presence of cholesterol ...

  3. Translocation of cell penetrating peptides and calcium-induced membrane fusion share same mechanism

    Czech Academy of Sciences Publication Activity Database

    Magarkar, Aniket; Allolio, Christoph; Jurkiewicz, Piotr; Baxová, Katarína; Šachl, Radek; Horinek, D.; Heinz, V.; Rachel, R.; Ziegler, C.; Jungwirth, Pavel

    2017-01-01

    Roč. 46, Suppl 1 (2017), S386 ISSN 0175-7571. [IUPAB congress /19./ and EBSA congress /11./. 16.07.2017-20.07.2017, Edinburgh] Institutional support: RVO:61388963 ; RVO:61388955 Keywords : membrane interactions * membrane fusion * cell penetration Subject RIV: BO - Biophysics

  4. Assessing the efficacy of vesicle fusion with planar membrane arrays using a mitochondrial porin as reporter

    International Nuclear Information System (INIS)

    Pszon-Bartosz, Kamila; Hansen, Jesper S.; Stibius, Karin B.; Groth, Jesper S.; Emneus, Jenny; Geschke, Oliver; Helix-Nielsen, Claus

    2011-01-01

    Research highlights: → We have established a vesicle fusion efficacy assay based on the major non-specific porin of Fusobacterium nucleatum (FomA). → Maximal fusion obtained was almost 150,000 porin insertions during 20 min. → Incorporation can be either first order or exponential kinetics which has implications for establishing protein delivery to biomimetic membranes. -- Abstract: Reconstitution of functionally active membrane protein into artificially made lipid bilayers is a challenge that must be overcome to create a membrane-based biomimetic sensor and separation device. In this study we address the efficacy of proteoliposome fusion with planar membrane arrays. We establish a protein incorporation efficacy assay using the major non-specific porin of Fusobacterium nucleatum (FomA) as reporter. We use electrical conductance measurements and fluorescence microscopy to characterize proteoliposome fusion with an array of planar membranes. We show that protein reconstitution in biomimetic membrane arrays may be quantified using the developed FomA assay. Specifically, we show that FomA vesicles are inherently fusigenic. Optimal FomA incorporation is obtained with a proteoliposome lipid-to-protein molar ratio (LPR) = 50 more than 10 5 FomA proteins could be incorporated in a bilayer array with a total membrane area of 2 mm 2 within 20 min. This novel assay for quantifying protein delivery into lipid bilayers may be a useful tool in developing biomimetic membrane applications.

  5. Assessing the efficacy of vesicle fusion with planar membrane arrays using a mitochondrial porin as reporter

    Energy Technology Data Exchange (ETDEWEB)

    Pszon-Bartosz, Kamila; Hansen, Jesper S. [Aquaporin A/S, Ole Maaloes Vej 3, DK-2200 Copenhagen N (Denmark); Technical University of Denmark, Department of Micro- and Nanotechnology, DK-2800 Kongens Lyngby (Denmark); Stibius, Karin B. [Aquaporin A/S, Ole Maaloes Vej 3, DK-2200 Copenhagen N (Denmark); Technical University of Denmark, Department of Physics, DTU Physics, DK-2800 Kongens Lyngby (Denmark); Groth, Jesper S. [Aquaporin A/S, Ole Maaloes Vej 3, DK-2200 Copenhagen N (Denmark); Emneus, Jenny; Geschke, Oliver [Technical University of Denmark, Department of Micro- and Nanotechnology, DK-2800 Kongens Lyngby (Denmark); Helix-Nielsen, Claus, E-mail: claus.helix.nielsen@fysik.dtu.dk [Aquaporin A/S, Ole Maaloes Vej 3, DK-2200 Copenhagen N (Denmark); Technical University of Denmark, Department of Physics, DTU Physics, DK-2800 Kongens Lyngby (Denmark)

    2011-03-04

    Research highlights: {yields} We have established a vesicle fusion efficacy assay based on the major non-specific porin of Fusobacterium nucleatum (FomA). {yields} Maximal fusion obtained was almost 150,000 porin insertions during 20 min. {yields} Incorporation can be either first order or exponential kinetics which has implications for establishing protein delivery to biomimetic membranes. -- Abstract: Reconstitution of functionally active membrane protein into artificially made lipid bilayers is a challenge that must be overcome to create a membrane-based biomimetic sensor and separation device. In this study we address the efficacy of proteoliposome fusion with planar membrane arrays. We establish a protein incorporation efficacy assay using the major non-specific porin of Fusobacterium nucleatum (FomA) as reporter. We use electrical conductance measurements and fluorescence microscopy to characterize proteoliposome fusion with an array of planar membranes. We show that protein reconstitution in biomimetic membrane arrays may be quantified using the developed FomA assay. Specifically, we show that FomA vesicles are inherently fusigenic. Optimal FomA incorporation is obtained with a proteoliposome lipid-to-protein molar ratio (LPR) = 50 more than 10{sup 5} FomA proteins could be incorporated in a bilayer array with a total membrane area of 2 mm{sup 2} within 20 min. This novel assay for quantifying protein delivery into lipid bilayers may be a useful tool in developing biomimetic membrane applications.

  6. Structure and orientation study of Ebola fusion peptide inserted in lipid membrane models.

    Science.gov (United States)

    Agopian, Audrey; Castano, Sabine

    2014-01-01

    The fusion peptide of Ebola virus comprises a highly hydrophobic sequence located downstream from the N-terminus of the glycoprotein GP2 responsible for virus-host membrane fusion. The internal fusion peptide of GP2 inserts into membranes of infected cell to mediate the viral and the host cell membrane fusion. Since the sequence length of Ebola fusion peptide is still not clear, we study in the present work the behavior of two fusion peptides of different lengths which were named EBO17 and EBO24 referring to their amino acid length. The secondary structure and orientation of both peptides in lipid model systems made of DMPC:DMPG:cholesterol:DMPE (6:2:5:3) were investigated using PMIRRAS and polarized ATR spectroscopy coupled with Brewster angle microscopy. The infrared results showed a structural flexibility of both fusion peptides which are able to transit reversibly from an α-helix to antiparallel β-sheets. Ellipsometry results corroborate together with isotherm measurements that EBO peptides interacting with lipid monolayer highly affected the lipid organization. When interacting with a single lipid bilayer, at low peptide content, EBO peptides insert as mostly α-helices mainly perpendicular into the lipid membrane thus tend to organize the lipid acyl chains. Inserted in multilamellar vesicles at higher peptide content, EBO peptides are mostly in β-sheet structures and induce a disorganization of the lipid chain order. In this paper, we show that the secondary structure of the Ebola fusion peptide is reversibly flexible between α-helical and β-sheet conformations, this feature being dependent on its concentration in lipids, eventually inducing membrane fusion. © 2013.

  7. Design, construction, and characterization of high-performance membrane fusion devices with target-selectivity.

    Science.gov (United States)

    Kashiwada, Ayumi; Yamane, Iori; Tsuboi, Mana; Ando, Shun; Matsuda, Kiyomi

    2012-01-31

    Membrane fusion proteins such as the hemagglutinin glycoprotein have target recognition and fusion accelerative domains, where some synergistically working elements are essential for target-selective and highly effective native membrane fusion systems. In this work, novel membrane fusion devices bearing such domains were designed and constructed. We selected a phenylboronic acid derivative as a recognition domain for a sugar-like target and a transmembrane-peptide (Leu-Ala sequence) domain interacting with the target membrane, forming a stable hydrophobic α-helix and accelerating the fusion process. Artificial membrane fusion behavior between the synthetic devices in which pilot and target liposomes were incorporated was characterized by lipid-mixing and inner-leaflet lipid-mixing assays. Consequently, the devices bearing both the recognition and transmembrane domains brought about a remarkable increase in the initial rate for the membrane fusion compared with the devices containing the recognition domain alone. In addition, a weakly acidic pH-responsive device was also constructed by replacing three Leu residues in the transmembrane-peptide domain by Glu residues. The presence of Glu residues made the acidic pH-dependent hydrophobic α-helix formation possible as expected. The target-selective liposome-liposome fusion was accelerated in a weakly acidic pH range when the Glu-substituted device was incorporated in pilot liposomes. The use of this pH-responsive device seems to be a potential strategy for novel applications in a liposome-based delivery system. © 2011 American Chemical Society

  8. Cytosol-dependent membrane fusion in ER, nuclear envelope and nuclear pore assembly: biological implications.

    Science.gov (United States)

    Rafikova, Elvira R; Melikov, Kamran; Chernomordik, Leonid V

    2010-01-01

    Endoplasmic reticulum and nuclear envelope rearrangements after mitosis are often studied in the reconstitution system based on Xenopus egg extract. In our recent work we partially replaced the membrane vesicles in the reconstitution mix with protein-free liposomes to explore the relative contributions of cytosolic and transmembrane proteins. Here we discuss our finding that cytosolic proteins mediate fusion between membranes lacking functional transmembrane proteins and the role of membrane fusion in endoplasmic reticulum and nuclear envelope reorganization. Cytosol-dependent liposome fusion has allowed us to restore, without adding transmembrane nucleoporins, functionality of nuclear pores, their spatial distribution and chromatin decondensation in nuclei formed at insufficient amounts of membrane material and characterized by only partial decondensation of chromatin and lack of nuclear transport. Both the mechanisms and the biological implications of the discovered coupling between spatial distribution of nuclear pores, chromatin decondensation and nuclear transport are discussed.

  9. Bacteriophage membrane protein P9 as a fusion partner for the efficient expression of membrane proteins in Escherichia coli.

    Science.gov (United States)

    Jung, Yuna; Jung, Hyeim; Lim, Dongbin

    2015-12-01

    Despite their important roles and economic values, studies of membrane proteins have been hampered by the difficulties associated with obtaining sufficient amounts of protein. Here, we report a novel membrane protein expression system that uses the major envelope protein (P9) of phage φ6 as an N-terminal fusion partner. Phage membrane protein P9 facilitated the synthesis of target proteins and their integration into the Escherichia coli cell membrane. This system was used to produce various multi-pass transmembrane proteins, including G-protein-coupled receptors, transporters, and ion channels of human origin. Green fluorescent protein fusion was used to confirm the correct folding of the expressed proteins. Of the 14 membrane proteins tested, eight were highly expressed, three were moderately expressed, and three were barely expressed in E. coli. Seven of the eight highly expressed proteins could be purified after extraction with the mild detergent lauryldimethylamine-oxide. Although a few proteins have previously been developed as fusion partners to augment membrane protein production, we believe that the major envelope protein P9 described here is better suited to the efficient expression of eukaryotic transmembrane proteins in E. coli. Copyright © 2015 Elsevier Inc. All rights reserved.

  10. Rational design of a fusion partner for membrane protein expression in E. coli.

    Science.gov (United States)

    Luo, Jianying; Choulet, Julie; Samuelson, James C

    2009-08-01

    We have designed a novel protein fusion partner (P8CBD) to utilize the co-translational SRP pathway in order to target heterologous proteins to the E. coli inner membrane. SRP-dependence was demonstrated by analyzing the membrane translocation of P8CBD-PhoA fusion proteins in wt and SRP-ffh77 mutant cells. We also demonstrate that the P8CBD N-terminal fusion partner promotes over-expression of a Thermotoga maritima polytopic membrane protein by replacement of the native signal anchor sequence. Furthermore, the yeast mitochondrial inner membrane protein Oxa1p was expressed as a P8CBD fusion and shown to function within the E. coli inner membrane. In this example, the mitochondrial targeting peptide was replaced by P8CBD. Several practical features were incorporated into the P8CBD expression system to aid in protein detection, purification, and optional in vitro processing by enterokinase. The basis of membrane protein over-expression toxicity is discussed and solutions to this problem are presented. We anticipate that this optimized expression system will aid in the isolation and study of various recombinant forms of membrane-associated protein.

  11. Physiological and molecular triggers for SARS-CoV membrane fusion and entry into host cells.

    Science.gov (United States)

    Millet, Jean Kaoru; Whittaker, Gary R

    2018-04-01

    During viral entry, enveloped viruses require the fusion of their lipid envelope with host cell membranes. For coronaviruses, this critical step is governed by the virally-encoded spike (S) protein, a class I viral fusion protein that has several unique features. Coronavirus entry is unusual in that it is often biphasic in nature, and can occur at or near the cell surface or in late endosomes. Recent advances in structural, biochemical and molecular biology of the coronavirus S protein has shed light on the intricacies of coronavirus entry, in particular the molecular triggers of coronavirus S-mediated membrane fusion. Furthermore, characterization of the coronavirus fusion peptide (FP), the segment of the fusion protein that inserts to a target lipid bilayer during membrane fusion, has revealed its particular attributes which imparts some of the unusual properties of the S protein, such as Ca 2+ -dependency. These unusual characteristics can explain at least in part the biphasic nature of coronavirus entry. In this review, using severe acute respiratory syndrome coronavirus (SARS-CoV) as model virus, we give an overview of advances in research on the coronavirus fusion peptide with an emphasis on its role and properties within the biological context of host cell entry. Copyright © 2017 Elsevier Inc. All rights reserved.

  12. Structure and interaction with lipid membrane models of Semliki Forest virus fusion peptide.

    Science.gov (United States)

    Agopian, A; Quetin, M; Castano, S

    2016-11-01

    Semliki Forest virus (SFV) is a well-characterized alphavirus that infects cells via endocytosis and an acid-triggered fusion step using class II fusion proteins. Membrane fusion is mediated by the viral spike protein, a heterotrimer of two transmembrane subunits, E1 and E2, and a peripheral protein, E3. Sequence analysis of the E1 ectodomain of a number of alphaviruses demonstrated the presence of a highly conserved hydrophobic domain on the E1 ectodomain. This sequence was proposed to be the fusion peptide of SFV and is believed to be the domain of E1 that interacts with the target membrane and triggers fusion. Here, we investigate the structure and the interaction with lipid membrane models of 76 YQCKVYTGVYPFMWGGAYCFC 96 sequence from SFV, named SFV21, using optical method (ellipsometry) and vibrational spectroscopiy approaches (Polarization Modulation infra-Red Reflection Absorption Spectroscopy, PMIRRAS, and polarized ATR-FTIR). We demonstrate a structural flexibility of SFV21 sequence whether the lateral pressure and the lipid environment. In a lipid environment that mimics eukaryotic cell membranes, a conformational transition from an α-helix to a β-sheet is induced in the presence of lipid by increasing the peptide to lipid ratio, which leads to important perturbations in the membrane organisation. Copyright © 2016 Elsevier B.V. All rights reserved.

  13. Assessing the efficacy of vesicle fusion with planar membrane arrays using a mitochondrial porin as reporter

    DEFF Research Database (Denmark)

    Pszon-Bartosz, Kamila Justyna; Hansen, Jesper S.; Stibius, Karin B.

    2011-01-01

    Reconstitution of functionally active membrane protein into artificially made lipid bilayers is a challenge that must be overcome to create a membrane-based biomimetic sensor and separation device. In this study we address the efficacy of proteoliposome fusion with planar membrane arrays. We...... establish a protein incorporation efficacy assay using the major non-specific porin of Fusobacterium nucleatum (FomA) as reporter. We use electrical conductance measurements and fluorescence microscopy to characterize proteoliposome fusion with an array of planar membranes. We show that protein...... reconstitution in biomimetic membrane arrays may be quantified using the developed FomA assay. Specifically, we show that FomA vesicles are inherently fusigenic. Optimal FomA incorporation is obtained with a proteoliposome lipid-to-protein molar ratio (LPR)=50 more than 105 FomA proteins could be incorporated...

  14. Inner/Outer nuclear membrane fusion in nuclear pore assembly: biochemical demonstration and molecular analysis.

    Science.gov (United States)

    Fichtman, Boris; Ramos, Corinne; Rasala, Beth; Harel, Amnon; Forbes, Douglass J

    2010-12-01

    Nuclear pore complexes (NPCs) are large proteinaceous channels embedded in double nuclear membranes, which carry out nucleocytoplasmic exchange. The mechanism of nuclear pore assembly involves a unique challenge, as it requires creation of a long-lived membrane-lined channel connecting the inner and outer nuclear membranes. This stabilized membrane channel has little evolutionary precedent. Here we mapped inner/outer nuclear membrane fusion in NPC assembly biochemically by using novel assembly intermediates and membrane fusion inhibitors. Incubation of a Xenopus in vitro nuclear assembly system at 14°C revealed an early pore intermediate where nucleoporin subunits POM121 and the Nup107-160 complex were organized in a punctate pattern on the inner nuclear membrane. With time, this intermediate progressed to diffusion channel formation and finally to complete nuclear pore assembly. Correct channel formation was blocked by the hemifusion inhibitor lysophosphatidylcholine (LPC), but not if a complementary-shaped lipid, oleic acid (OA), was simultaneously added, as determined with a novel fluorescent dextran-quenching assay. Importantly, recruitment of the bulk of FG nucleoporins, characteristic of mature nuclear pores, was not observed before diffusion channel formation and was prevented by LPC or OA, but not by LPC+OA. These results map the crucial inner/outer nuclear membrane fusion event of NPC assembly downstream of POM121/Nup107-160 complex interaction and upstream or at the time of FG nucleoporin recruitment.

  15. The destructive effect of botulinum neurotoxins on the SNARE protein: SNAP-25 and synaptic membrane fusion

    Directory of Open Access Journals (Sweden)

    Bin Lu

    2015-06-01

    Full Text Available Synaptic exocytosis requires the assembly of syntaxin 1A and SNAP-25 on the plasma membrane and synaptobrevin 2 (VAMP2 on the vesicular membrane to bridge the two opposite membranes. It is believed that the three SNARE proteins assemble in steps along the dynamic assembly pathway. The C-terminus of SNAP-25 is known to be the target of botulinum neurotoxins (BoNT/A and BoNT/E that block neurotransmitters release in vivo. In this study, we employed electron paramagnetic resonance (EPR spectroscopy to investigate the conformation of the SNAP-25 C-terminus in binary and ternary SNARE complexes. The fluorescence lipid mixing assay shows that the C-terminal of SNAP-25 is essential for membrane fusion, and that the truncated SNAP-25 mutants cleaved by BoNT/A and BoNT/E display different inhibition effects on membrane fusion: SNAP-25E (Δ26 abolishes the fusion activity of the SNARE complex, while SNAP-25A (Δ9 loses most of its function, although it can still form a SDS-resistant SNARE complex as the wild-type SNAP-25. CW-EPR spectra validate the unstable structures of the SNARE complex formed by SNAP-25 mutants. We propose that the truncated SNAP-25 mutants will disrupt the assembly of the SNARE core complex, and then inhibit the synaptic membrane fusion accordingly.

  16. The Fusion Loops of the Initial Prefusion Conformation of Herpes Simplex Virus 1 Fusion Protein Point Toward the Membrane

    Directory of Open Access Journals (Sweden)

    Juan Fontana

    2017-08-01

    Full Text Available All enveloped viruses, including herpesviruses, must fuse their envelope with the host membrane to deliver their genomes into target cells, making this essential step subject to interference by antibodies and drugs. Viral fusion is mediated by a viral surface protein that transits from an initial prefusion conformation to a final postfusion conformation. Strikingly, the prefusion conformation of the herpesvirus fusion protein, gB, is poorly understood. Herpes simplex virus (HSV, a model system for herpesviruses, causes diseases ranging from mild skin lesions to serious encephalitis and neonatal infections. Using cryo-electron tomography and subtomogram averaging, we have characterized the structure of the prefusion conformation and fusion intermediates of HSV-1 gB. To this end, we have set up a system that generates microvesicles displaying full-length gB on their envelope. We confirmed proper folding of gB by nondenaturing electrophoresis-Western blotting with a panel of monoclonal antibodies (MAbs covering all gB domains. To elucidate the arrangement of gB domains, we labeled them by using (i mutagenesis to insert fluorescent proteins at specific positions, (ii coexpression of gB with Fabs for a neutralizing MAb with known binding sites, and (iii incubation of gB with an antibody directed against the fusion loops. Our results show that gB starts in a compact prefusion conformation with the fusion loops pointing toward the viral membrane and suggest, for the first time, a model for gB’s conformational rearrangements during fusion. These experiments further illustrate how neutralizing antibodies can interfere with the essential gB structural transitions that mediate viral entry and therefore infectivity.

  17. Green fluorescence protein-based content-mixing assay of SNARE-driven membrane fusion.

    Science.gov (United States)

    Heo, Paul; Kong, Byoungjae; Jung, Young-Hun; Park, Joon-Bum; Shin, Jonghyeok; Park, Myungseo; Kweon, Dae-Hyuk

    2017-06-17

    Soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins mediate intracellular membrane fusion by forming a ternary SNARE complex. A minimalist approach utilizing proteoliposomes with reconstituted SNARE proteins yielded a wealth of information pinpointing the molecular mechanism of SNARE-mediated fusion and its regulation by accessory proteins. Two important attributes of a membrane fusion are lipid-mixing and the formation of an aqueous passage between apposing membranes. These two attributes are typically observed by using various fluorescent dyes. Currently available in vitro assay systems for observing fusion pore opening have several weaknesses such as cargo-bleeding, incomplete removal of unencapsulated dyes, and inadequate information regarding the size of the fusion pore, limiting measurements of the final stage of membrane fusion. In the present study, we used a biotinylated green fluorescence protein and streptavidin conjugated with Dylight 594 (DyStrp) as a Föster resonance energy transfer (FRET) donor and acceptor, respectively. This FRET pair encapsulated in each v-vesicle containing synaptobrevin and t-vesicle containing a binary acceptor complex of syntaxin 1a and synaptosomal-associated protein 25 revealed the opening of a large fusion pore of more than 5 nm, without the unwanted signals from unencapsulated dyes or leakage. This system enabled determination of the stoichiometry of the merging vesicles because the FRET efficiency of the FRET pair depended on the molar ratio between dyes. Here, we report a robust and informative assay for SNARE-mediated fusion pore opening. Copyright © 2017 Elsevier Inc. All rights reserved.

  18. The fusion-related hydrophobic domain of Sendai F protein can be moved through the cytoplasmic membrane of Escherichia coli.

    OpenAIRE

    Davis, N G; Hsu, M C

    1986-01-01

    Recent work on a prokaryotic membrane protein, gene III protein (pIII) of coliphage f1, showed that polypeptide segments of sufficient hydrophobicity functioned to stop transfer of the polypeptide across the cell membrane: strings of 16 or more hydrophobic amino acids sufficed. A fusion-related hydrophobic domain (FRHD) of Sendai F protein, a sequence of 26 consecutive uncharged residues, has been implicated in the fusion of the viral membrane envelope and the target-cell membrane through a h...

  19. Yeast Lipin 1 Orthologue Pah1p Regulates Vacuole Homeostasis and Membrane Fusion*

    Science.gov (United States)

    Sasser, Terry; Qiu, Quan-Sheng; Karunakaran, Surya; Padolina, Mark; Reyes, Anna; Flood, Blake; Smith, Sheena; Gonzales, Chad; Fratti, Rutilio A.

    2012-01-01

    Vacuole homotypic fusion requires a group of regulatory lipids that includes diacylglycerol, a fusogenic lipid that is produced through multiple metabolic pathways including the dephosphorylation of phosphatidic acid (PA). Here we examined the relationship between membrane fusion and PA phosphatase activity. Pah1p is the single yeast homologue of the Lipin family of PA phosphatases. Deletion of PAH1 was sufficient to cause marked vacuole fragmentation and abolish vacuole fusion. The function of Pah1p solely depended on its phosphatase activity as complementation studies showed that wild type Pah1p restored fusion, whereas the phosphatase dead mutant Pah1pD398E had no effect. We discovered that the lack of PA phosphatase activity blocked fusion by inhibiting the binding of SNAREs to Sec18p, an N-ethylmaleimide-sensitive factor homologue responsible for priming inactive cis-SNARE complexes. In addition, pah1Δ vacuoles were devoid of the late endosome/vacuolar Rab Ypt7p, the phosphatidylinositol 3-kinase Vps34p, and Vps39p, a subunit of the HOPS (homotypic fusion and vacuole protein sorting) tethering complex, all of which are required for vacuole fusion. The lack of Vps34p resulted in the absence of phosphatidylinositol 3-phosphate, a lipid required for SNARE activity and vacuole fusion. These findings demonstrate that Pah1p and PA phosphatase activity are critical for vacuole homeostasis and fusion. PMID:22121197

  20. Trans-complex formation by proteolipid channels in the terminal phase of membrane fusion

    DEFF Research Database (Denmark)

    Peters, C; Bayer, M J; Bühler, S

    2001-01-01

    -complex formation occurs downstream from trans-SNARE pairing, and depends on both the Rab-GTPase Ypt7 and calmodulin. The maintenance of existing complexes and completion of fusion are independent of trans-SNARE pairs. Reconstituted proteolipids form sealed channels, which can expand to form aqueous pores in a Ca2......+/calmodulin-dependent fashion. V0 trans-complexes may therefore form a continuous, proteolipid-lined channel at the fusion site. We propose that radial expansion of such a protein pore may be a mechanism for intracellular membrane fusion....

  1. The Heptad Repeat C Domain of the Respiratory Syncytial Virus Fusion Protein Plays a Key Role in Membrane Fusion.

    Science.gov (United States)

    Bermingham, Imogen M; Chappell, Keith J; Watterson, Daniel; Young, Paul R

    2018-02-15

    Respiratory syncytial virus (RSV) mediates host cell entry through the fusion (F) protein, which undergoes a conformational change to facilitate the merger of viral and host lipid membrane envelopes. The RSV F protein comprises a trimer of disulfide-bonded F 1 and F 2 subunits that is present on the virion surface in a metastable prefusion state. This prefusion form is readily triggered to undergo refolding to bring two heptad repeats (heptad repeat A [HRA] and HRB) into close proximity to form a six-helix bundle that stabilizes the postfusion form and provides the free energy required for membrane fusion. This process can be triggered independently of other proteins. Here, we have performed a comprehensive analysis of a third heptad repeat region, HRC (amino acids 75 to 97), an amphipathic α-helix that lies at the interface of the prefusion F trimer and is a major structural feature of the F 2 subunit. We performed alanine scanning mutagenesis from Lys-75 to Met-97 and assessed all mutations in transient cell culture for expression, proteolytic processing, cell surface localization, protein conformation, and membrane fusion. Functional characterization revealed a striking distribution of activity in which fusion-increasing mutations localized to one side of the helical face, while fusion-decreasing mutations clustered on the opposing face. Here, we propose a model in which HRC plays a stabilizing role within the globular head for the prefusion F trimer and is potentially involved in the early events of triggering, prompting fusion peptide release and transition into the postfusion state. IMPORTANCE RSV is recognized as the most important viral pathogen among pediatric populations worldwide, yet no vaccine or widely available therapeutic treatment is available. The F protein is critical for the viral replication process and is the major target for neutralizing antibodies. Recent years have seen the development of prefusion stabilized F protein-based approaches to

  2. Two coiled-coil domains of Chlamydia trachomatis IncA affect membrane fusion events during infection.

    Directory of Open Access Journals (Sweden)

    Erik Ronzone

    Full Text Available Chlamydia trachomatis replicates in a parasitophorous membrane-bound compartment called an inclusion. The inclusions corrupt host vesicle trafficking networks to avoid the degradative endolysosomal pathway but promote fusion with each other in order to sustain higher bacterial loads in a process known as homotypic fusion. The Chlamydia protein IncA (Inclusion protein A appears to play central roles in both these processes as it participates to homotypic fusion and inhibits endocytic SNARE-mediated membrane fusion. How IncA selectively inhibits or activates membrane fusion remains poorly understood. In this study, we analyzed the spatial and molecular determinants of IncA's fusogenic and inhibitory functions. Using a cell-free membrane fusion assay, we found that inhibition of SNARE-mediated fusion requires IncA to be on the same membrane as the endocytic SNARE proteins. IncA displays two coiled-coil domains showing high homology with SNARE proteins. Domain swap and deletion experiments revealed that although both these domains are capable of independently inhibiting SNARE-mediated fusion, these two coiled-coil domains cooperate in mediating IncA multimerization and homotypic membrane interaction. Our results support the hypothesis that Chlamydia employs SNARE-like virulence factors that positively and negatively affect membrane fusion and promote infection.

  3. Two coiled-coil domains of Chlamydia trachomatis IncA affect membrane fusion events during infection.

    Science.gov (United States)

    Ronzone, Erik; Paumet, Fabienne

    2013-01-01

    Chlamydia trachomatis replicates in a parasitophorous membrane-bound compartment called an inclusion. The inclusions corrupt host vesicle trafficking networks to avoid the degradative endolysosomal pathway but promote fusion with each other in order to sustain higher bacterial loads in a process known as homotypic fusion. The Chlamydia protein IncA (Inclusion protein A) appears to play central roles in both these processes as it participates to homotypic fusion and inhibits endocytic SNARE-mediated membrane fusion. How IncA selectively inhibits or activates membrane fusion remains poorly understood. In this study, we analyzed the spatial and molecular determinants of IncA's fusogenic and inhibitory functions. Using a cell-free membrane fusion assay, we found that inhibition of SNARE-mediated fusion requires IncA to be on the same membrane as the endocytic SNARE proteins. IncA displays two coiled-coil domains showing high homology with SNARE proteins. Domain swap and deletion experiments revealed that although both these domains are capable of independently inhibiting SNARE-mediated fusion, these two coiled-coil domains cooperate in mediating IncA multimerization and homotypic membrane interaction. Our results support the hypothesis that Chlamydia employs SNARE-like virulence factors that positively and negatively affect membrane fusion and promote infection.

  4. High-yield membrane protein expression from E. coli using an engineered outer membrane protein F fusion.

    Science.gov (United States)

    Su, Pin-Chuan; Si, William; Baker, Deidre L; Berger, Bryan W

    2013-04-01

    Obtaining high yields of membrane proteins necessary to perform detailed structural study is difficult due to poor solubility and variability in yields from heterologous expression systems. To address this issue, an Escherichia coli-based membrane protein overexpression system utilizing an engineered bacterial outer membrane protein F (pOmpF) fusion has been developed. Full-length human receptor activity-modifying protein 1 (RAMP1) was expressed using pOmpF, solubilized in FC15 and purified to homogeneity. Using circular dichroism and fluorescence spectroscopy, purified full-length RAMP1 is composed of approximately 90% α-helix, and retains its solubility and structure in FC15 over a wide range of temperatures (20-60°C). Thus, our approach provides a useful, complementary approach to achieve high-yield, full-length membrane protein overexpression for biophysical studies. Copyright © 2013 The Protein Society.

  5. Membrane fusion is induced by a distinct peptide sequence of the sea urchin fertilization protein bindin

    NARCIS (Netherlands)

    Ulrich, AS; Glabe, CG; Hoekstra, D

    1998-01-01

    Fertilization in the sea urchin is mediated by the membrane-associated acrosomal protein bindin, which plays a key role in the adhesion and fusion between sperm and egg. We have investigated the structure/function relationship of an 18-amino acid peptide fragment "B18," which represents the minimal

  6. Towards fully automated Identification of Vesicle-Membrane Fusion Events in TIRF Microscopy

    Science.gov (United States)

    Vallotton, Pascal; James, David E.; Hughes, William E.

    2007-11-01

    Total Internal Reflection Fluorescence Microscopy (TIRFM) is imposing itself as the tool of choice for studying biological activity in close proximity to the plasma membrane. For example, the exquisite selectivity of TIRFM allows monitoring the diffusion of GFP-phogrin vesicles and their recruitment to the plasma membrane in pancreatic β-cells. We present a novel computer vision system for automatically identifying the elusive fusion events of GFP-phogrin vesicles with the plasma membrane. Our method is based on robust object tracking and matched filtering. It should accelerate the quantification of TIRFM data and allow the extraction of more biological information from image data to support research in diabetes and obesity.

  7. Fusion of Sendai Virus with Vesicles of Oligomerizable Lipids: A Micro Calorimetric Analysis of Membrane Fusion

    OpenAIRE

    Ravoo, Bart Jan; Weringa, Wilke D.; Engberts, Jan B.F.N.

    2000-01-01

    Sendai virus fuses efficiently with small and large unilamellar vesicles of the lipid 1,2-di-n-hexadecyloxypropyl-4-(beta-nitrostyryl) phosphate (DHPBNS) at pH 7.4 and 37°C, as shown by lipid mixing assays and electron microscopy. However, fusion is strongly inhibited by oligomerization of the head groups of DHPBNS in the bilayer vesicles. The enthalpy associated with fusion of Sendai virus with DHPBNS vesicles was measured by isothermal titration microcalorimetry, comparing titrations of Sen...

  8. Influenza Hemifusion Phenotype Depends on Membrane Context: Differences in Cell-Cell and Virus-Cell Fusion.

    Science.gov (United States)

    Zawada, Katarzyna E; Okamoto, Kenta; Kasson, Peter M

    2018-03-02

    Influenza viral entry into the host cell cytoplasm is accomplished by a process of membrane fusion mediated by the viral hemagglutinin protein. Hemagglutinin acts in a pH-triggered fashion, inserting a short fusion peptide into the host membrane followed by refolding of a coiled-coil structure to draw the viral envelope and host membranes together. Mutations to this fusion peptide provide an important window into viral fusion mechanisms and protein-membrane interactions. Here, we show that a well-described fusion peptide mutant, G1S, has a phenotype that depends strongly on the viral membrane context. The G1S mutant is well known to cause a "hemifusion" phenotype based on experiments in transfected cells, where cells expressing G1S hemagglutinin can undergo lipid mixing in a pH-triggered fashion similar to virus but will not support fusion pores. We compare fusion by the G1S hemagglutinin mutant expressed either in cells or in influenza virions and show that this hemifusion phenotype occurs in transfected cells but that native virions are able to support full fusion, albeit at a slower rate and 10-100× reduced infectious titer. We explain this with a quantitative model where the G1S mutant, instead of causing an absolute block of fusion, alters the protein stoichiometry required for fusion. This change slightly slows fusion at high hemagglutinin density, as on the viral surface, but at lower hemagglutinin density produces a hemifusion phenotype. The quantitative model thus reproduces the observed virus-cell and cell-cell fusion phenotypes, yielding a unified explanation where membrane context can control the observed viral fusion phenotype. Copyright © 2018 Elsevier Ltd. All rights reserved.

  9. Study on a multi-component palladium alloy membrane for the fusion fuel cycle

    International Nuclear Information System (INIS)

    Yoshida, Hiroshi; Okuno, Kenji; Nagasaki, Takanori; Noda, Kenji; Ishii, Yoshinobu; Takeshita, Hidefumi.

    1985-11-01

    A feasibility study on the material integrity with respect to the hydride formation and helium damage of the palladium alloy membrane was performed for an application of the palladium diffuser to a fusion fuel cleanup process. This study was conducted under the Japan/US Fusion Cooperation Program. Experimental works on the crystallography, hydrogen solubility and 3 He release characteristics were carried out with a multi-component palladium alloy(Pd-25Ag.Au.Ru). The excellent hydrogen permeability and mechanical properties of the membrane made of this alloy had been confirmed by authors' previous study. Based on the present study, this alloy membrane has high resistivity to the hydrogen embrittlement, and swelling and fracture due to the helium bubble formation under the practical operating conditions of the diffuser. (author)

  10. Fusion of Sendai Virus with Vesicles of Oligomerizable Lipids : A Micro Calorimetric Analysis of Membrane Fusion

    NARCIS (Netherlands)

    Ravoo, Bart Jan; Weringa, Wilke D.; Engberts, Jan B.F.N.

    2000-01-01

    Sendai virus fuses efficiently with small and large unilamellar vesicles of the lipid 1,2-di-n-hexadecyloxypropyl-4-(beta-nitrostyryl) phosphate (DHPBNS) at pH 7.4 and 37°C, as shown by lipid mixing assays and electron microscopy. However, fusion is strongly inhibited by oligomerization of the head

  11. Membrane support of accelerated fuel capsules for inertial fusion energy reactors

    International Nuclear Information System (INIS)

    Petzoldt, R.W.; Moir, R.W.

    1993-01-01

    The use of a thin membrane to suspend an (inertial fusion energy) fuel capsule in a holder for injection into a reactor chamber is investigated. Capsule displacement and membrane deformation angle are calculated for an axisymmetric geometry for a range of membrane strain and capsule size. This information is used to calculate maximum target accelerations. Membranes must be thin (perhaps of order one micron) to minimize their effect on capsule implosion symmetry. For example, a 5 μm thick cryogenic mylar membrane is calculated to allow 1,000 m/s 2 acceleration of a 3 mm radius, 100 mg capsule. Vibration analysis (for a single membrane support) shows that if membrane vibration is not deliberately minimized, allowed acceleration may be reduced by a factor of four. A two membrane alternative geometry would allow several times greater acceleration. Therefore, alternative membrane geometry's should be used to provide greater target acceleration potential and reduce capsule displacement within the holder (for a given membrane thickness)

  12. Sequential analysis of trans-SNARE formation in intracellular membrane fusion.

    Directory of Open Access Journals (Sweden)

    Kannan Alpadi

    2012-01-01

    Full Text Available SNARE complexes are required for membrane fusion in the endomembrane system. They contain coiled-coil bundles of four helices, three (Q(a, Q(b, and Q(c from target (t-SNAREs and one (R from the vesicular (v-SNARE. NSF/Sec18 disrupts these cis-SNARE complexes, allowing reassembly of their subunits into trans-SNARE complexes and subsequent fusion. Studying these reactions in native yeast vacuoles, we found that NSF/Sec18 activates the vacuolar cis-SNARE complex by selectively displacing the vacuolar Q(a SNARE, leaving behind a Q(bcR subcomplex. This subcomplex serves as an acceptor for a Q(a SNARE from the opposite membrane, leading to Q(a-Q(bcR trans-complexes. Activity tests of vacuoles with diagnostic distributions of inactivating mutations over the two fusion partners confirm that this distribution accounts for a major share of the fusion activity. The persistence of the Q(bcR cis-complex and the formation of the Q(a-Q(bcR trans-complex are both sensitive to the Rab-GTPase inhibitor, GDI, and to mutations in the vacuolar tether complex, HOPS (HOmotypic fusion and vacuolar Protein Sorting complex. This suggests that the vacuolar Rab-GTPase, Ypt7, and HOPS restrict cis-SNARE disassembly and thereby bias trans-SNARE assembly into a preferred topology.

  13. Sequential Analysis of Trans-SNARE Formation in Intracellular Membrane Fusion

    Science.gov (United States)

    Alpadi, Kannan; Kulkarni, Aditya; Comte, Veronique; Reinhardt, Monique; Schmidt, Andrea; Namjoshi, Sarita; Mayer, Andreas; Peters, Christopher

    2012-01-01

    SNARE complexes are required for membrane fusion in the endomembrane system. They contain coiled-coil bundles of four helices, three (Qa, Qb, and Qc) from target (t)-SNAREs and one (R) from the vesicular (v)-SNARE. NSF/Sec18 disrupts these cis-SNARE complexes, allowing reassembly of their subunits into trans-SNARE complexes and subsequent fusion. Studying these reactions in native yeast vacuoles, we found that NSF/Sec18 activates the vacuolar cis-SNARE complex by selectively displacing the vacuolar Qa SNARE, leaving behind a QbcR subcomplex. This subcomplex serves as an acceptor for a Qa SNARE from the opposite membrane, leading to Qa-QbcR trans-complexes. Activity tests of vacuoles with diagnostic distributions of inactivating mutations over the two fusion partners confirm that this distribution accounts for a major share of the fusion activity. The persistence of the QbcR cis-complex and the formation of the Qa-QbcR trans-complex are both sensitive to the Rab-GTPase inhibitor, GDI, and to mutations in the vacuolar tether complex, HOPS (HOmotypic fusion and vacuolar Protein Sorting complex). This suggests that the vacuolar Rab-GTPase, Ypt7, and HOPS restrict cis-SNARE disassembly and thereby bias trans-SNARE assembly into a preferred topology. PMID:22272185

  14. Autographa californica multiple nucleopolyhedrovirus GP64 protein: Analysis of domain I and V amino acid interactions and membrane fusion activity

    Energy Technology Data Exchange (ETDEWEB)

    Yu, Qianlong [State Key Laboratory of Crop Stress Biology for Arid Areas, Key Laboratory of Northwest Loess Plateau Crop Pest Management of Ministry of Agriculture, College of Plant Protection, Northwest A& F University, Yangling, Shaanxi 712100 (China); Blissard, Gary W. [Boyce Thompson Institute, Cornell University, Ithaca, NY 14853, United State (United States); Liu, Tong-Xian [State Key Laboratory of Crop Stress Biology for Arid Areas, Key Laboratory of Northwest Loess Plateau Crop Pest Management of Ministry of Agriculture, College of Plant Protection, Northwest A& F University, Yangling, Shaanxi 712100 (China); Li, Zhaofei, E-mail: zhaofeili73@outlook.com [State Key Laboratory of Crop Stress Biology for Arid Areas, Key Laboratory of Northwest Loess Plateau Crop Pest Management of Ministry of Agriculture, College of Plant Protection, Northwest A& F University, Yangling, Shaanxi 712100 (China)

    2016-01-15

    The Autographa californica multiple nucleopolyhedrovirus GP64 is a class III viral fusion protein. Although the post-fusion structure of GP64 has been solved, its pre-fusion structure and the detailed mechanism of conformational change are unknown. In GP64, domain V is predicted to interact with two domain I segments that flank fusion loop 2. To evaluate the significance of the amino acids involved in these interactions, we examined 24 amino acid positions that represent interacting and conserved residues within domains I and V. In several cases, substitution of a single amino acid involved in a predicted interaction disrupted membrane fusion activity, but no single amino acid pair appears to be absolutely required. We identified 4 critical residues in domain V (G438, W439, T452, and T456) that are important for membrane fusion, and two residues (G438 and W439) that appear to be important for formation or stability of the pre-fusion conformation of GP64. - Highlights: • The baculovirus envelope glycoprotein GP64 is a class III viral fusion protein. • The detailed mechanism of conformational change of GP64 is unknown. • We analyzed 24 positions that might stabilize the post-fusion structure of GP64. • We identified 4 residues in domain V that were critical for membrane fusion. • Two residues are critical for formation of the pre-fusion conformation of GP64.

  15. Autographa californica multiple nucleopolyhedrovirus GP64 protein: Analysis of domain I and V amino acid interactions and membrane fusion activity

    International Nuclear Information System (INIS)

    Yu, Qianlong; Blissard, Gary W.; Liu, Tong-Xian; Li, Zhaofei

    2016-01-01

    The Autographa californica multiple nucleopolyhedrovirus GP64 is a class III viral fusion protein. Although the post-fusion structure of GP64 has been solved, its pre-fusion structure and the detailed mechanism of conformational change are unknown. In GP64, domain V is predicted to interact with two domain I segments that flank fusion loop 2. To evaluate the significance of the amino acids involved in these interactions, we examined 24 amino acid positions that represent interacting and conserved residues within domains I and V. In several cases, substitution of a single amino acid involved in a predicted interaction disrupted membrane fusion activity, but no single amino acid pair appears to be absolutely required. We identified 4 critical residues in domain V (G438, W439, T452, and T456) that are important for membrane fusion, and two residues (G438 and W439) that appear to be important for formation or stability of the pre-fusion conformation of GP64. - Highlights: • The baculovirus envelope glycoprotein GP64 is a class III viral fusion protein. • The detailed mechanism of conformational change of GP64 is unknown. • We analyzed 24 positions that might stabilize the post-fusion structure of GP64. • We identified 4 residues in domain V that were critical for membrane fusion. • Two residues are critical for formation of the pre-fusion conformation of GP64.

  16. Regulation of membrane fusion and secretory events in the sea urchin embryo

    Energy Technology Data Exchange (ETDEWEB)

    Roe, J.L.

    1990-01-01

    Membrane fusion and secretory events play a key role in fertilization and early development in the sea urchin embryo. To investigate the mechanism of membrane fusion, the effect of inhibitors of metalloendoprotease activity was studied on two model systems of cell fusion; fertilization and spiculogenesis by primary mesenchyme cells in the embryo. Both the zinc chelator, 1,10-phenanthroline, and peptide metalloprotease substrates were found to inhibit both fertilization and gamete fusion, while peptides that are not substrates of metalloproteases did not affect either process. Primary mesenchyme cells form the larval skeleton in the embryo by deposition of mineral and an organic matrix into a syncytial cavity formed by fusion of filopodia of these cells. Metalloprotease inhibitors were found to inhibit spiculogenesis both in vivo and in cultures of isolated primary mesenchyme cells, and the activity of a metalloprotease of the appropriate specificity was found in the primary mesenchyme cells. These two studies implicate the activity of a metalloprotease in a necessary step in membrane fusion. Following fertilization, exocytosis of the cortical granules results in the formation of the fertilization envelope and the hyaline layer, that surround the developing embryo. The hatching enzyme is secreted by the blastula stage sea urchin embryo, which proteolyzes the fertilization envelope surrounding the embryo, allowing the embryo to hatch. Using an assay that measures {sup 125}I-fertilization envelope degradation, the hatching enzyme was identified as a 33 kDa metalloprotease, and was purified by ion-exchange and affinity chromatography from the hatching media of Strongylocentrotus purpuratus embryos. The hatching enzyme showed a substrate preference for only a minor subset of fertilization envelope proteins.

  17. Tri-membrane nanoparticles produced by combining liposome fusion and a novel patchwork of bicelles to overcome endosomal and nuclear membrane barriers to cargo delivery.

    Science.gov (United States)

    Yamada, Asako; Mitsueda, Asako; Hasan, Mahadi; Ueda, Miho; Hama, Susumu; Warashina, Shota; Nakamura, Takashi; Harashima, Hideyoshi; Kogure, Kentaro

    2016-03-01

    Membrane fusion is a rational strategy for crossing intracellular membranes that present barriers to liposomal nanocarrier-mediated delivery of plasmid DNA into the nucleus of non-dividing cells, such as dendritic cells. Based on this strategy, we previously developed nanocarriers consisting of a nucleic acid core particle coated with four lipid membranes [Akita, et al., Biomaterials, 2009, 30, 2940-2949]. However, including the endosomal membrane and two nuclear membranes, cells possess three intracellular membranous barriers. Thus, after entering the nucleus, nanoparticles coated with four membranes would still have one lipid membrane remaining, and could impede cargo delivery. Until now, coating a core particle with an odd number of lipid membranes was challenging. To produce nanocarriers with an odd number of lipid membranes, we developed a novel coating method involving lipid nano-discs, also known as bicelles, as a material for packaging DNA in a carrier with an odd number of lipid membranes. In this procedure, bicelles fuse to form an outer coating that resembles a patchwork quilt, which allows the preparation of nanoparticles coated with only three lipid membranes. Moreover, the transfection activity of dendritic cells with these three-membrane nanoparticles was higher than that for nanoparticles coated with four lipid membranes. In summary, we developed novel nanoparticles coated with an odd number of lipid membranes using the novel "patchwork-packaging method" to deliver plasmid DNA into the nucleus via membrane fusion.

  18. Sequential Conformational Changes in the Morbillivirus Attachment Protein Initiate the Membrane Fusion Process

    Science.gov (United States)

    Ader-Ebert, Nadine; Khosravi, Mojtaba; Herren, Michael; Avila, Mislay; Alves, Lisa; Bringolf, Fanny; Örvell, Claes; Langedijk, Johannes P.; Zurbriggen, Andreas; Plemper, Richard K.; Plattet, Philippe

    2015-01-01

    Despite large vaccination campaigns, measles virus (MeV) and canine distemper virus (CDV) cause major morbidity and mortality in humans and animals, respectively. The MeV and CDV cell entry system relies on two interacting envelope glycoproteins: the attachment protein (H), consisting of stalk and head domains, co-operates with the fusion protein (F) to mediate membrane fusion. However, how receptor-binding by the H-protein leads to F-triggering is not fully understood. Here, we report that an anti-CDV-H monoclonal antibody (mAb-1347), which targets the linear H-stalk segment 126-133, potently inhibits membrane fusion without interfering with H receptor-binding or F-interaction. Rather, mAb-1347 blocked the F-triggering function of H-proteins regardless of the presence or absence of the head domains. Remarkably, mAb-1347 binding to headless CDV H, as well as standard and engineered bioactive stalk-elongated CDV H-constructs treated with cells expressing the SLAM receptor, was enhanced. Despite proper cell surface expression, fusion promotion by most H-stalk mutants harboring alanine substitutions in the 126-138 “spacer” section was substantially impaired, consistent with deficient receptor-induced mAb-1347 binding enhancement. However, a previously reported F-triggering defective H-I98A variant still exhibited the receptor-induced “head-stalk” rearrangement. Collectively, our data spotlight a distinct mechanism for morbillivirus membrane fusion activation: prior to receptor contact, at least one of the morbillivirus H-head domains interacts with the membrane-distal “spacer” domain in the H-stalk, leaving the F-binding site located further membrane-proximal in the stalk fully accessible. This “head-to-spacer” interaction conformationally stabilizes H in an auto-repressed state, which enables intracellular H-stalk/F engagement while preventing the inherent H-stalk’s bioactivity that may prematurely activate F. Receptor-contact disrupts the

  19. Nuclear inner membrane fusion facilitated by yeast Jem1p is required for spindle pole body fusion but not for the first mitotic nuclear division during yeast mating.

    Science.gov (United States)

    Nishikawa, Shuh-ichi; Hirata, Aiko; Endo, Toshiya

    2008-11-01

    During mating of budding yeast, Saccharomyces cerevisiae, two haploid nuclei fuse to produce a diploid nucleus. The process of nuclear fusion requires two J proteins, Jem1p in the endoplasmic reticulum (ER) lumen and Sec63p, which forms a complex with Sec71p and Sec72p, in the ER membrane. Zygotes of mutants defective in the functions of Jem1p or Sec63p contain two haploid nuclei that were closely apposed but failed to fuse. Here we analyzed the ultrastructure of nuclei in jem1 Delta and sec71 Delta mutant zygotes using electron microscope with the freeze-substituted fixation method. Three-dimensional reconstitution of nuclear structures from electron microscope serial sections revealed that Jem1p facilitates nuclear inner-membrane fusion and spindle pole body (SPB) fusion while Sec71p facilitates nuclear outer-membrane fusion. Two haploid SPBs that failed to fuse could duplicate, and mitotic nuclear division of the unfused haploid nuclei started in jem1 Delta and sec71 Delta mutant zygotes. This observation suggests that nuclear inner-membrane fusion is required for SPB fusion, but not for SPB duplication in the first mitotic cell division.

  20. Outer nuclear membrane fusion of adjacent nuclei in varicella-zoster virus-induced syncytia.

    Science.gov (United States)

    Wang, Wei; Yang, Lianwei; Huang, Xiumin; Fu, Wenkun; Pan, Dequan; Cai, Linli; Ye, Jianghui; Liu, Jian; Xia, Ningshao; Cheng, Tong; Zhu, Hua

    2017-12-01

    Syncytia formation has been considered important for cell-to-cell spread and pathogenesis of many viruses. As a syncytium forms, individual nuclei often congregate together, allowing close contact of nuclear membranes and possibly fusion to occur. However, there is currently no reported evidence of nuclear membrane fusion between adjacent nuclei in wild-type virus-induced syncytia. Varicella-zoster virus (VZV) is one typical syncytia-inducing virus that causes chickenpox and shingles in humans. Here, we report, for the first time, an interesting observation of apparent fusion of the outer nuclear membranes from juxtaposed nuclei that comprise VZV syncytia both in ARPE-19 human epithelial cells in vitro and in human skin xenografts in the SCID-hu mouse model in vivo. This work reveals a novel aspect of VZV-related cytopathic effect in the context of multinucleated syncytia. Additionally, the information provided by this study could be helpful for future studies on interactions of viruses with host cell nuclei. Copyright © 2017 Elsevier Inc. All rights reserved.

  1. A molecular model for membrane fusion based on solution studies of an amphiphilic peptide from HIV gp41.

    OpenAIRE

    Fujii, G.; Horvath, S.; Woodward, S.; Eiserling, F.; Eisenberg, D.

    1992-01-01

    The mechanism of protein-mediated membrane fusion and lysis has been investigated by solution-state studies of the effects of peptides on liposomes. A peptide (SI) corresponding to a highly amphiphilic C-terminal segment from the envelope protein (gp41) of the human immunodeficiency virus (HIV) was synthesized and tested for its ability to cause lipid membranes to fuse together (fusion) or to break open (lysis). These effects were compared to those produced by the lytic and fusogenic peptide ...

  2. Semliki Forest virus produced in the absence of the 6K protein has an altered spike structure as revealed by decreased membrane fusion capacity

    NARCIS (Netherlands)

    McInerney, GM; Smit, JM; Liljestrom, P; Wilschut, J

    2004-01-01

    We examined the kinetics of membrane fusion of wild type (wt) and Delta6K mutant Semliki Forest virus in a liposomal model system. The final extent of membrane fusion of the mutant (at pH 5.5) was approximately one third that of the wt virus, although the level of E1 (fusion protein) trimerization

  3. A new combination of membranes and membrane reactors for improved tritium management in breeder blanket of fusion machines

    International Nuclear Information System (INIS)

    Demange, D.; Staemmler, S.; Kind, M.

    2011-01-01

    Tritium used as fuel in future fusion machines will be produced within the breeder blanket. The tritium extraction system recovers the tritium to be routed into the inner-fuel cycle of the machine. Accurate and precise tritium accountancy between both systems is mandatory to ensure a reliable operation. Handling in the blanket huge helium flow rates containing tritium as traces in molecular and oxide forms is challenging both for the process and the accountancy. Alternative tritium processes based on combinations of membranes and membrane reactors are proposed to facilitate the tritium management. The PERMCAT process is based on counter-current isotope swamping in a palladium membrane reactor. It allows recovering tritium efficiently from any chemical species. It produces a pure hydrogen stream enriched in tritium of advantage for integration upstream of the accountancy stage. A pre-separation and pre-concentration stage using new zeolite membranes has been studied to optimize the whole process. Such a combination could improve the tritium processes and facilitate accountancy in DEMO.

  4. SNARE-fusion mediated insertion of membrane proteins into native and artificial membranes.

    Science.gov (United States)

    Nordlund, Gustav; Brzezinski, Peter; von Ballmoos, Christoph

    2014-07-02

    Membrane proteins carry out functions such as nutrient uptake, ATP synthesis or transmembrane signal transduction. An increasing number of reports indicate that cellular processes are underpinned by regulated interactions between these proteins. Consequently, functional studies of these networks at a molecular level require co-reconstitution of the interacting components. Here, we report a SNARE protein-based method for incorporation of multiple membrane proteins into artificial membrane vesicles of well-defined composition, and for delivery of large water-soluble substrates into these vesicles. The approach is used for in vitro reconstruction of a fully functional bacterial respiratory chain from purified components. Furthermore, the method is used for functional incorporation of the entire F1F0 ATP synthase complex into native bacterial membranes from which this component had been genetically removed. The novel methodology offers a tool to investigate complex interaction networks between membrane-bound proteins at a molecular level, which is expected to generate functional insights into key cellular functions.

  5. Spatiotemporal dynamics of membrane remodeling and fusion proteins during endocytic transport.

    Science.gov (United States)

    Arlt, Henning; Auffarth, Kathrin; Kurre, Rainer; Lisse, Dominik; Piehler, Jacob; Ungermann, Christian

    2015-04-01

    Organelles of the endolysosomal system undergo multiple fission and fusion events to combine sorting of selected proteins to the vacuole with endosomal recycling. This sorting requires a consecutive remodeling of the organelle surface in the course of endosomal maturation. Here we dissect the remodeling and fusion machinery on endosomes during the process of endocytosis. We traced selected GFP-tagged endosomal proteins relative to exogenously added fluorescently labeled α-factor on its way from the plasma membrane to the vacuole. Our data reveal that the machinery of endosomal fusion and ESCRT proteins has similar temporal localization on endosomes, whereas they precede the retromer cargo recognition complex. Neither deletion of retromer nor the fusion machinery with the vacuole affects this maturation process, although the kinetics seems to be delayed due to ESCRT deletion. Of importance, in strains lacking the active Rab7-like Ypt7 or the vacuolar SNARE fusion machinery, α-factor still proceeds to late endosomes with the same kinetics. This indicates that endosomal maturation is mainly controlled by the early endosomal fusion and remodeling machinery but not the downstream Rab Ypt7 or the SNARE machinery. Our data thus provide important further understanding of endosomal biogenesis in the context of cargo sorting. © 2015 Arlt et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

  6. Membrane cholesterol regulates lysosome-plasma membrane fusion events and modulates Trypanosoma cruzi invasion of host cells.

    Directory of Open Access Journals (Sweden)

    Bárbara Hissa

    Full Text Available BACKGROUND: Trypomastigotes of Trypanosoma cruzi are able to invade several types of non-phagocytic cells through a lysosomal dependent mechanism. It has been shown that, during invasion, parasites trigger host cell lysosome exocytosis, which initially occurs at the parasite-host contact site. Acid sphingomyelinase released from lysosomes then induces endocytosis and parasite internalization. Lysosomes continue to fuse with the newly formed parasitophorous vacuole until the parasite is completely enclosed by lysosomal membrane, a process indispensable for a stable infection. Previous work has shown that host membrane cholesterol is also important for the T. cruzi invasion process in both professional (macrophages and non-professional (epithelial phagocytic cells. However, the mechanism by which cholesterol-enriched microdomains participate in this process has remained unclear. METHODOLOGY/PRINCIPAL FINDING: In the present work we show that cardiomyocytes treated with MβCD, a drug able to sequester cholesterol from cell membranes, leads to a 50% reduction in invasion by T. cruzi trypomastigotes, as well as a decrease in the number of recently internalized parasites co-localizing with lysosomal markers. Cholesterol depletion from host membranes was accompanied by a decrease in the labeling of host membrane lipid rafts, as well as excessive lysosome exocytic events during the earlier stages of treatment. Precocious lysosomal exocytosis in MβCD treated cells led to a change in lysosomal distribution, with a reduction in the number of these organelles at the cell periphery, and probably compromises the intracellular pool of lysosomes necessary for T. cruzi invasion. CONCLUSION/SIGNIFICANCE: Based on these results, we propose that cholesterol depletion leads to unregulated exocytic events, reducing lysosome availability at the cell cortex and consequently compromise T. cruzi entry into host cells. The results also suggest that two different pools of

  7. Coiled-coils on lipid membranes : a new perspective on membrane fusion

    NARCIS (Netherlands)

    Rabe, Martin

    2014-01-01

    For decades a large amount of research has dealt with membrane interactions of peptides and proteins as well as peptide-peptide interactions to understand the mechanisms of essential biological processes such as protein-driven vesicle budding and fission, cell penetration and lysis by peptides, and

  8. Mechanism of HIV-1 Resistance to an Electronically Constrained α-Helical Peptide Membrane Fusion Inhibitor.

    Science.gov (United States)

    Wu, Xiyuan; Liu, Zixuan; Ding, Xiaohui; Yu, Danwei; Wei, Huamian; Qin, Bo; Zhu, Yuanmei; Chong, Huihui; Cui, Sheng; He, Yuxian

    2018-04-01

    SC29EK is an electronically constrained α-helical peptide HIV-1 fusion inhibitor that is highly effective against both wild-type and enfuvirtide (T20)-resistant viruses. In this study, we focused on investigating the mechanism of HIV-1 resistance to SC29EK by two approaches. First, SC29EK-escaping HIV-1 variants were selected and characterized. Three mutant viruses, which possessed two (N43K/E49A) or three (Q39R/N43K/N126K and N43K/E49A/N126K) amino acid substitutions in the N- and C-terminal repeat regions of gp41 were identified as conferring high resistance to SC29EK and cross-resistance to the first-generation (T20 and C34) and newly designed (sifuvirtide, MT-SC29EK, and 2P23) fusion inhibitors. The resistance mutations could reduce the binding stability of SC29EK, impair viral Env-mediated cell fusion and entry, and change the conformation of the gp41 core structure. Further, we determined the crystal structure of SC29EK in complex with a target mimic peptide, which revealed the critical intra- and interhelical interactions underlying the mode of action of SC29EK and the genetic pathway to HIV-1 resistance. Taken together, the present data provide new insights into the structure and function of gp41 and the structure-activity relationship (SAR) of viral fusion inhibitors. IMPORTANCE T20 is the only membrane fusion inhibitor available for treatment of viral infection, but it has relatively low anti-HIV activity and genetic barriers for resistance, thus calling for new drugs blocking the viral fusion process. As an electronically constrained α-helical peptide, SC29EK is highly potent against both wild-type and T20-resistant HIV-1 strains. Here, we report the characterization of HIV-1 variants resistant to SC29EK and the crystal structure of SC29EK. The key mutations mediating high resistance to SC29EK and cross-resistance to the first and new generations of fusion inhibitors as well as the underlying mechanisms were identified. The crystal structure of SC29EK

  9. Ebola virus glycoprotein needs an additional trigger, beyond proteolytic priming for membrane fusion.

    Directory of Open Access Journals (Sweden)

    Shridhar Bale

    2011-11-01

    Full Text Available Ebolavirus belongs to the family filoviridae and causes severe hemorrhagic fever in humans with 50-90% lethality. Detailed understanding of how the viruses attach to and enter new host cells is critical to development of medical interventions. The virus displays a trimeric glycoprotein (GP(1,2 on its surface that is solely responsible for membrane attachment, virus internalization and fusion. GP(1,2 is expressed as a single peptide and is cleaved by furin in the host cells to yield two disulphide-linked fragments termed GP1 and GP2 that remain associated in a GP(1,2 trimeric, viral surface spike. After entry into host endosomes, GP(1,2 is enzymatically cleaved by endosomal cathepsins B and L, a necessary step in infection. However, the functional effects of the cleavage on the glycoprotein are unknown.We demonstrate by antibody binding and Hydrogen-Deuterium Exchange Mass Spectrometry (DXMS of glycoproteins from two different ebolaviruses that although enzymatic priming of GP(1,2 is required for fusion, the priming itself does not initiate the required conformational changes in the ectodomain of GP(1,2. Further, ELISA binding data of primed GP(1,2 to conformational antibody KZ52 suggests that the low pH inside the endosomes also does not trigger dissociation of GP1 from GP2 to effect membrane fusion.The results reveal that the ebolavirus GP(1,2 ectodomain remains in the prefusion conformation upon enzymatic cleavage in low pH and removal of the glycan cap. The results also suggest that an additional endosomal trigger is necessary to induce the conformational changes in GP(1,2 and effect fusion. Identification of this trigger will provide further mechanistic insights into ebolavirus infection.

  10. Fusion

    International Nuclear Information System (INIS)

    Naraghi, M.

    1976-01-01

    It is proposed that Iran as a world's potential supplier of fossile fuel should participate in fusion research and gain experience in this new field. Fusion, as an ultimate source of energy in future, and the problems concerned with the fusion reactors are reviewed. Furthermore; plasma heating, magnetic and inertial confinement in a fusion reactor are discussed. A brief description of tokamak, theta pinch and magnetic mirror reactors is also included

  11. Inhibition of EBV-mediated membrane fusion by anti-gHgL antibodies

    Energy Technology Data Exchange (ETDEWEB)

    Sathiyamoorthy, Karthik; Jiang, Jiansen; Möhl, Britta S.; Chen, Jia; Zhou, Z. Hong; Longnecker, Richard; Jardetzky, Theodore S. (UCLA); (Stanford-MED); (NWU)

    2017-09-22

    Herpesvirus entry into cells requires the coordinated action of multiple virus envelope glycoproteins, including gH, gL, and gB. For EBV, the gp42 protein assembles into complexes with gHgL heterodimers and binds HLA class II to activate gB-mediated membrane fusion with B cells. EBV tropism is dictated by gp42 levels in the virion, as it inhibits entry into epithelial cells while promoting entry into B cells. The gHgL and gB proteins are targets of neutralizing antibodies and potential candidates for subunit vaccine development, but our understanding of their neutralizing epitopes and the mechanisms of inhibition remain relatively unexplored. Here we studied the structures and mechanisms of two anti-gHgL antibodies, CL40 and CL59, that block membrane fusion with both B cells and epithelial cells. We determined the structures of the CL40 and CL59 complexes with gHgL using X-ray crystallography and EM to identify their epitope locations. CL59 binds to the C-terminal domain IV of gH, while CL40 binds to a site occupied by the gp42 receptor binding domain. CL40 binding to gHgL/gp42 complexes is not blocked by gp42 and does not interfere with gp42 binding to HLA class II, indicating that its ability to block membrane fusion with B cells represents a defect in gB activation. These data indicate that anti-gHgL neutralizing antibodies can block gHgL-mediated activation of gB through different surface epitopes and mechanisms.

  12. Preliminary design of fusion reactor fuel cleanup system by palladium alloy membrane method

    International Nuclear Information System (INIS)

    Yoshida, Hiroshi; Konishi, Satoshi; Naruse, Yuji

    1981-10-01

    A design of palladium diffuser and Fuel Cleanup System (FCU) for D-T fusion reactor is proposed. Feasibility of palladium alloy membrane method is discussed based on the early studies by the authors. Operating conditions of the palladium diffuser are determined experimentally. Dimensions of the diffuser are estimated from computer simulation. FCU system is designed under the feed conditions of Tritium Systems Test Assembly (TSTA) at Los Alamos Scientific Laboratory. The system is composed of Pd-diffusers, catalytic oxidizer, freezer and zink beds, and has some advantages in system layout and operation. This design can readily be extended to other conditions of plasma exhaust gases. (author)

  13. Extended Synaptotagmin 1 Interacts with Herpes Simplex Virus 1 Glycoprotein M and Negatively Modulates Virus-Induced Membrane Fusion.

    Science.gov (United States)

    El Kasmi, Imane; Khadivjam, Bita; Lackman, Miki; Duron, Johanne; Bonneil, Eric; Thibault, Pierre; Lippé, Roger

    2018-01-01

    Enveloped viruses typically encode their own fusion machinery to enter cells. Herpesviruses are unusual, as they fuse with a number of cellular compartments throughout their life cycles. As uncontrolled fusion of the host membranes should be avoided in these events, tight regulation of the viral fusion machinery is critical. While studying herpes simplex virus 1 (HSV-1) glycoprotein gM, we identified the cellular protein E-Syt1 (extended synaptotagmin 1) as an interaction partner. The interaction took place in both infected and transfected cells, suggesting other viral proteins were not required for the interaction. Most interestingly, E-Syt1 is a member of the synaptotagmin family of membrane fusion regulators. However, the protein is known to promote the tethering of the endoplasmic reticulum (ER) to the plasma membrane. We now show that E-Syt1, along with the related E-Syt3, negatively modulates viral release into the extracellular milieu, cell-to-cell viral spread, and viral entry, all processes that implicate membrane fusion events. Similarly, these E-Syt proteins impacted the formation of virus-induced syncytia. Altogether, these findings hint at the modulation of the viral fusion machinery by the E-Syt family of proteins. IMPORTANCE Viruses typically encode their own fusion apparatus to enable them to enter cells. For many viruses, this means a single fusogenic protein. However, herpesviruses are large entities that express several accessory viral proteins to regulate their fusogenic activity. The present study hints at the additional participation of cellular proteins in this process, suggesting the host can also modulate viral fusion to some extent. Hence E-Syt proteins 1 and 3 seem to negatively modulate the different viral fusion events that take place during the HSV-1 life cycle. This could represent yet another innate immunity response to the virus. Copyright © 2017 American Society for Microbiology.

  14. Fusion

    CERN Document Server

    Mahaffey, James A

    2012-01-01

    As energy problems of the world grow, work toward fusion power continues at a greater pace than ever before. The topic of fusion is one that is often met with the most recognition and interest in the nuclear power arena. Written in clear and jargon-free prose, Fusion explores the big bang of creation to the blackout death of worn-out stars. A brief history of fusion research, beginning with the first tentative theories in the early 20th century, is also discussed, as well as the race for fusion power. This brand-new, full-color resource examines the various programs currently being funded or p

  15. Reassessment of the lineage fusion hypothesis for the origin of double membrane bacteria.

    Science.gov (United States)

    Swithers, Kristen S; Fournier, Gregory P; Green, Anna G; Gogarten, J Peter; Lapierre, Pascal

    2011-01-01

    In 2009, James Lake introduced a new hypothesis in which reticulate phylogeny reconstruction is used to elucidate the origin of gram-negative bacteria (Nature 460: 967-971). The presented data supported the gram-negative bacteria originating from an ancient endosymbiosis between the Actinobacteria and Clostridia. His conclusion was based on a presence-absence analysis of protein families that divided all prokaryotes into five groups: Actinobacteria, Double Membrane bacteria (DM), Clostridia, Archaea and Bacilli. Of these five groups, the DM are by far the largest and most diverse group compared to the other groupings. While the fusion hypothesis for the origin of double membrane bacteria is enticing, we show that the signal supporting an ancient symbiosis is lost when the DM group is broken down into smaller subgroups. We conclude that the signal detected in James Lake's analysis in part results from a systematic artifact due to group size and diversity combined with low levels of horizontal gene transfer.

  16. A molecular model for membrane fusion based on solution studies of an amphiphilic peptide from HIV gp41.

    Science.gov (United States)

    Fujii, G; Horvath, S; Woodward, S; Eiserling, F; Eisenberg, D

    1992-11-01

    The mechanism of protein-mediated membrane fusion and lysis has been investigated by solution-state studies of the effects of peptides on liposomes. A peptide (SI) corresponding to a highly amphiphilic C-terminal segment from the envelope protein (gp41) of the human immunodeficiency virus (HIV) was synthesized and tested for its ability to cause lipid membranes to fuse together (fusion) or to break open (lysis). These effects were compared to those produced by the lytic and fusogenic peptide from bee venom, melittin. Other properties studied included the changes in visible absorbance and mean particle size, and the secondary structure of peptides as judged by CD spectroscopy. Taken together, the observations suggest that protein-mediated membrane fusion is dependent not only on hydrophobic and electrostatic forces but also on the spatial arrangement of the amino acid residues to form an amphiphilic structure that promotes the mixing of the lipids between membranes. A speculative molecular model is proposed for membrane fusion by alpha-helical peptides, and its relationship to the forces involved in protein-membrane interactions is discussed.

  17. Performance of a palladium membrane reactor using a Ni catalyst for fusion fuel impurities processing

    International Nuclear Information System (INIS)

    Willm, R.S.; Okuno, K.

    1994-01-01

    The palladium membrane reactor (PMR) provides a means to recover hydrogen isotopes from impurities expected to be present in fusion reactor exhaust. This recovery is based on reactions such as water gas shift and steam reforming for which conversion is equilibrium limited. By including a selectively permeable membrane such as Pd/Ag in the catalyst bed, hydrogen isotopes can be removed from the reacting environment, thus promoting the reaction to complete conversion. Such a device has been built and operated at the Tritium Systems Test Assembly (TSTA) at Los Alamos National Laboratory (LANL). For the reactions listed above, earlier study with this unit has shown that hydrogen single-pass recoveries approaching 100% can be achieved. It was also determined that a nickel catalyst is a feasible choice for use with a PMR appropriate for fusion fuel impurities processing. The purpose of this study is to systematically assess the performance of the PMR using a nickel catalyst. Reactions which will be studied include the water gas shift reaction, steam reforming and methane cracking. Extended operation will be performed to detect performance degradation if it exists. The use of methane in these tests may lead to the formation of coke on the catalyst. Methods of removing the coke such as treatment with carbon dioxide or diluted oxygen will be examined

  18. Cdc42p and Rho1p are sequentially activated and mechanistically linked to vacuole membrane fusion

    International Nuclear Information System (INIS)

    Logan, Michael R.; Jones, Lynden; Eitzen, Gary

    2010-01-01

    Small monomeric GTPases act as molecular switches, regulating many biological functions via activation of membrane localized signaling cascades. Activation of their switch function is controlled by GTP binding and hydrolysis. Two Rho GTPases, Cdc42p and Rho1p, are localized to the yeast vacuole where they regulate membrane fusion. Here, we define a method to directly examine vacuole membrane Cdc42p and Rho1p activation based on their affinity to probes derived from effectors. Cdc42p and Rho1p showed unique temporal activation which aligned with distinct subreactions of in vitro vacuole fusion. Cdc42p was rapidly activated in an ATP-independent manner while Rho1p activation was kinetically slower and required ATP. Inhibitors that are known to block vacuole membrane fusion were examined for their effect on Cdc42p and Rho1p activation. Rdi1p, which inhibits the dissociation of GDP from Rho proteins, blocked both Cdc42p and Rho1p activation. Ligands of PI(4,5)P 2 specifically inhibited Rho1p activation while pre-incubation with U73122, which targets Plc1p function, increased Rho1p activation. These results define unique activation mechanisms for Cdc42p and Rho1p, which may be linked to the vacuole membrane fusion mechanism.

  19. Coiled-coil formation of the membrane-fusion K/E peptides viewed by electron paramagnetic resonance.

    Directory of Open Access Journals (Sweden)

    Pravin Kumar

    Full Text Available The interaction of the complementary K (Ac-(KIAALKE3-GW-NH2 and E (Ac-(EIAALEK3-GY-NH2 peptides, components of the zipper of an artificial membrane fusion system (Robson Marsden H. et al. Angew Chemie Int Ed. 2009 is investigated by electron paramagnetic resonance (EPR. By frozen solution continuous-wave EPR and double electron-electron resonance (DEER, the distance between spin labels attached to the K- and to the E-peptide is measured. Three constructs of spin-labelled K- and E-peptides are used in five combinations for low temperature investigations. The K/E heterodimers are found to be parallel, in agreement with previous studies. Also, K homodimers in parallel orientation were observed, a finding that was not reported before. Comparison to room-temperature, solution EPR shows that the latter method is less specific to detect this peptide-peptide interaction. Combining frozen solution cw-EPR for short distances (1.8 nm to 2.0 nm and DEER for longer distances thus proves versatile to detect the zipper interaction in membrane fusion. As the methodology can be applied to membrane samples, the approach presented suggests itself for in-situ studies of the complete membrane fusion process, opening up new avenues for the study of membrane fusion.

  20. Membrane fusion inducers, chloroquine and spermidine increase lipoplex-mediated gene transfection

    International Nuclear Information System (INIS)

    Wong-Baeza, Carlos; Bustos, Israel; Serna, Manuel; Tescucano, Alonso; Alcantara-Farfan, Veronica; Ibanez, Miguel; Montanez, Cecilia; Wong, Carlos; Baeza, Isabel

    2010-01-01

    Gene transfection into mammalian cells can be achieved with viral and non-viral vectors. Non-viral vectors, such as cationic lipids that form lipoplexes with DNA, are safer and more stable than viral vectors, but their transfection efficiencies are lower. Here we describe that the simultaneous treatment with a membrane fusion inducer (chlorpromazine or procainamide) plus the lysosomotropic agent chloroquine increases lipoplex-mediated gene transfection in human (HEK293 and C-33 A) and rat (PC12) cell lines (up to 9.2-fold), as well as in situ in BALB/c mice spleens and livers (up to 6-fold); and that the polyamine spermidine increases lipoplex-mediated gene transfection and expression in cell cultures. The use of these four drugs provides a novel, safe and relatively inexpensive way to considerably increase lipoplex-mediated gene transfection efficiency.

  1. Early Events in Chikungunya Virus Infection—From Virus CellBinding to Membrane Fusion

    Directory of Open Access Journals (Sweden)

    Mareike K. S. van Duijl-Richter

    2015-07-01

    Full Text Available Chikungunya virus (CHIKV is a rapidly emerging mosquito-borne alphavirus causing millions of infections in the tropical and subtropical regions of the world. CHIKV infection often leads to an acute self-limited febrile illness with debilitating myalgia and arthralgia. A potential long-term complication of CHIKV infection is severe joint pain, which can last for months to years. There are no vaccines or specific therapeutics available to prevent or treat infection. This review describes the critical steps in CHIKV cell entry. We summarize the latest studies on the virus-cell tropism, virus-receptor binding, internalization, membrane fusion and review the molecules and compounds that have been described to interfere with virus cell entry. The aim of the review is to give the reader a state-of-the-art overview on CHIKV cell entry and to provide an outlook on potential new avenues in CHIKV research.

  2. Cysteines and N-Glycosylation Sites Conserved among All Alphaherpesviruses Regulate Membrane Fusion in Herpes Simplex Virus 1 Infection.

    Science.gov (United States)

    Rider, Paul J F; Naderi, Misagh; Bergeron, Scott; Chouljenko, Vladimir N; Brylinski, Michal; Kousoulas, Konstantin G

    2017-11-01

    Neurotropism is a defining characteristic of alphaherpesvirus pathogenicity. Glycoprotein K (gK) is a conserved virion glycoprotein of all alphaherpesviruses that is not found in other herpesvirus subfamilies. The extracellular amino terminus of gK has been shown to be important to the ability of the prototypic alphaherpesvirus herpes simplex virus 1 (HSV-1) to enter neurons via axonal termini. Here, we determined the role of the two conserved N-linked glycosylation (N48 and N58) sites of gK in virus-induced cell fusion and replication. We found that N-linked glycosylation is important to the regulation of HSV-1-induced membrane fusion since mutating N58 to alanine (N58A) caused extensive virus-induced cell fusion. Due to the known contributions of N-linked glycosylation to protein processing and correct disulfide bond formation, we investigated whether the conserved extracellular cysteine residues within the amino terminus of gK contributed to the regulation of HSV-1-induced membrane fusion. We found that mutation of C37 and C114 residues led to a gK-null phenotype characterized by very small plaque formation and drastic reduction in infectious virus production, while mutation of C82 and C243 caused extensive virus-induced cell fusion. Comparison of N-linked glycosylation and cysteine mutant replication kinetics identified disparate effects on infectious virion egress from infected cells. Specifically, cysteine mutations caused defects in the accumulation of infectious virus in both the cellular and supernatant fractions, while glycosylation site mutants did not adversely affect virion egress from infected cells. These results demonstrate a critical role for the N glycosylation sites and cysteines for the structure and function of the amino terminus of gK. IMPORTANCE We have previously identified important entry and neurotropic determinants in the amino terminus of HSV-1 glycoprotein K (gK). Alphaherpesvirus-mediated membrane fusion is a complex and highly

  3. Fusion between perinuclear virions and the outer nuclear membrane requires the fusogenic activity of herpes simplex virus gB.

    Science.gov (United States)

    Wright, Catherine C; Wisner, Todd W; Hannah, Brian P; Eisenberg, Roselyn J; Cohen, Gary H; Johnson, David C

    2009-11-01

    Herpesviruses cross nuclear membranes (NMs) in two steps, as follows: (i) capsids assemble and bud through the inner NM into the perinuclear space, producing enveloped virus particles, and (ii) the envelopes of these virus particles fuse with the outer NM. Two herpes simplex virus (HSV) glycoproteins, gB and gH (the latter, likely complexed as a heterodimer with gL), are necessary for the second step of this process. Mutants lacking both gB and gH accumulate in the perinuclear space or in herniations (membrane vesicles derived from the inner NM). Both gB and gH/gL are also known to act directly in fusing the virion envelope with host cell membranes during HSV entry into cells, i.e., both glycoproteins appear to function directly in different aspects of the membrane fusion process. We hypothesized that HSV gB and gH/gL also act directly in the membrane fusion that occurs during virus egress from the nucleus. Previous studies of the role of gB and gH/gL in nuclear egress involved HSV gB and gH null mutants that could potentially also possess gross defects in the virion envelope. Here, we produced recombinant HSV-expressing mutant forms of gB with single amino acid substitutions in the hydrophobic "fusion loops." These fusion loops are thought to play a direct role in membrane fusion by insertion into cellular membranes. HSV recombinants expressing gB with any one of four fusion loop mutations (W174R, W174Y, Y179K, and A261D) were unable to enter cells. Moreover, two of the mutants, W174Y and Y179K, displayed reduced abilities to mediate HSV cell-to-cell spread, and W174R and A261D exhibited no spread. All mutant viruses exhibited defects in nuclear egress, enveloped virions accumulated in herniations and in the perinuclear space, and fewer enveloped virions were detected on cell surfaces. These results support the hypothesis that gB functions directly to mediate the fusion between perinuclear virus particles and the outer NM.

  4. Different sets of ER-resident J-proteins regulate distinct polar nuclear-membrane fusion events in Arabidopsis thaliana.

    Science.gov (United States)

    Maruyama, Daisuke; Yamamoto, Masaya; Endo, Toshiya; Nishikawa, Shuh-ichi

    2014-11-01

    Angiosperm female gametophytes contain a central cell with two polar nuclei. In many species, including Arabidopsis thaliana, the polar nuclei fuse during female gametogenesis. We previously showed that BiP, an Hsp70 in the endoplasmic reticulum (ER), was essential for membrane fusion during female gametogenesis. Hsp70 function requires partner proteins for full activity. J-domain containing proteins (J-proteins) are the major Hsp70 functional partners. A. thaliana ER contains three soluble J-proteins, AtERdj3A, AtERdj3B, and AtP58(IPK). Here, we analyzed mutants of these proteins and determined that double-mutant ovules lacking AtP58(IPK) and AtERdj3A or AtERdj3B were defective in polar nuclear fusion. Electron microscopy analysis identified that polar nuclei were in close contact, but no membrane fusion occurred in mutant ovules lacking AtP58(IPK) and AtERdj3A. The polar nuclear outer membrane appeared to be connected via the ER remaining at the inner unfused membrane in mutant ovules lacking AtP58(IPK) and AtERdj3B. These results indicate that ER-resident J-proteins, AtP58(IPK)/AtERdj3A and AtP58(IPK)/AtERdj3B, function at distinct steps of polar nuclear-membrane fusion. Similar to the bip1 bip2 double mutant female gametophytes, the aterdj3a atp58(ipk) double mutant female gametophytes defective in fusion of the outer polar nuclear membrane displayed aberrant endosperm proliferation after fertilization with wild-type pollen. However, endosperm proliferated normally after fertilization of the aterdj3b atp58(ipk) double mutant female gametophytes defective in fusion of the inner membrane. Our results indicate that the polar nuclear fusion defect itself does not cause an endosperm proliferation defect. © The Author 2014. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  5. Fusion

    Science.gov (United States)

    Herman, Robin

    1990-10-01

    The book abounds with fascinating anecdotes about fusion's rocky path: the spurious claim by Argentine dictator Juan Peron in 1951 that his country had built a working fusion reactor, the rush by the United States to drop secrecy and publicize its fusion work as a propaganda offensive after the Russian success with Sputnik; the fortune Penthouse magazine publisher Bob Guccione sank into an unconventional fusion device, the skepticism that met an assertion by two University of Utah chemists in 1989 that they had created "cold fusion" in a bottle. Aimed at a general audience, the book describes the scientific basis of controlled fusion--the fusing of atomic nuclei, under conditions hotter than the sun, to release energy. Using personal recollections of scientists involved, it traces the history of this little-known international race that began during the Cold War in secret laboratories in the United States, Great Britain and the Soviet Union, and evolved into an astonishingly open collaboration between East and West.

  6. Using Haloarcula marismortui bacteriorhodopsin as a fusion tag for enhancing and visible expression of integral membrane proteins in Escherichia coli.

    Directory of Open Access Journals (Sweden)

    Min-Feng Hsu

    Full Text Available Membrane proteins are key targets for pharmacological intervention because of their vital functions. Structural and functional studies of membrane proteins have been severely hampered because of the difficulties in producing sufficient quantities of properly folded and biologically active proteins. Here we generate a high-level expression system of integral membrane proteins in Escherichia coli by using a mutated bacteriorhodopsin (BR from Haloarcula marismortui (HmBRI/D94N as a fusion partner. A purification strategy was designed by incorporating a His-tag on the target membrane protein for affinity purification and an appropriate protease cleavage site to generate the final products. The fusion system can be used to detect the intended target membrane proteins during overexpression and purification either with the naked eye or by directly monitoring their characteristic optical absorption. In this study, we applied this approach to produce two functional integral membrane proteins, undecaprenyl pyrophosphate phosphatase and carnitine/butyrobetaine antiporter with significant yield enhancement. This technology could facilitate the development of a high-throughput strategy to screen for conditions that improve the yield of correctly folded target membrane proteins. Other robust BRs can also be incorporated in this system.

  7. Complementation between avirulent Newcastle disease virus and a fusion protein gene expressed from a retrovirus vector: requirements for membrane fusion.

    OpenAIRE

    Morrison, T; McQuain, C; McGinnes, L

    1991-01-01

    The cDNA derived from the fusion gene of the virulent AV strain of Newcastle disease virus (NDV) was expressed in chicken embryo cells by using a retrovirus vector. The fusion protein expressed in this system was transported to the cell surface and was efficiently cleaved into the disulfide-linked F1-F2 form found in infectious virions. The cells expressing the fusion gene grew normally and could be passaged many times. Monolayers of these cells would plaque, in the absence of trypsin, avirul...

  8. Reassessment of the lineage fusion hypothesis for the origin of double membrane bacteria.

    Directory of Open Access Journals (Sweden)

    Kristen S Swithers

    Full Text Available In 2009, James Lake introduced a new hypothesis in which reticulate phylogeny reconstruction is used to elucidate the origin of gram-negative bacteria (Nature 460: 967-971. The presented data supported the gram-negative bacteria originating from an ancient endosymbiosis between the Actinobacteria and Clostridia. His conclusion was based on a presence-absence analysis of protein families that divided all prokaryotes into five groups: Actinobacteria, Double Membrane bacteria (DM, Clostridia, Archaea and Bacilli. Of these five groups, the DM are by far the largest and most diverse group compared to the other groupings. While the fusion hypothesis for the origin of double membrane bacteria is enticing, we show that the signal supporting an ancient symbiosis is lost when the DM group is broken down into smaller subgroups. We conclude that the signal detected in James Lake's analysis in part results from a systematic artifact due to group size and diversity combined with low levels of horizontal gene transfer.

  9. Direct targeting of membrane fusion by SNARE mimicry: Convergent evolution of Legionella effectors.

    Science.gov (United States)

    Shi, Xingqi; Halder, Partho; Yavuz, Halenur; Jahn, Reinhard; Shuman, Howard A

    2016-08-02

    Legionella pneumophila, the Gram-negative pathogen causing Legionnaires' disease, infects host cells by hijacking endocytic pathways and forming a Legionella-containing vacuole (LCV) in which the bacteria replicate. To promote LCV expansion and prevent lysosomal targeting, effector proteins are translocated into the host cell where they alter membrane traffic. Here we show that three of these effectors [LegC2 (Legionella eukaryotic-like gene C2)/YlfB (yeast lethal factor B), LegC3, and LegC7/YlfA] functionally mimic glutamine (Q)-SNARE proteins. In infected cells, the three proteins selectively form complexes with the endosomal arginine (R)-SNARE vesicle-associated membrane protein 4 (VAMP4). When reconstituted in proteoliposomes, these proteins avidly fuse with liposomes containing VAMP4, resulting in a stable complex with properties resembling canonical SNARE complexes. Intriguingly, however, the LegC/SNARE hybrid complex cannot be disassembled by N-ethylmaleimide-sensitive factor. We conclude that LegCs use SNARE mimicry to divert VAMP4-containing vesicles for fusion with the LCV, thus promoting its expansion. In addition, the LegC/VAMP4 complex avoids the host's disassembly machinery, thus effectively trapping VAMP4 in an inactive state.

  10. Fusion proteins and select lipids cooperate as membrane receptors for the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) Vam7p.

    Science.gov (United States)

    Karunakaran, Vidya; Wickner, William

    2013-10-04

    Vam7p, the vacuolar soluble Qc-SNARE, is essential for yeast vacuole fusion. The large tethering complex, homotypic fusion and vacuole protein sorting complex (HOPS), and phosphoinositides, which interact with the Vam7p PX domain, have each been proposed to serve as its membrane receptors. Studies with the isolated organelle cannot determine whether these receptor elements suffice and whether ligands or mutations act directly or indirectly on Vam7p binding to the membrane. Using pure components that are active in reconstituted vacuolar fusion, we now find that Vam7p binds to membranes through its combined affinities for several vacuolar membrane constituents: HOPS, phosphatidylinositol 3-phosphate, SNAREs, and acidic phospholipids. Acidic lipids allow low concentrations of Vam7p to suffice for fusion; without acidic lipids, the block to fusion is partially bypassed by high concentrations of Vam7p.

  11. Crystal Structure of the Membrane Fusion Protein CusB from Escherichia coli

    Energy Technology Data Exchange (ETDEWEB)

    Su, Chih-Chia; Yang, Feng; Long, Feng; Reyon, Deepak; Routh, Mathew D.; Kuo, Dennis W.; Mokhtari, Adam K.; Van Ornam, Jonathan D.; Rabe, Katherine L.; Hoy, Julie A.; Lee, Young Jin; Rajashankar, Kanagalaghatta R.; Yu, Edward W.; (Cornell); (Iowa State)

    2010-03-29

    Gram-negative bacteria, such as Escherichia coli, frequently utilize tripartite efflux complexes belonging to the resistance-nodulation-division family to expel diverse toxic compounds from the cell. These systems contain a periplasmic membrane fusion protein (MFP) that is critical for substrate transport. We here present the x-ray structures of the CusB MFP from the copper/silver efflux system of E. coli. This is the first structure of any MFPs associated with heavy-metal efflux transporters. CusB bridges the inner-membrane efflux pump CusA and outer-membrane channel CusC to mediate resistance to Cu{sup +} and Ag{sup +} ions. Two distinct structures of the elongated molecules of CusB were found in the asymmetric unit of a single crystal, which suggests the flexible nature of this protein. Each protomer of CusB can be divided into four different domains, whereby the first three domains are mostly {beta}-strands and the last domain adopts an entirely helical architecture. Unlike other known structures of MFPs, the {alpha}-helical domain of CusB is folded into a three-helix bundle. This three-helix bundle presumably interacts with the periplasmic domain of CusC. The N- and C-termini of CusB form the first {beta}-strand domain, which is found to interact with the periplasmic domain of the CusA efflux pump. Atomic details of how this efflux protein binds Cu{sup +} and Ag{sup +} were revealed by the crystals of the CusB-Cu(I) and CusB-Ag(I) complexes. The structures indicate that CusB consists of multiple binding sites for these metal ions. These findings reveal novel structural features of an MFP in the resistance-nodulation-division efflux system and provide direct evidence that this protein specifically interacts with transported substrates.

  12. Susceptibility to virus-cell fusion at the plasma membrane is reduced through expression of HIV gp41 cytoplasmic domains

    International Nuclear Information System (INIS)

    Malinowsky, Katharina; Luksza, Julia; Dittmar, Matthias T.

    2008-01-01

    The cytoplasmic tail of the HIV transmembrane protein plays an important role in viral infection. In this study we analyzed the role of retroviral cytoplasmic tails in modulating the cytoskeleton and interfering with virus-cell fusion. HeLaP4 cells expressing different HIV cytoplasmic tail constructs showed reduced acetylated tubulin levels whereas the cytoplasmic tail of MLV did not alter microtubule stability indicating a unique function for the lentiviral cytoplasmic tail. The effect on tubulin is mediated through the membrane proximal region of the HIV cytoplasmic tail and was independent of membrane localization. Site-directed mutagenesis identified three motifs in the HIV-2 cytoplasmic tail required to effect the reduction in acetylated tubulin. Both the YxxΦ domain and amino acids 21 to 45 of the HIV-2 cytoplasmic tail need to be present to change the level of acetylated tubulin in transfected cells. T-cells stably expressing one HIV-2 cytoplasmic tail derived construct showed also a reduction in acetylated tubulin thus confirming the importance of this effect not only for HeLaP4 and 293T cells. Challenge experiments using transiently transfected HeLaP4 cells and T cells stably expressing an HIV cytoplasmic tail construct revealed both reduced virus-cell fusion and replication of HIV-1 NL4.3 compared to control cells. In the virus-cell fusion assay only virions pseudotyped with either HIV or MLV envelopes showed reduced fusion efficiency, whereas VSV-G pseudotyped virions where not affected by the expression of HIV derived cytoplasmic tail constructs, indicating that fusion at the plasma but not endosomal membrane is affected. Overexpression of human histone-deacetylase 6 (HDAC6) and constitutively active RhoA resulted in a reduction of acetylated tubulin and reduced virus-cell fusion as significant as that observed following expression of HIV cytoplasmic tail constructs. Inhibition of HDAC6 showed a strong increase in acetylated tubulin and increase of

  13. MITO-Porter: A liposome-based carrier system for delivery of macromolecules into mitochondria via membrane fusion.

    Science.gov (United States)

    Yamada, Yuma; Akita, Hidetaka; Kamiya, Hiroyuki; Kogure, Kentaro; Yamamoto, Takenori; Shinohara, Yasuo; Yamashita, Kikuji; Kobayashi, Hideo; Kikuchi, Hiroshi; Harashima, Hideyoshi

    2008-02-01

    Mitochondria are the principal producers of energy in higher cells. Mitochondrial dysfunction is implicated in a variety of human diseases, including cancer and neurodegenerative disorders. Effective medical therapies for such diseases will ultimately require targeted delivery of therapeutic proteins or nucleic acids to the mitochondria, which will be achieved through innovations in the nanotechnology of intracellular trafficking. Here we describe a liposome-based carrier that delivers its macromolecular cargo to the mitochondrial interior via membrane fusion. These liposome particles, which we call MITO-Porters, carry octaarginine surface modifications to stimulate their entry into cells as intact vesicles (via macropinocytosis). We identified lipid compositions for the MITO-Porter which promote both its fusion with the mitochondrial membrane and the release of its cargo to the intra-mitochondrial compartment in living cells. Thus, the MITO-Porter holds promise as an efficacious system for the delivery of both large and small therapeutic molecules into mitochondria.

  14. Reciprocal complementation of bovine parainfluenza virus type 3 lacking either the membrane or fusion gene.

    Science.gov (United States)

    Takada, Marina; Matsuura, Ryosuke; Kokuho, Takehiro; Tsuboi, Takamitsu; Kameyama, Ken-Ichiro; Takeuchi, Kaoru

    2017-11-01

    Two defective bovine parainfluenza virus type 3 (BPIV3) strains were generated, one lacking the membrane (M) protein gene and expressing EGFP (ΔM-EGFP) and the other lacking the fusion (F) protein gene and expressing mStrawberry (ΔF-mSB), by supplying deficient proteins in trans. When Madin-Darby bovine kidney (MDBK) cells were co-infected with ΔM-EGFP and ΔF-mSB at a multiplicity of infection (MOI) of 0.1, complemented viruses were easily obtained. Complemented viruses grew as efficiently as wild-type BPIV3 and could be passaged in MDBK cell cultures even at an MOI of 0.01, possibly due to multiploid virus particles containing genomes of both ΔM-EGFP and ΔF-mSB. This reciprocal complementation method using two defective viruses would be useful to express large or multiple proteins in cell cultures using paramyxovirus vectors. Copyright © 2017 Elsevier B.V. All rights reserved.

  15. Anti-tumor effects of gene therapy with GALV membrane fusion glycoprotein in lung adenocarcinoma.

    Science.gov (United States)

    Zhu, Bing; Yang, Jian-ru; Fu, Xin-ping; Jiang, Yue-quan

    2014-07-01

    This study examined the efficacy of gene therapy of lung adenocarcinoma using specifically controlled type I herpes simplex virus recombinant vector expressing Gibbon ape leukemia virus membrane fusion glycoprotein gene (GALV.fus). Recombinant HSV-I plasmid carrying target transgene was constructed, and recombinant viral vector was generated in Vero cells using Lipofectamine transfection. Viral vector was introduced into lung adenocarcinoma A549 cells or human fetal fibroblast HFL-I GNHu 5 cells, or inoculated into human lung adenocarcinoma xenografts in nude mice. The anti-tumor and cytotoxic effects of GALV-FMG, the transgene, were examined in these cell and animal models. Expression of GALV-FMG in xenographs achieved 100 % tumorigenicity. Recombinant HSV-I viral vector also exhibited significant tumor cell killing effect in vitro. Relative survival rates of tumor cells treated with GALV-FMG or control vectors were, respectively, 20 and 70 %. GALV.fus has a potent anti-tumor effect against lung cancer both in vitro and in vivo. This anti-tumor potential provides foundation for further studies with this vector.

  16. Convenient synthesis and application of versatile nucleic acid lipid membrane anchors in the assembly and fusion of liposomes

    DEFF Research Database (Denmark)

    Ries, Oliver; Löffler, Philipp M. G.; Vogel, Stefan

    2015-01-01

    or the construction of DNA origami structures. We herein present the synthesis and applications of versatile lipid membrane anchor building blocks suitable for solid phase oligonucleotide synthesis. These are readily synthesized in bulk in five to seven steps from commercially available precursors and can...... be incorporated at any position within an oligonucleotide without significantly altering duplex stability and structure as proven by thermal denaturation experiments and circular dichroism. Furthermore, applicability could be demonstrated by assembly and fusion of liposomes mediated by lipid...

  17. Vesicular PtdIns(3,4,5)P3 and Rab7 are key effectors of sea urchin zygote nuclear membrane fusion.

    Science.gov (United States)

    Lete, Marta G; Byrne, Richard D; Alonso, Alicia; Poccia, Dominic; Larijani, Banafshé

    2017-01-15

    Regulation of nuclear envelope dynamics is an important example of the universal phenomena of membrane fusion. The signalling molecules involved in nuclear membrane fusion might also be conserved during the formation of both pronuclear and zygote nuclear envelopes in the fertilised egg. Here, we determine that class-I phosphoinositide 3-kinases (PI3Ks) are needed for in vitro nuclear envelope formation. We show that, in vivo, PtdIns(3,4,5)P 3 is transiently located in vesicles around the male pronucleus at the time of nuclear envelope formation, and around male and female pronuclei before membrane fusion. We illustrate that class-I PI3K activity is also necessary for fusion of the female and male pronuclear membranes. We demonstrate, using coincidence amplified Förster resonance energy transfer (FRET) monitored using fluorescence lifetime imaging microscopy (FLIM), a protein-lipid interaction of Rab7 GTPase and PtdIns(3,4,5)P 3 that occurs during pronuclear membrane fusion to create the zygote nuclear envelope. We present a working model, which includes several molecular steps in the pathways controlling fusion of nuclear envelope membranes. © 2017. Published by The Company of Biologists Ltd.

  18. Determination of the topology of endoplasmic reticulum membrane proteins using redox-sensitive green-fluorescence protein fusions.

    Science.gov (United States)

    Tsachaki, Maria; Birk, Julia; Egert, Aurélie; Odermatt, Alex

    2015-07-01

    Membrane proteins of the endoplasmic reticulum (ER) are involved in a wide array of essential cellular functions. Identification of the topology of membrane proteins can provide significant insight into their mechanisms of action and biological roles. This is particularly important for membrane enzymes, since their topology determines the subcellular site where a biochemical reaction takes place and the dependence on luminal or cytosolic co-factor pools and substrates. The methods currently available for the determination of topology of proteins are rather laborious and require post-lysis or post-fixation manipulation of cells. In this work, we have developed a simple method for defining intracellular localization and topology of ER membrane proteins in living cells, based on the fusion of the respective protein with redox-sensitive green-fluorescent protein (roGFP). We validated the method and demonstrated that roGFP fusion proteins constitute a reliable tool for the study of ER membrane protein topology, using as control microsomal 11β-hydroxysteroid dehydrogenase (11β-HSD) proteins whose topology has been resolved, and comparing with an independent approach. We then implemented this method to determine the membrane topology of six microsomal members of the 17β-hydroxysteroid dehydrogenase (17β-HSD) family. The results revealed a luminal orientation of the catalytic site for three enzymes, i.e. 17β-HSD6, 7 and 12. Knowledge of the intracellular location of the catalytic site of these enzymes will enable future studies on their biological functions and on the role of the luminal co-factor pool. Copyright © 2015 Elsevier B.V. All rights reserved.

  19. The Structure of Herpesvirus Fusion Glycoprotein B-Bilayer Complex Reveals the Protein-Membrane and Lateral Protein-Protein Interaction

    NARCIS (Netherlands)

    Maurer, Ulrike E.; Zeev-Ben-Mordehai, Tzviya; Pandurangan, Arun Prasad; Cairns, Tina M.; Hannah, Brian P.; Whitbeck, J. Charles; Eisenberg, Roselyn J.; Cohen, Gary H.; Topf, Maya; Huiskonen, Juha T.; Gruenewald, Kay

    2013-01-01

    Glycoprotein B (gB) is a key component of the complex herpesvirus fusion machinery. We studied membrane interaction of two gB ectodomain forms and present an electron cryotomography structure of the gB-bilayer complex. The two forms differed in presence or absence of the membrane proximal region

  20. Molecular dynamics analysis of conformational change of paramyxovirus F protein during the initial steps of membrane fusion

    International Nuclear Information System (INIS)

    Martín-García, Fernando; Mendieta-Moreno, Jesús Ignacio; Mendieta, Jesús; Gómez-Puertas, Paulino

    2012-01-01

    Highlights: ► Initial conformational change of paramyxovirus F protein is caused only by mechanical forces. ► HRA region undergoes a structural change from a beta + alpha conformation to an extended coil and then to an all-alpha conformation. ► HRS domains of F protein form three single α-helices prior to generation of the coiled coil. -- Abstract: The fusion of paramyxovirus to the cell membrane is mediated by fusion protein (F protein) present in the virus envelope, which undergoes a dramatic conformational change during the process. Unlike hemagglutinin in orthomyxovirus, this change is not mediated by an alteration of environmental pH, and its cause remains unknown. Steered molecular dynamics analysis leads us to suggest that the conformational modification is mediated only by stretching mechanical forces once the transmembrane fusion peptide of the protein is anchored to the cell membrane. Such elongating forces will generate major secondary structure rearrangement in the heptad repeat A region of the F protein; from β-sheet conformation to an elongated coil and then spontaneously to an α-helix. In addition, it is proposed that the heptad repeat A region adopts a final three-helix coiled coil and that this structure appears after the formation of individual helices in each monomer.

  1. Conservation of proteo-lipid nuclear membrane fusion machinery during early embryogenesis.

    Science.gov (United States)

    Byrne, Richard D; Veeriah, Selvaraju; Applebee, Christopher J; Larijani, Banafshé

    2014-01-01

    The fusogenic lipid diacylglycerol is essential for remodeling gamete and zygote nuclear envelopes (NE) during early embryogenesis. It is unclear whether upstream signaling molecules are likewise conserved. Here we demonstrate PLCγ and its activator SFK1, which co-operate during male pronuclear envelope formation, also promote the subsequent male and female pronuclear fusion. PLCγ and SFK1 interact directly at the fusion site leading to PLCγ activation. This is accompanied by a spatially restricted reduction of PtdIns(4,5)P2. Consequently, pronuclear fusion is blocked by PLCγ or SFK1 inhibition. These findings identify new regulators of events in the early embryo and suggest a conserved "toolkit" of fusion machinery drives successive NE fusion events during embryogenesis.

  2. Targeting of a chimeric human histone fusion mRNA to membrane-bound polysomes in HeLa cells

    International Nuclear Information System (INIS)

    Zambetti, G.; Stein, J.; Stein, G.

    1987-01-01

    The subcellular location of histone mRNA-containing polysomes may play a key role in the posttranscriptional events that mediate histone mRNA turnover following inhibition of DNA synthesis. Previously, it has been shown that histone mRNA is found primarily on free polysomes that are associated with the cytoskeleton. The authors report here the construction of an Escherichia coli pBR322 β-lactamase signal peptide-human H3 histone fusion gene. The fusion transcript is targeted to membrane-bound polysomes and remains stable following interruption of DNA replication. Relocating mRNA within the cell may provide a procedure for studying the posttranscriptional regulation of gene expression

  3. NSF- and SNARE-mediated membrane fusion is required for nuclear envelope formation and completion of nuclear pore complex assembly in Xenopus laevis egg extracts.

    Science.gov (United States)

    Baur, Tina; Ramadan, Kristijan; Schlundt, Andreas; Kartenbeck, Jürgen; Meyer, Hemmo H

    2007-08-15

    Despite the progress in understanding nuclear envelope (NE) reformation after mitosis, it has remained unclear what drives the required membrane fusion and how exactly this is coordinated with nuclear pore complex (NPC) assembly. Here, we show that, like other intracellular fusion reactions, NE fusion in Xenopus laevis egg extracts is mediated by SNARE proteins that require activation by NSF. Antibodies against Xenopus NSF, depletion of NSF or the dominant-negative NSF(E329Q) variant specifically inhibited NE formation. Staging experiments further revealed that NSF was required until sealing of the envelope was completed. Moreover, excess exogenous alpha-SNAP that blocks SNARE function prevented membrane fusion and caused accumulation of non-flattened vesicles on the chromatin surface. Under these conditions, the nucleoporins Nup107 and gp210 were fully recruited, whereas assembly of FxFG-repeat-containing nucleoporins was blocked. Together, we define NSF- and SNARE-mediated membrane fusion events as essential steps during NE formation downstream of Nup107 recruitment, and upstream of membrane flattening and completion of NPC assembly.

  4. Membrane pumping technology, helium and hydrogen isotopes separation in the fusion hydrogen

    International Nuclear Information System (INIS)

    Pigarov, A.Yu.; Pistunovich, V.I.; Busnyuk, A.O.

    1994-01-01

    A gas pumping system for the ITER, improved by implementation of superpermeable membranes for selective hydrogen isotope exhaust, is considered. The study of the pumping capability of a niobium membrane for a hydrogen-helium mixture has been fulfilled. The membrane superpermeability can be only realized for atomic hydrogen. Helium does not pass through the membrane, and its presence does not affect the hydrogen pumping. A detailed Monte Carlo simulation of gas behavior for the experimental facility has been done. The probability of permeation for a hydrogen atom for one collision with the membrane is ∼0.1; the same probability of molecule permeation is ∼10 -5 . The probability for atomization, i.e. re-emission of an atomizer is ∼0.2; the probability of recombination of an atom is ∼0.2

  5. Gβγ directly modulates vesicle fusion by competing with synaptotagmin for binding to neuronal SNARE proteins embedded in membranes.

    Science.gov (United States)

    Zurawski, Zack; Page, Brian; Chicka, Michael C; Brindley, Rebecca L; Wells, Christopher A; Preininger, Anita M; Hyde, Karren; Gilbert, James A; Cruz-Rodriguez, Osvaldo; Currie, Kevin P M; Chapman, Edwin R; Alford, Simon; Hamm, Heidi E

    2017-07-21

    G i/o -coupled G protein-coupled receptors can inhibit neurotransmitter release at synapses via multiple mechanisms. In addition to Gβγ-mediated modulation of voltage-gated calcium channels (VGCC), inhibition can also be mediated through the direct interaction of Gβγ subunits with the soluble N -ethylmaleimide attachment protein receptor (SNARE) complex of the vesicle fusion apparatus. Binding studies with soluble SNARE complexes have shown that Gβγ binds to both ternary SNARE complexes, t-SNARE heterodimers, and monomeric SNAREs, competing with synaptotagmin 1(syt1) for binding sites on t-SNARE. However, in secretory cells, Gβγ, SNAREs, and synaptotagmin interact in the lipid environment of a vesicle at the plasma membrane. To approximate this environment, we show that fluorescently labeled Gβγ interacts specifically with lipid-embedded t-SNAREs consisting of full-length syntaxin 1 and SNAP-25B at the membrane, as measured by fluorescence polarization. Fluorescently labeled syt1 undergoes competition with Gβγ for SNARE-binding sites in lipid environments. Mutant Gβγ subunits that were previously shown to be more efficacious at inhibiting Ca 2+ -triggered exocytotic release than wild-type Gβγ were also shown to bind SNAREs at a higher affinity than wild type in a lipid environment. These mutant Gβγ subunits were unable to inhibit VGCC currents. Specific peptides corresponding to regions on Gβ and Gγ shown to be important for the interaction disrupt the interaction in a concentration-dependent manner. In in vitro fusion assays using full-length t- and v-SNAREs embedded in liposomes, Gβγ inhibited Ca 2+ /synaptotagmin-dependent fusion. Together, these studies demonstrate the importance of these regions for the Gβγ-SNARE interaction and show that the target of Gβγ, downstream of VGCC, is the membrane-embedded SNARE complex. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  6. Refining the Mechanisms of Heniparvirus-Mediated Membrane Fusion Through Mutagenesis of Hendra virus Envelope Glycoproteins

    Science.gov (United States)

    2007-09-06

    glycoprotein single point mutants…..67 Figure 9: Effects of multiple point mutations on fusion activity of HeV F……….……..69 Figure 10: Effects of multiple ...relapsing encephalitis (28). How or whether this latter manifestation of disease is at all analogous to Subacute Sclerosing Panencephalitis (SSPE), a...total DNA per T-25cm2 flask overnight followed by infection with wild-type vaccinia virus (strain WR) at a multiplicity of infection (MOI) of 10. At

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

    NARCIS (Netherlands)

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

    1988-01-01

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

  8. A high throughput Cre–lox activated viral membrane fusion assay identifies pharmacological inhibitors of HIV entry

    International Nuclear Information System (INIS)

    Esposito, Anthony M.; Cheung, Pamela; Swartz, Talia H.; Li, Hongru; Tsibane, Tshidi; Durham, Natasha D.; Basler, Christopher F.; Felsenfeld, Dan P.; Chen, Benjamin K.

    2016-01-01

    Enveloped virus entry occurs when viral and cellular membranes fuse releasing particle contents into the target cell. Human immunodeficiency virus (HIV) entry occurs by cell-free virus or virus transferred between infected and uninfected cells through structures called virological synapses. We developed a high-throughput cell-based assay to identify small molecule inhibitors of cell-free or virological synapse-mediated entry. An HIV clone carrying Cre recombinase as a Gag-internal gene fusion releases active Cre into cells upon viral entry activating a recombinatorial gene switch changing dsRed to GFP-expression. A screen of a 1998 known-biological profile small molecule library identified pharmacological HIV entry inhibitors that block both cell-free and cell-to-cell infection. Many top hits were noted as HIV inhibitors in prior studies, but not previously recognized as entry antagonists. Modest therapeutic indices for simvastatin and nigericin were observed in confirmatory HIV infection assays. This robust assay is adaptable to study HIV and heterologous viral pseudotypes. - Highlights: • Cre recombinase viral fusion assay screens cell-free or cell–cell entry inhibitors. • This Gag-iCre based assay is specific for the entry step of HIV replication. • Screened a library of known pharmacologic compounds for HIV fusion antagonists. • Many top hits were previously noted as HIV inhibitors, but here are classified as entry antagonists. Many top hits were previously noted as HIV inhibitors, but not as entry antagonists. • The assay is compatible with pseudotyping with HIV and heterologous viruses.

  9. A high throughput Cre–lox activated viral membrane fusion assay identifies pharmacological inhibitors of HIV entry

    Energy Technology Data Exchange (ETDEWEB)

    Esposito, Anthony M. [Division of Infectious Diseases, Department of Medicine, Icahn School of Medicine at Mount Sinai, Immunology Institute, New York, NY (United States); Cheung, Pamela [Integrated Screening Core, Icahn School of Medicine at Mount Sinai, New York, NY (United States); Swartz, Talia H.; Li, Hongru [Division of Infectious Diseases, Department of Medicine, Icahn School of Medicine at Mount Sinai, Immunology Institute, New York, NY (United States); Tsibane, Tshidi [Department of Microbiology, Icahn School of Medicine at Mount Sinai, New York, NY (United States); Durham, Natasha D. [Division of Infectious Diseases, Department of Medicine, Icahn School of Medicine at Mount Sinai, Immunology Institute, New York, NY (United States); Basler, Christopher F. [Department of Microbiology, Icahn School of Medicine at Mount Sinai, New York, NY (United States); Felsenfeld, Dan P. [Integrated Screening Core, Icahn School of Medicine at Mount Sinai, New York, NY (United States); Chen, Benjamin K., E-mail: benjamin.chen@mssm.edu [Division of Infectious Diseases, Department of Medicine, Icahn School of Medicine at Mount Sinai, Immunology Institute, New York, NY (United States)

    2016-03-15

    Enveloped virus entry occurs when viral and cellular membranes fuse releasing particle contents into the target cell. Human immunodeficiency virus (HIV) entry occurs by cell-free virus or virus transferred between infected and uninfected cells through structures called virological synapses. We developed a high-throughput cell-based assay to identify small molecule inhibitors of cell-free or virological synapse-mediated entry. An HIV clone carrying Cre recombinase as a Gag-internal gene fusion releases active Cre into cells upon viral entry activating a recombinatorial gene switch changing dsRed to GFP-expression. A screen of a 1998 known-biological profile small molecule library identified pharmacological HIV entry inhibitors that block both cell-free and cell-to-cell infection. Many top hits were noted as HIV inhibitors in prior studies, but not previously recognized as entry antagonists. Modest therapeutic indices for simvastatin and nigericin were observed in confirmatory HIV infection assays. This robust assay is adaptable to study HIV and heterologous viral pseudotypes. - Highlights: • Cre recombinase viral fusion assay screens cell-free or cell–cell entry inhibitors. • This Gag-iCre based assay is specific for the entry step of HIV replication. • Screened a library of known pharmacologic compounds for HIV fusion antagonists. • Many top hits were previously noted as HIV inhibitors, but here are classified as entry antagonists. Many top hits were previously noted as HIV inhibitors, but not as entry antagonists. • The assay is compatible with pseudotyping with HIV and heterologous viruses.

  10. Solid-state NMR spectroscopy of the HIV gp41 membrane fusion protein supports intermolecular antiparallel β sheet fusion peptide structure in the final six-helix bundle state.

    Science.gov (United States)

    Sackett, Kelly; Nethercott, Matthew J; Zheng, Zhaoxiong; Weliky, David P

    2014-03-06

    The HIV gp41 protein catalyzes fusion between viral and target cell membranes. Although the ~20-residue N-terminal fusion peptide (FP) region is critical for fusion, the structure of this region is not well characterized in large gp41 constructs that model the gp41 state at different times during fusion. This paper describes solid-state NMR (SSNMR) studies of FP structure in a membrane-associated construct (FP-Hairpin), which likely models the final fusion state thought to be thermostable trimers with six-helix bundle structure in the region C-terminal of the FP. The SSNMR data show that there are populations of FP-Hairpin with either α helical or β sheet FP conformation. For the β sheet population, measurements of intermolecular (13)C-(13)C proximities in the FP are consistent with a significant fraction of intermolecular antiparallel β sheet FP structure with adjacent strand crossing near L7 and F8. There appears to be negligible in-register parallel structure. These findings support assembly of membrane-associated gp41 trimers through interleaving of N-terminal FPs from different trimers. Similar SSNMR data are obtained for FP-Hairpin and a construct containing the 70 N-terminal residues of gp41 (N70), which is a model for part of the putative pre-hairpin intermediate state of gp41. FP assembly may therefore occur at an early fusion stage. On a more fundamental level, similar SSNMR data are obtained for FP-Hairpin and a construct containing the 34 N-terminal gp41 residues (FP34) and support the hypothesis that the FP is an autonomous folding domain. Copyright © 2013 Elsevier Ltd. All rights reserved.

  11. Reversible acid-induced inactivation of the membrane fusion protein of Semliki Forest virus

    NARCIS (Netherlands)

    Waarts, BL; Smit, JM; Aneke, OJC; McInerney, GM; Liljestrom, P; Bittman, R; Wilschut, J

    Previously, it has been shown that the exposure of Semliki Forest virus (SFV) to a mildly acidic environment induces a rapid and complete loss of the ability of the virus to bind and fuse to target membranes added subsequently. In the present study, incubation of SFV at low pH followed by a specific

  12. Huntingtin is required for ER-to-Golgi transport and for secretory vesicle fusion at the plasma membrane

    Directory of Open Access Journals (Sweden)

    Hemma Brandstaetter

    2014-12-01

    Full Text Available Huntingtin is a large membrane-associated scaffolding protein that associates with endocytic and exocytic vesicles and modulates their trafficking along cytoskeletal tracks. Although the progression of Huntington’s disease is linked to toxic accumulation of mutant huntingtin protein, loss of wild-type huntingtin function might also contribute to neuronal cell death, but its precise function is not well understood. Therefore, we investigated the molecular role of huntingtin in exocytosis and observed that huntingtin knockdown in HeLa cells causes a delay in endoplasmic reticulum (ER-to-Golgi transport and a reduction in the number of cargo vesicles leaving the trans-Golgi network. In addition, we found that huntingtin is required for secretory vesicle fusion at the plasma membrane. Similar defects in the early exocytic pathway were observed in primary fibroblasts from homozygous Htt140Q/140Q knock-in mice, which have the expansion inserted into the mouse huntingtin gene so lack wild-type huntingtin expression. Interestingly, heterozygous fibroblasts from a Huntington’s disease patient with a 180Q expansion displayed no obvious defects in the early secretory pathway. Thus, our results highlight the requirement for wild-type huntingtin at distinct steps along the secretory pathway.

  13. Herpes Simplex Virus 1 Glycoprotein M and the Membrane-Associated Protein UL11 Are Required for Virus-Induced Cell Fusion and Efficient Virus Entry

    Science.gov (United States)

    Kim, In-Joong; Chouljenko, Vladimir N.; Walker, Jason D.

    2013-01-01

    Herpes simplex virus 1 (HSV-1) facilitates virus entry into cells and cell-to-cell spread by mediating fusion of the viral envelope with cellular membranes and fusion of adjacent cellular membranes. Although virus strains isolated from herpetic lesions cause limited cell fusion in cell culture, clinical herpetic lesions typically contain large syncytia, underscoring the importance of cell-to-cell fusion in virus spread in infected tissues. Certain mutations in glycoprotein B (gB), gK, UL20, and other viral genes drastically enhance virus-induced cell fusion in vitro and in vivo. Recent work has suggested that gB is the sole fusogenic glycoprotein, regulated by interactions with the viral glycoproteins gD, gH/gL, and gK, membrane protein UL20, and cellular receptors. Recombinant viruses were constructed to abolish either gM or UL11 expression in the presence of strong syncytial mutations in either gB or gK. Virus-induced cell fusion caused by deletion of the carboxyl-terminal 28 amino acids of gB or the dominant syncytial mutation in gK (Ala to Val at amino acid 40) was drastically reduced in the absence of gM. Similarly, syncytial mutations in either gB or gK did not cause cell fusion in the absence of UL11. Neither the gM nor UL11 gene deletion substantially affected gB, gC, gD, gE, and gH glycoprotein synthesis and expression on infected cell surfaces. Two-way immunoprecipitation experiments revealed that the membrane protein UL20, which is found as a protein complex with gK, interacted with gM while gM did not interact with other viral glycoproteins. Viruses produced in the absence of gM or UL11 entered into cells more slowly than their parental wild-type virus strain. Collectively, these results indicate that gM and UL11 are required for efficient membrane fusion events during virus entry and virus spread. PMID:23678175

  14. Fusion Machinery

    DEFF Research Database (Denmark)

    Sørensen, Jakob Balslev; Milosevic, Ira

    2015-01-01

    SNARE proteins constitute the minimal machinery needed for membrane fusion. SNAREs operate by forming a complex, which pulls the lipid bilayers into close contact and provides the mechanical force needed for lipid bilayer fusion. At the chemical synapse, SNARE-complex formation between...... the vesicular SNARE VAMP2/synaptobrevin-2 and the target (plasma membrane) SNAREs SNAP25 and syntaxin-1 results in fusion and release of neurotransmitter, synchronized to the electrical activity of the cell by calcium influx and binding to synaptotagmin. Formation of the SNARE complex is tightly regulated...... and appears to start with syntaxin-1 bound to an SM (Sec1/Munc18-like) protein. Proteins of the Munc13-family are responsible for opening up syntaxin and allowing sequential binding of SNAP-25 and VAMP2/synaptobrevin-2. N- to C-terminal “zippering” of the SNARE domains leads to membrane fusion...

  15. Reassessment of the Lineage Fusion Hypothesis for the Origin of Double Membrane Bacteria

    OpenAIRE

    Swithers, Kristen S.; Fournier, Gregory P.; Green, Anna G.; Gogarten, J. Peter; Lapierre, Pascal

    2011-01-01

    In 2009, James Lake introduced a new hypothesis in which reticulate phylogeny reconstruction is used to elucidate the origin of gram-negative bacteria (Nature 460: 967-971). The presented data supported the gram-negative bacteria originating from an ancient endosymbiosis between the Actinobacteria and Clostridia. His conclusion was based on a presence-absence analysis of protein families that divided all prokaryotes into five groups: Actinobacteria, Double Membrane bacteria (DM), Clostridia, ...

  16. Mutations in the Fusion Protein of Measles Virus That Confer Resistance to the Membrane Fusion Inhibitors Carbobenzoxy-d-Phe-l-Phe-Gly and 4-Nitro-2-Phenylacetyl Amino-Benzamide.

    Science.gov (United States)

    Ha, Michael N; Delpeut, Sébastien; Noyce, Ryan S; Sisson, Gary; Black, Karen M; Lin, Liang-Tzung; Bilimoria, Darius; Plemper, Richard K; Privé, Gilbert G; Richardson, Christopher D

    2017-12-01

    The inhibitors carbobenzoxy (Z)-d-Phe-l-Phe-Gly (fusion inhibitor peptide [FIP]) and 4-nitro-2-phenylacetyl amino-benzamide (AS-48) have similar efficacies in blocking membrane fusion and syncytium formation mediated by measles virus (MeV). Other homologues, such as Z-d-Phe, are less effective but may act through the same mechanism. In an attempt to map the site of action of these inhibitors, we generated mutant viruses that were resistant to the inhibitory effects of Z-d-Phe-l-Phe-Gly. These 10 mutations were localized to the heptad repeat B (HRB) region of the fusion protein, and no changes were observed in the viral hemagglutinin, which is the receptor attachment protein. Mutations were validated in a luciferase-based membrane fusion assay, using transfected fusion and hemagglutinin expression plasmids or with syncytium-based assays in Vero, Vero-SLAM, and Vero-Nectin 4 cell lines. The changes I452T, D458N, D458G/V459A, N462K, N462H, G464E, and I483R conferred resistance to both FIP and AS-48 without compromising membrane fusion. The inhibitors did not block hemagglutinin protein-mediated binding to the target cell. Edmonston vaccine/laboratory and IC323 wild-type strains were equally affected by the inhibitors. Escape mutations were mapped upon a three-dimensional (3D) structure modeled from the published crystal structure of parainfluenzavirus 5 fusion protein. The most effective mutations were situated in a region located near the base of the globular head and its junction with the alpha-helical stalk of the prefusion protein. We hypothesize that the fusion inhibitors could interfere with the structural changes that occur between the prefusion and postfusion conformations of the fusion protein. IMPORTANCE Due to lapses in vaccination worldwide that have caused localized outbreaks, measles virus (MeV) has regained importance as a pathogen. Antiviral agents against measles virus are not commercially available but could be useful in conjunction with Me

  17. A single amino acid substitution modulates low-pH-triggered membrane fusion of GP64 protein in Autographa californica and Bombyx mori nucleopolyhedroviruses

    International Nuclear Information System (INIS)

    Katou, Yasuhiro; Yamada, Hayato; Ikeda, Motoko; Kobayashi, Michihiro

    2010-01-01

    We have previously shown that budded viruses of Bombyx mori nucleopolyhedrovirus (BmNPV) enter the cell cytoplasm but do not migrate into the nuclei of non-permissive Sf9 cells that support a high titer of Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV) multiplication. Here we show, using the syncytium formation assay, that low-pH-triggered membrane fusion of BmNPV GP64 protein (Bm-GP64) is significantly lower than that of AcMNPV GP64 protein (Ac-GP64). Mutational analyses of GP64 proteins revealed that a single amino acid substitution between Ac-GP64 H155 and Bm-GP64 Y153 can have significant positive or negative effects on membrane fusion activity. Studies using bacmid-based GP64 recombinant AcMNPV harboring point-mutated ac-gp64 and bm-gp64 genes showed that Ac-GP64 H155Y and Bm-GP64 Y153H substitutions decreased and increased, respectively, the multiplication and cell-to-cell spread of progeny viruses. These results indicate that Ac-GP64 H155 facilitates the low-pH-triggered membrane fusion reaction between virus envelopes and endosomal membranes.

  18. The Yeast ATP-binding Cassette (ABC) Transporter Ycf1p Enhances the Recruitment of the Soluble SNARE Vam7p to Vacuoles for Efficient Membrane Fusion*

    Science.gov (United States)

    Sasser, Terry L.; Lawrence, Gus; Karunakaran, Surya; Brown, Christopher; Fratti, Rutilio A.

    2013-01-01

    The Saccharomyces cerevisiae vacuole contains five ATP-binding cassette class C (ABCC) transporters, including Ycf1p, a family member that was originally characterized as a Cd2+ transporter. Ycf1p has also been found to physically interact with a wide array of proteins, including factors that regulate vacuole homeostasis. In this study, we examined the role of Ycf1p and other ABCC transporters in the regulation of vacuole homotypic fusion. We found that deletion of YCF1 attenuated in vitro vacuole fusion by up to 40% relative to wild-type vacuoles. Plasmid-expressed wild-type Ycf1p rescued the deletion phenotype; however, Ycf1p containing a mutation of the conserved Lys-669 to Met in the Walker A box of the first nucleotide-binding domain (Ycf1pK669M) was unable to complement the fusion defect of ycf1Δ vacuoles. This indicates that the ATPase activity of Ycf1p is required for its function in regulating fusion. In addition, we found that deleting YCF1 caused a striking decrease in vacuolar levels of the soluble SNARE Vam7p, whereas total cellular levels were not altered. The attenuated fusion of ycf1Δ vacuoles was rescued by the addition of recombinant Vam7p to in vitro experiments. Thus, Ycf1p contributes in the recruitment of Vam7p to the vacuole for efficient membrane fusion. PMID:23658021

  19. A soluble form of Epstein-Barr virus gH/gL inhibits EBV-induced membrane fusion and does not function in fusion

    Energy Technology Data Exchange (ETDEWEB)

    Rowe, Cynthia L. [Department of Microbiology and Immunology, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611 (United States); Connolly, Sarah A. [Department of Health Sciences, DePaul University, Chicago, IL 60614 (United States); Chen, Jia [Department of Microbiology and Immunology, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611 (United States); Jardetzky, Theodore S. [Department of Structural Biology, Stanford University School of Medicine, 371 Serra Mall, Stanford, CA 94305 (United States); Longnecker, Richard, E-mail: r-longnecker@northwestern.edu [Department of Microbiology and Immunology, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611 (United States)

    2013-02-05

    We investigated whether soluble EBV gH/gL (sgH/gL) functions in fusion and made a series of truncations of gH/gL domains based on the gH/gL crystal structure. We found sgH/gL failed to mediate cell-cell fusion both when co-expressed with the other entry glycoproteins and when added exogenously to fusion assays. Interestingly, sgH/gL inhibited cell-cell fusion in a dose dependent manner when co-expressed. sgH/gL from HSV was unable to inhibit EBV fusion, suggesting the inhibition was specific to EBV gH/gL. sgH/gL stably binds gp42, but not gB nor gH/gL. The domain mutants, DI/gL, DI-II/gL and DI-II-III/gL were unable to bind gp42. Instead, DI-II/gL, DI-II-III/gL and sgH/gL but not DI/gL decreased the expression of gp42, resulting in decreased overall fusion. Overall, our results suggest that domain IV may be required for proper folding and the transmembrane domain and cytoplasmic tail of EBV gH/gL are required for the most efficient fusion.

  20. Palmitoylation of SARS-CoV S protein is necessary for partitioning into detergent-resistant membranes and cell-cell fusion but not interaction with M protein

    International Nuclear Information System (INIS)

    McBride, Corrin E.; Machamer, Carolyn E.

    2010-01-01

    Coronaviruses are enveloped RNA viruses that generally cause mild disease in humans. However, the recently emerged coronavirus that caused severe acute respiratory syndrome (SARS-CoV) is the most pathogenic human coronavirus discovered to date. The SARS-CoV spike (S) protein mediates virus entry by binding cellular receptors and inducing fusion between the viral envelope and the host cell membrane. Coronavirus S proteins are palmitoylated, which may affect function. Here, we created a non-palmitoylated SARS-CoV S protein by mutating all nine cytoplasmic cysteine residues. Palmitoylation of SARS-CoV S was required for partitioning into detergent-resistant membranes and for cell-cell fusion. Surprisingly, however, palmitoylation of S was not required for interaction with SARS-CoV M protein. This contrasts with the requirement for palmitoylation of mouse hepatitis virus S protein for interaction with M protein and may point to important differences in assembly and infectivity of these two coronaviruses.

  1. Establishment of insect cell lines expressing green fluorescent protein on cell surface based on AcMNPV GP64 membrane fusion characteristic.

    Science.gov (United States)

    Qi, Ben-Xiang; Chen, Ying-Jian; Su, Rui; Li, Yi-Fei; Zheng, Gui-Ling; Li, Chang-You

    2017-10-01

    Displaying a protein on the surface of cells has been provided a very successful strategy to function research of exogenous proteins. Based on the membrane fusion characteristic of Autographa californica multiple nucleopolyhedrovirus envelope protein GP64, we amplified and cloned N-terminal signal peptide and C-terminal transmembrane domain as well as cytoplasmic tail domain of gp64 gene into vector pIZ/V5-His with multi-cloning sites to construct the cell surface expression vector pIZ/V5-gp64. To verify that the vector can be used to express proteins on the membrane of insect cells, a recombinant plasmid pIZ/V5-gp64-GFP was constructed by introducing the PCR amplified green fluorescent protein (GFP) gene and transfected into insect cell lines Sf9 and H5. The transected cells were screened with zeocin and cell cloning. PCR verification results showed that the GFP gene was successfully integrated into these cells. Green fluorescence in Sf9-GFP and H5-GFP cells was observed by using confocal laser scanning microscopy and immunofluorescence detection indicated that GFP protein was located on the cell membrane. Western blot results showed that a fusion protein GP64-GFP of about 40 kDa was expressed on the membrane of Sf9-GFP and H5-GFP cells. The expression system constructed in this paper can be used for localization and continuous expression of exogenous proteins on insect cell membrane.

  2. Unusual Self-Assembly of the Recombinant Chlamydia trachomatis Major Outer Membrane Protein-Based Fusion Antigen CTH522 Into Protein Nanoparticles

    DEFF Research Database (Denmark)

    Rose, Fabrice; Karlsen, Kasper; Jensen, Pernille

    2018-01-01

    but is a challenging vaccine candidate by being an integral membrane protein, and the immunogenicity depends on a correctly folded structure. We investigated the biophysical properties of the recombinant MOMP-based fusion antigen CTH522, which is tested in early human clinical trials. It consists of a truncated......Sexually transmitted Chlamydia trachomatis (Ct) infects more than 100 million people annually, and untreated chlamydia infections can cause severe complications. Therefore, there is an urgent need for a chlamydia vaccine. The Ct major outer membrane protein (MOMP) is highly immunogenic...

  3. Hexameric assembly of membrane fusion protein YknX of the sporulation delaying efflux pump from Bacillus amyloliquefaciens.

    Science.gov (United States)

    Xu, Yongbin; Jo, Inseong; Wang, Lulu; Chen, Jinli; Fan, Shengdi; Dong, Yuesheng; Quan, Chunshan; Ha, Nam-Chul

    2017-11-04

    Membrane fusion proteins (MFPs) play an essential role in the action of the drug efflux pumps and protein secretion systems in bacteria. The sporulation delaying protein (SDP) efflux pump YknWXYZ has been identified in diverse Bacillus species. The MFP YknX requires the ATP-binding cassette (ABC) transporter YknYZ and the Yip1 family protein YknW to form a functional complex. To date, the crystal structure, molecular function and mechanism of action of YknX remain unknown. In this study, to characterize the structural and biochemical roles of YknX in the functional assembly of YknWXYZ from B. amyloliquefaciens, we successfully obtained crystals of the YknX protein that diffracted X-rays to a resolution of 4.4 Å. We calculated an experimentally phased map using single-wavelength anomalous diffraction (SAD), revealing that YknX forms a hexameric assembly similar to that of MacA from Gram-negative bacteria. The hexameric assembly of YknX exhibited a funnel-like structure with a central channel and a conical mouth. Functional studies in vitro suggest that YknX can bind directly to peptidoglycan. Our study provides an improved understanding of the assembly of the YknWXYZ efflux pump and the role of YknX in the complex. Copyright © 2017 Elsevier Inc. All rights reserved.

  4. Mitochondrial matrix delivery using MITO-Porter, a liposome-based carrier that specifies fusion with mitochondrial membranes

    Energy Technology Data Exchange (ETDEWEB)

    Yasuzaki, Yukari; Yamada, Yuma [Laboratory for Molecular Design of Pharmaceutics, Faculty of Pharmaceutical Sciences, Hokkaido University, Kita-12, Nishi-6, Kita-ku, Sapporo 060-0812 (Japan); Harashima, Hideyoshi, E-mail: harasima@pharm.hokudai.ac.jp [Laboratory for Molecular Design of Pharmaceutics, Faculty of Pharmaceutical Sciences, Hokkaido University, Kita-12, Nishi-6, Kita-ku, Sapporo 060-0812 (Japan)

    2010-06-25

    Mitochondria are the principal producers of energy in cells of higher organisms. It was recently reported that mutations and defects in mitochondrial DNA (mtDNA) are associated with various mitochondrial diseases including a variety of neurodegenerative and neuromuscular diseases. Therefore, an effective mitochondrial gene therapy and diagnosis would be expected to have great medical benefits. To achieve this, therapeutic agents need to be delivered into the innermost mitochondrial space (mitochondrial matrix), which contains the mtDNA pool. We previously reported on the development of MITO-Porter, a liposome-based carrier that introduces macromolecular cargos into mitochondria via membrane fusion. In this study, we provide a demonstration of mitochondrial matrix delivery and the visualization of mitochondrial genes (mtDNA) in living cells using the MITO-Porter. We first prepared MITO-Porter containing encapsulated propidium iodide (PI), a fluorescent dye used to stain nucleic acids to detect mtDNA. We then confirmed the emission of red-fluorescence from PI by conjugation with mtDNA, when the carriers were incubated in the presence of isolated rat liver mitochondria. Finally, intracellular observation by confocal laser scanning microscopy clearly verified that the MITO-Porter delivered PI to the mitochondrial matrix.

  5. Mitochondrial matrix delivery using MITO-Porter, a liposome-based carrier that specifies fusion with mitochondrial membranes

    International Nuclear Information System (INIS)

    Yasuzaki, Yukari; Yamada, Yuma; Harashima, Hideyoshi

    2010-01-01

    Mitochondria are the principal producers of energy in cells of higher organisms. It was recently reported that mutations and defects in mitochondrial DNA (mtDNA) are associated with various mitochondrial diseases including a variety of neurodegenerative and neuromuscular diseases. Therefore, an effective mitochondrial gene therapy and diagnosis would be expected to have great medical benefits. To achieve this, therapeutic agents need to be delivered into the innermost mitochondrial space (mitochondrial matrix), which contains the mtDNA pool. We previously reported on the development of MITO-Porter, a liposome-based carrier that introduces macromolecular cargos into mitochondria via membrane fusion. In this study, we provide a demonstration of mitochondrial matrix delivery and the visualization of mitochondrial genes (mtDNA) in living cells using the MITO-Porter. We first prepared MITO-Porter containing encapsulated propidium iodide (PI), a fluorescent dye used to stain nucleic acids to detect mtDNA. We then confirmed the emission of red-fluorescence from PI by conjugation with mtDNA, when the carriers were incubated in the presence of isolated rat liver mitochondria. Finally, intracellular observation by confocal laser scanning microscopy clearly verified that the MITO-Porter delivered PI to the mitochondrial matrix.

  6. Herpes simplex virus glycoproteins gB and gH function in fusion between the virion envelope and the outer nuclear membrane.

    Science.gov (United States)

    Farnsworth, Aaron; Wisner, Todd W; Webb, Michael; Roller, Richard; Cohen, Gary; Eisenberg, Roselyn; Johnson, David C

    2007-06-12

    Herpesviruses must traverse the nuclear envelope to gain access to the cytoplasm and, ultimately, to exit cells. It is believed that herpesvirus nucleocapsids enter the perinuclear space by budding through the inner nuclear membrane (NM). To reach the cytoplasm these enveloped particles must fuse with the outer NM and the unenveloped capsids then acquire a second envelope in the trans-Golgi network. Little is known about the process by which herpesviruses virions fuse with the outer NM. Here we show that a herpes simplex virus (HSV) mutant lacking both the two putative fusion glycoproteins gB and gH failed to cross the nuclear envelope. Enveloped virions accumulated in the perinuclear space or in membrane vesicles that bulged into the nucleoplasm (herniations). By contrast, mutants lacking just gB or gH showed only minor or no defects in nuclear egress. We concluded that either HSV gB or gH can promote fusion between the virion envelope and the outer NM. It is noteworthy that fusion associated with HSV entry requires the cooperative action of both gB and gH, suggesting that the two types of fusion (egress versus entry) are dissimilar processes.

  7. TRAIL death receptor 4 signaling via lysosome fusion and membrane raft clustering in coronary arterial endothelial cells: evidence from ASM knockout mice.

    Science.gov (United States)

    Li, Xiang; Han, Wei-Qing; Boini, Krishna M; Xia, Min; Zhang, Yang; Li, Pin-Lan

    2013-01-01

    Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and its receptor, death receptor 4 (DR4), have been implicated in the development of endothelial dysfunction and atherosclerosis. However, the signaling mechanism mediating DR4 activation leading to endothelial injury remains unclear. We recently demonstrated that ceramide production via hydrolysis of membrane sphingomyelin by acid sphingomyelinase (ASM) results in membrane raft (MR) clustering and the formation of important redox signaling platforms, which play a crucial role in amplifying redox signaling in endothelial cells leading to endothelial dysfunction. The present study aims to investigate whether TRAIL triggers MR clustering via lysosome fusion and ASM activation, thereby conducting transmembrane redox signaling and changing endothelial function. Using confocal microscopy, we found that TRAIL induced MR clustering and co-localized with DR4 in coronary arterial endothelial cells (CAECs) isolated from wild-type (Smpd1 (+/+)) mice. Furthermore, TRAIL triggered ASM translocation, ceramide production, and NADPH oxidase aggregation in MR clusters in Smpd1 ( +/+ ) CAECs, whereas these observations were not found in Smpd1 (-/-) CAECs. Moreover, ASM deficiency reduced TRAIL-induced O(2) (-[Symbol: see text]) production in CAECs and abolished TRAIL-induced impairment on endothelium-dependent vasodilation in small resistance arteries. By measuring fluorescence resonance energy transfer, we found that Lamp-1 (lysosome membrane marker protein) and ganglioside G(M1) (MR marker) were trafficking together in Smpd1 (+/+) CAECs, which was absent in Smpd1 (-/-) CAECs. Consistently, fluorescence imaging of living cells with specific lysosome probes demonstrated that TRAIL-induced lysosome fusion with membrane was also absent in Smpd1 (-/-) CAECs. Taken together, these results suggest that ASM is essential for TRAIL-induced lysosomal trafficking, membrane fusion and formation of MR redox signaling platforms

  8. H1N1 Swine Influenza Viruses Differ from Avian Precursors by a Higher pH Optimum of Membrane Fusion

    Science.gov (United States)

    Baumann, Jan; Kouassi, Nancy Mounogou; Foni, Emanuela; Klenk, Hans-Dieter

    2015-01-01

    ABSTRACT The H1N1 Eurasian avian-like swine (EAsw) influenza viruses originated from an avian H1N1 virus. To characterize potential changes in the membrane fusion activity of the hemagglutinin (HA) during avian-to-swine adaptation of the virus, we studied EAsw viruses isolated in the first years of their circulation in pigs and closely related contemporary H1N1 viruses of wild aquatic birds. Compared to the avian viruses, the swine viruses were less sensitive to neutralization by lysosomotropic agent NH4Cl in MDCK cells, had a higher pH optimum of hemolytic activity, and were less stable at acidic pH. Eight amino acid substitutions in the HA were found to separate the EAsw viruses from their putative avian precursor; four substitutions—T492S, N722D, R752K, and S1132F—were located in the structural regions of the HA2 subunit known to play a role in acid-induced conformational transition of the HA. We also studied low-pH-induced syncytium formation by cell-expressed HA proteins and found that the HAs of the 1918, 1957, 1968, and 2009 pandemic viruses required a lower pH for fusion induction than did the HA of a representative EAsw virus. Our data show that transmission of an avian H1N1 virus to pigs was accompanied by changes in conformational stability and fusion promotion activity of the HA. We conclude that distinctive host-determined fusion characteristics of the HA may represent a barrier for avian-to-swine and swine-to-human transmission of influenza viruses. IMPORTANCE Continuing cases of human infections with zoonotic influenza viruses highlight the necessity to understand which viral properties contribute to interspecies transmission. Efficient binding of the HA to cellular receptors in a new host species is known to be essential for the transmission. Less is known about required adaptive changes in the membrane fusion activity of the HA. Here we show that adaptation of an avian influenza virus to pigs in Europe in 1980s was accompanied by mutations in

  9. pH Optimum of Hemagglutinin-Mediated Membrane Fusion Determines Sensitivity of Influenza A Viruses to the Interferon-Induced Antiviral State and IFITMs.

    Science.gov (United States)

    Gerlach, Thomas; Hensen, Luca; Matrosovich, Tatyana; Bergmann, Janina; Winkler, Michael; Peteranderl, Christin; Klenk, Hans-Dieter; Weber, Friedemann; Herold, Susanne; Pöhlmann, Stefan; Matrosovich, Mikhail

    2017-06-01

    The replication and pathogenicity of influenza A viruses (IAVs) critically depend on their ability to tolerate the antiviral interferon (IFN) response. To determine a potential role for the IAV hemagglutinin (HA) in viral sensitivity to IFN, we studied the restriction of IAV infection in IFN-β-treated human epithelial cells by using 2:6 recombinant IAVs that shared six gene segments of A/Puerto Rico/8/1934 virus (PR8) and contained HAs and neuraminidases of representative avian, human, and zoonotic H5N1 and H7N9 viruses. In A549 and Calu-3 cells, viruses displaying a higher pH optimum of HA-mediated membrane fusion, H5N1-PR8 and H7N9-PR8, were less sensitive to the IFN-induced antiviral state than their counterparts with HAs from duck and human viruses, which fused at a lower pH. The association between a high pH optimum of fusion and reduced IFN sensitivity was confirmed by using HA point mutants of A/Hong Kong/1/1968-PR8 that differed solely by their fusion properties. Furthermore, similar effects of the viral fusion pH on IFN sensitivity were observed in experiments with (i) primary human type II alveolar epithelial cells and differentiated cultures of human airway epithelial cells, (ii) nonrecombinant zoonotic and pandemic IAVs, and (iii) preparations of IFN-α and IFN-λ1. A higher pH of membrane fusion and reduced sensitivity to IFN correlated with lower restriction of the viruses in MDCK cells stably expressing the IFN-inducible transmembrane proteins IFITM2 and IFITM3, which are known to inhibit viral fusion. Our results reveal that the pH optimum of HA-driven membrane fusion of IAVs is a determinant of their sensitivity to IFN and IFITM proteins. IMPORTANCE The IFN system constitutes an important innate defense against viral infection. Substantial information is available on how IAVs avoid detection by sensors of the IFN system and disable IFN signaling pathways. Much less is known about the ability of IAVs to tolerate the antiviral activity of IFN

  10. Flagellar membrane fusion and protein exchange in trypanosomes; a new form of cell-cell communication? [version 1; referees: 2 approved

    Directory of Open Access Journals (Sweden)

    Simon Imhof

    2016-04-01

    Full Text Available Diverse structures facilitate direct exchange of proteins between cells, including plasmadesmata in plants and tunnelling nanotubes in bacteria and higher eukaryotes.  Here we describe a new mechanism of protein transfer, flagellar membrane fusion, in the unicellular parasite Trypanosoma brucei. When fluorescently tagged trypanosomes were co-cultured, a small proportion of double-positive cells were observed. The formation of double-positive cells was dependent on the presence of extracellular calcium and was enhanced by placing cells in medium supplemented with fresh bovine serum. Time-lapse microscopy revealed that double-positive cells arose by bidirectional protein exchange in the absence of nuclear transfer.  Furthermore, super-resolution microscopy showed that this process occurred in ≤1 minute, the limit of temporal resolution in these experiments. Both cytoplasmic and membrane proteins could be transferred provided they gained access to the flagellum. Intriguingly, a component of the RNAi machinery (Argonaute was able to move between cells, raising the possibility that small interfering RNAs are transported as cargo. Transmission electron microscopy showed that shared flagella contained two axonemes and two paraflagellar rods bounded by a single membrane. In some cases flagellar fusion was partial and interactions between cells were transient. In other cases fusion occurred along the entire length of the flagellum, was stable for several hours and might be irreversible. Fusion did not appear to be deleterious for cell function: paired cells were motile and could give rise to progeny while fused. The motile flagella of unicellular organisms are related to the sensory cilia of higher eukaryotes, raising the possibility that protein transfer between cells via cilia or flagella occurs more widely in nature.

  11. Evidence for a conserved inhibitory binding mode between the membrane fusion assembly factors Munc18 and syntaxin in animals.

    Science.gov (United States)

    Morey, Czuee; Kienle, C Nickias; Klöpper, Tobias H; Burkhardt, Pawel; Fasshauer, Dirk

    2017-12-15

    The membrane fusion necessary for vesicle trafficking is driven by the assembly of heterologous SNARE proteins orchestrated by the binding of Sec1/Munc18 (SM) proteins to specific syntaxin SNARE proteins. However, the precise mode of interaction between SM proteins and SNAREs is debated, as contrasting binding modes have been found for different members of the SM protein family, including the three vertebrate Munc18 isoforms. While different binding modes could be necessary, given their roles in different secretory processes in different tissues, the structural similarity of the three isoforms makes this divergence perplexing. Although the neuronal isoform Munc18a is well-established to bind tightly to both the closed conformation and the N-peptide of syntaxin 1a, thereby inhibiting SNARE complex formation, Munc18b and -c, which have a more widespread distribution, are reported to mainly interact with the N-peptide of their partnering syntaxins and are thought to instead promote SNARE complex formation. We have reinvestigated the interaction between Munc18c and syntaxin 4 (Syx4). Using isothermal titration calorimetry, we found that Munc18c, like Munc18a, binds to both the closed conformation and the N-peptide of Syx4. Furthermore, using a novel kinetic approach, we found that Munc18c, like Munc18a, slows down SNARE complex formation through high-affinity binding to syntaxin. This strongly suggests that secretory Munc18s in general control the accessibility of the bound syntaxin, probably preparing it for SNARE complex assembly. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  12. Rab3A is a new interacting partner of synaptotagmin I and may modulate synaptic membrane fusion through a competitive mechanism

    Energy Technology Data Exchange (ETDEWEB)

    Xie, Chunliang [Key Laboratory of Protein Chemistry and Developmental Biology of Ministry of Education, College of Life Sciences, Hunan Normal University, Changsha 410081 (China); Institute of Bast Fiber Crops, Chinese Academy of Agricultural Sciences, Changsha 410205 (China); Li, Jianglin; Guo, Tianyao; Yan, Yizhong; Tang, Cheng; Wang, Ying; Chen, Ping [Key Laboratory of Protein Chemistry and Developmental Biology of Ministry of Education, College of Life Sciences, Hunan Normal University, Changsha 410081 (China); Wang, Xianchun, E-mail: wang_xianchun@263.net [Key Laboratory of Protein Chemistry and Developmental Biology of Ministry of Education, College of Life Sciences, Hunan Normal University, Changsha 410081 (China); Liang, Songping, E-mail: liangsp@hunnu.edu.cn [Key Laboratory of Protein Chemistry and Developmental Biology of Ministry of Education, College of Life Sciences, Hunan Normal University, Changsha 410081 (China)

    2014-02-21

    Highlights: • Rab3A has been found to be a novel interacting protein of synaptotagmin I. • Rab3A binds to synaptotagmin I in a Ca{sup 2+}-independent manner. • KKKK motif in C2B domain of synaptotagmin I is a key site for Rab3A binding. • Rab3A competitively inhibits the binding of C2B in synaptotagmin I to syntaxin 1B. • Rab3A may regulate synaptic membrane fusion and exocytosis in a competitive manner. - Abstract: Rab3 and synaptotagmin have been reported to be the key proteins that have opposite actions but cooperatively play critical regulatory roles in selecting and limiting the number of vesicles released at central synapses. However, the exact mechanism has not been fully understood. In this study, Rab3A and synaptotagmin I, the most abundant isoforms of Rab3 and synaptotagmin, respectively, in brain were for the first time demonstrated to directly interact with each other in a Ca{sup 2+}-independent manner, and the KKKK motif in the C2B domain of synaptotagmin I was a key site for the Rab3A binding, which was further confirmed by the competitive inhibition of inositol hexakisphosphate. Further studies demonstrated that Rab3A competitively affected the synaptotagmin I interaction with syntaxin 1B that was involved in membrane fusion during the synaptic vesicle exocytosis. These data indicate that Rab3A is a new synaptotagmin I interacting partner and may participate in the regulation of synaptic membrane fusion and thus the vesicle exocytosis by competitively modulating the interaction of synaptotagmin with syntaxin of the t-SNARE complex in presynaptic membranes.

  13. Inner-membrane proteins PMI/TMEM11 regulate mitochondrial morphogenesis independently of the DRP1/MFN fission/fusion pathways.

    Science.gov (United States)

    Rival, Thomas; Macchi, Marc; Arnauné-Pelloquin, Laetitia; Poidevin, Mickael; Maillet, Frédéric; Richard, Fabrice; Fatmi, Ahmed; Belenguer, Pascale; Royet, Julien

    2011-03-01

    Mitochondria are highly dynamic organelles that can change in number and morphology during cell cycle, development or in response to extracellular stimuli. These morphological dynamics are controlled by a tight balance between two antagonistic pathways that promote fusion and fission. Genetic approaches have identified a cohort of conserved proteins that form the core of mitochondrial remodelling machineries. Mitofusins (MFNs) and OPA1 proteins are dynamin-related GTPases that are required for outer- and inner-mitochondrial membrane fusion respectively whereas dynamin-related protein 1 (DRP1) is the master regulator of mitochondrial fission. We demonstrate here that the Drosophila PMI gene and its human orthologue TMEM11 encode mitochondrial inner-membrane proteins that regulate mitochondrial morphogenesis. PMI-mutant cells contain a highly condensed mitochondrial network, suggesting that PMI has either a pro-fission or an anti-fusion function. Surprisingly, however, epistatic experiments indicate that PMI shapes the mitochondria through a mechanism that is independent of drp1 and mfn. This shows that mitochondrial networks can be shaped in higher eukaryotes by at least two separate pathways: one PMI-dependent and one DRP1/MFN-dependent.

  14. Zipping into fusion

    NARCIS (Netherlands)

    Zheng, Tingting

    2014-01-01

    Fusion of lipid bilayers in cells facilitates the active transport of chemicals. Non-viral membrane fusion is regulated by a cascade of proteins as the process is highly regulated both in space and time. In eukaryotic cells, the so-called SNARE protein complex is at the heart of fusion. However,

  15. Induction of heterosubtypic cross-protection against influenza by a whole inactivated virus vaccine: the role of viral membrane fusion activity.

    Directory of Open Access Journals (Sweden)

    Natalija Budimir

    Full Text Available BACKGROUND: The inability of seasonal influenza vaccines to effectively protect against infection with antigenically drifted viruses or newly emerging pandemic viruses underlines the need for development of cross-reactive influenza vaccines that induce immunity against a variety of virus subtypes. Therefore, potential cross-protective vaccines, e.g., whole inactivated virus (WIV vaccine, that can target conserved internal antigens such as the nucleoprotein (NP and/or matrix protein (M1 need to be explored. METHODOLOGY/PRINCIPAL FINDINGS: In the current study we show that a WIV vaccine, through induction of cross-protective cytotoxic T lymphocytes (CTLs, protects mice from heterosubtypic infection. This protection was abrogated after depletion of CD8+ cells in vaccinated mice, indicating that CTLs were the primary mediators of protection. Previously, we have shown that different procedures used for virus inactivation influence optimal activation of CTLs by WIV, most likely by affecting the membrane fusion properties of the virus. Specifically, inactivation with formalin (FA severely compromises fusion activity of the virus, while inactivation with β-propiolactone (BPL preserves fusion activity. Here, we demonstrate that vaccination of mice with BPL-inactivated H5N1 WIV vaccine induces solid protection from lethal heterosubtypic H1N1 challenge. By contrast, vaccination with FA-inactivated WIV, while preventing death after lethal challenge, failed to protect against development of disease and severe body weight loss. Vaccination with BPL-inactivated WIV, compared to FA-inactivated WIV, induced higher levels of specific CD8+ T cells in blood, spleen and lungs, and a higher production of granzyme B in the lungs upon H1N1 virus challenge. CONCLUSION/SIGNIFICANCE: The results underline the potential use of WIV as a cross-protective influenza vaccine candidate. However, careful choice of the virus inactivation procedure is important to retain membrane

  16. Induction of heterosubtypic cross-protection against influenza by a whole inactivated virus vaccine: the role of viral membrane fusion activity.

    Science.gov (United States)

    Budimir, Natalija; Huckriede, Anke; Meijerhof, Tjarko; Boon, Louis; Gostick, Emma; Price, David A; Wilschut, Jan; de Haan, Aalzen

    2012-01-01

    The inability of seasonal influenza vaccines to effectively protect against infection with antigenically drifted viruses or newly emerging pandemic viruses underlines the need for development of cross-reactive influenza vaccines that induce immunity against a variety of virus subtypes. Therefore, potential cross-protective vaccines, e.g., whole inactivated virus (WIV) vaccine, that can target conserved internal antigens such as the nucleoprotein (NP) and/or matrix protein (M1) need to be explored. In the current study we show that a WIV vaccine, through induction of cross-protective cytotoxic T lymphocytes (CTLs), protects mice from heterosubtypic infection. This protection was abrogated after depletion of CD8+ cells in vaccinated mice, indicating that CTLs were the primary mediators of protection. Previously, we have shown that different procedures used for virus inactivation influence optimal activation of CTLs by WIV, most likely by affecting the membrane fusion properties of the virus. Specifically, inactivation with formalin (FA) severely compromises fusion activity of the virus, while inactivation with β-propiolactone (BPL) preserves fusion activity. Here, we demonstrate that vaccination of mice with BPL-inactivated H5N1 WIV vaccine induces solid protection from lethal heterosubtypic H1N1 challenge. By contrast, vaccination with FA-inactivated WIV, while preventing death after lethal challenge, failed to protect against development of disease and severe body weight loss. Vaccination with BPL-inactivated WIV, compared to FA-inactivated WIV, induced higher levels of specific CD8+ T cells in blood, spleen and lungs, and a higher production of granzyme B in the lungs upon H1N1 virus challenge. The results underline the potential use of WIV as a cross-protective influenza vaccine candidate. However, careful choice of the virus inactivation procedure is important to retain membrane fusion activity and full immunogenicity of the vaccine.

  17. Cell fusion and nuclear fusion in plants.

    Science.gov (United States)

    Maruyama, Daisuke; Ohtsu, Mina; Higashiyama, Tetsuya

    2016-12-01

    Eukaryotic cells are surrounded by a plasma membrane and have a large nucleus containing the genomic DNA, which is enclosed by a nuclear envelope consisting of the outer and inner nuclear membranes. Although these membranes maintain the identity of cells, they sometimes fuse to each other, such as to produce a zygote during sexual reproduction or to give rise to other characteristically polyploid tissues. Recent studies have demonstrated that the mechanisms of plasma membrane or nuclear membrane fusion in plants are shared to some extent with those of yeasts and animals, despite the unique features of plant cells including thick cell walls and intercellular connections. Here, we summarize the key factors in the fusion of these membranes during plant reproduction, and also focus on "non-gametic cell fusion," which was thought to be rare in plant tissue, in which each cell is separated by a cell wall. Copyright © 2016 Elsevier Ltd. All rights reserved.

  18. Unusual Self-Assembly of the Recombinant Chlamydia trachomatis Major Outer Membrane Protein-Based Fusion Antigen CTH522 Into Protein Nanoparticles

    DEFF Research Database (Denmark)

    Rose, Fabrice; Karlsen, Kasper; Jensen, Pernille

    2018-01-01

    but is a challenging vaccine candidate by being an integral membrane protein, and the immunogenicity depends on a correctly folded structure. We investigated the biophysical properties of the recombinant MOMP-based fusion antigen CTH522, which is tested in early human clinical trials. It consists of a truncated......Sexually transmitted Chlamydia trachomatis (Ct) infects more than 100 million people annually, and untreated chlamydia infections can cause severe complications. Therefore, there is an urgent need for a chlamydia vaccine. The Ct major outer membrane protein (MOMP) is highly immunogenic......-defined secondary structural elements, and no thermal transitions were measurable. Chemical unfolding resulted monomers that upon removal of the denaturant self-assembled into higher order structures, comparable to the structure of the native protein. The conformation of CTH522 in nanoparticles is thus not entirely...

  19. Reconstituted Fusion Pore

    OpenAIRE

    Jeremic, Aleksandar; Kelly, Marie; Cho, Sang-Joon; Stromer, Marvin H.; Jena, Bhanu P.

    2003-01-01

    Fusion pores or porosomes are basket-like structures at the cell plasma membrane, at the base of which, membrane-bound secretory vesicles dock and fuse to release vesicular contents. Earlier studies using atomic force microscopy (AFM) demonstrated the presence of fusion pores at the cell plasma membrane in a number of live secretory cells, revealing their morphology and dynamics at nm resolution and in real time. ImmunoAFM studies demonstrated the release of vesicular contents through the por...

  20. Pandemic H1N1 influenza A directly induces a robust and acute inflammatory gene signature in primary human bronchial epithelial cells downstream of membrane fusion

    Energy Technology Data Exchange (ETDEWEB)

    Paquette, Stéphane G. [Division of Experimental Therapeutics, Toronto General Hospital Research Institute, University Health Network, Toronto, Ontario (Canada); Institute of Medical Science, Faculty of Medicine, University of Toronto, Toronto, Ontario (Canada); Banner, David [Division of Experimental Therapeutics, Toronto General Hospital Research Institute, University Health Network, Toronto, Ontario (Canada); Chi, Le Thi Bao [Department of Microbiology, Hue University of Medicine and Pharmacy, Thua Thien Hue (Viet Nam); Carlo Urbani Centre, Hue University of Medicine and Pharmacy, Thua Thien Hue (Viet Nam); Leon, Alberto J. [Division of Experimental Therapeutics, Toronto General Hospital Research Institute, University Health Network, Toronto, Ontario (Canada); International Institute of Infection and Immunity, Shantou University Medical College, Shantou, Guangdong (China); Xu, Luoling; Ran, Longsi [Division of Experimental Therapeutics, Toronto General Hospital Research Institute, University Health Network, Toronto, Ontario (Canada); Huang, Stephen S.H. [Division of Experimental Therapeutics, Toronto General Hospital Research Institute, University Health Network, Toronto, Ontario (Canada); Department of Immunology, Faculty of Medicine, University of Toronto, Toronto, Ontario (Canada); Farooqui, Amber [Division of Experimental Therapeutics, Toronto General Hospital Research Institute, University Health Network, Toronto, Ontario (Canada); International Institute of Infection and Immunity, Shantou University Medical College, Shantou, Guangdong (China); and others

    2014-01-05

    Pandemic H1N1 influenza A (H1N1pdm) elicits stronger pulmonary inflammation than previously circulating seasonal H1N1 influenza A (sH1N1), yet mechanisms of inflammatory activation in respiratory epithelial cells during H1N1pdm infection are unclear. We investigated host responses to H1N1pdm/sH1N1 infection and virus entry mechanisms in primary human bronchial epithelial cells in vitro. H1N1pdm infection rapidly initiated a robust inflammatory gene signature (3 h post-infection) not elicited by sH1N1 infection. Protein secretion inhibition had no effect on gene induction. Infection with membrane fusion deficient H1N1pdm failed to induce robust inflammatory gene expression which was rescued with restoration of fusion ability, suggesting H1N1pdm directly triggered the inflammatory signature downstream of membrane fusion. Investigation of intra-virion components revealed H1N1pdm viral RNA (vRNA) triggered a stronger inflammatory phenotype than sH1N1 vRNA. Thus, our study is first to report H1N1pdm induces greater inflammatory gene expression than sH1N1 in vitro due to direct virus–epithelial cell interaction. - Highlights: • We investigated H1N1pdm/sH1N1 infection in primary epithelial cells. • H1N1pdm directly initiated a robust inflammatory gene signature, sH1N1 did not. • H1N1pdm viral RNA triggered a stronger response than sH1N1. • H1N1pdm induces greater response due to direct virus–cell interaction. • These results have potential to impact vaccine and therapeutic development.

  1. Pandemic H1N1 influenza A directly induces a robust and acute inflammatory gene signature in primary human bronchial epithelial cells downstream of membrane fusion

    International Nuclear Information System (INIS)

    Paquette, Stéphane G.; Banner, David; Chi, Le Thi Bao; Leon, Alberto J.; Xu, Luoling; Ran, Longsi; Huang, Stephen S.H.; Farooqui, Amber

    2014-01-01

    Pandemic H1N1 influenza A (H1N1pdm) elicits stronger pulmonary inflammation than previously circulating seasonal H1N1 influenza A (sH1N1), yet mechanisms of inflammatory activation in respiratory epithelial cells during H1N1pdm infection are unclear. We investigated host responses to H1N1pdm/sH1N1 infection and virus entry mechanisms in primary human bronchial epithelial cells in vitro. H1N1pdm infection rapidly initiated a robust inflammatory gene signature (3 h post-infection) not elicited by sH1N1 infection. Protein secretion inhibition had no effect on gene induction. Infection with membrane fusion deficient H1N1pdm failed to induce robust inflammatory gene expression which was rescued with restoration of fusion ability, suggesting H1N1pdm directly triggered the inflammatory signature downstream of membrane fusion. Investigation of intra-virion components revealed H1N1pdm viral RNA (vRNA) triggered a stronger inflammatory phenotype than sH1N1 vRNA. Thus, our study is first to report H1N1pdm induces greater inflammatory gene expression than sH1N1 in vitro due to direct virus–epithelial cell interaction. - Highlights: • We investigated H1N1pdm/sH1N1 infection in primary epithelial cells. • H1N1pdm directly initiated a robust inflammatory gene signature, sH1N1 did not. • H1N1pdm viral RNA triggered a stronger response than sH1N1. • H1N1pdm induces greater response due to direct virus–cell interaction. • These results have potential to impact vaccine and therapeutic development

  2. Molecular dynamics simulations of T-2410 and T-2429 HIV fusion inhibitors interacting with model membranes: Insight into peptide behavior, structure and dynamics.

    Science.gov (United States)

    Mavioso, I C V C; de Andrade, V C R; Palace Carvalho, A J; Martins do Canto, A M T

    2017-09-01

    T-2410 and T-2429 are HIV fusion inhibitor peptides (FI) designed to present a higher efficiency even against HIV strains that developed resistance against other FIs. Similar peptides were shown to interact with model membranes both in the liquid disordered phase and in the liquid ordered state. Those results indicated that such interaction is important to function and could be correlated with their effectiveness. Extensive molecular dynamics simulations were carried out to investigate the interactions between both T-2410 and T-2429 with bilayers of pure 1-palmitoyl-2-oleoyl-phosphatidylcholine (POPC) and a mixture of POPC/cholesterol (Chol) (1:1). It was observed that both peptides interact strongly with both membrane systems, especially with the POPC/Chol systems, where these peptides show the highest number of H-bonds observed so far. T-2410 and T-2429 showed higher extent of interaction with bilayers when compared to T-20 or T-1249 in previous studies. This is most notable in POPC/Chol membranes where, although able to form H-bonds with Chol, they do so to a lesser extent than T-1249 does, the latter being the only FI peptide so far that was observed to form H-bonds with Chol. This behavior suggests that interaction of FI peptides with rigid Chol rich membranes may not be as dependent from peptide/Chol H-bond formation as previous results of T-1249 behavior led to believe. As in other similar peptides, the higher ability to interact with membranes shown by T-2410 and T2429 is probably correlated with its higher inhibitory efficiency. Copyright © 2017 Elsevier B.V. All rights reserved.

  3. Herpesvirus gB-induced fusion between the virion envelope and outer nuclear membrane during virus egress is regulated by the viral US3 kinase.

    Science.gov (United States)

    Wisner, Todd W; Wright, Catherine C; Kato, Akihisa; Kawaguchi, Yasushi; Mou, Fan; Baines, Joel D; Roller, Richard J; Johnson, David C

    2009-04-01

    Herpesvirus capsids collect along the inner surface of the nuclear envelope and bud into the perinuclear space. Enveloped virions then fuse with the outer nuclear membrane (NM). We previously showed that herpes simplex virus (HSV) glycoproteins gB and gH act in a redundant fashion to promote fusion between the virion envelope and the outer NM. HSV mutants lacking both gB and gH accumulate enveloped virions in herniations, vesicles that bulge into the nucleoplasm. Earlier studies had shown that HSV mutants lacking the viral serine/threonine kinase US3 also accumulate herniations. Here, we demonstrate that HSV gB is phosphorylated in a US3-dependent manner in HSV-infected cells, especially in a crude nuclear fraction. Moreover, US3 directly phosphorylated the gB cytoplasmic (CT) domain in in vitro assays. Deletion of gB in the context of a US3-null virus did not add substantially to defects in nuclear egress. The majority of the US3-dependent phosphorylation of gB involved the CT domain and amino acid T887, a residue present in a motif similar to that recognized by US3 in other proteins. HSV recombinants lacking gH and expressing either gB substitution mutation T887A or a gB truncated at residue 886 displayed substantial defects in nuclear egress. We concluded that phosphorylation of the gB CT domain is important for gB-mediated fusion with the outer NM. This suggested a model in which the US3 kinase is incorporated into the tegument layer (between the capsid and envelope) in HSV virions present in the perinuclear space. By this packaging, US3 might be brought close to the gB CT tail, leading to phosphorylation and triggering fusion between the virion envelope and the outer NM.

  4. Solid-state nuclear magnetic resonance measurements of HIV fusion peptide 13CO to lipid 31P proximities support similar partially inserted membrane locations of the α helical and β sheet peptide structures.

    Science.gov (United States)

    Gabrys, Charles M; Qiang, Wei; Sun, Yan; Xie, Li; Schmick, Scott D; Weliky, David P

    2013-10-03

    Fusion of the human immunodeficiency virus (HIV) membrane and the host cell membrane is an initial step of infection of the host cell. Fusion is catalyzed by gp41, which is an integral membrane protein of HIV. The fusion peptide (FP) is the ∼25 N-terminal residues of gp41 and is a domain of gp41 that plays a key role in fusion catalysis likely through interaction with the host cell membrane. Much of our understanding of the FP domain has been accomplished with studies of "HFP", i.e., a ∼25-residue peptide composed of the FP sequence but lacking the rest of gp41. HFP catalyzes fusion between membrane vesicles and serves as a model system to understand fusion catalysis. HFP binds to membranes and the membrane location of HFP is likely a significant determinant of fusion catalysis perhaps because the consequent membrane perturbation reduces the fusion activation energy. In the present study, many HFPs were synthesized and differed in the residue position that was (13)CO backbone labeled. Samples were then prepared that each contained a singly (13)CO labeled HFP incorporated into membranes that lacked cholesterol. HFP had distinct molecular populations with either α helical or oligomeric β sheet structure. Proximity between the HFP (13)CO nuclei and (31)P nuclei in the membrane headgroups was probed by solid-state NMR (SSNMR) rotational-echo double-resonance (REDOR) measurements. For many samples, there were distinct (13)CO shifts for the α helical and β sheet structures so that the proximities to (31)P nuclei could be determined for each structure. Data from several differently labeled HFPs were then incorporated into a membrane location model for the particular structure. In addition to the (13)CO labeled residue position, the HFPs also differed in sequence and/or chemical structure. "HFPmn" was a linear peptide that contained the 23 N-terminal residues of gp41. "HFPmn_V2E" contained the V2E mutation that for HIV leads to greatly reduced extent of fusion and

  5. Solid-State Nuclear Magnetic Resonance Measurements of HIV Fusion Peptide 13CO to Lipid 31P Proximities Support Similar Partially Inserted Membrane Locations of the α Helical and β Sheet Peptide Structures

    Science.gov (United States)

    Gabrys, Charles M.; Qiang, Wei; Sun, Yan; Xie, Li; Schmick, Scott D.; Weliky, David P.

    2013-10-01

    Fusion of the human immunodeficiency virus (HIV) membrane and the host cell membrane is an initial step of infection of the host cell. Fusion is catalyzed by gp41, which is an integral membrane protein of HIV. The fusion peptide (FP) is the -25 N-terminal residues of gp41 and is a domain of gp41 that plays a key role in fusion catalysis likely through interaction with the host cell membrane. Much of our understanding of the FP domain has been accomplished with studies of -HFP-, i.e., a -25-residue peptide composed of the FP sequence but lacking the rest of gp41. HFP catalyzes fusion between membrane vesicles and serves as a model system to understand fusion catalysis. HFP binds to membranes and the membrane location of HFP is likely a significant determinant of fusion catalysis perhaps because the consequent membrane perturbation reduces the fusion activation energy. In the present study, many HFPs were synthesized and differed in the residue position that was 13CO backbone labeled. Samples were then prepared that each contained a singly 13CO labeled HFP incorporated into membranes that lacked cholesterol. HFP had distinct molecular populations with either α helical or oligomeric - sheet structure. Proximity between the HFP 13CO nuclei and 31P nuclei in the membrane headgroups was probed by solid-state NMR (SSNMR) rotational-echo double-resonance (REDOR) measurements. For many samples, there were distinct 13CO shifts for the α helical and - sheet structures so that the proximities to 31P nuclei could be determined for each structure. Data from several differently labeled HFPs were then incorporated into a membrane location model for the particular structure. In addition to the 13CO labeled residue position, the HFPs also differed in sequence and/or chemical structure. -HFPmn- was a linear peptide that contained the 23 N-terminal residues of gp41. -HFPmn_V2E- contained the V2E mutation that for HIV leads to greatly reduced extent of fusion and infection. The

  6. Functional fluorescent protein insertions in herpes simplex virus gB report on gB conformation before and after execution of membrane fusion.

    Directory of Open Access Journals (Sweden)

    John R Gallagher

    2014-09-01

    Full Text Available Entry of herpes simplex virus (HSV into a target cell requires complex interactions and conformational changes by viral glycoproteins gD, gH/gL, and gB. During viral entry, gB transitions from a prefusion to a postfusion conformation, driving fusion of the viral envelope with the host cell membrane. While the structure of postfusion gB is known, the prefusion conformation of gB remains elusive. As the prefusion conformation of gB is a critical target for neutralizing antibodies, we set out to describe its structure by making genetic insertions of fluorescent proteins (FP throughout the gB ectodomain. We created gB constructs with FP insertions in each of the three globular domains of gB. Among 21 FP insertion constructs, we found 8 that allowed gB to remain membrane fusion competent. Due to the size of an FP, regions in gB that tolerate FP insertion must be solvent exposed. Two FP insertion mutants were cell-surface expressed but non-functional, while FP insertions located in the crown were not surface expressed. This is the first report of placing a fluorescent protein insertion within a structural domain of a functional viral fusion protein, and our results are consistent with a model of prefusion HSV gB constructed from the prefusion VSV G crystal structure. Additionally, we found that functional FP insertions from two different structural domains could be combined to create a functional form of gB labeled with both CFP and YFP. FRET was measured with this construct, and we found that when co-expressed with gH/gL, the FRET signal from gB was significantly different from the construct containing CFP alone, as well as gB found in syncytia, indicating that this construct and others of similar design are likely to be powerful tools to monitor the conformation of gB in any model system accessible to light microscopy.

  7. Membrane fusion activity of Semliki forest virus in a liposomal model system : Specific inhibition by Zn2+ ions

    NARCIS (Netherlands)

    Corver, J; Snippe, H; Kraaijeveld, C; Wilschut, J

    1997-01-01

    Semliki Forest virus (SFV) has been shown previously to fuse efficiently with cholesterol-and sphingolipid-containing liposomal model membranes in a low-pH-dependent manner. Several steps can be distinguished in this process, including low-pH-induced irreversible binding of the virus to the

  8. Mutation of the dengue virus type 2 envelope protein heparan sulfate binding sites or the domain III lateral ridge blocks replication in Vero cells prior to membrane fusion

    International Nuclear Information System (INIS)

    Roehrig, John T.; Butrapet, Siritorn; Liss, Nathan M.; Bennett, Susan L.; Luy, Betty E.; Childers, Thomas; Boroughs, Karen L.; Stovall, Janae L.; Calvert, Amanda E.; Blair, Carol D.; Huang, Claire Y.-H.

    2013-01-01

    Using an infectious cDNA clone we engineered seven mutations in the putative heparan sulfate- and receptor-binding motifs of the envelope protein of dengue virus serotype 2, strain 16681. Four mutant viruses, KK122/123EE, E202K, G304K, and KKK305/307/310EEE, were recovered following transfection of C6/36 cells. A fifth mutant, KK291/295EE, was recovered from C6/36 cells with a compensatory E295V mutation. All mutants grew in and mediated fusion of virus-infected C6/36 cells, but three of the mutants, KK122/123EE, E202K, G304K, did not grow in Vero cells without further modification. Two Vero cell lethal mutants, KK291/295EV and KKK307/307/310EEE, failed to replicate in DC-SIGN-transformed Raji cells and did not react with monoclonal antibodies known to block DENV attachment to Vero cells. Additionally, both mutants were unable to initiate negative-strand vRNA synthesis in Vero cells by 72 h post-infection, suggesting that the replication block occurred prior to virus-mediated membrane fusion. - Highlights: • Heparan sulfate- and receptor-binding motifs of DENV2 envelope protein were mutated. • Four mutant viruses were isolated—all could fuse C6/36 cells. • Two of these mutants were lethal in Vero cells without further modification. • Lethal mutations were KK291/295EV and KKK305/307/310EEE. • Cell attachment was implicated as the replication block for both mutants

  9. Mutation of the dengue virus type 2 envelope protein heparan sulfate binding sites or the domain III lateral ridge blocks replication in Vero cells prior to membrane fusion

    Energy Technology Data Exchange (ETDEWEB)

    Roehrig, John T., E-mail: jtr1@cdc.gov [Division of Vector-Borne Diseases, Centers for Disease Control and Prevention, Fort Collins, CO 80521 (United States); Butrapet, Siritorn; Liss, Nathan M. [Division of Vector-Borne Diseases, Centers for Disease Control and Prevention, Fort Collins, CO 80521 (United States); Bennett, Susan L. [Arthropod-borne and Infectious Diseases Laboratory, Department of Microbiology, Immunology, and Pathology, Colorado State University, Fort Collins, CO 80523 (United States); Luy, Betty E.; Childers, Thomas; Boroughs, Karen L.; Stovall, Janae L.; Calvert, Amanda E. [Division of Vector-Borne Diseases, Centers for Disease Control and Prevention, Fort Collins, CO 80521 (United States); Blair, Carol D. [Arthropod-borne and Infectious Diseases Laboratory, Department of Microbiology, Immunology, and Pathology, Colorado State University, Fort Collins, CO 80523 (United States); Huang, Claire Y.-H. [Division of Vector-Borne Diseases, Centers for Disease Control and Prevention, Fort Collins, CO 80521 (United States)

    2013-07-05

    Using an infectious cDNA clone we engineered seven mutations in the putative heparan sulfate- and receptor-binding motifs of the envelope protein of dengue virus serotype 2, strain 16681. Four mutant viruses, KK122/123EE, E202K, G304K, and KKK305/307/310EEE, were recovered following transfection of C6/36 cells. A fifth mutant, KK291/295EE, was recovered from C6/36 cells with a compensatory E295V mutation. All mutants grew in and mediated fusion of virus-infected C6/36 cells, but three of the mutants, KK122/123EE, E202K, G304K, did not grow in Vero cells without further modification. Two Vero cell lethal mutants, KK291/295EV and KKK307/307/310EEE, failed to replicate in DC-SIGN-transformed Raji cells and did not react with monoclonal antibodies known to block DENV attachment to Vero cells. Additionally, both mutants were unable to initiate negative-strand vRNA synthesis in Vero cells by 72 h post-infection, suggesting that the replication block occurred prior to virus-mediated membrane fusion. - Highlights: • Heparan sulfate- and receptor-binding motifs of DENV2 envelope protein were mutated. • Four mutant viruses were isolated—all could fuse C6/36 cells. • Two of these mutants were lethal in Vero cells without further modification. • Lethal mutations were KK291/295EV and KKK305/307/310EEE. • Cell attachment was implicated as the replication block for both mutants.

  10. Acid-induced movements in the glycoprotein shell of an alphavirus turn the spikes into membrane fusion mode

    OpenAIRE

    Haag, Lars; Garoff, Henrik; Xing, Li; Hammar, Lena; Kan, Sin-Tau; Cheng, R.Holland

    2002-01-01

    In the icosahedral (T = 4) Semliki Forest virus, the envelope protomers, i.e. E1–E2 heterodimers, make one-to-one interactions with capsid proteins below the viral lipid bilayer, transverse the membrane and form an external glycoprotein shell with projections. The shell is organized by protomer domains interacting as hexamers and pentamers around shell openings at icosahedral 2- and 5-fold axes, respectively, and the projections by other domains associating as trimers at 3- and quasi 3-fold a...

  11. Avian sarcoma and leukosis virus-receptor interactions: From classical genetics to novel insights into virus-cell membrane fusion

    International Nuclear Information System (INIS)

    Barnard, R.J.O.; Elleder, D.; Young, J.A.T.

    2006-01-01

    For over 40 years, avian sarcoma and leukosis virus (ASLV)-receptor interactions have been employed as a useful model system to study the mechanism of retroviral entry into cells. Pioneering studies on this system focused upon the genetic basis of the differential susceptibilities of different lines of chickens to infection by distinct subgroups of ASLV. These studies led to the definition of three distinct autosomal recessive genes that were predicted to encode cellular receptors for different viral subgroups. They also led to the concept of viral interference, i.e. the mechanism by which infection by one virus can render cells resistant to reinfection by other viruses that use the same cellular receptor. Here, we review the contributions that analyses of the ASLV-receptor system have made in unraveling the mechanisms of retroviral entry into cells and focus on key findings such as identification and characterization of the ASLV receptor genes and the subsequent elucidation of an unprecedented mechanism of virus-cell fusion. Since many of the initial findings on this system were published in the early volumes of Virology, this subject is especially well suited to this special anniversary issue of the journal

  12. A trivalent Apx-fusion protein delivered by E. coli outer membrane vesicles induce protection against Actinobacillus pleuropneumoniae of serotype 1 and 7 challenge in a murine model.

    Science.gov (United States)

    Xu, Kui; Zhao, Qin; Wen, Xintian; Wu, Rui; Wen, Yiping; Huang, Xiaobo; Huang, Yong; Yan, Qigui; Han, Xinfeng; Ma, Xiaoping; Chang, Yung-Fu; Cao, Sanjie

    2018-01-01

    Actinobacillus pleuropneumoniae (APP) causes serious economic losses in the swine industry, and is the etiologic agent of porcine pleuropneumonia. In this study we have engineered a trivalent Apx fusion protein enclosed in outer membrane vesicles (Apxr-OMV) and studied its immunoprotective efficacy against APP serotypes 1 and 7 challenge in mice. The results showed that the IgG levels in the Apxr-OMVs immune group were significantly higher than those of the negative control (P < 0.05). Up-regulation of both Th1 (IFN-γ, IL-2) and Th2 (IL-4) cytokines were detected in splenocytes of Apxr-OMVs immune group. The survival rates 87.5% and 62.5% were observed against APP strain 1516 of serotype 7 and APP strain 2701 of serotype 1 in the groups of Apxr-OMVs immune group, respectively. Histopathological lesions of the pulmonary structure alveoli were found to be minimal in APX-OMV group challenged with APP serotypes 1 and 7. These results strongly indicated that engineered OMVs could effectively induce specific humoral or cellular immune responses. Moreover, Apxr-OMVs used as novel vaccine provides cross-protective immunity against different serotype 1 and 7 of APP infection in a mouse model. In contrast, the OMV-empty and PBS as negative controls or inactivated strain of APP-2701 and APP-1516 as positive controls for the animal study cannot provide protection or cross-protection.

  13. MacA, a periplasmic membrane fusion protein of the macrolide transporter MacAB-TolC, binds lipopolysaccharide core specifically and with high affinity.

    Science.gov (United States)

    Lu, Shuo; Zgurskaya, Helen I

    2013-11-01

    The Escherichia coli MacAB-TolC transporter has been implicated in efflux of macrolide antibiotics and secretion of enterotoxin STII. In this study, we found that purified MacA, a periplasmic membrane fusion protein, contains one tightly bound rough core lipopolysaccharide (R-LPS) molecule per MacA molecule. R-LPS was bound specifically to MacA protein with affinity exceeding that of polymyxin B. Sequence analyses showed that MacA contains two high-density clusters of positively charged amino acid residues located in the cytoplasmic N-terminal domain and the periplasmic C-terminal domain. Substitutions in the C-terminal cluster reducing the positive-charge density completely abolished binding of R-LPS. At the same time, these substitutions significantly reduced the functionality of MacA in the protection of E. coli against macrolides in vivo and in the in vitro MacB ATPase stimulation assays. Taken together, our results suggest that R-LPS or a similar glycolipid is a physiological substrate of MacAB-TolC.

  14. Cell fusions in mammals

    DEFF Research Database (Denmark)

    Larsson, Lars-Inge; Bjerregaard, Bolette; Talts, Jan Fredrik

    2008-01-01

    Cell fusions are important to fertilization, placentation, development of skeletal muscle and bone, calcium homeostasis and the immune defense system. Additionally, cell fusions participate in tissue repair and may be important to cancer development and progression. A large number of factors appear...... to regulate cell fusions, including receptors and ligands, membrane domain organizing proteins, proteases, signaling molecules and fusogenic proteins forming alpha-helical bundles that bring membranes close together. The syncytin family of proteins represent true fusogens and the founding member, syncytin-1......, has been documented to be involved in fusions between placental trophoblasts, between cancer cells and between cancer cells and host ells. We review the literature with emphasis on the syncytin family and propose that syncytins may represent universal fusogens in primates and rodents, which work...

  15. The role of the N-terminal segment of CCR5 in HIV-1 Env-mediated membrane fusion and the mechanism of virus adaptation to CCR5 lacking this segment

    Directory of Open Access Journals (Sweden)

    Kabat David

    2007-08-01

    Full Text Available Abstract Background HIV-1 envelope glycoprotein (Env induces membrane fusion as a result of sequential binding to CD4 and chemokine receptors (CCR5 or CXCR4. The critical determinants of CCR5 coreceptor function are the N-terminal domain (Nt and the second extracellular loop. However, mutations in gp120 adapt HIV-1 to grow on cells expressing the N-terminally truncated CCR5(Δ18 (Platt et al., J. Virol. 2005, 79: 4357–68. Results We have functionally characterized the adapted Env (designated Env(NYP using a quantitative cell-cell fusion assay. The rate of fusion with target cells expressing wild-type CCR5 and the resistance to fusion inhibitors was virtually identical for wild-type Env and Env(NYP, implying that the coreceptor affinity had not increased as a result of adaptation. In contrast, Env(NYP-induced fusion with cells expressing CCR5(Δ18 occurred at a slower rate and was extremely sensitive to the CCR5 binding inhibitor, Sch-C. Resistance to Sch-C drastically increased after pre-incubation of Env(NYP- and CCR5(Δ18-expressing cells at a temperature that was not permissive to fusion. This indicates that ternary Env(NYP-CD4-CCR5(Δ18 complexes accumulate at sub-threshold temperature and that low-affinity interactions with the truncated coreceptor are sufficient for triggering conformational changes in the gp41 of Env(NYP but not in wild-type Env. We also demonstrated that the ability of CCR5(Δ18 to support fusion and infection mediated by wild-type Env can be partially reconstituted in the presence of a synthetic sulfated peptide corresponding to the CCR5 Nt. Pre-incubation of wild-type Env- and CCR5(Δ18-expressing cells with the sulfated peptide at sub-threshold temperature markedly increased the efficiency of fusion. Conclusion We propose that, upon binding the Nt region of CCR5, wild-type Env acquires the ability to productively engage the extracellular loop(s of CCR5 – an event that triggers gp41 refolding and membrane merger

  16. Surface Density of the Hendra G Protein Modulates Hendra F Protein-Promoted Membrane Fusion: Role for Hendra G Protein Trafficking and Degradation

    OpenAIRE

    Whitman, Shannon D.; Dutch, Rebecca Ellis

    2007-01-01

    Hendra virus, like most paramyxoviruses, requires both a fusion (F) and attachment (G) protein for promotion of cell-cell fusion. Recent studies determined that Hendra F is proteolytically processed by the cellular protease cathepsin L after endocytosis. This unique cathepsin L processing results in a small percentage of Hendra F on the cell surface. To determine how the surface densities of the two Hendra glycoproteins affect fusion promotion, we performed experiments that varied the levels ...

  17. Spinal fusion

    Science.gov (United States)

    ... Herniated disk - fusion; Spinal stenosis - fusion; Laminectomy - fusion Patient Instructions Bathroom safety - adults Preventing falls Preventing falls - what to ask your doctor Spine surgery - discharge Surgical wound care - open Images Scoliosis Spinal ...

  18. A cytochrome c fusion protein domain for convenient detection, quantification, and enhanced production of membrane proteins in Escherichia coli--expression and characterization of cytochrome-tagged Complex I subunits.

    Science.gov (United States)

    Gustavsson, Tobias; Trane, Maria; Moparthi, Vamsi K; Miklovyte, Egle; Moparthi, Lavanya; Górecki, Kamil; Leiding, Thom; Arsköld, Sindra Peterson; Hägerhäll, Cecilia

    2010-08-01

    Overproduction of membrane proteins can be a cumbersome task, particularly if high yields are desirable. NADH:quinone oxidoreductase (Complex I) contains several very large membrane-spanning protein subunits that hitherto have been impossible to express individually in any appreciable amounts in Escherichia coli. The polypeptides contain no prosthetic groups and are poorly antigenic, making optimization of protein production a challenging task. In this work, the C-terminal ends of the Complex I subunits NuoH, NuoL, NuoM, and NuoN from E. coli Complex I and the bona fide antiporters MrpA and MrpD were genetically fused to the cytochrome c domain of Bacillus subtilis cytochrome c(550). Compared with other available fusion-protein tagging systems, the cytochrome c has several advantages. The heme is covalently bound, renders the proteins visible by optical spectroscopy, and can be used to monitor, quantify, and determine the orientation of the polypeptides in a plethora of experiments. For the antiporter-like subunits NuoL, NuoM, and NuoN and the real antiporters MrpA and MrpD, unprecedented amounts of holo-cytochrome fusion proteins could be obtained in E. coli. The NuoHcyt polypeptide was also efficiently produced, but heme insertion was less effective in this construct. The cytochrome c(550) domain in all the fusion proteins exhibited normal spectra and redox properties, with an E(m) of about +170 mV. The MrpA and MrpD antiporters remained functional after being fused to the cytochrome c-tag. Finally, a his-tag could be added to the cytochrome domain, without any perturbations to the cytochrome properties, allowing efficient purification of the overexpressed fusion proteins.

  19. Cell fusion by ionizing radiation

    International Nuclear Information System (INIS)

    Khair, M.B.

    1993-08-01

    The relevance and importance of cell fusion are illustrated by the notion that current interest in this phenomenon is shared by scientists in quite varied disciplines. The diversity of cellular membrane fusion phenomena could provoke one to think that there must be a multitude of mechanisms that can account for such diversity. But, in general, the mechanism for the fusion reaction itself could be very similar in many, or even all, cases. Cell fusion can be induced by several factors such as virus Sendai, polyethylene glycol, electric current and ionizing radiation. This article provides the reader with short view of recent progress in research on cell fusion and gives some explanations about fusion mechanisms. This study shows for the first time, the results of the cell fusion induced by ionizing radiations that we have obtained in our researches and the work performed by other groups. (author). 44 refs

  20. EMP Fusion

    OpenAIRE

    KUNTAY, Isık

    2010-01-01

    This paper introduces a novel fusion scheme, called EMP Fusion, which has the promise of achieving breakeven and realizing commercial fusion power. The method is based on harnessing the power of an electromagnetic pulse generated by the now well-developed flux compression technology. The electromagnetic pulse acts as a means of both heating up the plasma and confining the plasma, eliminating intermediate steps. The EMP Fusion device is simpler compared to other fusion devices and this reduces...

  1. Fusion of Enveloped Viruses in Endosomes

    Science.gov (United States)

    White, Judith M.; Whittaker, Gary R.

    2016-01-01

    Ari Helenius launched the field of enveloped virus fusion in endosomes with a seminal paper in the Journal of Cell Biology in 1980. In the intervening years a great deal has been learned about the structures and mechanisms of viral membrane fusion proteins as well as about the endosomes in which different enveloped viruses fuse and the endosomal cues that trigger fusion. We now recognize three classes of viral membrane fusion proteins based on structural criteria and four mechanisms of fusion triggering. After reviewing general features of viral membrane fusion proteins and viral fusion in endosomes, we delve into three characterized mechanisms for viral fusion triggering in endosomes: by low pH, by receptor binding plus low pH, and by receptor binding plus the action of a protease. We end with a discussion of viruses that may employ novel endosomal fusion triggering mechanisms. A key take home message is that enveloped viruses that enter cells by fusing in endosomes traverse the endocytic pathway until they reach an endosome that has all of the environmental conditions (pH, proteases, ions, intracellular receptors, and lipid composition) to (if needed) prime and (in all cases) trigger the fusion protein and to support membrane fusion. PMID:26935856

  2. Solid-state nuclear magnetic resonance (NMR) spectroscopy of human immunodeficiency virus gp41 protein that includes the fusion peptide: NMR detection of recombinant Fgp41 in inclusion bodies in whole bacterial cells and structural characterization of purified and membrane-associated Fgp41.

    Science.gov (United States)

    Vogel, Erica P; Curtis-Fisk, Jaime; Young, Kaitlin M; Weliky, David P

    2011-11-22

    Human immunodeficiency virus (HIV) infection of a host cell begins with fusion of the HIV and host cell membranes and is mediated by the gp41 protein, a single-pass integral membrane protein of HIV. The 175 N-terminal residues make up the ectodomain that lies outside the virus. This work describes the production and characterization of an ectodomain construct containing the 154 N-terminal gp41 residues, including the fusion peptide (FP) that binds to target cell membranes. The Fgp41 sequence was derived from one of the African clade A strains of HIV-1 that have been less studied than European/North American clade B strains. Fgp41 expression at a level of ~100 mg/L of culture was evidenced by an approach that included amino acid type (13)CO and (15)N labeling of recombinant protein and solid-state NMR (SSNMR) spectroscopy of lyophilized whole cells. The approach did not require any protein solubilization or purification and may be a general approach for detection of recombinant protein. The purified Fgp41 yield was ~5 mg/L of culture. SSNMR spectra of membrane-associated Fgp41 showed high helicity for the residues C-terminal of the FP. This was consistent with a "six-helix bundle" (SHB) structure that is the final gp41 state during membrane fusion. This observation and negligible Fgp41-induced vesicle fusion supported a function for SHB gp41 of membrane stabilization and fusion arrest. SSNMR spectra of residues in the membrane-associated FP provided evidence of a mixture of molecular populations with either helical or β-sheet FP conformation. These and earlier SSNMR data strongly support the existence of these populations in the SHB state of membrane-associated gp41. © 2011 American Chemical Society

  3. Loss-of-function of the ciliopathy protein Cc2d2a disorganizes the vesicle fusion machinery at the periciliary membrane and indirectly affects Rab8-trafficking in zebrafish photoreceptors.

    Science.gov (United States)

    Ojeda Naharros, Irene; Gesemann, Matthias; Mateos, José M; Barmettler, Gery; Forbes, Austin; Ziegler, Urs; Neuhauss, Stephan C F; Bachmann-Gagescu, Ruxandra

    2017-12-01

    Ciliopathies are human disorders caused by dysfunction of primary cilia, ubiquitous organelles involved in transduction of environmental signals such as light sensation in photoreceptors. Concentration of signal detection proteins such as opsins in the ciliary membrane is achieved by RabGTPase-regulated polarized vesicle trafficking and by a selective barrier at the ciliary base, the transition zone (TZ). Dysfunction of the TZ protein CC2D2A causes Joubert/Meckel syndromes in humans and loss of ciliary protein localization in animal models, including opsins in retinal photoreceptors. The link between the TZ and upstream vesicle trafficking has been little explored to date. Moreover, the role of the small GTPase Rab8 in opsin-carrier vesicle (OCV) trafficking has been recently questioned in a mouse model. Using correlative light and electron microscopy and live imaging in zebrafish photoreceptors, we provide the first live characterization of Rab8-mediated trafficking in photoreceptors in vivo. Our results support a possibly redundant role for both Rab8a/b paralogs in OCV trafficking, based on co-localization of Rab8 and opsins in vesicular structures, and joint movement of Rab8-tagged particles with opsin. We further investigate the role of the TZ protein Cc2d2a in Rab8-mediated trafficking using cc2d2a zebrafish mutants and identify a requirement for Cc2d2a in the latest step of OCV trafficking, namely vesicle fusion. Progressive accumulation of opsin-containing vesicles in the apical portion of photoreceptors lacking Cc2d2a is caused by disorganization of the vesicle fusion machinery at the periciliary membrane with mislocalization and loss of the t-SNAREs SNAP25 and Syntaxin3 and of the exocyst component Exoc4. We further observe secondary defects on upstream Rab8-trafficking with cytoplasmic accumulation of Rab8. Taken together, our results support participation of Rab8 in OCV trafficking and identify a novel role for the TZ protein Cc2d2a in fusion of incoming

  4. From biological membranes to biomimetic model membranes

    Directory of Open Access Journals (Sweden)

    Eeman, M.

    2010-01-01

    Full Text Available Biological membranes play an essential role in the cellular protection as well as in the control and the transport of nutrients. Many mechanisms such as molecular recognition, enzymatic catalysis, cellular adhesion and membrane fusion take place into the biological membranes. In 1972, Singer et al. provided a membrane model, called fluid mosaic model, in which each leaflet of the bilayer is formed by a homogeneous environment of lipids in a fluid state including globular assembling of proteins and glycoproteins. Since its conception in 1972, many developments were brought to this model in terms of composition and molecular organization. The main development of the fluid mosaic model was made by Simons et al. (1997 and Brown et al. (1997 who suggested that membrane lipids are organized into lateral microdomains (or lipid rafts with a specific composition and a molecular dynamic that are different to the composition and the dynamic of the surrounding liquid crystalline phase. The discovery of a phase separation in the plane of the membrane has induced an explosion in the research efforts related to the biology of cell membranes but also in the development of new technologies for the study of these biological systems. Due to the high complexity of biological membranes and in order to investigate the biological processes that occur on the membrane surface or within the membrane lipid bilayer, a large number of studies are performed using biomimicking model membranes. This paper aims at revisiting the fundamental properties of biological membranes in terms of membrane composition, membrane dynamic and molecular organization, as well as at describing the most common biomimicking models that are frequently used for investigating biological processes such as membrane fusion, membrane trafficking, pore formation as well as membrane interactions at a molecular level.

  5. Optimizing E. coli-based membrane protein production using Lemo21(DE3) or pReX and GFP-Fusions

    NARCIS (Netherlands)

    Kuipers, Grietje; Peschke, Markus; Ismail, Nurzian Bernsel; Hjelm, Anna; Schlegel, Susan; Vikström, David; Luirink, Joen; de Gier, Jan Willem

    2017-01-01

    Optimizing the conditions for the production of membrane proteins in E. coli is usually a laborious and time-consuming process. Combining the Lemo21(DE3) strain or the pReX T7-based expression vector with membrane proteins C-terminally fused to Green Fluorescent Protein (GFP) greatly facilitates the

  6. MOF-mixed matrix membranes: Precise dispersion of MOF particles with better compatibility via a particle fusion approach for enhanced gas separation properties

    NARCIS (Netherlands)

    Shahid, S.; Nijmeijer, Dorothea C.; Nehache, Sabrina; Vankelecom, Ivo; Deratani, Andre; Quemener, Damien

    2015-01-01

    Mixed matrix membranes (MMMs) incorporating conventional fillers frequently suffer from insufficient adhesion between the polymer matrix and the fillers. This often results in the formation of non-selective voids at the filler/polymer interface, which decreases the performance of the membrane. A

  7. Fusion power

    International Nuclear Information System (INIS)

    Hancox, R.

    1981-01-01

    The principles of fusion power, and its advantages and disadvantages, are outlined. Present research programmes and future plans directed towards the development of a fusion power reactor, are summarized. (U.K.)

  8. Fusion rings and fusion ideals

    DEFF Research Database (Denmark)

    Andersen, Troels Bak

    This dissertation investigates fusion rings, which are Grothendieck groups of rigid, monoidal, semisimple, abelian categories. Special interest is in rational fusion rings, i.e., fusion rings which admit a finite basis, for as commutative rings they may be presented as quotients of polynomial rings...... by the so-called fusion ideals. The fusion rings of Wess-Zumino-Witten models have been widely studied and are well understood in terms of precise combinatorial descriptions and explicit generating sets of the fusion ideals. They also appear in another, more general, setting via tilting modules for quantum...

  9. Mechanics of post-fusion exocytotic vesicle

    Science.gov (United States)

    Stephens, Thomas; Wu, Zhanghan; Liu, Jian

    2017-06-01

    Exocytosis is an important cellular process controlled by metabolic signaling. It involves vesicle fusion to the plasma membrane, followed by the opening of a fusion pore, and the subsequent release of the vesicular lumen content into the extracellular space. While most modeling efforts focus on the events leading to membrane fusion, how the vesicular membrane remodels after fusing to plasma membrane remains unclear. This latter event dictates the nature and the efficiency of exocytotic vesicular secretions, and is thus critical for exocytotic function. We provide a generic membrane mechanical model to systematically study the fate of post-fusion vesicles. We show that while membrane stiffness favors full-collapse vesicle fusion into the plasma membrane, the intravesicular pressure swells the vesicle and causes the fusion pore to shrink. Dimensions of the vesicle and its associated fusion pore further modulate this mechanical antagonism. We systematically define the mechanical conditions that account for the full spectrum of the observed vesicular secretion modes. Our model therefore can serve as a unified theoretical framework that sheds light on the elaborate control mechanism of exocytosis.

  10. Fusion: introduction

    International Nuclear Information System (INIS)

    Decreton, M.

    2006-01-01

    The article gives an overview and introduction to the activities of SCK-CEN's research programme on fusion. The decision to construct the ITER international nuclear fusion experiment in Cadarache is highlighted. A summary of the Belgian contributions to fusion research is given with particular emphasis on studies of radiation effects on diagnostics systems, radiation effects on remote handling sensing systems, fusion waste management and socio-economic studies

  11. Fusion Canada

    International Nuclear Information System (INIS)

    1987-07-01

    This first issue of a quarterly newsletter announces the startup of the Tokamak de Varennes, describes Canada's national fusion program, and outlines the Canadian Fusion Fuels Technology Program. A map gives the location of the eleven principal fusion centres in Canada. (L.L.)

  12. Fusion neutronics

    CERN Document Server

    Wu, Yican

    2017-01-01

    This book provides a systematic and comprehensive introduction to fusion neutronics, covering all key topics from the fundamental theories and methodologies, as well as a wide range of fusion system designs and experiments. It is the first-ever book focusing on the subject of fusion neutronics research. Compared with other nuclear devices such as fission reactors and accelerators, fusion systems are normally characterized by their complex geometry and nuclear physics, which entail new challenges for neutronics such as complicated modeling, deep penetration, low simulation efficiency, multi-physics coupling, etc. The book focuses on the neutronics characteristics of fusion systems and introduces a series of theories and methodologies that were developed to address the challenges of fusion neutronics, and which have since been widely applied all over the world. Further, it introduces readers to neutronics design’s unique principles and procedures, experimental methodologies and technologies for fusion systems...

  13. Survival of the fittest: positive selection of CD4+ T cells expressing a membrane-bound fusion inhibitor following HIV-1 infection.

    Science.gov (United States)

    Kimpel, Janine; Braun, Stephen E; Qiu, Gang; Wong, Fay Eng; Conolle, Michelle; Schmitz, Jörn E; Brendel, Christian; Humeau, Laurent M; Dropulic, Boro; Rossi, John J; Berger, Annemarie; von Laer, Dorothee; Johnson, R Paul

    2010-08-23

    Although a variety of genetic strategies have been developed to inhibit HIV replication, few direct comparisons of the efficacy of these inhibitors have been carried out. Moreover, most studies have not examined whether genetic inhibitors are able to induce a survival advantage that results in an expansion of genetically-modified cells following HIV infection. We evaluated the efficacy of three leading genetic strategies to inhibit HIV replication: 1) an HIV-1 tat/rev-specific small hairpin (sh) RNA; 2) an RNA antisense gene specific for the HIV-1 envelope; and 3) a viral entry inhibitor, maC46. In stably transduced cell lines selected such that >95% of cells expressed the genetic inhibitor, the RNA antisense envelope and viral entry inhibitor maC46 provided the strongest inhibition of HIV-1 replication. However, when mixed populations of transduced and untransduced cells were challenged with HIV-1, the maC46 fusion inhibitor resulted in highly efficient positive selection of transduced cells, an effect that was evident even in mixed populations containing as few as 1% maC46-expressing cells. The selective advantage of the maC46 fusion inhibitor was also observed in HIV-1-infected cultures of primary T lymphocytes as well as in HIV-1-infected humanized mice. These results demonstrate robust inhibition of HIV replication with the fusion inhibitor maC46 and the antisense Env inhibitor, and importantly, a survival advantage of cells expressing the maC46 fusion inhibitor both in vitro and in vivo. Evaluation of the ability of genetic inhibitors of HIV-1 replication to confer a survival advantage on genetically-modified cells provides unique information not provided by standard techniques that may be important in the in vivo efficacy of these genes.

  14. Specific interaction of CXCR4 with CD4 and CD8α: Functional analysis of the CD4/CXCR4 interaction in the context of HIV-1 envelope glycoprotein-mediated membrane fusion

    International Nuclear Information System (INIS)

    Basmaciogullari, Stephane; Pacheco, Beatriz; Bour, Stephan; Sodroski, Joseph

    2006-01-01

    We investigated possible interactions between HIV-1 receptor (CD4) and the main coreceptors CXCR4 and CCR5. We found that CD4 and CXCR4 coexpressed in 293T cells form a complex that can be immunoprecipitated with antibodies directed against the extracellular domain of either protein. Mutagenesis revealed that the CD4/CXCR4 interaction maps to two previously uncharacterized basic motifs in the cytoplasmic domain of CD4. HIV-1 envelope glycoprotein-mediated membrane fusion was found to be independent of the ability of CD4 and CXCR4 to interact, whether fusion was studied in a virus-cell or a cell-cell model. However, this interaction might explain the adaptation of HIV-1 to CXCR4 as an alternative to CCR5. We found that CXCR4 also interacts with the cytoplasmic domain of CD8α in a way that is similar to the CD4/CXCR4 interaction. The CD4/CXCR4 and CD8α/CXCR4 interactions may thus be involved in cellular signaling pathways shared by the CD4 and CD8α molecules

  15. Fusion breeder

    International Nuclear Information System (INIS)

    Moir, R.W.

    1982-01-01

    The fusion breeder is a fusion reactor designed with special blankets to maximize the transmutation by 14 MeV neutrons of uranium-238 to plutonium or thorium to uranium-233 for use as a fuel for fission reactors. Breeding fissile fuels has not been a goal of the US fusion energy program. This paper suggests it is time for a policy change to make the fusion breeder a goal of the US fusion program and the US nuclear energy program. The purpose of this paper is to suggest this policy change be made and tell why it should be made, and to outline specific research and development goals so that the fusion breeder will be developed in time to meet fissile fuel needs

  16. Subcellular localization of a PhoE-LacZ fusion protein in E. coli by protease accessibility experiments reveals an inner-membrane-spanning form of the protein

    NARCIS (Netherlands)

    Tommassen, J.P.M.; Kroon, T. de

    1987-01-01

    Protease accessibility experiments were employed to localize a PhoE-LacZ hybrid protein, encompassing a large N-terminal fragment of the outer membrane PhoE protein of E. coli, fused to β-galactosidase, at the subcellular level. In previous studies, this protein was shown to co-fractionate with the

  17. Controlled fusion; La fusion controlee

    Energy Technology Data Exchange (ETDEWEB)

    Bobin, J.L

    2005-07-01

    During the last fifty years the researches on controlled thermonuclear fusion reached great performance in the magnetic confinement (tokamaks) as in the inertial confinement (lasers). But the state of the art is not in favor of the apparition of the fusion in the energy market before the second half of the 21 century. To explain this opinion the author presents the fusion reactions of light nuclei and the problems bound to the magnetic confinement. (A.L.B.)

  18. Fusion devices

    International Nuclear Information System (INIS)

    Fowler, T.K.

    1977-01-01

    Three types of thermonuclear fusion devices currently under development are reviewed for an electric utilities management audience. Overall design features of laser fusion, tokamak, and magnetic mirror type reactors are described and illustrated. Thrusts and trends in current research on these devices that promise to improve performance are briefly reviewed. Twenty photographs and drawings are included

  19. EPR Spectroscopy Targets Structural Changes in the E. coli Membrane Fusion CusB upon Cu(I) Binding.

    Science.gov (United States)

    Meir, Aviv; Abdelhai, Ahmad; Moskovitz, Yoni; Ruthstein, Sharon

    2017-06-20

    Bacterial cells have developed sophisticated systems to deal with the toxicity of metal ions. Escherichia coli CusCFBA is a complex efflux system, responsible for transferring Cu(I) and Ag(I) ions; this system, located in the periplasm, involves four proteins, CusA, CusB, CusC, and CusF. CusA, CusB, and CusC are connected to one another in an oligomerization ratio of 3:6:3 CusA/CusB/CusC to form the CusCBA periplasm membrane transporter. CusB is an adaptor protein that connects the two membrane proteins CusA (inner membrane) and CusC (outer membrane). CusF is a metallochaperone that transfers Cu(I) and Ag(I) to the CusCBA transporter from the periplasm. The crystal structures of CusB, CusC, CusF, and the CusBA complex have been resolved, shedding some light on the efflux mechanism underlying this intriguing system. However, since CusB is an adaptor protein, its role in operating this system is significant, and should be understood in detail. Here, we utilize EPR spectroscopy to target the conformational changes that take place in the full CusB protein upon binding Cu(I). We reveal that CusB is a dimer in solution, and that the orientation of one molecule with respect to the other molecule changes upon Cu(I) coordination, resulting in a more compact CusB structure. These structural and topological changes upon Cu(I) binding probably play the role of a switch for opening the channel and transferring metal ions from CusB to CusC and out of the cell. Copyright © 2017 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  20. Membrane-Based Inverse Transition Cycling: An Improved Means for Purifying Plant-Derived Recombinant Protein-Elastin-Like Polypeptide Fusions

    Directory of Open Access Journals (Sweden)

    Udo Conrad

    2011-04-01

    Full Text Available Elastin-like peptide (ELP was fused to two different avian flu H5N1 antigens and expressed in transgenic tobacco plants. The presence of the ELP tag enhanced the accumulation of the heterologous proteins in the tobacco leaves. An effective membrane-based Inverse Transition Cycling was developed to recover the ELPylated antigens and antibodies from plant material. The functionality of both the ELPylated neuraminidase and an ELPylated nanobody was demonstrated.

  1. Quantitative analysis of lipid droplet fusion: inefficient steady state fusion but rapid stimulation by chemical fusogens.

    Directory of Open Access Journals (Sweden)

    Samantha Murphy

    2010-12-01

    Full Text Available Lipid droplets (LDs are dynamic cytoplasmic organelles containing neutral lipids and bounded by a phospholipid monolayer. Previous studies have suggested that LDs can undergo constitutive homotypic fusion, a process linked to the inhibitory effects of fatty acids on glucose transporter trafficking. Using strict quantitative criteria for LD fusion together with refined light microscopic methods and real-time analysis, we now show that LDs in diverse cell types show low constitutive fusogenic activity under normal growth conditions. To investigate the possible modulation of LD fusion, we screened for agents that can trigger fusion. A number of pharmacological agents caused homotypic fusion of lipid droplets in a variety of cell types. This provided a novel cell system to study rapid regulated fusion between homotypic phospholipid monolayers. LD fusion involved an initial step in which the two adjacent membranes became continuous (<10 s, followed by the slower merging (100 s of the neutral lipid cores to produce a single spherical LD. These fusion events were accompanied by changes to the LD surface organization. Measurements of LDs undergoing homotypic fusion showed that fused LDs maintained their initial volume, with a corresponding decrease in surface area suggesting rapid removal of membrane from the fused LD. This study provides estimates for the level of constitutive LD fusion in cells and questions the role of LD fusion in vivo. In addition, it highlights the extent of LD restructuring which occurs when homotypic LD fusion is triggered in a variety of cell types.

  2. Membrane fusion proteins of type I secretion system and tripartite efflux pumps share a binding motif for TolC in gram-negative bacteria.

    Directory of Open Access Journals (Sweden)

    Minho Lee

    Full Text Available The Hly translocator complex of Escherichia coli catalyzes type I secretion of the toxin hemolysin A (HlyA. In this complex, HlyB is an inner membrane ABC (ATP Binding Cassette-type transporter, TolC is an outer membrane channel protein, and HlyD is a periplasmic adaptor anchored in the inner membrane that bridges HlyB to TolC. This tripartite organization is reminiscent of that of drug efflux systems such as AcrA-AcrB-TolC and MacA-MacB-TolC of E. coli. We have previously shown the crucial role of conserved residues located at the hairpin tip region of AcrA and MacA adaptors during assembly of their cognate systems. In this study, we investigated the role of the putative tip region of HlyD using HlyD mutants with single amino acid substitutions at the conserved positions. In vivo and in vitro data show that all mutations abolished HlyD binding to TolC and resulted in the absence of HlyA secretion. Together, our results suggest that, similarly to AcrA and MacA, HlyD interacts with TolC in a tip-to-tip manner. A general model in which these conserved interactions induce opening of TolC during drug efflux and type I secretion is discussed.

  3. Membrane Fusion Proteins of Type I Secretion System and Tripartite Efflux Pumps Share a Binding Motif for TolC in Gram-Negative Bacteria

    Science.gov (United States)

    Yoon, Bo-Young; Song, Saemee; Lee, Kangseok; Ha, Nam-Chul

    2012-01-01

    The Hly translocator complex of Escherichia coli catalyzes type I secretion of the toxin hemolysin A (HlyA). In this complex, HlyB is an inner membrane ABC (ATP Binding Cassette)-type transporter, TolC is an outer membrane channel protein, and HlyD is a periplasmic adaptor anchored in the inner membrane that bridges HlyB to TolC. This tripartite organization is reminiscent of that of drug efflux systems such as AcrA-AcrB-TolC and MacA-MacB-TolC of E. coli. We have previously shown the crucial role of conserved residues located at the hairpin tip region of AcrA and MacA adaptors during assembly of their cognate systems. In this study, we investigated the role of the putative tip region of HlyD using HlyD mutants with single amino acid substitutions at the conserved positions. In vivo and in vitro data show that all mutations abolished HlyD binding to TolC and resulted in the absence of HlyA secretion. Together, our results suggest that, similarly to AcrA and MacA, HlyD interacts with TolC in a tip-to-tip manner. A general model in which these conserved interactions induce opening of TolC during drug efflux and type I secretion is discussed. PMID:22792337

  4. Osteoclast Fusion

    DEFF Research Database (Denmark)

    Marie Julie Møller, Anaïs; Delaissé, Jean-Marie; Søe, Kent

    2017-01-01

    Investigations addressing the molecular keys of osteoclast fusion are primarily based on end-point analyses. No matter if investigations are performed in vivo or in vitro the impact of a given factor is predominantly analyzed by counting the number of multi-nucleated cells, the number of nuclei per...... multinucleated cell or TRAcP activity. But end-point analyses do not show how the fusion came about. This would not be a problem if fusion of osteoclasts was a random process and occurred by the same molecular mechanism from beginning to end. However, we and others have in the recent period published data...... suggesting that fusion partners may specifically select each other and that heterogeneity between the partners seems to play a role. Therefore, we set out to directly test the hypothesis that fusion factors have a heterogenic involvement at different stages of nuclearity. Therefore, we have analyzed...

  5. Fusion physics

    CERN Document Server

    Lackner, Karl; Tran, Minh Quang

    2012-01-01

    This publication is a comprehensive reference for graduate students and an invaluable guide for more experienced researchers. It provides an introduction to nuclear fusion and its status and prospects, and features specialized chapters written by leaders in the field, presenting the main research and development concepts in fusion physics. It starts with an introduction to the case for the development of fusion as an energy source. Magnetic and inertial confinement are addressed. Dedicated chapters focus on the physics of confinement, the equilibrium and stability of tokamaks, diagnostics, heating and current drive by neutral beam and radiofrequency waves, and plasma–wall interactions. While the tokamak is a leading concept for the realization of fusion, other concepts (helical confinement and, in a broader sense, other magnetic and inertial configurations) are also addressed in the book. At over 1100 pages, this publication provides an unparalleled resource for fusion physicists and engineers.

  6. Molecular dynamics simulations of lipid vesicle fusion in atomic detail

    NARCIS (Netherlands)

    Knecht, Volker; Marrink, Siewert-Jan

    The fusion of a membrane-bounded vesicle with a target membrane is a key step in intracellular trafficking, exocytosis, and drug delivery. Molecular dynamics simulations have been used to study the fusion of small unilamellar vesicles composed of a dipalmitoyl-phosphatidylcholine (DPPC)/palmitic

  7. Role of the synaptobrevin C terminus in fusion pore formation

    DEFF Research Database (Denmark)

    Ngatchou, Annita N; Kisler, Kassandra; Fang, Qinghua

    2010-01-01

    stimulation, the SNARE complex pulls the C terminus of sybII deeper into the vesicle membrane. We propose that this movement disrupts the vesicular membrane continuity leading to fusion pore formation. In contrast to current models, the experiments suggest that fusion pore formation begins with molecular......Neurotransmitter release is mediated by the SNARE proteins synaptobrevin II (sybII, also known as VAMP2), syntaxin, and SNAP-25, generating a force transfer to the membranes and inducing fusion pore formation. However, the molecular mechanism by which this force leads to opening of a fusion pore...

  8. Tension-induced vesicle fusion: pathways and pore dynamics

    DEFF Research Database (Denmark)

    Shillcock, Julian C.

    2008-01-01

    The dynamics of tension-induced fusion of two vesicles is studied using dissipative particle dynamics (DPD) simulations. The vesicle membranes use an improved DPD parameter set that results in their sustaining only a 10–30% relative area stretch before rupturing on the microsecond timescale...... fusion time on membrane tension implies that the fusion process is completed by overcoming two energy barriers with scales of 13kBT and 11kBT. The fusion pore radius as a function of time has also been extracted from the simulations, and provides a quantitative measure of the fusion dynamics which...

  9. Efficient Activation of Human T Cells of Both CD4 and CD8 Subsets by Urease-Deficient Recombinant Mycobacterium bovis BCG That Produced a Heat Shock Protein 70-M. tuberculosis-Derived Major Membrane Protein II Fusion Protein

    Science.gov (United States)

    Mukai, Tetsu; Tsukamoto, Yumiko; Maeda, Yumi; Tamura, Toshiki

    2014-01-01

    For the purpose of obtaining Mycobacterium bovis bacillus Calmette-Guérin (BCG) capable of activating human naive T cells, urease-deficient BCG expressing a fusion protein composed of Mycobacterium tuberculosis-derived major membrane protein II (MMP-II) and heat shock protein 70 (HSP70) of BCG (BCG-DHTM) was produced. BCG-DHTM secreted the HSP70-MMP-II fusion protein and effectively activated human monocyte-derived dendritic cells (DCs) by inducing phenotypic changes and enhanced cytokine production. BCG-DHTM-infected DCs activated naive T cells of both CD4 and naive CD8 subsets, in an antigen (Ag)-dependent manner. The T cell activation induced by BCG-DHTM was inhibited by the pretreatment of DCs with chloroquine. The naive CD8+ T cell activation was mediated by the transporter associated with antigen presentation (TAP) and the proteosome-dependent cytosolic cross-priming pathway. Memory CD8+ T cells and perforin-producing effector CD8+ T cells were efficiently produced from the naive T cell population by BCG-DHTM stimulation. Single primary infection with BCG-DHTM in C57BL/6 mice efficiently produced T cells responsive to in vitro secondary stimulation with HSP70, MMP-II, and M. tuberculosis-derived cytosolic protein and inhibited the multiplication of subsequently aerosol-challenged M. tuberculosis more efficiently than did vector control BCG. These results indicate that the introduction of MMP-II and HSP70 into urease-deficient BCG may be useful for improving BCG for control of tuberculosis. PMID:24152387

  10. Deduced sequences of the membrane fusion and attachment proteins of canine distemper viruses isolated from dogs and wild animals in Korea.

    Science.gov (United States)

    Bae, Chae-Wun; Lee, Joong-Bok; Park, Seung-Yong; Song, Chang-Seon; Lee, Nak-Hyung; Seo, Kun-Ho; Kang, Young-Sun; Park, Choi-Kyu; Choi, In-Soo

    2013-08-01

    Canine distemper virus (CDV) causes highly contagious respiratory, gastrointestinal, and neurological diseases in wild and domestic animal species. Despite a broad vaccination campaign, the disease is still a serious problem worldwide. In this study, six field CDV strains were isolated from three dogs, two raccoon dogs, and one badger in Korea. The full sequence of the genes encoding fusion (F) and hemagglutinin (H) proteins were compared with those of other CDVs including field and vaccine strains. The phylogenetic analysis for the F and H genes indicated that the two CDV strains isolated from dogs were most closely related to Chinese strains in the Asia-1 genotype. Another four strains were closely related to Japanese strains in the Asia-2 genotype. The six currently isolated strains shared 90.2-92.1% and 88.2-91.8% identities with eight commercial vaccine strains in their nucleotide and amino acid sequences of the F protein, respectively. They also showed 90.1-91.4% and 87.8-90.7% identities with the same vaccine strains in their nucleotide and deduced amino acid sequences of the H protein, respectively. Different N-linked glycosylation sites were identified in the F and H genes of the six isolates from the prototype vaccine strain Onderstepoort. Collectively, these results demonstrate that at least two different CDV genotypes currently exist in Korea. The considerable genetic differences between the vaccine strains and wild-type isolates would be a major factor of the incomplete protection of dogs from CDV infections.

  11. Cell-Delivered Entry Inhibitors for HIV-1: CCR5 Downregulation and Blocking Virus/Membrane Fusion in Defending the Host Cell Population.

    Science.gov (United States)

    Symonds, Geoff; Bartlett, Jeffrey S; Kiem, Hans-Peter; Tsie, Marlene; Breton, Louis

    2016-12-01

    HIV-1 infection requires the presence of the CD4 receptor on the target cell surface and a coreceptor, predominantly CC-chemokine receptor 5 (CCR5). It has been shown that individuals who are homozygous for a defective CCR5 gene are protected from HIV-1 infection. A novel self-inactivating lentiviral vector LVsh5/C46 (Cal-1) has been engineered to block HIV-1 infection with two viral entry inhibitors, conferring resistance to HIV-1 infection from both CCR5 and CXCR4 tropic strains. Cal-1 encodes a short hairpin RNA (sh5) to downregulate CCR5 and C46, an HIV-1 fusion inhibitor. Gene therapy by Cal-1 is aimed at transducing CD4 + T cells and CD34 + hematopoietic stem/progenitor cells in an autologous transplant setting. Pre-clinical safety and efficacy studies in vitro and in vivo (humanized mouse model and nonhuman primates) have shown that Cal-1 is safe with no indication of any toxicity risk and acts to decrease viral load and increase CD4 counts. Two clinical trials are underway using Cal-1: a phase I/II study to assess safety and feasibility in an adult HIV-1-positive population not on antiretroviral therapy (ART); and a second Fred Hutchinson Investigator Initiated phase I study to assess safety and feasibility in adults with HIV-1-associated non-Hodgkin or Hodgkin lymphoma.

  12. Cold fusion

    International Nuclear Information System (INIS)

    Suh, Suk Yong; Sung, Ki Woong; Kang, Joo Sang; Lee, Jong Jik

    1995-02-01

    So called 'cold fusion phenomena' are not confirmed yet. Excess heat generation is very delicate one. Neutron generation is most reliable results, however, the records are erratic and the same results could not be repeated. So there is no reason to exclude the malfunction of testing instruments. The same arguments arise in recording 4 He, 3 He, 3 H, which are not rich in quantity basically. An experiment where plenty of 4 He were recorded is attached in appendix. The problem is that we are trying to search cold fusion which is permitted by nature or not. The famous tunneling effect in quantum mechanics will answer it, however, the most fusion rate is known to be negligible. The focus of this project is on the theme that how to increase that negligible fusion rate. 6 figs, 4 tabs, 1512 refs. (Author)

  13. Cold fusion

    Energy Technology Data Exchange (ETDEWEB)

    Suh, Suk Yong; Sung, Ki Woong; Kang, Joo Sang; Lee, Jong Jik [Korea Atomic Energy Research Institute, Taejon (Korea, Republic of)

    1995-02-01

    So called `cold fusion phenomena` are not confirmed yet. Excess heat generation is very delicate one. Neutron generation is most reliable results, however, the records are erratic and the same results could not be repeated. So there is no reason to exclude the malfunction of testing instruments. The same arguments arise in recording {sup 4}He, {sup 3}He, {sup 3}H, which are not rich in quantity basically. An experiment where plenty of {sup 4}He were recorded is attached in appendix. The problem is that we are trying to search cold fusion which is permitted by nature or not. The famous tunneling effect in quantum mechanics will answer it, however, the most fusion rate is known to be negligible. The focus of this project is on the theme that how to increase that negligible fusion rate. 6 figs, 4 tabs, 1512 refs. (Author).

  14. The hallmarks of cell-cell fusion.

    Science.gov (United States)

    Hernández, Javier M; Podbilewicz, Benjamin

    2017-12-15

    Cell-cell fusion is essential for fertilization and organ development. Dedicated proteins known as fusogens are responsible for mediating membrane fusion. However, until recently, these proteins either remained unidentified or were poorly understood at the mechanistic level. Here, we review how fusogens surmount multiple energy barriers to mediate cell-cell fusion. We describe how early preparatory steps bring membranes to a distance of ∼10 nm, while fusogens act in the final approach between membranes. The mechanical force exerted by cell fusogens and the accompanying lipidic rearrangements constitute the hallmarks of cell-cell fusion. Finally, we discuss the relationship between viral and eukaryotic fusogens, highlight a classification scheme regrouping a superfamily of fusogens called Fusexins, and propose new questions and avenues of enquiry. © 2017. Published by The Company of Biologists Ltd.

  15. Fusion energy

    International Nuclear Information System (INIS)

    Anon.

    1979-01-01

    The efforts of the Chemical Technology Division in fusion energy include the areas of fuel handling, processing, and containment. Current studies are concerned largely with the development of vacuum pumps for fusion reactors and experiments and with development and evaluation of techniques for recovering tritium from solid or liquid breeding blankets. In addition, a small effort is devoted to support of the ORNL design of a major Tokamak experiment, The Next Step (TNS)

  16. Rho GTPase activity modulates paramyxovirus fusion protein-mediated cell-cell fusion

    International Nuclear Information System (INIS)

    Schowalter, Rachel M.; Wurth, Mark A.; Aguilar, Hector C.; Lee, Benhur; Moncman, Carole L.; McCann, Richard O.; Dutch, Rebecca Ellis

    2006-01-01

    The paramyxovirus fusion protein (F) promotes fusion of the viral envelope with the plasma membrane of target cells as well as cell-cell fusion. The plasma membrane is closely associated with the actin cytoskeleton, but the role of actin dynamics in paramyxovirus F-mediated membrane fusion is unclear. We examined cell-cell fusion promoted by two different paramyxovirus F proteins in three cell types in the presence of constitutively active Rho family GTPases, major cellular coordinators of actin dynamics. Reporter gene and syncytia assays demonstrated that expression of either Rac1 V12 or Cdc42 V12 could increase cell-cell fusion promoted by the Hendra or SV5 glycoproteins, though the effect was dependent on the cell type expressing the viral glycoproteins. In contrast, RhoA L63 decreased cell-cell fusion promoted by Hendra glycoproteins but had little affect on SV5 F-mediated fusion. Also, data suggested that GTPase activation in the viral glycoprotein-containing cell was primarily responsible for changes in fusion. Additionally, we found that activated Cdc42 promoted nuclear rearrangement in syncytia

  17. Destination fusion

    International Nuclear Information System (INIS)

    Shivakumar, B.; Shapira, D.; Stelson, P.H.; Beckerman, M.; Harmon, B.A.; Teh, K.; Ayik, S.

    1986-01-01

    The orbiting yields of reaction products from the 28 Si + 14 N interaction have been measured. The relative magnitudes of the orbiting yields indicate that the dinuclear complex (DNCs) formed in such interactions live sufficiently long to permit the equilibration of charge and mass. Since both the 16 O and the 12 C orbiting yields exceed the 14 N, it appears that there is no preferred directions for mass flow between the interacting nuclei. Since the orbiting yields are typically 10% of the fusion yield, and we believe the orbiting process reflects how the DNCs formed in such collisions evolve towards fusion, it seems apt to conclude that fusion occurs not through a process of continual particle exchange whereby one nucleus is gradually consumed by the other, but by a dinuclear system that retains it mass asymmetry. 17 refs., 2 figs., 1 tab

  18. Hendra virus fusion protein transmembrane domain contributes to pre-fusion protein stability.

    Science.gov (United States)

    Webb, Stacy; Nagy, Tamas; Moseley, Hunter; Fried, Michael; Dutch, Rebecca

    2017-04-07

    Enveloped viruses utilize fusion (F) proteins studding the surface of the virus to facilitate membrane fusion with a target cell membrane. Fusion of the viral envelope with a cellular membrane is required for release of viral genomic material, so the virus can ultimately reproduce and spread. To drive fusion, the F protein undergoes an irreversible conformational change, transitioning from a metastable pre-fusion conformation to a more thermodynamically stable post-fusion structure. Understanding the elements that control stability of the pre-fusion state and triggering to the post-fusion conformation is important for understanding F protein function. Mutations in F protein transmembrane (TM) domains implicated the TM domain in the fusion process, but the structural and molecular details in fusion remain unclear. Previously, analytical ultracentrifugation was utilized to demonstrate that isolated TM domains of Hendra virus F protein associate in a monomer-trimer equilibrium (Smith, E. C., Smith, S. E., Carter, J. R., Webb, S. R., Gibson, K. M., Hellman, L. M., Fried, M. G., and Dutch, R. E. (2013) J. Biol. Chem. 288, 35726-35735). To determine factors driving this association, 140 paramyxovirus F protein TM domain sequences were analyzed. A heptad repeat of β-branched residues was found, and analysis of the Hendra virus F TM domain revealed a heptad repeat leucine-isoleucine zipper motif (LIZ). Replacement of the LIZ with alanine resulted in dramatically reduced TM-TM association. Mutation of the LIZ in the whole protein resulted in decreased protein stability, including pre-fusion conformation stability. Together, our data suggest that the heptad repeat LIZ contributed to TM-TM association and is important for F protein function and pre-fusion stability. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  19. Fusion cuisine

    DEFF Research Database (Denmark)

    Peters, Chris; Broersma, Marcel

    2018-01-01

    define the culinary tradition of journalism studies in the first place. In so doing, we offer a recipe for journalism studies fusion cooking that: 1) considers technological change (audiences’ diets); 2) analyses institutional change (audiences’ supermarket of information); and 3) evaluates journalism...

  20. Magnetic fusion

    International Nuclear Information System (INIS)

    2002-01-01

    This document is a detailed lecture on thermonuclear fusion. The basic physics principles are recalled and the technological choices that have led to tokamaks or stellarators are exposed. Different aspects concerning thermonuclear reactors such as safety, economy and feasibility are discussed. Tore-supra is described in details as well as the ITER project

  1. Fusion systems

    OpenAIRE

    Aschbacher, Michael; Oliver, Bob

    2016-01-01

    This is a survey article on the theory of fusion systems, a relatively new area of mathematics with connections to local finite group theory, algebraic topology, and modular representation theory. We first describe the general theory and then look separately at these connections.

  2. Fusion of small unilamellar vesicles with viable EDTA-treated Escherichia coli cells.

    OpenAIRE

    Marvin, H J; ter Beest, M B; Hoekstra, D; Witholt, B

    1989-01-01

    Fusion characteristics of EDTA-treated Escherichia coli cells with small unilamellar vesicles were investigated, using a membrane fusion assay based on resonance energy transfer. Ca2+-EDTA treatments of Escherichia coli O111:B4 (wild type), E. coli C600 (rough), and E. coli D21f2 (deep rough) which permeabilize the outer membrane by inducing the release of lipopolysaccharide and outer membrane proteins resulted in fusion activity of the intact and viable bacteria with small unilamellar vesicl...

  3. Gene expression profiling of Drosophila tracheal fusion cells.

    Science.gov (United States)

    Chandran, Rachana R; Iordanou, Ekaterini; Ajja, Crystal; Wille, Michael; Jiang, Lan

    2014-07-01

    The Drosophila trachea is a premier genetic system to investigate the fundamental mechanisms of tubular organ formation. Tracheal fusion cells lead the branch fusion process to form an interconnected tubular network. Therefore, fusion cells in the Drosophila trachea will be an excellent model to study branch fusion in mammalian tubular organs, such as kidneys and blood vessels. The fusion process is a dynamic cellular process involving cell migration, adhesion, vesicle trafficking, cytoskeleton rearrangement, and membrane fusion. To understand how these cellular events are coordinated, we initiated the critical step to assemble a gene expression profile of fusion cells. For this study, we analyzed the expression of 234 potential tracheal-expressed genes in fusion cells during fusion cell development. 143 Tracheal genes were found to encode transcription factors, signal proteins, cytoskeleton and matrix proteins, transporters, and proteins with unknown function. These genes were divided into four subgroups based on their levels of expression in fusion cells compared to neighboring non-fusion cells revealed by in situ hybridization: (1) genes that have relative high abundance in fusion cells, (2) genes that are dynamically expressed in fusion cells, (3) genes that have relative low abundance in fusion cells, and (4) genes that are expressed at similar levels in fusion cells and non-fusion tracheal cells. This study identifies the expression profile of fusion cells and hypothetically suggests genes which are necessary for the fusion process and which play roles in distinct stages of fusion, as indicated by the location and timing of expression. These data will provide the basis for a comprehensive understanding of the molecular and cellular mechanisms of branch fusion. Copyright © 2014 Elsevier B.V. All rights reserved.

  4. Cold fusion, Alchemist's dream

    International Nuclear Information System (INIS)

    Clayton, E.D.

    1989-09-01

    In this report the following topics relating to cold fusion are discussed: muon catalysed cold fusion; piezonuclear fusion; sundry explanations pertaining to cold fusion; cosmic ray muon catalysed cold fusion; vibrational mechanisms in excited states of D 2 molecules; barrier penetration probabilities within the hydrogenated metal lattice/piezonuclear fusion; branching ratios of D 2 fusion at low energies; fusion of deuterons into 4 He; secondary D+T fusion within the hydrogenated metal lattice; 3 He to 4 He ratio within the metal lattice; shock induced fusion; and anomalously high isotopic ratios of 3 He/ 4 He

  5. Cold fusion, Alchemist's dream

    Energy Technology Data Exchange (ETDEWEB)

    Clayton, E.D.

    1989-09-01

    In this report the following topics relating to cold fusion are discussed: muon catalysed cold fusion; piezonuclear fusion; sundry explanations pertaining to cold fusion; cosmic ray muon catalysed cold fusion; vibrational mechanisms in excited states of D{sub 2} molecules; barrier penetration probabilities within the hydrogenated metal lattice/piezonuclear fusion; branching ratios of D{sub 2} fusion at low energies; fusion of deuterons into {sup 4}He; secondary D+T fusion within the hydrogenated metal lattice; {sup 3}He to {sup 4}He ratio within the metal lattice; shock induced fusion; and anomalously high isotopic ratios of {sup 3}He/{sup 4}He.

  6. Magnetic fusion; La fusion magnetique

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    2002-07-01

    This document is a detailed lecture on thermonuclear fusion. The basic physics principles are recalled and the technological choices that have led to tokamaks or stellarators are exposed. Different aspects concerning thermonuclear reactors such as safety, economy and feasibility are discussed. Tore-supra is described in details as well as the ITER project.

  7. Vesicle Fusion Mediated by Solanesol-Anchored DNA.

    Science.gov (United States)

    Flavier, Kristina M; Boxer, Steven G

    2017-09-19

    Fusion between two lipid bilayers is one of the central processes in cell biology, playing a key role in endocytosis, exocytosis, and vesicle transport. We have previously developed a model system that uses the hybridization of complementary DNA strands to model the formation of the SNARE four-helix bundle that mediates synaptic vesicle fusion and used it to study vesicle fusion to a tethered lipid bilayer. Using single vesicle assays, 70% of observed fusion events in the DNA-lipid system are arrested at the hemifusion stage, whereas only 5% eventually go to full fusion. This may be because the diglycerol ether that anchors the DNA in the membrane spans only half the bilayer: upon hemifusion and mixing of the outer leaflets, the DNA-lipid is free to diffuse into the target membrane and away from the vesicle. Here, we test the hypothesis that the length of the membrane anchor may impact the outcome by comparing single leaflet-spanning DNA-lipid mediated vesicle fusion with fusion mediated by DNA anchored by solanesol, a C45 isoprenoid of sufficient length to span the bilayer. When the solanesol anchor was present on the incoming vesicles, target membrane, or both, ∼2-3 times as much full fusion was observed as in the DNA-lipid mediated system, as measured by lipid mixing or content transfer. These results indicate that a transmembrane anchor increases the efficiency of full fusion. Copyright © 2017 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  8. Laser fusion

    International Nuclear Information System (INIS)

    Eliezer, S.

    1982-02-01

    In this paper, the physics of laser fusion is described on an elementary level. The irradiated matter consists of a dense inner core surrounded by a less dense plasma corona. The laser radiation is mainly absorbed in the outer periphery of the plasma. The absorbed energy is transported inward to the ablation surface where plasma flow is created. Due to this plasma flow, a sequence of inward going shock waves and heat waves are created, resulting in the compression and heating of the core to high density and temperature. The interaction physics between laser and matter leading to thermonuclear burn is summarized by the following sequence of events: Laser absorption → Energy transport → Compression → Nuclear Fusion. This scenario is shown in particular for a Nd:laser with a wavelength of 1 μm. The wavelength scaling of the physical processes is also discussed. In addition to the laser-plasma physics, the Nd high power pulsed laser is described. We give a very brief description of the oscillator, the amplifiers, the spatial filters, the isolators and the diagnostics involved. Last, but not least, the concept of reactors for laser fusion and the necessary laser system are discussed. (author)

  9. Distinct Requirements for HIV-Cell Fusion and HIV-mediated Cell-Cell Fusion*

    Science.gov (United States)

    Kondo, Naoyuki; Marin, Mariana; Kim, Jeong Hwa; Desai, Tanay M.; Melikyan, Gregory B.

    2015-01-01

    Whether HIV-1 enters cells by fusing with the plasma membrane or with endosomes is a subject of active debate. The ability of HIV-1 to mediate fusion between adjacent cells, a process referred to as “fusion-from-without” (FFWO), shows that this virus can fuse with the plasma membrane. To compare FFWO occurring at the cell surface with HIV-cell fusion through a conventional entry route, we designed an experimental approach that enabled the measurements of both processes in the same sample. The following key differences were observed. First, a very small fraction of viruses fusing with target cells participated in FFWO. Second, whereas HIV-1 fusion with adherent cells was insensitive to actin inhibitors, post-CD4/coreceptor binding steps during FFWO were abrogated. A partial dependence of HIV-cell fusion on actin remodeling was observed in CD4+ T cells, but this effect appeared to be due to the actin dependence of virus uptake. Third, deletion of the cytoplasmic tail of HIV-1 gp41 dramatically enhanced the ability of the virus to promote FFWO, while having a modest effect on virus-cell fusion. Distinct efficiencies and actin dependences of FFWO versus HIV-cell fusion are consistent with the notion that, except for a minor fraction of particles that mediate fusion between the plasma membranes of adjacent cells, HIV-1 enters through an endocytic pathway. We surmise, however, that cell-cell contacts enabling HIV-1 fusion with the plasma membrane could be favored at the sites of high density of target cells, such as lymph nodes. PMID:25589785

  10. Radioactivity and fusion energy

    International Nuclear Information System (INIS)

    Kudo, H.

    1995-01-01

    Nuclear fusion is expected to give an ultimate solution to energy problems over the long term. From recent progress in developing technology for fusion reactors, we can anticipate a prototype fusion reactor by 2030. This review article describes the present status of nuclear fusion research, including muon catalyzed fusion (μCF) which attracts quite new physical interest. Tritium is an essential component of fusion reactors, because the first-stage fusion reactors will utilize a mixture of deuterium and tritium as their fuel. The knowledge about tritium as well as the fusion-neutron induced radioactivity is summarized in terms of nuclear fusion research. (orig.)

  11. Distinct roles for key karyogamy proteins during yeast nuclear fusion.

    Science.gov (United States)

    Melloy, Patricia; Shen, Shu; White, Erin; Rose, Mark D

    2009-09-01

    During yeast mating, cell fusion is followed by the congression and fusion of the two nuclei. Proteins required for nuclear fusion are found at the surface (Prm3p) and within the lumen (Kar2p, Kar5p, and Kar8p) of the nuclear envelope (NE). Electron tomography (ET) of zygotes revealed that mutations in these proteins block nuclear fusion with different morphologies, suggesting that they act in different steps of fusion. Specifically, prm3 zygotes were blocked before formation of membrane bridges, whereas kar2, kar5, and kar8 zygotes frequently contained them. Membrane bridges were significantly larger and occurred more frequently in kar2 and kar8, than in kar5 mutant zygotes. The kinetics of NE fusion in prm3, kar5, and kar8 mutants, measured by live-cell fluorescence microscopy, were well correlated with the size and frequency of bridges observed by ET. However the kar2 mutant was defective for transfer of NE lumenal GFP, but not diffusion within the lumen, suggesting that transfer was blocked at the NE fusion junction. These observations suggest that Prm3p acts before initiation of outer NE fusion, Kar5p may help dilation of the initial fusion pore, and Kar2p and Kar8p act after outer NE fusion, during inner NE fusion.

  12. C-terminal tyrosine residues modulate the fusion activity of the Hendra virus fusion protein.

    Science.gov (United States)

    Popa, Andreea; Pager, Cara Teresia; Dutch, Rebecca Ellis

    2011-02-15

    The paramyxovirus family includes important human pathogens such as measles, mumps, respiratory syncytial virus, and the recently emerged, highly pathogenic Hendra and Nipah viruses. The viral fusion (F) protein plays critical roles in infection, promoting both the virus-cell membrane fusion events needed for viral entry as well as cell-cell fusion events leading to syncytia formation. We describe the surprising finding that addition of the short epitope HA tag to the cytoplasmic tail (CT) of the Hendra virus F protein leads to a significant increase in the extent of cell-cell membrane fusion. This increase was not due to alterations in surface expression, cleavage state, or association with lipid microdomains. Addition of a Myc tag of similar length did not alter Hendra F protein fusion activity, indicating that the observed stimulation was not solely a result of lengthening the CT. Three tyrosine residues within the HA tag were critical for the increase in the extent of fusion, suggesting C-terminal tyrosines may modulate Hendra fusion activity. The effects of addition of the HA tag varied with other fusion proteins, as parainfluenza virus 5 F-HA showed a decreased level of surface expression and no stimulation of fusion. These results indicate that additions to the C-terminal end of the F protein CT can modulate protein function in a sequence specific manner, reinforcing the need for careful analysis of epitope-tagged glycoproteins. In addition, our results implicate C-terminal tyrosine residues in the modulation of the membrane fusion reaction promoted by these viral glycoproteins.

  13. Dynamic clustering and dispersion of lipid rafts contribute to fusion competence of myogenic cells

    International Nuclear Information System (INIS)

    Mukai, Atsushi; Kurisaki, Tomohiro; Sato, Satoshi B.; Kobayashi, Toshihide; Kondoh, Gen; Hashimoto, Naohiro

    2009-01-01

    Recent research indicates that the leading edge of lamellipodia of myogenic cells (myoblasts and myotubes) contains presumptive fusion sites, yet the mechanisms that render the plasma membrane fusion-competent remain largely unknown. Here we show that dynamic clustering and dispersion of lipid rafts contribute to both cell adhesion and plasma membrane union during myogenic cell fusion. Adhesion-complex proteins including M-cadherin, β-catenin, and p120-catenin accumulated at the leading edge of lamellipodia, which contains the presumptive fusion sites of the plasma membrane, in a lipid raft-dependent fashion prior to cell contact. In addition, disruption of lipid rafts by cholesterol depletion directly prevented the membrane union of myogenic cell fusion. Time-lapse recording showed that lipid rafts were laterally dispersed from the center of the lamellipodia prior to membrane fusion. Adhesion proteins that had accumulated at lipid rafts were also removed from the presumptive fusion sites when lipid rafts were laterally dispersed. The resultant lipid raft- and adhesion complex-free area at the leading edge fused with the opposing plasma membrane. These results demonstrate a key role for dynamic clustering/dispersion of lipid rafts in establishing fusion-competent sites of the myogenic cell membrane, providing a novel mechanistic insight into the regulation of myogenic cell fusion.

  14. Dynamic clustering and dispersion of lipid rafts contribute to fusion competence of myogenic cells

    Energy Technology Data Exchange (ETDEWEB)

    Mukai, Atsushi [Department of Regenerative Medicine, National Institute for Longevity Sciences, National Center for Geriatrics and Gerontology, 36-3 Gengo, Morioka, Oobu, Aichi 474-8522 (Japan); Kurisaki, Tomohiro [Department of Growth Regulation, Institute for Frontier Medical Sciences, Kyoto University, 53 Shogoin-Kawahara-cho, Sakyo-ku, Kyoto 606-8507 (Japan); Sato, Satoshi B. [Research Center for Low Temperature and Material Sciences, Kyoto University, Yoshida-honmachi, Kyoto 606-8501 (Japan); Kobayashi, Toshihide [Lipid Biology Laboratory, Discovery Research Institute, RIKEN, Wako, Saitama 351-0198 (Japan); Kondoh, Gen [Laboratory of Animal Experiments for Regeneration, Institute for Frontier Medical Sciences, Kyoto University, 53 Shogoin-Kawahara-cho, Sakyo-ku, Kyoto 606-8507 (Japan); Hashimoto, Naohiro, E-mail: nao@nils.go.jp [Department of Regenerative Medicine, National Institute for Longevity Sciences, National Center for Geriatrics and Gerontology, 36-3 Gengo, Morioka, Oobu, Aichi 474-8522 (Japan)

    2009-10-15

    Recent research indicates that the leading edge of lamellipodia of myogenic cells (myoblasts and myotubes) contains presumptive fusion sites, yet the mechanisms that render the plasma membrane fusion-competent remain largely unknown. Here we show that dynamic clustering and dispersion of lipid rafts contribute to both cell adhesion and plasma membrane union during myogenic cell fusion. Adhesion-complex proteins including M-cadherin, {beta}-catenin, and p120-catenin accumulated at the leading edge of lamellipodia, which contains the presumptive fusion sites of the plasma membrane, in a lipid raft-dependent fashion prior to cell contact. In addition, disruption of lipid rafts by cholesterol depletion directly prevented the membrane union of myogenic cell fusion. Time-lapse recording showed that lipid rafts were laterally dispersed from the center of the lamellipodia prior to membrane fusion. Adhesion proteins that had accumulated at lipid rafts were also removed from the presumptive fusion sites when lipid rafts were laterally dispersed. The resultant lipid raft- and adhesion complex-free area at the leading edge fused with the opposing plasma membrane. These results demonstrate a key role for dynamic clustering/dispersion of lipid rafts in establishing fusion-competent sites of the myogenic cell membrane, providing a novel mechanistic insight into the regulation of myogenic cell fusion.

  15. PI(4,5)P2 regulates myoblast fusion through Arp2/3 regulator localization at the fusion site

    Science.gov (United States)

    Bothe, Ingo; Deng, Su; Baylies, Mary

    2014-01-01

    Cell-cell fusion is a regulated process that requires merging of the opposing membranes and underlying cytoskeletons. However, the integration between membrane and cytoskeleton signaling during fusion is not known. Using Drosophila, we demonstrate that the membrane phosphoinositide PI(4,5)P2 is a crucial regulator of F-actin dynamics during myoblast fusion. PI(4,5)P2 is locally enriched and colocalizes spatially and temporally with the F-actin focus that defines the fusion site. PI(4,5)P2 enrichment depends on receptor engagement but is upstream or parallel to actin remodeling. Regulators of actin branching via Arp2/3 colocalize with PI(4,5)P2 in vivo and bind PI(4,5)P2 in vitro. Manipulation of PI(4,5)P2 availability leads to impaired fusion, with a reduction in the F-actin focus size and altered focus morphology. Mechanistically, the changes in the actin focus are due to a failure in the enrichment of actin regulators at the fusion site. Moreover, improper localization of these regulators hinders expansion of the fusion interface. Thus, PI(4,5)P2 enrichment at the fusion site encodes spatial and temporal information that regulates fusion progression through the localization of activators of actin polymerization. PMID:24821989

  16. Gene transfer mediated by fusion protein hemagglutinin reconstituted in cationic lipid vesicles

    NARCIS (Netherlands)

    Schoen, P; Chonn, A; Cullis, PR; Wilschut, J; Scherrer, P

    Hemagglutinin, the membrane fusion protein of influenza virus,is known to mediate a low-pH-dependent fusion reaction between the viral envelope and the limiting membrane of the endosomal cell compartment following cellular uptake of the virus particles by receptor-mediated endocytosis. Here we

  17. Catalysed fusion

    CERN Document Server

    Farley, Francis

    2012-01-01

    A sizzling romance and a romp with subatomic particles at CERN. Love, discovery and adventure in the city where nations meet and beams collide. Life in a large laboratory. As always, the challenges are the same. Who leads? Who follows? Who succeeds? Who gets the credit? Who gets the women or the men? Young Jeremy arrives in CERN and joins the quest for green energy. Coping with baffling jargon and manifold dangers, he is distracted by radioactive rats, lovely ladies and an unscrupulous rival. Full of doubts and hesitations, he falls for a dazzling Danish girl, who leads him astray. His brilliant idea leads to a discovery and a new route to cold fusion. But his personal life is scrambled. Does it bring fame or failure? Tragedy or triumph?

  18. Efficient liposome fusion mediated by lipid-nucleic acid conjugates

    DEFF Research Database (Denmark)

    Ries, O; Löffler, P M G; Rabe, A

    2017-01-01

    The fusion of biomembranes with release of encapsulated content in a controlled way is crucial for cell signaling, endo- and exocytosis and intracellular trafficking. Programmable fusion of liposomes and an efficient mixing of their contents have the potential to enable the study of chemical...... and enzymatic processes in a confined environment and under crowded conditions outside biological systems. We report on DNA-controlled fusion of lipid bilayer membranes using lipid-nucleic acid conjugates (LiNAs) to mediate lipid and content mixing of liposomes. Screening of different membrane anchor and linker...

  19. The MARVEL domain protein, Singles Bar, is required for progression past the pre-fusion complex stage of myoblast fusion.

    Science.gov (United States)

    Estrada, Beatriz; Maeland, Anne D; Gisselbrecht, Stephen S; Bloor, James W; Brown, Nicholas H; Michelson, Alan M

    2007-07-15

    Multinucleated myotubes develop by the sequential fusion of individual myoblasts. Using a convergence of genomic and classical genetic approaches, we have discovered a novel gene, singles bar (sing), that is essential for myoblast fusion. sing encodes a small multipass transmembrane protein containing a MARVEL domain, which is found in vertebrate proteins involved in processes such as tight junction formation and vesicle trafficking where--as in myoblast fusion--membrane apposition occurs. sing is expressed in both founder cells and fusion competent myoblasts preceding and during myoblast fusion. Examination of embryos injected with double-stranded sing RNA or embryos homozygous for ethane methyl sulfonate-induced sing alleles revealed an identical phenotype: replacement of multinucleated myofibers by groups of single, myosin-expressing myoblasts at a stage when formation of the mature muscle pattern is complete in wild-type embryos. Unfused sing mutant myoblasts form clusters, suggesting that early recognition and adhesion of these cells are unimpaired. To further investigate this phenotype, we undertook electron microscopic ultrastructural studies of fusing myoblasts in both sing and wild-type embryos. These experiments revealed that more sing mutant myoblasts than wild-type contain pre-fusion complexes, which are characterized by electron-dense vesicles paired on either side of the fusing plasma membranes. In contrast, embryos mutant for another muscle fusion gene, blown fuse (blow), have a normal number of such complexes. Together, these results lead to the hypothesis that sing acts at a step distinct from that of blow, and that sing is required on both founder cell and fusion-competent myoblast membranes to allow progression past the pre-fusion complex stage of myoblast fusion, possibly by mediating fusion of the electron-dense vesicles to the plasma membrane.

  20. Fusion systems engineering

    International Nuclear Information System (INIS)

    Anon.

    1977-01-01

    Summaries of research are included for each of the following topics: (1) fusion reactor systems studies, (2) development of blanket processing technology for fusion reactors, (3) safety studies of fusion concepts, (4) the MACK/MACKLIB system for nuclear response functions, and (5) energy storage and power supply systems for fusion reactors

  1. Towards nuclear fusion reactors

    International Nuclear Information System (INIS)

    1993-11-01

    The results of nuclear fusion researches in JAERI are summarized. In this report, following themes are collected: the concept of fusion reactor (including ITER), fusion reactor safety, plasma confinement, fusion reactor equipment, and so on. Includes glossary. (J.P.N.)

  2. Membrane dynamics

    DEFF Research Database (Denmark)

    Bendix, Pól Martin

    2015-01-01

    Current topics include membrane-protein interactions with regard to membrane deformation or curvature sensing by BAR domains. Also, we study the dynamics of membrane tubes of both cells and simple model membrane tubes. Finally, we study membrane phase behavior which has important implications...

  3. Fusion fuel and renewables

    International Nuclear Information System (INIS)

    Entler, Slavomir

    2015-01-01

    It is shown that fusion fuel meets all aspects applied when defining renewables. A table of definitions of renewables is presented. The sections of the paper are as follows: An industrial renewable source; Nuclear fusion; Current situation in research; Definitions of renewable sources; Energy concept of nuclear fusion; Fusion fuel; Natural energy flow; Environmental impacts; Fusion fuel assessment; Sustainable power; and Energy mix from renewables. (P.A.)

  4. Fusion systems engineering

    International Nuclear Information System (INIS)

    Anon.

    1978-01-01

    Research during this report period has covered the following areas: (1) fusion reactor systems studies, (2) development of blanket processing technology for fusion reactors, (3) safety studies of fusion concepts, (4) MACKLIB-IV, a new library of nuclear response functions, (5) energy storage and power supply requirements for commercial fusion reactors, (6) blanket/shield design evaluation for commercial fusion reactors, and (7) cross section measurements, evaluations, and techniques

  5. Fusion energy

    International Nuclear Information System (INIS)

    1990-09-01

    The main purpose of the International Thermonuclear Experimental Reactor (ITER) is to develop an experimental fusion reactor through the united efforts of many technologically advanced countries. The ITER terms of reference, issued jointly by the European Community, Japan, the USSR, and the United States, call for an integrated international design activity and constitute the basis of current activities. Joint work on ITER is carried out under the auspices of the International Atomic Energy Agency (IAEA), according to the terms of quadripartite agreement reached between the European Community, Japan, the USSR, and the United States. The site for joint technical work sessions is at the MaxPlanck Institute of Plasma Physics. Garching, Federal Republic of Germany. The ITER activities have two phases: a definition phase performed in 1988 and the present design phase (1989--1990). During the definition phase, a set of ITER technical characteristics and supporting research and development (R ampersand D) activities were developed and reported. The present conceptual design phase of ITER lasts until the end of 1990. The objectives of this phase are to develop the design of ITER, perform a safety and environmental analysis, develop site requirements, define future R ampersand D needs, and estimate cost, manpower, and schedule for construction and operation. A final report will be submitted at the end of 1990. This paper summarizes progress in the ITER program during the 1989 design phase

  6. Localization of a region in the fusion protein of avian metapneumovirus that modulates cell-cell fusion.

    Science.gov (United States)

    Wei, Yongwei; Feng, Kurtis; Yao, Xiangjie; Cai, Hui; Li, Junan; Mirza, Anne M; Iorio, Ronald M; Li, Jianrong

    2012-11-01

    The genus Metapneumovirus within the subfamily Pneumovirinae of the family Paramyxoviridae includes two members, human metapneumovirus (hMPV) and avian metapneumovirus (aMPV), causing respiratory tract infections in humans and birds, respectively. Paramyxoviruses enter host cells by fusing the viral envelope with a host cell membrane. Membrane fusion of hMPV appears to be unique, in that fusion of some hMPV strains requires low pH. Here, we show that the fusion (F) proteins of aMPV promote fusion in the absence of the attachment protein and low pH is not required. Furthermore, there are notable differences in cell-cell fusion among aMPV subtypes. Trypsin was required for cell-cell fusion induced by subtype B but not subtypes A and C. The F protein of aMPV subtype A was highly fusogenic, whereas those from subtypes B and C were not. By construction and evaluation of chimeric F proteins composed of domains from the F proteins of subtypes A and B, we localized a region composed of amino acid residues 170 to 338 in the F protein that is responsible for the hyperfusogenic phenotype of the F from subtype A. Further mutagenesis analysis revealed that residues R295, G297, and K323 in this region collectively contributed to the hyperfusogenicity. Taken together, we have identified a region in the aMPV F protein that modulates the extent of membrane fusion. A model for fusion consistent with these data is presented.

  7. Hemi-fused structure mediates and controls fusion and fission in live cells.

    Science.gov (United States)

    Zhao, Wei-Dong; Hamid, Edaeni; Shin, Wonchul; Wen, Peter J; Krystofiak, Evan S; Villarreal, Seth A; Chiang, Hsueh-Cheng; Kachar, Bechara; Wu, Ling-Gang

    2016-06-23

    Membrane fusion and fission are vital for eukaryotic life. For three decades, it has been proposed that fusion is mediated by fusion between the proximal leaflets of two bilayers (hemi-fusion) to produce a hemi-fused structure, followed by fusion between the distal leaflets, whereas fission is via hemi-fission, which also produces a hemi-fused structure, followed by full fission. This hypothesis remained unsupported owing to the lack of observation of hemi-fusion or hemi-fission in live cells. A competing fusion hypothesis involving protein-lined pore formation has also been proposed. Here we report the observation of a hemi-fused Ω-shaped structure in live neuroendocrine chromaffin cells and pancreatic β-cells, visualized using confocal and super-resolution stimulated emission depletion microscopy. This structure is generated from fusion pore opening or closure (fission) at the plasma membrane. Unexpectedly, the transition to full fusion or fission is determined by competition between fusion and calcium/dynamin-dependent fission mechanisms, and is notably slow (seconds to tens of seconds) in a substantial fraction of the events. These results provide key missing evidence in support of the hemi-fusion and hemi-fission hypothesis in live cells, and reveal the hemi-fused intermediate as a key structure controlling fusion and fission, as fusion and fission mechanisms compete to determine the transition to fusion or fission.

  8. Side chain packing below the fusion peptide strongly modulates triggering of the Hendra virus F protein.

    Science.gov (United States)

    Smith, Everett Clinton; Dutch, Rebecca Ellis

    2010-10-01

    Triggering of the Hendra virus fusion (F) protein is required to initiate the conformational changes which drive membrane fusion, but the factors which control triggering remain poorly understood. Mutation of a histidine predicted to lie near the fusion peptide to alanine greatly reduced fusion despite wild-type cell surface expression levels, while asparagine substitution resulted in a moderate restoration in fusion levels. Slowed kinetics of six-helix bundle formation, as judged by sensitivity to heptad repeat B-derived peptides, was observed for all H372 mutants. These data suggest that side chain packing beneath the fusion peptide is an important regulator of Hendra virus F triggering.

  9. Solid-State Nuclear Magnetic Resonance Investigation of the Structural Topology and Lipid Interactions of a Viral Fusion Protein Chimera Containing the Fusion Peptide and Transmembrane Domain.

    Science.gov (United States)

    Yao, Hongwei; Lee, Myungwoon; Liao, Shu-Yu; Hong, Mei

    2016-12-13

    The fusion peptide (FP) and transmembrane domain (TMD) of viral fusion proteins play important roles during virus-cell membrane fusion, by inducing membrane curvature and transient dehydration. The structure of the water-soluble ectodomain of viral fusion proteins has been extensively studied crystallographically, but the structures of the FP and TMD bound to phospholipid membranes are not well understood. We recently investigated the conformations and lipid interactions of the separate FP and TMD peptides of parainfluenza virus 5 (PIV5) fusion protein F using solid-state nuclear magnetic resonance. These studies provide structural information about the two domains when they are spatially well separated in the fusion process. To investigate how these two domains are structured relative to each other in the postfusion state, when the ectodomain forms a six-helix bundle that is thought to force the FP and TMD together in the membrane, we have now expressed and purified a chimera of the FP and TMD, connected by a Gly-Lys linker, and measured the chemical shifts and interdomain contacts of the protein in several lipid membranes. The FP-TMD chimera exhibits α-helical chemical shifts in all the membranes examined and does not cause strong curvature of lamellar membranes or membranes with negative spontaneous curvature. These properties differ qualitatively from those of the separate peptides, indicating that the FP and TMD interact with each other in the lipid membrane. However, no 13 C- 13 C cross peaks are observed in two-dimensional correlation spectra, suggesting that the two helices are not tightly associated. These results suggest that the ectodomain six-helix bundle does not propagate into the membrane to the two hydrophobic termini. However, the loosely associated FP and TMD helices are found to generate significant negative Gaussian curvature to membranes that possess spontaneous positive curvature, consistent with the notion that the FP-TMD assembly may

  10. Membrane topology analysis of HIV-1 envelope glycoprotein gp41

    Directory of Open Access Journals (Sweden)

    Xiao Dan

    2010-11-01

    Full Text Available Abstract Background The gp41 subunit of the HIV-1 envelope glycoprotein (Env has been widely regarded as a type I transmembrane protein with a single membrane-spanning domain (MSD. An alternative topology model suggested multiple MSDs. The major discrepancy between the two models is that the cytoplasmic Kennedy sequence in the single MSD model is assigned as the extracellular loop accessible to neutralizing antibodies in the other model. We examined the membrane topology of the gp41 subunit in both prokaryotic and mammalian systems. We attached topological markers to the C-termini of serially truncated gp41. In the prokaryotic system, we utilized a green fluorescent protein (GFP that is only active in the cytoplasm. The tag protein (HaloTag and a membrane-impermeable ligand specific to HaloTag was used in the mammalian system. Results In the absence of membrane fusion, both the prokaryotic and mammalian systems (293FT cells supported the single MSD model. In the presence of membrane fusion in mammalian cells (293CD4 cells, the data obtained seem to support the multiple MSD model. However, the region predicted to be a potential MSD is the highly hydrophilic Kennedy sequence and is least likely to become a MSD based on several algorithms. Further analysis revealed the induction of membrane permeability during membrane fusion, allowing the membrane-impermeable ligand and antibodies to cross the membrane. Therefore, we cannot completely rule out the possible artifacts. Addition of membrane fusion inhibitors or alterations of the MSD sequence decreased the induction of membrane permeability. Conclusions It is likely that a single MSD model for HIV-1 gp41 holds true even in the presence of membrane fusion. The degree of the augmentation of membrane permeability we observed was dependent on the membrane fusion and sequence of the MSD.

  11. Induction of cell-cell fusion by ectromelia virus is not inhibited by its fusion inhibitory complex

    Directory of Open Access Journals (Sweden)

    Fuchs Pinhas

    2009-09-01

    Full Text Available Abstract Background Ectromelia virus, a member of the Orthopox genus, is the causative agent of the highly infectious mousepox disease. Previous studies have shown that different poxviruses induce cell-cell fusion which is manifested by the formation of multinucleated-giant cells (polykaryocytes. This phenomenon has been widely studied with vaccinia virus in conditions which require artificial acidification of the medium. Results We show that Ectromelia virus induces cell-cell fusion under neutral pH conditions and requires the presence of a sufficient amount of viral particles on the plasma membrane of infected cells. This could be achieved by infection with a replicating virus and its propagation in infected cells (fusion "from within" or by infection with a high amount of virus particles per cell (fusion "from without". Inhibition of virus maturation or inhibition of virus transport on microtubules towards the plasma membrane resulted in a complete inhibition of syncytia formation. We show that in contrast to vaccinia virus, Ectromelia virus induces cell-cell fusion irrespectively of its hemagglutination properties and cell-surface expression of the orthologs of the fusion inhibitory complex, A56 and K2. Additionally, cell-cell fusion was also detected in mice lungs following lethal respiratory infection. Conclusion Ectromelia virus induces spontaneous cell-cell fusion in-vitro and in-vivo although expressing an A56/K2 fusion inhibitory complex. This syncytia formation property cannot be attributed to the 37 amino acid deletion in ECTV A56.

  12. Mitochondrial Fusion Proteins and Human Diseases

    Directory of Open Access Journals (Sweden)

    Michela Ranieri

    2013-01-01

    Full Text Available Mitochondria are highly dynamic, complex organelles that continuously alter their shape, ranging between two opposite processes, fission and fusion, in response to several stimuli and the metabolic demands of the cell. Alterations in mitochondrial dynamics due to mutations in proteins involved in the fusion-fission machinery represent an important pathogenic mechanism of human diseases. The most relevant proteins involved in the mitochondrial fusion process are three GTPase dynamin-like proteins: mitofusin 1 (MFN1 and 2 (MFN2, located in the outer mitochondrial membrane, and optic atrophy protein 1 (OPA1, in the inner membrane. An expanding number of degenerative disorders are associated with mutations in the genes encoding MFN2 and OPA1, including Charcot-Marie-Tooth disease type 2A and autosomal dominant optic atrophy. While these disorders can still be considered rare, defective mitochondrial dynamics seem to play a significant role in the molecular and cellular pathogenesis of more common neurodegenerative diseases, for example, Alzheimer’s and Parkinson’s diseases. This review provides an overview of the basic molecular mechanisms involved in mitochondrial fusion and focuses on the alteration in mitochondrial DNA amount resulting from impairment of mitochondrial dynamics. We also review the literature describing the main disorders associated with the disruption of mitochondrial fusion.

  13. Formation of Cell Membrane Component Domains in Artificial Lipid Bilayer.

    Science.gov (United States)

    Tero, Ryugo; Fukumoto, Kohei; Motegi, Toshinori; Yoshida, Miyu; Niwano, Michio; Hirano-Iwata, Ayumi

    2017-12-20

    The lipid bilayer environment around membrane proteins strongly affects their structure and functions. Here, we aimed to study the fusion of proteoliposomes (PLs) derived from cultured cells with an artificial lipid bilayer membrane and the distribution of the PL components after the fusion. PLs, which were extracted as a crude membrane fraction from Chinese hamster ovary (CHO) cells, formed isolated domains in a supported lipid bilayer (SLB), comprising phosphatidylcholine (PC), phosphatidylethanolamine (PE), and cholesterol (Chol), after the fusion. Observation with a fluorescence microscope and an atomic force microscope showed that the membrane fusion occurred selectively at microdomains in the PC + PE + Chol-SLB, and that almost all the components of the PL were retained in the domain. PLs derived from human embryonic kidney 293 (HEK) cells also formed isolated domains in the PC + PE + Chol-SLB, but their fusion kinetics was different from that of the CHO-PLs. We attempted to explain the mechanism of the PL-SLB fusion and the difference between CHO- and HEK-PLs, based on a kinetic model. The domains that contained the whole cell membrane components provided environments similar to that of natural cell membranes, and were thus effective for studying membrane proteins using artificial lipid bilayer membranes.

  14. Structural characterization of Mumps virus fusion protein core

    International Nuclear Information System (INIS)

    Liu Yueyong; Xu Yanhui; Lou Zhiyong; Zhu Jieqing; Hu Xuebo; Gao, George F.; Qiu Bingsheng; Rao Zihe; Tien, Po

    2006-01-01

    The fusion proteins of enveloped viruses mediating the fusion between the viral and cellular membranes comprise two discontinuous heptad repeat (HR) domains located at the ectodomain of the enveloped glycoproteins. The crystal structure of the fusion protein core of Mumps virus (MuV) was determined at 2.2 A resolution. The complex is a six-helix bundle in which three HR1 peptides form a central highly hydrophobic coiled-coil and three HR2 peptides pack against the hydrophobic grooves on the surface of central coiled-coil in an oblique antiparallel manner. Fusion core of MuV, like those of simian virus 5 and human respiratory syncytium virus, forms typical 3-4-4-4-3 spacing. The similar charecterization in HR1 regions, as well as the existence of O-X-O motif in extended regions of HR2 helix, suggests a basic rule for the formation of the fusion core of viral fusion proteins

  15. Review of fusion synfuels

    International Nuclear Information System (INIS)

    Fillo, J.A.

    1980-01-01

    Thermonuclear fusion offers an inexhaustible source of energy for the production of hydrogen from water. Depending on design, electric generation efficiencies of approx. 40 to 60% and hydrogen production efficiencies by high-temperature electrolysis of approx. 50 to 65% are projected for fusion reactors using high-temperatures blankets. Fusion/coal symbiotic systems appear economically promising for the first generation of commercial fusion synfuels plants. Coal production requirements and the environmental effects of large-scale coal usage would be greatly reduced by a fusion/coal system. In the long term, there could be a gradual transition to an inexhaustible energy system based solely on fusion

  16. Fusion of Semliki Forest virus with cholesterol-containing liposomes at low pH

    DEFF Research Database (Denmark)

    Wilschut, J; Corver, J; Nieva, J L

    1995-01-01

    on the presence of cholesterol in the target membrane. In this paper, we show that fusion of SFV with cholesterol-containing liposomes depends on sphingomyelin (SM) or other sphingolipids in the target membrane, ceramide representing the sphingolipid minimally required for mediating the process. The action...... of the sphingolipid is confined to the actual fusion event, cholesterol being necessary and sufficient for low-pH-dependent binding of the virus to target membranes. The 3-hydroxyl group on the sphingosine backbone plays a key role in the SFV fusion reaction, since 3-deoxy-sphingomyelin does not support the process....... This, and the remarkably low levels of sphingolipid required for half-maximal fusion (1-2 mol%), suggest that the sphingolipid does not play a structural role in SFV fusion, but rather acts as a cofactor, possibly through activation of the viral fusion protein. Domain formation between cholesterol...

  17. `Full fusion' is not ineluctable during vesicular exocytosis of neurotransmitters by endocrine cells

    Science.gov (United States)

    Oleinick, Alexander; Svir, Irina; Amatore, Christian

    2017-01-01

    Vesicular exocytosis is an essential and ubiquitous process in neurons and endocrine cells by which neurotransmitters are released in synaptic clefts or extracellular fluids. It involves the fusion of a vesicle loaded with chemical messengers with the cell membrane through a nanometric fusion pore. In endocrine cells, unless it closes after some flickering (`Kiss-and-Run' events), this initial pore is supposed to expand exponentially, leading to a full integration of the vesicle membrane into the cell membrane-a stage called `full fusion'. We report here a compact analytical formulation that allows precise measurements of the fusion pore expansion extent and rate to be extracted from individual amperometric spike time courses. These data definitively establish that, during release of catecholamines, fusion pores enlarge at most to approximately one-fifth of the radius of their parent vesicle, hence ruling out the ineluctability of `full fusion'.

  18. Remotely controlled fusion of selected vesicles and living cells: a key issue review

    Science.gov (United States)

    Bahadori, Azra; Moreno-Pescador, Guillermo; Oddershede, Lene B.; Bendix, Poul M.

    2018-03-01

    Remote control over fusion of single cells and vesicles has a great potential in biological and chemical research allowing both transfer of genetic material between cells and transfer of molecular content between vesicles. Membrane fusion is a critical process in biology that facilitates molecular transport and mixing of cellular cytoplasms with potential formation of hybrid cells. Cells precisely regulate internal membrane fusions with the aid of specialized fusion complexes that physically provide the energy necessary for mediating fusion. Physical factors like membrane curvature, tension and temperature, affect biological membrane fusion by lowering the associated energy barrier. This has inspired the development of physical approaches to harness the fusion process at a single cell level by using remotely controlled electromagnetic fields to trigger membrane fusion. Here, we critically review various approaches, based on lasers or electric pulses, to control fusion between individual cells or between individual lipid vesicles and discuss their potential and limitations for present and future applications within biochemistry, biology and soft matter.

  19. Biobased Membrane

    NARCIS (Netherlands)

    Koenders, E.A.B.; Zlopasa, J.; Picken, S.J.

    2015-01-01

    The present invention is in the field of a composition for forming a bio-compatible membrane applicable to building material, such as concrete, cement, etc., to a meth od of applying said composition for forming a bio-compatible membrane, a biocompatible membrane, use of said membrane for various

  20. Fusion Canada issue 20

    International Nuclear Information System (INIS)

    1993-03-01

    Fusion Canada's publication of the National Fusion Program. Included in this issue is the CFFTP Industrial Impact Study, CCFM/TdeV Update:helium pumping, research funds, and deuterium in beryllium - high temperature behaviour. 3 figs

  1. Laser fusion: an overview

    International Nuclear Information System (INIS)

    Boyer, K.

    1975-01-01

    The laser fusion concept is described along with developments in neodymium and carbon dioxide lasers. Fuel design and fabrication are reviewed. Some spin-offs of the laser fusion program are discussed. (U.S.)

  2. Fusion reactor design studies

    International Nuclear Information System (INIS)

    Emmert, G.A.; Kulcinski, G.L.; Santarius, J.F.

    1990-01-01

    This report discusses the following topics on the ARIES tokamak: systems; plasma power balance; impurity control and fusion ash removal; fusion product ripple loss; energy conversion; reactor fueling; first wall design; shield design; reactor safety; and fuel cost and resources

  3. Canada's Fusion Program

    International Nuclear Information System (INIS)

    Jackson, D. P.

    1990-01-01

    Canada's fusion strategy is based on developing specialized technologies in well-defined areas and supplying these technologies to international fusion projects. Two areas are specially emphasized in Canada: engineered fusion system technologies, and specific magnetic confinement and materials studies. The Canadian Fusion Fuels Technology Project focuses on the first of these areas. It tritium and fusion reactor fuel systems, remote maintenance and related safety studies. In the second area, the Centre Canadian de fusion magnetique operates the Tokamak de Varennes, the main magnetic fusion device in Canada. Both projects are partnerships linking the Government of Canada, represented by Atomic Energy of Canada Limited, and provincial governments, electrical utilities, universities and industry. Canada's program has extensive international links, through which it collaborates with the major world fusion programs, including participation in the International Thermonuclear Experimental Reactor project

  4. Fusion Canada issue 27

    International Nuclear Information System (INIS)

    1995-03-01

    A short bulletin from the National Fusion Program highlighting in this issue ITER reactor siting, a major upgrade for TdeV tokamak, Ceramic Breeders: new tritium mapping technique and Joint Fusion Symposium. 2 figs

  5. Gallium nitride electrodes for membrane-based electrochemical biosensors.

    Science.gov (United States)

    Schubert, T; Steinhoff, G; von Ribbeck, H-G; Stutzmannn, M; Eickhoff, M; Tanaka, M

    2009-10-01

    We report on the deposition of planar lipid bilayers (supported membranes) on gallium nitride (GaN) electrodes for potential applications as membrane-based biosensors. The kinetics of the lipid membrane formation upon vesicle fusion were monitored by simultaneous measurements of resistance and capacitance of the membrane using AC impedance spectroscopy in the frequency range between 50 mHz and 50 kHz. We could identify a two-step process of membrane spreading and self-healing. Despite its relatively low resistance, the membrane can be modeled by a parallel combination of an ideal resistor and capacitor, indicating that the membrane efficiently blocks the diffusion of ions.

  6. Membrane tethering complexes in the endosomal system

    Directory of Open Access Journals (Sweden)

    Anne eSpang

    2016-05-01

    Full Text Available Vesicles that are generated by endocytic events at the plasma membrane are destined to early endosomes. A prerequisite for proper fusion is the tethering of two membrane entities. Tethering of vesicles to early endosomes is mediated by the CORVET complex, while fusion of late endosomes with lysosomes depends on the HOPS complex. Recycling through the TGN and to the plasma membrane is facilitated by the GARP and EARP complexes, respectively. However, there are other tethering functions in the endosomal system as there are multiple pathways through which proteins can be delivered from endosomes to either the TGN or the plasma membrane. Furthermore, complexes that may be part of novel tethering complexes have been recently identified. Thus it is likely that more tethering factors exist. In this review, I will provide an overview of different tethering complexes of the endosomal system and discuss how they may provide specificity in membrane traffic.

  7. A fusion-inhibiting peptide against Rift Valley fever virus inhibits multiple, diverse viruses.

    Directory of Open Access Journals (Sweden)

    Jeffrey W Koehler

    Full Text Available For enveloped viruses, fusion of the viral envelope with a cellular membrane is critical for a productive infection to occur. This fusion process is mediated by at least three classes of fusion proteins (Class I, II, and III based on the protein sequence and structure. For Rift Valley fever virus (RVFV, the glycoprotein Gc (Class II fusion protein mediates this fusion event following entry into the endocytic pathway, allowing the viral genome access to the cell cytoplasm. Here, we show that peptides analogous to the RVFV Gc stem region inhibited RVFV infectivity in cell culture by inhibiting the fusion process. Further, we show that infectivity can be inhibited for diverse, unrelated RNA viruses that have Class I (Ebola virus, Class II (Andes virus, or Class III (vesicular stomatitis virus fusion proteins using this single peptide. Our findings are consistent with an inhibition mechanism similar to that proposed for stem peptide fusion inhibitors of dengue virus in which the RVFV inhibitory peptide first binds to both the virion and cell membranes, allowing it to traffic with the virus into the endocytic pathway. Upon acidification and rearrangement of Gc, the peptide is then able to specifically bind to Gc and prevent fusion of the viral and endocytic membranes, thus inhibiting viral infection. These results could provide novel insights into conserved features among the three classes of viral fusion proteins and offer direction for the future development of broadly active fusion inhibitors.

  8. Cold fusion research

    International Nuclear Information System (INIS)

    1989-11-01

    I am pleased to forward to you the Final Report of the Cold Fusion Panel. This report reviews the current status of cold fusion and includes major chapters on Calorimetry and Excess Heat, Fusion Products and Materials Characterization. In addition, the report makes a number of conclusions and recommendations, as requested by the Secretary of Energy

  9. User's perspective on fusion

    International Nuclear Information System (INIS)

    Ashworth, C.P.

    1976-01-01

    The need in fusion, from the electric utilities viewpoint, is for fusion to be a real option, not huge, complicated nuclear plants costing $10 billion each and requiring restructuring the energy industry to provide and use them. A course for future fusion reactor work in order to be a real option is discussed. The advantages of alternate concepts to the tokamak are presented

  10. Fusion Canada issue 9

    International Nuclear Information System (INIS)

    1989-11-01

    A short bulletin from the National Fusion Program. Included in this issue is a report on availability of Canadian Tritium, an ITER update, a CCFM update on Tokamak and the new team organization, an international report on Fusion in Canada and a Laser Fusion Project at the University of Toronto. 3 figs

  11. Fusion Canada issue 17

    International Nuclear Information System (INIS)

    1992-05-01

    A short bulletin from the National Fusion Program. Included in this issue is a report on increased funding for the Canadian Fusion Program, news of the compact Toroid fuelling gun, an update on Tokamak de Varennes, the Canada - U.S. fusion meeting, measurements of plasma flow velocity, and replaceable Tokamak divertors. 4 figs

  12. The UKAEA's fusion programme

    International Nuclear Information System (INIS)

    Sweetman, D.R.

    1989-01-01

    D.R. Sweetman, director of the UKAEA fusion programme, reviews the current state of the work being performed on the UK-Euratom fusion programme. The JET programme, Tokamak programme, reversed field pinch programme, fusion technology and funding are all discussed. (author)

  13. The actin cytoskeleton inhibits pore expansion during PIV5 fusion protein-promoted cell-cell fusion

    International Nuclear Information System (INIS)

    Wurth, Mark A.; Schowalter, Rachel M.; Smith, Everett Clinton; Moncman, Carole L.; Ellis Dutch, Rebecca; McCann, Richard O.

    2010-01-01

    Paramyxovirus fusion (F) proteins promote both virus-cell fusion, required for viral entry, and cell-cell fusion, resulting in syncytia formation. We used the F-actin stabilizing drug, jasplakinolide, and the G-actin sequestrant, latrunculin A, to examine the role of actin dynamics in cell-cell fusion mediated by the parainfluenza virus 5 (PIV5) F protein. Jasplakinolide treatment caused a dose-dependent increase in cell-cell fusion as measured by both syncytia and reporter gene assays, and latrunculin A treatment also resulted in fusion stimulation. Treatment with jasplakinolide or latrunculin A partially rescued a fusion pore opening defect caused by deletion of the PIV5 F protein cytoplasmic tail, but these drugs had no effect on fusion inhibited at earlier stages by either temperature arrest or by a PIV5 heptad repeat peptide. These data suggest that the cortical actin cytoskeleton is an important regulator of fusion pore enlargement, an energetically costly stage of viral fusion protein-mediated membrane merger.

  14. Fusion technology 1992

    International Nuclear Information System (INIS)

    Ferro, C.; Gasparatto, M.; Knoepfel, H.

    1993-01-01

    The aim of the biennial series of symposia on the title subject, organized by the European Fusion Laboratories, is the exchange of information on the design, construction and operation of fusion experiments and on the technology being developed for the next step devices and fusion reactors. The coverage of the volume includes the technological aspects of fusion reactors in relation to new developments, this forming a guideline for the definition of future work. These proceedings comprise three volumes and contain both the invited lectures and contributed papers presented at the symposium which was attended by 569 participants from around the globe. The 343 papers, including 12 invited papers, characterize the increasing interest of industry in the fusion programme, giving a broad and current overview on the progress and trends fusion technology is experiencing now, as well as indicating the future for fusion devices

  15. Two single mutations in the fusion protein of Newcastle disease virus confer hemagglutinin-neuraminidase independent fusion promotion and attenuate the pathogenicity in chickens

    Science.gov (United States)

    The fusion (F) protein of Newcastle disease virus (NDV) plays an important role in viral infection and pathogenicity through mediating membrane fusion between the virion and host cells in the presence of the hemagglutinin-neuraminidase (HN). Previously, we obtained a velogenic NDV genotype VII muta...

  16. Membranous nephropathy

    Science.gov (United States)

    ... check for hepatitis B, hepatitis C, and syphilis Complement levels Cryoglobulin test Treatment The goal of treatment ... not as helpful for people with membranous nephropathy. Medicines used treat membranous nephropathy include: Angiotensin-converting enzyme ( ...

  17. Peptide-Mediated Liposome Fusion: The Effect of Anchor Positioning

    Directory of Open Access Journals (Sweden)

    Niek S. A. Crone

    2018-01-01

    Full Text Available A minimal model system for membrane fusion, comprising two complementary peptides dubbed “E” and “K” joined to a cholesterol anchor via a polyethyleneglycol spacer, has previously been developed in our group. This system promotes the fusion of large unilamellar vesicles and facilitates liposome-cell fusion both in vitro and in vivo. Whilst several aspects of the system have previously been investigated to provide an insight as to how fusion is facilitated, anchor positioning has not yet been considered. In this study, the effects of placing the anchor at either the N-terminus or in the center of the peptide are investigated using a combination of circular dichroism spectroscopy, dynamic light scattering, and fluorescence assays. It was discovered that anchoring the “K” peptide in the center of the sequence had no effect on its structure, its ability to interact with membranes, or its ability to promote fusion, whereas anchoring the ‘E’ peptide in the middle of the sequence dramatically decreases fusion efficiency. We postulate that anchoring the ‘E’ peptide in the middle of the sequence disrupts its ability to form homodimers with peptides on the same membrane, leading to aggregation and content leakage.

  18. Economics of fusion research

    International Nuclear Information System (INIS)

    1977-01-01

    This report provides the results of a study of methods of economic analysis applied to the evaluation of fusion research. The study recognizes that a hierarchy of economic analyses of research programs exists: standard benefit-cost analysis, expected value of R and D information, and expected utility analysis. It is shown that standard benefit-cost analysis, as commonly applied to research programs, is inadequate for the evaluation of a high technology research effort such as fusion research. A methodology for performing an expected value analysis is developed and demonstrated and an overview of an approach to perform an expected utility analysis of fusion research is presented. In addition, a potential benefit of fusion research, not previously identified, is discussed and rough estimates of its magnitude are presented. This benefit deals with the effect of a fusion research program on optimal fossil fuel consumption patterns. The results of this study indicate that it is both appropriate and possible to perform an expected value analysis of fusion research in order to assess the economics of a fusion research program. The results indicate further that the major area of benefits of fusion research is likely due to the impact of a fusion research program on optimal fossil fuel consumption patterns and it is recommended that this benefit be included in future assessments of fusion research economics

  19. Economics of fusion research

    Energy Technology Data Exchange (ETDEWEB)

    None, None

    1977-10-15

    This report provides the results of a study of methods of economic analysis applied to the evaluation of fusion research. The study recognizes that a hierarchy of economic analyses of research programs exists: standard benefit-cost analysis, expected value of R and D information, and expected utility analysis. It is shown that standard benefit-cost analysis, as commonly applied to research programs, is inadequate for the evaluation of a high technology research effort such as fusion research. A methodology for performing an expected value analysis is developed and demonstrated and an overview of an approach to perform an expected utility analysis of fusion research is presented. In addition, a potential benefit of fusion research, not previously identified, is discussed and rough estimates of its magnitude are presented. This benefit deals with the effect of a fusion research program on optimal fossil fuel consumption patterns. The results of this study indicate that it is both appropriate and possible to perform an expected value analysis of fusion research in order to assess the economics of a fusion research program. The results indicate further that the major area of benefits of fusion research is likely due to the impact of a fusion research program on optimal fossil fuel consumption patterns and it is recommended that this benefit be included in future assessments of fusion research economics.

  20. Annexin A4 and A6 induce membrane curvature and constriction during cell membrane repair

    DEFF Research Database (Denmark)

    Boye, Theresa Louise; Maeda, Kenji; Pezeshkian, Weria

    2017-01-01

    that annexin A4 binds to artificial membranes and generates curvature force initiated from free edges, whereas annexin A6 induces constriction force. In cells, plasma membrane injury and Ca2+ influx recruit annexin A4 to the vicinity of membrane wound edges where its homo-trimerization leads to membrane......Efficient cell membrane repair mechanisms are essential for maintaining membrane integrity and thus for cell life. Here we show that the Ca2+- and phospholipid-binding proteins annexin A4 and A6 are involved in plasma membrane repair and needed for rapid closure of micron-size holes. We demonstrate...... curvature near the edges. We propose that curvature force is utilized together with annexin A6-mediated constriction force to pull the wound edges together for eventual fusion. We show that annexin A4 can counteract various plasma membrane disruptions including holes of several micrometers indicating...

  1. Membrane Biophysics

    CERN Document Server

    Ashrafuzzaman, Mohammad

    2013-01-01

    Physics, mathematics and chemistry all play a vital role in understanding the true nature and functioning of biological membranes, key elements of living processes. Besides simple spectroscopic observations and electrical measurements of membranes we address in this book the phenomena of coexistence and independent existence of different membrane components using various theoretical approaches. This treatment will be helpful for readers who want to understand biological processes by applying both simple observations and fundamental scientific analysis. It provides a deep understanding of the causes and effects of processes inside membranes, and will thus eventually open new doors for high-level pharmaceutical approaches towards fighting membrane- and cell-related diseases.

  2. Fusion of Nonionic Vesicles

    DEFF Research Database (Denmark)

    Bulut, Sanja; Oskolkova, M. Z.; Schweins, R.

    2010-01-01

    We present an experimental study of vesicle fusion using light and neutron scattering to monitor fusion events. Vesicles are reproducibly formed with an extrusion procedure using an single amphiphile triethylene glycol mono-n-decyl ether in water. They show long-term stability for temperatures...... around 20 C, but at temperatures above 26 C we observe an increase in the scattered intensity due to fusion. The system is unusually well suited for the study of basic mechanisms of vesicle fusion. The vesicles are flexible with a bending rigidity of only a few k(H)T. The monolayer spontaneous curvature......, Ho, depends strongly on temperature in a known way and is thus tunable. For temperatures where H-0 > 0 vesicles tyre long-term stable, while in the range H-0 fusion rate increases the more negative the Spontaneous curvature Through a quantitative;analysis of the fusion rate we arrive tit...

  3. Document controlled fusion

    International Nuclear Information System (INIS)

    Abou, C.; Demarthon, F.; Ter Minassian, V.

    2004-01-01

    Since the years 30, the magnetic confinement is one of the researches programs to control the nuclear fusion. This document presents in a first part the historical aspects of the researches on the controlled fusion and in the second part the nuclear fusion. The nuclear fusion forcing two atomic nuclei to fuse together by reproducing the conditions of the thermonuclear reactions that make the stars burn. This technology is a potential source of inexhaustible energy for the future. Then are presented the tokamak which make possible to confine an extremely hot gaseous mixture (plasma that is over one hundred million degrees) in a vacuum chamber and the ITER project (superconductor tokamak) that will make it possible to attain the stage in which the plasma maintains the fusion reaction itself and therefore produces more energy than it consumes. The last part presents the projects of new fusion reactors. (A.L.B.)

  4. Muon Catalyzed Fusion

    Science.gov (United States)

    Armour, Edward A.G.

    2007-01-01

    Muon catalyzed fusion is a process in which a negatively charged muon combines with two nuclei of isotopes of hydrogen, e.g, a proton and a deuteron or a deuteron and a triton, to form a muonic molecular ion in which the binding is so tight that nuclear fusion occurs. The muon is normally released after fusion has taken place and so can catalyze further fusions. As the muon has a mean lifetime of 2.2 microseconds, this is the maximum period over which a muon can participate in this process. This article gives an outline of the history of muon catalyzed fusion from 1947, when it was first realised that such a process might occur, to the present day. It includes a description of the contribution that Drachrnan has made to the theory of muon catalyzed fusion and the influence this has had on the author's research.

  5. High- and low-affinity sites for sodium in delta-OR-G(i)1 alpha (Cys(351)-Ile(351)) fusion protein stably expressed in HEK293 cells; functional significance and correlation with biophysical state of plasma membrane

    Czech Academy of Sciences Publication Activity Database

    Vošahlíková, Miroslava; Jurkiewicz, Piotr; Roubalová, Lenka; Hof, Martin; Svoboda, Petr

    2014-01-01

    Roč. 387, č. 5 (2014), s. 487-502 ISSN 0028-1298 R&D Projects: GA ČR(CZ) GAP207/12/0919; GA ČR(CZ) GBP304/12/G069 Institutional support: RVO:67985823 ; RVO:61388955 Keywords : delta - opioid receptor * monovalent ions * agonist and antagonist binding * [35S]GTPγS binding * membrane biophysics * Laurdan fluorescence Subject RIV: CE - Biochemistry Impact factor: 2.471, year: 2014

  6. αvβ6- and αvβ8-integrins serve as interchangeable receptors for HSV gH/gL to promote endocytosis and activation of membrane fusion.

    Directory of Open Access Journals (Sweden)

    Tatiana Gianni

    Full Text Available Herpes simplex virus (HSV--and herpesviruses in general--encode for a multipartite entry/fusion apparatus. In HSV it consists of the HSV-specific glycoprotein D (gD, and three additional glycoproteins, gH/gL and gB, conserved across the Herpesviridae family and responsible for the execution of fusion. According to the current model, upon receptor binding, gD propagates the activation to gH/gL and to gB in a cascade fashion. Questions remain about how the cascade of activation is controlled and how it is synchronized with virion endocytosis, to avoid premature activation and exhaustion of the glycoproteins. We considered the possibility that such control might be carried out by as yet unknown receptors. Indeed, receptors for HSV gB, but not for gH/gL, have been described. In other members of the Herpesviridae family, such as Epstein-Barr virus, integrin receptors bind gH/gL and trigger conformational changes in the glycoproteins. We report that αvβ6- and αvβ8-integrins serve as receptors for HSV entry into experimental models of keratinocytes and other epithelial and neuronal cells. Evidence rests on loss of function experiments, in which integrins were blocked by antibodies or silenced, and gain of function experiments in which αvβ6-integrin was expressed in integrin-negative cells. αvβ6- and αvβ8-integrins acted independently and are thus interchangeable. Both bind gH/gL with high affinity. The interaction profoundly affects the route of HSV entry and directs the virus to acidic endosomes. In the case of αvβ8, but not αvβ6-integrin, the portal of entry is located at lipid microdomains and requires dynamin 2. Thus, a major role of αvβ6- or αvβ8-integrin in HSV infection appears to be to function as gH/gL receptors and to promote virus endocytosis. We propose that placing the gH/gL activation under the integrin trigger point enables HSV to synchronize virion endocytosis with the cascade of glycoprotein activation that culminates

  7. Status of fusion maintenance

    International Nuclear Information System (INIS)

    Fuller, G.M.

    1984-01-01

    Effective maintenance will be an essential ingredient in determining fusion system productivity. This level of productivity will result only after close attention is paid to the entire system as an entity and appropriate integration of the elements is made. The status of fusion maintenance is reviewed in the context of the entire system. While there are many challenging developmental tasks ahead in fusion maintenance, the required technologies are available in several high-technology industries, including nuclear fission

  8. Perfect focusing fusion system

    International Nuclear Information System (INIS)

    Miyamoto, G.; Takeda, T.; Iwata, G.; Mori, S.; Inoue, K.; Tanaka, M.

    1994-01-01

    We propose new perfect focusing (perfo) fusion systems in which ion- or atom-beam is used as a target for the fusion reaction, and ions (perfo particles) of different species moving in the perfo field collide with them. The 'efficiency' defined as the ratio of the fusion energy output to the radiation loss is ∼300, ∼20, and ∼4 for the T+D, 3 He+D, and 11 B+H reactions, respectively. (author)

  9. Magnetic-fusion program

    International Nuclear Information System (INIS)

    1980-08-01

    In February 1980, the Director of Energy Research requested that the Energy Research Advisory Board (ERAB) review the Department of Energy (DOE) Magnetic Fusion Program. Of particular concern to the DOE was the judicious choice of the next major steps toward demonstration of economic power production from fusion. Of equal concern was the overall soundness of the DOE Magnetic Fusion Program: its pace, scope, and funding profiles. Their finding and recommendations are included

  10. Maximum Likelihood Fusion Model

    Science.gov (United States)

    2014-08-09

    Symposium of Robotics Re- search. Sienna, Italy: Springer, 2003. [12] D. Hall and J. Llinas, “An introduction to multisensor data fusion ,” Proceed- ings of...a data fusion approach for combining Gaussian metric models of an environment constructed by multiple agents that operate outside of a global... data fusion , hypothesis testing,maximum likelihood estimation, mobile robot navigation REPORT DOCUMENTATION PAGE 11. SPONSOR/MONITOR’S REPORT

  11. Fusion research principles

    CERN Document Server

    Dolan, Thomas James

    2013-01-01

    Fusion Research, Volume I: Principles provides a general description of the methods and problems of fusion research. The book contains three main parts: Principles, Experiments, and Technology. The Principles part describes the conditions necessary for a fusion reaction, as well as the fundamentals of plasma confinement, heating, and diagnostics. The Experiments part details about forty plasma confinement schemes and experiments. The last part explores various engineering problems associated with reactor design, vacuum and magnet systems, materials, plasma purity, fueling, blankets, neutronics

  12. Fusion facility siting considerations

    International Nuclear Information System (INIS)

    Bussell, G.T.

    1985-01-01

    Inherent in the fusion program's transition from hydrogen devices to commercial power machines is a general increase in the size and scope of succeeding projects. This growth will lead to increased emphasis on safety, environmental impact, and the external effects of fusion in general, and of each new device in particular. A critically important consideration in this regard is site selection. The purpose of this paper is to examine major siting issues that may affect the economics, safety, and environmental impact of fusion

  13. Magnetic fusion technology

    CERN Document Server

    Dolan, Thomas J

    2014-01-01

    Magnetic Fusion Technology describes the technologies that are required for successful development of nuclear fusion power plants using strong magnetic fields. These technologies include: ? magnet systems, ? plasma heating systems, ? control systems, ? energy conversion systems, ? advanced materials development, ? vacuum systems, ? cryogenic systems, ? plasma diagnostics, ? safety systems, and ? power plant design studies. Magnetic Fusion Technology will be useful to students and to specialists working in energy research.

  14. Cell membrane structures during exocytosis.

    Science.gov (United States)

    Savigny, Pascale; Evans, John; McGrath, Kathryn M

    2007-08-01

    Exocytosis is a key biological process that controls the neurotransmission and release of hormones from cells. In endocrine cells, hormones are packed into secretory vesicles and released into the extracellular environment via openings in the plasma membrane, a few hundred nanometers wide, which form as a result of fusion of the membranes of the granule and cell. The complex processes and dynamics that result in the formation of the fusion pore, as well as its structure, remain scantly understood. A number of different exocytosis mechanisms have been postulated. Furthermore, the possibility exists that several mechanisms occur simultaneously. We present here an investigation of the cell membrane dynamics during exocytosis in anterior pituitary cells, especially gonadotropes, which secrete LH, a hormone central to ovulation. Gonadotrope enrichment was achieved using immunolabeled magnetic nanobeads. Three complementary imaging techniques were used to realize a fine structure study of the dynamics of the exocytosis-like sites occurring during secretion. Living pituitary and gonadotrope-enriched cells were imaged with atomic force microscopy, as well as cells that had been fixed to obtain better resolution. Atomic force microscopy, along with scanning and transmission electron microscopy, studies of these cells revealed that there are at least two different site configurations: simple single fusion pores and a complex association of pores consisting of a simple primary site combined with secondary attachments.

  15. Fusion of Semliki Forest virus with cholesterol-containing liposomes at low pH

    DEFF Research Database (Denmark)

    Wilschut, J; Corver, J; Nieva, J L

    1995-01-01

    . This, and the remarkably low levels of sphingolipid required for half-maximal fusion (1-2 mol%), suggest that the sphingolipid does not play a structural role in SFV fusion, but rather acts as a cofactor, possibly through activation of the viral fusion protein. Domain formation between cholesterol......Semliki Forest virus (SFV) utilizes a membrane fusion strategy to introduce its genome into the host cell. After binding to cell-surface receptors, virus particles are internalized through receptor-mediated endocytosis and directed to the endosomal cell compartment. Subsequently, triggered...... by the acid pH in the lumen of the endosomes, the viral envelope fuses with the endosomal membrane. As a result of this fusion reaction the viral RNA gains access to the cell cytosol. Low-pH-induced fusion of SFV, in model systems as well as in cells, has been demonstrated previously to be strictly dependent...

  16. Inertial confinement fusion (ICF)

    International Nuclear Information System (INIS)

    Nuckolls, J.

    1977-01-01

    The principal goal of the inertial confinement fusion program is the development of a practical fusion power plant in this century. Rapid progress has been made in the four major areas of ICF--targets, drivers, fusion experiments, and reactors. High gain targets have been designed. Laser, electron beam, and heavy ion accelerator drivers appear to be feasible. Record-breaking thermonuclear conditions have been experimentally achieved. Detailed diagnostics of laser implosions have confirmed predictions of the LASNEX computer program. Experimental facilities are being planned and constructed capable of igniting high gain fusion microexplosions in the mid 1980's. A low cost long lifetime reactor design has been developed

  17. lysosome tethering and fusion

    Indian Academy of Sciences (India)

    AMIT TULI

    Molecular mechanisms regulating endosome- lysosome tethering and fusion. Mahak Sharma. Assistant Professor & Wellcome Trust-DBT Intermediate Fellow. Department of Biological Sciences. IISER-Mohali ...

  18. COTS Fusion Tracker Evaluation

    National Research Council Canada - National Science Library

    Gertz, J

    2002-01-01

    .... This effort included cataloging the companies that have available ATC fusion trackers, acquiring executable tracker images from as many as possible of these trackers, running the commercial tracker...

  19. Laser fusion program overview

    International Nuclear Information System (INIS)

    Emmett, J.L.

    1977-01-01

    This program is structured to proceed through a series of well defined fusion milestones to proof of the scientific feasibility, of laser fusion with the Shiva Nova system. Concurrently, those key technical areas, such as advanced lasers, which are required to progress beyond proof of feasibility, are being studied. We have identified and quantified the opportunities and key technical issues in military applications, such as weapons effects simulations, and in civilian applications, such as central-station electric power production. We summarize the current status and future plans for the laser fusion program at LLL, emphasizing the civilian applications of laser fusion

  20. Magnetic-confinement fusion

    Science.gov (United States)

    Ongena, J.; Koch, R.; Wolf, R.; Zohm, H.

    2016-05-01

    Our modern society requires environmentally friendly solutions for energy production. Energy can be released not only from the fission of heavy nuclei but also from the fusion of light nuclei. Nuclear fusion is an important option for a clean and safe solution for our long-term energy needs. The extremely high temperatures required for the fusion reaction are routinely realized in several magnetic-fusion machines. Since the early 1990s, up to 16 MW of fusion power has been released in pulses of a few seconds, corresponding to a power multiplication close to break-even. Our understanding of the very complex behaviour of a magnetized plasma at temperatures between 150 and 200 million °C surrounded by cold walls has also advanced substantially. This steady progress has resulted in the construction of ITER, a fusion device with a planned fusion power output of 500 MW in pulses of 400 s. ITER should provide answers to remaining important questions on the integration of physics and technology, through a full-size demonstration of a tenfold power multiplication, and on nuclear safety aspects. Here we review the basic physics underlying magnetic fusion: past achievements, present efforts and the prospects for future production of electrical energy. We also discuss questions related to the safety, waste management and decommissioning of a future fusion power plant.

  1. Frontiers in fusion research

    CERN Document Server

    Kikuchi, Mitsuru

    2011-01-01

    Frontiers in Fusion Research provides a systematic overview of the latest physical principles of fusion and plasma confinement. It is primarily devoted to the principle of magnetic plasma confinement, that has been systematized through 50 years of fusion research. Frontiers in Fusion Research begins with an introduction to the study of plasma, discussing the astronomical birth of hydrogen energy and the beginnings of human attempts to harness the Sun's energy for use on Earth. It moves on to chapters that cover a variety of topics such as: * charged particle motion, * plasma kinetic theory, *

  2. Magnetic fusion reactor economics

    International Nuclear Information System (INIS)

    Krakowski, R.A.

    1995-01-01

    An almost primordial trend in the conversion and use of energy is an increased complexity and cost of conversion systems designed to utilize cheaper and more-abundant fuels; this trend is exemplified by the progression fossil fission → fusion. The present projections of the latter indicate that capital costs of the fusion ''burner'' far exceed any commensurate savings associated with the cheapest and most-abundant of fuels. These projections suggest competitive fusion power only if internal costs associate with the use of fossil or fission fuels emerge to make them either uneconomic, unacceptable, or both with respect to expensive fusion systems. This ''implementation-by-default'' plan for fusion is re-examined by identifying in general terms fusion power-plant embodiments that might compete favorably under conditions where internal costs (both economic and environmental) of fossil and/or fission are not as great as is needed to justify the contemporary vision for fusion power. Competitive fusion power in this context will require a significant broadening of an overly focused program to explore the physics and simbiotic technologies leading to more compact, simplified, and efficient plasma-confinement configurations that reside at the heart of an attractive fusion power plant

  3. Identification of cancer fusion drivers using network fusion centrality

    OpenAIRE

    Wu, Chia-Chin; Kannan, Kalpana; Lin, Steven; Yen, Laising; Milosavljevic, Aleksandar

    2013-01-01

    Summary: Gene fusions are being discovered at an increasing rate using massively parallel sequencing technologies. Prioritization of cancer fusion drivers for validation cannot be performed using traditional single-gene based methods because fusions involve portions of two partner genes. To address this problem, we propose a novel network analysis method called fusion centrality that is specifically tailored for prioritizing gene fusions. We first propose a domain-based fusion model built on ...

  4. Phosphatidylserine directly and positively regulates fusion of myoblasts into myotubes

    International Nuclear Information System (INIS)

    Jeong, Jaemin; Conboy, Irina M.

    2011-01-01

    Highlights: → PS broadly and persistently trans-locates to the outer leaflet of plasma membrane during myoblast fusion into myotubes. → Robust myotubes are formed when PS liposomes are added exogenously. → PS increases the width of de novo myotubes and the numbers of myonuclei, but not the myotube length. → Annexin V or PS antibody inhibits myotube formation by masking exposed PS. -- Abstract: Cell membrane consists of various lipids such as phosphatidylserine (PS), phosphatidylcholine (PC), and phosphatidylethanolamine (PE). Among them, PS is a molecular marker of apoptosis, because it is located to the inner leaflet of plasma membrane generally but it is moved to the outer leaflet during programmed cell death. The process of apoptosis has been implicated in the fusion of muscle progenitor cells, myoblasts, into myotubes. However, it remained unclear whether PS regulates muscle cell differentiation directly. In this paper, localization of PS to the outer leaflet of plasma membrane in proliferating primary myoblasts and during fusion of these myoblasts into myotubes is validated using Annexin V. Moreover, we show the presence of PS clusters at the cell-cell contact points, suggesting the importance of membrane ruffling and PS exposure for the myogenic cell fusion. Confirming this conclusion, experimentally constructed PS, but not PC liposomes dramatically enhance the formation of myotubes from myoblasts, thus demonstrating a direct positive effect of PS on the muscle cell fusion. In contrast, myoblasts exposed to PC liposomes produce long myotubes with low numbers of myonuclei. Moreover, pharmacological masking of PS on the myoblast surface inhibits fusion of these cells into myotubes in a dose-dependent manner.

  5. Structure and Function Study of HIV and Influenza Fusion Proteins

    Science.gov (United States)

    Liang, Shuang

    Human immunodeficiency virus (HIV) and influenza virus are membrane-enveloped viruses causing acquired immunodeficiency syndrome (AIDS) and flu. The initial step of HIV and influenza virus infection is fusion between viral and host cell membrane catalyzed by the viral fusion protein gp41 and hemagglutinin (HA) respectively. However, the structure of gp41 and HA as well as the infection mechanism are still not fully understood. This work addresses (1) full length gp41 ectodomain and TM domain structure and function and (2) IFP membrane location and IFP-membrane interaction. My studies of gp41 protein and IFP can provide better understanding of the membrane fusion mechanism and may aid development of anti-viral therapeutics and vaccine. The full length ectodomain and transmembrane domain of gp41 and shorter constructs were expressed, purified and solubilized at physiology conditions. The constructs adopt overall alpha helical structure in SDS and DPC detergents, and showed hyperthermostability with Tm > 90 °C. The oligomeric states of these proteins vary in different detergent buffer: predominant trimer for all constructs and some hexamer fraction for HM and HM_TM protein in SDS at pH 7.4; and mixtures of monomer, trimer, and higher-order oligomer protein in DPC at pH 4.0 and 7.4. Substantial protein-induced vesicle fusion was observed, including fusion of neutral vesicles at neutral pH, which are the conditions similar HIV/cell fusion. Vesicle fusion by a gp41 ectodomain construct has rarely been observed under these conditions, and is aided by inclusion of both the FP and TM, and by protein which is predominantly trimer rather than monomer. Current data was integrated with existing data, and a structural model was proposed. Secondary structure and conformation of IFP is a helix-turn-helix structure in membrane. However, there has been arguments about the IFP membrane location. 13C-2H REDOR solid-state NMR is used to solve this problem. The IFP adopts major alpha

  6. Mitochondrial morphological features are associated with fission and fusion events.

    Directory of Open Access Journals (Sweden)

    Laura M Westrate

    Full Text Available Mitochondria are dynamic organelles that undergo constant remodeling through the regulation of two opposing processes, mitochondrial fission and fusion. Although several key regulators and physiological stimuli have been identified to control mitochondrial fission and fusion, the role of mitochondrial morphology in the two processes remains to be determined. To address this knowledge gap, we investigated whether morphological features extracted from time-lapse live-cell images of mitochondria could be used to predict mitochondrial fate. That is, we asked if we could predict whether a mitochondrion is likely to participate in a fission or fusion event based on its current shape and local environment. Using live-cell microscopy, image analysis software, and supervised machine learning, we characterized mitochondrial dynamics with single-organelle resolution to identify features of mitochondria that are predictive of fission and fusion events. A random forest (RF model was trained to correctly classify mitochondria poised for either fission or fusion based on a series of morphological and positional features for each organelle. Of the features we evaluated, mitochondrial perimeter positively correlated with mitochondria about to undergo a fission event. Similarly mitochondrial solidity (compact shape positively correlated with mitochondria about to undergo a fusion event. Our results indicate that fission and fusion are positively correlated with mitochondrial morphological features; and therefore, mitochondrial fission and fusion may be influenced by the mechanical properties of mitochondrial membranes.

  7. Localization of a Region in the Fusion Protein of Avian Metapneumovirus That Modulates Cell-Cell Fusion

    Science.gov (United States)

    Wei, Yongwei; Feng, Kurtis; Yao, Xiangjie; Cai, Hui; Li, Junan; Mirza, Anne M.; Iorio, Ronald M.

    2012-01-01

    The genus Metapneumovirus within the subfamily Pneumovirinae of the family Paramyxoviridae includes two members, human metapneumovirus (hMPV) and avian metapneumovirus (aMPV), causing respiratory tract infections in humans and birds, respectively. Paramyxoviruses enter host cells by fusing the viral envelope with a host cell membrane. Membrane fusion of hMPV appears to be unique, in that fusion of some hMPV strains requires low pH. Here, we show that the fusion (F) proteins of aMPV promote fusion in the absence of the attachment protein and low pH is not required. Furthermore, there are notable differences in cell-cell fusion among aMPV subtypes. Trypsin was required for cell-cell fusion induced by subtype B but not subtypes A and C. The F protein of aMPV subtype A was highly fusogenic, whereas those from subtypes B and C were not. By construction and evaluation of chimeric F proteins composed of domains from the F proteins of subtypes A and B, we localized a region composed of amino acid residues 170 to 338 in the F protein that is responsible for the hyperfusogenic phenotype of the F from subtype A. Further mutagenesis analysis revealed that residues R295, G297, and K323 in this region collectively contributed to the hyperfusogenicity. Taken together, we have identified a region in the aMPV F protein that modulates the extent of membrane fusion. A model for fusion consistent with these data is presented. PMID:22915815

  8. Controlled Nuclear Fusion.

    Science.gov (United States)

    Glasstone, Samuel

    This publication is one of a series of information booklets for the general public published by The United States Atomic Energy Commission. Among the topics discussed are: Importance of Fusion Energy; Conditions for Nuclear Fusion; Thermonuclear Reactions in Plasmas; Plasma Confinement by Magnetic Fields; Experiments With Plasmas; High-Temperature…

  9. Fusion Canada issue 4

    International Nuclear Information System (INIS)

    1988-05-01

    A short bulletin from the National Fusion Program. Included in this issue is a technical update on Tokamak de Varennes, a report on the Beatrix II Breeding Materials Test Program, the Tritium glovebox system for UPM, Saudi Arabia, a broad update of the Canadian Fusion Fuels Technology Project is also included. 1 fig

  10. The fusion reactor

    International Nuclear Information System (INIS)

    Brennan, M.H.

    1974-01-01

    Basic principles of the fusion reactor are outlined. Plasma heating and confinement schemes are described. These confinement systems include the linear Z pinch, magnetic mirrors and Tokamaks. A fusion reactor is described and a discussion is given of its environmental impact and its fuel situation. (R.L.)

  11. Magnetic Fusion Program Plan

    International Nuclear Information System (INIS)

    1985-02-01

    This Plan reflects the present conditions of the energy situation and is consistent with national priorities for the support of basic and applied research. It is realistic in taking advantage of the technical position that the United States has already established in fusion research to make cost-effective progress toward the development of fusion power as a future energy option

  12. Fusion reactor materials

    International Nuclear Information System (INIS)

    Sethi, V.K.; Scholz, R.; Nolfi, F.V. Jr.; Turner, A.P.L.

    1980-01-01

    Data are given for each of the following areas: (1) effects of irradiation on fusion reactor materials, (2) hydrogen permeation and materials behavior in alloys, (3) carbon coatings for fusion applications, (4) surface damage of TiB 2 coatings under energetic D + and 4 He + irradiations, and (5) neutron dosimetry

  13. Fusion Canada issue 12

    International Nuclear Information System (INIS)

    1990-10-01

    A short bulletin from the National Fusion Program. Included in this issue is a report on Darlington's Tritium Removal Facility, work at universities on Deuterium Diffusivity in Beryllium, Fusion Studies, confinement research and the operation of divertors at Tokamak de Varennes. 5 figs

  14. Coatings for laser fusion

    International Nuclear Information System (INIS)

    Lowdermilk, W.H.

    1981-01-01

    Optical coatings are used in lasers systems for fusion research to control beam propagation and reduce surface reflection losses. The performance of coatings is important in the design, reliability, energy output, and cost of the laser systems. Significant developments in coating technology are required for future lasers for fusion research and eventual power reactors

  15. Two Horizons of Fusion

    Science.gov (United States)

    Lo, Mun Ling; Chik, Pakey Pui Man

    2016-01-01

    In this paper, we aim to differentiate the internal and external horizons of "fusion." "Fusion" in the internal horizon relates to the structure and meaning of the object of learning as experienced by the learner. It clarifies the interrelationships among an object's critical features and aspects. It also illuminates the…

  16. Sensor Data Fusion

    DEFF Research Database (Denmark)

    Plascencia, Alfredo; Stepán, Petr

    2006-01-01

    The main contribution of this paper is to present a sensor fusion approach to scene environment mapping as part of a Sensor Data Fusion (SDF) architecture. This approach involves combined sonar array with stereo vision readings.  Sonar readings are interpreted using probability density functions...

  17. Fusion Canada issue 15

    International Nuclear Information System (INIS)

    1991-10-01

    A short bulletin from the National Fusion Program. Included in this issue is a report on the 1996 IAEA Fusion Conference site, operations at the Tokamak de Varennes including divertor pumping of impurities and pumping of carbon monoxide and methane, a discussion of the CFFTP and it's role. 1 fig

  18. The IGNITEX fusion project

    International Nuclear Information System (INIS)

    Carrera, R.

    1987-01-01

    The author discusses the recently proposed fusion ignition experiment, IGNITEX. He emphasizes the basic ideas of this concept rather than the specific details of the physics and engineering aspects of the experiment. This concept is a good example of the importance of maintaining an adequate balance between the basic scientific progress in fusion physics and the new technologies that are becoming available in order to make fusion work. The objective of the IGNITEX project is to produce and control ignited plasmas for scientific study in the simplest and least expensive way possible. Being able to study this not-yet-produced regime of plasma operation is essential to fusion research. Two years after the fission nuclear reaction was discovered, a non-self-sustained fission reaction was produced in a laboratory, and in one more year a self-sustained reaction was achieved at the University of Chicago. However, after almost forty years of fusion research, a self-sustained fusion reaction has yet not been produced in a laboratory experiment. This fact indicates the greater difficulty of the fusion experiment. Because of the difficulty involved in the production of a self-sustained fusion reaction, it is necessary to propose such an experiment with maximum ignition margins, maximum simplicity, and minimum financial risk

  19. Some fusion perspectives

    International Nuclear Information System (INIS)

    McNally, J.R. Jr.

    1977-01-01

    Some of the concepts of nuclear fusion reactions, advanced fusion fuels, environmental impacts, etc., are explored using the following general outline: I. Principles of Fusion (Nuclear Fuels and Reactions, Lawson Condition, n tau vs T, Nuclear Burn Characteristics); II. Magnetic Mirror Possibilities (the Ion Layer and Electron Layer, Exponential Build-up at MeV energies, Lorentz trapping at GeV energies); III. Pellet Fuel Fusion Prospects (Advanced Pellet Fuel Fusion Prospects, Burn Characteristics and Applications, Excitation-heating Prospects for Runaway Ion Temperatures). Inasmuch as the outline is very skeletal, a significant research and development effort may be in order to evaluate these prospects in more detail and hopefully ''harness the H-bomb'' for peaceful applications, the author concludes. 28 references

  20. Fusion fuel blanket technology

    International Nuclear Information System (INIS)

    Hastings, I.J.; Gierszewski, P.

    1987-05-01

    The fusion blanket surrounds the burning hydrogen core of a fusion reactor. It is in this blanket that most of the energy released by the nuclear fusion of deuterium-tritium is converted into useful product, and where tritium fuel is produced to enable further operation of the reactor. As fusion research turns from present short-pulse physics experiments to long-burn engineering tests in the 1990's, energy removal and tritium production capabilities become important. This technology will involve new materials, conditions and processes with applications both to fusion and beyond. In this paper, we introduce features of proposed blanket designs and update and status of international research. In focusing on the Canadian blanket technology program, we discuss the aqueous lithium salt blanket concept, and the in-reactor tritium recovery test program

  1. Fusion safety data base

    International Nuclear Information System (INIS)

    Laats, E.T.; Hardy, H.A.

    1983-01-01

    The purpose of this Fusion Safety Data Base Program is to provide a repository of data for the design and development of safe commercial fusion reactors. The program is sponsored by the United States Department of Energy (DOE), Office of Fusion Energy. The function of the program is to collect, examine, permanently store, and make available the safety data to the entire US magnetic-fusion energy community. The sources of data will include domestic and foreign fusion reactor safety-related research programs. Any participant in the DOE Program may use the Data Base Program from his terminal through user friendly dialog and can view the contents in the form of text, tables, graphs, or system diagrams

  2. Compact fusion reactors

    CERN Multimedia

    CERN. Geneva

    2015-01-01

    Fusion research is currently to a large extent focused on tokamak (ITER) and inertial confinement (NIF) research. In addition to these large international or national efforts there are private companies performing fusion research using much smaller devices than ITER or NIF. The attempt to achieve fusion energy production through relatively small and compact devices compared to tokamaks decreases the costs and building time of the reactors and this has allowed some private companies to enter the field, like EMC2, General Fusion, Helion Energy, Lawrenceville Plasma Physics and Lockheed Martin. Some of these companies are trying to demonstrate net energy production within the next few years. If they are successful their next step is to attempt to commercialize their technology. In this presentation an overview of compact fusion reactor concepts is given.

  3. Measles Virus Fusion Protein: Structure, Function and Inhibition

    Directory of Open Access Journals (Sweden)

    Philippe Plattet

    2016-04-01

    Full Text Available Measles virus (MeV, a highly contagious member of the Paramyxoviridae family, causes measles in humans. The Paramyxoviridae family of negative single-stranded enveloped viruses includes several important human and animal pathogens, with MeV causing approximately 120,000 deaths annually. MeV and canine distemper virus (CDV-mediated diseases can be prevented by vaccination. However, sub-optimal vaccine delivery continues to foster MeV outbreaks. Post-exposure prophylaxis with antivirals has been proposed as a novel strategy to complement vaccination programs by filling herd immunity gaps. Recent research has shown that membrane fusion induced by the morbillivirus glycoproteins is the first critical step for viral entry and infection, and determines cell pathology and disease outcome. Our molecular understanding of morbillivirus-associated membrane fusion has greatly progressed towards the feasibility to control this process by treating the fusion glycoprotein with inhibitory molecules. Current approaches to develop anti-membrane fusion drugs and our knowledge on drug resistance mechanisms strongly suggest that combined therapies will be a prerequisite. Thus, discovery of additional anti-fusion and/or anti-attachment protein small-molecule compounds may eventually translate into realistic therapeutic options.

  4. Ångstrom-size exocytotic fusion pore: Implications for pituitary hormone secretion.

    Science.gov (United States)

    Kreft, Marko; Jorgačevski, Jernej; Stenovec, Matjaž; Zorec, Robert

    2018-03-05

    In the past, vesicle content release was thought to occur immediately and completely after triggering of exocytosis. However, vesicles may merge with the plasma membrane to form an Ångstrom diameter fusion pore that prevents the exit of secretions from the vesicle lumen. The advantage of such a narrow pore is to minimize the delay between the trigger and the release. Instead of stimulating a sequence of processes, leading to vesicle merger with the plasma membrane and a formation of a fusion pore, the stimulus only widens the pre-established fusion pore. The fusion pore may be stable and may exhibit repetitive opening of the vesicle lumen to the cell exterior accompanied by a content discharge. Such release of vesicle content is partial (subquantal), and depends on fusion pore open time, diameter and the diffusibility of the cargo. Such transient mode of fusion pore opening was not confirmed until the development of the membrane capacitance patch-clamp technique, which enables high-resolution measurement of changes in membrane surface area. It allows millisecond dwell-time measurements of fusion pores with subnanometer diameters. Currently, the soluble N-ethylmaleimide-sensitive factor-attachment protein receptor (SNARE) proteins are considered to be key entities in end-stage exocytosis, and the SNARE complex assembly/disassembly may regulate the fusion pore. Moreover, lipids or other membrane constituents with anisotropic (non-axisymmetric) geometry may also favour the establishment of stable narrow fusion pores, if positioned in the neck of the fusion pore. Copyright © 2017 Elsevier B.V. All rights reserved.

  5. US fusion community discussion on fusion strategies

    International Nuclear Information System (INIS)

    Marton, W.A.

    1998-01-01

    On April 26 - May 1, 1998, a US Fusion Community Forum for Major Next-Step Experiments was held at Madison, Wisconsin, USA. Both the Single Integrated Step strategy and the Multiple Machine strategy have substantial support from the about 180 scientists and engineers who participated

  6. Membrane paradigm

    International Nuclear Information System (INIS)

    Price, R.H.; Thorne, K.S.

    1986-01-01

    The membrane paradigm is a modified frozen star approach to modeling black holes, with particles and fields assuming a complex, static, boundary-layer type structure (membrane) near the event horizon. The membrane has no effects on the present or future evolution of particles and fields above itself. The mathematical representation is a combination of a formalism containing terms for the shear and bulk viscosity, surface pressure, momentum, temperature, entropy, etc., of the horizon and the 3+1 formalism. The latter model considers a family of three-dimensional spacelike hypersurfaces in one-dimensional time. The membrane model considers a magnetic field threading the hole and undergoing torque from the hole rotation. The field is cleaned by the horizon and distributed over the horizon so that ohmic dissipation is minimized. The membrane paradigm is invalid inside the horizon, but is useful for theoretically probing the properties of slowly evolving black holes

  7. Membrane processes

    Science.gov (United States)

    Staszak, Katarzyna

    2017-11-01

    The membrane processes have played important role in the industrial separation process. These technologies can be found in all industrial areas such as food, beverages, metallurgy, pulp and paper, textile, pharmaceutical, automotive, biotechnology and chemical industry, as well as in water treatment for domestic and industrial application. Although these processes are known since twentieth century, there are still many studies that focus on the testing of new membranes' materials and determining of conditions for optimal selectivity, i. e. the optimum transmembrane pressure (TMP) or permeate flux to minimize fouling. Moreover the researchers proposed some calculation methods to predict the membrane processes properties. In this article, the laboratory scale experiments of membrane separation techniques, as well their validation by calculation methods are presented. Because membrane is the "heart" of the process, experimental and computational methods for its characterization are also described.

  8. A small molecule fusion inhibitor of dengue virus

    NARCIS (Netherlands)

    Poh, Mee Kian; Yip, Andy; Zhang, Summer; Priestle, John P.; Ma, Ngai Ling; Smit, Jolanda M.; Wischut, Jan; Shi, Pei-Yong; Wenk, Markus R.; Schul, Wouter

    2009-01-01

    The dengue virus envelope protein plays an essential role in viral entry by mediating fusion between the viral and host membranes. The crystal structure of the envelope protein shows a pocket (located at a "hinge" between Domains I and II) that can be occupied by ligand n-octyl-beta-D-glucoside

  9. Materials for fusion reactors

    International Nuclear Information System (INIS)

    Ehrlich, K.; Kaletta, D.

    1978-03-01

    The following report describes five papers which were given during the IMF seminar series summer 1977. The purpose of this series was to discuss especially the irradiation behaviour of materials intended for the first wall of future fusion reactors. The first paper deals with the basic understanding of plasma physics relating to the fusion reactor and presents the current state of art of fusion technology. The next two talks discuss the metals intended for the first wall and structural components of a fusion reactor. Since 14 MeV neutrons play an important part in the process of irradiation damage their role is discussed in detail. The question which machines are presently available to simulate irradiation damage under conditions similar to the ones found in a fusion reactor are investigated in the fourth talk which also presents the limitations of the different methods of simulation. In this context also discussed is the importance future intensive neutron sources and materials test reactors will have for this problem area. The closing paper has as a theme the review of the present status of research of metallic and non-metallic materials in view of the quite different requirements for different fusion systems; a closing topic is the world supply on rare materials required for fusion reactors. (orig) [de

  10. Energy from nuclear fusion

    International Nuclear Information System (INIS)

    Pinkau, K.

    1997-01-01

    Nuclear fusion research is conducted for the long-term objective of developing a power plant generating energy from the fusion of atomic nuclei. In order for the fusion fire to be ignited the fuel, a hydrogen plasma, must be confined in magnetic fields and heated to high temperatures - a design principle resulting in good safety characteristics and environmental compatibility. As the source materials required for the fusion process are available in almost unlimited quantities and are distributed all over the world, nuclear fusion could make a sizeable contribution towards future energy supplies. Since its beginnings in the early fifties, fusion research has approached its ambitious goal in painstaking, detailed work. Sometimes unnoticed by the public, these activities have made considerable progress especially in the past few years. Such formerly critical problems as plasma heating, thermal insulation, prevention of plasma impurities, and energy extraction can now be considered nearly solved. It has been possible in the meantime to generate fusion powers of several megawatt. The results obtained so far allow a test reactor to be planned which, for the first time, is to produce a self-sustaining plasma with powers in the gigawatt range. (orig.) [de

  11. Fusion Studies in Japan

    Science.gov (United States)

    Ogawa, Yuichi

    2016-05-01

    A new strategic energy plan decided by the Japanese Cabinet in 2014 strongly supports the steady promotion of nuclear fusion development activities, including the ITER project and the Broader Approach activities from the long-term viewpoint. Atomic Energy Commission (AEC) in Japan formulated the Third Phase Basic Program so as to promote an experimental fusion reactor project. In 2005 AEC has reviewed this Program, and discussed on selection and concentration among many projects of fusion reactor development. In addition to the promotion of ITER project, advanced tokamak research by JT-60SA, helical plasma experiment by LHD, FIREX project in laser fusion research and fusion engineering by IFMIF were highly prioritized. Although the basic concept is quite different between tokamak, helical and laser fusion researches, there exist a lot of common features such as plasma physics on 3-D magnetic geometry, high power heat load on plasma facing component and so on. Therefore, a synergetic scenario on fusion reactor development among various plasma confinement concepts would be important.

  12. Role of Conserved Histidine Residues in the Low-pH Dependence of the Semliki Forest Virus Fusion Protein▿

    OpenAIRE

    Qin, Zhao-ling; Zheng, Yan; Kielian, Margaret

    2009-01-01

    A wide variety of enveloped viruses infects cells by taking advantage of the low pH in the endocytic pathway to trigger virus-membrane fusion. For alphaviruses such as Semliki Forest virus (SFV), acidic pH initiates a series of conformational changes in the heterodimeric virus envelope proteins E1 and E2. Low pH dissociates the E2/E1 dimer, releasing the membrane fusion protein E1. E1 inserts into the target membrane and refolds to a trimeric hairpin conformation, thus driving the fusion reac...

  13. Beam dancer fusion device

    International Nuclear Information System (INIS)

    Maier, H.B.

    1984-01-01

    To accomplish fusion of two or more fusion fuel elements numerous minute spots of energy or laser light are directed to a micro target area, there to be moved or danced about by a precision mechanical controlling apparatus at the source of the laser light or electromagnetic energy beams, so that merging and coinciding patterns of light or energy beams can occur around the area of the fuel atoms or ions. The projecting of these merging patterns may be considered as target searching techniques to locate responsive clusters of fuel elements and to compress such elements into a condition in which fusion may occur. Computerized programming may be used

  14. Mirror fusion reactors

    International Nuclear Information System (INIS)

    Anon.

    1978-01-01

    Conceptual design studies were made of fusion reactors based on the three current mirror-confinement concepts: the standard mirror, the tandem mirror, and the field-reversed mirror. Recent studies of the standard mirror have emphasized its potential as a fusion-fission hybrid reactor, designed to produce fuel for fission reactors. We have designed a large commercial hybrid and a small pilot-plant hybrid based on standard mirror confinement. Tandem mirror designs include a commercial 1000-MWe fusion power plant and a nearer term tandem mirror hybrid. Field-reversed mirror designs include a multicell commercial reactor producing 75 MWe and a single-cell pilot plant

  15. Fusion Reactor Materials

    International Nuclear Information System (INIS)

    Decreton, M.

    2002-01-01

    The objective of SCK-CEN's programme on fusion reactor materials is to contribute to the knowledge on the radiation-induced behaviour of fusion reactor materials and components as well as to help the international community in building the scientific and technical basis needed for the construction of the future reactor. Ongoing projects include: the study of the mechanical and chemical (corrosion) behaviour of structural materials under neutron irradiation and water coolant environment; the investigation of the characteristics of irradiated first wall material such as beryllium; investigations on the management of materials resulting from the dismantling of fusion reactors including waste disposal. Progress and achievements in these areas in 2001 are discussed

  16. Fusion Reactor Materials

    International Nuclear Information System (INIS)

    Decreton, M.

    2001-01-01

    The objective of SCK-CEN's programme on fusion reactor materials is to contribute to the knowledge on the behaviour of fusion reactor materials and components during and after irradiation. Ongoing projects include: the study of the mechanical behaviour of structural materials under neutron irradiation; the investigation of the characteristics of irradiated first wall material such as beryllium; the detection of abrupt electrical degradation of insulating ceramics under high temperature and neutron irradiation; and the study of dismantling and waste disposal strategy for fusion reactors. Progress and achievements in these areas in 2000 are discussed

  17. Mirror fusion reactors

    International Nuclear Information System (INIS)

    Carlson, G.A.; Moir, R.W.

    1978-01-01

    We have carried out conceptual design studies of fusion reactors based on the three current mirror confinement concepts: the standard mirror, the tandem mirror, and the field-reversed mirror. Recent studies of the standard mirror have emphasized its potential as a fusion-fission hybrid reactor, designed to produce fission fuel for fission reactors. We have designed a large commercial hybrid based on standard mirror confinement, and also a small pilot plant hybrid. Tandem mirror designs include a commercial 1000 MWe fusion power plant and a nearer term tandem mirror hybrid. Field-reversed mirror designs include a multicell commercial reactor producing 75 MWe and a single cell pilot plant

  18. Remote sensing image fusion

    CERN Document Server

    Alparone, Luciano; Baronti, Stefano; Garzelli, Andrea

    2015-01-01

    A synthesis of more than ten years of experience, Remote Sensing Image Fusion covers methods specifically designed for remote sensing imagery. The authors supply a comprehensive classification system and rigorous mathematical description of advanced and state-of-the-art methods for pansharpening of multispectral images, fusion of hyperspectral and panchromatic images, and fusion of data from heterogeneous sensors such as optical and synthetic aperture radar (SAR) images and integration of thermal and visible/near-infrared images. They also explore new trends of signal/image processing, such as

  19. Induction of Cell-Cell Fusion by Ebola Virus Glycoprotein: Low pH Is Not a Trigger.

    Directory of Open Access Journals (Sweden)

    Ruben M Markosyan

    2016-01-01

    Full Text Available Ebola virus (EBOV is a highly pathogenic filovirus that causes hemorrhagic fever in humans and animals. Currently, how EBOV fuses its envelope membrane within an endosomal membrane to cause infection is poorly understood. We successfully measure cell-cell fusion mediated by the EBOV fusion protein, GP, assayed by the transfer of both cytoplasmic and membrane dyes. A small molecule fusion inhibitor, a neutralizing antibody, as well as mutations in EBOV GP known to reduce viral infection, all greatly reduce fusion. By monitoring redistribution of small aqueous dyes between cells and by electrical capacitance measurements, we discovered that EBOV GP-mediated fusion pores do not readily enlarge-a marked difference from the behavior of other viral fusion proteins. EBOV GP must be cleaved by late endosome-resident cathepsins B or L in order to become fusion-competent. Cleavage of cell surface-expressed GP appears to occur in endosomes, as evidenced by the fusion block imposed by cathepsin inhibitors, agents that raise endosomal pH, or an inhibitor of anterograde trafficking. Treating effector cells with a recombinant soluble cathepsin B or thermolysin, which cleaves GP into an active form, increases the extent of fusion, suggesting that a fraction of surface-expressed GP is not cleaved. Whereas the rate of fusion is increased by a brief exposure to acidic pH, fusion does occur at neutral pH. Importantly, the extent of fusion is independent of external pH in experiments in which cathepsin activity is blocked and EBOV GP is cleaved by thermolysin. These results imply that low pH promotes fusion through the well-known pH-dependent activity of cathepsins; fusion induced by cleaved EBOV GP is a process that is fundamentally independent of pH. The cell-cell fusion system has revealed some previously unappreciated features of EBOV entry, which could not be readily elucidated in the context of endosomal entry.

  20. Structure information from fusion barriers

    Indian Academy of Sciences (India)

    effects on the fusion excitation function. However, a simultaneous analysis of the fusion, elastic and quasi-elastic channels would fix the structure and the reaction unambiguously. Keywords. Heavy ion fusion; fusion barrier distributions; nuclear structure; coupled reaction chan- nel calculations. PACS Nos 25.70.Bc; 25.70.

  1. Why and how of fusion

    International Nuclear Information System (INIS)

    Miley, G.H.

    1977-01-01

    The potential advantages of fusion power are listed. The approaches to plasma containment are mentioned and the status of the fusion program is described. The ERDA and EPRI programs are discussed. The Fusion Energy Foundation's activities are mentioned. Fusion research at the U. of Ill. is described briefly

  2. Paramyxovirus Fusion and Entry: Multiple Paths to a Common End

    Science.gov (United States)

    Chang, Andres; Dutch, Rebecca E.

    2012-01-01

    The paramyxovirus family contains many common human pathogenic viruses, including measles, mumps, the parainfluenza viruses, respiratory syncytial virus, human metapneumovirus, and the zoonotic henipaviruses, Hendra and Nipah. While the expression of a type 1 fusion protein and a type 2 attachment protein is common to all paramyxoviruses, there is considerable variation in viral attachment, the activation and triggering of the fusion protein, and the process of viral entry. In this review, we discuss recent advances in the understanding of paramyxovirus F protein-mediated membrane fusion, an essential process in viral infectivity. We also review the role of the other surface glycoproteins in receptor binding and viral entry, and the implications for viral infection. Throughout, we concentrate on the commonalities and differences in fusion triggering and viral entry among the members of the family. Finally, we highlight key unanswered questions and how further studies can identify novel targets for the development of therapeutic treatments against these human pathogens. PMID:22590688

  3. Paramyxovirus Fusion and Entry: Multiple Paths to a Common End

    Directory of Open Access Journals (Sweden)

    Rebecca E. Dutch

    2012-04-01

    Full Text Available The paramyxovirus family contains many common human pathogenic viruses, including measles, mumps, the parainfluenza viruses, respiratory syncytial virus, human metapneumovirus, and the zoonotic henipaviruses, Hendra and Nipah. While the expression of a type 1 fusion protein and a type 2 attachment protein is common to all paramyxoviruses, there is considerable variation in viral attachment, the activation and triggering of the fusion protein, and the process of viral entry. In this review, we discuss recent advances in the understanding of paramyxovirus F protein-mediated membrane fusion, an essential process in viral infectivity. We also review the role of the other surface glycoproteins in receptor binding and viral entry, and the implications for viral infection. Throughout, we concentrate on the commonalities and differences in fusion triggering and viral entry among the members of the family. Finally, we highlight key unanswered questions and how further studies can identify novel targets for the development of therapeutic treatments against these human pathogens.

  4. Primordial membranes

    DEFF Research Database (Denmark)

    Hanczyc, Martin M; Monnard, Pierre-Alain

    2017-01-01

    Cellular membranes, which are self-assembled bilayer structures mainly composed of lipids, proteins and conjugated polysaccharides, are the defining feature of cell physiology. It is likely that the complexity of contemporary cells was preceded by simpler chemical systems or protocells during...... the various evolutionary stages that led from inanimate to living matter. It is also likely that primitive membranes played a similar role in protocell 'physiology'. The composition of such ancestral membranes has been proposed as mixtures of single hydrocarbon chain amphiphiles, which are simpler versions...

  5. Nuclear fusion during yeast mating occurs by a three-step pathway.

    Science.gov (United States)

    Melloy, Patricia; Shen, Shu; White, Erin; McIntosh, J Richard; Rose, Mark D

    2007-11-19

    In Saccharomyces cerevisiae, mating culminates in nuclear fusion to produce a diploid zygote. Two models for nuclear fusion have been proposed: a one-step model in which the outer and inner nuclear membranes and the spindle pole bodies (SPBs) fuse simultaneously and a three-step model in which the three events occur separately. To differentiate between these models, we used electron tomography and time-lapse light microscopy of early stage wild-type zygotes. We observe two distinct SPBs in approximately 80% of zygotes that contain fused nuclei, whereas we only see fused or partially fused SPBs in zygotes in which the site of nuclear envelope (NE) fusion is already dilated. This demonstrates that SPB fusion occurs after NE fusion. Time-lapse microscopy of zygotes containing fluorescent protein tags that localize to either the NE lumen or the nucleoplasm demonstrates that outer membrane fusion precedes inner membrane fusion. We conclude that nuclear fusion occurs by a three-step pathway.

  6. Fusion Revisits CERN

    CERN Multimedia

    2001-01-01

    It's going to be a hot summer at CERN. At least in the Main Building, where from 13 July to 20 August an exhibition is being hosted on nuclear fusion, the energy of the Stars. Nuclear fusion is the engine driving the stars but also a potential source of energy for mankind. The exhibition shows the different nuclear fusion techniques and research carried out on the subject in Europe. Inaugurated at CERN in 1993, following collaboration between Lausanne's CRPP-EPFL and CERN, with input from Alessandro Pascolini of Italy's INFN, this exhibition has travelled round Europe before being revamped and returning to CERN. 'Fusion, Energy of the Stars', from 13 July onwards, Main Building

  7. Aneutronic Fusion Spacecraft Architecture

    Data.gov (United States)

    National Aeronautics and Space Administration — Description: provide framework to realize fusion propulsion for long-range space travel; analyze "hybrid" schemes with a solar or fission primary energy source along...

  8. Fusion plasma physics

    CERN Document Server

    Stacey, Weston M

    2012-01-01

    This revised and enlarged second edition of the popular textbook and reference contains comprehensive treatments of both the established foundations of magnetic fusion plasma physics and of the newly developing areas of active research. It concludes with a look ahead to fusion power reactors of the future. The well-established topics of fusion plasma physics -- basic plasma phenomena, Coulomb scattering, drifts of charged particles in magnetic and electric fields, plasma confinement by magnetic fields, kinetic and fluid collective plasma theories, plasma equilibria and flux surface geometry, plasma waves and instabilities, classical and neoclassical transport, plasma-materials interactions, radiation, etc. -- are fully developed from first principles through to the computational models employed in modern plasma physics. The new and emerging topics of fusion plasma physics research -- fluctuation-driven plasma transport and gyrokinetic/gyrofluid computational methodology, the physics of the divertor, neutral ...

  9. Fusion cost normalization

    International Nuclear Information System (INIS)

    Schulte, S.C.; Willke, T.L.

    1978-01-01

    The categorization and accounting methods described in this paper provide a common format that can be used to assess the economic character of magnetically confined fusion reactor design concepts. The format was developed with assistance from the fusion economics community, thus ensuring that the methods meet with the approval of potential users. The format will aid designers in the preparation of design concept cost estimates and also provide policy makers with a tool to assist in appraising which design concepts may be economically promising. Adherence to the format when evaluating prospective fusion reactor design concepts will result in the identification of the more promising concepts, thus enabling the fusion power alternatives with better economic potential to be quickly and efficiently developed

  10. Fusion reactor materials

    International Nuclear Information System (INIS)

    Rowcliffe, A.F.; Burn, G.L.; Knee', S.S.; Dowker, C.L.

    1994-02-01

    This is the fifteenth in a series of semiannual technical progress reports on fusion reactor materials. This report combines research and development activities which were previously reported separately in the following progress reports: Alloy Development for Irradiation Performance; Damage Analysis and Fundamental Studies; Special purpose Materials. These activities are concerned principally with the effects of the neutronic and chemical environment on the properties and performance of reactor materials; together they form one element of the overall materials programs being conducted in support of the Magnetic Fusion Energy Program of the U.S. Department of Energy. The Fusion Reactor Materials Program is a national effort involving several national laboratories, universities, and industries. The purpose of this series of reports is to provide a working technical record for the use of the program participants, and to provide a means of communicating the efforts of materials scientists to the rest of the fusion community, both nationally and worldwide

  11. Complimentary Advanced Fusion Exploration

    National Research Council Canada - National Science Library

    Alford, Mark G; Jones, Eric C; Bubalo, Adnan; Neumann, Melissa; Greer, Michael J

    2005-01-01

    .... The focus areas were in the following regimes: multi-tensor homographic computer vision image fusion, out-of-sequence measurement and track data handling, Nash bargaining approaches to sensor management, pursuit-evasion game theoretic modeling...

  12. Fusion technology programme

    International Nuclear Information System (INIS)

    Finken, D.

    1984-04-01

    KfK participates to the Fusion Technology Programme of the European Community. Most of the work in progress addresses the Next European Torus (NET) and the long term technology aspects as defined in the 82/86 programme. A minor part serves to preparation of future contributions and to design studies on fusion concepts in a wider perspective. The Fusion Technology Programme of Euratom covers mainly aspects of nuclear engineering. Plasma engineering, heating, refueling and vacuum technology are at present part of the Physics Programme. In view of NET, integration of the different areas of work will be mandatory. KfK is therefore prepared to address technical aspects beyond the actual scope of the physics experiments. The technology tasks are reported project wise under title and code of the Euratom programme. Most of the projects described here are shared with other European fusion laboratories as indicated in the table annexed to this report. (orig./GG)

  13. International aspects of fusion

    International Nuclear Information System (INIS)

    Stacey, W.M. Jr.

    1979-12-01

    International collaborative efforts in magnetic confinement fusion in which the USA is involved are reviewed. These efforts are carried under the auspices of international agencies and through bilateral agreements

  14. Fusion safety program plan

    International Nuclear Information System (INIS)

    Crocker, J.G.; Holland, D.F.; Herring, J.S.

    1980-09-01

    The program plan consists of research that has been divided into 13 different areas. These areas focus on the radioactive inventories that are expected in fusion reactors, the energy sources potentially available to release a portion of these inventories, and analysis and design techniques to assess and ensure that the safety risks associated with operation of magnetic fusion facilities are acceptably low. The document presents both long-term program requirements that must be fulfilled as part of the commercialization of fusion power and a five-year plan for each of the 13 different program areas. Also presented is a general discussion of magnetic fusion reactor safety, a method for establishing priorities in the program, and specific priority ratings for each task in the five-year plan

  15. Fusion-breeder program

    International Nuclear Information System (INIS)

    Moir, R.W.

    1982-01-01

    The various approaches to a combined fusion-fission reactor for the purpose of breeding 239 Pu and 233 U are described. Design aspects and cost estimates for fuel production and electricity generation are discussed

  16. Fusion technology (FT)

    International Nuclear Information System (INIS)

    1978-01-01

    The annual report of tha fusion technology (FT) working group discusses the projects carried out by the participating institutes in the fields of 1) fuel injection and plasma heating, 2) magnetic field technology, and 3) systems investigations. (HK) [de

  17. Cold nuclear fusion device

    International Nuclear Information System (INIS)

    Ogino, Shinji.

    1991-01-01

    Selection of cathode material is a key to the attainment of cold nuclear fusion. However, there are only few reports on the cathode material at present and an effective development has been demanded. The device comprises an anode and a cathode and an electrolytic bath having metal salts dissolved therein and containing heavy water in a glass container. The anode is made of gold or platinum and the cathode is made of metals of V, Sr, Y, Nb, Hf or Ta, and a voltage of 3-25V is applied by way of a DC power source between them. The metal comprising V, Sr, Y, Nb, Hf or Ta absorbs deuterium formed by electrolysis of heavy water effectively to cause nuclear fusion reaction at substantially the same frequency and energy efficiency as palladium and titanium. Accordingly, a cold nuclear fusion device having high nuclear fusion generation frequency can be obtained. (N.H.)

  18. Egg CD9 protein tides correlated with sperm oscillations tune the gamete fusion ability in mammal.

    Science.gov (United States)

    Ravaux, Benjamin; Favier, Sophie; Perez, Eric; Gourier, Christine

    2018-01-23

    Mammalian fertilization involves membrane events -adhesion, fusion, sperm engulfment, membrane block to polyspermy- whose causes remain largely unknown. Recently, specific oscillations of the sperm in contact with the egg were shown to be necessary for fusion. Using a microfluidic chip to impose the venue for the encounter of two gametes allowed real-time observation of the membrane remodelling occurring at the sperm/egg interface. The spatiotemporal mapping of egg CD9 revealed that this protein concentrates at the egg/sperm interface as a result of sperm oscillations, until a CD9-rich platform is nucleated on which fusion immediately takes place. Within 2 to 5 minutes after fusion, most of the CD9 leaves the egg for the external aqueous medium. Then an egg membrane wave engulfs the sperm head in approximately 25 minutes. These results show that sperm oscillations initiate the CD9 recruitment that causes gamete fusion after which CD9 and associated proteins leave the membrane in a process likely to contribute to block polyspermy. They highlight that the gamete fusion story in mammals is an unexpected interplay between mechanical constraints and proteins. © The Author(s) (2018). Published by Oxford University Press on behalf of Journal of Molecular Cell Biology, IBCB, SIBS, CAS. All rights reserved.

  19. Conference on Norwegian fusion research

    International Nuclear Information System (INIS)

    The question of instituting a systematic research programme in Norway on aspects of thermonuclear and plasma physics has been raised. The conference here reported was intended to provide basic information on the status of fusion research internationally and to discuss a possible Norwegian programme. The main contributions covered the present status of fusion research, international cooperation, fusion research in small countries and minor laboratories, fusion research in Denmark and Sweden, and a proposed fusion experiment in Bergen. (JIW)

  20. Bringing together fusion research

    International Nuclear Information System (INIS)

    Leiser, M.

    1982-01-01

    The increasing involvement of the IAEA in fusion, together with the growing efforts devoted to this area, are described. The author puts forward the idea that one of the most important aspects of this involvement is in providing a world-wide forum for scientists. The functions of the IFRC (International Fusion Research Council) as an advisory group are outlined, and the role played by IFRC in the definition and objectives of INTOR (International Tokamak Reactor) are briefly described

  1. Fusion Simulation Program

    International Nuclear Information System (INIS)

    Greenwald, Martin

    2011-01-01

    Many others in the fusion energy and advanced scientific computing communities participated in the development of this plan. The core planning team is grateful for their important contributions. This summary is meant as a quick overview the Fusion Simulation Program's (FSP's) purpose and intentions. There are several additional documents referenced within this one and all are supplemental or flow down from this Program Plan. The overall science goal of the DOE Office of Fusion Energy Sciences (FES) Fusion Simulation Program (FSP) is to develop predictive simulation capability for magnetically confined fusion plasmas at an unprecedented level of integration and fidelity. This will directly support and enable effective U.S. participation in International Thermonuclear Experimental Reactor (ITER) research and the overall mission of delivering practical fusion energy. The FSP will address a rich set of scientific issues together with experimental programs, producing validated integrated physics results. This is very well aligned with the mission of the ITER Organization to coordinate with its members the integrated modeling and control of fusion plasmas, including benchmarking and validation activities. (1). Initial FSP research will focus on two critical Integrated Science Application (ISA) areas: ISA1, the plasma edge; and ISA2, whole device modeling (WDM) including disruption avoidance. The first of these problems involves the narrow plasma boundary layer and its complex interactions with the plasma core and the surrounding material wall. The second requires development of a computationally tractable, but comprehensive model that describes all equilibrium and dynamic processes at a sufficient level of detail to provide useful prediction of the temporal evolution of fusion plasma experiments. The initial driver for the whole device model will be prediction and avoidance of discharge-terminating disruptions, especially at high performance, which are a critical

  2. Perspectives of fusion power

    International Nuclear Information System (INIS)

    Jensen, V.O.

    1984-01-01

    New and practically inexhaustible sources of energy must be developed for the period when oil, coal and uranium will become scarce and expensive. Nuclear fusion holds great promise as one of these practically inexhaustible energy sources. Based on the deuteriumtritium reaction with tritium obtained from naturally occuring lithium, which is also widely available in Europe, the accessible energy resources in the world are 3.10 12 to 3.10 16 toe; based on the deuterium-deuterium reaction, the deuterium content of the oceans corresponds to 10 20 toe. It is presently envisaged that in order to establish fusion as a large-scale energy source, three major thresholds must be reached: - Scientific feasibility, - Technical feasibility, i.e. the proof that the basic technical problems of the fusion reactor can be solved. - Commercial feasibility, i.e. proof that fusion power reactors can be built on an industrial scale, can be operated reliably and produce usable energy at prices competitive with other energy sources. From the above it is clear that the route to commercial fusion will be long and costly and involve the solution of extremely difficult technical problems. In view of the many steps which have to be taken, it appears unlikely that commercial fusion power will be in general use within the next 50 years and by that time world-wide expenditure on research, development and demonstration may well have exceeded 100 Bio ECU. (author)

  3. Status of fusion technology

    International Nuclear Information System (INIS)

    Mohan, Ashok

    1978-01-01

    The current status of fusion technology is surveyed. Limited reserves of fossil fuel and dangers of proliferation from nuclear reactors have brought into focus the need to develop an optional energy source. Fusion is being looked upon as an optional energy source which is free from environmental hazards unlike fossil fuels and nuclear reactors. Investments in R and D of fusion energy have increased rapidly in USA, Japan, USSR and European countries. Out of the various fusion fuels known, a mixture of D and T is widely chosen. The main problem in fusion technology is the confinement of plasma for a time sufficient to start the fusion reaction. This can be done magnetically or inertially. The three approaches to magnetic confinement are : (1) tokamak, (2) mirror and (3) pinch. Inertial confinement makes use of lasers or electron beams or ion beams. Both the methods of confinement i.e. magnetic and inertial have problems which are identified and their nature is discussed. (M.G.B.)

  4. Energy from inertial fusion

    International Nuclear Information System (INIS)

    1995-03-01

    This book contains 22 articles on inertial fusion energy (IFE) research and development written in the framework of an international collaboration of authors under the guidance of an advisory group on inertial fusion energy set up in 1991 to advise the IAEA. It describes the actual scientific, engineering and technological developments in the field of inertial confinement fusion (ICF). It also identifies ways in which international co-operation in ICF could be stimulated. The book is intended for a large audience and provides an introduction to inertial fusion energy and an overview of the various technologies needed for IFE power plants to be developed. It contains chapters on (i) the fundamentals of IFE; (ii) inertial confinement target physics; (iii) IFE power plant design principles (requirements for power plant drivers, solid state laser drivers, gas laser drivers, heavy ion drivers, and light ion drivers, target fabrication and positioning, reaction chamber systems, power generation and conditioning and radiation control, materials management and target materials recovery), (iv) special design issues (radiation damage in structural materials, induced radioactivity, laser driver- reaction chamber interfaces, ion beam driver-reaction chamber interfaces), (v) inertial fusion energy development strategy, (vi) safety and environmental impact, (vii) economics and other figures of merit; (viii) other uses of inertial fusion (both those involving and not involving implosions); and (ix) international activities. Refs, figs and tabs

  5. Controlled thermonuclear fusion: research on magnetic fusion

    International Nuclear Information System (INIS)

    Paris, P.J.

    1988-12-01

    Recent progress in thermonuclear fusion research indicates that the scientists' schedule for the demonstration of the scientific feasibility will be kept and that break-even will be attained in the course of the next decade. To see the implementation of ignition, however, the generation of future experiments must be awaited. These projects are currently under study. With technological research going on in parallel, they should at the same time contribute to the design of a reactor. Fusion reactors will be quite different from the fission nuclear reactors we know, and the waste of the plants will also be of a different nature. It is still too early to define the precise design of a fusion reactor. On the basis of a toric machine concept like that of the tokamak, we can, however, envisage that the problems with which we are confronted will be solved one after the other. As we have just seen, these will be the objectives of the future experimental installations where ignition will be possible and where the flux of fast neutrons will be so strong that they will allow the study of low-activation materials which will be used in the structure of the reactor. But this is also a task in which from now onwards numerous laboratories in Europe and in the world participate. The works are in fact punctiform, and often the mutual incidences can only be determined by an approach simulated by numerical codes. (author) 19 figs., 6 tabs., 8 refs

  6. Osteoclast Fusion is Based on Heterogeneity Between Fusion Partners

    DEFF Research Database (Denmark)

    Hobolt-Pedersen, Anne-Sofie; Delaissé, Jean-Marie; Søe, Kent

    2014-01-01

    Bone-resorbing osteoclasts are formed through fusion of mononucleated precursors. Their choice of partners during the fusion process remains unclear. We hypothesized that osteoclasts are selective in their choice of fusion partner and that this selectivity is based on heterogeneity among the cells...... with respect to their maturation stage and their expression and cellular organization of fusion factors. Support for this hypothesis was found from immunofluorescence staining of the osteoclast fusion factors CD47, dendritic cell-specific transmembrane protein (DC-STAMP), and syncytin-1. These stainings...... fusion steps was also suggested from experiments with a CD47 blocking antibody, which resulted in an inhibition of the fusion of small osteoclasts. Conversely, blocking of connexin 43 affected the fusion of larger osteoclasts with four or more nuclei. The suggestion that different fusion factors function...

  7. The rigid amphipathic fusion inhibitor dUY11 acts through photosensitization of viruses.

    Science.gov (United States)

    Vigant, Frederic; Hollmann, Axel; Lee, Jihye; Santos, Nuno C; Jung, Michael E; Lee, Benhur

    2014-02-01

    Rigid amphipathic fusion inhibitors (RAFIs) are lipophilic inverted-cone-shaped molecules thought to antagonize the membrane curvature transitions that occur during virus-cell fusion and are broad-spectrum antivirals against enveloped viruses (Broad-SAVE). Here, we show that RAFIs act like membrane-binding photosensitizers: their antiviral effect is dependent on light and the generation of singlet oxygen ((1)O(2)), similar to the mechanistic paradigm established for LJ001, a chemically unrelated class of Broad-SAVE. Photosensitization of viral membranes is a common mechanism that underlies these Broad-SAVE.

  8. Advanced fusion reactor

    Energy Technology Data Exchange (ETDEWEB)

    Tomita, Yukihiro [National Inst. for Fusion Science, Toki, Gifu (Japan)

    2003-04-01

    The main subjects on fusion research are now on D-T fueled fusion, mainly due to its high fusion reaction rate. However, many issues are still remained on the wall loading by the 14 MeV neutrons. In the case of D-D fueled fusion, the neutron wall loading is still remained, though the technology related to tritium breeding is not needed. The p-{sup 6}Li and p-{sup 11}B fueled fusions are not estimated to be the next generation candidate until the innovated plasma confinement technologies come in useful to achieve the high performance plasma parameters. The fusion reactor of D-{sup 3}He fuels has merits on the smaller neutron wall loading and tritium handling. However, there are difficulties on achieving the high temperature plasma more than 100 keV. Furthermore the high beta plasma is needed to decrease synchrotron radiation loss. In addition, the efficiency of the direct energy conversion from protons coming out from fusion reaction is one of the key parameters in keeping overall power balance. Therefore, open magnetic filed lines should surround the plasma column. In this paper, we outlined the design of the commercial base reactor (ARTEMIS) of 1 GW electric output power configured by D-{sup 3}He fueled FRC (Field Reversed Configuration). The ARTEMIS needs 64 kg of {sup 3}He per a year. On the other hand, 1 million tons of {sup 3}He is estimated to be in the moon. The {sup 3}He of about 10{sup 23} kg are to exist in gaseous planets such as Jupiter and Saturn. (Y. Tanaka)

  9. Identification of cancer fusion drivers using network fusion centrality

    Science.gov (United States)

    Wu, Chia-Chin; Kannan, Kalpana; Lin, Steven; Yen, Laising; Milosavljevic, Aleksandar

    2013-01-01

    Summary: Gene fusions are being discovered at an increasing rate using massively parallel sequencing technologies. Prioritization of cancer fusion drivers for validation cannot be performed using traditional single-gene based methods because fusions involve portions of two partner genes. To address this problem, we propose a novel network analysis method called fusion centrality that is specifically tailored for prioritizing gene fusions. We first propose a domain-based fusion model built on the theory of exon/domain shuffling. The model leads to a hypothesis that a fusion is more likely to be an oncogenic driver if its partner genes act like hubs in a network because the fusion mutation can deregulate normal functions of many other genes and their pathways. The hypothesis is supported by the observation that for most known cancer fusion genes, at least one of the fusion partners appears to be a hub in a network, and even for many fusions both partners appear to be hubs. Based on this model, we construct fusion centrality, a multi-gene-based network metric, and use it to score fusion drivers. We show that the fusion centrality outperforms other single gene-based methods. Specifically, the method successfully predicts most of 38 newly discovered fusions that had validated oncogenic importance. To our best knowledge, this is the first network-based approach for identifying fusion drivers. Availability: Matlab code implementing the fusion centrality method is available upon request from the corresponding authors. Contact: perwu777@gmail.com Supplementary information: Supplementary data are available at Bioinformatics online. PMID:23505294

  10. Fast Vesicle Fusion in Living Cells Requires at Least Three SNARE Complexes

    DEFF Research Database (Denmark)

    Mohrmann, Ralf; de Wit, Heidi; Verhage, Matthijs

    2010-01-01

    chromaffin cells. Simultaneous expression of wild-type SNAP-25 and a mutant unable to support exocytosis progressively altered fusion kinetics and fusion pore opening, indicating that both proteins assemble into heteromeric fusion complexes. Expressing different wild-type:mutant ratios revealed a third power......Exocytosis requires formation of SNARE complexes between vesicle- and target-membranes. Recent assessments in reduced model systems have produced divergent estimates of the number of SNARE complexes needed for fusion. Here, we used a titration approach to answer this question in intact, cultured...... relationship for fast (synchronous) fusion and a near-linear relationship for overall release. Thus, fast fusion typically observed in synapses and neurosecretory cells requires at least three functional SNARE complexes, while slower release might occur with fewer. Heterogeneity in SNARE-complex number may...

  11. Coatings for fusion reactor environments

    International Nuclear Information System (INIS)

    Mattox, D.M.

    1979-01-01

    The internal surfaces of a tokamak fusion reactor control the impurity injection and gas recycling into the fusion plasma. Coating of internal surfaces may provide a desirable and possibly necessary design flexibility for achieving the temperatures, ion densities and containment times necessary for net energy production from fusion reactions to take place. In this paper the reactor environments seen by various componentare reviewed along with possible materials responses. Characteristics of coating-substrate systems, important to fusion applications, are delineated and the present status of coating development for fusion applications is reviewed. Coating development for fusion applications is just beginning and poses a unique and important challenge for materials development

  12. Fusion technology programme

    International Nuclear Information System (INIS)

    Finken, D.

    1986-05-01

    In 1982, KfK joined the fusion programme of EURATOM as a further association introducing its experience in nuclear technology. KfK closely cooperates with IPP Garching, the two institutions forming a research unit aiming at planning and realization of future development steps of fusion. KfK has combined its forces in the Nuclear Fusion Project (PKF) with participation of several KfK departments to the project tasks. Previous work of KfK in magnetic fusion has addressed mainly superconducting magnets, plasma heating by cluster ions and studies on structural materials. At present, emphasis of our work has concentrated increasingly on the nuclear part, i.e. the first wall and blanket structures and the elements of the tritium extraction and purification system. Associated to this component development are studies of remote maintenance and safety. Most of the actual work addresses NET, the next step to a demonstration of fusion feasibility. NET is supposed to follow JET, the operating plasma physics experiment of Euratom, on the 1990's. Detailed progress of the work in the past half year is described in this report. (orig./GG)

  13. Myoblast fusion in Drosophila

    International Nuclear Information System (INIS)

    Haralalka, Shruti; Abmayr, Susan M.

    2010-01-01

    The body wall musculature of a Drosophila larva is composed of an intricate pattern of 30 segmentally repeated muscle fibers in each abdominal hemisegment. Each muscle fiber has unique spatial and behavioral characteristics that include its location, orientation, epidermal attachment, size and pattern of innervation. Many, if not all, of these properties are dictated by founder cells, which determine the muscle pattern and seed the fusion process. Myofibers are then derived from fusion between a specific founder cell and several fusion competent myoblasts (FCMs) fusing with as few as 3-5 FCMs in the small muscles on the most ventral side of the embryo and as many as 30 FCMs in the larger muscles on the dorsal side of the embryo. The focus of the present review is the formation of the larval muscles in the developing embryo, summarizing the major issues and players in this process. We have attempted to emphasize experimentally-validated details of the mechanism of myoblast fusion and distinguish these from the theoretically possible details that have not yet been confirmed experimentally. We also direct the interested reader to other recent reviews that discuss myoblast fusion in Drosophila, each with their own perspective on the process . With apologies, we use gene nomenclature as specified by Flybase (http://flybase.org) but provide Table 1 with alternative names and references.

  14. Material for fusion reactor

    International Nuclear Information System (INIS)

    Abhishek, Anuj; Ranjan, Prem

    2011-01-01

    To make nuclear fusion power a reality, the scientists are working restlessly to find the materials which can confine the power generated by the fusion of two atomic nuclei. A little success in this field has been achieved, though there are still miles to go. Fusion reaction is a special kind of reaction which must occur at very high density and temperature to develop extremely large amount of energy, which is very hard to control and confine within using the present techniques. As a whole it requires the physical condition that rarely exists on the earth to carry out in an efficient manner. As per the growing demand and present scenario of the world energy, scientists are working round the clock to make effective fusion reactions to real. In this paper the work presently going on is considered in this regard. The progress of the Joint European Torus 2010, ITER 2005, HiPER and minor works have been studied to make the paper more object oriented. A detailed study of the technological and material requirement has been discussed in the paper and a possible suggestion is provided to make a contribution in the field of building first ever nuclear fusion reactor

  15. Fusion energy option

    International Nuclear Information System (INIS)

    Schmid, L.C.

    The potential of fusion energy contributing to the energy needs is discussed. Controlled thermonuclear reactions hold the promise of an abundant source of fuel used to produce electrical energy for the future in an environmentally acceptable way. Once feasibility questions are answered and engineering problems are resolved, it should be possible to produce energy in a form that can use current methods of electrical generation to convert it into a useful form. If the fusion system is operated with only deuterium as fuel, the deuterium available from a pail of water would produce energy equivalent to that produced by 600 gallons of gasoline. The water in the ocean could provide energy for billions of years at the current rate of consumption. Experimental results are currently confirming the theoretical predictions and the schedule for fusion development is shown to be completion of feasibility experiments in 1974; physics test reactors appearing in 1983; experimental power reactors being built in 1990; and the demonstration plant on-line in 1994. The process of producing fusion power, fusion research needs, and problems to be solved are reviewed

  16. Fusion, magnetic confinement

    Energy Technology Data Exchange (ETDEWEB)

    Berk, H.L.

    1992-08-06

    An overview is presented of the principles of magnetic confinement of plasmas for the purpose of achieving controlled fusion conditions. Sec. 1 discusses the different nuclear fusion reactions which can be exploited in prospective fusion reactors and explains why special technologies need to be developed for the supply of tritium or {sup 3}He, the probable fuels. In Sec. 2 the Lawson condition, a criterion that is a measure of the quality of confinement relative to achieving fusion conditions, is explained. In Sec. 3 fluid equations are used to describe plasma confinement. Specific confinement configurations are considered. In Sec. 4 the orbits of particle sin magneti and electric fields are discussed. In Sec. 5 stability considerations are discussed. It is noted that confinement systems usually need to satisfy stability constraints imposed by ideal magnetohydrodynamic (MHD) theory. The paper culminates with a summary of experimental progress in magnetic confinement. Present experiments in tokamaks have reached the point that the conditions necessary to achieve fusion are being satisfied.

  17. Reconstitution of high affinity α2 adrenergic agonist binding by fusion with a pertussis toxin substrate

    International Nuclear Information System (INIS)

    Kim, M.H.; Neubig, R.R.

    1986-01-01

    High affinity α 2 adrenergic agonist binding is thought to occur via a coupling of the α 2 receptor with N/sub i/, the inhibitory guanyl nucleotide binding protein. Human platelet membranes pretreated at pH 11.5 exhibit a selective inactivation of agonist binding and N/sub i/. To further study the mechanism of agonist binding, alkali treated membranes (ATM) were mixed with membranes pretreated with 10 μM phenoxybenzamine to block α 2 receptors (POB-M). The combined membrane pellet was incubated in 50% polyethylene glycol (PEG) to promote membrane-membrane fusion and assayed for binding to the α 2 agonist [ 3 H]UK 14,304 (UK) and the antagonist [ 3 H] yohimbine. PEG treatment resulted in a 2-4 fold enhancement of UK binding whereas yohimbine binding was unchanged. No enhancement of UK binding was observed in the absence of PEG treatment. The reconstitution was dependent on the addition of POB-M. They found that a 1:1 ratio of POB-M:ATM was optimal. Reconstituted binding was inhibited by GppNHp. Fusion of rat C6 glioma cell membranes, which do not contain α 2 receptors, also enhanced agonist binding to ATM. Fusion of C6 membranes from cells treated with pertussis toxin did not enhance [ 3 H] UK binding. These data show that a pertussis toxin sensitive membrane component, possibly N/sub i/, can reconstitute high affinity α 2 agonist binding

  18. Structural and Functional Studies on the Fusion and Attachment Envelope Glycoproteins of Nipah Virus and Hendra Virus

    Science.gov (United States)

    2003-01-01

    subcloned into the vaccinia virus promoter driven expression vector pMCO2 (90) and recombinant vaccinia viruses were generated using standard...characteristics and properties of these novel viruses , and may provide insights into membrane fusion mechanisms, the virus infection process, and towards...these novel viruses , and may provide insights into membrane fusion mechanisms, the virus infection process, and towards the development of therapeutics

  19. Kar5p is required for multiple functions in both inner and outer nuclear envelope fusion in Saccharomyces cerevisiae.

    Science.gov (United States)

    Rogers, Jason V; Rose, Mark D

    2014-12-02

    During mating in the budding yeast Saccharomyces cerevisiae, two haploid nuclei fuse via two sequential membrane fusion steps. SNAREs (i.e., soluble N-ethylmaleimide-sensitive factor attachment protein receptors) and Prm3p mediate outer nuclear membrane fusion, but the inner membrane fusogen remains unknown. Kar5p is a highly conserved transmembrane protein that localizes adjacent to the spindle pole body (SPB), mediates nuclear envelope fusion, and recruits Prm3p adjacent to the SPB. To separate Kar5p's functions, we tested localization, Prm3p recruitment, and nuclear fusion efficiency in various kar5 mutants. All domains and the conserved cysteine residues were essential for nuclear fusion. Several kar5 mutant proteins localized properly but did not mediate Prm3p recruitment; other kar5 mutant proteins localized and recruited Prm3p but were nevertheless defective for nuclear fusion, demonstrating additional functions beyond Prm3p recruitment. We identified one Kar5p domain required for SPB localization, which is dependent on the half-bridge protein Mps3p. Electron microscopy revealed a kar5 mutant that arrests with expanded nuclear envelope bridges, suggesting that Kar5p is required after outer nuclear envelope fusion. Finally, a split-GFP assay demonstrated that Kar5p localizes to both the inner and outer nuclear envelope. These insights suggest a mechanism by which Kar5p mediates inner nuclear membrane fusion. Copyright © 2015 Rogers and Rose.

  20. Low-pH-dependent fusion of sindbis virus with receptor-free cholesterol- and sphingolipid-containing liposomes

    NARCIS (Netherlands)

    Smit, JM; Bittman, R; Wilschut, J

    1999-01-01

    There is controversy as to whether the cell entry mechanism of Sindbis virus (SIN) involves direct fusion of the viral envelope with the plasma membrane at neutral pH Dr uptake by receptor-mediated endocytosis and subsequent low-pH-induced fusion from within acidic endosomes. Here, we studied the

  1. Unraveling a three-step spatiotemporal mechanism of triggering of receptor-induced Nipah virus fusion and cell entry.

    Directory of Open Access Journals (Sweden)

    Qian Liu

    Full Text Available Membrane fusion is essential for entry of the biomedically-important paramyxoviruses into their host cells (viral-cell fusion, and for syncytia formation (cell-cell fusion, often induced by paramyxoviral infections [e.g. those of the deadly Nipah virus (NiV]. For most paramyxoviruses, membrane fusion requires two viral glycoproteins. Upon receptor binding, the attachment glycoprotein (HN/H/G triggers the fusion glycoprotein (F to undergo conformational changes that merge viral and/or cell membranes. However, a significant knowledge gap remains on how HN/H/G couples cell receptor binding to F-triggering. Via interdisciplinary approaches we report the first comprehensive mechanism of NiV membrane fusion triggering, involving three spatiotemporally sequential cell receptor-induced conformational steps in NiV-G: two in the head and one in the stalk. Interestingly, a headless NiV-G mutant was able to trigger NiV-F, and the two head conformational steps were required for the exposure of the stalk domain. Moreover, the headless NiV-G prematurely triggered NiV-F on virions, indicating that the NiV-G head prevents premature triggering of NiV-F on virions by concealing a F-triggering stalk domain until the correct time and place: receptor-binding. Based on these and recent paramyxovirus findings, we present a comprehensive and fundamentally conserved mechanistic model of paramyxovirus membrane fusion triggering and cell entry.

  2. Prospects of fusion energy system

    International Nuclear Information System (INIS)

    Kohzaki, Yasuji; Seki, Yasushi; Motojima, Osamu

    1993-01-01

    Nuclear fusion energy that collects large expectation as the energy system of 21st century adopts the tokamak with DT fuel as the main line to advance the research and development, and succeeded in the confinement of plasma that nearly satisfies the condition of zero power output. However, as for nuclear fusion energy, other various generation and utilization forms are conceivable. At present, there are many subjects before the practical use, but as to nuclear fusion energy system which is considered to contribute greatly to mankind when it will be practically used in future, it is significant to clarify the present state of the research and the subjects of the research for the realization. Tokamak type fusion reactor, helical type fusion reactor, D-He-3 FRC fusion reactor, inertial fusion reactor,fusion-fission hybrid reactor, nuclear fusion rocket, muon catalytic nuclear fusion, normal temperature nuclear fusion and so on are described. As the final summary, on the basis of the concepts of individual nuclear fusion reactors, what possibility nuclear fusion energy has as a whole is considered, and the way of advancing the development hereafter is summarized. (K.I.)

  3. Heavy ion inertial fusion

    International Nuclear Information System (INIS)

    Keefe, D.; Sessler, A.M.

    1980-01-01

    Inertial fusion has not yet been as well explored as magnetic fusion but can offer certain advantages as an alternative source of electric energy for the future. Present experiments use high-power beams from lasers and light-ion diodes to compress the deuterium-tritium (D-T) pellets but these will probably be unsuitable for a power plant. A more promising method is to use intense heavy-ion beams from accelerator systems similar to those used for nuclear and high-energy physics; the present paper addresses itself to this alternative. As will be demonstrated the very high beam power needed poses new design questions, from the ion-source through the accelerating system, the beam transport system, to the final focus. These problems will require extensive study, both theoretically and experimentally, over the next several years before an optimum design for an inertial fusion driver can be arrived at. (Auth.)

  4. Fusion reactor wastes

    International Nuclear Information System (INIS)

    Young, J.R.

    1976-01-01

    The fusion reactor currently is being developed as a clean source of electricity with an essentially infinite source of fuel. These reactors are visualized as using a fusion reaction to generate large quantities of high temperature energy which can be used as process heat or for the generation of electricity. The energy would be created primarily as the kinetic energy of neutrons or other reaction products. Neutron energy could be converted to high-temperature heat by moderation and capture of the neutrons. The energy of other reaction products could be converted to high-temperature heat by capture, or directly to electricity by direct conversion electrostatic equipment. An analysis to determine the wastes released as a result of operation of fusion power plants is presented

  5. Canadian fusion program

    International Nuclear Information System (INIS)

    Brown, T.S.

    1982-06-01

    The National Research Council of Canada is establishing a coordinated national program of fusion research and development that is planned to grow to a total annual operating level of about $20 million in 1985. The long-term objective of the program is to put Canadian industry in a position to manufacture sub-systems and components of fusion power reactors. In the near term the program is designed to establish a minimum base of scientific and technical expertise sufficient to make recognized contributions and thereby gain access to the international effort. The Canadian program must be narrowly focussed on a few specializations where Canada has special indigenous skills or technologies. The programs being funded are the Tokamak de Varennes, the Fusion Fuels Technology Project centered on tritium management, and high-power gas laser technology and associated diagnostic instrumentation

  6. Fusion research at ORNL

    Energy Technology Data Exchange (ETDEWEB)

    1982-03-01

    The ORNL Fusion Program includes the experimental and theoretical study of two different classes of magnetic confinement schemes - systems with helical magnetic fields, such as the tokamak and stellarator, and the ELMO Bumpy Torus (EBT) class of toroidally linked mirror systems; the development of technologies, including superconducting magnets, neutral atomic beam and radio frequency (rf) heating systems, fueling systems, materials, and diagnostics; the development of databases for atomic physics and radiation effects; the assessment of the environmental impact of magnetic fusion; and the design of advanced demonstration fusion devices. The program involves wide collaboration, both within ORNL and with other institutions. The elements of this program are shown. This document illustrates the program's scope; and aims by reviewing recent progress.

  7. On impact fusion

    International Nuclear Information System (INIS)

    Winterberg, F.

    1997-01-01

    Impact fusion is a promising, but much less developed road towards inertial confinement fusion. It offers an excellent solution to the so-called stand-off problem for thermonuclear microexplosions but is confronted with the challenge to accelerate macroscopic particles to the needed high velocities of 10 2 -10 3 km/s. To reach these velocities, two ways have been studied in the past. The electric acceleration of a beam of microparticles, with the particles as small as large clusters, and the magnetic acceleration of gram-size ferromagnetic or superconducting projectiles. For the generation of an intense burst of soft X-rays used for the indirect drive, impact fusion may offer new promising possibilities

  8. Ceramics for fusion devices

    International Nuclear Information System (INIS)

    Clinard, F.W. Jr.

    1984-01-01

    Ceramics are required for a number of applications in fusion devices, among the most critical of which are magnetic coil insulators, windows for RF heating systems, and structural uses. Radiation effects dominate consideration of candidate materials, although good pre-irradiation properties are a requisite. Materials and components can be optimized by careful control of chemical and microstructural content, and application of brittle material design and testing techniques. Future directions for research and development should include further extension of the data base in the areas of electrical, structural, and thermal properties; establishment of a fission neutron/fusion neutron correlation including transmutation gas effects; and development of new materials tailored to meet the specific needs of fusion reactors

  9. Mirror Fusion Test Facility

    International Nuclear Information System (INIS)

    Dixon, F.P.

    1977-01-01

    On October 1, 1977 work began at LLL on the Mirror Fusion Test Facility (MFTF), an advanced experimental fusion device. Scheduled for operation in late 1981, MFTF is designed as an intermediate step between present mirror machines, such as 2XIIB, and an experimental fusion reactor. This design incorporates improved technology and a better theoretical understanding of how neutral beam injection, plasma guns, and gas injection into the plasma region compensate for cooling and particle losses. With the new facility, we expect to achieve a confinement factor (n tau) of 10 12 particles . sm/cm 3 --a tenfold increase over 2XIIB n tau values--and to increase plasma temperature to over 500 million K. The following article describes this new facility and reports on progress in some of the R and D projects that are providing the technological base for its construction

  10. Peaceful Uses of Fusion

    Science.gov (United States)

    Teller, E.

    1958-07-03

    Applications of thermonuclear energy for peaceful and constructive purposes are surveyed. Developments and problems in the release and control of fusion energy are reviewed. It is pointed out that the future of thermonuclear power reactors will depend upon the construction of a machine that produces more electric energy than it consumes. The fuel for thermonuclear reactors is cheap and practically inexhaustible. Thermonuclear reactors produce less dangerous radioactive materials than fission reactors and, when once brought under control, are not as likely to be subject to dangerous excursions. The interaction of the hot plasma with magnetic fields opens the way for the direct production of electricity. It is possible that explosive fusion energy released underground may be harnessed for the production of electricity before the same feat is accomplished in controlled fusion processes. Applications of underground detonations of fission devices in mining and for the enhancement of oil flow in large low-specific-yield formations are also suggested.

  11. Ceramics for fusion applications

    International Nuclear Information System (INIS)

    Clinard, F.W. Jr.

    1987-01-01

    Ceramics are required for a variety of uses in both near-term fusion devices and in commercial powerplants. These materials must retain adequate structural and electrical properties under conditions of neutron, particle and ionizing irradiation; thermal and applied stresses; and physical and chemical sputtering. Ceramics such as Al 2 O 3 , MgAl 2 O 4 , BeO, Si 3 N 4 and SiC are currently under study for fusion applications, and results to date show widely-varying responses to the fusion environment. Materials can be identified today that will meet initial operating requirements, but improvements in physical properties are needed to achieve satisfactory lifetimes for critical applications. (author)

  12. Fusion Reactor Materials

    Energy Technology Data Exchange (ETDEWEB)

    Decreton, M

    2002-04-01

    The objective of SCK-CEN's programme on fusion reactor materials is to contribute to the knowledge on the radiation-induced behaviour of fusion reactor materials and components as well as to help the international community in building the scientific and technical basis needed for the construction of the future reactor. Ongoing projects include: the study of the mechanical and chemical (corrosion) behaviour of structural materials under neutron irradiation and water coolant environment; the investigation of the characteristics of irradiated first wall material such as beryllium; investigations on the management of materials resulting from the dismantling of fusion reactors including waste disposal. Progress and achievements in these areas in 2001 are discussed.

  13. Ceramics for fusion applications

    International Nuclear Information System (INIS)

    Clinard, F.W. Jr.

    1986-01-01

    Ceramics are required for a variety of uses in both near-term fusion devices and in commercial powerplants. These materials must retain adequate structural and electrical properties under conditions of neutron, particle, and ionizing irradiation; thermal and applied stresses; and physical and chemical sputtering. Ceramics such as Al 2 O 3 , MgAl 2 O 4 , BeO, Si 3 N 4 and SiC are currently under study for fusion applications, and results to date show widely-varying response to the fusion environment. Materials can be identified today which will meet initial operating requirements, but improvements in physical properties are needed to achieve satisfactory lifetimes for critical applications

  14. Intense fusion neutron sources

    Science.gov (United States)

    Kuteev, B. V.; Goncharov, P. R.; Sergeev, V. Yu.; Khripunov, V. I.

    2010-04-01

    The review describes physical principles underlying efficient production of free neutrons, up-to-date possibilities and prospects of creating fission and fusion neutron sources with intensities of 1015-1021 neutrons/s, and schemes of production and application of neutrons in fusion-fission hybrid systems. The physical processes and parameters of high-temperature plasmas are considered at which optimal conditions for producing the largest number of fusion neutrons in systems with magnetic and inertial plasma confinement are achieved. The proposed plasma methods for neutron production are compared with other methods based on fusion reactions in nonplasma media, fission reactions, spallation, and muon catalysis. At present, intense neutron fluxes are mainly used in nanotechnology, biotechnology, material science, and military and fundamental research. In the near future (10-20 years), it will be possible to apply high-power neutron sources in fusion-fission hybrid systems for producing hydrogen, electric power, and technological heat, as well as for manufacturing synthetic nuclear fuel and closing the nuclear fuel cycle. Neutron sources with intensities approaching 1020 neutrons/s may radically change the structure of power industry and considerably influence the fundamental and applied science and innovation technologies. Along with utilizing the energy produced in fusion reactions, the achievement of such high neutron intensities may stimulate wide application of subcritical fast nuclear reactors controlled by neutron sources. Superpower neutron sources will allow one to solve many problems of neutron diagnostics, monitor nano-and biological objects, and carry out radiation testing and modification of volumetric properties of materials at the industrial level. Such sources will considerably (up to 100 times) improve the accuracy of neutron physics experiments and will provide a better understanding of the structure of matter, including that of the neutron itself.

  15. Insulators for fusion applications

    International Nuclear Information System (INIS)

    1987-04-01

    Design studies for fusion devices and reactors have become more detailed in recent years and with this has come a better understanding of requirements and operating conditions for insulators in these machines. Ceramic and organic insulators are widely used for many components of fusion devices and reactors namely: radio frequency (RF) energy injection systems (BeO, Al 2 O 3 , Mg Al 2 O 4 , Si 3 N 4 ); electrical insulation for the torus structure (SiC, Al 2 O 3 , MgO, Mg Al 2 O 4 , Si 4 Al 2 O 2 N 6 , Si 3 N 4 , Y 2 O 3 ); lightly-shielded magnetic coils (MgO, MgAl 2 O 4 ); the toroidal field coil (epoxies, polyimides), neutron shield (B 4 C, TiH 2 ); high efficiency electrical generation; as well as the generation of very high temperatures for high efficiency hydrogen production processes (ZrO 2 and Al 2 O 3 - mat, graphite and carbon - felt). Timely development of insulators for fusion applications is clearly necessary. Those materials to be used in fusion machines should show high resistance to radiation damage and maintain their structural integrity. Now the need is urgent for a variety of radiation resistant materials, but much effort in these areas is required for insulators to be considered seriously by the design community. This document contains 14 papers from an IAEA meeting. It was the objective of this meeting to identify existing problems in analysing various situations of applications and requirements of electrical insulators and ceramics in fusion and to recommend strategies and different stages of implementation. This meeting was endorsed by the International Fusion Research Council

  16. International fusion research

    International Nuclear Information System (INIS)

    Pease, R.S.

    1983-01-01

    Nuclear energy of the light elements deuterium and lithium can be released if the 100 MK degree temperature required for deuterium-tritium thermonuclear fusion reactions can be achieved together with sufficient thermal insulation for a net energy yield. Progress of world-wide research shows good prospect for these physical conditions being achieved by the use of magnetic field confinement and of rapidly developing heating methods. Tokamak systems, alternative magnetic systems and inertial confinement progress are described. International co-operation features a number of bilateral agreements between countries: the Euratom collaboration which includes the Joint European Torus, a joint undertaking of eleven Western European nations of Euratom, established to build and operate a major confinement experiment; the development of co-operative projects within the OECD/IEA framework; the INTOR workshop, a world-wide study under IAEA auspices of the next major step in fusion research which might be built co-operatively; and assessments of the potential of nuclear fusion by the IAEA and the International Fusion Research Council. The INTOR (International Tokamak Reactor) studies have outlined a major plant of the tokamak type to study the engineering and technology of fusion reactor systems, which might be constructed on a world-wide basis to tackle and share the investment risks of the developments which lie ahead. This paper summarizes the recent progress of research on controlled nuclear fusion, featuring those areas where international co-operation has played an important part, and describes the various arrangements by which this international co-operation is facilitated. (author)

  17. Neutrons and fusion

    International Nuclear Information System (INIS)

    Maynard, C.W.

    1976-01-01

    The production of energy from fusion reactions does not require neutrons in the fundamental sense that they are required in a fission reactor. Nevertheless, the dominant fusion reaction, that between deuterium and tritium, yields a 14 MeV neutron. To contrast a fusion reactor based on this reaction with the fission case, 3 x 10 20 such neutrons produced per gigawatt of power. This is four times as many neutrons as in an equivalent fission reactor and they carry seven times the energy of the fission neutrons. Thus, they dominate the energy recovery problem and create technological problems comparable to the original plasma confinement problem as far as a practical power producing device is concerned. Further contrasts of the fusion and fission cases are presented to establish the general role of neutrons in fusion devices. Details of the energy deposition processes are discussed and those reactions necessary for producing additional tritium are outlined. The relatively high energy flux with its large intensity will activate almost any materials of which the reactor may be composed. This activation is examined from the point of view of decay heat, radiological safety, and long-term storage. In addition, a discussion of the deleterious effects of neutron interactions on materials is given in some detail; this includes the helium and hydrogen producing reactions and displacement rate of the lattice atoms. The various materials that have been proposed for structural purposes, for breeding, reflecting, and moderating neutrons, and for radiation shielding are reviewed from the nuclear standpoint. The specific reactions of interest are taken up for various materials and finally a report is given on the status and prospects of data for fusion studies

  18. Regulation of Exocytotic Fusion Pores by SNARE Protein Transmembrane Domains

    Directory of Open Access Journals (Sweden)

    Zhenyong Wu

    2017-10-01

    Full Text Available Calcium-triggered exocytotic release of neurotransmitters and hormones from neurons and neuroendocrine cells underlies neuronal communication, motor activity and endocrine functions. The core of the neuronal exocytotic machinery is composed of soluble N-ethyl maleimide sensitive factor attachment protein receptors (SNAREs. Formation of complexes between vesicle-attached v- and plasma-membrane anchored t-SNAREs in a highly regulated fashion brings the membranes into close apposition. Small, soluble proteins called Complexins (Cpx and calcium-sensing Synaptotagmins cooperate to block fusion at low resting calcium concentrations, but trigger release upon calcium increase. A growing body of evidence suggests that the transmembrane domains (TMDs of SNARE proteins play important roles in regulating the processes of fusion and release, but the mechanisms involved are only starting to be uncovered. Here we review recent evidence that SNARE TMDs exert influence by regulating the dynamics of the fusion pore, the initial aqueous connection between the vesicular lumen and the extracellular space. Even after the fusion pore is established, hormone release by neuroendocrine cells is tightly controlled, and the same may be true of neurotransmitter release by neurons. The dynamics of the fusion pore can regulate the kinetics of cargo release and the net amount released, and can determine the mode of vesicle recycling. Manipulations of SNARE TMDs were found to affect fusion pore properties profoundly, both during exocytosis and in biochemical reconstitutions. To explain these effects, TMD flexibility, and interactions among TMDs or between TMDs and lipids have been invoked. Exocytosis has provided the best setting in which to unravel the underlying mechanisms, being unique among membrane fusion reactions in that single fusion pores can be probed using high-resolution methods. An important role will likely be played by methods that can probe single fusion pores

  19. Confinement inertial fusion. Power reactors of nuclear fusion by lasers

    International Nuclear Information System (INIS)

    Velarde, G.; Ahnert, C.; Aragones, J.M.; Leira, G; Martinez-Val, J.M.

    1980-01-01

    The energy crisis and the need of the nuclear fusion energy are analized. The nuclear processes in the laser interation with the ablator material are studied, as well as the thermohydrodinamic processes in the implossion, and the neutronics of the fusion. The fusion reactor components are described and the economic and social impact of its introduction in the future energetic strategies.(author)

  20. Nuclear fusion: Pursuing the Soft [Symposium on fusion technology] option

    International Nuclear Information System (INIS)

    Kenward, M.

    1991-01-01

    Fusion research has come a long way since the fusion community held the first Symposium on fusion technology (Soft) in Britain 30 years ago. Some of the recent achievements of the Jet project are reported from this year's symposium, the 16th in the series, held in London at the beginning of September. (author)

  1. Fusion Propulsion Study

    Science.gov (United States)

    1989-07-01

    surround ICF rocket concept. Calculated results of a typical propellant surround concept are shown in Figures 16 and 17 With this system, it appears...However, the time available to burn the fusion fuel to the specified burnup is also reduced. This is a minor problem with D-T fuel, but, as will be seen...TABLE 19 TCT FOR INPUT LIST NO. NAME DESCRIPTIQN 1 THRUST ROCKET THRUST, NEWTONS 2 ISP SPECIFIC IMPULSE, SECONDS 3 RATE FUSION REP RATE, HZ 4 BURNUP

  2. Cold fusion in perspective

    International Nuclear Information System (INIS)

    Sanford, L.

    1989-01-01

    Since early April a great deal of excitement has been created over the Fleischmann/Pons cold fusion experiment, which if it performs as advertised, could turn out to be mankind's best hope of heading off the energy crisis scheduled for early in the next century. Dozens of groups around the world are now attempting to duplicate the experiment to see if Fleischmann and Pons' discovery is an experimental mistake, an unknown electrochemical effect or a new kind of fusion reaction. This article puts the experiment into the perspective of today and looks at how it might affect the energy scene tomorrow if it should turn out to be commercially exploitable. (author)

  3. Vacuum fusion of uranium

    International Nuclear Information System (INIS)

    Stohr, J.A.

    1957-01-01

    After having outlined that vacuum fusion and moulding of uranium and of its alloys have some technical and economic benefits (vacuum operations avoid uranium oxidation and result in some purification; precision moulding avoids machining, chip production and chemical reprocessing of these chips; direct production of the desired shape is possible by precision moulding), this report presents the uranium fusion unit (its low pressure enclosure and pumping device, the crucible-mould assembly, and the MF supply device). The author describes the different steps of cast production, and briefly comments the obtained results

  4. Fusion technology programme

    International Nuclear Information System (INIS)

    Finken, D.

    1985-05-01

    In the current Fusion Technology Programme of the European Community the KfK association is working at present on 16 R and D contracts. Most of the work is strongly oriented towards the Next European Torus. Direct support to NET is given by three KfK delegates being member of the NET study group. In addition to the R and D contracts the association is working on 11 NET study contracts. Though KfK contributes to all areas defined in fusion technology, the main emphasis is put on superconducting magnet and breeding blanket development. Other important fields are tritium technology, materials research, and remote handling. (orig./GG)

  5. Atomic data for fusion

    International Nuclear Information System (INIS)

    Hunter, H.T.; Kirkpatrick, M.I.; Alvarez, I.; Cisneros, C.; Phaneuf, R.A.; Barnett, C.F.

    1990-07-01

    This report provides a handbook of recommended cross-section and rate-coefficient data for inelastic collisions between hydrogen, helium and lithium atoms, molecules and ions, and encompasses more than 400 different reactions of primary interest in fusion research. Published experimental and theoretical data have been collected and evaluated, and the recommended data are presented in tabular, graphical and parametrized form. Processes include excitation and spectral line emission, charge exchange, ionization, stripping, dissociation and particle interchange reactions. The range of collision energies is appropriate to applications in fusion-energy research

  6. Fusion Reactor Materials

    International Nuclear Information System (INIS)

    Decreton, M.

    2000-01-01

    SCK-CEN's research and development programme on fusion reactor materials includes: (1) the study of the mechanical behaviour of structural materials under neutron irradiation (including steels, inconel, molybdenum, chromium); (2) the determination and modelling of the characteristics of irradiated first wall materials such as beryllium; (3) the detection of abrupt electrical degradation of insulating ceramics under high temperature and neutron irradiation; (4) the study of the dismantling and waste disposal strategy for fusion reactors.; (5) a feasibility study for the testing of blanket modules under neutron radiation. Main achievements in these topical areas in the year 1999 are summarised

  7. Fusion Reactor Materials

    Energy Technology Data Exchange (ETDEWEB)

    Decreton, M

    2000-07-01

    SCK-CEN's research and development programme on fusion reactor materials includes: (1) the study of the mechanical behaviour of structural materials under neutron irradiation (including steels, inconel, molybdenum, chromium); (2) the determination and modelling of the characteristics of irradiated first wall materials such as beryllium; (3) the detection of abrupt electrical degradation of insulating ceramics under high temperature and neutron irradiation; (4) the study of the dismantling and waste disposal strategy for fusion reactors.; (5) a feasibility study for the testing of blanket modules under neutron radiation. Main achievements in these topical areas in the year 1999 are summarised.

  8. Atomic data for fusion

    Energy Technology Data Exchange (ETDEWEB)

    Hunter, H.T.; Kirkpatrick, M.I.; Alvarez, I.; Cisneros, C.; Phaneuf, R.A. (eds.); Barnett, C.F.

    1990-07-01

    This report provides a handbook of recommended cross-section and rate-coefficient data for inelastic collisions between hydrogen, helium and lithium atoms, molecules and ions, and encompasses more than 400 different reactions of primary interest in fusion research. Published experimental and theoretical data have been collected and evaluated, and the recommended data are presented in tabular, graphical and parametrized form. Processes include excitation and spectral line emission, charge exchange, ionization, stripping, dissociation and particle interchange reactions. The range of collision energies is appropriate to applications in fusion-energy research.

  9. Bringing fusion electric power closer

    International Nuclear Information System (INIS)

    Kintner, E.

    1977-01-01

    A review of the controlled fusion research program is given. The tokamak research program is described. Beam injection heating, control systems, and the safety of fusion reactors are topics that are also discussed

  10. Fusion in the energy system

    DEFF Research Database (Denmark)

    Fusion energy is the fundamental energy source of the Universe, as the energy of the Sun and the stars are produced by fusion of e.g. hydrogen to helium. Fusion energy research is a strongly international endeavor aiming at realizing fusion energy production in power plants on Earth. Reaching...... this goal, mankind will have a sustainable base load energy source with abundant resources, having no CO2 release, and with no longlived radioactive waste. This presentation will describe the basics of fusion energy production and the status and future prospects of the research. Considerations...... of integration into the future electricity system and socio-economic studies of fusion energy will be presented, referring to the programme of Socio-Economic Research on Fusion (SERF) under the European Fusion Energy Agreement (EFDA)....

  11. Atomic physics issues in fusion

    International Nuclear Information System (INIS)

    Post, D.E.

    1982-01-01

    Atomic physics issues have played a large role in controlled fusion research. A general introduction to the present role of atomic processes in both inertial and magnetic controlled fusion work is presented. (Auth.)

  12. The quest for fusion energy

    International Nuclear Information System (INIS)

    Johnson, J.L.

    1997-10-01

    A brief history of the magnetic fusion program from the point of view of a stellarator enthusiast who worked at a major tokamak laboratory. The reason that success in the magnetic fusion energy program is essential is presented. (author)

  13. Accelerator and fusion research division

    International Nuclear Information System (INIS)

    1992-12-01

    This report contains brief discussions on research topics in the following area: Heavy-Ion Fusion Accelerator Research; Magnetic Fusion Energy; Advanced Light Source; Center for Beam Physics; Superconducting Magnets; and Bevalac Operations

  14. Fusion technology status and requirements

    International Nuclear Information System (INIS)

    Thomassen, K.I.

    1982-01-01

    This paper summarizes the status of fusion technology and discusses the requirements to be met in order to build a demonstration fusion plant. Strategies and programmatic considerations in pursuing engineering feasibility are also outlined

  15. Polyclonal and monoclonal antibodies specific for the six-helix bundle of the human respiratory syncytial virus fusion glycoprotein as probes of the protein post-fusion conformation

    International Nuclear Information System (INIS)

    Palomo, Concepción; Mas, Vicente; Vázquez, Mónica; Cano, Olga; Luque, Daniel; Terrón, María C.; Calder, Lesley J.; Melero, José A.

    2014-01-01

    Human respiratory syncytial virus (hRSV) has two major surface glycoproteins (G and F) anchored in the lipid envelope. Membrane fusion promoted by hRSV F occurs via refolding from a pre-fusion form to a highly stable post-fusion state involving large conformational changes of the F trimer. One of these changes results in assembly of two heptad repeat sequences (HRA and HRB) into a six-helix bundle (6HB) motif. To assist in distinguishing pre- and post-fusion conformations of hRSV F , we have prepared polyclonal (α-6HB) and monoclonal (R145) rabbit antibodies specific for the 6HB. Among other applications, these antibodies were used to explore the requirements of 6HB formation by isolated protein segments or peptides and by truncated mutants of the F protein. Site-directed mutagenesis and electron microscopy located the R145 epitope in the post-fusion hRSV F at a site distantly located from previously mapped epitopes, extending the repertoire of antibodies that can decorate the F molecule. - Highlights: • Antibodies specific for post-fusion respiratory syncytial virus fusion protein are described. • Polyclonal antibodies were obtained in rabbit inoculated with chimeric heptad repeats. • Antibody binding required assembly of a six-helix bundle in the post-fusion protein. • A monoclonal antibody with similar structural requirements is also described. • Binding of this antibody to the post-fusion protein was visualized by electron microscopy

  16. Polyclonal and monoclonal antibodies specific for the six-helix bundle of the human respiratory syncytial virus fusion glycoprotein as probes of the protein post-fusion conformation

    Energy Technology Data Exchange (ETDEWEB)

    Palomo, Concepción; Mas, Vicente; Vázquez, Mónica; Cano, Olga [Unidad de Biología Viral, Centro Nacional de Microbiología, Madrid (Spain); CIBER de Enfermedades Respiratorias, Instituto de Salud Carlos III, Majadahonda, 28220 Madrid (Spain); Luque, Daniel; Terrón, María C. [Unidad de Microscopía Electrónica y Confocal, Centro Nacional de Microbiología, Instituto de Salud Carlos III, Majadahonda, 28220 Madrid (Spain); Calder, Lesley J. [National Institute for Medical Research, MRC, Mill Hill, London NW7 1AA (United Kingdom); Melero, José A., E-mail: jmelero@isciii.es [Unidad de Biología Viral, Centro Nacional de Microbiología, Madrid (Spain); CIBER de Enfermedades Respiratorias, Instituto de Salud Carlos III, Majadahonda, 28220 Madrid (Spain)

    2014-07-15

    Human respiratory syncytial virus (hRSV) has two major surface glycoproteins (G and F) anchored in the lipid envelope. Membrane fusion promoted by hRSV{sub F} occurs via refolding from a pre-fusion form to a highly stable post-fusion state involving large conformational changes of the F trimer. One of these changes results in assembly of two heptad repeat sequences (HRA and HRB) into a six-helix bundle (6HB) motif. To assist in distinguishing pre- and post-fusion conformations of hRSV{sub F}, we have prepared polyclonal (α-6HB) and monoclonal (R145) rabbit antibodies specific for the 6HB. Among other applications, these antibodies were used to explore the requirements of 6HB formation by isolated protein segments or peptides and by truncated mutants of the F protein. Site-directed mutagenesis and electron microscopy located the R145 epitope in the post-fusion hRSV{sub F} at a site distantly located from previously mapped epitopes, extending the repertoire of antibodies that can decorate the F molecule. - Highlights: • Antibodies specific for post-fusion respiratory syncytial virus fusion protein are described. • Polyclonal antibodies were obtained in rabbit inoculated with chimeric heptad repeats. • Antibody binding required assembly of a six-helix bundle in the post-fusion protein. • A monoclonal antibody with similar structural requirements is also described. • Binding of this antibody to the post-fusion protein was visualized by electron microscopy.

  17. Robotic membranes

    DEFF Research Database (Denmark)

    Ramsgaard Thomsen, Mette

    2008-01-01

    , Vivisection and Strange Metabolisms, were developed at the Centre for Information Technology and Architecture (CITA) at the Royal Danish Academy of Fine Arts in Copenhagen as a means of engaging intangible digital data with tactile physical material. As robotic membranes, they are a dual examination...

  18. Amphipathic properties of HIV-1 gp41 fusion inhibitors.

    Science.gov (United States)

    Gochin, Miriam; Zhou, Guangyan

    2011-12-01

    Small molecule inhibition of HIV fusion has been an elusive goal, despite years of effort by both pharmaceutical and academic laboratories. In this review, we will discuss the amphipathic properties of both peptide and small molecule inhibitors of gp41-mediated fusion. Many of the peptides and small molecules that have been developed target a large hydrophobic pocket situated within the grooves of the coiled coil, a potential hotspot for inhibiting the trimer of hairpin formation that accompanies fusion. Peptide studies reveal molecular properties required for effective inhibition, including elongated structure and lipophilic or amphiphilic nature. The characteristics of peptides that bind in this pocket provide features that should be considered in small molecule development. Additionally, a novel site for small molecule inhibition of fusion has recently been suggested, involving residues of the loop and fusion peptide. We will review the small molecule structures that have been developed, evidence pointing to their mechanism of action and strategies towards improving their affinity. The data points to the need for a strongly amphiphilic character of the inhibitors, possibly as a means to mediate the membrane - protein interaction that occurs in gp41 in addition to the protein - protein interaction that accompanies the fusion-activating conformational transition.

  19. Studies on virus-induced cell fusion. Progress report, August 1, 1977--June 30, 1978

    Energy Technology Data Exchange (ETDEWEB)

    Person, S.

    1978-07-01

    We have previously postulated that wild-type Herpes Simplex Virus type I (HSV-1) infections are characterized by the presence of a fusion factor and a fusion inhibitor activity. The fusion inhibitor presumably is dominant so that a small fraction of cells fuse in a typical wild-type infection. Furthermore, the syn mutants isolated in our laboratory are thought to cause extensive cell fusion because the production of active fusion inhibitor in cell membranes is delayed. If mutations existed that altered both the fusion factor and fusion inhibitor activity then separate viruses containing these two mutations might be able to complement each other, each supplying the defective gene product missing in the other virus. This would produce a wild type and not a syncytial mutant response. Complementation tests between two viruses, tsB5 and syn 20, which are thought to contain defects in the production of active fusion factor and fusion inhibitor activity, respectively, were done. A wild-type response was observed indicating that the mutations affecting fusion were in two separate genes.

  20. Fusion reactor development: A review

    International Nuclear Information System (INIS)

    Anon.

    1983-01-01

    This paper is a review of the current prospects for fusion reactor development based upon the present status in plasma physics research, fusion technology development and reactor conceptual design for the tokamak magnetic confinement concept. Recent advances in tokamak plasma research and fusion technology development are summarized. The direction and conclusions of tokamak reactor conceptual design are discussed. The status of alternate magnetic confinement concept research is reviewed briefly. A feasible timetable for the development of fusion reactors is presented

  1. Fusion engineering device design description

    International Nuclear Information System (INIS)

    Flanagan, C.A.; Steiner, D.; Smith, G.E.

    1981-12-01

    The US Magnetic Fusion Engineering Act of 1980 calls for the operation of a Fusion Engineering Device (FED) by 1990. It is the intent of the Act that the FED, in combination with other testing facilities, will establish the engineering feasibility of magnetic fusion energy. During 1981, the Fusion Engineering Design Center (FEDC), under the guidance of a Technical Management Board (TMB), developed a baseline design for the FED. This design is summarized herein

  2. Fusion engineering device design description

    Energy Technology Data Exchange (ETDEWEB)

    Flanagan, C.A.; Steiner, D.; Smith, G.E.

    1981-12-01

    The US Magnetic Fusion Engineering Act of 1980 calls for the operation of a Fusion Engineering Device (FED) by 1990. It is the intent of the Act that the FED, in combination with other testing facilities, will establish the engineering feasibility of magnetic fusion energy. During 1981, the Fusion Engineering Design Center (FEDC), under the guidance of a Technical Management Board (TMB), developed a baseline design for the FED. This design is summarized herein.

  3. Fusion Engineering Device design description

    Energy Technology Data Exchange (ETDEWEB)

    Flanagan, C.A.; Steiner, D.; Smith, G.E.

    1981-12-01

    The US Magnetic Fusion Engineering Act of 1980 calls for the operation of a Fusion Engineering Device (FED) by 1990. It is the intent of the Act that the FED, in combination with other testing facilities, will establish the engineering feasibility of magnetic fusion energy. During 1981, the Fusion Engineering Design Center (FEDC), under the guidance of a Technical Management Board (TMB), developed a baseline design for the FED. This design is summarized herein.

  4. Graphite for fusion energy applications

    International Nuclear Information System (INIS)

    Eatherly, W.P.; Clausing, R.E.; Strehlow, R.A.; Kennedy, C.R.; Mioduszewski, P.K.

    1987-03-01

    Graphite is in widespread and beneficial use in present fusion energy devices. This report reflects the view of graphite materials scientists on using graphite in fusion devices. Graphite properties are discussed with emphasis on application to fusion reactors. This report is intended to be introductory and descriptive and is not intended to serve as a definitive information source

  5. Incomplete fusion reactions in Ho

    Indian Academy of Sciences (India)

    This is confirmed by the predictions of breakup fusion model of the incomplete fusion. Keywords. Heavy ion ... Several models are used to explain these ICF namely, sum rule model [8], breakup fusion model [9], promptly emitted .... The solid lines are eye guides to the experimental points. Figure 1. Excitation functions of ...

  6. Structure information from fusion barriers

    Indian Academy of Sciences (India)

    It is shown that the analysis of fusion barrier distributions is not always an unambiguous test or a 'fingerprint' of the structure information of the colliding nuclei. Examples are presented with same fusion barrier distributions for nuclei having different structures. The fusion excitation functions for 16O+208Pb, using the coupled ...

  7. Fusion reactor radioactive waste management

    International Nuclear Information System (INIS)

    Kaser, J.D.; Postma, A.K.; Bradley, D.J.

    1976-01-01

    Quantities and compositions of non-tritium radioactive waste are estimated for some current conceptual fusion reactor designs, and disposal of large amounts of radioactive waste appears necessary. Although the initial radioactivity of fusion reactor and fission reactor wastes are comparable, the radionuclides in fusion reactor wastes are less hazardous and have shorter half-lives. Areas requiring further research are discussed

  8. HIV-1 gp41 Fusion Intermediate: A Target for HIV Therapeutics

    Directory of Open Access Journals (Sweden)

    Chungen Pan

    2010-02-01

    Full Text Available Human immunodeficiency virus (HIV-1 infection is initiated by the binding of gp120 envelope glyco-protein to its cell receptor (CD4 and a coreceptor (CXCR4 or CCR5, followed by a series of conformational changes in the gp41 transmembrane subunit. These changes include insertion of fusion peptide into the target cell membrane and association of C-heptad repeat (CHR peptide with the N-heptad repeat (NHR trimer, a pre-hairpin fusion intermediate. A stable six-helix bundle core is then formed, bringing the viral envelope and target cell membrane into close proximity for fusion. Peptides derived from the CHR region, such as T20 and C34, inhibit HIV-1 fusion by interacting with the gp41 fusion intermediate. A number of anti-HIV-1 peptides and small molecule compounds targeting the gp41 NHR-trimer have been identified. By combining HIV fusion/entry inhibitors targeting different sites in the gp41 fusion intermediate, a potent synergistic effect takes place, resulting in a potential new therapeutic strategy for the HIV infection/AIDS. Here, we present an overview of the current development of anti-HIV drugs, particularly those targeting the gp41 fusion intermediate.

  9. Plasma membrane disruption: repair, prevention, adaptation

    Science.gov (United States)

    McNeil, Paul L.; Steinhardt, Richard A.

    2003-01-01

    Many metazoan cells inhabit mechanically stressful environments and, consequently, their plasma membranes are frequently disrupted. Survival requires that the cell rapidly repair or reseal the disruption. Rapid resealing is an active and complex structural modification that employs endomembrane as its primary building block, and cytoskeletal and membrane fusion proteins as its catalysts. Endomembrane is delivered to the damaged plasma membrane through exocytosis, a ubiquitous Ca2+-triggered response to disruption. Tissue and cell level architecture prevent disruptions from occurring, either by shielding cells from damaging levels of force, or, when this is not possible, by promoting safe force transmission through the plasma membrane via protein-based cables and linkages. Prevention of disruption also can be a dynamic cell or tissue level adaptation triggered when a damaging level of mechanical stress is imposed. Disease results from failure of either the preventive or resealing mechanisms.

  10. Fusion development and technology

    International Nuclear Information System (INIS)

    Montgomery, D.B.

    1991-01-01

    This report discusses the following topics: superconducting magnet technology high field superconductors; advanced magnetic system and divertor development; poloidal field coils; gyrotron development; commercial reactor studies -- Aries; ITER physics; ITER superconducting PF scenario and magnet analysis; and safety, environmental and economic factors in fusion development

  11. Fusion og frasigelse

    DEFF Research Database (Denmark)

    Hansen, Lone L.

    2012-01-01

    Artiklen analyserer, i hvilket omfang reglerne om beskyttelse af lønmodtagere ved virksomhedsoverdragelse også finder anvendelse ved selskabsretliug fusion og spaltning. Der sættes fokus på forskrifterne om erhververens frasigelse af overdragerens kollektive overenskomster, og det efterprøves, om...

  12. Bouillabaisse sushi fusion power

    CERN Multimedia

    2004-01-01

    "If avant-garde cuisine is any guide, Japanese-French fusion does not work all that well. And the interminable discussions over the International Thermonuclear Experimental Reactor (ITER) suggest that what is true of cooking is true of physics" (1 page)

  13. Fusion technology programme

    International Nuclear Information System (INIS)

    Finken, D.

    1985-10-01

    KfK is involved in the European Fusion Programme predominantly in the NET and Fusion Technology part. The following fields of activity are covered: Studies for NET, alternative confinement concepts, and needs and issues of integral testing. Research on structural materials. Development of superconducting magnets. Gyrotron development (part of the Physics Programme). Nuclear technology (breeding materials, blanket design, tritium technology, safety and environmental aspects of fusion, remote maintenance). Reported here are status and results of work under contracts with the CEC within the NET and Technology Programme. The aim of the major part of this R and D work is the support of NET, some areas (e.g. materials, safety and environmental impact, blanket design) have a wider scope and address problems of a demonstration reactor. In the current working period, several new proposals have been elaborated to be implemented into the 85/89 Euratom Fusion Programme. New KfK contributions relate to materials research (dual beam and fast reactor irradiations, ferritic steels), to blanket engineering (MHD-effects) and to safety studies (e.g. magnet safety). (orig./GG)

  14. Rencontre on fusion technology

    International Nuclear Information System (INIS)

    Read, S.F.J.

    1979-02-01

    This report of a rencontre held to consider the technology of magnetic confinement fusion devices gives the agenda for the meeting and lists those topics which were identified as areas of research. These topics included materials, tritium, structures and heat transfer, neutronics and nuclear data, and corrosion problems. (UK)

  15. Intelligence Fusion [video

    OpenAIRE

    Center for Homeland Defense and Security Naval Postgraduate School; France, Paul

    2016-01-01

    Paul France is the Homeland Security Field Operations Manager for the state of Wisconsin. He is a current participant in the Naval Postgraduate School's Homeland Security Master's Program and is working on a thesis entitled 'Preventing Terrorism through Information Sharing Using TEW Systems and Intelligence Fusion Centers.

  16. Fusion Canada issue 21

    International Nuclear Information System (INIS)

    1993-08-01

    A short bulletin from the National Fusion Program highlighting in this issue Europe proposes Canada's participation in ITER, tritium for JET, CCFM/TdeV-Tokamak helium pumping and TdeV update, ITER-related R and D at CFFTP, ITER Deputy Director visits Canada, NFP Director to Chair IFRC, Award for Akira Hirose. 3 figs

  17. Fusion meets electronics

    Czech Academy of Sciences Publication Activity Database

    Řípa, Milan

    2011-01-01

    Roč. 1, č. 2 (2011), s. 8-8 ISSN 1818-5355 Institutional research plan: CEZ:AV0Z20430508 Keywords : fusion * AMPER 2011 * ITER * COMPASS * IPP * CR Subject RIV: BL - Plasma and Gas Discharge Physics http://www.jet.efda.org/ multimedia /newsletter/current/

  18. Magnetic fusion energy

    International Nuclear Information System (INIS)

    McNamara, B.

    1977-01-01

    A brief review of fusion research during the last 20 years is given. Some highlights of theoretical plasma physics are presented. The role that computational plasma physics is playing in analyzing and understanding the experiments of today is discussed. The magnetic mirror program is reviewed

  19. Fusion energy studies

    International Nuclear Information System (INIS)

    Anon.

    1978-01-01

    The following topics are considered: (1) cryosorption vacuum pumping for fusion reactors, (2) TNS support studies, (3) tritium recovery from irradiated Li-Al and SAP, (4) actinide oxides, nitrides, and carbides, and (5) transition metal-actinide-C phase equilibria

  20. Fusion reactor materials

    Energy Technology Data Exchange (ETDEWEB)

    none,

    1989-01-01

    This paper discuses the following topics on fusion reactor materials: irradiation, facilities, test matrices, and experimental methods; dosimetry, damage parameters, and activation calculations; materials engineering and design requirements; fundamental mechanical behavior; radiation effects; development of structural alloys; solid breeding materials; and ceramics.

  1. Fusion Canada issue 24

    International Nuclear Information System (INIS)

    1994-04-01

    A short bulletin from the National Fusion Program highlighting in this issue the opening of the Garching ITER site, a CCFM/TdeV update,a tritium release field test, measuring radial profile of plasma current density on TdeV, power supplies for TdeV, 5th International Tritium Technology conference, and 1994 basic tritium course. 4 figs

  2. International fusion og spaltning

    DEFF Research Database (Denmark)

    Hansen, Lone L.

    Bogen analyserer de nye muligheder fra 2007 i europæisk ret med hensyn til fusion eller spaltning mellem aktieselskaber og anpartsselskaber med hjemsted i forskellige europæiske lande. Bogen gennemgår de nye muligheder for strukturændringer, der herved er opstået mulighed for, og den sætter fokus...

  3. Controlled nuclear fusion apparatus

    International Nuclear Information System (INIS)

    Bussard, R.W.; Coppi, B.

    1982-01-01

    A fusion power generating device is disclosed having a relatively small and inexpensive core region which may be contained within an energy absorbing blanket region. The fusion power core region contains apparatus of the toroidal type for confining a high density plasma. The fusion power core is removable from the blanket region and may be disposed and/or recycled for subsequent use within the same blanket region. Thermonuclear ignition of the plasma is obtained by feeding neutral fusible gas into the plasma in a controlled manner such that charged particle heating produced by the fusion reaction is utilized to bootstrap the device to a region of high temperatures and high densities wherein charged particle heating is sufficient to overcome radiation and thermal conductivity losses. The high density plasma produces a large radiation and particle flux on the first wall of the plasma core region thereby necessitating replacement of the core from the blanket region from time to time. A series of potentially disposable and replaceable central core regions are disclosed for a large-scale economical electrical power generating plant

  4. lysosome tethering and fusion

    Indian Academy of Sciences (India)

    AMIT TULI

    LYSOSOME. MTOC. LATE ENDOSOME. Arl8b promotes the assembly of the HOPS complex on the lysosomes to mediate late endosome-lysosome fusion and cargo delivery to lysosomes. Khatter D et al., J Cell Science 2015. Khatter D et al., Cellular Logistics 2015 ...

  5. Membrane anchorage brings about fusogenic properties in a short synthetic peptide

    NARCIS (Netherlands)

    Pecheur, EI; Hoekstra, D; SainteMarie, J; Maurin, L; Bienvenue, A; Philippot, [No Value

    1997-01-01

    The fusogenic properties of an amphipathic net-negative peptide (wae 11), consisting of 11 amino acid residues, were studied. We demonstrate that, whereas the free peptide displays no significant fusion activity, membrane fusion is strongly promoted when the peptide is anchored to a liposomal

  6. Controlled thermonuclear fusion reactors

    International Nuclear Information System (INIS)

    Walstrom, P.L.

    1976-01-01

    Controlled production of energy by fusion of light nuclei has been the goal of a large portion of the physics community since the 1950's. In order for a fusion reaction to take place, the fuel must be heated to a temperature of 100 million degrees Celsius. At this temperature, matter can exist only in the form of an almost fully ionized plasma. In order for the reaction to produce net power, the product of the density and energy confinement time must exceed a minimum value of 10 20 sec m -3 , the so-called Lawson criterion. Basically, two approaches are being taken to meet this criterion: inertial confinement and magnetic confinement. Inertial confinement is the basis of the laser fusion approach; a fuel pellet is imploded by intense laser beams from all sides and ignites. Magnetic confinement devices, which exist in a variety of geometries, rely upon electromagnetic forces on the charged particles of the plasma to keep the hot plasma from expanding. Of these devices, the most encouraging results have been achieved with a class of devices known as tokamaks. Recent successes with these devices have given plasma physicists confidence that scientific feasibility will be demonstrated in the next generation of tokamaks; however, an even larger effort will be required to make fusion power commercially feasible. As a result, emphasis in the controlled thermonuclear research program is beginning to shift from plasma physics to a new branch of nuclear engineering which can be called fusion engineering, in which instrumentation and control engineers will play a major role. Among the new problem areas they will deal with are plasma diagnostics and superconducting coil instrumentation

  7. Controlled thermonuclear fusion reactors

    Energy Technology Data Exchange (ETDEWEB)

    Walstrom, P.L.

    1976-01-01

    Controlled production of energy by fusion of light nuclei has been the goal of a large portion of the physics community since the 1950's. In order for a fusion reaction to take place, the fuel must be heated to a temperature of 100 million degrees Celsius. At this temperature, matter can exist only in the form of an almost fully ionized plasma. In order for the reaction to produce net power, the product of the density and energy confinement time must exceed a minimum value of 10/sup 20/ sec m/sup -3/, the so-called Lawson criterion. Basically, two approaches are being taken to meet this criterion: inertial confinement and magnetic confinement. Inertial confinement is the basis of the laser fusion approach; a fuel pellet is imploded by intense laser beams from all sides and ignites. Magnetic confinement devices, which exist in a variety of geometries, rely upon electromagnetic forces on the charged particles of the plasma to keep the hot plasma from expanding. Of these devices, the most encouraging results have been achieved with a class of devices known as tokamaks. Recent successes with these devices have given plasma physicists confidence that scientific feasibility will be demonstrated in the next generation of tokamaks; however, an even larger effort will be required to make fusion power commercially feasible. As a result, emphasis in the controlled thermonuclear research program is beginning to shift from plasma physics to a new branch of nuclear engineering which can be called fusion engineering, in which instrumentation and control engineers will play a major role. Among the new problem areas they will deal with are plasma diagnostics and superconducting coil instrumentation.

  8. Lysosomal fusion and SNARE function are impaired by cholesterol accumulation in lysosomal storage disorders.

    Science.gov (United States)

    Fraldi, Alessandro; Annunziata, Fabio; Lombardi, Alessia; Kaiser, Hermann-Josef; Medina, Diego Luis; Spampanato, Carmine; Fedele, Anthony Olind; Polishchuk, Roman; Sorrentino, Nicolina Cristina; Simons, Kai; Ballabio, Andrea

    2010-11-03

    The function of lysosomes relies on the ability of the lysosomal membrane to fuse with several target membranes in the cell. It is known that in lysosomal storage disorders (LSDs), lysosomal accumulation of several types of substrates is associated with lysosomal dysfunction and impairment of endocytic membrane traffic. By analysing cells from two severe neurodegenerative LSDs, we observed that cholesterol abnormally accumulates in the endolysosomal membrane of LSD cells, thereby reducing the ability of lysosomes to efficiently fuse with endocytic and autophagic vesicles. Furthermore, we discovered that soluble N-ethylmaleimide-sensitive factor attachment protein (SNAP) receptors (SNAREs), which are key components of the cellular membrane fusion machinery are aberrantly sequestered in cholesterol-enriched regions of LSD endolysosomal membranes. This abnormal spatial organization locks SNAREs in complexes and impairs their sorting and recycling. Importantly, reducing membrane cholesterol levels in LSD cells restores normal SNARE function and efficient lysosomal fusion. Our results support a model by which cholesterol abnormalities determine lysosomal dysfunction and endocytic traffic jam in LSDs by impairing the membrane fusion machinery, thus suggesting new therapeutic targets for the treatment of these disorders.

  9. Directions for improved fusion reactors

    International Nuclear Information System (INIS)

    Krakowski, R.A.; Miller, R.L.; Delene, J.G.

    1986-01-01

    Conceptual fusion reactor studies over the past 10 to 15 years have projected systems that may be too large, complex, and costly to be of commercial interest. One main direction for improved fusion reactors points towards smaller, higher-power-density approaches. First-order economic issues (i.e., unit direct cost and cost of electricity) are used to support the need for more compact fusion reactors. A generic fusion physics/engineering/costing model is used to provide a quantiative basis for these arguments for specific fusion concepts

  10. Controlled fusion and plasma physics

    CERN Document Server

    Miyamoto, Kenro

    2006-01-01

    Resulting from ongoing, international research into fusion processes, the International Tokamak Experimental Reactor (ITER) is a major step in the quest for a new energy source.The first graduate-level text to cover the details of ITER, Controlled Fusion and Plasma Physics introduces various aspects and issues of recent fusion research activities through the shortest access path. The distinguished author breaks down the topic by first dealing with fusion and then concentrating on the more complex subject of plasma physics. The book begins with the basics of controlled fusion research, foll

  11. Crystal structure of Venezuelan hemorrhagic fever virus fusion glycoprotein reveals a class 1 postfusion architecture with extensive glycosylation.

    Science.gov (United States)

    Parsy, Marie-Laure; Harlos, Karl; Huiskonen, Juha T; Bowden, Thomas A

    2013-12-01

    Guanarito virus (GTOV) is an emergent and deadly pathogen. We present the crystal structure of the glycosylated GTOV fusion glycoprotein to 4.1-Å resolution in the postfusion conformation. Our structure reveals a classical six-helix bundle and presents direct verification that New World arenaviruses exhibit class I viral membrane fusion machinery. The structure provides visualization of an N-linked glycocalyx coat, and consideration of glycan dynamics reveals extensive coverage of the underlying protein surface, following virus-host membrane fusion.

  12. Enhanced expression of rabies virus surface G-protein in Escherichia coli using SUMO fusion.

    Science.gov (United States)

    Singh, Ankit; Yadav, Dinesh; Rai, Krishan Mohan; Srivastava, Meenal; Verma, Praveen C; Singh, Pradhyumna K; Tuli, Rakesh

    2012-01-01

    Fusion systems are known to increase the expression of difficult to express recombinant proteins in soluble form to facilitate their purification. Rabies glycoprotein was also tough to express at sufficient level in soluble form in both E. coli and plant. The present work was aimed to over-express and purify this membrane protein from soluble extract of E. coli. Fusion of Small Ubiqutin like Modifier (SUMO) with rabies glycoprotein increased ~1.5 fold higher expression and ~3.0 fold solubility in comparison to non-fused in E. coli. The SUMO fusion also simplified the purification process. Previously engineered rabies glycoprotein gene in tobacco plants provides complete protection to mice, but the expression was very low for purification. Our finding demonstrated that the SUMO-fusion was useful for enhancing expression and solubility of the membrane protein and again proves to be a good alternative technology for applications in biomedical and pharmaceutical research.

  13. Nuclear fusion and genome encounter during yeast zygote formation.

    Science.gov (United States)

    Tartakoff, Alan Michael; Jaiswal, Purnima

    2009-06-01

    When haploid cells of Saccharomyces cerevisiae are crossed, parental nuclei congress and fuse with each other. To investigate underlying mechanisms, we have developed assays that evaluate the impact of drugs and mutations. Nuclear congression is inhibited by drugs that perturb the actin and tubulin cytoskeletons. Nuclear envelope (NE) fusion consists of at least five steps in which preliminary modifications are followed by controlled flux of first outer and then inner membrane proteins, all before visible dilation of the waist of the nucleus or coalescence of the parental spindle pole bodies. Flux of nuclear pore complexes occurs after dilation. Karyogamy requires both the Sec18p/NSF ATPase and ER/NE luminal homeostasis. After fusion, chromosome tethering keeps tagged parental genomes separate from each other. The process of NE fusion and evidence of genome independence in yeast provide a prototype for understanding related events in higher eukaryotes.

  14. Fusion research activities in China

    International Nuclear Information System (INIS)

    Deng Xiwen

    1998-01-01

    The fusion program in China has been executed in most areas of magnetic confinement fusion for more than 30 years. Basing on the situation of the power supply requirements of China, the fusion program is becoming an important and vital component of the nuclear power program in China. This paper reviews the status of fusion research and next step plans in China. The motivation and goal of the Chinese fusion program is explained. Research and development on tokamak physics and engineering in the southwestern institute of physics (SWIP) and the institute of plasma physics of Academic Sinica (ASIPP) are introduced. A fusion breeder program and a pure fusion reactor design program have been supported by the state science and technology commission (SSTC) and the China national nuclear corporation (CNNC), respectively. Some features and progress of fusion reactor R and D activities are reviewed. Non fusion applications of plasma science are an important part of China fusion research; a brief introduction about this area is given. Finally, an introductional collaboration network on fusion research activities in China is reported. (orig.)

  15. The path to fusion power.

    Science.gov (United States)

    Llewellyn Smith, Chris; Ward, David

    2007-04-15

    Fusion is potentially an environmentally responsible and intrinsically safe source of essentially limitless power. It should be possible to build viable fusion power stations, and it looks as if the cost of fusion power will be reasonable. But time is needed to further develop the technology and to test in power station conditions the materials that would be used in their construction. Assuming no major adverse surprises, an orderly fusion development programme could lead to a prototype fusion power station putting electricity into the grid within 30 years, with commercial fusion power following some 10 or more years later. In the second half of the century, fusion could therefore be an important part of the portfolio of measures that are needed to cope with rising demand for energy in an environmentally responsible manner. In this paper, we describe the basics of fusion, its potential attractions, the status of fusion R&D, the remaining challenges and how they will be tackled at the International Tokamak Experimental Reactor and the proposed International Fusion Materials Irradiation Facility, and the timetable for the subsequent commercialization of fusion power.

  16. Physics of magnetic confinement fusion

    Science.gov (United States)

    Wagner, F.

    2013-06-01

    Fusion is the energy source of the universe. The local conditions in the core of the Sun allow the transfer of mass into energy, which is finally released in the form of radiation. Technical fusion melts deuterons and tritons to helium releasing large amounts of energy per fusion process. Because of the conditions for fusion, which will be deduced, the fusion fuel is in the plasma state. Here we report on the confinement of fusion plasmas by magnetic fields. Different confinement concepts — tokamaks and stellarators — will be introduced and described. The first fusion reactor, ITER, and the most modern stellarator, Wendelstein 7-X, are under construction. Their basic features and objectives will be presented.

  17. Physics of magnetic confinement fusion

    Directory of Open Access Journals (Sweden)

    Wagner F.

    2013-06-01

    Full Text Available Fusion is the energy source of the universe. The local conditions in the core of the Sun allow the transfer of mass into energy, which is finally released in the form of radiation. Technical fusion melts deuterons and tritons to helium releasing large amounts of energy per fusion process. Because of the conditions for fusion, which will be deduced, the fusion fuel is in the plasma state. Here we report on the confinement of fusion plasmas by magnetic fields. Different confinement concepts — tokamaks and stellarators — will be introduced and described. The first fusion reactor, ITER, and the most modern stellarator, Wendelstein 7-X, are under construction. Their basic features and objectives will be presented.

  18. Fusion of Selected Cells and Vesicles Mediated by Optically Trapped Plasmonic Nanoparticles

    DEFF Research Database (Denmark)

    Bahadori, Azra

    . In this work, we introduce a novel and extremely flexible physical method which can trigger membrane fusion in a highly selective manner not only between synthetic GUVs of different compositions, but also between live cells which remain viable after fusion. Optical tweezers’ laser (1064 nm) is used to position......Selective fusion of two membrane surrounded volumes is of great interest in nanochemistry and nanomedicine as it can pave the way for performing controlled nanoscale chemical reactions and for delivering a cargo (e.g., chemicals, genetic regulatory factors, etc.) to a desired living cell...... the two desired cells and/or GUVs next to each other and in immediate contact. Then, the same laser is placed in the contact zone between the two adjacent membranes until one or more gold nanoparticles diffuse into the focus. Gold nanoparticles absorb part of near infrared light and dissipate the absorbed...

  19. Fusion theory and computations

    International Nuclear Information System (INIS)

    Bers, A.; Coppi, B.; Davidson, R.; Dupree, T.; Freidberg, J.; Molvig, K.; McCune, J.

    1979-01-01

    It is proposed to carry out theoretical studies of the equilibrium, stability, transport and heating properties of high-temperature fusion plasmas. Continued emphasis will be placed on the effective interface of fusion theory and computations with the local Alcator, Versator, Constance and Torex experimental programs. The proposed research includes but will not be limited to the following types of studies: (a) investigation of RF heating of toroidal plasmas, (b) investigation of the MHD equilibrium and stability properties of tokamak plasmas, (c) develop the basic understanding of a wide variety of non-linear and turbulent phenomena, including stochastic magnetic fields, clumps and nonlinear saturation of linear instabilities, (d) investigate the effects of ambipolar fields on transport and stability properties of toroidal plasmas. Investigate high-beat stability properties of tandem-mirror systems, and (e) investigation of the MHD equilibrium and stability properties of Torsatron/Stellarator configurations

  20. Fusion through the NET

    International Nuclear Information System (INIS)

    Spears, B.

    1987-01-01

    The paper concerns the next generation of fusion machines which are intended to demonstrate the technical viability of fusion. In Europe, the device that will follow on from JET is known as NET - the Next European Torus. If the design programme for NET proceeds, Europe could start to build the machine in 1994. The present JET programme hopes to achieve breakeven in the early 1990's. NET hopes to reach ignition in the next century, and so lay the foundation for a demonstration reactor. A description is given of the technical specifications of the components of NET, including: the first wall, the divertors to protect the wall, the array of magnets that provide the fields containing the plasma, the superconducting magnets, and the shield of the machine. NET's research programme is briefly outlined, including the testing programme to optimise conditions in the machine to achieve ignition, and its safety work. (U.K.)

  1. Fusion technology programme

    International Nuclear Information System (INIS)

    Finken, D.

    1984-10-01

    The KfK-Association has continued work on 17 R and D contracts of the Fusion Technology Programme. An effort of 94 manyears per year is at present contributed by 10 KfK departments, covering all aereas defined in the Fusion Technology Programme. The dominant part of the work is directed towards the need of the NET design or supporting experiments. Some additional effort addresses long term technological issues and system studies relevant to DEMO or confinement schemes alternative to tokamaks. Direct contribution to the NET team has increased by augmentation of NET study contracts and delegation of personnel, three KfK delegates being at present members of the NET team. In reverse, specifications and design guidelines worked out by NET have started to have an impact on the current R and D-work in the laboratory. (orig./GG)

  2. DNA fusion gene vaccines

    DEFF Research Database (Denmark)

    Holst, Peter Johannes; Bassi, Maria Rosaria; Thomsen, Allan Randrup

    2010-01-01

    DNA vaccines are versatile and safe, but limited immunogenicity has prevented their use in the clinical setting. Experimentally, immunogenicity may be enhanced by the use of new delivery technologies, by coadministration of cytokines and pathogen-associated molecular patterns, or by fusion...... of antigens into molecular domains that enhance antigen presentation. More specifically, the immunogenicity of DNA vaccines may benefit from increased protein synthesis, increased T-cell help and MHC class I presentation, and the addition of a range of specific cytokines and pathogen-associated molecular...... with these modifications, it is likely that the primary use of DNA vaccines may be as primers for viral-vectored vaccines, rather than as single agents. This review discusses the approaches used to enhance DNA vaccine immunogenicity, with a primary focus on fusion strategies that enhance antigen presentation....

  3. Fusion: hopes and doubts

    International Nuclear Information System (INIS)

    Bruhns, H.

    1990-01-01

    The author examines current public and political attitudes to fossil fuel and nuclear fission generated electricity. He concludes that the nuclear fusion programme, currently funded and managed jointly in Europe, offers an environmentally acceptable alternative. He explains progress to date at the Joint European Torus at Culham in the United Kingdom and points out that, while much admirable work has been carried out, further work to reach plasma ignition must be in a bigger, more costly new facility yet to be built. Political and economic opposition to high cost nuclear research is highlighted, especially in West Germany. The author concludes by discussing the advantages of power generation by nuclear fusion from the safety, operational and economic points of view. (UK)

  4. Fluorination by fusion

    International Nuclear Information System (INIS)

    Gray, J.H.

    1986-01-01

    LECO crucibles and incinerator ash are two waste categories that cannot be discarded due to the presence of insoluble transuranics. Current chemical processing methods are not too effective, requiring a number of repeated operations in order to dissolve more than half the transuranics. An alternate dissolution approach has been developed involving the use of ammonium bifluoride. Low temperature fusion of the waste with ammonium bifluoride is followed by dissolution of the fused material in boiling nitric acid solutions. Greater than 60% of the transuranics contained in LECO crucibles and greater than 95% of the transuranics mixed with the incinerator ash are dissolved after a single fusion and dissolution step. Fluorination of the transuranics along with other impurities appears to render the waste material soluble in nitric acid

  5. Pulsed fusion reactors

    International Nuclear Information System (INIS)

    1975-01-01

    This summer school specialized in examining specific fusion center systems. Papers on scientific feasibility are first presented: confinement of high-beta plasma, liners, plasma focus, compression and heating and the use of high power electron beams for thermonuclear reactors. As for technological feasibility, lectures were on the theta-pinch toroidal reactors, toroidal diffuse pinch, electrical engineering problems in pulsed magnetically confined reactors, neutral gas layer for heat removal, the conceptual design of a series of laser fusion power plants with ''Saturn'', implosion experiments and the problem of the targets, the high brightness lasers for plasma generation, and topping and bottoming cycles. Some problems common to pulsed reactors were examined: energy storage and transfer, thermomechanical and erosion effects in the first wall and blanket, the problems of tritium production, radiation damage and neutron activation in blankets, and the magnetic and inertial confinement

  6. Heavy ion fusion III

    International Nuclear Information System (INIS)

    Hammer, D.; Max, C.; Perkins, F.; Rosenbluth, M.

    1987-03-01

    This report updates Heavy Ion Fusion, JSR-82-302, dated January, 1983. During the last four years, program management and direction has been changed and the overall Inertial Confinement Program has been reviewed. This report therefore concentrates on accelerator physics issues, how the program has addressed those issues during the last four years, and how it will be addressing them in the future. 8 refs., 3 figs

  7. (Fusion energy research)

    Energy Technology Data Exchange (ETDEWEB)

    Phillips, C.A. (ed.)

    1988-01-01

    This report discusses the following topics: principal parameters achieved in experimental devices (FY88); tokamak fusion test reactor; Princeton beta Experiment-Modification; S-1 Spheromak; current drive experiment; x-ray laser studies; spacecraft glow experiment; plasma deposition and etching of thin films; theoretical plasma; tokamak modeling; compact ignition tokamak; international thermonuclear experimental reactor; Engineering Department; Project Planning and Safety Office; quality assurance and reliability; and technology transfer.

  8. Fusion Reactor Materials

    International Nuclear Information System (INIS)

    Moons, F.

    1998-01-01

    SCK-CEN's programme on fusion reactor materials includes studies (1) to investigate fracture mechanics of neutron-irradiated beryllium; (2) to describe the helium behaviour in irradiated beryllium at atomic scale; (3) to define the kinetics of beryllium reacting with air or steam; (3) to perform a feasibility study for the testing of integrated blanket modules under neutron irradiation. Progress and achievements in 1997 are reported

  9. [Fusion energy research

    International Nuclear Information System (INIS)

    Phillips, C.A.

    1988-01-01

    This report discusses the following topics: principal parameters achieved in experimental devices (FY88); tokamak fusion test reactor; Princeton beta Experiment-Modification; S-1 Spheromak; current drive experiment; x-ray laser studies; spacecraft glow experiment; plasma deposition and etching of thin films; theoretical plasma; tokamak modeling; compact ignition tokamak; international thermonuclear experimental reactor; Engineering Department; Project Planning and Safety Office; quality assurance and reliability; and technology transfer

  10. Fusion development and technology

    International Nuclear Information System (INIS)

    Montgomery, D.B.

    1992-01-01

    This report discusses the following: superconducting magnet technology; high field superconductors; advanced magnetic system and divertor development; poloidal field coils; gyrotron development; commercial reactor studies--aries; ITER physics: alpha physics and alcator R ampersand D for ITER; lower hybrid current drive and heating in the ITER device; ITER superconducting PF scenario and magnet analysis; ITER systems studies; and safety, environmental and economic factors in fusion development

  11. Fusion development and technology

    Energy Technology Data Exchange (ETDEWEB)

    Montgomery, D.B.

    1992-01-01

    This report discusses the following: superconducting magnet technology; high field superconductors; advanced magnetic system and divertor development; poloidal field coils; gyrotron development; commercial reactor studies--aries; ITER physics: alpha physics and alcator R D for ITER; lower hybrid current drive and heating in the ITER device; ITER superconducting PF scenario and magnet analysis; ITER systems studies; and safety, environmental and economic factors in fusion development.

  12. Inertial confinement fusion

    International Nuclear Information System (INIS)

    Nuckolls, J.H.; Wood, L.L.

    1988-01-01

    Edward Teller has been a strong proponent of harnessing nuclear explosions for peaceful purposes. There are two approaches: Plowshare, which utilizes macro- explosions, and inertial confinement fusion, which utilizes microexplosions. The development of practical fusion power plants is a principal goal of the inertial program. It is remarkable that Teller's original thermonuclear problem, how to make super high yield nuclear explosions, and the opposite problem, how to make ultra low yield nuclear explosions, may both be solved by Teller's radiation implosion scheme. This paper reports on the essential physics of these two thermonuclear domains, which are separated by nine orders of magnitude in yield, provided by Teller's similarity theorem and its exceptions. Higher density makes possible thermonuclear burn of smaller masses of fuel. The leverage is high: the scale of the explosion diminishes with the square of the increase in density. The extraordinary compressibility of matter, first noticed by Teller during the Los Alamos atomic bomb program, provides an almost incredible opportunity to harness fusion. The energy density of thermonuclear fuels isentropically compressed to super high-- -densities---even to ten thousand times solid density---is small compared to the energy density at thermonuclear ignition temperatures. In small masses of fuel imploded to these super high matter densities, the energy required to achieve ignition may be greatly reduced by exploiting thermonuclear propagation from a relatively small hot spot

  13. Future of fusion implementation

    International Nuclear Information System (INIS)

    Beardsworth, E.; Powell, J.R.

    1978-01-01

    For fusion to become available for commercial use in the 21st century, R and D must be undertaken now. But it is hard to justify these expenditures with a cost/benefit oriented assessment methodology, because of both the time-frame and the uncertainty of the future benefits. Focusing on the factors most relevant for current consideration of fusion's commercial prospects, i.e., consumption levels and the outcomes for fission, solar, and coal, many possible futures of the US energy system are posited and analyzed under various assumptions about costs. The Reference Energy System approach was modified to establish both an appropriate degree of detail and explicit time dependence, and a computer code used to organize the relevant data and to perform calculations of system cost (annual and discounted present value), resource use, and residuals that are implied by the consumptions levels and technology mix in each scenario. Not unreasonable scenarios indicate benefits in the form of direct cost savings, which may well exceed R and D costs, which could be attributed to the implementation of fusion

  14. Heavy ion fusion

    International Nuclear Information System (INIS)

    Hofmann, Ingo

    1993-01-01

    With controlled thermonuclear fusion holding out the possibility of a prolific and clean new source of energy, the goal remains elusive after many years of continual effort. While the conventional Tokamak route with magnetic confinement continues to hit the headlines, other alternatives are now becoming competitive. One possible solution is to confine the thermonuclear fuel pellet by high power beams. Current research and perspectives for future work in such inertial confinement was the subject of the 'Prospects for Heavy Ion Fusion' European Research Conference held in Aghia Pelaghia, Crete, last year. Its main focus was on the potential of heavy ion accelerators as well as recent advances in target physics with high power lasers and light ion beams. Carlo Rubbia declared that high energy accelerators, with their high efficiency, are the most promising approach to economical fusion energy production. However the need for cost saving in the driver accelerator requires new ideas in target design tailored to the particularities of heavy ion beams, which need to be pushed to the limits of high current and phase space density at the same time

  15. Dynamic assembly of brambleberry mediates nuclear envelope fusion during early development.

    Science.gov (United States)

    Abrams, Elliott W; Zhang, Hong; Marlow, Florence L; Kapp, Lee; Lu, Sumei; Mullins, Mary C

    2012-08-03

    To accommodate the large cells following zygote formation, early blastomeres employ modified cell divisions. Karyomeres are one such modification, mitotic intermediates wherein individual chromatin masses are surrounded by nuclear envelope; the karyomeres then fuse to form a single mononucleus. We identified brambleberry, a maternal-effect zebrafish mutant that disrupts karyomere fusion, resulting in formation of multiple micronuclei. As karyomeres form, Brambleberry protein localizes to the nuclear envelope, with prominent puncta evident near karyomere-karyomere interfaces corresponding to membrane fusion sites. brambleberry corresponds to an unannotated gene with similarity to Kar5p, a protein that participates in nuclear fusion in yeast. We also demonstrate that Brambleberry is required for pronuclear fusion following fertilization in zebrafish. Our studies provide insight into the machinery required for karyomere fusion and suggest that specialized proteins are necessary for proper nuclear division in large dividing blastomeres. Copyright © 2012 Elsevier Inc. All rights reserved.

  16. A new NFIA:RAF1 fusion activating the MAPK pathway in pilocytic astrocytoma

    DEFF Research Database (Denmark)

    Yde, Christina Westmose; Sehested, Astrid; Regué, Àngels Mateu

    2016-01-01

    are more rarely found. However, in some cases, no apparent cancer driver events can be identified. Here, we describe a novel fusion between the transcription factor nuclear factor 1A (NFIA) and Raf-1 proto-oncogene (RAF1) in a 5-year old boy with PA. The novel fusion was identified as part......Pilocytic astrocytoma (PA) is one of the most common brain cancers among children and activation of the Mitogen-Activated Protein Kinase (MAPK) pathway is considered the hallmark. In the majority of cases, oncogenic BRAF fusions or BRAF V600E mutations are observed, while RAF1 or NF1 alterations...... of a comprehensive genomic tumor profiling. We show that the NFIA:RAF1 fusion results in constitutive Raf1 kinase activity, leading to activation of downstream MEK1/2 cascade and increased proliferation of cancer cells. The NFIA:RAF1 fusion displayed distinct subcellular localization towards the plasma membrane...

  17. Prm3p is a pheromone-induced peripheral nuclear envelope protein required for yeast nuclear fusion.

    Science.gov (United States)

    Shen, Shu; Tobery, Cynthia E; Rose, Mark D

    2009-05-01

    Nuclear membrane fusion is the last step in the mating pathway of the yeast Saccharomyces cerevisiae. We adapted a bioinformatics approach to identify putative pheromone-induced membrane proteins potentially required for nuclear membrane fusion. One protein, Prm3p, was found to be required for nuclear membrane fusion; disruption of PRM3 caused a strong bilateral defect, in which nuclear congression was completed but fusion did not occur. Prm3p was localized to the nuclear envelope in pheromone-responding cells, with significant colocalization with the spindle pole body in zygotes. A previous report, using a truncated protein, claimed that Prm3p is localized to the inner nuclear envelope. Based on biochemistry, immunoelectron microscopy and live cell microscopy, we find that functional Prm3p is a peripheral membrane protein exposed on the cytoplasmic face of the outer nuclear envelope. In support of this, mutations in a putative nuclear localization sequence had no effect on full-length protein function or localization. In contrast, point mutations and deletions in the highly conserved hydrophobic carboxy-terminal domain disrupted both protein function and localization. Genetic analysis, colocalization, and biochemical experiments indicate that Prm3p interacts directly with Kar5p, suggesting that nuclear membrane fusion is mediated by a protein complex.

  18. ER-associated SNAREs and Sey1p mediate nuclear fusion at two distinct steps during yeast mating.

    Science.gov (United States)

    Rogers, Jason V; Arlow, Tim; Inkellis, Elizabeth R; Koo, Timothy S; Rose, Mark D

    2013-12-01

    During yeast mating, two haploid nuclei fuse membranes to form a single diploid nucleus. However, the known proteins required for nuclear fusion are unlikely to function as direct fusogens (i.e., they are unlikely to directly catalyze lipid bilayer fusion) based on their predicted structure and localization. Therefore we screened known fusogens from vesicle trafficking (soluble N-ethylmaleimide-sensitive factor attachment protein receptors [SNAREs]) and homotypic endoplasmic reticulum (ER) fusion (Sey1p) for additional roles in nuclear fusion. Here we demonstrate that the ER-localized SNAREs Sec20p, Ufe1p, Use1p, and Bos1p are required for efficient nuclear fusion. In contrast, Sey1p is required indirectly for nuclear fusion; sey1Δ zygotes accumulate ER at the zone of cell fusion, causing a block in nuclear congression. However, double mutants of Sey1p and Sec20p, Ufe1p, or Use1p, but not Bos1p, display extreme ER morphology defects, worse than either single mutant, suggesting that retrograde SNAREs fuse ER in the absence of Sey1p. Together these data demonstrate that SNAREs mediate nuclear fusion, ER fusion after cell fusion is necessary to complete nuclear congression, and there exists a SNARE-mediated, Sey1p-independent ER fusion pathway.

  19. Inertial fusion energy; L'energie de fusion inertielle

    Energy Technology Data Exchange (ETDEWEB)

    Decroisette, M.; Andre, M.; Bayer, C.; Juraszek, D. [CEA Bruyeres-le-Chatel, Dir. des Systemes d' Information (CEA/DIF), 91 (France); Le Garrec, B. [CEA Centre d' Etudes Scientifiques et Techniques d' Aquitaine, 33 - Le Barp (France); Deutsch, C. [Paris-11 Univ., 91 - Orsay (France); Migus, A. [Institut d' Optique Centre scientifique, 91 - Orsay (France)

    2005-07-01

    We first recall the scientific basis of inertial fusion and then describe a generic fusion reactor with the different components: the driver, the fusion chamber, the material treatment unit, the target factory and the turbines. We analyse the options proposed at the present time for the driver and for target irradiation scheme giving the state of art for each approach. We conclude by the presentation of LMJ (laser Megajoule) and NIF (national ignition facility) projects. These facilities aim to demonstrate the feasibility of laboratory DT ignition, first step toward Inertial Fusion Energy. (authors)

  20. Localized cyclic AMP-dependent protein kinase activity is required for myogenic cell fusion

    International Nuclear Information System (INIS)

    Mukai, Atsushi; Hashimoto, Naohiro

    2008-01-01

    Multinucleated myotubes are formed by fusion of mononucleated myogenic progenitor cells (myoblasts) during terminal skeletal muscle differentiation. In addition, myoblasts fuse with myotubes, but terminally differentiated myotubes have not been shown to fuse with each other. We show here that an adenylate cyclase activator, forskolin, and other reagents that elevate intracellular cyclic AMP (cAMP) levels induced cell fusion between small bipolar myotubes in vitro. Then an extra-large myotube, designated a 'myosheet,' was produced by both primary and established mouse myogenic cells. Myotube-to-myotube fusion always occurred between the leading edge of lamellipodia at the polar end of one myotube and the lateral plasma membrane of the other. Forskolin enhanced the formation of lamellipodia where cAMP-dependent protein kinase (PKA) was accumulated. Blocking enzymatic activity or anchoring of PKA suppressed forskolin-enhanced lamellipodium formation and prevented fusion of multinucleated myotubes. Localized PKA activity was also required for fusion of mononucleated myoblasts. The present results suggest that localized PKA plays a pivotal role in the early steps of myogenic cell fusion, such as cell-to-cell contact/recognition through lamellipodium formation. Furthermore, the localized cAMP-PKA pathway might be involved in the specification of the fusion-competent areas of the plasma membrane in lamellipodia of myogenic cells