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Sample records for intracellular actin pedestals

  1. Intracellular transport based on actin polymerization.

    PubMed

    Khaitlina, S Yu

    2014-09-01

    In addition to the intracellular transport of particles (cargo) along microtubules, there are in the cell two actin-based transport systems. In the actomyosin system the transport is driven by myosin, which moves the cargo along actin microfilaments. This transport requires the hydrolysis of ATP in the myosin molecule motor domain that induces conformational changes in the molecule resulting in the myosin movement along the actin filament. The other actin-based transport system of the cell does not involve myosin or other motor proteins. This system is based on a unidirectional actin polymerization, which depends on ATP hydrolysis in actin polymers and is initiated by proteins bound to the surface of transported particles. Obligatory components of the actin-based transport are proteins of the WASP/Scar family and a complex of Arp2/3 proteins. Moreover, the actin-based systems often contain dynamin and cortactin. It is known that a system of actin filaments formed on the surface of particles, the so-called "comet-like tail", is responsible for intracellular movements of pathogenic bacteria, micropinocytotic vesicles, clathrin-coated vesicles, and phagosomes. This movement is reproduced in a cell-free system containing extract of Xenopus oocytes. The formation of a comet-like structure capable of transporting vesicles from the plasma membrane into the cell depth has been studied in detail by high performance electron microscopy combined with electron tomography. A similar mechanism provides the movement of vesicles containing membrane rafts enriched with sphingolipids and cholesterol, changes in position of the nuclear spindle at meiosis, and other processes. This review will consider current ideas about actin polymerization and its regulation by actin-binding proteins and show how these mechanisms are realized in the intracellular actin-based vesicular transport system.

  2. Crk Adaptors Negatively Regulate Actin Polymerization in Pedestals Formed by Enteropathogenic Escherichia coli (EPEC) by Binding to Tir Effector

    PubMed Central

    Martín-Villa, José Manuel; Benito-León, María; Martinez-Quiles, Narcisa

    2014-01-01

    Infections by enteropathogenic Escherichia coli (EPEC) cause diarrhea linked to high infant mortality in developing countries. EPEC adheres to epithelial cells and induces the formation of actin pedestals. Actin polymerization is driven fundamentally through signaling mediated by Tir bacterial effector protein, which inserts in the plasma membrane of the infected cell. Tir binds Nck adaptor proteins, which in turn recruit and activate N-WASP, a ubiquitous member of the Wiskott-Aldrich syndrome family of proteins. N-WASP activates the Arp2/3 complex to promote actin polymerization. Other proteins aside from components of the Tir-Nck-N-WASP pathway are recruited to the pedestals but their functions are unknown. Here we investigate the function of two alternatively spliced isoforms of Crk adaptors (CrkI/II) and the paralog protein CrkL during pedestal formation by EPEC. We found that the Crk isoforms act as redundant inhibitors of pedestal formation. The SH2 domain of CrkII and CrkL binds to phosphorylated tyrosine 474 of Tir and competes with Nck to bind Tir, preventing its recruitment to pedestals and thereby inhibiting actin polymerization. EPEC infection induces phosphorylation of the major regulatory tyrosine in CrkII and CrkL, possibly preventing the SH2 domain of these proteins from interacting with Tir. Phosphorylated CrkII and CrkL proteins localize specifically to the plasma membrane in contact with EPEC. Our study uncovers a novel role for Crk adaptors at pedestals, opening a new perspective in how these oncoproteins regulate actin polymerization. PMID:24675776

  3. Crk adaptors negatively regulate actin polymerization in pedestals formed by enteropathogenic Escherichia coli (EPEC) by binding to Tir effector.

    PubMed

    Nieto-Pelegrin, Elvira; Meiler, Eugenia; Martín-Villa, José Manuel; Benito-León, María; Martinez-Quiles, Narcisa

    2014-03-01

    Infections by enteropathogenic Escherichia coli (EPEC) cause diarrhea linked to high infant mortality in developing countries. EPEC adheres to epithelial cells and induces the formation of actin pedestals. Actin polymerization is driven fundamentally through signaling mediated by Tir bacterial effector protein, which inserts in the plasma membrane of the infected cell. Tir binds Nck adaptor proteins, which in turn recruit and activate N-WASP, a ubiquitous member of the Wiskott-Aldrich syndrome family of proteins. N-WASP activates the Arp2/3 complex to promote actin polymerization. Other proteins aside from components of the Tir-Nck-N-WASP pathway are recruited to the pedestals but their functions are unknown. Here we investigate the function of two alternatively spliced isoforms of Crk adaptors (CrkI/II) and the paralog protein CrkL during pedestal formation by EPEC. We found that the Crk isoforms act as redundant inhibitors of pedestal formation. The SH2 domain of CrkII and CrkL binds to phosphorylated tyrosine 474 of Tir and competes with Nck to bind Tir, preventing its recruitment to pedestals and thereby inhibiting actin polymerization. EPEC infection induces phosphorylation of the major regulatory tyrosine in CrkII and CrkL, possibly preventing the SH2 domain of these proteins from interacting with Tir. Phosphorylated CrkII and CrkL proteins localize specifically to the plasma membrane in contact with EPEC. Our study uncovers a novel role for Crk adaptors at pedestals, opening a new perspective in how these oncoproteins regulate actin polymerization.

  4. Actin Pedestal Formation by Enterohemorrhagic Escherichia coli Enhances Bacterial Host Cell Attachment and Concomitant Type III Translocation

    PubMed Central

    Battle, Scott E.; Brady, Michael J.; Vanaja, Sivapriya Kailasan; Leong, John M.

    2014-01-01

    Attachment of enterohemorrhagic Escherichia coli (EHEC) to intestinal epithelial cells is critical for colonization and is associated with localized actin assembly beneath bound bacteria. The formation of these actin “pedestals” is dependent on the translocation of effectors into mammalian cells via a type III secretion system (T3SS). Tir, an effector required for pedestal formation, localizes in the host cell plasma membrane and promotes attachment of bacteria to mammalian cells by binding to the EHEC outer surface protein Intimin. Actin pedestal formation has been shown to foster intestinal colonization by EHEC in some animal models, but the mechanisms responsible for this remain undefined. Investigation of the role of Tir-mediated actin assembly promoting host cell binding is complicated by other, potentially redundant EHEC-encoded binding pathways, so we utilized cell binding assays that specifically detect binding mediated by Tir-Intimin interaction. We also assessed the role of Tir-mediated actin assembly in two-step assays that temporally segregated initial translocation of Tir from subsequent Tir-Intimin interaction, thereby permitting the distinction of effects on translocation from effects on cell attachment. In these experimental systems, we compromised Tir-mediated actin assembly by chemically inhibiting actin assembly or by infecting mammalian cells with EHEC mutants that translocate Tir but are specifically defective in Tir-mediated pedestal formation. We found that an inability of Tir to promote actin assembly resulted in a significant and striking decrease in bacterial binding mediated by Tir and Intimin. Bacterial mutants defective for pedestal formation translocated type III effectors to mammalian cells with reduced efficiency, but the decrease in translocation could be entirely accounted for by the decrease in host cell attachment. PMID:24958711

  5. Actin-Based Motility of Intracellular Microbial Pathogens

    PubMed Central

    Goldberg, Marcia B.

    2001-01-01

    A diverse group of intracellular microorganisms, including Listeria monocytogenes, Shigella spp., Rickettsia spp., and vaccinia virus, utilize actin-based motility to move within and spread between mammalian host cells. These organisms have in common a pathogenic life cycle that involves a stage within the cytoplasm of mammalian host cells. Within the cytoplasm of host cells, these organisms activate components of the cellular actin assembly machinery to induce the formation of actin tails on the microbial surface. The assembly of these actin tails provides force that propels the organisms through the cell cytoplasm to the cell periphery or into adjacent cells. Each of these organisms utilizes preexisting mammalian pathways of actin rearrangement to induce its own actin-based motility. Particularly remarkable is that while all of these microbes use the same or overlapping pathways, each intercepts the pathway at a different step. In addition, the microbial molecules involved are each distinctly different from the others. Taken together, these observations suggest that each of these microbes separately and convergently evolved a mechanism to utilize the cellular actin assembly machinery. The current understanding of the molecular mechanisms of microbial actin-based motility is the subject of this review. PMID:11729265

  6. Enterohemorrhagic E. coli Requires N-WASP for Efficient Type III Translocation but Not for EspFU-Mediated Actin Pedestal Formation

    PubMed Central

    Brady, Michael J.; Skehan, Brian; Battle, Scott E.; Robbins, Douglas; Kapoor, Archana; Hecht, Gail; Snapper, Scott B.; Leong, John M.

    2010-01-01

    Upon infection of mammalian cells, enterohemorrhagic E. coli (EHEC) O157:H7 utilizes a type III secretion system to translocate the effectors Tir and EspFU (aka TccP) that trigger the formation of F-actin-rich ‘pedestals’ beneath bound bacteria. EspFU is localized to the plasma membrane by Tir and binds the nucleation-promoting factor N-WASP, which in turn activates the Arp2/3 actin assembly complex. Although N-WASP has been shown to be required for EHEC pedestal formation, the precise steps in the process that it influences have not been determined. We found that N-WASP and actin assembly promote EHEC-mediated translocation of Tir and EspFU into mammalian host cells. When we utilized the related pathogen enteropathogenic E. coli to enhance type III translocation of EHEC Tir and EspFU, we found surprisingly that actin pedestals were generated on N-WASP-deficient cells. Similar to pedestal formation on wild type cells, Tir and EspFU were the only bacterial effectors required for pedestal formation, and the EspFU sequences required to interact with N-WASP were found to also be essential to stimulate this alternate actin assembly pathway. In the absence of N-WASP, the Arp2/3 complex was both recruited to sites of bacterial attachment and required for actin assembly. Our results indicate that actin assembly facilitates type III translocation, and reveal that EspFU, presumably by recruiting an alternate host factor that can signal to the Arp2/3 complex, exhibits remarkable versatility in its strategies for stimulating actin polymerization. PMID:20808845

  7. Nck adaptors, besides promoting N-WASP mediated actin-nucleation activity at pedestals, influence the cellular levels of enteropathogenic Escherichia coli Tir effector

    PubMed Central

    Nieto-Pelegrin, Elvira; Kenny, Brendan; Martinez-Quiles, Narcisa

    2014-01-01

    Enteropathogenic Escherichia coli (EPEC) binding to human intestinal cells triggers the formation of disease-associated actin rich structures called pedestals. The latter process requires the delivery, via a Type 3 secretion system, of the translocated Intimin receptor (Tir) protein into the host plasma membrane where binding of a host kinase-modified form to the bacterial surface protein Intimin triggers pedestal formation. Tir-Intimin interaction recruits the Nck adaptor to a Tir tyrosine phosphorylated residue where it activates neural Wiskott-Aldrich syndrome protein (N-WASP); initiating the major pathway to actin polymerization mediated by the actin-related protein (Arp) 2/3 complex. Previous studies with Nck-deficient mouse embryonic fibroblasts (MEFs) identified a key role for Nck in pedestal formation, presumed to reflect a lack of N-WASP activation. Here, we show the defect relates to reduced amounts of Tir within Nck-deficient cells. Indeed, Tir delivery and, thus, pedestal formation defects were much greater for MEFs than HeLa (human epithelial) cells. Crucially, the levels of two other effectors (EspB/EspF) within Nck-deficient MEFs were not reduced unlike that of Map (Mitochondrial associated protein) which, like Tir, requires CesT chaperone function for efficient delivery. Interestingly, drugs blocking various host protein degradation pathways failed to increase Tir cellular levels unlike an inhibitor of deacetylase activity (Trichostatin A; TSA). Treatments with TSA resulted in significant recovery of Tir levels, potentiation of actin polymerization and improvement in bacterial attachment to cells. Our findings have important implications for the current model of Tir-mediated actin polymerization and opens new lines of research in this area. PMID:25482634

  8. Nck adaptors, besides promoting N-WASP mediated actin-nucleation activity at pedestals, influence the cellular levels of enteropathogenic Escherichia coli Tir effector.

    PubMed

    Nieto-Pelegrin, Elvira; Kenny, Brendan; Martinez-Quiles, Narcisa

    2014-01-01

    Enteropathogenic Escherichia coli (EPEC) binding to human intestinal cells triggers the formation of disease-associated actin rich structures called pedestals. The latter process requires the delivery, via a Type 3 secretion system, of the translocated Intimin receptor (Tir) protein into the host plasma membrane where binding of a host kinase-modified form to the bacterial surface protein Intimin triggers pedestal formation. Tir-Intimin interaction recruits the Nck adaptor to a Tir tyrosine phosphorylated residue where it activates neural Wiskott-Aldrich syndrome protein (N-WASP); initiating the major pathway to actin polymerization mediated by the actin-related protein (Arp) 2/3 complex. Previous studies with Nck-deficient mouse embryonic fibroblasts (MEFs) identified a key role for Nck in pedestal formation, presumed to reflect a lack of N-WASP activation. Here, we show the defect relates to reduced amounts of Tir within Nck-deficient cells. Indeed, Tir delivery and, thus, pedestal formation defects were much greater for MEFs than HeLa (human epithelial) cells. Crucially, the levels of two other effectors (EspB/EspF) within Nck-deficient MEFs were not reduced unlike that of Map (Mitochondrial associated protein) which, like Tir, requires CesT chaperone function for efficient delivery. Interestingly, drugs blocking various host protein degradation pathways failed to increase Tir cellular levels unlike an inhibitor of deacetylase activity (Trichostatin A; TSA). Treatments with TSA resulted in significant recovery of Tir levels, potentiation of actin polymerization and improvement in bacterial attachment to cells. Our findings have important implications for the current model of Tir-mediated actin polymerization and opens new lines of research in this area.

  9. Insulin receptor tyrosine kinase substrate links the E. coli O157:H7 actin assembly effectors Tir and EspF(U) during pedestal formation.

    PubMed

    Vingadassalom, Didier; Kazlauskas, Arunas; Skehan, Brian; Cheng, Hui-Chun; Magoun, Loranne; Robbins, Douglas; Rosen, Michael K; Saksela, Kalle; Leong, John M

    2009-04-21

    Enterohemorrhagic Escherichia coli O157:H7 translocates 2 effectors to trigger localized actin assembly in mammalian cells, resulting in filamentous actin "pedestals." One effector, the translocated intimin receptor (Tir), is localized in the plasma membrane and clustered upon binding the bacterial outer membrane protein intimin. The second, the proline-rich effector EspF(U) (aka TccP) activates the actin nucleation-promoting factor WASP/N-WASP, and is recruited to sites of bacterial attachment by a mechanism dependent on an Asn-Pro-Tyr (NPY(458)) sequence in the Tir C-terminal cytoplasmic domain. Tir, EspF(U), and N-WASP form a complex, but neither EspF(U) nor N-WASP bind Tir directly, suggesting involvement of another protein in complex formation. Screening of the mammalian SH3 proteome for the ability to bind EspF(U) identified the SH3 domain of insulin receptor tyrosine kinase substrate (IRTKS), a factor known to regulate the cytoskeleton. Derivatives of WASP, EspF(U), and the IRTKS SH3 domain were capable of forming a ternary complex in vitro, and replacement of the C terminus of Tir with the IRTKS SH3 domain resulted in a fusion protein competent for actin assembly in vivo. A second domain of IRTKS, the IRSp53/MIM homology domain (IMD), bound to Tir in a manner dependent on the C-terminal NPY(458) sequence, thereby recruiting IRTKS to sites of bacterial attachment. Ectopic expression of either the IRTKS SH3 domain or the IMD, or genetic depletion of IRTKS, blocked pedestal formation. Thus, enterohemorrhagic E. coli translocates 2 effectors that bind to distinct domains of a common host factor to promote the formation of a complex that triggers robust actin assembly at the plasma membrane.

  10. Microtubule and Actin Interplay Drive Intracellular c-Src Trafficking.

    PubMed

    Arnette, Christopher; Frye, Keyada; Kaverina, Irina

    2016-01-01

    The proto-oncogene c-Src is involved in a variety of signaling processes. Therefore, c-Src spatiotemporal localization is critical for interaction with downstream targets. However, the mechanisms regulating this localization have remained elusive. Previous studies have shown that c-Src trafficking is a microtubule-dependent process that facilitates c-Src turnover in neuronal growth cones. As such, microtubule depolymerization lead to the inhibition of c-Src recycling. Alternatively, c-Src trafficking was also shown to be regulated by RhoB-dependent actin polymerization. Our results show that c-Src vesicles primarily exhibit microtubule-dependent trafficking; however, microtubule depolymerization does not inhibit vesicle movement. Instead, vesicular movement becomes both faster and less directional. This movement was associated with actin polymerization directly at c-Src vesicle membranes. Interestingly, it has been shown previously that c-Src delivery is an actin polymerization-dependent process that relies on small GTPase RhoB at c-Src vesicles. In agreement with this finding, microtubule depolymerization induced significant activation of RhoB, together with actin comet tail formation. These effects occurred downstream of GTP-exchange factor, GEF-H1, which was released from depolymerizing MTs. Accordingly, GEF-H1 activity was necessary for actin comet tail formation at the Src vesicles. Our results indicate that regulation of c-Src trafficking requires both microtubules and actin polymerization, and that GEF-H1 coordinates c-Src trafficking, acting as a molecular switch between these two mechanisms.

  11. Non-lytic, actin-based exit of intracellular parasites from C. elegans intestinal cells.

    PubMed

    Estes, Kathleen A; Szumowski, Suzannah C; Troemel, Emily R

    2011-09-01

    The intestine is a common site for invasion by intracellular pathogens, but little is known about how pathogens restructure and exit intestinal cells in vivo. The natural microsporidian parasite N. parisii invades intestinal cells of the nematode C. elegans, progresses through its life cycle, and then exits cells in a transmissible spore form. Here we show that N. parisii causes rearrangements of host actin inside intestinal cells as part of a novel parasite exit strategy. First, we show that N. parisii infection causes ectopic localization of the normally apical-restricted actin to the basolateral side of intestinal cells, where it often forms network-like structures. Soon after this actin relocalization, we find that gaps appear in the terminal web, a conserved cytoskeletal structure that could present a barrier to exit. Reducing actin expression creates terminal web gaps in the absence of infection, suggesting that infection-induced actin relocalization triggers gap formation. We show that terminal web gaps form at a distinct stage of infection, precisely timed to precede spore exit, and that all contagious animals exhibit gaps. Interestingly, we find that while perturbations in actin can create these gaps, actin is not required for infection progression or spore formation, but actin is required for spore exit. Finally, we show that despite large numbers of spores exiting intestinal cells, this exit does not cause cell lysis. These results provide insight into parasite manipulation of the host cytoskeleton and non-lytic escape from intestinal cells in vivo.

  12. Intracellular calcium rise is not a necessary step for the stimulated actin polymerization

    SciTech Connect

    Yassin, R.

    1986-03-01

    Stimulation of rabbit peritoneal neutrophils by many chemotactic (formyl Methionyl-Leucyl-Phenylalanine (fMLP), Leukotriene B/sub 4/ (LTB/sub 4/)) and non-chemotactic (phorbol 12-myristate, 13-acetate (PMA), platelet activating factor (PAF), and the calcium ionophore A23187) factors produces rapid and dose dependent increases in the amount of actin associated with the cytoskeleton. The stimulated increase in cytoskeletal actin does not appear to require a rise in the intracellular concentration of free calcium. The increase in cytoskeletal actin produced by A23187 is transient and does not depend on the presence of calcium in the suspending medium. In the presence of extracellular calcium, the effect of the ionophore is biphasic with respect to concentration. The increases in actin association with cytoskeletal produced by fMLP, LTB/sub 4/, and A23187 but not by PMA, are inhibited by hyperosmolarity and pertussis toxin pretreatment. On the other hand, the addition of hyperosmolarity or pertussis toxin has small effect on the rise in the intracellular calcium produced by A23187. The results presented here suggest that an increase in the intracellular concentration of free calcium is not necessary for the stimulated increases in cytoskeletal actin.

  13. Viral infectivity and intracellular distribution of matrix (M) protein of canine distemper virus are affected by actin filaments.

    PubMed

    Klauschies, F; Gützkow, T; Hinkelmann, S; von Messling, V; Vaske, B; Herrler, G; Haas, L

    2010-09-01

    To investigate the role of cytoskeletal components in canine distemper virus (CDV) replication, various agents were used that interfere with turnover of actin filaments and microtubules. Only inhibition of actin filaments significantly reduced viral infectivity. Analysis of the intracellular localization of the viral matrix (M) protein revealed that it aligned along actin filaments. Treatment with actin filament-disrupting drugs led to a marked intracellular redistribution of M protein during infection as well as transfection. In contrast, the localization of the CDV fusion (F) protein was not significantly changed during transfection. Thus, a M protein-actin filament interaction appears to be important for generation of infectious CDV.

  14. Pedestal Grinder.

    ERIC Educational Resources Information Center

    Engelbrecht, Nancy; And Others

    These instructional materials provide an orientation to the pedestal grinder for use at the postsecondary level. The first of eight sections defines 14 important terms. The second section outlines 16 rules for safe use of the pedestal grinder. The third section covers grinding wheels for five different types of materials. The fourth section…

  15. Hijacking Host Cell Highways: Manipulation of the Host Actin Cytoskeleton by Obligate Intracellular Bacterial Pathogens.

    PubMed

    Colonne, Punsiri M; Winchell, Caylin G; Voth, Daniel E

    2016-01-01

    Intracellular bacterial pathogens replicate within eukaryotic cells and display unique adaptations that support key infection events including invasion, replication, immune evasion, and dissemination. From invasion to dissemination, all stages of the intracellular bacterial life cycle share the same three-dimensional cytosolic space containing the host cytoskeleton. For successful infection and replication, many pathogens hijack the cytoskeleton using effector proteins introduced into the host cytosol by specialized secretion systems. A subset of effectors contains eukaryotic-like motifs that mimic host proteins to exploit signaling and modify specific cytoskeletal components such as actin and microtubules. Cytoskeletal rearrangement promotes numerous events that are beneficial to the pathogen, including internalization of bacteria, structural support for bacteria-containing vacuoles, altered vesicular trafficking, actin-dependent bacterial movement, and pathogen dissemination. This review highlights a diverse group of obligate intracellular bacterial pathogens that manipulate the host cytoskeleton to thrive within eukaryotic cells and discusses underlying molecular mechanisms that promote these dynamic host-pathogen interactions.

  16. Host actin polymerization tunes the cell division cycle of an intracellular pathogen.

    PubMed

    Siegrist, M Sloan; Aditham, Arjun K; Espaillat, Akbar; Cameron, Todd A; Whiteside, Sarah A; Cava, Felipe; Portnoy, Daniel A; Bertozzi, Carolyn R

    2015-04-28

    Growth and division are two of the most fundamental capabilities of a bacterial cell. While they are well described for model organisms growing in broth culture, very little is known about the cell division cycle of bacteria replicating in more complex environments. Using a D-alanine reporter strategy, we found that intracellular Listeria monocytogenes (Lm) spend a smaller proportion of their cell cycle dividing compared to Lm growing in broth culture. This alteration to the cell division cycle is independent of bacterial doubling time. Instead, polymerization of host-derived actin at the bacterial cell surface extends the non-dividing elongation period and compresses the division period. By decreasing the relative proportion of dividing Lm, actin polymerization biases the population toward cells with the highest propensity to form actin tails. Thus, there is a positive-feedback loop between the Lm cell division cycle and a physical interaction with the host cytoskeleton.

  17. Hijacking Host Cell Highways: Manipulation of the Host Actin Cytoskeleton by Obligate Intracellular Bacterial Pathogens

    PubMed Central

    Colonne, Punsiri M.; Winchell, Caylin G.; Voth, Daniel E.

    2016-01-01

    Intracellular bacterial pathogens replicate within eukaryotic cells and display unique adaptations that support key infection events including invasion, replication, immune evasion, and dissemination. From invasion to dissemination, all stages of the intracellular bacterial life cycle share the same three-dimensional cytosolic space containing the host cytoskeleton. For successful infection and replication, many pathogens hijack the cytoskeleton using effector proteins introduced into the host cytosol by specialized secretion systems. A subset of effectors contains eukaryotic-like motifs that mimic host proteins to exploit signaling and modify specific cytoskeletal components such as actin and microtubules. Cytoskeletal rearrangement promotes numerous events that are beneficial to the pathogen, including internalization of bacteria, structural support for bacteria-containing vacuoles, altered vesicular trafficking, actin-dependent bacterial movement, and pathogen dissemination. This review highlights a diverse group of obligate intracellular bacterial pathogens that manipulate the host cytoskeleton to thrive within eukaryotic cells and discusses underlying molecular mechanisms that promote these dynamic host-pathogen interactions. PMID:27713866

  18. Intracellular photoactivation of caged cGMP induces myosin II and actin responses in motile cells.

    PubMed

    Pfannes, Eva K B; Anielski, Alexander; Gerhardt, Matthias; Beta, Carsten

    2013-12-01

    Cyclic GMP (cGMP) is a ubiquitous second messenger in eukaryotic cells. It is assumed to regulate the association of myosin II with the cytoskeleton of motile cells. When cells of the social amoeba Dictyostelium discoideum are exposed to chemoattractants or to increased osmotic stress, intracellular cGMP levels rise, preceding the accumulation of myosin II in the cell cortex. To directly investigate the impact of intracellular cGMP on cytoskeletal dynamics in a living cell, we released cGMP inside the cell by laser-induced photo-cleavage of a caged precursor. With this approach, we could directly show in a live cell experiment that an increase in intracellular cGMP indeed induces myosin II to accumulate in the cortex. Unexpectedly, we observed for the first time that also the amount of filamentous actin in the cell cortex increases upon a rise in the cGMP concentration, independently of cAMP receptor activation and signaling. We discuss our results in the light of recent work on the cGMP signaling pathway and suggest possible links between cGMP signaling and the actin system.

  19. Insulin stimulates actin comet tails on intracellular GLUT4-containing compartments in differentiated 3T3L1 adipocytes.

    PubMed

    Kanzaki, M; Watson, R T; Khan, A H; Pessin, J E

    2001-12-28

    Incubation of isolated GLUT4-containing vesicles with Xenopus oocyte extracts resulted in a guanosine 5'-[gamma-thio]triphosphate (GTP gamma S) and sodium orthovanadate stimulation of actin comet tails. The in vitro actin-based GLUT4 vesicle motility was inhibited by both latrunculin B and a dominant-interfering N-WASP mutant, N-WASP/Delta VCA. Preparations of gently sheared (broken) 3T3L1 adipocytes also displayed GTP gamma S and sodium orthovanadate stimulation of actin comet tails on GLUT4 intracellular compartments. Furthermore, insulin pretreatment of intact adipocytes prior to gently shearing also resulted in a marked increase in actin polymerization and actin comet tailing on GLUT4 vesicles. In addition, the insulin stimulation of actin comet tails was completely inhibited by Clostridum difficile toxin B, demonstrating a specific role for a Rho family member small GTP-binding protein. Expression of N-WASP/Delta VCA in intact cells had little effect on adipocyte cortical actin but partially inhibited insulin-stimulated GLUT4 translocation. Taken together, these data demonstrate that insulin can induce GLUT4 vesicle actin comet tails that are necessary for the efficient translocation of GLUT4 from intracellular storage sites to the plasma membrane.

  20. Intracellular Theileria annulata Promote Invasive Cell Motility through Kinase Regulation of the Host Actin Cytoskeleton

    PubMed Central

    Ma, Min; Baumgartner, Martin

    2014-01-01

    The intracellular, protozoan Theileria species parasites are the only eukaryotes known to transform another eukaryotic cell. One consequence of this parasite-dependent transformation is the acquisition of motile and invasive properties of parasitized cells in vitro and their metastatic dissemination in the animal, which causes East Coast Fever (T. parva) or Tropical Theileriosis (T. annulata). These motile and invasive properties of infected host cells are enabled by parasite-dependent, poorly understood F-actin dynamics that control host cell membrane protrusions. Herein, we dissected functional and structural alterations that cause acquired motility and invasiveness of T. annulata-infected cells, to understand the molecular basis driving cell dissemination in Tropical Theileriosis. We found that chronic induction of TNFα by the parasite contributes to motility and invasiveness of parasitized host cells. We show that TNFα does so by specifically targeting expression and function of the host proto-oncogenic ser/thr kinase MAP4K4. Blocking either TNFα secretion or MAP4K4 expression dampens the formation of polar, F-actin-rich invasion structures and impairs cell motility in 3D. We identified the F-actin binding ERM family proteins as MAP4K4 downstream effectors in this process because TNFα-induced ERM activation and cell invasiveness are sensitive to MAP4K4 depletion. MAP4K4 expression in infected cells is induced by TNFα-JNK signalling and maintained by the inhibition of translational repression, whereby both effects are parasite dependent. Thus, parasite-induced TNFα promotes invasive motility of infected cells through the activation of MAP4K4, an evolutionary conserved kinase that controls cytoskeleton dynamics and cell motility. Hence, MAP4K4 couples inflammatory signaling to morphodynamic processes and cell motility, a process exploited by the intracellular Theileria parasite to increase its host cell's dissemination capabilities. PMID:24626571

  1. Dual chemotaxis signalling regulates Dictyostelium development: intercellular cyclic AMP pulses and intracellular F-actin disassembly waves induce each other.

    PubMed

    Vicker, Michael G; Grutsch, James F

    2008-10-01

    Aggregating Dictyostelium discoideum amoebae periodically emit and relay cAMP, which regulates their chemotaxis and morphogenesis into a multicellular, differentiated organism. Cyclic AMP also stimulates F-actin assembly and chemotactic pseudopodium extension. We used actin-GFP expression to visualise for the first time intracellular F-actin assembly as a spatio-temporal indicator of cell reactions to cAMP, and thus the kinematics of cell communication, in aggregating streams. Every natural cAMP signal pulse induces an autowave of F-actin disassembly, which propagates from each cell's leading end to its trailing end at a linear rate, much slower than the calculated and measured velocities of cAMP diffusion in aggregating Dictyostelium. A sequence of transient reactions follows behind the wave, including anterior F-actin assembly, chemotactic pseudopodium extension and cell advance at the cell front and, at the back, F-actin assembly, extension of a small retrograde pseudopodium (forcing a brief cell retreat) and chemotactic stimulation of the following cell, yielding a 20s cAMP relay delay. These dynamics indicate that stream cell behaviour is mediated by a dual signalling system: a short-range cAMP pulse directed from one cell tail to an immediately following cell front and a slower, long-range wave of intracellular F-actin disassembly, each inducing the other.

  2. Calcium inhibition as an intracellular signal for actin-myosin interaction.

    PubMed

    Kohama, Kazuhiro

    2016-01-01

    Intracellular signaling pathways include both the activation and the inhibition of biological processes. The activation of Ca(2+) regulation of actin-myosin interactions was examined first, whereas it took 20 years for the author to clarify the inhibitory mode by using Physarum polycephalum, a lower eukaryote. This review describes the investigation of the inhibitory mode since 1980. The inhibitory effect of Ca(2+) on myosin was detected chemically by ATPase assays and mechanically by in vitro motility assays. The Ca(2+)-binding ability of Physarum myosin is as high as that of scallop myosin. Ca(2+) inhibits Physarum myosin, whereas it activates scallop myosin. We cloned cDNA of the myosin heavy chain and light chains to express a hybrid of Physarum and scallop myosin, and found that the Ca-binding light chain (CaLc), which belongs to an alkali light chain class, plays a major role in Ca inhibition. The role of CaLc was confirmed by mutating its EF-hand, Ca-binding structure and expressing Physarum myosin as a recombinant protein. Thus, the data obtained by classical protein purification were confirmed by the results obtained with the modern recombinant techniques. However, there are some discrepancies that remain to be solved as described in Section XII.

  3. Effects of Roundup and glyphosate formulations on intracellular transport, microtubules and actin filaments in Xenopus laevis melanophores.

    PubMed

    Hedberg, Daniel; Wallin, Margareta

    2010-04-01

    Glyphosate containing herbicides, such as Roundup, are commonly used and generally considered to be safe. However, some toxic effects are found on amphibians in vivo and human and mouse cells in vitro. In this study the effects of Roundup, glyphosate, glyphosateisopropylamine and isopropylamine were studied on intracellular transport by measuring aggregation capacity in Xenopus laevis melanophores. The chemicals inhibited retrograde transport of melanosomes in the range of 0.5-5mM. Cellular morphology and localization of microtubules and actin filaments were affected as determined by immunocytochemistry. Both glyphosate and Roundup decreased pH in the media. Acidic pH inhibited melanosome transport and altered microtubule and actin morphology in the absence of chemicals, while transport inhibiting concentrations of glyphosate, Roundup and glyphosateisopropylamine disassembled both microtubules and actin filaments. At physiological pH the effects of Roundup decreased whereas glyphosate failed to inhibit transport. Physiological pH decreases glyphosate lipophilicity and its diffusion into the cytoplasm. The Roundup formulation contains surfactants, such as POEA (polyetylated tallow amine) that increases membrane permeability allowing cellular uptake at physiological pH. Our results show that the effects of glyphosate containing compounds are pH-dependent and that they inhibit intracellular transport through disassembly of the cytoskeleton possibly by interfering with intracellular Ca(2+)-balance. Copyright (c) 2010 Elsevier Ltd. All rights reserved.

  4. Bio-mimetic surface engineering of plasmid-loaded nanoparticles for active intracellular trafficking by actin comet-tail motility.

    PubMed

    Ng, Chee Ping; Goodman, Thomas T; Park, In-Kyu; Pun, Suzie H

    2009-02-01

    Intracellular transport after endosomal escape presents one of the major barriers for efficient non-viral gene delivery because plasmid DNA and synthetic nanoparticulate carriers suffer from significantly restricted diffusion in the cytoplasm. We postulate that forces generated by actin polymerization, a mechanism used by several bacterial pathogens such as Listeria monocytogenes, can be harnessed to propel nanoparticles within the cytoplasm and thereby overcome diffusional limitations associated with gene transport in the cell cytoplasm. In this work, we synthesized and characterized plasmid DNA-containing nanoparticles modified with ActA protein, the single protein in L. monocytogenes responsible for activating actin polymerization and initiating actin comet-tail propulsion. The motility of the ActA-modified nanoparticles was assessed in Xenopus laevis cytoplasmic extract supplemented with fluorescently labeled actin. Nanoparticle motility was monitored using multi-color, time-lapse fluorescence microscopy for the formation of actin comet tails attached to the fluorescently labeled vehicle. We observed particle motility with velocities approximately 0.06 microm/s with anionic-charged plasmid carriers formed from either poly(lactic-co-glycolic acid) (PLGA) or 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) liposomes, but interestingly not with cationic particles assembled by encapsulation of plasmid with either polyethylenimine (PEI) or 1,2-dioleoyl-3-trimethylammonium-propane/1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOTAP/DOPE) lipids. Control particles coated with albumin instead of ActA also showed no motility. Taken together, we have demonstrated the feasibility of translating the comet-tail propulsion mechanism to synthetic drug carriers as a potential approach to overcome intracellular transport barriers, and also have identified appropriate gene delivery systems that can be employed for this mechanism.

  5. Pedestal stability comparison and ITER pedestal prediction

    NASA Astrophysics Data System (ADS)

    Snyder, P. B.; Aiba, N.; Beurskens, M.; Groebner, R. J.; Horton, L. D.; Hubbard, A. E.; Hughes, J. W.; Huysmans, G. T. A.; Kamada, Y.; Kirk, A.; Konz, C.; Leonard, A. W.; Lönnroth, J.; Maggi, C. F.; Maingi, R.; Osborne, T. H.; Oyama, N.; Pankin, A.; Saarelma, S.; Saibene, G.; Terry, J. L.; Urano, H.; Wilson, H. R.

    2009-08-01

    The pressure at the top of the edge transport barrier (or 'pedestal height') strongly impacts fusion performance, while large edge localized modes (ELMs), driven by the free energy in the pedestal region, can constrain material lifetimes. Accurately predicting the pedestal height and ELM behavior in ITER is an essential element of prediction and optimization of fusion performance. Investigation of intermediate wavelength MHD modes (or 'peeling-ballooning' modes) has led to an improved understanding of important constraints on the pedestal height and the mechanism for ELMs. The combination of high-resolution pedestal diagnostics, including substantial recent improvements, and a suite of highly efficient stability codes, has made edge stability analysis routine on several major tokamaks, contributing both to understanding, and to experimental planning and performance optimization. Here we present extensive comparisons of observations to predicted edge stability boundaries on several tokamaks, both for the standard (Type I) ELM regime, and for small ELM and ELM-free regimes. We further discuss a new predictive model for the pedestal height and width (EPED1), developed by self-consistently combining a simple width model with peeling-ballooning stability calculations. This model is tested against experimental measurements, and used in initial predictions of the pedestal height for ITER.

  6. Pedestal stability comparison and ITER pedestal prediction

    SciTech Connect

    Snyder, P.; Alba, N; Beurskens, M.; Horton, L D

    2009-01-01

    The pressure at the top of the edge transport barrier (or 'pedestal height') strongly impacts fusion performance, while large edge localized modes (ELMs), driven by the free energy in the pedestal region, can constrain material lifetimes. Accurately predicting the pedestal height and ELM behavior in ITER is an essential element of prediction and optimization of fusion performance. Investigation of intermediate wavelength MHD modes (or 'peeling ballooning' modes) has led to an improved understanding of important constraints on the pedestal height and the mechanism for ELMs. The combination of high-resolution pedestal diagnostics, including substantial recent improvements, and a suite of highly efficient stability codes, has made edge stability analysis routine on several major tokamaks, contributing both to understanding, and to experimental planning and performance optimization. Here we present extensive comparisons of observations to predicted edge stability boundaries on several tokamaks, both for the standard (Type I) ELM regime, and for small ELM and ELM-free regimes. We further discuss a new predictive model for the pedestal height and width (EPED1), developed by self-consistently combining a simple width model with peeling-ballooning stability calculations. This model is tested against experimental measurements, and used in initial predictions of the pedestal height for ITER.

  7. Actin-based mechanisms for light-dependent intracellular positioning of nuclei and chloroplasts in Arabidopsis.

    PubMed

    Iwabuchi, Kosei; Takagi, Shingo

    2010-08-01

    The plant organelles, chloroplast and nucleus, change their position in response to light. In Arabidopsis thaliana leaf cells, chloroplasts and nuclei are distributed along the inner periclinal wall in darkness. In strong blue light, they become positioned along the anticlinal wall, while in weak blue light, only chloroplasts are accumulated along the inner and outer periclinal walls. Blue-light dependent positioning of both organelles is mediated by the blue-light receptor phototropin and controlled by the actin cytoskeleton. Interestingly, however, it seems that chloroplast movement requires short, fine actin filaments organized at the chloroplast edge, whereas nuclear movement does cytoplasmic, thick actin bundles intimately associated with the nucleus. Although there are many similarities between photo-relocation movements of chloroplasts and nuclei, plant cells appear to have evolved distinct mechanisms to regulate actin organization required for driving the movements of these organelles.

  8. Actin-based motility of the intracellular pathogen Listeria monocytogenes: assessing the inhibitory specificity of ABM-1 peptide analogues.

    PubMed

    Purich, D L; Southwick, F S

    1999-06-01

    Actin-Based Motility motifs [ABM-1 sequence = (D/E)FPPPPX(D/E), where X = P or T, and ABM-2 sequence = XPPPPP, where X denotes G, A, L, P, and S] facilitate assembly of an activated motility complex. Potent inhibition of intracellular motility of pathogens by ABM-1 and ABM-2 peptide analogues has served as a criterion for investigating actin-based motility. To assess the specificity of ABM-1 peptide inhibitors, we microinjected proline-rich peptides into Listeria-infected PtK2 host cells. Use of a combinatorial ABM-1 peptide library (empirical formula = D1E2F2P4T1) demonstrated that high-potency inhibition requires a precise sequence, and not merely a particular amino acid composition. Calculated concentrations of specific sequences in this library indicate that the entire (D/E)FPPPPX(D/E) motif is needed to achieve high-affinity inhibition in living cells. The failure of the well known proline-rich SH3 binding antagonists VSL-12 or APP-12 to inhibit Listeria motility also indicates that SH3 interactions are unlikely to control actin-based motility directly.

  9. Pedestal Crater Development

    NASA Image and Video Library

    2015-07-01

    In this image, we see an approximately 500-meter crater that is fairly fresh (in geological terms), but the ejecta is already high-standing. Could this be an indication of early stage of pedestal development? A pedestal crater is when the ejecta from an impact settles around the new crater and is more erosion-resistant than the surrounding terrain. Over time, the surrounding terrain erodes much faster than the ejecta; in fact, some pedestal craters are measured to be hundreds of meters above the surrounding area. HiRISE has imaged many other pedestal craters before, and the ejecta isn't always symmetrical, as in this observation. http://photojournal.jpl.nasa.gov/catalog/PIA19849

  10. Effect of the Fructus Ligustri Lucidi extract and its monomers quercetin and oleanolic acid on the adhesion and migration of melanocytes and intracellular actin

    PubMed Central

    WU, YANHUA; LI, QILIN; LI, XIANGJUN; HE, DANHUA; NIU, MU; LU, XIAOJUAN; LI, HUI

    2016-01-01

    The present study aimed to investigate the effects of the Fructus Ligustri Lucidi (FLL) extract and its monomers quercetin and oleanolic acid on the adhesion and migration of human epidermal melanocytes (MCs) and intracellular actin. The human epidermal MCs were cultured and identified. The cells were treated with different concentrations of FLL extract, quercetin and oleanolic acid. The adhesion and migration abilities of the cells were determined by the fibronectin-coated culture experiment and Transwell assay, respectively. The structure and distribution of intracellular actin were observed by confocal laser microscopy, with semi-quantitative analysis. Results showed that compared with the control group, 0.0375–0.3 mg/ml of the FLL extract and 40 µM quercetin significantly improved the adhesion rate of MCs (P<0.05). The numbers of MCs permeating the microporous membrane in the 0.15 mg/ml FLL extract and 12 µM oleanolic acid groups were 43.7 and 30.3, respectively, significantly higher compared to the control group (P<0.01). In the control group, the intracellular actin was less, and the stress fiber structure was not clear. In the 0.15 mg/ml FLL extract, 12 µM oleanolic acid and 40 µM quercetin groups, there were numerous bunched stress fibers, indicating the aggregation of filamentous fibrous actin. The mean optical densities of actin expression in the 0.15 mg/ml FLL extract, 12 µM oleanolic acid and 40 µM quercetin groups were significantly higher compared to the control group (P<0.05). The FLL extract has a significant stimulatory effect on the adhesion and migration of human epidermal MCs. The mechanism may be associated with the promotion of intracellular actin cytoskeleton aggregation. PMID:27123251

  11. Transition to superdiffusive behavior in intracellular actin-based transport mediated by molecular motors

    NASA Astrophysics Data System (ADS)

    Bruno, L.; Levi, V.; Brunstein, M.; Despósito, M. A.

    2009-07-01

    Intracellular transport of large cargoes, such as organelles, vesicles, or large proteins, is a complex dynamical process that involves the interplay of adenosine triphosphate-consuming molecular motors, cytoskeleton filaments, and the viscoelastic cytoplasm. In this work we investigate the motion of pigment organelles (melanosomes) driven by myosin-V motors in Xenopus laevis melanocytes using a high-spatio-temporal resolution tracking technique. By analyzing the obtained trajectories, we show that the melanosomes mean-square displacement undergoes a transition from a subdiffusive to a superdiffusive behavior. A stochastic theoretical model, which explicitly considers the collective action of the molecular motors, is introduced to generalize the interpretation of our data. Starting from a generalized Langevin equation, we derive an analytical expression for the mean square displacement, which also takes into account the experimental noise. By fitting theoretical expressions to experimental data we were able to discriminate the exponents that characterize the passive and active contributions to the dynamics and to estimate the “global” motor forces correctly. Then, our model gives a quantitative description of active transport in living cells with a reduced number of parameters.

  12. Transition to superdiffusive behavior in intracellular actin-based transport mediated by molecular motors.

    PubMed

    Bruno, L; Levi, V; Brunstein, M; Despósito, M A

    2009-07-01

    Intracellular transport of large cargoes, such as organelles, vesicles, or large proteins, is a complex dynamical process that involves the interplay of adenosine triphosphate-consuming molecular motors, cytoskeleton filaments, and the viscoelastic cytoplasm. In this work we investigate the motion of pigment organelles (melanosomes) driven by myosin-V motors in Xenopus laevis melanocytes using a high-spatio-temporal resolution tracking technique. By analyzing the obtained trajectories, we show that the melanosomes mean-square displacement undergoes a transition from a subdiffusive to a superdiffusive behavior. A stochastic theoretical model, which explicitly considers the collective action of the molecular motors, is introduced to generalize the interpretation of our data. Starting from a generalized Langevin equation, we derive an analytical expression for the mean square displacement, which also takes into account the experimental noise. By fitting theoretical expressions to experimental data we were able to discriminate the exponents that characterize the passive and active contributions to the dynamics and to estimate the "global" motor forces correctly. Then, our model gives a quantitative description of active transport in living cells with a reduced number of parameters.

  13. Pedestal Crater and Yardangs

    NASA Technical Reports Server (NTRS)

    2003-01-01

    MGS MOC Release No. MOC2-444, 6 August 2003

    This April 2003 Mars Global Surveyor (MGS) Mars Orbiter Camera (MOC) image shows a small meteor impact crater that has been modified by wind erosion. Two things happened after the crater formed. First, the upper few meters of surface material into which the meteor impacted was later eroded away by wind. The crater ejecta formed a protective armor that kept the material under the ejecta from been blown away. This caused the crater and ejecta to appear as if standing upon a raised platform--a feature that Mars geologists call a pedestal crater. Next, the pedestal crater was buried beneath several meters of new sediment, and then this material was eroded away by wind to form the array of sharp ridges that run across the pedestal crater's surface. These small ridges are known as yardangs. This picture is illuminated by sunlight from the upper left; it is located in west Daedalia Planum near 14.6oS, 131.9oW.

  14. Magnetohydrodynamic mechanism for pedestal formation.

    PubMed

    Guazzotto, L; Betti, R

    2011-09-16

    Time-dependent two-dimensional magnetohydrodynamic simulations are carried out for tokamak plasmas with edge poloidal flow. Differently from conventional equilibrium theory, a density pedestal all around the edge is obtained when the poloidal velocity exceeds the poloidal sound speed. The outboard pedestal is induced by the transonic discontinuity, the inboard one by mass redistribution. The density pedestal follows the formation of a highly sheared flow at the transonic surface. These results may be relevant to the L-H transition and pedestal formation in high performance tokamak plasmas.

  15. Microinstabilities in the pedestal region

    NASA Astrophysics Data System (ADS)

    Dickinson, David; Dudson, Benjamin; Wilson, Howard; Roach, Colin

    2014-10-01

    The regulation of transport at the pedestal top is important for the inter-ELM pedestal dynamics. Linear gyrokinetic analysis of the pedestal region during an ELM cycle on MAST has shown kinetic ballooning modes to be unstable at the knee of the pressure profile and in the steep pedestal region whilst microtearing modes (MTMs) dominate in the shallow gradient region inboard of the pedestal top. The transition between these instabilities at the pedestal knee has been observed in low and high collisionality MAST pedestals, and is likely to play an important role in the broadening of the pedestal. Nonlinear simulations are needed in this region to understand the microturbulence, the corresponding transport fluxes, and to gain further insight into the processes underlying the pedestal evolution. Such gyrokinetic simulations are numerically challenging and recent upgrades to the GS2 gyrokinetic code help improve their feasibility. We are also exploring reduced models that capture the relevant physics using the plasma simulation framework BOUT + + . An electromagnetic gyrofluid model has recently been implemented with BOUT + + that has significantly reduced computational cost compared to the gyrokinetic simulations against which it will be benchmarked. This work was funded by the RCUK Energy programme, EURATOM and a EUROFusion fellowship WP14-FRF-CCFE/Dickinson and was carried out using: HELIOS at IFERC, Japan; ARCHER (EPSRC Grant No. EP/L000237/1); HECToR (EPSRC Grant No. EP/H002081/1).

  16. Burkholderia pseudomallei type III secreted protein BipC: role in actin modulation and translocation activities required for the bacterial intracellular lifecycle

    PubMed Central

    Kang, Wen Tyng; Vellasamy, Kumutha Malar; Rajamani, Lakshminarayanan; Beuerman, Roger W.

    2016-01-01

    Melioidosis, an infection caused by the facultative intracellular pathogen Burkholderia pseudomallei, has been classified as an emerging disease with the number of patients steadily increasing at an alarming rate. B. pseudomalleipossess various virulence determinants that allow them to invade the host and evade the host immune response, such as the type III secretion systems (TTSS). The products of this specialized secretion system are particularly important for the B. pseudomallei infection. Lacking in one or more components of the TTSS demonstrated different degrees of defects in the intracellular lifecycle of B. pseudomallei. Further understanding the functional roles of proteins involved in B. pseudomallei TTSS will enable us to dissect the enigma of B. pseudomallei-host cell interaction. In this study, BipC (a translocator), which was previously reported to be involved in the pathogenesis of B. pseudomallei, was further characterized using the bioinformatics and molecular approaches. The bipCgene, coding for a putative invasive protein, was first PCR amplified from B. pseudomallei K96243 genomic DNA and cloned into an expression vector for overexpression in Escherichia coli. The soluble protein was subsequently purified and assayed for actin polymerization and depolymerization. BipC was verified to subvert the host actin dynamics as demonstrated by the capability to polymerize actin in vitro. Homology modeling was also attempted to predict the structure of BipC. Overall, our findings identified that the protein encoded by the bipC gene plays a role as an effector involved in the actin binding activity to facilitate internalization of B. pseudomalleiinto the host cells. PMID:28028452

  17. Paleoclassical Model of Pedestal Structure

    NASA Astrophysics Data System (ADS)

    Callen, J. D.

    2010-11-01

    Predictions are developed for the structure of plasma parameter profiles of H-mode pedestals in transport quasi-equilibrium in tokamak plasmas. They are based on assuming paleoclassical radial plasma transport processes dominate throughout the pedestal. The natural level of paleoclassical density transport is large in the pedestal compared to edge fueling due to neutral recycling. Thus, in this model the pedestal density profile is determined not by edge source fueling but rather by the density profile needed for the outward paleoclassical diffusive flux to be nearly balanced by the inward paleoclassical pinch flow. Specific predictions are given for the electron temperature and density gradients, profiles and magnitudes in the pedestal. The transition into ETG-driven anomalous radial electron heat transport in the core plasma determines the height of the electron pressure pedestal. Also, the profile of the toroidal plasma rotation in the pedestal is predicted. Model predictions are found to agree quantitatively (within a factor of 2) with the interpretive transport results obtained for the 98889 DIII-D pedestal [1]. 6pt[1] J.D. Callen et al., Nucl. Fusion 50, 064004 (2010).

  18. Tir Is Essential for the Recruitment of Tks5 to Enteropathogenic Escherichia coli Pedestals

    PubMed Central

    Jensen, Helene H.; Pedersen, Hans N.; Stenkjær, Eva; Pedersen, Gitte A.; Login, Frédéric H.; Nejsum, Lene N.

    2015-01-01

    Enteropathogenic Escherichia coli (EPEC) is a bacterial pathogen that infects the epithelial lining of the small intestine and causes diarrhea. Upon attachment to the intestinal epithelium, EPEC uses a Type III Secretion System to inject its own high affinity receptor Translocated intimin receptor (Tir) into the host cell. Tir facilitates tight adhesion and recruitment of actin-regulating proteins leading to formation of an actin pedestal beneath the infecting bacterium. The pedestal has several similarities with podosomes, which are basolateral actin-rich extensions found in some migrating animal cells. Formation of podosomes is dependent upon the early podosome-specific scavenger protein Tks5, which is involved in actin recruitment. Although Tks5 is expressed in epithelial cells, and podosomes and EPEC pedestals share many components in their structure and mechanism of formation, the potential role of Tks5 in EPEC infections has not been studied. The aim of this study was to determine the subcellular localization of Tks5 in epithelial cells and to investigate if Tks5 is recruited to the EPEC pedestal. In an epithelial MDCK cell line stably expressing Tks5-EGFP, Tks5 localized to actin bundles. Upon infection, EPEC recruited Tks5-EGFP. Tir, but not Tir phosphorylation was essential for the recruitment. Time-lapse microscopy revealed that Tks5-EGFP was recruited instantly upon EPEC attachment to host cells, simultaneously with actin and N-WASp. EPEC infection of cells expressing a ΔPX-Tks5 deletion version of Tks5 showed that EPEC was able to both infect and form pedestals when the PX domain was deleted from Tks5. Future investigations will clarify the role of Tks5 in EPEC infection and pedestal formation. PMID:26536015

  19. Tir Is Essential for the Recruitment of Tks5 to Enteropathogenic Escherichia coli Pedestals.

    PubMed

    Jensen, Helene H; Pedersen, Hans N; Stenkjær, Eva; Pedersen, Gitte A; Login, Frédéric H; Nejsum, Lene N

    2015-01-01

    Enteropathogenic Escherichia coli (EPEC) is a bacterial pathogen that infects the epithelial lining of the small intestine and causes diarrhea. Upon attachment to the intestinal epithelium, EPEC uses a Type III Secretion System to inject its own high affinity receptor Translocated intimin receptor (Tir) into the host cell. Tir facilitates tight adhesion and recruitment of actin-regulating proteins leading to formation of an actin pedestal beneath the infecting bacterium. The pedestal has several similarities with podosomes, which are basolateral actin-rich extensions found in some migrating animal cells. Formation of podosomes is dependent upon the early podosome-specific scavenger protein Tks5, which is involved in actin recruitment. Although Tks5 is expressed in epithelial cells, and podosomes and EPEC pedestals share many components in their structure and mechanism of formation, the potential role of Tks5 in EPEC infections has not been studied. The aim of this study was to determine the subcellular localization of Tks5 in epithelial cells and to investigate if Tks5 is recruited to the EPEC pedestal. In an epithelial MDCK cell line stably expressing Tks5-EGFP, Tks5 localized to actin bundles. Upon infection, EPEC recruited Tks5-EGFP. Tir, but not Tir phosphorylation was essential for the recruitment. Time-lapse microscopy revealed that Tks5-EGFP was recruited instantly upon EPEC attachment to host cells, simultaneously with actin and N-WASp. EPEC infection of cells expressing a ΔPX-Tks5 deletion version of Tks5 showed that EPEC was able to both infect and form pedestals when the PX domain was deleted from Tks5. Future investigations will clarify the role of Tks5 in EPEC infection and pedestal formation.

  20. Cytosolic Extract Induces Tir Translocation and Pedestals in EPEC-Infected Red Blood Cells

    PubMed Central

    Swimm, Alyson I; Kalman, Daniel

    2008-01-01

    Enteropathogenic Escherichia coli (EPEC) are deadly contaminants in water and food, and induce protrusion of actin-filled membranous pedestals beneath themselves upon attachment to intestinal epithelia. Pedestal formation requires clustering of Tir and subsequent recruitment of cellular tyrosine kinases including Abl, Arg, and Etk as well as signaling molecules Nck, N-WASP, and Arp2/3 complex. We have developed a cytosolic extract-based cellular system that recapitulates actin pedestal formation in permeabilized red blood cells (RBC) infected with EPEC. RBC support attachment of EPEC and translocation of virulence factors, but not pedestal formation. We show here that extract induces a rapid Ca++-dependent release of Tir from the EPEC Type III secretion system, and that cytoplasmic factor(s) present in the extract facilitate translocation of Tir into the RBC plasma membrane. We show that Abl and related kinases in the extract phosphorylate Tir and that actin polymerization can be reconstituted in infected RBC following addition of cytosolic extract. Reconstitution requires the bacterial virulence factors Tir and intimin, and phosphorylation of Tir on tyrosine residue 474 results in the recruitment of Nck, N-WASP, and Arp2/3 complex beneath attached bacteria at sites of actin polymerization. Together these data describe a biochemical system for dissection of host components that mediate Type III secretion and the mechanisms by which complexes of proteins are recruited to discrete sites within the plasma membrane to initiate localized actin polymerization and morphological changes. PMID:18208322

  1. Cytosolic extract induces Tir translocation and pedestals in EPEC-infected red blood cells.

    PubMed

    Swimm, Alyson I; Kalman, Daniel

    2008-01-01

    Enteropathogenic Escherichia coli (EPEC) are deadly contaminants in water and food, and induce protrusion of actin-filled membranous pedestals beneath themselves upon attachment to intestinal epithelia. Pedestal formation requires clustering of Tir and subsequent recruitment of cellular tyrosine kinases including Abl, Arg, and Etk as well as signaling molecules Nck, N-WASP, and Arp2/3 complex. We have developed a cytosolic extract-based cellular system that recapitulates actin pedestal formation in permeabilized red blood cells (RBC) infected with EPEC. RBC support attachment of EPEC and translocation of virulence factors, but not pedestal formation. We show here that extract induces a rapid Ca++-dependent release of Tir from the EPEC Type III secretion system, and that cytoplasmic factor(s) present in the extract facilitate translocation of Tir into the RBC plasma membrane. We show that Abl and related kinases in the extract phosphorylate Tir and that actin polymerization can be reconstituted in infected RBC following addition of cytosolic extract. Reconstitution requires the bacterial virulence factors Tir and intimin, and phosphorylation of Tir on tyrosine residue 474 results in the recruitment of Nck, N-WASP, and Arp2/3 complex beneath attached bacteria at sites of actin polymerization. Together these data describe a biochemical system for dissection of host components that mediate Type III secretion and the mechanisms by which complexes of proteins are recruited to discrete sites within the plasma membrane to initiate localized actin polymerization and morphological changes.

  2. Analysis of pedestal plasma transport

    SciTech Connect

    Callen, J. D.; Groebner, R.; Osborne, T.H.; Canik, John; Owen, Larry W; Pankin, A. Y.; Rafiq, T.; Rognlien, T. D.; Stacey, W. M.

    2010-01-01

    An H-mode edge pedestal plasma transport benchmarking exercise was undertaken for a single DIII-D pedestal. Transport modelling codes used include 1.5D interpretive (ONETWO, GTEDGE), 1.5D predictive (ASTRA) and 2D ones (SOLPS, UEDGE). The particular DIII-D discharge considered is 98889, which has a typical low density pedestal. Profiles for the edge plasma are obtained from Thomson and charge-exchange recombination data averaged over the last 20% of the average 33.53 ms repetition time between type I edge localized modes. The modelled density of recycled neutrals is largest in the divertor X-point region and causes the edge plasma source rate to vary by a factor similar to 10(2) on the separatrix. Modelled poloidal variations in the densities and temperatures on flux surfaces are small on all flux surfaces up to within about 2.6 mm (rho(N) > 0.99) of the mid-plane separatrix. For the assumed Fick's-diffusion-type laws, the radial heat and density fluxes vary poloidally by factors of 2-3 in the pedestal region; they are largest on the outboard mid-plane where flux surfaces are compressed and local radial gradients are largest. Convective heat flows are found to be small fractions of the electron (less than or similar to 10%) and ion (less than or similar to 25%) heat flows in this pedestal. Appropriately averaging the transport fluxes yields interpretive 1.5D effective diffusivities that are smallest near the mid-point of the pedestal. Their 'transport barrier' minima are about 0.3 (electron heat), 0.15 (ion heat) and 0.035 (density) m(2) s(-1). Electron heat transport is found to be best characterized by electron-temperature-gradient-induced transport at the pedestal top and paleoclassical transport throughout the pedestal. The effective ion heat diffusivity in the pedestal has a different profile from the neoclassical prediction and may be smaller than it. The very small effective density diffusivity may be the result of an inward pinch flow nearly balancing a

  3. Intracellular motility and the evolution of the actin cytoskeleton during development of the male gametophyte of wheat (Triticum aestivum L.)

    PubMed Central

    Heslop-Harrison, J.; Heslop-Harrison, Y.

    1997-01-01

    The uniaperturate pollen of wheat is dispersed in a partially hydrated condition. Amyloplasts are concentrated in the apertural hemisphere where they surround the two sperms, while vigorously moving polysaccharide-containing wall precursor bodies (P-particles) together with the vegetative nucleus occupy the other. This disposition is the product of a post-meiotic developmental sequence apparently peculiar to the grasses. During vacuolation of the spore after release from the tetrad, the nucleus is displaced to the pole of the cell opposite the site of the germination aperture, already defined in the tetrad. Following pollen mitosis, the vegetative nucleus migrates along the wall of the vegetative cell towards the aperture, leaving the generative cell at the opposite pole isolated by a callose wall. As the vacuole is resorbed, the generative cell rounds up, loses its wall and follows the vegetative nucleus, passing along the wall of the vegetative cell towards the aperture where it eventually divides to produce the two sperms. Throughout this period of nucleus and cell manoeuvrings, minor inclusions of the vegetative cell cytoplasm, including mitochondria, lipid globuli and developing amyloplasts, move randomly. Coordinated vectorial movement begins after the main period of starch accumulation, when the amyloplasts migrate individually into the apertural hemisphere of the grain, a final redistribution betokening the attainment of germinability. In the present paper we correlate aspects of the evolution of the actin cytoskeleton with these events in the developing grain, and relate the observations to published evidence from another monocotyledonous species concerning the timing of the expression of actin genes during male gametophyte development, as revealed in the synthesis of actin mRNA.

  4. Identification of a Novel Member of the Chloride Intracellular Channel Gene Family (CLIC5) That Associates with the Actin Cytoskeleton of Placental Microvilli

    PubMed Central

    Berryman, Mark; Bretscher, Anthony

    2000-01-01

    The chloride intracellular channel (CLIC) gene family has been implicated in chloride ion transport within various subcellular compartments. We report here the molecular, biochemical, and cellular characterization of a new member of this gene family termed CLIC5. CLIC5 was isolated from extracts of placental microvilli as a component of a multimeric complex consisting of several known cytoskeletal proteins, including actin, ezrin, α-actinin, gelsolin, and IQGAP1. We cloned human cDNAs and generated antibodies specific for CLIC5, CLIC1/NCC27, and CLIC4/huH1/p64H1. CLIC5 shares 52–76% overall identity with human CLIC1, CLIC2, CLIC3, and CLIC4. Northern blot analysis showed that CLIC5 has a distinct pattern of expression compared with CLIC1 and CLIC4. Immunoblot analysis of extracts from placental tissues demonstrated that CLIC4 and CLIC5 are enriched in isolated placental microvilli, whereas CLIC1 is not. Moreover, in contrast to CLIC1 and CLIC4, CLIC5 is associated with the detergent-insoluble cytoskeletal fraction of microvilli. Indirect immunofluorescence microscopy revealed that CLIC4 and CLIC5 are concentrated within the apical region of the trophoblast, whereas CLIC1 is distributed throughout the cytoplasm. These studies suggest that CLIC1, CLIC4, and CLIC5 play distinct roles in chloride transport and that CLIC5 interacts with the cortical actin cytoskeleton in polarized epithelial cells. PMID:10793131

  5. A GTD analysis of ogive pedestal

    NASA Technical Reports Server (NTRS)

    Lai, Kin-Yue Albert; Burnside, Walter D.

    1987-01-01

    The metal ogive pedestal is claimed to have low radar cross section and low observability features. This study uses the Geometric Theory of Diffraction (GTD) to analyze the pedestal scattering for three cases: direct backscattered field, backscattered field structure, and target/pedestal multiple scattering. This study can be used to evaluate the various ways that the metal conical ogive pedestal can effect the performance of a high quality radar cross section measurement system.

  6. Pedestal substrate for coated optics

    DOEpatents

    Hale, Layton C.; Malsbury, Terry N.; Patterson, Steven R.

    2001-01-01

    A pedestal optical substrate that simultaneously provides high substrate dynamic stiffness, provides low surface figure sensitivity to mechanical mounting hardware inputs, and constrains surface figure changes caused by optical coatings to be primarily spherical in nature. The pedestal optical substrate includes a disk-like optic or substrate section having a top surface that is coated, a disk-like base section that provides location at which the substrate can be mounted, and a connecting cylindrical section between the base and optics or substrate sections. The connecting cylindrical section may be attached via three spaced legs or members. However, the pedestal optical substrate can be manufactured from a solid piece of material to form a monolith, thus avoiding joints between the sections, or the disk-like base can be formed separately and connected to the connecting section. By way of example, the pedestal optical substrate may be utilized in the fabrication of optics for an extreme ultraviolet (EUV) lithography imaging system, or in any optical system requiring coated optics and substrates with reduced sensitivity to mechanical mounts.

  7. Bacterial Actins.

    PubMed

    Izoré, Thierry; van den Ent, Fusinita

    2017-01-01

    A diverse set of protein polymers, structurally related to actin filaments contributes to the organization of bacterial cells as cytomotive or cytoskeletal filaments. This chapter describes actin homologs encoded by bacterial chromosomes. MamK filaments, unique to magnetotactic bacteria, help establishing magnetic biological compasses by interacting with magnetosomes. Magnetosomes are intracellular membrane invaginations containing biomineralized crystals of iron oxide that are positioned by MamK along the long-axis of the cell. FtsA is widespread across bacteria and it is one of the earliest components of the divisome to arrive at midcell, where it anchors the cell division machinery to the membrane. FtsA binds directly to FtsZ filaments and to the membrane through its C-terminus. FtsA shows altered domain architecture when compared to the canonical actin fold. FtsA's subdomain 1C replaces subdomain 1B of other members of the actin family and is located on the opposite side of the molecule. Nevertheless, when FtsA assembles into protofilaments, the protofilament structure is preserved, as subdomain 1C replaces subdomain IB of the following subunit in a canonical actin filament. MreB has an essential role in shape-maintenance of most rod-shaped bacteria. Unusually, MreB filaments assemble from two protofilaments in a flat and antiparallel arrangement. This non-polar architecture implies that both MreB filament ends are structurally identical. MreB filaments bind directly to membranes where they interact with both cytosolic and membrane proteins, thereby forming a key component of the elongasome. MreB filaments in cells are short and dynamic, moving around the long axis of rod-shaped cells, sensing curvature of the membrane and being implicated in peptidoglycan synthesis.

  8. Why is Actin Patchy?

    NASA Astrophysics Data System (ADS)

    Carlsson, Anders

    2009-03-01

    The intracellular protein actin, by reversibly polymerizing into filaments, generates forces for motion and shape changes of many types of biological cells. Fluorescence imaging studies show that actin often occurs in the form of localized patches of size roughly one micrometer at the cell membrane. Patch formation is most prevalent when the free-actin concentration is low. I investigate possible mechanisms for the formation of actin patches by numerically simulating the ``dendritic nucleation'' model of actin network growth. The simulations include filament growth, capping, branching, severing, and debranching. The attachment of membrane-bound activators to actin filaments, and subsequent membrane diffusion of unattached activators, are also included. It is found that as the actin concentration increases from zero, the actin occurs in patches at lower actin concentrations, and the size of the patches increases with increasing actin concentration. At a critical value of the actin concentration, the system undergoes a transition to complete coverage. The results are interpreted within the framework of reaction-diffusion equations in two dimensions.

  9. Role of electron temperature in the particle transport in the pedestal during pedestal evolution

    NASA Astrophysics Data System (ADS)

    Willensdorfer, M.; Fable, E.; Wolfrum, E.; Aumayr, F.; Fischer, R.; Reimold, F.; Ryter, F.

    2015-08-01

    The effect of the electron temperature (Te) on the edge particle transport in the pedestal is analyzed during the density build-up after the L-H transition. Electron cyclotron resonance heating was used to vary the pedestal temperature during the density evolution between subsequent H-mode phases. Although the pedestal Te and its gradients could be varied by a factor of 2, almost no change in the edge density evolution is observed within the measurement uncertainties. ASTRA was used to interpret the measurements and to analyze the dependence of the pedestal particle transport on the Te profile. Thermo-diffusion seems to play a minor role in the pedestal.

  10. Microtearing Instability In The ITER Pedestal

    SciTech Connect

    Wong, K. L.; Mikkelsen, D. R.; Rewoldt, G. M.; Budny, R.

    2010-12-01

    Unstable microtearing modes are discovered by the GS2 gyrokinetic siimulation code, in the pedestal region of a simulated ITER H-mode plasma with approximately 400 WM DT fusion power. Existing nonlinear theory indicates that these instabilities should produce stochastic magnetic fields and broaden the pedestal. The resulted electron thermal conductivity is estimated and the implications of these findings are discussed.

  11. Pedestal crater deposits as seen by SHARAD

    NASA Astrophysics Data System (ADS)

    Nunes, D. C.; Fisher, B.; Smrekar, S. E.; Plaut, J. J.; Holt, J. W.; Phillips, R. J.; Seu, R.; Head, J. W.

    2009-12-01

    Early in the exploration of Mars, Mariner 9 revealed a distinct morphologic class of craters that lie above the surrounding terrain atop steep-sided mesas roughly concentric to the craters. The predominant interpretation of these features argues that the crest of a pedestal represents the level of the surface at the time of the impact event that formed the central crater. Subsequent deflation lowered the surface pervasively, where as the pedestal remained due to increased resistance to removal via armoring or lag layers provided by the ejected blanket. Different models have been proposed to explain pedestal formation and persistence, and they generally involve the presence of a paleo-unit that was either friable or rich in volatiles. Given that pedestal craters occur dominantly in the mid-latitudes, with some cases also reported in the equatorial and polar regions, removal of extensive friable or volatile units at least as thick as the pedestals has occurred throughout Mars. We have analyzed data from the Shallow Radar (SHARAD) sounding instrument onboard the Mars Reconnaissance Orbiter to investigate the nature of the material composing the pedestals. Of the 60 pedestals examined, ranging a few to ~100 kilometers in diameter, we have identified several cases where the pedestal material is transparent to the SHARAD signal and a basal reflection is observed. These are cases where the pedestal is tens to ~ 100 kilometers in diameter. Subsurface reflections are not seen in every large pedestal, and for pedestals only a few kilometers wide we have not been able to unambiguously identify a basal reflector apart from surface clutter. In at least one case (in Malea Planum) internal layering between the surface and basal reflections is seen in SHARAD data. These layers appear to follow the shape of the basal topography and likely reflect past depositional events; layering is also seen at the edges of this pedestal in visible imagery from multiple orbiting cameras in this

  12. Linking microfilaments to intracellular membranes: the actin-binding and vesicle-associated protein comitin exhibits a mannose-specific lectin activity.

    PubMed Central

    Jung, E; Fucini, P; Stewart, M; Noegel, A A; Schleicher, M

    1996-01-01

    Comitin is a 24 kDa actin-binding protein from Dictyostelium discoideum that is located primarily on Golgi and vesicle membranes. We have probed the molecular basis of comitin's interaction with both actin and membranes using a series of truncation mutants obtained by expressing the appropriate cDNA in Escherichia coli. Comitin dimerizes in solution; its principle actin-binding activity is located between residues 90 and 135. The N-terminal 135 'core' residues of comitin contain a 3-fold sequence repeat that is homologous to several monocotyledon lectins and which retains key residues that determine these lectins' three-dimensional structure and mannose binding. These repeats of comitin appear to mediate its interaction with mannose residues in glycoproteins or glycolipids on the cytoplasmic surface of membrane vesicles from D.discoideum, and comitin can be released from membranes with mannose. Our data indicate that comitin binds to vesicle membranes via mannose residues and, by way of its interaction with actin, links these membranes to the cytoskeleton. Images PMID:8635456

  13. Actinic keratosis

    MedlinePlus

    Solar keratosis; Sun-induced skin changes - keratosis; Keratosis - actinic (solar); Skin lesion - actinic keratosis ... likely to develop it if you: Have fair skin, blue or green eyes, or blond or red ...

  14. Dynamics of Actin Stress Fibers and Focal Adhesions during Slow Migration in Swiss 3T3 Fibroblasts: Intracellular Mechanism of Cell Turning

    PubMed Central

    Miyoshi, Hiromi; Miura, Takuya; Tanaka, Hiroto; Tsubota, Ken-ichi; Liu, Hao

    2016-01-01

    To understand the mechanism regulating the spontaneous change in polarity that leads to cell turning, we quantitatively analyzed the dynamics of focal adhesions (FAs) coupling with the self-assembling actin cytoskeletal structure in Swiss 3T3 fibroblasts. Fluorescent images were acquired from cells expressing GFP-actin and RFP-zyxin by laser confocal microscopy. On the basis of the maximum area, duration, and relocation distance of FAs extracted from the RFP-zyxin images, the cells could be divided into 3 regions: the front region, intermediate lateral region, and rear region. In the intermediate lateral region, FAs appeared close to the leading edge and were stabilized gradually as its area increased. Simultaneously, bundled actin stress fibers (SFs) were observed vertically from the positions of these FAs, and they connected to the other SFs parallel to the leading edge. Finally, these connecting SFs fused to form a single SF with matured FAs at both ends. This change in SF organization with cell retraction in the first cycle of migration followed by a newly formed protrusion in the next cycle is assumed to lead to cell turning in migrating Swiss 3T3 fibroblasts. PMID:28119928

  15. Prediction and Optimization of the ITER Pedestal

    NASA Astrophysics Data System (ADS)

    Snyder, P. B.; Meneghini, O.; Beurskens, M. N. A.; Hughes, J. W.; Osborne, T. H.; Wilson, H. R.

    2016-10-01

    The structure of the edge transport barrier, or pedestal, plays an important role in ITER performance, with fusion power predicted to scale roughly with the square of the pedestal pressure. Understanding the physics of the pedestal is also critical to reliably suppressing or mitigating ELMs. We present tests of the EPED model, which predicts pedestal structure based on the intersection of two calculated criticality constraints, on more than 800 cases on existing tokamaks, and assess model accuracy across a range of parameters including normalized gyroradius. The EPED model is found to predict observations significantly more accurately than existing empirical pedestal models. The model is then used both independently, and coupled to core transport predictions from TGLF and NEO, using OMFIT, to predict and optimize ITER performance, including exploring possible operation in the Super H-Mode regime. Work supported by contracts DE-FG02-95ER54309, FC02-06ER54873, and DE-FC02-04ER54698.

  16. Role of the Arf6 GDP/GTP cycle and Arf6 GTPase-activating proteins in actin remodeling and intracellular transport.

    PubMed

    Klein, Stéphanie; Franco, Michel; Chardin, Pierre; Luton, Frédéric

    2006-05-05

    We have analyzed both biochemically and functionally a series of Arf6 mutants, providing new insights into the molecular mode of action of the small G protein Arf6. First, by comparing a fast-cycling mutant (Arf6(T157N)) and a GTPase-deficient mutant (Arf6(Q67L)), we established the necessity for completion of the Arf6 GDP/GTP cycle for recycling of major histocompatibility complex molecules to the plasma membrane. Second, we found that aluminum fluoride (AlF), known for inducing membrane protrusion in cells expressing exogenous wild-type Arf6, stabilized a functional wild-type Arf6.AlF(x) . GTPase-activating protein (GAP) complex in vitro and in vivo. We also found that the tandem mutation Q37E/S38I prevented the binding of two Arf GAPs, but not the effector ARHGAP10, and blocked the formation of membrane protrusion and actin reorganization. Together, our results with AlF(x) and Arf6(Q37E/S38I) demonstrate the critical role of the Arf6 GAPs as effectors for Arf6-regulated actin cytoskeleton remodeling. Finally, competition experiments conducted in vivo suggest the existence of a membrane receptor for GDP-bound Arf6.

  17. Bacterial nucleators: actin' on actin

    PubMed Central

    Bugalhão, Joana N.; Mota, Luís Jaime; Franco, Irina S.

    2015-01-01

    The actin cytoskeleton is a key target of numerous microbial pathogens, including protozoa, fungi, bacteria and viruses. In particular, bacterial pathogens produce and deliver virulence effector proteins that hijack actin dynamics to enable bacterial invasion of host cells, allow movement within the host cytosol, facilitate intercellular spread or block phagocytosis. Many of these effector proteins directly or indirectly target the major eukaryotic actin nucleator, the Arp2/3 complex, by either mimicking nucleation promoting factors or activating upstream small GTPases. In contrast, this review is focused on a recently identified class of effector proteins from Gram-negative bacteria that function as direct actin nucleators. These effector proteins mimic functional activities of formins, WH2-nucleators and Ena/VASP assembly promoting factors demonstrating that bacteria have coopted the complete set of eukaryotic actin assembly pathways. Structural and functional analyses of these nucleators have revealed several motifs and/or mechanistic activities that are shared with eukaryotic actin nucleators. However, functional effects of these proteins during infection extend beyond plain actin polymerization leading to interference with other host cell functions such as vesicle trafficking, cell cycle progression and cell death. Therefore, their use as model systems could not only help in the understanding of the mechanistic details of actin polymerization but also provide novel insights into the connection between actin dynamics and other cellular pathways. PMID:26416078

  18. [Actinic keratoses].

    PubMed

    Babilas, P; Landthaler, M; Szeimies, R-M

    2003-06-01

    Actinic keratoses are defined as proliferation of cytologically atypical keratinocytes in the zone of epidermal-dermal junction in photodamaged skin. In the northern hemisphere the prevalence of actinic keratoses ranges depending on different epidemiological studies from 11% to 25% for people aged 40 or older. The main cause of actinic keratoses is exposure to UVB radiation in sunlight UVB radiation induces mutations in the telomerase gene and in the tumor suppressor gene P53, which can also be detected in invasive squamous cell carcinoma. The only histological parameter to distinguish between actinic keratoses and SCC is the level of invasiveness. The risk for actinic keratoses to develop into SCC is about 16% over lo years. For this reason and because of the high prevalence of actinic keratoses, it has been suggested to replace the term,, actinic keratosis K with intraepidermal squamous cell carcinoma' to better characterize the lesion. In the following review recent aspects of pathogenesis and therapy of actinic keratoses are discussed.

  19. Actinous enigma or enigmatic actin

    PubMed Central

    Povarova, Olga I; Uversky, Vladimir N; Kuznetsova, Irina M; Turoverov, Konstantin K

    2014-01-01

    Being the most abundant protein of the eukaryotic cell, actin continues to keep its secrets for more than 60 years. Everything about this protein, its structure, functions, and folding, is mysteriously counterintuitive, and this review represents an attempt to solve some of the riddles and conundrums commonly found in the field of actin research. In fact, actin is a promiscuous binder with a wide spectrum of biological activities. It can exist in at least three structural forms, globular, fibrillar, and inactive (G-, F-, and I-actin, respectively). G-actin represents a thermodynamically instable, quasi-stationary state, which is formed in vivo as a result of the energy-intensive, complex posttranslational folding events controlled and driven by cellular folding machinery. The G-actin structure is dependent on the ATP and Mg2+ binding (which in vitro is typically substituted by Ca2+) and protein is easily converted to the I-actin by the removal of metal ions and by action of various denaturing agents (pH, temperature, and chemical denaturants). I-actin cannot be converted back to the G-form. Foldable and “natively folded” forms of actin are always involved in interactions either with the specific protein partners, such as Hsp70 chaperone, prefoldin, and the CCT chaperonin during the actin folding in vivo or with Mg2+ and ATP as it takes place in the G-form. We emphasize that the solutions for the mysteries of actin multifunctionality, multistructurality, and trapped unfolding can be found in the quasi-stationary nature of this enigmatic protein, which clearly possesses many features attributed to both globular and intrinsically disordered proteins. PMID:28232879

  20. Toxofilin, a Novel Actin-binding Protein from Toxoplasma gondii, Sequesters Actin Monomers and Caps Actin Filaments

    PubMed Central

    Poupel, Olivier; Boleti, Haralabia; Axisa, Sophie; Couture-Tosi, Evelyne; Tardieux, Isabelle

    2000-01-01

    Toxoplasma gondii relies on its actin cytoskeleton to glide and enter its host cell. However, T. gondii tachyzoites are known to display a strikingly low amount of actin filaments, which suggests that sequestration of actin monomers could play a key role in parasite actin dynamics. We isolated a 27-kDa tachyzoite protein on the basis of its ability to bind muscle G-actin and demonstrated that it interacts with parasite G-actin. Cloning and sequence analysis of the gene coding for this protein, which we named Toxofilin, showed that it is a novel actin-binding protein. In in vitro assays, Toxofilin not only bound to G-actin and inhibited actin polymerization as an actin-sequestering protein but also slowed down F-actin disassembly through a filament end capping activity. In addition, when green fluorescent protein-tagged Toxofilin was overexpressed in mammalian nonmuscle cells, the dynamics of actin stress fibers was drastically impaired, whereas green fluorescent protein-Toxofilin copurified with G-actin. Finally, in motile parasites, during gliding or host cell entry, Toxofilin was localized in the entire cytoplasm, including the rear end of the parasite, whereas in intracellular tachyzoites, especially before they exit from the parasitophorous vacuole of their host cell, Toxofilin was found to be restricted to the apical end. PMID:10637313

  1. Suppressors of Yeast Actin Mutations

    PubMed Central

    Novick, P.; Osmond, B. C.; Botstein, D.

    1989-01-01

    Suppressors of a temperature-sensitive mutation (act1-1) in the single actin gene of Saccharomyces cerevisiae were selected that had simultaneously acquired a cold-sensitive growth phenotype. Five genes, called SAC (suppressor of actin) were defined by complementation tests; both suppression and cold-sensitive phenotypes were recessive. Three of the genes (SAC1, SAC2 and SAC3) were subjected to extensive genetic and phenotypic analysis, including molecular cloning. Suppression was found to be allele-specific with respect to actin alleles. The sac mutants, even in ACT1(+) genetic backgrounds, displayed phenotypes similar to those of actin mutants, notably aberrant organization of intracellular actin and deposition of chitin at the cell surface. These results are interpreted as being consistent with the idea that the SAC genes encode proteins that interact with actin, presumably as components or controllers of the assembly or stability of the yeast actin cytoskeleton. Two unexpected properties of the SAC1 gene were noted. Disruptions of the gene indicated that its function is essential only at temperatures below about 17° and all sac1 alleles are inviable when combined with act1-2. These properties are interpreted in the context of the evolution of the actin cytoskeleton of yeast. PMID:2656401

  2. Chlamydia trachomatis Tarp harbors distinct G and F actin binding domains that bundle actin filaments.

    PubMed

    Jiwani, Shahanawaz; Alvarado, Stephenie; Ohr, Ryan J; Romero, Adriana; Nguyen, Brenda; Jewett, Travis J

    2013-02-01

    All species of Chlamydia undergo a unique developmental cycle that transitions between extracellular and intracellular environments and requires the capacity to invade new cells for dissemination. A chlamydial protein called Tarp has been shown to nucleate actin in vitro and is implicated in bacterial entry into human cells. Colocalization studies of ectopically expressed enhanced green fluorescent protein (EGFP)-Tarp indicate that actin filament recruitment is restricted to the C-terminal half of the effector protein. Actin filaments are presumably associated with Tarp via an actin binding alpha helix that is also required for actin nucleation in vitro, but this has not been investigated. Tarp orthologs from C. pneumoniae, C. muridarum, and C. caviae harbor between 1 and 4 actin binding domains located in the C-terminal half of the protein, but C. trachomatis serovar L2 has only one characterized domain. In this work, we examined the effects of domain-specific mutations on actin filament colocalization with EGFP-Tarp. We now demonstrate that actin filament colocalization with Tarp is dependent on two novel F-actin binding domains that endow the Tarp effector with actin-bundling activity. Furthermore, Tarp-mediated actin bundling did not require actin nucleation, as the ability to bundle actin filaments was observed in mutant Tarp proteins deficient in actin nucleation. These data shed molecular insight on the complex cytoskeletal rearrangements required for C. trachomatis entry into host cells.

  3. Scaling studies of H-mode pedestal characteristics on Experimental Advanced Superconducting Tokamak

    SciTech Connect

    Wang, Teng Fei E-mail: zangq@ipp.ac.cn; Zang, Qing E-mail: zangq@ipp.ac.cn; Han, Xiao Feng; Xiao, Shu Mei; Tian, Bao Gang; Hu, Ai Lan; Zhao, Jun Yu

    2016-03-15

    The characteristics of high-confinement mode (H-mode) pedestal are examined on the Experimental Advanced Superconducting Tokamak. It is found that they are closely dependent on each other between electron pedestal characteristics and global parameters for all types of edge localized mode (ELM). The scaling of pedestal temperature based on thermal conduction and pedestal pressure width is carried out. Based on pedestal pressure gradient and pedestal density, six pedestal pressure width models are applied to predict the pedestal temperature height of type I ELMy H-mode. Compared to experimental results, the normalized poloidal beta model is more consistent than other models.

  4. Compatibility of Detached Divertor Operation with Robust Edge Pedestal Performance

    NASA Astrophysics Data System (ADS)

    Leonard, A. W.; Osborne, T. H.; Snyder, P. B.; Makowski, M. A.; McLean, A. G.

    2014-10-01

    The compatibility of radiative detached divertor operation with the maintenance of a robust H-mode pedestal is examined in DIII-D. A density scan with deuterium injection into H-mode spanned a range of divertor conditions from fully attached, ~30 eV at the target, with little divertor radiation to a fully detached with Te < 5 eV throughout the divertor up to the X-point. Over this scan of pedestal density from n /nGW = 30% to 60% the pedestal Te was reduced from 800 eV to 350 eV, representing a ~20% reduction in pedestal pressure with a similar reduction in normalized energy confinement. The reduction in pedestal pressure at high density was found to be consistent with a reduced pedestal ELM MHD stability limit at high collisionality. The scaling of the pedestal top pressure with density was also consistent with the EPED model, which assumes an additional constraint on the local pressure gradient. The MHD stability limit at the highest collisionality depends on details of the ELM instability growth rate normalization. This result is encouraging for future burning plasmas where a low collisionality pedestal is expected to be maintained even for high density detached divertor operation. Work supported by the US DOE under DE-FC02-04ER54698 and DE-AC52-07NA27344.

  5. Actinic reticuloid

    SciTech Connect

    Marx, J.L.; Vale, M.; Dermer, P.; Ragaz, A.; Michaelides, P.; Gladstein, A.H.

    1982-09-01

    A 58-year-old man has his condition diagnosed as actinic reticuloid on the basis of clinical and histologic findings and phototesting data. He had clinical features resembling mycosis fungoides in light-exposed areas. Histologic findings disclosed a bandlike infiltrate with atypical mononuclear cells in the dermis and scattered atypical cells in the epidermis. Electron microscopy disclosed mononuclear cells with bizarre, convoluted nuclei, resembling cerebriform cells of Lutzner. Phototesting disclosed a diminished minimal erythemal threshold to UV-B and UV-A. Microscopic changes resembling actinic reticuloid were reproduced in this patient 24 and 72 hours after exposure to 15 minimal erythemal doses of UV-B.

  6. Compatibility of detached divertor operation with robust edge pedestal performance

    NASA Astrophysics Data System (ADS)

    Leonard, A. W.; Makowski, M. A.; McLean, A. G.; Osborne, T. H.; Snyder, P. B.

    2015-08-01

    The compatibility of detached radiative divertor operation with a robust H-mode pedestal is examined in DIII-D. A density scan produced low temperature plasmas at the divertor target, Te ⩽ 2 eV, with high radiation leading to a factor of ⩾4 drop in peak divertor heat flux. The cold radiative plasma was confined to the divertor and did not extend across the separatrix in X-point region. A robust H-mode pedestal was maintained with a small degradation in pedestal pressure at the highest densities. The response of the pedestal pressure to increasing density is reproduced by the EPED pedestal model. However, agreement of the EPED model with experiment at high density requires an assumption of reduced diamagnetic stabilization of edge Peeling-Ballooning modes.

  7. Repair of the DSS-14 Pedestal Concrete

    NASA Technical Reports Server (NTRS)

    Mcclure, D.

    1985-01-01

    About three years after the Goldstone Deep Space Station antenna was dedicated, grout under the hydrostatic bearing runner was found to be interacting with the runner, causing rust to form between the runner and the sole plates upon which it rests. The rust formed unevenly and the runner could not be kept flat so in 1969 the grout was removed and replaced with a Portland cement and sand dry pack grout that was less likely to produce rust. In the years that followed, oil leaking from the runner assembly caused progressive deterioration of the drypack grout. In 1982 over one thousand hours of spacecraft tracking time were lost due to this deterioration. A plan was developed to rehabilitate the bearing. The plan called for raising the rotating structure free from the concrete pedestal and placing it on three pairs of external support columns. With the weight of the structure transferred to the columns, the pads and runner could be removed and the repair started. The very successful repair included the replacement of a significant portion of the antenna pedestal.

  8. Repair of the DSS-14 Pedestal Concrete

    NASA Technical Reports Server (NTRS)

    Mcclure, D.

    1985-01-01

    About three years after the Goldstone Deep Space Station antenna was dedicated, grout under the hydrostatic bearing runner was found to be interacting with the runner, causing rust to form between the runner and the sole plates upon which it rests. The rust formed unevenly and the runner could not be kept flat so in 1969 the grout was removed and replaced with a Portland cement and sand dry pack grout that was less likely to produce rust. In the years that followed, oil leaking from the runner assembly caused progressive deterioration of the drypack grout. In 1982 over one thousand hours of spacecraft tracking time were lost due to this deterioration. A plan was developed to rehabilitate the bearing. The plan called for raising the rotating structure free from the concrete pedestal and placing it on three pairs of external support columns. With the weight of the structure transferred to the columns, the pads and runner could be removed and the repair started. The very successful repair included the replacement of a significant portion of the antenna pedestal.

  9. Intracellular Parasite Invasion Strategies

    NASA Astrophysics Data System (ADS)

    Sibley, L. D.

    2004-04-01

    Intracellular parasites use various strategies to invade cells and to subvert cellular signaling pathways and, thus, to gain a foothold against host defenses. Efficient cell entry, ability to exploit intracellular niches, and persistence make these parasites treacherous pathogens. Most intracellular parasites gain entry via host-mediated processes, but apicomplexans use a system of adhesion-based motility called ``gliding'' to actively penetrate host cells. Actin polymerization-dependent motility facilitates parasite migration across cellular barriers, enables dissemination within tissues, and powers invasion of host cells. Efficient invasion has brought widespread success to this group, which includes Toxoplasma, Plasmodium, and Cryptosporidium.

  10. The ability of an attaching and effacing pathogen to trigger localized actin assembly contributes to virulence by promoting mucosal attachment

    PubMed Central

    Mallick, Emily M.; Garber, John J.; Vanguri, Vijay K.; Balasubramanian, Sowmya; Blood, Timothy; Clark, Stacie; Vingadassalom, Didier; Louissaint, Christopher; McCormick, Beth; Snapper, Scott B.; Leong, John M.

    2014-01-01

    Enterohemorrhagic Escherichia coli (EHEC) adheres to intestinal epithelial cells, then stimulates the actin nucleation promoting factor N-WASP to induce localized actin assembly resulting in an actin “pedestal”, the function of which is poorly understood. EHEC also produces Shiga toxin (Stx), which penetrates the intestinal epithelium to cause a life-threatening renal and systemic disease. To assess the role of pedestal formation in colonization and disease, we utilized the murine pathogen Citrobacter rodentium, which also forms actin pedestals, and the genetically engineered C. rodentium (Φstx2dact), which additionally triggers Stx-mediated systemic disease. We found that an intestine-specific N-WASP-deficient (iNWKO) mouse suffered dramatically less colonization and disease than N-WASP-proficient littermate controls when infected with either strain. In addition, upon infection of wild type mice, mutants of C. rodentium or C. rodentium (Φstx2dact) that are specifically defective in pedestal formation demonstrated a relatively modest defect in cecal colonization and fecal shedding, but a more severe defect in colonization of the colonic mucosa. The C. rodentium (Φstx2dact) pedestal-defective mutant did not cause renal disease and, after normalizing for fecal bacterial load, was associated with a 16-fold lower risk of lethality. These findings suggest that the ability of an attaching and effacing pathogen to promote localized actin assembly contributes to virulence by promoting mucosal attachment. PMID:24780054

  11. Microtearing Turbulence Limiting the JET-ILW Pedestal

    NASA Astrophysics Data System (ADS)

    Hatch, David; Kotschenreuther, Michael; Mahajan, Swadesh; Valanju, Prashant; Liu, Xing; Goerler, Tobias; Jenko, Frank; Told, Daniel

    2015-11-01

    The gyrokinetic GENE code is used to model instabilities and transport in the JET-ILW (ITER like wall) pedestal. Local GENE simulations identify microtearing modes (MTM) as the dominant low-ky instability across most of the pedestal, with KBM unstable in a narrow region near the separatrix. Global simulations find that MTM growth rates are decreased by ExB shear, but to a lesser extent than electrostatic ITG/TEM-type modes, so that the MTM becomes relatively more prominent in the presence of ExB shear. A β scan demonstrates local KBM to be unstable across the pedestal at lower β (60% of the experimental value). As β approaches and surpasses the experimental value, the KBM become more stable, and are limited to progressively narrower regions of the pedestal (consistent with the concept of second stability), while the MTM becomes more unstable and spans most of the pedestal. The absence of KBM is even more pronounced in global simulations. Nonlinear simulations of MTM turbulence using the experimental profiles produce transport levels that are comparable to experimental expectations, establishing the MTM as the likely mechanism limiting pedestal profile evolution in JET-ILW pedestals. This work was supported by U.S. DOE Contract No. DE-FG02-04ER54742.

  12. Predictive modeling of pedestal structure in KSTAR using EPED model

    SciTech Connect

    Han, Hyunsun; Kim, J. Y.; Kwon, Ohjin

    2013-10-15

    A predictive calculation is given for the structure of edge pedestal in the H-mode plasma of the KSTAR (Korea Superconducting Tokamak Advanced Research) device using the EPED model. Particularly, the dependence of pedestal width and height on various plasma parameters is studied in detail. The two codes, ELITE and HELENA, are utilized for the stability analysis of the peeling-ballooning and kinetic ballooning modes, respectively. Summarizing the main results, the pedestal slope and height have a strong dependence on plasma current, rapidly increasing with it, while the pedestal width is almost independent of it. The plasma density or collisionality gives initially a mild stabilization, increasing the pedestal slope and height, but above some threshold value its effect turns to a destabilization, reducing the pedestal width and height. Among several plasma shape parameters, the triangularity gives the most dominant effect, rapidly increasing the pedestal width and height, while the effect of elongation and squareness appears to be relatively weak. Implication of these edge results, particularly in relation to the global plasma performance, is discussed.

  13. Predictive modeling of pedestal structure in KSTAR using EPED model

    NASA Astrophysics Data System (ADS)

    Han, Hyunsun; Kwon, Ohjin; Kim, J. Y.

    2013-10-01

    A predictive calculation is given for the structure of edge pedestal in the H-mode plasma of the KSTAR (Korea Superconducting Tokamak Advanced Research) device using the EPED model. Particularly, the dependence of pedestal width and height on various plasma parameters is studied in detail. The two codes, ELITE and HELENA, are utilized for the stability analysis of the peeling-ballooning and kinetic ballooning modes, respectively. Summarizing the main results, the pedestal slope and height have a strong dependence on plasma current, rapidly increasing with it, while the pedestal width is almost independent of it. The plasma density or collisionality gives initially a mild stabilization, increasing the pedestal slope and height, but above some threshold value its effect turns to a destabilization, reducing the pedestal width and height. Among several plasma shape parameters, the triangularity gives the most dominant effect, rapidly increasing the pedestal width and height, while the effect of elongation and squareness appears to be relatively weak. Implication of these edge results, particularly in relation to the global plasma performance, is discussed.

  14. Rho GTPases, phosphoinositides, and actin

    PubMed Central

    Croisé, Pauline; Estay-Ahumada, Catherine; Gasman, Stéphane; Ory, Stéphane

    2014-01-01

    Rho GTPases are well known regulators of the actin cytoskeleton that act by binding and activating actin nucleators. They are therefore involved in many actin-based processes, including cell migration, cell polarity, and membrane trafficking. With the identification of phosphoinositide kinases and phosphatases as potential binding partners or effectors, Rho GTPases also appear to participate in the regulation of phosphoinositide metabolism. Since both actin dynamics and phosphoinositide turnover affect the efficiency and the fidelity of vesicle transport between cell compartments, Rho GTPases have emerged as critical players in membrane trafficking. Rho GTPase activity, actin remodeling, and phosphoinositide metabolism need to be coordinated in both space and time to ensure the progression of vesicles along membrane trafficking pathways. Although most molecular pathways are still unclear, in this review, we will highlight recent advances made in our understanding of how Rho-dependent signaling pathways organize actin dynamics and phosphoinositides and how phosphoinositides potentially provide negative feedback to Rho GTPases during endocytosis, exocytosis and membrane exchange between intracellular compartments. PMID:24914539

  15. Actin dynamics and cofilin-actin rods in Alzheimer disease

    PubMed Central

    Bamburg, James R.; Bernstein, Barbara W.

    2017-01-01

    Cytoskeletal abnormalities and synaptic loss, typical of both familial and sporadic Alzheimer disease (AD), are induced by diverse stresses such as neuroinflammation, oxidative stress, and energetic stress, each of which may be initiated or enhanced by proinflammatory cytokines or amyloid-β (Aβ) peptides. Extracellular Aβ-containing plaques and intracellular phospho-tau-containing neurofibrillary tangles are postmortem pathologies required to confirm AD and have been the focus of most studies. However, AD brain, but not normal brain, also have increased levels of cytoplasmic rod-shaped bundles of filaments composed of ADF/cofilin-actin in a 1:1 complex (rods). Cofilin, the major ADF/cofilin isoform in mammalian neurons, severs actin filaments at low cofilin/actin ratios and stabilizes filaments at high cofilin/actin ratios. It binds cooperatively to ADP-actin subunits in F-actin. Cofilin is activated by dephosphorylation and may be oxidized in stressed neurons to form disulfide-linked dimers, required for bundling cofilin-actin filaments into stable rods. Rods form within neurites causing synaptic dysfunction by sequestering cofilin, disrupting normal actin dynamics, blocking transport, and exacerbating mitochondrial membrane potential loss. Aβ and proinflammatory cytokines induce rods through a cellular prion protein-dependent activation of NADPH oxidase and production of reactive oxygen species. Here we review recent advances in our understanding of cofilin biochemistry, rod formation, and the development of cognitive deficits. We will then discuss rod formation as a molecular pathway for synapse loss that may be common between all three prominent current AD hypotheses, thus making rods an attractive therapeutic target. PMID:26873625

  16. Actin: Structure, Function, Dynamics, and Interactions with Bacterial Toxins.

    PubMed

    Kühn, Sonja; Mannherz, Hans Georg

    Actin is one of the most abundant proteins in any eukaryotic cell and an indispensable component of the cytoskeleton. In mammalian organisms, six highly conserved actin isoforms can be distinguished, which differ by only a few amino acids. In non-muscle cells, actin polymerizes into actin filaments that form actin structures essential for cell shape stabilization, and participates in a number of motile activities like intracellular vesicle transport, cytokinesis, and also cell locomotion. Here, we describe the structure of monomeric and polymeric actin, the polymerization kinetics, and its regulation by actin-binding proteins. Probably due to its conserved nature and abundance, actin and its regulating factors have emerged as prefered targets of bacterial toxins and effectors, which subvert the host actin cytoskeleton to serve bacterial needs.

  17. Enhancement of the Bootstrap Current in a Tokamak Pedestal

    SciTech Connect

    Kagan, Grigory; Catto, Peter J.

    2010-07-23

    The strong radial electric field in a subsonic tokamak pedestal modifies the neoclassical ion parallel flow velocity, as well as the radial ion heat flux. Existing experimental evidence of the resulting alteration in the poloidal flow of a trace impurity is discussed. We then demonstrate that the modified parallel ion flow can noticeably enhance the pedestal bootstrap current when the background ions are in the banana regime. Only the coefficient of the ion temperature gradient drive term is affected. The revised expression for the pedestal bootstrap current is presented. The prescription for inserting the modification into any existing banana regime bootstrap current expression is given.

  18. Microtearing turbulence limiting the JET-ILW pedestal

    NASA Astrophysics Data System (ADS)

    Hatch, D. R.; Kotschenreuther, M.; Mahajan, S.; Valanju, P.; Jenko, F.; Told, D.; Görler, T.; Saarelma, S.

    2016-10-01

    The first nonlinear gyrokinetic turbulence simulations that quantitatively reproduce experimental transport levels in an H-mode pedestal are reported. In the JET-ILW (ITER-like wall) pedestal, the bulk of the transport in the steep gradient region is caused by the turbulence driven by the microtearing mode (MTM). Kinetic ballooning modes are found to be in a second-stability regime. With contributions from the neoclassical and electron temperature gradient driven transport, the MTM mechanism reproduces, quantitatively, the experimental power balance across most of the pedestal.

  19. Enhancement of the Bootstrap Current in a Tokamak Pedestal

    NASA Astrophysics Data System (ADS)

    Kagan, Grigory; Catto, Peter J.

    2010-07-01

    The strong radial electric field in a subsonic tokamak pedestal modifies the neoclassical ion parallel flow velocity, as well as the radial ion heat flux. Existing experimental evidence of the resulting alteration in the poloidal flow of a trace impurity is discussed. We then demonstrate that the modified parallel ion flow can noticeably enhance the pedestal bootstrap current when the background ions are in the banana regime. Only the coefficient of the ion temperature gradient drive term is affected. The revised expression for the pedestal bootstrap current is presented. The prescription for inserting the modification into any existing banana regime bootstrap current expression is given.

  20. ELM Suppression and Pedestal Structure in I-Mode Plasmas

    NASA Astrophysics Data System (ADS)

    Walk, John

    2013-10-01

    The I-mode regime is characterized by the formation of a temperature pedestal and enhanced energy confinement (H98 up to 1.2), without an accompanying density pedestal or drop in particle transport. Unlike ELMy H-modes, I-mode operation appears to have naturally-occurring suppression of large ELMs in addition to its highly favorable scalings of pedestal structure (and therefore overall performance). Instead, continuous Weakly Coherent Modes help to regulate density. Extensive study of the ELMy H-mode has led to the development of the EPED model, which utilizes calculations of coupled peeling-ballooning MHD modes and kinetic-ballooning mode (KBM) stability limits to predict the pedestal structure preceding an ELM crash. We apply similar tools to the structure and ELM stability of I-mode pedestals. Peeling-ballooning MHD calculations are completed using the ELITE code, showing I-mode pedestals to be generally MHD-stable. Under certain conditions, intermittent ELMs are observed in I-mode at reduced field, typically triggered by sawtooth crashes; modification of the temperature pedestal (and therefore the pressure profile stability) by sawtooth heat pulses is being examined in ELITE. Modeled stability to KBM turbulence in I-mode and ELMy H-mode suggests that typical I-modes are stable against KBM turbulence. Measured I-mode pedestals are significantly wider (more stable) than the width scaling with the square root of poloidal beta characteristic of the KBM-limited pedestals in ELMy H-mode. Finally, we explore scalings of pedestal structure with engineering parameters compared to ELMy H-modes on C-Mod. In particular, we focus on scalings of the pressure pedestal with heating power (and its relation to the favorable scaling of confinement with power in I-mode) and on relationships between heat flux and pedestal temperature gradients. This work is supported by DOE agreement DE-FC02-99ER54512. Theory work at General Atomics is supported by DOE agreement DE-FG02-99ER54309.

  1. A new F-actin structure in fungi: actin ring formation around the cell nucleus of Cryptococcus neoformans.

    PubMed

    Kopecká, Marie; Kawamoto, Susumu; Yamaguchi, Masashi

    2013-04-01

    The F-actin cytoskeleton of Cryptococcus neoformans is known to comprise actin cables, cortical patches and cytokinetic ring. Here, we describe a new F-actin structure in fungi, a perinuclear F-actin collar ring around the cell nucleus, by fluorescent microscopic imaging of rhodamine phalloidin-stained F-actin. Perinuclear F-actin rings form in Cryptococcus neoformans treated with the microtubule inhibitor Nocodazole or with the drug solvent dimethyl sulfoxide (DMSO) or grown in yeast extract peptone dextrose (YEPD) medium, but they are absent in cells treated with Latrunculin A. Perinuclear F-actin rings may function as 'funicular cabin' for the cell nucleus, and actin cables as intracellular 'funicular' suspending nucleus in the central position in the cell and moving nucleus along the polarity axis along actin cables.

  2. 51. (no plate) Lens, lens pedestal, mercury float, shade holder ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    51. (no plate) Lens, lens pedestal, mercury float, shade holder installation, drawing # 3101, sheet 2 of 2. Approved April 6, 1928. - Block Island Southeast Light, Spring Street & Mohegan Trail at Mohegan Bluffs, New Shoreham, Washington County, RI

  3. 50. (no plate) Lens, lens pedestal, mercury float, drawing # ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    50. (no plate) Lens, lens pedestal, mercury float, drawing # 3101, sheet 1 of 2. Approved April 6, 1928. - Block Island Southeast Light, Spring Street & Mohegan Trail at Mohegan Bluffs, New Shoreham, Washington County, RI

  4. 8. SOUTH PLANT SHELL OIL COMPANY FACILITIES, WITH PIPELINE PEDESTALS ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    8. SOUTH PLANT SHELL OIL COMPANY FACILITIES, WITH PIPELINE PEDESTALS IN FOREGROUND. VIEW TO SOUTHWEST. - Rocky Mountain Arsenal, Bounded by Ninety-sixth Avenue & Fifty-sixth Avenue, Buckley Road, Quebec Street & Colorado Highway 2, Commerce City, Adams County, CO

  5. 4. FORGE, ANVIL, PEDESTAL GRINDER, AND BELT DRIVES. NOTE WATERWHEEL ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    4. FORGE, ANVIL, PEDESTAL GRINDER, AND BELT DRIVES. NOTE WATERWHEEL NEEDLE VALVE CASTING HANGING ON THE WALL ABOVE THE FORGE. VIEW TO NORTH. - Santa Ana River Hydroelectric System, SAR-1 Machine Shop, Redlands, San Bernardino County, CA

  6. MLP SUPPORT PEDESTAL 4 AT PARKING SITE Cape Canaveral ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    MLP SUPPORT PEDESTAL 4 AT PARKING SITE - Cape Canaveral Air Force Station, Launch Complex 39, Mobile Launcher Platforms, Launcher Road, East of Kennedy Parkway North, Cape Canaveral, Brevard County, FL

  7. MLP SUPPORT PEDESTAL 2 AT PARKING SITE Cape Canaveral ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    MLP SUPPORT PEDESTAL 2 AT PARKING SITE - Cape Canaveral Air Force Station, Launch Complex 39, Mobile Launcher Platforms, Launcher Road, East of Kennedy Parkway North, Cape Canaveral, Brevard County, FL

  8. MLP SUPPORT PEDESTAL 1 AT PARKING SITE Cape Canaveral ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    MLP SUPPORT PEDESTAL 1 AT PARKING SITE - Cape Canaveral Air Force Station, Launch Complex 39, Mobile Launcher Platforms, Launcher Road, East of Kennedy Parkway North, Cape Canaveral, Brevard County, FL

  9. DETAIL VIEW OF A CAMERA PEDESTAL ON THE BLAST DECK ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    DETAIL VIEW OF A CAMERA PEDESTAL ON THE BLAST DECK OF THE 0 LEVEL - Cape Canaveral Air Force Station, Launch Complex 39, Mobile Launcher Platforms, Launcher Road, East of Kennedy Parkway North, Cape Canaveral, Brevard County, FL

  10. Non-local effects on pedestal kinetic ballooning mode stability

    NASA Astrophysics Data System (ADS)

    Saarelma, S.; Martin-Collar, J.; Dickinson, D.; McMillan, B. F.; Roach, C. M.; MAST Team; JET Contributors, The

    2017-06-01

    The H-mode pedestal height plays an important role in determining the global confinement of the tokamak plasma. In type I ELMy H-mode the ultimate limit for the pedestal pressure at constant width is set by the ideal MHD peeling-ballooning modes (PBMs) that are thought to be the trigger for the ELMs. However, the PBM criterion does not uniquely determine the pedestal. Increasing the width of the pedestal, the marginally peeling-ballooning stable pedestal height increases as well. The second criterion for the pedestal is set by the transport processes in the pedestal that limit the gradient between the ELMs. One candidate for driving this transport is the kinetic ballooning mode (KBM) that is driven by the pressure gradient (Snyder et al 2009 Nucl. Fusion 49 085035). The KBM growth rate increases very rapidly after the critical pressure gradient is exceeded leading to very stiff profiles with the pressure gradient clamped near to the stability limit. In the local linear gyrokinetic analysis of experimental MAST and JET plasmas we have found that, like the {n}=∞ ideal MHD ballooning modes, the KBMs can access locally so called second stability if the magnetic shear becomes low enough (Dickinson et al 2011 Plasma Phys. Control. Fusion 53 115010; Saarelma et al 2013 Nucl. Fusion 53 123012). However, in the pedestal region the local assumption that the equilibrium can be considered radially constant for the investigated modes is no longer justified. In this paper we revisit the KBM analysis using a global code ORB5 to investigate whether second stability access exists for KBMs. We find that counter to the local analysis, the global KBM stability is not sensitive to the magnetic shear in the pedestal region. At sufficiently high β (but still below the ideal peeling-ballooning limit) the pedestal region becomes KBM unstable regardless of the amount of bootstrap current assumed in the equilibrium reconstruction. However, just as in local analysis, the mode is stabilised

  11. Global performance enhancements via pedestal optimisation on ASDEX Upgrade

    NASA Astrophysics Data System (ADS)

    Dunne, M. G.; Frassinetti, L.; Beurskens, M. N. A.; Cavedon, M.; Fietz, S.; Fischer, R.; Giannone, L.; Huijsmans, G. T. A.; Kurzan, B.; Laggner, F.; McCarthy, P. J.; McDermott, R. M.; Tardini, G.; Viezzer, E.; Willensdorfer, M.; Wolfrum, E.; The EUROfusion MST1 Team; The ASDEX Upgrade Team

    2017-02-01

    Results of experimental scans of heating power, plasma shape, and nitrogen content are presented, with a focus on global performance and pedestal alteration. In detailed scans at low triangularity, it is shown that the increase in stored energy due to nitrogen seeding stems from the pedestal. It is also shown that the confinement increase is driven through the temperature pedestal at the three heating power levels studied. In a triangularity scan, an orthogonal effect of shaping and seeding is observed, where increased plasma triangularity increases the pedestal density, while impurity seeding (carbon and nitrogen) increases the pedestal temperature in addition to this effect. Modelling of these effects was also undertaken, with interpretive and predictive models being employed. The interpretive analysis shows a general agreement of the experimental pedestals in separate power, shaping, and seeding scans with peeling-ballooning theory. Predictive analysis was used to isolate the individual effects, showing that the trends of additional heating power and increased triangularity can be recoverd. However, a simple change of the effective charge in the plasma cannot explain the observed levels of confinement improvement in the present models.

  12. The Research of EAST Pedestal Structure and Preliminary Application

    NASA Astrophysics Data System (ADS)

    Wang, Tengfei; Zang, Qing; Han, Xiaofeng; Xiao, Shumei; Hu, Ailan; Zhao, Junyu

    2016-10-01

    The pedestal characteristic is an important basis for high confinement mode (H-mode) research. Because of the finite spatial resolution of Thomson scattering (TS) diagnostic on Experimental Advanced Superconducting Tokamak (EAST), it is necessary to characterize the pedestal with a suitable functional form. Based on simulated and experimental data of EAST, it is shown that the two-line method with a bilinear fitting has better reproducibility of pedestal parameters than hyperbolic tangent (tanh) and modified hyperbolic tangent (mtanh) methods. This method has been applied to EAST type I edge localized mode (ELM) discharges, and the electron pedestal density is found to be proportional to the line-averaged density and the edge pressure gradient is found to be proportional to the pedestal pressure. Furthermore, the ion poloidal gyro-radius has been identified as the suitable parameter to describe the pedestal pressure width. supported by National Natural Science Foundation of China (Nos. 11275233 and 11405206), and the National Magnetic Confinement Fusion Science Program of China (No. 2013GB112003), and Science Foundation of Institute of Plasma Physics, Chinese Academy of Sciences (No. DSJJ-15-JC01)

  13. Stability and ELM Characterization in I-Mode Pedestals

    NASA Astrophysics Data System (ADS)

    Walk, J. R.; Hughes, J. W.; Snyder, P. B.; Hubbard, A. E.; Terry, J. L.; White, A. E.; Whyte, D. G.; Baek, S. G.; Cziegler, I.; Edlund, E.

    2014-10-01

    The I-mode is a novel high-confinement regime explored on Alcator C-Mod, notable for its formation of an H-mode-like temperature pedestal without the accompanying density pedestal, maintaining L-mode particle confinement. I-mode exhibits a number of desirable properties for a reactor regime: among them, it naturally lacks large ELMs, avoiding the need for externally-applied ELM suppression. However, under certain conditions small, intermittent ELM-like events are seen. These events exhibit a range of phenomena in terms of edge and pedestal behavior, particularly for the ELM trigger - the majority of events are synchronized with the sawtooth heat pulse reaching the edge. The stationary pedestal structure is stable against peeling-ballooning MHD as calculated by ELITE in all cases, necessitating treatment of transient pedestal modification to characterize these events. We characterize these ELM events in terms of edge behavior, particularly the modification of the temperature pedestal, edge turbulence and fluctuations, and peeling-ballooning MHD stability. This work is supported by USDoE Award DE-FC02-99ER54512.

  14. Actin Automata: Phenomenology and Localizations

    NASA Astrophysics Data System (ADS)

    Adamatzky, Andrew; Mayne, Richard

    Actin is a globular protein which forms long filaments in the eukaryotic cytoskeleton, whose roles in cell function include structural support, contractile activity to intracellular signaling. We model actin filaments as two chains of one-dimensional binary-state semi-totalistic automaton arrays to describe hypothetical signaling events therein. Each node of the actin automaton takes state "0" (resting) or "1" (excited) and updates its state in discrete time depending on its neighbor's states. We analyze the complete rule space of actin automata using integral characteristics of space-time configurations generated by these rules and compute state transition rules that support traveling and mobile localizations. Approaches towards selection of the localization supporting rules using the global characteristics are outlined. We find that some properties of actin automata rules may be predicted using Shannon entropy, activity and incoherence of excitation between the polymer chains. We also show that it is possible to infer whether a given rule supports traveling or stationary localizations by looking at ratios of excited neighbors that are essential for generations of the localizations. We conclude by applying biomolecular hypotheses to this model and discuss the significance of our findings in context with cell signaling and emergent behavior in cellular computation.

  15. Signalling Pathways Controlling Cellular Actin Organization.

    PubMed

    Steffen, Anika; Stradal, Theresia E B; Rottner, Klemens

    2017-01-01

    The actin cytoskeleton is essential for morphogenesis and virtually all types of cell shape changes. Reorganization is per definition driven by continuous disassembly and re-assembly of actin filaments, controlled by major, ubiquitously operating machines. These are specifically employed by the cell to tune its activities in accordance with respective environmental conditions or to satisfy specific needs.Here we sketch some fundamental signalling pathways established to contribute to the reorganization of specific actin structures at the plasma membrane. Rho-family GTPases are at the core of these pathways, and dissection of their precise contributions to actin reorganization in different cell types and tissues will thus continue to improve our understanding of these important signalling nodes. Furthermore, we will draw your attention to the emerging theme of actin reorganization on intracellular membranes, its functional relation to Rho-GTPase signalling, and its relevance for the exciting phenomenon autophagy.

  16. Actinic Prurigo.

    PubMed

    Rodríguez-Carreón, Alma Angélica; Rodríguez-Lobato, Erika; Rodríguez-Gutiérrez, Georgina; Cuevas-González, Juan Carlos; Mancheno-Valencia, Alexandra; Solís-Arias, Martha Patricia; Vega-Memije, María Elisa; Hojyo-Tomoka, María Teresa; Domínguez-Soto, Luciano

    2015-01-01

    Actinic prurigo is an idiopathic photodermatosis that affects the skin, as well as the labial and conjunctival mucosa in indigenous and mestizo populations of Latin America. It starts predominantly in childhood, has a chronic course, and is exacerbated with solar exposure. Little is known of its pathophysiology, including the known mechanisms of the participation of HLA-DR4 and an abnormal immunologic response with increase of T CD4+ lymphocytes. The presence of IgE, eosinophils, and mast cells suggests that it is a hypersensitivity reaction (likely type IVa or b). The diagnosis is clinical, and the presence of lymphoid follicles in the mucosal histopathologic study of mucosa is pathognomonic. The best available treatment to date is thalidomide, despite its secondary effects.

  17. Dendritic Actin Nucleation Causes Traveling Waves and Patches

    NASA Astrophysics Data System (ADS)

    Carlsson, Anders

    2010-03-01

    Reversible polymerization of the intracellular protein actin into semiflexible filaments is crucial for cell motion and environmental sensing. Recent studies have shown that polymerized actin can spontaneously form traveling waves and/or moving patches. I investigate possible mechanisms for such phenomena by numerically simulating the ``dendritic nucleation'' model of actin network growth. The simulations treat the growth of an actin network on a flat portion of a cell membrane, using a stochastic-growth method which calculates an explicit three-dimensional network structure. The calculations treat processes including filament growth, capping, branching, severing, and Brownian motion. The dynamics of membrane proteins stimulating actin polymerization are also included: they diffuse in the membrane, and detach/deactivate in the presence of polymerized actin. The simulations show three types of polymerized-actin behavior: 1) traveling waves, 2) coherently moving patches, and 3) random fluctuations with occasional moving patches. Wave formation is favored at low free-actin concentrations by a long reattachment time for the membrane proteins, and by weakness of the attractive interaction between filaments and the membrane. Raising the free-actin concentration results in a randomly varying distribution of polymerized actin. Lowering the free-actin concentration below the optimal value for waves causes the waves to break up into patches which, however, move coherently. Effects of similar magnitude are predicted when other intracellular protein concentrations are varied. Diffusion of the membrane proteins slows the waves, and, if fast enough, stops them completely, resulting in the formation of a static spot.

  18. Turbulence driven particle pinch at the pedestal region in EAST

    NASA Astrophysics Data System (ADS)

    Yan, Ning; Xu, Sheng; Chen, Liang; Lan, Heng

    2015-11-01

    Existence of an anomalous inward particle flux in tokamak has been realized for a long time. Since particle transport up the density gradient particularly play a key role on the formation of edge pedestal in H-mode plasma, intensive efforts were made to identify the origin of particle pinch in tokamak. However, the mechanism of particle pinch is still a big challenge for plasma physics. In order to improve our understanding on inward particle pinch, turbulent transport have been investigated in boundary plasma with reciprocating Langmuir probe measurements on EAST. A broad turbulence is detected in pedestal after the L-H transition, which shows as fast (300kHz-500kHz) fluctuations in floating potential signals. It appears and gradually saturates after the dithering phase. The broad turbulence is ultimately terminated by the break out of ELMs. It is observed that most of the particle transport is outward directed before the emergence of broad turbulence. However, the particle transport is reversed to inward direction once the broad turbulence initializes in pedestal. Moreover, the edge pedestal starts to establish at the onset of the observed broad turbulence. It gradually stabilizes with the saturation of broad turbulence. During this period, the fluctuations and associated transport in the SOL are almost unaffected, which suggests a signature of particle pinch induced by the broad turbulence originating at the pedestal region. Institute of Plasma Physics, Chinese Academy of Sciences, Hefei, China

  19. Neoclassical transport in density pedestals with non-trace impurities

    NASA Astrophysics Data System (ADS)

    Buller, Stefan; Pusztai, Istvan; Landreman, Matt

    2016-10-01

    We study neoclassical transport in steady-state density pedestals with non-trace impurities using the Eulerian δf code Perfect, with an emphasis on radially global effects and the effects of impurities. To properly describe transport in a tokamak pedestal, radial coupling must be included, which strongly affects the transport. We find that radial coupling reduces the pedestal heat flux compared to local predictions. Furthermore, the influence of the pedestal persists several orbit widths into the core. The electron flux is significant in the pedestal, and global neoclassical transport is not intrinsically ambipolar. Thus, the impurity flux is not simply opposing the ion flux. The resulting radial current gives a torque that is balanced by a non-negligible radial transport of toroidal momentum. The effective Prandtl number is comparable to typical turbulent values in the core (0.1 - 0.3), and is sensitive to the impurity content. Global effects have a strong contribution to the poloidal flows of low- Z ions, which give rise to larger in-out flow asymmetries. Supported by the INCA Grant of Vetenskapsrådet (Dnr. 330-2014-6313). ML is supported by the USDoE Grants DEFG0293ER54197 and DEFC0208ER54964. The simulations used computational resources of Hebbe at C3SE (C3SE2016-1-10 & SNIC2016-1-161).

  20. A gyrokinetic perspective on the JET-ILW pedestal

    NASA Astrophysics Data System (ADS)

    Hatch, D. R.; Kotschenreuther, M.; Mahajan, S.; Valanju, P.; Liu, X.

    2017-03-01

    JET has been unable to recover historical confinement levels when operating with an ITER-like wall (ILW) due largely to the inaccessibility of high pedestal temperatures. Finding a path to overcome this challenge is of utmost importance for both a prospective JET DT campaign and for future ITER operation. Gyrokinetic simulations (using the Gene code) quantitatively capture experimental transport levels for a representative experimental discharge and qualitatively recover the major experimental trends. Microtearing turbulence is a major transport mechanisms for the low-temperature pedestals characteristic of unseeded JET-ILW discharges. At higher temperatures and/or lower {ρ\\ast} , we identify electrostatic ITG transport of a type that is strongly shear-suppressed on smaller machines. Consistent with observations, this transport mechanism is strongly reduced by the presence of a low-Z impurity (e.g. carbon or nitrogen at the level of {{Z}\\text{eff}}∼ 2 ), recovering the accessibility of high pedestal temperatures. Notably, simulations based on dimensionless {ρ\\ast} scans recover historical scaling behavior except in the unique JET-ILW parameter regime where ITG turbulence becomes important. Our simulations also elucidate the observed degradation of confinement caused by gas puffing, emphasizing the important role of the density pedestal structure. This study maps out important regions of parameter space, providing insights that may point to optimal physical regimes that can enable the recovery of high pedestal temperatures on JET.

  1. Actin and Actin-Binding Proteins.

    PubMed

    Pollard, Thomas D

    2016-08-01

    Organisms from all domains of life depend on filaments of the protein actin to provide structure and to support internal movements. Many eukaryotic cells use forces produced by actin polymerization for their motility, and myosin motor proteins use ATP hydrolysis to produce force on actin filaments. Actin polymerizes spontaneously, followed by hydrolysis of a bound adenosine triphosphate (ATP). Dissociation of the γ-phosphate prepares the polymer for disassembly. This review provides an overview of the properties of actin and shows how dozens of proteins control both the assembly and disassembly of actin filaments. These players catalyze nucleotide exchange on actin monomers, initiate polymerization, promote phosphate dissociation, cap the ends of polymers, cross-link filaments to each other and other cellular components, and sever filaments. Copyright © 2016 Cold Spring Harbor Laboratory Press; all rights reserved.

  2. Progress in characterizing pedestal stability on Alcator C-Mod

    NASA Astrophysics Data System (ADS)

    Hughes, J. W.; Churchill, R. M.; Cziegler, I.; Davis, E. M.; Dominguez, A.; Ennever, P.; Ernst, D.; Hubbard, A. E.; Lipschultz, B.; Ma, Y.; Walk, J. R.; Wolfe, S. M.; Snyder, P. B.; Osborne, T.; Xu, X.; Sugiyama, L.

    2011-10-01

    Experimental studies on Alcator C-Mod explore pedestal structure and edge relaxation mechanisms primarily in three high confinement regimes: ELMy H-mode, EDA H-mode and I-mode. Extensive scans of BT, IP, ne, Pnet and shaping parameters have been carried out in these regimes, allowing the characterization of the operating space for ELMs as well as benign pedestal relaxation mechanisms (e.g. quasi- and weakly coherent modes). Well resolved edge profiles and accurate equilibrium reconstructions are produced from typical discharges, enabling evaluation of edge stability using various computational tools, such as ELITE, BOUT++, M3D and GS2. Relationships among dominant edge instabilities, radial transport and pedestal structure will be discussed. Supported by US DoE Award DE-FC02-99-ER54512.

  3. MODEL OF THE TOKAMAK EDGE DENSITY PEDESTAL INCLUDING DIFFUSIVE NEUTRALS

    SciTech Connect

    BURRELL.KH

    2003-01-01

    OAK-B135 Several previous analytic models of the tokamak edge density pedestal have been based on diffusive transport of plasma plus free-streaming of neutrals. This latter neutral model includes only the effect of ionization and neglects charge exchange. The present work models the edge density pedestal using diffusive transport for both the plasma and the neutrals. In contrast to the free-streaming model, a diffusion model for the neutrals includes the effect of both charge exchange and ionization and is valid when charge exchange is the dominant interaction. Surprisingly, the functional forms for the electron and neutral density profiles from the present calculation are identical to the results of the previous analytic models. There are some differences in the detailed definition of various parameters in the solution. For experimentally relevant cases where ionization and charge exchange rate are comparable, both models predict approximately the same width for the edge density pedestal.

  4. Chromatic discrimination in the presence of incremental and decremental rod pedestals.

    PubMed

    Cao, Dingcai; Zele, Andrew J; Pokorny, Joel

    2008-01-01

    Signals from rods can alter chromatic discrimination. Here, chromatic discrimination ellipses were determined in the presence of rod incremental and decremental pedestals at mesopic light levels. The data were represented in a relative cone Troland space, normalized by discrimination thresholds measured along the cardinal axes without a rod pedestal. In the quadrant of cone space where L-cone relative to M-cone excitation increased, and S-cone excitation decreased, rod incremental pedestals degraded chromatic discrimination, and rod decremental pedestals improved chromatic discrimination. Discrimination in the other three quadrants of cone space was unaffected by the incremental or decremental rod pedestals. A second experiment measured chromatic discrimination under conditions where cone pedestals were matched to the appearances of the incremental and decremental rod pedestals. Based on the matching pedestal data, discrimination then could be measured independently along the cardinal axes using either chromatic [L/(L + M); S/(L + M)] or luminance (L + M) pedestal components. The discrimination data altered by the rod pedestals were similar to chromatic cone pedestals for L/M increment discrimination, but similar to luminance cone pedestals for S decrement discrimination. The results indicated that the rod and cone signals combined differently in determining chromatic discrimination for different post-receptoral pathways.

  5. Actin cytoskeletal defects in immunodeficiency

    PubMed Central

    Moulding, Dale A; Record, Julien; Malinova, Dessislava; Thrasher, Adrian J

    2013-01-01

    The importance of the cytoskeleton in mounting a successful immune response is evident from the wide range of defects that occur in actin-related primary immunodeficiencies (PIDs). Studies of these PIDs have revealed a pivotal role for the actin cytoskeleton in almost all stages of immune system function, from hematopoiesis and immune cell development, through to recruitment, migration, intercellular and intracellular signaling, and activation of both innate and adaptive immune responses. The major focus of this review is the immune defects that result from mutations in the Wiskott-Aldrich syndrome gene (WAS), which have a broad impact on many different processes and give rise to clinically heterogeneous immunodeficiencies. We also discuss other related genetic defects and the possibility of identifying new genetic causes of cytoskeletal immunodeficiency. PMID:24117828

  6. Actin dynamics shape microglia effector functions.

    PubMed

    Uhlemann, Ria; Gertz, Karen; Boehmerle, Wolfgang; Schwarz, Tobias; Nolte, Christiane; Freyer, Dorette; Kettenmann, Helmut; Endres, Matthias; Kronenberg, Golo

    2016-06-01

    Impaired actin filament dynamics have been associated with cellular senescence. Microglia, the resident immune cells of the brain, are emerging as a central pathophysiological player in neurodegeneration. Microglia activation, which ranges on a continuum between classical and alternative, may be of critical importance to brain disease. Using genetic and pharmacological manipulations, we studied the effects of alterations in actin dynamics on microglia effector functions. Disruption of actin dynamics did not affect transcription of genes involved in the LPS-triggered classical inflammatory response. By contrast, in consequence of impaired nuclear translocation of phospho-STAT6, genes involved in IL-4 induced alternative activation were strongly downregulated. Functionally, impaired actin dynamics resulted in reduced NO secretion and reduced release of TNFalpha and IL-6 from LPS-stimulated microglia and of IGF-1 from IL-4 stimulated microglia. However, pathological stabilization of the actin cytoskeleton increased LPS-induced release of IL-1beta and IL-18, which belong to an unconventional secretory pathway. Reduced NO release was associated with decreased cytoplasmic iNOS protein expression and decreased intracellular arginine uptake. Furthermore, disruption of actin dynamics resulted in reduced microglia migration, proliferation and phagocytosis. Finally, baseline and ATP-induced [Ca(2+)]int levels were significantly increased in microglia lacking gelsolin, a key actin-severing protein. Together, the dynamic state of the actin cytoskeleton profoundly and distinctly affects microglia behaviours. Disruption of actin dynamics attenuates M2 polarization by inhibiting transcription of alternative activation genes. In classical activation, the role of actin remodelling is complex, does not relate to gene transcription and shows a major divergence between cytokines following conventional and unconventional secretion.

  7. [Cytoskeletal actin and its associated proteins. Some examples in Protista].

    PubMed

    Guillén, N; Carlier, M F; Brugerolle, G; Tardieux, I; Ausseil, J

    1998-06-01

    Many processes, cell motility being an example, require cells to remodel the actin cytoskeleton in response to both intracellular and extracellular signals. Reorganization of the actin cytoskeleton involves the rapid disassembly and reassembly of actin filaments, a phenomenon regulated by the action of particular actin-binding proteins. In recent years, an interest in studying actin regulation in unicellular organisms has arisen. Parasitic protozoan are among these organisms and studies of the cytoskeleton functions of these protozoan are relevant related to either cell biology or pathogenicity. To discuss recent data in this field, a symposium concerning "Actin and actin-binding proteins in protists" was held on May 8-11 in Paris, France, during the XXXV meeting of the French Society of Protistology. As a brief summary of the symposium we report here findings concerning the in vitro actin dynamic assembly, as well as the characterization of several actin-binding proteins from the parasitic protozoan Entamoeba histolytica, Trichomonas vaginalis and Plasmodium knowlesi. In addition, localization of actin in non-pathogen protists such as Prorocentrum micans and Crypthecodinium cohnii is also presented. The data show that some actin-binding proteins facilitate organization of filaments into higher order structures as pseudopods, while others have regulatory functions, indicating very particular roles for actin-binding proteins. One of the proteins discussed during the symposium, the actin depolymerizing factor ADF, was shown to enhance the treadmilling rate of actin filaments. In vitro, ADF binds to the ADP-bound forms of G-actin and F-actin, thereby participating in and changing the rate of actin assembly. Biochemical approaches allowed the identification of a protein complex formed by HSP/C70-cap32-34 which might also be involved in depolymerization of F-actin in P. knowlesi. Molecular and cellular approaches were used to identify proteins such as ABP-120 and myosin

  8. Actin filament curvature biases branching direction

    PubMed Central

    Risca, Viviana I.; Wang, Evan B.; Chaudhuri, Ovijit; Chia, Jia Jun; Geissler, Phillip L.; Fletcher, Daniel A.

    2012-01-01

    Mechanical cues affect many important biological processes in metazoan cells, such as migration, proliferation, and differentiation. Such cues are thought to be detected by specialized mechanosensing molecules linked to the cytoskeleton, an intracellular network of protein filaments that provide mechanical rigidity to the cell and drive cellular shape change. The most abundant such filament, actin, forms branched networks nucleated by the actin-related protein (Arp) 2/3 complex that support or induce membrane protrusions and display adaptive behavior in response to compressive forces. Here we show that filamentous actin serves in a mechanosensitive capacity itself, by biasing the location of actin branch nucleation in response to filament bending. Using an in vitro assay to measure branching from curved sections of immobilized actin filaments, we observed preferential branch formation by the Arp2/3 complex on the convex face of the curved filament. To explain this behavior, we propose a fluctuation gating model in which filament binding or branch nucleation by Arp2/3 occur only when a sufficiently large, transient, local curvature fluctuation causes a favorable conformational change in the filament, and we show with Monte Carlo simulations that this model can quantitatively account for our experimental data. We also show how the branching bias can reinforce actin networks in response to compressive forces. These results demonstrate how filament curvature can alter the interaction of cytoskeletal filaments with regulatory proteins, suggesting that direct mechanotransduction by actin may serve as a general mechanism for organizing the cytoskeleton in response to force. PMID:22308368

  9. Control of bootstrap current in the pedestal region of tokamaks

    SciTech Connect

    Shaing, K. C.; Lai, A. L.

    2013-12-15

    The high confinement mode (H-mode) plasmas in the pedestal region of tokamaks are characterized by steep gradient of the radial electric field, and sonic poloidal U{sub p,m} flow that consists of poloidal components of the E×B flow and the plasma flow velocity that is parallel to the magnetic field B. Here, E is the electric field. The bootstrap current that is important for the equilibrium, and stability of the pedestal of H-mode plasmas is shown to have an expression different from that in the conventional theory. In the limit where ‖U{sub p,m}‖≫ 1, the bootstrap current is driven by the electron temperature gradient and inductive electric field fundamentally different from that in the conventional theory. The bootstrap current in the pedestal region can be controlled through manipulating U{sub p,m} and the gradient of the radial electric. This, in turn, can control plasma stability such as edge-localized modes. Quantitative evaluations of various coefficients are shown to illustrate that the bootstrap current remains finite when ‖U{sub p,m}‖ approaches infinite and to provide indications how to control the bootstrap current. Approximate analytic expressions for viscous coefficients that join results in the banana and plateau-Pfirsch-Schluter regimes are presented to facilitate bootstrap and neoclassical transport simulations in the pedestal region.

  10. Observation of Pedestal Plasma Turbulence on EAST Tokamak

    NASA Astrophysics Data System (ADS)

    Gao, Xiang; Zhang, Tao; Han, Xiang; Zhang, Shoubiao; Wang, Yumin; Liu, Zixi; Yang, Yao; Liu, Shaocheng; Shi, Nan; Ling, Bili; Li, Jiangang; The EAST Team

    2013-08-01

    Pedestal plasma turbulence was experimentally studied by microwave reflectometry on EAST tokamak. The characteristics of edge pedestal turbulence during dithering L-H transition, ELM-free H-mode phase and inter-ELM phase have recently been studied on EAST. An edge spatial structure of density fluctuation and its dithering temporal evolution is observed for the first time on the EAST tokamak during the L-H transition phase. A coherent mode usually appears during the ELM-free phase prior to the first ELM on EAST tokamak. The mode frequency gradually decreases as the pedestal evolves. Analysis shows that the coherent mode is in the pedestal region inside the separatrix. In plasma with type-III ELMs, a precursor mode before ELM is usually observed. The frequency of the precursor was initially about 150 kHz and gradually decreased till the next ELM. The mode amplitude increases or shows saturation before ELM. In the plasma with compound ELMs composed of high and low frequency ELMs, the precursor was also observed before the high frequency ELM while the harmonic oscillations with frequencies of 20 kHz, 40 kHz and 60 kHz appear before the low frequency ELM.

  11. Placed on a Pedestal: Famous Faces in Clay

    ERIC Educational Resources Information Center

    Walkup, Nancy

    2010-01-01

    Artists have created portraits of people for thousands of years. In sculpture, a portrait of a person's face often includes the neck and part of the shoulders and chest. These artworks are called portrait busts. In this article, the author describes how her fifth-grade students created clay portrait busts on pedestal columns. The objectives are…

  12. Placed on a Pedestal: Famous Faces in Clay

    ERIC Educational Resources Information Center

    Walkup, Nancy

    2010-01-01

    Artists have created portraits of people for thousands of years. In sculpture, a portrait of a person's face often includes the neck and part of the shoulders and chest. These artworks are called portrait busts. In this article, the author describes how her fifth-grade students created clay portrait busts on pedestal columns. The objectives are…

  13. Kinetic neoclassical transport in the H-mode pedestal

    SciTech Connect

    Battaglia, D. J.; Burrell, K. H.; Chang, C. S.; Ku, S.; deGrassie, J. S.; Grierson, B. A.

    2014-07-16

    Multi-species kinetic neoclassical transport through the QH-mode pedestal and scrapeoff layer on DIII-D is calculated using XGC0, a 5D full-f particle-in-cell drift-kinetic solver with self-consistent neutral recycling and sheath potentials. We achieved quantitative agreement between the fluxdriven simulation and the experimental electron density, impurity density and orthogonal measurements of impurity temperature and flow profiles by adding random-walk particle diffusion to the guiding-center drift motion. Furthermore, we computed the radial electric field (Er) that maintains ambipolar transport across flux surfaces and to the wall self-consistently on closed and open magnetic field lines, and is in excellent agreement with experiment. The Er inside the separatrix is the unique solution that balances the outward flux of thermal tail deuterium ions against the outward neoclassical electron flux and inward pinch of impurity and colder deuterium ions. Particle transport in the pedestal is primarily due to anomalous transport, while the ion heat and momentum transport is primarily due to the neoclassical transport. The full-f treatment quantifies the non-Maxwellian energy distributions that describe a number of experimental observations in low-collisionallity pedestals on DIII-D, including intrinsic co-Ip parallel flows in the pedestal, ion temperature anisotropy and large impurity temperatures in the scrape-off layer.

  14. Kinetic neoclassical transport in the H-mode pedestal

    SciTech Connect

    Battaglia, D. J.; Chang, C. S.; Ku, S.; Grierson, B. A.; Burrell, K. H.; Grassie, J. S. de

    2014-07-15

    Multi-species kinetic neoclassical transport through the QH-mode pedestal and scrape-off layer on DIII-D is calculated using XGC0, a 5D full-f particle-in-cell drift-kinetic solver with self-consistent neutral recycling and sheath potentials. Quantitative agreement between the flux-driven simulation and the experimental electron density, impurity density, and orthogonal measurements of impurity temperature and flow profiles is achieved by adding random-walk particle diffusion to the guiding-center drift motion. The radial electric field (E{sub r}) that maintains ambipolar transport across flux surfaces and to the wall is computed self-consistently on closed and open magnetic field lines and is in excellent agreement with experiment. The E{sub r} inside the separatrix is the unique solution that balances the outward flux of thermal tail deuterium ions against the outward neoclassical electron flux and inward pinch of impurity and colder deuterium ions. Particle transport in the pedestal is primarily due to anomalous transport, while the ion heat and momentum transport are primarily due to the neoclassical transport. The full-f treatment quantifies the non-Maxwellian energy distributions that describe a number of experimental observations in low-collisionallity pedestals on DIII-D, including intrinsic co-I{sub p} parallel flows in the pedestal, ion temperature anisotropy, and large impurity temperatures in the scrape-off layer.

  15. Kinetic neoclassical transport in the H-mode pedestal

    DOE PAGES

    Battaglia, D. J.; Burrell, K. H.; Chang, C. S.; ...

    2014-07-16

    Multi-species kinetic neoclassical transport through the QH-mode pedestal and scrapeoff layer on DIII-D is calculated using XGC0, a 5D full-f particle-in-cell drift-kinetic solver with self-consistent neutral recycling and sheath potentials. We achieved quantitative agreement between the fluxdriven simulation and the experimental electron density, impurity density and orthogonal measurements of impurity temperature and flow profiles by adding random-walk particle diffusion to the guiding-center drift motion. Furthermore, we computed the radial electric field (Er) that maintains ambipolar transport across flux surfaces and to the wall self-consistently on closed and open magnetic field lines, and is in excellent agreement with experiment. The Ermore » inside the separatrix is the unique solution that balances the outward flux of thermal tail deuterium ions against the outward neoclassical electron flux and inward pinch of impurity and colder deuterium ions. Particle transport in the pedestal is primarily due to anomalous transport, while the ion heat and momentum transport is primarily due to the neoclassical transport. The full-f treatment quantifies the non-Maxwellian energy distributions that describe a number of experimental observations in low-collisionallity pedestals on DIII-D, including intrinsic co-Ip parallel flows in the pedestal, ion temperature anisotropy and large impurity temperatures in the scrape-off layer.« less

  16. 24. A SECTION OF OPEN BALUSTRADE AND AN INTERMEDIATE PEDESTAL ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    24. A SECTION OF OPEN BALUSTRADE AND AN INTERMEDIATE PEDESTAL (CENTERED ON A PYLON) OVER THE CENTRAL PORTIONS OF THE BRIDGE. THE WALKS ARE CONCRETE, WITH AN INTEGRAL CURB. THE ROADWAY IS ASPHALT. - Main Street Bridge, Spanning East Fork Whitewater River, Richmond, Wayne County, IN

  17. Actin network disassembly powers dissemination of Listeria monocytogenes

    PubMed Central

    Talman, Arthur M.; Chong, Ryan; Chia, Jonathan; Svitkina, Tatyana; Agaisse, Hervé

    2014-01-01

    ABSTRACT Several bacterial pathogens hijack the actin assembly machinery and display intracellular motility in the cytosol of infected cells. At the cell cortex, intracellular motility leads to bacterial dissemination through formation of plasma membrane protrusions that resolve into vacuoles in adjacent cells. Here, we uncover a crucial role for actin network disassembly in dissemination of Listeria monocytogenes. We found that defects in the disassembly machinery decreased the rate of actin tail turnover but did not affect the velocity of the bacteria in the cytosol. By contrast, defects in the disassembly machinery had a dramatic impact on bacterial dissemination. Our results suggest a model of L. monocytogenes dissemination in which the disassembly machinery, through local recycling of the actin network in protrusions, fuels continuous actin assembly at the bacterial pole and concurrently exhausts cytoskeleton components from the network distal to the bacterium, which enables membrane apposition and resolution of protrusions into vacuoles. PMID:24155331

  18. Edge-localized mode avoidance and pedestal structure in I-mode plasmasa)

    NASA Astrophysics Data System (ADS)

    Walk, J. R.; Hughes, J. W.; Hubbard, A. E.; Terry, J. L.; Whyte, D. G.; White, A. E.; Baek, S. G.; Reinke, M. L.; Theiler, C.; Churchill, R. M.; Rice, J. E.; Snyder, P. B.; Osborne, T.; Dominguez, A.; Cziegler, I.

    2014-05-01

    I-mode is a high-performance tokamak regime characterized by the formation of a temperature pedestal and enhanced energy confinement, without an accompanying density pedestal or drop in particle and impurity transport. I-mode operation appears to have naturally occurring suppression of large Edge-Localized Modes (ELMs) in addition to its highly favorable scalings of pedestal structure and overall performance. Extensive study of the ELMy H-mode has led to the development of the EPED model, which utilizes calculations of coupled peeling-ballooning MHD modes and kinetic-ballooning mode (KBM) stability limits to predict the pedestal structure preceding an ELM crash. We apply similar tools to the structure and ELM stability of I-mode pedestals. Analysis of I-mode discharges prepared with high-resolution pedestal data from the most recent C-Mod campaign reveals favorable pedestal scalings for extrapolation to large machines—pedestal temperature scales strongly with power per particle Pnet/n ¯e, and likewise pedestal pressure scales as the net heating power (consistent with weak degradation of confinement with heating power). Matched discharges in current, field, and shaping demonstrate the decoupling of energy and particle transport in I-mode, increasing fueling to span nearly a factor of two in density while maintaining matched temperature pedestals with consistent levels of Pnet/n ¯e. This is consistent with targets for increased performance in I-mode, elevating pedestal βp and global performance with matched increases in density and heating power. MHD calculations using the ELITE code indicate that I-mode pedestals are strongly stable to edge peeling-ballooning instabilities. Likewise, numerical modeling of the KBM turbulence onset, as well as scalings of the pedestal width with poloidal beta, indicates that I-mode pedestals are not limited by KBM turbulence—both features identified with the trigger for large ELMs, consistent with the observed suppression of

  19. Resemblance of actin-binding protein/actin gels to covalently crosslinked networks

    NASA Astrophysics Data System (ADS)

    Janmey, Paul A.; Hvidt, Søren; Lamb, Jennifer; Stossel, Thomas P.

    1990-05-01

    THE maintainance of the shape of cells is often due to their surface elasticity, which arises mainly from an actin-rich cytoplasmic cortex1,2. On locomotion, phagocytosis or fission, however, these cells become partially fluid-like. The finding of proteins that can bind to actin and control the assembly of, or crosslink, actin filaments, and of intracellular messages that regulate the activities of some of these actin-binding proteins, indicates that such 'gel sol' transformations result from the rearrangement of cortical actin-rich networks3. Alternatively, on the basis of a study of the mechanical properties of mixtures of actin filaments and an Acanthamoeba actin-binding protein, α-actinin, it has been proposed that these transformations can be accounted for by rapid exchange of crosslinks between actin filaments4: the cortical network would be solid when the deformation rate is greater than the rate of crosslink exchange, but would deform or 'creep' when deformation is slow enough to permit crosslinker molecules to rearrange. Here we report, however, that mixtures of actin filaments and actin-binding protein (ABP), an actin crosslinking protein of many higher eukaryotes, form gels Theologically equivalent to covalently crosslinked networks. These gels do not creep in response to applied stress on a time scale compatible with most cell-surface movements. These findings support a more complex and controlled mechanism underlying the dynamic mechanical properties of cortical cytoplasm, and can explain why cells do not collapse under the constant shear forces that often exist in tissues.

  20. Thymosin beta4: actin regulation and more.

    PubMed

    Yarmola, Elena G; Klimenko, Evguenia S; Fujita, Go; Bubb, Michael R

    2007-09-01

    The intracellular function of thymosin beta(4) is not limited to simple sequestration of globular actin. Our recent studies revealed that thymosin beta(4) affects actin critical concentration and forms a ternary complex with actin and profilin. The consequences of this complex formation can be very significant. Our new data demonstrate that it is likely that profilin affects binding of thymosin beta(4) to actin in the ternary complex through allosteric changes in actin rather than through competition for the binding site. The N- and C-terminal thymosin beta(4) helices are known to be unstructured in aqueous solution and to adopt helical conformation in organic solvents or upon binding to actin. Osmolytes stabilize protein structure, and TMAO (trimethylamine N-oxide) specifically stabilizes hydrogen bonds. This increases affinity of intact thymosin beta(4) to actin significantly, but the increase is much less for thymosin beta(4) sulfoxide. Our data show that oxidation does not alter binding of profilin to form a ternary complex, and therefore it is very likely that there is no direct steric interference by methionine 6 of thymosin beta(4). Rather, since TMAO has little effect on thymosin beta(4) sulfoxide, this observation is consistent with the hypothesis that methionine oxidation prevents helix transition. The experiment with truncated versions of thymosin beta(4) also supports this hypothesis. Oxidation and formation of the helices are important for both intra- and extracellular properties of thymosin beta(4). We found that actin and, in lesser extent, profilin-actin complex protect thymosin beta(4) from oxidation.

  1. Pharmacological characterization of actin-binding (-)-doliculide.

    PubMed

    Foerster, Florian; Braig, Simone; Chen, Tao; Altmann, Karl-Heinz; Vollmar, Angelika M

    2014-09-15

    Natural compounds offer a broad spectrum of potential drug candidates against human malignancies. Several cytostatic drugs, which are in clinical use for decades, derive directly from natural sources or are synthetically optimized derivatives of natural lead structures. An eukaryote target molecule to which many natural derived anti-cancer drugs bind to is the microtubule network. Of similar importance for the cell is the actin cytoskeleton, responsible for cell movements, migration of cells and cytokinesis. Nature provides also a broad range of compounds directed against actin as intracellular target, but none of these actin-targeting compounds has ever been brought to clinical trials. One reason why actin-binding compounds have not yet been considered for further clinical investigations is that little is known about their pharmacological properties in cancer cells. Herein, we focused on the closer characterization of doliculide, an actin binding natural compound of marine origin in the breast cancer cell lines MCF7 and MDA-MB-231. We used fluorescence-recovery-after-photobleaching (FRAP) analysis to determine doliculide's early effects on the actin cytoskeleton and rhodamin-phalloidin staining for long-term effects on the actin CSK. After validating the disruption of the actin network, we further investigated the functional effects of doliculide. Doliculide treatment leads to inhibition of proliferation and impairs the migratory potential. Finally, we could also show that doliculide leads to the induction of apoptosis in both cell lines. Our data for the first time provide a closer characterization of doliculide in breast cancer cells and propagate doliculide for further investigations as lead structure and potential therapeutic option as actin-targeting compound.

  2. Global and pedestal confinement and pedestal structure in dimensionless collisionality scans of low-triangularity H-mode plasmas in JET-ILW

    NASA Astrophysics Data System (ADS)

    Frassinetti, L.; Beurskens, M. N. A.; Saarelma, S.; Boom, J. E.; Delabie, E.; Flanagan, J.; Kempenaars, M.; Giroud, C.; Lomas, P.; Meneses, L.; Maggi, C. S.; Menmuir, S.; Nunes, I.; Rimini, F.; Stefanikova, E.; Urano, H.; Verdoolaege, G.; Contributors, JET

    2017-01-01

    A dimensionless collisionality scan in low-triangularity plasmas in the Joint European Torus with the ITER-like wall (JET-ILW) has been performed. The increase of the normalized energy confinement (defined as the ratio between thermal energy confinement and Bohm confinement time) with decreasing collisionality is observed. Moreover, at low collisionality, a confinement factor H98, comparable to JET-C, is achieved. At high collisionality, the low normalized confinement is related to a degraded pedestal stability and a reduction in the density-profile peaking. The increase of normalized energy confinement is due to both an increase in the pedestal and in the core regions. The improvement in the pedestal is related to the increase of the stability. The improvement in the core is driven by (i) the core temperature increase via the temperature-profile stiffness and by (ii) the density-peaking increase driven by the low collisionality. Pedestal stability analysis performed with the ELITE (edge-localized instabilities in tokamak equilibria) code has a reasonable qualitative agreement with the experimental results. An improvement of the pedestal stability with decreasing collisionality is observed. The improvement is ascribed to the reduction of the pedestal width, the increase of the bootstrap current and the reduction of the relative shift between the positions of the pedestal density and pedestal temperature. The EPED1 model predictions for the pedestal pressure height are qualitatively well correlated with the experimental results. Quantitatively, EPED1 overestimates the experimental pressure by 15-35%. In terms of the pedestal width, a correct agreement (within 10-15%) between the EPED1 and the experimental width is found at low collisionality. The experimental pedestal width increases with collisionality. Nonetheless, an extrapolation to low-collisionality values suggests that the width predictions from the KBM constraint are reasonable for ITER.

  3. Circular F-actin bundles and a G-actin gradient in pollen and pollen tubes of Lilium davidii.

    PubMed

    Li, Y; Zee, S Y; Liu, Y M; Huang, B Q; Yen, L F

    2001-09-01

    The distribution of and relationship between F-actin and G-actin were investigated in pollen grains and pollen tubes of Lilium davidii Duch. using a confocal laser scanning microscope after fluorescence and immunofluorescence labeling. Circular F-actin bundles were found to be the main form of microfilament cytoskeleton in pollen grains and pollen tubes. Consistent with cytoplasmic streaming in pollen tubes, there were no obvious F-actin bundles in the 10- to 20-microm tip region of long pollen tubes, only a few short F-actin fragments. Labeling with fluorescein isothiocyanate (FITC)-DNase I at first established the presence of a tip-focused gradient of intracellular G-actin concentration at the extreme apex of the tube, the concentration of G-actin being about twice as high in the 10- to 20-microm region of the tip as in other regions of the pollen tube. We also found that the distribution of G-actin was related negatively to that of the F-actin in pollen tubes of L. davidii. Caffeine treatment caused the G-actin tip-focused gradient to disappear, and F-actin to extend into the pollen tube tip. Based on these results, we speculate that the circular F-actin bundles may be the track for bidirectional cytoplasmic streaming in pollen tubes, and that in the pollen tube tip most of the F-actin is depolymerized into G-actin, leading to the absence of F-actin bundles in this region.

  4. Quantitative Studies of Endothelial Cell Fibronectin and Filamentous Actin (F-Actin) Coalignment in Response to Shear Stress.

    PubMed

    Gong, Xianghui; Zhao, Xixi; Li, Bin; Sun, Yan; Liu, Meili; Huang, Yan; Jia, Xiaoling; Ji, Jing; Fan, Yubo

    2017-09-12

    Both fibronectin (FN) and filamentous actin (F-actin) fibers play a critical role for endothelial cells (ECs) in responding to shear stress and modulating cell alignment and functions. FN is dynamically coupled to the F-actin cytoskeleton via focal adhesions. However, it is unclear how ECs cooperatively remodel their subcellular FN matrix and intracellular F-actin cytoskeleton in response to shear stress. Current studies are hampered by the lack of a reliable and sensitive quantification method of FN orientation. In this study, we developed a MATLAB-based feature enhancement method to quantify FN and F-actin orientation. The role of F-actin in FN remodeling was also studied by treating ECs with cytochalasin D. We have demonstrated that FN and F-actin codistributed and coaligned parallel to the flow direction, and that F-actin alignment played an essential role in regulating FN alignment in response to shear stress. Our findings offer insight into how ECs cooperatively remodel their subcellular ECM and intracellular F-actin cytoskeleton in response to mechanical stimuli, and are valuable for vascular tissue engineering.

  5. Relative effects of increment and pedestal duration on the detection of intensity increments a

    PubMed Central

    Valente, Daniel L.; Patra, Harisadhan; Jesteadt, Walt

    2011-01-01

    The detection of a brief increment in the intensity of a longer duration pedestal is commonly used as a measure of intensity-resolution. Increment detection is known to improve with increasing duration of the increment and also with increasing duration of the pedestal, but the relative effects of these two parameters have not been explored in the same study. In several past studies of the effects of increment duration, pedestal duration was increased as increment duration increased. In the present study, increment and pedestal duration were independently manipulated. Increment-detection thresholds were determined for four subjects with normal-hearing using a 500- or 4000-Hz pedestal presented at 60 dB sound pressure level (SPL). Increment durations were 10, 20, 40, 80, 160, and 320 ms. Pedestal durations were 20, 40, 80, 160, and 320 ms. Each increment duration was combined with all pedestals of equal or greater duration. Multiple-regression analyses indicate that increment detection under these conditions is determined primarily by pedestal duration. Follow-up experiments ruled out effects of off-frequency listening or overshoot. The results suggest that effects of increment duration have been confounded by effects of pedestal duration in studies that co-varied increment and pedestal duration. Implications for models of temporal integration are discussed. PMID:21476665

  6. Advancing the predictive capability for pedestal structure through experiment and modeling

    NASA Astrophysics Data System (ADS)

    Hughes, Jerry

    2012-10-01

    Prospects for predictive capability of the edge pedestal in magnetic fusion devices have been dramatically enhanced due to recent research, which was conducted jointly by the US experimental and theory communities. Studies on the C-Mod, DIII-D and NSTX devices have revealed common features, including an upper limit on pedestal pressure in ELMy H-mode determined by instability to peeling-ballooning modes (PBMs), and pedestal width which scales approximately as βpol^1/2. The width dependence is consistent with a pedestal regulated by kinetic ballooning modes (KBMs). Signatures of KBMs have been actively sought both in experimental fluctuation measurements and in gyrokinetic simulations of the pedestal, with encouraging results. Studies of the temporal evolution of the pedestal during the ELM cycle reveal a tendency for the pressure gradient to saturate in advance of the ELM, with a steady growth in the pedestal width occurring prior to the ELM crash, which further supports a model for KBMs and PBMs working together to set the pedestal structure. Such a model, EPED, reproduces the pedestal height and width to better than 20% accuracy on existing devices over a range of more than 20 in pedestal pressure. Additional transport processes are assessed for their impact on pedestal structure, in particular the relative variation of the temperature and density pedestals due, for example, to differences in edge neutral sources. Such differences are observed in dimensionlessly matched discharges on C-Mod and DIII-D, despite their having similar calculated MHD stability and similar edge fluctuations. In certain high performance discharges, such as EDA H-mode, QH-mode and I-mode, pedestal relaxation is accomplished by continuous edge fluctuations, avoiding peeling-ballooning instabilities and associated ELMs. Progress in understanding these regimes will be reported.

  7. H-mode pedestal scaling in DIII-D, ASDEX Upgrade, and JET

    SciTech Connect

    Beurskens, M. N. A.; Lomas, P.; Saarelma, S.; Scannell, R.; Balboa, I.; Brix, M.; Flanagan, J.; Giroud, C.; Kempenaars, M.; Maddison, G.; McDonald, D.; Schneider, P. A.; Wolfrum, E.; Maggi, C. F.; Frassinetti, L.; Nunes, I.

    2011-05-15

    Multidevice pedestal scaling experiments in the DIII-D, ASDEX Upgrade (AUG), and JET tokamaks are presented in order to test two plasma physics pedestal width models. The first model proposes a scaling of the pedestal width {Delta}/a {proportional_to} {rho}*{sup 1/2} to {rho}* based on the radial extent of the pedestal being set by the point where the linear turbulence growth rate exceeds the ExB velocity. In the multidevice experiment where {rho}* at the pedestal top was varied by a factor of four while other dimensionless parameters where kept fixed, it has been observed that the temperature pedestal width in real space coordinates scales with machine size, and that therefore the gyroradius scaling suggested by the model is not supported by the experiments. The density pedestal width is not invariant with {rho}* which after comparison with a simple neutral fuelling model may be attributed to variations in the neutral fuelling patterns. The second model, EPED1, is based on kinetic ballooning modes setting the limit of the radial extent of the pedestal region and leads to {Delta}{sub {psi} {proportional_to}} {beta}{sub p}{sup 1/2}. All three devices show a scaling of the pedestal width in normalised poloidal flux as {Delta}{sub {psi} {proportional_to}} {beta}{sub p}{sup 1/2}, as described by the kinetic ballooning model; however, on JET and AUG, this could not be distinguished from an interpretation where the pedestal is fixed in real space. Pedestal data from all three devices have been compared with the predictive pedestal model EPED1 and the model produces pedestal height values that match the experimental data well.

  8. Scaling studies of the H-mode pedestal

    SciTech Connect

    Groebner, R.J.; Osborne, T.H.

    1998-01-01

    The structure and scaling of the H-mode pedestal are examined for discharges in the DIII-D tokamak. For typical conditions, the pedestal values of the ion and electron temperatures T{sub i} and T{sub e} are comparable. Measurements of main ion and C{sup 6+} profiles indicate that the ion pressure gradient in the barrier is 50%--100% of the electron pressure gradient for deuterium plasmas. The magnitude of the pressure gradient in the barrier often exceeds the predictions of infinite-n ballooning mode theory by a factor of two. Moreover, via the bootstrap current, the finite pressure gradient acts to entirely remove ballooning stability limits for typical discharges. For a large dataset, the width of the pressure barrier {delta} is best described by the dimensionless scaling {delta}/R {proportional_to} ({beta}{sub pol}{sup ped}){sup 0.4} where ({beta}{sub pol}{sup ped}) is the pedestal value of poloidal beta and R is the major radius. Scalings based on the poloidal ion gyroradius or the edge density gradient do not adequately describe overall trends in the data set and the propagation of the pressure barrier observed between edge-localized modes. The width of the T{sub i} barrier is quite variable and is not a good measure of the width of the pressure barrier.

  9. Gyrokinetic simulations of microturbulence in DIII-D tokamak pedestal

    NASA Astrophysics Data System (ADS)

    Holod, Ihor; Fulton, Daniel; Taimourzadeh, Sam; Lin, Zhihong; Nazikian, Raffi; Spong, Donald

    2015-11-01

    The characteristics of H-mode pedestal are generally believed to be constrained by current-driven peeling-ballooning modes and pressure-driven instabilities, such as kinetic ballooning mode (KBM). In this work we use global gyrokinetic code (GTC) to identify and study the edge pressure-driven instabilities in the H-mode pedestal using realistic geometry and plasma profiles of DIII-D shot 131997. In our simulations we observe the KBM mode marginally dominant in the steep gradient region (ψN = 0 . 98), in the range of kθ ~ 1 cm-1 which corresponds to the most unstable mode number in the nonlinearly saturated state. For shorter wavelengths the trapped electron mode becomes dominant since its linear growth rate increases with the mode number, while the KBM gets saturated. In the pedestal top region (ψN = 0 . 95) the ITG dominates. Resonant magnetic perturbations (RMP) are widely applied for ELM mitigation. During RMP suppression, the increase of edge turbulence is often observed. To understand this phenomena we use gyrokinetic simulations to address the direct effect of magnetic perturbations on the microturbulence. Simulations with 3D equilibrium reconstructed by VMEC code have been compared with toroidally averaged equilibrium, using identical pressure profiles. Work supported by DOE grant DE-SC0010416 and by General Atomics subcontract.

  10. A Gyrokinetic Perspective on the JET-ILW Pedestal

    NASA Astrophysics Data System (ADS)

    Hatch, David; Kotschenreuther, Mike; Mahajan, Swadesh; Valanju, Prashant; Liu, Xing

    2016-10-01

    Simulations using the GENE code based on JET-ILW profiles and equilibria quantitatively capture experimental transport levels for a representative experimental discharge and qualitatively recover the major experimental trends. Microtearing turbulence is a major transport mechanism for the low-temperature pedestals characteristic of unseeded JET-ILW discharges. At higher temperatures, we identify electrostatic ITG-like transport of a type that is strongly shear-suppressed on smaller machines. Consistent with observations, this transport mechanism is substantially reduced by the presence of a low-Z impurity (e.g., carbon or nitrogen at the level of Z-effective 2). Multiple transport mechanisms, including ITG, ETG, microtearing modes, and neoclassical transport are found to play important roles depending on the pedestal parameters. The picture that emerges involves several parameters-notably, rho*, Z-effective, pedestal top temperature, and separatrix density-mediating the relative roles of these transport mechanisms. This study maps out important regions of this parameter space, providing insights that may point to optimal physical regimes that can enable the recovery of high temperatures on JET.

  11. Coupled simulation of kinetic pedestal growth and MHD ELM crash

    SciTech Connect

    Park, G.; Cummings, J.; Chang, C. S.; Klasky, Scott A; Ku, S.; Podhorszki, Norbert; Pankin, A.; Samtaney, Ravi; Shoshani, A.; Snyder, P.; Strauss, H.; Sugiyama, L.; CPES Team, the

    2007-01-01

    Edge pedestal height and the accompanying ELM crash are critical elements of ITER physics yet to be understood and predicted through high performance computing. An entirely self-consistent first principles simulation is being pursued as a long term research goal, and the plan is planned for completion in time for ITER operation. However, a proof-of-principle work has already been established using a computational tool that employs the best first principles physics available at the present time. A kinetic edge equilibrium code XGC0, which can simulate the neoclassically dominant pedestal growth from neutral ionization (using a phenomenological residual turbulence diffusion motion superposed upon the neoclassical particle motion) is coupled to an extended MHD code M3D, which can perform the nonlinear ELM crash. The stability boundary of the pedestal is checked by an ideal MHD linear peeling-ballooning code, which has been validated against many experimental data sets for the large scale (type I) ELMs onset boundary. The coupling workflow and scientific results to be enabled by it are described.

  12. Coupled simulation of kinetic pedestal growth and MHD ELM crash

    SciTech Connect

    Park, G-Y; Cummings, J.; Chang, C S; Podhorszki, Norbert; Klasky, Scott A; Ku, S.; Pankin, A.; Samtaney, Ravi; Shoshani, A.; Snyder, P.; Sugiyama, L.

    2009-01-01

    Edge pedestal height and the accompanying ELM crash are critical elements of ITER physics yet to be understood and predicted through high performance computing. An entirely self-consistent first principles simulation is being pursued as a long term research goal, and the plan is planned for completion in time for ITER operation. However, a proof-of-principle work has already been established using a computational tool that employs the best first principles physics available at the present time. A kinetic edge equilibrium code XGC0, which can simulate the neoclassically dominant pedestal growth from neutral ionization (using a phenomenological residual turbulence diffusion motion superposed upon the neoclassical particle motion) is coupled to an extended MHD code M3D, which can perform the nonlinear ELM crash. The stability boundary of the pedestal is checked by an ideal MHD linear peeling-ballooning code, which has been validated against many experimental data sets for the large scale (type I) ELMs onset boundary. The coupling workflow and scientific results to be enabled by it are described.

  13. Enteropathogenic Escherichia coli and Vaccinia Virus Do Not Require the Family of WASP-Interacting Proteins for Pathogen-Induced Actin Assembly

    PubMed Central

    Garber, John J.; Takeshima, Fuminao; Antón, Inés M.; Oyoshi, Michiko K.; Lyubimova, Anna; Kapoor, Archana; Shibata, Tomoyuki; Chen, Feng; Alt, Frederick W.; Geha, Raif S.; Leong, John M.

    2012-01-01

    The human pathogens enteropathogenic Escherichia coli (EPEC) and vaccinia virus trigger actin assembly in host cells by activating the host adaptor Nck and the actin nucleation promoter neural Wiskott-Aldrich syndrome protein (N-WASP). EPEC translocates effector molecules into host cells via type III secretion, and the interaction between the translocated intimin receptor (Tir) and the bacterial membrane protein intimin stimulates Nck and N-WASP recruitment, leading to the formation of actin pedestals beneath adherent bacteria. Vaccinia virus also recruits Nck and N-WASP to generate actin tails that promote cell-to-cell spread of the virus. In addition to Nck and N-WASP, WASP-interacting protein (WIP) localizes to vaccinia virus tails, and inhibition of actin tail formation upon ectopic expression of WIP mutants led to the suggestion that WIP is required for this process. Similar studies of WIP mutants, however, did not affect the ability of EPEC to form actin pedestals, arguing against an essential role for WIP in EPEC-induced actin assembly. In this study, we demonstrate that Nck and N-WASP are normally recruited by vaccinia virus and EPEC in the absence of WIP, and neither WIP nor the WIP family members CR16 and WIRE/WICH are essential for pathogen induced actin assembly. In addition, although Nck binds EPEC Tir directly, N-WASP is required for its localization during pedestal formation. Overall, these data highlight similar pathogenic strategies shared by EPEC and vaccinia virus by demonstrating a requirement for both Nck and N-WASP, but not WIP or WIP family members in pathogen-induced actin assembly. PMID:22966049

  14. Enteropathogenic Escherichia coli and vaccinia virus do not require the family of WASP-interacting proteins for pathogen-induced actin assembly.

    PubMed

    Garber, John J; Takeshima, Fuminao; Antón, Inés M; Oyoshi, Michiko K; Lyubimova, Anna; Kapoor, Archana; Shibata, Tomoyuki; Chen, Feng; Alt, Frederick W; Geha, Raif S; Leong, John M; Snapper, Scott B

    2012-12-01

    The human pathogens enteropathogenic Escherichia coli (EPEC) and vaccinia virus trigger actin assembly in host cells by activating the host adaptor Nck and the actin nucleation promoter neural Wiskott-Aldrich syndrome protein (N-WASP). EPEC translocates effector molecules into host cells via type III secretion, and the interaction between the translocated intimin receptor (Tir) and the bacterial membrane protein intimin stimulates Nck and N-WASP recruitment, leading to the formation of actin pedestals beneath adherent bacteria. Vaccinia virus also recruits Nck and N-WASP to generate actin tails that promote cell-to-cell spread of the virus. In addition to Nck and N-WASP, WASP-interacting protein (WIP) localizes to vaccinia virus tails, and inhibition of actin tail formation upon ectopic expression of WIP mutants led to the suggestion that WIP is required for this process. Similar studies of WIP mutants, however, did not affect the ability of EPEC to form actin pedestals, arguing against an essential role for WIP in EPEC-induced actin assembly. In this study, we demonstrate that Nck and N-WASP are normally recruited by vaccinia virus and EPEC in the absence of WIP, and neither WIP nor the WIP family members CR16 and WIRE/WICH are essential for pathogen induced actin assembly. In addition, although Nck binds EPEC Tir directly, N-WASP is required for its localization during pedestal formation. Overall, these data highlight similar pathogenic strategies shared by EPEC and vaccinia virus by demonstrating a requirement for both Nck and N-WASP, but not WIP or WIP family members in pathogen-induced actin assembly.

  15. Actin dynamics in Phytophthora infestans; rapidly reorganizing cables and immobile, long-lived plaques.

    PubMed

    Meijer, Harold J G; Hua, Chenlei; Kots, Kiki; Ketelaar, Tijs; Govers, Francine

    2014-06-01

    The actin cytoskeleton is a dynamic but well-organized intracellular framework that is essential for proper functioning of eukaryotic cells. Here, we use the actin binding peptide Lifeact to investigate the in vivo actin cytoskeleton dynamics in the oomycete plant pathogen Phytophthora infestans. Lifeact-eGFP labelled thick and thin actin bundles and actin filament plaques allowing visualization of actin dynamics. All actin structures in the hyphae were cortically localized. In growing hyphae actin filament cables were axially oriented in the sub-apical region whereas in the extreme apex in growing hyphae, waves of fine F-actin polymerization were observed. Upon growth termination, actin filament plaques appeared in the hyphal tip. The distance between a hyphal tip and the first actin filament plaque correlated strongly with hyphal growth velocity. The actin filament plaques were nearly immobile with average lifetimes exceeding 1 h, relatively long when compared to the lifetime of actin patches known in other eukaryotes. Plaque assembly required ∼30 s while disassembly was accomplished in ∼10 s. Remarkably, plaque disassembly was not accompanied with internalization and the formation of endocytic vesicles. These findings suggest that the functions of actin plaques in oomycetes differ from those of actin patches present in other organisms.

  16. Degradation of picosecond temporal contrast of Ti:sapphire lasers with coherent pedestals.

    PubMed

    Khodakovskiy, Nikita; Kalashnikov, Mikhail; Gontier, Emilien; Falcoz, Franck; Paul, Pierre-Mary

    2016-10-01

    Recompressed pulses from Ti:sapphire chirped-pulse lasers are accompanied by a slowly decaying post-pulse pedestal that is coherent with the main pulse. The pedestal typically consists of numerous pulses with temporal separation in the picosecond range. The source of this artifact lies in the Ti:sapphire active medium itself, both in the Kerr-lens mode-locked oscillator and in subsequent amplifiers. In the presence of substantial self-phase modulation, after recompression the post-pedestal generates a mirror-symmetric pre-pulse pedestal. This pedestal severely degrades the leading edge of the output pulse. This degradation is far more limiting than the original post-pedestal and severely lowers the achievable temporal contrast.

  17. A Legionella Effector Disrupts Host Cytoskeletal Structure by Cleaving Actin

    PubMed Central

    Liu, Yao; Zhu, Wenhan; Tan, Yunhao; Nakayasu, Ernesto S.; Staiger, Christopher J.

    2017-01-01

    Legionella pneumophila, the etiological agent of Legionnaires’ disease, replicates intracellularly in protozoan and human hosts. Successful colonization and replication of this pathogen in host cells requires the Dot/Icm type IVB secretion system, which translocates approximately 300 effector proteins into the host cell to modulate various cellular processes. In this study, we identified RavK as a Dot/Icm substrate that targets the host cytoskeleton and reduces actin filament abundance in mammalian cells upon ectopic expression. RavK harbors an H95EXXH99 motif associated with diverse metalloproteases, which is essential for the inhibition of yeast growth and for the induction of cell rounding in HEK293T cells. We demonstrate that the actin protein itself is the cellular target of RavK and that this effector cleaves actin at a site between residues Thr351 and Phe352. Importantly, RavK-mediated actin cleavage also occurs during L. pneumophila infection. Cleavage by RavK abolishes the ability of actin to form polymers. Furthermore, an F352A mutation renders actin resistant to RavK-mediated cleavage; expression of the mutant in mammalian cells suppresses the cell rounding phenotype caused by RavK, further establishing that actin is the physiological substrate of RavK. Thus, L. pneumophila exploits components of the host cytoskeleton by multiple effectors with distinct mechanisms, highlighting the importance of modulating cellular processes governed by the actin cytoskeleton in the intracellular life cycle of this pathogen. PMID:28129393

  18. Extracellular motility and cell-to-cell transmission of enterohemorrhagic E. coli is driven by EspFU-mediated actin assembly

    PubMed Central

    Velle, Katrina B.

    2017-01-01

    Enteropathogenic and enterohemorrhagic Escherichia coli (EPEC and EHEC) are closely-related pathogens that attach tightly to intestinal epithelial cells, efface microvilli, and promote cytoskeletal rearrangements into protrusions called actin pedestals. To trigger pedestal formation, EPEC employs the tyrosine phosphorylated transmembrane receptor Tir, while EHEC relies on the multivalent scaffolding protein EspFU. The ability to generate these structures correlates with bacterial colonization in several animal models, but the precise function of pedestals in infection remains unclear. To address this uncertainty, we characterized the colonization properties of EPEC and EHEC during infection of polarized epithelial cells. We found that EPEC and EHEC both formed distinct bacterial communities, or “macrocolonies,” that encompassed multiple host cells. Tir and EspFU, as well as the host Arp2/3 complex, were all critical for the expansion of macrocolonies over time. Unexpectedly, EspFU accelerated the formation of larger macrocolonies compared to EPEC Tir, as EspFU-mediated actin assembly drove faster bacterial motility to cell junctions, where bacteria formed a secondary pedestal on a neighboring cell and divided, allowing one of the daughters to disengage and infect the second cell. Collectively, these data reveal that EspFU enhances epithelial colonization by increasing actin-based motility and promoting an efficient method of cell-to-cell transmission. PMID:28771584

  19. Correlation of Density Pedestal Width and Neutral Penetration

    NASA Astrophysics Data System (ADS)

    Xu, X. Q.; Nevins, W. M.; Cohen, R. H.; Rognlien, T. D.; Umansky, M. V.

    2003-10-01

    Pedestal studies in DIII-D and C-Mod find a good correlation between the width of the H-mode density barrier and the neutral penetration length.[1][2] These results suggest that the width may be set by the combined effects of neutral and plasma transport. This paper is a report on fluid simulations of boundary plasma using BOUT code [3] with neutral source added. Thus both neutral and plasma physics are treated. The plasma transport is self-consistently driven by boundary turbulence due to the resistive X-point mode, while neutral is described by a simple fluid diffusive model. The plasma profiles are evolved on the same time scale as the turbulence for the given heat source from the core plasma and particle source from the neutrals. For prescribed neutral profiles, we find the formation of a density pedestal inside the separatrix in the L-mode even though the calculated plasma diffusion coefficients are almost radially constant and without the formation of a temperature pedestal. These results support the hypothesis that particle fueling can provide the dominant control for the size of the H-mode density barrier. The width of the density barrier, and its relationship to pedestal height and neutral penetration length at the midplane, will be given by using hyperbolic-tangent fit to the simulation data. [1] R.J.Groebner, M.A.Mahdavi, A.W.Leonard, 19th IAEA Fusion Energy Conference(Lyon, France, 14 to 19 October 2002), IAEA-CN-94/EX/C2-3. [2] D.Mossessian, J.W.Hughes, M.Greenwald, et al., ``Local dimensionless identity method as a tool for studying H-mode pedestal'', The 9th Joint US-European TTF workshop (Madison, Wisconsin, April 2nd - 5th, 2003). [3] X.Q.Xu, R.H.Cohen, T.D.Rognlien and J.R.Myra, Physics of Plasma, Vol. 7, 1951-1958 (2000).

  20. [Nucleotide receptors and actin cytoskeleton dynamics].

    PubMed

    Kłopocka, Wanda; Korczyński, Jarosław

    2014-01-01

    Signaling cascades evoked by P2Y2 receptor plays an important role in the phenomena dependent on the actin cytoskeleton dynamics endocy-tosis, cell division, adhesion, intracellular transport and migration. P2Y2R coupled with G proteins, in response to ATP or UTP activates Rac1 and RhoA proteins important factors in actin cytoskeletal reorganization and regulates the level of phosphatidylinositol-4,5-bisphosphate (PIP2) that binds directly to a variety of actin regulatory proteins and modulates their function. The P2Y2 nucleotide receptor contains the integrin-binding domain enables it to interact selectively with α(v)β3 and α(v)β5 integrins and is required for G0-mediated Rac1 activation. Interaction with α(v)β5 is necessary for coupling the P2Y2 receptor to G12 and subsequent activation of RhoA.

  1. Cofilin mediates ATP depletion-induced endothelial cell actin alterations.

    PubMed

    Suurna, Maria V; Ashworth, Sharon L; Hosford, Melanie; Sandoval, Ruben M; Wean, Sarah E; Shah, Bijal M; Bamburg, James R; Molitoris, Bruce A

    2006-06-01

    Ischemia and sepsis lead to endothelial cell damage, resulting in compromised microvascular flow in many organs. Much remains to be determined regarding the intracellular structural events that lead to endothelial cell dysfunction. To investigate potential actin cytoskeletal-related mechanisms, ATP depletion was induced in mouse pancreatic microvascular endothelial cells (MS1). Fluorescent imaging and biochemical studies demonstrated a rapid and progressive increase in F-actin along with a decrease in G-actin at 60 min. Confocal microscopic analysis showed ATP depletion resulted in destruction of actin stress fibers and accumulation of F-actin aggregates. We hypothesized these actin alterations were secondary to dephosphorylation/activation of actin-depolymerizing factor (ADF)/cofilin proteins. Cofilin, the predominant isoform expressed in MS1 cells, was rapidly dephosphorylated/activated during ATP depletion. To directly investigate the role of cofilin activation on the actin cytoskeleton during ischemia, MS1 cells were infected with adenoviruses containing the cDNAs for wild-type Xenopus laevis ADF/cofilin green fluorescent protein [XAC(wt)-GFP], GFP, and the constitutively active and inactive isoforms XAC(S3A)-GFP and XAC(S3E)-GFP. The rate and extent of cortical actin destruction and actin aggregate formation were increased in ATP-depleted XAC(wt)-GFP- and XAC(S3A)-GFP-expressing cells, whereas increased actin stress fibers were observed in XAC(S3E)-GFP-expressing cells. To investigate the upstream signaling pathway of ADF/cofilin, LIM kinase 1-GFP (LIMK1-GFP) was expressed in MS1 cells. Cells expressing LIMK1-GFP protein had higher levels of phosphorylated ADF/cofilin, increased stress fibers, and delayed F-actin cytoskeleton destruction during ATP depletion. These results strongly support the importance of cofilin regulation in ischemia-induced endothelial cell actin cytoskeleton alterations leading to cell damage and microvascular dysfunction.

  2. Dynamical Evolution of Pedestal Parameters in ELMy H-mode in the National Spherical Torus Experiment

    SciTech Connect

    Diallo, A; Kubota, S; Sontag, A; Osborne, T; Podesta, M; Bell, R E; LeBlanc, B P; Menard, J

    2011-07-27

    Characterizations of the pedestal parameter dynamics throughout the edge localized modes(ELM) cycles are performed on the National Spherical Torus Experiment (NSTX, [M. Ono et al., Nucl. Fusion 40, 557 (2000)]). A clear buildup of the pedestal height is observed between ELMs for three di erent plasma currents, which tends to saturate prior to the onset of ELM at low and medium plasma current. Similarly, the pedestal width increases with no clear evidence of saturation during an ELM cycle. The maximum pedestal gradient increases as a function of plasma current, reaches a nominal value after the ELM crash, and remains constant until the end of the ELM cycle. The pedestal height just prior to the onset of ELM is shown to increase quadratically with plasma current. The pedestal width Δ is proportional to the square-root of the poloidal Β at the top of the pedestal. Coherent density uctuations strongly increasing at the plasma edge are observed to be maximum after the ELM crash and to decay during the rest of the ELM cycle. Finally, the pedestal parameters evolution during the ELM cycle as well as the scaling with Ip of the pedestal pressure prior to the onset ELM are found to be qualitatively consistent with the peeling ballooning theory.

  3. Pedestal structure and inter-ELM evolution for different main ion species in ASDEX Upgrade

    NASA Astrophysics Data System (ADS)

    Laggner, F. M.; Wolfrum, E.; Cavedon, M.; Mink, F.; Bernert, M.; Dunne, M. G.; Schneider, P. A.; Kappatou, A.; Birkenmeier, G.; Fischer, R.; Willensdorfer, M.; Aumayr, F.

    2017-05-01

    In tokamak plasmas with different main ion species, a change in confinement occurs, known as the isotope effect. Experiments comparing hydrogen (H), deuterium (D), and helium (4He) plasmas have been performed to identify processes that define the pedestal structure and evolution in between the crashes of edge localized modes (ELMs). The pedestal top electron densities and temperatures have been matched to compare the pedestal shape and stability. In the D and H discharges, the pedestal electron temperature profiles do not differ, whereas the density profile in H has shallower gradients. Furthermore, the heat flux across the pedestal in H is roughly a factor of two higher than in D. In 4He plasmas at similar stored energy, the pedestal top electron density is roughly a factor of 1.5 larger than in the references owing to the larger effective charge. The peeling-ballooning theory, which is independent of the main ion species mass, can sufficiently describe the pedestal stability in the hydrogenic plasmas. The inter-ELM pedestal evolution has the same sequence of recovery phases for all investigated species, giving evidence that similar mechanisms are acting in the pedestals. This is further supported by a similar evolution of the inter-ELM magnetic signature and the corresponding toroidal structure.

  4. Development and validation of a predictive model for the pedestal height

    SciTech Connect

    Snyder, P. B.; Groebner, R. J.; Leonard, A. W.; Osborne, T. H.; Wilson, H. R.

    2009-05-15

    The pressure at the top of the edge transport barrier (or 'pedestal height') strongly impacts tokamak fusion performance. Predicting the pedestal height in future devices such as ITER [ITER Physics Basis Editors, Nucl. Fusion 39, 2137 (1999)] remains an important challenge. While uncertainties remain, magnetohydrodynamic stability calculations at intermediate wavelength (the ''peeling-ballooning'' model), accounting for diamagnetic stabilization, have been largely successful in determining the observed maximum pedestal height, when the edge barrier width is taken as an input. Here, we develop a second relation between the pedestal width in normalized poloidal flux ({delta}) and pedestal height ({delta}=0.076{beta}{sub {theta}}{sub ,ped}{sup 1/2}), using an argument based upon kinetic ballooning mode turbulence and observation. Combining this relation with direct calculations of peeling-ballooning stability yields two constraints, which together determine both the height and width of the pedestal. The resulting model, EPED1, allows quantitative prediction of the pedestal height and width in both existing and future experiments. EPED1 is successfully tested both against a dedicated experiment on the DIII-D [J. L. Luxon, Nucl. Fusion 42, 614 (2002)] tokamak, in which predictions were made before the experiment, and against a broader DIII-D data set, including ITER demonstration discharges. EPED1 is found to quantitatively capture the observed complex dependencies of the pedestal height and width. An initial set of pedestal predictions for the ITER device is presented.

  5. Dynamical Evolution of Pedestal Parameters in ELMy H-mode in the National Spherical Torus Experiment

    SciTech Connect

    Diallo, A.; Maingi, Rajesh; Kubota, S.; Sontag, Aaron C; Osborne, T.; Podesta, M.; Bell, R. E.; LeBlanc, B. P.; Menard, J.; Sabbagh, S. A.

    2011-01-01

    Characterizations of the pedestal parameter dynamics throughout the edge localized mode (ELM) cycles are performed on the National Spherical Torus Experiment (NSTX, (Ono et al 2000 Nucl. Fusion 40 557)). A clear buildup of the pedestal height between ELMs is observed for three different plasma currents. This buildup tends to saturate at low and medium plasma currents. Similarly, the pedestal width increases with no clear evidence of saturation during an ELM cycle. The maximum pedestal gradient increases as a function of plasma current, reaches a nominal value after the ELM crash, and remains constant until the end of the ELM cycle. The pedestal height just prior to the onset of ELM is shown to increase quadratically with plasma current. The pedestal width (Delta) scales as Delta = 0.17 root beta(ped)(theta). with the poloidal beta at the top of the pedestal. Coherent density fluctuations strongly increasing at the plasma edge are observed to be maximum after the ELM crash and to decay during the rest of the ELM cycle. Finally, the evolution of the pedestal height and width during the ELM cycle as well as the scaling with I(p) of the pedestal pressure prior to the onset ELM are found to be qualitatively consistent with the peeling-ballooning theory.

  6. Electrostatics control actin filament nucleation and elongation kinetics.

    PubMed

    Crevenna, Alvaro H; Naredi-Rainer, Nikolaus; Schönichen, André; Dzubiella, Joachim; Barber, Diane L; Lamb, Don C; Wedlich-Söldner, Roland

    2013-04-26

    The actin cytoskeleton is a central mediator of cellular morphogenesis, and rapid actin reorganization drives essential processes such as cell migration and cell division. Whereas several actin-binding proteins are known to be regulated by changes in intracellular pH, detailed information regarding the effect of pH on the actin dynamics itself is still lacking. Here, we combine bulk assays, total internal reflection fluorescence microscopy, fluorescence fluctuation spectroscopy techniques, and theory to comprehensively characterize the effect of pH on actin polymerization. We show that both nucleation and elongation are strongly enhanced at acidic pH, with a maximum close to the pI of actin. Monomer association rates are similarly affected by pH at both ends, although dissociation rates are differentially affected. This indicates that electrostatics control the diffusional encounter but not the dissociation rate, which is critical for the establishment of actin filament asymmetry. A generic model of protein-protein interaction, including electrostatics, explains the observed pH sensitivity as a consequence of charge repulsion. The observed pH effect on actin in vitro agrees with measurements of Listeria propulsion in pH-controlled cells. pH regulation should therefore be considered as a modulator of actin dynamics in a cellular environment.

  7. Electrostatics Control Actin Filament Nucleation and Elongation Kinetics*

    PubMed Central

    Crevenna, Alvaro H.; Naredi-Rainer, Nikolaus; Schönichen, André; Dzubiella, Joachim; Barber, Diane L.; Lamb, Don C.; Wedlich-Söldner, Roland

    2013-01-01

    The actin cytoskeleton is a central mediator of cellular morphogenesis, and rapid actin reorganization drives essential processes such as cell migration and cell division. Whereas several actin-binding proteins are known to be regulated by changes in intracellular pH, detailed information regarding the effect of pH on the actin dynamics itself is still lacking. Here, we combine bulk assays, total internal reflection fluorescence microscopy, fluorescence fluctuation spectroscopy techniques, and theory to comprehensively characterize the effect of pH on actin polymerization. We show that both nucleation and elongation are strongly enhanced at acidic pH, with a maximum close to the pI of actin. Monomer association rates are similarly affected by pH at both ends, although dissociation rates are differentially affected. This indicates that electrostatics control the diffusional encounter but not the dissociation rate, which is critical for the establishment of actin filament asymmetry. A generic model of protein-protein interaction, including electrostatics, explains the observed pH sensitivity as a consequence of charge repulsion. The observed pH effect on actin in vitro agrees with measurements of Listeria propulsion in pH-controlled cells. pH regulation should therefore be considered as a modulator of actin dynamics in a cellular environment. PMID:23486468

  8. The yeast actin cytoskeleton.

    PubMed

    Mishra, Mithilesh; Huang, Junqi; Balasubramanian, Mohan K

    2014-03-01

    The actin cytoskeleton is a complex network of dynamic polymers, which plays an important role in various fundamental cellular processes, including maintenance of cell shape, polarity, cell division, cell migration, endocytosis, vesicular trafficking, and mechanosensation. Precise spatiotemporal assembly and disassembly of actin structures is regulated by the coordinated activity of about 100 highly conserved accessory proteins, which nucleate, elongate, cross-link, and sever actin filaments. Both in vivo studies in a wide range of organisms from yeast to metazoans and in vitro studies of purified proteins have helped shape the current understanding of actin dynamics and function. Molecular genetics, genome-wide functional analysis, sophisticated real-time imaging, and ultrastructural studies in concert with biochemical analysis have made yeast an attractive model to understand the actin cytoskeleton, its molecular dynamics, and physiological function. Studies of the yeast actin cytoskeleton have contributed substantially in defining the universal mechanism regulating actin assembly and disassembly in eukaryotes. Here, we review some of the important insights generated by the study of actin cytoskeleton in two important yeast models the budding yeast Saccharomyces cerevisiae and the fission yeast Schizosaccharomyces pombe. © 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

  9. Diclofenac Topical (actinic keratosis)

    MedlinePlus

    Solaraze® Gel ... Diclofenac topical gel (Solaraze) is used to treat actinic keratosis (flat, scaly growths on the skin caused by too much sun ... nonsteroidal anti-inflammatory drugs (NSAIDs). The way diclofenac gel works to treat actinic keratosis is not known. ...

  10. Role of Actin Polymerization in Cell Locomotion: Molecules and Models

    PubMed Central

    Bearer, E. L.

    2015-01-01

    Actin filaments forming at the anterior margin of a migrating cell are essential for the formation of filopodia, lamellipodia, and pseudopodia, the “feet” that the cell extends before it. These structures in turn are required for cell locomotion. Yet the molecular nature of the “nucleator” that seeds the polymerization of actin at the leading edge is unknown. Recent advances, including video microscopy of actin dynamics, discovery of proteins unique to the leading edge such as ponticulin, the Mab 2E4 antigen, and ABP 120, and novel experimental models of actin polymerization such as the actin-based movements of intracellular parasites, promise to shed light on this problem in the near future. PMID:8323743

  11. Unconventional actins and actin-binding proteins in human protozoan parasites.

    PubMed

    Gupta, C M; Thiyagarajan, S; Sahasrabuddhe, A A

    2015-06-01

    Actin and its regulatory proteins play a key role in several essential cellular processes such as cell movement, intracellular trafficking and cytokinesis in most eukaryotes. While these proteins are highly conserved in higher eukaryotes, a number of unicellular eukaryotic organisms contain divergent forms of these proteins which have highly unusual biochemical and structural properties. Here, we review the biochemical and structural properties of these unconventional actins and their core binding proteins which are present in commonly occurring human protozoan parasites. Copyright © 2015 Australian Society for Parasitology Inc. Published by Elsevier Ltd. All rights reserved.

  12. Fimbrin phosphorylation by metaphase Cdk1 regulates actin cable dynamics in budding yeast

    PubMed Central

    Miao, Yansong; Han, Xuemei; Zheng, Liangzhen; Xie, Ying; Mu, Yuguang; Yates, John R.; Drubin, David G.

    2016-01-01

    Actin cables, composed of actin filament bundles nucleated by formins, mediate intracellular transport for cell polarity establishment and maintenance. We previously observed that metaphase cells preferentially promote actin cable assembly through cyclin-dependent kinase 1 (Cdk1) activity. However, the relevant metaphase Cdk1 targets were not known. Here we show that the highly conserved actin filament crosslinking protein fimbrin is a critical Cdk1 target for actin cable assembly regulation in budding yeast. Fimbrin is specifically phosphorylated on threonine 103 by the metaphase cyclin–Cdk1 complex, in vivo and in vitro. On the basis of conformational simulations, we suggest that this phosphorylation stabilizes fimbrin's N-terminal domain, and modulates actin filament binding to regulate actin cable assembly and stability in cells. Overall, this work identifies fimbrin as a key target for cell cycle regulation of actin cable assembly in budding yeast, and suggests an underlying mechanism. PMID:27068241

  13. Fimbrin phosphorylation by metaphase Cdk1 regulates actin cable dynamics in budding yeast.

    PubMed

    Miao, Yansong; Han, Xuemei; Zheng, Liangzhen; Xie, Ying; Mu, Yuguang; Yates, John R; Drubin, David G

    2016-04-12

    Actin cables, composed of actin filament bundles nucleated by formins, mediate intracellular transport for cell polarity establishment and maintenance. We previously observed that metaphase cells preferentially promote actin cable assembly through cyclin-dependent kinase 1 (Cdk1) activity. However, the relevant metaphase Cdk1 targets were not known. Here we show that the highly conserved actin filament crosslinking protein fimbrin is a critical Cdk1 target for actin cable assembly regulation in budding yeast. Fimbrin is specifically phosphorylated on threonine 103 by the metaphase cyclin-Cdk1 complex, in vivo and in vitro. On the basis of conformational simulations, we suggest that this phosphorylation stabilizes fimbrin's N-terminal domain, and modulates actin filament binding to regulate actin cable assembly and stability in cells. Overall, this work identifies fimbrin as a key target for cell cycle regulation of actin cable assembly in budding yeast, and suggests an underlying mechanism.

  14. The centrosome is an actin-organizing center

    PubMed Central

    Farina, Francesca; Gaillard, Jérémie; Guérin, Christophe; Couté, Yohann; Sillibourne, James; Blanchoin, Laurent; Théry, Manuel

    2016-01-01

    Microtubules and actin filaments are the two main cytoskeleton networks supporting intracellular architecture and cell polarity. The centrosome nucleates and anchors microtubules and is therefore considered to be the main microtubule-organizing center. However, recurring, yet unexplained, observations have pointed towards a connection between the centrosome and actin filaments. Here we have used isolated centrosomes to demonstrate that the centrosome can directly promote actin filament assembly. A cloud of centrosome-associated actin filaments could be identified in living cells as well. Actin-filament nucleation at the centrosome was mediated by the nucleation promoting factor WASH in combination with the Arp2/3 complex. Pericentriolar material 1 (PCM1) appeared to modulate the centrosomal actin network by regulating Arp2/3 complex and WASH recruitment to the centrosome. Hence our results reveal an additional facet of the centrosome as an intracellular organizer and provide mechanistic insights into how the centrosome can function as an actin filament-organizing center. PMID:26655833

  15. The centrosome is an actin-organizing centre.

    PubMed

    Farina, Francesca; Gaillard, Jérémie; Guérin, Christophe; Couté, Yohann; Sillibourne, James; Blanchoin, Laurent; Théry, Manuel

    2016-01-01

    Microtubules and actin filaments are the two main cytoskeleton networks supporting intracellular architecture and cell polarity. The centrosome nucleates and anchors microtubules and is therefore considered to be the main microtubule-organizing centre. However, recurring, yet unexplained, observations have pointed towards a connection between the centrosome and actin filaments. Here we have used isolated centrosomes to demonstrate that the centrosome can directly promote actin-filament assembly. A cloud of centrosome-associated actin filaments could be identified in living cells as well. Actin-filament nucleation at the centrosome was mediated by the nucleation-promoting factor WASH in combination with the Arp2/3 complex. Pericentriolar material 1 (PCM1) seemed to modulate the centrosomal actin network by regulating Arp2/3 complex and WASH recruitment to the centrosome. Hence, our results reveal an additional facet of the centrosome as an intracellular organizer and provide mechanistic insights into how the centrosome can function as an actin-filament-organizing centre.

  16. Intracellular proteoglycans.

    PubMed Central

    Kolset, Svein Olav; Prydz, Kristian; Pejler, Gunnar

    2004-01-01

    Proteoglycans (PGs) are proteins with glycosaminoglycan chains, are ubiquitously expressed and have a wide range of functions. PGs in the extracellular matrix and on the cell surface have been the subject of extensive structural and functional studies. Less attention has so far been given to PGs located in intracellular compartments, although several reports suggest that these have biological functions in storage granules, the nucleus and other intracellular organelles. The purpose of this review is, therefore, to present some of these studies and to discuss possible functions linked to PGs located in different intracellular compartments. Reference will be made to publications relevant for the topics we present. It is beyond the scope of this review to cover all publications on PGs in intracellular locations. PMID:14759226

  17. Improved understanding of physics processes in pedestal structure, leading to improved predictive capability for ITER

    NASA Astrophysics Data System (ADS)

    Groebner, R. J.; Chang, C. S.; Hughes, J. W.; Maingi, R.; Snyder, P. B.; Xu, X. Q.; Boedo, J. A.; Boyle, D. P.; Callen, J. D.; Canik, J. M.; Cziegler, I.; Davis, E. M.; Diallo, A.; Diamond, P. H.; Elder, J. D.; Eldon, D. P.; Ernst, D. R.; Fulton, D. P.; Landreman, M.; Leonard, A. W.; Lore, J. D.; Osborne, T. H.; Pankin, A. Y.; Parker, S. E.; Rhodes, T. L.; Smith, S. P.; Sontag, A. C.; Stacey, W. M.; Walk, J.; Wan, W.; Wang, E. H.-J.; Watkins, J. G.; White, A. E.; Whyte, D. G.; Yan, Z.; Belli, E. A.; Bray, B. D.; Candy, J.; Churchill, R. M.; Deterly, T. M.; Doyle, E. J.; Fenstermacher, M. E.; Ferraro, N. M.; Hubbard, A. E.; Joseph, I.; Kinsey, J. E.; LaBombard, B.; Lasnier, C. J.; Lin, Z.; Lipschultz, B. L.; Liu, C.; Ma, Y.; McKee, G. R.; Ponce, D. M.; Rost, J. C.; Schmitz, L.; Staebler, G. M.; Sugiyama, L. E.; Terry, J. L.; Umansky, M. V.; Waltz, R. E.; Wolfe, S. M.; Zeng, L.; Zweben, S. J.

    2013-09-01

    Joint experiment/theory/modelling research has led to increased confidence in predictions of the pedestal height in ITER. This work was performed as part of a US Department of Energy Joint Research Target in FY11 to identify physics processes that control the H-mode pedestal structure. The study included experiments on C-Mod, DIII-D and NSTX as well as interpretation of experimental data with theory-based modelling codes. This work provides increased confidence in the ability of models for peeling-ballooning stability, bootstrap current, pedestal width and pedestal height scaling to make correct predictions, with some areas needing further work also being identified. A model for pedestal pressure height has made good predictions in existing machines for a range in pressure of a factor of 20. This provides a solid basis for predicting the maximum pedestal pressure height in ITER, which is found to be an extrapolation of a factor of 3 beyond the existing data set. Models were studied for a number of processes that are proposed to play a role in the pedestal ne and Te profiles. These processes include neoclassical transport, paleoclassical transport, electron temperature gradient turbulence and neutral fuelling. All of these processes may be important, with the importance being dependent on the plasma regime. Studies with several electromagnetic gyrokinetic codes show that the gradients in and on top of the pedestal can drive a number of instabilities.

  18. Interaction of microtubules with the actin cytoskeleton via cross-talk of EB1-containing +TIPs and γ-actin in epithelial cells

    PubMed Central

    Dugina, Vera; Alieva, Irina; Khromova, Natalya; Kireev, Igor; Gunning, Peter W.; Kopnin, Pavel

    2016-01-01

    Actin microfilaments and microtubules are both highly dynamic cytoskeleton components implicated in a wide range of intracellular processes as well as cell-cell and cell-substrate interactions. The interactions of actin filaments with the microtubule system play an important role in the assembly and maintenance of 3D cell structure. Here we demonstrate that cytoplasmic actins are differentially distributed in relation to the microtubule system. LSM, 3D-SIM, proximity ligation assay (PLA) and co-immunoprecipitation methods applied in combination with selective depletion of β- or γ-cytoplasmic actins revealed a selective interaction between microtubules and γ-, but not β-cytoplasmic actin via the microtubule +TIPs protein EB1. EB1-positive comet distribution analysis and quantification have shown more effective microtubule growth in the absence of β-actin. Our data represent the first demonstration that microtubule +TIPs protein EB1 interacts mainly with γ-cytoplasmic actin in epithelial cells. PMID:27683037

  19. Interaction of microtubules with the actin cytoskeleton via cross-talk of EB1-containing +TIPs and γ-actin in epithelial cells.

    PubMed

    Dugina, Vera; Alieva, Irina; Khromova, Natalya; Kireev, Igor; Gunning, Peter W; Kopnin, Pavel

    2016-11-08

    Actin microfilaments and microtubules are both highly dynamic cytoskeleton components implicated in a wide range of intracellular processes as well as cell-cell and cell-substrate interactions. The interactions of actin filaments with the microtubule system play an important role in the assembly and maintenance of 3D cell structure. Here we demonstrate that cytoplasmic actins are differentially distributed in relation to the microtubule system. LSM, 3D-SIM, proximity ligation assay (PLA) and co-immunoprecipitation methods applied in combination with selective depletion of β- or γ-cytoplasmic actins revealed a selective interaction between microtubules and γ-, but not β-cytoplasmic actin via the microtubule +TIPs protein EB1. EB1-positive comet distribution analysis and quantification have shown more effective microtubule growth in the absence of β-actin. Our data represent the first demonstration that microtubule +TIPs protein EB1 interacts mainly with γ-cytoplasmic actin in epithelial cells.

  20. Actin-binding proteins implicated in the formation of the punctate actin foci stimulated by the self-incompatibility response in Papaver.

    PubMed

    Poulter, Natalie S; Staiger, Christopher J; Rappoport, Joshua Z; Franklin-Tong, Vernonica E

    2010-03-01

    The actin cytoskeleton is a key target for signaling networks and plays a central role in translating signals into cellular responses in eukaryotic cells. Self-incompatibility (SI) is an important mechanism responsible for preventing self-fertilization. The SI system of Papaver rhoeas pollen involves a Ca(2+)-dependent signaling network, including massive actin depolymerization as one of the earliest cellular responses, followed by the formation of large actin foci. However, no analysis of these structures, which appear to be aggregates of filamentous (F-)actin based on phalloidin staining, has been carried out to date. Here, we characterize and quantify the formation of F-actin foci in incompatible Papaver pollen tubes over time. The F-actin foci increase in size over time, and we provide evidence that their formation requires actin polymerization. Once formed, these SI-induced structures are unusually stable, being resistant to treatments with latrunculin B. Furthermore, their formation is associated with changes in the intracellular localization of two actin-binding proteins, cyclase-associated protein and actin-depolymerizing factor. Two other regulators of actin dynamics, profilin and fimbrin, do not associate with the F-actin foci. This study provides, to our knowledge, the first insights into the actin-binding proteins and mechanisms involved in the formation of these intriguing structures, which appear to be actively formed during the SI response.

  1. SOL effects on the pedestal structure in DIII-D discharges

    DOE PAGES

    Sontag, Aaron C.; Chen, Xi; Canik, John; ...

    2017-05-24

    SOLPS analysis explains the differences in pedestal structure associated with different ion ∇B drift directions in DIII-D. Core transport models predict that fusion power scales roughly as the square of the pressure at the top of the pedestal, so understanding the effects that determine pedestal structure in steady-state operational scenarios is important to projecting scenarios developed in DIII-D to ITER and other devices. Both experiments and modeling indicate that scrape off layer (SOL) conditions are important in optimizing the pedestal structure for high-beta steady-state scenarios. The SOLPS code is used to provide interpretive analysis of the pedestal and SOL tomore » examine the nature of flows and fueling on the pedestal structure including the effects of drifts in the fluid model. This analysis shows that flows driven by the ion ∇B drift are outward when this drift is toward the x-point in a single-null divertor configuration (favorable ∇B direction for reduced H-mode power threshold), and inward when the drift is away from the x-point (unfavorable ∇B direction). It is hypothesized that these flows decrease the density gradient in the pedestal in the favorable direction, thereby stabilizing the kinetic ballooning mode (KBM) and increasing the pedestal width. Comparisons of pedestal structures in similarly shaped DIII-D steady-state plasmas confirm this change, showing increased density pedestal width and lower peak density and lower separatrix density with the favorable drift direction. The pedestal temperature is higher in the lower density case, resulting in an increased pedestal pressure, which indicates that the increased particle flux does not significantly degrade energy confinement. Modeling of cases with constant ∇B drift direction but changing between the more open lower divertor and more closed upper divertor show that there is increased fueling inside the pedestal with the more open geometry. As a result, the pedestal fueling rate

  2. Edge-localized mode avoidance and pedestal structure in I-mode plasmas

    SciTech Connect

    Walk, J. R. Hughes, J. W.; Hubbard, A. E.; Terry, J. L.; Whyte, D. G.; White, A. E.; Baek, S. G.; Reinke, M. L.; Theiler, C.; Churchill, R. M.; Rice, J. E.; Snyder, P. B.; Osborne, T.; Dominguez, A; Cziegler, I.

    2014-05-15

    I-mode is a high-performance tokamak regime characterized by the formation of a temperature pedestal and enhanced energy confinement, without an accompanying density pedestal or drop in particle and impurity transport. I-mode operation appears to have naturally occurring suppression of large Edge-Localized Modes (ELMs) in addition to its highly favorable scalings of pedestal structure and overall performance. Extensive study of the ELMy H-mode has led to the development of the EPED model, which utilizes calculations of coupled peeling-ballooning MHD modes and kinetic-ballooning mode (KBM) stability limits to predict the pedestal structure preceding an ELM crash. We apply similar tools to the structure and ELM stability of I-mode pedestals. Analysis of I-mode discharges prepared with high-resolution pedestal data from the most recent C-Mod campaign reveals favorable pedestal scalings for extrapolation to large machines—pedestal temperature scales strongly with power per particle P{sub net}/n{sup ¯}{sub e}, and likewise pedestal pressure scales as the net heating power (consistent with weak degradation of confinement with heating power). Matched discharges in current, field, and shaping demonstrate the decoupling of energy and particle transport in I-mode, increasing fueling to span nearly a factor of two in density while maintaining matched temperature pedestals with consistent levels of P{sub net}/n{sup ¯}{sub e}. This is consistent with targets for increased performance in I-mode, elevating pedestal β{sub p} and global performance with matched increases in density and heating power. MHD calculations using the ELITE code indicate that I-mode pedestals are strongly stable to edge peeling-ballooning instabilities. Likewise, numerical modeling of the KBM turbulence onset, as well as scalings of the pedestal width with poloidal beta, indicates that I-mode pedestals are not limited by KBM turbulence—both features identified with the trigger for large ELMs

  3. Contribution to the multi-machine pedestal scaling from the COMPASS tokamak

    NASA Astrophysics Data System (ADS)

    Komm, M.; Bílková, P.; Aftanas, M.; Berta, M.; Böhm, P.; Bogár, O.; Frassinetti, L.; Grover, O.; Háček, P.; Havlicek, J.; Hron, M.; Imríšek, M.; Krbec, J.; Mitošínková, K.; Naydenkova, D.; Pánek, R.; Peterka, M.; Snyder, P. B.; Stefanikova, E.; Stöckel, J.; Sos, M.; Urban, J.; Varju, J.; Vondráček, P.; Weinzettl, V.; the COMPASS Team

    2017-05-01

    First systematic measurements of pedestal structure during Ohmic and NBI-assisted Type I ELMy H-modes were performed on the COMPASS tokamak in two dedicated experimental campaigns during 2015 and 2016. By adjusting the NBI heating and a toroidal magnetic field, the electron pedestal temperature was increased from 200 eV up to 300 eV, which allowed reaching pedestal collisionality ν \\text{ped}\\ast   <  1 at q95 ~3. COMPASS has approached conditions for the Identity experiment done at JET & DIII-D, complementing the range of scanned ρ \\text{ped}\\ast . The pedestal pressure was successfully reproduced by the EPED model. The dependence of pedestal pressure width on ν \\text{ped}\\ast and β \\text{ped ~ }\\text{pol} is discussed.

  4. Collapse of density pedestal by giant ELM on JT-60U

    NASA Astrophysics Data System (ADS)

    Oyama, N.; Shinohara, K.; Kamada, Y.; Miura, Y.; Oikawa, T.; Takeji, S.

    2001-05-01

    In JT-60U ELMy H-mode discharges, the detailed behaviour of giant (type I) ELMs was measured using a heterodyne reflectometer system in order to understand the collapse mechanism of the pedestal structure and consequences on the core/edge plasma. The phase signal of the reflectometer exhibits the movement of the cutoff layer (density layer) due to the collapse of the pedestal in a density profile by an ELM. An ELM event can be classified into a precursor phase, collapse phase, recovery phase and a relaxation phase. A certain density layer measured near the shoulder of the pedestal moves about 7 cm inside the plasma in the collapse phase. The precursor oscillation in a pedestal density and the relationship between the collapse of the pedestal structure and a Dα burst are also studied.

  5. In-out impurity density asymmetry in the pedestal region of Alcator C-Mod

    NASA Astrophysics Data System (ADS)

    Churchill, R. M.; Lipschultz, B.; Theiler, C.; the Alcator C-Mod Team

    2013-12-01

    Measurements are presented of the boron (B5+) density in the pedestal region at the low-field side (LFS) midplane and the high-field side (HFS) midplane of Alcator C-Mod. In H-mode plasmas, a large (≳10x) in-out asymmetry in impurity density forms, with larger densities at the HFS. In contrast, there is no impurity density asymmetry in L-mode or I-mode plasmas. A comparison of pedestal parameters in H-mode plasmas shows that the HFS impurity density pedestal width and position remain fairly fixed over a range of plasma conditions, while the LFS pedestal width widens, and the pedestal position shifts in towards the core as the plasma current is lowered, indicating a change in the underlying transport.

  6. Theoretical explanation for strong poloidal impurity asymmetry in tokamak pedestals

    NASA Astrophysics Data System (ADS)

    Espinosa, Silvia

    2016-10-01

    Stronger impurity density in-out poloidal asymmetries than predicted by the most comprehensive neoclassical models have been measured in H-mode tokamak pedestals during the last decade. However, these pioneering theories neglect the impurity diamagnetic drift, while recent measurements indicate that it can be of the same order as the ExB drift that is retained. In order to keep both drifts self-consistently, stronger radial gradients of the impurity density must be allowed. As a result, radial impurity flow effects need to be included for the first time. These effects substantially alter the parallel impurity flow. The resulting modification in the impurity friction with the banana regime background ions then allows stronger poloidal variation of the impurity density, temperature and potential. Even the six-fold high field side accumulation of boron density measured on Alcator C-Mod can be explained without invoking anomalous transport. Moreover, the potential can no longer be assumed to be a flux function since the impurity density variation gives a poloidally varying potential that results in strong poloidal variation of the radial electric field. The fact that the magnitude of the negative radial electric field and the impurity temperature are both larger on the low field side is also correctly predicted. Finally, this pedestal neoclassical model with radial flows may provide insight on how to control impurity accumulation in JET. Supported by DOE Grant DE-FG0291ER54109 and La Caixa Fellowship.

  7. Removal of pedestals and directional ambiguity of optical anemometer signals.

    PubMed

    Durst, F; Zaré, M

    1974-11-01

    Laser Doppler anemometry permits, in principle, the measurement of both magnitude and direction of components of a particle's velocity vector. Most exiting anemometers, however, permit measurements only with a directional ambiguity of 180 degrees , resulting in errors in certain flow fields. Available methods of eliminating the directional ambiguity of Laser Doppler anemometers are reviewed, covering frequency shifting of the incident and scattered light beams, the use of beams with different polarization properties, and employment of multicolor laser beams. The advantages and disadvantages of existing methods are summarized, and suggestions for alterations are made. Different techniques used to remove the pedestal of laser Doppler anemometer signals are also reviewed. Optical techniques should be employed in any advanced optical anemometer system to avoid dynamic range limitations by electronic bandpass filters. Suggestions are made for advanced optical anemometers employing multielement avalanche photodiodes that can be used for simultaneous measurements of two velocity components. These anemometers incorporate devices to sense the direction of the velocity components and to eliminate optically the pedestal of laser Doppler signals.

  8. The Boundary/Pedestal Integrated Science Application for FSP

    SciTech Connect

    Rognlien, T. D.; Snyder, P. B.

    2011-06-17

    This Integrated Science Application (ISA) represents a combination of the former Integrated Boundary and Pedestal Science Drivers carried out over the last year as part of the Fusion Simulation Program’s planning phase. Those Science Driver plans, as well as four others, can be viewed on the website (http://fspscidri.web.lehigh.edu/index.php/Main_Page). The Boundary Science Driver report is also available as LLNL document LLNL-TR-471260. The plans described in those documents assumed ample resources would be available. This document represents a plan of vital importance for developing powerful simulation tools for magnetic fusion energy devices, but is of substantially less scope than the original Science Drivers because of budget limitations and the fact that here two Science Drivers are merged owing to the close proximity of the two regions that they model: (1) the warm plasma region known as the scrapeoff layer (SOL) where magnetic field lines directly contact material structures together with the associate plasma-wall interactions and (2) the adjacent hotter plasma region know as the pedestal, which is the beginning of the confining closed magnetic field line core.

  9. Differences in the H-mode pedestal width of temperature and density

    NASA Astrophysics Data System (ADS)

    Schneider, P. A.; Wolfrum, E.; Groebner, R. J.; Osborne, T. H.; Beurskens, M. N. A.; Dunne, M. G.; Ferron, J. R.; Günter, S.; Kurzan, B.; Lackner, K.; Snyder, P. B.; Zohm, H.; the ASDEX Upgrade Team; the DIII-D Team; EFDA Contributors, JET

    2012-10-01

    A pedestal database was built using data from type-I ELMy H-modes of ASDEX Upgrade, DIII-D and JET. ELM synchronized pedestal data were analysed with the two-line method. The two-line method is a bilinear fit which shows better reproducibility of pedestal parameters than a modified hyperbolic tangent fit. This was tested with simulated and experimental data. The influence of the equilibrium reconstruction on pedestal parameters was investigated with sophisticated reconstructions from CLISTE and EFIT including edge kinetic profiles. No systematic deviation between the codes could be observed. The flux coordinate system is influenced by machine size, poloidal field and plasma shape. This will change the representation of the width in different coordinates, in particular, the two normalized coordinates ΨN and r/a show a very different dependence on the plasma shape. The scalings derived for the pedestal width, Δ, of all machines suggest a different scaling for the electron temperature and the electron density. Both cases show similar dependence with machine size, poloidal magnetic field and pedestal electron temperature and density. The influence of ion temperature and toroidal magnetic field is different on each of \\Delta_{T_\\rme} and \\Delta_{n_\\rme} . In dimensionless form the density pedestal width in ΨN scales with \\rho^{0.6}_{i\\star} , the temperature pedestal width with \\beta_p,ped^{0.5} . Both widths also show a strong correlation with the plasma shape. The shape dependence originates from the coordinate transformation and is not visible in real space. The presented scalings predict that in ITER the temperature pedestal will be appreciably wider than the density pedestal.

  10. Actin Mechanics and Fragmentation*

    PubMed Central

    De La Cruz, Enrique M.; Gardel, Margaret L.

    2015-01-01

    Cell physiological processes require the regulation and coordination of both mechanical and dynamical properties of the actin cytoskeleton. Here we review recent advances in understanding the mechanical properties and stability of actin filaments and how these properties are manifested at larger (network) length scales. We discuss how forces can influence local biochemical interactions, resulting in the formation of mechanically sensitive dynamic steady states. Understanding the regulation of such force-activated chemistries and dynamic steady states reflects an important challenge for future work that will provide valuable insights as to how the actin cytoskeleton engenders mechanoresponsiveness of living cells. PMID:25957404

  11. Actin Polymerization is Stimulated by Actin Crosslinking Protein Palladin

    PubMed Central

    Gurung, Ritu; Yadav, Rahul; Brungardt, Joseph G.; Orlova, Albina; Egelman, Edward H.; Beck, Moriah R.

    2016-01-01

    The actin scaffold protein palladin regulates both normal cell migration and invasive cell motility, processes that require the coordinated regulation of actin dynamics. However, the potential effect of palladin on actin dynamics has remained elusive. Here we show that the actin binding immunoglobulin-like domain of palladin, which is directly responsible for both actin binding and bundling, also stimulates actin polymerization in vitro. Palladin eliminated the lag phase that is characteristic of the slow nucleation step of actin polymerization. Furthermore, palladin dramatically reduced depolymerization, slightly enhanced the elongation rate, and did not alter the critical concentration. Microscopy and in vitro crosslinking assays reveal differences in actin bundle architecture when palladin is incubated with actin before or after polymerization. These results suggest a model whereby palladin stimulates a polymerization-competent form of G-actin, akin to metal ions, either through charge neutralization or conformational changes. PMID:26607837

  12. Axonal actin in action: Imaging actin dynamics in neurons.

    PubMed

    Ladt, Kelsey; Ganguly, Archan; Roy, Subhojit

    2016-01-01

    Actin is a highly conserved, key cytoskeletal protein involved in numerous structural and functional roles. In neurons, actin has been intensively investigated in axon terminals-growth cones-and dendritic spines, but details about actin structure and dynamics in axon shafts have remained obscure for decades. A major barrier in the field has been imaging actin. Actin exists as soluble monomers (G-actin) as well as actin filaments (F-actin), and labeling actin with conventional fluorescent probes like GFP/RFP typically leads to a diffuse haze that makes it difficult to discern kinetic behaviors. In a recent publication, we used F-actin selective probes to visualize actin dynamics in axons, resolving striking actin behaviors that have not been described before. However, using these probes to visualize actin dynamics is challenging as they can cause bundling of actin filaments; thus, experimental parameters need to be strictly optimized. Here we describe some practical methodological details related to using these probes for visualizing F-actin dynamics in axons.

  13. Characterization of the Pedestal in I-Mode Plasmas on Alcator C-Mod

    NASA Astrophysics Data System (ADS)

    Walk, J. R.; Hughes, J. W.; Terry, J. L.; Hubbard, A. E.; Whyte, D. G.; White, A. E.; Marmar, E. S.; Rice, J. E.; Churchill, R. M.; Theiler, C.; Labombard, B.; Brunner, D.; Reinke, M. L.; Dominguez, A.; Sung, C.

    2012-10-01

    I-mode is an improved energy confinement regime on C-Mod, notably featuring an edge temperature pedestal without the accompanying density pedestal found in H-mode operation. I-mode has been shown to be capable of stationary operation, with the mode sustained for ˜10 energy confinement times, and to have a pedestal naturally lacking large ELMs. Combined with comparable performance to H-mode operation and weaker degradation with input power, this makes I-modes potentially favorable as a reactor regime. A full characterization using the suite of high-resolution edge diagnostics on C-Mod of the pedestal in reversed-field, lower-single-null I-modes (therefore with the ion ∇B drift away from the x-point) is presented here. The presented discharges primarily feature detailed scans of plasma current and ICRF heating power at several density values. The pedestal structure found in these I-modes is also contrasted with previous studies of the pedestal in EDA and ELMy H-modes, particularly potential scalings with βp,ped consistent with the KBM-limited pedestals found in these H-modes.

  14. Pedestal shape, stability and inter-ELM evolution for different main ion species in ASDEX Upgrade

    NASA Astrophysics Data System (ADS)

    Laggner, Florian M.

    2016-10-01

    In tokamak plasmas with different main ion species as hydrogen isotopes or helium, a change of confinement occurs, known as isotope effect. To identify the processes defining the pedestal structure and evolution, experiments comparing hydrogen (H), deuterium (D) and helium (He) plasmas have been performed. Their goal was to match the pedestal top electron density and temperatures and compare the pedestal shape and stability. A factor of almost 10 higher gas puff as well as a factor of 2 higher heating power were required in H to achieve the same pedestal top values as in the D reference. While the pedestal electron temperature profiles do not differ, the density profile in H has shallower gradients. These can be explained by a lower particle confinement in H, if the ionization source profile is assumed to be similar. In He plasmas owing to the larger effective charge, the stored energy at similar pedestal top electron density is roughly a factor of 1.5 smaller than in the references, leading to the absence of ELMs. In summary the experimental results suggest different particle and energy confinement for different main ion species, however, peeling-ballooning theory can sufficiently describe the pedestal stability and ELM behavior. This work has been carried out within the framework of the EUROfusion Consortium and has received funding from the Euratom research and training programme 2014-2018 under Grant Agreement No. 633053.

  15. Baseline neoclassical scaling law on H-mode pedestal width from XGC0 kinetic simulation

    NASA Astrophysics Data System (ADS)

    Park, Gunyoung; Chang, C. S.; Ku, S.

    2009-11-01

    In the H-mode pedestal before the ELM onset, nonlocal neoclassical self-organization is an important physical effect, to set the baseline pedestal width scaling law. Deviation from the neoclassical scaling will define the anomalous scaling. The neoclassical self-organization includes effects from the self-consistent radial electric field shear, strong magnetic field shear, ion-orbit loss across the last closed magnetic surface, finite ion banana width, particle source from neutral ionization, heat flux from the core plasma, and collisional transport. XGC0 code is used to perform an inter-machine study of the neoclassical pedestal scaling law between two representative devices DIII-D (low-B, low collisionality) and C- Mod (high-B, high collisionality). Anomalous scaling component in the experimental pedestal width data will be separated out from the neoclassical component. Prediction for ITER pedestal will be attempted based upon the combined neoclassical (theoretical) and anomalous (empirical) scaling laws obtained in this study. This ion-electron study indicates that the neoclassical pedestal width is broader than the previous ion only study results, closer to experimental pedestal width.

  16. Limits to the H-mode pedestal pressure gradient in DIII-D

    SciTech Connect

    Groebner, R. J.; Snyder, P. B.; Osborne, T. H.; Leonard, A. W.; Rhodes, T. L.; Zeng, L.; Unterberg, Ezekial A; Yan, Z.; Mckee, G. R.; Lasnier, C. J.; Boedo, J.A.; Watkins, J. G.

    2010-01-01

    The spatial and temporal evolution of the total pedestal pressure profile has been measured during the pedestal evolution between successive edge localized modes (ELMs) of type-I ELMing H-mode discharges in DIII-D. Measurements are used to test a model that predicts that kinetic ballooning modes (KBMs) provide a strong constraint on the pedestal pressure gradient obtained during an inter-ELM cycle and cause the pedestal width to scale as the square root of the pedestal poloidal beta. Discharges in two different parameter regimes are examined for evidence that the evolution of the pressure gradient reaches a limit prior to the onset of an ELM. Both discharges show evidence of rapid evolution of the pressure profile very early in the recovery phase from an ELM. In one discharge, the pressure gradient reached approximate steady state within similar to 3 ms after the ELM event. In the other discharge, the pressure gradient just inboard of the last closed flux surface reached steady state early in the ELM recovery phase even as the pedestal expanded into the core and the maximum pressure gradient continued to rise during the remainder of the ELM cycle. Simple quantitative theoretical metrics show that pressure gradients in both discharges reached levels that were large enough to excite KBMs. In addition, the peeling-ballooning theory for the onset of type-I ELMs and the EPED1 model for pedestal height and width make predictions consistent with the data of both discharges.

  17. Structure, Stability and ELM Dynamics of the H-Mode Pedestal in DIII-D

    SciTech Connect

    Fenstermacher, M E; Leonard, A W; Osborne, T H; Snyder, P B; Thomas, D M; Boedo, J A; Casper, T A; Colchin, R J; Groebner, R J; Groth, M; Kempenaars, M H; Loarte, A; Saibene, G; VanZeeland, M A; Zeng, L; Xu, X Q

    2004-10-13

    Experiments are described that have increased understanding of the transport and stability physics that set the H-mode edge pedestal width and height, determine the onset of Type-I edge localized modes (ELMs), and produce the nonlinear dynamics of the ELM perturbation in the pedestal and scrape-off layer (SOL). Predictive models now exist for the n{sub e} pedestal profile and the p{sub e} height at the onset of Type-I ELMs, and progress has been made toward predictive models of the T{sub e} pedestal width and nonlinear ELM evolution. Similarity experiments between DIII-D and JET suggested that neutral penetration physics dominates in the relationship between the width and height of the n{sub e} pedestal while plasma physics dominates in setting the T{sub e} pedestal width. Measured pedestal conditions including edge current at ELM onset agree with intermediate-n peeling-ballooning (P-B) stability predictions. Midplane ELM dynamics data show the predicted (P-B) structure at ELM onset, large rapid variations of the SOL parameters, and fast radial propagation in later phases, similar to features in nonlinear ELM simulations.

  18. Preservation of layered paleodeposits in high-latitude pedestal craters on Mars

    NASA Astrophysics Data System (ADS)

    Kadish, Seth J.; Head, James W.

    2011-06-01

    An outstanding question in Mars' climate history is whether or not pedestal craters represent the armored remnants of ice-rich paleodeposits. We address this question using new high-resolution images; in a survey of several hundred high-latitude pedestal craters, we have identified 12 examples in which visible and/or topographically expressed layers are exposed on the marginal scarp of the pedestal. One example, located on the south polar layered deposits, preserves ice-rich layers that have otherwise been completely removed from the polar cap. These observations provide empirical evidence that the pedestal crater formation mechanism is capable of armoring and preserving ice-rich layered paleodeposits. Although layered exposures have not yet been observed in mid-latitude pedestal craters, high-latitude instances of discontinuous, partially covered layers suggest that layers can be readily concealed, likely through mantling and/or mass wasting processes along the marginal scarp. This interpretation is supported by the observation that high-latitude pedestals with exposed layers along their margins are, on average, taller than mid-latitude examples, and have larger, steeper marginal scarps, which may help to maintain layer exposures. These observations favor the interpretation that mid- to high-latitude pedestal craters represent the armored remnants of ice- and dust-rich paleodeposits, which occurred transiently due to changes in the climate regime. Preservation of fine-scale layering of ice and dust at these latitudes implies that the climate change did not involve regional melting conditions.

  19. Curvature and torsion in growing actin networks

    NASA Astrophysics Data System (ADS)

    Shaevitz, Joshua W.; Fletcher, Daniel A.

    2008-06-01

    Intracellular pathogens such as Listeria monocytogenes and Rickettsia rickettsii move within a host cell by polymerizing a comet-tail of actin fibers that ultimately pushes the cell forward. This dense network of cross-linked actin polymers typically exhibits a striking curvature that causes bacteria to move in gently looping paths. Theoretically, tail curvature has been linked to details of motility by considering force and torque balances from a finite number of polymerizing filaments. Here we track beads coated with a prokaryotic activator of actin polymerization in three dimensions to directly quantify the curvature and torsion of bead motility paths. We find that bead paths are more likely to have low rather than high curvature at any given time. Furthermore, path curvature changes very slowly in time, with an autocorrelation decay time of 200 s. Paths with a small radius of curvature, therefore, remain so for an extended period resulting in loops when confined to two dimensions. When allowed to explore a three-dimensional (3D) space, path loops are less evident. Finally, we quantify the torsion in the bead paths and show that beads do not exhibit a significant left- or right-handed bias to their motion in 3D. These results suggest that paths of actin-propelled objects may be attributed to slow changes in curvature, possibly associated with filament debranching, rather than a fixed torque.

  20. Curvature and torsion in growing actin networks

    PubMed Central

    Shaevitz, Joshua W; Fletcher, Daniel A

    2011-01-01

    Intracellular pathogens such as Listeria monocytogenes and Rickettsia rickettsii move within a host cell by polymerizing a comet-tail of actin fibers that ultimately pushes the cell forward. This dense network of cross-linked actin polymers typically exhibits a striking curvature that causes bacteria to move in gently looping paths. Theoretically, tail curvature has been linked to details of motility by considering force and torque balances from a finite number of polymerizing filaments. Here we track beads coated with a prokaryotic activator of actin polymerization in three dimensions to directly quantify the curvature and torsion of bead motility paths. We find that bead paths are more likely to have low rather than high curvature at any given time. Furthermore, path curvature changes very slowly in time, with an autocorrelation decay time of 200 s. Paths with a small radius of curvature, therefore, remain so for an extended period resulting in loops when confined to two dimensions. When allowed to explore a three-dimensional (3D) space, path loops are less evident. Finally, we quantify the torsion in the bead paths and show that beads do not exhibit a significant left- or right-handed bias to their motion in 3D. These results suggest that paths of actin-propelled objects may be attributed to slow changes in curvature, possibly associated with filament debranching, rather than a fixed torque. PMID:18560043

  1. Bacillus anthracis Edema Toxin Impairs Neutrophil Actin-Based Motility▿

    PubMed Central

    Szarowicz, Sarah E.; During, Russell L.; Li, Wei; Quinn, Conrad P.; Tang, Wei-Jen; Southwick, Frederick S.

    2009-01-01

    Inhalation anthrax results in high-grade bacteremia and is accompanied by a delay in the rise of the peripheral polymorphonuclear neutrophil (PMN) count and a paucity of PMNs in the infected pleural fluid and mediastinum. Edema toxin (ET) is one of the major Bacillus anthracis virulence factors and consists of the adenylate cyclase edema factor (EF) and protective antigen (PA). Relatively low concentrations of ET (100 to 500 ng/ml of PA and EF) significantly impair human PMN chemokinesis, chemotaxis, and ability to polarize. These changes are accompanied by a reduction in chemoattractant-stimulated PMN actin assembly. ET also causes a significant decrease in Listeria monocytogenes intracellular actin-based motility within HeLa cells. These defects in actin assembly are accompanied by a >50-fold increase in intracellular cyclic AMP and a >4-fold increase in the phosphorylation of protein kinase A. We have previously shown that anthrax lethal toxin (LT) also impairs neutrophil actin-based motility (R. L. During, W. Li, B. Hao, J. M. Koenig, D. S. Stephens, C. P. Quinn, and F. S. Southwick, J. Infect. Dis. 192:837-845, 2005), and we now find that LT combined with ET causes an additive inhibition of PMN chemokinesis, polarization, chemotaxis, and FMLP (N-formyl-met-leu-phe)-induced actin assembly. We conclude that ET alone or combined with LT impairs PMN actin assembly, resulting in paralysis of PMN chemotaxis. PMID:19349425

  2. Lamellipodin promotes actin assembly by clustering Ena/VASP proteins and tethering them to actin filaments

    PubMed Central

    Hansen, Scott D; Mullins, R Dyche

    2015-01-01

    Enabled/Vasodilator (Ena/VASP) proteins promote actin filament assembly at multiple locations, including: leading edge membranes, focal adhesions, and the surface of intracellular pathogens. One important Ena/VASP regulator is the mig-10/Lamellipodin/RIAM family of adaptors that promote lamellipod formation in fibroblasts and drive neurite outgrowth and axon guidance in neurons. To better understand how MRL proteins promote actin network formation we studied the interactions between Lamellipodin (Lpd), actin, and VASP, both in vivo and in vitro. We find that Lpd binds directly to actin filaments and that this interaction regulates its subcellular localization and enhances its effect on VASP polymerase activity. We propose that Lpd delivers Ena/VASP proteins to growing barbed ends and increases their polymerase activity by tethering them to filaments. This interaction represents one more pathway by which growing actin filaments produce positive feedback to control localization and activity of proteins that regulate their assembly. DOI: http://dx.doi.org/10.7554/eLife.06585.001 PMID:26295568

  3. Histones bundle F-actin filaments and affect actin structure.

    PubMed

    Blotnick, Edna; Sol, Asaf; Muhlrad, Andras

    2017-01-01

    Histones are small polycationic proteins complexed with DNA located in the cell nucleus. Upon apoptosis they are secreted from the cells and react with extracellular polyanionic compounds. Actin which is a polyanionic protein, is also secreted from necrotic cells and interacts with histones. We showed that both histone mixture (histone type III) and the recombinant H2A histone bundles F-actin, increases the viscosity of the F-actin containing solution and polymerizes G-actin. The histone-actin bundles are relatively insensitive to increase of ionic strength, unlike other polycation, histatin, lysozyme, spermine and LL-37 induced F-actin bundles. The histone-actin bundles dissociate completely only in the presence of 300-400 mM NaCl. DNA, which competes with F-actin for histones, disassembles histone induced actin bundles. DNase1, which depolymerizes F- to G-actin, actively unbundles the H2A histone induced but slightly affects the histone mixture induced actin bundles. Cofilin decreases the amount of F-actin sedimented by low speed centrifugation, increases light scattering and viscosity of F-actin-histone mixture containing solutions and forms star like superstructures by copolymerizing G-actin with H2A histone. The results indicate that histones are tightly attached to F-actin by strong electrostatic and hydrophobic forces. Since both histones and F-actin are present in the sputum of patients with cystic fibrosis, therefore, the formation of the stable histone-actin bundles can contribute to the pathology of this disease by increasing the viscosity of the sputum. The actin-histone interaction in the nucleus might affect gene expression.

  4. Polycation induced actin bundles.

    PubMed

    Muhlrad, Andras; Grintsevich, Elena E; Reisler, Emil

    2011-04-01

    Three polycations, polylysine, the polyamine spermine and the polycationic protein lysozyme were used to study the formation, structure, ionic strength sensitivity and dissociation of polycation-induced actin bundles. Bundles form fast, simultaneously with the polymerization of MgATP-G-actins, upon the addition of polycations to solutions of actins at low ionic strength conditions. This indicates that nuclei and/or nascent filaments bundle due to attractive, electrostatic effect of polycations and the neutralization of repulsive interactions of negative charges on actin. The attractive forces between the filaments are strong, as shown by the low (in nanomolar range) critical concentration of their bundling at low ionic strength. These bundles are sensitive to ionic strength and disassemble partially in 100 mM NaCl, but both the dissociation and ionic strength sensitivity can be countered by higher polycation concentrations. Cys374 residues of actin monomers residing on neighboring filaments in the bundles can be cross-linked by the short span (5.4Å) MTS-1 (1,1-methanedyl bismethanethiosulfonate) cross-linker, which indicates a tight packing of filaments in the bundles. The interfilament cross-links, which connect monomers located on oppositely oriented filaments, prevent disassembly of bundles at high ionic strength. Cofilin and the polysaccharide polyanion heparin disassemble lysozyme induced actin bundles more effectively than the polylysine-induced bundles. The actin-lysozyme bundles are pathologically significant as both proteins are found in the pulmonary airways of cystic fibrosis patients. Their bundles contribute to the formation of viscous mucus, which is the main cause of breathing difficulties and eventual death in this disorder.

  5. Actin-Dependent Propulsion of Endosomes and Lysosomes by Recruitment of N-Wasp✪

    PubMed Central

    Taunton, Jack; Rowning, Brian A.; Coughlin, Margaret L.; Wu, Michael; Moon, Randall T.; Mitchison, Timothy J.; Larabell, Carolyn A.

    2000-01-01

    We examined the spatial and temporal control of actin assembly in living Xenopus eggs. Within minutes of egg activation, dynamic actin-rich comet tails appeared on a subset of cytoplasmic vesicles that were enriched in protein kinase C (PKC), causing the vesicles to move through the cytoplasm. Actin comet tail formation in vivo was stimulated by the PKC activator phorbol myristate acetate (PMA), and this process could be reconstituted in a cell-free system. We used this system to define the characteristics that distinguish vesicles associated with actin comet tails from other vesicles in the extract. We found that the protein, N-WASP, was recruited to the surface of every vesicle associated with an actin comet tail, suggesting that vesicle movement results from actin assembly nucleated by the Arp2/3 complex, the immediate downstream target of N-WASP. The motile vesicles accumulated the dye acridine orange, a marker for endosomes and lysosomes. Furthermore, vesicles associated with actin comet tails had the morphological features of multivesicular endosomes as revealed by electron microscopy. Endosomes and lysosomes from mammalian cells preferentially nucleated actin assembly and moved in the Xenopus egg extract system. These results define endosomes and lysosomes as recruitment sites for the actin nucleation machinery and demonstrate that actin assembly contributes to organelle movement. Conversely, by nucleating actin assembly, intracellular membranes may contribute to the dynamic organization of the actin cytoskeleton. PMID:10662777

  6. Tropomodulins: pointed-end capping proteins that regulate actin filament architecture in diverse cell types

    PubMed Central

    Yamashiro, Sawako; Gokhin, David S.; Kimura, Sumiko; Nowak, Roberta B.; Fowler, Velia M.

    2012-01-01

    Tropomodulins are a family of four proteins (Tmods 1–4) that cap the pointed ends of actin filaments in actin cytoskeletal structures in a developmentally regulated and tissue-specific manner. Unique among capping proteins, Tmods also bind tropomyosins (TMs), which greatly enhance the actin filament pointed-end capping activity of Tmods. Tmods are defined by a tropomyosin (TM)-regulated/Pointed-End Actin Capping (TM-Cap) domain in their unstructured N-terminal portion, followed by a compact, folded Leucine-Rich Repeat/Pointed-End Actin Capping (LRR-Cap) domain. By inhibiting actin monomer association and dissociation from pointed ends, Tmods regulate regulate actin dynamics and turnover, stabilizing actin filament lengths and cytoskeletal architecture. In this review, we summarize the genes, structural features, molecular and biochemical properties, actin regulatory mechanisms, expression patterns, and cell and tissue functions of Tmods. By understanding Tmods’ functions in the context of their molecular structure, actin regulation, binding partners, and related variants (leiomodins 1–3), we can draw broad conclusions that can explain the diverse morphological and functional phenotypes that arise from Tmod perturbation experiments in vitro and in vivo. Tmod-based stabilization and organization of intracellular actin filament networks provide key insights into how the emergent properties of the actin cytoskeleton drive tissue morphogenesis and physiology. PMID:22488942

  7. Cofilin is a Component of Intranuclear and Cytoplasmic Actin Rods Induced in Cultured Cells

    NASA Astrophysics Data System (ADS)

    Nishida, Eisuke; Iida, Kazuko; Yonezawa, Naoto; Koyasu, Shigeo; Yahara, Ichiro; Sakai, Hikoichi

    1987-08-01

    Incubation of cultured cells under specific conditions induces a dramatic change in the actin organization: induction of intranuclear and/or cytoplasmic actin rods (actin paracrystal-like intracellular structures). We have found that cofilin, a 21-kDa actin-binding protein, is a component of these rods. Antibodies directed against cofilin labeled intranuclear actin rods induced in cells treated with dimethyl sulfoxide or exposed to heat shock and also labeled cytoplasmic actin rods induced in cells incubated in specific salt buffers. Moreover, we found that these actin rods are not stained with fluorescent phalloidin derivatives at all and appear to be right-handed helices, different from straight bundles of F-actin such as stress fibers. In vitro experiments revealed that cofilin and phalloidin compete with each other for binding to F-actin. Since cofilin and phalloidin have the ability to stoichiometrically bind actin molecule in the filament in vitro, the above results seem to suggest that cofilin directly binds to actin molecule in nearly an equimolar ratio in these rods. We call these rods ``actin/cofilin rods.''

  8. Actin-dependent propulsion of endosomes and lysosomes byrecruitment of n-wasp

    SciTech Connect

    Taunton J; Rowning BA; Coughlin ML; Wu M; Moon RT; Mitchison TJ; Larabell CA

    2000-02-07

    We examined the spatial and temporal control of actin assembly in living Xenopus eggs. Within minutes of egg activation,dynamic actin-rich comet tails appeared on a subset of cytoplasmic vesicles that were enriched in protein kinase C (PKC), causing the vesicles to move through the cytoplasm. Actin comet tail formation in vivo was stimulated by the PKC activator phorbol myristate acetate (PMA),and this process could be reconstituted in a cell-free system. We used this system to define the characteristics that distinguish vesicles associated with actin comet tails from other vesicles in the extract. We found that the protein, N-WASP, was recruited to the surface of every vesicle associated with an actin comet tail, suggesting that vesicle movement results from actin assembly nucleated by the Arp2/3 complex, the immediate downstream target of N-WASP, The motile vesicles accumulated the dye acridine orange, a marker for endosomes and lysosomes. Furthermore, vesicles associated with actin comet tails had the morphological features of multivesicular endosomes as revealed by electron microscopy. Endosomes and lysosomes from mammalian cells preferentially nucleated actin assembly and moved in the Xenopus egg extract system. These results define endosomes and lysosomes as recruitment sites for the actin nucleation machinery and demonstrate that actin assembly contributes to organelle movement. Conversely, by nucleating actin assembly, intracellular membranes may contribute to the dynamic organization of the actin cytoskeleton.

  9. ECE-imaging of the H-mode pedestal (invited).

    PubMed

    Tobias, B J; Austin, M E; Boom, J E; Burrell, K H; Classen, I G J; Domier, C W; Luhmann, N C; Nazikian, R; Snyder, P B

    2012-10-01

    A synthetic diagnostic has been developed that reproduces the highly structured electron cyclotron emission (ECE) spectrum radiated from the edge region of H-mode discharges. The modeled dependence on local perturbations of the equilibrium plasma pressure allows for interpretation of ECE data for diagnosis of local quantities. Forward modeling of the diagnostic response in this region allows for improved mapping of the observed fluctuations to flux surfaces within the plasma, allowing for the poloidal mode number of coherent structures to be resolved. In addition, other spectral features that are dependent on both T(e) and n(e) contain information about pedestal structure and the electron energy distribution of localized phenomena, such as edge filaments arising during edge-localized mode (ELM) activity.

  10. ECE-imaging of the H-mode pedestal (invited)

    SciTech Connect

    Tobias, B. J.; Nazikian, R.; Austin, M. E.; Boom, J. E.; Classen, I. G. J.; Burrell, K. H.; Snyder, P. B.; Domier, C. W.; Luhmann, N. C. Jr.

    2012-10-15

    A synthetic diagnostic has been developed that reproduces the highly structured electron cyclotron emission (ECE) spectrum radiated from the edge region of H-mode discharges. The modeled dependence on local perturbations of the equilibrium plasma pressure allows for interpretation of ECE data for diagnosis of local quantities. Forward modeling of the diagnostic response in this region allows for improved mapping of the observed fluctuations to flux surfaces within the plasma, allowing for the poloidal mode number of coherent structures to be resolved. In addition, other spectral features that are dependent on both T{sub e} and n{sub e} contain information about pedestal structure and the electron energy distribution of localized phenomena, such as edge filaments arising during edge-localized mode (ELM) activity.

  11. Impact of zonal flows on edge pedestal collapse

    NASA Astrophysics Data System (ADS)

    Jhang, Hogun; Kaang, Helen H.; Kim, S. S.; Rhee, T.; Singh, R.; Hahm, T. S.

    2017-02-01

    We perform a computational study of the role of zonal flows in edge pedestal collapse on the basis of a nonlinear three-field reduced magnetohydrodynamic (MHD) model. A dramatic change of dynamics takes place when ideal ballooning modes are completely stabilized. Analyses show that a new instability is developed due to a strong excitation of zonal vorticity, resulting in a series of secondary crashes. The presence of subsidiary bursts after a main crash increases the effective crash time and energy loss. These simulation results resemble the behavior of compound edge localized modes (ELMs). Analyses in this paper indicate that a complete understanding of ELM crash dynamics requires the self-consistent inclusion of nonlinear zonal flows-MHD interaction and transport physics.

  12. Isotope and density profile effects on pedestal neoclassical transport

    NASA Astrophysics Data System (ADS)

    Buller, S.; Pusztai, I.

    2017-10-01

    Cross-field neoclassical transport of heat, particles and momentum is studied in sharp density pedestals, with a focus on isotope and profile effects, using a radially global approach. Global effects—which tend to reduce the peak ion heat flux, and shift it outward—increase with isotope mass for fixed profiles. The heat flux reduction exhibits a saturation with a favorable isotopic trend. A significant part of the heat flux can be convective even in pure plasmas, unlike in the plasma core, and it is sensitive to how momentum sources are distributed between the various species. In particular, if only ion momentum sources are allowed, in global simulations of pure plasmas the ion particle flux remains close to its local value, while this may not be the case for simulations with isotope mixtures or electron momentum sources. The radial angular momentum transport that is a finite orbit width effect, is found to be strongly correlated with heat sources.

  13. DYNA3D Finite Element Analysis of Steam Explosion Loads on a Pedestal Wall Design

    SciTech Connect

    Noble, C R

    2007-01-18

    The objective of this brief report is to document the ESBWR pedestal wall finite element analyses that were performed as a quick turnaround effort in July 2005 at Lawrence Livermore National Laboratory and describe the assumptions and failure criteria used for these analyses [Ref 4]. The analyses described within are for the pedestal wall design that included an internal steel liner. The goal of the finite element analyses was to assist in determining the load carrying capacity of the ESBWR pedestal wall subjected to an impulsive pressure generated by a steam explosion.

  14. Validation of the cuff pedestal technique for rapid eye movement sleep (REMs) deprivation by electrophysiological recordings.

    PubMed

    Hilakivi, I; Peder, M; Elomaa, E; Johansson, G

    1984-06-01

    Twenty-four-hour recordings of electrophysiological correlates of the sleep-waking cycle in the rat were performed during different stages of cuff pedestal treatment. It was found that rats adapted to live on pedestals with the cuff raised displayed undisturbed patterns of sleep and wakefulness. Lowering the cuff for three days resulted in virtually total disappearance of rapid eye movement sleep (REMs), while slow wave sleep (SWs) was only slightly reduced. Raising the cuff induced a prominent rebound increase of REMs. These results accord with data obtained by means of the conventional flowerpot procedure and corroborate the validity of the cuff pedestal technique.

  15. Possible association of actin filaments with chloroplasts of spinach mesophyll cells in vivo and in vitro.

    PubMed

    Kumatani, T; Sakurai-Ozato, N; Miyawaki, N; Yokota, E; Shimmen, T; Terashima, I; Takagi, S

    2006-11-01

    In palisade mesophyll cells of spinach (Spinacia oleracea L.) kept under low-intensity white light, chloroplasts were apparently immobile and seemed to be surrounded by fine bundles of actin filaments. High-intensity blue light induced actin-dependent chloroplast movement concomitant with the appearance of a couple of long, straight bundles of actin filaments in each cell, whereas high-intensity red light was essentially ineffective in inducing these responses. The actin organization observed under low-intensity white light has been postulated to function in anchoring chloroplasts at proper intracellular positions through direct interaction with the chloroplasts. Intact chloroplasts, which retained their outer envelopes, were isolated after homogenization of leaves and Percoll centrifugation. No endogenous actin was detected by immunoblotting in the final intact-chloroplast fraction prepared from the leaves kept under low-intensity white light or in darkness. In cosedimentation assays with exogenously added skeletal muscle filamentous actin, however, actin was detected in the intact-chloroplast fraction precipitated after low-speed centrifugation. The association of actin with chloroplasts was apparently dependent on incubation time and chloroplast density. After partial disruption of the outer envelope of isolated chloroplasts by treatment with trypsin, actin was no longer coprecipitated. The results suggest that chloroplasts in spinach leaves can directly interact with actin, and that this interaction may be involved in the regulation of intracellular positioning of chloroplasts.

  16. Villin Severing Activity Enhances Actin-based Motility In Vivo

    PubMed Central

    Revenu, Céline; Courtois, Matthieu; Michelot, Alphée; Sykes, Cécile; Louvard, Daniel

    2007-01-01

    Villin, an actin-binding protein associated with the actin bundles that support microvilli, bundles, caps, nucleates, and severs actin in a calcium-dependant manner in vitro. We hypothesized that the severing activity of villin is responsible for its reported role in enhancing cell plasticity and motility. To test this hypothesis, we chose a loss of function strategy and introduced mutations in villin based on sequence comparison with CapG. By pyrene-actin assays, we demonstrate that this mutant has a strongly reduced severing activity, whereas nucleation and capping remain unaffected. The bundling activity and the morphogenic effects of villin in cells are also preserved in this mutant. We thus succeeded in dissociating the severing from the three other activities of villin. The contribution of villin severing to actin dynamics is analyzed in vivo through the actin-based movement of the intracellular bacteria Shigella flexneri in cells expressing villin and its severing variant. The severing mutations abolish the gain of velocity induced by villin. To further analyze this effect, we reconstituted an in vitro actin-based bead movement in which the usual capping protein is replaced by either the wild type or the severing mutant of villin. Confirming the in vivo results, villin-severing activity enhances the velocity of beads by more than two-fold and reduces the density of actin in the comets. We propose a model in which, by severing actin filaments and capping their barbed ends, villin increases the concentration of actin monomers available for polymerization, a mechanism that might be paralleled in vivo when an enterocyte undergoes an epithelio-mesenchymal transition. PMID:17182858

  17. Villin severing activity enhances actin-based motility in vivo.

    PubMed

    Revenu, Céline; Courtois, Matthieu; Michelot, Alphée; Sykes, Cécile; Louvard, Daniel; Robine, Sylvie

    2007-03-01

    Villin, an actin-binding protein associated with the actin bundles that support microvilli, bundles, caps, nucleates, and severs actin in a calcium-dependant manner in vitro. We hypothesized that the severing activity of villin is responsible for its reported role in enhancing cell plasticity and motility. To test this hypothesis, we chose a loss of function strategy and introduced mutations in villin based on sequence comparison with CapG. By pyrene-actin assays, we demonstrate that this mutant has a strongly reduced severing activity, whereas nucleation and capping remain unaffected. The bundling activity and the morphogenic effects of villin in cells are also preserved in this mutant. We thus succeeded in dissociating the severing from the three other activities of villin. The contribution of villin severing to actin dynamics is analyzed in vivo through the actin-based movement of the intracellular bacteria Shigella flexneri in cells expressing villin and its severing variant. The severing mutations abolish the gain of velocity induced by villin. To further analyze this effect, we reconstituted an in vitro actin-based bead movement in which the usual capping protein is replaced by either the wild type or the severing mutant of villin. Confirming the in vivo results, villin-severing activity enhances the velocity of beads by more than two-fold and reduces the density of actin in the comets. We propose a model in which, by severing actin filaments and capping their barbed ends, villin increases the concentration of actin monomers available for polymerization, a mechanism that might be paralleled in vivo when an enterocyte undergoes an epithelio-mesenchymal transition.

  18. Growing actin networks regulated by obstacle size and shape

    NASA Astrophysics Data System (ADS)

    Gong, Bo; Lin, Ji; Qian, Jin

    2017-01-01

    Growing actin networks provide the driving force for the motility of cells and intracellular pathogens. Based on the molecular-level processes of actin polymerization, branching, capping, and depolymerization, we have developed a modeling framework to simulate the stochastic and cooperative behaviors of growing actin networks in propelling obstacles, with an emphasis on the size and shape effects on work capacity and filament orientation in the growing process. Our results show that the characteristic size of obstacles changes the protrusion power per unit length, without influencing the orientation distribution of actin filaments in growing networks. In contrast, the geometry of obstacles has a profound effect on filament patterning, which influences the orientation of filaments differently when the drag coefficient of environment is small, intermediate, or large. We also discuss the role of various parameters, such as the aspect ratio of obstacles, branching rate, and capping rate, in affecting the protrusion power of network growth.

  19. Primary immunodeficiencies due to abnormalities of the actin cytoskeleton.

    PubMed

    Burns, Siobhan O; Zarafov, Anton; Thrasher, Adrian J

    2017-01-01

    Primary immunodeficiencies (PIDs) are inherited conditions where components of the immune system are missing or dysfunctional. Over 300 genes have been causally linked to monogenic forms of PID, including a number that regulate the actin cytoskeleton. The majority of cytoskeletal defects disrupt assembly and disassembly of filamentous actin in multiple immune cell lineages impacting functions such as cell migration and adhesion, pathogen uptake, intercellular communication, intracellular signalling, and cell division. In the past 24 months, new actin defects have been identified through next generation sequencing technologies. Substantial progress has also been made in understanding the pathogenic mechanisms that contribute to immunological dysfunction, and also how the cytoskeleton participates in normal physiological immune processes. This review summarises recent advances in the field, raising awareness of these conditions and our current understanding of their presentation. Description of further cases and new conditions will extend the clinical phenotype of actin-related disorders, and will promote the development of more effective and targeted therapies.

  20. Intracellular ROS

    PubMed Central

    Leshem, Yehoram

    2007-01-01

    Intracellular localization of stress induced reactive oxygen species (ROS) has emerged as an important aspect towards understanding of cellular responses to environmental stimuli. Our recent study published in the PNAS (103:18008–13)1 shows that NaCl-induced ROS appear within endosomes on the way to tonoplast as part of the vacuolar vesicle trafficking. In addition to showing ROS damage to the tonoplast, this finding may shed light upon recently reported aspects of root water relations during salt stress, suggesting a new signaling role for intracellular ROS in Arabidopsis root cells, during salt stress: ROS that are compartmentalized in endosomes are delivered by the vacuolar vesicle trafficking pathway to the tonoplast, resulting in oxidative gating of TIPs water channels. The closure of the tonoplast aquaporins contributes to the observed reduction in root hydraulic conductivity during salt stress. PMID:19704741

  1. Nuclear actin and protein 4.1: Essential interactions during nuclear assembly in vitro

    SciTech Connect

    Krauss, Sharon Wald; Chen, Cynthia; Penman, Sheldon; Heald, Rebecca

    2003-06-11

    Structural protein 4.1, which has crucial interactions within the spectin-actin lattice of the human red cell membrane skeleton, also is widely distributed at diverse intracellular sites in nucleated cells. We previously showed that 4.1 is essential for assembly of functional nuclei in vitro and that the capacity of 4.1 to bind actin is required. Here we report that 4.1 and actin colocalize in mammalian cell nuclei using fluorescence microscopy and, by higher resolution cell whole mount electron microscopy, are associated on nuclear filaments. We also devised a cell-free assay using Xenopus egg extract containing fluorescent actin to follow actin during nuclear assembly. By directly imaging actin under non-perturbing conditions, the total nuclear actin population is retained and is visualized in situ relative to intact chromatin. We detected actin initially when chromatin and nuclear pores began assembling. As the nuclear lamina assembled, but preceding DNA synthesis, a discrete actin network formed throughout the nucleus. Protein 4.1 epitopes also were detected when actin began to accumulate in nuclei, producing a diffuse coincident pattern. As nuclei matured, actin was detected both coincident with and also independent of 4.1 epitopes. To test whether acquisition of nuclear actin is required for nuclear assembly, the actin inhibitor latrunculin A was added to Xenopus egg extracts during nuclear assembly. Latrunculin A strongly perturbed nuclear assembly and produced distorted nuclear structures containing neither actin nor protein 4.1. Our results suggest that actin as well as 4.1 is necessary for nuclear assembly and that 4.1-actin interactions may be critical.

  2. A Legionella Effector Disrupts Host Cytoskeletal Structure by Cleaving Actin

    DOE PAGES

    Liu, Yao; Zhu, Wenhan; Tan, Yunhao; ...

    2017-01-27

    Legionella pneumophila, the etiological agent of Legionnaires' disease, replicates intracellularly in protozoan and human hosts. Successful colonization and replication of this pathogen in host cells requires the Dot/Icm type IVB secretion system, which translocates approximately 300 effector proteins into the host cell to modulate various cellular processes. In this study, we identified RavK as a Dot/Icm substrate that targets the host cytoskeleton and reduces actin filament abundance in mammalian cells upon ectopic expression. RavK harbors an H95EXXH99 motif associated with diverse metalloproteases, which is essential for the inhibition of yeast growth and for the induction of cell rounding in HEK293Tmore » cells. We demonstrate that the actin protein itself is the cellular target of RavK and that this effector cleaves actin at a site between residues Thr351 and Phe352. Importantly, RavK-mediated actin cleavage also occurs during L. pneumophila infection. Cleavage by RavK abolishes the ability of actin to form polymers. Furthermore, an F352A mutation renders actin resistant to RavK-mediated cleavage; expression of the mutant in mammalian cells suppresses the cell rounding phenotype caused by RavK, further establishing that actin is the physiological substrate of RavK. Furthermore, L. pneumophila exploits components of the host cytoskeleton by multiple effectors with distinct mechanisms, highlighting the importance of modulating cellular processes governed by the actin cytoskeleton in the intracellular life cycle of this pathogen.« less

  3. Theory and Modeling of ELMs and Constraints on the H-Mode Pedestal

    NASA Astrophysics Data System (ADS)

    Snyder, P. B.; Ferron, J. R.; Lao, L. L.; Leonard, A. W.; Osborne, T. H.; Turnbull, A. D.; Wilson, H. R.; Webster, A. J.; Xu, X. Q.; Mossessian, D.; Murakami, M.

    2002-11-01

    We present a theory of edge localized modes (ELMs) and constraints on the H-mode pedestal, based on the stability of intermediate wavelength peeling-ballooning modes driven by the strong pressure gradient and resulting bootstrap current in the pedestal region. Detailed studies of ideal MHD pedestal stability bounds are presented using the ELITE code, and are compared to data from multiple tokamaks. Observed ELM onset times and characteristics, as well as variation in pedestal behavior with discharge shape and collisionality, are studied. In addition, the impact of diamagnetic stabilization and rotation shear are assessed, and progress on dynamic modeling of the ELM cycle which couples transport codes to stability calculations and ELM crash models is briefly discussed. Nonlinear simulations using the BOUT code are used to provide further insight on non-ideal effects and ELM crash dynamics.

  4. Characteristics of edge pedestals in LHW and NBI heated H-mode plasmas on EAST

    NASA Astrophysics Data System (ADS)

    Zang, Q.; Wang, T.; Liang, Y.; Sun, Y.; Chen, H.; Xiao, S.; Han, X.; Hu, A.; Hsieh, C.; Zhou, H.; Zhao, J.; Zhang, T.; Gong, X.; Hu, L.; Liu, F.; Hu, C.; Gao, X.; Wan, B.; the EAST Team

    2016-10-01

    By using the recently developed Thomson scattering diagnostic, the pedestal structure of the H-mode with neutral beam injection (NBI) or/and lower hybrid wave (LHW) heating on EAST (Experimental Advanced Superconducting Tokamak) is analyzed in detail. We find that a higher ratio of the power of the NBI to the total power of the NBI and the lower hybrid wave (LHW) will produce a large and regular different edge-localized mode (ELM), and a lower ratio will produce a small and irregular ELM. The experiments show that the mean pedestal width has good correlation with β \\text{p,\\text{ped}}0.5 , The pedestal width appears to be wider than that on other similar machines, which could be due to lithium coating. However, it is difficult to draw any conclusion of correlation between ρ * and the pedestal width for limited ρ * variation and scattered distribution. It is also found that T e/\

  5. Impact of fuelling and impurity on pedestal dynamics and instabilities in the HL-2A tokamak

    NASA Astrophysics Data System (ADS)

    Zhong, W. L.; Zou, X. L.; Gao, J. M.; Shi, Z. B.; Feng, B. B.; Cui, Z. Y.; Xu, M.; Shen, Y.; Dong, J. Q.; Ding, X. T.; Duan, X. R.; Liu, Yong; HL-2A Team

    2017-01-01

    In recent experiments of the HL-2A tokamak, the effect on the pedestal dynamics by the plasma fuelling with supersonic molecular beam injection (SMBI) has been intensively investigated. Experimental results in several tokamaks suggested that SMBI is a promising technique for ELM mitigation. In addition to the fuelling, the impact of impurities on the pedestal dynamics and instabilities has been investigated in HL-2A. Experimental results have shown that during the H-mode phase, a broadband electromagnetic (EM) turbulence was driven by peaked impurity density profile at the edge plasma region, and governed by double critical gradients of the impurity density. The absolute value of the threshold in positive gradient region is much lower than that in the negative region. This strong asymmetry in the critical gradients has been predicted by theoretical simulation. The results reveal that pedestal dynamics and heat loads can be actively controlled by exciting or changing pedestal instabilities.

  6. Neoclassical ion heat flux and poloidal flow in a tokamak pedestal

    NASA Astrophysics Data System (ADS)

    Kagan, Grigory; Catto, Peter J.

    2010-05-01

    In the core of a tokamak, turbulent transport normally dominates over neoclassical. The situation could be different in a high confinement (or H) mode pedestal, where the former may be suppressed by a strongly sheared equilibrium electric field. On the other hand, this very field makes conventional neoclassical results inapplicable in the pedestal by significantly modifying ion drift orbits. We present the first calculation of the banana regime neoclassical ion heat flux and poloidal flow in the pedestal accounting for the strong E × B drift inherent to this tokamak region. Interestingly, we find that due to the electric field the pedestal poloidal ion flow can change its direction as compared with its core counterpart. This result elucidates the discrepancy between the conventional banana regime predictions and recent experimental measurements of the impurity flow performed at Alcator C-Mod.

  7. Finite element analysis of bridge steel pedestal anchor bolts in reinforced concrete

    NASA Astrophysics Data System (ADS)

    Logan Hancock, B.; Hite Head, Monique

    2009-10-01

    Steel pedestals are short, column-like structures currently being used to elevate highway bridges to reduce the risk of collisions with over-height vehicles. Previous full-scale experimental research has been done to examine the efficacy of these steel pedestals and their components under quasi-static loading to evaluate any added instability in the event of an earthquake. As part of the Undergraduate Summer Research Grant (USRG) program at Texas A&M University, this specific project was focused on observing the behavior of the post-installed steel pedestal anchor bolts under applied shear and tensile loading using the finite element (FE) software Abaqus. The results from some of the preliminary analyses are compared to theoretical anchorage calculations with the aim of producing a benchmark for future steel pedestal anchor bolt embedment design. Future research improvements regarding FE modeling and structural design suggestions are proposed as well.

  8. RefilinB (FAM101B) targets FilaminA to organize perinuclear actin networks and regulates nuclear shape

    PubMed Central

    Gay, Olivia; Gilquin, Benoît; Nakamura, Fumihiko; Jenkins, Zandra A.; McCartney, Rosannah; Krakow, Deborah; Deshiere, Alexandre; Assard, Nicole; Hartwig, John H.; Robertson, Stephen P.; Baudier, Jacques

    2011-01-01

    The intracellular localization and shape of the nucleus plays a central role in cellular and developmental processes. In fibroblasts, nuclear movement and shape are controlled by a specific perinuclear actin network made of contractile actin filament bundles called transmembrane actin-associated nuclear (TAN) lines that form a structure called the actin cap. The identification of regulatory proteins associated with this specific actin cytoskeletal dynamic is a priority for understanding actin-based changes in nuclear shape and position in normal and pathological situations. Here, we first identify a unique family of actin regulators, the refilin proteins (RefilinA and RefilinB), that stabilize specifically perinuclear actin filament bundles. We next identify the actin-binding filamin A (FLNA) protein as the downstream effector of refilins. Refilins act as molecular switches to convert FLNA from an actin branching protein into one that bundles. In NIH 3T3 fibroblasts, the RefilinB/FLNA complex organizes the perinuclear actin filament bundles forming the actin cap. Finally, we demonstrate that in epithelial normal murine mammary gland (NmuMG) cells, the RefilinB/FLNA complex controls formation of a new perinuclear actin network that accompanies nuclear shape changes during the epithelial–mesenchymal transition (EMT). Our studies open perspectives for further functional analyses of this unique actin-based network and shed light on FLNA function during development and in human syndromes associated with FLNA mutations. PMID:21709252

  9. Oscillatory Increases in Alkalinity Anticipate Growth and May Regulate Actin Dynamics in Pollen Tubes of Lily[W][OA

    PubMed Central

    Lovy-Wheeler, Alenka; Kunkel, Joseph G.; Allwood, Ellen G.; Hussey, Patrick J.; Hepler, Peter K.

    2006-01-01

    Lily (Lilium formosanum or Lilium longiflorum) pollen tubes, microinjected with a low concentration of the pH-sensitive dye bis-carboxyethyl carboxyfluorescein dextran, show oscillating pH changes in their apical domain relative to growth. An increase in pH in the apex precedes the fastest growth velocities, whereas a decline follows growth, suggesting a possible relationship between alkalinity and cell extension. A target for pH may be the actin cytoskeleton, because the apical cortical actin fringe resides in the same region as the alkaline band in lily pollen tubes and elongation requires actin polymerization. A pH-sensitive actin binding protein, actin-depolymerizing factor (ADF), together with actin-interacting protein (AIP) localize to the cortical actin fringe region. Modifying intracellular pH leads to reorganization of the actin cytoskeleton, especially in the apical domain. Acidification causes actin filament destabilization and inhibits growth by 80%. Upon complete growth inhibition, the actin fringe is the first actin cytoskeleton component to disappear. We propose that during normal growth, the pH increase in the alkaline band stimulates the fragmenting activity of ADF/AIP, which in turn generates more sites for actin polymerization. Increased actin polymerization supports faster growth rates and a proton influx, which inactivates ADF/AIP, decreases actin polymerization, and retards growth. As pH stabilizes and increases, the activity of ADF/AIP again increases, repeating the cycle of events. PMID:16920777

  10. Pedestal structure and stability in H-mode and I-mode: a comparative study on Alcator C-Mod

    NASA Astrophysics Data System (ADS)

    Hughes, J. W.; Snyder, P. B.; Walk, J. R.; Davis, E. M.; Diallo, A.; LaBombard, B.; Baek, S. G.; Churchill, R. M.; Greenwald, M.; Groebner, R. J.; Hubbard, A. E.; Lipschultz, B.; Marmar, E. S.; Osborne, T.; Reinke, M. L.; Rice, J. E.; Theiler, C.; Terry, J.; White, A. E.; Whyte, D. G.; Wolfe, S.; Xu, X. Q.

    2013-04-01

    New experimental data from the Alcator C-Mod tokamak are used to benchmark predictive modelling of the edge pedestal in various high-confinement regimes, contributing to greater confidence in projection of pedestal height and width in ITER and reactors. ELMy H-modes operate near stability limits for ideal peeling-ballooning modes, as shown by calculations with the ELITE code. Experimental pedestal width in ELMy H-mode scales as the square root of βpol at the pedestal top, i.e. the dependence expected from theory if kinetic ballooning modes (KBMs) were responsible for limiting the pedestal width. A search for KBMs in experiment has revealed a short-wavelength electromagnetic fluctuation in the pedestal that is a candidate driver for inter-edge localized mode (ELM) pedestal regulation. A predictive pedestal model (EPED) has been tested on an extended set of ELMy H-modes from C-Mod, reproducing pedestal height and width reasonably well across the data set, and extending the tested range of EPED to the highest absolute pressures available on any existing tokamak and to within a factor of three of the pedestal pressure targeted for ITER. In addition, C-Mod offers access to two regimes, enhanced D-alpha (EDA) H-mode and I-mode, that have high pedestals, but in which large ELM activity is naturally suppressed and, instead, particle and impurity transport are regulated continuously. Pedestals of EDA H-mode and I-mode discharges are found to be ideal magnetohydrodynamic (MHD) stable with ELITE, consistent with the general absence of ELM activity. Invocation of alternative physics mechanisms may be required to make EPED-like predictions of pedestals in these kinds of intrinsically ELM-suppressed regimes, which would be very beneficial to operation in burning plasma devices.

  11. Pedestal properties of H-modes with negative triangularity using the EPED-CH model

    NASA Astrophysics Data System (ADS)

    Merle, A.; Sauter, O.; Medvedev, S. Yu

    2017-10-01

    The EPED model has been designed to predict the pedestal height and width from a minimal set of parameters and using the stability of the pedestal region for global MHD peeling-ballooning (P-B) modes as well as local kinetic ballooning modes (KBMs). This approach has been validated for type-I ELMy H-modes and quiescent H-modes (QH) but can also be used for other types of H-modes where it usually sets an upper limit on the achievable pedestal height. Using the recently developed EPED-like model called EPED-CH and based on the equilibrium codes CHEASE and CAXE and the MHD stability code KINX, we investigate in this work the effect of negative triangularity on the pedestal structure. Our simulation results confirm the experimental results from TCV where a reduction of the pedestal height was observed when going from positive to negative top triangularity. This was interpreted as a degradation of the peeling-ballooning stability due to the closed access to the second stability region for ballooning modes in the case of negative triangularity. This effect is further enhanced by the coupling to the KBM stability criterion in EPED simulations. The novel concept of the negative triangularity tokamak (a DEMO-sized machine) is also investigated. Again a strong reduction of the pedestal height and width is observed going from positive to negative triangularity for up-down symmetric equilibria. The pedestal height is also reduced going to more up-down asymmetric cases. The beneficial effect of the global β value on the pedestal height, which is linked to the second stability access, is strongly reduced for negative triangularity.

  12. Two-dimensional magnetohydrodynamic simulations of poloidal flows in tokamaks and MHD pedestal

    SciTech Connect

    Guazzotto, L.; Betti, R.

    2011-09-15

    Poloidal rotation is routinely observed in present-day tokamak experiments, in particular near the plasma edge and in the high-confinement mode of operation. According to the magnetohydrodynamic (MHD) equilibrium theory [R. Betti and J. P. Freidberg, Phys. Plasmas 7, 2439 (2000)], radial discontinuities form when the poloidal velocity exceeds the poloidal sound speed (or rather, more correctly, the poloidal magneto-slow speed). Two-dimensional compressible magnetohydrodynamic simulations show that the transonic discontinuities develop on a time scale of a plasma poloidal revolution to form an edge density pedestal and a localized velocity shear layer at the pedestal location. While such an MHD pedestal surrounds the entire core, the outboard side of the pedestal is driven by the transonic discontinuity while the inboard side is caused by a poloidal redistribution of the mass. The MHD simulations use a smooth momentum source to drive the poloidal flow. Soon after the flow exceeds the poloidal sound speed, the density pedestal and the velocity shear layer form and persist into a quasi steady state. These results may be relevant to the L-H transition, the early stages of the pedestal and edge transport barrier formation.

  13. Probe Measurements in the H-mode Pedestal Region in the Pegasus Toroidal Experiment

    NASA Astrophysics Data System (ADS)

    Bodner, G. M.; Bongard, M. W.; Fonck, R. J.; Thome, K. E.; Thompson, D. S.

    2014-10-01

    In near-unity aspect ratio Pegasus discharges, Ohmic heating and high-field-side fueling together trigger an L-H mode transition in both limited and diverted configurations. H-mode plasmas are predicted to exhibit pedestals in both the pressure and current density profiles. Operation at A ~ 1 allows for the use of local magnetic and Langmuir probes in the pedestal region. A current pedestal is routinely observed in Pegasus H-mode plasmas, but not in L-mode plasmas or during ELMs. Conventionally, edge pedestal measurements are observed in the edge pressure profile. A triple Langmuir probe has recently been installed in order to investigate the structure of the edge pressure pedestal in Pegasus H-mode discharges and complement the current density profile measurements. Local density and temperature measurements will be collected using the triple Langmuir probe at varying spatial locations to identify edge pressure profiles. These pressure profiles will be measured in both the L-mode and H-mode regimes. The triple probe will additionally be used to observe the turbulence levels before, during, and after the L-H mode transition. Complete density and temperature profiles including the pedestal will be obtained using a combination of Langmuir probe and Thomson scattering measurements. Work supported by US DOE Grant DE-FG02-96ER54375.

  14. Martian pedestal craters: Marginal sublimation pits implicate a climate-related formation mechanism

    NASA Astrophysics Data System (ADS)

    Kadish, Seth J.; Head, James W.; Barlow, Nadine G.; Marchant, David R.

    2008-08-01

    Pedestal craters on Mars are defined by an outward-facing scarp forming a plateau perched tens of meters above the surrounding terrain. Their origin has been attributed to impact armoring of the surface and subsequent removal of inter-crater terrain by either eolian deflation or sublimation of an ice-rich substrate. We identified 2696 pedestal craters between ~60°N and 60°S latitude; 98% are poleward of 33°N and 40°S. The majority of pedestal crater margins are smoothly sloped, but ~3%, concentrated in Utopia Planitia and Malea Planum, display distinctive marginal pits. These pedestal crater scarps are anomalously tall (usually >80-100 m) and the pits resemble sublimation depressions seen on Earth and elsewhere on Mars, providing evidence for sublimation of volatiles in the scarp, where the armored surface has tapered. The pitted scarps provide insight into the origin of the general pedestal crater population, favoring formation via deposition of a volatile-rich substrate, impact armoring, and sublimation of intervening volatiles. Crater densities and overlapping pedestal craters suggest multiple periods of emplacement and loss of these climate-related, latitude-dependent deposits throughout the Amazonian.

  15. Two-dimensional magnetohydrodynamic simulations of poloidal flows in tokamaks and MHD pedestal

    NASA Astrophysics Data System (ADS)

    Guazzotto, L.; Betti, R.

    2011-09-01

    Poloidal rotation is routinely observed in present-day tokamak experiments, in particular near the plasma edge and in the high-confinement mode of operation. According to the magnetohydrodynamic (MHD) equilibrium theory [R. Betti and J. P. Freidberg, Phys. Plasmas 7, 2439 (2000)], radial discontinuities form when the poloidal velocity exceeds the poloidal sound speed (or rather, more correctly, the poloidal magneto-slow speed). Two-dimensional compressible magnetohydrodynamic simulations show that the transonic discontinuities develop on a time scale of a plasma poloidal revolution to form an edge density pedestal and a localized velocity shear layer at the pedestal location. While such an MHD pedestal surrounds the entire core, the outboard side of the pedestal is driven by the transonic discontinuity while the inboard side is caused by a poloidal redistribution of the mass. The MHD simulations use a smooth momentum source to drive the poloidal flow. Soon after the flow exceeds the poloidal sound speed, the density pedestal and the velocity shear layer form and persist into a quasi steady state. These results may be relevant to the L-H transition, the early stages of the pedestal and edge transport barrier formation.

  16. Directed actin assembly and motility.

    PubMed

    Boujemaa-Paterski, Rajaa; Galland, Rémi; Suarez, Cristian; Guérin, Christophe; Théry, Manuel; Blanchoin, Laurent

    2014-01-01

    The actin cytoskeleton is a key component of the cellular architecture. However, understanding actin organization and dynamics in vivo is a complex challenge. Reconstitution of actin structures in vitro, in simplified media, allows one to pinpoint the cellular biochemical components and their molecular interactions underlying the architecture and dynamics of the actin network. Previously, little was known about the extent to which geometrical constraints influence the dynamic ultrastructure of these networks. Therefore, in order to study the balance between biochemical and geometrical control of complex actin organization, we used the innovative methodologies of UV and laser patterning to design a wide repertoire of nucleation geometries from which we assembled branched actin networks. Using these methods, we were able to reconstitute complex actin network organizations, closely related to cellular architecture, to precisely direct and control their 3D connections. This methodology mimics the actin networks encountered in cells and can serve in the fabrication of innovative bioinspired systems.

  17. Amplification of actin polymerization forces

    PubMed Central

    Dmitrieff, Serge; Nédélec, François

    2016-01-01

    The actin cytoskeleton drives many essential processes in vivo, using molecular motors and actin assembly as force generators. We discuss here the propagation of forces caused by actin polymerization, highlighting simple configurations where the force developed by the network can exceed the sum of the polymerization forces from all filaments. PMID:27002174

  18. Amplification of actin polymerization forces.

    PubMed

    Dmitrieff, Serge; Nédélec, François

    2016-03-28

    The actin cytoskeleton drives many essential processes in vivo, using molecular motors and actin assembly as force generators. We discuss here the propagation of forces caused by actin polymerization, highlighting simple configurations where the force developed by the network can exceed the sum of the polymerization forces from all filaments.

  19. Unique Properties of Eukaryote-Type Actin and Profilin Horizontally Transferred to Cyanobacteria

    PubMed Central

    Guljamow, Arthur; Delissen, Friedmar; Baumann, Otto; Thünemann, Andreas F.; Dittmann, Elke

    2012-01-01

    A eukaryote-type actin and its binding protein profilin encoded on a genomic island in the cyanobacterium Microcystis aeruginosa PCC 7806 co-localize to form a hollow, spherical enclosure occupying a considerable intracellular space as shown by in vivo fluorescence microscopy. Biochemical and biophysical characterization reveals key differences between these proteins and their eukaryotic homologs. Small-angle X-ray scattering shows that the actin assembles into elongated, filamentous polymers which can be visualized microscopically with fluorescent phalloidin. Whereas rabbit actin forms thin cylindrical filaments about 100 µm in length, cyanobacterial actin polymers resemble a ribbon, arrest polymerization at 5-10 µm and tend to form irregular multi-strand assemblies. While eukaryotic profilin is a specific actin monomer binding protein, cyanobacterial profilin shows the unprecedented property of decorating actin filaments. Electron micrographs show that cyanobacterial profilin stimulates actin filament bundling and stabilizes their lateral alignment into heteropolymeric sheets from which the observed hollow enclosure may be formed. We hypothesize that adaptation to the confined space of a bacterial cell devoid of binding proteins usually regulating actin polymerization in eukaryotes has driven the co-evolution of cyanobacterial actin and profilin, giving rise to an intracellular entity. PMID:22253827

  20. Actinic keratosis. Current treatment options.

    PubMed

    Jeffes, E W; Tang, E H

    2000-01-01

    Actinic keratoses are hyperkeratotic skin lesions that represent focal abnormal proliferation of epidermal keratinocytes. Some actinic keratoses evolve into squamous cell carcinoma of the skin, while others resolve spontaneously. The conversion rate of actinic keratosis to squamous cell carcinoma is not accurately known, but appears to be in the range of 0.25 to 1% per year. Although there is a low rate of conversion of actinic keratoses to squamous cell carcinoma, 60% of squamous cell carcinomas of the skin probably arise from actinic keratoses. The main cause of actinic keratoses in otherwise healthy Caucasians appears to be the sun. Therapy for actinic keratoses begins with prevention which starts with sun avoidance and physical protection. Sunprotection with sunscreens actually slows the return of actinic keratoses in patients already getting actinic keratoses. Interestingly, a few studies are available that demonstrate that a high fat diet is associated with the production of more actinic keratoses than is a low fat diet. One of the mainstays of therapy has been local destruction of the actinic keratoses with cryotherapy, and curettage and electrodesiccation. A new addition to this group of therapies to treat individual actinic keratoses is photodynamic therapy with topical aminolevulinic acid and light. In patients who have numerous actinic keratoses in an area of severely sun damaged skin, therapies which are applied to the whole actinic keratosis area are used. The goal of treating such an area of skin is to treat all of the early as well as the numerous clinically evident actinic keratoses at the same time. The classical approaches for treating areas of photodamaged skin without treating actinic keratoses individually include: the use of topically applied fluorouracil cream, dermabrasion, and cutaneous peels with various agents like trichloroacetic acid. Both topically as well as orally administered retinoids have been used to treat actinic keratoses but

  1. Cytoplasmic Actin Is an Extracellular Insect Immune Factor which Is Secreted upon Immune Challenge and Mediates Phagocytosis and Direct Killing of Bacteria, and Is a Plasmodium Antagonist

    PubMed Central

    Sandiford, Simone L.; Dong, Yuemei; Pike, Andrew; Blumberg, Benjamin J.; Bahia, Ana C.; Dimopoulos, George

    2015-01-01

    Actin is a highly versatile, abundant, and conserved protein, with functions in a variety of intracellular processes. Here, we describe a novel role for insect cytoplasmic actin as an extracellular pathogen recognition factor that mediates antibacterial defense. Insect actins are secreted from cells upon immune challenge through an exosome-independent pathway. Anopheles gambiae actin interacts with the extracellular MD2-like immune factor AgMDL1, and binds to the surfaces of bacteria, mediating their phagocytosis and direct killing. Globular and filamentous actins display distinct functions as extracellular immune factors, and mosquito actin is a Plasmodium infection antagonist. PMID:25658622

  2. Control of actin-based motility through localized actin binding

    PubMed Central

    Banigan, Edward J.; Lee, Kun-Chun; Liu, Andrea J.

    2014-01-01

    A wide variety of cell biological and biomimetic systems use actin polymerization to drive motility. It has been suggested that an object such as a bacterium can propel itself by self-assembling a high concentration of actin behind it if it is repelled by actin. However, it is also known that it is essential for the moving object to bind actin. Therefore, a key question is how the actin tail can propel an object when it both binds and repels the object. We present a physically consistent Brownian dynamics model for actin-based motility that includes the minimal components of the dendritic nucleation model and allows for both attractive and repulsive interactions between actin and a moveable disk. We find that the concentration gradient of filamentous actin generated by polymerization is sufficient to propel the object, even with moderately strong binding interactions. Additionally, actin binding can act as a biophysical cap, and may directly control motility through modulation of network growth. Overall, this mechanism is robust in that it can drive motility against a load up to a stall pressure that depends on the Young’s modulus of the actin network and can explain several aspects of actin-based motility. PMID:24225232

  3. Global and pedestal confinement in JET with a Be/W metallic wall

    NASA Astrophysics Data System (ADS)

    Beurskens, M. N. A.; Frassinetti, L.; Challis, C.; Giroud, C.; Saarelma, S.; Alper, B.; Angioni, C.; Bilkova, P.; Bourdelle, C.; Brezinsek, S.; Buratti, P.; Calabro, G.; Eich, T.; Flanagan, J.; Giovannozzi, E.; Groth, M.; Hobirk, J.; Joffrin, E.; Leyland, M. J.; Lomas, P.; de la Luna, E.; Kempenaars, M.; Maddison, G.; Maggi, C.; Mantica, P.; Maslov, M.; Matthews, G.; Mayoral, M.-L.; Neu, R.; Nunes, I.; Osborne, T.; Rimini, F.; Scannell, R.; Solano, E. R.; Snyder, P. B.; Voitsekhovitch, I.; de Vries, Peter; Contributors, JET-EFDA

    2014-04-01

    Type I ELMy H-mode operation in JET with the ITER-like Be/W wall (JET-ILW) generally occurs at lower pedestal pressures compared to those with the full carbon wall (JET-C). The pedestal density is similar but the pedestal temperature where type I ELMs occur is reduced and below to the so-called critical type I-type III transition temperature reported in JET-C experiments. Furthermore, the confinement factor H98(y,2) in type I ELMy H-mode baseline plasmas is generally lower in JET-ILW compared to JET-C at low power fractions Ploss/Pthr,08 < 2 (where Ploss is (Pin - dW/dt), and Pthr,08 the L-H power threshold from Martin et al 2008 (J. Phys. Conf. Ser. 123 012033)). Higher power fractions have thus far not been achieved in the baseline plasmas. At Ploss/Pthr,08 > 2, the confinement in JET-ILW hybrid plasmas is similar to that in JET-C. A reduction in pedestal pressure is the main reason for the reduced confinement in JET-ILW baseline ELMy H-mode plasmas where typically H98(y,2) = 0.8 is obtained, compared to H98(y,2) = 1.0 in JET-C. In JET-ILW hybrid plasmas a similarly reduced pedestal pressure is compensated by an increased peaking of the core pressure profile resulting in H98(y,2) ⩽ 1.25. The pedestal stability has significantly changed in high triangularity baseline plasmas where the confinement loss is also most apparent. Applying the same stability analysis for JET-C and JET-ILW, the measured pedestal in JET-ILW is stable with respect to the calculated peeling-ballooning stability limit and the ELM collapse time has increased to 2 ms from typically 200 µs in JET-C. This indicates that changes in the pedestal stability may have contributed to the reduced pedestal confinement in JET-ILW plasmas. A comparison of EPED1 pedestal pressure prediction with JET-ILW experimental data in over 500 JET-C and JET-ILW baseline and hybrid plasmas shows a good agreement with 0.8 < (measured pped)/(predicted pped,EPED) < 1.2, but that the role of triangularity is generally

  4. Visualization of actin polymerization in invasive structures of macrophages and carcinoma cells using photoconvertible β-actin-Dendra2 fusion proteins.

    PubMed

    Dovas, Athanassios; Gligorijevic, Bojana; Chen, Xiaoming; Entenberg, David; Condeelis, John; Cox, Dianne

    2011-02-14

    Actin polymerization controls a range of cellular processes, from intracellular trafficking to cell motility and invasion. Generation and elongation of free barbed ends defines the regions of actively polymerizing actin in cells and, consequently, is of importance in the understanding of the mechanisms through which actin dynamics are regulated. Herein we present a method that does not involve cell permeabilization and provides direct visualization of growing barbed ends using photoswitchable β-actin-Dendra2 constructs expressed in murine macrophage and rat mammary adenocarcinoma cell lines. The method exploits the ability of photoconverted (red) G-actin species to become incorporated into pre-existing (green) actin filaments, visualized in two distinct wavelengths using TIRF microscopy. In growing actin filaments, photoconverted (red) monomers are added to the barbed end while only green monomers are recycled from the pointed end. We demonstrate that incorporation of actin into intact podosomes of macrophages occurs constitutively and is amenable to inhibition by cytochalasin D indicating barbed end incorporation. Additionally, actin polymerization does not occur in quiescent invadopodial precursors of carcinoma cells suggesting that the filaments are capped and following epidermal growth factor stimulation actin incorporation occurs in a single but extended peak. Finally, we show that Dendra2 fused to either the N- or the C-terminus of β-actin profoundly affects its localization and incorporation in distinct F-actin structures in carcinoma cells, thus influencing the ability of monomers to be photoconverted. These data support the use of photoswitchable actin-Dendra2 constructs as powerful tools in the visualization of free barbed ends in living cells.

  5. REM sleep deprivation attenuates actin-binding protein cortactin: a link between sleep and hippocampal plasticity.

    PubMed

    Davis, Christopher J; Meighan, Peter C; Taishi, Ping; Krueger, James M; Harding, Joseph W; Wright, John W

    2006-06-12

    Rapid eye-movement sleep (REMS) is thought to affect synaptic plasticity. Cortactin is a cytoskeletal protein critically involved in the regulation of actin branching and stabilization including the actin backbone of dendritic spines. Hippocampal cortactin levels, phosphorylation, and processing appear to be altered during learning and long-term potentiation (LTP); consistent with a role for cortactin in the dendritic restructuring that accompanies synaptic plasticity. In this study juvenile male Sprague-Dawley rats were selectively REMS-deprived (RD) for 48 h by the flowerpot method. Cage control (CC) and large pedestal control (PC) animals were used for comparison. Animals were euthanized immediately, or 12 h, after removal from the pedestal. The hippocampus was dissected, flash-frozen, and stored for subsequent Western blot or quantitative RT-PCR analysis of cortactin. Cortactin mRNA/cDNA levels initially rose in PC and RD rats but returned to CC levels by 12 h after removal from the pedestal. Predictably cortactin protein levels were initially unchanged but were up-regulated after 12 h. The PC group had more total and tyrosine-phosphorylated cortactin protein expression than the RD and CC groups. This increase in cortactin was likely due to the exposure of the rats to the novel environment of the deprivation chambers thus triggering plasticity events. The lack of REMS, however, severely hampered cortactin protein up-regulation and phosphorylation observed in the PC group suggesting an attenuation of plasticity-related events. Thus, these data support a functional link between REMS and cytoskeletal reorganization in the hippocampus, a process that is essential for synaptic plasticity.

  6. A study of poloidal asymmetries in the pedestal region

    NASA Astrophysics Data System (ADS)

    Churchill, R. M.; Lipschultz, B.; Lisgo, S.; Reimold, F.; Goldstein, J.; Alcator C-Mod Team Team

    2011-10-01

    Simultaneous CXRS measurements of boron density, velocity, and temperature in the pedestal region (0 . 8 < r / a < 1 . 05) at the low- and high-field sides (LFS and HFS) of Alcator C-Mod allow studies of variations in boron density and total velocity on a flux surface. While previous studies used different neutral sources (a 50keV hydrogen neutral beam at the LFS and a thermal D2 gas puff at the HFS) to localize CXRS measurements we have recently expanded our diagnostic set to allow thermal gas CXRS at both locations, thus removing uncertainties due to different measurement techniques and cross-section. We have also upgraded our modelling capability to utilize the DIVIMP code, which uses the plasma-neutral code combination of OSM-EIRENE to determine the local neutral density. Comparisons between CXRS methods using different neutral sources will be shown as well as an exploration of whether the constants K(ψ) and ω(ψ) in the description of V =K/(ψ) n B + ω (ψ)R2 ∇ ϕ are constant on a flux surface as typically assumed. The above comparisons will be shown for a variety of H-mode and I-mode plasmas. Supported by USDoE award DE-FC02-99ER54512.

  7. Laser heated pedestal growth system commissioning and fiber processing

    NASA Astrophysics Data System (ADS)

    Buric, Michael; Yip, M. J.; Chorpening, Ben; Ohodnicki, Paul

    2016-05-01

    A new Laser Heated Pedestal Growth system was designed and fabricated using various aspects of effective legacy designs for the growth of single-crystal high-temperature-compatible optical fibers. The system is heated by a 100-watt, DC driven, CO2 laser with PID power control. Fiber diameter measurements are performed using a telecentric video system which identifies the molten zone and utilizes edge detection algorithms to report fiber-diameter. Beam shaping components include a beam telescope; along with gold-coated reflaxicon, turning, and parabolic focusing mirrors consistent with similar previous systems. The optical system permits melting of sapphire-feedstock up to 1.5mm in diameter for growth. Details regarding operational characteristics are reviewed and properties of single-crystal sapphire fibers produced by the system are evaluated. Aspects of the control algorithm efficacy will be discussed, along with relevant alternatives. Finally, some new techniques for in-situ processing making use of the laser-heating system are discussed. Ex-situ fiber modification and processing are also examined for improvements in fiber properties.

  8. Intracellular microlasers

    NASA Astrophysics Data System (ADS)

    Humar, Matjaž; Hyun Yun, Seok

    2015-09-01

    Optical microresonators, which confine light within a small cavity, are widely exploited for various applications ranging from the realization of lasers and nonlinear devices to biochemical and optomechanical sensing. Here we use microresonators and suitable optical gain materials inside biological cells to demonstrate various optical functions in vitro including lasing. We explore two distinct types of microresonator—soft and hard—that support whispering-gallery modes. Soft droplets formed by injecting oil or using natural lipid droplets support intracellular laser action. The laser spectra from oil-droplet microlasers can chart cytoplasmic internal stress (˜500 pN μm-2) and its dynamic fluctuations at a sensitivity of 20 pN μm-2 (20 Pa). In a second form, whispering-gallery modes within phagocytized polystyrene beads of different sizes enable individual tagging of thousands of cells easily and, in principle, a much larger number by multiplexing with different dyes.

  9. System-wide organization of actin cytoskeleton determines organelle transport in hypocotyl plant cells

    PubMed Central

    Nowak, Jacqueline; Ivakov, Alexander; Somssich, Marc; Persson, Staffan; Nikoloski, Zoran

    2017-01-01

    The actin cytoskeleton is an essential intracellular filamentous structure that underpins cellular transport and cytoplasmic streaming in plant cells. However, the system-level properties of actin-based cellular trafficking remain tenuous, largely due to the inability to quantify key features of the actin cytoskeleton. Here, we developed an automated image-based, network-driven framework to accurately segment and quantify actin cytoskeletal structures and Golgi transport. We show that the actin cytoskeleton in both growing and elongated hypocotyl cells has structural properties facilitating efficient transport. Our findings suggest that the erratic movement of Golgi is a stable cellular phenomenon that might optimize distribution efficiency of cell material. Moreover, we demonstrate that Golgi transport in hypocotyl cells can be accurately predicted from the actin network topology alone. Thus, our framework provides quantitative evidence for system-wide coordination of cellular transport in plant cells and can be readily applied to investigate cytoskeletal organization and transport in other organisms. PMID:28655850

  10. Interaction of Phalloidin with Actin

    PubMed Central

    Lengsfeld, Anneliese M.; Löw, Irmentraut; Wieland, Theodor; Dancker, Peter; Hasselbach, Wilhelm

    1974-01-01

    Phalloidin, a toxic bicyclic peptide of rapid action from the toadstool, Amanita phalloides, gives rise to polymerization of G-actin to filamentous structures (Ph-actin) in a medium of low ionic strength. Ph-actin closely resembles the microfilaments found in liver membrane fractions (Ph-filaments) after in vivo or in vitro poisoning. Both phalloidin induced filaments are resistant to 0.6 M KI in contrast to F-actin, and become decorated by heavy meromyosin. After preincubation with cytochalasin B significantly fewer actin filaments are observed. Images PMID:4368830

  11. Metallothionein immunolocalization in actinic skin nonmelanoma carcinomas.

    PubMed

    Borges Júnior, Paulo C; Ribeiro, Rosy I M A; Cardoso, Sérgio V; Berbet, Alceu L C; Rocha, Ademir; Espindola, Foued S; Loyola, Adriano M

    2007-06-01

    Basal cell carcinoma (BCC) and squamous cell carcinoma (SCC) are the most frequent skin cancer. Its pathogeny is linked to genotoxic effects of actinic radiation exposure, especially to ultraviolet wavelength. Metallothionein (MT) is a low-molecular weight protein with high affinity for heavy metal. Its intracellular function has been related to heavy metals and free-radical detoxification, although many studies linked MT to protective action against actinic mutagenesis. In other way, overexpression in malignant tumors has been related to worse prognosis. We aimed to evaluate MT immunohistochemical expression in skin cancer associated to actinic radiation. Twenty-six BCC cases, 20 SCC, and 6 normal skin fragments were investigated. Immunohistochemical assay were performed by streptavidin-biotin-peroxidase technique with standard monoclonal antibody (E9). In normal skin, immunostaining was observed in basal layer of the epithelium. In the epithelium adjacent to tumors, suprabasal layer was also intensely labeled. Mean MT immunostaining indices were 18.5+21.2% for BCC and 69.1+14.4% for SCC. This difference was statistically significant. Higher MT expression in SCC as compared with BCC suggests association with tumoral aggressiveness.

  12. Clamped-filament elongation model for actin-based motors.

    PubMed Central

    Dickinson, Richard B; Purich, Daniel L

    2002-01-01

    Although actin-based motility drives cell crawling and intracellular locomotion of organelles and certain pathogens, the underlying mechanism of force generation remains a mystery. Recent experiments demonstrated that Listeria exhibit episodes of 5.4-nm stepwise motion corresponding to the periodicity of the actin filament subunits, and extremely small positional fluctuations during the intermittent pauses [S. C. Kuo and J. L. McGrath. 2000. Nature. 407:1026-1029]. These findings suggest that motile bacteria remain firmly bound to actin filament ends as they elongate, a behavior that appears to rule out previous models for actin-based motility. We propose and analyze a new mechanochemical model (called the "Lock, Load & Fire" mechanism) for force generation by means of affinity-modulated, clamped-filament elongation. During the locking step, the filament's terminal ATP-containing subunit binds tightly to a clamp situated on the surface of a motile object; in the loading step, actin.ATP monomer(s) bind to the filament end, an event that triggers the firing step, wherein ATP hydrolysis on the clamped subunit attenuates the filament's affinity for the clamp. This last step initiates translocation of the new ATP-containing terminus to the clamp, whereupon another cycle begins anew. This model explains how surface-tethered filaments can grow while exerting flexural or tensile force on the motile surface. Moreover, stochastic simulations of the model reproduce the signature motions of Listeria. This elongation motor, which we term actoclampin, exploits actin's intrinsic ATPase activity to provide a simple, high-fidelity enzymatic reaction cycle for force production that does not require elongating filaments to dissociate from the motile surface. This mechanism may operate whenever actin polymerization is called upon to generate the forces that drive cell crawling or intracellular organelle motility. PMID:11806905

  13. Cofilin 1-Mediated Biphasic F-Actin Dynamics of Neuronal Cells Affect Herpes Simplex Virus 1 Infection and Replication

    PubMed Central

    Xiang, Yangfei; Zheng, Kai; Ju, Huaiqiang; Wang, Shaoxiang; Pei, Ying; Ding, Weichao; Chen, Zhenping; Wang, Qiaoli; Qiu, Xianxiu; Zhong, Meigong; Zeng, Fanli; Ren, Zhe; Qian, Chuiwen; Liu, Ge

    2012-01-01

    Herpes simplex virus 1 (HSV-1) invades the nervous system and causes pathological changes. In this study, we defined the remodeling of F-actin and its possible mechanisms during HSV-1 infection of neuronal cells. HSV-1 infection enhanced the formation of F-actin-based structures in the early stage of infection, which was followed by a continuous decrease in F-actin during the later stages of infection. The disruption of F-actin dynamics by chemical inhibitors significantly reduced the efficiency of viral infection and intracellular HSV-1 replication. The active form of the actin-depolymerizing factor cofilin 1 was found to increase at an early stage of infection and then to continuously decrease in a manner that corresponded to the remodeling pattern of F-actin, suggesting that cofilin 1 may be involved in the biphasic F-actin dynamics induced by HSV-1 infection. Knockdown of cofilin 1 impaired HSV-1-induced F-actin assembly during early infection and inhibited viral entry; however, overexpression of cofilin 1 did not affect F-actin assembly or viral entry during early infection but decreased intracellular viral reproduction efficiently. Our results, for the first time, demonstrated the biphasic F-actin dynamics in HSV-1 neuronal infection and confirmed the association of F-actin with the changes in the expression and activity of cofilin 1. These results may provide insight into the mechanism by which HSV-1 productively infects neuronal cells and causes pathogenesis. PMID:22623803

  14. Actin dynamics at the Golgi complex in mammalian cells.

    PubMed

    Egea, Gustavo; Lázaro-Diéguez, Francisco; Vilella, Montserrat

    2006-04-01

    Secretion and endocytosis are highly dynamic processes that are sensitive to external stimuli. Thus, in multicellular organisms, different cell types utilize specialised pathways of intracellular membrane traffic to facilitate specific physiological functions. In addition to the complex internal molecular factors that govern sorting functions and fission or fusion of transport carriers, the actin cytoskeleton plays an important role in both the endocytic and secretory pathways. The interaction between the actin cytoskeleton and membrane trafficking is not restricted to transport processes: it also appears to be directly involved in the biogenesis of Golgi-derived transport carriers (budding and fission processes) and in the maintenance of the unique flat shape of Golgi cisternae.

  15. Regulators of Actin Dynamics in Gastrointestinal Tract Tumors

    PubMed Central

    Steinestel, Konrad; Wardelmann, Eva; Hartmann, Wolfgang; Grünewald, Inga

    2015-01-01

    Reorganization of the actin cytoskeleton underlies cell migration in a wide variety of physiological and pathological processes, such as embryonic development, wound healing, and tumor cell invasion. It has been shown that actin assembly and disassembly are precisely regulated by intracellular signaling cascades that respond to changes in the cell microenvironment, ligand binding to surface receptors, or oncogenic transformation of the cell. Actin-nucleating and actin-depolymerizing (ANFs/ADFs) and nucleation-promoting factors (NPFs) regulate cytoskeletal dynamics at the leading edge of migrating cells, thereby modulating cell shape; these proteins facilitate cellular movement and mediate degradation of the surrounding extracellular matrix by secretion of lytic proteases, thus eliminating barriers for tumor cell invasion. Accordingly, expression and activity of these actin-binding proteins have been linked to enhanced metastasis and poor prognosis in a variety of malignancies. In this review, we will summarize what is known about expression patterns and the functional role of actin regulators in gastrointestinal tumors and evaluate first pharmacological approaches to prevent invasion and metastatic dissemination of malignant cells. PMID:26345720

  16. Cell elasticity is regulated by the tropomyosin isoform composition of the actin cytoskeleton.

    PubMed

    Jalilian, Iman; Heu, Celine; Cheng, Hong; Freittag, Hannah; Desouza, Melissa; Stehn, Justine R; Bryce, Nicole S; Whan, Renee M; Hardeman, Edna C; Fath, Thomas; Schevzov, Galina; Gunning, Peter W

    2015-01-01

    The actin cytoskeleton is the primary polymer system within cells responsible for regulating cellular stiffness. While various actin binding proteins regulate the organization and dynamics of the actin cytoskeleton, the proteins responsible for regulating the mechanical properties of cells are still not fully understood. In the present study, we have addressed the significance of the actin associated protein, tropomyosin (Tpm), in influencing the mechanical properties of cells. Tpms belong to a multi-gene family that form a co-polymer with actin filaments and differentially regulate actin filament stability, function and organization. Tpm isoform expression is highly regulated and together with the ability to sort to specific intracellular sites, result in the generation of distinct Tpm isoform-containing actin filament populations. Nanomechanical measurements conducted with an Atomic Force Microscope using indentation in Peak Force Tapping in indentation/ramping mode, demonstrated that Tpm impacts on cell stiffness and the observed effect occurred in a Tpm isoform-specific manner. Quantitative analysis of the cellular filamentous actin (F-actin) pool conducted both biochemically and with the use of a linear detection algorithm to evaluate actin structures revealed that an altered F-actin pool does not absolutely predict changes in cell stiffness. Inhibition of non-muscle myosin II revealed that intracellular tension generated by myosin II is required for the observed increase in cell stiffness. Lastly, we show that the observed increase in cell stiffness is partially recapitulated in vivo as detected in epididymal fat pads isolated from a Tpm3.1 transgenic mouse line. Together these data are consistent with a role for Tpm in regulating cell stiffness via the generation of specific populations of Tpm isoform-containing actin filaments.

  17. Cell Elasticity Is Regulated by the Tropomyosin Isoform Composition of the Actin Cytoskeleton

    PubMed Central

    Jalilian, Iman; Heu, Celine; Cheng, Hong; Freittag, Hannah; Desouza, Melissa; Stehn, Justine R.; Bryce, Nicole S.; Whan, Renee M.; Hardeman, Edna C.

    2015-01-01

    The actin cytoskeleton is the primary polymer system within cells responsible for regulating cellular stiffness. While various actin binding proteins regulate the organization and dynamics of the actin cytoskeleton, the proteins responsible for regulating the mechanical properties of cells are still not fully understood. In the present study, we have addressed the significance of the actin associated protein, tropomyosin (Tpm), in influencing the mechanical properties of cells. Tpms belong to a multi-gene family that form a co-polymer with actin filaments and differentially regulate actin filament stability, function and organization. Tpm isoform expression is highly regulated and together with the ability to sort to specific intracellular sites, result in the generation of distinct Tpm isoform-containing actin filament populations. Nanomechanical measurements conducted with an Atomic Force Microscope using indentation in Peak Force Tapping in indentation/ramping mode, demonstrated that Tpm impacts on cell stiffness and the observed effect occurred in a Tpm isoform-specific manner. Quantitative analysis of the cellular filamentous actin (F-actin) pool conducted both biochemically and with the use of a linear detection algorithm to evaluate actin structures revealed that an altered F-actin pool does not absolutely predict changes in cell stiffness. Inhibition of non-muscle myosin II revealed that intracellular tension generated by myosin II is required for the observed increase in cell stiffness. Lastly, we show that the observed increase in cell stiffness is partially recapitulated in vivo as detected in epididymal fat pads isolated from a Tpm3.1 transgenic mouse line. Together these data are consistent with a role for Tpm in regulating cell stiffness via the generation of specific populations of Tpm isoform-containing actin filaments. PMID:25978408

  18. Actin stress in cell reprogramming

    PubMed Central

    Guo, Jun; Wang, Yuexiu; Sachs, Frederick; Meng, Fanjie

    2014-01-01

    Cell mechanics plays a role in stem cell reprogramming and differentiation. To understand this process better, we created a genetically encoded optical probe, named actin–cpstFRET–actin (AcpA), to report forces in actin in living cells in real time. We showed that stemness was associated with increased force in actin. We reprogrammed HEK-293 cells into stem-like cells using no transcription factors but simply by softening the substrate. However, Madin-Darby canine kidney (MDCK) cell reprogramming required, in addition to a soft substrate, Harvey rat sarcoma viral oncogene homolog expression. Replating the stem-like cells on glass led to redifferentiation and reduced force in actin. The actin force probe was a FRET sensor, called cpstFRET (circularly permuted stretch sensitive FRET), flanked by g-actin subunits. The labeled actin expressed efficiently in HEK, MDCK, 3T3, and bovine aortic endothelial cells and in multiple stable cell lines created from those cells. The viability of the cell lines demonstrated that labeled actin did not significantly affect cell physiology. The labeled actin distribution was similar to that observed with GFP-tagged actin. We also examined the stress in the actin cross-linker actinin. Actinin force was not always correlated with actin force, emphasizing the need for addressing protein specificity when discussing forces. Because actin is a primary structural protein in animal cells, understanding its force distribution is central to understanding animal cell physiology and the many linked reactions such as stress-induced gene expression. This new probe permits measuring actin forces in a wide range of experiments on preparations ranging from isolated proteins to transgenic animals. PMID:25422450

  19. Recruitment Kinetics of Tropomyosin Tpm3.1 to Actin Filament Bundles in the Cytoskeleton Is Independent of Actin Filament Kinetics.

    PubMed

    Appaduray, Mark A; Masedunskas, Andrius; Bryce, Nicole S; Lucas, Christine A; Warren, Sean C; Timpson, Paul; Stear, Jeffrey H; Gunning, Peter W; Hardeman, Edna C

    2016-01-01

    The actin cytoskeleton is a dynamic network of filaments that is involved in virtually every cellular process. Most actin filaments in metazoa exist as a co-polymer of actin and tropomyosin (Tpm) and the function of an actin filament is primarily defined by the specific Tpm isoform associated with it. However, there is little information on the interdependence of these co-polymers during filament assembly and disassembly. We addressed this by investigating the recovery kinetics of fluorescently tagged isoform Tpm3.1 into actin filament bundles using FRAP analysis in cell culture and in vivo in rats using intracellular intravital microscopy, in the presence or absence of the actin-targeting drug jasplakinolide. The mobile fraction of Tpm3.1 is between 50% and 70% depending on whether the tag is at the C- or N-terminus and whether the analysis is in vivo or in cultured cells. We find that the continuous dynamic exchange of Tpm3.1 is not significantly impacted by jasplakinolide, unlike tagged actin. We conclude that tagged Tpm3.1 may be able to undergo exchange in actin filament bundles largely independent of the assembly and turnover of actin.

  20. Recruitment Kinetics of Tropomyosin Tpm3.1 to Actin Filament Bundles in the Cytoskeleton Is Independent of Actin Filament Kinetics

    PubMed Central

    Appaduray, Mark A.; Masedunskas, Andrius; Lucas, Christine A.; Warren, Sean C.; Timpson, Paul; Stear, Jeffrey H.

    2016-01-01

    The actin cytoskeleton is a dynamic network of filaments that is involved in virtually every cellular process. Most actin filaments in metazoa exist as a co-polymer of actin and tropomyosin (Tpm) and the function of an actin filament is primarily defined by the specific Tpm isoform associated with it. However, there is little information on the interdependence of these co-polymers during filament assembly and disassembly. We addressed this by investigating the recovery kinetics of fluorescently tagged isoform Tpm3.1 into actin filament bundles using FRAP analysis in cell culture and in vivo in rats using intracellular intravital microscopy, in the presence or absence of the actin-targeting drug jasplakinolide. The mobile fraction of Tpm3.1 is between 50% and 70% depending on whether the tag is at the C- or N-terminus and whether the analysis is in vivo or in cultured cells. We find that the continuous dynamic exchange of Tpm3.1 is not significantly impacted by jasplakinolide, unlike tagged actin. We conclude that tagged Tpm3.1 may be able to undergo exchange in actin filament bundles largely independent of the assembly and turnover of actin. PMID:27977753

  1. Disseminated superficial actinic porokeratosis.

    PubMed

    Rouhani, Panta; Fischer, Max; Meehan, Shane; Pomeranz, Miriam Keltz

    2012-12-15

    Disseminated superficial actinic porokeratosis, which was described in 1966, is characterized by small, atrophic patches with distinctive keratin rims that occur on sun-exposed areas of the extremities, shoulders, and back. The diagnosis is based on the histopathologic finding of a cornoid lamella, absence of a granular layer, and often a thin epidermis. It is associated with exposure to ultraviolet radiation. Gene studies suggest a pathway defect in which several mutations in keratinocyte proliferation and differentiation lead to development of porokeratosis.

  2. Ca2+ regulation of gelsolin activity: binding and severing of F-actin.

    PubMed Central

    Kinosian, H J; Newman, J; Lincoln, B; Selden, L A; Gershman, L C; Estes, J E

    1998-01-01

    Regulation of the F-actin severing activity of gelsolin by Ca2+ has been investigated under physiologic ionic conditions. Tryptophan fluorescence intensity measurements indicate that gelsolin contains at least two Ca2+ binding sites with affinities of 2.5 x 10(7) M-1 and 1.5 x 10(5) M-1. At F-actin and gelsolin concentrations in the range of those found intracellularly, gelsolin is able to bind F-actin with half-maximum binding at 0.14 microM free Ca2+ concentration. Steady-state measurements of gelsolin-induced actin depolymerization suggest that half-maximum depolymerization occurs at approximately 0.4 microM free Ca2+ concentration. Dynamic light scattering measurements of the translational diffusion coefficient for actin filaments and nucleated polymerization assays for number concentration of actin filaments both indicate that severing of F-actin occurs slowly at micromolar free Ca2+ concentrations. The data suggest that binding of Ca2+ to the gelsolin-F-actin complex is the rate-limiting step for F-actin severing by gelsolin; this Ca2+ binding event is a committed step that results in a Ca2+ ion bound at a high-affinity, EGTA-resistant site. The very high affinity of gelsolin for the barbed end of an actin filament drives the binding reaction equilibrium toward completion under conditions where the reaction rate is slow. PMID:9826630

  3. Quantitative Analysis of Statics and Dynamics of Actin Cables in Fission Yeast

    NASA Astrophysics Data System (ADS)

    Yusuf, Eddy; Wu, Jian-Qiu; Vavylonis, Dimitrios

    2010-03-01

    The assembly of actin and tubulin proteins into long filaments and bundles, i.e. closely-packed filaments, underlies important cellular processes such as cell motility, intracellular transport, and cell division. Recent theoretical and experimental work has addressed the nonequilibrium dynamics of single microtubules within live cells [1]. Actin filaments usually form dense networks that prevents microscopic imaging of individual filaments or bundles. Here, we studied actin dynamics using fission yeast that has low-density actin cytoskeleton consisting of actin cables (actin bundles aligned along the long axis of the cell) and ``actin patches.'' Yeast cells expressing GFP-CHD were imaged by 3D confocal microscopy. Stretching open active contours [2] were used to segment and track individual actin cables. We analyzed their curvature distribution, the tangent correlation, and the temporal bending amplitude fluctuations. We contrast our findings to equilibrium fluctuating semiflexible polymers and to microtubules in cells. We calculate the important time and length scales for the actin cables. We also discuss our findings within the broad context of understanding actin assembly in cells. [1] C. P. Brangwynne et. al., Phys. Rev. Lett. 100, 118104 (2008) [2] H. Li et. al., Proc. of the IEEE Int'l Symposium on Biomedical Imaging: From Nano to Macro, ISBI'09

  4. Role of actin cortex in the subplasmalemmal transport of secretory granules in PC-12 cells.

    PubMed Central

    Lang, T; Wacker, I; Wunderlich, I; Rohrbach, A; Giese, G; Soldati, T; Almers, W

    2000-01-01

    In neuroendocrine PC-12 cells, evanescent-field fluorescence microscopy was used to track motions of green fluorescent protein (GFP)-labeled actin or GFP-labeled secretory granules in a thin layer of cytoplasm where cells adhered to glass. The layer contained abundant filamentous actin (F-actin) locally condensed into stress fibers. More than 90% of the granules imaged lay within the F-actin layer. One-third of the granules did not move detectably, while two-thirds moved randomly; the average diffusion coefficient was 23 x 10(-4) microm(2)/s. A small minority (<3%) moved rapidly and in a directed fashion over distances more than a micron. Staining of F-actin suggests that such movement occurred along actin bundles. The seemingly random movement of most other granules was not due to diffusion since it was diminished by the myosin inhibitor butanedione monoxime, and blocked by chelating intracellular Mg(2+) and replacing ATP with AMP-PNP. Mobility was blocked also when F-actin was stabilized with phalloidin, and was diminished when the actin cortex was degraded with latrunculin B. We conclude that the movement of granules requires metabolic energy, and that it is mediated as well as limited by the actin cortex. Opposing actions of the actin cortex on mobility may explain why its degradation has variable effects on secretion. PMID:10827968

  5. Integrity of the actin cytoskeleton of host macrophages is essential for Leishmania donovani infection.

    PubMed

    Roy, Saptarshi; Kumar, G Aditya; Jafurulla, Md; Mandal, Chitra; Chattopadhyay, Amitabha

    2014-08-01

    Visceral leishmaniasis is a vector-borne disease caused by an obligate intracellular protozoan parasite Leishmania donovani. The molecular mechanism involved in internalization of Leishmania is poorly understood. The entry of Leishmania involves interaction with the plasma membrane of host cells. We have previously demonstrated the requirement of host membrane cholesterol in the binding and internalization of L. donovani into macrophages. In the present work, we explored the role of the host actin cytoskeleton in leishmanial infection. We observed a dose-dependent reduction in the attachment of Leishmania promastigotes to host macrophages upon destabilization of the actin cytoskeleton by cytochalasin D. This is accompanied by a concomitant reduction in the intracellular amastigote load. We utilized a recently developed high resolution microscopy-based method to quantitate cellular F-actin content upon treatment with cytochalasin D. A striking feature of our results is that binding of Leishmania promastigotes and intracellular amastigote load show close correlation with cellular F-actin level. Importantly, the binding of Escherichia coli remained invariant upon actin destabilization of host cells, thereby implying specific involvement of the actin cytoskeleton in Leishmania infection. To the best of our knowledge, these novel results constitute the first comprehensive demonstration on the specific role of the host actin cytoskeleton in Leishmania infection. Our results could be significant in developing future therapeutic strategies to tackle leishmaniasis.

  6. Actin Filaments Play a Critical Role in Vacuolar Trafficking at the Golgi Complex in Plant Cells

    PubMed Central

    Kim, Hyeran; Park, Misoon; Kim, Soo Jin; Hwang, Inhwan

    2005-01-01

    Actin filaments are thought to play an important role in intracellular trafficking in various eukaryotic cells. However, their involvement in intracellular trafficking in plant cells has not been clearly demonstrated. Here, we investigated the roles actin filaments play in intracellular trafficking in plant cells using latrunculin B (Lat B), an inhibitor of actin filament assembly, or actin mutants that disrupt actin filaments when overexpressed. Lat B and actin2 mutant overexpression inhibited the trafficking of two vacuolar reporter proteins, sporamin:green fluorescent protein (GFP) and Arabidopsis thaliana aleurain-like protein:GFP, to the central vacuole; instead, a punctate staining pattern was observed. Colocalization experiments with various marker proteins indicated that these punctate stains corresponded to the Golgi complex. The A. thaliana vacuolar sorting receptor VSR-At, which mainly localizes to the prevacuolar compartment, also accumulated at the Golgi complex in the presence of Lat B. However, Lat B had no effect on the endoplasmic reticulum (ER) to Golgi trafficking of sialyltransferase or retrograde Golgi to ER trafficking. Lat B also failed to influence the Golgi to plasma membrane trafficking of H+-ATPase:GFP or the secretion of invertase:GFP. Based on these observations, we propose that actin filaments play a critical role in the trafficking of proteins from the Golgi complex to the central vacuole. PMID:15722471

  7. Separation of Particle and Energy Transport in the H- and QH-mode Pedestal

    NASA Astrophysics Data System (ADS)

    Battaglia, D. J.; Chang, C. S.; Diallo, A.; Grierson, B. A.; Burrell, K. H.; Groebner, R. J.

    2014-10-01

    Net particle transport through the H-mode pedestal is dictated by anomalous transport mechanisms; however, a significant fraction of the energy transport is governed by enhanced transport of high-energy ions on collisionless orbits. The pedestal radial electric field (Er) is constrained to the value that balances this ion flux with a pinch of colder main ions and impurities as demonstrated using XGC0, a self-consistent full-f multi-species neoclassical calculation that includes neutral recycling and transport. These calculations resolve how edge modes can increase the anomalous particle transport with only a small effect on energy transport, the observed scaling of the height of the density pedestal with Ip, and the structure of Er in the pedestal. Quantitative agreement between XGC0 and the unique features of QH-mode, such as Ti anisotropy, large scrape-off layer Ti and intrinsic co-Ip edge rotation provide confidence that the simulation captures the kinetic effects in the pedestal that drive the neoclassical energy transport. Work supported in part by the US DOE under DE-AC02-09CH11466 and DE-FC02-04ER54698.

  8. Anomalous transport in the H-mode pedestal of Alcator C-Mod discharges

    NASA Astrophysics Data System (ADS)

    Pankin, A. Y.; Hughes, J. W.; Greenwald, M. J.; Kritz, A. H.; Rafiq, T.

    2017-02-01

    Anomalous transport in the H-mode pedestal region of five Alcator C-Mod discharges, representing a collisionality scan is analyzed. The understanding of anomalous transport in the pedestal region is important for the development of a comprehensive model for the H-mode pedestal slope. In this research, a possible role of the drift resistive inertial ballooning modes (Rafiq et al 2010 Phys. Plasmas 17 082511) in the edge of Alcator C-Mod discharges is analyzed. The stability analysis, carried out using the TRANSP code, indicates that the DRIBM modes are strongly unstable in Alcator C-Mod discharges with large electron collisionality. An improved interpretive analysis of H-mode pedestal experimental data is carried out utilizing the additive flux minimization technique (Pankin et al 2013 Phys. Plasmas 20 102501) together with the guiding-center neoclassical kinetic XGC0 code. The neoclassical and neutral physics are simulated in the XGC0 code and the anomalous fluxes are computed using the additive flux minimization technique. The anomalous fluxes are reconstructed and compared with each other for the collisionality scan Alcator C-Mod discharges. It is found that the electron thermal anomalous diffusivities at the pedestal top increase with the electron collisionality. This dependence can also point to the drift resistive inertial ballooning modes as the modes that drive the anomalous transport in the plasma edge of highly collisional discharges.

  9. Multi-device studies of pedestal physics and confinement in the I-mode regime

    DOE PAGES

    Hubbard, A. E.; Osborne, T.; Ryter, F.; ...

    2016-07-05

    This paper describes joint ITPA studies of the I-mode regime, which features an edge thermal barrier together with L-mode-like particle and impurity transport and no Edge Localized Modes (ELMs). The regime has been demonstrated on the Alcator C-Mod, ASDEX Upgrade and DIII-D tokamaks, over a wide range of device parameters and pedestal conditions. Dimensionless parameters at the pedestal show overlap across devices and extend to low collisionality. When they are matched, pedestal temperature profiles are also similar. Pedestals are stable to peeling ballooning modes, consistent with lack of ELMs. Access to Imode is independent of heating method (neutral beam injection,more » Ion Cyclotron and/or Electron Cyclotron Resonance Heating). Normalized energy confinement H98,y2 ≥ 1 has been achieved for a range of 3 ≤ q95 ≤ 4.9 and scales favourably with power. Changes in turbulence in the pedestal region accompany the transition from L-mode to I-mode. The L-I threshold increases with plasma density and current, and with device size, but has a weak dependence on toroidal magnetic field BT. The upper limit of power for I-modes, which is set by I-H transitions, increases with BT and the power range is largest on Alcator C-Mod at B > 5 T. Finally, issues for extrapolation to ITER and other future fusion devices are discussed.« less

  10. Multi-device studies of pedestal physics and confinement in the I-mode regime

    SciTech Connect

    Hubbard, A. E.; Osborne, T.; Ryter, F.; Austin, M.; Barrera Orte, L.; Churchill, R. M.; Cziegler, I.; Fenstermacher, M.; Fischer, R.; Gerhardt, S.; Groebner, R.; Gohil, P.; Happel, T.; Hughes, J. W.; Loarte, A.; Maingi, R.; Manz, P.; Marinoni, A.; Marmar, E. S.; McDermott, R. M.; McKee, G.; Rhodes, T. L.; Rice, J. E.; Schmitz, L.; Theiler, C.; Viezzer, E.; Walk, J. R.; White, A.; Whyte, D.; Wolfe, S.; Wolfrum, E.; Yan, Z.

    2016-07-05

    This paper describes joint ITPA studies of the I-mode regime, which features an edge thermal barrier together with L-mode-like particle and impurity transport and no Edge Localized Modes (ELMs). The regime has been demonstrated on the Alcator C-Mod, ASDEX Upgrade and DIII-D tokamaks, over a wide range of device parameters and pedestal conditions. Dimensionless parameters at the pedestal show overlap across devices and extend to low collisionality. When they are matched, pedestal temperature profiles are also similar. Pedestals are stable to peeling ballooning modes, consistent with lack of ELMs. Access to Imode is independent of heating method (neutral beam injection, Ion Cyclotron and/or Electron Cyclotron Resonance Heating). Normalized energy confinement H98,y2 ≥ 1 has been achieved for a range of 3 ≤ q95 ≤ 4.9 and scales favourably with power. Changes in turbulence in the pedestal region accompany the transition from L-mode to I-mode. The L-I threshold increases with plasma density and current, and with device size, but has a weak dependence on toroidal magnetic field BT. The upper limit of power for I-modes, which is set by I-H transitions, increases with BT and the power range is largest on Alcator C-Mod at B > 5 T. Finally, issues for extrapolation to ITER and other future fusion devices are discussed.

  11. Optimization of CFETR baseline performance by controlling rotation shear and pedestal collisionality through integrated modeling

    NASA Astrophysics Data System (ADS)

    Jian, Xiang; Chen, Jiale; Chan, Vincent S.; Zhuang, Ge; Li, Guoqiang; Deng, Zhao; Shi, Nan; Xu, Guoliang; Staebler, Gary M.; Guo, Wenfeng

    2017-04-01

    The optimization of a CFETR baseline scenario (Chan et al 2015 Nucl. Fusion 55 023017) with an electron cyclotron (EC) wave and neutral beam (NB) is performed using a multi-dimensional code suite. TGLF and NEO are used to calculate turbulent and neoclassical transport. The evaluation of sources and sinks, as well as the current evolution, are performed using ONETWO, and the equilibrium is updated using EFIT. The pedestal is consistent with the EPED model. Rotation shear is controlled using NB. It has been found that both fusion gain Q and NB power deposited in the edge increase with decreasing NB energy, with NB providing current drive, torque, energy and particle source simultaneously. By using an optimized combination of two NBs, Q can be kept at a high level while the NB edge power is reduced. Pedestal collisionality is controlled to find an optimization path for Q by trading off between the pedestal density and temperature with the pedestal pressure fixed. It has been found that Q increases with pedestal collisionality, while the density peaking factor (DPF) remains almost unchanged. The invariance of DPF can be explained by the change of the dominant type of turbulence from the core to the edge (i.e. trapped electron mode in the core and ion temperature gradient mode at the edge), and collisionality has the opposite effect on particle transport for these two modes. A weaker dependence of DPF on collisionality makes a higher density operation more favorable for fusion gain.

  12. Flux Surface Variation of Impurity Density and Flows in the Pedestal Region

    NASA Astrophysics Data System (ADS)

    Churchill, Michael; Lipschultz, Bruce; Theiler, Christian; Alcator C-Mod Team

    2013-10-01

    Measured impurity density and flows in the pedestal region of Alcator C-Mod can deviate significantly on a flux surface from current model predictions. Comparing localized measurements at the low-field side (LFS) midplane and the high-field side (HFS) midplane, boron (B5 +) impurity density asymmetries larger than 10x are observed in H-mode plasmas, with larger densities at the HFS. The LFS density pedestal varies in position and width with varying plasma conditions, while the HFS impurity density profile remains rather fixed. Impurity density asymmetries are not observed in plasmas with small gradients, i.e L-mode, suggesting the drive for the asymmetry may be the strong gradients in the H-mode pedestal region. However, impurity density asymmetries are also absent in I-mode plasmas, despite the presence of a strong radial gradient in temperature (with no main ion density pedestal). This indicates an interplay between the gradient scale lengths of the main ion density and temperature in the drive of the impurity density asymmetry. Impurity flows in the pedestal show the opposite behavior; flows measured in H-mode plasmas are close to the expected in-out variation, while in I-mode they deviate significantly. Supported by USDoE award DE-FC02-99ER54512.

  13. Impact of the Pedestal on Global Performance and Confinement Scalings in I-mode

    NASA Astrophysics Data System (ADS)

    Walk, John; Hughes, Jerry; Hubbard, Amanda; Whyte, Dennis; White, Anne; Alcator C-Mod Team

    2015-11-01

    The I-mode is a novel high-confinement regime pioneered on Alcator C-Mod, notable for its strong temperature pedestal without the accompanying density pedestal found in conventional H-modes. This separation in transport channels gives the desired improved energy confinement while maintaining low particle confinement, avoiding excessive impurity accumulation. Moreover, I-mode operation is naturally free of deleterious Edge-Localized Modes (ELMs). Recent experiments on Alcator C-Mod have characterized the pedestal structure in I-mode. The impact of the pedestal response (particularly to fueling and heating power) and core profile stiffness on global performance and confinement have demonstrated confinement metrics competitive with H-mode operation on Alcator C-Mod, and consistent with concepts for I-mode access & operation on ITER. Following the practice of the ITER89 and ITER98 scaling laws for L- and H-mode energy confinement, an initial, illustrative attempt at an I-mode confinement scaling has also been developed. The initial characterization from C-Mod data is consistent with the observed pedestal properties in I-mode, particularly the weak degradation of energy confinement with heating power, and comparatively strong positive response to fueling and increased magnetic field. Supported by U.S. Department of Energy award DE-FC02-99ER54512, using Alcator C-Mod, a DOE Office of Science User Facility.

  14. Neoclassical ion heat flux and poloidal flow in a tokamak pedestal

    NASA Astrophysics Data System (ADS)

    Kagan, Grigory; Catto, Peter J.

    2009-11-01

    In the core of a tokamak, neoclassical transport normally dominates over classical while itself being dominated by turbulent transport. The situation may be different in a high confinement (or H) mode pedestal, where the latter is effectively suppressed by a strongly sheared equilibrium electric field. On the other hand, this very field makes conventional neoclassical results inapplicable in the pedestal by significantly modifying ion drift orbits. We present the first calculation of the banana regime neoclassical ion heat flux and poloidal flow in the pedestal accounting for the strong ExB drift inherent to this tokamak region. Interestingly, the fact that ion heat conductivity depends on the local values of the electric field and its shear allows us to hypothesize about possible shapes of the global electric field and density profiles in the pedestal. We also find that due to the electric field the pedestal poloidal ion flow is likely to change its direction as compared to its core counterpart. This result elucidates the discrepancy between the conventional banana regime predictions and recent experimental measurements of the impurity flow performed at Alcator C-Mod.

  15. Transport in the plateau regime in a tokamak pedestal

    SciTech Connect

    Seol, J.; Shaing, K. C.

    2012-07-15

    In a tokamak H-mode, a strong E Multiplication-Sign B flow shear is generated during the L-H transition. Turbulence in a pedestal is suppressed significantly by this E Multiplication-Sign B flow shear. In this case, neoclassical transport may become important. The neoclassical fluxes are calculated in the plateau regime with the parallel plasma flow using their kinetic definitions. In an axisymmetric tokamak, the neoclassical particles fluxes can be decomposed into the banana-plateau flux and the Pfirsch-Schlueter flux. The banana-plateau particle flux is driven by the parallel viscous force and the Pfirsch-Schlueter flux by the poloidal variation of the friction force. The combined quantity of the radial electric field and the parallel flow is determined by the flux surface averaged parallel momentum balance equation rather than requiring the ambipolarity of the total particle fluxes. In this process, the Pfirsch-Schlueter flux does not appear in the flux surface averaged parallel momentum equation. Only the banana-plateau flux is used to determine the parallel flow in the form of the flux surface averaged parallel viscosity. The heat flux, obtained using the solution of the parallel momentum balance equation, decreases exponentially in the presence of sonic M{sub p} without any enhancement over that in the standard neoclassical theory. Here, M{sub p} is a combination of the poloidal E Multiplication-Sign B flow and the parallel mass flow. The neoclassical bootstrap current in the plateau regime is presented. It indicates that the neoclassical bootstrap current also is related only to the banana-plateau fluxes. Finally, transport fluxes are calculated when M{sub p} is large enough to make the parallel electron viscosity comparable with the parallel ion viscosity. It is found that the bootstrap current has a finite value regardless of the magnitude of M{sub p}.

  16. Neural Wiskott-Aldrich syndrome protein is implicated in the actin-based motility of Shigella flexneri.

    PubMed Central

    Suzuki, T; Miki, H; Takenawa, T; Sasakawa, C

    1998-01-01

    Shigella, the causative agent of bacillary dysentery, is capable of directing its own movement in the cytoplasm of infected epithelial cells. The bacterial surface protein VirG recruits host components mediating actin polymerization, which is thought to serve as the propulsive force. Here, we show that neural Wiskott-Aldrich syndrome protein (N-WASP), which is a critical target for filopodium formation downstream of Cdc42, is required for assembly of the actin tail generated by intracellular S.flexneri. N-WASP accumulates at the front of the actin tail and is capable of interacting with VirG in vitro and in vivo, a phenomenon that is not observed in intracellular Listeria monocytogenes. The verprolin-homology region in N-WASP was required for binding to the glycine-rich repeats domain of VirG, an essential domain for recruitment of F-actin on intracellular S.flexneri. Overexpression of a dominant-negative N-WASP mutant greatly inhibited formation of the actin tail by intracellular S.flexneri. Furthermore, depletion of N-WASP from Xenopus egg extracts shut off Shigella actin tail assembly, and this was restored upon addition of N-WASP protein, suggesting that N-WASP is a critical host factor for the assembly of the actin tail by intracellular Shigella. PMID:9582270

  17. On the recovery of pedestal temperature of JET-ILW plasmas with injection of low-Z impurities

    NASA Astrophysics Data System (ADS)

    Giroud, Carine

    2016-10-01

    The pedestal confinement has significantly decreased in JET with its metallic ITER-like wall with reference to the carbon wall phase of JET (JET-C). A reduction in pedestal temperature is observed in all scenarios regardless of the level of D-gas injection or value of βN. In particular, the JET-ILW 2.5MA/2.7T high- δ (δ = 0.4) plasmas at nped/nGW >= 0.7, discharges most comparable with JET-C, the pedestal pressure has reduced by 40% with a decrease in pedestal temperature from 0.9keV to 0.5keV with the change of wall. The pedestal stability has been modified with the new wall: the reference JET-C plasmas pedestals had an operational point in the corner of the Peeling-Ballooning (PB) diagram, with pressure limited by intermediate n-numbers (n =5-20), whereas the JET-ILW unseeded plasmas have a lower pressure gradient limited by high n-numbers >=70 (ballooning modes). Seeding N, a low-Z impurity, almost recovers the thermal stored energy, pedestal pressure and pedestal temperature to JET-C levels and with an operation point in the corner of the PB diagram. The mechanisms linked to the pedestal recovery with N are likely related to the mechanisms leading to a decrease in pedestal temperature in the absence of C in the plasma composition. The improved pedestal stability with N is not solely linked to the ideal linear PB stability since N-seeded plasmas in JET-ILW can be in type-III ELM regime and have a higher pedestal pressure than unseeded type-I ELMy H-mode. An increased pedestal pressure via an inward movement of the pedestal pressure from the separatrix is not observed with N seeding. However, we have identified two mechanisms responsible. A first initial mechanism linked to the change in ELM energy losses which raises modestly the average global beta by 10% but allows in return a second mechanism to take place. The considered high- δ plasmas can then benefit, if in type-I ELM regime, from the virtuous cycle (2nd mechanisms) of an increased Shafranov shift

  18. Graphene Oxide Nanosheets Retard Cellular Migration via Disruption of Actin Cytoskeleton.

    PubMed

    Tian, Xin; Yang, Zaixing; Duan, Guangxin; Wu, Anqing; Gu, Zonglin; Zhang, Leili; Chen, Chunying; Chai, Zhifang; Ge, Cuicui; Zhou, Ruhong

    2017-01-01

    Graphene and graphene-based nanomaterials are broadly used for various biomedical applications due to their unique physiochemical properties. However, how graphene-based nanomaterials interact with biological systems has not been thoroughly studied. This study shows that graphene oxide (GO) nanosheets retard A549 lung carcinoma cell migration through nanosheet-mediated disruption of intracellular actin filaments. After GO nanosheets treatment, A549 cells display slower migration and the structure of the intracellular actin filaments is dramatically changed. It is found that GO nanosheets are capable of absorbing large amount of actin and changing the secondary structures of actin monomers. Large-scale all-atom molecular dynamics simulations further reveal the interactions between GO nanosheets and actin filaments at molecular details. GO nanosheets can insert into the interstrand gap of actin tetramer (helical repeating unit of actin filament) and cause the separation of the tetramer which eventually leads to the disruption of actin filaments. These findings offer a novel mechanism of GO nanosheet induced biophysical responses and provide more insights into their potential for biomedical applications.

  19. Integrated fusion simulation with self-consistent core-pedestal coupling

    SciTech Connect

    Meneghini, O.; Snyder, P. B.; Smith, S. P.; Candy, J.; Staebler, G. M.; Belli, E. A.; Lao, L. L.; Park, J. M.; Green, D. L.; Elwasif, W.; Grierson, B. A.; Holland, C.

    2016-04-20

    In this study, accurate prediction of fusion performance in present and future tokamaks requires taking into account the strong interplay between core transport, pedestal structure, current profile and plasma equilibrium. An integrated modeling workflow capable of calculating the steady-state self- consistent solution to this strongly-coupled problem has been developed. The workflow leverages state-of-the-art components for collisional and turbulent core transport, equilibrium and pedestal stability. Validation against DIII-D discharges shows that the workflow is capable of robustly pre- dicting the kinetic profiles (electron and ion temperature and electron density) from the axis to the separatrix in good agreement with the experiments. An example application is presented, showing self-consistent optimization for the fusion performance of the 15 MA D-T ITER baseline scenario as functions of the pedestal density and ion effective charge Zeff.

  20. Integrated fusion simulation with self-consistent core-pedestal coupling

    DOE PAGES

    Meneghini, O.; Snyder, P. B.; Smith, S. P.; ...

    2016-04-20

    In this study, accurate prediction of fusion performance in present and future tokamaks requires taking into account the strong interplay between core transport, pedestal structure, current profile and plasma equilibrium. An integrated modeling workflow capable of calculating the steady-state self- consistent solution to this strongly-coupled problem has been developed. The workflow leverages state-of-the-art components for collisional and turbulent core transport, equilibrium and pedestal stability. Validation against DIII-D discharges shows that the workflow is capable of robustly pre- dicting the kinetic profiles (electron and ion temperature and electron density) from the axis to the separatrix in good agreement with the experiments.more » An example application is presented, showing self-consistent optimization for the fusion performance of the 15 MA D-T ITER baseline scenario as functions of the pedestal density and ion effective charge Zeff.« less

  1. Optimization of CFETR confinement by controlling rotation shear and pedestal collisionality

    NASA Astrophysics Data System (ADS)

    Jian, Xiang; Chen, Jiale; Chan, Vincent; Li, Guoqiang; Zhuang, Ge; Cfetr Physics Team

    2016-10-01

    Optimization of a CFETR baseline scenario (Chan et al. 2015 Nucl. Fusion. 55) with EC and NB H&CD is performed using a multi-dimensional code suite. Rotation shear is controlled using NB, with injection angle being constrained to avoid edge heating and to maintain q_min >2. The NB power is adjusted to keep the plasma fully non-inductive. The NB energy that maximize the fusion gain Q is identified. Trade-off between the pedestal density and temperature is performed with the pedestal pressure fixed. It is found that Q increases with pedestal density, while the density peaking factor (DPF) remains unchanged. Linear analysis shows that the transport is dominated by TEM and ITG turbulence. Collisionality affects these turbulences in such a way that the induced changes in DPF cancel out. A weaker dependence of DPF makes higher density operation more favorable for fusion gain.

  2. Integrated fusion simulation with self-consistent core-pedestal coupling

    NASA Astrophysics Data System (ADS)

    Meneghini, O.; Snyder, P. B.; Smith, S. P.; Candy, J.; Staebler, G. M.; Belli, E. A.; Lao, L. L.; Park, J. M.; Green, D. L.; Elwasif, W.; Grierson, B. A.; Holland, C.

    2016-04-01

    Accurate prediction of fusion performance in present and future tokamaks requires taking into account the strong interplay between core transport, pedestal structure, current profile, and plasma equilibrium. An integrated modeling workflow capable of calculating the steady-state self-consistent solution to this strongly coupled problem has been developed. The workflow leverages state-of-the-art components for collisional and turbulent core transport, equilibrium and pedestal stability. Testing against a DIII-D discharge shows that the workflow is capable of robustly predicting the kinetic profiles (electron and ion temperature and electron density) from the axis to the separatrix in a good agreement with the experiments. An example application is presented, showing self-consistent optimization for the fusion performance of the 15 MA D-T ITER baseline scenario as functions of the pedestal density and ion effective charge Zeff .

  3. Integrated fusion simulation with self-consistent core-pedestal coupling

    SciTech Connect

    Meneghini, O.; Snyder, P. B.; Smith, S. P.; Candy, J.; Staebler, G. M.; Belli, E. A.; Lao, L. L.; Park, J. M.; Green, D. L.; Elwasif, W.; Grierson, B. A.; Holland, C.

    2016-04-20

    In this study, accurate prediction of fusion performance in present and future tokamaks requires taking into account the strong interplay between core transport, pedestal structure, current profile and plasma equilibrium. An integrated modeling workflow capable of calculating the steady-state self- consistent solution to this strongly-coupled problem has been developed. The workflow leverages state-of-the-art components for collisional and turbulent core transport, equilibrium and pedestal stability. Validation against DIII-D discharges shows that the workflow is capable of robustly pre- dicting the kinetic profiles (electron and ion temperature and electron density) from the axis to the separatrix in good agreement with the experiments. An example application is presented, showing self-consistent optimization for the fusion performance of the 15 MA D-T ITER baseline scenario as functions of the pedestal density and ion effective charge Zeff.

  4. An Understanding of H-Mode Pedestal Instabilities in the DIII-D Tokamak

    SciTech Connect

    Ferron, J.R.; Snyder, P.B.

    2005-10-15

    The experimental and modeling results on H-mode edge-localized mode (ELM) instabilities from the DIII-D tokamak project are reviewed. This work has led to the conclusion that the most common type of ELM, called Type I, is triggered by a coupled peeling-ballooning instability driven by the pressure gradient and current density in the H-mode edge pedestal region. Good agreement is found between theoretically predicted stability boundaries and toroidal mode numbers for this instability and experimental observations of edge pedestal parameters and ELM amplitude and frequency as a function of discharge shape and edge-region collisionality. The range of toroidal mode numbers for which there is access to a second stability regime is shown to play an important role. This model of H-mode edge stability has been used to predict the pedestal parameters for ITER and FIRE.

  5. Calcium channel and glutamate receptor activities regulate actin organization in salamander retinal neurons

    PubMed Central

    Cristofanilli, Massimiliano; Akopian, Abram

    2006-01-01

    Intracellular Ca2+ regulates a variety of neuronal functions, including neurotransmitter release, protein phosphorylation, gene expression and synaptic plasticity. In a variety of cell types, including neurons, Ca2+ is involved in actin reorganization, resulting in either actin polymerization or depolymerization. Very little, however, is known about the relationship between Ca2+ and the actin cytoskeleton organization in retinal neurons. We studied the effect of high-K+-induced depolarization on F-actin organization in salamander retina and found that Ca2+ influx through voltage-gated L-type channels causes F-actin disruption, as assessed by 53 ± 5% (n = 23, P < 0.001) reduction in the intensity of staining with Alexa-Fluor488-phalloidin, a compound that permits visualization and quantification of polymerized actin. Calcium-induced F-actin depolymerization was attenuated in the presence of protein kinase C antagonists, chelerythrine or bis-indolylmaleimide hydrochloride (GF 109203X). In addition, phorbol 12-myristate 13-acetate (PMA), but not 4α-PMA, mimicked the effect of Ca2+ influx on F-actin. Activation of ionotropic AMPA and NMDA glutamate receptors also caused a reduction in F-actin. No effect on F-actin was exerted by caffeine or thapsigargin, agents that stimulate Ca2+ release from internal stores. In whole-cell recording from a slice preparation, light-evoked ‘off’ but not ‘on’ EPSCs in ‘on–off’ ganglion cells were reduced by 60 ± 8% (n = 8, P < 0.01) by cytochalasin D. These data suggest that elevation of intracellular Ca2+ during excitatory synaptic activity initiates a cascade for activity-dependent actin remodelling, which in turn may serve as a feedback mechanism to attenuate excitotoxic Ca2+ accumulation induced by synaptic depolarization. PMID:16777935

  6. The role of actin networks in cellular mechanosensing

    NASA Astrophysics Data System (ADS)

    Azatov, Mikheil

    behavior as in cancer metastasis. In addition to stiffness, the local geometry or topography of the surface has been shown to modulate the movement, morphology, and cytoskeletal organization of cells. However, the effect of topography on fluctuations of intracellular structures, which arise from motor driven activity on a viscoelastic actin network are not known. I have used nanofabricated substrates with parallel ridges to show that the cell shape, the actin cytoskeleton and focal adhesions all align along the direction of the ridges, exhibiting a biphasic dependence on the spacing between ridges. I further demonstrated that palladin bands along actin stress fibers undergo a complex diffusive motion with velocities aligned along the direction of ridges. These results provide insight into the mechanisms of cellular mechanosensing of the environment, suggesting a complex interplay between the actin cytoskeleton and cellular adhesions in coordinating cellular response to surface topography. Overall, this work has advanced our understanding of mechanisms that govern cellular responses to their physical environment.

  7. Computational Tension Mapping of Adherent Cells Based on Actin Imaging

    PubMed Central

    Manifacier, Ian; Milan, Jean-Louis; Jeanneau, Charlotte; Chmilewsky, Fanny; Chabrand, Patrick; About, Imad

    2016-01-01

    Forces transiting through the cytoskeleton are known to play a role in adherent cell activity. Up to now few approaches haves been able to determine theses intracellular forces. We thus developed a computational mechanical model based on a reconstruction of the cytoskeleton of an adherent cell from fluorescence staining of the actin network and focal adhesions (FA). Our custom made algorithm converted the 2D image of an actin network into a map of contractile interactions inside a 2D node grid, each node representing a group of pixels. We assumed that actin filaments observed under fluorescence microscopy, appear brighter when thicker, we thus presumed that nodes corresponding to pixels with higher actin density were linked by stiffer interactions. This enabled us to create a system of heterogeneous interactions which represent the spatial organization of the contractile actin network. The contractility of this interaction system was then adapted to match the level of force the cell truly exerted on focal adhesions; forces on focal adhesions were estimated from their vinculin expressed size. This enabled the model to compute consistent mechanical forces transiting throughout the cell. After computation, we applied a graphical approach on the original actin image, which enabled us to calculate tension forces throughout the cell, or in a particular region or even in single stress fibers. It also enabled us to study different scenarios which may indicate the mechanical role of other cytoskeletal components such as microtubules. For instance, our results stated that the ratio between intra and extra cellular compression is inversely proportional to intracellular tension. PMID:26812601

  8. Computational Tension Mapping of Adherent Cells Based on Actin Imaging.

    PubMed

    Manifacier, Ian; Milan, Jean-Louis; Jeanneau, Charlotte; Chmilewsky, Fanny; Chabrand, Patrick; About, Imad

    2016-01-01

    Forces transiting through the cytoskeleton are known to play a role in adherent cell activity. Up to now few approaches haves been able to determine theses intracellular forces. We thus developed a computational mechanical model based on a reconstruction of the cytoskeleton of an adherent cell from fluorescence staining of the actin network and focal adhesions (FA). Our custom made algorithm converted the 2D image of an actin network into a map of contractile interactions inside a 2D node grid, each node representing a group of pixels. We assumed that actin filaments observed under fluorescence microscopy, appear brighter when thicker, we thus presumed that nodes corresponding to pixels with higher actin density were linked by stiffer interactions. This enabled us to create a system of heterogeneous interactions which represent the spatial organization of the contractile actin network. The contractility of this interaction system was then adapted to match the level of force the cell truly exerted on focal adhesions; forces on focal adhesions were estimated from their vinculin expressed size. This enabled the model to compute consistent mechanical forces transiting throughout the cell. After computation, we applied a graphical approach on the original actin image, which enabled us to calculate tension forces throughout the cell, or in a particular region or even in single stress fibers. It also enabled us to study different scenarios which may indicate the mechanical role of other cytoskeletal components such as microtubules. For instance, our results stated that the ratio between intra and extra cellular compression is inversely proportional to intracellular tension.

  9. Microturbulence in DIII-D tokamak pedestal. III. Effects of collisions

    NASA Astrophysics Data System (ADS)

    Liao, X.; Lin, Z.; Holod, I.; Xiao, Y.; Li, B.; Snyder, P. B.

    2016-12-01

    Gyrokinetic simulations of the H-mode pedestal in DIII-D discharge 145701 find that the kinetic ballooning mode (KBM) is the most unstable mode for low toroidal numbers (n ≤ 25) and that the trapped electron mode (TEM) dominates over the KBM at higher toroidal mode numbers for realistic pressure gradients in the pedestal. Collisions reduce the TEM growth rate but have little effects on the KBM. KBM has the conventional ballooning mode structure peaking at the outer mid-plane, while TEM has an unconventional mode structure peaking at the top and bottom of the poloidal plane.

  10. Retired NASA F-18 being lowered on to pedestal mount at Lancaster California Municipal Baseball Stad

    NASA Technical Reports Server (NTRS)

    1997-01-01

    As news media and city officials watch from the balcony of the city baseball stadium in Lancaster, California, a crane gently positions an F/A-18 Hornet aircraft for mounting on a steel pedestal. The F/A-18 was recently retired by NASA's Dryden Flight Research Center, Edwards, California, after being flown as a safety chase and support aircraft over the past nine years. The aircraft is now mounted nose skyward on the 28-foot-tall pedestal in front of the Lancaster Municipal Stadium, know as 'The Hangar.' The stadium is the home field of the Lancaster Jethawks, a Class-A farm team of the Seattle Mariners.

  11. Microturbulence in DIII-D tokamak pedestal. IV. Electrostatic turbulent transport

    NASA Astrophysics Data System (ADS)

    Liao, X.; Lin, Z.; Holod, I.; Li, B.; Sun, G. Y.

    2016-12-01

    Gyrokinetic simulations of electrostatic microturbulence in the edge plasmas of DIII-D shot 131997 find that the geodesic acoustic mode (GAM) is generated after nonlinear saturation both at the pedestal top and in the peak gradient region, and in turn, regulates the turbulence in both regions. Collisions significantly reduce the GAM amplitude and the associated GAM modulation of the turbulence in both regions. Collisions have little effects on the ion temperature gradient turbulent transport level at the pedestal top. On the other hand, collisional damping of the TEM significantly reduces the turbulent transport level in the peak gradient region.

  12. Using LGI experiments to achieve better understanding of pedestal-edge coupling in NSTX-U

    SciTech Connect

    Wang, Zhehui

    2015-02-23

    PowerPoint presentation. Latest advances in granule or dust injection technologies, fast and high-resolution imaging, together with micro-/nano-structured material fabrication, provide new opportunities to examine plasma-material interaction (PMI) in magnetic fusion environment. Some of our previous work in these areas is summarized. The upcoming LGI experiments in NSTX-U will shed new light on granular matter transport in the pedestal-edge region. In addition to particle control, these results can also be used for code validation and achieving better understanding of pedestal-edge coupling in fusion plasmas in both NSTX-U and others.

  13. Retired NASA F-18 being lowered on to pedestal mount at Lancaster California Municipal Baseball Stad

    NASA Technical Reports Server (NTRS)

    1997-01-01

    As news media and city officials watch from the balcony of the city baseball stadium in Lancaster, California, a crane gently positions an F/A-18 Hornet aircraft for mounting on a steel pedestal. The F/A-18 was recently retired by NASA's Dryden Flight Research Center, Edwards, California, after being flown as a safety chase and support aircraft over the past nine years. The aircraft is now mounted nose skyward on the 28-foot-tall pedestal in front of the Lancaster Municipal Stadium, know as 'The Hangar.' The stadium is the home field of the Lancaster Jethawks, a Class-A farm team of the Seattle Mariners.

  14. Quasi-coherent fluctuations limiting the pedestal growth on Alcator C-Mod: Experiment and modelling

    SciTech Connect

    Diallo, A.; Hughes, J. W.; Baek, S. -G.; LaBombard, Brian; Terry, J.; Cziegler, I.; Hubbard, A.; Davis, E.; Walk, J.; Delgado-Aparicio, L.; Reinke, M. L.; Theiler, C.; Churchhill, R. M.; Edlund, E. M.; Canik, John; Snyder, P.; Greenwald, M.; White, A.

    2015-04-09

    Performance predictions for future fusion devices rely on an accurate model of the pedestal structure. The candidate for predictive pedestal structure is EPED, and it is imperative to test the underlying hypotheses to further gain confidence for ITER projections. Here, we present experimental work testing one of the EPED hypotheses, namely the existence of a soft limit set by microinstabilities such as the kinetic ballooning mode. Furthermore, this work extends recent work on Alactor C-Mod (Diallo et al 2014 Phys. Rev. Lett. 112 115001), to include detailed measurements of the edge fluctuations and comparisons of edge simulation codes and experimental observations.

  15. Design of the exhaust device for light vehicle engine pedestal experiment

    NASA Astrophysics Data System (ADS)

    Sun, Shuguang

    2017-01-01

    In view of the shortcomings and the insufficiency of the existing exhaust device for light vehicle engine pedestal experiment, improvement scheme is proposed to design a suitable multi-type exhaust device for light vehicle engine pedestal experiment, which has flex space and a certain degree of freedom in six directions x, y, z, x, y, z, so the problem of interference during the process of installation can be solved, the cost on research and development and test can be reduced and the development cycle can be shorten and it can also be multi-usage.

  16. Actin-based motility drives baculovirus transit to the nucleus and cell surface.

    PubMed

    Ohkawa, Taro; Volkman, Loy E; Welch, Matthew D

    2010-07-26

    Most viruses move intracellularly to and from their sites of replication using microtubule-based mechanisms. In this study, we show that nucleocapsids of the baculovirus Autographa californica multiple nucleopolyhedrovirus undergo intracellular motility driven by actin polymerization. Motility requires the viral P78/83 capsid protein and the host Arp2/3 complex. Surprisingly, the virus directs two sequential and coordinated phases of actin-based motility. Immediately after cell entry, motility enables exploration of the cytoplasm and collision with the nuclear periphery, speeding nuclear entry and the initiation of viral gene expression. Nuclear entry itself requires transit through nuclear pore complexes. Later, after the onset of early gene expression, motility is required for accumulation of a subpopulation of nucleocapsids in the tips of actin-rich surface spikes. Temporal coordination of actin-based nuclear and surface translocation likely enables rapid transmission to neighboring cells during infection in insects and represents a distinctive evolutionary strategy for overcoming host defenses.

  17. Actin Filaments and Myosin I Alpha Cooperate with Microtubules for the Movement of LysosomesV⃞

    PubMed Central

    Cordonnier, Marie-Neige; Dauzonne, Daniel; Louvard, Daniel; Coudrier, Evelyne

    2001-01-01

    An earlier report suggested that actin and myosin I alpha (MMIα), a myosin associated with endosomes and lysosomes, were involved in the delivery of internalized molecules to lysosomes. To determine whether actin and MMIα were involved in the movement of lysosomes, we analyzed by time-lapse video microscopy the dynamic of lysosomes in living mouse hepatoma cells (BWTG3 cells), producing green fluorescent protein actin or a nonfunctional domain of MMIα. In GFP-actin cells, lysosomes displayed a combination of rapid long-range directional movements dependent on microtubules, short random movements, and pauses, sometimes on actin filaments. We showed that the inhibition of the dynamics of actin filaments by cytochalasin D increased pauses of lysosomes on actin structures, while depolymerization of actin filaments using latrunculin A increased the mobility of lysosomes but impaired the directionality of their long-range movements. The production of a nonfunctional domain of MMIα impaired the intracellular distribution of lysosomes and the directionality of their long-range movements. Altogether, our observations indicate for the first time that both actin filaments and MMIα contribute to the movement of lysosomes in cooperation with microtubules and their associated molecular motors. PMID:11739797

  18. Gelsolin, a protein that caps the barbed ends and severs actin filaments, enhances the actin-based motility of Listeria monocytogenes in host cells.

    PubMed

    Laine, R O; Phaneuf, K L; Cunningham, C C; Kwiatkowski, D; Azuma, T; Southwick, F S

    1998-08-01

    The actin-based motility of Listeria monocytogenes requires the addition of actin monomers to the barbed or plus ends of actin filaments. Immunofluorescence micrographs have demonstrated that gelsolin, a protein that both caps barbed ends and severs actin filaments, is concentrated directly behind motile bacteria at the junction between the actin filament rocket tail and the bacterium. In contrast, CapG, a protein that strictly caps actin filaments, fails to localize near intracellular Listeria. To explore the effect of increasing concentrations of gelsolin on bacterial motility, NIH 3T3 fibroblasts stably transfected with gelsolin cDNA were infected with Listeria. The C5 cell line containing 2.25 times control levels of gelsolin supported significantly higher velocities of bacterial movement than did control fibroblasts (mean +/- standard error of the mean, 0.09 +/- 0.003 micro(m)/s [n = 176] versus 0.05 +/- 0.003 micro(m)/s [n = 65]). The rate of disassembly of the Listeria-induced actin filament rocket tail was found to be independent of gelsolin content. Therefore, if increases in gelsolin content result in increases in Listeria-induced rocket tail assembly rates, a positive correlation between gelsolin content and tail length would be expected. BODIPY-phalloidin staining of four different stably transfected NIH 3T3 fibroblast cell lines confirmed this expectation (r = 0.92). Rocket tails were significantly longer in cells with a high gelsolin content. Microinjection of gelsolin 1/2 (consisting of the amino-terminal half of native gelsolin) also increased bacterial velocity by more than 2.2 times. Microinjection of CapG had no effect on bacterial movement. Cultured skin fibroblasts derived from gelsolin-null mice were capable of supporting intracellular Listeria motility at velocities comparable to those supported by wild-type skin fibroblasts. These experiments demonstrated that the surface of Listeria contains a polymerization zone that can block the barbed

  19. Gelsolin, a Protein That Caps the Barbed Ends and Severs Actin Filaments, Enhances the Actin-Based Motility of Listeria monocytogenes in Host Cells

    PubMed Central

    Laine, Roney O.; Phaneuf, Katherine L.; Cunningham, Casey C.; Kwiatkowski, David; Azuma, Toshi; Southwick, Frederick S.

    1998-01-01

    The actin-based motility of Listeria monocytogenes requires the addition of actin monomers to the barbed or plus ends of actin filaments. Immunofluorescence micrographs have demonstrated that gelsolin, a protein that both caps barbed ends and severs actin filaments, is concentrated directly behind motile bacteria at the junction between the actin filament rocket tail and the bacterium. In contrast, CapG, a protein that strictly caps actin filaments, fails to localize near intracellular Listeria. To explore the effect of increasing concentrations of gelsolin on bacterial motility, NIH 3T3 fibroblasts stably transfected with gelsolin cDNA were infected with Listeria. The C5 cell line containing 2.25 times control levels of gelsolin supported significantly higher velocities of bacterial movement than did control fibroblasts (mean ± standard error of the mean, 0.09 ± 0.003 μm/s [n = 176] versus 0.05 ± 0.003 μm/s [n = 65]). The rate of disassembly of the Listeria-induced actin filament rocket tail was found to be independent of gelsolin content. Therefore, if increases in gelsolin content result in increases in Listeria-induced rocket tail assembly rates, a positive correlation between gelsolin content and tail length would be expected. BODIPY-phalloidin staining of four different stably transfected NIH 3T3 fibroblast cell lines confirmed this expectation (r = 0.92). Rocket tails were significantly longer in cells with a high gelsolin content. Microinjection of gelsolin 1/2 (consisting of the amino-terminal half of native gelsolin) also increased bacterial velocity by more than 2.2 times. Microinjection of CapG had no effect on bacterial movement. Cultured skin fibroblasts derived from gelsolin-null mice were capable of supporting intracellular Listeria motility at velocities comparable to those supported by wild-type skin fibroblasts. These experiments demonstrated that the surface of Listeria contains a polymerization zone that can block the barbed

  20. Antenna Mechanism of Length Control of Actin Cables

    PubMed Central

    Mohapatra, Lishibanya; Goode, Bruce L.; Kondev, Jane

    2015-01-01

    Actin cables are linear cytoskeletal structures that serve as tracks for myosin-based intracellular transport of vesicles and organelles in both yeast and mammalian cells. In a yeast cell undergoing budding, cables are in constant dynamic turnover yet some cables grow from the bud neck toward the back of the mother cell until their length roughly equals the diameter of the mother cell. This raises the question: how is the length of these cables controlled? Here we describe a novel molecular mechanism for cable length control inspired by recent experimental observations in cells. This “antenna mechanism” involves three key proteins: formins, which polymerize actin, Smy1 proteins, which bind formins and inhibit actin polymerization, and myosin motors, which deliver Smy1 to formins, leading to a length-dependent actin polymerization rate. We compute the probability distribution of cable lengths as a function of several experimentally tuneable parameters such as the formin-binding affinity of Smy1 and the concentration of myosin motors delivering Smy1. These results provide testable predictions of the antenna mechanism of actin-cable length control. PMID:26107518

  1. Participation of Actin on Giardia lamblia Growth and Encystation

    PubMed Central

    Castillo-Romero, Araceli; Leon-Avila, Gloria; Perez Rangel, Armando; Cortes Zarate, Rafael; Garcia Tovar, Carlos; Hernandez, Jose Manuel

    2009-01-01

    Background Microfilaments play a determinant role in different cell processes such as: motility, cell division, phagocytosis and intracellular transport; however, these structures are poorly understood in the parasite Giardia lamblia. Methodology and Principal Findings By confocal microscopy using TRITC-phalloidin, we found structured actin distributed in the entire trophozoite, the label stand out at the ventral disc, median body, flagella and around the nuclei. During Giardia encystation, a sequence of morphological changes concurrent to modifications on the distribution of structured actin and in the expression of actin mRNA were observed. To elucidate whether actin participates actively on growth and encystation, cells were treated with Cytochalasin D, Latrunculin A and Jasplakinolide and analyzed by confocal and scanning electron microscopy. All drugs caused a growth reduction (27 to 45%) and changes on the distribution of actin. Besides, 60 to 80% of trophozoites treated with the drugs, exhibited damage at the caudal region, alterations in the flagella and wrinkles-like on the plasma membrane. The drugs also altered the cyst-yield and the morphology, scanning electron microscopy revealed diminished cytokinesis, cysts with damages in the wall and alterations in the size and on the intermembranal space. Furthermore, the drugs caused a significant reduction of the intensity of flourescence-labeled CWP1 on ESV and on cyst wall, this was coincident with a reduction of CWP1 gene expression (34%). Conclusions and Significance All our results, indicated an important role of actin in the morphology, growth and encystation and indirectly suggested an actin role in gene expression. PMID:19774081

  2. Ring closure in actin polymers

    NASA Astrophysics Data System (ADS)

    Sinha, Supurna; Chattopadhyay, Sebanti

    2017-03-01

    We present an analysis for the ring closure probability of semiflexible polymers within the pure bend Worm Like Chain (WLC) model. The ring closure probability predicted from our analysis can be tested against fluorescent actin cyclization experiments. We also discuss the effect of ring closure on bend angle fluctuations in actin polymers.

  3. Coupling of the hydration water dynamics and the internal dynamics of actin detected by quasielastic neutron scattering

    SciTech Connect

    Fujiwara, Satoru; Plazanet, Marie; Oda, Toshiro

    2013-02-15

    Highlights: ► Quasielastic neutron scattering spectra of F-actin and G-actin were measured. ► Analysis of the samples in D{sub 2}O and H{sub 2}O provided the spectra of hydration water. ► The first layer hydration water around F-actin is less mobile than around G-actin. ► This difference in hydration water is in concert with the internal dynamics of actin. ► Water outside the first layer behaves bulk-like but influenced by the first layer. -- Abstract: In order to characterize dynamics of water molecules around F-actin and G-actin, quasielastic neutron scattering experiments were performed on powder samples of F-actin and G-actin, hydrated either with D{sub 2}O or H{sub 2}O, at hydration ratios of 0.4 and 1.0. By combined analysis of the quasielastic neutron scattering spectra, the parameter values characterizing the dynamics of the water molecules in the first hydration layer and those of the water molecules outside of the first layer were obtained. The translational diffusion coefficients (D{sub T}) of the hydration water in the first layer were found to be 1.2 × 10{sup −5} cm{sup 2}/s and 1.7 × 10{sup −5} cm{sup 2}/s for F-actin and G-actin, respectively, while that for bulk water was 2.8 × 10{sup −5} cm{sup 2}/s. The residence times were 6.6 ps and 5.0 ps for F-actin and G-actin, respectively, while that for bulk water was 0.62 ps. These differences between F-actin and G-actin, indicating that the hydration water around G-actin is more mobile than that around F-actin, are in concert with the results of the internal dynamics of F-actin and G-actin, showing that G-actin fluctuates more rapidly than F-actin. This implies that the dynamics of the hydration water is coupled to the internal dynamics of the actin molecules. The D{sub T} values of the water molecules outside of the first hydration layer were found to be similar to that of bulk water though the residence times are strongly affected by the first hydration layer. This supports the

  4. A Steric Antagonism of Actin Polymerization by a Salmonella Virulence Protein

    SciTech Connect

    Margarit,S.; Davidson, W.; Frego, L.; Stebbins, F.

    2006-01-01

    Salmonella spp. require the ADP-ribosyltransferase activity of the SpvB protein for intracellular growth and systemic virulence. SpvB covalently modifies actin, causing cytoskeletal disruption and apoptosis. We report here the crystal structure of the catalytic domain of SpvB, and we show by mass spectrometric analysis that SpvB modifies actin at Arg177, inhibiting its ATPase activity. We also describe two crystal structures of SpvB-modified, polymerization-deficient actin. These structures reveal that ADP-ribosylation does not lead to dramatic conformational changes in actin, suggesting a model in which this large family of toxins inhibits actin polymerization primarily through steric disruption of intrafilament contacts.

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

    PubMed

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

    2016-03-29

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

  6. Toxoplasma gondii F-actin forms an extensive filamentous network required for material exchange and parasite maturation

    PubMed Central

    Periz, Javier; Whitelaw, Jamie; Harding, Clare; Gras, Simon; Del Rosario Minina, Mario Igor; Latorre-Barragan, Fernanda; Lemgruber, Leandro; Reimer, Madita Alice; Insall, Robert; Heaslip, Aoife; Meissner, Markus

    2017-01-01

    Apicomplexan actin is important during the parasite's life cycle. Its polymerization kinetics are unusual, permitting only short, unstable F-actin filaments. It has not been possible to study actin in vivo and so its physiological roles have remained obscure, leading to models distinct from conventional actin behaviour. Here a modified version of the commercially available actin-chromobody was tested as a novel tool for visualising F-actin dynamics in Toxoplasma gondii. Cb labels filamentous actin structures within the parasite cytosol and labels an extensive F-actin network that connects parasites within the parasitophorous vacuole and allows vesicles to be exchanged between parasites. In the absence of actin, parasites lack a residual body and inter-parasite connections and grow in an asynchronous and disorganized manner. Collectively, these data identify new roles for actin in the intracellular phase of the parasites lytic cycle and provide a robust new tool for imaging parasitic F-actin dynamics. DOI: http://dx.doi.org/10.7554/eLife.24119.001 PMID:28322189

  7. A prophage-encoded actin-like protein required for efficient viral DNA replication in bacteria

    PubMed Central

    Donovan, Catriona; Heyer, Antonia; Pfeifer, Eugen; Polen, Tino; Wittmann, Anja; Krämer, Reinhard; Frunzke, Julia; Bramkamp, Marc

    2015-01-01

    In host cells, viral replication is localized at specific subcellular sites. Viruses that infect eukaryotic and prokaryotic cells often use host-derived cytoskeletal structures, such as the actin skeleton, for intracellular positioning. Here, we describe that a prophage, CGP3, integrated into the genome of Corynebacterium glutamicum encodes an actin-like protein, AlpC. Biochemical characterization confirms that AlpC is a bona fide actin-like protein and cell biological analysis shows that AlpC forms filamentous structures upon prophage induction. The co-transcribed adaptor protein, AlpA, binds to a consensus sequence in the upstream promoter region of the alpAC operon and also interacts with AlpC, thus connecting circular phage DNA to the actin-like filaments. Transcriptome analysis revealed that alpA and alpC are among the early induced genes upon excision of the CGP3 prophage. Furthermore, qPCR analysis of mutant strains revealed that both AlpA and AlpC are required for efficient phage replication. Altogether, these data emphasize that AlpAC are crucial for the spatio-temporal organization of efficient viral replication. This is remarkably similar to actin-assisted membrane localization of eukaryotic viruses that use the actin cytoskeleton to concentrate virus particles at the egress sites and provides a link of evolutionary conserved interactions between intracellular virus transport and actin. PMID:25916847

  8. A prophage-encoded actin-like protein required for efficient viral DNA replication in bacteria.

    PubMed

    Donovan, Catriona; Heyer, Antonia; Pfeifer, Eugen; Polen, Tino; Wittmann, Anja; Krämer, Reinhard; Frunzke, Julia; Bramkamp, Marc

    2015-05-26

    In host cells, viral replication is localized at specific subcellular sites. Viruses that infect eukaryotic and prokaryotic cells often use host-derived cytoskeletal structures, such as the actin skeleton, for intracellular positioning. Here, we describe that a prophage, CGP3, integrated into the genome of Corynebacterium glutamicum encodes an actin-like protein, AlpC. Biochemical characterization confirms that AlpC is a bona fide actin-like protein and cell biological analysis shows that AlpC forms filamentous structures upon prophage induction. The co-transcribed adaptor protein, AlpA, binds to a consensus sequence in the upstream promoter region of the alpAC operon and also interacts with AlpC, thus connecting circular phage DNA to the actin-like filaments. Transcriptome analysis revealed that alpA and alpC are among the early induced genes upon excision of the CGP3 prophage. Furthermore, qPCR analysis of mutant strains revealed that both AlpA and AlpC are required for efficient phage replication. Altogether, these data emphasize that AlpAC are crucial for the spatio-temporal organization of efficient viral replication. This is remarkably similar to actin-assisted membrane localization of eukaryotic viruses that use the actin cytoskeleton to concentrate virus particles at the egress sites and provides a link of evolutionary conserved interactions between intracellular virus transport and actin. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

  9. Gyrokinetic simulation of I-mode C-Mod pedestal using GENE

    NASA Astrophysics Data System (ADS)

    Liu, Xing; David, H.; Kotschenreuther, M.; Mahajan, S.; Huges, J.; Hubbard, A.; Valanju, P.

    2016-10-01

    Naturally stable to ELMs, and with widths larger than EPED predictions, the I-modes are an excellent laboratory for investigating the role of drift microinstabilities in pedestal formation since I-mode pedestal are not ``limited'' by MHD instabilities-Peeling Ballooning or the Kinetic Ballooning. Because the Weakly Coherent Mode (WCM) is shown to be correlated, primarily, to particle transport, the pedestal heat transport, in some sense, must be controlled by drift-type modes. We present here a study based on gyrokinetic simulations (using GENE) to model heat transport in the I-mode pedestals in C-Mod. Nonlinear ETG simulations, found to be streamer-dominated, can match experimental heat flux with profile adjustment well within experimental error bars. The ETG simulations reveal very notable fine-scale structure (in the parallel direction) of the eigenfunctions in both linear and nonlinear simulations. Simulations, varying impurity level (Zeff) and temperature and density profiles (within experimental error bars), are used to probe the sensitivity of ETG heat transport to the most important input parameters. Efforts to identify an instability corresponding to the WCM will also be discussed. Work supported by USDOE Grant DE-FG02-04ER54742.

  10. The role of the density profile in the ASDEX-Upgrade pedestal structure

    NASA Astrophysics Data System (ADS)

    Dunne, M. G.; Potzel, S.; Reimold, F.; Wischmeier, M.; Wolfrum, E.; Frassinetti, L.; Beurskens, M.; Bilkova, P.; Cavedon, M.; Fischer, R.; Kurzan, B.; Laggner, F. M.; McDermott, R. M.; Tardini, G.; Trier, E.; Viezzer, E.; Willensdorfer, M.; The EUROfusion MST1 Team; The ASDEX-Upgrade Team

    2017-01-01

    Experimental evidence for the impact of a region of high density localised in the high-field side scrape-off layer (the HFSHD) on plasma confinement is shown in various dedicated experiments on ASDEX Upgrade (AUG). Increasing main ion fuelling is shown to increase the separatrix density and shift the density profile outwards. Predictive pedestal modelling of this shift indicates a 25% decrease in the attainable pedestal top pressure, which compares well with experimental observations in the gas scan. Since the HFSHD can be mitigated by applying nitrogen seeding, a combined scan in fuelling rate, heating power, and nitrogen seeding is presented. Significant increases in the achievable pedestal top pressure are observed with seeding, in particular at high heating powers, and are correlated with inward shifted density profiles and a reduction of the HFSHD and separatrix density. Interpretive linear stability analysis also confirms the impact of a radially shifted pressure profile on peeling-ballooning stability, with an inward shift allowing access to higher pressure gradients and pedestal widths.

  11. A Scaling for the H--mode Edge Pedestal in DIII--D

    NASA Astrophysics Data System (ADS)

    Yushmanov, P.; Groebner, R. J.; Lao, L. L.; Carlstrom, T. N.

    1996-11-01

    A database characterizing the edge pedestal of H--mode plasmas in a wide variety of DIII--D operational conditions is being collected and analyzed by regression methods. The edge density and pressure pedestals are described by the amplitude and the width of the high gradient region. The set of regression parameters includes global discharge characteristics such as toroidal magnetic field, plasma current, stored energy, average density, energy flux through the separatrix as well as local values of electron and ion temperatures and plasma density at the inner side of the edge pedestal. The parameters of the magnetic configuration are described by the critical pressure gradient and the width of the second stability region in the vicinity of the separatrix. These parameters are obtained from magnetic and kinetic measurements with the use of the EFIT equilibrium code and the CAMINO stability code. The goal of this work is to develop a scaling for the H--mode edge pedestal in terms of local plasma parameters which may be used for comparison with the data from other tokamaks and for developing ITER confinement models.

  12. Magnetohydrodynamic interference with the edge pedestal motional Stark effect diagnostic on DIII-D

    SciTech Connect

    King, J. D.; Makowski, M. A.; Holcomb, C. T.; Allen, S. L.; Hill, D. N.; Meyer, W. H.; Geer, R.; La Haye, R. J.; Petty, C. C.; Van Zeeland, M. A.; Turco, F.; Rhodes, T. L.; Morse, E. C.

    2011-03-15

    Accurate measurement of internal magnetic field direction using motional Stark effect (MSE) polarimetry in the edge pedestal is desired for nearly all tokamak scenario work. A newly installed 500 kHz 32-channel digitizer on the MSE diagnostic of DIII-D allows full spectral information of the polarimeter signal to be recovered for the first time. Fourier analysis of this data has revealed magnetohydrodynamic (MHD) fluctuations in the plasma edge pedestal at {rho}{>=} 0.92. By correlating edge localized mode fluctuations seen on lock-in amplifier outputs with MSE spectrograms, it has been shown that edge pedestal tearing mode fluctuations cause interference with MSE second harmonic instrument frequencies. This interference results in unrecoverable errors in the real-time polarization angle measurement that are more than an order of magnitude larger than typical polarimeter uncertainties. These errors can cause as much as a 38% difference in local q. By using a redundant measure of the linear polarization found at the fourth harmonic photo-elastic modulator (PEM) frequency, MHD interference can be avoided. However, because of poorer signal-to-noise the fourth harmonic signal computed polarization angle shows no improvement over the MHD polluted second harmonics. MHD interference could be avoided in future edge pedestal tokamak polarimeters by utilizing PEMs with higher fundamental frequencies and a greater separation between their frequencies.

  13. Pedestal and edge electrostatic turbulence characteristics from an XGC1 gyrokinetic simulation

    NASA Astrophysics Data System (ADS)

    Churchill, R. M.; Chang, C. S.; Ku, S.; Dominski, J.

    2017-10-01

    Understanding the multi-scale neoclassical and turbulence physics in the edge region (pedestal + scrape-off layer (SOL)) is required in order to reliably predict performance in future fusion devices. We explore turbulent characteristics in the edge region from a multi-scale neoclassical and turbulent XGC1 gyrokinetic simulation in a DIII-D like tokamak geometry, here excluding neutrals and collisions. For an H-mode type plasma with steep pedestal, it is found that the electron density fluctuations increase towards the separatrix, and stay high well into the SOL, reaching a maximum value of δ {n}e/{\\bar{n}}e˜ 0.18. Blobs are observed, born around the magnetic separatrix surface and propagate radially outward with velocities generally less than 1 km s-1. Strong poloidal motion of the blobs is also present, near 20 km s-1, consistent with E × B rotation. The electron density fluctuations show a negative skewness in the closed field-line pedestal region, consistent with the presence of ‘holes’, followed by a transition to strong positive skewness across the separatrix and into the SOL. These simulations indicate that not only neoclassical phenomena, but also turbulence, including the blob-generation mechanism, can remain important in the steep H-mode pedestal and SOL. Qualitative comparisons will be made to experimental observations.

  14. 49. (no plate) 1,000 poind Mercury Float Pedestal and clock ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    49. (no plate) 1,000 poind Mercury Float Pedestal and clock case sheet no. 1 of 4 drawing # 5765. Approved 2-3-1916. - Block Island Southeast Light, Spring Street & Mohegan Trail at Mohegan Bluffs, New Shoreham, Washington County, RI

  15. SOL effects on the pedestal structure in DIII-D discharges

    NASA Astrophysics Data System (ADS)

    Sontag, A. C.; Chen, X.; Canik, J.; Leonard, A.; Lore, J. D.; Moser, A. L.; Murakami, M.; Park, J. M.; Petty, C.

    2017-07-01

    Analysis with the SOLPS code suite (Schneider R. et al 1992 J. Nucl. Mater. 196 810; Wiesen S. et al 2015 J. Nucl. Mater. 463 480-4 Bonnin X. et al 2016 Plasma Fusion Res. 11 1403102) explains the differences in pedestal structure associated with different ion \

  16. Neoclassical toroidal viscosity torque in tokamak edge pedestal induced by external resonant magnetic perturbation

    NASA Astrophysics Data System (ADS)

    Yan, Xingting; Zhu, Ping; Sun, Youwen

    2017-08-01

    The characteristic profile and magnitude are predicted in theory for the neoclassical toroidal viscosity (NTV) torque induced by the plasma response to the resonant magnetic perturbation (RMP) in a tokamak with an edge pedestal. For a low-β equilibrium, the NTV torque is dominated by the toroidal component with the same dominant toroidal mode number of RMPs. The NTV torque profile is found to be localized, whose peak location is determined by profiles of both the equilibrium temperature (pressure) and the plasma response. In general, the peak of the NTV torque profile is found to track the pedestal position. The magnitude of NTV torque strongly depends on the β value at the top of pedestal, which suggests a more significant role of NTV torque in higher plasma β regimes. For a fixed plasma β, decreasing density hence increasing temperature can also enhance the amplitude of NTV torque due to the reduced collisionality in the 1/ν regime. Based on those findings, we identify the tokamak operation regimes where the significance of NTV torque in the edge pedestal induced by RMP can approach those from other momentum sources such as the neutral beam injections.

  17. H-mode pedestal turbulence in DIII-D and NSTX using BOUT++

    NASA Astrophysics Data System (ADS)

    Xu, X. Q.; Dudson, B. D.; Joseph, I.; Groebner, R. J.; Maingi, R.

    2011-10-01

    In this work, we will report BOUT++ simulations for H-mode pedestal instabilities and turbulent transport. For DIII-D H-mode discharges, the BOUT++ peeling-ballooning ELM model including electron inertia was used to analyze the ideal linear stability and ELM dynamics. The beta scan is carried out from a series of self-consistent MHD equilibria generated from EFIT by varying pressure and/or current. For typical tokamak pedestal plasmas with high temperature and low collisionality, we found that the collisionless ballooning modes driven by electron inertia are unstable in the H-mode pedestal and have a lower beta threshold than ideal peeling-ballooning modes, which are the triggers for Edge Localized Modes. The growth rate of electron inertia ballooning modes is found to increase with the magnitude of the electron skin depth de=c/ωpe. Thus, collisionless (electron inertia) ballooning modes might be responsible for H-mode turbulence transport when the pedestal is stable to peeling-ballooning modes. BOUT++ calculations also show that NSTX Elm stability boundaries are sensitive to flow shear profile. Attempts are underway to calculate nonlinear turbulence and transport in H-mode discharges due to the non-ideal effects. Performed for USDoE by LLNL under Contract DE-AC52-07NA27344.

  18. Neoclassical Toroidal Viscosity Torque Induced by Plasma Response in a Low- β Tokamak with Edge Pedestal

    NASA Astrophysics Data System (ADS)

    Yan, Xingting; Zhu, Ping; Sun, Youwen

    2016-10-01

    The characteristic profile and magnitude are predicted in theory for the neoclassical toroidal viscosity (NTV) torque induced by the plasma response to the resonant magnetic perturbation (RMP) in a tokamak with an edge pedestal, using the newly developed module coupling the NIMROD and the NTVTOK codes. For a low β equilibrium, the NTV torque is mainly induced by the dominant toroidal mode of plasma response. The NTV torque profile is radially localized and peaked, which is determined by profiles of both the equilibrium temperature and the plasma response fields. In general, the peak of NTV torque profile is found to trace the pedestal location. The magnitude of NTV torque is extremely sensitive to the β of pedestal top; for a given plasma response, the peak value of NTV torque can increase by three orders of magnitude, when the pedestal β increases by only one order of magnitude. This suggests a more significant role of NTV torque in higher plasma β regimes. Supported by the National Magnetic Confinement Fusion Program of China under Grant Nos. 2014GB124002 and 2015GB101004, and the 100 Talent Program of the Chinese Academy of Sciences.

  19. Reaction-diffusion waves of actin filament polymerization/depolymerization in Dictyostelium pseudopodium extension and cell locomotion.

    PubMed

    Vicker, M G

    2000-04-14

    Cell surface movements and the intracellular spatial patterns and dynamics of actin filament (F-actin) were investigated in living and formalin-fixed cells of Dictyostelium discoideum by confocal microscopy. Excitation waves of F-actin assembly developed and propagated several micrometers at up to 26 microm/min in cells which had been intracellularly loaded with fluorescently labeled actin monomer. Wave propagation and extinction corresponded with the initiation and attenuation of pseudopodium extension and cell advance, respectively. The identification of chemical waves was supported by the ring, sphere, spiral and scroll wave patterns, which were observed in the extensions of fixed cells stained with phalloidin-rhodamine, and by the similar, asymmetrical [F-actin] distribution in wavefronts in living and fixed cells. These F-actin patterns and dynamics in Dictyostelium provide evidence for a new supramolecular state of actin, which propagates as a self-organized, reaction-diffusion wave of reversible F-actin assembly and affects pseudopodium extension. Actin's properties of oscillation and self-organization might also fundamentally determine the nature of the eukaryotic cell's reactions of adaptation, timing and signal response.

  20. LIM Kinase 1 Modulates Cortical Actin and CXCR4 Cycling and Is Activated by HIV-1 to Initiate Viral Infection*

    PubMed Central

    Vorster, Paul J.; Guo, Jia; Yoder, Alyson; Wang, Weifeng; Zheng, Yanfang; Xu, Xuehua; Yu, Dongyang; Spear, Mark; Wu, Yuntao

    2011-01-01

    Almost all viral pathogens utilize a cytoskeleton for their entry and intracellular transport. In HIV-1 infection, binding of the virus to blood resting CD4 T cells initiates a temporal course of cortical actin polymerization and depolymerization, a process mimicking the chemotactic response initiated from chemokine receptors. The actin depolymerization has been suggested to promote viral intracellular migration through cofilin-mediated actin treadmilling. However, the role of the virus-mediated actin polymerization in HIV infection is unknown, and the signaling molecules involved remain unidentified. Here we describe a pathogenic mechanism for triggering early actin polymerization through HIV-1 envelope-mediated transient activation of the LIM domain kinase (LIMK), a protein that phosphorylates cofilin. We demonstrate that HIV-mediated LIMK activation is through gp120-triggered transient activation of the Rack-PAK-LIMK pathway, and that knockdown of LIMK through siRNA decreases filamentous actin, increases CXCR4 trafficking, and diminishes viral DNA synthesis. These results suggest that HIV-mediated early actin polymerization may directly regulate the CXCR4 receptor during viral entry and is involved in viral DNA synthesis. Furthermore, we also demonstrate that in resting CD4 T cells, actin polymerization can be triggered through transient treatment with a pharmacological agent, okadaic acid, that activates LIMK and promotes HIV latent infection of resting CD4 T cells. Taken together, our results suggest that HIV hijacks LIMK to control the cortical actin dynamics for the initiation of viral infection of CD4 T cells. PMID:21321123

  1. Actin binding proteins and spermiogenesis

    PubMed Central

    Mruk, Dolores D

    2011-01-01

    Drebrin E, an actin-binding protein lacking intrinsic activity in the regulation of actin dynamics (e.g., polymerization, capping, nucleation, branching, cross-linking, bundling and severing), is known to recruit actin regulatory proteins to a specific cellular site. Herein, we critically evaluate recent findings in the field which illustrate that drebrin E works together with two other actin-binding proteins, namely Arp3 (actin-related protein 3, a component of the Arp2/3 complex that simultaneously controls actin nucleation for polymerization and branching of actin filaments) and Eps8 (epidermal growth factor receptor pathway substrate 8 that controls capping of the barbed ends of actin filaments, as well as actin filament bundling) to regulate the homeostasis of F-actin filament bundles at the ectoplasmic specialization (ES), a testis-specific atypical adherens junction (AJ) in the seminiferous epithelium. This is mediated by the strict temporal and spatial expression of these three actin-binding proteins at the apical and basal ES at the Sertoli cell-spermatid (step 8–19) and Sertoli-Sertoli cell interface, respectively, during the seminiferous epithelial cycle of spermatogenesis. In this Commentary, we put forth a possible model by which drebrin E may be acting as a platform upon which proteins (e.g., Arp3) that are needed to alter the conformation of actin filament bundles at the ES can be recruited to the site, thus facilitating changes in cell shape and cell position in the epithelium during spermiogenesis and spermiation. In short, drebrin E may be acting as a “logistic” distribution center to manage different regulatory proteins at the apical ES, thereby regulating the dynamics of actin filament bundles and modulating the plasticity of the apical ES. This would allow adhesion to be altered continuously throughout the epithelial cycle to accommodate spermatid movement in the seminiferous epithelium during spermiogenesis and spermiation. We also

  2. Influence of plasma pedestal profiles on access to ELM-free regimes in ITER

    SciTech Connect

    Medvedev, S. Yu. Ivanov, A. A. Martynov, A. A. Poshekhonov, Yu. Yu.; Konovalov, S. V.; Polevoi, A. R.

    2016-05-15

    The influence of current density and pressure gradient profiles in the pedestal on the access to the regimes free from edge localized modes (ELMs) like quiescent H-mode in ITER is investigated. Using the simulator of MHD modes localized near plasma boundary based on the KINX code, calculations of the ELM stability were performed for the ITER plasma in scenarios 2 and 4 under variations of density and temperature profiles with the self-consistent bootstrap current in the pedestal. Low pressure gradient values at the separatrix, the same position of the density and temperature pedestals and high poloidal beta values facilitate reaching high current density in the pedestal and a potential transition into the regime with saturated large scale kink modes. New version of the localized MHD mode simulator allows one to compute the growth rates of ideal peeling-ballooning modes with different toroidal mode numbers and to determine the stability region taking into account diamagnetic stabilization. The edge stability diagrams computations and sensitivity studies of the stability limits to the value of diamagnetic frequency show that diamagnetic stabilization of the modes with high toroidal mode numbers can help to access the quiescent H-mode even with high plasma density but only with low pressure gradient values at the separatrix. The limiting pressure at the top of the pedestal increases for higher plasma density. With flat density profile the access to the quiescent H-mode is closed even with diamagnetic stabilization taken into account, while toroidal mode numbers of the most unstable peeling-ballooning mode decrease from n = 10−40 to n = 3−20.

  3. Pressure-Gradient-Limiting Instability Dynamics in the H-mode Pedestal on DIII-D

    NASA Astrophysics Data System (ADS)

    Yan, Z.

    2010-11-01

    Detailed 2D measurements of long-wavelength density fluctuations in the pedestal region with beam emission spectroscopy during the inter-ELM phase indicate two distinct bands of fluctuations propagating in opposite poloidal directions in the plasma frame: one lower frequency band (20-150 kHz) advects in the ion-diamagnetic drift direction (ion mode), and a higher frequency band (200-400 kHz) advects in the electron diamagnetic drift direction (electron mode). Interestingly, the mode amplitudes are modulated with the ELM cycle with the ion mode having some features qualitatively similar to those predicted for kinetic ballooning modes (KBM). Experiments have focused on determining the role of current and pressure gradient-driven instabilities in determining the H-mode pedestal structure. Detailed analysis of the temporal evolution reveals complex dynamics. The ion mode amplitude increases rapidly after an ELM and then saturates, consistent with the dynamics of the pedestal electron pressure, while the electron mode is quasi-stationary between ELMs. The decorrelation time of the ion mode is <5,s (τcxcs/a<=1), the radial correlation length is of order 10,ρi and the poloidal wave-number kθρi˜0.1. The mode velocity is comparable to the diamagnetic velocity. In related Quiescent H-mode experiments, pedestals with high electron pressure and high ExB shearing rates exhibit a set of high-frequency coherent modes propagating in the ion diamagnetic direction. These modes also exhibit KBM-like characteristics, but do not develop into fully turbulent structures. Numerical simulations are in progress to help identify the underlying instabilities and nature of these modes, and ultimately help validate nonlinear models of the H-mode pedestal structure.

  4. Multi-device studies of pedestal physics and confinement in the I-mode regime

    NASA Astrophysics Data System (ADS)

    Hubbard, A. E.; Osborne, T.; Ryter, F.; Austin, M.; Barrera Orte, L.; Churchill, R. M.; Cziegler, I.; Fenstermacher, M.; Fischer, R.; Gerhardt, S.; Groebner, R.; Gohil, P.; Happel, T.; Hughes, J. W.; Loarte, A.; Maingi, R.; Manz, P.; Marinoni, A.; Marmar, E. S.; McDermott, R. M.; McKee, G.; Rhodes, T. L.; Rice, J. E.; Schmitz, L.; Theiler, C.; Viezzer, E.; Walk, J. R.; White, A.; Whyte, D.; Wolfe, S.; Wolfrum, E.; Yan, Z.; Alcator C-Mod, the; Upgrade, ASDEX; DIII-D Teams

    2016-08-01

    This paper describes joint ITPA studies of the I-mode regime, which features an edge thermal barrier together with L-mode-like particle and impurity transport and no edge localized modes (ELMs). The regime has been demonstrated on the Alcator C-Mod, ASDEX Upgrade and DIII-D tokamaks, over a wide range of device parameters and pedestal conditions. Dimensionless parameters at the pedestal show overlap across devices and extend to low collisionality. When they are matched, pedestal temperature profiles are also similar. Pedestals are stable to peeling-ballooning modes, consistent with lack of ELMs. Access to I-mode is independent of heating method (neutral beam injection, ion cyclotron and/or electron cyclotron resonance heating). Normalized energy confinement H 98,y2  ⩾  1 has been achieved for a range of 3  ⩽  q 95  ⩽  4.9 and scales favourably with power. Changes in turbulence in the pedestal region accompany the transition from L-mode to I-mode. The L-I threshold increases with plasma density and current, and with device size, but has a weak dependence on toroidal magnetic field B T. The upper limit of power for I-modes, which is set by I-H transitions, increases with B T and the power range is largest on Alcator C-Mod at B  >  5 T. Issues for extrapolation to ITER and other future fusion devices are discussed.

  5. Bacterial Actins and Their Interactors.

    PubMed

    Gayathri, Pananghat

    2017-01-01

    Bacterial actins polymerize in the presence of nucleotide (preferably ATP), form a common arrangement of monomeric interfaces within a protofilament, and undergo ATP hydrolysis-dependent change in stability of the filament-all of which contribute to performing their respective functions. The relative stability of the filament in the ADP-bound form compared to that of ATP and the rate of addition of monomers at the two ends decide the filament dynamics. One of the major differences between eukaryotic actin and bacterial actins is the variety in protofilament arrangements and dynamics exhibited by the latter. The filament structure and the polymerization dynamics enable them to perform various functions such as shape determination in rod-shaped bacteria (MreB), cell division (FtsA), plasmid segregation (ParM family of actin-like proteins), and organelle positioning (MamK). Though the architecture and dynamics of a few representative filaments have been studied, information on the effect of interacting partners on bacterial actin filament dynamics is not very well known. The chapter reviews some of the structural and functional aspects of bacterial actins, with special focus on the effect that interacting partners exert on the dynamics of bacterial actins, and how these assist them to carry out the functions within the bacterial cell.

  6. Novel actin depolymerizing macrolide aplyronine A.

    PubMed

    Saito, S; Watabe, S; Ozaki, H; Kigoshi, H; Yamada, K; Fusetani, N; Karaki, H

    1996-09-01

    Aplyronine A is a macrolide isolated from Aplysia kurodai. By monitoring fluorescent intensity of pyrenyl-actin, it was found that aplyronine A inhibited both the velocity and the degree of actin polymerization. Aplyronine A also quickly depolymerized F-actin. The kinetics of depolymerization suggest that aplyronine A severs F-actin. The relationship between the concentration of total actin and F-actin at different concentrations of aplyronine A suggests that aplyronine A forms a 1:1 complex with G-actin. From these results, it is concluded that aplyronine A inhibits actin polymerization and depolymerizes F-actin by nibbling. Comparison of the chemical structure of aplyronine A and another actin-depolymerizing macrolide, mycalolide B, suggests that the side-chain but not the macrolide ring of aplyronine A may account for its actin binding and severing activity.

  7. erythro-9-[3-(2-Hydroxynonyl)]adenine is an effective inhibitor of cell motility and actin assembly.

    PubMed Central

    Schliwa, M; Ezzell, R M; Euteneuer, U

    1984-01-01

    erythro-9-[3-(2-Hydroxynonyl)]adenine (EHNA) has been reported previously to be an agent that arrests sperm motility by inhibiting the axonemal dynein ATPase activity and has been used to probe the involvement of putative cytoplasmic dyneins in mitosis and intracellular organelle transport. We report here that EHNA profoundly and reversibly affects several actin-dependent processes, both in vivo and in vitro. It induces dramatic changes in actin organization in cultured cells, inhibits cell translocation, blocks actin-dependent cytoplasmic streaming, interferes with actin-dependent gelation of cytoplasmic extracts, and inhibits actin assembly. Just as the cytochalasins, EHNA appears to be a highly effective inhibitor of actin-based motility, whose effects in complex biological systems should be interpreted with caution. Images PMID:6385006

  8. Actin cytoskeleton: putting a CAP on actin polymerization.

    PubMed

    Stevenson, V A; Theurkauf, W E

    2000-10-05

    Two recent studies have identified a Drosophila homolog of cyclase-associated protein (CAP) as a developmentally important negative regulator of actin polymerization that may also directly mediate signal transduction.

  9. Formin' actin in the nucleus.

    PubMed

    Baarlink, Christian; Grosse, Robert

    2014-01-01

    Many if not most proteins can, under certain conditions, change cellular compartments, such as, for example, shuttling from the cytoplasm to the nucleus. Thus, many proteins may exert functions in various and very different subcellular locations, depending on the signaling context. A large amount of actin regulatory proteins has been detected in the mammalian cell nucleus, although their potential roles are much debated and are just beginning to emerge. Recently, members of the formin family of actin nucleators were also reported to dynamically localize to the nuclear environment. Here we discuss our findings that specific diaphanous-related formins can promote nuclear actin assembly in a signal-dependent manner.

  10. Fluorescent beads disintegrate actin networks.

    PubMed

    Golde, Tom; Schuldt, Carsten; Schnauß, Jörg; Strehle, Dan; Glaser, Martin; Käs, Josef

    2013-10-01

    We studied the influence of fluorescent polystyrene beads on both entangled and cross-linked actin networks. Thermal bead fluctuations were observed via video particle tracking and analyzed with one-point microrheology. Illumination of fluorescent beads with their appropriate excitation wavelength leads to a drastic softening of actin gels. Other wavelengths and bright field microscopy do not increase thermal bead fluctuations. This effect cannot be significantly reduced by adding common oxygen scavengers. We conclude that the usage of fluorescent beads impairs results when studying the microrheology of actin networks.

  11. Actin organization associated with the expression of multidrug resistant phenotype in osteosarcoma cells and the effect of actin depolymerization on drug resistance.

    PubMed

    Takeshita, H; Kusuzaki, K; Ashihara, T; Gebhardt, M C; Mankin, H J; Hirasawa, Y

    1998-04-10

    We have previously reported that P-glycoprotein (Pgp)-overexpressing multidrug resistant (MDR) osteosarcoma cells were functionally more differentiated than their parent cells. The present study showed that in the parent cells, the actin filaments were sparsely distributed or were diffusely spread throughout the cytoplasm, whereas the MDR osteosarcoma cells exhibited a remarkable increase in well-organized actin stress fibers. Furthermore, dihydrocytochalasin B, a specific inhibitor of actin polymerization, dramatically disrupted this network of stress fibers, increased the intracellular accumulation of doxorubicin (DOX) and modified the resistance against DOX. These results indicate that the organization of actin filaments associated with cellular differentiation may be involved in the expression of Pgp function in the MDR osteosarcoma cells.

  12. Investigation of the plasma shaping effects on the H-mode pedestal structure using coupled kinetic neoclassical/MHD stability simulations

    NASA Astrophysics Data System (ADS)

    Pankin, A. Y.; Rafiq, T.; Kritz, A. H.; Park, G. Y.; Snyder, P. B.; Chang, C. S.

    2017-06-01

    The effects of plasma shaping on the H-mode pedestal structure are investigated. High fidelity kinetic simulations of the neoclassical pedestal dynamics are combined with the magnetohydrodynamic (MHD) stability conditions for triggering edge localized mode (ELM) instabilities that limit the pedestal width and height in H-mode plasmas. The neoclassical kinetic XGC0 code [Chang et al., Phys. Plasmas 11, 2649 (2004)] is used in carrying out a scan over plasma elongation and triangularity. As plasma profiles evolve, the MHD stability limits of these profiles are analyzed with the ideal MHD ELITE code [Snyder et al., Phys. Plasmas 9, 2037 (2002)]. Simulations with the XGC0 code, which includes coupled ion-electron dynamics, yield predictions for both ion and electron pedestal profiles. The differences in the predicted H-mode pedestal width and height for the DIII-D discharges with different elongation and triangularities are discussed. For the discharges with higher elongation, it is found that the gradients of the plasma profiles in the H-mode pedestal reach semi-steady states. In these simulations, the pedestal slowly continues to evolve to higher pedestal pressures and bootstrap currents until the peeling-ballooning stability conditions are satisfied. The discharges with lower elongation do not reach the semi-steady state, and ELM crashes are triggered at earlier times. The plasma elongation is found to have a stronger stabilizing effect than the plasma triangularity. For the discharges with lower elongation and lower triangularity, the ELM frequency is large, and the H-mode pedestal evolves rapidly. It is found that the temperature of neutrals in the scrape-off-layer (SOL) region can affect the dynamics of the H-mode pedestal buildup. However, the final pedestal profiles are nearly independent of the neutral temperature. The elongation and triangularity affect the pedestal widths of plasma density and electron temperature profiles differently. This provides a new

  13. Papaverine Prevents Vasospasm by Regulation of Myosin Light Chain Phosphorylation and Actin Polymerization in Human Saphenous Vein

    PubMed Central

    Hocking, Kyle M.; Putumbaka, Gowthami; Wise, Eric S.; Cheung-Flynn, Joyce; Brophy, Colleen M.; Komalavilas, Padmini

    2016-01-01

    Objective Papaverine is used to prevent vasospasm in human saphenous veins (HSV) during vein graft preparation prior to implantation as a bypass conduit. Papaverine is a nonspecific inhibitor of phosphodiesterases, leading to increases in both intracellular cGMP and cAMP. We hypothesized that papaverine reduces force by decreasing intracellular calcium concentrations ([Ca2+]i) and myosin light chain phosphorylation, and increasing actin depolymerization via regulation of actin regulatory protein phosphorylation. Approach and Results HSV was equilibrated in a muscle bath, pre-treated with 1 mM papaverine followed by 5 μM norepinephrine, and force along with [Ca2+]i levels were concurrently measured. Filamentous actin (F-actin) level was measured by an in vitro actin assay. Tissue was snap frozen to measure myosin light chain and actin regulatory protein phosphorylation. Pre-treatment with papaverine completely inhibited norepinephrine-induced force generation, blocked increases in [Ca2+]i and led to a decrease in the phosphorylation of myosin light chain. Papaverine pre-treatment also led to increased phosphorylation of the heat shock-related protein 20 (HSPB6) and the vasodilator stimulated phosphoprotein (VASP), as well as decreased filamentous actin (F-actin) levels suggesting depolymerization of actin. Conclusions These results suggest that papaverine-induced force inhibition of HSV involves [Ca2+]i-mediated inhibition of myosin light chain phosphorylation and actin regulatory protein phosphorylation-mediated actin depolymerization. Thus, papaverine induces sustained inhibition of contraction of HSV by the modulation of both myosin cross-bridge formation and actin cytoskeletal dynamics and is a pharmacological alternative to high pressure distention to prevent vasospasm. PMID:27136356

  14. Free Actin Impairs Macrophage Bacterial Defenses via Scavenger Receptor MARCO Interaction, with Reversal by Plasma Gelsolin.

    PubMed

    Ordija, Christine M; Chiou, Terry Ting-Yu; Yang, Zhiping; Deloid, Glen M; de Oliveira Valdo, Melina; Wang, Zhi; Bedugnis, Alice; Noah, Terry L; Jones, Samuel; Koziel, Henry; Kobzik, Lester

    2017-04-06

    Lung injury can release intracellular actin into the alveolar milieu, and is also associated with increased susceptibility to secondary infections. We investigated the effect of free (extracellular) actin on lung macrophage host defense functions. Western blot analysis demonstrated free actin release into the lung lavage fluids of mouse models of ozone injury, influenza infection and secondary pneumococcal pneumonia, and in samples from patients following burn and inhalation injury. Using levels comparable to those observed in lung injury, we found that free actin markedly inhibited murine lung macrophage binding and uptake in vitro of S. pneumoniae, S. aureus and E. coli e.g., S. pneumoniae, mean % inhibition, actin vs vehicle: 85 ± 0.3 (SD), n = 22, p <.001). Similar effects were observed on the ability of primary human macrophages to bind and ingest fluorescent S. aureus (~75 % inhibition). Plasma gelsolin (pGSN), a protein that functions to bind and cleave actin, restored bacterial binding and uptake by both murine and human macrophages. Scavenger receptor inhibitors reduced binding of fluorescent actin by murine macrophages (fluorescence index (x 10-3) after incubation with vehicle, actin, or actin + polyinosinic acid, respectively: 0.8 ± 0.7, 101.7 ± 50.7, 52.7 ± 16.9, n = 5-6, p < 0.05). In addition, actin binding was reduced in a MARCO / SR-AI/II deficient cell line and by normal AMs obtained from MARCO -/- mice. After release from injured cells during lung injury, free actin likely contributes to impaired host defense by blocking scavenger receptor binding of bacteria. This mechanism for increased risk of secondary infections after lung injury or inflammation may represent another target for therapeutic intervention with pGSN.

  15. Dual pools of actin at presynaptic terminals.

    PubMed

    Bleckert, Adam; Photowala, Huzefa; Alford, Simon

    2012-06-01

    We investigated actin's function in vesicle recycling and exocytosis at lamprey synapses and show that FM1-43 puncta and phalloidin-labeled filamentous actin (F-actin) structures are colocalized, yet recycling vesicles are not contained within F-actin clusters. Additionally, phalloidin also labels a plasma membrane-associated cortical actin. Injection of fluorescent G-actin revealed activity-independent dynamic actin incorporation into presynaptic synaptic vesicle clusters but not into cortical actin. Latrunculin-A, which sequesters G-actin, dispersed vesicle-associated actin structures and prevented subsequent labeled G-actin and phalloidin accumulation at presynaptic puncta, yet cortical phalloidin labeling persisted. Dispersal of presynaptic F-actin structures by latrunculin-A did not disrupt vesicle clustering or recycling or alter the amplitude or kinetics of excitatory postsynaptic currents (EPSCs). However, it slightly enhanced release during repetitive stimulation. While dispersal of presynaptic actin puncta with latrunculin-A failed to disperse synaptic vesicles or inhibit synaptic transmission, presynaptic phalloidin injection blocked exocytosis and reduced endocytosis measured by action potential-evoked FM1-43 staining. Furthermore, phalloidin stabilization of only cortical actin following pretreatment with latrunculin-A was sufficient to inhibit synaptic transmission. Conversely, treatment of axons with jasplakinolide, which induces F-actin accumulation but disrupts F-actin structures in vivo, resulted in increased synaptic transmission accompanied by a loss of phalloidin labeling of cortical actin but no loss of actin labeling within vesicle clusters. Marked synaptic deficits seen with phalloidin stabilization of cortical F-actin, in contrast to the minimal effects of disruption of a synaptic vesicle-associated F-actin, led us to conclude that two structurally and functionally distinct pools of actin exist at presynaptic sites.

  16. Chemotaxis and Actin Oscillations

    NASA Astrophysics Data System (ADS)

    Bodenschatz, Eberhard; Hsu, Hsin-Fang; Negrete, Jose; Beta, Carsten; Pumir, Alain; Gholami, Azam; Tarantola, Marco; Westendorf, Christian; Zykov, Vladimir

    Recently, self-oscillations of the cytoskeletal actin have been observed in Dictyostelium, a model system for studying chemotaxis. Here we report experimental results on the self-oscillation mechanism and the role of regulatory proteins and myosin II. We stimulate cells rapidly and periodically by using photo un-caging of the chemoattractant in a micro-fluidic device and measured the cellular responses. We found that the response amplitude grows with stimulation strength only in a very narrow region of stimulation, after which the response amplitude reaches a plateau. Moreover, the frequency-response is not constant but rather varies with the strength of external stimuli. To understand the underlying mechanism, we analyzed the polymerization and de-polymerization time in the single cell level. Despite of the large cell-to-cell variability, we found that the polymerization time is independent of external stimuli and the de-polymerization time is prolonged as the stimulation strength increases. Our conclusions will be summarized and the role of noise in the signaling network will be discussed. German Science Foundation CRC 937.

  17. [Photodynamic therapy for actinic cheilitis].

    PubMed

    Castaño, E; Comunión, A; Arias, D; Miñano, R; Romero, A; Borbujo, J

    2009-12-01

    Actinic cheilitis is a subtype of actinic keratosis that mainly affects the lower lip and has a higher risk of malignant transformation. Its location on the labial mucosa influences the therapeutic approach. Vermilionectomy requires local or general anesthetic and is associated with a risk of an unsightly scar, and the treatment with 5-fluorouracil or imiquimod lasts for several weeks and the inflammatory reaction can be very intense. A number of authors have used photodynamic therapy as an alternative to the usual treatments. We present 3 patients with histologically confirmed actinic cheilitis treated using photodynamic therapy with methyl aminolevulinic acid as the photosensitizer and red light at 630 nm. The clinical response was good, with no recurrences after 3 to 6 months of follow-up. Our experience supports the use of photodynamic therapy as a good alternative for the treatment of actinic cheilitis.

  18. Thermal unfolding and aggregation of actin.

    PubMed

    Levitsky, Dmitrii I; Pivovarova, Anastasiya V; Mikhailova, Valeria V; Nikolaeva, Olga P

    2008-09-01

    Actin is one of the most abundant proteins in nature. It is found in all eukaryotes and plays a fundamental role in many diverse and dynamic cellular processes. Also, actin is one of the most ubiquitous proteins because actin-like proteins have recently been identified in bacteria. Actin filament (F-actin) is a highly dynamic structure that can exist in different conformational states, and transitions between these states may be important in cytoskeletal dynamics and cell motility. These transitions can be modulated by various factors causing the stabilization or destabilization of actin filaments. In this review, we look at actin stabilization and destabilization as expressed by changes in the thermal stability of actin; specifically, we summarize and analyze the existing data on the thermal unfolding of actin as measured by differential scanning calorimetry. We also analyze in vitro data on the heat-induced aggregation of actin, the process that normally accompanies actin thermal denaturation. In this respect, we focus on the effects of small heat shock proteins, which can prevent the aggregation of thermally denatured actin with no effect on actin thermal unfolding. As a result, we have proposed a mechanism describing the thermal denaturation and aggregation of F-actin. This mechanism explains some of the special features of the thermal unfolding of actin filaments, including the effects of their stabilization and destabilization; it can also explain how small heat shock proteins protect the actin cytoskeleton from damage caused by the accumulation of large insoluble aggregates under heat shock conditions.

  19. Logical gates in actin monomer.

    PubMed

    Adamatzky, Andrew

    2017-09-18

    We evaluate information processing capacity of a single actin molecule by calculating distributions of logical gates implemented by the molecule via propagating patterns of excitation. We represent a filamentous actin molecule as an excitable automaton network (F-actin automaton). where every atom updates its state depending on states of atoms its connected to with chemical bonds (hard neighbours) and atoms being in physical proximity to the atom (soft neighbours). A resting atom excites if a sum of its excited hard neighbours and a weighted sum of its soft neighbours belong to some specified interval. We demonstrate that F-actin automata implement OR, AND, XOR and AND-NOT gates via interacting patterns of excitation. Gate AND is the most common gate and gate XOR is the rarest. Using the architectures of gates discovered we implement one bit half-adder and controlled-not circuits in the F-actin automata. Speed and space values of the F-actin molecular computers are discussed.

  20. Intermediate-k density and magnetic field fluctuations during inter-ELM pedestal evolution in MAST

    NASA Astrophysics Data System (ADS)

    Hillesheim, J. C.; Dickinson, D.; Roach, C. M.; Saarelma, S.; Scannell, R.; Kirk, A.; Crocker, N. A.; Peebles, W. A.; Meyer, H.; the MAST Team

    2016-01-01

    Measurements of local density and magnetic field fluctuations near the pedestal top, conditionally averaged over the edge localized mode (ELM) cycle, have been made in Mega Amp Spherical Tokamak (MAST). A Doppler backscattering (DBS) system installed at MAST was used to measure intermediate-k ≤ft({{k}\\bot}{ρi}≈ 3~\\text{to}~4\\right) density fluctuations at the top of the pedestal. A novel diagnostic technique combining DBS with cross-polarization scattering (CP-DBS) enabled magnetic field fluctuations to also be locally measured at similar wave numbers. Polarization isolation and other effects for CP-DBS are discussed. Both measurements were used in a series of high-β ≤ft({βn}≈ 4.0\\right. -4.5) MAST plasmas with large type-I ELMs with an ˜ 8~\\text{to}~9~\\text{ms} period where microtearing modes (MTMs) had been predicted to be unstable in similar conditions (Dickinson 2012 Phys. Rev. Lett. 108 135002). The measured density fluctuation level increased by a factor of about 4 between 2 and 4 ms after the ELM, which was correlated with the recovery of the density profile while the temperature pedestal height continued to increase slowly. Magnetic field fluctuations showed different temporal behaviors, slowly increasing throughout the ELM cycle as the local β increased. Linear GS2 calculations show both MTM and electron temperature gradient (ETG) modes unstable at similar wave numbers as the measurements (although with more overlap between ETG wave numbers and diagnostic spectral resolution) at the top of the pedestal, along with kinetic ballooning modes are unstable lower in the pedestal (at larger wavelengths). The inferred ratio of fluctuation levels from experiment was ≤ft(δ B/B\\right)/≤ft(δ n/n\\right)≈ 1/20 . The comparable ratios from GS2 were ≤ft(δ B/B\\right)/≤ft(δ n/n\\right)≈ 0.4 for the MTM and ≤ft(δ B/B\\right)/≤ft(δ n/n\\right)≈ 0.02 for the ETG. Both the experimental wave number range and the fluctuation ratio

  1. Kinetic Instabilities that Limit β in the Edge of a Tokamak Plasma: A Picture of an H-Mode Pedestal

    NASA Astrophysics Data System (ADS)

    Dickinson, D.; Roach, C. M.; Saarelma, S.; Scannell, R.; Kirk, A.; Wilson, H. R.

    2012-03-01

    Plasma equilibria reconstructed from the Mega-Amp Spherical Tokamak have sufficient resolution to capture plasma evolution during the short period between edge-localized modes (ELMs). Immediately after the ELM, steep gradients in pressure, P, and density, ne, form pedestals close to the separatrix, and they then expand into the core. Local gyrokinetic analysis over the ELM cycle reveals the dominant microinstabilities at perpendicular wavelengths of the order of the ion Larmor radius. These are kinetic ballooning modes in the pedestal and microtearing modes in the core close to the pedestal top. The evolving growth rate spectra, supported by gyrokinetic analysis using artificial local equilibrium scans, suggest a new physical picture for the formation and arrest of this pedestal.

  2. Kinetic instabilities that limit β in the edge of a tokamak plasma: a picture of an H-mode pedestal.

    PubMed

    Dickinson, D; Roach, C M; Saarelma, S; Scannell, R; Kirk, A; Wilson, H R

    2012-03-30

    Plasma equilibria reconstructed from the Mega-Amp Spherical Tokamak have sufficient resolution to capture plasma evolution during the short period between edge-localized modes (ELMs). Immediately after the ELM, steep gradients in pressure, P, and density, n(e), form pedestals close to the separatrix, and they then expand into the core. Local gyrokinetic analysis over the ELM cycle reveals the dominant microinstabilities at perpendicular wavelengths of the order of the ion Larmor radius. These are kinetic ballooning modes in the pedestal and microtearing modes in the core close to the pedestal top. The evolving growth rate spectra, supported by gyrokinetic analysis using artificial local equilibrium scans, suggest a new physical picture for the formation and arrest of this pedestal.

  3. Polymer dynamics and fluid flow in actin-based cell motility

    NASA Astrophysics Data System (ADS)

    Theriot, Julie

    2005-03-01

    In living cells, nonequilibrium protein polymerization reactions are frequently used to convert chemical energy into mechanical energy and thereby generate useful force for cellular movements. We have examined the polymer and fluid dynamics in two biological cases where the assembly of branched actin filament networks generates force: the intracellular movement of the bacterial pathogen Listeria monocytogenes, and the extension of the leading edge of skin epithelial cells during wound-healing. In both cases, net actin filament assembly occurs at the front of the network structure and net disassembly occurs at the rear. Actin protein subunits and other network components must be recycled through the fluid phase to the front of the polymerizing network in order for forward movement to continue at steady state. For actin-based movement of Listeria monocytogenes, we have found that actin recycling is not rate-limiting; instead, the speed of movement is governed by the cooperative dissociation of groups of noncovalent protein-protein bonds attaching the filamentous network to the bacterial surface. In contrast, rapid actin-based extension at the leading edge of moving epithelial cells is associated with unusual perturbations in intracellular fluid flow.

  4. Effect of Actin Filament on Deformation-Induced Ca2+ Response in Osteoblast-Like Cells

    NASA Astrophysics Data System (ADS)

    Adachi, Taiji; Murai, Takayuki; Hoshiai, Sodai; Tomita, Yoshihiro

    Under the influence of mechanical environment, bone structure is formed and maintained by adaptive remodeling that involves osteoclastic resorption and osteoblastic formation. In the mechanotransduction system in osteoblasts, it is believed that intracellular calcium plays a fundamental role and cytoskeletal actin filament is a crucial component for the signal transduction process. To clarify the role of actin filament in deformation-induced Ca2+ signaling, osteoblast-like cells (MC3T3-E1) with different actin filament densities controlled by cytochalasin D were subjected to tensile strain in vitro. The change in intracellular Ca2+ concentration labeled by fluo-3 was observed using a confocal laser-scanning microscope. As a result, the disruption of the actin filament was found to significantly suppress the deformation-induced Ca2+ response that was regulated according to the degree of actin filament organization. This result indicates that the actin filament is indispensable for the quantitative regulation of Ca2+ signaling in response to a mechanical stimulus in osteoblasts.

  5. The trinity of the cortical actin in the initiation of HIV-1 infection

    PubMed Central

    2012-01-01

    For an infecting viral pathogen, the actin cortex inside the host cell is the first line of intracellular components that it encounters. Viruses devise various strategies to actively engage or circumvent the actin structure. In this regard, the human immunodeficiency virus-1 (HIV-1) exemplifies command of cellular processes to take control of actin dynamics for the initiation of infection. It has becomes increasingly evident that cortical actin presents itself both as a barrier to viral intracellular migration and as a necessary cofactor that the virus must actively engage, particularly, in the infection of resting CD4 blood T cells, the primary targets of HIV-1. The coercion of this most fundamental cellular component permits infection by facilitating entry, reverse transcription, and nuclear migration, three essential processes for the establishment of viral infection and latency in blood T cells. It is the purpose of this review to examine, in detail, the manifestation of viral dependence on the actin cytoskeleton, and present a model of how HIV utilizes actin dynamics to initiate infection. PMID:22640593

  6. Antibody against the actin-binding protein depactin attenuates Ca2+ signaling in starfish eggs.

    PubMed

    Chun, Jong T; Vasilev, Filip; Santella, Luigia

    2013-11-15

    Being present in starfish oocytes, the cofilin/ADF (actin-depolymerizing factor) family protein depactin severs actin filaments. Previously, we reported that exogenous cofilin microinjected into starfish eggs significantly augmented the Ca(2+) release in response to inositol 1,4,5-trisphosphate (InsP3) or fertilizing sperm, raising the possibility that intracellular Ca(2+) signaling could be modulated by the actin cytoskeleton. In this communication, we have targeted the endogenous depactin by use of the specific antibody that was raised against its actin-binding domain. The anti-depactin antibody microinjected into the starfish oocytes and eggs effectively altered the structure of the actin cytoskeleton, and significantly delayed the meiotic progression induced by 1-methyladenine. When microinjected into the mature eggs, the anti-depactin antibody markedly reduced the amplitude of the Ca(2+) response in a dose-dependent manner, corroborating the results of our previous study with cofilin. In addition, the eggs microinjected with the anti-depactin antibody displayed reduced rate of successful elevation of the fertilization envelope and an elevated tendency of polyspermic interaction. Taken together, our data suggest that the actin cytoskeleton is implicated not only in meiotic maturation and intracellular Ca(2+) signaling, but also in the fine regulation of gametes interaction and cortical granules exocytosis. Copyright © 2013 Elsevier Inc. All rights reserved.

  7. Cofilin-mediated actin dynamics promotes actin bundle formation during Drosophila bristle development

    PubMed Central

    Wu, Jing; Wang, Heng; Guo, Xuan; Chen, Jiong

    2016-01-01

    The actin bundle is an array of linear actin filaments cross-linked by actin-bundling proteins, but its assembly and dynamics are not as well understood as those of the branched actin network. Here we used the Drosophila bristle as a model system to study actin bundle formation. We found that cofilin, a major actin disassembly factor of the branched actin network, promotes the formation and positioning of actin bundles in the developing bristles. Loss of function of cofilin or AIP1, a cofactor of cofilin, each resulted in increased F-actin levels and severe defects in actin bundle organization, with the defects from cofilin deficiency being more severe. Further analyses revealed that cofilin likely regulates actin bundle formation and positioning by the following means. First, cofilin promotes a large G-actin pool both locally and globally, likely ensuring rapid actin polymerization for bundle initiation and growth. Second, cofilin limits the size of a nonbundled actin-myosin network to regulate the positioning of actin bundles. Third, cofilin prevents incorrect assembly of branched and myosin-associated actin filament into bundles. Together these results demonstrate that the interaction between the dynamic dendritic actin network and the assembling actin bundles is critical for actin bundle formation and needs to be closely regulated. PMID:27385345

  8. Support pedestals for interconnecting a cover and nozzle band wall in a gas turbine nozzle segment

    DOEpatents

    Yu, Yufeng Phillip; Itzel, Gary Michael; Webbon, Waylon Willard; Bagepalli, Radhakrishna; Burdgick, Steven Sebastian; Kellock, Iain Robertson

    2002-01-01

    A gas turbine nozzle segment has outer and inner band portions. Each band portion includes a nozzle wall, a cover and an impingement plate between the cover and nozzle wall defining two cavities on opposite sides of the impingement plate. Cooling steam is supplied to one cavity for flow through the apertures of the impingement plate to cool the nozzle wall. Structural pedestals interconnect the cover and nozzle wall and pass through holes in the impingement plate to reduce localized stress otherwise resulting from a difference in pressure within the chamber of the nozzle segment and the hot gas path and the fixed turbine casing surrounding the nozzle stage. The pedestals may be cast or welded to the cover and nozzle wall.

  9. NASTRAN structural model for the large ground antenna pedestal with applications to hydrostatic bearing of film

    NASA Technical Reports Server (NTRS)

    Chian, C. T.

    1986-01-01

    Investigations were conducted on the 64-meter antenna hydrostatic bearing oil film thickness under a variety of loads and elastic moduli. These parametric studies used a NASTRAN pedestal structural model to determine the deflections under the hydrostatic bearing pad. The deflections formed the input for a computer program to determine the hydrostratic bearing oil film thickness. For the future 64-meter to 70-meter antenna extension and for the 2.2-meter (86-in.) haunch concrete replacement cases, the program predicted safe oil film thickness (greater than 0.13 mm (0.005 in.) at the corners of the pad). The effects of varying moduli of elasticity for different sections of the pedestal and the film height under stressed runner conditions were also studied.

  10. Ballooning Stability Of Tokamak Pedestals In The Presence Of Applied 3D Magnetic Perturbations

    NASA Astrophysics Data System (ADS)

    Cote, T. B.; Hegna, C. C.; Willensdorfer, M.; Strumberger, E.; Suttrop, W.; Zohm, H.

    2016-10-01

    Applied 3d magnetic perturbations can destabilize ideal mhd ballooning modes in tokamak pedestals. In this work, we describe techniques for studying infinite-n ballooning stability of 3d equilibria deduced from vmec calculations. Full magnetic profiles from vmec are used to construct local equilibria for flux surfaces in and around the edge pedestal region. These local equilibrium calculations are coupled with ideal ballooning stability analysis to determine stability of the system for given rmp configurations. This theoretical development is motivated by recent asdex-u experiments, where toroidally localized high-n mhd activity is observed in the presence of applied 3d fields. We will attempt to explain these observations. Supported by US DOE under Grant No. DE-FG02-86ER53218.

  11. Test and model correlation of the atmospheric emission photometric imager fiberglass pedestal

    NASA Technical Reports Server (NTRS)

    Lee, H. M., III; Barker, L. A.

    1990-01-01

    The correlation is presented of the static loads testing and finite element modeling for the fiberglass pedestal used on the Atmospheric Emission Photometric Imaging (AEPI) experiment. This payload is to be launched in the space shuttle as part of the ATLAS-1 experiment. Strain gauge data from rosettes around the highly loaded base are compared to the same load case run for the Spacelab 1 testing done in 1981. Correlation of the model and test data was accomplished through comparison of the composite stress invariant using the expected flight loads for the ATLAS-1 mission. Where appropriate, the Tsai-Wu failure criteria was used in the development of the key margins of safety. Margins of safety are all positive for the pedestal and are reported.

  12. Characterization and parametric dependencies of low wavenumber pedestal turbulence in the National Spherical Torus Experiment

    SciTech Connect

    Smith, D. R.; Fonck, R. J.; McKee, G. R.; Thompson, D. S.; Bell, R. E.; Diallo, A.; Guttenfelder, W.; Kaye, S. M.; LeBlanc, B. P.; Podesta, M.

    2013-05-15

    The spherical torus edge region is among the most challenging regimes for plasma turbulence simulations. Here, we measure the spatial and temporal properties of ion-scale turbulence in the steep gradient region of H-mode pedestals during edge localized mode-free, MHD quiescent periods in the National Spherical Torus Experiment. Poloidal correlation lengths are about 10 ρ{sub i}, and decorrelation times are about 5 a/c{sub s}. Next, we introduce a model aggregation technique to identify parametric dependencies among turbulence quantities and transport-relevant plasma parameters. The parametric dependencies show the most agreement with transport driven by trapped-electron mode, kinetic ballooning mode, and microtearing mode turbulence, and the least agreement with ion temperature gradient turbulence. In addition, the parametric dependencies are consistent with turbulence regulation by flow shear and the empirical relationship between wider pedestals and larger turbulent structures.

  13. Effect of Pedestal Temperature on Bonding Strength and Deformation Characteristics for 5N Copper Wire Bonding

    NASA Astrophysics Data System (ADS)

    Singh, Gurbinder; Haseeb, A. S. M. A.

    2016-06-01

    In recent years, copper has increasingly been used to replace gold to create wire-bonded interconnections in microelectronics. While engineers and researchers in the semiconductor packaging field are continuously working on this transition from gold to copper wires to reduce costs, the challenge remains in producing robust and reliable joints for semiconductor devices. This research paper investigates the effect of pedestal temperature on bonding strength and deformation for 99.999% purity (5N) copper wire bonding on nickel-palladium-gold (NiPdAu) bond pads. With increasing pedestal temperature, significant thinning of the copper ball bond can be achieved, resulting in higher as-bonded ball shear strengths while producing no pad damage. This can be helpful for low-k devices with thin structures, so as to prevent the use of excessive bond force and ultrasonic energy during copper wire bonding.

  14. Predictive modelling of the impact of a radiative divertor on pedestal confinement on ASDEX Upgrade

    NASA Astrophysics Data System (ADS)

    Dunne, Mike; Potzel, Steffen; Wischmeier, Marco; Wolfrum, Elisabeth; Frassinetti, Lorenzo; Reimold, Felix; Eurofusion Mst1 Team; ASDEX Upgrade Team

    2015-11-01

    In future devices, tailoring of the edge density profile and radiation profile for power exhaust control via a deuterium gas puff and extrinsic impurity seeding will be necessary. It has been observed on present day machines that high D fuelling can reduce the plasma stored energy while adding impurity seeding can act to improve confinement by up to 40%. This study presents a combination of observations and modelling completed on AUG with the aim of determining the mechanisms behind the confinement degradation with a gas puff and improvement with impurity seeding. In particular, predictive modelling, based on the EPED pedestal model, has been extensively used. Alterations of the temperature and density at the separatrix are found to have large impacts on pedestal stability. Measured changes in divertor properties are used to inform the direction and magnitude of these alterations, with experimentally relevant confinement changes being recovered via pressure profile shifts. http://www.euro-fusionscipub.org/mst1

  15. Unipolar reorganization of F-actin layer at bacterial division and bundling of actin filaments by plastin correlate with movement of Shigella flexneri within HeLa cells.

    PubMed Central

    Prévost, M C; Lesourd, M; Arpin, M; Vernel, F; Mounier, J; Hellio, R; Sansonetti, P J

    1992-01-01

    Shigella flexneri causes bacillary dysentery, an invasive disease of colonic epithelial cells in humans. The capacity of bacteria, once they have entered into a cell and escaped the phagocytic vacuole, to spread intracellularly and directly to adjacent cells without further extracellular passage is a key factor in invasion of the epithelial layer. Movement of intracellular bacteria is dependent upon the polymerization of actin; concentration of the formed filaments to one end of the bacterium is associated with initiation of movement. This movement may lead to the formation of a protrusion at the cell surface through which the bacterium passes to an adjacent cell. Development of these protrusions in infected HeLa cells is described, with emphasis on two critical observations. First, initiation of movement is coupled with bacterial division since elongation of the bacterial body is associated with relocalization of the previously uniformly distributed layer of actin to one pole of the bacterium. Second, the actin-bundling protein plastin appears to bundle the actin filaments just posterior to the bacterium, producing an ongoing contraction of the cylindrical actin tail that may be associated with forward movement of the bacterium within the protrusion. Images PMID:1398922

  16. High-Precision Dispensing of Nanoliter Biofluids on Glass Pedestal Arrays for Ultrasensitive Biomolecule Detection.

    PubMed

    Chen, Xiaoxiao; Liu, Yang; Xu, QianFeng; Zhu, Jing; Poget, Sébastien F; Lyons, Alan M

    2016-05-04

    Precise dispensing of nanoliter droplets is necessary for the development of sensitive and accurate assays, especially when the availability of the source solution is limited. Conventional approaches are limited by imprecise positioning, large shear forces, surface tension effects, and high costs. To address the need for precise and economical dispensing of nanoliter volumes, we developed a new approach where the dispensed volume is dependent on the size and shape of defined surface features, thus freeing the dispensing process from pumps and fine-gauge needles requiring accurate positioning. The surface we fabricated, called a nanoliter droplet virtual well microplate (nVWP), achieves high-precision dispensing (better than ±0.5 nL or ±1.6% at 32 nL) of 20-40 nL droplets using a small source drop (3-10 μL) on isolated hydrophilic glass pedestals (500 μm on a side) bonded to arrays of polydimethylsiloxane conical posts. The sharp 90° edge of the glass pedestal pins the solid-liquid-vapor triple contact line (TCL), averting the wetting of the glass sidewalls while the fluid is prevented from receding from the edge. This edge creates a sufficiently large energy barrier such that microliter water droplets can be poised on the glass pedestals, exhibiting contact angles greater >150°. This approach relieves the stringent mechanical alignment tolerances required for conventional dispensing techniques, shifting the control of dispensed volume to the area circumscribed by the glass edge. The effects of glass surface chemistry and dispense velocity on droplet volume were studied using optical microscopy and high-speed video. Functionalization of the glass pedestal surface enabled the selective adsorption of specific peptides and proteins from synthetic and natural biomolecule mixtures, such as venom. We further demonstrate how the nVWP dispensing platform can be used for a variety of assays, including sensitive detection of proteins and peptides by fluorescence

  17. Actin Recruitment to the Chlamydia Inclusion Is Spatiotemporally Regulated by a Mechanism That Requires Host and Bacterial Factors

    PubMed Central

    Chin, Elizabeth; Kirker, Kelly; Zuck, Meghan; James, Garth; Hybiske, Kevin

    2012-01-01

    The ability to exit host cells at the end of their developmental growth is a critical step for the intracellular bacterium Chlamydia. One exit strategy, extrusion, is mediated by host signaling pathways involved with actin polymerization. Here, we show that actin is recruited to the chlamydial inclusion as a late event, occurring after 20 hours post-infection (hpi) and only within a subpopulation of cells. This event increases significantly in prevalence and extent from 20 to 68 hpi, and actin coats strongly correlated with extrusions. In contrast to what has been reported for other intracellular pathogens, actin nucleation on Chlamydia inclusions did not ‘flash’, but rather exhibited moderate depolymerization dynamics. By using small molecule agents to selectively disrupt host signaling pathways involved with actin nucleation, modulate actin polymerization dynamics and also to disable the synthesis and secretion of chlamydial proteins, we further show that host and bacterial proteins are required for actin coat formation. Transient disruption of either host or bacterial signaling pathways resulted in rapid loss of coats in all infected cells and a reduction in extrusion formation. Inhibition of Chlamydia type III secretion also resulted in rapid loss of actin association on inclusions, thus implicating chlamydial effector proteins(s) as being central factors for engaging with host actin nucleating factors, such as formins. In conclusion, our data illuminate the host and bacterial driven process by which a dense actin matrix is dynamically nucleated and maintained on the Chlamydia inclusion. This late stage event is not ubiquitous for all infected cells in a population, and escalates in prevalence and extent throughout the developmental cycle of Chlamydia, culminating with their exit from the host cell by extrusion. The initiation of actin recruitment by Chlamydia appears to be novel, and may serve as an upstream determinant of the extrusion mechanism. PMID

  18. Roles of actin cytoskeleton for regulation of chloroplast anchoring.

    PubMed

    Sakai, Yuuki; Takagi, Shingo

    2017-08-22

    Chloroplasts are known to maintain specific intracellular distribution patterns under specific environmental conditions, enabling the optimal performance of photosynthesis. To this end, chloroplasts are anchored in the cortical cytoplasm. In leaf epidermal cells of aquatic monocot Vallisneria, we recently demonstrated that the anchored chloroplasts are rapidly de-anchored upon irradiation with high-intensity blue light and that the process is probably mediated by the blue-light receptor phototropins. Chloroplast de-anchoring is a necessary step rendering the previously anchored chloroplasts mobile to allow their migration. In this article, based on the results obtained in Vallisneria together with those in other plant species, we briefly discussed possible modes of regulation of chloroplast anchoring and de-anchoring by actin cytoskeleton. The topics include roles of photoreceptor systems, actin-filament-dependent and -independent chloroplast anchoring, and independence of chloroplast de-anchoring from actomyosin and microtubule systems.

  19. Mechanics of biomimetic systems propelled by actin comet tails

    NASA Astrophysics Data System (ADS)

    Kang, Hyeran; Tambe, Dhananjay; Shenoy, Vivek; Tang, Jay

    2009-03-01

    The motility of intracellular bacterial pathogens such as Listeria monocytogenes is driven by filamentous actin comet tails in a variety of trajectories. Here, we present the in vitro study on the actin-based movements using spherical beads of different sizes coated with VCA protein, a partial domain of N-Wasp, in platelet extracts. Long term two-dimensional trajectories of the spherical beads motility show characteristic difference than those observed for bacteria, which have both elongated shape and asymmetric expression of the polymerization inducing enzyme. The trajectories also vary sensitively with the bead size and shape. These results provide a useful test to our new analytical model including the rotation of the bead relative to the tail.

  20. Plant actin cytoskeleton re-modeling by plant parasitic nematodes.

    PubMed

    Engler, Janice de Almeida; Rodiuc, Natalia; Smertenko, Andrei; Abad, Pierre

    2010-03-01

    The cytoskeleton is an important component of the plant's defense mechanism against the attack of pathogenic organisms. Plants however, are defenseless against parasitic root-knot and cyst nematodes and respond to the invasion by the development of a special feeding site that supplies the parasite with nutrients required for the completion of its life cycle. Recent studies of nematode invasion under treatment with cytoskeletal drugs and in mutant plants where normal functions of the cytoskeleton have been affected, demonstrate the importance of the cytoskeleton in the establishment of a feeding site and successful nematode reproduction. It appears that in the case of microfilaments, nematodes hijack the intracellular machinery that regulates actin dynamics and modulate the organization and properties of the actin filament network. Intervening with this process reduces the nematode infection efficiency and inhibits its life cycle. This discovery uncovers a new pathway that can be exploited for the protection of plants against nematodes.

  1. Self-consistent core-pedestal transport simulations with neural network accelerated models

    DOE PAGES

    Meneghini, Orso; Smith, Sterling P.; Snyder, Philip B.; ...

    2017-07-12

    Fusion whole device modeling simulations require comprehensive models that are simultaneously physically accurate, fast, robust, and predictive. In this paper we describe the development of two neural-network (NN) based models as a means to perform a snon-linear multivariate regression of theory-based models for the core turbulent transport fluxes, and the pedestal structure. Specifically, we find that a NN-based approach can be used to consistently reproduce the results of the TGLF and EPED1 theory-based models over a broad range of plasma regimes, and with a computational speedup of several orders of magnitudes. These models are then integrated into a predictive workflowmore » that allows prediction with self-consistent core-pedestal coupling of the kinetic profiles within the last closed flux surface of the plasma. Finally, the NN paradigm is capable of breaking the speed-accuracy trade-off that is expected of traditional numerical physics models, and can provide the missing link towards self-consistent coupled core-pedestal whole device modeling simulations that are physically accurate and yet take only seconds to run.« less

  2. Impact of plasma core profiles on MHD stability at tokamak edge pedestal

    NASA Astrophysics Data System (ADS)

    Aiba, N.; Urano, H.

    2014-11-01

    Impact of plasma core profiles on magnetohydrodynamics (MHD) stability at tokamak edge pedestal is investigated numerically to extend an operation regime for small amplitude grassy edge localized mode (ELM). With the hypotheses that pedestal pressure profile can be predicted with the EPED1 model and the trigger of grassy ELM is an ideal ballooning mode, the impacts of plasma poloidal beta and plasma internal inductance on edge MHD stability are investigated, the parameters of which are related to plasma core profiles and are important parameters for grassy ELMy H-modes in JET quasi-double null plasma. The numerical results indicate that a ballooning mode can be destabilized by decreasing poloidal beta and/or internal inductance. In contrast, it is confirmed that pedestal density, which is also an important parameter for realizing grassy ELMy H-mode, can stabilize a ballooning mode. In combination with these trends, it is possible to relax the necessary conditions for grassy ELMy H-mode by adjusting the parameters carefully, though this relaxation destabilizes type-I ELM more easily due to the increase in edge current density.

  3. Preliminary experience using a cochlear implant with a novel linear pedestal design.

    PubMed

    Parkes, William J; Gnanasegaram, Joshua J; Cushing, Sharon L; James, Adrian L; Gordon, Karen A; Papsin, Blake C

    2017-02-01

    To assess the safety and efficiency of cochlear implantation using a novel device with a linear silastic pedestal (2 mm wide, 2 mm deep, 10 mm long) on the flat undersurface. Operative times required to drill a linear groove (LG) for the new linear pedestal design were prospectively accrued for 46 implantations in 30 children (median age 3). Intra-operative safety was assessed during each case. Instances of dural exposure in the base of the LG were noted. Length of stay was also recorded as a secondary measure of efficiency. Across all surgeons, the mean time needed to create the LG was 1.9 ± 1.5 min (±SD) with a median time of 1.5 min (95% Cl: 1-2 min). The range in time was 1-10 min. No intraoperative complications occurred. Intended device positioning was confirmed with on-table post-operative x-rays in all cases. 43% of patients were discharged on the day of surgery. The novel linear pedestal design allows for deliberate device placement while adding little additional operative time and complexity, an improvement on our current standard of care. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  4. Enhanced H-mode pedestals with lithium injection in DIII-D

    DOE PAGES

    Osborne, Thomas H.; Jackson, Gary L.; Yan, Zheng; ...

    2015-05-08

    Periods of edge localized mode (ELM)-free H-mode with increased pedestal pressure and width were observed in the DIII-D tokamak when density fluctuations localized to the region near the separatrix were present. Injection of a powder of 45 μm diameter lithium particles increased the duration of the enhanced pedestal phases to up to 350 ms, and also increased the likelihood of a transition to the enhanced phase. Lithium injection at a level sufficient for triggering the extended enhanced phases resulted in significant lithium in the plasma core, but carbon and other higher Z impurities as well as radiated power levels weremore » reduced. Recycling of the working deuterium gas appeared unaffected by this level of lithium injection. The ion scale, kθ ρs ~ 0.1–0.2, density fluctuations propagated in the electron drift direction with f ~ 80 kHz and occurred in bursts every ~1 ms. The fluctuation bursts correlated with plasma loss resulting in a flattening of the pressure profile in a region near the separatrix. This localized flattening 2 allowed higher overall pedestal pressure at the peeling-ballooning stability limit and higher pressure than expected under the EPED model due to reduction of the pressure gradient below the “ballooning critical profile”. Furthermore, reduction of the ion pressure by lithium dilution may contribute to the long ELM-free periods.« less

  5. MHD Analysis of the Tokamak Edge Pedestal in the Low Collisionality Regime

    NASA Astrophysics Data System (ADS)

    Snyder, P. B.; Wilson, H. R.; Konz, C.

    2005-10-01

    The peeling-ballooning model proposes that intermediate wavelength MHD instabilities are responsible for edge localized modes (ELMs) and impose constraints on the pedestal height. In typical discharges with ELMs, the pedestal goes unstable to coupled peeling-ballooning or pure ballooning modes shortly before an ELM is observed. However, at very low collisionality, the bootstrap current in the pedestal region can be large, even very near the separatrix, and the discharge can be most unstable to current-driven kink/peeling modes, typically at relatively low mode number (n˜,1-10). Recently, interesting ELM-free regimes, including both Quiescent (QH) and Resonant Magnetic Perturbation (RMP) H-mode, have been observed to occur in this low collisionality regime. Here we systematically explore MHD stability in this regime, including the effects of a conducting wall and sheared toroidal flow. We consider the implications for both RMP and QH discharges, including possible connections between the EHO observed in QH mode and low-n kink/peeling modes.

  6. Characterization of broadband fluctuations in wide-pedestal QH-mode plasmas on DIII-D

    NASA Astrophysics Data System (ADS)

    Muscatello, C. M.; Burrell, K. H.; Luhmann, N. C., Jr.; McKee, G. R.; Tobias, B.

    2016-10-01

    Edge broadband fluctuations observed in wide pedestal quiescent H-mode plasmas may play an important role in driving transport necessary for stabilizing the edge to kink-peeling modes, thought to lead to ELMs. Density fluctuation measurements from BES and MIR independently observe periodic bursts in the pedestal that show up spectrally as broadband fluctuations. The period of the fluctuation bursts correlate with the period of enhanced bicoherence in the frequency range of the fluctuations, suggesting nonlinear coupling of turbulence. Time-delay estimation analysis of the 2D BES data shows strong evidence of a low-frequency zonal flow in the pedestal with a period matching that of the bursts. The carbon pressure gradient and E × B velocity, determined from CER, and ECE emission also oscillate with the same period. This behavior can be described as a quasi-stationary, limit-cycle oscillation and modeled by a set of predator-prey equations relating the zonal flow, equilibrium flow, and turbulence amplitude. Supported by the US DOE under DE-FC02-04ER54698, DE-FG02-99ER54531, DE-AC02-09CH11466.

  7. High resolution equilibrium calculations of pedestal and SOL plasma in tokamaks

    NASA Astrophysics Data System (ADS)

    Medvedev, S. Yu; Martynov, A. A.; Drozdov, V. V.; Ivanov, A. A.; Poshekhonov, Yu Yu

    2017-02-01

    For integrated modeling of equilibrium, stability and dynamics of the divertor tokamak plasma with scrape-off layer (SOL) high resolution equilibrium calculations are needed. A new version of the CAXE equilibrium code computes the tokamak equilibrium on a numerical grid adaptive to magnetic surfaces both in the plasma region with closed flux surfaces and in the SOL region with open magnetic lines. The plasma profiles can be prescribed independently in each region with nested flux surfaces, and realistic SOL profiles with very short pressure drop off length can be accurately treated. The influence of the finite current density in SOL on the connection length is studied. From the point of view of the MHD equilibrium and stability modeling, self-consistent calculations of diverted tokamak configurations with finite current density at the separatrix require taking into account plasma outside the separatrix. Calculated high resolution equilibria provide an input to new versions of the ideal MHD stability codes treating tokamak plasma with SOL. The study of the influence of the pressure gradient profile in the pedestal plasma inside and outside the separatrix on the pedestal height limit set by external kink-ballooning mode stability is presented. Another possible application of the high resolution pedestal and SOL equilibrium code is a coupling to the SOLPS code with a purpose to increase equilibrium accuracy and support self-consistent plasma flow/equilibrium modeling.

  8. Self-consistent core-pedestal transport simulations with neural network accelerated models

    NASA Astrophysics Data System (ADS)

    Meneghini, O.; Smith, S. P.; Snyder, P. B.; Staebler, G. M.; Candy, J.; Belli, E.; Lao, L.; Kostuk, M.; Luce, T.; Luda, T.; Park, J. M.; Poli, F.

    2017-08-01

    Fusion whole device modeling simulations require comprehensive models that are simultaneously physically accurate, fast, robust, and predictive. In this paper we describe the development of two neural-network (NN) based models as a means to perform a snon-linear multivariate regression of theory-based models for the core turbulent transport fluxes, and the pedestal structure. Specifically, we find that a NN-based approach can be used to consistently reproduce the results of the TGLF and EPED1 theory-based models over a broad range of plasma regimes, and with a computational speedup of several orders of magnitudes. These models are then integrated into a predictive workflow that allows prediction with self-consistent core-pedestal coupling of the kinetic profiles within the last closed flux surface of the plasma. The NN paradigm is capable of breaking the speed-accuracy trade-off that is expected of traditional numerical physics models, and can provide the missing link towards self-consistent coupled core-pedestal whole device modeling simulations that are physically accurate and yet take only seconds to run.

  9. Enhanced H-mode pedestals with lithium injection in DIII-D

    SciTech Connect

    Osborne, Thomas H.; Jackson, Gary L.; Yan, Zheng; Maingi, Rajesh; Mansfield, Dennis K.; Grierson, Brian A.; Chrobak, Chris P.; McLean, Adam G.; Allen, Steve L.; Battaglia, Devon J.; Briesemeister, Alexis R.; Fenstermacher, Max E.; McKee, George R.; Snyder, Philip B.

    2015-05-08

    Periods of edge localized mode (ELM)-free H-mode with increased pedestal pressure and width were observed in the DIII-D tokamak when density fluctuations localized to the region near the separatrix were present. Injection of a powder of 45 μm diameter lithium particles increased the duration of the enhanced pedestal phases to up to 350 ms, and also increased the likelihood of a transition to the enhanced phase. Lithium injection at a level sufficient for triggering the extended enhanced phases resulted in significant lithium in the plasma core, but carbon and other higher Z impurities as well as radiated power levels were reduced. Recycling of the working deuterium gas appeared unaffected by this level of lithium injection. The ion scale, kθ ρs ~ 0.1–0.2, density fluctuations propagated in the electron drift direction with f ~ 80 kHz and occurred in bursts every ~1 ms. The fluctuation bursts correlated with plasma loss resulting in a flattening of the pressure profile in a region near the separatrix. This localized flattening 2 allowed higher overall pedestal pressure at the peeling-ballooning stability limit and higher pressure than expected under the EPED model due to reduction of the pressure gradient below the “ballooning critical profile”. Furthermore, reduction of the ion pressure by lithium dilution may contribute to the long ELM-free periods.

  10. Enhanced H-mode pedestals with lithium injection in DIII-D

    SciTech Connect

    Osborne, T. H.; Jackson, G. L.; Yan, Z.; Maingi, R.; Mansfield, D. K.; Grierson, Brian A.; Chrobak, C. P.; McLean, A. G.; Allen, Steve L.; Battaglia, D. J.; Briesemeister, Alexis R.; Fenstermacher, M. E.; Mckee, G. R.; Snyder, P. B.

    2015-05-08

    Periods of edge localized mode (ELM)-free H-mode with increased pedestal pressure and width were observed in the DIII-D tokamak when density fluctuations localized to the region near the separatrix were present. Injection of a powder of 45 m diameter lithium particles increased the duration of the enhanced pedestal phases to up to 350 ms, and also increased the likelihood of a transition to the enhanced phase. Lithium injection at a level sufficient for triggering the extended enhanced phases resulted in significant lithium in the plasma core, but carbon and other higher Z impurities as well as radiated power levels were reduced. Recycling of the working deuterium gas appeared unaffected by this level of lithium injection. The ion scale, kθϱs ~ 0.1 0.2, density fluctuations propagated in the electron drift direction with f ~ 80 kHz and occurred in bursts every ~1 ms. The fluctuation bursts correlated with plasma loss resulting in a flattening of the pressure profile in a region near the separatrix. This localized flattening allowed higher overall pedestal pressure at the peeling ballooning stability limit and higher pressure than expected under the EPED model due to reduction of the pressure gradient below the 'ballooning critical profile'. Furthermore, reduction of the ion pressure by lithium dilution may contribute to the long ELM-free periods.

  11. The relationships between edge localized modes suppression, pedestal profiles and lithium wall coatings in NSTX

    SciTech Connect

    Boyle, D. P.; Maingi, R.; Snyder, P. B.; Manickam, J.; Osborne, T.H.; Bell, R. E.; LeBlanc, B. P.

    2011-01-01

    Recently in the National Spherical Torus Experiment (NSTX), increasing lithium wall coatings suppressed edge localized modes (ELMs), gradually but not quite monotonically. This work details profile and stability analysis as ELMs disappeared throughout the lithium scan. While the quantity of lithium deposited between discharges did not uniquely determine the presence of ELMs, profile analysis demonstrated that lithium was correlated with wider density and pressure pedestals with peak gradients farther from the separatrix. Moreover, the ELMy and ELM-free discharges were cleanly separated by their density and pedestal widths and peak gradient locations. Ultimately, ELMs were only suppressed when lithium caused the density pedestal to widen and shift inward. These changes in the density gradient were directly reflected in the pressure gradient and calculated bootstrap current. This supports the theory that ELMs in NSTX are caused by peeling and/or ballooning modes, as kink/peeling modes are stabilized when the edge current and pressure gradient shift away from the separatrix. Edge stability analysis using ELITE corroborated this picture, as reconstructed equilibria from ELM-free discharges were generally farther from their kink/peeling stability boundaries than ELMy discharges. We conclude that density profile control provided by lithium is the key first step to ELM suppression in NSTX.

  12. The Relationships Between ELM Suppression, Pedestal Profiles, and Lithium Wall Coatings in NSTX

    SciTech Connect

    D.P. Boyle, R. Maingi, P.B. Snyder, J. Manickam, T.H. Osborne, R.E. Bell, B.P. LeBlanc, and the NSTX Team

    2012-08-17

    Recently in the National Spherical Torus Experiment (NSTX), increasing lithium wall coatings suppressed edge localized modes (ELMs), gradually but not quite monotonically. This work details profile and stability analysis as ELMs disappeared throughout the lithium scan. While the quantity of lithium deposited between discharges did not uniquely determine the presence of ELMs, profile analysis demonstrated that lithium was correlated to wider density and pressure pedestals with peak gradients farther from the separatrix. Moreover, the ELMy and ELM-free discharges were cleanly separated by their density and pedestal widths and peak gradient locations. Ultimately, ELMs were only suppressed when lithium caused the density pedestal to widen and shift inward. These changes in the density gradient were directly reflected in the pressure gradient and calculated bootstrap current. This supports the theory that ELMs in NSTX are caused by peeling and/or ballooning modes, as kink/peeling modes are stabilized when the edge current and pressure gradient shift away from the separatrix. Edge stability analysis using ELITE corroborated this picture, as reconstructed equilibria from ELM-free discharges were generally farther from their kink/peeling stability boundaries than ELMy discharges. We conclude that density profile control provided by lithium is the key first step to ELM suppression in NSTX

  13. Nucleus-associated actin in Amoeba proteus.

    PubMed

    Berdieva, Mariia; Bogolyubov, Dmitry; Podlipaeva, Yuliya; Goodkov, Andrew

    2016-10-01

    The presence, spatial distribution and forms of intranuclear and nucleus-associated cytoplasmic actin were studied in Amoeba proteus with immunocytochemical approaches. Labeling with different anti-actin antibodies and staining with TRITC-phalloidin and fluorescent deoxyribonuclease I were used. We showed that actin is abundant within the nucleus as well as in the cytoplasm of A. proteus cells. According to DNase I experiments, the predominant form of intranuclear actin is G-actin which is associated with chromatin strands. Besides, unpolymerized actin was shown to participate in organization of a prominent actin layer adjacent to the outer surface of nuclear envelope. No significant amount of F-actin was found in the nucleus. At the same time, the amoeba nucleus is enclosed in a basket-like structure formed by circumnuclear actin filaments and bundles connected with global cytoplasmic actin cytoskeleton. A supposed architectural function of actin filaments was studied by treatment with actin-depolymerizing agent latrunculin A. It disassembled the circumnuclear actin system, but did not affect the intranuclear chromatin structure. The results obtained for amoeba cells support the modern concept that actin is involved in fundamental nuclear processes that have evolved in the cells of multicellular organisms.

  14. Boolean gates on actin filaments

    NASA Astrophysics Data System (ADS)

    Siccardi, Stefano; Tuszynski, Jack A.; Adamatzky, Andrew

    2016-01-01

    Actin is a globular protein which forms long polar filaments in the eukaryotic cytoskeleton. Actin networks play a key role in cell mechanics and cell motility. They have also been implicated in information transmission and processing, memory and learning in neuronal cells. The actin filaments have been shown to support propagation of voltage pulses. Here we apply a coupled nonlinear transmission line model of actin filaments to study interactions between voltage pulses. To represent digital information we assign a logical TRUTH value to the presence of a voltage pulse in a given location of the actin filament, and FALSE to the pulse's absence, so that information flows along the filament with pulse transmission. When two pulses, representing Boolean values of input variables, interact, then they can facilitate or inhibit further propagation of each other. We explore this phenomenon to construct Boolean logical gates and a one-bit half-adder with interacting voltage pulses. We discuss implications of these findings on cellular process and technological applications.

  15. The ATP binding cassette transporter, ABCG1, localizes to cortical actin filaments

    PubMed Central

    Pandzic, Elvis; Gelissen, Ingrid C.; Whan, Renee; Barter, Philip J.; Sviridov, Dmitri; Gaus, Katharina; Rye, Kerry-Anne; Cochran, Blake J.

    2017-01-01

    The ATP-binding cassette sub-family G member 1 (ABCG1) exports cellular cholesterol to high-density lipoproteins (HDL). However, a number of recent studies have suggested ABCG1 is predominantly localised to intracellular membranes. In this study, we found that ABCG1 was organized into two distinct cellular pools: one at the plasma membrane and the other associated with the endoplasmic reticulum (ER). The plasma membrane fraction was organized into filamentous structures that were associated with cortical actin filaments. Inhibition of actin polymerization resulted in complete disruption of ABCG1 filaments. Cholesterol loading of the cells increased the formation of the filamentous ABCG1, the proximity of filamentous ABCG1 to actin filaments and the diffusion rate of membrane associated ABCG1. Our findings suggest that the actin cytoskeleton plays a critical role in the plasma membrane localization of ABCG1. PMID:28165022

  16. Effects of actin-binding proteins on the thermal stability of monomeric actin.

    PubMed

    Pivovarova, Anastasia V; Chebotareva, Natalia A; Kremneva, Elena V; Lappalainen, Pekka; Levitsky, Dmitrii I

    2013-01-08

    Differential scanning calorimetry (DSC) was applied to investigate the thermal unfolding of rabbit skeletal muscle G-actin in its complexes with actin-binding proteins, cofilin, twinfilin, and profilin. The results show that the effects of these proteins on the thermal stability of G-actin depend on the nucleotide, ATP or ADP, bound in the nucleotide-binding cleft between actin subdomains 2 and 4. Interestingly, cofilin binding stabilizes both ATP-G-actin and ADP-G-actin, whereas twinfilin increases the thermal stability of the ADP-G-actin but not that of the ATP-G-actin. By contrast, profilin strongly decreases the thermal stability of the ATP-G-actin but has no appreciable effect on the ADP-G-actin. Comparison of these DSC results with literature data reveals a relationship between the effects of actin-binding proteins on the thermal unfolding of G-actin, stabilization or destabilization, and their effects on the rate of nucleotide exchange in the nucleotide-binding cleft, decrease or increase. These results suggest that the thermal stability of G-actin depends, at least partially, on the conformation of the nucleotide-binding cleft: the actin molecule is more stable when the cleft is closed, while an opening of the cleft leads to significant destabilization of G-actin. Thus, DSC studies of the thermal unfolding of G-actin can provide new valuable information about the conformational changes induced by actin-binding proteins in the actin molecule.

  17. G-actin regulates rapid induction of actin nucleation by mDia1 to restore cellular actin polymers.

    PubMed

    Higashida, Chiharu; Suetsugu, Shiro; Tsuji, Takahiro; Monypenny, James; Narumiya, Shuh; Watanabe, Naoki

    2008-10-15

    mDia1 belongs to the formin family of proteins that share FH1 and FH2 domains. Although formins play a critical role in the formation of many actin-based cellular structures, the physiological regulation of formin-mediated actin assembly within the cell is still unknown. Here we show that cells possess an acute actin polymer restoration mechanism involving mDia1. By using single-molecule live-cell imaging, we found that several treatments including low-dose G-actin-sequestering drugs and unpolymerizable actin mutants activate mDia1 to initiate fast directional movement. The FH2 region, the core domain for actin nucleation, is sufficient to respond to latrunculin B (LatB) to increase its actin nucleation frequency. Simulation analysis revealed an unexpected paradoxical effect of LatB that leads to a several fold increase in free G-actin along with an increase in total G-actin. These results indicate that in cells, the actin nucleation frequency of mDia1 is enhanced not only by Rho, but also strongly through increased catalytic efficiency of the FH2 domain. Consistently, frequent actin nucleation by mDia1 was found around sites of vigorous actin disassembly. Another major actin nucleator, the Arp2/3 complex, was not affected by the G-actin increase induced by LatB. Taken together, we propose that transient accumulation of G-actin works as a cue to promote mDia1-catalyzed actin nucleation to execute rapid reassembly of actin filaments.

  18. Evolution of intracellular pathogens.

    PubMed

    Casadevall, Arturo

    2008-01-01

    The evolution of intracellular pathogens is considered in the context of ambiguities in basic definitions and the diversity of host-microbe interactions. Intracellular pathogenesis is a subset of a larger world of host-microbe interactions that includes amoeboid predation and endosymbiotic existence. Intracellular pathogens often reveal genome reduction. Despite the uniqueness of each host-microbe interaction, there are only a few general solutions to the problem of intracellular survival, especially in phagocytic cells. Similarities in intracellular pathogenic strategies between phylogenetically distant microbes suggest convergent evolution. For discerning such patterns, it is useful to consider whether the microbe is acquired from another host or directly from the environment. For environmentally acquired microbes, biotic pressures, such as amoeboid predators, may select for the capacity for virulence. Although often viewed as a specialized adaptation, the capacity for intracellular survival may be widespread among microbes, thus questioning whether the intracellular lifestyle warrants a category of special distinctiveness.

  19. Technical advance: identification of plant actin-binding proteins by F-actin affinity chromatography

    NASA Technical Reports Server (NTRS)

    Hu, S.; Brady, S. R.; Kovar, D. R.; Staiger, C. J.; Clark, G. B.; Roux, S. J.; Muday, G. K.

    2000-01-01

    Proteins that interact with the actin cytoskeleton often modulate the dynamics or organization of the cytoskeleton or use the cytoskeleton to control their localization. In plants, very few actin-binding proteins have been identified and most are thought to modulate cytoskeleton function. To identify actin-binding proteins that are unique to plants, the development of new biochemical procedures will be critical. Affinity columns using actin monomers (globular actin, G-actin) or actin filaments (filamentous actin, F-actin) have been used to identify actin-binding proteins from a wide variety of organisms. Monomeric actin from zucchini (Cucurbita pepo L.) hypocotyl tissue was purified to electrophoretic homogeneity and shown to be native and competent for polymerization to actin filaments. G-actin, F-actin and bovine serum albumin affinity columns were prepared and used to separate samples enriched in either soluble or membrane-associated actin-binding proteins. Extracts of soluble actin-binding proteins yield distinct patterns when eluted from the G-actin and F-actin columns, respectively, leading to the identification of a putative F-actin-binding protein of approximately 40 kDa. When plasma membrane-associated proteins were applied to these columns, two abundant polypeptides eluted selectively from the F-actin column and cross-reacted with antiserum against pea annexins. Additionally, a protein that binds auxin transport inhibitors, the naphthylphthalamic acid binding protein, which has been previously suggested to associate with the actin cytoskeleton, was eluted in a single peak from the F-actin column. These experiments provide a new approach that may help to identify novel actin-binding proteins from plants.

  20. Technical advance: identification of plant actin-binding proteins by F-actin affinity chromatography

    NASA Technical Reports Server (NTRS)

    Hu, S.; Brady, S. R.; Kovar, D. R.; Staiger, C. J.; Clark, G. B.; Roux, S. J.; Muday, G. K.

    2000-01-01

    Proteins that interact with the actin cytoskeleton often modulate the dynamics or organization of the cytoskeleton or use the cytoskeleton to control their localization. In plants, very few actin-binding proteins have been identified and most are thought to modulate cytoskeleton function. To identify actin-binding proteins that are unique to plants, the development of new biochemical procedures will be critical. Affinity columns using actin monomers (globular actin, G-actin) or actin filaments (filamentous actin, F-actin) have been used to identify actin-binding proteins from a wide variety of organisms. Monomeric actin from zucchini (Cucurbita pepo L.) hypocotyl tissue was purified to electrophoretic homogeneity and shown to be native and competent for polymerization to actin filaments. G-actin, F-actin and bovine serum albumin affinity columns were prepared and used to separate samples enriched in either soluble or membrane-associated actin-binding proteins. Extracts of soluble actin-binding proteins yield distinct patterns when eluted from the G-actin and F-actin columns, respectively, leading to the identification of a putative F-actin-binding protein of approximately 40 kDa. When plasma membrane-associated proteins were applied to these columns, two abundant polypeptides eluted selectively from the F-actin column and cross-reacted with antiserum against pea annexins. Additionally, a protein that binds auxin transport inhibitors, the naphthylphthalamic acid binding protein, which has been previously suggested to associate with the actin cytoskeleton, was eluted in a single peak from the F-actin column. These experiments provide a new approach that may help to identify novel actin-binding proteins from plants.

  1. The nitrate reductase inhibitor, tungsten, disrupts actin microfilaments in Zea mays L.

    PubMed

    Adamakis, Ioannis-Dimosthenis S; Panteris, Emmanuel; Eleftheriou, Eleftherios P

    2014-05-01

    Tungsten is a widely used inhibitor of nitrate reductase, applied to diminish the nitric oxide levels in plants. It was recently shown that tungsten also has heavy metal attributes. Since information about the toxic effects of tungsten on actin is limited, and considering that actin microfilaments are involved in the entry of tungsten inside plant cells, the effects of tungsten on them were studied in Zea mays seedlings. Treatments with sodium tungstate for 3, 6, 12 or 24 h were performed on intact seedlings and seedlings with truncated roots. Afterwards, actin microfilaments in meristematic root and leaf tissues were stained with fluorescent phalloidin, and the specimens were examined by confocal laser scanning microscopy. While the actin microfilament network was well organized in untreated seedlings, in tungstate-treated ones it was disrupted in a time-dependent manner. In protodermal root cells, the effects of tungsten were stronger as cortical microfilaments were almost completely depolymerized and the intracellular ones appeared highly bundled. Fluorescence intensity measurements confirmed the above results. In the meristematic leaf tissue of intact seedlings, no depolymerization of actin microfilaments was noticed. However, when root tips were severed prior to tungstate application, both cortical and endoplasmic actin networks of leaf cells were disrupted and bundled after 24 h of treatment. The differential response of root and leaf tissues to tungsten toxicity may be due to differential penetration and absorption, while the effects on actin microfilaments could not be attributed to the nitric oxide depletion by tungsten.

  2. Choosing orientation: influence of cargo geometry and ActA polarization on actin comet tails

    PubMed Central

    Lacayo, Catherine I.; Soneral, Paula A. G.; Zhu, Jie; Tsuchida, Mark A.; Footer, Matthew J.; Soo, Frederick S.; Lu, Yu; Xia, Younan; Mogilner, Alexander; Theriot, Julie A.

    2012-01-01

    Networks of polymerizing actin filaments can propel intracellular pathogens and drive movement of artificial particles in reconstituted systems. While biochemical mechanisms activating actin network assembly have been well characterized, it remains unclear how particle geometry and large-scale force balance affect emergent properties of movement. We reconstituted actin-based motility using ellipsoidal beads resembling the geometry of Listeria monocytogenes. Beads coated uniformly with the L. monocytogenes ActA protein migrated equally well in either of two distinct orientations, with their long axes parallel or perpendicular to the direction of motion, while intermediate orientations were unstable. When beads were coated with a fluid lipid bilayer rendering ActA laterally mobile, beads predominantly migrated with their long axes parallel to the direction of motion, mimicking the orientation of motile L. monocytogenes. Generating an accurate biophysical model to account for our observations required the combination of elastic-propulsion and tethered-ratchet actin-polymerization theories. Our results indicate that the characteristic orientation of L. monocytogenes must be due to polarized ActA rather than intrinsic actin network forces. Furthermore, viscoelastic stresses, forces, and torques produced by individual actin filaments and lateral movement of molecular complexes must all be incorporated to correctly predict large-scale behavior in the actin-based movement of nonspherical particles. PMID:22219381

  3. Actin Cytoskeleton Manipulation by Effector Proteins Secreted by Diarrheagenic Escherichia coli Pathotypes

    PubMed Central

    Navarro-Garcia, Fernando; Serapio-Palacios, Antonio; Ugalde-Silva, Paul; Tapia-Pastrana, Gabriela; Chavez-Dueñas, Lucia

    2013-01-01

    The actin cytoskeleton is a dynamic structure necessary for cell and tissue organization, including the maintenance of epithelial barriers. Disruption of the epithelial barrier coincides with alterations of the actin cytoskeleton in several disease states. These disruptions primarily affect the paracellular space, which is normally regulated by tight junctions. Thereby, the actin cytoskeleton is a common and recurring target of bacterial virulence factors. In order to manipulate the actin cytoskeleton, bacteria secrete and inject toxins and effectors to hijack the host cell machinery, which interferes with host-cell pathways and with a number of actin binding proteins. An interesting model to study actin manipulation by bacterial effectors is Escherichia coli since due to its genome plasticity it has acquired diverse genetic mobile elements, which allow having different E. coli varieties in one bacterial species. These E. coli pathotypes, including intracellular and extracellular bacteria, interact with epithelial cells, and their interactions depend on a specific combination of virulence factors. In this paper we focus on E. coli effectors that mimic host cell proteins to manipulate the actin cytoskeleton. The study of bacterial effector-cytoskeleton interaction will contribute not only to the comprehension of the molecular causes of infectious diseases but also to increase our knowledge of cell biology. PMID:23509714

  4. The Actin Filament-Binding Protein Coronin Regulates Motility in Plasmodium Sporozoites

    PubMed Central

    Bane, Kartik S.; Singer, Mirko; Reinig, Miriam; Klug, Dennis; Heiss, Kirsten; Baum, Jake; Mueller, Ann-Kristin; Frischknecht, Friedrich

    2016-01-01

    Parasites causing malaria need to migrate in order to penetrate tissue barriers and enter host cells. Here we show that the actin filament-binding protein coronin regulates gliding motility in Plasmodium berghei sporozoites, the highly motile forms of a rodent malaria-causing parasite transmitted by mosquitoes. Parasites lacking coronin show motility defects that impair colonization of the mosquito salivary glands but not migration in the skin, yet result in decreased transmission efficiency. In non-motile sporozoites low calcium concentrations mediate actin-independent coronin localization to the periphery. Engagement of extracellular ligands triggers an intracellular calcium release followed by the actin-dependent relocalization of coronin to the rear and initiation of motility. Mutational analysis and imaging suggest that coronin organizes actin filaments for productive motility. Using coronin-mCherry as a marker for the presence of actin filaments we found that protein kinase A contributes to actin filament disassembly. We finally speculate that calcium and cAMP-mediated signaling regulate a switch from rapid parasite motility to host cell invasion by differentially influencing actin dynamics. PMID:27409081

  5. Bacterial Actins? An Evolutionary Perspective

    NASA Technical Reports Server (NTRS)

    Doolittle, Russell F.; York, Amanda L.

    2003-01-01

    According to the conventional wisdom, the existence of a cytoskeleton in eukaryotes and its absence in prokaryotes constitute a fundamental divide between the two domains of life. An integral part of the dogma is that a cytoskeleton enabled an early eukaryote to feed upon prokaryotes, a consequence of which was the occasional endosymbiosis and the eventual evolution of organelles. Two recent papers present compelling evidence that actin, one of the principal components of a cytoskeleton, has a homolog in Bacteria that behaves in many ways like eukaryotic actin. Sequence comparisons reveml that eukaryotic actin and the bacterial homolog (mreB protein), unlike many other proteins common to eukaryotes and Bacteria, have very different and more highly extended evolutionary histories.

  6. Bacterial Actins? An Evolutionary Perspective

    NASA Technical Reports Server (NTRS)

    Doolittle, Russell F.; York, Amanda L.

    2003-01-01

    According to the conventional wisdom, the existence of a cytoskeleton in eukaryotes and its absence in prokaryotes constitute a fundamental divide between the two domains of life. An integral part of the dogma is that a cytoskeleton enabled an early eukaryote to feed upon prokaryotes, a consequence of which was the occasional endosymbiosis and the eventual evolution of organelles. Two recent papers present compelling evidence that actin, one of the principal components of a cytoskeleton, has a homolog in Bacteria that behaves in many ways like eukaryotic actin. Sequence comparisons reveml that eukaryotic actin and the bacterial homolog (mreB protein), unlike many other proteins common to eukaryotes and Bacteria, have very different and more highly extended evolutionary histories.

  7. Impact of impurity seeding and divertor conditions on transitions, pedestal structure and ELMs

    NASA Astrophysics Data System (ADS)

    Dunne, M. G.

    2017-02-01

    Future devices will require a high scrape-off later (SOL) density and impurity seeding to avoid high-Z sputtering. However, these operational parameters are not included in present-day scaling laws, making extrapolations to larger devices difficult. As such, understanding the physics of such effects is vital in order to design the operational scenarios most favorable to high fusion gain. This review presents the favorable lowering of L-H transition power by changing to metal walled devices and sumarises the effects currently thought to be responsible for how SOL geometry can play a role in determining this threshold. Experimental observations on changes to the pedestal structure with main ion fuelling and low-, medium-, and high-Z impurity seeding are presented. These results, from several devices, show that main ion fuelling or high density operation can result in a lower pedestal top pressure, and hence reduced stored energy, while impurity seeding can recover this lost pressure. Particular focus is given to nitrogen seeded discharges and the recovery of pedestal parameters (notably high {{T}\\text{e,\\text{ped}}} ) in JET and AUG since the changeover to metal walls in these devices. Lithium seeding is also emerging as a strong actuator in pedestal dynamics, with results ranging from a prolonged inter-ELM period to completely ELM-free scenarios on different devices. ELM dynamics are also presented in each section, with nitrogen seeding offering a probe into the structure of the ELM and demonstrating the difference between the initial ELM crash, likely due to a sharp MHD event, and a prolonged second phase, the origin of which remains unkown. Finally, modelling of the pedestal in impurity seeded scenarios reveals a common effect in the position of the density profile. Either through mode excitation near to the separatrix or an altered fuelling profile, seeding of impurities results in an inward shift of the density profile. This inward shift improves MHD stability

  8. Effects of solution crowding on actin polymerization reveal the energetic basis for nucleotide-dependent filament stability

    PubMed Central

    Frederick, Kendra B.; Sept, David; De La Cruz, Enrique M.

    2008-01-01

    Actin polymerization is a fundamental cellular process involved in cell structure maintenance, force generation, and motility. Phosphate release from filament subunits following ATP hydrolysis destabilizes the filament lattice and increases the critical concentration (Cc) for assembly. The structural differences between ATP- and ADP-actin are still debated, as well as the energetic factors that underlie nucleotide-dependent filament stability, particularly under crowded intracellular conditions. Here, we investigate the effect of crowding agents on ATP- and ADP-actin polymerization, and find that ATP-actin polymerization is largely unaffected by solution crowding, while crowding agents lower the Cc of ADP-actin in a concentration-dependent manner. The stabilities of ATP- and ADP-actin filaments are comparable in the presence of physiological amounts (~30% w/v) and types (sorbitol) of low molecular weight crowding agents. Crowding agents act to stabilize ADP-F-actin by slowing subunit dissociation. These observations suggest that nucleotide hydrolysis and phosphate release per se do not introduce intrinsic differences in the in vivo filament stability. Rather, the preferential disassembly of ADP-actin filaments in cells is driven through interactions with regulatory proteins. Interpretation of the experimental data according to osmotic stress theory implicates water as an allosteric regulator of actin activity and hydration as the molecular basis for nucleotide-dependent filament stability. PMID:18374941

  9. Regulation of the actin cycle in vivo by actin filament severing

    PubMed Central

    McGrath, James L.; Osborn, Eric A.; Tardy, Yanik S.; Dewey, C. Forbes; Hartwig, John H.

    2000-01-01

    Cycling of actin subunits between monomeric and filamentous phases is essential for cell crawling behavior. We investigated actin filament turnover rates, length, number, barbed end exposure, and binding of cofilin in bovine arterial endothelial cells moving at different speeds depending on their position in a confluent monolayer. Fast-translocating cells near the wound edge have short filament lifetimes compared with turnover values that proportionately increase in slower moving cells situated at increasing distances from the wound border. Contrasted with slow cells exhibiting slow actin filament turnover speeds, fast cells have less polymerized actin, shorter actin filaments, more free barbed ends, and less actin-associated cofilin. Cultured primary fibroblasts manifest identical relationships between speed and actin turnover as the endothelial cells, and fast fibroblasts expressing gelsolin have higher actin turnover rates than slow fibroblasts that lack this actin-severing protein. These results implicate actin filament severing as an important control mechanism for actin cycling in cells. PMID:10823888

  10. Actin polymerization is stimulated by actin cross-linking protein palladin.

    PubMed

    Gurung, Ritu; Yadav, Rahul; Brungardt, Joseph G; Orlova, Albina; Egelman, Edward H; Beck, Moriah R

    2016-02-15

    The actin scaffold protein palladin regulates both normal cell migration and invasive cell motility, processes that require the co-ordinated regulation of actin dynamics. However, the potential effect of palladin on actin dynamics has remained elusive. In the present study, we show that the actin-binding immunoglobulin-like domain of palladin, which is directly responsible for both actin binding and bundling, also stimulates actin polymerization in vitro. Palladin eliminated the lag phase that is characteristic of the slow nucleation step of actin polymerization. Furthermore, palladin dramatically reduced depolymerization, slightly enhanced the elongation rate, and did not alter the critical concentration. Microscopy and in vitro cross-linking assays reveal differences in actin bundle architecture when palladin is incubated with actin before or after polymerization. These results suggest a model whereby palladin stimulates a polymerization-competent form of globular or monomeric actin (G-actin), akin to metal ions, either through charge neutralization or through conformational changes.

  11. Optimal treatment of actinic keratoses

    PubMed Central

    Uhlenhake, Elizabeth E

    2013-01-01

    The most compelling reason and primary goal of treating actinic keratoses is to prevent malignant transformation into invasive squamous cell carcinoma, and although there are well established guidelines outlining treatment modalities and regimens for squamous cell carcinoma, the more commonly encountered precancerous actinic lesions have no such standard. Many options are available with variable success and patient compliance rates. Prevention of these lesions is key, with sun protection being a must in treating aging patients with sun damage as it is never too late to begin protecting the skin. PMID:23345970

  12. Formation and Destabilization of Actin Filaments with Tetramethylrhodamine-Modified Actin

    PubMed Central

    Kudryashov, Dmitry S.; Phillips, Martin; Reisler, Emil

    2004-01-01

    Actin labeling at Cys374 with tethramethylrhodamine derivatives (TMR-actin) has been widely used for direct observation of the in vitro filaments growth, branching, and treadmilling, as well as for the in vivo visualization of actin cytoskeleton. The advantage of TMR-actin is that it does not lock actin in filaments (as rhodamine-phalloidin does), possibly allowing for its use in investigating the dynamic assembly behavior of actin polymers. Although it is established that TMR-actin alone is polymerization incompetent, the impact of its copolymerization with unlabeled actin on filament structure and dynamics has not been tested yet. In this study, we show that TMR-actin perturbs the filaments structure when copolymerized with unlabeled actin; the resulting filaments are more fragile and shorter than the control filaments. Due to the increased severing of copolymer filaments, TMR-actin accelerates the polymerization of unlabeled actin in solution also at mole ratios lower than those used in most fluorescence microscopy experiments. The destabilizing and severing effect of TMR-actin is countered by filament stabilizing factors, phalloidin, S1, and tropomyosin. These results point to an analogy between the effects of TMR-actin and severing proteins on F-actin, and imply that TMR-actin may be inappropriate for investigations of actin filaments dynamics. PMID:15298916

  13. Fascin regulates nuclear actin during Drosophila oogenesis

    PubMed Central

    Kelpsch, Daniel J.; Groen, Christopher M.; Fagan, Tiffany N.; Sudhir, Sweta; Tootle, Tina L.

    2016-01-01

    Drosophila oogenesis provides a developmental system with which to study nuclear actin. During Stages 5–9, nuclear actin levels are high in the oocyte and exhibit variation within the nurse cells. Cofilin and Profilin, which regulate the nuclear import and export of actin, also localize to the nuclei. Expression of GFP-tagged Actin results in nuclear actin rod formation. These findings indicate that nuclear actin must be tightly regulated during oogenesis. One factor mediating this regulation is Fascin. Overexpression of Fascin enhances nuclear GFP-Actin rod formation, and Fascin colocalizes with the rods. Loss of Fascin reduces, whereas overexpression of Fascin increases, the frequency of nurse cells with high levels of nuclear actin, but neither alters the overall nuclear level of actin within the ovary. These data suggest that Fascin regulates the ability of specific cells to accumulate nuclear actin. Evidence indicates that Fascin positively regulates nuclear actin through Cofilin. Loss of Fascin results in decreased nuclear Cofilin. In addition, Fascin and Cofilin genetically interact, as double heterozygotes exhibit a reduction in the number of nurse cells with high nuclear actin levels. These findings are likely applicable beyond Drosophila follicle development, as the localization and functions of Fascin and the mechanisms regulating nuclear actin are widely conserved. PMID:27535426

  14. Stationary QH-mode plasmas with high and wide pedestal at low rotation on DIII-D

    NASA Astrophysics Data System (ADS)

    Chen, Xi; Burrell, K. H.; Osborne, T. H.; Solomon, W. M.; Barada, K.; Garofalo, A. M.; Groebner, R. J.; Luhmann, N. C.; McKee, G. R.; Muscatello, C. M.; Ono, M.; Petty, C. C.; Porkolab, M.; Rhodes, T. L.; Rost, J. C.; Snyder, P. B.; Staebler, G. M.; Tobias, B. J.; Yan, Z.; the DIII-D Team

    2017-02-01

    A stationary, quiescent H-mode (QH-mode) regime with a wide pedestal and improved confinement at low rotation has been discovered on DIII-D with reactor relevant edge parameters and no ELMs. As the injected neutral beam torque is ramped down and the edge E  ×  B rotation shear reduces, the transition from standard QH to the wide pedestal QH-mode occurs. At the transition, the coherent edge harmonic oscillations (EHO) that usually regulate the standard QH edge cease and broadband edge MHD modes appear along with a rapid increase in the pedestal pressure height (by  ⩽60%) and width (by  ⩽50%). We posit that the enhanced edge turbulence-driven transport, enabled by the lower edge E  ×  B flow shear due to lower torque reduces the pedestal gradient and, combined with the high edge instability limit provided by the balanced double-null plasma shape, permits the development of a broader and thus higher pedestal that is turbulence-transport-limited. Even with the significantly enhanced pedestal pressure, the edge operating point is below the peeling ballooning mode stability boundary and thus without ELMs. Improved transport in the outer core region (0.8  ⩽  ρ  ⩽0.9) owing to increased E  ×  B flow shear in that region and the enhanced pedestal boost the overall confinement by up to 45%. These findings advance the physics basis for developing stationary ELM-free high-confinement operation at low rotation for future burning plasma where similar collisionality and rotation levels are expected.

  15. Stationary QH-mode plasmas with high and wide pedestal at low rotation on DIII-D

    DOE PAGES

    Chen, Xi; Burrell, K. H.; Osborne, T. H.; ...

    2016-09-30

    A stationary, quiescent H-mode (QH-mode) regime with a wide pedestal and improved confinement at low rotation has been discovered on DIII-D with reactor relevant edge parameters and no ELMs. As the injected neutral beam torque is ramped down and the edge ExB rotation shear reduces, the transition from standard QH to the wide pedestal QH-mode occurs. And at the transition, the coherent edge harmonic oscillations (EHO) that usually regulate the standard QH edge cease and broadband edge MHD modes appear along with a rapid increase in the pedestal pressure height (by ≤60%) and width (by ≤50%). We posit that themore » enhanced edge turbulence-driven transport, enabled by the lower edge ExB flow shear due to lower torque reduces the pedestal gradient and, combined with the high edge instability limit provided by the balanced double-null plasma shape, permits the development of a broader and thus higher pedestal that is turbulence-transport-limited. Even with the significantly enhanced pedestal pressure, the edge operating point is below the peeling ballooning mode stability boundary and thus without ELMs. Improved transport in the outer core region (0.8≤ρ≤0.9) owing to increased ExB flow shear in that region and the enhanced pedestal boost the overall confinement by up to 45%. Our findings advance the physics basis for developing stationary ELM-free high-confinement operation at low rotation for future burning plasma where similar collisionality and rotation levels are expected.« less

  16. Stationary QH-mode plasmas with high and wide pedestal at low rotation on DIII-D

    SciTech Connect

    Chen, Xi; Burrell, K. H.; Osborne, T. H.; Solomon, W. M.; Barada, K.; Garofalo, A. M.; Groebner, R. J.; Luhmann, N. C.; McKee, G. R.; Muscatello, C. M.; Ono, M.; Petty, C. C.; Porkolab, M.; Rhodes, T. L.; Rost, J. C.; Snyder, P. B.; Staebler, G. M.; Tobias, B. J.; Yan, Z.

    2016-09-30

    A stationary, quiescent H-mode (QH-mode) regime with a wide pedestal and improved confinement at low rotation has been discovered on DIII-D with reactor relevant edge parameters and no ELMs. As the injected neutral beam torque is ramped down and the edge ExB rotation shear reduces, the transition from standard QH to the wide pedestal QH-mode occurs. And at the transition, the coherent edge harmonic oscillations (EHO) that usually regulate the standard QH edge cease and broadband edge MHD modes appear along with a rapid increase in the pedestal pressure height (by ≤60%) and width (by ≤50%). We posit that the enhanced edge turbulence-driven transport, enabled by the lower edge ExB flow shear due to lower torque reduces the pedestal gradient and, combined with the high edge instability limit provided by the balanced double-null plasma shape, permits the development of a broader and thus higher pedestal that is turbulence-transport-limited. Even with the significantly enhanced pedestal pressure, the edge operating point is below the peeling ballooning mode stability boundary and thus without ELMs. Improved transport in the outer core region (0.8≤ρ≤0.9) owing to increased ExB flow shear in that region and the enhanced pedestal boost the overall confinement by up to 45%. Our findings advance the physics basis for developing stationary ELM-free high-confinement operation at low rotation for future burning plasma where similar collisionality and rotation levels are expected.

  17. Strategies for Intracellular Survival of Burkholderia pseudomallei

    PubMed Central

    Allwood, Elizabeth M.; Devenish, Rodney J.; Prescott, Mark; Adler, Ben; Boyce, John D.

    2011-01-01

    Burkholderia pseudomallei is the causative agent of melioidosis, a disease with high mortality that is prevalent in tropical regions of the world. A key component of the pathogenesis of melioidosis is the ability of B. pseudomallei to enter, survive, and replicate within mammalian host cells. For non-phagocytic cells, bacterial adhesins have been identified both on the bacterial surface and associated with Type 4 pili. Cell invasion involves components of one or more of the three Type 3 Secretion System clusters, which also mediate, at least in part, the escape of bacteria from the endosome into the cytoplasm, where bacteria move by actin-based motility. The mechanism of actin-based motility is not clearly understood, but appears to differ from characterized mechanisms in other bacterial species. A small proportion of intracellular bacteria is targeted by host cell autophagy, involving direct recruitment of LC3 to endosomes rather than through uptake by canonical autophagosomes. However, the majority of bacterial cells are able to circumvent autophagy and other intracellular defense mechanisms such as the induction of inducible nitric oxide synthase, and then replicate in the cytoplasm and spread to adjacent cells via membrane fusion, resulting in the formation of multi-nucleated giant cells. A potential role for host cell ubiquitin in the autophagic response to bacterial infection has recently been proposed. PMID:22007185

  18. Proinflammatory cytokines provoke oxidative damage to actin in neuronal cells mediated by Rac1 and NADPH oxidase.

    PubMed

    Barth, Brian M; Stewart-Smeets, Shelli; Kuhn, Thomas B

    2009-06-01

    The proinflammatory cytokines TNFalpha and Il-1beta orchestrate the progression of CNS inflammation, which substantially contributes to neurodegeneration in many CNS pathologies. TNFalpha and Il-1beta stimulate actin filament reorganization in non-neuronal cells often accompanied by the formation of reactive oxygen species (ROS). Actin filament dynamics is vital for cellular plasticity, mitochondrial function, and gene expression despite being highly susceptible to oxidative damage. We demonstrated that, in neuronal cells, TNFalpha and Il-1beta stimulate a transient, redox-dependent reorganization of the actin cytoskeleton into lamellipodia under the regulation of Rac1 and a neuronal NADPH oxidase as the source of ROS. The persistent presence of intracellular ROS provoked oxidative damage (carbonylation) to actin coinciding with the loss of lamellipodia and arrest of cellular plasticity. Inhibition of NADPH oxidase activity or Rac1 abolished the adverse effects of cytokines. These findings suggest that oxidative damage to the neuronal actin cytoskeleton could represent a key step in CNS neurodegeneration.

  19. Three-dimensional structure of actin filaments and of an actin gel made with actin-binding protein

    PubMed Central

    1983-01-01

    Purified muscle actin and mixtures of actin and actin-binding protein were examined in the transmission electron microscope after fixation, critical point drying, and rotary shadowing. The three-dimensional structure of the protein assemblies was analyzed by a computer-assisted graphic analysis applicable to generalized filament networks. This analysis yielded information concerning the frequency of filament intersections, the filament length between these intersections, the angle at which filaments branch at these intersections, and the concentration of filaments within a defined volume. Purified actin at a concentration of 1 mg/ml assembled into a uniform mass of long filaments which overlap at random angles between 0 degrees and 90 degrees. Actin in the presence of macrophage actin-binding protein assembled into short, straight filaments, organized in a perpendicular branching network. The distance between branch points was inversely related to the molar ratio of actin-binding protein to actin. This distance was what would be predicted if actin filaments grew at right angles off of nucleation sites on the two ends of actin-binding protein dimers, and then annealed. The results suggest that actin in combination with actin-binding protein self-assembles to form a three- dimensional network resembling the peripheral cytoskeleton of motile cells. PMID:6682423

  20. Three-dimensional structure of actin filaments and of an actin gel made with actin-binding protein.

    PubMed

    Niederman, R; Amrein, P C; Hartwig, J

    1983-05-01

    Purified muscle actin and mixtures of actin and actin-binding protein were examined in the transmission electron microscope after fixation, critical point drying, and rotary shadowing. The three-dimensional structure of the protein assemblies was analyzed by a computer-assisted graphic analysis applicable to generalized filament networks. This analysis yielded information concerning the frequency of filament intersections, the filament length between these intersections, the angle at which filaments branch at these intersections, and the concentration of filaments within a defined volume. Purified actin at a concentration of 1 mg/ml assembled into a uniform mass of long filaments which overlap at random angles between 0 degrees and 90 degrees. Actin in the presence of macrophage actin-binding protein assembled into short, straight filaments, organized in a perpendicular branching network. The distance between branch points was inversely related to the molar ratio of actin-binding protein to actin. This distance was what would be predicted if actin filaments grew at right angles off of nucleation sites on the two ends of actin-binding protein dimers, and then annealed. The results suggest that actin in combination with actin-binding protein self-assembles to form a three-dimensional network resembling the peripheral cytoskeleton of motile cells.

  1. Regulation of Actin Cytoskeleton Dynamics in Cells

    PubMed Central

    Lee, Sung Haeng; Dominguez, Roberto

    2014-01-01

    The dynamic remolding of the actin cytoskeleton is a critical part of most cellular activities, and malfunction of cytoskeletal proteins results in various human diseases. The transition between two forms of actin, monomeric or G-actin and filamentous or F-actin, is tightly regulated in time and space by a large number of signaling, scaffolding and actin-binding proteins (ABPs). New ABPs are constantly being discovered in the post-genomic era. Most of these proteins are modular, integrating actin binding, protein-protein interaction, membrane-binding, and signaling domains. In response to extracellular signals, often mediated by Rho family GTPases, ABPs control different steps of actin cytoskeleton assembly, including filament nucleation, elongation, severing, capping, and depolymerization. This review summarizes structure-function relationships among ABPs in the regulation of actin cytoskeleton assembly. PMID:20446344

  2. Red light, Phot1 and JAC1 modulate Phot2-dependent reorganization of chloroplast actin filaments and chloroplast avoidance movement.

    PubMed

    Ichikawa, Satoshi; Yamada, Noboru; Suetsugu, Noriyuki; Wada, Masamitsu; Kadota, Akeo

    2011-08-01

    The phototropin (phot)-dependent intracellular relocation of chloroplasts is a ubiquitous phenomenon in plants. We have previously revealed the involvement of a short cp-actin (chloroplast actin) filament-based mechanism in this movement. Here, the reorganization of cp-actin filaments during the avoidance movement of chloroplasts was analyzed in higher time resolution under blue GFP (green fluorescent protein) excitation light in an actin filament-visualized line of Arabidopsis thaliana. Under standard background red light of 89 μmol m(-2) s(-1), cp-actin filaments transiently disappeared at approximately 30 s and reappeared in a biased configuration on chloroplasts approximately 70 s after blue excitation light irradiation. The timing of biased cp-actin reappearance was delayed under the background of strong red light or in the absence of red light. Consistently, chloroplast movement was delayed under these conditions. In phot1 mutants, acceleration of both the disappearance and reappearance of cp-actin filaments occurred, indicating an inhibitory action of phot1 on reorganization of cp-actin filaments. Avoidance movements began sooner in phot1 than in wild-type plants. No reorganization of cp-actin filaments was seen in phot2 or phot1phot2 mutants lacking phot2, which is responsible for avoidance movements. Surprisingly, jac1 (j-domain protein required for chloroplast accumulation response 1) mutants, lacking the accumulation response, showed no avoidance movements under the whole-cell irradiation condition for GFP observation. Cp-actin filaments in jac1 did not show a biased distribution, with a small or almost no transient decrease in the number. These results indicate a close association between the biased distribution of cp-actin filaments and chloroplast movement. Further, JAC1 is suggested to function in the biased cp-actin filament distribution by regulating their appearance and disappearance.

  3. Nuclear F-actin enhances the transcriptional activity of β-catenin by increasing its nuclear localization and binding to chromatin.

    PubMed

    Yamazaki, Shota; Yamamoto, Koji; de Lanerolle, Primal; Harata, Masahiko

    2016-04-01

    Actin plays multiple roles both in the cytoplasm and in the nucleus. Cytoplasmic actin, in addition to its structural role in the cytoskeleton, also contributes to the subcellular localization of transcription factors by interacting with them or their partners. The transcriptional cofactor β-catenin, which acts as an intracellular transducer of canonical Wnt signaling, indirectly associates with the cytoplasmic filamentous actin (F-actin). Recently, it has been observed that F-actin is transiently formed within the nucleus in response to serum stimulation and integrin signaling, and also during gene reprogramming. Despite these earlier observations, information about the function of nuclear F-actin is poorly defined. Here, by facilitating the accumulation of nuclear actin artificially, we demonstrate that polymerizing nuclear actin enhanced the nuclear accumulation and transcriptional function of β-catenin. Our results also show that the nuclear F-actin colocalizes with β-catenin and enhances the binding of β-catenin to the downstream target genes of the Wnt/β-catenin signaling pathway, including the genes for the cell cycle regulators c-myc and cyclin D, and the OCT4 gene. Nuclear F-actin itself also associated with these genes. Since Wnt/β-catenin signaling has important roles in cell differentiation and pluripotency, our observations suggest that nuclear F-actin formed during these biological processes is involved in regulating Wnt/β-catenin signaling.

  4. Actin Filament Cables in Drosophila Nurse Cells Are Composed of Modules That Slide Passively Past One Another during Dumping

    PubMed Central

    Guild, Gregory M.; Connelly, Patricia S.; Shaw, Michael K.; Tilney, Lewis G.

    1997-01-01

    At a late stage in Drosophila oogenesis, nurse cells rapidly expel their cytoplasm into the oocyte via intracellular bridges by a process called nurse cell dumping. Before dumping, numerous cables composed of actin filaments appear in the cytoplasm and extend inward from the plasma membrane toward the nucleus. This actin cage prevents the nucleus, which becomes highly lobed, from physically blocking the intracellular bridges during dumping. Each cable is composed of a linear series of modules composed of ∼25 cross-linked actin filaments. Adjacent modules overlap in the cable like the units of an extension ladder. During cable formation, individual modules are nucleated from the cell surface as microvilli, released, and then cross-linked to an adjacent forming module. The filaments in all the modules in a cable are unidirectionally polarized. During dumping as the volume of the cytoplasm decreases, the nucleus to plasma membrane distance decreases, compressing the actin cables that shorten as adjacent modules slide passively past one another just as the elements of an extension ladder slide past one another for storage. In Drosophila, the modular construction of actin cytoskeletons seems to be a generalized strategy. The behavior of modular actin cytoskeletons has implications for other actin-based cytoskeletal systems, e.g., those involved in Listeria movement, in cell spreading, and in retrograde flow in growth cones and fibroblasts. PMID:9265646

  5. Actinic cheilitis in dental practice.

    PubMed

    Savage, N W; McKay, C; Faulkner, C

    2010-06-01

    Actinic cheilitis is a potentially premalignant condition involving predominantly the vermilion of the lower lip. The aim of the current paper was to review the clinical presentation of actinic cheilitis and demonstrate the development of management plans using a series of cases. These are designed to provide immediate treatment where required but also to address the medium and long-term requirements of the patient. The authors suggest that the clinical examination of lips and the assessment of actinic cheilitis and other lip pathology become a regular part of the routine soft tissue examination undertaken as a part of the periodic examination of dental patients. Early recognition of actinic cheilitis can allow the development of strategies for individual patients that prevent progression. These are based on past sun exposure, future lifestyle changes and the daily use of emollient sunscreens, broad-brimmed hats and avoidance of sun exposure during the middle of the day. This is a service that is not undertaken as a matter of routine in general medical practice as patients are not seen with the regularity of dental patients and generally not under the ideal examination conditions available in the dental surgery.

  6. Plant actin controls membrane permeability.

    PubMed

    Hohenberger, Petra; Eing, Christian; Straessner, Ralf; Durst, Steffen; Frey, Wolfgang; Nick, Peter

    2011-09-01

    The biological effects of electric pulses with low rise time, high field strength, and durations in the nanosecond range (nsPEFs) have attracted considerable biotechnological and medical interest. However, the cellular mechanisms causing membrane permeabilization by nanosecond pulsed electric fields are still far from being understood. We investigated the role of actin filaments for membrane permeability in plant cells using cell lines where different degrees of actin bundling had been introduced by genetic engineering. We demonstrate that stabilization of actin increases the stability of the plasma membrane against electric permeabilization recorded by penetration of Trypan Blue into the cytoplasm. By use of a cell line expressing the actin bundling WLIM domain under control of an inducible promotor we can activate membrane stabilization by the glucocorticoid analog dexamethasone. By total internal reflection fluorescence microscopy we can visualize a subset of the cytoskeleton that is directly adjacent to the plasma membrane. We conclude that this submembrane cytoskeleton stabilizes the plasma membrane against permeabilization through electric pulses. Copyright © 2011 Elsevier B.V. All rights reserved.

  7. Force of an Actin Spring

    PubMed Central

    Shin, Jennifer H.; Tam, Barney K.; Brau, Ricardo R.; Lang, Matthew J.; Mahadevan, L.; Matsudaira, Paul

    2007-01-01

    Cellular movements are produced by forces. Typically, cytoskeletal proteins such as microtubules and actin filaments generate forces via polymerization or in conjunction with molecular motors. However, the fertilization of a Limulus polyphemus egg involves a third type of actin-based cellular engine—a biological spring. During the acrosome reaction, a 60-μm long coiled and twisted bundle of actin filaments straightens and extends from a sperm cell, penetrating the vitelline layer surrounding the egg. A subtle overtwist of 0.2°/subunit underlies the mechanochemical basis for the extension of this actin spring. Upon calcium activation, this conformational strain energy is converted to mechanical work, generating the force required to extend the bundle through the vitelline layer. In this article, we stall the extension of the acrosome bundle in agarose gels of different concentrations. From the stall forces, we estimate a maximum force of 2 nN and a puncturing pressure of 1.6 MPa. We show the maximum force of extension is three times larger than the force required to puncture the vitelline layer. Thus, the elastic strain energy stored in the acrosome bundle is more than sufficient to power the acrosome reaction through the egg envelope. PMID:17351007

  8. Antibodies to Actin in Autoimmune Neutropenia

    DTIC Science & Technology

    1990-02-01

    protein as actin. Purified Acanthamoeba actin by anti-neutrophil antibodies in autoimmune neutropenia, comigrated with the protein and was specifically...anti-rabbit IgG were obtained from ICN Immunobiolog- formed using purified Acanthamoeba actin (gift of Dr Blair Bowers. icals, Naperville, IL. Cells...preparations𔃼 1 - was the protein recognized by these anti-neutrophil antibody 6 .2- positive sera, lgG, and F(ab’) 2. Purified Acanthamoeba actin

  9. Association of actin with alpha crystallins

    NASA Technical Reports Server (NTRS)

    Gopalakrishnan, S.; Boyle, D.; Takemoto, L.; Spooner, B. S. (Principal Investigator)

    1993-01-01

    The alpha crystallins are cytosolic proteins that co-localize and co-purify with actin-containing microfilaments. Affinity column chromatography employing both covalently-coupled actin or alpha crystallin was used to demonstrate specific and saturable binding of actin with alpha crystallin. This conclusion was confirmed by direct visualization of alpha aggregates bound to actin polymerized in vitro. The significance of this interaction in relation to the functional properties of these two polypeptides will be discussed.

  10. An actin cytoskeleton with evolutionarily conserved functions in the absence of canonical actin-binding proteins

    PubMed Central

    Paredez, Alexander R.; Assaf, Zoe June; Sept, David; Timofejeva, Ljudmilla; Dawson, Scott C.; Wang, Chung-Ju Rachel; Cande, W. Z.

    2011-01-01

    Giardia intestinalis, a human intestinal parasite and member of what is perhaps the earliest-diverging eukaryotic lineage, contains the most divergent eukaryotic actin identified to date and is the first eukaryote known to lack all canonical actin-binding proteins (ABPs). We sought to investigate the properties and functions of the actin cytoskeleton in Giardia to determine whether Giardia actin (giActin) has reduced or conserved roles in core cellular processes. In vitro polymerization of giActin produced filaments, indicating that this divergent actin is a true filament-forming actin. We generated an anti-giActin antibody to localize giActin throughout the cell cycle. GiActin localized to the cortex, nuclei, internal axonemes, and formed C-shaped filaments along the anterior of the cell and a flagella-bundling helix. These structures were regulated with the cell cycle and in encysting cells giActin was recruited to the Golgi-like cyst wall processing vesicles. Knockdown of giActin demonstrated that giActin functions in cell morphogenesis, membrane trafficking, and cytokinesis. Additionally, Giardia contains a single G protein, giRac, which affects the Giardia actin cytoskeleton independently of known target ABPs. These results imply that there exist ancestral and perhaps conserved roles for actin in core cellular processes that are independent of canonical ABPs. Of medical significance, the divergent giActin cytoskeleton is essential and commonly used actin-disrupting drugs do not depolymerize giActin structures. Therefore, the giActin cytoskeleton is a promising drug target for treating giardiasis, as we predict drugs that interfere with the Giardia actin cytoskeleton will not affect the mammalian host. PMID:21444821

  11. An actin cytoskeleton with evolutionarily conserved functions in the absence of canonical actin-binding proteins.

    PubMed

    Paredez, Alexander R; Assaf, Zoe June; Sept, David; Timofejeva, Ljudmilla; Dawson, Scott C; Wang, Chung-Ju Rachel; Cande, W Z

    2011-04-12

    Giardia intestinalis, a human intestinal parasite and member of what is perhaps the earliest-diverging eukaryotic lineage, contains the most divergent eukaryotic actin identified to date and is the first eukaryote known to lack all canonical actin-binding proteins (ABPs). We sought to investigate the properties and functions of the actin cytoskeleton in Giardia to determine whether Giardia actin (giActin) has reduced or conserved roles in core cellular processes. In vitro polymerization of giActin produced filaments, indicating that this divergent actin is a true filament-forming actin. We generated an anti-giActin antibody to localize giActin throughout the cell cycle. GiActin localized to the cortex, nuclei, internal axonemes, and formed C-shaped filaments along the anterior of the cell and a flagella-bundling helix. These structures were regulated with the cell cycle and in encysting cells giActin was recruited to the Golgi-like cyst wall processing vesicles. Knockdown of giActin demonstrated that giActin functions in cell morphogenesis, membrane trafficking, and cytokinesis. Additionally, Giardia contains a single G protein, giRac, which affects the Giardia actin cytoskeleton independently of known target ABPs. These results imply that there exist ancestral and perhaps conserved roles for actin in core cellular processes that are independent of canonical ABPs. Of medical significance, the divergent giActin cytoskeleton is essential and commonly used actin-disrupting drugs do not depolymerize giActin structures. Therefore, the giActin cytoskeleton is a promising drug target for treating giardiasis, as we predict drugs that interfere with the Giardia actin cytoskeleton will not affect the mammalian host.

  12. Actin crosslinkers: repairing the sense of touch.

    PubMed

    Sun, Sean X; Walcott, Sam

    2010-10-26

    Cells use actin bundles infused with myosin to exert contractile forces on the extracellular environment. This active tension is essential for cellular mechanosensation. Now, the role of actin crosslinkers in stabilizing and repairing the actin bundles is coming into clearer view.

  13. Linear gyrokinetic simulations of microinstabilities within the pedestal region of H-mode NSTX discharges in a highly shaped geometry

    DOE PAGES

    Coury, M.; Guttenfelder, W.; Mikkelsen, D. R.; ...

    2016-06-30

    Linear (local) gyrokinetic predictions of edge microinstabilities in highly shaped, lithiated and non-lithiated NSTX discharges are reported using the gyrokinetic code GS2. Microtearing modes dominate the non-lithiated pedestal top. The stabilization of these modes at the lithiated pedestal top enables the electron temperature pedestal to extend further inwards, as observed experimentally. Kinetic ballooning modes are found to be unstable mainly at the mid-pedestal of both types of discharges, with un- stable trapped electron modes nearer the separatrix region. At electron wavelengths, ETG modes are found to be unstable from mid-pedestal outwards for ηe, exp ~2.2 with higher growth rates formore » the lithiated discharge. Near the separatrix, the critical temperature gradient for driving ETG modes is reduced in the presence of lithium, re ecting the reduction of the lithiated density gradients observed experimentally. A preliminary linear study in the edge of non-lithiated discharges shows that the equilibrium shaping alters the electrostatic modes stability, found more unstable at high plasma shaping.« less

  14. Linear gyrokinetic simulations of microinstabilities within the pedestal region of H-mode NSTX discharges in a highly shaped geometry

    SciTech Connect

    Coury, M.; Guttenfelder, W.; Mikkelsen, D. R.; Canik, J. M.; Canal, G. P.; Diallo, A.; Kaye, S.; Kramer, G. J.; Maingi, R.

    2016-06-30

    Linear (local) gyrokinetic predictions of edge microinstabilities in highly shaped, lithiated and non-lithiated NSTX discharges are reported using the gyrokinetic code GS2. Microtearing modes dominate the non-lithiated pedestal top. The stabilization of these modes at the lithiated pedestal top enables the electron temperature pedestal to extend further inwards, as observed experimentally. Kinetic ballooning modes are found to be unstable mainly at the mid-pedestal of both types of discharges, with un- stable trapped electron modes nearer the separatrix region. At electron wavelengths, ETG modes are found to be unstable from mid-pedestal outwards for ηe, exp ~2.2 with higher growth rates for the lithiated discharge. Near the separatrix, the critical temperature gradient for driving ETG modes is reduced in the presence of lithium, re ecting the reduction of the lithiated density gradients observed experimentally. A preliminary linear study in the edge of non-lithiated discharges shows that the equilibrium shaping alters the electrostatic modes stability, found more unstable at high plasma shaping.

  15. Linear gyrokinetic simulations of microinstabilities within the pedestal region of H-mode NSTX discharges in a highly shaped geometry

    SciTech Connect

    Coury, M.; Guttenfelder, W.; Mikkelsen, D. R.; Canik, J. M.; Canal, G. P.; Diallo, A.; Kaye, S.; Kramer, G. J.; Maingi, R.

    2016-06-30

    Linear (local) gyrokinetic predictions of edge microinstabilities in highly shaped, lithiated and non-lithiated NSTX discharges are reported using the gyrokinetic code GS2. Microtearing modes dominate the non-lithiated pedestal top. The stabilization of these modes at the lithiated pedestal top enables the electron temperature pedestal to extend further inwards, as observed experimentally. Kinetic ballooning modes are found to be unstable mainly at the mid-pedestal of both types of discharges, with un- stable trapped electron modes nearer the separatrix region. At electron wavelengths, ETG modes are found to be unstable from mid-pedestal outwards for ηe, exp ~2.2 with higher growth rates for the lithiated discharge. Near the separatrix, the critical temperature gradient for driving ETG modes is reduced in the presence of lithium, re ecting the reduction of the lithiated density gradients observed experimentally. A preliminary linear study in the edge of non-lithiated discharges shows that the equilibrium shaping alters the electrostatic modes stability, found more unstable at high plasma shaping.

  16. Fabrication and optical characterization of pedestal micro-structures on DUV210 polymer: waveguides structures towards micro-resonators

    NASA Astrophysics Data System (ADS)

    Specht, Marion; Huby, Nolwenn; Lhermite, Hervé; Castro-Beltran, Rigoberto; Loas, Goulchen; Bêche, Bruno

    2015-07-01

    The paper presents the global design, fabrication and optical characterizations of pedestal waveguides and 2D microresonators (MRs) made of DUV210 polymer. These particular geometries are achieved thanks to specific deep-UV lithography procedures allowing sub-lambda development coupled to chemical etching able to shape such pedestal configuration on various optical microstructures. Scanning electron microscopy images have confirmed their features. Moreover, such families of pedestal structures were characterized experimentally, including the optical losses measurements for the waveguides and the optical resonance responses for two kinds of microresonators. Optical studies of single mode propagation losses measurements have been performed by a cut back method leading to values close to 20 dB/cm at a 635 nm wavelength. Additionally, resonant spectral analysis has been performed into pedestal rings and racetracks microresonators with a broadband laser source centered at 795 nm, demonstrating the presence of expected theoretical resonances. Free spectral range (FSR) values of 2.8 nm and 2.51 nm have been measured for these new designed pedestal resonators on DUV210 related to quality factor values superior to 520 and 610 respectively.

  17. Yersinia effector YopO uses actin as bait to phosphorylate proteins that regulate actin polymerization.

    PubMed

    Lee, Wei Lin; Grimes, Jonathan M; Robinson, Robert C

    2015-03-01

    Pathogenic Yersinia species evade host immune systems through the injection of Yersinia outer proteins (Yops) into phagocytic cells. One Yop, YopO, also known as YpkA, induces actin-filament disruption, impairing phagocytosis. Here we describe the X-ray structure of Yersinia enterocolitica YopO in complex with actin, which reveals that YopO binds to an actin monomer in a manner that blocks polymerization yet allows the bound actin to interact with host actin-regulating proteins. SILAC-MS and biochemical analyses confirm that actin-polymerization regulators such as VASP, EVL, WASP, gelsolin and the formin diaphanous 1 are directly sequestered and phosphorylated by YopO through formation of ternary complexes with actin. This leads to a model in which YopO at the membrane sequesters actin from polymerization while using the bound actin as bait to recruit, phosphorylate and misregulate host actin-regulating proteins to disrupt phagocytosis.

  18. Yersinia effector YopO uses actin as bait to phosphorylate proteins that regulate actin polymerization

    PubMed Central

    Lee, Wei Lin; Grimes, Jonathan M; Robinson, Robert C

    2016-01-01

    Pathogenic Yersinia species evade host immune systems through the injection of Yersinia outer proteins (Yops) into phagocytic cells. One Yop, YopO, also known as YpkA, induces actin-filament disruption, impairing phagocytosis. Here we describe the X-ray structure of Yersinia enterocolitica YopO in complex with actin, which reveals that YopO binds to an actin monomer in a manner that blocks polymerization yet allows the bound actin to interact with host actin-regulating proteins. SILAC-MS and biochemical analyses confirm that actin-polymerization regulators such as VASP, EVL, WASP, gelsolin and the formin diaphanous 1 are directly sequestered and phosphorylated by YopO through formation of ternary complexes with actin. This leads to a model in which YopO at the membrane sequesters actin from polymerization while using the bound actin as bait to recruit, phosphorylate and misregulate host actin-regulating proteins to disrupt phagocytosis. PMID:25664724

  19. Intracellular proliferation of S. aureus in osteoblasts and effects of rifampicin and gentamicin on S. aureus intracellular proliferation and survival.

    PubMed

    Mohamed, W; Sommer, U; Sethi, S; Domann, E; Thormann, U; Schütz, I; Lips, K S; Chakraborty, T; Schnettler, R; Alt, V

    2014-10-23

    Staphylococcus aureus is the most clinically relevant pathogen regarding implant-associated bone infection and its capability to invade osteoblasts is well known. The aim of this study was to investigate firstly whether S. aureus is not only able to invade but also to proliferate within osteoblasts, secondly to delineate the mechanism of invasion and thirdly to clarify whether rifampicin or gentamicin can inhibit intracellular proliferation and survival of S. aureus. The SAOS-2 osteoblast-like cell line and human primary osteoblasts were infected with S. aureus EDCC5055 and S. aureus Rosenbach 1884. Both S. aureus strains were able to invade efficiently and to proliferate within human osteoblasts. Immunofluorescence microscopy showed intracellular invasion of S. aureus and transmission electron microscopy images could demonstrate bacterial division as a sign of intracellular proliferation as well as cytosolic bacterial persistence. Cytochalasin D, the major actin depolymerisation agent, was able to significantly reduce S. aureus invasion, suggesting that invasion was enabled by promoting actin rearrangement at the cell surface. 7.5 μg/mL of rifampicin was able to inhibit bacterial survival in SAOS-2 cells with almost complete elimination of bacteria after 4 h. Gentamicin could also kill intracellular S. aureus in a dose-dependent manner, an effect that was significantly lower than that observed using rifampicin. In conclusion, S. aureus is not only able to invade but also to proliferate in osteoblasts. Invasion seems to be associated with actin rearrangement at the cell surface. Rifampicin is effective in intracellular eradication of S. aureus whereas gentamicin only poorly eliminates intracellularly replicating bacteria.

  20. Measurement of edge currents in DIII-D and their implication for pedestal stabilitya)

    NASA Astrophysics Data System (ADS)

    Thomas, D. M.; Leonard, A. W.; Groebner, R. J.; Osborne, T. H.; Casper, T. A.; Snyder, P. B.; Lao, L. L.

    2005-05-01

    The present performance limits of tokamak discharges are strongly coupled to the stability and transport properties of the edge plasma. Both experimental and modeling efforts have shown a clear connection between the edge pressure pedestal height and core plasma confinement. The key to understanding the stability and performance limits of the pedestal revolves around an accurate knowledge of the plasma current in this region. Using the Zeeman effect in an injected 30keV lithium beam, we have measured the currents in the edge of the DIII-D [J. L. Luxon, Nucl. Fusion 42, 6114 (2002)] tokamak for various confinement modes. This method of determining j(r ) is insensitive to the large electric fields which coexist in the pedestal region and which complicate motional Stark effect measurements. For the high confinement cases, where substantial pedestal pressures exist, we find large (~MA/m2), localized (ΔR ˜1-2cm) currents in the pedestal region, located near the maximum in the pressure gradient. These values are consistent with calculations of edge bootstrap current using the neoclassical NCLASS [W. A. Houlberg, K. C. Shaing, S. P. Hirshman, and M. C. Zarnstorff, Phys. Plasmas 4, 3230 (1997)] and Sauter [O. Sauter, C. Angioni, and Y. R. Lin-Lin, Phys. Plasmas 6, 2834 (1999)] models and the measured pedestal density and temperature profiles. The apparent consistency of the measured jEDGE with neoclassical predictions occurs despite the violation of one of the fundamental tenets of the theory, namely, ɛ =ρi/LP≪1, where ρi is the ion poloidal gyroradius and Lp is the pressure gradient scale length. The measured jEDGE has also been used to generate self-consistent reconstructions using the free boundary equilibrium solvers CORSICA [T. A. Casper, T. B. Kaiser, R. A. Jong, L. L. LoDestro, J. Moller, and L. D. Pearlstein, Plasma Phys. Controlled Fusion 45, 1193 (2003)] and EFIT [L. L. Lao, H. E. St. John, R. D. Stambough, A. G. Kellman, and W. Pfeiffer, Nucl. Fusion 25

  1. Measurement of edge currents in DIII-D and their implication for pedestal stability

    SciTech Connect

    Thomas, D.M.; Leonard, A.W.; Groebner, R.J.; Osborne, T.H.; Casper, T.A.; Snyder, P.B.; Lao, L.L.

    2005-05-15

    The present performance limits of tokamak discharges are strongly coupled to the stability and transport properties of the edge plasma. Both experimental and modeling efforts have shown a clear connection between the edge pressure pedestal height and core plasma confinement. The key to understanding the stability and performance limits of the pedestal revolves around an accurate knowledge of the plasma current in this region. Using the Zeeman effect in an injected 30 keV lithium beam, we have measured the currents in the edge of the DIII-D [J. L. Luxon, Nucl. Fusion 42, 6114 (2002)] tokamak for various confinement modes. This method of determining j(r) is insensitive to the large electric fields which coexist in the pedestal region and which complicate motional Stark effect measurements. For the high confinement cases, where substantial pedestal pressures exist, we find large ({approx}MA/m{sup 2}), localized ({delta}R{approx}1-2 cm) currents in the pedestal region, located near the maximum in the pressure gradient. These values are consistent with calculations of edge bootstrap current using the neoclassical NCLASS [W. A. Houlberg, K. C. Shaing, S. P. Hirshman, and M. C. Zarnstorff, Phys. Plasmas 4, 3230 (1997)] and Sauter [O. Sauter, C. Angioni, and Y. R. Lin-Lin, Phys. Plasmas 6, 2834 (1999)] models and the measured pedestal density and temperature profiles. The apparent consistency of the measured j{sub EDGE} with neoclassical predictions occurs despite the violation of one of the fundamental tenets of the theory, namely, {epsilon}={rho}{sub i}/L{sub P}<<1, where {rho}{sub i} is the ion poloidal gyroradius and L{sub p} is the pressure gradient scale length. The measured j{sub EDGE} has also been used to generate self-consistent reconstructions using the free boundary equilibrium solvers CORSICA [T. A. Casper, T. B. Kaiser, R. A. Jong, L. L. LoDestro, J. Moller, and L. D. Pearlstein, Plasma Phys. Controlled Fusion 45, 1193 (2003)] and EFIT [L. L. Lao, H. E. St

  2. H-Mode Turbulence, Power Threshold, ELM, and Pedestal Studies in NSTX

    SciTech Connect

    R. Maingi; C.E. Bush; E.D. Fredrickson; D.A. Gates; S.M. Kaye; B.P. LeBlanc; J.E. Menard; H. Meyer; D. Mueller; N. Nishino; A.L. Roquemore; S.A. Sabbagh; K. Tritz; S.J. Zweben; M.G. Bell; R.E. Bell; T. Biewer; J.A. Boedo; D.W. Johnson; R. Kaita; H.W. Kugel; R.J. Maqueda; T. Munsat; R. Raman; V.A. Soukhanovskii; T. Stevenson; D. Stutman

    2004-10-28

    High-confinement mode (H-mode) operation plays a crucial role in NSTX [National Spherical Torus Experiment] research, allowing higher beta limits due to reduced plasma pressure peaking, and long-pulse operation due to high bootstrap current fraction. Here, new results are presented in the areas of edge localized modes (ELMs), H-mode pedestal physics, L-H turbulence, and power threshold studies. ELMs of several other types (as observed in conventional aspect ratio tokamaks) are often observed: (1) large, Type I ELMs, (2) ''medium'' Type II/III ELMs, and (3) giant ELMs which can reduce stored energy by up to 30% in certain conditions. In addition, many high-performance discharges in NSTX have tiny ELMs (newly termed Type V), which have some differences as compared with ELM types in the published literature. The H-mode pedestal typically contains between 25-33% of the total stored energy, and the NSTX pedestal energy agrees reasonably well with a recent international multi-machine scaling. We find that the L-H transition occurs on a {approx}100 {micro}sec timescale as viewed by a gas puff imaging diagnostic, and that intermittent quiescent periods precede the final transition. A power threshold identity experiment between NSTX and MAST shows comparable loss power at the L-H transition in balanced double-null discharges. Both machines require more power for the L-H transition as the balance is shifted toward lower single null. High field side gas fueling enables more reliable H-mode access, but does not always lead to a lower power threshold e.g., with a reduction of the duration of early heating. Finally the edge plasma parameters just before the L-H transition were compared with theories of the transition. It was found that while some theories can separate well-developed L- and H-mode data, they have little predictive value.

  3. Acoustic transfer of protein crystals from agarose pedestals to micromeshes for high-throughput screening.

    PubMed

    Cuttitta, Christina M; Ericson, Daniel L; Scalia, Alexander; Roessler, Christian G; Teplitsky, Ella; Joshi, Karan; Campos, Olven; Agarwal, Rakhi; Allaire, Marc; Orville, Allen M; Sweet, Robert M; Soares, Alexei S

    2015-01-01

    Acoustic droplet ejection (ADE) is an emerging technology with broad applications in serial crystallography such as growing, improving and manipulating protein crystals. One application of this technology is to gently transfer crystals onto MiTeGen micromeshes with minimal solvent. Once mounted on a micromesh, each crystal can be combined with different chemicals such as crystal-improving additives or a fragment library. Acoustic crystal mounting is fast (2.33 transfers s(-1)) and all transfers occur in a sealed environment that is in vapor equilibrium with the mother liquor. Here, a system is presented to retain crystals near the ejection point and away from the inaccessible dead volume at the bottom of the well by placing the crystals on a concave agarose pedestal (CAP) with the same chemical composition as the crystal mother liquor. The bowl-shaped CAP is impenetrable to crystals. Consequently, gravity will gently move the crystals into the optimal location for acoustic ejection. It is demonstrated that an agarose pedestal of this type is compatible with most commercially available crystallization conditions and that protein crystals are readily transferred from the agarose pedestal onto micromeshes with no loss in diffraction quality. It is also shown that crystals can be grown directly on CAPs, which avoids the need to transfer the crystals from the hanging drop to a CAP. This technology has been used to combine thermolysin and lysozyme crystals with an assortment of anomalously scattering heavy atoms. The results point towards a fast nanolitre method for crystal mounting and high-throughput screening.

  4. Acoustic transfer of protein crystals from agarose pedestals to micromeshes for high-throughput screening

    SciTech Connect

    Cuttitta, Christina M.; Ericson, Daniel L.; Scalia, Alexander; Roessler, Christian G.; Teplitsky, Ella; Joshi, Karan; Campos, Olven; Agarwal, Rakhi; Allaire, Marc; Orville, Allen M.; Sweet, Robert M.; Soares, Alexei S.

    2015-01-01

    An acoustic high-throughput screening method is described for harvesting protein crystals and combining the protein crystals with chemicals such as a fragment library. Acoustic droplet ejection (ADE) is an emerging technology with broad applications in serial crystallography such as growing, improving and manipulating protein crystals. One application of this technology is to gently transfer crystals onto MiTeGen micromeshes with minimal solvent. Once mounted on a micromesh, each crystal can be combined with different chemicals such as crystal-improving additives or a fragment library. Acoustic crystal mounting is fast (2.33 transfers s{sup −1}) and all transfers occur in a sealed environment that is in vapor equilibrium with the mother liquor. Here, a system is presented to retain crystals near the ejection point and away from the inaccessible dead volume at the bottom of the well by placing the crystals on a concave agarose pedestal (CAP) with the same chemical composition as the crystal mother liquor. The bowl-shaped CAP is impenetrable to crystals. Consequently, gravity will gently move the crystals into the optimal location for acoustic ejection. It is demonstrated that an agarose pedestal of this type is compatible with most commercially available crystallization conditions and that protein crystals are readily transferred from the agarose pedestal onto micromeshes with no loss in diffraction quality. It is also shown that crystals can be grown directly on CAPs, which avoids the need to transfer the crystals from the hanging drop to a CAP. This technology has been used to combine thermolysin and lysozyme crystals with an assortment of anomalously scattering heavy atoms. The results point towards a fast nanolitre method for crystal mounting and high-throughput screening.

  5. Acoustic transfer of protein crystals from agarose pedestals to micromeshes for high-throughput screening

    PubMed Central

    Cuttitta, Christina M.; Ericson, Daniel L.; Scalia, Alexander; Roessler, Christian G.; Teplitsky, Ella; Joshi, Karan; Campos, Olven; Agarwal, Rakhi; Allaire, Marc; Orville, Allen M.; Sweet, Robert M.; Soares, Alexei S.

    2015-01-01

    Acoustic droplet ejection (ADE) is an emerging technology with broad applications in serial crystallography such as growing, improving and manipulating protein crystals. One application of this technology is to gently transfer crystals onto MiTeGen micromeshes with minimal solvent. Once mounted on a micromesh, each crystal can be combined with different chemicals such as crystal-improving additives or a fragment library. Acoustic crystal mounting is fast (2.33 transfers s−1) and all transfers occur in a sealed environment that is in vapor equilibrium with the mother liquor. Here, a system is presented to retain crystals near the ejection point and away from the inaccessible dead volume at the bottom of the well by placing the crystals on a concave agarose pedestal (CAP) with the same chemical composition as the crystal mother liquor. The bowl-shaped CAP is impenetrable to crystals. Consequently, gravity will gently move the crystals into the optimal location for acoustic ejection. It is demonstrated that an agarose pedestal of this type is compatible with most commercially available crystallization conditions and that protein crystals are readily transferred from the agarose pedestal onto micromeshes with no loss in diffraction quality. It is also shown that crystals can be grown directly on CAPs, which avoids the need to transfer the crystals from the hanging drop to a CAP. This technology has been used to combine thermolysin and lysozyme crystals with an assortment of anomalously scattering heavy atoms. The results point towards a fast nanolitre method for crystal mounting and high-throughput screening. PMID:25615864

  6. Acoustic transfer of protein crystals from agarose pedestals to micromeshes for high-throughput screening

    SciTech Connect

    Cuttitta, Christina M.; Ericson, Daniel L.; Scalia, Alexander; Roessler, Christian G.; Teplitsky, Ella; Joshi, Karan; Campos, Olven; Agarwal, Rakhi; Allaire, Marc; Orville, Allen M.; Sweet, Robert M.; Soares, Alexei S.

    2014-06-01

    Acoustic droplet ejection (ADE) is an emerging technology with broad applications in serial crystallography such as growing, improving and manipulating protein crystals. One application of this technology is to gently transfer crystals onto MiTeGen micromeshes with minimal solvent. Once mounted on a micromesh, each crystal can be combined with different chemicals such as crystal-improving additives or a fragment library. Acoustic crystal mounting is fast (2.33 transfers s-1) and all transfers occur in a sealed environment that is in vapor equilibrium with the mother liquor. Here, a system is presented to retain crystals near the ejection point and away from the inaccessible dead volume at the bottom of the well by placing the crystals on a concave agarose pedestal (CAP) with the same chemical composition as the crystal mother liquor. The bowl-shaped CAP is impenetrable to crystals. Consequently, gravity will gently move the crystals into the optimal location for acoustic ejection. It is demonstrated that an agarose pedestal of this type is compatible with most commercially available crystallization conditions and that protein crystals are readily transferred from the agarose pedestal onto micromeshes with no loss in diffraction quality. It is also shown that crystals can be grown directly on CAPs, which avoids the need to transfer the crystals from the hanging drop to a CAP. This technology has been used to combine thermolysin and lysozyme crystals with an assortment of anomalously scattering heavy atoms. The results point towards a fast nanolitre method for crystal mounting and high-throughput screening.

  7. Feasibility study for a new high resolution Thomson scattering system for the ASDEX Upgrade pedestal

    NASA Astrophysics Data System (ADS)

    Tsalas, M.; Kantor, M. Yu; Maj, O.; Bilato, R.; de Vries, P. C.; Donné, A. J. H.; Herrmann, A.; Kurzan, B.; Wolfrum, E.; the ASDEX Upgrade Team

    2012-03-01

    A new Thomson scattering diagnostic is proposed for the study of fast plasma dynamics in the pedestal of ASDEX Upgrade. The diagnostic will measure electron temperature and density profiles over a ~ 3 cm wide area in the edge transport barrier region, with ~ 1-2 mm spatial resolution and ~ 10 kHz sampling rate. A challenging goal of the project is the study of the bootstrap current in the plasma pedestal by measuring the distortion and shift of the electron distribution along the toroidal direction. Expected spatial and time resolutions of the current density measurements are ~ 3 mm and ~ 1 ms correspondingly. The new diagnostic will be used to study the fast dynamic behaviour of the pedestal bootstrap current, where models indicate that it plays a key role in regulating edge stability, e.g. during ELMs. The diagnostic design is based on the intra-cavity multi-pass system currently in operation in TEXTOR, which uses a probing ruby laser, a grating spectrometer and two fast CMOS cameras for scattered light detection, and has achieved measuring accuracies of the order of ~ 1% for ne and ~ 2% for Te. Parts of that system will be reused in ASDEX Upgrade (some with significant modifications), but the laser multi-pass and light collection systems are entirely redesigned. Restrictions in space and line-of-sight availability have led to the adoption of a design which uses in-vessel multi-pass mirrors and light collection optics, requiring a number of innovative technical solutions to permit remote laser alignment and light collection. We give an overview of the project, discuss the underlying physics basis and present a number of technical solutions employed.

  8. Acoustic transfer of protein crystals from agarose pedestals to micromeshes for high-throughput screening

    DOE PAGES

    Cuttitta, Christina M.; Ericson, Daniel L.; Scalia, Alexander; ...

    2014-06-01

    Acoustic droplet ejection (ADE) is an emerging technology with broad applications in serial crystallography such as growing, improving and manipulating protein crystals. One application of this technology is to gently transfer crystals onto MiTeGen micromeshes with minimal solvent. Once mounted on a micromesh, each crystal can be combined with different chemicals such as crystal-improving additives or a fragment library. Acoustic crystal mounting is fast (2.33 transfers s-1) and all transfers occur in a sealed environment that is in vapor equilibrium with the mother liquor. Here, a system is presented to retain crystals near the ejection point and away from themore » inaccessible dead volume at the bottom of the well by placing the crystals on a concave agarose pedestal (CAP) with the same chemical composition as the crystal mother liquor. The bowl-shaped CAP is impenetrable to crystals. Consequently, gravity will gently move the crystals into the optimal location for acoustic ejection. It is demonstrated that an agarose pedestal of this type is compatible with most commercially available crystallization conditions and that protein crystals are readily transferred from the agarose pedestal onto micromeshes with no loss in diffraction quality. It is also shown that crystals can be grown directly on CAPs, which avoids the need to transfer the crystals from the hanging drop to a CAP. This technology has been used to combine thermolysin and lysozyme crystals with an assortment of anomalously scattering heavy atoms. The results point towards a fast nanolitre method for crystal mounting and high-throughput screening.« less

  9. Mechanism of Actin-Based Motility

    NASA Astrophysics Data System (ADS)

    Pantaloni, Dominique; Le Clainche, Christophe; Carlier, Marie-France

    2001-05-01

    Spatially controlled polymerization of actin is at the origin of cell motility and is responsible for the formation of cellular protrusions like lamellipodia. The pathogens Listeria monocytogenes and Shigella flexneri, which undergo actin-based propulsion, are acknowledged models of the leading edge of lamellipodia. Actin-based motility of the bacteria or of functionalized microspheres can be reconstituted in vitro from only five pure proteins. Movement results from the regulated site-directed treadmilling of actin filaments, consistent with observations of actin dynamics in living motile cells and with the biochemical properties of the components of the synthetic motility medium.

  10. How cellular membrane properties are affected by the actin cytoskeleton.

    PubMed

    Lemière, J; Valentino, F; Campillo, C; Sykes, C

    2016-11-01

    Lipid membranes define the boundaries of living cells and intracellular compartments. The dynamic remodelling of these membranes by the cytoskeleton, a very dynamic structure made of active biopolymers, is crucial in many biological processes such as motility or division. In this review, we present some aspects of cellular membranes and how they are affected by the presence of the actin cytoskeleton. We show that, in parallel with the direct study of membranes and cytoskeleton in vivo, biomimetic in vitro systems allow reconstitution of biological processes in a controlled environment. In particular, we show that liposomes, or giant unilamellar vesicles, encapsulating a reconstituted actin network polymerizing at their membrane are suitable models of living cells and can be used to decipher the relative contributions of membrane and actin on the mechanical properties of the cellular interface. Copyright © 2016 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.

  11. Propulsion and Comet-Tail Formation by Actin Polymerization

    NASA Astrophysics Data System (ADS)

    Katz, Jonathan; Carlsson, Anders

    2002-03-01

    We have developed and calculated a model of the propulsion of intracellular parasites by actin polymerization. A disc or sphere is uniformly coated with sites at which F-actin is bound and G-actin may intercalate. Intercalation displaces the sphere and polymer along the polymeric axis, in opposite directions, by amounts inversely proportional to their viscous drags. As the sphere is displaced and rotates the polymers are subject to drag in the flow field. We consider two cases: 1. The polymers remain perpendicular to the surface of the sphere, but are free to slide along it; 2. The polymers are hinged at fixed points of attachment. In each case fluctuations in displacement (resulting from stochastic choice of intercalation sites) grow as motion in one direction sweeps the polymers back, so further intercalation augments the directed motion. Numerical simulation shows that the second case leads to runaway (pinwheel) rotation and little translation, but that in the first case the sphere is effectively propelled after a latency time, and a comet tail is formed.

  12. Regulatory mimicry in Listeria monocytogenes actin-based motility

    PubMed Central

    Chong, Ryan; Swiss, Rachel; Briones, Gabriel; Stone, Kathryn L.; Gulcicek, Erol E.; Agaisse, Hervé

    2009-01-01

    Summary The actin-based motility of the intracellular pathogen Listeria monocytogenes relies on ActA, a bacterial factor mimicking the activity of host cell nucleation-promoting factors of the WASP/WAVE family. The activity of WASP and WAVE is tightly regulated in cells. However, it is not known whether the activity of ActA is regulated upon L. monocytogenes infection. Here, we used an RNAi-based genetic approach in combination with computer-assisted image analysis to investigate the role of host factors in L. monocytogenes spread from cell to cell. We showed that the host cell serine/threonine kinase CK2 is required for efficient actin tail formation. We demonstrated that, similar to WASP and WAVE, the affinity of ActA for the ARP2/3 complex is regulated by CK2-mediated phosphorylation. We also demonstrated the importance of this regulatory mechanism in a mouse model of infection. Our work suggests that ActA is a bacterial virulence factor that not only displays a structural mimic of the VCA domain of WASP/WAVE family members, but also co-opted CK2 as the host cell factor regulating its activity, a form of mimicry that we refer to as regulatory mimicry. We present comparative evidence supporting the notion that unrelated pathogens displaying actin-based motility may have evolved a similar strategy. PMID:19748468

  13. Bootstrap Current for the Edge Pedestal Plasma in a Diverted Tokamak Geometry

    SciTech Connect

    Koh, S.; Chang, C. S.; Ku, S.; Menard, J. E.; Weitzner, H.; Choe, W.

    2012-08-10

    The edge bootstrap current plays a critical role in the equilibrium and stability of the steep edge pedestal plasma. The pedestal plasma has an unconventional and difficult neoclassical property, as compared with the core plasma. It has a narrow passing particle region in velocity space that can be easily modified or destroyed by Coulomb collisions. At the same time, the edge pedestal plasma has steep pressure and electrostatic potential gradients whose scale-lengths are comparable with the ion banana width, and includes a magnetic separatrix surface, across which the topological properties of the magnetic field and particle orbits change abruptly. A driftkinetic particle code XGC0, equipped with a mass-momentum-energy conserving collision operator, is used to study the edge bootstrap current in a realistic diverted magnetic field geometry with a self-consistent radial electric field. When the edge electrons are in the weakly collisional banana regime, surprisingly, the present kinetic simulation confirms that the existing analytic expressions [represented by O. Sauter et al. , Phys. Plasmas 6 , 2834 (1999)] are still valid in this unconventional region, except in a thin radial layer in contact with the magnetic separatrix. The agreement arises from the dominance of the electron contribution to the bootstrap current compared with ion contribution and from a reasonable separation of the trapped-passing dynamics without a strong collisional mixing. However, when the pedestal electrons are in plateau-collisional regime, there is significant deviation of numerical results from the existing analytic formulas, mainly due to large effective collisionality of the passing and the boundary layer trapped particles in edge region. In a conventional aspect ratio tokamak, the edge bootstrap current from kinetic simulation can be significantly less than that from the Sauter formula if the electron collisionality is high. On the other hand, when the aspect ratio is close to unity

  14. Bootstrap current for the edge pedestal plasma in a diverted tokamak geometry

    SciTech Connect

    Koh, S.; Choe, W.; Chang, C. S.; Ku, S.; Menard, J. E.; Weitzner, H.

    2012-07-15

    The edge bootstrap current plays a critical role in the equilibrium and stability of the steep edge pedestal plasma. The pedestal plasma has an unconventional and difficult neoclassical property, as compared with the core plasma. It has a narrow passing particle region in velocity space that can be easily modified or destroyed by Coulomb collisions. At the same time, the edge pedestal plasma has steep pressure and electrostatic potential gradients whose scale-lengths are comparable with the ion banana width, and includes a magnetic separatrix surface, across which the topological properties of the magnetic field and particle orbits change abruptly. A drift-kinetic particle code XGC0, equipped with a mass-momentum-energy conserving collision operator, is used to study the edge bootstrap current in a realistic diverted magnetic field geometry with a self-consistent radial electric field. When the edge electrons are in the weakly collisional banana regime, surprisingly, the present kinetic simulation confirms that the existing analytic expressions [represented by O. Sauter et al., Phys. Plasmas 6, 2834 (1999)] are still valid in this unconventional region, except in a thin radial layer in contact with the magnetic separatrix. The agreement arises from the dominance of the electron contribution to the bootstrap current compared with ion contribution and from a reasonable separation of the trapped-passing dynamics without a strong collisional mixing. However, when the pedestal electrons are in plateau-collisional regime, there is significant deviation of numerical results from the existing analytic formulas, mainly due to large effective collisionality of the passing and the boundary layer trapped particles in edge region. In a conventional aspect ratio tokamak, the edge bootstrap current from kinetic simulation can be significantly less than that from the Sauter formula if the electron collisionality is high. On the other hand, when the aspect ratio is close to unity

  15. Retired NASA F-18 being mounted on pedestal mount at Lancaster California Municipal Baseball Stadium

    NASA Technical Reports Server (NTRS)

    1997-01-01

    Workers carefully align a mounting bracket attached to an F/A-18 Hornet aircraft with the top of a pedestal in front of the municipal baseball stadium in the city of Lancaster, California. The Blue-and-white twin-jet aircraft, formerly flown as a safety chase and support aircraft by NASA's Dryden Flight Research Center, Edwards, California, was loaned to the city for display following its recent retirement. Known as 'The Hangar,' the stadium is the home field of the Lancaster Jethawks, a Class-A farm team of the Seattle Mariners.

  16. Retired NASA F-18 being lowered on to pedestal mount at Lancaster California Municipal Baseball Stad

    NASA Technical Reports Server (NTRS)

    1997-01-01

    A large crane gingerly lowers an F/A-18 Hornet aircraft onto a 28-foot-tall pedestal in front of the municipal baseball stadium in the city of Lancaster, California. The blue-and-white F/A-18 was recently loaned to the city by NASA's Dryden Flight Research Center, Edwards, California. NASA Dryden had flown the twin-jet aircraft as a safety chase and support aircraft over the past nine years. The stadium, known as 'The Hangar,' is the home field of the Lancaster Jethawks, a Class-A farm team of the Seattle Mariners.

  17. Retired NASA F-18 being mounted on pedestal mount at Lancaster California Municipal Baseball Stadium

    NASA Technical Reports Server (NTRS)

    1997-01-01

    An F/A-18 Hornet aircraft formerly flown by NASA's Dryden Flight Research Center, Edwards, California, is sandwiched between two groups of workers as they mount it atop a pedestal at the municipal baseball stadium in the city of Lancaster, California. NASA Dryden had flown the blue-and-white twin-jet as a safety chase and support aircraft for about nine years prior to its recent retirement. The aircraft is now in loan to the city for public display. Known as 'The Hangar,' the stadium is the home field of the Lancaster Jethawks, a Class-A farm team of the Seattle Mariners.

  18. Pedestal Fueling Simulations with a Coupled Kinetic-kinetic Plasma-neutral Transport Code

    SciTech Connect

    D.P. Stotler, C.S. Chang, S.H. Ku, J. Lang and G.Y. Park

    2012-08-29

    A Monte Carlo neutral transport routine, based on DEGAS2, has been coupled to the guiding center ion-electron-neutral neoclassical PIC code XGC0 to provide a realistic treatment of neutral atoms and molecules in the tokamak edge plasma. The DEGAS2 routine allows detailed atomic physics and plasma-material interaction processes to be incorporated into these simulations. The spatial pro le of the neutral particle source used in the DEGAS2 routine is determined from the uxes of XGC0 ions to the material surfaces. The kinetic-kinetic plasma-neutral transport capability is demonstrated with example pedestal fueling simulations.

  19. TEMPEST Simulations of Collisionless Damping of Geodesic-Acoustic Mode in Edge Plasma Pedestal

    SciTech Connect

    Xu, X; Xiong, Z; Nevins, W; McKee, G

    2007-05-31

    The fully nonlinear 4D TEMPEST gyrokinetic continuum code produces frequency, collisionless damping of geodesic-acoustic mode (GAM) and zonal flow with fully nonlinear Boltzmann electrons for the inverse aspect ratio {epsilon}-scan and the tokamak safety factor q-scan in homogeneous plasmas. The TEMPEST simulation shows that GAM exists in edge plasma pedestal for steep density and temperature gradients, and an initial GAM relaxes to the standard neoclassical residual, rather than Rosenbluth-Hinton residual due to the presence of ion-ion collisions. The enhanced GAM damping explains experimental BES measurements on the edge q scaling of the GAM amplitude.

  20. Method of pedestal and common-mode noise correction for switched-capacitor analog memories

    DOEpatents

    Britton, C.L.

    1996-12-31

    A method and apparatus are disclosed for correcting common-mode noise and pedestal noise in a multichannel array of switched-capacitor analog memories wherein each analog memory is connected to an associated analog-to-digital converter. The apparatus comprises a single differential element in two different embodiments. In a first embodiment, the differential element is a reference analog memory connected to a buffer. In the second embodiment, the differential element is a reference analog memory connected to a reference analog-to-digital connected to an array of digital summing circuits. 4 figs.

  1. Method of pedestal and common-mode noise correction for switched-capacitor analog memories

    DOEpatents

    Britton, C.L.

    1997-09-23

    A method and apparatus are disclosed for correcting common-mode noise and pedestal noise in a multichannel array of switched-capacitor analog memories wherein each analog memory is connected to an associated analog-to-digital converter. The apparatus comprises a single differential element in two different embodiments. In a first embodiment, the differential element is a reference analog memory connected to a buffer. In the second embodiment, the differential dement is a reference analog memory connected to a reference analog-to-digital connected to an array of digital summing circuits. 4 figs.

  2. TEMPEST Simulations of Collisionless Damping of Geodesic-Acoustic Mode in Edge Plasma Pedestal

    SciTech Connect

    Xu, X Q; Xiong, Z; Nevins, W M; McKee, G R

    2007-05-30

    The fully nonlinear (full-f) 4D TEMPEST gyrokinetic continuum code produces frequency, collisionless damping of GAM and zonal flow with fully nonlinear Boltzmann electrons for the inverse aspect ratio {epsilon}-scan and the tokamak safety factor q-scan in homogeneous plasmas. The TEMPEST simulation shows that GAM exists in edge plasma pedestal for steep density and temperature gradients, and an initial GAM relaxes to the standard neoclassical residual, rather than Rosenbluth-Hinton residual due to the presence of ion-ion collisions. The enhanced GAM damping explains experimental BES measurements on the edge q scaling of the GAM amplitude.

  3. Evaluation of DSS-14 pedestal-review of top surface repair procedures

    NASA Technical Reports Server (NTRS)

    Oesterle, R. G.; Musser, D. W.; Salse, E. A. B.

    1983-01-01

    Proposed repair procedures for the top surface of the pedestal supporting the hydrostatic bearing runner for the 64m Antenna are presented. These procedures included: (1) removal of existing grout and concrete to approximately 8 in. below original concrete surface using a presplitting technique with expansive cement followed by secondary breaking; (2) preparation of exposed concrete surface including an epoxy bonding agent; and (3) replacement of material removed with 8 in. of new concrete surface including an epoxy bonding agent; and (4) replacement of material removed with 8 in. of new concrete and 4 in. of new grout.

  4. Retired NASA F-18 being mounted on pedestal mount at Lancaster California Municipal Baseball Stadium

    NASA Technical Reports Server (NTRS)

    1997-01-01

    Workers carefully align a mounting bracket attached to an F/A-18 Hornet aircraft with the top of a pedestal in front of the municipal baseball stadium in the city of Lancaster, California. The Blue-and-white twin-jet aircraft, formerly flown as a safety chase and support aircraft by NASA's Dryden Flight Research Center, Edwards, California, was loaned to the city for display following its recent retirement. Known as 'The Hangar,' the stadium is the home field of the Lancaster Jethawks, a Class-A farm team of the Seattle Mariners.

  5. Retired NASA F-18 being mounted on pedestal mount at Lancaster California Municipal Baseball Stadium

    NASA Technical Reports Server (NTRS)

    1997-01-01

    An F/A-18 Hornet aircraft formerly flown by NASA's Dryden Flight Research Center, Edwards, California, is sandwiched between two groups of workers as they mount it atop a pedestal at the municipal baseball stadium in the city of Lancaster, California. NASA Dryden had flown the blue-and-white twin-jet as a safety chase and support aircraft for about nine years prior to its recent retirement. The aircraft is now in loan to the city for public display. Known as 'The Hangar,' the stadium is the home field of the Lancaster Jethawks, a Class-A farm team of the Seattle Mariners.

  6. Retired NASA F-18 being lowered on to pedestal mount at Lancaster California Municipal Baseball Stad

    NASA Technical Reports Server (NTRS)

    1997-01-01

    A large crane gingerly lowers an F/A-18 Hornet aircraft onto a 28-foot-tall pedestal in front of the municipal baseball stadium in the city of Lancaster, California. The blue-and-white F/A-18 was recently loaned to the city by NASA's Dryden Flight Research Center, Edwards, California. NASA Dryden had flown the twin-jet aircraft as a safety chase and support aircraft over the past nine years. The stadium, known as 'The Hangar,' is the home field of the Lancaster Jethawks, a Class-A farm team of the Seattle Mariners.

  7. Method of pedestal and common-mode noise correction for switched-capacitor analog memories

    DOEpatents

    Britton, Charles L.

    1997-01-01

    A method and apparatus for correcting common-mode noise and pedestal noise in a multichannel array of switched-capacitor analog memories wherein each analog memory is connected to an associated analog-to-digital converter. The apparatus comprises a single differential element in two different embodiments. In a first embodiment, the differential element is a reference analog memory connected to a buffer. In the second embodiment, the differential dement is a reference analog memory connected to a reference analog-to-digital connected to an array of digital summing circuits.

  8. Method of pedestal and common-mode noise correction for switched-capacitor analog memories

    DOEpatents

    Britton, Charles L.

    1996-01-01

    A method and apparatus for correcting common-mode noise and pedestal noise in a multichannel array of switched-capacitor analog memories wherein each analog memory is connected to an associated analog-to-digital converter. The apparatus comprises a single differential element in two different embodiments. In a first embodiment, the differential element is a reference analog memory connected to a buffer. In the second embodiment, the differential element is a reference analog memory connected to a reference analog-to-digital connected to an array of digital summing circuits.

  9. Detailed heat transfer coefficient measurements and thermal analysis at engine conditions of a pedestal with fillet radii

    NASA Astrophysics Data System (ADS)

    Wang, Z.; Ireland, P. T.; Jones, T. V.

    1995-04-01

    The heat transfer coefficient over the surface of a pedestal with fillet radii has been measured using thermochromic liquid crystals and the transient heat transfer method. The tests were performed at engine representative Reynolds numbers for a geometry typical of those used in turbine blade cooling systems. The heat conduction process that occurs in the engine was subsequently modeled numerically with a finite element discretization of the solid pedestal. The measured heat transfer coefficients were used to derive the exact boundary conditions applicable to the engine. The temperature field within the pedestal, calculated using the correct heat transfer coefficient distribution, is compared to that calculated using an area-averaged heat transfer coefficient. Metal temperature differences of 90 K are predicted across the blade wall.

  10. Investigation of inter-ELM ion heat transport in the H-mode pedestal of ASDEX Upgrade plasmas

    NASA Astrophysics Data System (ADS)

    Viezzer, E.; Fable, E.; Cavedon, M.; Angioni, C.; Dux, R.; Laggner, F. M.; Bernert, M.; Burckhart, A.; McDermott, R. M.; Pütterich, T.; Ryter, F.; Willensdorfer, M.; Wolfrum, E.; the ASDEX Upgrade Team; the EUROfusion MST1 Team

    2017-02-01

    The ion heat transport in the pedestal of H-mode plasmas is investigated in various H-mode discharges with different pedestal ion collisionalities. Interpretive modelling suggests that in all analyzed discharges the ion heat diffusivity coefficient, {χ\\text{i}} , in the pedestal is close to the neoclassical prediction within the experimental uncertainties. The impact of changing the deposition location of the electron cyclotron resonance heating on the ion heat transport has been studied. The effect on the background profiles is small. The pre-ELM (edge localized modes) edge profiles as well as the behaviour of the electron temperature and density, ion temperature and impurity toroidal rotation during the ELM cycle are very similar in discharges with on- and off-axis ECRH heating. No significant deviation of {χ\\text{i}} from neoclassics is observed when changing the ECRH deposition location to the plasma edge.

  11. Nuclear Actin in Development and Transcriptional Reprogramming.

    PubMed

    Misu, Shinji; Takebayashi, Marina; Miyamoto, Kei

    2017-01-01

    Actin is a highly abundant protein in eukaryotic cells and dynamically changes its polymerized states with the help of actin-binding proteins. Its critical function as a constituent of cytoskeleton has been well-documented. Growing evidence demonstrates that actin is also present in nuclei, referred to as nuclear actin, and is involved in a number of nuclear processes, including transcriptional regulation and chromatin remodeling. The contribution of nuclear actin to transcriptional regulation can be explained by its direct interaction with transcription machineries and chromatin remodeling factors and by controlling the activities of transcription factors. In both cases, polymerized states of nuclear actin affect the transcriptional outcome. Nuclear actin also plays an important role in activating strongly silenced genes in somatic cells for transcriptional reprogramming. When these nuclear functions of actin are considered, it is plausible to speculate that nuclear actin is also implicated in embryonic development, in which numerous genes need to be activated in a well-coordinated manner. In this review, we especially focus on nuclear actin's roles in transcriptional activation, reprogramming and development, including stem cell differentiation and we discuss how nuclear actin can be an important player in development and cell differentiation.

  12. Actin Dynamics: From Nanoscale to Microscale

    PubMed Central

    Carlsson, Anders E.

    2010-01-01

    The dynamic nature of actin in cells manifests itself in many ways: Polymerization near the cell edge is balanced by depolymerization in the interior, externally induced actin polymerization is followed by depolymerization, and spontaneous oscillations of the cell periphery are frequently seen. I discuss how mathematical modeling relates quantitative measures of actin dynamics to the rates of underlying molecular level processes. The rate of actin incorporation at the leading edge of a moving cell is roughly consistent with existing theories, and the factors determining the characteristic time of actin polymerization are fairly well understood. However, our understanding of actin disassembly is limited, in particular the interplay between severing and depolymerization and the role of specific combinations of proteins in implementing disassembly events. The origins of cell-edge oscillations, and their possible relation to actin waves, are a fruitful area of future research. PMID:20462375

  13. Repeated Cycles of Rapid Actin Assembly and Disassembly on Epithelial Cell PhagosomesV⃞

    PubMed Central

    Yam, Patricia T.; Theriot, Julie A.

    2004-01-01

    We have found that early in infection of the intracellular pathogen Listeria monocytogenes in Madin-Darby canine kidney epithelial cells expressing actin conjugated to green fluorescent protein, F-actin rapidly assembles (∼25 s) and disassembles (∼30 s) around the bacteria, a phenomenon we call flashing. L. monocytogenes strains unable to perform actin-based motility or unable to escape the phagosome were capable of flashing, suggesting that the actin assembly occurs on the phagosome membrane. Cycles of actin assembly and disassembly could occur repeatedly on the same phagosome. Indirect immunofluorescence showed that most bacteria were fully internalized when flashing occurred, suggesting that actin flashing does not represent phagocytosis. Escherichia coli expressing invA, a gene product from Yersinia pseudotuberculosis that mediates cellular invasion, also induced flashing. Furthermore, polystyrene beads coated with E-cadherin or transferrin also induced flashing after internalization. This suggests that flashing occurs downstream of several distinct molecular entry mechanisms and may be a general consequence of internalization of large objects by epithelial cells. PMID:15456901

  14. Calcium-mediated actin reset (CaAR) mediates acute cell adaptations

    PubMed Central

    Wales, Pauline; Schuberth, Christian E; Aufschnaiter, Roland; Fels, Johannes; García-Aguilar, Ireth; Janning, Annette; Dlugos, Christopher P; Schäfer-Herte, Marco; Klingner, Christoph; Wälte, Mike; Kuhlmann, Julian; Menis, Ekaterina; Hockaday Kang, Laura; Maier, Kerstin C; Hou, Wenya; Russo, Antonella; Higgs, Henry N; Pavenstädt, Hermann; Vogl, Thomas; Roth, Johannes; Qualmann, Britta; Kessels, Michael M; Martin, Dietmar E; Mulder, Bela; Wedlich-Söldner, Roland

    2016-01-01

    Actin has well established functions in cellular morphogenesis. However, it is not well understood how the various actin assemblies in a cell are kept in a dynamic equilibrium, in particular when cells have to respond to acute signals. Here, we characterize a rapid and transient actin reset in response to increased intracellular calcium levels. Within seconds of calcium influx, the formin INF2 stimulates filament polymerization at the endoplasmic reticulum (ER), while cortical actin is disassembled. The reaction is then reversed within a few minutes. This Calcium-mediated actin reset (CaAR) occurs in a wide range of mammalian cell types and in response to many physiological cues. CaAR leads to transient immobilization of organelles, drives reorganization of actin during cell cortex repair, cell spreading and wound healing, and induces long-lasting changes in gene expression. Our findings suggest that CaAR acts as fundamental facilitator of cellular adaptations in response to acute signals and stress. DOI: http://dx.doi.org/10.7554/eLife.19850.001 PMID:27919320

  15. Actin dynamics in the regulation of endothelial barrier functions and neutrophil recruitment during endotoxemia and sepsis.

    PubMed

    Schnoor, Michael; García Ponce, Alexander; Vadillo, Eduardo; Pelayo, Rosana; Rossaint, Jan; Zarbock, Alexander

    2017-02-02

    Sepsis is a leading cause of death worldwide. Increased vascular permeability is a major hallmark of sepsis. Dynamic alterations in actin fiber formation play an important role in the regulation of endothelial barrier functions and thus vascular permeability. Endothelial integrity requires a delicate balance between the formation of cortical actin filaments that maintain endothelial cell contact stability and the formation of actin stress fibers that generate pulling forces, and thus compromise endothelial cell contact stability. Current research has revealed multiple molecular pathways that regulate actin dynamics and endothelial barrier dysfunction during sepsis. These include intracellular signaling proteins of the small GTPases family (e.g., Rap1, RhoA and Rac1) as well as the molecules that are directly acting on the actomyosin cytoskeleton such as myosin light chain kinase and Rho kinases. Another hallmark of sepsis is an excessive recruitment of neutrophils that also involves changes in the actin cytoskeleton in both endothelial cells and neutrophils. This review focuses on the available evidence about molecules that control actin dynamics and regulate endothelial barrier functions and neutrophil recruitment. We also discuss treatment strategies using pharmaceutical enzyme inhibitors to target excessive vascular permeability and leukocyte recruitment in septic patients.

  16. Diffusion Rate Limitations in Actin-Based Propulsion of Hard and Deformable Particles

    PubMed Central

    Dickinson, Richard B.; Purich, Daniel L.

    2006-01-01

    The mechanism by which actin polymerization propels intracellular vesicles and invasive microorganisms remains an open question. Several recent quantitative studies have examined propulsion of biomimetic particles such as polystyrene microspheres, phospholipid vesicles, and oil droplets. In addition to allowing quantitative measurement of parameters such as the dependence of particle speed on its size, these systems have also revealed characteristic behaviors such a saltatory motion of hard particles and oscillatory deformation of soft particles. Such measurements and observations provide tests for proposed mechanisms of actin-based motility. In the actoclampin filament end-tracking motor model, particle-surface-bound filament end-tracking proteins are involved in load-insensitive processive insertion of actin subunits onto elongating filament plus-ends that are persistently tethered to the surface. In contrast, the tethered-ratchet model assumes working filaments are untethered and the free-ended filaments grow as thermal ratchets in a load-sensitive manner. This article presents a model for the diffusion and consumption of actin monomers during actin-based particle propulsion to predict the monomer concentration field around motile particles. The results suggest that the various behaviors of biomimetic particles, including dynamic saltatory motion of hard particles and oscillatory vesicle deformations, can be quantitatively and self-consistently explained by load-insensitive, diffusion-limited elongation of (+)-end-tethered actin filaments, consistent with predictions of the actoclampin filament-end tracking mechanism. PMID:16731556

  17. Investigation of the plasma shaping effects on the H-mode pedestal structure using coupled kinetic neoclassical/MHD stability simulations

    DOE PAGES

    Pankin, A. Y.; Rafiq, T.; Kritz, A. H.; ...

    2017-06-08

    The effects of plasma shaping on the H-mode pedestal structure are investigated. High fidelity kinetic simulations of the neoclassical pedestal dynamics are combined with the magnetohydrodynamic (MHD) stability conditions for triggering edge localized mode (ELM) instabilities that limit the pedestal width and height in H-mode plasmas. We use the neoclassical kinetic XGC0 code [Chang et al., Phys. Plasmas 11, 2649 (2004)] to carry out a scan over plasma elongation and triangularity. As plasma profiles evolve, the MHD stability limits of these profiles are analyzed with the ideal MHD ELITE code [Snyder et al., Phys. Plasmas 9, 2037 (2002)]. In simulationsmore » with the XGC0 code, which includes coupled ion-electron dynamics, yield predictions for both ion and electron pedestal profiles. The differences in the predicted H-mode pedestal width and height for the DIII-D discharges with different elongation and triangularities are discussed. For the discharges with higher elongation, it is found that the gradients of the plasma profiles in the H-mode pedestal reach semi-steady states. In these simulations, the pedestal slowly continues to evolve to higher pedestal pressures and bootstrap currents until the peeling-ballooning stability conditions are satisfied. The discharges with lower elongation do not reach the semi-steady state, and ELM crashes are triggered at earlier times. The plasma elongation is found to have a stronger stabilizing effect than the plasma triangularity. For the discharges with lower elongation and lower triangularity, the ELM frequency is large, and the H-mode pedestal evolves rapidly. It is found that the temperature of neutrals in the scrape-off-layer (SOL) region can affect the dynamics of the H-mode pedestal buildup. But the final pedestal profiles are nearly independent of the neutral temperature. The elongation and triangularity affect the pedestal widths of plasma density and electron temperature profiles differently. This provides a new

  18. Formation of actin filament bundles in the ring canals of developing Drosophila follicles

    PubMed Central

    1996-01-01

    Growing the intracellular bridges that connect nurse cells with each o ther and to the developing oocyte is vital for egg development. These ring canals increase from 0.5 microns in diameter at stage 2 to 10 microns in diameter at stage 11. Thin sections cut horizontally as you would cut a bagel, show that there is a layer of circumferentially oriented actin filaments attached to the plasma membrane at the periphery of each canal. By decoration with subfragment 1 of myosin we find actin filaments of mixed polarities in the ring such as found in the "contractile ring" formed during cytokinesis. In vertical sections through the canal the actin filaments appear as dense dots. At stage 2 there are 82 actin filaments in the ring, by stage 6 there are 717 and by stage 10 there are 726. Taking into account the diameter, this indicates that there is 170 microns of actin filaments/canal at stage 2 (pi x 0.5 microns x 82), 14,000 microns at stage 9 and approximately 23,000 microns at stage 11 or one inch of actin filament! The density of actin filaments remains unchanged throughout development. What is particularly striking is that by stages 4-5, the ring of actin filaments has achieved its maximum thickness, even though the diameter has not yet increased significantly. Thereafter, the diameter increases. Throughout development, stages 2-11, the canal length also increases. Although the density (number of actin filaments/micron2) through a canal remains constant from stage 5 on, the actin filaments appear as a net of interconnected bundles. Further information on this net of bundles comes from studying mutant animals that lack kelch, a protein located in the ring canal that has homology to the actin binding protein, scruin. In this mutant, the actin filaments form normally but individual bundles that comprise the fibers of the net are not bound tightly together. Some bundles enter into the ring canal lumen but do not completely occlude the lumen. all these observations lay

  19. Activation of endothelial nitric oxide synthase is dependent on its interaction with globular actin in human umbilical vein endothelial cells.

    PubMed

    Mi, Qiongyu; Chen, Nan; Shaifta, Yasin; Xie, Liping; Lu, Hui; Liu, Zhen; Chen, Qi; Hamid, Colleen; Becker, Silke; Ji, Yong; Ferro, Albert

    2011-09-01

    Endothelial nitric oxide synthase (eNOS) has been reported to associate with globular actin, and this association increases eNOS activity. Adenosine, histamine, salbutamol and thrombin cause activation of eNOS through widely different mechanisms. Whether these eNOS agonists can regulate eNOS activity through affecting its association with actin is unknown. As previously reported, we confirmed in cultured human umbilical vein endothelial cells (HUVEC) that histamine and thrombin increased intracellular Ca(2+) whereas adenosine and salbutamol did not, and that these four agonists caused different effects on actin filament structure. Nevertheless, despite their divergent effects on intracellular Ca(2+) and on actin filament structure, we found by immunoprecipitation that adenosine, histamine, salbutamol and thrombin all caused an increase in association between eNOS and globular actin. This increase of association was inhibited by pre-treatment with phalloidin, an actin filament stabilizer. All of these agonists also increased phosphorylation of eNOS on serine residue 1177, eNOS activity, and cyclic guanosine-3', 5'-monophosphate, and these increases were all attenuated by phalloidin. Agonist-induced phosphorylation of eNOS on serine 1177 was attenuated by Akt inhibition, whereas association of eNOS with actin was not. We also found, in HEK-293 cells transfected with the eNOS mutants eNOS-S1177A or eNOS-S1177D, that the association between eNOS and globular actin was decreased as compared to cells transfected with wild-type eNOS. We conclude that association of globular actin with eNOS plays an essential and necessary role in agonist-induced eNOS activation, through enabling its phosphorylation by Akt at serine residue 1177.

  20. Hexosamine biosynthesis pathway flux contributes to insulin resistance via altering membrane phosphatidylinositol 4,5-bisphosphate and cortical filamentous actin.

    PubMed

    Bhonagiri, Padma; Pattar, Guruprasad R; Horvath, Emily M; Habegger, Kirk M; McCarthy, Alicia M; Elmendorf, Jeffrey S

    2009-04-01

    We recently found that plasma membrane phosphatidylinositol 4,5-bisphosphate (PIP(2))-regulated filamentous actin (F-actin) polymerization was diminished in hyperinsulinemic cell culture models of insulin resistance. Here we delineated whether increased glucose flux through the hexosamine biosynthesis pathway (HBP) causes the PIP(2)/F-actin dysregulation and insulin resistance induced by hyperinsulinemia. Increased HBP activity was detected in 3T3-L1 adipocytes cultured under conditions closely resembling physiological hyperinsulinemia (5 nm insulin for 12 h) and in cells where HBP activity was amplified by 2 mm glucosamine (GlcN). Both the physiological hyperinsulinemia and experimental GlcN challenge induced comparable losses of PIP(2) and F-actin. In addition to protecting against the insulin-induced membrane/cytoskeletal abnormality and insulin-resistant state, exogenous PIP(2) corrected the GlcN-induced insult on these parameters. Moreover, in accordance with HBP flux directly weakening PIP(2)/F-actin structure, inhibition of the rate-limiting HBP enzyme (glutamine:fructose-6-phosphate amidotransferase) restored PIP(2)-regulated F-actin structure and insulin responsiveness. Conversely, overexpression of glutamine:fructose-6-phosphate amidotransferase was associated with a loss of detectable plasma membrane PIP(2) and insulin sensitivity. A slight decrease in intracellular ATP resulted from amplifying HBP by hyperinsulinemia and GlcN. However, experimental maintenance of the intracellular ATP pool under both conditions with inosine did not reverse the PIP(2)/F-actin-based insulin-resistant state. Furthermore, less invasive challenges with glucose, in the absence of insulin, also led to PIP(2)/F-actin dysregulation. Accordingly, we suggest that the functionality of cell systems dependent on PIP(2) and/or F-actin status, such as the glucose transport system, can be critically compromised by inappropriate HBP activity.

  1. Hexosamine Biosynthesis Pathway Flux Contributes to Insulin Resistance via Altering Membrane Phosphatidylinositol 4,5-Bisphosphate and Cortical Filamentous Actin

    PubMed Central

    Bhonagiri, Padma; Pattar, Guruprasad R.; Horvath, Emily M.; Habegger, Kirk M.; McCarthy, Alicia M.; Elmendorf, Jeffrey S.

    2009-01-01

    We recently found that plasma membrane phosphatidylinositol 4,5-bisphosphate (PIP2)-regulated filamentous actin (F-actin) polymerization was diminished in hyperinsulinemic cell culture models of insulin resistance. Here we delineated whether increased glucose flux through the hexosamine biosynthesis pathway (HBP) causes the PIP2/F-actin dysregulation and insulin resistance induced by hyperinsulinemia. Increased HBP activity was detected in 3T3-L1 adipocytes cultured under conditions closely resembling physiological hyperinsulinemia (5 nm insulin for 12 h) and in cells where HBP activity was amplified by 2 mm glucosamine (GlcN). Both the physiological hyperinsulinemia and experimental GlcN challenge induced comparable losses of PIP2 and F-actin. In addition to protecting against the insulin-induced membrane/cytoskeletal abnormality and insulin-resistant state, exogenous PIP2 corrected the GlcN-induced insult on these parameters. Moreover, in accordance with HBP flux directly weakening PIP2/F-actin structure, inhibition of the rate-limiting HBP enzyme (glutamine:fructose-6-phosphate amidotransferase) restored PIP2-regulated F-actin structure and insulin responsiveness. Conversely, overexpression of glutamine:fructose-6-phosphate amidotransferase was associated with a loss of detectable plasma membrane PIP2 and insulin sensitivity. A slight decrease in intracellular ATP resulted from amplifying HBP by hyperinsulinemia and GlcN. However, experimental maintenance of the intracellular ATP pool under both conditions with inosine did not reverse the PIP2/F-actin-based insulin-resistant state. Furthermore, less invasive challenges with glucose, in the absence of insulin, also led to PIP2/F-actin dysregulation. Accordingly, we suggest that the functionality of cell systems dependent on PIP2 and/or F-actin status, such as the glucose transport system, can be critically compromised by inappropriate HBP activity. PMID:19036880

  2. Intracellular Pressure Dynamics in Blebbing Cells.

    PubMed

    Strychalski, Wanda; Guy, Robert D

    2016-03-08

    Blebs are pressure-driven protrusions that play an important role in cell migration, particularly in three-dimensional environments. A bleb is initiated when the cytoskeleton detaches from the cell membrane, resulting in the pressure-driven flow of cytosol toward the area of detachment and local expansion of the cell membrane. Recent experiments involving blebbing cells have led to conflicting hypotheses regarding the timescale of intracellular pressure propagation. The interpretation of one set of experiments supports a poroelastic model of the cytoplasm that leads to slow pressure equilibration when compared to the timescale of bleb expansion. A different study concludes that pressure equilibrates faster than the timescale of bleb expansion. To address this discrepancy, a dynamic computational model of the cell was developed that includes mechanics of and the interactions among the cytoplasm, the actin cortex, the cell membrane, and the cytoskeleton. The model results quantify the relationship among cytoplasmic rheology, pressure, and bleb expansion dynamics, and provide a more detailed picture of intracellular pressure dynamics. This study shows the elastic response of the cytoplasm relieves pressure and limits bleb size, and that both permeability and elasticity of the cytoplasm determine bleb expansion time. Our model with a poroelastic cytoplasm shows that pressure disturbances from bleb initiation propagate faster than the timescale of bleb expansion and that pressure equilibrates slower than the timescale of bleb expansion. The multiple timescales in intracellular pressure dynamics explain the apparent discrepancy in the interpretation of experimental results.

  3. Intracellular Pressure Dynamics in Blebbing Cells

    PubMed Central

    Strychalski, Wanda; Guy, Robert D.

    2016-01-01

    Blebs are pressure-driven protrusions that play an important role in cell migration, particularly in three-dimensional environments. A bleb is initiated when the cytoskeleton detaches from the cell membrane, resulting in the pressure-driven flow of cytosol toward the area of detachment and local expansion of the cell membrane. Recent experiments involving blebbing cells have led to conflicting hypotheses regarding the timescale of intracellular pressure propagation. The interpretation of one set of experiments supports a poroelastic model of the cytoplasm that leads to slow pressure equilibration when compared to the timescale of bleb expansion. A different study concludes that pressure equilibrates faster than the timescale of bleb expansion. To address this discrepancy, a dynamic computational model of the cell was developed that includes mechanics of and the interactions among the cytoplasm, the actin cortex, the cell membrane, and the cytoskeleton. The model results quantify the relationship among cytoplasmic rheology, pressure, and bleb expansion dynamics, and provide a more detailed picture of intracellular pressure dynamics. This study shows the elastic response of the cytoplasm relieves pressure and limits bleb size, and that both permeability and elasticity of the cytoplasm determine bleb expansion time. Our model with a poroelastic cytoplasm shows that pressure disturbances from bleb initiation propagate faster than the timescale of bleb expansion and that pressure equilibrates slower than the timescale of bleb expansion. The multiple timescales in intracellular pressure dynamics explain the apparent discrepancy in the interpretation of experimental results. PMID:26958893

  4. Control of actin filament length by phosphorylation of fragmin-actin complex

    PubMed Central

    1990-01-01

    Fragmin is a Ca2(+)-sensitive F-actin-severing protein purified from a slime mold, Physarum polycephalum (Hasegawa, T., S. Takahashi, H. Hayashi, and S. Hatano. 1980. Biochemistry. 19:2677-2683). It binds to G-actin to form a 1:1 fragmin/actin complex in the presence of micromolar free Ca2+. The complex nucleates actin polymerization and caps the barbed end of the short F-actin (Sugino, H., and S. Hatano. 1982. Cell Motil. 2:457-470). Subsequent removal of Ca2+, however, hardly dissociates the complex. This complex nucleates actin polymerization and caps the F-actin regardless of Ca2+ concentration. Here we report that this activity of fragmin-actin complex can be abolished by phosphorylation of actin of the complex. When crude extract from Physarum plasmodium was incubated with 5 mM ATP and 1 mM EGTA, the activities of the complex decreased to a great extent. The inactivation of the complex in the crude extract was not observed in the presence of Ca2+. In addition, the activities of the complex inactivated in the crude extract were restored under conditions suitable for phosphatase reactions. We purified factors that inactivated fragmin-actin complex from the crude extract. These factors phosphorylated actin of the complex, and the activities of the complex decreased with an increased level of phosphorylation of the complex. These factors, termed actin kinase, also inactivated the complex that capped the barbed end of short F-actin, leading to elongation of the short F-actin to long F-actin. Thus the length of F-actin can be controlled by phosphorylation of fragmin-actin complex by actin kinase. PMID:2202733

  5. Nucleocapsid of Tomato spotted wilt tospovirus forms mobile particles that traffic on an actin/endoplasmic reticulum network driven by myosin XI-K.

    PubMed

    Feng, Zhike; Chen, Xiaojiao; Bao, Yiqun; Dong, Jiahong; Zhang, Zhongkai; Tao, Xiaorong

    2013-12-01

    A number of viral proteins from plant viruses, other than movement proteins, have been shown to traffic intracellularly along actin filaments and to be involved in viral infection. However, there has been no report that a viral capsid protein may traffic within a cell by utilizing the actin/endoplasmic reticulum (ER) network. We used Tomato spotted wilt tospovirus (TSWV) as a model virus to study the cell biological properties of a nucleocapsid (N) protein. We found that TSWV N protein was capable of forming highly motile cytoplasmic inclusions that moved along the ER and actin network. The disruption of actin filaments by latrunculin B, an actin-depolymerizing agent, almost stopped the intracellular movement of N inclusions, whereas treatment with a microtubule-depolymerizing reagent, oryzalin, did not. The over-expression of a myosin XI-K tail, functioning in a dominant-negative manner, completely halted the movement of N inclusions. Latrunculin B treatment strongly inhibited the formation of TSWV local lesions in Nicotiana tabacum cv Samsun NN and delayed systemic infection in N. benthamiana. Collectively, our findings provide the first evidence that the capsid protein of a plant virus has the novel property of intracellular trafficking. The findings add capsid protein as a new class of viral protein that traffics on the actin/ER system.

  6. Evolution of edge pedestal transport between edge-localized modes in DIII-D

    SciTech Connect

    Floyd, J.-P.; Stacey, W. M.; Mellard, S. C.; Groebner, R. J.

    2015-02-15

    Evolution of measured profiles of densities, temperatures, and velocities in the edge pedestal region between successive ELM (edge-localized mode) events are analyzed and interpreted in terms of the constraints imposed by particle, momentum and energy balance in order to gain insights regarding the underlying evolution of transport processes in the edge pedestal between ELMs in a series of DIII-D [J. Luxon, Nucl. Fusion 42, 614 (2002)] discharges. The data from successive inter-ELM periods during an otherwise steady-state phase of the discharges were combined into a composite inter-ELM period for the purpose of increasing the number of data points in the analysis. Variation of diffusive and non-diffusive (pinch) particle, momentum, and energy transport over the inter-ELM period are interpreted using the GTEDGE code for discharges with plasma currents from 0.5 to 1.5 MA and inter-ELM periods from 50 to 220 ms. Diffusive transport is dominant for ρ < 0.925, while non-diffusive and diffusive transport are very large and nearly balancing in the sharp gradient region 0.925 < ρ < 1.0. During the inter-ELM period, diffusive transport increases slightly more than non-diffusive transport, increasing total outward transport. Both diffusive and non-diffusive transport have a strong inverse correlation with plasma current.

  7. Progress in understanding the enhanced pedestal H-mode in NSTX

    SciTech Connect

    Gerhardt, S. P.; Canik, J. M.; Maingi, R.; Battaglia, D.; Bell, R. E.; Guttenfelder, W.; LeBlanc, B. P.; Smith, D. R.; Yuh, H.; Sabbagh, S.

    2014-08-01

    The paper describes the enhanced pedestal (EP) H-mode observed in the National Spherical Torus Experiment (NSTX). The defining characteristics of EP H-mode are given, namely i)transition after the L- to H-mode transition, ii) region of very steep ion temperature gradient, and iii) associated region of strong rotational shear. A newly observed long-pulse EP H-mode example shows quiescent behavior for as long as the heating and current drive sources are maintained. Cases are shown where the region of steep ion temperature gradient is located at the very edge, and cases where it is shifted up to 10 cm inward from the plasma edge; these cases are united by a common dependence of the ion temperature gradient on the toroidal rotation frequency shear. EP H-mode examples have been observed across a wide range of q95 and pedestal collisionality. No strong changes in the fluctuation amplitudes have been observed following the eP H-mode transition, and transport analysis indicates that the ion t hermal transport is comparable to or less than anticipated from a simple neoclassical transport model. Cases are shown where EP H-modes were reliably generated, through these low-q95 examples were difficult to sustain. A case where an externally triggered ELM precipitates the transition to EP H-mode is also shown, though an initial experiment designed to trigger EP-H-modes in this fashion was successful.

  8. Development of a Lithium Beam Probe and Measurement of Density Pedestal in JT-60U

    NASA Astrophysics Data System (ADS)

    Kojima, Atsushi; Kamiya, Kensaku; Fujita, Takaaki; Kubo, Hirotaka; Iguchi, Harukazu; Oyama, Naoyuki; Suzuki, Takahiro; Kamada, Yutaka; JT-60 Team

    A lithium beam probe (LiBP) has been developed for the measurement of electron density profiles with highly spatial and temporal resolutions in JT-60U. Using an electron beam heating ion source with a capability of 10 mA extraction, a 5.5 mA beam has been injected to the plasmas. It corresponds to the equivalent neutral beam current of 2 mA. A spectrum width of the beam emission has been small enough to separate Zeeman splitting. By use of the LiBP, time evolutions of pedestal density profiles during type I and grassy edge localized modes (ELMs) have been obtained for the first time. After a type I ELM crash, the drop of the line-integrated density measured by an interferometer delays by 2 ms later than that of the pedestal density. Comparing the line-integrated density to the line integration of the edge density profile measured by the LiBP, it is found that the recovery from the type I ELM crash is correlated with the reduction of core plasma density. As for grassy ELMs, grassy ELMs have smaller density crashes than that of type I ELMs, which is mainly derived from the narrower ELM affected area.

  9. Improved kinetic neoclassical transport calculation for a low-collisionality QH-mode pedestal

    SciTech Connect

    Battaglia, D. J.; Burrell, K. H.; Chang, C. S.; deGrassie, J. S.; Grierson, B. A.; Groebner, R. J.; Hager, R.

    2016-07-15

    The role of neoclassical, anomalous and neutral transport to the overall H-mode pedestal and scrape-off layer (SOL) structure in an ELM-free QH-mode discharge on DIII-D is explored using XGC0, a 5D full-f multi-species particle-in-cell drift-kinetic solver with self-consistent neutral recycling and sheath potentials. The work in this paper builds on previous work aimed at achieving quantitative agreement between the flux-driven simulation and the experimental electron density, impurity density and orthogonal measurements of impurity temperature and flow profiles. Improved quantitative agreement is achieved by performing the calculations with a more realistic electron mass, larger neutral density and including finite-Larmor-radius corrections self-consistently in the drift-kinetic motion of the particles. Consequently, the simulations provide stronger evidence that the radial electric field (Er) in the pedestal is primarily established by the required balance between the loss of high-energy tail main ions against a pinch of colder main ions and impurities. The kinetic loss of a small population of ions carrying a large proportion of energy and momentum leads to a separation of the particle and energy transport rates and introduces a source of intrinsic edge torque. Ion orbit loss and finite orbit width effects drive the energy distributions away from Maxwellian, and describe the anisotropy, poloidal asymmetry and local minimum near the separatrix observed in the Ti profile.

  10. Pedestal density fluctuation dynamics during the inter-ELM cycle in DIII-D a)

    NASA Astrophysics Data System (ADS)

    Yan, Z.; McKee, G. R.; Groebner, R. J.; Snyder, P. B.; Osborne, T. H.; Beurskens, M. N.; Burrell, K. H.

    2011-05-01

    Detailed 2D measurements of long-wavelength density fluctuations in the pedestal region with beam emission spectroscopy during the period between edge localized modes (ELMs) indicate two distinct bands of fluctuations propagating in opposite poloidal directions in the plasma frame: one lower frequency band (50-150 kHz) advects in the ion-diamagnetic drift direction (ion mode) and a higher frequency band (200-400 kHz) advects in the electron diamagnetic drift direction (electron mode). The ion mode amplitude is modulated with the ELM cycle: it increases rapidly after an ELM and then saturates, similar to the evolution of the pedestal electron pressure and density gradients. The electron mode, in contrast, has no significant time evolution between ELMs. The decorrelation time of the ion mode is <5 μs [τc(cs/csa a)≤1], the radial correlation length is of order 10 ρi and has poloidal wave-number kθρi~0.1, and the mode advects at near the ion diamagnetic velocity in the plasma frame. These spatiotemporal dynamics are qualitatively similar to features predicted for kinetic ballooning modes.

  11. Calibration of Model for Tokamak H-mode Pedestal and ELMs

    NASA Astrophysics Data System (ADS)

    MacDonald, C.; Bateman, G.; Kritz, A. H.; McElhenny, J.; Osborne, T.; Pankin, A. Y.

    2004-11-01

    Experimental data is used to calibrate a model for the pedestal and Edge Localized Modes (ELMs) implemented in the ASTRA integrated code. The model is calibrated to predict the frequency of the ELMs and the height of the electron and ion temperature pedestals just before an ELM crash. Detailed comparisons are made with experimental data from the DIII-D 98889 discharge, in which the noise in the data is reduced by overlaying the plasma profiles from a sequence of consecutive, nearly identical ELM cycles. The model includes neoclassical transport and transport driven by ion drift modes, resistive ballooning modes, and the electron gradient temperature mode. The criterion for triggering ELM crashes allows for access to second stability. The calibration is carried out by adjusting: (1) The flow shear rates for individual modes of long wavelength turbulent transport; (2) the stability criterion that is used to trigger ELM crashes; and (3) the shapes of the plasma profiles and plasma energy lost after each ELM crash. The calibration is presented as well as the sensitivity to the coefficients in the model.

  12. OVERVIEW OF H-MODE PEDESTAL RESEARCH ON DIII-D

    SciTech Connect

    T.H. OSBORNE; K.H. BURRELL; T.N. CARLSTROM; M.S. CHU; E.J. DOYLE; J.R. FERRON; R.J. GROEBNER; R.J. LA HAYE; L.L. LAO; A.W. LEONARD; M.A. MAHDAVI; G.R. PORTER; P.B. SNYDER; E.J. STRAIT; G.M. STAEBLER; D.M. THOMAS; A.D. TURNBULL; M.R. WADE; THE DIII-D TEAM

    2001-07-01

    Developing an understanding of the processes that control the H-mode transport barrier is motivated by the significant impact this small region (typically <2% of the minor radius) can have on overall plasma performance. Conditions at the inner edge of the H-mode transport barrier can strongly influence the overall energy confinement, and the maximum density, and therefore fusion power, that can be achieved with the typically flat H-mode density profiles [1,2]. The ELM instability, which usually regulates the pressure gradient in the H-mode edge, can result in large power loads to, and erosion of, the divertor targets in a reactor scale device [3]. The goal of H-mode pedestal research at DIII-D is to: (1) develop a physics based model that would allow prediction of the conditions at the top of the H-mode pedestal, (2) develop an understanding of processes which control Type I ELM effects in the core and divertor, and (3) explore alternatives to the Type I ELM regime.

  13. Pedestal density fluctuation dynamics during the inter-ELM cycle in DIII-D

    SciTech Connect

    Yan, Z.; McKee, G. R.; Groebner, R. J.; Snyder, P. B.; Osborne, T. H.; Burrell, K. H.; Beurskens, M. N.

    2011-05-15

    Detailed 2D measurements of long-wavelength density fluctuations in the pedestal region with beam emission spectroscopy during the period between edge localized modes (ELMs) indicate two distinct bands of fluctuations propagating in opposite poloidal directions in the plasma frame: one lower frequency band (50-150 kHz) advects in the ion-diamagnetic drift direction (ion mode) and a higher frequency band (200-400 kHz) advects in the electron diamagnetic drift direction (electron mode). The ion mode amplitude is modulated with the ELM cycle: it increases rapidly after an ELM and then saturates, similar to the evolution of the pedestal electron pressure and density gradients. The electron mode, in contrast, has no significant time evolution between ELMs. The decorrelation time of the ion mode is <5 {mu}s[{tau}{sub c}(c{sub s}/c{sub s}aa){<=}1], the radial correlation length is of order 10 {rho}{sub i} and has poloidal wave-number k{sub {theta}{rho}i{approx}}0.1, and the mode advects at near the ion diamagnetic velocity in the plasma frame. These spatiotemporal dynamics are qualitatively similar to features predicted for kinetic ballooning modes.

  14. Improved kinetic neoclassical transport calculation for a low-collisionality QH-mode pedestal

    NASA Astrophysics Data System (ADS)

    Battaglia, D. J.; Burrell, K. H.; Chang, C. S.; deGrassie, J. S.; Grierson, B. A.; Groebner, R. J.; Hager, R.

    2016-08-01

    The role of neoclassical, anomalous and neutral transport to the overall H-mode pedestal and scrape-off layer (SOL) structure in an ELM-free QH-mode discharge on DIII-D is explored using XGC0, a 5D full-f multi-species particle-in-cell drift-kinetic solver with self-consistent neutral recycling and sheath potentials. The work in this paper builds on previous work aimed at achieving quantitative agreement between the flux-driven simulation and the experimental electron density, impurity density and orthogonal measurements of impurity temperature and flow profiles. Improved quantitative agreement is achieved by performing the calculations with a more realistic electron mass, larger neutral density and including finite-Larmor-radius corrections self-consistently in the drift-kinetic motion of the particles. Consequently, the simulations provide stronger evidence that the radial electric field ({{E}\\text{r}} ) in the pedestal is primarily established by the required balance between the loss of high-energy tail main ions against a pinch of colder main ions and impurities. The kinetic loss of a small population of ions carrying a large proportion of energy and momentum leads to a separation of the particle and energy transport rates and introduces a source of intrinsic edge torque. Ion orbit loss and finite orbit width effects drive the energy distributions away from Maxwellian, and describe the anisotropy, poloidal asymmetry and local minimum near the separatrix observed in the {{T}i} profile.

  15. Improved kinetic neoclassical transport calculation for a low-collisionality QH-mode pedestal

    SciTech Connect

    Battaglia, D. J.; Burrell, K. H.; Chang, C. S.; deGrassie, J. S.; Grierson, B. A.; Groebner, R. J.; Hager, R.

    2016-07-15

    The role of neoclassical, anomalous and neutral transport to the overall H-mode pedestal and scrape-off layer (SOL) structure in an ELM-free QH-mode discharge on DIII-D is explored using XGC0, a 5D full-f multi-species particle-in-cell drift-kinetic solver with self-consistent neutral recycling and sheath potentials. The work in this paper builds on previous work aimed at achieving quantitative agreement between the flux-driven simulation and the experimental electron density, impurity density and orthogonal measurements of impurity temperature and flow profiles. Improved quantitative agreement is achieved by performing the calculations with a more realistic electron mass, larger neutral density and including finite-Larmor-radius corrections self-consistently in the drift-kinetic motion of the particles. Consequently, the simulations provide stronger evidence that the radial electric field (E-r) in the pedestal is primarily established by the required balance between the loss of high-energy tail main ions against a pinch of colder main ions and impurities. The kinetic loss of a small population of ions carrying a large proportion of energy and momentum leads to a separation of the particle and energy transport rates and introduces a source of intrinsic edge torque. Ion orbit loss and finite orbit width effects drive the energy distributions away from Maxwellian, and describe the anisotropy, poloidal asymmetry and local minimum near the separatrix observed in the T-i profile.

  16. Progress in understanding the enhanced pedestal H-mode in NSTX

    DOE PAGES

    Gerhardt, S. P.; Canik, J. M.; Maingi, R.; ...

    2014-08-01

    The paper describes the enhanced pedestal (EP) H-mode observed in the National Spherical Torus Experiment (NSTX). The defining characteristics of EP H-mode are given, namely i)transition after the L- to H-mode transition, ii) region of very steep ion temperature gradient, and iii) associated region of strong rotational shear. A newly observed long-pulse EP H-mode example shows quiescent behavior for as long as the heating and current drive sources are maintained. Cases are shown where the region of steep ion temperature gradient is located at the very edge, and cases where it is shifted up to 10 cm inward from themore » plasma edge; these cases are united by a common dependence of the ion temperature gradient on the toroidal rotation frequency shear. EP H-mode examples have been observed across a wide range of q95 and pedestal collisionality. No strong changes in the fluctuation amplitudes have been observed following the eP H-mode transition, and transport analysis indicates that the ion t hermal transport is comparable to or less than anticipated from a simple neoclassical transport model. Cases are shown where EP H-modes were reliably generated, through these low-q95 examples were difficult to sustain. A case where an externally triggered ELM precipitates the transition to EP H-mode is also shown, though an initial experiment designed to trigger EP-H-modes in this fashion was successful.« less

  17. Neural-network accelerated fusion simulation with self-consistent core-pedestal coupling

    NASA Astrophysics Data System (ADS)

    Meneghini, O.; Candy, J.; Snyder, P. B.; Staebler, G.; Belli, E.

    2016-10-01

    Practical fusion Whole Device Modeling (WDM) simulations require the ability to perform predictions that are fast, but yet account for the sensitivity of the fusion performance to the boundary constraint that is imposed by the pedestal structure of H-mode plasmas due to the stiff core transport models. This poster presents the development of a set of neural-network (NN) models for the pedestal structure (as predicted by the EPED model), and the neoclassical and turbulent transport fluxes (as predicted by the NEO and TGLF codes, respectively), and their self-consistent coupling within the TGYRO transport code. The results are benchmarked with the ones obtained via the coupling scheme described in [Meneghini PoP 2016]. By substituting the most demanding codes with their NN-accelerated versions, the solution can be found at a fraction of the computation cost of the original coupling scheme, thereby combining the accuracy of a high-fidelity model with the fast turnaround time of a reduced model. Work supported by U.S. DOE DE-FC02-04ER54698 and DE-FG02-95ER54309.

  18. Collisional Ion and Electron Scale Gyrokinetic Simulations in the Tokamak Pedestal

    NASA Astrophysics Data System (ADS)

    Belli, E. A.; Candy, J.; Snyder, P. B.

    2016-10-01

    A new gyrokinetic solver, CGYRO, has been developed for precise studies of high collisionality regimes, such as the H-mode pedestal and L-mode edge. Building on GYRO and NEO, CGYRO uses the same velocity-space coordinates as NEO to optimize the accuracy of the collision dynamics and allow for advanced operators beyond the standard Lorentz pitch-angle scattering model. These advanced operators include energy diffusion and finite-FLR collisional effects. The code is optimized for multiscale (coupled electron and ion turbulence scales) simulations, employing a new spatial discretization and array distribution scheme that targets scalability on next-generation (exascale) HPC systems. In this work, CGYRO is used to study the complex spectrum of modes in the pedestal region. The onset of the linear KBM with full collisional effects is assessed to develop an improved KBM/RBM model for EPED. The analysis is extended to high k to explore the role of electron-scale (ETG-range) physics. Comparisons with new analytic collisional theories are made. Inclusion of sonic toroidal rotation (including full centrifugal effects) for studies including heavy wall impurities is also reported. Work supported in part by the US DOE under DE-FC02-06ER54873 and DE-FC02-08ER54963.

  19. TccP2-mediated subversion of actin dynamics by EPEC 2 - a distinct evolutionary lineage of enteropathogenic Escherichia coli.

    PubMed

    Whale, Andrew D; Hernandes, Rodrigo T; Ooka, Tadasuke; Beutin, Lothar; Schüller, Stephanie; Garmendia, Junkal; Crowther, Lynette; Vieira, Mônica A M; Ogura, Yoshitoshi; Krause, Gladys; Phillips, Alan D; Gomes, Tania A T; Hayashi, Tetsuya; Frankel, Gad

    2007-06-01

    Enteropathogenic Escherichia coli (EPEC) is a major cause of infantile diarrhoea in developing countries. While colonizing the gut mucosa, EPEC triggers extensive actin-polymerization activity at the site of intimate bacterial attachment, which is mediated by avid interaction between the outer-membrane adhesin intimin and the type III secretion system (T3SS) effector Tir. The prevailing dogma is that actin polymerization by EPEC is achieved following tyrosine phosphorylation of Tir, recruitment of Nck and activation of neuronal Wiskott-Aldrich syndrome protein (N-WASP). In closely related enterohaemorrhagic E. coli (EHEC) O157 : H7, actin polymerization is triggered following recruitment of the T3SS effector TccP/EspF(U) (instead of Nck) and local activation of N-WASP. In addition to tccP, typical EHEC O157 : H7 harbour a pseudogene (tccP2). However, it has recently been found that atypical, sorbitol-fermenting EHEC O157 carries functional tccP and tccP2 alleles. Interestingly, intact tccP2 has been identified in the incomplete genome sequence of the prototype EPEC strain B171 (serotype O111 : H-), but it is missing from another prototype EPEC strain E2348/69 (O127 : H7). E2348/69 and B171 belong to two distinct evolutionary lineages of EPEC, termed EPEC 1 and EPEC 2, respectively. Here, it is reported that while both EPEC 1 and EPEC 2 triggered actin polymerization via the Nck pathway, tccP2 was found in 26 of 27 (96.2 %) strains belonging to EPEC 2, and in none of the 34 strains belonging to EPEC 1. It was shown that TccP2 was: (i) translocated by the locus of enterocyte effacement-encoded T3SS; (ii) localized at the tip of the EPEC 2-induced actin-rich pedestals in infected HeLa cells and human intestinal in vitro organ cultures ex vivo; and (iii) essential for actin polymerization in infected Nck-/- cells. Therefore, unlike strains belonging to EPEC 1, strains belonging to EPEC 2 can trigger actin polymerization using both Nck and TccP2 actin

  20. Native globular actin has a thermodynamically unstable quasi-stationary structure with elements of intrinsic disorder.

    PubMed

    Kuznetsova, Irina M; Povarova, Olga I; Uversky, Vladimir N; Turoverov, Konstantin K

    2016-02-01

    The native form of globular actin, G-actin, is formed in vivo as a result of complex post-translational folding processes that require ATP energy expenditure and are assisted by the 70 kDa heat shock protein, prefoldin and chaperonin containing TCP-1. G-actin is stabilized by the binding of one ATP molecule and one Ca(2+) ion (or Mg(2+) in vivo). Chemical denaturants, heating or Ca(2+) removal transform native actin (N) into 'inactivated actin' (I), a compact oligomer comprising 14-16 subunits. Viscogenic and crowding agents slow this process but do not stop it. The lack of calcium in the solution accelerates the spontaneous N → I transition. Thus, native G-actin has a kinetically stable (as a result of the high free energy barrier between the N and I states) but thermodynamically unstable structure, which, in the absence of Ca(2+) or other bivalent metal ions, spontaneously converts to the thermodynamically stable I state. It was noted that native actin has much in common with intrinsically disordered proteins: it has functionally important disordered regions; it is constantly in complex with one of its numerous partners; and it plays key roles in many cellular processes, in a manner similar to disordered hub proteins. By analyzing actin folding in vivo and unfolding in vitro, we advanced the hypothesis that proteins in a native state may have a thermodynamically unstable quasi-stationary structure. The kinetically stable native state of these proteins appears forcibly under the influence of intracellular folding machinery. The denaturation of such proteins is always irreversible because the inactivated state, for which the structure is determined by the amino acid sequence of a protein, comprises the thermodynamically stable state under physiological conditions.

  1. Early events of fertilization in sea urchin eggs are sensitive to actin-binding organic molecules.

    PubMed

    Chun, Jong T; Limatola, Nunzia; Vasilev, Filip; Santella, Luigia

    2014-08-01

    We previously demonstrated that many aspects of the intracellular Ca(2+) increase in fertilized eggs of starfish are significantly influenced by the state of the actin cytoskeleton. In addition, the actin cytoskeleton appeared to play comprehensive roles in modulating cortical granules exocytosis and sperm entry during the early phase of fertilization. In the present communication, we have extended our work to sea urchin which is believed to have bifurcated from the common ancestor in the phylogenetic tree some 500 million years ago. To corroborate our earlier findings in starfish, we have tested how the early events of fertilization in sea urchin eggs are influenced by four different actin-binding drugs that promote either depolymerization or stabilization of actin filaments. We found that all the actin drugs commonly blocked sperm entry in high doses and significantly reduced the speed of the Ca(2+) wave. At low doses, however, cytochalasin B and phalloidin increased the rate of polyspermy. Overall, certain aspects of Ca(2+) signaling in these eggs were in line with the morphological changes induced by the actin drugs. That is, the time interval between the cortical flash and the first Ca(2+) spot at the sperm interaction site (the latent period) was significantly prolonged in the eggs pretreated with cytochalasin B or latrunculin A, whereas the Ca(2+) decay kinetics after the peak was specifically attenuated in the eggs pretreated with jasplakinolide or phalloidin. In addition, the sperm interacting with the eggs pretreated with actin drugs often generated multiple Ca(2+) waves, but tended to fail to enter the egg. Thus, our results indicated that generation of massive Ca(2+) waves is neither indicative of sperm entry nor sufficient for cortical granules exocytosis in the inseminated sea urchin eggs, whereas the structure and functionality of the actin cytoskeleton are the major determining factors in the two processes. Copyright © 2014 Elsevier Inc. All rights

  2. Mesoscopic model of actin-based propulsion.

    PubMed

    Zhu, Jie; Mogilner, Alex

    2012-01-01

    Two theoretical models dominate current understanding of actin-based propulsion: microscopic polymerization ratchet model predicts that growing and writhing actin filaments generate forces and movements, while macroscopic elastic propulsion model suggests that deformation and stress of growing actin gel are responsible for the propulsion. We examine both experimentally and computationally the 2D movement of ellipsoidal beads propelled by actin tails and show that neither of the two models can explain the observed bistability of the orientation of the beads. To explain the data, we develop a 2D hybrid mesoscopic model by reconciling these two models such that individual actin filaments undergoing nucleation, elongation, attachment, detachment and capping are embedded into the boundary of a node-spring viscoelastic network representing the macroscopic actin gel. Stochastic simulations of this 'in silico' actin network show that the combined effects of the macroscopic elastic deformation and microscopic ratchets can explain the observed bistable orientation of the actin-propelled ellipsoidal beads. To test the theory further, we analyze observed distribution of the curvatures of the trajectories and show that the hybrid model's predictions fit the data. Finally, we demonstrate that the model can explain both concave-up and concave-down force-velocity relations for growing actin networks depending on the characteristic time scale and network recoil. To summarize, we propose that both microscopic polymerization ratchets and macroscopic stresses of the deformable actin network are responsible for the force and movement generation.

  3. Nuclear Actin in Development and Transcriptional Reprogramming

    PubMed Central

    Misu, Shinji; Takebayashi, Marina; Miyamoto, Kei

    2017-01-01

    Actin is a highly abundant protein in eukaryotic cells and dynamically changes its polymerized states with the help of actin-binding proteins. Its critical function as a constituent of cytoskeleton has been well-documented. Growing evidence demonstrates that actin is also present in nuclei, referred to as nuclear actin, and is involved in a number of nuclear processes, including transcriptional regulation and chromatin remodeling. The contribution of nuclear actin to transcriptional regulation can be explained by its direct interaction with transcription machineries and chromatin remodeling factors and by controlling the activities of transcription factors. In both cases, polymerized states of nuclear actin affect the transcriptional outcome. Nuclear actin also plays an important role in activating strongly silenced genes in somatic cells for transcriptional reprogramming. When these nuclear functions of actin are considered, it is plausible to speculate that nuclear actin is also implicated in embryonic development, in which numerous genes need to be activated in a well-coordinated manner. In this review, we especially focus on nuclear actin’s roles in transcriptional activation, reprogramming and development, including stem cell differentiation and we discuss how nuclear actin can be an important player in development and cell differentiation. PMID:28326098

  4. Mesoscopic Model of Actin-Based Propulsion

    PubMed Central

    Zhu, Jie; Mogilner, Alex

    2012-01-01

    Two theoretical models dominate current understanding of actin-based propulsion: microscopic polymerization ratchet model predicts that growing and writhing actin filaments generate forces and movements, while macroscopic elastic propulsion model suggests that deformation and stress of growing actin gel are responsible for the propulsion. We examine both experimentally and computationally the 2D movement of ellipsoidal beads propelled by actin tails and show that neither of the two models can explain the observed bistability of the orientation of the beads. To explain the data, we develop a 2D hybrid mesoscopic model by reconciling these two models such that individual actin filaments undergoing nucleation, elongation, attachment, detachment and capping are embedded into the boundary of a node-spring viscoelastic network representing the macroscopic actin gel. Stochastic simulations of this ‘in silico’ actin network show that the combined effects of the macroscopic elastic deformation and microscopic ratchets can explain the observed bistable orientation of the actin-propelled ellipsoidal beads. To test the theory further, we analyze observed distribution of the curvatures of the trajectories and show that the hybrid model's predictions fit the data. Finally, we demonstrate that the model can explain both concave-up and concave-down force-velocity relations for growing actin networks depending on the characteristic time scale and network recoil. To summarize, we propose that both microscopic polymerization ratchets and macroscopic stresses of the deformable actin network are responsible for the force and movement generation. PMID:23133366

  5. Labeling F-actin barbed ends with rhodamine-actin in permeabilized neuronal growth cones.

    PubMed

    Marsick, Bonnie M; Letourneau, Paul C

    2011-03-17

    The motile tips of growing axons are called growth cones. Growth cones lead navigating axons through developing tissues by interacting with locally expressed molecular guidance cues that bind growth cone receptors and regulate the dynamics and organization of the growth cone cytoskeleton. The main target of these navigational signals is the actin filament meshwork that fills the growth cone periphery and that drives growth cone motility through continual actin polymerization and dynamic remodeling. Positive or attractive guidance cues induce growth cone turning by stimulating actin filament (F-actin) polymerization in the region of the growth cone periphery that is nearer the source of the attractant cue. This actin polymerization drives local growth cone protrusion, adhesion of the leading margin and axonal elongation toward the attractant. Actin filament polymerization depends on the availability of sufficient actin monomer and on polymerization nuclei or actin filament barbed ends for the addition of monomer. Actin monomer is abundantly available in chick retinal and dorsal root ganglion (DRG) growth cones. Consequently, polymerization increases rapidly when free F-actin barbed ends become available for monomer addition. This occurs in chick DRG and retinal growth cones via the local activation of the F-actin severing protein actin depolymerizing factor (ADF/cofilin) in the growth cone region closer to an attractant. This heightened ADF/cofilin activity severs actin filaments to create new F-actin barbed ends for polymerization. The following method demonstrates this mechanism. Total content of F-actin is visualized by staining with fluorescent phalloidin. F-actin barbed ends are visualized by the incorporation of rhodamine-actin within growth cones that are permeabilized with the procedure described in the following, which is adapted from previous studies of other motile cells. When rhodamine-actin is added at a concentration above the critical concentration

  6. The Interaction of Vinculin with Actin

    PubMed Central

    Golji, Javad; Mofrad, Mohammad R. K.

    2013-01-01

    Vinculin can interact with F-actin both in recruitment of actin filaments to the growing focal adhesions and also in capping of actin filaments to regulate actin dynamics. Using molecular dynamics, both interactions are simulated using different vinculin conformations. Vinculin is simulated either with only its vinculin tail domain (Vt), with all residues in its closed conformation, with all residues in an open I conformation, and with all residues in an open II conformation. The open I conformation results from movement of domain 1 away from Vt; the open II conformation results from complete dissociation of Vt from the vinculin head domains. Simulation of vinculin binding along the actin filament showed that Vt alone can bind along the actin filaments, that vinculin in its closed conformation cannot bind along the actin filaments, and that vinculin in its open I conformation can bind along the actin filaments. The simulations confirm that movement of domain 1 away from Vt in formation of vinculin 1 is sufficient for allowing Vt to bind along the actin filament. Simulation of Vt capping actin filaments probe six possible bound structures and suggest that vinculin would cap actin filaments by interacting with both S1 and S3 of the barbed-end, using the surface of Vt normally occluded by D4 and nearby vinculin head domain residues. Simulation of D4 separation from Vt after D1 separation formed the open II conformation. Binding of open II vinculin to the barbed-end suggests this conformation allows for vinculin capping. Three binding sites on F-actin are suggested as regions that could link to vinculin. Vinculin is suggested to function as a variable switch at the focal adhesions. The conformation of vinculin and the precise F-actin binding conformation is dependent on the level of mechanical load on the focal adhesion. PMID:23633939

  7. Small GTPases promote actin coat formation on microsporidian pathogens traversing the apical membrane of Caenorhabditis elegans intestinal cells

    PubMed Central

    Szumowski, Suzannah C.; Estes, Kathleen A.; Popovich, John J.; Botts, Michael R.; Sek, Grace; Troemel, Emily R.

    2017-01-01

    Summary Many intracellular pathogens co-opt actin in host cells, but little is known about these interactions in vivo. We study the in vivo trafficking and exit of the microsporidian Nematocida parisii, which is an intracellular pathogen that infects intestinal cells of the nematode Caenorhabditis elegans. We recently demonstrated that N. parisii uses directional exocytosis to escape out of intestinal cells into the intestinal tract. Here, we show that an intestinal-specific isoform of C. elegans actin called ACT-5 forms coats around membrane compartments that contain single exocytosing spores, and that these coats appear to form after fusion with the apical membrane. We performed a genetic screen for host factors required for actin coat formation and identified small GTPases important for this process. Through analysis of animals defective in these factors, we found that actin coats are not required for pathogen exit although they may boost exocytic output. Later during infection, we find that ACT-5 also forms coats around membrane-bound vesicles that contain multiple spores. These vesicles are likely formed by clathrin-dependent compensatory endocytosis to retrieve membrane material that has been trafficked to the apical membrane as part of the exocytosis process. These findings provide insight into microsporidia interaction with host cells, and provide novel in vivo examples of the manner in which intracellular pathogens co-opt host actin during their life cycle. PMID:26147591

  8. Live Cell Imaging Reveals Structural Associations between the Actin and Microtubule Cytoskeleton in Arabidopsis [W] [OA

    PubMed Central

    Sampathkumar, Arun; Lindeboom, Jelmer J.; Debolt, Seth; Gutierrez, Ryan; Ehrhardt, David W.; Ketelaar, Tijs; Persson, Staffan

    2011-01-01

    In eukaryotic cells, the actin and microtubule (MT) cytoskeletal networks are dynamic structures that organize intracellular processes and facilitate their rapid reorganization. In plant cells, actin filaments (AFs) and MTs are essential for cell growth and morphogenesis. However, dynamic interactions between these two essential components in live cells have not been explored. Here, we use spinning-disc confocal microscopy to dissect interaction and cooperation between cortical AFs and MTs in Arabidopsis thaliana, utilizing fluorescent reporter constructs for both components. Quantitative analyses revealed altered AF dynamics associated with the positions and orientations of cortical MTs. Reorganization and reassembly of the AF array was dependent on the MTs following drug-induced depolymerization, whereby short AFs initially appeared colocalized with MTs, and displayed motility along MTs. We also observed that light-induced reorganization of MTs occurred in concert with changes in AF behavior. Our results indicate dynamic interaction between the cortical actin and MT cytoskeletons in interphase plant cells. PMID:21693695

  9. Adherens and tight junctions: structure, function and connections to the actin cytoskeleton.

    PubMed

    Hartsock, Andrea; Nelson, W James

    2008-03-01

    Adherens junctions and Tight junctions comprise two modes of cell-cell adhesion that provide different functions. Both junctional complexes are proposed to associate with the actin cytoskeleton, and formation and maturation of cell-cell contacts involves reorganization of the actin cytoskeleton. Adherens junctions initiate cell-cell contacts, and mediate the maturation and maintenance of the contact. Adherens junctions consist of the transmembrane protein E-cadherin, and intracellular components, p120-catenin, beta-catenin and alpha-catenin. Tight junctions regulate the paracellular pathway for the movement of ions and solutes in-between cells. Tight junctions consist of the transmembrane proteins occludin and claudin, and the cytoplasmic scaffolding proteins ZO-1, -2, and -3. This review discusses the binding interactions of the most studied proteins that occur within each of these two junctional complexes and possible modes of regulation of these interactions, and the different mechanisms that connect and regulate interactions with the actin cytoskeleton.

  10. Actin cortex architecture regulates cell surface tension.

    PubMed

    Chugh, Priyamvada; Clark, Andrew G; Smith, Matthew B; Cassani, Davide A D; Dierkes, Kai; Ragab, Anan; Roux, Philippe P; Charras, Guillaume; Salbreux, Guillaume; Paluch, Ewa K

    2017-06-01

    Animal cell shape is largely determined by the cortex, a thin actin network underlying the plasma membrane in which myosin-driven stresses generate contractile tension. Tension gradients result in local contractions and drive cell deformations. Previous cortical tension regulation studies have focused on myosin motors. Here, we show that cortical actin network architecture is equally important. First, we observe that actin cortex thickness and tension are inversely correlated during cell-cycle progression. We then show that the actin filament length regulators CFL1, CAPZB and DIAPH1 regulate mitotic cortex thickness and find that both increasing and decreasing thickness decreases tension in mitosis. This suggests that the mitotic cortex is poised close to a tension maximum. Finally, using a computational model, we identify a physical mechanism by which maximum tension is achieved at intermediate actin filament lengths. Our results indicate that actin network architecture, alongside myosin activity, is key to cell surface tension regulation.

  11. Elasticity, adhesion and actin based propulsion

    NASA Astrophysics Data System (ADS)

    Gopinathan, Ajay

    2006-03-01

    When a cells crawls, its shape re-organizes via polymerization and depolymerization of actin filaments. The growing ends of the filaments are oriented towards the outside of the cell, and their polymerization pushes the cell membrane forwards. The same mechanism comes into play when the bacterial pathogen Listeria monocytogenes infects a cell. The bacterium hijacks the host cell's actin machinery to create an actin network (the actin comet tail) that propels the bacterium through cells and into neighboring cells. We propose a mechanism for how polymerization gives rise to motility that incorporates the effects of inhomogeneous polymerization. We treat the actin comet tail as an elastic continuum tethered to the rear of the bacterium. The interplay of polymerization and tethering gives rise to inhomogeneous stresses calculated with a finite element analysis. We quantitatively reproduce many distinctive features of actin propulsion that have been observed experimentally, including stepped motion, hopping, tail shape and the propulsion of flat surfaces.

  12. Actin dynamics in mouse fibroblasts in microgravity

    NASA Astrophysics Data System (ADS)

    Moes, Maarten J. A.; Bijvelt, Jose J.; Boonstra, Johannes

    2007-09-01

    After stimulating with the growth factor PDGF, cells exhibit abundant membrane ruffling and other morphological changes under normal gravity conditions. These morphological changes are largely determined by the actin microfilament system. Now these actin dynamics were studied under microgravity conditions in mouse fibroblasts during the DELTA mission. The aim of the present study was to describe the actin morphology in detail, to establish the effect of PDGF on actin morphology and to study the role of several actin-interacting proteins involved in introduced actin dynamics in microgravity. Identical experiments were conducted at 1G on earth as a reference. No results in microgravity were obtained due to a combination of malfunctioning hardware and unfulfilled temperature requirements.

  13. Polymerization of actin by positively charged liposo