Sample records for cell-surface tethered modulators

  1. Membrane Tether Formation on a Cell Surface with Reservoir

    Institute of Scientific and Technical Information of China (English)

    JIANG Yu-Qiang; GUO Hong-Lian; LIU Chun-Xiang; LI Zhao-Lin; CHENG Bing-Ying; ZHANG Dao-Zhong; JIA Suo-Tang


    @@ We propose a mathematical model to analyse the membrane tether formation process on a cell surface with reservoir. Based on the experimental results, the membrane reservoir density of breast cancer cell was obtained,p = 8.02. The membrane surface viscosity between membrane and environment η is 0.021(pN.s/μm3), and the static force F0 = 5.71 pN.

  2. Tethering of Ficolin-1 to cell surfaces through recognition of sialic acid by the fibrinogen-like domain

    DEFF Research Database (Denmark)

    Honore, C.; Rorvig, S.; Hummelshoj, T.;


    surface is restricted to monocytes and granulocytes. Ficolin-1 is tethered to the cell surface of these cells through its fibrinogen-like domain, and the ligand involved in the binding of Ficolin-1 is shown to be sialic acid. Moreover, rFicolin-1 bound activated but not resting T lymphocytes. Together......, these results demonstrate a novel self-recognition mechanism of leukocytes mediated by the fibrinogen-like domain of Ficolin-1....

  3. BST2/CD317 counteracts human coronavirus 229E productive infection by tethering virions at the cell surface

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Shiu-Mei [Department of Medical Research and Education, Taipei Veterans General Hospital and Institute of Clinical Medicine, Taipei 11217, Taiwan (China); Institute of Clinical Medicine, National Yang-Ming University School of Medicine, Taipei, Taiwan (China); Huang, Kuo-Jung [Department of Medical Research and Education, Taipei Veterans General Hospital and Institute of Clinical Medicine, Taipei 11217, Taiwan (China); Wang, Chin-Tien, E-mail: [Department of Medical Research and Education, Taipei Veterans General Hospital and Institute of Clinical Medicine, Taipei 11217, Taiwan (China); Institute of Clinical Medicine, National Yang-Ming University School of Medicine, Taipei, Taiwan (China)


    Bone marrow stromal antigen 2 (BST2), an interferon-inducible antiviral factor, has been shown to block the release of various enveloped viruses from cells. It has also been identified as an innate immune system component. Most enveloped viruses subject to BST2 restriction bud at the plasma membrane. Here we report our findings that (a) the production of human coronavirus 229E (HCoV-229E) progeny viruses, whose budding occurs at the ER-Golgi intermediate compartment (ERGIC), markedly decreases in the presence of BST2; and (b) BST2 knockdown expression results in enhanced HCoV-229E virion production. Electron microscopy analyses indicate that HCoV-229E virions are tethered to cell surfaces or intracellular membranes by BST2. Our results suggest that BST2 exerts a broad blocking effect against enveloped virus release, regardless of whether budding occurs at the plasma membrane or intracellular compartments. - Highlights: • BST2 knockdown expression results in enhanced HCoV-229E egress. • HCoV-229E virions are tethered to cell surfaces or intracellular membranes by BST2. • HCoV-229E infection at high MOI can significantly downregulate HeLa BST2 and rescue HIV-1 egress.

  4. The Biological Function of the Prion Protein: A Cell Surface Scaffold of Signaling Modules (United States)

    Linden, Rafael


    The prion glycoprotein (PrPC) is mostly located at the cell surface, tethered to the plasma membrane through a glycosyl-phosphatydil inositol (GPI) anchor. Misfolding of PrPC is associated with the transmissible spongiform encephalopathies (TSEs), whereas its normal conformer serves as a receptor for oligomers of the β-amyloid peptide, which play a major role in the pathogenesis of Alzheimer’s Disease (AD). PrPC is highly expressed in both the nervous and immune systems, as well as in other organs, but its functions are controversial. Extensive experimental work disclosed multiple physiological roles of PrPC at the molecular, cellular and systemic levels, affecting the homeostasis of copper, neuroprotection, stem cell renewal and memory mechanisms, among others. Often each such process has been heralded as the bona fide function of PrPC, despite restricted attention paid to a selected phenotypic trait, associated with either modulation of gene expression or to the engagement of PrPC with a single ligand. In contrast, the GPI-anchored prion protein was shown to bind several extracellular and transmembrane ligands, which are required to endow that protein with the ability to play various roles in transmembrane signal transduction. In addition, differing sets of those ligands are available in cell type- and context-dependent scenarios. To account for such properties, we proposed that PrPC serves as a dynamic platform for the assembly of signaling modules at the cell surface, with widespread consequences for both physiology and behavior. The current review advances the hypothesis that the biological function of the prion protein is that of a cell surface scaffold protein, based on the striking similarities of its functional properties with those of scaffold proteins involved in the organization of intracellular signal transduction pathways. Those properties are: the ability to recruit spatially restricted sets of binding molecules involved in specific signaling

  5. The ATPases of cohesin interface with regulators to modulate cohesin-mediated DNA tethering. (United States)

    Çamdere, Gamze; Guacci, Vincent; Stricklin, Jeremiah; Koshland, Douglas


    Cohesin tethers together regions of DNA, thereby mediating higher order chromatin organization that is critical for sister chromatid cohesion, DNA repair and transcriptional regulation. Cohesin contains a heterodimeric ATP-binding Cassette (ABC) ATPase comprised of Smc1 and Smc3 ATPase active sites. These ATPases are required for cohesin to bind DNA. Cohesin's DNA binding activity is also promoted by the Eco1 acetyltransferase and inhibited by Wpl1. Recently we showed that after cohesin stably binds DNA, a second step is required for DNA tethering. This second step is also controlled by Eco1 acetylation. Here, we use genetic and biochemical analyses to show that this second DNA tethering step is regulated by cohesin ATPase. Furthermore, our results also suggest that Eco1 promotes cohesion by modulating the ATPase cycle of DNA-bound cohesin in a state that is permissive for DNA tethering and refractory to Wpl1 inhibition.

  6. Phosphorylation and chronic agonist treatment atypically modulate GABAB receptor cell surface stability. (United States)

    Fairfax, Benjamin P; Pitcher, Julie A; Scott, Mark G H; Calver, Andrew R; Pangalos, Menelas N; Moss, Stephen J; Couve, Andrés


    GABA(B) receptors are heterodimeric G protein-coupled receptors that mediate slow synaptic inhibition in the central nervous system. The dynamic control of the cell surface stability of GABA(B) receptors is likely to be of fundamental importance in the modulation of receptor signaling. Presently, however, this process is poorly understood. Here we demonstrate that GABA(B) receptors are remarkably stable at the plasma membrane showing little basal endocytosis in cultured cortical and hippocampal neurons. In addition, we show that exposure to baclofen, a well characterized GABA(B) receptor agonist, fails to enhance GABA(B) receptor endocytosis. Lack of receptor internalization in neurons correlates with an absence of agonist-induced phosphorylation and lack of arrestin recruitment in heterologous systems. We also demonstrate that chronic exposure to baclofen selectively promotes endocytosis-independent GABA(B) receptor degradation. The effect of baclofen can be attenuated by activation of cAMP-dependent protein kinase or co-stimulation of beta-adrenergic receptors. Furthermore, we show that increased degradation rates are correlated with reduced receptor phosphorylation at serine 892 in GABA(B)R2. Our results support a model in which GABA(B)R2 phosphorylation specifically stabilizes surface GABA(B) receptors in neurons. We propose that signaling pathways that regulate cAMP levels in neurons may have profound effects on the tonic synaptic inhibition by modulating the availability of GABA(B) receptors.

  7. The cell surface receptor Slamf6 modulates innate immune responses during Citrobacter rodentium-induced colitis. (United States)

    van Driel, Boaz; Wang, Guoxing; Liao, Gongxian; Halibozek, Peter J; Keszei, Marton; O'Keeffe, Michael S; Bhan, Atul K; Wang, Ninghai; Terhorst, Cox


    The homophilic cell surface receptors CD150 (Slamf1) and CD352 (Slamf6) are known to modulate adaptive immune responses. Although the Th17 response was enhanced in Slamf6(-/-) C57BL/6 mice upon oral infection with Citrobacter rodentium, the pathologic consequences are indistinguishable from an infection of wild-type C57BL/6 mice. Using a reporter-based binding assay, we show that Slamf6 can engage structures on the outer cell membrane of several Gram(-) bacteria. Therefore, we examined whether Slamf6, like Slamf1, is also involved in innate responses to bacteria and regulates peripheral inflammation by assessing the outcome of C. rodentium infections in Rag(-/-) mice. Surprisingly, the pathology and immune responses in the lamina propria of C. rodentium-infected Slamf6(-/-) Rag(-/-) mice were markedly reduced as compared with those of Rag(-/-) mice. Infiltration of inflammatory phagocytes into the lamina propria was consistently lower in Slamf6(-/-) Rag(-/-) mice than in Rag(-/-) animals. Concomitant with the reduced systemic translocation of the bacteria was an enhanced production of IL-22, suggesting that Slamf6 suppresses a mucosal protective program. Furthermore, administering a mAb (330) that inhibits bacterial interactions with Slamf6 to Rag(-/-) mice ameliorated the infection compared with a control antibody. We conclude that Slamf6-mediated interactions of colonic innate immune cells with specific Gram(-) bacteria reduce mucosal protection and enhance inflammation, contributing to lethal colitis that is caused by C. rodentium infections in Rag(-/-) mice.

  8. Graphene Oxide Modulates B Cell Surface Phenotype and Impairs Immunoglobulin Secretion in Plasma Cell. (United States)

    Xu, Shaohai; Xu, Shengmin; Chen, Shaopeng; Fan, Huadong; Luo, Xun; Yang, Xiaoyao; Wang, Jun; Yuan, Hang; Xu, An; Wu, Lijun


    Since discovery, graphene oxide (GO) has been used in all aspects of human life and revealed promising applications in biomedicine. Nevertheless, the potential risks of GO were always being revealed. Although GO was found to induce immune cell death and innate immune response, little is known regarding its toxicity to the specific adaptive immune system that is crucial for protecting against exotic invasion. The B-cell mediated adaptive immune system, which composed of highly specialized cells (B and plasma cell) and specific immune response (antibody response) is the focus in our present study. Using diverse standard immunological techniques, we found that GO modulated B cell surface phenotype, both costimulatory molecules (CD80, CD86 and especially CD40) and antigen presenting molecules (both classical and nonclassical) under the condition without causing cell death. Meanwhile, the terminal differentiated immunoglobulin (Ig) secreting plasma cell was affected by GO, which displayed a less secretion of Ig and more severe ER stress caused by the retention of the secreted form of Ig in cell compartment. The combined data reveal that GO has a particular adverse effect to B cell and the humoral immunity, directly demonstrating the potential risk of GO to the specific adaptive immunity.

  9. The modulation of cell surface cAMP receptors from Dictyostelium disscoideum by ammonium sulfate

    NARCIS (Netherlands)

    Haastert, Peter J.M. van


    Dictyostelium discoideum cells contain a heterogeneous population of cell surface cAMP receptors with components possessing different affinities (Kd between 15 and 450 nM) and different off-rates of the cAMP-receptor complex (t½ between 0.7 and 150 s). The association of cAMP to the receptor and the

  10. Tumor suppressor protein SMAR1 modulates the roughness of cell surface: combined AFM and SEM study

    Directory of Open Access Journals (Sweden)

    Mamgain Hitesh


    Full Text Available Abstract Background Imaging tools such as scanning electron microscope (SEM and atomic force microscope (AFM can be used to produce high-resolution topographic images of biomedical specimens and hence are well suited for imaging alterations in cell morphology. We have studied the correlation of SMAR1 expression with cell surface smoothness in cell lines as well as in different grades of human breast cancer and mouse tumor sections. Methods We validated knockdown and overexpression of SMAR1 using RT-PCR as well as Western blotting in human embryonic kidney (HEK 293, human breast cancer (MCF-7 and mouse melanoma (B16F1 cell lines. The samples were then processed for cell surface roughness studies using atomic force microscopy (AFM and scanning electron microscopy (SEM. The same samples were used for microarray analysis as well. Tumors sections from control and SMAR1 treated mice as well as tissues sections from different grades of human breast cancer on poly L-lysine coated slides were used for AFM and SEM studies. Results Tumor sections from mice injected with melanoma cells showed pronounced surface roughness. In contrast, tumor sections obtained from nude mice that were first injected with melanoma cells followed by repeated injections of SMAR1-P44 peptide, exhibited relatively smoother surface profile. Interestingly, human breast cancer tissue sections that showed reduced SMAR1 expression exhibited increased surface roughness compared to the adjacent normal breast tissue. Our AFM data establishes that treatment of cells with SMAR1-P44 results into increase in cytoskeletal volume that is supported by comparative gene expression data showing an increase in the expression of specific cytoskeletal proteins compared to the control cells. Altogether, these findings indicate that tumor suppressor function of SMAR1 might be exhibited through smoothening of cell surface by regulating expression of cell surface proteins. Conclusion Tumor suppressor

  11. Guanine Nucleotides Modulate Cell Surface cAMP-Binding Sites in Membranes from Dictyostelium discoideum

    NARCIS (Netherlands)

    Haastert, Peter J.M. van


    D. discoideum contains kinetically distinguishable cell surface cAMP binding sites. One class, S, is slowly dissociating and has high affinity for cAMP (Kd = 15 nM, t½ = 15 s). A second class is fast dissociating (t½ about 1 s) and is composed of high affinity binding sites H (Kd ≈ 60 nM), and low a

  12. Modulation of cell surface GABA B receptors by desensitization,trafficking and regulated degradation

    Institute of Scientific and Technical Information of China (English)

    Dietmar; Benke; Khaled; Zemoura; Patrick; J; Maier


    Inhibitory neurotransmission ensures normal brain function by counteracting and integrating excitatory activity.-Aminobutyric acid (GABA) is the main inhibitory neurotransmitter in the mammalian central nervous system,and mediates its effects via two classes of receptors:the GABA A and GABA B receptors.GABA A receptors are heteropentameric GABA-gated chloride channels and responsible for fast inhibitory neurotransmission.GABA B receptors are heterodimeric G protein coupled receptors (GPCR) that mediate slow and prolonged inhibitory transmission.The extent of inhibitory neurotransmission is determined by a variety of factors,such as the degree of transmitter release and changes in receptor activity by posttranslational modifications (e.g.,phosphorylation),as well as by the number of receptors present in the plasma membrane available for signal transduction.The level of GABA B receptors at the cell surface critically depends on the residence time at the cell surface and finally the rates of endocytosis and degradation.In this review we focus primarily on recent advances in the understanding of trafficking mechanisms that determine the expression level of GABA B receptors in the plasma membrane,and thereby signaling strength.

  13. The Clostridium difficile Protease Cwp84 Modulates both Biofilm Formation and Cell-Surface Properties.

    Directory of Open Access Journals (Sweden)

    Véronique Pantaléon

    Full Text Available Clostridium difficile is responsible for 15-20% of antibiotic-associated diarrheas, and nearly all cases of pseudomembranous colitis. Among the cell wall proteins involved in the colonization process, Cwp84 is a protease that cleaves the S-layer protein SlpA into two subunits. A cwp84 mutant was previously shown to be affected for in vitro growth but not in its virulence in a hamster model. In this study, the cwp84 mutant elaborated biofilms with increased biomass compared with the parental strain, allowing the mutant to grow more robustly in the biofilm state. Proteomic analyses of the 630Δerm bacteria growing within the biofilm revealed the distribution of abundant proteins either in cell surface, matrix or supernatant fractions. Of note, the toxin TcdA was found in the biofilm matrix. Although the overall proteome differences between the cwp84 mutant and the parental strains were modest, there was still a significant impact on bacterial surface properties such as altered hydrophobicity. In vitro and in vivo competition assays revealed that the mutant was significantly impaired for growth only in the planktonic state, but not in biofilms or in vivo. Taken together, our results suggest that the phenotypes in the cwp84 mutant come from either the accumulation of uncleaved SlpA, or the ability of Cwp84 to cleave as yet undetermined proteins.

  14. Monocyte cell surface glycosaminoglycans positively modulate IL-4-induced differentiation toward dendritic cells.

    NARCIS (Netherlands)

    Dekker, E. den; Grefte, S.; Huijs, T.; Dam, G.B. ten; Versteeg, E.M.M.; Berk, L.C.J. van den; Bladergroen, B.A.; Kuppevelt, A.H.M.S.M. van; Figdor, C.G.; Torensma, R.


    IL-4 induces the differentiation of monocytes toward dendritic cells (DCs). The activity of many cytokines is modulated by glycosaminoglycans (GAGs). In this study, we explored the effect of GAGs on the IL-4-induced differentiation of monocytes toward DCs. IL-4 dose-dependently up-regulated the expr

  15. The modulation of biodistribution of stem cells by anchoring lipid-conjugated heparin on the cell surface. (United States)

    Kim, Jong Chul; Tae, Giyoong


    Heparin is a bioactive glycosaminoglycan that can interact with various extracellular matrix (ECM) proteins and growth factors. Lipid-conjugated heparin was synthesized, and was used to coat adipose-derived stem cells (ADSCs) by physical insertion on the cell membrane. Coating of lipid-conjugated heparin with two lipid moieties on ADSCs was stable for 24h in vitro. Biodistribution of heparin-coated ADSCs upon intravenous injection in mice was analyzed by In-Vivo Imaging System (IVIS), and showed enhanced accumulation in the liver and spleen while reduced entrapment in the lung. Thus, the coating of ADSCs with lipid-conjugated heparin could significantly modulate the biodistribution of cells.

  16. β1- and β3- voltage-gated sodium channel subunits modulate cell surface expression and glycosylation of Nav1.7 in HEK293 cells

    Directory of Open Access Journals (Sweden)

    Cedric James Laedermann


    Full Text Available Voltage-gated sodium channels (Navs are glycoproteins composed of a pore-forming α-subunit and associated β-subunits that regulate Nav α-subunit plasma membrane density and biophysical properties. Glycosylation of the Nav α-subunit also directly affects Navs gating. β-subunits and glycosylation thus comodulate Nav α-subunit gating. We hypothesized that β-subunits could directly influence α-subunit glycosylation. Whole-cell patch clamp of HEK293 cells revealed that both β1- and β3-subunits coexpression shifted V1/2 of steady-state activation and inactivation and increased Nav1.7-mediated INa density. Biotinylation of cell surface proteins, combined with the use of deglycosydases, confirmed that Nav1.7 α-subunits exist in multiple glycosylated states. The α-subunit intracellular fraction was found in a core-glycosylated state, migrating at approximately 250 kDa. At the plasma membrane, in addition to the core-glycosylated form, a fully glycosylated form of Nav1.7 (~280 kDa was observed. This higher band shifted to an intermediate band (~260 kDa when β1-subunits were coexpressed, suggesting that the β1-subunit promotes an alternative glycosylated form of Nav1.7. Furthermore, the β1-subunit increased the expression of this alternative glycosylated form and the β3-subunit increased the expression of the core-glycosylated form of Nav1.7. This study describes a novel role for β1- and β3-subunits in the modulation of Nav1.7 α-subunit glycosylation and cell surface expression.

  17. Site-directed mutagenesis of HIV-1 vpu gene demonstrates two clusters of replication-defective mutants with distinct ability to down-modulate cell surface CD4 and tetherin

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    Masako Nomaguchi


    Full Text Available HIV-1 Vpu acts positively on viral infectivity by mediating CD4 degradation in endoplasmic reticulum and enhances virion release by counteracting a virion release restriction factor, tetherin. In order to define the impact of Vpu activity on HIV-1 replication, we have generated a series of site-specific proviral vpu mutants. Of fifteen mutants examined, seven exhibited a replication-defect similar to that of a vpu-deletion mutant in a lymphocyte cell line H9. These mutations clustered in narrow regions within transmembrane domain (TMD and cytoplasmic domain (CTD. Replication-defective mutants displayed the reduced ability to enhance virion release from a monolayer cell line HEp2 without exception. Upon transfection with Vpu expression vectors, neither TMD mutants nor CTD mutants blocked CD4 expression at the cell surface in another monolayer cell line MAGI. While TMD mutants were unable to down-modulate cell surface tetherin in HEp2 cells, CTD mutants did quite efficiently. Confocal microscopy analysis revealed the difference of intracellular localization between TMD and CTD mutants. In total, replication capability of HIV-1 carrying vpu mutations correlates well with the ability of Vpu to enhance virion release and to impede the cell surface expression of CD4 but not with the ability to down-modulate cell surface tetherin. Our results here suggest that efficient viral replication requires not only down-regulation of cell surface tetherin but also its degradation.

  18. Mechanistic target of rapamycin (mTOR) regulates trophoblast folate uptake by modulating the cell surface expression of FR-α and the RFC. (United States)

    Rosario, Fredrick J; Powell, Theresa L; Jansson, Thomas


    Folate deficiency in fetal life is strongly associated with structural malformations and linked to intrauterine growth restriction. In addition, limited availability of methyl donors, such as folate, during pregnancy may result in abnormal gene methylation patterns and contribute to developmental programming. The fetus is dependent on placental transfer of folate, however the molecular mechanisms regulating placental folate transport are unknown. We used cultured primary human trophoblast cells to test the hypothesis that mechanistic target of rapamycin complex 1 (mTORC1) and 2 (mTORC2) regulate folate transport by post-translational mechanisms. Silencing raptor (inhibits mTORC1) or rictor (inhibits mTORC2) markedly decreased basal folate uptake. Folate uptake stimulated by insulin + IGF-1 was mediated by mTORC2 but did not involve mTORC1. mTORC1 or mTORC2 silencing markedly decreased the plasma membrane expression of FR-α and RFC transporter isoforms without affecting global protein expression. Inhibition of the ubiquitin ligase Nedd4-2 had no effect on folate transport. In conclusion, we report for the first time that mTORC1/C2 are positive regulators of cellular folate uptake by modulating the cell surface abundance of specific transporter isoforms. We propose that regulation of placental folate transport by mTOR signaling provide a direct link between placental function, gene methylation and fetal programming.

  19. Alkyl hydroperoxide reductase has a role in oxidative stress resistance and in modulating changes in cell-surface properties in Azospirillum brasilense Sp245. (United States)

    Wasim, Mariam; Bible, Amber N; Xie, Zhihong; Alexandre, Gladys


    An ahpC mutant derivative of Azospirillum brasilense Sp245 (strain SK586) that encodes an alkyl hydroperoxide reductase was found to be more sensitive to oxidative stress caused by organic hydroperoxides compared with the wild-type. In addition, the ahpC mutant strain had multiple defects in a large array of cellular functions that were consistent with alteration of cell-surface properties, such as cell morphology in stationary phase, Calcofluor White-, Congo Red- and lectin-binding abilities, as well as cell-to-cell aggregation and flocculation. All phenotypes of the ahpC mutant were complemented by in trans expression of AhpC, and overexpression of AhpC in the wild-type strain was found to affect the same set of phenotypes, suggesting that the pleiotropic effects were caused by the ahpC mutation. SK586 was also found to be fully motile, but it lost motility at a higher rate than the wild-type during growth, such that most SK586 cells were non-motile in stationary phase. Despite these defects, the mutant did not differ from the wild-type in short-term colonization of sterile wheat roots when inoculated alone, and in competition with the wild-type strain; this implied that AhpC activity may not endow the cells with a competitive advantage in colonization under these conditions. Although the exact function of AhpC in affecting these phenotypes remains to be determined, changes in cell morphology, surface properties, cell-to-cell aggregation and flocculation are common adaptive responses to various stresses in bacteria, and the data obtained here suggest that AhpC contributes to modulating such stress responses in A. brasilense.

  20. MicroRNA miR-182 cluster mediated modulation of RECK without changes in cell surface membrane type-1 matrix metalloproteinase (MT1-MMP). (United States)

    Silva, Milagros; Hernandez, Maria E; Rojas, Fausto; Li, Lihua; Subramanian, Subbaya; Wilson, Michael J


    Cell surface localized membrane type 1-matrix metalloproteinase (MT1-MMP) plays an important role in physiological and pathological processes and its function can be regulated by proteins such as RECK. We examined the ability of miR-182 (one of the miR-183 cluster miRNAs), which can target RECK, to control cell surface MT1-MMP activity. Expression of RECK mRNA and protein was increased with anti-miRs to miR-182, miR-183 or miR-96 in HT1080 fibrosarcoma cells, but, decreased RECK mRNA and increased its protein in the benign prostatic hyperplasia cell line BPH-1. Treatment of BPH-1 and HT-1080 cells with the anti-miRs did not change the level of cell surface MT1-MMP activity, nor their rate of migration in an in vitro wound-healing assay. Trichostatin A (TSA) did not increase the level of RECK, but blocked cell surface MT1-MMP activity and decreased cell motility. Anti-miRs mediated increased RECK levels did not interfere with cell surface MT1-MMP function, and TSA may block cell surface localization of MT1-MMP by a mechanism independent of RECK.

  1. A space tethered towing method using tension and platform thrusts (United States)

    Meng, Zhongjie; Wang, Bingheng; Huang, Panfeng


    Orbit maneuver via tether is a promising countermeasure for space debris removal and satellite orbit transfer. A space tethered towing method is explored that utilizes thrust to fulfill transfer and bounded tension to stabilize tether heading. For this purpose, a time-energy optimal orbit is designed by Gauss pseudospectral method. The theoretical attitude commands are obtained by equilibria analysis. An effective attitude control strategy is presented where the commands are optimized first and then feedback controller is designed. To deal with the underactuated problem with tension constraint, hierarchical sliding mode theory is employed and an adaptive anti-windup module is added to mitigate the actuator saturation. Simulation results show that the target is towed effectively by the thrusts, and a smooth tracking for the commands of tether length and in-plane tether heading is guaranteed by the bounded tension. In addition, the designed controller also presents appreciable robustness to model error and determination error.

  2. GRASP: A multitasking tether

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    Catherine eRabouille


    Full Text Available Originally identified as Golgi stacking factors in vitro, the Golgi reassembly stacking protein (GRASP family has been shown to act as membrane tethers with multiple cellular roles. As an update to previous comprehensive reviews of the GRASP family (Vinke et al., 2011 (Giuliani et al., 2011;Jarvela and Linstedt, 2012, we outline here the latest findings concerning their diverse roles. New insights into the mechanics of GRASP-mediated tethering come from recent crystal structures. The models of how GRASP65 and GRASP55 tether membranes relate directly to their role in Golgi ribbon formation in mammalian cells and the unlinking of the ribbon at the onset of mitosis. However, it is also clear that GRASPs act outside the Golgi with roles at the ER and ER exit sites (ERES. Furthermore, the proteins of this family display other roles upon cellular stress, especially in mediating unconventional secretion of both transmembrane proteins (Golgi bypass and cytoplasmic proteins (through secretory autophagosomes.

  3. Molecular Structure of Membrane Tethers


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


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

  4. Mapping cell surface adhesion by rotation tracking and adhesion footprinting (United States)

    Li, Isaac T. S.; Ha, Taekjip; Chemla, Yann R.


    Rolling adhesion, in which cells passively roll along surfaces under shear flow, is a critical process involved in inflammatory responses and cancer metastasis. Surface adhesion properties regulated by adhesion receptors and membrane tethers are critical in understanding cell rolling behavior. Locally, adhesion molecules are distributed at the tips of membrane tethers. However, how functional adhesion properties are globally distributed on the individual cell’s surface is unknown. Here, we developed a label-free technique to determine the spatial distribution of adhesive properties on rolling cell surfaces. Using dark-field imaging and particle tracking, we extract the rotational motion of individual rolling cells. The rotational information allows us to construct an adhesion map along the contact circumference of a single cell. To complement this approach, we also developed a fluorescent adhesion footprint assay to record the molecular adhesion events from cell rolling. Applying the combination of the two methods on human promyelocytic leukemia cells, our results surprisingly reveal that adhesion is non-uniformly distributed in patches on the cell surfaces. Our label-free adhesion mapping methods are applicable to the variety of cell types that undergo rolling adhesion and provide a quantitative picture of cell surface adhesion at the functional and molecular level.

  5. Tethered float liquid level sensor

    Energy Technology Data Exchange (ETDEWEB)

    Daily, III, William Dean


    An apparatus for sensing the level of a liquid includes a float, a tether attached to the float, a pulley attached to the tether, a rotation sensor connected to the pulley that senses vertical movement of said float and senses the level of the liquid.

  6. Apoplastic venom allergen-like proteins of cyst nematodes modulate the activation of basal plant innate immunity by cell surface receptors. (United States)

    Lozano-Torres, Jose L; Wilbers, Ruud H P; Warmerdam, Sonja; Finkers-Tomczak, Anna; Diaz-Granados, Amalia; van Schaik, Casper C; Helder, Johannes; Bakker, Jaap; Goverse, Aska; Schots, Arjen; Smant, Geert


    Despite causing considerable damage to host tissue during the onset of parasitism, nematodes establish remarkably persistent infections in both animals and plants. It is thought that an elaborate repertoire of effector proteins in nematode secretions suppresses damage-triggered immune responses of the host. However, the nature and mode of action of most immunomodulatory compounds in nematode secretions are not well understood. Here, we show that venom allergen-like proteins of plant-parasitic nematodes selectively suppress host immunity mediated by surface-localized immune receptors. Venom allergen-like proteins are uniquely conserved in secretions of all animal- and plant-parasitic nematodes studied to date, but their role during the onset of parasitism has thus far remained elusive. Knocking-down the expression of the venom allergen-like protein Gr-VAP1 severely hampered the infectivity of the potato cyst nematode Globodera rostochiensis. By contrast, heterologous expression of Gr-VAP1 and two other venom allergen-like proteins from the beet cyst nematode Heterodera schachtii in plants resulted in the loss of basal immunity to multiple unrelated pathogens. The modulation of basal immunity by ectopic venom allergen-like proteins in Arabidopsis thaliana involved extracellular protease-based host defenses and non-photochemical quenching in chloroplasts. Non-photochemical quenching regulates the initiation of the defense-related programmed cell death, the onset of which was commonly suppressed by venom allergen-like proteins from G. rostochiensis, H. schachtii, and the root-knot nematode Meloidogyne incognita. Surprisingly, these venom allergen-like proteins only affected the programmed cell death mediated by surface-localized immune receptors. Furthermore, the delivery of venom allergen-like proteins into host tissue coincides with the enzymatic breakdown of plant cell walls by migratory nematodes. We, therefore, conclude that parasitic nematodes most likely utilize

  7. Space Station tethered elevator system (United States)

    Haddock, Michael H.; Anderson, Loren A.; Hosterman, K.; Decresie, E.; Miranda, P.; Hamilton, R.


    The optimized conceptual engineering design of a space station tethered elevator is presented. The tethered elevator is an unmanned, mobile structure which operates on a ten-kilometer tether spanning the distance between Space Station Freedom and a platform. Its capabilities include providing access to residual gravity levels, remote servicing, and transportation to any point along a tether. The report discusses the potential uses, parameters, and evolution of the spacecraft design. Emphasis is placed on the elevator's structural configuration and three major subsystem designs. First, the design of elevator robotics used to aid in elevator operations and tethered experimentation is presented. Second, the design of drive mechanisms used to propel the vehicle is discussed. Third, the design of an onboard self-sufficient power generation and transmission system is addressed.

  8. Enzymatic activity of soluble and membrane tethered peptide pro-hormone convertase 1. (United States)

    Bruzzaniti, Angela; Mains, Richard E


    Pro-hormone convertases PC1 and PC2 perform endoproteolytic cleavages of precursors in peptide-containing secretory granules. PC1 and PC2 are soluble, secreted with bioactive peptides. Evolutionarily related PCs have membrane tethers, not secreted. We tethered PC1 to the transmembrane-cytoplasmic domains (CD) of a granule enzyme (peptidylglycine-alpha-amidating monooxygenase; PAM) and Golgi-localized PC8. The tethered PC1 is far more stable to elevated temperature and denaturants than soluble PC1, and more active. Both tethers allow PC1 to visit the cell surface transiently, cleaving soluble molecules outside the cell. Both membrane-bound PC1 chimeras cleave membrane PAM into soluble active fragments when PAM is expressed on adjacent cells.

  9. Tether Elevator Crawler Systems (TECS) (United States)

    Swenson, Frank R.


    One of the needs of the experimenters on the space station is access to steady and controlled-variation microgravity environments. A method of providing these environments is to place the experiment on a tether attached to the space station. This provides a high degree of isolation from structural oscillations and vibrations. Crawlers can move these experiments along the tethers to preferred locations, much like an elevator. This report describes the motion control laws developed for these crawlers and the testing of laboratory models of these tether elevator crawlers.

  10. Modulation of BAG3 Expression and Proteasomal Activity by sAPPα Does Not Require Membrane-Tethered Holo-APP. (United States)

    Kundu, Arpita; Milosch, Nelli; Antonietti, Patrick; Baumkötter, Frederik; Zymny, Andreas; Müller, Ulrike C; Kins, Stefan; Hajieva, Parvana; Behl, Christian; Kögel, Donat


    Maintenance of intracellular proteostasis is essential for neuronal function, and emerging data support the view that disturbed proteostasis plays an important role in brain aging and the pathogenesis of age-related neurodegenerative disorders such as Alzheimer's disease (AD). sAPPalpha (sAPPα), the extracellularly secreted N-terminal alpha secretase cleavage product of the amyloid precursor protein (APP), has an established function in neuroprotection. Recently, we provided evidence that membrane-bound holo-APP functionally cooperates with sAPPα to mediate neuroprotection via activation of the Akt survival signaling pathway and sAPPα directly affects proteostasis. Here, we demonstrate that in addition to its anti-apoptotic function, sAPPα has effects on neuronal proteostasis under conditions of proteasomal stress. In particular, recombinant sAPPα significantly suppressed MG132-triggered expression of the co-chaperone BAG3 and aggresome formation, and it partially rescued proteasomal activity in a dose-dependent manner in SH-SY5Y neuroblastoma cells. In analogy, sAPPα was able to inhibit MG132-induced BAG3 expression in primary hippocampal neurons. Strikingly, these sAPPα-induced changes were unaltered in APP-depleted SH-SY5Y cells and APP-deficient neurons, demonstrating that holo-APP is not required for this particular function of sAPPα. Importantly, recombinant sAPPbeta (sAPPβ) failed to modulate BAG3 expression and proteostasis in APP-proficient wild-type (wt) cells, indicating that these biological effects are highly selective for sAPPα. In conclusion, we demonstrate that modulation of proteostasis is a distinct biological function of sAPPα and does not require surface-bound holo-APP. Our data shed new light on the physiological functions of APP and the interplay between APP processing and proteostasis during brain aging.

  11. Interplay of matrix stiffness and protein tethering in stem cell differentiation (United States)

    Wen, Jessica H.; Vincent, Ludovic G.; Fuhrmann, Alexander; Choi, Yu Suk; Hribar, Kolin C.; Taylor-Weiner, Hermes; Chen, Shaochen; Engler, Adam J.


    Stem cells regulate their fate by binding to, and contracting against, the extracellular matrix. Recently, it has been proposed that in addition to matrix stiffness and ligand type, the degree of coupling of fibrous protein to the surface of the underlying substrate, that is, tethering and matrix porosity, also regulates stem cell differentiation. By modulating substrate porosity without altering stiffness in polyacrylamide gels, we show that varying substrate porosity did not significantly change protein tethering, substrate deformations, or the osteogenic and adipogenic differentiation of human adipose-derived stromal cells and marrow-derived mesenchymal stromal cells. Varying protein-substrate linker density up to 50-fold changed tethering, but did not affect osteogenesis, adipogenesis, surface-protein unfolding or underlying substrate deformations. Differentiation was also unaffected by the absence of protein tethering. Our findings imply that the stiffness of planar matrices regulates stem cell differentiation independently of protein tethering and porosity.

  12. The Tethered Moon (United States)

    Zahnle, Kevin; Lupu, Roxana Elena; Dubrovolskis, A. R.


    that the Moon's orbit evolves is limited by the modest radiative cooling rate of Earth's atmosphere, which in effect tethers the Moon to the Earth. Consequently the Moon's orbit evolves orders of magnitude more slowly than in conventional models. Slow orbital evolution promotes capture by orbital resonances that may have been important in the Earth-Moon system

  13. Enabling Tethered Exploration on Mars Project (United States)

    National Aeronautics and Space Administration — Strong science motivations exist for exploring hard to reach terrain on Mars and the leading systems proposed to do so require tethers. While tethers are used...

  14. Tethered satellite design (United States)

    Manarini, G.


    The capability of the satellite to perform a variety of space operations to be accomplished from the shuttle is reviewed considering use of the satellite with man-in-loop and closed loop modes and deployment (toward or away from Earth, up to 100 km), stationkeeping, retrieval and control of the satellite. Scientific payloads are to be used to perform experiments and scientific investigation for applications such as magnetometry, electrodynamics, atmospheric science, chemical release, communications, plasmaphysics, dynamic environment, and power and thrust generation. The TSS-S will be reused for at least 3 missions after reconfiguration and refurbishment by changing the peculiar mission items such as thermal control, fixed boom for experiments, aerodynamic tail for yaw attitude control, external skin, experiments, and any other feature. The TSS-S is to be composed of three modules in order to allow independent integration of a single module and to facilitate the refurbishment and reconfiguration between flights. The three modules are service, auxiliary propulsion, and payload modules.

  15. Membrane tethering complexes in the endosomal system

    Directory of Open Access Journals (Sweden)

    Anne eSpang


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

  16. Numerical modelling of elastic space tethers

    DEFF Research Database (Denmark)

    Kristiansen, Kristian Uldall; Palmer, P. L.; Roberts, R. M.


    In this paper the importance of the ill-posedness of the classical, non-dissipative massive tether model on an orbiting tether system is studied numerically. The computations document that via the regularisation of bending resistance a more reliable numerical integrator can be produced. Furthermore......, the numerical experiments of an orbiting tether system show that bending may introduce significant forces in some regions of phase space. Finally, numerical evidence for the existence of an almost invariant slow manifold of the singularly perturbed, regularised, non-dissipative massive tether model is provided...

  17. The constitutive equation for membrane tether extraction. (United States)

    Chen, Yong; Yao, Da-Kang; Shao, Jin-Yu


    Membrane tethers or nanotubes play a critical role in a variety of cellular and subcellular processes such as leukocyte rolling and intercellular mass transport. The current constitutive equations that describe the relationship between the pulling force and the tether velocity during tether extraction have serious limitations. In this article, we propose a new phenomenological constitutive equation that captures all known characteristics of nanotube formation, including nonlinearity, nonzero threshold force, and possible negative tether velocity. We used tether extraction from endothelial cells as a prototype to illustrate how to obtain the material constants in the constitutive equation. With the micropipette aspiration technique, we measured tether pulling forces at both positive and negative tether velocities. We also determined the threshold force of 55 pN experimentally for the first time. This new constitutive equation unites two established ones and provides us a unified platform to better understand not only the physiological role of tether extraction during leukocyte rolling and intercellular or intracellular transport, but also the physics of membrane tether growth or retraction.

  18. Mobile tethering: Overview, perspectives and challengess

    NARCIS (Netherlands)

    Constantinescu, M.; Onur, E.; Durmus, Y.; Nikou, S.; Reuver, M. de; Bouwman, H.; Djurica, M.; Glatz, P.M.


    Purpose: The purpose of this paper is to analyze mobile tethering from technological and social perspectives. Mobile tethering allows us to share cellular data connection with others over WiFi, Bluetooth or USB. Although the technology is ready and has promising outcomes, service providers and the u

  19. Biophysical assay for tethered signaling reactions reveals tether-controlled activity for the phosphatase SHP-1 (United States)

    Goyette, Jesse; Salas, Citlali Solis; Coker-Gordon, Nicola; Bridge, Marcus; Isaacson, Samuel A.; Allard, Jun; Dushek, Omer


    Tethered enzymatic reactions are ubiquitous in signaling networks but are poorly understood. A previously unreported mathematical analysis is established for tethered signaling reactions in surface plasmon resonance (SPR). Applying the method to the phosphatase SHP-1 interacting with a phosphorylated tether corresponding to an immune receptor cytoplasmic tail provides five biophysical/biochemical constants from a single SPR experiment: two binding rates, two catalytic rates, and a reach parameter. Tether binding increases the activity of SHP-1 by 900-fold through a binding-induced allosteric activation (20-fold) and a more significant increase in local substrate concentration (45-fold). The reach parameter indicates that this local substrate concentration is exquisitely sensitive to receptor clustering. We further show that truncation of the tether leads not only to a lower reach but also to lower binding and catalysis. This work establishes a new framework for studying tethered signaling processes and highlights the tether as a control parameter in clustered receptor signaling.

  20. Seismic Response of Submerged Floating Tunnel Tether

    Institute of Scientific and Technical Information of China (English)

    SU Zhi-bin; SUN Sheng-nan


    A mathematical equation for vibration of submerged floating tunnel tether under the effects of earthquake and parametric excitation is presented.Multi-step Galerkin method is used to simplify this equation and the fourth-order Runge-Kuta integration method is used for numerical analysis.Finally,vibration response of submerged floating tunnel tether subjected to earthquake and parametric excitation is analyzed in a few numerical examples.The results show that the vibration response of tether varies with the seismic wave type; the steady maximum mid-span displacement of tether subjected to seismic wave keeps constant when parametric resonance takes place; the transient maximum mid-span displacement of tether is related to the peak value of input seismic wave acceleration.

  1. Eukaryotic membrane tethers revisited using magnetic tweezers (United States)

    Hosu, Basarab G.; Sun, Mingzhai; Marga, Françoise; Grandbois, Michel; Forgacs, Gabor


    Membrane nanotubes, under physiological conditions, typically form en masse. We employed magnetic tweezers (MTW) to extract tethers from human brain tumor cells and compared their biophysical properties with tethers extracted after disruption of the cytoskeleton and from a strongly differing cell type, Chinese hamster ovary cells. In this method, the constant force produced with the MTW is transduced to cells through super-paramagnetic beads attached to the cell membrane. Multiple sudden jumps in bead velocity were manifest in the recorded bead displacement-time profiles. These discrete events were interpreted as successive ruptures of individual tethers. Observation with scanning electron microscopy supported the simultaneous existence of multiple tethers. The physical characteristics, in particular, the number and viscoelastic properties of the extracted tethers were determined from the analytic fit to bead trajectories, provided by a standard model of viscoelasticity. Comparison of tethers formed with MTW and atomic force microscopy (AFM), a technique where the cantilever-force transducer is moved at constant velocity, revealed significant differences in the two methods of tether formation. Our findings imply that extreme care must be used to interpret the outcome of tether pulling experiments performed with single molecular techniques (MTW, AFM, optical tweezers, etc). First, the different methods may be testing distinct membrane structures with distinct properties. Second, as soon as a true cell membrane (as opposed to that of a vesicle) can attach to a substrate, upon pulling on it, multiple nonspecific membrane tethers may be generated. Therefore, under physiological conditions, distinguishing between tethers formed through specific and nonspecific interactions is highly nontrivial if at all possible.

  2. A Tether-Based Variable-Gravity Research Facility Concept (United States)

    Sorensen, Kirk


    The recent announcement of a return to the Moon and a mission to Mars has made the question of human response to lower levels of gravity more important. Recent advances in tether technology spurred by NASA s research in MXER tethers has led to a re-examination of the concept of a variable-gravity research facility (xGRF) for human research in low Earth orbit. Breakthroughs in simplified inertial tracking have made it possible to consider eliminating the despun section of previous designs. This, in turn, improves the prospect of a facility based entirely around a tether, with the human module on one end and a countermass on the other. With such a configuration, propellantless spinup and spindown is also possible based on the conservation of angular momentum from a gravity-gradient configuration to a spinning configuration. This not only saves large amounts of propellant but vastly simplifies crew and consumable resupply operations, since these can now be done in a microgravity configuration. The importance of the science to be obtained and the performance improvements in this new design argue strongly for further investigation.

  3. Activation and routing of membrane-tethered prohormone convertases 1 and 2. (United States)

    Bruzzaniti, A; Marx, R; Mains, R E


    Many peptide hormones and neuropeptides are processed by members of the subtilisin-like family of prohormone convertases (PCs), which are either soluble or integral membrane proteins. PC1 and PC2 are soluble PCs that are primarily localized to large dense core vesicles in neurons and endocrine cells. We examined whether PC1 and PC2 were active when expressed as membrane-tethered proteins, and how tethering to membranes alters the biosynthesis, enzymatic activity, and intracellular routing of these PCs. PC1 and PC2 chimeras were constructed using the transmembrane domain and cytoplasmic domain of the amidating enzyme, peptidylglycine alpha-amidating monooxygenase (PAM). The membrane-tethered PCs were rerouted from large dense core vesicles to the Golgi region. In addition, the chimeras were transiently expressed at the cell surface and rapidly internalized to the Golgi region in a fashion similar to PAM. Membrane-tethered PC1 and PC2 exhibited changes in pro-domain maturation rates, N-glycosylation, and in the pH and calcium optima required for maximal enzymatic activity against a fluorogenic substrate. In addition, the PC chimeras efficiently cleaved endogenous pro-opiomelanocortin to the correct bioactive peptides. The PAM transmembrane domain/cytoplasmic domain also prevented stimulated secretion of pro-opiomelanocortin products in AtT-20 cells.

  4. Tethered Lubricants for Small Systems

    Energy Technology Data Exchange (ETDEWEB)

    Lynden A. Archer


    The objective of this research project is two-fold. First, to fundamentally understand friction and relaxation dynamics of polymer chains near surfaces; and second, to develop novel self-lubricated substrates suitable for MEMS devices. During the three-year performance period of this study the PI and his students have shown using theory and experiments that systematic introduction of disorder into tethered lubricant coatings (e.g. by using self-assembled monolayer (SAM) mixtures or SAMs with nonlinear, branched architectures) can be used to significantly reduce the friction coefficient of a surface. They have also developed a simple procedure based on dielectric spectroscopy for quantifying the effect of surface disorder on molecular relaxation in lubricant coatings. Details of research accomplishments in each area of the project are described in the body of the report.

  5. Golgi GRASPs: moonlighting membrane tethers

    Directory of Open Access Journals (Sweden)

    Jarvela T


    Full Text Available Timothy Jarvela, Adam D LinstedtDepartment of Biological Sciences, Carnegie Mellon University, Pittsburgh, PA, USAAbstract: The identification of mammalian Golgi reassembly stacking proteins (GRASPs 15 years ago was followed by experiments implicating them in diverse functions, including two differing structural roles in Golgi biogenesis and at least two distinct roles in the secretion of proteins. GRASP55 and GRASP65 are localized to cis and medial/trans Golgi cisternae, respectively. They are both required for stacking of Golgi membranes in a Golgi reassembly assay. Depletion of either GRASP from cultured cells prevents the linking of Golgi membranes into their normal ribbon-like network. While GRASPs are not required for transport of secretory cargo per se, they are required for ER-to-Golgi transport of certain specific cargo, such as those containing a C-terminal valine motif. Surprisingly, GRASPs also promote secretion of cargo by the so-called unconventional secretory pathway, which bypasses the Golgi apparatus where the GRASPs reside. Furthermore, regulation of GRASP activity is now recognized for its connections to cell cycle control, development, and disease. Underlying these diverse activities is the structurally conserved N-terminal GRASP domain whose crystal structure was recently determined. It consists of a tandem array of atypical PSD95–DlgA–Zo–1 (PDZ domains, which are well-known protein–protein interaction motifs. The GRASP PDZ domains are used to localize the proteins to the Golgi as well as GRASP-mediated membrane tethering and cargo interactions. These activities are regulated, in part, by phosphorylation of the large unstructured C-terminal domain.Keywords: GRASP, review, membrane, tether, PDZ domain, secretory chaperone, unconventional secretion

  6. The space station tethered elevator system (United States)

    Anderson, Loren A.


    The optimized conceptual engineering design of a space station tethered elevator is presented. The elevator is an unmanned mobile structure which operates on a ten kilometer tether spanning the distance between the Space Station and a tethered platform. Elevator capabilities include providing access to residual gravity levels, remote servicing, and transportation to any point along a tether. The potential uses, parameters, and evolution of the spacecraft design are discussed. Engineering development of the tethered elevator is the result of work conducted in the following areas: structural configurations; robotics, drive mechanisms; and power generation and transmission systems. The structural configuration of the elevator is presented. The structure supports, houses, and protects all systems on board the elevator. The implementation of robotics on board the elevator is discussed. Elevator robotics allow for the deployment, retrieval, and manipulation of tethered objects. Robotic manipulators also aid in hooking the elevator on a tether. Critical to the operation of the tethered elevator is the design of its drive mechanisms, which are discussed. Two drivers, located internal to the elevator, propel the vehicle along a tether. These modular components consist of endless toothed belts, shunt-wound motors, regenerative power braking, and computer controlled linear actuators. The designs of self-sufficient power generation and transmission systems are reviewed. Thorough research indicates all components of the elevator will operate under power provided by fuel cells. The fuel cell systems will power the vehicle at seven kilowatts continuously and twelve kilowatts maximally. A set of secondary fuel cells provides redundancy in the unlikely event of a primary system failure. Power storage exists in the form of Nickel-Hydrogen batteries capable of powering the elevator under maximum loads.

  7. The cell surface of Trypanosoma cruzi

    Directory of Open Access Journals (Sweden)

    Wanderley de Souza


    Full Text Available The cell surface of trypanosomatids is formed by the plasma membrane and a layer of sub-pellicular microtubules which are connected to the plasma membrane. The plasma membrane is composed by proteins, lipids and carbohydrates which form the glycocalix. In this paper we will review briefly aspects related to the organization of the cell surface of Trypanosoma cruzi.

  8. FCAPD Protective Coating for Space Tethers Project (United States)

    National Aeronautics and Space Administration — Alameda Applied Sciences Corporation (AASC) proposes to demonstrate extended service lifetime of space tethers in the Low Earth Orbit (LEO) environment by using...

  9. Cell-surface remodelling during mammalian erythropoiesis. (United States)

    Wraith, D C; Chesterton, C J


    Current evidence suggests that the major cell-surface modification occurring during mammalian erythropoiesis could be generated by two separate mechanisms: either selective loss of membrane proteins during enucleation or endocytosis at the subsequent reticulocyte and erythrocyte stages. The former idea was tested by collecting developing rabbit erythroid cells before and after the enucleation step and comparing their cell-surface protein composition via radiolabelling and electrophoresis. Few changes were observed. Our data thus lend support to the endocytosis mechanism.

  10. Biotechnology Applications of Tethered Lipid Bilayer Membranes

    Directory of Open Access Journals (Sweden)

    Joshua A. Jackman


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

  11. Parametric Vibration of Submerged Floating Tunnel Tether Under Random Excitation

    Institute of Scientific and Technical Information of China (English)

    SUN Sheng-nan; SU Zhi-bin


    For the study of the parametric vibration response of submerged floating tunnel tether under random excitation, a nonlinear random parametric vibration equation of coupled tether and tube of submerged floating tunnel is set up. Subsequently, vibration response of tether in the tether-tube system is analyzed by Monte Carlo method. It may be concluded that when the tube is subjected to zero-mean Gaussian white noise random excitation, the displacement and velocity root mean square responses of tether reach the peak if the circular frequency of tube doubles that of tether; the displacernent and velocity root mean square responses of tether increase as the random excitation root mean square increases; owing to the damping force of water, the displacement and velocity root mean square responses of tether decrease rapidly compared with tether in air; increasing the damping of the tether or tube reduces the displacement and velocity root mean square responses of tether; the large-amplitude vibration of tether may be avoided by locating dampers on the tether or tube.

  12. Tethering complexes in the Arabidopsis endomembrane system

    Directory of Open Access Journals (Sweden)

    Nemanja eVukasinovic


    Full Text Available AbstractTargeting of endomembrane transport containers is of the utmost importance for proper land plant growth and development. Given the immobility of plant cells, localized membrane vesicle secretion and recycling are amongst the main processes guiding proper cell, tissue and whole plant morphogenesis. Cell wall biogenesis and modification are dependent on vectorial membrane traffic, not only during normal development, but also in stress responses and in plant defence against pathogens and/or symbiosis. It is surprising how little we know about these processes in plants, from small GTPase regulation to the tethering complexes that act as their effectors. Tethering factors are single proteins or protein complexes mediating first contact between the target membrane and arriving membrane vesicles. In this review we focus on the tethering complexes of the best-studied plant model – Arabidopsis thaliana. Genome-based predictions indicate the presence of all major tethering complexes in plants that are known from a hypothetical last eukaryotic common ancestor (LECA. The evolutionary multiplication of paralogs of plant tethering complex subunits has produced the massively expanded EXO70 family, indicating a subfunctionalization of the terminal exocytosis machinery in land plants. Interpretation of loss of function (LOF mutant phenotypes has to consider that related, yet clearly functionally-specific complexes often share some common core subunits. It is therefore impossible to conclude with clarity which version of the complex is responsible for the phenotypic deviations observed. Experimental interest in the analysis of plant tethering complexes is growing and we hope to contribute with this review by attracting even more attention to this fascinating field of plant cell biology.

  13. Tetherin is an exosomal tether (United States)

    Edgar, James R; Manna, Paul T; Nishimura, Shinichi; Banting, George; Robinson, Margaret S


    Exosomes are extracellular vesicles that are released when endosomes fuse with the plasma membrane. They have been implicated in various functions in both health and disease, including intercellular communication, antigen presentation, prion transmission, and tumour cell metastasis. Here we show that inactivating the vacuolar ATPase in HeLa cells causes a dramatic increase in the production of exosomes, which display endocytosed tracers, cholesterol, and CD63. The exosomes remain clustered on the cell surface, similar to retroviruses, which are attached to the plasma membrane by tetherin. To determine whether tetherin also attaches exosomes, we knocked it out and found a 4-fold reduction in plasma membrane-associated exosomes, with a concomitant increase in exosomes discharged into the medium. This phenotype could be rescued by wild-type tetherin but not tetherin lacking its GPI anchor. We propose that tetherin may play a key role in exosome fate, determining whether they participate in long-range or short-range interactions. DOI: PMID:27657169

  14. Impaired tethering and fusion of GLUT4 vesicles in insulin-resistant human adipose cells. (United States)

    Lizunov, Vladimir A; Lee, Jo-Ping; Skarulis, Monica C; Zimmerberg, Joshua; Cushman, Samuel W; Stenkula, Karin G


    Systemic glucose homeostasis is profoundly influenced by adipose cell function. Here we investigated GLUT4 dynamics in living adipose cells from human subjects with varying BMI and insulin sensitivity index (Si) values. Cells were transfected with hemagglutinin (HA)-GLUT4-green fluorescent protein (GFP)/mCherry (red fluorescence), and were imaged live using total internal reflection fluorescence and confocal microscopy. HA-GLUT4-GFP redistribution to the plasma membrane (PM) was quantified by surface-exposed HA epitope. In the basal state, GLUT4 storage vesicle (GSV) trafficking to and fusion with the PM were invariant with donor subject Si, as was total cell-surface GLUT4. In cells from insulin-sensitive subjects, insulin augmented GSV tethering and fusion approximately threefold, resulting in a corresponding increase in total PM GLUT4. However, with decreasing Si, these effects diminished progressively. All insulin-induced effects on GLUT4 redistribution and trafficking correlated strongly with Si and only weakly with BMI. Thus, while basal GLUT4 dynamics and total cell-surface GLUT4 are intact in human adipose cells, independent of donor Si, cells from insulin-resistant donors show markedly impaired GSV tethering and fusion responses to insulin, even after overnight culture. This altered insulin responsiveness is consistent with the hypothesis that adipose cellular dysfunction is a primary contributor to systemic metabolic dysfunction.

  15. Functions of proteoglycans at the cell surface

    DEFF Research Database (Denmark)

    Höök, M; Woods, A; Johansson, S;


    Proteoglycans (primarily heparan sulphate proteoglycans) are found at the surface of most adherent eukaryotic cells. Earlier studies suggest that these molecules can be associated with the cell surface principally by two different mechanisms. Proteoglycans may occur as membrane......-intercalated glycoproteins, where the core protein of the proteoglycan is anchored in the lipid interior of the plasma membrane, or they may be bound via the polysaccharide components of the molecule to specific anchoring proteins present at the cell surface. A number of functions have been proposed for cell surface......-associated proteoglycans, including: regulation of cell-substrate adhesion; regulation of cell proliferation; participation in the binding and uptake of extracellular components; and participation in the regulation of extracellular matrix formation. Evidence is discussed suggesting that the cell-associated heparan...

  16. Dynamics and control of tethered spacecraft - A brief overview (United States)

    Modi, V. J.; Lakshmanan, P. K.; Misra, A. K.


    Work related to the dynamics of tether connected orbiting systems and their control during deployment, operational, and retrieval phases is briefly reviewed. In particular, attention is given to the modeling of tether dynamics and control, end bodies tethered at point masses, deployment dynamics and tension control, and thruster and offset control. Directions of future efforts aimed at gaining a better understanding of the performance of tethered systems are outlined.

  17. 77 FR 50956 - Exclusion of Tethered Launches From Licensing Requirements (United States)


    ... one tether capable of bearing the maximum force exerted on the tether system, regardless of the number...\\ Nicholas E. Martino, Design and Analysis Guidelines for Launch Vehicle Tether Systems, Aerospace Report No. ATR-2008 (5377)- 1, The Aerospace Corporation (Sept. 30, 2007). This report is available in the...

  18. Ionic-Liquid-Tethered Nanoparticles: Hybrid Electrolytes

    KAUST Repository

    Moganty, Surya S.


    A new class of solventless electrolytes was created by tethering ionic liquids to hard inorganic ZrO2 nanostructures (see picture; NIM=nanoscale ionic material). These hybrid fluids exhibit exceptional redox stability windows, excellent thermal stability, good lithium transference numbers, long-term interfacial stability in the presence of a lithium anode and, when doped with lithium salt, reasonable ionic conductivities.

  19. Yielding elastic tethers stabilize robust cell adhesion.

    Directory of Open Access Journals (Sweden)

    Matt J Whitfield


    Full Text Available Many bacteria and eukaryotic cells express adhesive proteins at the end of tethers that elongate reversibly at constant or near constant force, which we refer to as yielding elasticity. Here we address the function of yielding elastic adhesive tethers with Escherichia coli bacteria as a model for cell adhesion, using a combination of experiments and simulations. The adhesive bond kinetics and tether elasticity was modeled in the simulations with realistic biophysical models that were fit to new and previously published single molecule force spectroscopy data. The simulations were validated by comparison to experiments measuring the adhesive behavior of E. coli in flowing fluid. Analysis of the simulations demonstrated that yielding elasticity is required for the bacteria to remain bound in high and variable flow conditions, because it allows the force to be distributed evenly between multiple bonds. In contrast, strain-hardening and linear elastic tethers concentrate force on the most vulnerable bonds, which leads to failure of the entire adhesive contact. Load distribution is especially important to noncovalent receptor-ligand bonds, because they become exponentially shorter lived at higher force above a critical force, even if they form catch bonds. The advantage of yielding is likely to extend to any blood cells or pathogens adhering in flow, or to any situation where bonds are stretched unequally due to surface roughness, unequal native bond lengths, or conditions that act to unzip the bonds.

  20. Tethered "kiteplane" design for the Laddermill project

    NARCIS (Netherlands)

    Breukels, J.; Ockels, W.


    The Laddermill is an innovative concept for generating energy from wind using large kite-like wings on a tether. The wings are able to fly in both the regime of airplanes and kites. We therefore call these structures "kiteplanes". By providing a recurring motion with a large lift during ascending an

  1. Structure and functions of fungal cell surfaces (United States)

    Nozawa, Y.


    A review with 24 references on the biochemistry, molecular structure, and function of cell surfaces of fungi, especially dermatophytes: the chemistry and structure of the cell wall, the effect of polyene antibiotics on the morphology and function of cytoplasmic membranes, and the chemical structure and function of pigments produced by various fungi are discussed.

  2. Coordinated coupling control of tethered space robot using releasing characteristics of space tether (United States)

    Huang, Panfeng; Zhang, Fan; Xu, Xiudong; Meng, Zhongjie; Liu, Zhengxiong; Hu, Yongxin


    Tethered space robot (TSR) is a new concept of space robot, which is released from the platform satellite, and retrieved via connected tether after space debris capture. In this paper, we propose a new coordinate control scheme for optimal trajectory and attitude tracking, and use releasing motor torque to instead the tension force, since it is difficult to track in practical. Firstly, the 6-DOF dynamics model of TSR is derived, in which the dynamics of tether releasing system is taken into account. Then, we propose and design the coordinated coupled controller, which is composed of a 6-DOF sliding mode controller and a PD controller tether's releasing. Thrust is treated as control input of the 6-DOF sliding mode controller to control the in-plane and out-of-plane angle of the tether and attitude angles of the TSR. The torque of releasing motor is used as input of PD controller, which controls the length rate of space tether. After the verification of the control scheme, finally, the simulation experiment is presented in order to validate the effectiveness of this control method. The results show that TSR can track the optimal approaching trajectory accurately. Simultaneously, the attitude angles can be changed to the desired attitude angles in control period, and the terminal accuracy is ±0.3°.

  3. Surfactant Behavior of Amphiphilic Polymer-Tethered Nanoparticles. (United States)

    Zhang, Yue; Zhao, Hanying


    In recent years, an emerging research area has been the surfactant behavior of polymer-tethered nanoparticles. In this feature article, we have provided a general introduction to the synthesis, self-assembly, and interfacial activity of polymer-tethered inorganic nanoparticles, polymer-tethered organic nanoparticles, and polymer-tethered natural nanoparticles. In addition, applications of the polymer-tethered nanoparticles in colloidal and materials science are briefly reviewed. All research demonstrates that amphiphilic polymer-tethered nanoparticles exhibit surfactant behavior and can be used as elemental building blocks for the fabrication of advanced structures by the self-assembly approach. The polymer-tethered nanoparticles provide new opportunities to engineer materials and biomaterials possessing specific functionality and physical properties.

  4. Bacterial cell surface structures in Yersinia enterocolitica. (United States)

    Białas, Nataniel; Kasperkiewicz, Katarzyna; Radziejewska-Lebrecht, Joanna; Skurnik, Mikael


    Yersinia enterocolitica is a widespread member of the family of Enterobacteriaceae that contains both non-virulent and virulent isolates. Pathogenic Y. enterocolitica strains, especially belonging to serotypes O:3, O:5,27, O:8 and O:9 are etiologic agents of yersiniosis in animals and humans. Y. enterocolitica cell surface structures that play a significant role in virulence have been subject to many investigations. These include outer membrane (OM) glycolipids such as lipopolysaccharide (LPS) and enterobacterial common antigen (ECA) and several cell surface adhesion proteins present only in virulent Y. enterocolitica, i.e., Inv, YadA and Ail. While the yadA gene is located on the Yersinia virulence plasmid the Ail, Inv, LPS and ECA are chromosomally encoded. These structures ensure the correct architecture of the OM, provide adhesive properties as well as resistance to antimicrobial peptides and to host innate immune response mechanisms.

  5. Research on the tether assisted observation of an asteroid (United States)

    Zhong, Rui; Wang, Yue


    The exploration of asteroids attracts much attention due to its potential in both scientific research and engineering application. However, the observation of an asteroid is a difficult task as the gravitational attraction of the asteroid is limited and complex. This paper proposes a concept of keeping probes in hovering above the asteroid by space tethers. The dynamics of a tethered probe attached to the surface of an asteroid is analyzed and the equations of motion are derived using Lagrange's equation. Then the equilibrium points of the dynamic system are calculated. The equilibrium tether libration angles are determined by the tether length and tether attaching location, while subjected to the constraint of positive tether tension. Afterwards, the stability of the equilibrium points are studied based on Lyapunov's theory. The variation of the equilibrium points with respect to the tether attaching location is numerically analyzed in the scenarios of different tether lengths. A parametric study of the stability of the equilibrium points is also provided. Finally, the dynamic behavior of a tethered probe perturbed from the equilibrium states is simulated to verify the proposed tether assisted technology for the observation of the asteroid.

  6. Probes for anionic cell surface detection (United States)

    Smith, Bradley D.


    Embodiments of the present invention are generally directed to compositions comprising a class of molecular probes for detecting the presence of anionic cell surfaces. Embodiments include compositions that are enriched for these compositions and preparations, particularly preparations suitable for use as laboratory/clinical reagents and diagnostic indicators, either alone or as part of a kit. An embodiment of the invention provides for a highly selective agent useful in the discernment and identification of dead or dying cells, such as apoptotic cells, in a relatively calcium-free environment. An embodiment of the invention provides a selective agent for the identification of bacteria in a mixed population of bacterial cells and nonbacterial cells.

  7. Nanotomography of Cell Surfaces with Evanescent Fields

    Directory of Open Access Journals (Sweden)

    Michael Wagner


    Full Text Available The technique of variable-angle total internal reflection fluorescence microscopy (TIRFM and its application to nanotomography of cell surfaces are described. Present applications include (1 3D imaging of chromosomes in their metaphase to demonstrate axial resolution in the nanometre range, (2 measurements of cell-substrate topology, which upon cholesterol depletion shows some loosening of cell-substrate contacts, and (3 measurements of cell topology upon photodynamic therapy (PDT, which demonstrate cell swelling and maintenance of focal contacts. The potential of the method for in vitro diagnostics, but also some requirements and limitations are discussed.

  8. The stochastic dynamics of tethered microcantilevers in a viscous fluid

    Energy Technology Data Exchange (ETDEWEB)

    Robbins, Brian A.; Paul, Mark R. [Department of Mechanical Engineering, Virginia Tech, Blacksburg, Virginia 24061 (United States); Radiom, Milad; Ducker, William A. [Department of Chemical Engineering, Virginia Tech, Blacksburg, Virginia 24061 (United States); Walz, John Y. [Department of Chemical Engineering, University of Kentucky, Lexington, Kentucky 40506 (United States)


    We explore and quantify the coupled dynamics of a pair of micron scale cantilevers immersed in a viscous fluid that are also directly tethered to one another at their tips by a spring force. The spring force, for example, could represent the molecular stiffness or elasticity of a biomolecule or material tethered between the cantilevers. We use deterministic numerical simulations with the fluctuation-dissipation theorem to compute the stochastic dynamics of the cantilever pair for the conditions of experiment when driven only by Brownian motion. We validate our approach by comparing directly with experimental measurements in the absence of the tether which shows excellent agreement. Using numerical simulations, we quantify the correlated dynamics of the cantilever pair over a range of tether stiffness. Our results quantify the sensitivity of the auto- and cross-correlations of equilibrium fluctuations in cantilever displacement to the stiffness of the tether. We show that the tether affects the magnitude of the correlations which can be used in a measurement to probe the properties of an attached tethering substance. For the configurations of current interest using micron scale cantilevers in water, we show that the magnitude of the fluid coupling between the cantilevers is sufficiently small such that the influence of the tether can be significant. Our results show that the cross-correlation is more sensitive to tether stiffness than the auto-correlation indicating that a two-cantilever measurement has improved sensitivity when compared with a measurement using a single cantilever.

  9. Lyapunov Orbits in the Jupiter System Using Electrodynamic Tethers (United States)

    Bokelmann, Kevin; Russell, Ryan P.; Lantoine, Gregory


    Various researchers have proposed the use of electrodynamic tethers for power generation and capture from interplanetary transfers. The effect of tether forces on periodic orbits in Jupiter-satellite systems are investigated. A perturbation force is added to the restricted three-body problem model and a series of simplifications allows development of a conservative system that retains the Jacobi integral. Expressions are developed to find modified locations of equilibrium positions. Modified families of Lyapunov orbits are generated as functions of tether size and Jacobi integral. Zero velocity curves and stability analyses are used to evaluate the dynamical properties of tether-modified orbits.

  10. Dynamic control of the space tethered system (United States)

    Malashin, A. A.; Smirnov, N. N.; Bryukvina, O. Yu.; Dyakov, P. A.


    We discuss the problem of simultaneous dynamical stabilization and suppression of transverse and longitudinal vibrations of the space tethered system deployed along a certain trajectory. The dynamics of the system is described by a system of nonlinear partial differential equations for the longitudinal and transverse waves and we consider a non-classical version of the problem with one moving boundary. We formulate a mathematical model and perform the analytic and numerical analysis of the boundary control problem based on the Lyapunov method. A scheme of the deployment mechanism is suggested. It includes a control torque and transverse displacement of the boundary and ensures stable deployment of the whole system.

  11. Stress, endogenous opioids and stereotypies in tethered pigs

    NARCIS (Netherlands)

    Loijens, L.W.S.


    Tether housing of female pigs in narrow, individual boxes represents a chronic stressor for the animals. Pigs that are housed tethered often develop behavioural disturbances, such as stereotypies, and changes in physiological regulation. The results of the studies described in the present thesis con

  12. Climber Motion Optimization for the Tethered Space Elevator

    NARCIS (Netherlands)

    Williams, P.; Ockels, W.J.

    The tethered space elevator could provide a revolutionary means for enabling cheap transportation to geostationary altitude and beyond. Assuming that such a system can be built, one of the dynamic design problems is determining a means of moving the elevator along the tether so as to minimize the re

  13. Lipid Gymnastics: Tethers and Fingers in membrane (United States)

    Tayebi, Lobat; Miller, Gregory; Parikh, Atul


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

  14. Tethered Contactless Mobile Nuclear Environment Monitoring Robot

    Energy Technology Data Exchange (ETDEWEB)

    Choi, S. Y.; Lee, E. S.; Lee, Kun J.; Kim, Su H.; Rim, C. T. [Korea Advanced Institute of Science and Technology, Daejeon (Korea, Republic of)


    In fact, the nuclear environment monitoring is significantly crucial for early detection of NPP accident, radiological emergency, the estimation of radiation exposure to nearby residents as well as the long term radioactivity. In the UAE, the nuclear environment monitoring is, however, quite challenging because sampling locations are far from NPPs and the outdoor temperature and humidity are very high for NPP workers to collect soil, air, and water samples. Therefore, nuclear environment monitoring robots (Nubos) are strongly needed for the NPPs in the UAE. The Nubos can be remotely controlled to collect samples in extreme environment instead of NPP workers. Moreover, the Nubos can be unmanned ground vehicles (UGVs), unmanned aerial vehicles (UAVs) and unmanned marine vehicles (UMVs) to collect soil, air, and water samples, respectively. In this paper, the prototype development of UGV type Nubos using power cable for a long distance power delivery, called Tethered contactless mobile Nubo is introduced and validated by experiments. In this paper, the prototype development of Tethered Contactless Mobile (TeCoM) Nubo, which can be powered continuously within several km distance and avoid tangled cable, and the indoor test are finished. As further works, outdoor demonstration and a grand scale R and D proposal of practical Nubo will be proceeded.

  15. Formation of Tethers from Spreading Cellular Aggregates. (United States)

    Beaune, Grégory; Winnik, Françoise M; Brochard-Wyart, Françoise


    Membrane tubes are commonly extruded from cells and vesicles when a point-like force is applied on the membrane. We report here the unexpected formation of membrane tubes from lymph node cancer prostate (LNCaP) cell aggregates in the absence of external applied forces. The spreading of LNCaP aggregates deposited on adhesive glass substrates coated with fibronectin is very limited because cell-cell adhesion is stronger than cell-substrate adhesion. Some cells on the aggregate periphery are very motile and try to escape from the aggregate, leading to the formation of membrane tubes. Tethered networks and exchange of cargos between cells were observed as well. Growth of the tubes is followed by either tube retraction or tube rupture. Hence, even very cohesive cells are successful in escaping aggregates, which may lead to epithelial mesenchymal transition and tumor metastasis. We interpret the dynamics of formation and retraction of tubes in the framework of membrane mechanics.

  16. The cell surface proteome of Entamoeba histolytica. (United States)

    Biller, Laura; Matthiesen, Jenny; Kühne, Vera; Lotter, Hannelore; Handal, Ghassan; Nozaki, Tomoyoshi; Saito-Nakano, Yumiko; Schümann, Michael; Roeder, Thomas; Tannich, Egbert; Krause, Eberhard; Bruchhaus, Iris


    Surface molecules are of major importance for host-parasite interactions. During Entamoeba histolytica infections, these interactions are predicted to be of prime importance for tissue invasion, induction of colitis and liver abscess formation. To date, however, little is known about the molecules involved in these processes, with only about 20 proteins or protein families found exposed on the E. histolytica surface. We have therefore analyzed the complete surface proteome of E. histolytica. Using cell surface biotinylation and mass spectrometry, 693 putative surface-associated proteins were identified. In silico analysis predicted that ∼26% of these proteins are membrane-associated, as they contain transmembrane domains and/or signal sequences, as well as sites of palmitoylation, myristoylation, or prenylation. An additional 25% of the identified proteins likely represent nonclassical secreted proteins. Surprisingly, no membrane-association sites could be predicted for the remaining 49% of the identified proteins. To verify surface localization, 23 proteins were randomly selected and analyzed by immunofluorescence microscopy. Of these 23 proteins, 20 (87%) showed definite surface localization. These findings indicate that a far greater number of E. histolytica proteins than previously supposed are surface-associated, a phenomenon that may be based on the high membrane turnover of E. histolytica.

  17. Molecular Dynamics Modeling of Tethered Organics in Confined Spaces

    Energy Technology Data Exchange (ETDEWEB)

    Waksburg, Avi [Monash University, Australia; Nguyen, M [Monash University, Australia; Chaffe, Alan [Monash University, Australia; Kidder, Michelle [ORNL; Buchanan III, A C [ORNL; Britt, Phillip F [ORNL


    A computational method for constructing and evaluating the dynamic behaviour of functionalised hexagonal mesoporous silica (HMS) MCM-41 models is reported. HMS with three pore diameters (1.7, 2.2 and 2.9 nm) were prepared, and, from these, two series of derivative structures were constructed - one with 1,3-diphenylpropyl (DPP) tethers and the other with smaller dimethylsilyl (DMS) tethers attached to the mesopores internal surfaces. Comparison with experimental data shows that simulation results correctly predict the maximum tether density that can be achieved for each tether and each pore diameter. For the smaller pore models, the extent of DPP functionalisation that can be achieved is limited by the available pore volume. However, for the larger pore model, the extent of functionalisation is limited by access to potentially reactive sites on the pore surface. The dynamic behaviour of the models was investigated over a range of temperatures (240-648 K). At lower temperatures (< 400 K), the mobility of DPP tethers in the 2.9 nm model is actually less than that observed in either the 2.2 nm model or the 1.7 nm model due to the extensive non-bonded interactions that are able to develop between tethers and the silica surface at this diameter. At higher temperatures, the free ends of these tethers break away from the surface, extend further into the pore space and the DPP mobility in the 2.9 nm model is higher than in the smaller pore systems.

  18. Tethered actuator for vibration control of space structures (United States)

    Fujii, H. A.; Sugimoto, Y.; Watanabe, T.; Kusagaya, T.


    Effectiveness of a micro-tension actuator for vibration control of such flexible space structures as the tethered space solar power satellites is experimentally studied on the ground. A flexible leverage is employed as the micro-tension actuator in order to control the microtension of tether. The flexible leverage is connected through a tether to the flexible beam as an experimental model of the flexible solar panel with the low first modal frequency of order 1 Hz. The nonlinearity of the flexible tether is taken into account for the vibration control since the tether becomes ineffective when it slacks, i.e., when it is tension-free. The feedback controller is designed by means of the Mission Function control algorithm. Flexural rigidity of the flexible leverage plays an important role in the vibration suppression and is studied experimentally to shed light on the effectiveness of the leverages with five different kinds of rigidity. The experimental results show not only the effect of the flexural rigidity of the flexible leverage on the control performance of the vibration suppression but also the importance of selection of the rigidity to control the vibration of tethered flexible space structures through the microtension of tethers in space.

  19. Airborne Internet Providing Tethered Balloon System

    Directory of Open Access Journals (Sweden)

    Suvriti Dhawan1


    Full Text Available In this paper we shall introduce a new system for providing wireless network communication over a specified area using ’lighter than air’ balloons. This technology will replace the existing fiber optic network system. This will be done by using a tethered balloon along with the payload (containing a receiver, a transmitter and a radio communication device.This payload will be suspended from the ground at an altitude (depending on the area of coverage required. Users under this area will be able to access this system directly for internet connectivity. This system can be used over large areas like universities, companies and societies to provide internet facility to their users through Wi-Fi or over an area where the user is specified (commercial purposes. Currently Google is working on similar idea called the ’Google Loon’ in which they use high altitude balloons which float at an altitude twice as high as air planes and the weather. They recently tested this system over New-Zealand by providing internet to their pilot testers on ground. Their balloons not being stationary, move with directional winds and have to be replaced one after the other to maintain consistency. This can be a huge problem over the areas where upper atmospheric winds are not in favorable direction. We can resolve this problem by using our stationary tethered balloon system which can communicate with the loon balloons to provide internet facility over a desired area. Moreover when our balloon will communicate with the loon balloon it will increase the coverage area as the loon balloon has to communicate to a point which is above the ground. Our system will not only replace the existing fiber optic system but it will also be selfsustaining i.e. It will generate its own power using solar panels.

  20. Interaction of the Space Shuttle on-orbit autopilot with tether dynamics (United States)

    Bergmann, Edward V.


    The effect of Orbiter flight control on tether dynamics is studied by simulation. Open-loop effects of Orbiter jet firing on tether dynamics are shown, and the potential for closed-loop interaction between tether dynamics and Orbiter flight control is determined. The significance of these effects on Orbiter flight control and tether control is assessed.

  1. Passivity-Based Control of a Rigid Electrodynamic Tether

    DEFF Research Database (Denmark)

    Larsen, Martin Birkelund; Blanke, Mogens


    how these periodic solutions can be stabilized by controlling only the current through the tether. A port-controlled Hamiltonian formulation is employed to describe the tethered satellite system and a passive input-output connection is utilized in the control design. The control law consists of two...... parts, a feedback connection, which stabilizes the open-loop equilibrium, and a bias term, which is able to drive the system trajectory away from this equilibrium, a feature necessary to obtain orbit adjustment capabilities of the electrodynamic tether. It is then shown how the periodic solutions...

  2. Tethers as Debris: Simulating Impacts of Kevlar Tethers on Shuttle Tiles (United States)

    Evans, Steven W.


    In a previous paper I examined the effects of impacts of polymer tethers on aluminum plates using the SPHC hydrodynamic code. In this paper I apply tether models to a new target - models of Space Shuttle tiles developed during the STS 107 accident investigation. In this three-dimensional simulation, a short tether fragment strikes a single tile supported on an aluminum backing plate. A tile of the LI-900 material is modeled. Penetration and damage to the tile and the backwall are characterized for three normal impact velocities. The tether is modeled as a bundle of eight 1-mm strands, with the bundle having dimensions 2-mm x 4-mm x 20-cm. The bulk material properties used are those of Kevlar(TradeMark) 49, for which a Mie-Gruneisen multiphase equation of state (eos) is used. In addition, the strength model is applied in a linear sense, such that tensile loads along the strand length are supported, but there is no strength in the lateral directions. Tile models include the various layers making up the tile structure. The outermost layer is a relatively dense borosilicate glass, known as RCG, 0.5-mm thick. The RCG layer is present on the top and four sides of the tile. Below this coating is the bulk of the tile, 1.8- in thick, made of LI-900, a product consisting of rigidized fiberous silica with a density of 9 lWft3. Below the main insulating layer is a bottom layer of the same material that has been treated to increase its density by approximately 69% to improve its strength. This densified layer is bonded to a Strain Isolation Pad (SIP), modeled as a refractory felt fabric. The SIP is bonded to an aluminum 2024 wall 0.1-in thick. The tile and backwall materials use a Me-Gruneisen multiphase eos, with the exception of the SIP felt, which uses a fabric equation of state. Fabrics must be crushed to the full bulk material density before bulk material properties and a Mie-Gruneisen eos are applied. Tether fragment impact speeds of 3,7, and 10 km/s are simulated, with

  3. Interaction between mouse adenovirus type 1 and cell surface heparan sulfate proteoglycans.

    Directory of Open Access Journals (Sweden)

    Liesbeth Lenaerts

    Full Text Available Application of human adenovirus type 5 (Ad5 derived vectors for cancer gene therapy has been limited by the poor cell surface expression, on some tumor cell types, of the primary Ad5 receptor, the coxsackie-adenovirus-receptor (CAR, as well as the accumulation of Ad5 in the liver following interaction with blood coagulation factor X (FX and subsequent tethering of the FX-Ad5 complex to heparan sulfate proteoglycan (HSPG on liver cells. As an alternative vector, mouse adenovirus type 1 (MAV-1 is particularly attractive, since this non-human adenovirus displays pronounced endothelial cell tropism and does not use CAR as a cellular attachment receptor. We here demonstrate that MAV-1 uses cell surface heparan sulfate proteoglycans (HSPGs as primary cellular attachment receptor. Direct binding of MAV-1 to heparan sulfate-coated plates proved to be markedly more efficient compared to that of Ad5. Experiments with modified heparins revealed that the interaction of MAV-1 to HSPGs depends on their N-sulfation and, to a lesser extent, 6-O-sulfation rate. Whereas the interaction between Ad5 and HSPGs was enhanced by FX, this was not the case for MAV-1. A slot blot assay demonstrated the ability of MAV-1 to directly interact with FX, although the amount of FX complexed to MAV-1 was much lower than observed for Ad5. Analysis of the binding of MAV-1 and Ad5 to the NCI-60 panel of different human tumor cell lines revealed the preference of MAV-1 for ovarian carcinoma cells. Together, the data presented here enlarge our insight into the HSPG receptor usage of MAV-1 and support the development of an MAV-1-derived gene vector for human cancer therapy.

  4. Cell Surface-based Sensing with Metallic Nanoparticles


    Jiang, Ziwen; Le, Ngoc D. B.; Gupta, Akash; Rotello, Vincent M.


    Metallic nanoparticles provide versatile scaffolds for biosensing applications. In this review, we focus on the use of metallic nanoparticles for cell surface sensings. Examples of the use of both specific recognition and array-based “chemical nose” approaches to cell surface sensing will be discussed.

  5. Temperature of electro dynamic tether for space debris removal and its effect on Lorentz force



    Electrodynamics tether (EDT) systems is expected as an effective system for deorbiting space debris. EDT systems is a high efficiency propulsion system using the Lorentz force generated by the interference with the earth's magnetic field and the current through the tether. The conductivity of the tether varies with temperacture of tether in orbit, and it affects the Lorentz force that EDT systems can generate. Tether temperature in orbit depends on the thermal optical property. However, the s...

  6. Fiber Optic Shape Sensing for Tethered Marsupial Rovers Project (United States)

    National Aeronautics and Space Administration — Building upon the successful proof of concept work in Phase I, Luna Innovations Incorporated is proposing to design, build, and test a sensing tether for marsupial...

  7. Development of a Tethered Formation Flight Testbed for ISS Project (United States)

    National Aeronautics and Space Administration — The development of a testbed for the development and demonstration of technologies needed by tethered formation flying satellites is proposed. Such a testbed would...

  8. Pilot Hartsfield in sleep restraint tethered to forward middeck lockers (United States)


    Pilot Hartsfield demonstrates the sleeping accomodations onboard the Earth-orbiting Columbia, Orbiter Vehicle (OV) 102. The sleep restraint is located in the middeck area of the spacecraft and is tethered to forward middeck lockers.

  9. Fiber Optic Shape Sensing for Tethered Marsupial Rovers Project (United States)

    National Aeronautics and Space Administration — Luna Innovations Incorporated is proposing to design, build, and test a shape, length, and tension sensing tether for robotic exploration and sample-gathering...

  10. Stationary Tether Device for Buoy Apparatus and System for Using (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — A rigid, neutrally buoyant hydrodynamicaly-faired tether and associated fastening hardware that loosely holds a bathymetric float at a predetermined distance from a...

  11. The role of hydrogen bonding in tethered polymer layers


    Ren, C; Nap, R. J.; Szleifer, I.


    A molecular theory to study the properties of end tethered polymer layers, in which the polymers have the ability to form hydrogen bonds with water is presented. The approach combines the ideas of the single-chain mean-field theory to treat tethered layers with the approach of Dormidontova (Macromolecules, 2002 35,987) to include hydrogen bonds. The generalization includes the consideration of position dependent polymer-water and water-water hydrogen bonds. The theory is applied to model poly...

  12. Dynamics and control of tethered spacecraft during deployment and retrieval (United States)

    Modi, V. J.; Lakshmanan, P. K.; Misra, A. K.

    The potential of tether-connected orbiting systems has led to numerous studies of their dynamics and control during deployment, operational (stationkeeeping), and retrieval phases. This paper examines some of the important aspects of the studies, including the modeling of tether dynamics and control, and system dynamics and control. Significant conclusions based on these studies are discussesd, and future research that would aid in a better understanding of the system performance is outlined.

  13. Dynamics of single-stranded DNA tethered to a solid (United States)

    Radiom, Milad; Paul, Mark R.; Ducker, William A.


    Tethering is used to deliver specific biological and industrial functions. For example, single-stranded DNA (ssDNA) is tethered to polymerases and long sequences of double-stranded DNA (dsDNA) during replication, and to solids in DNA microarrays. However, tethering ssDNA to a large object limits not only the available ssDNA conformations, but also the range of time-scales over which the mechanical responses of ssDNA are important. In this work we examine the effect of tethering by measurement of the mechanical response of ssDNA that is tethered at each end to two separate atomic force microscope cantilevers in aqueous solution. Thermal motion of the cantilevers drives the ends of the ssDNA chain at frequencies near 2 kHz. The presence of a tethered molecule makes a large difference to the asymmetric cross-correlation of two cantilevers, which enables resolution of the mechanical properties in our experiments. By analysis of the correlated motion of the cantilevers we extract the friction and stiffness of the ssDNA. We find that the measured friction is much larger than the friction that is usually associated with the unencumbered motion of ssDNA. We also find that the measured relaxation time, ∼30 μs, is much greater than prior measurements of the free-molecule relaxation time. We attribute the difference to the loss of conformational possibilities as a result of constraining the ends of the ssDNA.

  14. Rate limit of protein elastic response is tether dependent (United States)

    Berkovich, Ronen; Hermans, Rodolfo I.; Popa, Ionel; Stirnemann, Guillaume; Garcia-Manyes, Sergi; Berne, Bruce J.; Fernandez, Julio M.


    The elastic restoring force of tissues must be able to operate over the very wide range of loading rates experienced by living organisms. It is surprising that even the fastest events involving animal muscle tissues do not surpass a few hundred hertz. We propose that this limit is set in part by the elastic dynamics of tethered proteins extending and relaxing under a changing load. Here we study the elastic dynamics of tethered proteins using a fast force spectrometer with sub-millisecond time resolution, combined with Brownian and Molecular Dynamics simulations. We show that the act of tethering a polypeptide to an object, an inseparable part of protein elasticity in vivo and in experimental setups, greatly reduces the attempt frequency with which the protein samples its free energy. Indeed, our data shows that a tethered polypeptide can traverse its free-energy landscape with a surprisingly low effective diffusion coefficient Deff ∼ 1,200 nm2/s. By contrast, our Molecular Dynamics simulations show that diffusion of an isolated protein under force occurs at Deff ∼ 108 nm2/s. This discrepancy is attributed to the drag force caused by the tethering object. From the physiological time scales of tissue elasticity, we calculate that tethered elastic proteins equilibrate in vivo with Deff ∼ 104–106 nm2/s which is two to four orders magnitude smaller than the values measured for untethered proteins in bulk. PMID:22895787

  15. The Golgin Family of Coiled-Coil Tethering Proteins

    Directory of Open Access Journals (Sweden)

    Tomasz M Witkos


    Full Text Available The golgins are a family of predominantly coiled-coil proteins that are localized to the Golgi apparatus. Golgins are present in all eukaryotes, suggesting an evolutionary conserved function. Golgins are anchored to the Golgi membrane by their carboxy terminus and are predicted to adopt an extended conformation that projects into the surrounding cytoplasm. This arrangement is ideal for the capture or tethering of nearby membranes or cytoskeletal elements. Golgin-mediated tethering is thought to be important for vesicular traffic at the Golgi apparatus, the maintenance of Golgi architecture, as well as the positioning of the Golgi apparatus within cells. In addition to acting as tethers, some golgins can also sequester various factors at the Golgi membrane, allowing for the spatiotemporal regulation of downstream cellular functions. Although it is now established that golgins are membrane and cytoskeleton tethers, the mechanisms underlying tethering remain poorly defined. Moreover, the importance of golgin-mediated tethering in a physiological context remains to be fully explored. This review will describe our current understanding of golgin function, highlighting recent progress that has been made, and goes on to discuss outstanding questions and potential avenues for future research with regard to this family of conserved Golgi-associated proteins.

  16. Anchoring a Leviathan: How the Nuclear Membrane Tethers the Genome. (United States)

    Czapiewski, Rafal; Robson, Michael I; Schirmer, Eric C


    It is well established that the nuclear envelope has many distinct direct connections to chromatin that contribute to genome organization. The functional consequences of genome organization on gene regulation are less clear. Even less understood is how interactions of lamins and nuclear envelope transmembrane proteins (NETs) with chromatin can produce anchoring tethers that can withstand the physical forces of and on the genome. Chromosomes are the largest molecules in the cell, making megadalton protein structures like the nuclear pore complexes and ribosomes seem small by comparison. Thus to withstand strong forces from chromosome dynamics an anchoring tether is likely to be much more complex than a single protein-protein or protein-DNA interaction. Here we will briefly review known NE-genome interactions that likely contribute to spatial genome organization, postulate in the context of experimental data how these anchoring tethers contribute to gene regulation, and posit several hypotheses for the physical nature of these tethers that need to be investigated experimentally. Significantly, disruption of these anchoring tethers and the subsequent consequences for gene regulation could explain how mutations in nuclear envelope proteins cause diseases ranging from muscular dystrophy to lipodystrophy to premature aging progeroid syndromes. The two favored hypotheses for nuclear envelope protein involvement in disease are (1) weakening nuclear and cellular mechanical stability, and (2) disrupting genome organization and gene regulation. Considerable experimental support has been obtained for both. The integration of both mechanical and gene expression defects in the disruption of anchoring tethers could provide a unifying hypothesis consistent with both.

  17. A Method to Predict the Orbital Lifetimes of Free Tethers and Tether-Trailing Satellites using Artificial Neural Networks (United States)


    completely developed the method in a memoir on the perturbations of comets moving in elliptical orbits.76 As a result, orbital element variational...6848.48 70.09 160.98 159.94 -0.65 159.84 -0.71 " lAits with Free Tethers. i = 28.5 0. Results obtained for free tethers originating in nonstandard orbits

  18. Adult idiopathic scoliosis: the tethered spine. (United States)

    Whyte Ferguson, Lucy


    This article reports on an observational and treatment study using three case histories to describe common patterns of muscle and fascial asymmetry in adults with idiopathic scoliosis (IS) who have significant scoliotic curvatures that were not surgically corrected and who have chronic pain. Rather than being located in the paraspinal muscles, the myofascial trigger points (TrPs) apparently responsible for the pain were located at some distance from the spine, yet referred pain to locations throughout the thoracolumbar spine. Asymmetries in these muscles appear to tether the spine in such a way that they contribute to scoliotic curvatures. Evaluation also showed that each of these individuals had major ligamentous laxity and this may also have contributed to development of scoliotic curvatures. Treatment focused on release of TrPs found to refer pain into the spine, release of related fascia, and correction of related joint dysfunction. Treatment resulted in substantial relief of longstanding chronic pain. Treatment thus validated the diagnostic hypothesis that myofascial and fascial asymmetries were to some extent responsible for pain in adults with significant scoliotic curvatures. Treatment of these patterns of TrPs and muscle and fascial asymmetries and related joint dysfunction was also effective in relieving pain in each of these individuals after they were injured in auto accidents. Treatment of myofascial TrPs and asymmetrical fascial tension along with treatment of accompanying joint dysfunction is proposed as an effective approach to treating both chronic and acute pain in adults with scoliosis that has not been surgically corrected.

  19. Calreticulin: Roles in Cell-Surface Protein Expression

    Directory of Open Access Journals (Sweden)

    Yue Jiang


    Full Text Available In order to perform their designated functions, proteins require precise subcellular localizations. For cell-surface proteins, such as receptors and channels, they are able to transduce signals only when properly targeted to the cell membrane. Calreticulin is a multi-functional chaperone protein involved in protein folding, maturation, and trafficking. However, evidence has been accumulating that calreticulin can also negatively regulate the surface expression of certain receptors and channels. In these instances, depletion of calreticulin enhances cell-surface expression and function. In this review, we discuss the role of calreticulin with a focus on its negative effects on the expression of cell-surface proteins.

  20. Cell surface engineering of yeast for applications in white biotechnology. (United States)

    Kuroda, Kouichi; Ueda, Mitsuyoshi


    Cell surface engineering is a promising strategy for the molecular breeding of whole-cell biocatalysts. By using this strategy, yeasts can be constructed by the cell surface display of functional proteins; these yeasts are referred to as arming yeasts. Because reactions using arming yeasts as whole-cell biocatalysts occur on the cell surface, materials that cannot enter the cell can be used as reaction substrates. Numerous arming yeasts have therefore been constructed for a wide range of uses such as biofuel production, synthesis of valuable chemicals, adsorption or degradation of environmental pollutants, recovery of rare metal ions, and biosensors. Here, we review the science of yeast cell surface modification as well as current applications and future opportunities.

  1. Phase coupling of a circadian neuropeptide with rest/activity rhythms detected using a membrane-tethered spider toxin.

    Directory of Open Access Journals (Sweden)

    Ying Wu


    Full Text Available Drosophila clock neurons are self-sustaining cellular oscillators that rely on negative transcriptional feedback to keep circadian time. Proper regulation of organismal rhythms of physiology and behavior requires coordination of the oscillations of individual clock neurons within the circadian control network. Over the last decade, it has become clear that a key mechanism for intercellular communication in the circadian network is signaling between a subset of clock neurons that secrete the neuropeptide pigment dispersing factor (PDF and clock neurons that possess its G protein-coupled receptor (PDFR. Furthermore, the specific hypothesis has been proposed that PDF-secreting clock neurons entrain the phase of organismal rhythms, and the cellular oscillations of other clock neurons, via the temporal patterning of secreted PDF signals. In order to test this hypothesis, we have devised a novel technique for altering the phase relationship between circadian transcriptional feedback oscillation and PDF secretion by using an ion channel-directed spider toxin to modify voltage-gated Na(+ channel inactivation in vivo. This technique relies on the previously reported "tethered-toxin" technology for cell-autonomous modulation of ionic conductances via heterologous expression of subtype-specific peptide ion channel toxins as chimeric fusion proteins tethered to the plasma membrane with a glycosylphosphatidylinositol (GPI anchor. We demonstrate for the first time, to our knowledge, the utility of the tethered-toxin technology in a transgenic animal, validating four different tethered spider toxin ion channel modifiers for use in Drosophila. Focusing on one of these toxins, we show that GPI-tethered Australian funnel-web spider toxin delta-ACTX-Hv1a inhibits Drosophila para voltage-gated Na(+ channel inactivation when coexpressed in Xenopus oocytes. Transgenic expression of membrane-tethered delta-ACTX-Hv1a in vivo in the PDF-secreting subset of clock neurons

  2. A mass spectrometric-derived cell surface protein atlas.

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    Damaris Bausch-Fluck

    Full Text Available Cell surface proteins are major targets of biomedical research due to their utility as cellular markers and their extracellular accessibility for pharmacological intervention. However, information about the cell surface protein repertoire (the surfaceome of individual cells is only sparsely available. Here, we applied the Cell Surface Capture (CSC technology to 41 human and 31 mouse cell types to generate a mass-spectrometry derived Cell Surface Protein Atlas (CSPA providing cellular surfaceome snapshots at high resolution. The CSPA is presented in form of an easy-to-navigate interactive database, a downloadable data matrix and with tools for targeted surfaceome rediscovery ( The cellular surfaceome snapshots of different cell types, including cancer cells, resulted in a combined dataset of 1492 human and 1296 mouse cell surface glycoproteins, providing experimental evidence for their cell surface expression on different cell types, including 136 G-protein coupled receptors and 75 membrane receptor tyrosine-protein kinases. Integrated analysis of the CSPA reveals that the concerted biological function of individual cell types is mainly guided by quantitative rather than qualitative surfaceome differences. The CSPA will be useful for the evaluation of drug targets, for the improved classification of cell types and for a better understanding of the surfaceome and its concerted biological functions in complex signaling microenvironments.

  3. Effect of chromosome tethering on nuclear organization in yeast.

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    Barış Avşaroğlu

    Full Text Available Interphase chromosomes in Saccharomyces cerevisiae are tethered to the nuclear envelope at their telomeres and to the spindle pole body (SPB at their centromeres. Using a polymer model of yeast chromosomes that includes these interactions, we show theoretically that telomere attachment to the nuclear envelope is a major determinant of gene positioning within the nucleus only for genes within 10 kb of the telomeres. We test this prediction by measuring the distance between the SPB and the silent mating locus (HML on chromosome III in wild-type and mutant yeast strains that contain altered chromosome-tethering interactions. In wild-type yeast cells we find that disruption of the telomere tether does not dramatically change the position of HML with respect to the SPB, in agreement with theoretical predictions. Alternatively, using a mutant strain with a synthetic tether that localizes an HML-proximal site to the nuclear envelope, we find a significant change in the SPB-HML distance, again as predicted by theory. Our study quantifies the importance of tethering at telomeres on the organization of interphase chromosomes in yeast, which has been shown to play a significant role in determining chromosome function such as gene expression and recombination.

  4. Tethered Nanoparticle–Polymer Composites: Phase Stability and Curvature

    KAUST Repository

    Srivastava, Samanvaya


    Phase behavior of poly(ethylene glycol) (PEG) tethered silica nanoparticles dispersed in PEG hosts is investigated using small-angle X-ray scattering. Phase separation in dispersions of densely grafted nanoparticles is found to display strikingly different small-angle X-ray scattering signatures in comparison to phase-separated composites comprised of bare or sparsely grafted nanoparticles. A general diagram for the dispersion state and phase stability of polymer tethered nanoparticle-polymer composites incorporating results from this as well as various other contemporary studies is presented. We show that in the range of moderate to high grafting densities the dispersion state of nanoparticles in composites is largely insensitive to the grafting density of the tethered chains and chemistry of the polymer host. Instead, the ratio of the particle diameter to the size of the tethered chain and the ratio of the molecular weights of the host and tethered polymer chains (P/N) are shown to play a dominant role. Additionally, we find that well-functionalized nanoparticles form stable dispersions in their polymer host beyond the P/N limit that demarcates the wetting/dewetting transition in polymer brushes on flat substrates interacting with polymer melts. A general strategy for achieving uniform nanoparticle dispersion in polymers is proposed. © 2012 American Chemical Society.

  5. Clinical and MR findings of tethered cord syndrome

    Energy Technology Data Exchange (ETDEWEB)

    Chung, Ho In; Kim, Hyae Young; Chung, Eun Chul; Suh, Jeong Soo [Ewha Womans University College of Medicine, Seoul (Korea, Republic of); Lim, Hyo Keun [Hallym University College of Medicine, Seoul (Korea, Republic of); Lee, Seoung Ro [Hanyang University College of Medicine, Seoul (Korea, Republic of); Lee, Young Seok [Chung Ang Gil Hospital, Incheon (Korea, Republic of)


    Tethered cord syndrome(TCS)is defined as low position of the conus medullaris by the abnormally fixed spinal cord with progressive neurologic deficit. To evaluate the findings of TCS at MRI and its diagnostic value, we performed a retrospective analysis of MRI of 30 patients with emphasis on clinical manifestation, level of conus medullaris, cause of tethering, and associated findings. Clinical presentation included back mass(26 cases), neurogenic bladder(5 cases), urinary incontinences(5 cases), progressive constipation(2 cases), skin dimpling(1 case), gait disturbance(1 case) and club foot (1 case). Neurologic deficit was developed 11 cases(40%) and mean age of these patients at the time of diagnosis was 8.6 years. The most common cause of tethering was lipoma(63%). The tips of conus medullaris were below the level of the second lumbar spine ia all patients. The causes of tethering were lipomatous components(spinal lipoma and lipomyelomenigocele) in 67% myelomeningocele in 20%, presacral mass in 7%, thickened filum terminale in 3% and postoperative change in 3%. Associated anomalies included syringomyelia(20%) and hydrocephalus was associated in 3 out 5 patients who underwent brain MRI. MRI clearly delineated the location of conus, tethering of the filum terminate with their causes and associated abnormalities. MRI examination is a very useful diagnostic tool for the early evaluation of TCS and the postoperative follow up.

  6. Multiple membrane tethers probed by atomic force microscopy. (United States)

    Sun, Mingzhai; Graham, John S; Hegedüs, Balazs; Marga, Françoise; Zhang, Ying; Forgacs, Gabor; Grandbois, Michel


    Using the atomic force microscope to locally probe the cell membrane, we observed the formation of multiple tethers (thin nanotubes, each requiring a similar pulling force) as reproducible features within force profiles recorded on individual cells. Forces obtained with Chinese hamster ovary cells, a malignant human brain tumor cell line, and human endothelial cells (EA hy926) were found to be 28 +/- 10 pN, 29 +/- 9 pN, and 29 +/- 10 pN, respectively, independent of the nature of attachment to the cantilever. The rather large variation of the tether pulling forces measured at several locations on individual cells points to the existence of heterogeneity in the membrane properties of a morphologically homogeneous cell. Measurement of the summary lengths of the simultaneously extracted tethers provides a measure of the size of the available membrane reservoir through which co-existing tethers are associated. As expected, partial disruption of the actin cytoskeleton and removal of the hyaluronan backbone of the glycocalyx were observed to result in a marked decrease (30-50%) in the magnitude and a significant sharpening of the force distribution indicating reduced heterogeneity of membrane properties. Taken together, our results demonstrate the ability of the plasma membrane to locally produce multiple interdependent tethers-a process that could play an important role in the mechanical association of cells with their environment.

  7. Tether-directed synthesis of highly substituted oxasilacycles via an intramolecular allylation employing allylsilanes

    Directory of Open Access Journals (Sweden)

    Cox Liam R


    Full Text Available Abstract Background Using a silyl tether to unite an aldehyde electrophile and allylsilane nucleophile into a single molecule allows a subsequent Lewis-acid-mediated allylation to proceed in an intramolecular sense and therefore receive all the benefits associated with such processes. However, with the ability to cleave the tether post allylation, a product that is the result of a net intermolecular reaction can be obtained. In the present study, four diastereoisomeric β-silyloxy-α-methyl aldehydes, which contain an allylsilane tethered through the β-carbinol centre, have been prepared, in order to probe how the relative configuration of the two stereogenic centres affects the efficiency and selectivity of the intramolecular allylation. Results Syn-aldehydes, syn-4a and syn-4b, both react poorly, affording all four possible diastereoisomeric oxasilacycle products. In contrast, the anti aldehydes anti-4a and anti-4b react analogously to substrates that lack substitution at the α-site, affording only two of the four possible allylation products. Conclusion The outcome of the reaction with anti-aldehydes is in accord with reaction proceeding through a chair-like transition state (T.S.. In these systems, the sense of 1,3-stereoinduction can be rationalised by the aldehyde electrophile adopting a pseudoaxial orientation, which will minimise dipole-dipole interactions in the T.S. The 1,4-stereoinduction in these substrates is modest and seems to be modulated by the R substituent in the starting material. In the case of the syn-substrates, cyclisation through a chair T.S. is unlikely as this would require the methyl substituent α to the reacting carbonyl group to adopt an unfavourable pseudoaxial position. It is therefore proposed that these substrates react through poorly-defined T.S.s and consequently exhibit essentially no stereoselectivity.

  8. Deployment dynamics of tethered-net for space debris removal (United States)

    Shan, Minghe; Guo, Jian; Gill, Eberhard


    A tethered-net is a promising method for space debris capturing. However, its deployment dynamics is complex because of the flexibility, and its dependency of the deployment parameters is insufficiently understood. To investigate the deployment dynamics of tethered-net, four critical deployment parameters, namely maximum net area, deployment time, traveling distance and effective period are identified in this paper, and the influence of initial deployment conditions on these four parameters is investigated. Besides, a comprehensive study on a model for the tethered-net based on absolute nodal coordinates formulation (ANCF) is provided. Simulations show that the results based on the ANCF modeling method present a good agreement with that based on the conventional mass-spring modeling method. Moreover, ANCF model is capable of describing the flexibility between two nodes on the net. However, it is more computationally expensive.

  9. Orbit Maneuver of Spinning Tether via Tidal Force

    CERN Document Server

    Baoyin, Hexi; Li, Junfeng


    Recently, the spinning tethered system is regarded as a typical and fundamental space structure attracting great interest of the aerospace engineers, and has been discussed primarily for specific space missions in past decades, including on-orbit capture and propellantless orbit transfer etc. The present work studies the dynamical behaviours of a fast spinning tethered binary system under central gravitational field, and derives principles of the basic laws of orbital maneuver. Considering the characteristics of coupled librational and orbital motions, an averaging method is introduced to deal with the slow-fast system equation, thus a definite equivalent model is derived. The general orbit motion is completely determined analytically, including the orbit geometry, periodicity, conversations and moving region etc. Since the possibility of orbit control using tether reaction has been proved by previous studies, special attention is paid to the transportation mode of angular momentum and mechanical energy betwe...

  10. Surfactant mediated morphological tethering of Cu2O nanoparticles (United States)

    Sharma, Poonam


    This communication describes a very simple and reproducible methodology to study the self-assembly of nanoparticles functionalized with a non-ionic tethering agent attached to the surface of the nanoparticle seeds. The synthesis starts with the [Cu(OH)4]2- species acting as a template, with varying concentration of the tethering agent Triton X-100 (TX100). The morphological alteration is systematically investigated. The effect of surfactant micelles, growth reaction time, and solution temperature has a tremendous impact on the morphology of the nanocrystals that govern the controlled synthesis of different shapes of nanostructures. The initial morphology of the nanocrystals is polyhedron in the absence of a tethering additive. The addition of TX100 suppresses the polymorph phase morphology and enhances the non-uniform spherical morphology of the nanocrystals. The surface modification effect enhances the morphological alteration, which potentially makes it applicable to various industrial uses such as water cleaning, hydrogen production, and third-generation solar cells.

  11. Anchoring a Leviathan: How the Nuclear Membrane Tethers the Genome

    Directory of Open Access Journals (Sweden)

    Rafal eCzapiewski


    Full Text Available It is well established that the nuclear envelope has many distinct direct connections to chromatin that contribute to genome organization. The functional consequences of genome organization on gene regulation are less clear. Even less understood is how interactions of lamins and nuclear envelope transmembrane proteins (NETs with chromatin can produce anchoring tethers that can withstand the physical forces of and on the genome. Chromosomes are the largest molecules in the cell, making megadalton protein structures like the nuclear pore complexes and ribosomes seem small by comparison. Thus to withstand strong forces from chromosome dynamics an anchoring tether is likely to be much more complex than a single protein-protein or protein-DNA interaction. Here we will briefly review known NE-genome interactions that likely contribute to spatial genome organization, postulate in the context of experimental data how these anchoring tethers contribute to gene regulation, and posit several hypotheses for the physical nature of these tethers that need to be investigated experimentally. Significantly, disruption of these anchoring tethers and the subsequent consequences for gene regulation could explain how mutations in nuclear envelope proteins cause diseases ranging from muscular dystrophy to lipodystrophy to premature ageing progeroid syndromes. The two favored hypotheses for nuclear envelope protein involvement in disease are 1 weakening nuclear and cellular mechanical stability, and 2 disrupting genome organization and gene regulation. Considerable experimental support has been obtained for both. The integration of both mechanical and gene expression defects in the disruption of anchoring tethers could provide a unifying hypothesis consistent with both.

  12. Simulation and Analysis of Tethering Behavior of Neutrophils with Pseudopods.

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    Anne D Rocheleau

    Full Text Available P-selectin and P-selectin glycoprotein ligand-1 (PSGL-1 play important roles in mediating the inflammatory cascade. Selectin kinetics, together with neutrophil hydrodynamics, regulate the fundamental adhesion cascade of cell tethering and rolling on the endothelium. The current study uses the Multiscale Adhesive Dynamics computational model to simulate, for the first time, the tethering and rolling behavior of pseudopod-containing neutrophils as mediated by P-selectin/PSGL-1 bonds. This paper looks at the effect of including P-selectin/PSGL-1 adhesion kinetics. The parameters examined included the shear rate, adhesion on-rate, initial neutrophil position, and receptor number sensitivity. The outcomes analyzed included types of adhesive behavior observed, tether rolling distance and time, number of bonds formed during an adhesive event, contact area, and contact time. In contrast to the hydrodynamic model, P-selectin/PSGL-1 binding slows the neutrophil's translation in the direction of flow and causes the neutrophil to swing around perpendicular to flow. Several behaviors were observed during the simulations, including tethering without firm adhesion, tethering with downstream firm adhesion, and firm adhesion upon first contact with the endothelium. These behaviors were qualitatively consistent with in vivo data of murine neutrophils with pseudopods. In the simulations, increasing shear rate, receptor count, and bond formation rate increased the incidence of firm adhesion upon first contact with the endothelium. Tethering was conserved across a range of physiological shear rates and was resistant to fluctuations in the number of surface PSGL-1 molecules. In simulations where bonding occurred, interaction with the side of the pseudopod, rather than the tip, afforded more surface area and greater contact time with the endothelial wall.

  13. Microarray analysis of genes associated with cell surface NIS protein levels in breast cancer

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    Richardson Andrea L


    Full Text Available Abstract Background Na+/I- symporter (NIS-mediated iodide uptake allows radioiodine therapy for thyroid cancer. NIS is also expressed in breast tumors, raising potential for radionuclide therapy of breast cancer. However, NIS expression in most breast cancers is low and may not be sufficient for radionuclide therapy. We aimed to identify biomarkers associated with NIS expression such that mechanisms underlying NIS modulation in human breast tumors may be elucidated. Methods Published oligonucleotide microarray data within the National Center for Biotechnology Information Gene Expression Omnibus database were analyzed to identify gene expression tightly correlated with NIS mRNA level among human breast tumors. NIS immunostaining was performed in a tissue microarray composed of 28 human breast tumors which had corresponding oligonucleotide microarray data available for each tumor such that gene expression associated with cell surface NIS protein level could be identified. Results and Discussion NIS mRNA levels do not vary among breast tumors or when compared to normal breast tissues when detected by Affymetrix oligonucleotide microarray platforms. Cell surface NIS protein levels are much more variable than their corresponding NIS mRNA levels. Despite a limited number of breast tumors examined, our analysis identified cysteinyl-tRNA synthetase as a biomarker that is highly associated with cell surface NIS protein levels in the ER-positive breast cancer subtype. Conclusions Further investigation on genes associated with cell surface NIS protein levels within each breast cancer molecular subtype may lead to novel targets for selectively increasing NIS expression/function in a subset of breast cancers patients.

  14. Rate limit of protein elastic response is tether dependent



    The elastic restoring force of tissues must be able to operate over the very wide range of loading rates experienced by living organisms. It is surprising that even the fastest events involving animal muscle tissues do not surpass a few hundred hertz. We propose that this limit is set in part by the elastic dynamics of tethered proteins extending and relaxing under a changing load. Here we study the elastic dynamics of tethered proteins using a fast force spectrometer with sub-millisecond tim...

  15. Modeling of tethered satellite formations using graph theory

    DEFF Research Database (Denmark)

    Larsen, Martin Birkelund; Smith, Roy S; Blanke, Mogens


    satellite formation and proposes a method to deduce the equations of motion for the attitude dynamics of the formation in a compact form. The use of graph theory and Lagrange mechanics together allows a broad class of formations to be described using the same framework. A method is stated for finding...... could form stable formations in space are cumbersome when done at a case to case basis, and a common framework providing a basic model of the dynamics of tethered satellite formations can therefore be advantageous. This paper suggests the use of graph theoretical quantities to describe a tethered...

  16. Role of vesicle tethering factors in the ER-Golgi membrane traffic


    Sztul, Elizabeth; Lupashin, Vladimir


    Tethers are a diverse group of loosely related proteins and protein complexes grouped into 3 families based on structural and functional similarities. A well-accepted role for tethering factors is the initial attachment of transport carriers to acceptor membranes prior to fusion. However, accumulating evidence indicates that tethers are more than static bridges. Tethers have been shown to interact with components of the fusion machinery and with components involved in vesicle formation. Tethe...

  17. The golgin tether giantin regulates the secretory pathway by controlling stack organization within Golgi apparatus. (United States)

    Koreishi, Mayuko; Gniadek, Thomas J; Yu, Sidney; Masuda, Junko; Honjo, Yasuko; Satoh, Ayano


    Golgins are coiled-coil proteins that play a key role in the regulation of Golgi architecture and function. Giantin, the largest golgin in mammals, forms a complex with p115, rab1, GM130, and soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs), thereby facilitating vesicle tethering and fusion processes around the Golgi apparatus. Treatment with the microtubule destabilizing drug nocodazole transforms the Golgi ribbon into individual Golgi stacks. Here we show that siRNA-mediated depletion of giantin resulted in more dispersed Golgi stacks after nocodazole treatment than by control treatment, without changing the average cisternal length. Furthermore, depletion of giantin caused an increase in cargo transport that was associated with altered cell surface protein glycosylation. Drosophila S2 cells are known to have dispersed Golgi stacks and no giantin homolog. The exogenous expression of mammalian giantin cDNA in S2 cells resulted in clustered Golgi stacks, similar to the Golgi ribbon in mammalian cells. These results suggest that the spatial organization of the Golgi ribbon is mediated by giantin, which also plays a role in cargo transport and sugar modifications.

  18. The golgin tether giantin regulates the secretory pathway by controlling stack organization within Golgi apparatus.

    Directory of Open Access Journals (Sweden)

    Mayuko Koreishi

    Full Text Available Golgins are coiled-coil proteins that play a key role in the regulation of Golgi architecture and function. Giantin, the largest golgin in mammals, forms a complex with p115, rab1, GM130, and soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs, thereby facilitating vesicle tethering and fusion processes around the Golgi apparatus. Treatment with the microtubule destabilizing drug nocodazole transforms the Golgi ribbon into individual Golgi stacks. Here we show that siRNA-mediated depletion of giantin resulted in more dispersed Golgi stacks after nocodazole treatment than by control treatment, without changing the average cisternal length. Furthermore, depletion of giantin caused an increase in cargo transport that was associated with altered cell surface protein glycosylation. Drosophila S2 cells are known to have dispersed Golgi stacks and no giantin homolog. The exogenous expression of mammalian giantin cDNA in S2 cells resulted in clustered Golgi stacks, similar to the Golgi ribbon in mammalian cells. These results suggest that the spatial organization of the Golgi ribbon is mediated by giantin, which also plays a role in cargo transport and sugar modifications.

  19. Expanding the diversity of unnatural cell surface sialic acids

    Energy Technology Data Exchange (ETDEWEB)

    Luchansky, Sarah J.; Goon, Scarlett; Bertozzi, Carolyn R.


    Novel chemical reactivity can be introduced onto cell surfaces through metabolic oligosaccharide engineering. This technique exploits the substrate promiscuity of cellular biosynthetic enzymes to deliver unnatural monosaccharides bearing bioorthogonal functional groups into cellular glycans. For example, derivatives of N-acetylmannosamine (ManNAc) are converted by the cellular biosynthetic machinery into the corresponding sialic acids and subsequently delivered to the cell surface in the form of sialoglycoconjugates. Analogs of N-acetylglucosamine (GlcNAc) and N-acetylgalactosamine (GalNAc) are also metabolized and incorporated into cell surface glycans, likely through the sialic acid and GalNAc salvage pathways, respectively. Furthermore, GlcNAc analogs can be incorporated into nucleocytoplasmic proteins in place of {beta}-O-GlcNAc residues. These pathways have been exploited to integrate unique electrophiles such as ketones and azides into the target glycoconjugate class. These functional groups can be further elaborated in a chemoselective fashion by condensation with hydrazides and by Staudinger ligation, respectively, thereby introducing detectable probes onto the cell. In conclusion, sialic acid derivatives are efficient vehicles for delivery of bulky functional groups to cell surfaces and masking of their hydroxyl groups improves their cellular uptake and utilization. Furthermore, the successful introduction of photoactivatable aryl azides into cell surface glycans opens up new avenues for studying sialic acid-binding proteins and elucidating the role of sialic acid in essential processes such as signaling and cell adhesion.

  20. Cell surface engineering of industrial microorganisms for biorefining applications. (United States)

    Tanaka, Tsutomu; Kondo, Akihiko


    In order to decrease carbon emissions and negative environmental impacts of various pollutants, biofuel/biochemical production should be promoted for replacing fossil-based industrial processes. Utilization of abundant lignocellulosic biomass as a feedstock has recently become an attractive option. In this review, we focus on recent efforts of cell surface display using industrial microorganisms such as Escherichia coli and yeast. Cell surface display is used primarily for endowing cellulolytic activity on the host cells, and enables direct fermentation to generate useful fuels and chemicals from lignocellulosic biomass. Cell surface display systems are systematically summarized, and the drawbacks/perspectives as well as successful application of surface display for industrial biotechnology are discussed.

  1. Tethered Nanoparticle -Polymer Composites: Phase behavior and rheology (United States)

    Mangal, Rahul; Archer, Lynden A.


    Polymer nanocomposites with particle radius (a) approaching the radius of gyration (Rg) of entangled host polymer have been reported to exhibit an unusual negative reinforcement effect, which leads to an anomalous reduction in relative an anomalous reduction in relative viscosity at low particle loadings (φ) . This so-called Non-Einsteinian flow behavior is understood to be sensitive to the dispersion state of particles in host polymer. We studied suspensions of SiO2 nanoparticles tethered with polethylene glycol (PEG) in polymethylmethacralate (PMMA) with molecular weights (Mw) from 17 KDa to 280 KDa. Due to strong enthalpic interactions between PEG and PMMA (χ = -0.65), nanoparticles are expected to be well-dispersed, independent of Mw of PMMA. Using small angle x-ray scattering measurements we show that the phase stability of suspensions depends on Mw of the tethered PEG, host PMMA, and φ. Particles functionalized with low molecular weight PEG aggregate at low φ, but disperse at high φ. In contrast, nanoparticles functionalized with higher molecular weight PEG are well dispersed for host chain lengths (P) to tethered chain length (N), (P/N), is as high as 160. The stability boundary of these suspensions extends well beyond expectations for nanocomposites based on tethered PEG chains suspended in PEG. Through in-depth analysis of rheology and x-ray photon correlation spectra we explore the fundamental origins of non-Einsteinian flow behavior. King Abdullah University of Science and Technology (KAUST), Advanced Photon Source (APS).

  2. Sounding rocket experiment of bare electrodynamic tether system


    Fujii, Hironori; Watanabe, Takeo; Kojima, Hirohisa; OYAMA, Koh-ichiro; Kusagaya, Tairo; Yamagiwa, Yoshiki; Ohtsu, Hirotaka; Cho, Mengu; Sasaki, Susumu; Tanaka, Koji; Williams, John; Rubin, Binyamin; Les Jhonson, Charles; Khazanov, George; Sanmartín Losada, Juan Ramón


    An overview of asounding rocket S-520-25th, project on space tether technology experiment is presented.The project is prepared by an international research group consisting of Japanese,European,American,andAustralianresearchers.The sounding rocket will be assembled by the ISAS/JAXA and will be launched in the summer of 2009.

  3. Fortissimo: A Japanese Space Test Of Bare Wire Anode Tethers (United States)

    Johnson, Les; Fujii, H. A.; Sanmartin, J. R.


    A Japanese led international team is developing a suborbital test of orbital-motion-limited (OML) bare wire anode current collection for application to electrodynamic tether (EDT) propulsion. The tether is a tape with a width of 25 mm, thickness of 0.05 mm, and is 300 m in length. This will be the first space test of OML theory. The mission will launch in the summer of 2009 using an S520 Sounding Rocket. During ascent, and above approx. 100 km in attitude, the tape tether will be deployed at a rate of approx. 8 m/s. Once deployed, the tape tether will serve as an anode, collecting ionospheric electrons. The electrons will be expelled into space by a hollow cathode device, thereby completing the circuit and allowing current to flow. The total amount of current collected will be used to assess the validity of OML theory. This paper will describe the objectives of the proposed mission, the technologies to be employed, and the application of the results to future space missions using EDTs for propulsion or power generation.

  4. Hierarchical Structure in Semicrystalline Polymers Tethered to Nanospheres

    KAUST Repository

    Kim, Sung A


    We report on structural and dynamic transitions of polymers tethered to nanoparticles. In particular, we use X-ray diffraction, vibrational spectroscopy, and thermal measurements to investigate multiscale structure and dynamic transitions of poly(ethylene glycol) (PEG) chains densely grafted to SiO2 nanoparticles. The approach used for synthesizing these hybrid particles leads to homogeneous SiO2-PEG composites with polymer grafting densities as high as 1.5 chains/nm2, which allows the hybrid materials to exist as self-suspended suspensions with distinct hierarchical structure and thermal properties. On angstrom and nanometer length scales, the tethered PEG chains exhibit more dominant TTG conformations and helix unit cell structure, in comparison to the untethered polymer. The nanoparticle tethered PEG chains are also reported to form extended crystallites on tens of nanometers length scales and to exhibit more stable crystalline structure on small dimensions. On length scales comparable to the size of each hybrid SiO 2-PEG unit, the materials are amorphous presumably as a result of the difficulty fitting the nanoparticle anchors into the PEG crystal lattice. This structural change produces large effects on the thermal transitions of PEG molecules tethered to nanoparticles. © 2014 American Chemical Society.

  5. Tethered acoustic doppler current profiler platforms for measuring streamflow (United States)

    Rehmel, Michael S.; Stewart, James A.; Morlock, Scott E.


    The U.S. Geological Survey tested and refined tethered-platform designs for measuring streamflow. Platform specifications were developed, radio-modem telemetry of acoustic Doppler current profiler (ADCP) data and potential platform-hull sources were investigated, and hulls were tested and evaluated.

  6. Dynamic analysis and trajectory tracking of a tethered space robot (United States)

    Soltani, Mehrzad; Keshmiri, Mehdi; Misra, Arun K.


    Dynamic analysis and trajectory tracking of a Tethered Space Robot (TSR) is investigated in this paper. A hybrid controller is used to perform the control task. It consists of two components, the first one deals with librational motion of the tether, while the second one takes care of the manipulator motion. A Nonlinear Model Predictive Control (NMPC) approach is used to control the tether libration; for this purpose, the libration is described by a single degree of freedom and the tether length rate is employed as the input to suppress the librational motion. A modified Computed Torque Method (CTM) is used to control the manipulator motion. The dynamic interaction between the manipulator motion and the librational motion is considered both in the system dynamics and control of the system. Using numerical simulations, performance of the proposed control system is evaluated for end-effector positioning as well as for trajectory tracking for two cases: a Low Earth Orbit (LEO) and the Geostationary Earth Orbit (GEO).

  7. Tether de-orbiting of satellites at end of mission (United States)

    Sanmartin, Juan R.; Sánchez-Torres, Antonio


    The accumulation of space debris around the Earth has become critical for Space security. The BETs project, financed by the European Commission through its FP7-Space program, is focusing on preventing generation of new debris by de-orbiting satellites at end of mission. The de-orbiting system considered, involving an electrodynamic bare tape-tether, uses no propellant and no power supply, while generating power for on-board use during de-orbiting. As an example, preliminary results are here presented on a specific orbit/satellite case: 1300 km altitude and 65 degrees inclination, and 500 kg mass. Design tether dimensions are 8 km length, 1.5 cm width, and 0.05 mm thickness; subsystem masses are limited to twice tether mass. Simple calculations, using orbit-averaging, solar mid-cycle phase, and ionospheric and geomagnetic field models, yield 2.6 months time for de-orbiting down to 200 km, with a probability of about 1 percent of debris cutting the tape. References: Sanmartin, J.R., Lorenzini, E.C., and Martinez-Sanchez, M., Electrodynamic Tether Applications and Constraints, J. Space. Rockets 47, 442-456, 2010. Sanmartin, J.R. et al., A universal system to de-orbit satellites at end of life, Journal of Space Technology and Science, to appear.

  8. Position Control of an X4-Flyer Using a Tether

    Directory of Open Access Journals (Sweden)

    , Keigo Watanabe


    Full Text Available In Japan, aging of infrastructures, such as roads, bridges, and water and sewer services, etc. poses a problem, and it is required to extend the life-span of such infrastructures by maintenance. Among infrastructures, especially bridges are periodically inspected by short range visual observations, which check the damage and deterioration of the surface. However, since there are some cases where the short range visual observation is difficult, an alternative method is required so as to replace the short range visual observation with it. So, "X4-Flyer" is very attractive because of realizing a movement at high altitude easily. The objective of this study is to develop a tethered X4- Flyer, so that the conventional short range visual observation of bridges is replaced by it. In this paper, a method for the measurement and control of the position is described by using a tether for controlling the position of the X4-Flyer. In addition, it is checked whether the tethered X4-Flyer can control the position using the proposed method or not, letting it fly in a state in which a tether is being attached.

  9. Chemistry and material science at the cell surface

    Directory of Open Access Journals (Sweden)

    Weian Zhao


    Full Text Available Cell surfaces are fertile ground for chemists and material scientists to manipulate or augment cell functions and phenotypes. This not only helps to answer basic biology questions but also has diagnostic and therapeutic applications. In this review, we summarize the most recent advances in the engineering of the cell surface. In particular, we focus on the potential applications of surface engineered cells for 1 targeting cells to desirable sites in cell therapy, 2 programming assembly of cells for tissue engineering, 3 bioimaging and sensing, and ultimately 4 manipulating cell biology.

  10. T-Rex: A Japanese Space Tether Experiment (United States)

    Johnson, Les


    Electrodynamic tether (EDT) thrusters work by virtue of the force a magnetic field exerts on a wire carrying an electrical current. The force, which acts on any charged particle moving through a magnetic field (including the electrons moving in a current-carrying wire), were concisely expressed by Lorentz in 1895 in an equation that now bears his name. The force acts in a direction perpendicular to both the direction of current flow and the magnetic field vector. Electric motors make use of this force: a wire loop in a magnetic field is made to rotate by the torque the Lorentz Force exerts on it due to an alternating current in the loop times so as to keep the torque acting in the same sense. The motion of the loop is transmitted to a shaft, thus providing work. Although the working principle of EDT thrusters is not new, its application to space transportation may be significant. In essence, an EDT thruster is just a clever way of getting an electrical current to flow in a long orbiting wire (the tether) so that the Earth s magnetic field will accelerate the wire and, consequently the payload attached to the wire. The direction of current flow in the tether, either toward or away from the Earth along the local vertical, determines whether the magnetic force will raise or lower the orbit. The bias voltage of a vertically deployed metal tether, which results just from its orbital motion (assumed eastward) through Earth s magnetic field, is positive with respect to the ambient plasma at the top and negative at the bottom. This polarization is due to the action of the Lorentz force on the electrons in the tether. Thus, the natural current flow is the result of negative electrons being attracted to the upper end and then returned to the plasma at the lower end. The magnetic force in this case has a component opposite to the direction of motion, and thus leads to a lowering of the orbit and eventually to re-entry. In this generator mode of operation the Lorentz Force

  11. Modelling a tethered mammalian sperm cell undergoing hyperactivation

    KAUST Repository

    Curtis, M.P.


    The beat patterns of mammalian sperm flagella can be categorised into two different types. The first involves symmetric waves propagating down the flagellum with a net linear propulsion of the sperm cell. The second, hyperactive, waveform is classified by vigorous asymmetric waves of higher amplitude, lower wavenumber and frequency propagating down the flagellum resulting in highly curved trajectories. The latter beat pattern is part of the capacitation process whereby sperm prepare for the prospective penetration of the zona pellucida and fusion with the egg. Hyperactivation is often observed to initiate as sperm escape from epithelial and ciliary bindings formed within the isthmic regions of the female oviducts, leading to a conjecture in the literature that this waveform is mechanically important for sperm escape. Hence, we explore the mechanical effects of hyperactivation on a tethered sperm, focussing on a Newtonian fluid. Using a resistive force theory model we demonstrate that hyperactivation can indeed generate forces that pull the sperm away from a tethering point and consequently a hyperactivated sperm cell bound to an epithelial surface need not always be pushed by its flagellum. More generally, directions of the forces generated by tethered flagella are insensitive to reductions in beat frequency and the detailed flagellar responses depend on the nature of the binding at the tethering point. Furthermore, waveform asymmetry and amplitude increases enhance the tendency for a tethered flagellum to start tugging on its binding. The same is generally predicted to be true for reductions in the wavenumber of the flagellum beat, but not universally so, emphasising the dynamical complexity of flagellar force generation. Finally, qualitative observations drawn from experimental data of human sperm bound to excised female reproductive tract are also presented and are found to be consistent with the theoretical predictions. © 2012 Elsevier Ltd.

  12. The International Tethered Cord Partnership: Beginnings, process, and status (United States)

    Mulholland, Celene B.; Aranda, Guzmán; Arredondo, Luis Angel; Calgua, Erwin; Contreras, Fernando; Espinoza, Dulce Maria; Gonzalez, Juan Bosco; Hoil, Jose A.; Komolafe, Edward; Lazareff, Jorge A.; Liu, Yunhui; Soto-Mancilla, Juan Luis; Mannucci, Graciela; Nan, Bao; Portillo, Santiago; Zhao, Hongyu


    Background: Spina bifida presents a significant cause of childhood morbidity in lower- and middle-income nations. Unfortunately, there is a paucity of literature examining outcomes among children with spina bifida in these countries. The goal of the International Tethered Cord Parternship is twofold: (1) to establish an international surveillance database to examine the correlation between time of repair and clinical outcomes in children with spina bifida and tethered cord; and (2) to foster collaboration among international institutions around pediatric neurosurgical concerns. Methods: Twelve institutions in 7 countries committed to participating in the International Tethered Cord Partnership. A neurosurgeon at each institution will evaluate all children presenting with spina bifida and/or tethered cord using the survey instrument after appropriate consent is obtained. The instrument was developed collaboratively and based on previous measures of motor and sensory function, ambulation, and continence. All institutions who have begun collecting data received appropriate Institutional Review Board approval. All data will be entered into a Health Insurance Portability and Accountability Act (HIPAA) compliant database. In addition, a participant restricted internet forum was created to foster communication and includes non–project-specific communications, such as case and journal article discussion. Results: From October 2010 to December 2010, 82 patients were entered from the various study sites. Conclusion: To our knowledge this is the first international pediatric neurosurgical database focused on clinical outcomes and predictors of disease progression. The collaborative nature of the project will not only increase knowledge of spina bifida and tethered cord, but also foster discussion and further collaboration between neurosurgeons internationally. PMID:21541204

  13. Relaxation Dynamics of Nanoparticle-Tethered Polymer Chains

    KAUST Repository

    Kim, Sung A


    © 2015 American Chemical Society. Relaxation dynamics of nanoparticle-tethered cis-1,4-polyisoprene (PI) are investigated using dielectric spectroscopy and rheometry. A model system composed of polymer chains densely grafted to spherical SiO2 nanoparticles to form self-suspended suspensions facilitates detailed studies of slow global chain and fast segmental mode dynamics under surface and geometrical confinement-from experiments performed in bulk materials. We report that unentangled polymer molecules tethered to nanoparticles relax far more slowly than their tethered entangled counterparts. Specifically, at fixed grafting density we find, counterintuitively, that increasing the tethered polymer molecular weight up to values close to the entanglement molecular weight speeds up chain relaxation dynamics. Decreasing the polymer grafting density for a fixed molecular weight has the opposite effect: it dramatically slows down chain relaxation, increases interchain coupling, and leads to a transition in rheological response from simple fluid behavior to viscoelastic fluid behavior for tethered PI chains that are unentangled by conventional measures. Increasing the measurement temperature produces an even stronger elastic response and speeds up molecular relaxation at a rate that decreases with grafting density and molecular weight. These observations are discussed in terms of chain confinement driven by crowding between particles and by the existence of an entropic attractive force produced by the space-filling constraint on individual chains in a self-suspended material. Our results indicate that the entropic force between densely grafted polymer molecules couples motions of individual chains in an analogous manner to reversible cross-links in associating polymers.

  14. Autonomous Vision-Based Tethered-Assisted Rover Docking (United States)

    Tsai, Dorian; Nesnas, Issa A.D.; Zarzhitsky, Dimitri


    Many intriguing science discoveries on planetary surfaces, such as the seasonal flows on crater walls and skylight entrances to lava tubes, are at sites that are currently inaccessible to state-of-the-art rovers. The in situ exploration of such sites is likely to require a tethered platform both for mechanical support and for providing power and communication. Mother/daughter architectures have been investigated where a mother deploys a tethered daughter into extreme terrains. Deploying and retracting a tethered daughter requires undocking and re-docking of the daughter to the mother, with the latter being the challenging part. In this paper, we describe a vision-based tether-assisted algorithm for the autonomous re-docking of a daughter to its mother following an extreme terrain excursion. The algorithm uses fiducials mounted on the mother to improve the reliability and accuracy of estimating the pose of the mother relative to the daughter. The tether that is anchored by the mother helps the docking process and increases the system's tolerance to pose uncertainties by mechanically aligning the mating parts in the final docking phase. A preliminary version of the algorithm was developed and field-tested on the Axel rover in the JPL Mars Yard. The algorithm achieved an 80% success rate in 40 experiments in both firm and loose soils and starting from up to 6 m away at up to 40 deg radial angle and 20 deg relative heading. The algorithm does not rely on an initial estimate of the relative pose. The preliminary results are promising and help retire the risk associated with the autonomous docking process enabling consideration in future martian and lunar missions.

  15. One kilometer (1 km) electric solar wind sail tether produced automatically. (United States)

    Seppänen, Henri; Rauhala, Timo; Kiprich, Sergiy; Ukkonen, Jukka; Simonsson, Martin; Kurppa, Risto; Janhunen, Pekka; Hæggström, Edward


    We produced a 1 km continuous piece of multifilament electric solar wind sail tether of μm-diameter aluminum wires using a custom made automatic tether factory. The tether comprising 90,704 bonds between 25 and 50 μm diameter wires is reeled onto a metal reel. The total mass of 1 km tether is 10 g. We reached a production rate of 70 m/24 h and a quality level of 1‰ loose bonds and 2‰ rebonded ones. We thus demonstrated that production of long electric solar wind sail tethers is possible and practical.

  16. Determination of single-bond association kinetics by tethered particle motion: concept and simulations

    CERN Document Server

    Merkus, Koen E; Storm, Cornelis


    Tethered particle motion (TPM) --- the motion of a micro- or nanoparticle tethered to a substrate by a macromolecule --- is a system that has proven extremely useful for its ability to reveal physical features of the tether, because the thermal motion of the bound particle reports sensitively on parameters like the length, the rigidity, or the folding state of its tether. In this paper, we discuss a novel application of TPM, surveying its utility in probing the kinetics of single secondary bonds: bonds that form and break between the tethered particle and a substrate due, for instance, to receptor/ligand pairs on particle and substrate. Much like the tether itself affects the motion pattern, so do the presence and absence of such secondary connections. Keeping the tether properties constant, we demonstrate how raw positional TPM data may be parsed to generate detailed insights into the association and dissociation kinetics of single secondary bonds. We do this using coarse-grained molecular dynamics simulatio...

  17. DNA-mediated electron transfer in DNA duplexes tethered to gold electrodes via phosphorothioated dA tags. (United States)

    Campos, Rui; Kotlyar, Alexander; Ferapontova, Elena E


    The efficiency of DNA-based bioelectronic devices strongly depends on the way DNA molecules are linked to the electronic component. Commonly, DNA is tethered to metal electrodes via an alkanethiol linker representing an additional barrier for electron transport. Here we demonstrate that the replacement of the alkanethiol linker for a phosphorothioated adenosine tag increases the rate of DNA-mediated electron transfer (ET) up to 259 s(-1), representing the highest hitherto reported rate of electrochemically-modulated ET, and improves the stability of DNA-electrode surface binding. Both results offer pronounced technological and scientific benefits for DNA-based electronics.

  18. Analysis of cell surface antigens by Surface Plasmon Resonance imaging

    NARCIS (Netherlands)

    Stojanovic, I.; Schasfoort, R.B.M.; Terstappen, L.W.M.M.


    Surface Plasmon Resonance (SPR) is most commonly used to measure bio-molecular interactions. SPR is used significantly less frequent for measuring whole cell interactions. Here we introduce a method to measure whole cells label free using the specific binding of cell surface antigens expressed on th

  19. Immunogold labels: cell-surface markers in atomic force microscopy

    NARCIS (Netherlands)

    Putman, Constant A.J.; Grooth, de Bart G.; Hansma, Paul K.; Hulst, van Niek F.; Greve, Jan


    The feasibility of using immunogold labels as cell-surface markers in atomic force microscopy is shown in this paper. The atomic force microscope (AFM) was used to image the surface of immunogold-labeled human lymphocytes. The lymphocytes were isolated from whole blood and labeled by an indirect imm

  20. Tethered Ozonesonde Measurements During FRAPPE July-August 2014 (United States)

    Oltmans, S. J.; Johnson, B.; Sterling, C. W.; Cullis, P.; Hall, E. G.; Jordan, A. F.; Wendell, J.; Schnell, R. C.; McClure-Begley, A.; Thompson, A. M.


    O3 and temperature profiles were measured from tethered ozonesondes from surface to 400 m above ground level on 9 days during the summer of 2014 Colorado Front Range Air Pollution and Photochemistry Experiment (FRAPPE). The portable tethered ozonesonde system was set up at one of 3 sites located next to a Colorado Department of Public Health and Environment surface monitoring station. The day and site chosen were based on the previous day O3 and weather forecast. Measurements typically began at 8:30 AM and ended at 4:30 PM, averaging 40 profiles in one day. The ozonesonde when sampling at the surface consistently read within 0-3 ppbv of the surface monitor at each of the sites with a typical daytime range of 20-90 ppbv. The hourly values were averaged at 50 meter intervals showing O3 production rates were consistently around 8 ppbv per hour from 50 to 300 meters above ground level. On sunny, light wind days the O3 mixing ratio reached a maximum of 80-90 ppbv between 14:00 and 15:00 local time. The generally constant mixing ratio with height and highest mixing ratios above the surface indicate that photochemical O3 production was taking place throughout the profile. Continuous O3 profiles from a tall tower (5 and 300 m) and daily ozonesondes tracked O3 variability through the experiment. High O3 at each site was associated with different local wind directions. At Ft. Collins winds were generally out of the southeast, at Chatfield from the northeast, and at City Park Golf Course more variable. The tether system was developed at NOAA/ESRL to provide a cost effective method to measure O3 profiles on a continuous basis. The tether system consisted of a deep sea fishing pole, electric motor driving the reel with light-weight fishing line attached to the balloon ozonesonde, a tether control box, and laptop. The in house software package monitored data and controlled the tether speed and turn-around point based on real time GPS altitude from the transmitting radiosonde.

  1. Characterization of atrial natriuretic peptide degradation by cell-surface peptidase activity on endothelial cells (United States)

    Frost, S. J.; Whitson, P. A.


    Atrial natriuretic peptide (ANP) is a fluid-regulating peptide hormone that promotes vasorelaxation, natriuresis, and diuresis. The mechanisms for the release of ANP and for its clearance from the circulation play important roles in modulating its biological effects. Recently, we have reported that the cell surface of an endothelial cell line, CPA47, could degrade 125I-ANP in the presence of EDTA. In this study, we have characterized this degradation of 125I-ANP. The kinetics of ANP degradation by the surface of CPA47 cells were first order, with a Km of 320 +/- 60 nM and Vmax of 35 +/- 14 pmol of ANP degraded/10 min/10(5) cells at pH 7.4. ANP is degraded by the surface of CPA47 cells over a broad pH range from 7.0-8.5. Potato carboxypeptidase inhibitor and bestatin inhibited 125I-ANP degradation, suggesting that this degradative activity on the surface of CPA47 cells has exopeptidase characteristics. The selectivity of CPA47 cell-surface degradation of ANP was demonstrated when 125I-ANP degradation was inhibited in the presence of neuropeptide Y and angiotensin I and II but not bradykinin, bombesin, endothelin-1, or substance P. The C-terminal amino acids phe26 and tyr28 were deduced to be important for ANP interaction with the cell-surface peptidase(s) based on comparison of the IC50 of various ANP analogues and other natriuretic peptides for the inhibition of ANP degradation. These data suggest that a newly characterized divalent cation-independent exopeptidase(s) that selectively recognizes ANP and some other vasoactive peptides exists on the surface of endothelial cells.

  2. Tethered elevator: A unique opportunity for space processing (United States)

    Monti, R.


    The latest fluid dynamic and material science experiments in the microgravity environment have emphasized the importance of the residual gravity level and of the g-jitter on fluid physics phenomena. The tethered elevator presents the possibility of providing variable g-levels (both steady and g-jitter) around a very low steady g-level (that can be realized when the elevator is near the center of mass of the space station-tether complex). When positioning a variable periodic oscillation to the payload a clean g-jitter disturbance can be obtained that would not be otherwise obtainable by other systems. These two possibilities make the elevator a facility to help resolve a number of still open questions that are preventing wider utilization of the space environment in the microgravity area.

  3. Long distance cell communication using spherical tether balloons (United States)

    Manchanda, R. K.; Rajagopalan, Vasudevan; Vasudevan, Rajagopalan; Mehrotra, R. K.; Sreenivasan, S.; Pawaskar, M.; Subba Rao Jonnalagadda, Venkata; Buduru, Suneelkumar; Kulkarni, P. M.

    A proof-of-concept experiment was conducted for long-range cell communication for rural tele-phony and internet. We designed and fabricated a spherical tether balloon to carry the con-ventional micro base transceiver station (BTS) along with three slotted antenna to cover 2-pi radius. AC power and optical fiber were anchored along with the tether line. A special fre-quency license was obtained from Wireless Planning Commission (WPC) wing of Department of Telecommunication (DoT), India for the period of experiment so as not to affect the opera-tional networks. The experiments were carried out for different BTS heights up to 500 meter. Signal measurement both in data mode and voice quality were done in different quadrant using mobile vans. This paper describes the methodology (under patenting) and utility of technique for operational application.

  4. Crowded, Confined, and Frustrated: Dynamics of Molecules Tethered to Nanoparticles

    KAUST Repository

    Agarwal, Praveen


    Above a critical chemistry-dependent molecular weight, all polymer molecules entangle and, as a result, exhibit slow dynamics, enhanced viscosity, and elasticity. Herein we report on the dynamics of low molecular weight polymers tethered to nanoparticles and find that even conventionally unentangled chains manifest dynamical features similar to entangled, long-chain molecules. Our findings are shown to imply that crowding and confinement of polymers on particles produce topological constraints analogous to those in entangled systems. © 2012 American Physical Society.

  5. Currents between tethered electrodes in a magnetized laboratory plasma (United States)

    Stenzel, R. L.; Urrutia, J. M.


    Laboratory experiments on important plasma physics issues of electrodynamic tethers were performed. These included current propagation, formation of wave wings, limits of current collection, nonlinear effects and instabilities, charging phenomena, and characteristics of transmission lines in plasmas. The experiments were conducted in a large afterglow plasma. The current system was established with a small electron-emitting hot cathode tethered to an electron-collecting anode, both movable across the magnetic field and energized by potential difference up to V approx.=100 T(sub e). The total current density in space and time was obtained from complete measurements of the perturbed magnetic field. The fast spacecraft motion was reproduced in the laboratory by moving the tethered electrodes in small increments, applying delayed current pulses, and reconstructing the net field by a linear superposition of locally emitted wavelets. With this technique, the small-amplitude dc current pattern is shown to form whistler wings at each electrode instead of the generally accepted Alfven wings. For the beam electrode, the whistler wing separates from the field-aligned beam which carries no net current. Large amplitude return currents to a stationary anode generate current-driven microinstabilities, parallel electric fields, ion depletions, current disruptions and time-varying electrode charging. At appropriately high potentials and neutral densities, excess neutrals are ionized near the anode. The anode sheath emits high-frequency electron transit-time oscillations at the sheath-plasma resonance. The beam generates Langmuir turbulence, ion sound turbulence, electron heating, space charge fields, and Hall currents. An insulated, perfectly conducting transmission line embedded in the plasma becomes lossy due to excitation of whistler waves and magnetic field diffusion effects. The implications of the laboratory observations on electrodynamic tethers in space are discussed.

  6. Effect of Chromosome Tethering on Nuclear Organization in Yeast


    Barış Avşaroğlu; Gabriel Bronk; Susannah Gordon-Messer; Jungoh Ham; Debra A Bressan; Haber, James E; Jane Kondev


    Interphase chromosomes in Saccharomyces cerevisiae are tethered to the nuclear envelope at their telomeres and to the spindle pole body (SPB) at their centromeres. Using a polymer model of yeast chromosomes that includes these interactions, we show theoretically that telomere attachment to the nuclear envelope is a major determinant of gene positioning within the nucleus only for genes within 10 kb of the telomeres. We test this prediction by measuring the distance between the SPB and the sil...

  7. Electrodynamic tethers for exploration of Jupiter and its icy moons


    Sanmartín Losada, Juan Ramón


    Use of electrodynamic bare tethers in exploring the Jovian system by tapping its rotational energy for power and propulsion is studied. The position of perijove and apojove in elliptical orbits, relative to the synchronous orbit at 2.24 times Jupiter’s radius, is exploited to conveniently make the induced Lorentz force to be drag or thrust, while generating power, and navigating the system. Capture and evolution to a low elliptical orbit near Jupiter, and capture into low circular orbits at m...

  8. CD44 is the principal cell surface receptor for hyaluronate. (United States)

    Aruffo, A; Stamenkovic, I; Melnick, M; Underhill, C B; Seed, B


    CD44 is a broadly distributed cell surface protein thought to mediate cell attachment to extracelular matrix components or specific cell surface ligands. We have created soluble CD44-immunoglobulin fusion proteins and characterized their reactivity with tissue sections and lymph node high endothelial cells in primary culture. The CD44 target on high endothelial cells is sensitive to enzymes that degrade hyaluronate, and binding of soluble CD44 is blocked by low concentrations of hyaluronate or high concentrations of chondroitin 4- and 6-sulfates. A mouse anti-hamster hyaluonate receptor antibody reacts with COS cells expressing hamster CD44 cDNA. In sections of all tissues examined, including lymph nodes and Peyer's patches, predigestion with hyaluronidase eliminated CD44 binding.

  9. Soluble and cell surface receptors for tumor necrosis factor

    DEFF Research Database (Denmark)

    Wallach, D; Engelmann, H; Nophar, Y


    Tumor necrosis factor (TNF) initiates its multiple effects on cell function by binding at a high affinity to specific cell surface receptors. Two different molecular species of these receptors, which are expressed differentially in different cells, have been identified. The cDNAs of both receptor...... have recently been cloned. Antibodies to one of these receptor species (the p55, type I receptor) can trigger a variety of TNF like effects by cross-linking of the receptor molecules. Thus, it is not TNF itself but its receptors that provide the signal for the response to this cytokine...... in certain pathological situations. Release of the soluble receptors from the cells seems to occur by proteolytic cleavage of the cell surface forms and appears to be a way of down-regulating the cell response to TNF. Because of their ability to bind TNF, the soluble receptors exert an inhibitory effect...

  10. Investigation of force approximations in tethered cells simulations

    CERN Document Server

    Zakrisson, Johan; Axner, Ove; Andersson, Magnus


    Simulations of tethered cells in viscous sub-layers are frequently performed using the Stokes drag force, but without taking into account contributions from surface corrections, lift forces, buoyancy, the Basset force, the cells finite inertia, or added mass. In this work, we investigate to which extent such contributions influence, under a variety of hydrodynamic conditions, the force at the anchor point of a tethered cell and the survival probability of a bacterium that is attached to a host by either a slip or a catch bond via a tether with a few different biomechanical properties. We show that a consequence of not including some of these contributions is that the force to which a bond is exposed can be significantly underestimated; in general by ~32-46 %, where the influence of the surface corrections dominate (the parallel and normal correction coefficients contribute with ~5-8 or 23-26 %, respectively). The Basset force is a major contributor, up to 20 %, for larger cells and shear rates. The lift force...

  11. Recent Insights into Cell Surface Heparan Sulphate Proteoglycans and Cancer

    DEFF Research Database (Denmark)

    Couchman, John R; Multhaupt, Hinke; Sanderson, Ralph D


    A small group of cell surface receptors are proteoglycans, possessing a core protein with one or more covalently attached glycosaminoglycan chains. They are virtually ubiquitous and their chains are major sites at which protein ligands of many types interact. These proteoglycans can signal and re...... or fragmented proteoglycans into exosomes that can be paracrine effectors or biomarkers, and lateral interactions between some proteoglycans and calcium channels that impact the actin cytoskeleton....

  12. Recent Insights into Cell Surface Heparan Sulphate Proteoglycans and Cancer

    DEFF Research Database (Denmark)

    Couchman, John R; Multhaupt, Hinke; Sanderson, Ralph D


    behaviour. Here, we review some recent advances, emphasising that many tumour-related functions of proteoglycans are revealed only after their modification in processes subsequent to synthesis and export to the cell surface. These include enzymes that modify heparan sulphate structure, recycling of whole...... or fragmented proteoglycans into exosomes that can be paracrine effectors or biomarkers, and lateral interactions between some proteoglycans and calcium channels that impact the actin cytoskeleton....

  13. Live Cell Surface Labeling with Fluorescent Ag Nanocluster Conjugates†


    Yu, Junhua; Choi, Sungmoon; Richards, Chris I.; Antoku, Yasuko; Dickson, Robert M


    DNA-encapsulated silver clusters are readily conjugated to proteins and serve as alternatives to organic dyes and semiconductor quantum dots. Stable and bright on the bulk and single molecule levels, Ag nanocluster fluorescence is readily observed when staining live cell surfaces. Being significantly brighter and more photostable than organics and much smaller than quantum dots with a single point of attachment, these nanomaterials offer promising new approaches for bulk and single molecule b...

  14. Interaction of KSHV with Host Cell Surface Receptors and Cell Entry

    Directory of Open Access Journals (Sweden)

    Mohanan Valiya Veettil


    Full Text Available Virus entry is a complex process characterized by a sequence of events. Since the discovery of KSHV in 1994, tremendous progress has been made in our understanding of KSHV entry into its in vitro target cells. KSHV entry is a complex multistep process involving viral envelope glycoproteins and several cell surface molecules that is utilized by KSHV for its attachment and entry. KSHV has a broad cell tropism and the attachment and receptor engagement on target cells have an important role in determining the cell type-specific mode of entry. KSHV utilizes heparan sulfate, integrins and EphrinA2 molecules as receptors which results in the activation of host cell pre-existing signal pathways that facilitate the subsequent cascade of events resulting in the rapid entry of virus particles, trafficking towards the nucleus followed by viral and host gene expression. KSHV enters human fibroblast cells by dynamin dependant clathrin mediated endocytosis and by dynamin independent macropinocytosis in dermal endothelial cells. Once internalized into endosomes, fusion of the viral envelope with the endosomal membranes in an acidification dependent manner results in the release of capsids which subsequently reaches the nuclear pore vicinity leading to the delivery of viral DNA into the nucleus. In this review, we discuss the principal mechanisms that enable KSHV to interact with the host cell surface receptors as well as the mechanisms that are required to modulate cell signaling machinery for a successful entry.

  15. Cargo binding promotes KDEL receptor clustering at the mammalian cell surface. (United States)

    Becker, Björn; Shaebani, M Reza; Rammo, Domenik; Bubel, Tobias; Santen, Ludger; Schmitt, Manfred J


    Transmembrane receptor clustering is a ubiquitous phenomenon in pro- and eukaryotic cells to physically sense receptor/ligand interactions and subsequently translate an exogenous signal into a cellular response. Despite that receptor cluster formation has been described for a wide variety of receptors, ranging from chemotactic receptors in bacteria to growth factor and neurotransmitter receptors in mammalian cells, a mechanistic understanding of the underlying molecular processes is still puzzling. In an attempt to fill this gap we followed a combined experimental and theoretical approach by dissecting and modulating cargo binding, internalization and cellular response mediated by KDEL receptors (KDELRs) at the mammalian cell surface after interaction with a model cargo/ligand. Using a fluorescent variant of ricin toxin A chain as KDELR-ligand (eGFP-RTA(H/KDEL)), we demonstrate that cargo binding induces dose-dependent receptor cluster formation at and subsequent internalization from the membrane which is associated and counteracted by anterograde and microtubule-assisted receptor transport to preferred docking sites at the plasma membrane. By means of analytical arguments and extensive numerical simulations we show that cargo-synchronized receptor transport from and to the membrane is causative for KDELR/cargo cluster formation at the mammalian cell surface.

  16. Research Resource: Androgen Receptor Activity Is Regulated Through the Mobilization of Cell Surface Receptor Networks. (United States)

    Hsiao, Jordy J; Ng, Brandon H; Smits, Melinda M; Martinez, Harryl D; Jasavala, Rohini J; Hinkson, Izumi V; Fermin, Damian; Eng, Jimmy K; Nesvizhskii, Alexey I; Wright, Michael E


    The aberrant expression of androgen receptor (AR)-dependent transcriptional programs is a defining pathology of the development and progression of prostate cancers. Transcriptional cofactors that bind AR are critical determinants of prostate tumorigenesis. To gain a deeper understanding of the proteins linked to AR-dependent gene transcription, we performed a DNA-affinity chromatography-based proteomic screen designed to identify proteins involved in AR-mediated gene transcription in prostate tumor cells. Functional experiments validated the coregulator roles of known AR-binding proteins in AR-mediated transcription in prostate tumor cells. More importantly, novel coregulatory functions were detected in components of well-established cell surface receptor-dependent signal transduction pathways. Further experimentation demonstrated that components of the TNF, TGF-β, IL receptor, and epidermal growth factor signaling pathways modulated AR-dependent gene transcription and androgen-dependent proliferation in prostate tumor cells. Collectively, our proteomic dataset demonstrates that the cell surface receptor- and AR-dependent pathways are highly integrated, and provides a molecular framework for understanding how disparate signal-transduction pathways can influence AR-dependent transcriptional programs linked to the development and progression of human prostate cancers.

  17. Structure of a bacterial cell surface decaheme electron conduit. (United States)

    Clarke, Thomas A; Edwards, Marcus J; Gates, Andrew J; Hall, Andrea; White, Gaye F; Bradley, Justin; Reardon, Catherine L; Shi, Liang; Beliaev, Alexander S; Marshall, Matthew J; Wang, Zheming; Watmough, Nicholas J; Fredrickson, James K; Zachara, John M; Butt, Julea N; Richardson, David J


    Some bacterial species are able to utilize extracellular mineral forms of iron and manganese as respiratory electron acceptors. In Shewanella oneidensis this involves decaheme cytochromes that are located on the bacterial cell surface at the termini of trans-outer-membrane electron transfer conduits. The cell surface cytochromes can potentially play multiple roles in mediating electron transfer directly to insoluble electron sinks, catalyzing electron exchange with flavin electron shuttles or participating in extracellular intercytochrome electron exchange along "nanowire" appendages. We present a 3.2-Å crystal structure of one of these decaheme cytochromes, MtrF, that allows the spatial organization of the 10 hemes to be visualized for the first time. The hemes are organized across four domains in a unique crossed conformation, in which a staggered 65-Å octaheme chain transects the length of the protein and is bisected by a planar 45-Å tetraheme chain that connects two extended Greek key split β-barrel domains. The structure provides molecular insight into how reduction of insoluble substrate (e.g., minerals), soluble substrates (e.g., flavins), and cytochrome redox partners might be possible in tandem at different termini of a trifurcated electron transport chain on the cell surface.

  18. Engineering novel cell surface chemistry for selective tumor cell targeting

    Energy Technology Data Exchange (ETDEWEB)

    Bertozzi, C.R. [Univ. of California, Berkeley, CA (United States)]|[Lawrence Berkeley National Lab., CA (United States)


    A common feature of many different cancers is the high expression level of the two monosaccharides sialic acid and fucose within the context of cell-surface associated glycoconjugates. A correlation has been made between hypersialylation and/or hyperfucosylation and the highly metastatic phenotype. Thus, a targeting strategy based on sialic acid or fucose expression would be a powerful tool for the development of new cancer cell-selective therapies and diagnostic agents. We have discovered that ketone groups can be incorporated metabolically into cell-surface associated sialic acids. The ketone is can be covalently ligated with hydrazide functionalized proteins or small molecules under physiological conditions. Thus, we have discovered a mechanism to selectively target hydrazide conjugates to highly sialylated cells such as cancer cells. Applications of this technology to the generation of novel cancer cell-selective toxins and MRI contrast reagents will be discussed, in addition to progress towards the use of cell surface fucose residues as vehicles for ketone expression.

  19. In vitro DNA tethering of HIV-1 integrase by the transcriptional coactivator LEDGF/p75. (United States)

    McNeely, Melissa; Hendrix, Jelle; Busschots, Katrien; Boons, Eline; Deleersnijder, Angélique; Gerard, Melanie; Christ, Frauke; Debyser, Zeger


    Although LEDGF/p75 is believed to act as a cellular cofactor of lentiviral integration by tethering integrase (IN) to chromatin, there is no good in vitro model to analyze this functionality. We designed an AlphaScreen assay to study how LEDGF/p75 modulates the interaction of human immunodeficiency virus type 1 IN with DNA. IN bound with similar affinity to DNA mimicking the long terminal repeat or to random DNA. While LEDGF/p75 bound DNA strongly, a mutant of LEDGF/p75 with compromised nuclear localization signal (NLS)/AT hook interacted weakly, and the LEDGF/p75 PWWP domain did not interact, corroborating previous reports on the role of NLS and AT hooks in charge-dependent DNA binding. LEDGF/p75 stimulated IN binding to DNA 10-fold to 30-fold. Stimulation of IN-DNA binding required a direct interaction between IN and the C-terminus of LEDGF/p75. Addition of either the C-terminus of LEDGF/p75 (amino acids 325-530) or LEDGF/p75 mutated in the NLS/AT hooks interfered with IN binding to DNA. Our results are consistent with an in vitro model of LEDGF/p75-mediated tethering of IN to DNA. The inhibition of IN-DNA interaction by the LEDGF/p75 C-terminus may provide a novel strategy for the inhibition of HIV IN activity and may explain the potent inhibition of HIV replication observed after the overexpression of C-terminal fragments in cell culture.

  20. Heparan sulfate proteoglycans on the cell surface: versatile coordinators of cellular functions

    DEFF Research Database (Denmark)

    Tumova, S; Woods, A; Couchman, J R


    Heparan sulfate proteoglycans are complex molecules composed of a core protein with covalently attached glycosaminoglycan chains. While the protein part determines localization of the proteoglycan on the cell surfaces or in the extracellular matrix, the glycosaminoglycan component, heparan sulfate......, mediates interactions with a variety of extracellular ligands such as growth factors and adhesion molecules. Through these interactions, heparan sulfate proteoglycans participate in many events during cell adhesion, migration, proliferation and differentiation. We are determining the multitude...... of proteoglycan functions, as their intricate roles in many pathways are revealed. They act as coreceptors for growth factors, participate in signalling during cell adhesion, modulate the activity of a broad range of molecules, and partake in many developmental and pathological processes, including tumorigenesis...

  1. Tethered elevator and platforms as space station facilities: Systems studies and demonstrative experiments (United States)


    Several key concepts of the science and applications tethered platforms were studied. Some conclusions reached are herein listed. Tether elevator and platform could improve the space station scientific and applicative capabilities. The space elevator presents unique characteristics as microgravity facility and as a tethered platform servicing vehicle. Pointing platforms could represent a new kind of observation facility for large class of payloads. The dynamical, control and technological complexity of these concepts advised demonstrative experiments. The on-going tethered satellite system offers the opportunity to perform such experiments. And feasibility studies are in progress.

  2. Configuration maintaining control of three-body ring tethered system based on thrust compensation (United States)

    Huang, Panfeng; Liu, Binbin; Zhang, Fan


    Space multi-tethered systems have shown broad prospects in remote observation missions. This paper mainly focuses on the dynamics and configuration maintaining control of space spinning three-body ring tethered system for such mission. Firstly, we establish the spinning dynamic model of the three-body ring tethered system considering the elasticity of the tether using Newton-Euler method, and then validate the suitability of this model by numerical simulation. Subsequently, LP (Likins-Pringle) initial equilibrium conditions for the tethered system are derived based on rigid body's equilibrium theory. Simulation results show that tether slack, snapping and interaction between the tethers exist in the three-body ring system, and its' configuration can not be maintained without control. Finally, a control strategy based on thrust compensation, namely thrust to simulate tether compression under LP initial equilibrium conditions is designed to solve the configuration maintaining control problem. Control effects are verified by numerical simulation compared with uncontrolled situation. Simulation results show that the configuration of the three-body ring tethered system could maintain under this active control strategy.

  3. Microfluidics-based side view flow chamber reveals tether-to-sling transition in rolling neutrophils. (United States)

    Marki, Alex; Gutierrez, Edgar; Mikulski, Zbigniew; Groisman, Alex; Ley, Klaus


    Neutrophils rolling at high shear stress (above 6 dyn/cm(2)) form tethers in the rear and slings in the front. Here, we developed a novel photo-lithographically fabricated, silicone(PDMS)-based side-view flow chamber to dynamically visualize tether and sling formation. Fluorescently membrane-labeled mouse neutrophils rolled on P-selectin substrate at 10 dyn/cm(2). Most rolling cells formed 5 tethers that were 2-30 μm long. Breaking of a single tether caused a reproducible forward microjump of the cell, showing that the tether was load-bearing. About 15% of all tether-breaking events resulted in slings. The tether-to-sling transition was fast (<100 ms) with no visible material extending above the rolling cell, suggesting a very low bending modulus of the tether. The sling downstream of the rolling cell aligned according to the streamlines before landing on the flow chamber. These new observations explain how slings form from tethers and provide insight into their biomechanical properties.

  4. Novel roaming and stationary tethered aerial robots for continuous mobile missions in nuclear power plants

    Energy Technology Data Exchange (ETDEWEB)

    Gu, Beom W.; Choi, Su Y.; Rim, Chun T. [Dept. of Nuclear and Quantum Engineering, KAIST, Daejeon (Korea, Republic of); Choi, Young Soo [Korea Atomic Energy Research Institute, Daejeon (Korea, Republic of); Cai, Guowei; Seneviratne, Lakmal [Dept. of Aerospace Engineering, Khalifa University, Abu Dhabi (United Arab Emirates)


    In this paper, new tethered aerial robots including roaming tethered aerial robots (RTARs) for radioactive material sampling and stationary tethered aerial robots (STARs) for environment monitoring are proposed to meet extremely-long-endurance missions of nuclear power plants. The flight of the proposed tethered aerial robots may last for a few days or even a few months as long as the tethered cable provides continuous power. A high voltage AC or DC power system was newly adopted to reduce the mass of the tethered cable. The RTAR uses a tethered cable spooled from the aerial robot and an aerial tension control system. The aerial tension control system provides the appropriate tension to the tethered cable, which is accordingly laid down on the ground as the RTAR roams. The STAR includes a tethered cable spooled from the ground and a ground tension control system, which enables the STAR to reach high altitudes. Prototypes of the RTAR and STAR were designed and successfully demonstrated in outdoor environments, where the load power, power type, operating frequency, and flight attitude of the RTAR and STAR were: 180 W, AC 100 kHz, and 20 m; and 300 W, AC or DC 100 kHz, and 80 m, respectively.

  5. Tether-mission design for multiple flybys of moon Europa (United States)

    Sanmartin, J. R. S.; Charro, M. C.; Sanchez-Arriaga, G. S. A.; Sanchez-Torres, A. S. T.


    A tether mission to carry out multiple flybys of Jovian moon Europa is here presented. There is general agreement on elliptic-orbit flybys of Europa resulting in cost to attain given scientific goals lower than if actually orbiting the moon, tethers being naturally fit to fly-by rather than orbit moons1. The present mission is similar in this respect to the Clipper mission considered by NASA, the basic difference lying in location of periapsis, due to different emphasis on mission-challenge metrics. Clipper minimizes damaging radiation-dose by avoiding the Jupiter neighborhood and its very harsh environment; periapsis would be at Europa, apoapsis as far as moon Callisto. As in all past outer-planet missions, Clipper faces, however, critical power and propulsion needs. On the other hand, tethers can provide both propulsion and power, but must reach near the planet to find high plasma density and magnetic field values, leading to high induced tether current, and Lorentz drag and power. The bottom line is a strong radiation dose under the very intense Radiation Belts of Jupiter. Mission design focuses on limiting dose. Perijove would be near Jupiter, at about 1.2-1.3 Jovian radius, apojove about moon Ganymede, corresponding to 1:1 resonance with Europa, so as to keep dose down: setting apojove at Europa, for convenient parallel flybys, would require two perijove passes per flyby (the Ganymede apojove, resulting in high eccentricity, about 0.86, is also less requiring on tether operations). Mission is designed to attain reductions in eccentricity per perijove pass as high as Δe ≈ - 0.04. Due the low gravity-gradient, tether spinning is necessary to keep it straight, plasma contactors placed at both ends taking active turns at being cathodic. Efficiency of capture of the incoming S/C by the tether is gauged by the ratio of S/C mass to tether mass; efficiency is higher for higher tape-tether length and lower thickness and perijove. Low tether bowing due to the Lorentz

  6. Evaluation of Relative Yeast Cell Surface Hydrophobicity Measured by Flow Cytometry

    Directory of Open Access Journals (Sweden)

    Lisa Colling


    Full Text Available Objective: To develop an efficient method for evaluating cell surface hydrophobicity and to apply the method to demonstrate the effects of fungal growth conditions on cell surface properties.

  7. Cell surface topology creates high Ca2+ signalling microdomains

    DEFF Research Database (Denmark)

    Brasen, Jens Christian; Olsen, Lars Folke; Hallett, Maurice B


    of a smooth cell surface predicts only moderate localized effects, the more realistic "wrinkled" surface topology predicts that Ca2+ concentrations up to 80 microM can persist within the folds of membranes for significant times. This intra-wrinkle location may account for 5% of the total cell volume. Using...... different geometries of wrinkles, our simulations show that high Ca2+ microdomains will be generated most effectively by long narrow membrane wrinkles of similar dimensions to those found experimentally. This is a new concept which has not previously been considered, but which has ramifications as the intra-wrinkle...

  8. Ice-Tethered Profiler Contributions to the Arctic Observing Network (United States)

    Toole, J.; Krishfield, R.; Proshutinsky, A.; Timmermans, M.


    One of the hoped-for legacies of the International Polar Year is a sustained observational program such as the Arctic Observing Network to document and build understanding of future climate and ecosystem change. In the spirit of the now-operational international Argo float program, investigators from North America, Europe and Japan are collaborating to deploy drifting, ice-based observatory instrument systems on and below floes in the Arctic to sample the polar atmosphere-ice-ocean system and to make the resulting data available to researchers world-wide in real time. One element of these observatories is the WHOI Ice-Tethered Profiler, first deployed in August 2004. The ITP consists of a surface float and electronics package that sits atop an ice floe, a weighted, plastic-jacketed wire-rope tether extending from the surface float through the ice and down to 750-800 m depth, and a profiling vehicle with sensor package that moves up and down the tether. To date, 30 ITP systems (funded by research programs in 5 countries) have been deployed in the Arctic that together have returned more than 10,000 high-vertical-resolution temperature and salinity profiles spanning approximately 7 to 760 m depth over all seasons. Examples of the science being conducted with these data will be presented, along with performance statistics for the ITP instruments and engineering improvements/enhancements that are being implemented. Plans for sustaining the ITP contribution to the Arctic Observing Network will also be reviewed and future international collaborations invited.

  9. Cell surface hydrophobicity is conveyed by S-layer proteins - A study in recombinant lactobacilli

    NARCIS (Netherlands)

    Mei, H.C. van der; Belt-Gritter, B. van de; Pouwels, P.H.; Martinez, B.; Busscher, H.J.


    Cell surface hydrophobicity is one of the most important factors controlling adhesion of microorganisms to surfaces. In this paper, cell surface properties of lactobacilli and recombinant lactobacilli with and without a surface layer protein (SLP) associated with cell surface hydrophobicity were det

  10. Tangling of Tethered Swimmers: Interactions between Two Nematodes (United States)

    Backholm, Matilda; Schulman, Rafael D.; Ryu, William S.; Dalnoki-Veress, Kari


    The tangling of two tethered microswimming worms serving as the ends of "active strings" is investigated experimentally and modeled analytically. C. elegans nematodes of similar size are caught by their tails using micropipettes and left to swim and interact at different separations over long times. The worms are found to tangle in a reproducible and statistically predictable manner, which is modeled based on the relative motion of the worm heads. Our results provide insight into the intricate tangling interactions present in active biological systems.

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

    Energy Technology Data Exchange (ETDEWEB)

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


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

  12. Tethered spinal cord syndrome with symptomatic onset in adulthood

    Institute of Scientific and Technical Information of China (English)

    HE Shi-sheng; ZHAO Ying-chuan; SHI Zhi-cai; LI Ming; HOU Tie-sheng; ZHANG Ye; WU Yun-gang


    @@ Tethered spinal cord syndrome(TCS)is a condition of overstretching or compression of the caudal part of the spinal cord caused by various spinal lesions,such as a tight filum terminale or an intraspinal lipoma.~(1-9) Though it is a well-recognized cause of neurological deterioration in childhood,its symptomatic onset in adulthood is uncommon.~(10-23) Eleven cases of TCS are presented here.In addition,their related clinical features,surgical procedures and outcomes are investigated.

  13. Tethered Transition Metals Promoted Photocatalytic System for Efficient Hydrogen Evolutions

    KAUST Repository

    Takanabe, Kazuhiro


    The present invention is directed, at least in part, to a process for improving the efficiency of a photocatalyst (a semiconductor photocatalyst) by tethering (depositing) a metal (e.g., metal ions of a late transition metal, such as nickel) to the semiconductor (photocatalyst) surface through the use of an organic ligand. More specifically, 1,2-ethanedithiol (EDT) functions as an excellent molecular linker (organic ligand) to attach a transition metal complex (e.g., nickel (Ni.sup.2+ ions)) to the semiconductor surface, which can be in the form of a cadmium sulfide surface. The photocatalyst has particular utility in generating hydrogen from H.sub.2S.

  14. Bacterial Cell Surface Adsorption of Rare Earth Elements (United States)

    Jiao, Y.; Park, D.; Reed, D.; Fujita, Y.; Yung, M.; Anderko, A.; Eslamimanesh, A.


    Rare earth elements (REE) play a critical role in many emerging clean energy technologies, including high-power magnets, wind turbines, solar panels, hybrid/electric vehicle batteries and lamp phosphors. In order to sustain demand for such technologies given current domestic REE shortages, there is a need to develop new approaches for ore processing/refining and recycling of REE-containing materials. To this end, we have developed a microbially-mediated bioadsorption strategy with application towards enrichment of REE from complex mixtures. Specifically, the bacterium Caulobacter crescentus was genetically engineered to display lanthanide binding tags (LBTs), short peptides that possess high affinity and specificity for rare earth elements, on its cell surface S-layer protein. Under optimal conditions, LBT-displayed cells adsorbed greater than 5-fold more REE than control cells lacking LBTs. Competition binding experiments with a selection of REEs demonstrated that our engineered cells could facilitate separation of light- from heavy- REE. Importantly, binding of REE onto our engineered strains was much more favorable compared to non-REE metals. Finally, REE bound to the cell surface could be stripped off using citrate, providing an effective and non-toxic REE recovery method. Together, this data highlights the potential of our approach for selective REE enrichment from REE containing mixtures.

  15. Methods To Identify Aptamers against Cell Surface Biomarkers

    Directory of Open Access Journals (Sweden)

    Frédéric Ducongé


    Full Text Available Aptamers are nucleic acid-based ligands identified through a process of molecular evolution named SELEX (Systematic Evolution of Ligands by Exponential enrichment. During the last 10-15 years, numerous aptamers have been developed specifically against targets present on or associated with the surface of human cells or infectious pathogens such as viruses, bacteria, fungi or parasites. Several of the aptamers have been described as potent probes, rivalling antibodies, for use in flow cytometry or microscopy. Some have also been used as drugs by inhibiting or activating functions of their targets in a manner similar to neutralizing or agonistic antibodies. Additionally, it is straightforward to conjugate aptamers to other agents without losing their affinity and they have successfully been used in vitro and in vivo to deliver drugs, siRNA, nanoparticles or contrast agents to target cells. Hence, aptamers identified against cell surface biomarkers represent a promising class of ligands. This review presents the different strategies of SELEX that have been developed to identify aptamers for cell surface-associated proteins as well as some of the methods that are used to study their binding on living cells.

  16. Documentation for Woods Hole Oceanographic Institution Ice-Tethered Profiler records archived at NCEI (NCEI Accession 0156293) (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — This accession contains documentation about the data format and processing of Ice-Tethered Profiler data. The Ice-Tethered Profiler (ITP) system consists of a small...

  17. Analytical investigation of the dynamics of tethered constellations in earth orbit (United States)

    Lorenzini, Enrico C.; Gullahorn, Gordon E.; Estes, Robert D.


    This Quarterly Report on Tethering in Earth Orbit deals with three topics: (1) Investigation of the propagation of longitudinal and transverse waves along the upper tether. Specifically, the upper tether is modeled as three massive platforms connected by two perfectly elastic continua (tether segments). The tether attachment point to the station is assumed to vibrate both longitudinally and transversely at a given frequency. Longitudinal and transverse waves propagate along the tethers affecting the acceleration levels at the elevator and at the upper platform. The displacement and acceleration frequency-response functions at the elevator and at the upper platform are computed for both longitudinal and transverse waves. An analysis to optimize the damping time of the longitudinal dampers is also carried out in order to select optimal parameters. The analytical evaluation of the performance of tuned vs. detuned longitudinal dampers is also part of this analysis. (2) The use of the Shuttle primary Reaction Control System (RCS) thrusters for blowing away a recoiling broken tether is discussed. A microcomputer system was set up to support this operation. (3) Most of the effort in the tether plasma physics study was devoted to software development. A particle simulation code has been integrated into the Macintosh II computer system and will be utilized for studying the physics of hollow cathodes.

  18. Stabilization of periodic solutions in a tethered satellite system by damping injection

    DEFF Research Database (Denmark)

    Larsen, Martin Birkelund; Blanke, Mogens


    presents a control design for stabilizing these periodic solutions. The design consists of a control law for stabilizing the open-loop equilibrium and a bias term which forces the system trajectory away from the equilibrium. The tether needs to be positioned away from open-loop equilibrium for the tether...

  19. Analysis of thermionic bare tether operation regimes in passive mode (United States)

    Sanmartín, J. R.; Chen, Xin; Sánchez-Arriaga, G.


    A thermionic bare tether (TBT) is a long conductor coated with a low work-function material. In drag mode, a tether segment extending from anodic end A to a zero-bias point B, with the standard Orbital-motion-limited current collection, is followed by a complex cathodic segment. In general, as bias becomes more negative in moving from B to cathodic end C, one first finds space-charge-limited (SCL) emission covering up to some intermediate point B*, then full Richardson-Dushman (RD) emission reaching from B* to end C. An approximate analytical study, which combines the current and voltage profile equations with results from asymptotic studies of the Vlasov-Poisson system for emissive probes, is carried out to determine the parameter domain covering two limit regimes, which are effectively controlled by just two dimensionless parameters involving ambient plasma and TBT material properties. In one such limit regime, no point B* is reached and thus no full RD emission develops. In an opposite regime, SCL segment BB* is too short to contribute significantly to the current balance.

  20. Cellulose Microfibril Formation by Surface-Tethered Cellulose Synthase Enzymes. (United States)

    Basu, Snehasish; Omadjela, Okako; Gaddes, David; Tadigadapa, Srinivas; Zimmer, Jochen; Catchmark, Jeffrey M


    Cellulose microfibrils are pseudocrystalline arrays of cellulose chains that are synthesized by cellulose synthases. The enzymes are organized into large membrane-embedded complexes in which each enzyme likely synthesizes and secretes a β-(1→4) glucan. The relationship between the organization of the enzymes in these complexes and cellulose crystallization has not been explored. To better understand this relationship, we used atomic force microscopy to visualize cellulose microfibril formation from nickel-film-immobilized bacterial cellulose synthase enzymes (BcsA-Bs), which in standard solution only form amorphous cellulose from monomeric BcsA-B complexes. Fourier transform infrared spectroscopy and X-ray diffraction techniques show that surface-tethered BcsA-Bs synthesize highly crystalline cellulose II in the presence of UDP-Glc, the allosteric activator cyclic-di-GMP, as well as magnesium. The cellulose II cross section/diameter and the crystal size and crystallinity depend on the surface density of tethered enzymes as well as the overall concentration of substrates. Our results provide the correlation between cellulose microfibril formation and the spatial organization of cellulose synthases.

  1. Piperidinium tethered nanoparticle-hybrid electrolyte for lithium metal batteries

    KAUST Repository

    Korf, Kevin S.


    We report on the synthesis of novel piperidinium-based ionic liquid tethered nanoparticle hybrid electrolytes and investigate their physical and electrochemical properties. Hybrid electrolytes based on the ionic liquid 1-methyl-1-propylpiperidinium bis(trifluoromethanesulfone) imide covalently tethered to silica nanoparticles (SiO2-PP-TFSI) were blended with propylene carbonate-1 M lithium bis(trifluoromethanesulfone) imide (LiTFSI). We employed NMR analysis to confirm the successful creation of the hybrid material. Dielectric and rheological measurements show that these electrolytes exhibit exceptional room-temperature DC ionic conductivity (10-2 to 10 -3 S cm-1) as well as high shear mechanical moduli (105 to 106 Pa). Lithium transference numbers were found to increase with particle loading and to reach values as high as 0.22 at high particle loadings where the particle jam to form a soft glassy elastic medium. Analysis of lithium electrodeposits obtained in the hybrid electrolytes using SEM and EDX spectra show that the SiO2-PP-TFSI nanoparticles are able to smooth lithium deposition and inhibit lithium dendrite proliferation in Li metal batteries. LTOSiO2-PP-TFSI/PC in 1 M LiTFSILi half-cells based on the SiO2-PP-TFSI hybrid electrolytes exhibit attractive voltage profiles and trouble-free extended cycling behavior over more than 1000 cycles of charge and discharge. This journal is © the Partner Organisations 2014.

  2. Flight mechanics applications for tethers in space: Cooperative Italian-US programs (United States)

    Bevilacqua, Franco; Merlina, Pietro; Anderson, John L.


    Since the 1974 proposal by Giuseppe Colombo to fly a tethered subsatellite from the Shuttle Orbiter, the creative thinking of many scientists and engineers from Italy and U.S. has generated a broad range of potential tether applications in space. Many of these applications have promise for enabling innovative research and operational activities relating to flight mechanics in earth orbit and at suborbital altitudes. From a flight mechanics standpoint the most interesting of the currently proposed flight demonstrations are: the second Tethered Satellite System experiment which offers both the potential for aerothermodynamics and hypersonics research and for atmospheric science research; the Tethered Initiated Space Recovery System which would enable orbital deboost and recovery of a re-entry vehicle and waste removal from a space station; and the Tether Elevator/Crawler System which would provide a variable microgravity environment and space station center of mass management. The outer atmospheric and orbital flight mechanics characteristics of these proposed tether flight demonstrations are described. The second Tethered Satellite System mission will deploy the tethered satellite earthward and will bring it as low as 130 km from ground and thus into the transition region between the atmosphere (non-ionized) and the partially ionized ionosphere. The atmospheric flight mechanics of the tethered satellite is discussed and simulation results are presented. The Tether Initiated Space Recovery System experiment will demonstrate the ability of a simple tether system to deboost and recover a reentry vehicle. The main feature of this demonstration is the utilization of a Small Expendable Deployment System (SEDS) and the low-tension deployment assumed to separate the reentry vehicle from the Shuttle. This low-tension deployment maneuver is discussed and its criticalities are outlined. The Tether Elevator/Crawler System is a new space element able to move in a controlled way

  3. Effects of tethering alkyl chains for amphiphilic ruthenium complex dyes on their adsorption to titanium oxide and photovoltaic properties. (United States)

    Ni, Jen-Shyang; Hung, Chun-Yi; Liu, Ken-Yen; Chang, Yu-Hsun; Ho, Kuo-Chuan; Lin, King-Fu


    Ruthenium (II) complex dye, Ru(4,4'-dicarboxyl-2,2'-bipyridine)(4-nonyl-2,2'-bipyridine) (NCS)(2), (denoted as RuC9) tethering single alkyl chain was synthesized and well characterized. Its adsorption behavior onto the mesoporous TiO(2) and photovoltaic properties were compared with Z907 which has similar chemical structure but tethers two alkyl chains. RuC9 dyes tend to aggregate into vesicles in the acetonitrile/t-butanol co-solvent as a result of the amphiphilic structure, whereas Z907 dyes aggregate into lamellae. The dye-sensitized solar cell (DSSC) with RuC9 dye showed higher short-circuit photocurrent than that with Z907, attributing to its higher molar optical extinction coefficient and more adsorption amount onto the mesoporous TiO(2). However, the DSSC with Z907 dye has higher open-circuit photovoltage and power conversion efficiency, presumably due to the fact that Z907 with more alkyl chains formed a molecular layer with higher hydrophobicity. It reduced the charge recombination in the interface between the dye-sensitized mesoporous TiO(2) and electrolyte as verified by the electrochemical impedance spectroscopy and intensity modulated photocurrent and photovoltage spectroscopies.

  4. Transport vesicle tethering at the trans Golgi network: coiled coil proteins in action

    Directory of Open Access Journals (Sweden)

    Pak-yan Patricia Cheung


    Full Text Available The Golgi complex is decorated with so-called Golgin proteins that share a common feature: a large proportion of their amino acid sequences are predicted to form coiled-coil structures. The possible presence of extensive coiled coils implies that these proteins are highly elongated molecules that can extend a significant distance from the Golgi surface. This property would help them to capture or trap inbound transport vesicles and to tether Golgi mini-stacks together. This review will summarize our current understanding of coiled coil tethers that are needed for the receipt of transport vesicles at the trans Golgi network. How do long tethering proteins actually catch vesicles? Golgi-associated, coiled coil tethers contain numerous binding sites for small GTPases, SNARE proteins, and vesicle coat proteins. How are these interactions coordinated and are any or all of them important for the tethering process? Progress towards understanding these questions and remaining, unresolved mysteries will be discussed.

  5. Shortest Path Planning for a Tethered Robot or an Anchored Cable

    Energy Technology Data Exchange (ETDEWEB)

    Xavier, P.G.


    We consider the problem of planning shortest paths for a tethered robot with a finite length tether in a 2D environment with polygonal obstacles. We present an algorithm that runs in time O((k{sub 1} + 1){sup 2}n{sup 4}) and finds the shortest path or correctly determines that none exists that obeys the constraints; here n is the number obstacle vertices, and k{sub 1} is the number loops in the initial configuration of the tether. The robot may cross its tether but nothing can cross obstacles, which cause the tether to bend. The algorithm applies as well for planning a shortest path for the free end of an anchored cable.

  6. Cell surface carbohydrates as prognostic markers in human carcinomas

    DEFF Research Database (Denmark)

    Dabelsteen, Erik


    Tumour development is usually associated with changes in cell surface carbohydrates. These are often divided into changes related to terminal carbohydrate structures, which include incomplete synthesis and modification of normally existing carbohydrates, and changes in the carbohydrate core...... structure. The latter includes chain elongation of both glycolipids and proteins, increased branching of carbohydrates in N-linked glycoproteins, and blocked synthesis of carbohydrates in O-linked mucin-like glycoproteins. In mature organisms, expression of distinct carbohydrates is restricted to specific...... cell types; within a given tissue, variation in expression may be related to cell maturation. Tumour-associated carbohydrate structures often reflect a certain stage of cellular development; most of these moieties are structures normally found in other adult or embryonic tissues. There is no unique...

  7. Mechanotransduction Across the Cell Surface and Through the Cytoskeleton (United States)

    Wang, Ning; Butler, James P.; Ingber, Donald E.


    Mechanical stresses were applied directly to cell surface receptors with a magnetic twisting device. The extracellular matrix receptor, integrin β_1, induced focal adhesion formation and supported a force-dependent stiffening response, whereas nonadhesion receptors did not. The cytoskeletal stiffness (ratio of stress to strain) increased in direct proportion to the applied stress and required intact microtubules and intermediate filaments as well as microfilaments. Tensegrity models that incorporate mechanically interdependent struts and strings that reorient globally in response to a localized stress mimicked this response. These results suggest that integrins act as mechanoreceptors and transmit mechanical signals to the cytoskeleton. Mechanotransduction, in turn, may be mediated simultaneously at multiple locations inside the cell through force-induced rearrangements within a tensionally integrated cytoskeleton.

  8. Vaccines based on the cell surface carbohydrates of pathogenic bacteria

    Directory of Open Access Journals (Sweden)

    Jones Christopher


    Full Text Available Glycoconjugate vaccines, in which a cell surface carbohydrate from a micro-organism is covalently attached to an appropriate carrier protein are proving to be the most effective means to generate protective immune responses to prevent a wide range of diseases. The technology appears to be generic and applicable to a wide range of pathogens, as long as antibodies against surface carbohydrates help protect against infection. Three such vaccines, against Haemophilus influenzae type b, Neisseria meningitidis Group C and seven serotypes of Streptococcus pneumoniae, have already been licensed and many others are in development. This article discusses the rationale for the development and use of glycoconjugate vaccines, the mechanisms by which they elicit T cell-dependent immune responses and the implications of this for vaccine development, the role of physicochemical methods in the characterisation and quality control of these vaccines, and the novel products which are under development.

  9. Novel eukaryotic enzymes modifying cell-surface biopolymers

    Directory of Open Access Journals (Sweden)

    Aravind L


    Full Text Available Abstract Background Eukaryotic extracellular matrices such as proteoglycans, sclerotinized structures, mucus, external tests, capsules, cell walls and waxes contain highly modified proteins, glycans and other composite biopolymers. Using comparative genomics and sequence profile analysis we identify several novel enzymes that could be potentially involved in the modification of cell-surface glycans or glycoproteins. Results Using sequence analysis and conservation we define the acyltransferase domain prototyped by the fungal Cas1p proteins, identify its active site residues and unify them to the superfamily of classical 10TM acyltransferases (e.g. oatA. We also identify a novel family of esterases (prototyped by the previously uncharacterized N-terminal domain of Cas1p that have a similar fold as the SGNH/GDSL esterases but differ from them in their conservation pattern. Conclusions We posit that the combined action of the acyltransferase and esterase domain plays an important role in controlling the acylation levels of glycans and thereby regulates their physico-chemical properties such as hygroscopicity, resistance to enzymatic hydrolysis and physical strength. We present evidence that the action of these novel enzymes on glycans might play an important role in host-pathogen interaction of plants, fungi and metazoans. We present evidence that in plants (e.g. PMR5 and ESK1 the regulation of carbohydrate acylation by these acylesterases might also play an important role in regulation of transpiration and stress resistance. We also identify a subfamily of these esterases in metazoans (e.g. C7orf58, which are fused to an ATP-grasp amino acid ligase domain that is predicted to catalyze, in certain animals, modification of cell surface polymers by amino acid or peptides. Reviewers This article was reviewed by Gaspar Jekely and Frank Eisenhaber

  10. Novel eukaryotic enzymes modifying cell-surface biopolymers (United States)


    Background Eukaryotic extracellular matrices such as proteoglycans, sclerotinized structures, mucus, external tests, capsules, cell walls and waxes contain highly modified proteins, glycans and other composite biopolymers. Using comparative genomics and sequence profile analysis we identify several novel enzymes that could be potentially involved in the modification of cell-surface glycans or glycoproteins. Results Using sequence analysis and conservation we define the acyltransferase domain prototyped by the fungal Cas1p proteins, identify its active site residues and unify them to the superfamily of classical 10TM acyltransferases (e.g. oatA). We also identify a novel family of esterases (prototyped by the previously uncharacterized N-terminal domain of Cas1p) that have a similar fold as the SGNH/GDSL esterases but differ from them in their conservation pattern. Conclusions We posit that the combined action of the acyltransferase and esterase domain plays an important role in controlling the acylation levels of glycans and thereby regulates their physico-chemical properties such as hygroscopicity, resistance to enzymatic hydrolysis and physical strength. We present evidence that the action of these novel enzymes on glycans might play an important role in host-pathogen interaction of plants, fungi and metazoans. We present evidence that in plants (e.g. PMR5 and ESK1) the regulation of carbohydrate acylation by these acylesterases might also play an important role in regulation of transpiration and stress resistance. We also identify a subfamily of these esterases in metazoans (e.g. C7orf58), which are fused to an ATP-grasp amino acid ligase domain that is predicted to catalyze, in certain animals, modification of cell surface polymers by amino acid or peptides. Reviewers This article was reviewed by Gaspar Jekely and Frank Eisenhaber PMID:20056006

  11. Substrate recognition by the cell surface palmitoyl transferase DHHC5. (United States)

    Howie, Jacqueline; Reilly, Louise; Fraser, Niall J; Vlachaki Walker, Julia M; Wypijewski, Krzysztof J; Ashford, Michael L J; Calaghan, Sarah C; McClafferty, Heather; Tian, Lijun; Shipston, Michael J; Boguslavskyi, Andrii; Shattock, Michael J; Fuller, William


    The cardiac phosphoprotein phospholemman (PLM) regulates the cardiac sodium pump, activating the pump when phosphorylated and inhibiting it when palmitoylated. Protein palmitoylation, the reversible attachment of a 16 carbon fatty acid to a cysteine thiol, is catalyzed by the Asp-His-His-Cys (DHHC) motif-containing palmitoyl acyltransferases. The cell surface palmitoyl acyltransferase DHHC5 regulates a growing number of cellular processes, but relatively few DHHC5 substrates have been identified to date. We examined the expression of DHHC isoforms in ventricular muscle and report that DHHC5 is among the most abundantly expressed DHHCs in the heart and localizes to caveolin-enriched cell surface microdomains. DHHC5 coimmunoprecipitates with PLM in ventricular myocytes and transiently transfected cells. Overexpression and silencing experiments indicate that DHHC5 palmitoylates PLM at two juxtamembrane cysteines, C40 and C42, although C40 is the principal palmitoylation site. PLM interaction with and palmitoylation by DHHC5 is independent of the DHHC5 PSD-95/Discs-large/ZO-1 homology (PDZ) binding motif, but requires a ∼ 120 amino acid region of the DHHC5 intracellular C-tail immediately after the fourth transmembrane domain. PLM C42A but not PLM C40A inhibits the Na pump, indicating PLM palmitoylation at C40 but not C42 is required for PLM-mediated inhibition of pump activity. In conclusion, we demonstrate an enzyme-substrate relationship for DHHC5 and PLM and describe a means of substrate recruitment not hitherto described for this acyltransferase. We propose that PLM palmitoylation by DHHC5 promotes phospholipid interactions that inhibit the Na pump.

  12. Engineering the cell surface display of cohesins for assembly of cellulosome-inspired enzyme complexes on Lactococcus lactis

    Directory of Open Access Journals (Sweden)

    Wieczorek Andrew S


    Full Text Available Abstract Background The assembly and spatial organization of enzymes in naturally occurring multi-protein complexes is of paramount importance for the efficient degradation of complex polymers and biosynthesis of valuable products. The degradation of cellulose into fermentable sugars by Clostridium thermocellum is achieved by means of a multi-protein "cellulosome" complex. Assembled via dockerin-cohesin interactions, the cellulosome is associated with the cell surface during cellulose hydrolysis, forming ternary cellulose-enzyme-microbe complexes for enhanced activity and synergy. The assembly of recombinant cell surface displayed cellulosome-inspired complexes in surrogate microbes is highly desirable. The model organism Lactococcus lactis is of particular interest as it has been metabolically engineered to produce a variety of commodity chemicals including lactic acid and bioactive compounds, and can efficiently secrete an array of recombinant proteins and enzymes of varying sizes. Results Fragments of the scaffoldin protein CipA were functionally displayed on the cell surface of Lactococcus lactis. Scaffolds were engineered to contain a single cohesin module, two cohesin modules, one cohesin and a cellulose-binding module, or only a cellulose-binding module. Cell toxicity from over-expression of the proteins was circumvented by use of the nisA inducible promoter, and incorporation of the C-terminal anchor motif of the streptococcal M6 protein resulted in the successful surface-display of the scaffolds. The facilitated detection of successfully secreted scaffolds was achieved by fusion with the export-specific reporter staphylococcal nuclease (NucA. Scaffolds retained their ability to associate in vivo with an engineered hybrid reporter enzyme, E. coli β-glucuronidase fused to the type 1 dockerin motif of the cellulosomal enzyme CelS. Surface-anchored complexes exhibited dual enzyme activities (nuclease and β-glucuronidase, and were

  13. Recognition and tethering of transport vesicles at the Golgi apparatus. (United States)

    Witkos, Tomasz M; Lowe, Martin


    The Golgi apparatus occupies a central position within the secretory pathway where it is a hub for vesicle trafficking. Distinct classes of transport vesicles traffic diverse cargoes into and out of this organelle, as well as between the different Golgi subcompartments. A key feature of Golgi trafficking is the specific recognition of transport vesicles at the different regions of the Golgi apparatus, required for the correct cargo delivery. Specificity is ensured by coiled-coil golgins and multi-subunit tethering complexes (MTCs), which act together to capture vesicles and promote their subsequent fusion with the Golgi membrane. In this review we discuss our current understanding of how golgins and MTCs function together to mediate the specific recognition of vesicles at the Golgi apparatus.

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

    Directory of Open Access Journals (Sweden)

    Jakob Andersson


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

  15. DOE Geothermal Data Repository - Tethering Data to Information: Preprint

    Energy Technology Data Exchange (ETDEWEB)

    Weers, J.; Anderson, A.


    Data are not inherently information. Without context, data are just numbers, figures, names, or points on a line. By assigning context to data, we can validate ideas, form opinions, and generate knowledge. This is an important distinction to information scientists, as we recognize that the context in which we keep our data plays a big part in generating its value. The mechanisms used to assign this context often include their own data, supplemental to the data being described and defining semantic relationships, commonly referred to as metadata. This paper provides the status of the DOE Geothermal Data Repository (DOE GDR), including recent efforts to tether data submissions to information, discusses the important distinction between data and information, outlines a path to generate useful knowledge from raw data, and details the steps taken in order to become a node on the National Geothermal Data System (NGDS).

  16. Kv7 channels can function without constitutive calmodulin tethering.

    Directory of Open Access Journals (Sweden)

    Juan Camilo Gómez-Posada

    Full Text Available M-channels are voltage-gated potassium channels composed of Kv7.2-7.5 subunits that serve as important regulators of neuronal excitability. Calmodulin binding is required for Kv7 channel function and mutations in Kv7.2 that disrupt calmodulin binding cause Benign Familial Neonatal Convulsions (BFNC, a dominantly inherited human epilepsy. On the basis that Kv7.2 mutants deficient in calmodulin binding are not functional, calmodulin has been defined as an auxiliary subunit of Kv7 channels. However, we have identified a presumably phosphomimetic mutation S511D that permits calmodulin-independent function. Thus, our data reveal that constitutive tethering of calmodulin is not required for Kv7 channel function.

  17. Kv7 Channels Can Function without Constitutive Calmodulin Tethering (United States)

    Alberdi, Araitz; Alaimo, Alessandro; Etxeberría, Ainhoa; Fernández-Orth, Juncal; Zamalloa, Teresa; Roura-Ferrer, Meritxell; Villace, Patricia; Areso, Pilar; Casis, Oscar; Villarroel, Alvaro


    M-channels are voltage-gated potassium channels composed of Kv7.2-7.5 subunits that serve as important regulators of neuronal excitability. Calmodulin binding is required for Kv7 channel function and mutations in Kv7.2 that disrupt calmodulin binding cause Benign Familial Neonatal Convulsions (BFNC), a dominantly inherited human epilepsy. On the basis that Kv7.2 mutants deficient in calmodulin binding are not functional, calmodulin has been defined as an auxiliary subunit of Kv7 channels. However, we have identified a presumably phosphomimetic mutation S511D that permits calmodulin-independent function. Thus, our data reveal that constitutive tethering of calmodulin is not required for Kv7 channel function. PMID:21980481

  18. Controlled Activation of Protein Rotational Dynamics Using Smart Hydrogel Tethering

    Energy Technology Data Exchange (ETDEWEB)

    Beech, Brenda M.; Xiong, Yijia; Boschek, Curt B.; Baird, Cheryl L.; Bigelow, Diana J.; Mcateer, Kathleen; Squier, Thomas C.


    Stimulus-responsive hydrogel materials that stabilize and control protein dynamics have the potential to enable a range of applications to take advantage of the inherent specificity and catalytic efficiencies of proteins. Here we describe the modular construction of a hydrogel using an engineered calmodulin (CaM) within a polyethylene glycol (PEG) matrix that involves the reversible tethering of proteins through an engineered CaM-binding sequence. For these measurements, maltose binding protein (MBP) was isotopically labeled with [13C] and [15N], permitting dynamic structural measurements using TROSY-HSQC NMR spectroscopy. Upon initial formation of hydrogels protein dynamics are suppressed, with concomitant increases in protein stability. Relaxation of the hydrogel matrix following transient heating results in the activation of protein dynamics and restoration of substrate-induced large-amplitude domain motions necessary for substrate binding.

  19. Excess area dependent scaling behavior of nano-sized membrane tethers

    CERN Document Server

    Ramakrishnan, N; Eckmann, David M; Ayyaswamy, Portnovo S; Baumgart, Tobias; Pucadyil, Thomas; Patil, Shivprasad; Weaver, Valerie M; Radhakrishnan, Ravi


    Thermal fluctuations in cell membranes manifest as an excess area (${\\cal A}_{\\rm ex}$) which governs a multitude of physical process at the sub-micron scale. We present a theoretical framework, based on an in silico tether pulling method, which may be used to reliably estimate ${\\cal A}_{\\rm ex}$ in live cells. The tether forces estimated from our simulations compare well with our experimental measurements for tethers extracted from ruptured GUVs and HeLa cells. We demonstrate the significance and validity of our method by showing that all our calculations along with experiments of tether extraction in 15 different cell types collapse onto two unified scaling relationships mapping tether force, tether radius, bending stiffness $\\kappa$, and membrane tension $\\sigma$. We show that $R_{\\rm bead}$, the size of the wetting region, is an important determinant of the radius of the extracted tether, which is equal to $\\xi=\\sqrt{\\kappa/2\\sigma}$ (a characteristic length scale of the membrane) for $R_{\\rm bead}\\xi$. ...

  20. The Effect of Tethers on Artificial Cell Membranes: A Coarse-Grained Molecular Dynamics Study (United States)

    Hoiles, William; Gupta, Rini; Cornell, Bruce; Krishnamurthy, Vikram


    Tethered bilayer lipid membranes (tBLMs) provide a stable platform for modeling the dynamics and order of biological membranes where the tethers mimic the cytoskeletal supports present in biological cell membranes. In this paper coarse-grained molecular dynamics (CGMD) is applied to study the effects of tethers on lipid membrane properties. Using results from the CGMD model and the overdamped Fokker-Planck equation, we show that the diffusion tensor and particle density of water in the tBLM is spatially dependent. Further, it is shown that the membrane thickness, lipid diffusion, defect density, free energy of lipid flip-flop, and membrane dielectric permittivity are all dependent on the tether density. The numerically computed results from the CGMD model are in agreement with the experimentally measured results from tBLMs containing different tether densities and lipids derived from Archaebacteria. Additionally, using experimental measurements from Escherichia coli bacteria and Saccharomyces Cerevisiae yeast tethered membranes, we illustrate how previous molecular dynamics results can be combined with the proposed model to estimate the dielectric permittivity and defect density of these membranes as a function of tether density. PMID:27736860

  1. Three-dimensional deployment of electro-dynamic tether via tension and current control with constraints (United States)

    Wen, Hao; Jin, Dongping; Hu, Haiyan


    The concept of space tether has found a great deal of promising applications in space engineering. A prerequisite of any space tether mission is to deploy its tether to a commanded length. This paper aims to achieving the three-dimensional deployment of an electro-dynamic tether system in a propellant-free manner via the feedback control of the tension and electric current in the tether. The proposed controller is formulated in an analytical form with an extremely low level of computational load, and can explicitly account for the physical bounds of the tether tension and electric current by using a pair of strictly increasing saturation functions. In addition, the Lyapunov analysis is made to gain an insight into the stability characteristics of the proposed control strategy. To facilitate the theoretical analysis, the dynamic model of the system is developed under the widely used dumbbell assumption, along with the geomagnetic field modeled using a tilted dipole approximation. Finally, numerical case studies on a representative electro-dynamic tether system are conducted to evaluate the performance of the proposed controller and the influence of the actuating conditions and orbital inclinations.

  2. Experimental studies on the human gait using a tethered pelvic assist device (T-PAD). (United States)

    Vashista, Vineet; Mustafa, S K; Agrawal, Sunil K


    This paper presents the prototype of a novel tethered pelvic assist device (T-PAD). This is a purely passive device, consisting of a set of elastic tethers with one end attached to a hip brace worn by a subject walking on a treadmill, and the other end attached to a fixed frame surrounding the subject. T-PAD offers the flexibility of varying the assistance required on the pelvis by changing the configuration of the tether attachment locations, number of tethers and tether elasticity. Experimental studies were conducted using a full and a partial pelvic constraint configuration of T-PAD, with varying tether elasticity. The studies were aimed at observing the effect of T-PAD on the human gait. Results show that T-PAD reduced the range-of-motion for the pelvic angles with increase of tether elasticity. However, it had mixed effects on the range-of-motion of the hip angles, but negligible effect on the knee and ankle joint angles. Overall, T-PAD shows potential as a low-cost pelvic support device with pelvic motion control capabilities, and can work in tandem with existing gait trainers.

  3. Heterobifunctional crosslinkers for tethering single ligand molecules to scanning probes

    Energy Technology Data Exchange (ETDEWEB)

    Riener, Christian K.; Kienberger, Ferry; Hahn, Christoph D.; Buchinger, Gerhard M.; Egwim, Innocent O.C.; Haselgruebler, Thomas; Ebner, Andreas; Romanin, Christoph; Klampfl, Christian; Lackner, Bernd; Prinz, Heino; Blaas, Dieter; Hinterdorfer, Peter; Gruber, Hermann J


    Single molecule recognition force microscopy (SMRFM) is a versatile atomic force microscopy (AFM) method to probe specific interactions of cognitive molecules on the single molecule level. It allows insights to be gained into interaction potentials and kinetic barriers and is capable of mapping interaction sites with nm positional accuracy. These applications require a ligand to be attached to the AFM tip, preferably by a distensible poly(ethylene glycol) (PEG) chain between the measuring tip and the ligand molecule. The PEG chain greatly facilitates specific binding of the ligand to immobile receptor sites on the sample surface. The present study contributes to tip-PEG-ligand tethering in three ways: (i) a convenient synthetic route was found to prepare NH{sub 2}-PEG-COOH which is the key intermediate for long heterobifunctional crosslinkers; (ii) a variety of heterobifunctional PEG derivatives for tip-PEG-ligand linking were prepared from NH{sub 2}-PEG-COOH; (iii) in particular, a new PEG crosslinker with one thiol-reactive end and one terminal nitrilotriacetic acid (NTA) group was synthesized and successfully used to tether His{sub 6}-tagged protein molecules to AFM tips via noncovalent NTA-Ni{sup 2+}-His{sub 6} bridges. The new crosslinker was applied to link a recombinant His{sub 6}-tagged fragment of the very-low density lipoprotein receptor to the AFM tip whereupon specific docking to the capsid of human rhinovirus particles was observed by force microscopy. In a parallel study, the specific interaction of the small GTPase Ran with the nuclear import receptor importin {beta}1 was studied in detail by SMRFM, using the new crosslinker to link His{sub 6}-tagged Ran to the measuring tip [Nat. Struct. Biol. (2003), 10, 553-557].

  4. Minimally invasive tethered cord release in children: A technical note

    Directory of Open Access Journals (Sweden)

    S. Kağan Başarslan


    Full Text Available Tethered cord release is commonly performed in pediatric neurosurgery. Nowadays, minimally invasive procedures are created growing interest due to its highly tolerable nature for surgery. It has been main purpose a minimal damaging on access route and maximum protection of normal structures in surgery. We present a surgical treatment of tethered cord syndrome, by which is provided the cord releasing unlike the many methods being applied with tissue removal. The main advantage of performing this surgery through 2 cm hole is to avoid removing ligamentum flavum and bony structure like lamina in addition to reduce the length of the incision and the related scar tissue. J Clin Exp Invest 2014; 5 (1: 115-117 Technical note: the patient was taken on the operating table in the sitting-prone position, and L5-S1 distance was determined by fluoroscopy. The skin and subcutaneous tissues was passed via a 2 cm vertical incision settled in 0.5 cm laterally from midline. L5-S1 distance and its covering ligamentum flavum are displayed by the guidance of L5 lamina. Williams’s retractor was placed in the distance after fetching microscope. The foregoing procedures are the same with microdiscectomic surgery. By a vertical incision made on the flavum, its both layer was lifted up and hanged with simple suture on the back tissue for a comfortable exposure of the Dura. Thecal sac was opened by 0.5 cm long vertical incision on the Dura after obtaining secure CSF drainage with the help of yellow-tipped syringe needle. With finding by a nerve hook, the phylum was burned and released securely. Then the Dura was sutured primarily for the closure by means of microsurgery instruments, and flavum was laid on it again.

  5. Equine herpesvirus type 4 UL56 and UL49.5 proteins downregulate cell surface major histocompatibility complex class I expression independently of each other. (United States)

    Said, Abdelrahman; Azab, Walid; Damiani, Armando; Osterrieder, Nikolaus


    Major histocompatibility complex class I (MHC-I) molecules are critically important in the host defense against various pathogens through presentation of viral peptides to cytotoxic T lymphocytes (CTLs), a process resulting in the destruction of virus-infected cells. Herpesviruses interfere with CTL-mediated elimination of infected cells by various mechanisms, including inhibition of peptide transport and loading, perturbation of MHC-I trafficking, and rerouting and proteolysis of cell surface MHC-I. In this study, we show that equine herpesvirus type 4 (EHV-4) modulates MHC-I cell surface expression through two different mechanisms. First, EHV-4 can lead to a significant downregulation of MHC-I expression at the cell surface through the product of ORF1, a protein expressed with early kinetics from a gene that is homologous to herpes simplex virus 1 UL56. The EHV-4 UL56 protein reduces cell surface MHC-I as early as 4 h after infection. Second, EHV-4 can interfere with MHC-I antigen presentation, starting at 6 h after infection, by inhibition of the transporter associated with antigen processing (TAP) through its UL49.5 protein. Although pUL49.5 has no immediate effect on overall surface MHC-I levels in infected cells, it blocks the supply of antigenic peptides to the endoplasmic reticulum (ER) and transport of peptide-loaded MHC-I to the cell surface. Taken together, our results show that EHV-4 encodes at least two viral immune evasion proteins: pUL56 reduces MHC-I molecules on the cell surface at early times after infection, and pUL49.5 interferes with MHC-I antigen presentation by blocking peptide transport in the ER.

  6. Cell Surface Binding and Internalization of Aβ Modulated by Degree of Aggregation

    Directory of Open Access Journals (Sweden)

    David A. Bateman


    Full Text Available The amyloid peptides, Aβ40 and Aβ42, are generated through endoproteolytic cleavage of the amyloid precursor protein. Here we have developed a model to investigate the interaction of living cells with various forms of aggregated Aβ40/42. After incubation at endosomal pH 6, we observed a variety of Aβ conformations after 3 (Aβ3, 24 (Aβ24, and 90 hours (Aβ90. Both Aβ4224 and Aβ4024 were observed to rapidly bind and internalize into differentiated PC12 cells, leading to accumulation in the lysosome. In contrast, Aβ40/4290 were both found to only weakly associate with cells, but were observed as the most aggregated using dynamic light scattering and thioflavin-T. Internalization of Aβ40/4224 was inhibited with treatment of monodansylcadaverine, an endocytosis inhibitor. These studies indicate that the ability of Aβ40/42 to bind and internalize into living cells increases with degree of aggregation until it reaches a maximum beyond which its ability to interact with cells diminishes drastically.

  7. Distribution, Arrangement and Interconnectedness of Cell Surface Receptor sites in the body of an Organism

    Directory of Open Access Journals (Sweden)



    Full Text Available Cell surface receptors have been identified as the sites of disease infectivity in living organisms in a previous study. Drugs used for the treatment or cure of infections have to eliminate infections through attacking infective organisms at the cell surface receptors to which the infective organisms are attached. Problem statement: The present study examines a wide sample of living things to get more information on the relationship of one cell surface receptor to other cell surface receptors in the body of an organism. Approach: The arrangement of cell surface receptors on the external covering of a few samples of fruits, leaves, stems, dry wood of a plant; wall gecko and some parts of the human body, were examined and photographed. Transverse and/or Longitudinal sections of soursop fruit and sycamore fruit were also examined and photographed. The five different coverings of the fleshy part of a coconut were also photographed. The photographs were studied to note the relationship of disease infection attached to cell surface receptors on the external surface of an organ to disease infection on the innermost covering of the same organ. Results: The results of the study showed that all living things had ubiquitous distribution of cell surface receptors which are usually observable with the unaided eye as dots or spots on the external covering of an organ, tissue or cell. The dots or receptor sites of cell surface receptors in the study are arranged in lines which were perpendicular, oblique, transverse or arranged in any other lineal geometrical form. The lineally arranged cell surface receptors were noted to be connected by grooves, channels or pipes which joined other receptor channels or intersected with them. Smaller cell surface receptor channels emptied into bigger channels or continued as small sized channels that ran side by side in a connective tissue bundle. These connective tissue bundles that carried many independent small-sized cell

  8. Nonlinear Control of Electrodynamic Tether in Equatorial or Somewhat Inclined Orbits

    DEFF Research Database (Denmark)

    Larsen, Martin Birkelund; Blanke, Mogens


    This paper applies different control design methods to a tethered satellite system (TSS) to investigate essential control properties of this under-actuated and nonlinear system. When the tether position in the orbit plane is controlled by the tether current, out of orbit plane motions occur...... as an unwanted side effect, due to nonlinear interaction with the Earth’s magnetic field. This paper focus on the uncontrollable out-of-plane motions and the robustness against B-field uncertainty associated with each of three popular controller design methodologies for nonlinear systems: linear quadratic...

  9. Characterization and use of crystalline bacterial cell surface layers (United States)

    Sleytr, Uwe B.; Sára, Margit; Pum, Dietmar; Schuster, Bernhard


    Crystalline bacterial cell surface layers (S-layers) are one of the most common outermost cell envelope components of prokaryotic organisms (archaea and bacteria). S-layers are monomolecular arrays composed of a single protein or glycoprotein species and represent the simplest biological membranes developed during evolution. S-layers as the most abundant of prokaryotic cellular proteins are appealing model systems for studying the structure, synthesis, genetics, assembly and function of proteinaceous supramolecular structures. The wealth of information existing on the general principle of S-layers have revealed a broad application potential. The most relevant features exploited in applied S-layer research are: (i) pores passing through S-layers show identical size and morphology and are in the range of ultrafiltration membranes; (ii) functional groups on the surface and in the pores are aligned in well-defined positions and orientations and accessible for chemical modifications and binding functional molecules in very precise fashion; (iii) isolated S-layer subunits from a variety of organisms are capable of recrystallizing as closed monolayers onto solid supports (e.g., metals, polymers, silicon wafers) at the air-water interface, on lipid films or onto the surface of liposomes; (iv) functional domains can be incorporated in S-layer proteins by genetic engineering. Thus, S-layer technologies particularly provide new approaches for biotechnology, biomimetics, molecular nanotechnology, nanopatterning of surfaces and formation of ordered arrays of metal clusters or nanoparticles as required for nanoelectronics.

  10. RPE cell surface proteins in normal and dystrophic rats

    Energy Technology Data Exchange (ETDEWEB)

    Clark, V.M.; Hall, M.O.


    Membrane-bound proteins in plasma membrane enriched fractions from cultured rat RPE were analyzed by two-dimensional gel electrophoresis. Membrane proteins were characterized on three increasingly specific levels. Total protein was visualized by silver staining. A maximum of 102 separate proteins were counted in silver-stained gels. Glycoproteins were labeled with 3H-glucosamine or 3H-fucose and detected by autoradiography. Thirty-eight fucose-labeled and 61-71 glucosamine-labeled proteins were identified. All of the fucose-labeled proteins were labeled with glucosamine-derived radioactivity. Proteins exposed at the cell surface were labeled by lactoperoxidase-catalyzed radioiodination prior to preparation of membranes for two-dimensional analysis. Forty separate 125I-labeled surface proteins were resolved by two-dimensional electrophoresis/autoradiography. Comparison with the glycoprotein map showed that a number of these surface labeled proteins were glycoproteins. Two-dimensional maps of total protein, fucose-labeled, and glucosamine-labeled glycoproteins, and 125I-labeled surface proteins of membranes from dystrophic (RCS rdy-p+) and normal (Long Evans or RCS rdy+p+) RPE were compared. No differences in the total protein or surface-labeled proteins were observed. However, the results suggest that a 183K glycoprotein is more heavily glycosylated with glucosamine and fucose in normal RPE membranes as compared to membranes from dystrophic RPE.

  11. Antifouling property of highly oleophobic substrates for solar cell surfaces (United States)

    Fukada, Kenta; Nishizawa, Shingo; Shiratori, Seimei


    Reduction of solar cell conversion efficiency by bird spoor or oil smoke is a common issue. Maintaining the surface of solar cells clean to retain the incident light is of utmost importance. In this respect, there has been growing interest in the area of superhydrophobicity for developing water repelling and self-cleaning surfaces. This effect is inspired by lotus leaves that have micro papillae covered with hydrophobic wax nanostructures. Superhydrophobic surfaces on transparent substrates have been developed for removing contaminants from solar cell surfaces. However, oil cannot be removed by superhydrophobic effect. In contrast, to prevent bird spoor, a highly oleophobic surface is required. In a previous study, we reported transparent-type fabrics comprising nanoparticles with a nano/micro hierarchical structure that ensured both oleophobicity and transparency. In the current study, we developed new highly oleophobic stripes that were constructed into semi-transparent oleophobic surfaces for solar cells. Solar cell performance was successfully maintained; the total transmittance was a key factor for determining conversion efficiency.

  12. Studies of cell-surface glorin receptors, glorin degradation, and glorin-induced cellular responses during development of Polysphondylium violaceum. (United States)

    De Wit, R J; van Bemmelen, M X; Penning, L C; Pinas, J E; Calandra, T D; Bonner, J T


    The chemoattractant mediating cell aggregation in the slime mold Polysphondylium violaceum is N-propionyl-gamma-L-glutamyl-L-ornithine-delta-lactam ethylester (glorin). Here we examine the binding properties of tritiated glorin to intact P. violaceum cells. Scatchard analysis of binding data yielded slightly curvilinear plots with Kd values in the range of 20 and 100 nM. The number of glorin receptors increased from 35,000 in the vegetative stage to 45,000 per cell during aggregation. Later, during culmination receptor numbers decreased to undetectable levels (less than 1000). The receptor binding kinetics show binding equilibrium within 30 s at 0 degrees C, and ligand dissociation occurs from two kinetically distinct receptors whose half-times were 2 s for 72% of the bound glorin and 28 s for the remainder. The enzymatic degradation of glorin did not affect binding data during incubations of up to 1 min at 0 degrees C. Two glorinase activities were observed. An ornithine delta-lactam cleaving activity with a Km of ca. 10(-4) M and a propionic acid removing activity (Km 10(-5) M), both of which were detected mainly on the cell surface. Cleavage of the lactam occurred at a higher rate than removal of propionic acid. Lactam-cleaved glorin showed no chemotactic activity nor did it bind to cell-surface glorin receptors. Cell-surface-bound glorinase activity and glorin-induced cGMP synthesis were developmentally regulated, peaking at aggregation. In the most sensitive stage half-maximal responses (cGMP synthesis, chemotaxis, light-scattering) were elicited in the 10-100 nM range. Neither cAMP synthesis nor glorin-induced glorin synthesis was observed. Guanine nucleotides specifically modulated glorin receptor binding on isolated membranes, and, conversely, glorin modulated GTP gamma S binding to membrane preparations. Our results support the notion that glorin mediates chemotactic cell aggregation in P. violaceum acting via cell-surface receptors, G-proteins, and c

  13. Materials for Advancement of MXER Tether Design (1000-549) Project (United States)

    National Aeronautics and Space Administration — There exist a need to develop, identify, and classify various materials that can be used in the fabrication of electrodynamic tethers for various applications. These...

  14. Construction and Structural Analysis of Tethered Lipid Bilayer Containing Photosynthetic Antenna Proteins for Functional Analysis

    Energy Technology Data Exchange (ETDEWEB)

    Sumino, Ayumi; Dewa, Takehisa; Takeuchi, Toshikazu; Sugiura, Ryuta; Sasaki, Nobuaki; Misawa, Nobuo; Tero, Ryugo; Urisu, Tsuneo; Gardiner, Alastair T; Cogdell, Richard J; Hashimoto, Hideki; Nango, Mamoru


    The construction and structural analysis of a tethered planar lipid bilayer containing bacterial photosynthetic membrane proteins, light-harvesting complex 2 (LH2), and light-harvesting core complex (LH1-RC) is described and establishes this system as an experimental platform for their functional analysis. The planar lipid bilayer containing LH2 and/or LH1-RC complexes was successfully formed on an avidin-immobilized coverglass via an avidin-biotin linkage. Atomic force microscopy (AFM) showed that a smooth continuous membrane was formed there. Lateral diffusion of these membrane proteins, observed by a fluorescence recovery after photobleaching (FRAY), is discussed in terms of the membrane architecture. Energy transfer from LH2 to LH1-RC within the tethered membrane architecture. Energy transfer from LH2 to LH1-RC within the tethered membrane was observed by steady-state fluorescence spectroscopy, indicating that the tethered membrane can mimic the natural situation.

  15. An Effective Approach Control Scheme for the Tethered Space Robot System

    Directory of Open Access Journals (Sweden)

    Zhongjie Meng


    Full Text Available The tethered space robot system (TSR, which is composed of a platform, a gripper and a space tether, has great potential in future space missions. Given the relative motion among the platform, tether, gripper and the target, an integrated approach model is derived. Then, a novel coordinated approach control scheme is presented, in which the tether tension, thrusters and the reaction wheel are all utilized. It contains the open-loop trajectory optimization, the feedback trajectory control and attitude control. The numerical simulation results show that the rendezvous between TSR and the target can be realized by the proposed coordinated control scheme, and the propellant consumption is efficiently reduced. Moreover, the control scheme performs well in the presence of the initial state’s perturbations, actuator characteristics and sensor errors.

  16. Three-dimensional multi-tethered satellite formation with the elements moving along Lissajous curves (United States)

    Yarotsky, D.; Sidorenko, V.; Pritykin, D.


    This note presents a novel approach to maintain three-dimensional multi-tethered satellite formation in space. For a formation consisting of a main body connected by tethers with several deputy satellites (the so-called "hub-and-spoke" configuration) we demonstrate that under proper choice of the system's parameters the deputy satellites can move along Lissajous curves in the plane normal to the local vertical with all tethers stretched; the total force due to the tension forces acting on the main satellite is balanced in a way allowing it to be in relative equilibrium strictly below or strictly above the system's center of mass. We analyze relations between the system's essential parameters and obtain conditions under which the proposed motion does take place. We also study analytically the motion stability for different configurations and whether the deputy satellites can collide or the tethers can entangle. Our theoretical findings are corroborated and validated by numerical experiments.

  17. Advanced Particle-in-Cell (PIC) Tools for Simulation of Electrodynamic Tether Plasma Interactions Project (United States)

    National Aeronautics and Space Administration — Electrodynamic tethers are optimally suited for use in Low-Earth-Orbit (LEO) to generate thrust or drag maneuver satellites. LEO region is polluted with space debris...

  18. Adaptive sliding mode control of tethered satellite deployment with input limitation (United States)

    Ma, Zhiqiang; Sun, Guanghui


    This paper proposes a novel adaptive sliding mode tension control method for the deployment of tethered satellite, where the input tension limitation is taken into account. The underactuated governing equations of the tethered satellites system are firstly derived based on Lagrangian mechanics theory. Considering the fact that the tether can only resist axial stretching, the tension input is modelled as input limitation. New adaptive sliding mode laws are addressed to guarantee the stability of the tethered satellite deployment with input disturbance, meanwhile to eliminate the effect of the limitation features of the tension input. Compared with the classic control strategy, the newly proposed adaptive sliding mode control law can deploy the satellite with smaller overshoot of the in-plane angle and implement the tension control reasonably and effectively in engineering practice. The numerical results validate the effectiveness of the proposed methods.

  19. UV-Curable Hybrid Nanocomposite Coating to Protect Tether Polymer Materials Project (United States)

    National Aeronautics and Space Administration — To address the NASA need for coatings to protect and strengthen tether materials for Momentum-exchange Electrodynamic Reboost (MXER) technology, Luminit, LLC,...

  20. Modeling and Control of Electrodynamic Tethers - an Energy and Topology Approach

    DEFF Research Database (Denmark)

    Larsen, Martin Birkelund

    of propellant a spacecraft need to bring from Earth can be reduced. In this thesis the modeling and control of electrodynamic tethers are investigated, both when a single tether is used to connect two spacecrafts, and when the tethers are used i more general formations of spacecrafts. One of the main challenges......, and separate derivations of the dynamical equations can be cumbersome. It can therefore be advantageous to be able to model a formation independent of its topology, i.e. the way tethers and satellites are interconnected. The thesis treats a class of formations in a generic framework, using graph theory...... to describe the topology of the formations. The framework can be used both to deduce the equations of motion for the attitude motion of the formation and for control design regarding the same motion. The main part of the thesis consists of five scientific papers which have been submitted for international...

  1. Myocardial Infarction Alters Adaptation of the Tethered Mitral Valve

    NARCIS (Netherlands)

    Dal-Bianco, Jacob P; Aikawa, Elena; Bischoff, Joyce; Guerrero, J Luis; Hjortnaes, Jesper; Beaudoin, Jonathan; Szymanski, Catherine; Bartko, Philipp E; Seybolt, Margo M; Handschumacher, Mark D; Sullivan, Suzanne; Garcia, Michael L; Mauskapf, Adam; Titus, James S; Wylie-Sears, Jill; Irvin, Whitney S; Chaput, Miguel; Messas, Emmanuel; Hagège, Albert A; Carpentier, Alain; Levine, Robert A


    BACKGROUND: In patients with myocardial infarction (MI), leaflet tethering by displaced papillary muscles induces mitral regurgitation (MR), which doubles mortality. Mitral valves (MVs) are larger in such patients but fibrosis sets in counterproductively. The investigators previously reported that e

  2. Materials for advancement of MXER tether design (1000-371) Project (United States)

    National Aeronautics and Space Administration — There exist a need to develop, identify, and classify various materials that can be used in the fabrication of electrodynamic tethers for various applications. These...

  3. An efficient delivery of DAMPs on the cell surface by the unconventional secretion pathway

    Energy Technology Data Exchange (ETDEWEB)

    Zhu, Haiyan; Wang, Lan; Ruan, Yuanyuan; Zhou, Lei; Zhang, Dongmei [Department of Biochemistry and Molecular Biology, Shanghai Medical Collage, Fudan University, Shanghai 200032 (China); Min, Zhihui [Biomedical Research Center, Zhongshan Hospital, Fudan University, Shanghai 200032 (China); Xie, Jianhui [Department of Biochemistry and Molecular Biology, Shanghai Medical Collage, Fudan University, Shanghai 200032 (China); Yu, Min, E-mail: [Department of Biochemistry and Molecular Biology, Shanghai Medical Collage, Fudan University, Shanghai 200032 (China); Gu, Jianxin [Department of Biochemistry and Molecular Biology, Shanghai Medical Collage, Fudan University, Shanghai 200032 (China)


    Research highlights: {yields} Hsp60 transported to cell surface through the classical secretory pathway was modified with N-glycosylation. {yields} HSAPB-N18 could efficiently deliver Hsp60 to the cell surface via the unconventional secretory pathway. {yields} Cell surface Hsp60 delivered by HASPB-N18 has a proper conformation. {yields} HASPB-N18 is an efficient delivery signal for other DAMP molecules such as Hsp70 and HMGB1. -- Abstract: Damage-associated molecular patterns (DAMPs) are signals released from dying cells evoking the immune system response in several inflammatory disorders. In normal situations, many of DAMPs are nuclear or cytosolic proteins with defined intracellular function, but they could be found on the cell surface following tissue injury. The biological function of the translocated DAMPs is still not well known and an efficient delivery of these molecules on the cell surface is required to clarify their biological effects. In this study, we demonstrated that an unclassical secretory signal peptide, N-terminal 18 amino acids of HASPB (HASPB-N18), could efficiently deliver Hsp60, Hsp70, and HMGB1 on the cell surface. Furthermore, the delivery of these molecules on the cell surface by HASPB-N18 is not limited to a special cell line because several cell lines could use this delivery signal to deliver these molecules on the cell surface. Moreover, we demonstrated that Hsp60 on the cell surface delivered by HASPB-N18 could be recognized by a soluble form of LOX-1, which implies that DAMPs on the cell surface delivered by HASPB-N18 have a proper conformation during transport. Therefore, delivery of DAMPs by HASPB-N18 is a reliable model to further understand the biological significance of DAMPs on the cell surface.

  4. Developmentally regulated epitopes of cell surface arabinogalactan proteins and their relation to root tissue pattern formation. (United States)

    Knox, J P; Linstead, P J; Peart, J; Cooper, C; Roberts, K


    Two polymorphic forms of an extracellular arabinogalactan protein (AGP1 and AGP2), obtained from the conditioned media of two carrot suspension-cultured cell lines, have been identified in terms of binding of the anti-plasma membrane antibodies JIM4 and MAC207. AGP1 and AGP2 have been used as immunogens to generate further anti-AGP monoclonal antibodies. JIM14 identified an epitope carried by AGP2 and also by glycoproteins of low molecular weight localized to the plant cell wall. In addition, further antibodies (JIM13 and JIM15) identified carbohydrate epitopes of the AGPs that also occur on plasma membrane glycoproteins and are expressed by patterns of cells that reflect cell position at the carrot root apex. Indirect immunofluorescence microscopy indicated that JIM13 recognized the surface of cells forming the epidermis and cells marking the region and axis of the future xylem. JIM15 recognized a pattern of cells directly complementary to the JIM13 pattern. The panel of anti-AGP monoclonal antibodies now available indicates groups of cells within the root meristem that may reflect an early pre-pattern of the tissues of the mature root structure and suggests extensive modulation of cell surface AGPs during cell development and the positioning of cells within the apex.

  5. A membrane-tethered transcription factor ANAC089 negatively regulates floral initiation in Arabidopsis thaliana

    Institute of Scientific and Technical Information of China (English)


    The plant-specific NAC (NAM, ATAF1/2,and CUC2) transcription factors have a regulatory function in developmental processes and stress responses. Notably a group of NAC members named NTLs (NTM1-Like) are membrane-tethered, ensuring plants rapidly respond to developmental changes and environmental stimuli. Our results indicated that ANAC089 was a membrane-tethered transcription factor and its truncated form was responsible for the physiological function in flowering time control.

  6. Design and Synthesis of Novel Isoxazole Tethered Quinone-Amino Acid Hybrids

    Directory of Open Access Journals (Sweden)

    P. Ravi Kumar


    Full Text Available A new series of isoxazole tethered quinone-amino acid hybrids has been designed and synthesized involving 1,3-dipolar cycloaddition reaction followed by an oxidation reaction using cerium ammonium nitrate (CAN. Using this method, for the first time various isoxazole tethered quinone-phenyl alanine and quinone-alanine hybrids were synthesized from simple commercially available 4-bromobenzyl bromide, propargyl bromide, and 2,5-dimethoxybenzaldehyde in good yield.

  7. Dissecting docking and tethering of secretory vesicles at the target membrane (United States)

    Toonen, Ruud F; Kochubey, Olexiy; de Wit, Heidi; Gulyas-Kovacs, Attila; Konijnenburg, Bas; Sørensen, Jakob B; Klingauf, Jurgen; Verhage, Matthijs


    Secretory vesicles dock at their target in preparation for fusion. Using single-vesicle total internal reflection fluorescence microscopy in chromaffin cells, we show that most approaching vesicles dock only transiently, but that some are captured by at least two different tethering modes, weak and strong. Both vesicle delivery and tethering depend on Munc18-1, a known docking factor. By decreasing the amount of cortical actin by Latrunculin A application, morphological docking can be restored artificially in docking-deficient munc18-1 null cells, but neither strong tethering nor fusion, demonstrating that morphological docking is not sufficient for secretion. Deletion of the t-SNARE and Munc18-1 binding partner syntaxin, but not the v-SNARE synaptobrevin/VAMP, also reduces strong tethering and fusion. We conclude that docking vesicles either undock immediately or are captured by minimal tethering machinery and converted in a munc18-1/syntaxin-dependent, strongly tethered, fusion-competent state. PMID:16902411

  8. Tethered motion of uniflagellated bacteria at the liquid-solid surface (United States)

    Bell, Jordan; Tang, Jay

    Direct evidence of the bacterial flagellar motor's rotation was first noted when multiflagellated bacterial cells were observed (under the optical microscope) to rotate when tethered to glass by a single flagellum. The tethered cell assay has continued to play a significant role throughout the subsequent studies of motor characteristics and behavior. Such studies have expanded to include uniflagellated bacteria, such as Vibrio alginolyticus, Pseudomonas aeruginosa, and Caulobacter crescentus. Here we show that such cells are not necessarily tethered by their flagellum, but rather elsewhere on the cell body. The observed cell body rotation is actually due to the flagellum either ``rolling'' against the glass surface, or pushing the cell body at the flagellar base. These motions are directly observed for Vibrio alginolyticus with darkfield microscopy. Additionally, our recently measured distributions of intervals between motor switches for tethered Caulobacter crescentus also confirm this more complicated mode of tethering. Therefore, the rotational speed of tethered uniflagellated bacteria may not equate to that of the motor itself, as is commonly assumed.

  9. The mechanics of motorised momentum exchange tethers when applied to active debris removal from LEO (United States)

    Caldecott, Ralph; Kamarulzaman, Dayangku N. S.; Kirrane, James P.; Cartmell, Matthew P.; Ganilova, Olga A.


    The concept of momentum exchange when applied to space tethers for propulsion is well established, and a considerable body of literature now exists on the on-orbit modelling, the dynamics, and also the control of a large range of tether system applications. The authors consider here a new application for the Motorised Momentum Exchange Tether by highlighting three key stages of development leading to a conceptualisation that can subsequently be developed into a technology for Active Debris Removal. The paper starts with a study of the on-orbit mechanics of a full sized motorised tether in which it is shown that a laden and therefore highly massasymmetrical tether can still be forced to spin, and certainly to librate, thereby confirming its possible usefulness for active debris removal (ADR). The second part of the paper concentrates on the modelling of the centripetal deployment of a symmetrical MMET in order to get it initialized for debris removal operations, and the third and final part of the paper provides an entry into scale modelling for low cost mission design and testing. It is shown that the motorised momentum exchange tether offers a potential solution to the removal of large pieces of orbital debris, and that dynamic methodologies can be implemented to in order to optimise the emergent design.

  10. X-ray photoelectron spectroscopy for the study of microbial cell surfaces

    NARCIS (Netherlands)

    van der Mei, Henderina C; de Vries, Jacob; Busscher, Hendrik J


    X-ray photoelectron spectroscopy (XPS) is well known for the characterisation of material surfaces, but at first glance, is an unexpected technique to study the composition of microbial cell surfaces. Despite the fact that intimate contact between materials and microbial cell surfaces occurs in many

  11. Targeting Cell Surface Proteins in Molecular Photoacoustic Imaging to Detect Ovarian Cancer Early (United States)


    10-1-0422 TITLE: Targeting Cell Surface Proteins in Molecular Photoacoustic Imaging to Detect Ovarian Cancer Early PRINCIPAL...DATES COVERED 1 July 2010 - 30 June 2013 4. TITLE AND SUBTITLE Targeting Cell Surface Proteins in Molecular 5a. CONTRACT NUMBER Photoacoustic ...upon request). Aim 2) Prioritize ovarian cancer-associated surface proteins for their utility as molecular photoacoustic imaging targets and

  12. A reference guide to microbial cell surface hydrophobicity based on contact angles

    NARCIS (Netherlands)

    van der Mei, HC; Busscher, HJ; Bos, R.R.M.


    Acid-base interactions form the origin of the hydrophobicity of microbial cell-surfaces and can be quantitated from contact angle measurements on microbial lawns with water, formamide, methyleneiodide and/or alpha-bromonaphthalene. This review provides a reference guide to microbial cell surface hyd

  13. Regulation of ADAM12 cell-surface expression by protein kinase C epsilon

    DEFF Research Database (Denmark)

    Sundberg, Christina; Thodeti, Charles Kumar; Kveiborg, Marie;


    as a constitutively active protein. However, little is known about the regulation of ADAM12 cell-surface translocation. Here, we used human RD rhabdomyosarcoma cells, which express ADAM12 at the cell surface, in a temporal pattern. We report that protein kinase C (PKC) epsilon induces ADAM12 translocation to the cell...

  14. Cell surface expression of glycosylated, nonglycosylated, and truncated forms of a cytoplasmic protein pyruvate kinase. (United States)

    Hiebert, S W; Lamb, R A


    The soluble cytoplasmic protein pyruvate kinase (PK) has been expressed at the cell surface in a membrane-anchored form (APK). The hybrid protein contains the NH2-terminal signal/anchor domain of a class II integral membrane protein (hemagglutinin/neuraminidase, of the paramyxovirus SV5) fused to the PK NH2 terminus. APK contains a cryptic site that is used for N-linked glycosylation but elimination of this site by site-specific mutagenesis does not prevent cell surface localization. Truncated forms of the APK molecule, with up to 80% of the PK region of APK removed, can also be expressed at the cell surface. These data suggest that neither the complete PK molecule nor its glycosylation are necessary for intracellular transport of PK to the cell surface, and it is possible that specific signals may not be needed in the ectodomain of this hybrid protein to specify cell surface localization.

  15. Lipoproteins tethered dendrimeric nanoconstructs for effective targeting to cancer cells (United States)

    Jain, Anupriya; Jain, Keerti; Mehra, Neelesh Kumar; Jain, N. K.


    In the present investigation, poly (propylene imine) dendrimers up to fifth generation (PPI G5.0) were synthesized using ethylene diamine and acrylonitrile. Lipoproteins (high-density lipoprotein; HDL and low-density lipoprotein; LDL) were isolated from human plasma by discontinuous density gradient ultracentrifugation, characterized and tethered to G5.0 PPI dendrimers to construct LDL- and HDL-conjugated dendrimeric nanoconstructs for tumor-specific delivery of docetaxel. Developed formulations showed sustained release characteristics in in vitro drug release and in vivo pharmacokinetic studies. The cancer targeting potential of lipoprotein coupled dendrimers was investigated by ex vivo cytotoxicity and cell uptake studies using human hepatocellular carcinoma cell lines (HepG2 cells) and biodistribution studies in albino rats of Sprague-Dawley strain. Lipoprotein anchored dendrimeric nanoconstructs showed significant uptake by cancer cells as well as higher biodistribution of docetaxel to liver and spleen. It is concluded that these precisely synthesized engineered dendrimeric nanoconstructs could serve as promising drug carrier for fighting with the fatal disease, i.e., cancer, attributed to their defined targeting and therapeutic potential.

  16. Lipoproteins tethered dendrimeric nanoconstructs for effective targeting to cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Jain, Anupriya; Jain, Keerti, E-mail:; Mehra, Neelesh Kumar, E-mail:; Jain, N. K., E-mail: [Dr. H. S. Gour University, Pharmaceutics Research Laboratory, Department of Pharmaceutical Sciences (India)


    In the present investigation, poly (propylene imine) dendrimers up to fifth generation (PPI G5.0) were synthesized using ethylene diamine and acrylonitrile. Lipoproteins (high-density lipoprotein; HDL and low-density lipoprotein; LDL) were isolated from human plasma by discontinuous density gradient ultracentrifugation, characterized and tethered to G5.0 PPI dendrimers to construct LDL- and HDL-conjugated dendrimeric nanoconstructs for tumor-specific delivery of docetaxel. Developed formulations showed sustained release characteristics in in vitro drug release and in vivo pharmacokinetic studies. The cancer targeting potential of lipoprotein coupled dendrimers was investigated by ex vivo cytotoxicity and cell uptake studies using human hepatocellular carcinoma cell lines (HepG2 cells) and biodistribution studies in albino rats of Sprague-Dawley strain. Lipoprotein anchored dendrimeric nanoconstructs showed significant uptake by cancer cells as well as higher biodistribution of docetaxel to liver and spleen. It is concluded that these precisely synthesized engineered dendrimeric nanoconstructs could serve as promising drug carrier for fighting with the fatal disease, i.e., cancer, attributed to their defined targeting and therapeutic potential.

  17. Detergent interaction with tethered bilayer lipid membranes for protein reconstitution (United States)

    Broccio, Matteo; Zan Goh, Haw; Loesche, Mathias


    Tethered bilayer lipid membranes (tBLMs) are self-assembled biomimetic structures in which the membrane is separated from a solid substrate by a nm-thick hydrated submembrane space. These model systems are being used in binding studies of peripheral proteins and exotoxins. Here we aim at their application for the reconstitution of water-insoluble integral membrane proteins. As an alternative to fusion of preformed proteoliposomes we study the direct reconstitution of such proteins for applications in biosensing and pharmaceutical screening. For reconstitution, highly insulating tBLMs (R˜10^5-10^6 φ) were temporarily incubated with a detergent to screen for conditions that keep the detergent-saturated membranestable and ready to incorporate detergent-solubilized proteins. We assess the electrical characteristics, i.e. specific resistance and capacitance, by means of electrochemical impedance spectroscopy (EIS) under timed incubation with decylmaltoside and dodecylmaltoside detergents in a regime around their critical micelle concentration, 1.8 mM and 0.17 mM respectively and demonstrate the restoration of the tBLM upon detergent removal. Thereby a range of concentration and incubation times was identified, that represents optimal conditions for the subsequent membrane protein reconstitution.

  18. Ectopically tethered CP190 induces large-scale chromatin decondensation (United States)

    Ahanger, Sajad H.; Günther, Katharina; Weth, Oliver; Bartkuhn, Marek; Bhonde, Ramesh R.; Shouche, Yogesh S.; Renkawitz, Rainer


    Insulator mediated alteration in higher-order chromatin and/or nucleosome organization is an important aspect of epigenetic gene regulation. Recent studies have suggested a key role for CP190 in such processes. In this study, we analysed the effects of ectopically tethered insulator factors on chromatin structure and found that CP190 induces large-scale decondensation when targeted to a condensed lacO array in mammalian and Drosophila cells. In contrast, dCTCF alone, is unable to cause such a decondensation, however, when CP190 is present, dCTCF recruits it to the lacO array and mediates chromatin unfolding. The CP190 induced opening of chromatin may not be correlated with transcriptional activation, as binding of CP190 does not enhance luciferase activity in reporter assays. We propose that CP190 may mediate histone modification and chromatin remodelling activity to induce an open chromatin state by its direct recruitment or targeting by a DNA binding factor such as dCTCF.


    Institute of Scientific and Technical Information of China (English)

    Zhen-mei Liu; Zhi-kang Xu; Mathias Ulbricht


    Two kinds of polypeptides were tethered onto the surface of polypropylene microporous membrane (PPMM)through a ring opening polymerization of L-glutamate N-carboxyanhydride initiated by amino groups which were introduced by ammonia plasma and γ-aminopropyl triethanoxysilane treatments. X-ray photoelectron spectroscopy (XPS), attenuated total reflectance Fourier transform infrared spectroscopy (FT-IR/ATR), scanning electron microscopy (SEM), together with water contact angle measurements were used to characterize the modified membranes. XPS analyses and FT-IR/ATR spectra demonstrated that polypeptides are actually grafted onto the membrane surface. The wettability of the membrane surface increases at first and then decreases with the increase in grafting degrees of polypeptide. Platelet adhesion and murine macrophage attachment experiments reveal an enhanced hemocompatibility for the polypeptide modified PPMMs. All these results give evidence that polypeptide grafting can simultaneously improve the hemocompatibility as well as reserve the hydrophobicity for the membrane, which will provide a potential approach to improve the performance of polypropylene hollow fiber microporous membrane used in artificial oxygenator.

  20. The Neurorepellent Slit2 Inhibits Postadhesion Stabilization of Monocytes Tethered to Vascular Endothelial Cells. (United States)

    Mukovozov, Ilya; Huang, Yi-Wei; Zhang, Qiuwang; Liu, Guang Ying; Siu, Allan; Sokolskyy, Yaroslav; Patel, Sajedabanu; Hyduk, Sharon J; Kutryk, Michael J B; Cybulsky, Myron I; Robinson, Lisa A


    The secreted neurorepellent Slit2, acting through its transmembrane receptor, Roundabout (Robo)-1, inhibits chemotaxis of varied cell types, including leukocytes, endothelial cells, and vascular smooth muscle cells, toward diverse attractants. The role of Slit2 in regulating the steps involved in recruitment of monocytes in vascular inflammation is not well understood. In this study, we showed that Slit2 inhibited adhesion of monocytic cells to activated human endothelial cells, as well as to immobilized ICAM-1 and VCAM-1. Microfluidic live cell imaging showed that Slit2 inhibited the ability of monocytes tethered to endothelial cells to stabilize their actin-associated anchors and to resist detachment in response to increasing shear forces. Transfection of constitutively active plasmids revealed that Slit2 inhibited postadhesion stabilization of monocytes on endothelial cells by preventing activation of Rac1. We further found that Slit2 inhibited chemotaxis of monocytes toward CXCL12 and CCL2. To determine whether Slit2 and Robo-1 modulate pathologic monocyte recruitment associated with vascular inflammation and cardiovascular disease, we tested PBMC from patients with coronary artery disease. PBMC from these patients had reduced surface levels of Robo-1 compared with healthy age- and sex-matched subjects, and Slit2 failed to inhibit chemotaxis of PBMC of affected patients, but not healthy control subjects, toward CCL2. Furthermore, administration of Slit2 to atherosclerosis-prone LDL receptor-deficient mice inhibited monocyte recruitment to nascent atherosclerotic lesions. These results demonstrate that Slit2 inhibits chemotaxis of monocytes, as well as their ability to stabilize adhesions and resist detachment forces. Slit2 may represent a powerful new tool to inhibit pathologic monocyte recruitment in vascular inflammation and atherosclerosis.

  1. Space demostration of bare electrodynamic tape-tether technology on the sounding rocket S520-25


    Fujii, Hironori; Watanabe, Takeo; Sahara, Hironori; Kojima, Hirohisa; Takehara, Shoichiro; Yamagiwa, Yoshiki; Sasaki, Susumu; Abe, Takumi; Tanaka, Koji; Oyama, Khoichiro; Jhonson, Les; Khazanov, V.; Sanmartín Losada, Juan Ramón; Charro, Mario; Kruijff, Michiel


    A spaceflight validation of bare electro dynamic tape tether technology was conducted. A S520-25 sounding rocket was launched successfully at 05:00am on 31 August 2010 and successfully deployed 132.6m of tape tether over 120 seconds in a ballistic flight. The electrodynamic performance of the bare tape tether employed as an atmospheric probe was measured. Flight results are introduced through the present progressive report of the demonstration and the results of flight experiment are ex...

  2. T-REX: Bare electro-dynamic tape-tether technology experimetn on sounding rocket S520


    Watanabe, Takeo; Fujii, Hironori; Kusagaya, Tairo; Sahara, Hironori; Kojima, Hirohisa; Takehara, Shoichiro; Yamagiwa, Yoshiki; Sasaki, Susumu; Abe, Takumi; Tanaka, Koji; Oyama, Khoichiro; Ebinuma, Takuji; Johson, Les; Khazanov, George; Sanmartín Losada, Juan Ramón


    The project to verify the performance of space tether technology was successfully demonstrated by the launch of the sounding rocket S520 the 25tu. The project is the space demonstration of science and engineering technologies of a bare tape electrodynamic tether (EDT) in the international campaign between Japan, USA, Europe and Australia. Method of "Inverse ORIGAMI (Tape tether folding)" was employed in order to deploy the bare tape EDT in a short period time of the suborbital flight. The ...

  3. Centralized Dynamics and Control of Novel Orbiting Formations of Tethered Spacecraft (United States)

    Quadrelli, Marco B.; Hadaegh, Fred Y.

    acting as leader of the tethered formation. An application of this problem arises when a distributed sensor array formed by a chain of tethered data-gathering vehicles is being commanded to reconfigure from a remote location by the formation leader. Another application is in radar mapping where multiple free-flying vehicles synthesize multiple apertures with the main tethered vehicle for increased coverage. In this way, a centralized control architecture distributes the information flow among the members of the sensor array. Defining an orbiting formation as an ensemble of orbiting spacecraft performing a cooperative task, we point out that, until now, only spacecraft modeled as rigid bodies have been analyzed in the literature of orbiting formations and constellations. After the formation is in place, one may identify what is known as the virtual truss, i.e. the connection between the elements of the formation, which provides structural rigidity on account of the information flow between them. Our problem is different than conventional formation dynamics problems in that the presence of a tethered spacecraft within the formation demands an investigation of the dynamics coupling between spacecraft caused by tether viscoelasticity. The dynamics model takes into account the orbital and spacecraft dynamics of each vehicle. The control architecture features a separated spacecraft, which has visibility to the entire group of tethered vehicles. This vehicle is the leader of the formation, and ensures that the spacecraft on the tether remain connected and move according to a pre-specified program. The control system design consists of a proportional-derivative feedback plus acceleration feedforward. This ensures that modeling errors are compensated appropriately, and that the commanded slew is tracked accurately. The leader is also where the centralized estimator is located. This estimator continuously updates the state of the formation and estimates inter

  4. Ketone isosteres of 2-N-acetamidosugars as substrates for metabolic cell surface engineering

    Energy Technology Data Exchange (ETDEWEB)

    Hang, Howard C.; Bertozzi, Carolyn R.


    Novel chemical reactivity can be engendered on cell surfaces by the metabolic incorporation of unnatural sugars into cell surface glycoconjuagtes. 2-N-Acetamido sugars such as GalNAc and GlcNAc are abundant components of cell surface glycoconjugates, and hence attractive targets for metabolic cell surface engineering. Here we report (1) the synthesis of isosteric analogs bearing a ketone group in place of the N-acetamido group, and (2) evaluation of their metabolic incorporation into mammalian cell surface glycans. A ketone isostere of GalNAc was metabolized by CHO cells through the salvage pathway and delivered to O-linked glycoproteins on the cell surface. Its residence at the core position of O-linked glycans is suggested by studies with a-benzyl GalNAc, an inhibitor of O-linked oligosaccharide extension. A mutant CHO cell line lacking endogenous UDP-GalNAc demonstrated enhanced metabolism of the GalNAc analog, suggesting that competition with native intermediates might limits enzymatic transformation in mammalian cells. A ketone isostere of GlcNAc could not be detected on CHO or human cell surfaces after incubation. Thus, the enzymes in the GlcNAc salvage pathway might be less permissive of unnatural substrates than those comprising the GalNAc salvage pathway. Alternatively, high levels of endogenous GlcNAc derivatives might compete with the ketone isostere and prevent its incorporation into oligosaccharides.

  5. Biogenic nonmethane hydrocarbon emissions estimated from tethered balloon observations (United States)

    Davis, K. J.; Lenschow, D. H.; Zimmerman, P. R.


    A new technique for estimating surface fluxes of trace gases, the mixed-layer gradient technique, is used to calculate isoprene and terpene emissions from forests. The technique is applied to tethered balloon measurements made over the Amazon forest and a pine-oak forest in Alabama at altitudes up to 300 m. The observations were made during the dry season Amazon Boundary Layer Experiment (ABLE 2A) and the Rural Oxidants in the Southern Environment 1990 experiment (ROSE I). Results from large eddy simulations of scalar transport in the clear convective boundary layer are used to infer fluxes from the balloon profiles. Profiles from the Amazon give a mean daytime emission of 3630 +/- 1400 micrograms isoprene sq m/h, where the uncertainty represents the standard deviation of the mean of eight flux estimates. Twenty profiles from Alabama give emissions of 4470 +/- 3300 micrograms isoprene sq m/h, 1740 +/- 1060 micrograms alpha-pinene sq m/h, and 790 +/- 560 micrograms beta-pinene sq m/h, respectively. These results are in agreement with emissions derived from chemical budgets. The emissions may be overestimated because of uncertainty about how to incorporate the effects of the canopy on the mixed-layer gradients. The large variability in these emission estimates is probably due to the relatively short sampling times of the balloon profiles, though spatially heterogeneous emissions may also play a role. Fluxes derived using this technique are representative of an upwind footprint of several kilometers and are independent of hydrocarbon oxidation rate and mean advection.

  6. Phosphatidylserine liposomes can be tethered by caldesmon to actin filaments. (United States)

    Makuch, R; Zasada, A; Mabuchi, K; Krauze, K; Wang, C L; Dabrowska, R


    Rotary shadowing electron microscopy revealed that attachment of caldesmon to phosphatidylserine (PS) liposomes was mainly through its C-terminal end. To determine the PS-binding sites of caldesmon, we have made use of synthetic peptides covering the two C-terminal calmodulin binding sites and a recombinant fragment corresponding to the N-terminal end of the C-terminal domain that contains an amphipathic helix. Interactions of these peptides with the PS liposomes were studied by nondenaturing gel electrophoresis and fluorescence spectroscopy. The results showed that both calmodulin-binding sites of caldesmon were able to interact with PS. The affinity (Kd) of PS for these sites was in the range of 1.8-14.3 x 10(-5) M, compared to 0.69 x 10(-5) M for the whole caldesmon molecule. Fragments located outside of calmodulin-binding sites bound PS weakly (3.85 x 10(-4) M) and thus may contain a second class of lipid-binding sites. Binding of PS induced conformational changes in regions other than the C-terminal PS-binding sites, as evidenced by the changes in the susceptibility to proteolytic cleavages. Most significantly, the presence of caldesmon greatly increased binding of PS to F-actin, suggesting that caldesmon may tether PS liposomes to actin filaments. These results raise the possibility that caldesmon-lipid interactions could play a functionally important role in the assembly of contractile filaments near the membranes. Images FIGURE 2 FIGURE 4 FIGURE 6 PMID:9284327

  7. Template-directed ligation of tethered mononucleotides by t4 DNA ligase for kinase ribozyme selection.

    Directory of Open Access Journals (Sweden)

    David G Nickens

    Full Text Available BACKGROUND: In vitro selection of kinase ribozymes for small molecule metabolites, such as free nucleosides, will require partition systems that discriminate active from inactive RNA species. While nucleic acid catalysis of phosphoryl transfer is well established for phosphorylation of 5' or 2' OH of oligonucleotide substrates, phosphorylation of diffusible small molecules has not been demonstrated. METHODOLOGY/PRINCIPAL FINDINGS: This study demonstrates the ability of T4 DNA ligase to capture RNA strands in which a tethered monodeoxynucleoside has acquired a 5' phosphate. The ligation reaction therefore mimics the partition step of a selection for nucleoside kinase (deoxyribozymes. Ligation with tethered substrates was considerably slower than with nicked, fully duplex DNA, even though the deoxynucleotides at the ligation junction were Watson-Crick base paired in the tethered substrate. Ligation increased markedly when the bridging template strand contained unpaired spacer nucleotides across from the flexible tether, according to the trends: A(2>A(1>A(3>A(4>A(0>A(6>A(8>A(10 and T(2>T(3>T(4>T(6 approximately T(1>T(8>T(10. Bridging T's generally gave higher yield of ligated product than bridging A's. ATP concentrations above 33 microM accumulated adenylated intermediate and decreased yields of the gap-sealed product, likely due to re-adenylation of dissociated enzyme. Under optimized conditions, T4 DNA ligase efficiently (>90% joined a correctly paired, or TratioG wobble-paired, substrate on the 3' side of the ligation junction while discriminating approximately 100-fold against most mispaired substrates. Tethered dC and dG gave the highest ligation rates and yields, followed by tethered deoxyinosine (dI and dT, with the slowest reactions for tethered dA. The same kinetic trends were observed in ligase-mediated capture in complex reaction mixtures with multiple substrates. The "universal" analog 5-nitroindole (dNI did not support ligation when

  8. A preliminary report on the use of laser-Doppler flowmetry during tethered spinal cord release. (United States)

    Schneider, S J; Rosenthal, A D; Greenberg, B M; Danto, J


    Neurological deterioration in the tethered cord syndrome has been postulated to result from a compromise of blood flow in the distal spinal cord. In order to evaluate vascular perfusion in human subjects, a new technique of laser-Doppler flowmetry was used to monitor continuously the microcirculation of the distal spinal cord during surgery for tethered cord release in 10 children. For further comparison, five children undergoing selective dorsal rhizotomy were also monitored. In the tethered cord syndrome group, spinal cord blood flow before untethering was a mean of 12.6 ml/min per 100 g of tissue and increased in all cases after release to a mean of 29.4 ml/min per 100 g of tissue. All patients improved neurologically. The selective dorsal rhizotomy group had a preoperative mean spinal cord blood flow of 30.8 ml/min per 100 g of tissue, which was not altered by the operative procedure. Significant improvement occurs in distal spinal cord blood flow after tethered cord release, which may be representative of an important mechanism in the pathophysiology of the tethered cord syndrome.

  9. Tethered spheroids as an in vitro hepatocyte model for drug safety screening. (United States)

    Xia, Lei; Sakban, Rashidah Binte; Qu, Yinghua; Hong, Xin; Zhang, Wenxia; Nugraha, Bramasta; Tong, Wen Hao; Ananthanarayanan, Abhishek; Zheng, Baixue; Chau, Ian Yin-Yan; Jia, Ruirui; McMillian, Michael; Silva, Jose; Dallas, Shannon; Yu, Hanry


    Hepatocyte spheroids mimic many in vivo liver-tissue phenotypes but increase in size during extended culture which limits their application in drug testing applications. We have developed an improved hepatocyte 3D spheroid model, namely tethered spheroids, on RGD and galactose-conjugated membranes using an optimized hybrid ratio of the two bioactive ligands. Cells in the spheroid configuration maintained 3D morphology and uncompromised differentiated hepatocyte functions (urea and albumin production), while the spheroid bottom was firmly tethered to the substratum maintaining the spheroid size in multi-well plates. The oblate shape of the tethered spheroids, with an average height of 32 μm, ensured efficient nutrient, oxygen and drug access to all the cells within the spheroid structure. Cytochrome P450 induction by prototypical inducers was demonstrated in the tethered spheroids and was comparable or better than that observed with hepatocyte sandwich cultures. These data suggested that tethered 3D hepatocyte spheroids may be an excellent alternative to 2D hepatocyte culture models for drug safety applications.

  10. Electron Emitter for small-size Electrodynamic Space Tether using MEMS Technology

    DEFF Research Database (Denmark)

    Fleron, René A. W.; Blanke, Mogens


    Adjustment of the orbit of a spacecraft using the forces created by an electro-dynamic space-tether has been shown as a theoretic possibility in recent literature. Practical implementation is being pursued for larger scale missions where a hot filament device controls electron emission and the cu......Adjustment of the orbit of a spacecraft using the forces created by an electro-dynamic space-tether has been shown as a theoretic possibility in recent literature. Practical implementation is being pursued for larger scale missions where a hot filament device controls electron emission...... and the current flowing in the electrodynamic space tether. Applications to small spacecraft, or space debris in the 1–10 kg range, possess difficulties with electron emission technology, as low power emitting devices are needed. This paper addresses the system concepts of a small spacecraft electrodynamic tether...... system with focus on electron emitter design and manufacture using micro-electro-mechanical- system (MEMS) technology. The paper addresses the system concepts of a small size electrodynamic tether mission and shows a novel electron emitter for the 1-2 mA range where altitude can be effectively affected...

  11. Myo1c binding to submembrane actin mediates insulin-induced tethering of GLUT4 vesicles. (United States)

    Boguslavsky, Shlomit; Chiu, Tim; Foley, Kevin P; Osorio-Fuentealba, Cesar; Antonescu, Costin N; Bayer, K Ulrich; Bilan, Philip J; Klip, Amira


    GLUT4-containing vesicles cycle between the plasma membrane and intracellular compartments. Insulin promotes GLUT4 exocytosis by regulating GLUT4 vesicle arrival at the cell periphery and its subsequent tethering, docking, and fusion with the plasma membrane. The molecular machinery involved in GLUT4 vesicle tethering is unknown. We show here that Myo1c, an actin-based motor protein that associates with membranes and actin filaments, is required for insulin-induced vesicle tethering in muscle cells. Myo1c was found to associate with both mobile and tethered GLUT4 vesicles and to be required for vesicle capture in the total internal reflection fluorescence (TIRF) zone beneath the plasma membrane. Myo1c knockdown or overexpression of an actin binding-deficient Myo1c mutant abolished insulin-induced vesicle immobilization, increased GLUT4 vesicle velocity in the TIRF zone, and prevented their externalization. Conversely, Myo1c overexpression immobilized GLUT4 vesicles in the TIRF zone and promoted insulin-induced GLUT4 exposure to the extracellular milieu. Myo1c also contributed to insulin-dependent actin filament remodeling. Thus we propose that interaction of vesicular Myo1c with cortical actin filaments is required for insulin-mediated tethering of GLUT4 vesicles and for efficient GLUT4 surface delivery in muscle cells.

  12. Wellcome Prize Lecture. Cell surface, ion-sensing receptors. (United States)

    Riccardi, Daniela


    Changes in extracellular calcium (Ca(2+)o) concentration ([Ca2+]o) affect kidney function both under basal and hormone-stimulated conditions. The molecular identification of an extracellular Ca(2+)-sensing receptor (CaR) has confirmed a direct role of Ca(2+)o on parathyroid and kidney function (i.e. independent of calciotropic hormones) as a modulator of Ca2+ homeostasis. In addition, evidence accumulated over the last 10 years has shown that CaR is also expressed in regions outside the calcium homeostatic system where its role is largely undefined but seems to be linked to regulation of local ionic homeostasis. The parathyroid and kidney CaRs are 1081 and 1079 amino acids long, respectively, and belong to the type III family of G protein-coupled receptors (GPCRs), which includes other CaRs, metabotropic glutamate receptors and putative vomeronasal organ receptors. For the CaR, its low (millimolar) affinity for Ca2+, its positive cooperativity and its large ion-sensing extracellular domain, indicate that the receptor is more sensitive to changes in net cationic charge rather than to a specific ligand. Mg2+, trivalent cations of the lanthanide series and polyvalent cations such as spermine and aminoglycoside antibiotics can all activate the receptor in vitro with EC50 values in the micromolar range for trivalent and polyvalent cations or in the millimolar range for Ca2+ and Mg2+. In addition to true CaR agonists, CaR sensitivity to Ca(2+)o is also susceptible to allosteric modulation by ionic strength, L-amino acids and by pharmacological agents. This review will address endogenous and exogenous CaR agonists, the role of the receptor in the calcium homeostatic system and some speculation on possible role(s) of the CaR in regions not involved in mineral ion homeostasis.

  13. Cell surface receptors for signal transduction and ligand transport: a design principles study.

    Directory of Open Access Journals (Sweden)

    Harish Shankaran


    Full Text Available Receptors constitute the interface of cells to their external environment. These molecules bind specific ligands involved in multiple processes, such as signal transduction and nutrient transport. Although a variety of cell surface receptors undergo endocytosis, the systems-level design principles that govern the evolution of receptor trafficking dynamics are far from fully understood. We have constructed a generalized mathematical model of receptor-ligand binding and internalization to understand how receptor internalization dynamics encodes receptor function and regulation. A given signaling or transport receptor system represents a particular implementation of this module with a specific set of kinetic parameters. Parametric analysis of the response of receptor systems to ligand inputs reveals that receptor systems can be characterized as being: i avidity-controlled where the response control depends primarily on the extracellular ligand capture efficiency, ii consumption-controlled where the ability to internalize surface-bound ligand is the primary control parameter, and iii dual-sensitivity where both the avidity and consumption parameters are important. We show that the transferrin and low-density lipoprotein receptors are avidity-controlled, the vitellogenin receptor is consumption-controlled, and the epidermal growth factor receptor is a dual-sensitivity receptor. Significantly, we show that ligand-induced endocytosis is a mechanism to enhance the accuracy of signaling receptors rather than merely serving to attenuate signaling. Our analysis reveals that the location of a receptor system in the avidity-consumption parameter space can be used to understand both its function and its regulation.

  14. Distinct roles for dystroglycan, beta1 integrin and perlecan in cell surface laminin organization

    DEFF Research Database (Denmark)

    Henry, M D; Satz, J S; Brakebusch, C;


    Dystroglycan (DG) is a cell surface receptor for several extracellular matrix (ECM) molecules including laminins, agrin and perlecan. Recent data indicate that DG function is required for the formation of basement membranes in early development and the organization of laminin on the cell surface....... Here we show that DG-mediated laminin clustering on mouse embryonic stem (ES) cells is a dynamic process in which clusters are consolidated over time into increasingly more complex structures. Utilizing various null-mutant ES cell lines, we define roles for other molecules in this process. In beta1...... integrin-deficient ES cells, laminin-1 binds to the cell surface, but fails to organize into more morphologically complex structures. This result indicates that beta1 integrin function is required after DG function in the cell surface-mediated laminin assembly process. In perlecan-deficient ES cells...

  15. Proteomic inventory of "anchorless" proteins on the colon adenocarcinoma cell surface.

    NARCIS (Netherlands)

    Tjalsma, H.; Pluk, W.J.G.; Heuvel, L.P.W.J. van den; Peters, W.H.M.; Roelofs, R.H.W.M.; Swinkels, D.W.


    Surface proteins play important pathophysiological roles in health and disease, and accumulating proteomics-based studies suggest that several "non-membrane" proteins are sorted to the cell surface by unconventional mechanisms. Importantly, these proteins may comprise attractive therapeutic targets

  16. Isolation of cell surface proteins for mass spectrometry-based proteomics. (United States)

    Elschenbroich, Sarah; Kim, Yunee; Medin, Jeffrey A; Kislinger, Thomas


    Defining the cell surface proteome has profound importance for understanding cell differentiation and cell-cell interactions, as well as numerous pathogenic abnormalities. Owing to their hydrophobic nature, plasma membrane proteins that reside on the cell surface pose analytical challenges and, despite efforts to overcome difficulties, remain under-represented in proteomic studies. Limitations in the classically employed ultracentrifugation-based approaches have led to the invention of more elaborate techniques for the purification of cell surface proteins. Three of these methods--cell surface coating with cationic colloidal silica beads, biotinylation and chemical capture of surface glycoproteins--allow for marked enrichment of this subcellular proteome, with each approach offering unique advantages and characteristics for different experiments. In this article, we introduce the principles of each purification method and discuss applications from the recent literature.

  17. Adsorption of dirhamnolipid on four microorganisms and the effect on cell surface hydrophobicity. (United States)

    Zhong, Hua; Zeng, Guang Ming; Yuan, Xing Zhong; Fu, Hai Yan; Huang, Guo He; Ren, Fang Yi


    In this study, adsorption of dirhamnolipid biosurfactant on a Gram-negative Pseudomonas aeruginosa, two Gram-positive Bacillus subtilis, and a yeast, Candida lipolytica, was investigated, and the causality between the adsorption and change of cell surface hydrophobicity was discussed. The adsorption was not only specific to the microorganisms but also depended on the physiological status of the cells. Components of the biosurfactant with different rhamnosyl number or aliphatic chain length also exhibited slight difference in adsorption manner. The adsorption indeed caused the cell surface hydrophobicity to change regularly; however, the changes depended on both the concentrations of rhamnolipid solutions applied and the adsorbent physiological conditions. Orientation of rhamnolipid monomers on cell surface and micelle deposition are supposed to be the basic means of adsorption to change cell hydrophobicity at low and high rhamnolipid concentrations, respectively. This study proposed the possibility to modify cell surface hydrophobicity with biosurfactant of low concentrations, which may be of importance in in situ soil remediation.

  18. Effect of Direct Electric Current on the Cell Surface Properties of Phenol-Degrading Bacteria


    Luo, Qishi; Wang, Hui; Zhang, Xihui; Qian, Yi


    The change in cell surface properties in the presence of electric currents is of critical concern when the potential to manipulate bacterial movement with electric fields is evaluated. In this study, the effects of different direct electric currents on the cell surface properties involved in bacterial adhesion were investigated by using a mixed phenol-degrading bacterial culture in the exponential growth phase. The traits investigated were surface hydrophobicity (measured by adherence to n-oc...

  19. Exocellular esterase and emulsan release from the cell surface of Acinetobacter calcoaceticus.


    Shabtai, Y; Gutnick, D L


    An esterase activity has been found, both in the cell-free growth medium and on the cell surface of the hydrocarbon-degrading Acinetobacter calcoaceticus RAG-1. The enzyme catalyzed the hydrolysis of acetyl and other acyl groups from triglycerides and aryl and alkyl esters. Emulsan, the extracellular heteropolysaccharide bioemulsifier produced by strain RAG-1, was also a substrate for the enzyme. Gel filtration showed that the cell-free enzyme was released from the cell surface either emulsan...

  20. Cell surface profiling using high-throughput flow cytometry : a platform for biomarker discovery and analysis of cellular heterogeneity

    NARCIS (Netherlands)

    Gedye, Craig A; Hussain, Ali; Paterson, Joshua; Smrke, Alannah; Saini, Harleen; Sirskyj, Danylo; Pereira, Keira; Lobo, Nazleen; Stewart, Jocelyn; Go, Christopher; Ho, Jenny; Medrano, Mauricio; Hyatt, Elzbieta; Yuan, Julie; Lauriault, Stevan; Meyer, Mona; Kondratyev, Maria; van den Beucken, Twan; Jewett, Michael; Dirks, Peter; Guidos, Cynthia J; Danska, Jayne; Wang, Jean; Wouters, Bradly; Neel, Benjamin; Rottapel, Robert; Ailles, Laurie E


    Cell surface proteins have a wide range of biological functions, and are often used as lineage-specific markers. Antibodies that recognize cell surface antigens are widely used as research tools, diagnostic markers, and even therapeutic agents. The ability to obtain broad cell surface protein profil

  1. DMPD: Innate immune sensing of pathogens and danger signals by cell surface Toll-likereceptors. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 17275324 Innate immune sensing of pathogens and danger signals by cell surface Toll... Show Innate immune sensing of pathogens and danger signals by cell surface Toll-likereceptors. PubmedID 172...75324 Title Innate immune sensing of pathogens and danger signals by cell surface

  2. Self-Assembled Array of Tethered Manganese Oxide Nanoparticles for the Next Generation of Energy Storage (United States)

    Stevens, Tyler E.; Pearce, Charles J.; Whitten, Caleah N.; Grant, Richard P.; Monson, Todd C.


    Many challenges must be overcome in order to create reliable electrochemical energy storage devices with not only high energy but also high power densities. Gaps exist in both battery and supercapacitor technologies, with neither one satisfying the need for both large power and energy densities in a single device. To begin addressing these challenges (and others), we report a process to create a self-assembled array of electrochemically active nanoparticles bound directly to a current collector using extremely short (2 nm or less) conductive tethers. The tethered array of nanoparticles, MnO in this case, bound directly to a gold current collector via short conducting linkages eliminates the need for fillers, resulting in a material which achieves 99.9% active material by mass (excluding the current collector). This strategy is expected to be both scalable as well as effective for alternative tethers and metal oxide nanoparticles. PMID:28287183

  3. A perspective from transport protein particle: vesicle tether and human diseases. (United States)

    Li, Chunman; Yu, Sidney


    Vesicle-mediated transport of proteins is a highly regulated, multi-step process. When the vesicle is approaching its target membrane compartment, many factors are required to provide specificity and tethering between the incoming vesicle and the target membrane, before vesicle fusion can occur. Tethering factors, which include multisubunit complexes, coiled-coil proteins, with the help of small GTPases, provide the initial interaction between the vesicle and its target membrane. Of the multisubunit tethering factors, the transport protein particle (TRAPP) complexes function in a number of trafficking steps, including endoplasmic reticulum (ER)-to-Golgi transport, intra- and post-Golgi traffic and autophagosome formation. In this review, we summarize the updated progress in structure and function of TRAPP complexes as well as human diseases caused by genetic mutations in TRAPP.

  4. Electrodynamic-Tether Magnetosphere Interaction From Capture to Low Jovian Orbit of its Spacecraft (United States)

    Sanmartin, J. R.; Charro, M.; Lorenzini, E. C.; Bombardelli, C.; Bramanti, C.


    An orbiting conductive tether provides a dissipative mechanism in planets that have magnetic field and ionosphere/magnetosphere. The Jovian system is a particularly appropriate place for use of an electrodynamic tether because the magnetic field is intense, the stationary orbit is close to the planet, and moon Io provides a dense plasma torus farther away. The interaction of the tether with the magnetized plasma is analyzed under a variety of conditions, since the spacecraft is captured into an equatorial, highly elliptical orbit with perijove inside the stationary orbit, till the spacecraft reaches a low circular orbit around Jupiter, below the radiation belts. The radiation dose accumulated as the apojove distance is reduced through of sequence of perijove passes, is studied.

  5. Sliding tethered ligands add topological interactions to the toolbox of ligand-receptor design (United States)

    Bauer, Martin; Kékicheff, Patrick; Iss, Jean; Fajolles, Christophe; Charitat, Thierry; Daillant, Jean; Marques, Carlos M.


    Adhesion in the biological realm is mediated by specific lock-and-key interactions between ligand-receptor pairs. These complementary moieties are ubiquitously anchored to substrates by tethers that control the interaction range and the mobility of the ligands and receptors, thus tuning the kinetics and strength of the binding events. Here we add sliding anchoring to the toolbox of ligand-receptor design by developing a family of tethered ligands for which the spacer can slide at the anchoring point. Our results show that this additional sliding degree of freedom changes the nature of the adhesive contact by extending the spatial range over which binding may sustain a significant force. By introducing sliding tethered ligands with self-regulating length, this work paves the way for the development of versatile and reusable bio-adhesive substrates with potential applications for drug delivery and tissue engineering.

  6. Quantitative comparison of a human cancer cell surface proteome between interphase and mitosis. (United States)

    Özlü, Nurhan; Qureshi, Mohammad H; Toyoda, Yusuke; Renard, Bernhard Y; Mollaoglu, Gürkan; Özkan, Nazlı E; Bulbul, Selda; Poser, Ina; Timm, Wiebke; Hyman, Anthony A; Mitchison, Timothy J; Steen, Judith A


    The cell surface is the cellular compartment responsible for communication with the environment. The interior of mammalian cells undergoes dramatic reorganization when cells enter mitosis. These changes are triggered by activation of the CDK1 kinase and have been studied extensively. In contrast, very little is known of the cell surface changes during cell division. We undertook a quantitative proteomic comparison of cell surface-exposed proteins in human cancer cells that were tightly synchronized in mitosis or interphase. Six hundred and twenty-eight surface and surface-associated proteins in HeLa cells were identified; of these, 27 were significantly enriched at the cell surface in mitosis and 37 in interphase. Using imaging techniques, we confirmed the mitosis-selective cell surface localization of protocadherin PCDH7, a member of a family with anti-adhesive roles in embryos. We show that PCDH7 is required for development of full mitotic rounding pressure at the onset of mitosis. Our analysis provided basic information on how cell cycle progression affects the cell surface. It also provides potential pharmacodynamic biomarkers for anti-mitotic cancer chemotherapy.

  7. Differential regulation of synaptic vesicle tethering and docking by UNC-18 and TOM-1

    Directory of Open Access Journals (Sweden)

    Elena O Gracheva


    Full Text Available The assembly of SNARE complexes between syntaxin, SNAP-25 and synaptobrevin is required to prime synaptic vesicles for fusion. Since Munc18 and tomosyn compete for syntaxin interactions, the interplay between these proteins is predicted to be important in regulating synaptic transmission. We explored this possibility, by examining genetic interactions between C. elegans unc-18(Munc18, unc-64(syntaxin and tom-1(tomosyn. We have previously demonstrated that unc-18 mutants have reduced synaptic transmission, whereas tom-1 mutants exhibit enhanced release. Here we show that the unc-18 mutant release defect is associated with loss of two morphologically distinct vesicle pools; those tethered within 25nm of the plasma membrane and those docked with the plasma membrane. In contrast, priming defective unc-13 mutants accumulate tethered vesicles, while docked vesicles are greatly reduced, indicating tethering is UNC-18-dependent and occurs in the absence of priming. C. elegans unc-64 mutants phenocopy unc-18 mutants, losing both tethered and docked vesicles, whereas overexpression of open syntaxin preferentially increases vesicle docking, suggesting UNC-18/closed syntaxin interactions are responsible for vesicle tethering. Given the competition between vertebrate tomosyn and Munc18, for syntaxin binding, we hypothesized that C. elegans TOM-1 may inhibit both UNC-18-dependent vesicle targeting steps. Consistent with this hypothesis, tom-1 mutants exhibit enhanced UNC-18 plasma membrane localization and a concomitant increase in both tethered and docked synaptic vesicles. Furthermore, in tom-1;unc-18 double mutants the docked, primed vesicle pool is preferentially rescued relative to unc-18 single mutants. Together these data provide evidence for the differential regulation of two vesicle targeting steps by UNC-18 and TOM-1 through competitive interactions with syntaxin

  8. Conserved TCP domain of Sas-4/CPAP is essential for pericentriolar material tethering during centrosome biogenesis. (United States)

    Zheng, Xiangdong; Gooi, Li Ming; Wason, Arpit; Gabriel, Elke; Mehrjardi, Narges Zare; Yang, Qian; Zhang, Xingrun; Debec, Alain; Basiri, Marcus L; Avidor-Reiss, Tomer; Pozniakovsky, Andrei; Poser, Ina; Saric, Tomo; Hyman, Anthony A; Li, Haitao; Gopalakrishnan, Jay


    Pericentriolar material (PCM) recruitment to centrioles forms a key step in centrosome biogenesis. Deregulation of this process leads to centrosome aberrations causing disorders, one of which is autosomal recessive primary microcephaly (MCPH), a neurodevelopmental disorder where brain size is reduced. During PCM recruitment, the conserved centrosomal protein Sas-4/CPAP/MCPH6, known to play a role in centriole formation, acts as a scaffold for cytoplasmic PCM complexes to bind and then tethers them to centrioles to form functional centrosomes. To understand Sas-4's tethering role, we determined the crystal structure of its T complex protein 10 (TCP) domain displaying a solvent-exposed single-layer of β-sheets fold. This unique feature of the TCP domain suggests that it could provide an "extended surface-like" platform to tether the Sas-4-PCM scaffold to a centriole. Functional studies in Drosophila, human cells, and human induced pluripotent stem cell-derived neural progenitor cells were used to test this hypothesis, where point mutations within the 9-10th β-strands (β9-10 mutants including a MCPH-associated mutation) perturbed PCM tethering while allowing Sas-4/CPAP to scaffold cytoplasmic PCM complexes. Specifically, the Sas-4 β9-10 mutants displayed perturbed interactions with Ana2, a centrosome duplication factor, and Bld-10, a centriole microtubule-binding protein, suggesting a role for the β9-10 surface in mediating protein-protein interactions for efficient Sas-4-PCM scaffold centriole tethering. Hence, we provide possible insights into how centrosomal protein defects result in human MCPH and how Sas-4 proteins act as a vehicle to tether PCM complexes to centrioles independent of its well-known role in centriole duplication.

  9. Cell patterning on polylactic acid through surface-tethered oligonucleotides. (United States)

    Matsui, Toshiki; Arima, Yusuke; Takemoto, Naohiro; Iwata, Hiroo


    Polylactic acid (PLA) is a candidate material to prepare scaffolds for 3-D tissue regeneration. However, cells do not adhere or proliferate well on the surface of PLA because it is hydrophobic. We report a simple and rapid method for inducing cell adhesion to PLA through DNA hybridization. Single-stranded DNA (ssDNA) conjugated to poly(ethylene glycol) (PEG) and to a terminal phospholipid (ssDNA-PEG-lipid) was used for cell surface modification. Through DNA hybridization, modified cells were able to attach to PLA surfaces modified with complementary sequence (ssDNA'). Different cell types can be attached to PLA fibers and films in a spatially controlled manner by using ssDNAs with different sequences. In addition, they proliferate well in a culture medium supplemented with fetal bovine serum. The coexisting modes of cell adhesion through DNA hybridization and natural cytoskeletal adhesion machinery revealed no serious effects on cell growth. The combination of a 3-D scaffold made of PLA and cell immobilization on the PLA scaffold through DNA hybridization will be useful for the preparation of 3-D tissue and organs.

  10. Drop Tower tests in preparation of a Tethered Electromagnetic Docking space demonstration (United States)

    Olivieri, Lorenzo; Francesconi, Alessandro; Antonello, Andrea; Bettiol, Laura; Branz, Francesco; Duzzi, Matteo; Mantellato, Riccardo; Sansone, Francesco; Savioli, Livia


    A group of students of the University of Padova is recently developing some technologies to implement a Tethered Electromagnetic Docking (TED) experiment, a novel system for close rendezvous and mating manoeuvres between two spacecraft, consisting in a small tethered probe ejected by the chaser and magnetically guided by a receiving electromagnet mounted on the target. Because of the generated magnetic field, automatic self-alignment and mating are possible; then, as the tether is rewinded, the chaser is able to dock with the target. This concept allows to simplify standard docking procedures, thanks to the reduction of proximity navigation and guidance requirements, as well as consequent fuel reduction. Other interesting applications are expected, from active debris removal to space tugging; in particular, the utilization of the tethered connection for detumbling operations is considered. The realization of a space demonstrator requires a preliminary verification of the critical technologies employed in TED, in particular the magnetic guidance and the probe deploy and retrieve; in the framework of ESA "Drop your Thesis!" 2014 and 2016 campaigns the experiments FELDs (Flexible Electromagnetic Leash Docking system) and STAR (System for Tether Automatic Retrieval) have been focused on the test of such critical elements in the relevant microgravity environment of ZARM Drop Tower in Bremen. In particular, FELDs consisted in a simplified model of TED with a magnetic target interface, a passive tethered probe and its launch system: the experiment allowed to assess the passive self-alignment of the probe with respect to the target and to study the effect of friction between the tether and the release system. Similarly, STAR is investigating the tether actively controlled deployment and retrieval, with the experiment campaign planned on November 2016. In addition, another microgravity experiment is in preparation for the investigation of active magnetic navigation: PACMAN

  11. Towing Asteroids with Gravity Tractors Enhanced by Tethers and Solar Sails (United States)

    Shen, Haijun; Roithmayr, Carlos M.


    Material collected from an asteroid's surface can be used to increase gravitational attraction between the asteroid and a Gravity Tractor (GT); the spacecraft therefore operates more effectively and is referred to as an Enhanced Gravity Tractor (EGT). The use of tethers and solar sails to further improve effectiveness and simplify operations is investigated. By employing a tether, the asteroidal material can be placed close to the asteroid while the spacecraft is stationed farther away, resulting in a better safety margin and improved thruster efficiency. A solar sail on a spacecraft can naturally provide radial offset and inter-spacecraft separation required for multiple EGTs.

  12. USGS tethered ACP platforms: New design means more safety and accuracy (United States)

    Morlock, S.E.; Stewart, J.A.; Rehmel, M.S.


    The US Geological Survey has developed an innovative tethered platform that supports an Acoustic Current Profiler (ACP) in making stream-flow measurements (use of the term ACP in this article refers to a class of instruments and not a specific brand name or model). The tethered platform reduces the hazards involved in conventional methods of stream-flow measurement. The use of the platform reduces or eliminates time spent by personnel in streams and boats or on bridges and cableway and stream-flow measurement accuracy is increased.

  13. System Dynamics and Feedforward Control for Tether-Net Space Robot System


    Bin Liang; Yue Qiu; Guang Zhai; Cheng Li


    A new concept using flexible tether-net system to capture space debris is presented in this paper. With a mass point assumption the tether-net system dynamic model is established in orbital frame by applying Lagrange Equations. In order to investigate the net in-plane trajectories during after cast, the non-control R-bar and V-bar captures are simulated with ignoring the out-of-plane libration, the effect of in-plane libration on the trajectories of the capture net is demonstrated by simulati...

  14. Calculating the electromagnetic field on the earth due to an electrodynamic tethered system in the ionosphere (United States)

    Estes, Robert D.


    A method is presented for calculating the electromagnetic wave field on the earth's surface associated with the operation of an electrodynamic tethered satellite system of constant or slowly varying current in the upper ionosphere. The wave field at the ionospheric boundary and on the earth's surface is obtained by numerical integration. The results suggest that the ionospheric waves do not propagate into the atmosphere and that the image of the Alfven wings from a steady-current tether should be greatly broadened on the earth's surface.

  15. Wall effect on fluid-structure interactions of a tethered bluff body (United States)

    Sharma, Sumant; Raghav, Vrishank; Komerath, Narayanan; Smith, Marilyn


    Wind tunnel experiments have shown an unexplained amplification of the free motion of a tethered bluff body in a small wind tunnel relative to that in a large wind tunnel. The influence of wall proximity on fluid-structure interaction is explored using a compound pendulum motion in the plane orthogonal to a steady freestream with a doublet model for aerodynamic forces. Wall proximity amplifies a purely symmetric single degree of freedom oscillation with the addition of an out-of-phase force. The success of this simple level of simulation enables progress to develop metrics for unsteady wall interference in dynamic testing of tethered bluff bodies.

  16. Control by damping Injection of Electrodynamic Tether System in an Inclined Orbit

    DEFF Research Database (Denmark)

    Larsen, Martin Birkelund; Blanke, Mogens


    Control of a satellite system with an electrodynamic tether as actuator is a time-periodic and underactuated control problem. This paper considers the tethered satellite in a Hamiltonian framework and determines a port-controlled Hamiltonian formulation that adequately describes the nonlinear...... dynamical system. Based on this model, a nonlinear controller is designed that will make the system asymptotically stable around its open-loop equilibrium. The control scheme handles the time-varying nature of the system in a suitable manner resulting in a large operational region. The performance...

  17. GRP78 and Cripto Form a Complex at the Cell Surface and Collaborate To Inhibit Transforming Growth Factor β Signaling and Enhance Cell Growth▿ (United States)

    Shani, Gidi; Fischer, Wolfgang H.; Justice, Nicholas J.; Kelber, Jonathan A.; Vale, Wylie; Gray, Peter C.


    Cripto is a multifunctional cell surface protein with important roles in vertebrate embryogenesis and the progression of human tumors. While Cripto has been shown to modulate multiple signaling pathways, its binding partners do not appear to fully explain its molecular actions. Therefore, we conducted a screen aimed at identifying novel Cripto-interacting proteins. This screen led to our identification of glucose-regulated protein 78 (GRP78), an endoplasmic reticulum (ER) chaperone that is also expressed at the surfaces of tumor cells. Here we demonstrate that Cripto and GRP78 interact at the cell surfaces of multiple cell lines and that their interaction is independent of prior association within the ER. Interestingly, short hairpin RNA knockdown of endogenous GRP78 resulted in enhanced transforming growth factor β (TGF-β) signaling, indicating that like Cripto, GRP78 inhibits this pathway. We further show that when coexpressed, GRP78 and Cripto collaborate to antagonize TGF-β responses, including Smad phosphorylation and growth inhibition of prostate cancer cells grown under anchorage-dependent or -independent conditions. Finally, we provide evidence that cells coexpressing GRP78 and Cripto grow much more rapidly in soft agar than do cells expressing either protein individually. Together, our results indicate that these proteins bind at the cell surface to enhance tumor growth via the inhibition of TGF-β signaling. PMID:17991893

  18. Role of Site-Specific N-Glycans Expressed on GluA2 in the Regulation of Cell Surface Expression of AMPA-Type Glutamate Receptors. (United States)

    Takeuchi, Yusuke; Morise, Jyoji; Morita, Ippei; Takematsu, Hiromu; Oka, Shogo


    The AMPA-type glutamate receptor (AMPAR), which is a tetrameric complex composed of four subunits (GluA1-4) with several combinations, mediates the majority of rapid excitatory synaptic transmissions in the nervous system. Cell surface expression levels of AMPAR modulate synaptic plasticity, which is considered one of the molecular bases for learning and memory formation. To date, a unique trisaccharide (HSO3-3GlcAβ1-3Galβ1-4GlcNAc), human natural killer-1 (HNK-1) carbohydrate, was found expressed specifically on N-linked glycans of GluA2 and regulated the cell surface expression of AMPAR and the spine maturation process. However, evidence that the HNK-1 epitope on N-glycans of GluA2 directly affects these phenomena is lacking. Moreover, it is thought that other N-glycans on GluA2 also have potential roles in the regulation of AMPAR functions. In the present study, using a series of mutants lacking potential N-glycosylation sites (N256, N370, N406, and N413) within GluA2, we demonstrated that the mutant lacking the N-glycan at N370 strongly suppressed the intracellular trafficking of GluA2 from the endoplasmic reticulum (ER) in HEK293 cells. Cell surface expression of GluA1, which is a major subunit of AMPAR in neurons, was also suppressed by co-expression of the GluA2 N370S mutant. The N370S mutant and wild-type GluA2 were co-immunoprecipitated with GluA1, suggesting that N370S was properly associated with GluA1. Moreover, we found that N413 was the main potential site of the HNK-1 epitope that promoted the interaction of GluA2 with N-cadherin, resulting in enhanced cell surface expression of GluA2. The HNK-1 epitope on N-glycan at the N413 of GluA2 was also involved in the cell surface expression of GluA1. Thus, our data suggested that site-specific N-glycans on GluA2 regulate the intracellular trafficking and cell surface expression of AMPAR.

  19. Role of Site-Specific N-Glycans Expressed on GluA2 in the Regulation of Cell Surface Expression of AMPA-Type Glutamate Receptors.

    Directory of Open Access Journals (Sweden)

    Yusuke Takeuchi

    Full Text Available The AMPA-type glutamate receptor (AMPAR, which is a tetrameric complex composed of four subunits (GluA1-4 with several combinations, mediates the majority of rapid excitatory synaptic transmissions in the nervous system. Cell surface expression levels of AMPAR modulate synaptic plasticity, which is considered one of the molecular bases for learning and memory formation. To date, a unique trisaccharide (HSO3-3GlcAβ1-3Galβ1-4GlcNAc, human natural killer-1 (HNK-1 carbohydrate, was found expressed specifically on N-linked glycans of GluA2 and regulated the cell surface expression of AMPAR and the spine maturation process. However, evidence that the HNK-1 epitope on N-glycans of GluA2 directly affects these phenomena is lacking. Moreover, it is thought that other N-glycans on GluA2 also have potential roles in the regulation of AMPAR functions. In the present study, using a series of mutants lacking potential N-glycosylation sites (N256, N370, N406, and N413 within GluA2, we demonstrated that the mutant lacking the N-glycan at N370 strongly suppressed the intracellular trafficking of GluA2 from the endoplasmic reticulum (ER in HEK293 cells. Cell surface expression of GluA1, which is a major subunit of AMPAR in neurons, was also suppressed by co-expression of the GluA2 N370S mutant. The N370S mutant and wild-type GluA2 were co-immunoprecipitated with GluA1, suggesting that N370S was properly associated with GluA1. Moreover, we found that N413 was the main potential site of the HNK-1 epitope that promoted the interaction of GluA2 with N-cadherin, resulting in enhanced cell surface expression of GluA2. The HNK-1 epitope on N-glycan at the N413 of GluA2 was also involved in the cell surface expression of GluA1. Thus, our data suggested that site-specific N-glycans on GluA2 regulate the intracellular trafficking and cell surface expression of AMPAR.

  20. Alteration in cell surface properties of Burkholderia spp. during surfactant-aided biodegradation of petroleum hydrocarbons

    Energy Technology Data Exchange (ETDEWEB)

    Mohanty, Sagarika; Mukherji, Suparna [Indian Institute of Technology Bombay, Mumbai (India). Centre for Environmental Science and Engineering (CESE)


    Chemical surfactants may impact microbial cell surface properties, i.e., cell surface hydrophobicity (CSH) and cell surface charge, and may thus affect the uptake of components from non-aqueous phase liquids (NAPLs). This work explored the impact of Triton X-100, Igepal CA 630, and Tween 80 (at twice the critical micelle concentration, CMC) on the cell surface characteristics of Burkholderia cultures, Burkholderia cepacia (ES1, aliphatic degrader) and Burkholderia multivorans (NG1, aromatic degrader), when grown on a six-component model NAPL. In the presence of Triton X-100, NAPL biodegradation was enhanced from 21% to 60% in B. cepacia and from 18% to 53% in B. multivorans. CSH based on water contact angle (50-52 ) was in the same range for both strains while zeta potential at neutral pH was -38 and -31 mV for B. cepacia and B. multivorans, respectively. In the presence of Triton X-100, their CSH increased to greater than 75 and the zeta potential decreased. This induced a change in the mode of uptake and initiated aliphatic hydrocarbon degradation by B. multivorans and increased the rate of aliphatic hydrocarbon degradation in B. cepacia. Igepal CA 630 and Tween 80 also altered the cell surface properties. For B. cepacia grown in the presence of Triton X-100 at two and five times its CMC, CSH increased significantly in the log growth phase. Growth in the presence of the chemical surfactants also affected the abundance of chemical functional groups on the cell surface. Cell surface changes had maximum impact on NAPL degradation in the presence of emulsifying surfactants, Triton X-100 and Igepal CA630.

  1. Different roles of cell surface and exogenous glycosaminoglycans in controlling gene delivery by arginine-rich peptides with varied distribution of arginines. (United States)

    Naik, Rangeetha J; Chatterjee, Anindo; Ganguli, Munia


    The role of cell surface and exogenous glycosaminoglycans (GAGs) in DNA delivery by cationic peptides is controlled to a large extent by the peptide chemistry and the nature of its complex with DNA. We have previously shown that complexes formed by arginine homopeptides with DNA adopt a GAG-independent cellular internalization mechanism and show enhanced gene delivery in presence of exogenous GAGs. In contrast, lysine complexes gain cellular entry primarily by a GAG-dependent pathway and are destabilized by exogenous GAGs. The aim of the current study was to elucidate the factors governing the role of cell surface and soluble glycosaminoglycans in DNA delivery by sequences of arginine-rich peptides with altered arginine distributions (compared to homopeptide). Using peptides with clustered arginines which constitute known heparin-binding motifs and a control peptide with arginines alternating with alanines, we show that complexes formed by these peptides do not require cell surface GAGs for cellular uptake and DNA delivery. However, the charge distribution and the spacing of arginine residues affects DNA delivery efficiency of these peptides in presence of soluble GAGs, since these peptides show only a marginal increase in transfection in presence of exogenous GAGs unlike that observed with arginine homopeptides. Our results indicate that presence of arginine by itself drives these peptides to a cell surface GAG-independent route of entry to efficiently deliver functional DNA into cells in vitro. However, the inherent stability of the complexes differ when the distribution of arginines in the peptides is altered, thereby modulating its interaction with exogenous GAGs.

  2. Self-assembled vesicles of urea-tethered foldamers as hydrophobic drug carriers. (United States)

    Ingole, Tukaram S; Kale, Sangram S; Santhosh Babu, Sukumaran; Sanjayan, Gangadhar J


    Molecular self-assembly of nonamphiphilic α,β-hybrid foldamers based on urea-tethered anthranilic acid-proline (Ant-Pro) foldamers is reported. These self-assembled hollow vesicular architectures can take up and release the anticancer hydrophobic drug curcumin.

  3. Folding of multidomain proteins: biophysical consequences of tethering even in apparently independent folding. (United States)

    Arviv, Oshrit; Levy, Yaakov


    Most eukaryotic and a substantial fraction of prokaryotic proteins are composed of more than one domain. The tethering of these evolutionary, structural, and functional units raises, among others, questions regarding the folding process of conjugated domains. Studying the folding of multidomain proteins in silico enables one to identify and isolate the tethering-induced biophysical determinants that govern crosstalks generated between neighboring domains. For this purpose, we carried out coarse-grained and atomistic molecular dynamics simulations of two two-domain constructs from the immunoglobulin-like β-sandwich fold. Each of these was experimentally shown to behave as the "sum of its parts," that is, the thermodynamic and kinetic folding behavior of the constituent domains of these constructs seems to occur independently, with the folding of each domain uncoupled from the folding of its partner in the two-domain construct. We show that the properties of the individual domains can be significantly affected by conjugation to another domain. The tethering may be accompanied by stabilizing as well as destabilizing factors whose magnitude depends on the size of the interface, the length, and the flexibility of the linker, and the relative stability of the domains. Accordingly, the folding of a multidomain protein should not be viewed as the sum of the folding patterns of each of its parts, but rather, it involves abrogating several effects that lead to this outcome. An imbalance between these effects may result in either stabilization or destabilization owing to the tethering.

  4. Applications of a dynamic tethering system to enable the deep space cam jointed observation bot (United States)

    Leake, Skye; McGuire, Thomas; Parsons, Michael; Hirsch, Michael P.; Straub, Jeremy


    A device capable of creating tethers for use with spacecraft that are made from a diverse material palette could serve many functions. These functions include supporting applications such as data transfer, power generation, and resource collection. Applications that are currently being considered include use in a system for orientation, data transfer, and power delivery and use as part of a free-moving camera system which would be used in proximity to a spacecraft for capturing images and video for promotional and preforming diagnostic and "self-check" operations. Materials that have been considered for use in such a tethering device have different physical attributes in order to facilitate supporting the widest possible degree of applications for use in scientific, remote sensing, power generation, and electromagnetic applications methods for the parent spacecraft. Physical properties that have been considered include: rigidity, conductivity, heat dissipation, and opacity. The proposed dynamic tethering system would be driven by 3D printing technologies. This prospective application of 3D printing remains relatively unexplored. This provides great opportunities for knowledge expansion and the development of dynamic tethers for use capturing video footage and pictures, and for other scientific endeavors.

  5. New amidines from intramolecular cyclization in triflic acid of nitroketene aminals with a tethered phenyl ring

    Indian Academy of Sciences (India)

    Soro Yaya; Bamba Fanté; Siaka Sorho; Coustard Jean-Marie; Adima A Augustin


    Nitroketene aminals with a tethered phenyl group underwent an intramolecular cyclization in trifluoromethanesulfonic acid to afford the corresponding N-(3-ethyl-hydrohydroxyiminobenzocycloalkenylidene) methylamine trifluoromethanesulfonate. The yields were fair to good excepted for the starting compound 1-[N-ethyl-N-(2-phenylethyl)amino]-1-methylamino-2-nitroethene.

  6. Progressive kyphoscoliosis associated with tethered cord treated by posterior vertebral column resection: a case report. (United States)

    Matsumoto, Morio; Watanabe, Kota; Tsuji, Takashi; Ishii, Ken; Takaishi, Hironari; Nakamura, Masaya; Toyama, Yoshiaki; Chiba, Kazuhiro


    STUDY DESIGN.: A case report. OBJECTIVES.: To report a case of progressive kyphoscoliosis associated with a tethered cord that was corrected by posterior vertebral column resection after complicated untethering surgery. SUMMARY OF BACKGROUND DATA.: There have been few clinical reports on posterior vertebral column resection conducted for severe deformity associated with a tethered cord. METHODS.: A patient with progressive kyphoscoliosis associated with a tethered cord first underwent untethering surgery, resulting in neurologic deterioration. Posterior vertebral column resection was performed to correct the kyphoscoliosis while shortening the spinal column to prevent the spinal cord from stretch injury. RESULTS.: Good correction of kyphoscoliosis was obtained without further neurologic deterioration. The Cobb angles of scoliosis was 103 degrees before surgery and 25 degrees after surgery (correction rate; 75.7%), and that of kyphosis was 90 degrees and 36 degrees , respectively (correction rate; 60.0%). CONCLUSION.: Correction of progressive kyphoscoliosis associated with a tethered cord can be achieved successfully by posterior vertebral column resection even after complicated untethering surgery.

  7. Study of the triple-mass Tethered Satellite System under aerodynamic drag and J2 perturbations (United States)

    Razzaghi, Pourya; Assadian, Nima


    The dynamics of multi-tethered satellite formations consisting of three masses are studied in this paper. The triple-mass triple-tethered satellite system is modeled under the low Earth orbit perturbations of drag and Earth's oblateness and its equilibrium conditions are derived. It is modeled as three equal end-masses connected by a uniform-mass straight tether. The lengths of tethers are supposed to be constant and in this manner the angles of the plane consisting the masses are taken as the state variables of the system. The governing equations of motion are derived using Lagrangian approach. The aerodynamic drag perturbation is expressed as an external non-conservative force and the Earth oblateness (J2 perturbation) is considered as a term of potential energy. The equilibrium conditions of this system are found and their stability is investigated through the linear stability theory. Then, the results are verified by using a nonlinear simulation for three types of equilibrium conditions.

  8. Molecular Iodine-Mediated Cyclization of Tethered Heteroatom-Containing Alkenyl or Alkynyl Systems

    Directory of Open Access Journals (Sweden)

    Malose Jack Mphahlele


    Full Text Available Molecular iodine has established itself as a readily available and easy-to-handle electrophilic and oxidizing reagent used in various organic transformations. In this review attention is focused on the use of molecular iodine in promoting cyclization (iodocyclization and cyclodehydroiodination of tethered heteroatom-containing alkenyl or alkynyl systems.

  9. Bare-tether cathodic contact through thermionic emission by low-work-function materials

    Energy Technology Data Exchange (ETDEWEB)

    Chen Xin; Sanmartin, J. R. [Departamento de Fisica Aplicada, Escuela Tecnica Superior de Ingenieros Aeronauticos, Universidad Politecnica de Madrid, Plaza Cardenal Cisneros 3, 28040 Madrid (Spain)


    A new material, C12A7:e{sup -} electride, which might present a work function as low as 0.6 eV and moderately high temperature stability, was recently proposed as coating for floating bare tethers. Arising from heating under space operation, current is emitted by thermionic emission along a thus coated cathodic segment. A preliminary study on the space-charge-limited (SCL) double layer in front of the cathodic segment is presented using Langmuir's SCL electron current between cylindrical electrodes and orbital-motion-limited ion-collection sheath. A detailed calculation of current and bias profiles along the entire tether length is carried out with ohmic effects and the transition from SCL to full Richardson-Dushman emission included. Analysis shows that in the simplest drag mode, under typical orbital and tether conditions, thermionic emission leads to a short cathodic section and may eliminate the need for an active cathodic device and its corresponding gas feed requirements and power subsystem, which results in a truly 'propellant-less' tether system for such basic applications as de-orbiting low earth orbit satellites.

  10. Optimal Cross-Wind Towing and Power Generation with Tethered Kites

    NARCIS (Netherlands)

    Williams, P.; Lansdorp, B.; Ockels, W.


    Non-powered flight vehicles such as kites can provide a means of transmitting wind energy from higher altitudes to the ground via tethers. Although there have been many proposals for systems to extract wind energy from higher altitudes, this paper focuses on the use of a light lifting body at the en

  11. Titanium-tethered vancomycin prevents resistance to rifampicin in Staphylococcus aureus in vitro.

    Directory of Open Access Journals (Sweden)

    Martin Rottman

    Full Text Available Rifampicin is currently recognized as the most potent drug against Gram positive implant related infections. The use of rifampicin is limited by the emergence of bacterial resistance, which is often managed by coadministration of a second antibiotic. The purpose of this study was to determine the effectiveness of soluble rifampicin in combination with vancomycin tethered to titanium metal as a means to control bacterial growth and resistance in vitro. Bacterial growth was inhibited when the vancomycin-tethered titanium discs were treated with Staphylococcus aureus inocula of ≤2×10⁶ CFU, however inocula greater than 2×10⁶ CFU/disc adhered and survived. The combination of surface-tethered vancomycin with soluble rifampicin enhanced the inhibitory effect of rifampicin for an inoculum of 10⁶ CFU/cm² by one dilution (combination MIC of 0.008 mg/L versus 0.015 mg/L for rifampicin alone. Moreover, surface tethered vancomycin prevented the emergence of a rifampicin resistant population in an inoculum of 2×10⁸ CFU.

  12. Growth Factor Tethering to Protein Nanoparticles via Coiled-Coil Formation for Targeted Drug Delivery. (United States)

    Assal, Yasmine; Mizuguchi, Yoshinori; Mie, Masayasu; Kobatake, Eiry


    Protein-based nanoparticles are attractive carriers for drug delivery because they are biodegradable and can be genetically designed. Moreover, modification of protein-based nanoparticles with cell-specific ligands allows for active targeting abilities. Previously, we developed protein nanoparticles comprising genetically engineered elastin-like polypeptides (ELPs) with fused polyaspartic acid tails (ELP-D). Epidermal growth factor (EGF) was displayed on the surface of the ELP-D nanoparticles via genetic design to allow for active cell-targeting abilities. Herein, we focused on the coiled-coil structural motif as a means for noncovalent tethering of growth factor to ELP-D. Specifically, two peptides known to form a heterodimer via a coiled-coil structural motif were fused to ELP-D and single-chain vascular endothelial growth factor (scVEGF121), to facilitate noncovalent tethering upon formation of the heterodimer coiled-coil structure. Drug-loaded growth factor-tethered ELP-Ds were found to be effective against cancer cells by provoking cell apoptosis. These results demonstrate that tethering growth factor to protein nanoparticles through coiled-coil formation yields a promising biomaterial candidate for targeted drug delivery.

  13. Summary of Data Analysis of the YES2 Tethered SpaceMail Experiment

    NARCIS (Netherlands)

    Kruijff, M.; Van der Heide, E.J.; Ockels, W.J.; Gill, E.K.A.


    The 2nd Young Engineers' Satellite (YES2) is a 36 kg student-built experiment that piggybacked on the Foton-M3 microgravity platform in September 2007. YES2 intended to demonstrate tethered SpaceMail, a concept for frequent sample return originally proposed for the International Space Station (ISS).

  14. Investigation of the Cell Surface Proteome of Human Periodontal Ligament Stem Cells

    Directory of Open Access Journals (Sweden)

    Jimin Xiong


    Full Text Available The present study examined the cell surface proteome of human periodontal ligament stem cells (PDLSC compared to human fibroblasts. Cell surface proteins were prelabelled with CyDye before processing to extract the membrane lysates, which were separated using 2D electrophoresis. Selected differentially expressed protein “spots” were identified using Mass spectrometry. Four proteins were selected for validation: CD73, CD90, Annexin A2, and sphingosine kinase 1 previously associated with mesenchymal stem cells. Flow cytometric analysis found that CD73 and CD90 were highly expressed by human PDLSC and gingival fibroblasts but not by keratinocytes, indicating that these antigens could be used as potential markers for distinguishing between mesenchymal cells and epithelial cell populations. Annexin A2 was also found to be expressed at low copy number on the cell surface of human PDLSC and gingival fibroblasts, while human keratinocytes lacked any cell surface expression of Annexin A2. In contrast, sphingosine kinase 1 expression was detected in all the cell types examined using immunocytochemical analysis. These proteomic studies form the foundation to further define the cell surface protein expression profile of PDLSC in order to better characterise this cell population and help develop novel strategies for the purification of this stem cell population.

  15. Visualization of sialic acid produced on bacterial cell surfaces by lectin staining. (United States)

    Kajiwara, Hitomi; Toda, Munetoyo; Mine, Toshiki; Nakada, Hiroshi; Wariishi, Hiroyuki; Yamamoto, Takeshi


    Oligosaccharides containing N-acetylneuraminic acid on the cell surface of some pathogenic bacteria are important for host-microbe interactions. N-acetylneuraminic acid (Neu5Ac) plays a major role in the pathogenicity of bacterial pathogens. For example, cell surface sialyloligosaccharide moieties of the human pathogen Haemophilus influenzae are involved in virulence and adhesion to host cells. In this study, we have established a method of visualizing Neu5Ac linked to a glycoconjugate on the bacterial cell surface based on lectin staining. Photobacterium damselae strain JT0160, known to produce a-2,6-sialyltransferase, was revealed to possess Neu5Ac by HPLC. Using the strain, a strong Sambucus sieboldiana lectin-binding signal was detected. The bacteria producing α-2,6-sialyltransferases could be divided into two groups: those with a lot of α-2,6-linked Neu5Ac on the cell surface and those with a little. In the present study, we developed a useful method for evaluating the relationship between Neu5Ac expression on the cell surface and the degree of virulence of marine bacteria.

  16. Cell surface hydrophobicity and charge of Staphylococcus aureus and coagulase-negative staphylococci from bovine mastitis. (United States)

    Mamo, W; Rozgonyi, F; Brown, A; Hjertén, S; Wadström, T


    The effects of seven growth media on cell surface hydrophobicity of a collection of Staphylococcus aureus and coagulase-negative staphylococci isolated from bovine mastitis were compared in the salt-aggregation test. Thirty-three per cent of Staph. aureus strains showed extremely high cell surface hydrophobicity (auto-aggregated) and 28% were moderately hydrophobic while 26% were hydrophilic after growth on horse blood agar at 37 degrees C for 18 h. There were great variations in the proportion and degree of the hydrophobicity depending on the medium used. Cultivations on/in capsule-inducing media caused a shift from a high to a low degree of hydrophobicity, although a microscopically detectable capsule or slime layer was seen in only one strain. This strain and encapsulated reference strains had a hydrophilic cell surface and migrated faster in free zone electrophoresis than cells of unencapsulated strains. Cells of strains grown on staphylococcus medium 110 agar migrated faster than those grown on horse blood agar regardless of their capsule production. Coagulase-negative staphylococci showed uniformly hydrophilic cell surface after cultivation on horse blood agar, but not when grown in tryptic soy broth or proteose peptone broth. It was concluded that most of the Staph. aureus strains from bovine mastitis under a variety of growth conditions in stationary phase culture constantly expressed hydrophobic cell surface.

  17. Membrane tethering of APP c-terminal fragments is a prerequisite for T668 phosphorylation preventing nuclear sphere generation. (United States)

    Bukhari, Hassan; Kolbe, Katharina; Leonhardt, Gregor; Loosse, Christina; Schröder, Elisabeth; Knauer, Shirley; Marcus, Katrin; Müller, Thorsten


    A central molecular hallmark of Alzheimer's disease (AD) is the β- and γ-secretase-mediated cleavage of the amyloid precursor protein (APP), which causes the generation of different c-terminal fragments like C99, AICD57, or AICD50 that fully or in part contain the APP transmembrane domain. In this study, we demonstrate that membrane-tethered C99 is phosphorylated by JNK3A at residue T668 (APP695 numbering) to a higher extent than AICD57, whereas AICD50 is not capable of being phosphorylated. The modification decreases the turnover of APP, while the blockade of APP cleavage increases APP phosphorylation. Generation of nuclear spheres, complexes consisting of the translocated AICD, FE65 and other proteins, is significantly reduced as soon as APP c-terminal fragments are accessible for phosphorylation. This APP modification, which we identified as significantly reduced in high plaque-load areas of the human brain, is linearly dependent on the level of APP expression. Accordingly, we show that APP abundance is likewise capable of modulating nuclear sphere generation. Thus, the precise and complex regulation of APP phosphorylation, abundance, and cleavage impacts the generation of nuclear spheres, which are under discussion of being of relevance in neurodegeneration and dementia. Future pharmacological manipulation of nuclear sphere generation may be a promising approach for AD treatment.

  18. Quantifying interspecific variation in dispersal ability of noctuid moths using an advanced tethered flight technique. (United States)

    Jones, Hayley B C; Lim, Ka S; Bell, James R; Hill, Jane K; Chapman, Jason W


    Dispersal plays a crucial role in many aspects of species' life histories, yet is often difficult to measure directly. This is particularly true for many insects, especially nocturnal species (e.g. moths) that cannot be easily observed under natural field conditions. Consequently, over the past five decades, laboratory tethered flight techniques have been developed as a means of measuring insect flight duration and speed. However, these previous designs have tended to focus on single species (typically migrant pests), and here we describe an improved apparatus that allows the study of flight ability in a wide range of insect body sizes and types. Obtaining dispersal information from a range of species is crucial for understanding insect population dynamics and range shifts. Our new laboratory tethered flight apparatus automatically records flight duration, speed, and distance of individual insects. The rotational tethered flight mill has very low friction and the arm to which flying insects are attached is extremely lightweight while remaining rigid and strong, permitting both small and large insects to be studied. The apparatus is compact and thus allows many individuals to be studied simultaneously under controlled laboratory conditions. We demonstrate the performance of the apparatus by using the mills to assess the flight capability of 24 species of British noctuid moths, ranging in size from 12-27 mm forewing length (~40-660 mg body mass). We validate the new technique by comparing our tethered flight data with existing information on dispersal ability of noctuids from the published literature and expert opinion. Values for tethered flight variables were in agreement with existing knowledge of dispersal ability in these species, supporting the use of this method to quantify dispersal in insects. Importantly, this new technology opens up the potential to investigate genetic and environmental factors affecting insect dispersal among a wide range of species.

  19. The Relationship Between Propulsive Force in Tethered Swimming and 200-m Front Crawl Performance. (United States)

    Santos, Karini B; Bento, Paulo C B; Pereira, Gleber; Rodacki, André L F


    Santos, KB, Bento, PCB, Pereira, G, and Rodacki, ALF. The relationship between propulsive force in tethered swimming and 200-m front crawl performance. J Strength Cond Res 30(9): 2500-2507, 2016-The aims of this study were to determine whether propulsive force (peak force, mean force, impulse, and rate of force development) and stroke rate change during 2 minutes of front crawl tethered swimming and to correlate them with the stroke rate and swimming velocity in 200-m front crawl swimming. Twenty-one swimmers (21.6 ± 4.8 years, 1.78 ± 0.06 m, 71.7 ± 8.1 kg), with 200-m front crawl swimming performance equivalent to 78% of the world record (140.4 ± 10.1 seconds), were assessed during 2 minutes of maximal front crawl tethered swimming (propulsive forces and stroke rate) and 200-m front crawl swimming (stroke rate and clean velocity). Propulsive forces decreased between the beginning and the middle instants (∼20%; p ≤ 0.05) but remained stable between the middle and the end instants (∼6%; p > 0.05). The peak force was positively correlated with the clean velocity in the 200-m front crawl swimming (mean r = 0.61; p < 0.02). The stroke rates of the tethered swimming and 200-m front crawl swimming were positively correlated (r = 45; p≤ 0.01) at the middle instant. Therefore, the propulsive force and stroke rate changed throughout the 2 minutes of tethered swimming, and the peak force is the best propulsive force variable tested that correlated with 200-m front crawl swimming performance.

  20. Cell surface estrogen receptor alpha is upregulated during subchronic metabolic stress and inhibits neuronal cell degeneration.

    Directory of Open Access Journals (Sweden)

    Cristiana Barbati

    Full Text Available In addition to the classical nuclear estrogen receptor, the expression of non-nuclear estrogen receptors localized to the cell surface membrane (mER has recently been demonstrated. Estrogen and its receptors have been implicated in the development or progression of numerous neurodegenerative disorders. Furthermore, the pathogenesis of these diseases has been associated with disturbances of two key cellular programs: apoptosis and autophagy. An excess of apoptosis or a defect in autophagy has been implicated in neurodegeneration. The aim of this study was to clarify the role of ER in determining neuronal cell fate and the possible implication of these receptors in regulating either apoptosis or autophagy. The human neuronal cell line SH-SY5Y and mouse neuronal cells in primary culture were thus exposed to chronic minimal peroxide treatment (CMP, a form of subcytotoxic minimal chronic stress previously that mimics multiple aspects of long-term cell stress and represents a limited molecular proxy for neurodegenerative processes. We actually found that either E2 or E2-bovine serum albumin construct (E2BSA, i.e. a non-permeant form of E2 was capable of modulating intracellular cell signals and regulating cell survival and death. In particular, under CMP, the up-regulation of mERα, but not mERβ, was associated with functional signals (ERK phosphorylation and p38 dephosphorylation compatible with autophagic cytoprotection triggering and leading to cell survival. The mERα trafficking appeared to be independent of the microfilament system cytoskeletal network but was seemingly associated with microtubular apparatus network, i.e., to MAP2 molecular chaperone. Importantly, antioxidant treatments, administration of siRNA to ERα, or the presence of antagonist of ERα hindered these events. These results support that the surface expression of mERα plays a pivotal role in determining cell fate, and that ligand-induced activation of mER signalling exerts a

  1. Cell-surface expression of Hsp70 on hematopoietic cancer cells after inhibition of HDAC activity

    DEFF Research Database (Denmark)

    Jensen, Helle

    -derived antigenic peptides, a function which is currently explored in immunotherapeutic approaches against cancer. Additionally, membrane-bound Hsp70 can stimulate antigen presenting cells to release proinflammatory cytokines and can provide a target structure for NK cell-mediated lysis. Human cancer cells...... frequently express Hsp70 on their cell surface, whereas the corresponding normal tissues do not. In addition, several clinically applied reagents, such as alkyl-lysophospholipides, chemotherapeutic agents, and anti-inflammatory reagents, have been found to enhance Hsp70 cell surface expression on cancer...... cells. We have found that inhibition of histone deacetylase (HDAC) activity leads to surface expression of Hsp70 on various hematopoietic cancer cells, an occurance that was not observed on naïve or activated peripheral blood cells. HDAC-inhibitor mediated Hsp70 cell surface expression was confined...

  2. Significance of nano- and microtopography for cell-surface interactions in orthopaedic implants. (United States)

    Jäger, M; Zilkens, C; Zanger, K; Krauspe, R


    Cell-surface interactions play a crucial role for biomaterial application in orthopaedics. It is evident that not only the chemical composition of solid substances influence cellular adherence, migration, proliferation and differentiation but also the surface topography of a biomaterial. The progressive application of nanostructured surfaces in medicine has gained increasing interest to improve the cytocompatibility and osteointegration of orthopaedic implants. Therefore, the understanding of cell-surface interactions is of major interest for these substances. In this review, we elucidate the principle mechanisms of nano- and microscale cell-surface interactions in vitro for different cell types onto typical orthopaedic biomaterials such as titanium (Ti), cobalt-chrome-molybdenum (CoCrMo) alloys, stainless steel (SS), as well as synthetic polymers (UHMWPE, XLPE, PEEK, PLLA). In addition, effects of nano- and microscaled particles and their significance in orthopaedics were reviewed. The significance for the cytocompatibility of nanobiomaterials is discussed critically.

  3. Cell surface activation of progelatinase A (proMMP—2) and cell migration

    Institute of Scientific and Technical Information of China (English)



    Gelatinase A (MMP-2) is considered to play a critical role in cell migration and invasion.The proteinase is cerceted from the cell as an inactive zymogen.In vivo it is postulated that activation of progelationase A (proMMP-2) takes place on the cell surface mediated by membrane-type matrix metalloproteinases (MT-MMPs).Recent studies have demonstrated that proMMP-2 is recruited to the cell surface by interacting with tissue inhibitor of metalloproteinases-2 (TIMP-2) bound to MT1-MMP by forming a ternary complex.Free MT1-MMP closely located to the ternary complex then activates proMMP-2 on the cell surface.MT1-MMP is found in cultured invasive cancer cells at the invadopodia.The MT-MMP/TIMP-2/MMP-2 system thus provides localized expression of proteolysis of the extracellular matrix required for cell migration.

  4. Cell surface differences of Naegleria fowleri and Naegleria lovaniensis exposed with surface markers. (United States)

    González-Robles, Arturo; Castañón, Guadalupe; Cristóbal-Ramos, Ana Ruth; Hernández-Ramírez, Verónica Ivonne; Omaña-Molina, Maritza; Martínez-Palomo, Adolfo


    Differences in the distribution of diverse cell surface coat markers were found between Naegleria fowleri and Naegleria lovaniensis. The presence of carbohydrate-containing components in the cell coat of the two species was detected by selective staining with ruthenium red and alcian blue. Using both markers, N. fowleri presented a thicker deposit than N. lovaniensis. The existence of exposed mannose or glucose residues was revealed by discriminatory agglutination with the plant lectin Concanavalin A. These sugar residues were also visualized at the cell surface of these parasites either by transmission electron microscopy or by fluorescein-tagged Concanavalin A. Using this lectin cap formation was induced only in N. fowleri. The anionic sites on the cell surface detected by means of cationized ferritin were more apparent in N. fowleri. Biotinylation assays confirmed that even though the two amoebae species have some analogous plasma membrane proteins, there is a clear difference in their composition.

  5. Nucleolin on the cell surface as a new molecular target for gastric cancer treatment. (United States)

    Watanabe, Tatsuro; Hirano, Kazuya; Takahashi, Atsushi; Yamaguchi, Kensei; Beppu, Masatoshi; Fujiki, Hirota; Suganuma, Masami


    Nucleolin is an abundant non-ribosomal protein found in nucleolus and a major component of silver-stained nucleolar organizer region (AgNOR), a histopathological marker of cancer which is highly elevated in cancer cells. We recently reported that nucleolin on the cell surface of mouse gastric cancer cells acts as a receptor for tumor necrosis factor-alpha-inducing protein (Tipalpha), a new carcinogenic factor of Helicobacter pylori. In this study, we first examined the localization of nucleolin on cell surface of five gastric cancer cell lines by cell fractionation and flow cytometry: We found that large amounts of nucleolin were present on surface of MKN-45, KATOIII, MKN-74, and AGS cells, with smaller amounts on surface of MKN-1 cells. The membrane fraction of normal epithelial cells of mouse glandular stomach did not contain much nucleolin, suggesting that translocation of nucleolin to the cell surface occurs during carcinogenesis, making for easier binding with Tipalpha. AS1411, a nucleolin targeted DNA aptamer, inhibited growth of gastric cancer cell lines in this order of potency: MKN-45>KATOIII>AGS>MKN-74=MKN-1, associated with induction of S-phase cell cycle arrest. Fluorescein isothiocyanate (FITC)-AS1411 was more rapidly incorporated into MKN-45 and AGS than into MKN-1 cells, based on varying amounts of cell surface nucleolin. We think that AS1411 first binds to nucleolin on the cell surface and that the binding complex is then incorporated into the cells. All results indicate that nucleolin on the cell surface is a new and promising therapeutic target for treatment of gastric cancer.

  6. Adhesion defective BHK cell mutant has cell surface heparan sulfate proteoglycan of altered properties

    DEFF Research Database (Denmark)

    Couchman, J R; Austria, R; Woods, A;


    In the light of accumulating data that implicate cell surface heparan sulfate proteoglycans (HSPGs) with a role in cell interactions with extracellular matrix molecules such as fibronectin, we have compared the properties of these molecules in wild-type BHK cells and an adhesion-defective ricin......-resistant mutant (RicR14). Our results showed that the mutant, unlike BHK cells, cannot form focal adhesions when adherent to planar substrates in the presence of serum. Furthermore, while both cell lines possess similar amounts of cell surface HSPG with hydrophobic properties, that of RicR14 cells had decreased...

  7. The Parkinson disease-related protein DJ-1 counteracts mitochondrial impairment induced by the tumour suppressor protein p53 by enhancing endoplasmic reticulum-mitochondria tethering. (United States)

    Ottolini, Denis; Calì, Tito; Negro, Alessandro; Brini, Marisa


    DJ-1 was first identified as an oncogene. More recently, mutations in its gene have been found causative for autosomal recessive familial Parkinson disease. Numerous studies support the DJ-1 role in the protection against oxidative stress and maintenance of mitochondria structure; however, the mechanism of its protective function remains largely unknown. We investigated whether mitochondrial Ca(2+) homeostasis, a key parameter in cell physiology, could be a target for DJ-1 action. Here, we show that DJ-1 modulates mitochondrial Ca(2+) transients induced upon cell stimulation with an 1,4,5-inositol-tris-phosphate agonist by favouring the endoplasmic reticulum (ER)-mitochondria tethering. A reduction of DJ-1 levels results in mitochondria fragmentation and decreased mitochondrial Ca(2+) uptake in stimulated cells. To functionally couple these effects with the well-recognized cytoprotective role of DJ-1, we investigated its action in respect to the tumour suppressor p53. p53 overexpression in HeLa cells impairs their ability to accumulate Ca(2+) in the mitochondrial matrix, causes alteration of the mitochondrial morphology and reduces ER-mitochondria contact sites. Mitochondrial impairments are independent from Drp1 activation, since the co-expression of the dominant negative mutant of Drp1 failed to abolish them. DJ-1 overexpression prevents these alterations by re-establishing the ER-mitochondria tethering. Similarly, the co-expression of the pro-fusion protein Mitofusin 2 blocks the effects induced by p53 on mitochondria, confirming that the modulation of the ER-mitochondria contact sites is critical to mitochondria integrity. Thus, the impairment of ER-mitochondria communication, as a consequence of DJ-1 loss-of-function, may be detrimental for mitochondria-related processes and be at the basis of mitochondrial dysfunction observed in Parkinson disease.

  8. Characterization of cell surface and extracellular matrix remodeling of Azospirillum brasilense chemotaxis-like 1 signal transduction pathway mutants by atomic force microscopy. (United States)

    Edwards, Amanda Nicole; Siuti, Piro; Bible, Amber N; Alexandre, Gladys; Retterer, Scott T; Doktycz, Mitchel J; Morrell-Falvey, Jennifer L


    To compete in complex microbial communities, bacteria must sense environmental changes and adjust cellular functions for optimal growth. Chemotaxis-like signal transduction pathways are implicated in the regulation of multiple behaviors in response to changes in the environment, including motility patterns, exopolysaccharide production, and cell-to-cell interactions. In Azospirillum brasilense, cell surface properties, including exopolysaccharide production, are thought to play a direct role in promoting flocculation. Recently, the Che1 chemotaxis-like pathway from A. brasilense was shown to modulate flocculation, suggesting an associated modulation of cell surface properties. Using atomic force microscopy, distinct changes in the surface morphology of flocculating A. brasilense Che1 mutant strains were detected. Whereas the wild-type strain produces a smooth mucosal extracellular matrix after 24 h, the flocculating Che1 mutant strains produce distinctive extracellular fibril structures. Further analyses using flocculation inhibition, lectin-binding assays, and comparison of lipopolysaccharides profiles suggest that the extracellular matrix differs between the cheA1 and the cheY1 mutants, despite an apparent similarity in the macroscopic floc structures. Collectively, these data indicate that disruption of the Che1 pathway is correlated with distinctive changes in the extracellular matrix, which likely result from changes in surface polysaccharides structure and/or composition.

  9. Characterization of cell surface and extracellular matrix remodeling of Azospirillum brasilense chemotaxis-like 1 signal transduction pathway mutants by atomic force microscopy

    Energy Technology Data Exchange (ETDEWEB)

    Doktycz, Mitchel John [ORNL; Morrell-Falvey, Jennifer L [ORNL


    To compete in complex microbial communities, bacteria must sense environmental changes and adjust cellular functions for optimal growth. Chemotaxis-like signal transduction pathways are implicated in the regulation of multiple behaviors in response to changes in the environment, including motility patterns, exopolysaccharide production, and cell-to-cell interactions. In Azospirillum brasilense, cell surface properties, including exopolysaccharide production, are thought to play a direct role in promoting flocculation. Recently, the Che1 chemotaxis-like pathway from A. brasilense was shown to modulate flocculation, suggesting an associated modulation of cell surface properties. Using atomic force microscopy, distinct changes in the surface morphology of flocculating A. brasilense Che1 mutant strains were detected. Whereas the wild-type strain produces a smooth mucosal extracellular matrix after 24 h, the flocculating Che1 mutant strains produce distinctive extracellular fibril structures. Further analyses using flocculation inhibition, lectin-binding assays, and comparison of lipopolysaccharides profiles suggest that the extracellular matrix differs between the cheA1 and the cheY1 mutants, despite an apparent similarity in the macroscopic floc structures. Collectively, these data indicate that disruption of the Che1 pathway is correlated with distinctive changes in the extracellular matrix, which likely result from changes in surface polysaccharides structure and/or composition.

  10. Design, synthesis, and evaluation of mitomycin-tethered phosphorothioate oligodeoxynucleotides. (United States)

    Huh, N; Rege, A A; Yoo, B; Kogan, T P; Kohn, H


    separate from 22. In the second procedure, phosphorothioate oligodexynucleotides that contained a hexylamino spacer at the 5'termini were coupled to 10-des(carbamoyloxy)-10-isothiocyanatoporfiromycin (9). Compound 9 was prepared in four steps from 11. Mesylation (methanesulfonyl chloride/pyridine) of 11 gave the C(10) mesylate 13, which was then treated with NaN3 (dimethylformamide, 90 degrees C) to give 10-des(carbamoyloxy)-10-azidoporfiromycin (14). Catalytic reduction (PtO2, H2) of 14 in pyridine afforded C(10) amine 15. Treatment of 15 with di-2-pyridyl thionocarbonate provided the desired 10-des(carbamoyloxy)-10-isothiocyanatoporfiromycin (9). Compound 9 readily coupled with 17 and base in both methylene chloride and aqueous buffered solutions to give 25. Use of the 5'hexylaminophosphorothioate oligodeoxynucleotides 32-35 in place of 17 gave the conjugated adducts 28-31, respectively, in a 12% to near-quantitative yield. The products were purified by semipreparative HPLC. Antisense agents 28-31 were designed to target a 30-base-long region from the coding region of the human FGFR1 gene. One adduct, 29, reduced the number of FGFR1 receptors in human aortic smooth cells for bFGF on the cell surface, which suggested down-regulation of FGFR1 gene expression. Further, 29 inhibited cultured human aortic smooth muscle cell proliferation and was less cytotoxic than porfiromycin (2). The biological assay data suggest that the phosphorothioate oligodexynucleotide porfiromycin conjugates may be more target selective and less toxic than either mitomycin or porfiromycin and thus be promising therapeutic agents.

  11. Cell-surface metalloprotease ADAM12 is internalized by a clathrin- and Grb2-dependent mechanism

    DEFF Research Database (Denmark)

    Hansen, Dorte Stautz; Leyme, Anthony; Grandal, Michael Vibo;


    ADAM12 (A Disintegrin And Metalloprotease 12), a member of the ADAMs family of transmembrane proteins, is involved in ectodomain shedding, cell-adhesion and signaling, with important implications in cancer. Therefore, mechanisms that regulate the levels and activity of ADAM12 at the cell-surface ...

  12. Cell Surface Enzymatic Engineering-Based Approaches to Improve Cellular Therapies

    KAUST Repository

    AbuElela, Ayman


    The cell surface represents the interface between the cell and its environment. As such, the cell surface controls cell–cell interactions and functions such as adhesion and migration, and will transfer external cues to regulate processes such as survival, death, and differentiation. Redefining the cell surface by temporarily (or permanently) modifying the molecular landscape of the plasma membrane affects the way in which the cell interacts with its environment and influences the information that is relayed into the cell along downstream signaling pathways. This chapter outlines the role of key enzymes, the glycosyltransferases, in posttranslationally modifying proteins and lipids to fine-tune cells, ability to migrate. These enzymes are critical in controlling the formation of a platform structure, sialyl Lewis x (sLex), on circulating cells that plays a central role in the recognition and recruitment by selectin counter receptors on endothelial cells that line blood vessels of tissues throughout the body. By developing methods to manipulate the activity of these enzymes and hence the cell surface structures that result, treatments can be envisioned that direct the migration of therapeutic cells to specific locations throughout the body and also to inhibit metastasis of detrimental cells such as circulating tumor cells.

  13. Extraction of cell surface-associated proteins from living yeast cells.

    NARCIS (Netherlands)

    F.M. Klis; M. de Jong; S. Brul; P.W.J. de Groot


    To extract cell surface-associated proteins from living fungal cells, reducing agents such as beta-mercaptoethanol and dithiothreitol are often used. We show here that both compounds are moderately lipophilic and may perturb the plasma membrane, thus causing the release of cytosolic proteins, especi

  14. Genetic and proteomic evidences support the localization of yeast enolase in the cell surface

    DEFF Research Database (Denmark)

    López-Villar, Elena; Monteoliva, Lucía; Larsen, Martin Røssel;


    Although enolase, other glycolytic enzymes, and a variety of cytoplasmic proteins lacking an N-terminal secretion signal have been widely described as located at the cell surface in yeast and in mammalian cells, their presence in this external location is still controversial. Here, we report that...

  15. Breast and ovarian cancers: a survey and possible roles for the cell surface heparan sulfate proteoglycans

    DEFF Research Database (Denmark)

    Yoneda, Atsuko; Lendorf, Maria E; Couchman, John R;


    of breast cancer may also develop ovarian cancer. Here, the authors review the different tumor markers of breast and ovarian carcinoma and discuss the expression, mutations, and possible roles of cell surface heparan sulfate proteoglycans during tumorigenesis of these carcinomas. The focus is on two groups...

  16. Growth condition-dependent cell surface proteome analysis of Enterococcus faecium

    NARCIS (Netherlands)

    Sinnige, Jan C; de Been, Mark; Zhou, Miaomiao; Bonten, Marc J M; Willems, Rob J L; Top, Janetta


    The last 30 years Enterococcus faecium has become an important nosocomial pathogen in hospitals worldwide. The aim of this study was to obtain insight in the cell surface proteome of E. faecium when grown in laboratory and clinically relevant conditions. Enterococcus faecium E1162, a clinical blood

  17. Oxidation state of chromium associated with cell surfaces of Shewanella oneidensis during chromate reduction

    Energy Technology Data Exchange (ETDEWEB)

    Neal, Andrew L.; Lowe, Kristine; Daulton, Tyrone L.; Jones-Meehan, Joanne; Little, Brenda J


    Employing electron energy loss spectroscopy (EELS) and X-ray photoelectron spectroscopy (XPS), we demonstrate that in both aerobic and anaerobic culture Shewanella oneidensis cells are capable of chromate reduction. No Cr(VI) or Cr(V) species were identified at the cell surfaces in Cr 2p{sub 3/}ore photoelectron spectra. More chromium was associated with cell surfaces recovered from anaerobic medium than aerobic. Multiplet-splitting models derived for Cr(III) and Cr(IV) were employed to determine contributions from each ion to Cr 2p{sub 3/2} photopeaks collected from the various cell treatments. Whilst in all cases Cr(III) was the major ion associated with cell surfaces, a significant contribution was identified due to Cr(IV) in anaerobically grown cells. The Cr(IV) contribution was far less when cells were grown aerobically. Moreover, when anaerobically grown cells were exposed to oxygen very little re-oxidation of Cr-precipitates occurred, the precipitates were again identified as a mixture of Cr(III) and Cr(IV). A positive relationship was observed between amounts of chromium and phosphorous associated with cell surfaces resulting from the various treatments, suggesting the precipitates included Cr(III)-phosphate. The fact that Cr(IV) remained associated with precipitates following re-oxidation suggests that under anaerobic conditions the intermediate ion is afforded sufficient stability to be incorporated within the precipitate matrix and thus conferred a degree of protection from oxidation.

  18. The ligand-binding domain of the cell surface receptor for urokinase-type plasminogen activator

    DEFF Research Database (Denmark)

    Behrendt, N; Ploug, M; Patthy, L;


    part of the intact receptor, probably including the whole sequence 1-87, and contained N-linked carbohydrate. After detergent phase separation in the Triton X-114 system, the fragment was present in the water phase where its binding activity could be demonstrated in the absence of the rest...... applications in interfering with cell-surface plasmin-mediated proteolysis....

  19. Cell surface heparan sulfate proteoglycans control adhesion and invasion of breast carcinoma cells

    DEFF Research Database (Denmark)

    Lim, Hooi Ching; Multhaupt, Hinke A. B.; Couchman, John R.


    phenotype of mammary carcinoma cells. Finally, both syndecan-2 and caveolin-2 were upregulated in tissue arrays from breast cancer patients compared to normal mammary tissue. Moreover their expression levels were correlated in triple negative breast cancers. Conclusion: Cell surface proteoglycans, notably...

  20. Cell surface physico chemistry alters biofilm development of Pseudomonas aeruginosa lipopolysaccharide mutants

    NARCIS (Netherlands)

    Flemming, CA; Palmer, RJ; Arrage, AA; Van der Mei, HC; White, DC


    The hydrophobic and electrostatic characteristics of bacterial cell surfaces were compared with attachment proclivity and biomass accumulation over time between wildtype Pseudomonas aeruginosa serotype O6 (possesses A and B band LPS), and three LPS-deficient mutants, vi;. A28 (A(+)B(-)), R5 (A(+)B(-

  1. Comprehensive proteomic analysis of Trypanosoma cruzi epimastigote cell surface proteins by two complementary methods

    DEFF Research Database (Denmark)

    Queiroz, Rayner M L; Charneau, Sébastien; Motta, Flávia N;


    Trypanosoma cruzi is a protozoan that causes Chagas' disease, a neglected infectious illness that affects millions of people, mostly in Latin America. Here, the cell surface subproteome of the T. cruzi epimastigote life form was characterized. In order to prepare samples enriched in epimastigote...

  2. Selective labelling of cell-surface proteins using CyDye DIGE Fluor minimal dyes. (United States)

    Hagner-McWhirter, Asa; Winkvist, Maria; Bourin, Stephanie; Marouga, Rita


    Surface proteins are central to the cell's ability to react to its environment and to interact with neighboring cells. They are known to be inducers of almost all intracellular signaling. Moreover, they play an important role in environmental adaptation and drug treatment, and are often involved in disease pathogenesis and pathology (1). Protein-protein interactions are intrinsic to signaling pathways, and to gain more insight in these complex biological processes, sensitive and reliable methods are needed for studying cell surface proteins. Two-dimensional (2-D) electrophoresis is used extensively for detection of biomarkers and other targets in complex protein samples to study differential changes. Cell surface proteins, partly due to their low abundance (1 2% of cellular proteins), are difficult to detect in a 2-D gel without fractionation or some other type of enrichment. They are also often poorly represented in 2-D gels due to their hydrophobic nature and high molecular weight (2). In this study, we present a new protocol for intact cells using CyDye DIGE Fluor minimal dyes for specific labeling and detection of this important group of proteins. The results showed specific labeling of a large number of cell surface proteins with minimal labeling of intracellular proteins. This protocol is rapid, simple to use, and all three CyDye DIGE Fluor minimal dyes (Cy 2, Cy 3 and Cy 5) can be used to label cell-surface proteins. These features allow for multiplexing using the 2-D Fluorescence Difference Gel Electrophoresis (2-D DIGE) with Ettan DIGE technology and analysis of protein expression changes using DeCyder 2-D Differential Analysis Software. The level of cell-surface proteins was followed during serum starvation of CHO cells for various lengths of time (see Table 1). Small changes in abundance were detected with high accuracy, and results are supported by defined statistical methods.

  3. ProtEx: a novel technology to display exogenous proteins on the cell surface for immunomodulation. (United States)

    Singh, Narendra P; Yolcu, Esma S; Askenasy, Nadir; Shirwan, Haval


    Gene therapy as an immunomodulatory approach has the potential to treat various inherited and acquired immune-based human diseases. However, its clinical application has several challenges, varying from the efficiency of gene transfer, control of gene expression, cell and tissue targeting, and safety concerns associated with the introduction of exogenous DNA into cells/tissues. Gene therapy is also a time- and labor-intensive procedure. As an alternative, we recently developed a novel technology, ProtEx, that allows for rapid, efficient, and durable display of exogenous proteins on the surface of cells, tissues, and organs without detectable toxicity. This technology exploits the strong binding affinity (Kd = 10(-15) M) of streptavidin with biotin and involves generation of chimeric molecules composed of the extracellular portions of immunological proteins of interest and a modified form of streptavidin, biotinylation of biological surfaces, and decoration of the modified surface with chimeric proteins. Biotin persists on the cell surface for weeks both in vitro and in vivo, thereby providing a platform to display exogenous proteins with extended cell surface kinetics. Two chimeric proteins, rat FasL (SA-FasL) and human CD80 (CD80-SA), were generated and tested for cell surface display and immunomodulatory functions. SA-FasL and CD80-SA molecules persisted on the surface of various cell types for extended periods, varying from days to weeks in vitro and in vivo. The cell surface kinetics, however, were protein and cell type dependent. SA-FasL showed potent apoptotic activity against Fas+ cells as a soluble protein or displayed on the cell surface and effectively blocked alloreactive responses. The display of CD80-SA on the surface of tumor cells, however, converted them into antigen-presenting cells for effective stimulation of autologous and allogeneic T-cell responses. ProtEx technology, therefore, represents a practical and effective alternative to DNA

  4. The HOPS/Class C Vps Complex Tethers High-Curvature Membranes via a Direct Protein-Membrane Interaction. (United States)

    Ho, Ruoya; Stroupe, Christopher


    Membrane tethering is a physical association of two membranes before their fusion. Many membrane tethering factors have been identified, but the interactions that mediate inter-membrane associations remain largely a matter of conjecture. Previously, we reported that the homotypic fusion and protein sorting/Class C vacuolar protein sorting (HOPS/Class C Vps) complex, which has two binding sites for the yeast vacuolar Rab GTPase Ypt7p, can tether two low-curvature liposomes when both membranes bear Ypt7p. Here, we show that HOPS tethers highly curved liposomes to Ypt7p-bearing low-curvature liposomes even when the high-curvature liposomes are protein-free. Phosphorylation of the curvature-sensing amphipathic lipid-packing sensor (ALPS) motif from the Vps41p HOPS subunit abrogates tethering of high-curvature liposomes. A HOPS complex without its Vps39p subunit, which contains one of the Ypt7p binding sites in HOPS, lacks tethering activity, though it binds high-curvature liposomes and Ypt7p-bearing low-curvature liposomes. Thus, HOPS tethers highly curved membranes via a direct protein-membrane interaction. Such high-curvature membranes are found at the sites of vacuole tethering and fusion. There, vacuole membranes bend sharply, generating large areas of vacuole-vacuole contact. We propose that HOPS localizes via the Vps41p ALPS motif to these high-curvature regions. There, HOPS binds via Vps39p to Ypt7p in an apposed vacuole membrane.

  5. A tethered bilayer sensor containing alamethicin channels and its detection of amiloride based inhibitors. (United States)

    Yin, Ping; Burns, Christopher J; Osman, Peter D J; Cornell, Bruce A


    Alamethicin, a small transmembrane peptide, inserts into a tethered bilayer membrane (tBLM) to form ion channels, which we have investigated using electrical impedance spectroscopy. The number of channels formed is dependent on the incubation time, concentration of the alamethicin and the application of DC voltage. The properties of the ion channels when formed in tethered bilayers are similar to those for such channels assembled into black lipid membranes (BLMs). Furthermore, amiloride and certain analogs can inhibit the channel pores, formed in the tBLMs. The potency and concentration of the inhibitors can be determined by measuring the change of impedance. Our work illustrates the possibility of using a synthetic tBLM for the study of small peptide voltage dependent ion channels. A potential application of such a device is as a screening tool in drug discovery processes.

  6. Force-free measurements of the conformations of DNA molecules tethered to a wall (United States)

    Lindner, Moshe; Nir, Guy; Medalion, Shlomi; Dietrich, Heidelinde R. C.; Rabin, Yitzhak; Garini, Yuval


    Using an optimized combination of tethered particle motion method, total internal reflection, and a gold nanobead, we measured the three-dimensional distribution of the free end of a tethered DNA molecule. The distribution along the axial z direction (perpendicular to the surface) is found to be Rayleigh-like, in agreement with wormlike chain and freely jointed chain simulations. Using these simulations, we show that the presence of the wall increases the correlations between the orientations of neighboring chain segments compared to free DNA. While the measured and the simulated planar (xy) distributions always agree with that of a Gaussian-random-walk (GRW) model, for short DNA lengths (1 μm) studied in our experiment, the corresponding axial (z) distributions deviate from those predicted for a GRW confined to half-space.

  7. Experimental verification of chaotic control of an underactuated tethered satellite system (United States)

    Pang, Zhaojun; Jin, Dongping


    This paper studies chaotic control of a tethered satellite system (TSS) driven only by a momentum-exchange device during its attitude adjustment. In dealing with such the underactuated system, an extended time-delay autosynchronization (ETDAS) is employed to stabilize the chaotic motion to a periodic motion. To obtain the control domains of the ETDAS method, a stability analysis of the controlled tethered satellite system in elliptical orbit is implemented. According to the principle of dynamic similarity, then, ground-based experiment setups are proposed and designed to emulate the in-plane motions of the TSS. Representative experiments are presented to demonstrate the effectiveness of the ETDAS scheme in controlling the chaotic motion of the underactuated TSS.

  8. Use of Site-Specifically Tethered Chemical Nucleases to Study Macromolecular Reactions

    Directory of Open Access Journals (Sweden)

    Mukherjee Srabani


    Full Text Available During a complex macromolecular reaction multiple changes in molecular conformation and interactions with ligands may occur. X-ray crystallography may provide only a limited set of snapshots of these changes. Solution methods can augment such structural information to provide a more complete picture of a macromolecular reaction. We analyzed the changes in protein conformation and protein:nucleic acid interactions which occur during transcription initiation by using a chemical nuclease tethered to cysteines introduced site-specifically into the RNA polymerase of bacteriophage T7 (T7 RNAP. Changes in cleavage patterns as the polymerase steps through transcription reveal a series of structural transitions which mediate transcription initiation. Cleavage by tethered chemical nucleases is seen to be a powerful method for revealing the conformational dynamics of macromolecular reactions, and has certain advantages over cross-linking or energy transfer approaches.

  9. Loss of cell-surface laminin anchoring promotes tumor growth and is associated with poor clinical outcomes. (United States)

    Akhavan, Armin; Griffith, Obi L; Soroceanu, Liliana; Leonoudakis, Dmitri; Luciani-Torres, Maria Gloria; Daemen, Anneleen; Gray, Joe W; Muschler, John L


    Perturbations in the composition and assembly of extracellular matrices (ECM) contribute to progression of numerous diseases, including cancers. Anchoring of laminins at the cell surface enables assembly and signaling of many ECMs, but the possible contributions of altered laminin anchoring to cancer progression remain undetermined. In this study, we investigated the prominence and origins of defective laminin anchoring in cancer cells and its association with cancer subtypes and clinical outcomes. We found loss of laminin anchoring to be widespread in cancer cells. Perturbation of laminin anchoring originated from several distinct defects, which all led to dysfunctional glycosylation of the ECM receptor dystroglycan. In aggressive breast and brain cancers, defective laminin anchoring was often due to suppressed expression of the glycosyltransferase LARGE. Reduced expression of LARGE characterized a broad array of human tumors in which it was associated with aggressive cancer subtypes and poor clinical outcomes. Notably, this defect robustly predicted poor survival in patients with brain cancers. Restoring LARGE expression repaired anchoring of exogenous and endogenous laminin and modulated cell proliferation and tumor growth. Together, our findings suggest that defects in laminin anchoring occur commonly in cancer cells, are characteristic of aggressive cancer subtypes, and are important drivers of disease progression.

  10. Characterization of heparin-binding site of tissue transglutaminase: its importance in cell surface targeting, matrix deposition, and cell signaling. (United States)

    Wang, Zhuo; Collighan, Russell J; Pytel, Kamila; Rathbone, Daniel L; Li, Xiaoling; Griffin, Martin


    Tissue transglutaminase (TG2) is a multifunctional Ca(2+)-activated protein cross-linking enzyme secreted into the extracellular matrix (ECM), where it is involved in wound healing and scarring, tissue fibrosis, celiac disease, and metastatic cancer. Extracellular TG2 can also facilitate cell adhesion important in wound healing through a nontransamidating mechanism via its association with fibronectin, heparan sulfates (HS), and integrins. Regulating the mechanism how TG2 is translocated into the ECM therefore provides a strategy for modulating these physiological and pathological functions of the enzyme. Here, through molecular modeling and mutagenesis, we have identified the HS-binding site of TG2 (202)KFLKNAGRDCSRRSSPVYVGR(222). We demonstrate the requirement of this binding site for translocation of TG2 into the ECM through a mechanism involving cell surface shedding of HS. By synthesizing a peptide NPKFLKNAGRDCSRRSS corresponding to the HS-binding site within TG2, we also demonstrate how this mimicking peptide can in isolation compensate for the RGD-induced loss of cell adhesion on fibronectin via binding to syndecan-4, leading to activation of PKCα, pFAK-397, and ERK1/2 and the subsequent formation of focal adhesions and actin cytoskeleton organization. A novel regulatory mechanism for TG2 translocation into the extracellular compartment that depends upon TG2 conformation and the binding of HS is proposed.

  11. Supramolecularly-knitted Tethered Oligopeptide/Single-walled Carbon Nanotube Organogels (United States)

    Zou, Jiong; He, Xun; Fan, Jingwei; Raymond, Jeffery E.


    A facile polymerization of an allyl-functional N-carboxyanhydride (NCA) monomer is utilized to construct an A-B-A type triblock structure containing β-sheet-rich oligomeric peptide segments tethered by a poly(ethylene oxide) chain, which are capable of dispersing and gelating single-walled carbon nanotubes (SWCNTs) noncovalently in organic solvents, resulting in significant enhancement of the mechanical properties of polypeptide-based organogels. PMID:24961389

  12. Diagnosis and surgical treatment of terminal syringomyelia within spinal cord combined with tethered cord syndrome


    Jing-cheng XIE; Wang, Zhen-Yu; Chen, Xiao-Dong


    Objective To summarize the clinical manifestations, imaging characteristics and experience of surgical treatment of spinal cord terminal syringomyelia with tethered cord syndrome (TCS).  Methods and Results Clinical data of 10 patients with spinal cord syringomyelia combined with TCS surgically treated under microscope from January 1999 to March 2014 in our hospital were retrospectively analyzed. There were 3 males and 7 females with average age of 15.06 years old (ranged from 2 to 35 y...

  13. Turbulence fluxes and variances measured with a sonic anemometer mounted on a tethered balloon


    Canut, Guylaine; Couvreux, Fleur; Lothon, Marie; Legain, Dominique; Piguet, Bruno; Lampert, Astrid; Maurel, William; Moulin, Eric


    This study presents the first deployment in field campaigns of a balloon-borne turbulence probe, developed with a sonic anemometer and an inertial motion sensor suspended below a tethered balloon. This system measures temperature and horizontal and vertical wind at high frequency and allows the estimation of heat and momentum fluxes as well as turbulent kinetic energy in the lower part of the boundary layer. The system was validated during three field experiments with differ...

  14. Lifting options for stratospheric aerosol geoengineering: advantages of tethered balloon systems. (United States)

    Davidson, Peter; Burgoyne, Chris; Hunt, Hugh; Causier, Matt


    The Royal Society report 'Geoengineering the Climate' identified solar radiation management using albedo-enhancing aerosols injected into the stratosphere as the most affordable and effective option for geoengineering, but did not consider in any detail the options for delivery. This paper provides outline engineering analyses of the options, both for batch-delivery processes, following up on previous work for artillery shells, missiles, aircraft and free-flying balloons, as well as a more lengthy analysis of continuous-delivery systems that require a pipe connected to the ground and supported at a height of 20 km, either by a tower or by a tethered balloon. Towers are shown not to be practical, but a tethered balloon delivery system, with high-pressure pumping, appears to have much lower operating and capital costs than all other delivery options. Instead of transporting sulphuric acid mist precursors, such a system could also be used to transport slurries of high refractive index particles such as coated titanium dioxide. The use of such particles would allow useful experiments on opacity, coagulation and atmospheric chemistry at modest rates so as not to perturb regional or global climatic conditions, thus reducing scale-up risks. Criteria for particle choice are discussed, including the need to minimize or prevent ozone destruction. The paper estimates the time scales and relatively modest costs required if a tethered balloon system were to be introduced in a measured way with testing and development work proceeding over three decades, rather than in an emergency. The manufacture of a tether capable of sustaining the high tensions and internal pressures needed, as well as strong winds, is a significant challenge, as is the development of the necessary pumping and dispersion technologies. The greatest challenge may be the manufacture and launch of very large balloons, but means have been identified to significantly reduce the size of such balloons or aerostats.

  15. A Quantitative Study of Tethered Chains in Various Solution Conditions Using Langmuir Diblock Copolymer Monolayers

    Energy Technology Data Exchange (ETDEWEB)

    Kent, Michael S.


    This article summarizes our investigations of tethered chain systems using Langmuir monolayer of polydimethysiloxane-poly styrene (PDMS-PS) diblock copolymers on organic liquids. In this system, the PDMS block adsorbs to the air surface while the PS block dangles into the subphase liquid. The air surface can be made either repulsive or attractive for the tethered PS chain segments by choosing a subphase liquid which has a surface tension lower or greater than that of PS, respectively. The segment profile of the PS block is determined by neutron reflection as a function of the surface density, the molecular weights of the PS and PDMS blocks, and the solution conditions. We cover the range of reduced surface density (SIGMA) characteristic of the large body of data in the literature for systems of chains tethered onto solid surfaces from dilute solution in good or theta solvent conditions (SIGMA < 12). We emphasize quantitative comparisons with analytical profile forms and scaling predictions. We find that the strong-stretching limit invoked in analytical SCF and scaling theories is not valid over this Z range. On the other hand, over a large portion of this range (SIGMA < 5) tethered layers are well described by a renormalization group theory addressing weakly interacting or noninteracting chains. Simultaneous with the study of the profile form, the free energy of the chains is examined through the surface tension. A strong increase in the surface pressure is observed with increasing surface density which determines the maximum surface density which can be achieved. This apparently nonequilibrium effect is attributed to steric interactions and limited lateral interpenetration. This effect may explain several outstanding discrepancies regarding the adsorption of end-functionalized chains and diblock copolymers onto solid surfaces.

  16. A Correlational Analysis of Tethered Swimming, Swim Sprint Performance and Dry-land Power Assessments. (United States)

    Loturco, I; Barbosa, A C; Nocentini, R K; Pereira, L A; Kobal, R; Kitamura, K; Abad, C C C; Figueiredo, P; Nakamura, F Y


    Swimmers are often tested on both dry-land and in swimming exercises. The aim of this study was to test the relationships between dry-land, tethered force-time curve parameters and swimming performances in distances up to 200 m. 10 young male high-level swimmers were assessed using the maximal isometric bench-press and quarter-squat, mean propulsive power in jump-squat, squat and countermovement jumps (dry-land assessments), peak force, average force, rate of force development (RFD) and impulse (tethered swimming) and swimming times. Pearson product-moment correlations were calculated among the variables. Peak force and average force were very largely correlated with the 50- and 100-m swimming performances (r=- 0.82 and -0.74, respectively). Average force was very-largely/largely correlated with the 50- and 100-m performances (r=- 0.85 and -0.67, respectively). RFD and impulse were very-largely correlated with the 50-m time (r=- 0.72 and -0.76, respectively). Tethered swimming parameters were largely correlated (r=0.65 to 0.72) with mean propulsive power in jump-squat, squat-jump and countermovement jumps. Finally, mean propulsive power in jump-squat was largely correlated (r=- 0.70) with 50-m performance. Due to the significant correlations between dry-land assessments and tethered/actual swimming, coaches are encouraged to implement strategies able to increase leg power in sprint swimmers.

  17. Characteristic Changes in Cell Surface Glycosylation Accompany Intestinal Epithelial Cell (IEC) Differentiation: High Mannose Structures Dominate the Cell Surface Glycome of Undifferentiated Enterocytes. (United States)

    Park, Dayoung; Brune, Kristin A; Mitra, Anupam; Marusina, Alina I; Maverakis, Emanual; Lebrilla, Carlito B


    Changes in cell surface glycosylation occur during the development and differentiation of cells and have been widely correlated with the progression of several diseases. Because of their structural diversity and sensitivity to intra- and extracellular conditions, glycans are an indispensable tool for analyzing cellular transformations. Glycans present on the surface of intestinal epithelial cells (IEC) mediate interactions with billions of native microorganisms, which continuously populate the mammalian gut. A distinct feature of IECs is that they differentiate as they migrate upwards from the crypt base to the villus tip. In this study, nano-LC/ESI QTOF MS profiling was used to characterize the changes in glycosylation that correspond to Caco-2 cell differentiation. As Caco-2 cells differentiate to form a brush border membrane, a decrease in high mannose type glycans and a concurrent increase in fucosylated and sialylated complex/hybrid type glycans were observed. At day 21, when cells appear to be completely differentiated, remodeling of the cell surface glycome ceases. Differential expression of glycans during IEC maturation appears to play a key functional role in regulating the membrane-associated hydrolases and contributes to the mucosal surface innate defense mechanisms. Developing methodologies to rapidly identify changes in IEC surface glycans may lead to a rapid screening approach for a variety of disease states affecting the GI tract.

  18. Triggered Ca2+ influx is required for extended synaptotagmin 1-induced ER-plasma membrane tethering. (United States)

    Idevall-Hagren, Olof; Lü, Alice; Xie, Beichen; De Camilli, Pietro


    The extended synaptotagmins (E-Syts) are ER proteins that act as Ca(2+)-regulated tethers between the ER and the plasma membrane (PM) and have a putative role in lipid transport between the two membranes. Ca(2+) regulation of their tethering function, as well as the interplay of their different domains in such function, remains poorly understood. By exposing semi-intact cells to buffers of variable Ca(2+) concentrations, we found that binding of E-Syt1 to the PI(4,5)P2-rich PM critically requires its C2C and C2E domains and that the EC50 of such binding is in the low micromolar Ca(2+) range. Accordingly, E-Syt1 accumulation at ER-PM contact sites occurred only upon experimental manipulations known to achieve these levels of Ca(2+) via its influx from the extracellular medium, such as store-operated Ca(2+) entry in fibroblasts and membrane depolarization in β-cells. We also show that in spite of their very different physiological functions, membrane tethering by E-Syt1 (ER to PM) and by synaptotagmin (secretory vesicles to PM) undergo a similar regulation by plasma membrane lipids and cytosolic Ca(2+).

  19. A Structure-Based Mechanism for Vesicle Capture by the Multisubunit Tethering Complex Dsl1

    Energy Technology Data Exchange (ETDEWEB)

    Ren, Y.; Yip, C; Tripathi, A; Huie, D; Jeffrey, P; Walz, T; Hughson, F


    Vesicle trafficking requires membrane fusion, mediated by SNARE proteins, and upstream events that probably include tethering, an initial long-range attachment between a vesicle and its target organelle. Among the factors proposed to mediate tethering are a set of multisubunit tethering complexes (MTCs). The Dsl1 complex, with only three subunits, is the simplest known MTC and is essential for the retrograde traffic of COPI-coated vesicles from the Golgi to the ER. To elucidate structural principles underlying MTC function, we have determined the structure of the Dsl1 complex, revealing a tower containing at its base the binding sites for two ER SNAREs and at its tip a flexible lasso for capturing vesicles. The Dsl1 complex binds to individual SNAREs via their N-terminal regulatory domains and also to assembled SNARE complexes; moreover, it is capable of accelerating SNARE complex assembly. Our results suggest that even the simplest MTC may be capable of orchestrating vesicle capture, uncoating, and fusion.

  20. The Atg1-kinase complex tethers Atg9-vesicles to initiate autophagy (United States)

    Rao, Yijian; Perna, Marco G.; Hofmann, Benjamin; Beier, Viola; Wollert, Thomas


    Autophagosomes are double-membrane vesicles that sequester cytoplasmic material for lysosomal degradation. Their biogenesis is initiated by recruitment of Atg9-vesicles to the phagophore assembly site. This process depends on the regulated activation of the Atg1-kinase complex. However, the underlying molecular mechanism remains unclear. Here we reconstitute this early step in autophagy from purified components in vitro. We find that on assembly from its cytoplasmic subcomplexes, the Atg1-kinase complex becomes activated, enabling it to recruit and tether Atg9-vesicles. The scaffolding protein Atg17 targets the Atg1-kinase complex to autophagic membranes by specifically recognizing the membrane protein Atg9. This interaction is inhibited by the two regulatory subunits Atg31 and Atg29. Engagement of the Atg1-Atg13 subcomplex restores the Atg9-binding and membrane-tethering activity of Atg17. Our data help to unravel the mechanism that controls Atg17-mediated tethering of Atg9-vesicles, providing the molecular basis to understand initiation of autophagosome-biogenesis.

  1. A highly versatile adaptor protein for the tethering of growth factors to gelatin-based biomaterials. (United States)

    Addi, Cyril; Murschel, Frédéric; Liberelle, Benoît; Riahi, Nesrine; De Crescenzo, Gregory


    In the field of tissue engineering, the tethering of growth factors to tissue scaffolds in an oriented manner can enhance their activity and increase their half-life. We chose to investigate the capture of the basic Fibroblast Growth Factor (bFGF) and the Epidermal Growth Factor (EGF) on a gelatin layer, as a model for the functionalization of collagen-based biomaterials. Our strategy relies on the use of two high affinity interactions, that is, the one between two distinct coil peptides as well as the one occurring between a collagen-binding domain (CBD) and gelatin. We expressed a chimeric protein to be used as an adaptor that comprises one of the coil peptides and a CBD derived from the human fibronectin. We proved that it has the ability to bind simultaneously to a gelatin substrate and to form a heterodimeric coiled-coil domain with recombinant growth factors being tagged with the complementary coil peptide. The tethering of the growth factors was characterized by ELISA and surface plasmon resonance-based biosensing. The bioactivity of the immobilized bFGF and EGF was evaluated by a human umbilical vein endothelial cell proliferation assay and a vascular smooth muscle cell survival assay. We found that the tethering of EGF preserved its mitogenic and anti-apoptotic activity. In the case of bFGF, when captured via our adaptor protein, changes in its natural mode of interaction with gelatin were observed.

  2. Management of adult tethered cord syndrome: Our experience and review of literature

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    Kanwaljeet Garg


    Full Text Available Background: Tethered cord syndrome (TCS is a complex clinicopathologic entity, mostly described in children with limited number of studies describing in adults. This unique and rare subgroup of patients presents with characteristic features of TCS, but unlike children, pain is a predominant clinical symptom. Materials and Methods: Case records of 24 patients aged ≥16 years who had undergone surgery with a diagnosis of TCS between 2001 and 2011 were reviewed. Patients who have underwent surgery earlier for tethered cord or for diastematomyelia/spinal dysraphism and patients who had radiological evidence of tethering elements like lipoma of the cord on magnetic resonance imaging (MRI were excluded from the study. Results: Low backache was the most common presenting symptom. At the time of final follow-up, 15 (83.3% patients had shown improvement in backache. Weakness improved by at least one grade in seven (77.8% patients. Bladder symptoms improved in six (50% patients. Conclusion: In case of symptomatic patient with low-lying cord, detethering is an advisable option.

  3. History of the current understanding and management of tethered spinal cord. (United States)

    Safavi-Abbasi, Sam; Mapstone, Timothy B; Archer, Jacob B; Wilson, Christopher; Theodore, Nicholas; Spetzler, Robert F; Preul, Mark C


    An understanding of the underlying pathophysiology of tethered cord syndrome (TCS) and modern management strategies have only developed within the past few decades. Current understanding of this entity first began with the understanding and management of spina bifida; this later led to the gradual recognition of spina bifida occulta and the symptoms associated with tethering of the filum terminale. In the 17th century, Dutch anatomists provided the first descriptions and initiated surgical management efforts for spina bifida. In the 19th century, the term "spina bifida occulta" was coined and various presentations of spinal dysraphism were appreciated. The association of urinary, cutaneous, and skeletal abnormalities with spinal dysraphism was recognized in the 20th century. Early in the 20th century, some physicians began to suspect that traction on the conus medullaris caused myelodysplasia-related symptoms and that prophylactic surgical management could prevent the occurrence of clinical manifestations. It was not, however, until later in the 20th century that the term "tethered spinal cord" and the modern management of TCS were introduced. This gradual advancement in understanding at a time before the development of modern imaging modalities illustrates how, over the centuries, anatomists, pathologists, neurologists, and surgeons used clinical examination, a high level of suspicion, and interest in the subtle and overt clinical appearances of spinal dysraphism and TCS to advance understanding of pathophysiology, clinical appearance, and treatment of this entity. With the availability of modern imaging, spinal dysraphism can now be diagnosed and treated as early as the intrauterine stage.

  4. Controlled-surface-wettability-based fabrication of hydrogel substrates with matrix tethering density variations (United States)

    Rahman, Md. Mahmudur; Lee, Donghee; Bhagirath, Divya; Zhao, Xiangshan; Band, Vimla; Ryu, Sangjin


    It is widely accepted that cells behave differently responding to the stiffness of extracellular matrix (ECM). Such observations were made by culturing cells on hydrogel substrates of tunable stiffness. However, it was recently proposed that cells actually sense how strongly they are tethered to ECM, not the local stiffness of ECM. To investigate the hypothesis, we develop constant-stiffness hydrogel substrates with varying matrix tethering density (the number of anchoring sites between the gel and the ECM protein molecules). We fabricate polyacrylamide gel of static stiffness and conjugate ECM proteins to the gel using a cross-linker. When treating the gel with the cross-linker, we control positioning of cross-linker solutions with different concentrations using superhydrophobic barriers on glass, functionalize the gel by pressing it to the aligned cross-linker solutions, and conjugate an ECM protein of constant concentration to the gel. We expect that the gel will be functionalized to different degrees depending on the concentration distribution of the cross-linker and thus the gel will have variations of matrix tethering density even with constant ECM protein concentration. We acknowledge support from Bioengineering for Human Health grant of UNL-UNMC.

  5. MiniCORVET is a Vps8-containing early endosomal tether in Drosophila (United States)

    Lőrincz, Péter; Lakatos, Zsolt; Varga, Ágnes; Maruzs, Tamás; Simon-Vecsei, Zsófia; Darula, Zsuzsanna; Benkő, Péter; Csordás, Gábor; Lippai, Mónika; Andó, István; Hegedűs, Krisztina; Medzihradszky, Katalin F; Takáts, Szabolcs; Juhász, Gábor


    Yeast studies identified two heterohexameric tethering complexes, which consist of 4 shared (Vps11, Vps16, Vps18 and Vps33) and 2 specific subunits: Vps3 and Vps8 (CORVET) versus Vps39 and Vps41 (HOPS). CORVET is an early and HOPS is a late endosomal tether. The function of HOPS is well known in animal cells, while CORVET is poorly characterized. Here we show that Drosophila Vps8 is highly expressed in hemocytes and nephrocytes, and localizes to early endosomes despite the lack of a clear Vps3 homolog. We find that Vps8 forms a complex and acts together with Vps16A, Dor/Vps18 and Car/Vps33A, and loss of any of these proteins leads to fragmentation of endosomes. Surprisingly, Vps11 deletion causes enlargement of endosomes, similar to loss of the HOPS-specific subunits Vps39 and Lt/Vps41. We thus identify a 4 subunit-containing miniCORVET complex as an unconventional early endosomal tether in Drosophila. DOI: PMID:27253064

  6. TI tether rig for solving secular spinrate change problem of electric sail

    CERN Document Server

    Janhunen, Pekka


    The electric solar wind sail (E-sail) is a way to propel a spacecraft by using the natural solar wind as a thrust source. The problem of secular spinrate change was identified earlier which is due to the orbital Coriolis effect and tends to slowly increase or decrease the sail's spinrate, depending on which way the sail is inclined with respect to the solar wind. Here we present an E-sail design and its associated control algorithm which enable spinrate control during propulsive flight by the E-sail effect itself. In the design, every other maintether ("T-tether") is galvanically connected through the remote unit with the two adjacent auxtethers, while the other maintethers ("I-tethers") are insulated from the tethers. This enables one to effectively control the maintether and auxtether voltages separately, which in turn enables spinrate control. We use a detailed numerical simulation to show that the algorithm can fully control the E-sail's spin state in real solar wind. The simulation includes a simple and ...

  7. Tethered Satellites as an Enabling Platform for Operational Space Weather Monitoring Systems (United States)

    Gilchrist, Brian E.; Krause, Linda Habash; Gallagher, Dennis Lee; Bilen, Sven Gunnar; Fuhrhop, Keith; Hoegy, Walt R.; Inderesan, Rohini; Johnson, Charles; Owens, Jerry Keith; Powers, Joseph; Voronka, Nestor; Williams, Scott


    Tethered satellites offer the potential to be an important enabling technology to support operational space weather monitoring systems. Space weather "nowcasting" and forecasting models rely on assimilation of near-real-time (NRT) space environment data to provide warnings for storm events and deleterious effects on the global societal infrastructure. Typically, these models are initialized by a climatological model to provide "most probable distributions" of environmental parameters as a function of time and space. The process of NRT data assimilation gently pulls the climate model closer toward the observed state (e.g., via Kalman smoothing) for nowcasting, and forecasting is achieved through a set of iterative semi-empirical physics-based forward-prediction calculations. Many challenges are associated with the development of an operational system, from the top-level architecture (e.g., the required space weather observatories to meet the spatial and temporal requirements of these models) down to the individual instruments capable of making the NRT measurements. This study focuses on the latter challenge: we present some examples of how tethered satellites (from 100s of m to 20 km) are uniquely suited to address certain shortfalls in our ability to measure critical environmental parameters necessary to drive these space weather models. Examples include long baseline electric field measurements, magnetized ionospheric conductivity measurements, and the ability to separate temporal from spatial irregularities in environmental parameters. Tethered satellite functional requirements are presented for two examples of space environment observables.

  8. Postoperative epidural hematoma contributes to delayed upper cord tethering after decompression of Chiari malformation type I

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    Antonio Lopez-Gonzalez


    Full Text Available Background: Symptomatic arachnoiditis after posterior fossa surgical procedures such as decompression of Chiari malformation is a possible complication. Clinical presentation is generally insidious and delayed by months or years. It causes disturbances in the normal flow of cerebrospinal fluid and enlargement of a syrinx cavity in the upper spinal cord. Surgical de-tethering has favorable results with progressive collapse of the syrinx and relief of the associated symptoms. Case Description: A 30-year-old male with Chiari malformation type I was treated by performing posterior fossa bone decompression, dura opening and closure with a suturable bovine pericardium dural graft. Postoperative period was uneventful until the fifth day in which the patient suffered intense headache and progressive loose of consciousness caused by an acute posterior fossa epidural hematoma. It was quickly removed with complete clinical recovering. One year later, the patient experienced progressive worsened of his symptoms. Upper spinal cord tethering was diagnosed and a new surgery for debridement was required. Conclusions: The epidural hematoma compressing the dural graft against the neural structures contributes to the upper spinal cord tethering and represents a nondescribed cause of postoperative fibrosis, adhesion formation, and subsequent recurrent hindbrain compression.

  9. Specific Measurement of Tethered Running Kinetics and its Relationship to Repeated Sprint Ability. (United States)

    Sousa, Filipe; Dos Reis, Ivan; Ribeiro, Luiz; Martins, Luiz; Gobatto, Claudio


    Repeated sprint ability has been widely studied by researchers, however, analysis of the relationship between most kinetic variables and the effect of fatigue is still an ongoing process. To search for the best biomechanical parameter to evaluate repeated sprint ability, several kinetic variables were measured in a tethered field running test and compared regarding their sensitivity to fatigue and correlation with time trials in a free running condition. Nine male sprint runners (best average times: 100 m = 10.45 ± 0.07 s; 200 m = 21.36 ± 0.17 s; 400 m = 47.35 ± 1.09 s) completed two test sessions on a synthetic track. Each session consisted of six 35 m sprints interspersed by 10 s rest under tethered field running or free running conditions. Force, power, work, an impulse and a rate of force development were all directly measured using the sensors of a new tethered running apparatus, and a one-way ANOVA with Scheffé post-hoc test used to verify differences between sprints (p sprints. These three variables presented low to moderate correlations with free running performance (r between 0.01 and -0.35). Maximum and mean power presented the strongest correlations with free running performance (r = -0.71 and -0.76, respectively; p sprint ability than the other studied variables.

  10. Novel, UV-curable coatings containing a tethered biocide: Synthesis, characterization, and antimicrobial activity. (United States)

    Chen, Zhigang; Chisholm, Bret J; Stafslien, Shane; He, Jie; Patel, Sandeep


    Cationic, UV -curable coatings containing the tethered biocide, triclosan, were produced and their antimicrobial activity toward Staphylococcus epidermidis and Escherichia coli determined. Two polysiloxanes functionalized with both cycloaliphatic epoxy and triclosan were synthesized using hydrosilylation. The functionalized polysiloxanes, with varied concentration of pendant triclosan, were used to produce UV-curable coatings with reasonably good coating properties. Fourier transform (FT)-Raman spectroscopy showed that the tethered triclosan moieties self-concentrate on the coating surface. Using biological assays, it was determined that the coatings possessed nearly 100% antimicrobial activity toward the Gram-positive bacterium, S. epidermidis, without leaching toxic components. For the Gram-negative bacterium, E. coli, 60-80% reduction in biofilm retention was observed for all the coatings. Interestingly, the coatings were lesser effective in reducing E. coli cell viability suggesting that the tethered triclosan were able to substantially reduce the production of the biofilm extracellular matrix with minimal adverse affect on the bacterial cells attached to the coating surfaces. The high specificity of the coatings toward S. epidermidis indicates that a novel mode of contact-active antimicrobial activity was achieved through the disruption of processes unique to the Gram-positive cell wall. These novel UV-curable coatings have potential applications in inhibiting implantable biomedical device associated infections.

  11. Modelling the Bioelectronic Interface in Engineered Tethered Membranes: From Biosensing to Electroporation. (United States)

    Hoiles, William; Krishnamurthy, Vikram; Cornell, Bruce


    This paper studies the construction and predictive models of three novel measurement platforms: (i) a Pore Formation Measurement Platform (PFMP) for detecting the presence of pore forming proteins and peptides, (ii) the Ion Channel Switch (ICS) biosensor for detecting the presence of analyte molecules in a fluid chamber, and (iii) an Electroporation Measurement Platform (EMP) that provides reliable measurements of the electroporation phenomenon. Common to all three measurement platforms is that they are comprised of an engineered tethered membrane that is formed via a rapid solvent exchange technique allowing the platform to have a lifetime of several months. The membrane is tethered to a gold electrode bioelectronic interface that includes an ionic reservoir separating the membrane and gold surface, allowing the membrane to mimic the physiological response of natural cell membranes. The electrical response of the PFMP, ICS, and EMP are predicted using continuum theories for electrodiffusive flow coupled with boundary conditions for modelling chemical reactions and electrical double layers present at the bioelectronic interface. Experimental measurements are used to validate the predictive accuracy of the dynamic models. These include using the PFMP for measuring the pore formation dynamics of the antimicrobial peptide PGLa and the protein toxin Staphylococcal α-Hemolysin; the ICS biosensor for measuring nano-molar concentrations of streptavidin, ferritin, thyroid stimulating hormone (TSH), and human chorionic gonadotropin (pregnancy hormone hCG); and the EMP for measuring electroporation of membranes with different tethering densities, and membrane compositions.

  12. Cell Surface Receptor Theory of Disease Infectivity; Body's Defence and Normal Body Functioning in Living Things

    Directory of Open Access Journals (Sweden)



    Full Text Available Problem statement: A study of the pattern of Candida spp. infection of the human body and the mode and pattern of reaction of the human body to this infection showed that disease infectivity and self healing by plants followed the same procedures and patterns. Approach: A comparism of these procedures and patterns of natural self- healing of disease infection by the human body and plants/plant parts with the cutaneous Candida spp. killing and elimination procedures and patterns of Vernonia amygdalina leaf extract, showed that cell surface receptors are the sites through which disease infects the body and also the sites at which the body is defended. They are also the sites where activities which result in normal body functioning are carried out. The mode and patterns of Cutaneous Candida infection in a human subject and its containment by the body was examined and photographed. The disease infection and self healing procedures and patterns of plants were also examined in comparism with those of their healthy counterparts and photographed. The findings from the observations on disease infectivity and natural body’s defence patterns and procedures of the plant parts studied and those of the human body in reaction to Candida spp. infection were compared with those of the Candida spp. killing procedures and patterns of aqueous and Arachis hypogeal oil extract of Vernonia amygdalina leaf. Results: The findings of this study also showed that disease-infective organisms gain access to the body of a host through attachment to the cell surface receptors of that host which are placed linearly and are interconnected by channels. The results of the study also indicated that living organisms have a main endogenous substance that mediates both their body’s defence and their normal physiological functioning which is therefore the owner of the cell surface receptor. Other endogenous substances which participate in normal body functioning/body’s defence or in

  13. Nanoscale analysis of caspofungin-induced cell surface remodelling in Candida albicans (United States)

    El-Kirat-Chatel, Sofiane; Beaussart, Audrey; Alsteens, David; Jackson, Desmond N.; Lipke, Peter N.; Dufrêne, Yves F.


    The advent of fungal pathogens that are resistant to the classic repertoire of antifungal drugs has increased the need for new therapeutic agents. A prominent example of such a novel compound is caspofungin, known to alter cell wall biogenesis by inhibiting β-1,3-d-glucan synthesis. Although much progress has been made in understanding the mechanism of action of caspofungin, little is known about its influence on the biophysical properties of the fungal cells. Here, we use atomic force microscopy (AFM) to demonstrate that caspofungin induces major remodelling of the cell surface properties of Candida albicans. Caspofungin causes major morphological and structural alterations of the cells, which correlate with a decrease of the cell wall mechanical strength. Moreover, we find that the drug induces the massive exposure of the cell adhesion protein Als1 on the cell surface and leads to increased cell surface hydrophobicity, two features that trigger cell aggregation. This behaviour is not observed in yeast species lacking Als1, demonstrating the key role that the protein plays in determining the aggregation phenotype of C. albicans. The results show that AFM opens up new avenues for understanding the molecular bases of microbe-drug interactions and for developing new therapeutic agents.The advent of fungal pathogens that are resistant to the classic repertoire of antifungal drugs has increased the need for new therapeutic agents. A prominent example of such a novel compound is caspofungin, known to alter cell wall biogenesis by inhibiting β-1,3-d-glucan synthesis. Although much progress has been made in understanding the mechanism of action of caspofungin, little is known about its influence on the biophysical properties of the fungal cells. Here, we use atomic force microscopy (AFM) to demonstrate that caspofungin induces major remodelling of the cell surface properties of Candida albicans. Caspofungin causes major morphological and structural alterations of the

  14. Role of N-glycosylation in cell surface expression and protection against proteolysis of the intestinal anion exchanger SLC26A3. (United States)

    Hayashi, Hisayoshi; Yamashita, Yukari


    SLC26A3 is a Cl(-)/HCO(3)(-) exchanger that plays a major role in Cl(-) absorption from the intestine. Its mutation causes congenital chloride-losing diarrhea. It has been shown that SLC26A3 are glycosylated, with the attached carbohydrate being extracellular and perhaps modulating function. However, the role of glycosylation has yet to be clearly determined. We used the approaches of biochemical modification and site-directed mutagenesis to prevent glycosylation. Deglycosylation experiments with glycosidases indicated that the mature glycosylated form of SLC26A3 exists at the plasma membrane, and a putative large second extracellular loop contains all of the N-linked carbohydrates. Deglycosylation of SLC26A3 causes depression of transport activity compared with wild-type, although robust intracellular pH changes were still observed, suggesting that N-glycosylation is not absolutely necessary for transport activity. To localize glycosylation sites, we mutated the five consensus sites by replacing asparagine (N) with glutamine. Immnoblotting suggests that SLC26A3 is glycosylated at N153, N161, and N165. Deglycosylation of SLC26A3 causes a defect in cell surface processing with decreased cell surface expression. We also assessed whether SLC26A3 is protected from tryptic digestion. While the mature glycosylated SLC26A3 showed little breakdown after treatment with trypsin, deglycosylated SLC26A3 exhibited increased susceptibility to trypsin, suggesting that the oligosaccharides protect SLC26A3 from tryptic digestion. In conclusion, our data indicate that N-glycosylation of SLC26A3 is important for cell surface expression and for protection from proteolytic degradation that may contribute to the understanding of pathogenesis of congenital disorders of glycosylation.

  15. Cell surface domain specific postsynaptic currents evoked by identified GABAergic neurones in rat hippocampus in vitro. (United States)

    Maccaferri, G; Roberts, J D; Szucs, P; Cottingham, C A; Somogyi, P


    of IPSCs evoked by BiCs and O-BiCs showed the largest cell to cell variation, and a single interneurone could evoke both small and slow as well as large and relatively fast IPSCs. 6. The kinetic properties of the somatically recorded postsynaptic current are correlated with the innervated cell surface domain. A significant correlation of rise and decay times for the overall population of unitary IPSCs suggests that electrotonic filtering of distal responses is a major factor for the location and cell type specific differences of unitary IPSCs, but molecular heterogeneity of postsynaptic GABAA receptors may also contribute to the observed kinetic differences. Furthermore, domain specific differences in the short-term plasticity of the postsynaptic response indicate a differentiation of interneurones in activity-dependent responses.

  16. Optimum design of amphiphilic polymers bearing hydrophobic groups for both cell surface ligand presentation and intercellular cross-linking. (United States)

    Takeo, Masafumi; Li, Cuicui; Matsuda, Masayoshi; Nagai, Hiroko; Hatanaka, Wataru; Yamamoto, Tatsuhiro; Kishimura, Akihiro; Mori, Takeshi; Katayama, Yoshiki


    Amphiphilic polymers bearing hydrophobic alkyl groups are expected to be applicable for both ligand presentation on the cell surface and intercellular crosslinking. To explore the optimum design for each application, we synthesized eight different acyl-modified dextrans with varying molecular weight, alkyl length, and alkyl modification degree. We found that the behenate-modified polymers retained on the cell surface longer than the palmitate-modified ones. Since the polymers were also modified with biotin, streptavidin can be presented on the cell surface through biotin-streptavidin recognition. The duration of streptavidin on the cell surface is longer in the behenate-modified polymer than the palmitate-modified one. As for the intercellular crosslinking, the palmitate-modified polymers were more efficient than the behenate-modified polymers. The findings in this research will be helpful to design the acyl-modified polymers for the cell surface engineering.

  17. Cell-Surface Protein Profiling Identifies Distinctive Markers of Progenitor Cells in Human Skeletal Muscle

    Directory of Open Access Journals (Sweden)

    Akiyoshi Uezumi


    Full Text Available Skeletal muscle contains two distinct stem/progenitor populations. One is the satellite cell, which acts as a muscle stem cell, and the other is the mesenchymal progenitor, which contributes to muscle pathogeneses such as fat infiltration and fibrosis. Detailed and accurate characterization of these progenitors in humans remains elusive. Here, we performed comprehensive cell-surface protein profiling of the two progenitor populations residing in human skeletal muscle and identified three previously unrecognized markers: CD82 and CD318 for satellite cells and CD201 for mesenchymal progenitors. These markers distinguish myogenic and mesenchymal progenitors, and enable efficient isolation of the two types of progenitors. Functional study revealed that CD82 ensures expansion and preservation of myogenic progenitors by suppressing excessive differentiation, and CD201 signaling favors adipogenesis of mesenchymal progenitors. Thus, cell-surface proteins identified here are not only useful markers but also functionally important molecules, and provide valuable insight into human muscle biology and diseases.

  18. SERS imaging of cell-surface biomolecules metabolically labeled with bioorthogonal Raman reporters. (United States)

    Xiao, Ming; Lin, Liang; Li, Zefan; Liu, Jie; Hong, Senlian; Li, Yaya; Zheng, Meiling; Duan, Xuanming; Chen, Xing


    Live imaging of biomolecules with high specificity and sensitivity as well as minimal perturbation is essential for studying cellular processes. Here, we report the development of a bioorthogonal surface-enhanced Raman scattering (SERS) imaging approach that exploits small Raman reporters for visualizing cell-surface biomolecules. The cells were cultured and imaged by SERS microscopy on arrays of Raman-enhancing nanoparticles coated on silicon wafers or glass slides. The Raman reporters including azides, alkynes, and carbondeuterium bonds are small in size and spectroscopically bioorthogonal (background-free). We demonstrated that various cell-surface biomolecules including proteins, glycans, and lipids were metabolically incorporated with the corresponding precursors bearing a Raman reporter and visualized by SERS microscopy. The coupling of SERS microscopy with bioorthogonal Raman reporters expands the capabilities of live-cell microscopy beyond the modalities of fluorescence and label-free imaging.

  19. [The influence of cell surface hydrophobicity Candida sp. on biofilm formation on different biomaterials]. (United States)

    Ciok-Pater, Emilia; Gospodarek, Eugenia; Prazyńska, Małgorzata; Bogiel, Tomasz


    The ability of yeasts to form biofilm is believed to play an important role in patomechanism of fungal infection. Candida sp. is considered to form biofilm on surfaces of biomaterials used in production of catheters, drains and prosthesis. Therefore this may lead to serious problems in patients with biomaterials used for diagnostic or therapeutic purposes. The aim of the study was to evaluate the influence of cell surface hydrophobicity (CSH) of Candida sp. on biofilm formation on different biomaterials. CSH was evaluated by two methods: Salt Aggregation Test (SAT) and Microbe Adhesion to Hydrocarbon Test (MATH). Biofilm formation on different biomaterials was measured by Richard's method after 72 hour incubation at 37 degrees C. Candida biofilm formation occurred more frequently in case of strains exhibiting hydrophobic than hydrophilic properties of cell surface. The statistically significant correlation between CSH and ability of biofilm formation on different biomaterials was observed (p < 0.05).

  20. Mammalian carboxylesterase (CES) releases GPI-anchored proteins from the cell surface upon lipid raft fluidization. (United States)

    Orihashi, Kaoru; Tojo, Hiromasa; Okawa, Katsuya; Tashima, Yuko; Morita, Takashi; Kondoh, Gen


    Mammalian carboxylesterase (CES) is well known as a biotransformation enzyme for prodrugs and xenobiotics. Here, we purified CES as a GPI-anchored protein (GPI-AP)-releasing factor (GPIase) that releases such protein from the cell surface. All five isoforms of CES showed this activity to various degrees. When the serine residue of the catalytic triad for esterase was replaced by alanine, esterase activity was completely disrupted, while full GPIase activity remained, suggesting that these two activities are exhibited via different mechanisms. CES6, a new class of mammalian CES, exhibited the highest GPIase activity and released specific GPI-APs from the cell surface after lipid raft fluidization. The released product contained a GPI component, indicating that GPI-AP was released by cleavage in GPI. These results revealed for the first time that CES recognizes and catalyzes macromolecule GPI-AP as well as small molecules.

  1. Lack of increased expression of cell surface markers for circulating fibrocyte progenitors in limited scleroderma. (United States)

    Russo, R; Medbury, H; Guiffre, A; Englert, H; Manolios, N


    The aetiology and pathogenesis of scleroderma is incompletely understood. Recently, a cell called the fibrocyte has been shown to be derived from circulating monocytes with the ability to produce collagen. The aim of this study was to evaluate differences in the cell surface characteristics of circulating fibrocyte progenitors (monocytes) in patients with limited scleroderma compared to controls. A case-control study was performed in eight patients with limited scleroderma, which were matched with eight controls. Three-colour flow cytometry was used to assess the relative expression of cell surface markers. Statistical analysis then compared the relative expression between the two groups. In this preliminary study, there were no significant differences in the expression of circulating monocyte surface molecules involved with cell transformation, function, or migration presumed to give rise to fibrocytes, in a population of patients with limited scleroderma. Various explanations for the results are discussed.

  2. Hydrogen-bonded LbL Shells for Living Cell Surface Engineering (United States)


    phospholipids into the lipid bilayer membrane, and cell decoration with/inclusion into biodegradable gel microparti- cles.7–11 However, for these strategies...polymer membrane. The poly(allylamine hydrochloride)/poly(styrene sulfonate) ( PAH /PSS) coating is the mostly explored poly- electrolyte pair used to...this approach for cell surface engineering.42,43 As suggested, overall toxicity of the PAH /PSS LbL shells originates from the positive charge of

  3. Revealing the fate of cell surface human P-glycoprotein (ABCB1): The Lysosomal Degradation Pathway (United States)

    Katayama, Kazuhiro; Kapoor, Khyati; Ohnuma, Shinobu; Patel, Atish; Swaim, William; Ambudkar, Indu S.; Ambudkar, Suresh V.


    P-glycoprotein (P-gp) transports a variety of chemically dissimilar amphipathic compounds including anticancer drugs. Although mechanisms of P-gp drug transport are widely studied, the pathways involving its internalization are poorly understood. The present study is aimed at elucidating the pathways involved in degradation of cell surface P-gp. The fate of P-gp at the cell surface was determined by biotinylating cell surface proteins followed by flow cytometry and Western blotting. Our data shows that the half-life of endogenously expressed P-gp is 26.7 ± 1.1 h in human colorectal cancer HCT-15 cells. Treatment of cells with Bafilomycin A1 (BafA1) a vacuolar H+ ATPase inhibitor increased the half-life of P-gp at the cell surface to 36.1± 0.5 h. Interestingly, treatment with the proteasomal inhibitors MG132, MG115 or lactacystin alone did not alter the half-life of the protein. When cells were treated with both lysosomal and proteasomal inhibitors (BafA1 and MG132), the half-life was further prolonged to 39-50 h. Functional assays done with rhodamine 123 or calcein-AM, fluorescent substrates of P-gp, indicated that the transport function of P-gp was not affected by either biotinylation or treatment with BafA1 or proteasomal inhibitors. Immunofluorescence studies done with the antibody against lysosomal marker LAMP1 and the P-gp-specific antibody UIC2 in permeabilized cells indicated that intracellular P-gp is primarily localized in the lysosomal compartment. Our results suggest that the lysosomal degradation system could be targeted to increase the sensitivity of P-gp expressing cancer cells towards chemotherapeutic drugs. PMID:26057472

  4. The Mesenchymal Precursor Cell Marker Antibody STRO-1 Binds to Cell Surface Heat Shock Cognate 70. (United States)

    Fitter, Stephen; Gronthos, Stan; Ooi, Soo Siang; Zannettino, Andrew C W


    Since its discovery more than 25 years ago, the STRO-1 antibody has played a fundamental role in defining the hierarchical nature of mesenchymal precursor cells (MPC) and their progeny. STRO-1 antibody binding remains a hallmark of immature pluripotent MPC. Despite the significance of STRO-1 in the MPC field, the identity of the antigen has remained elusive. Using a combination of two-dimensional gel electrophoresis, coupled with Western blotting and Tandem mass spectroscopy, we have identified the STRO-1 antigen as heat shock cognate 70 (HSC70;HSPA8). STRO-1 binds to immune-precipitated HSC70 and siRNA-mediated knock down of HSPA8 reduced STRO-1 binding. STRO-1 surface binding does not correlate with HSC70 expression and sequestration of cholesterol reduces STRO-1 surface binding, suggesting that the plasma membrane lipid composition may be an important determinant in the presentation of HSC70 on the cell surface. HSC70 is present on the surface of STRO-1(+) but not STRO-1(-) cell lines as assessed by cell surface biotinylation and recombinant HSC70 blocks STRO-1 binding to the cell surface. The STRO-1 epitope on HSC70 was mapped to the ATPase domain using a series of deletion mutants in combination with peptide arrays. Deletion of the first four amino acids of the consensus epitope negated STRO-1 binding. Notably, in addition to HSC70, STRO-1 cross-reacts with heat shock protein 70 (HSP70), however all the clonogenic cell activity is restricted to the STRO-1(BRIGHT) /HSP70(-) fraction. These results provide important insight into the properties that define multipotent MPC and provide the impetus to explore the role of cell surface HSC70 in MPC biology. Stem Cells 2016.

  5. Syndecans as cell surface receptors: Unique structure equates with functional diversity

    DEFF Research Database (Denmark)

    Choi, Youngsil; Chung, Heesung; Jung, Heyjung


    An increasing number of functions for syndecan cell surface heparan sulfate proteoglycans have been proposed over the last decade. Moreover, aberrant syndecan regulation has been found to play a critical role in multiple pathologies, including cancers, as well as wound healing and inflammation...... glycosaminoglycan chains, especially heparan sulfate. This heterodisperse polysaccharide has the potential to interact with many ligands from diverse protein families. Here, we relate the structural features of syndecans to some of their known functions....

  6. Revealing the fate of cell surface human P-glycoprotein (ABCB1): The lysosomal degradation pathway. (United States)

    Katayama, Kazuhiro; Kapoor, Khyati; Ohnuma, Shinobu; Patel, Atish; Swaim, William; Ambudkar, Indu S; Ambudkar, Suresh V


    P-glycoprotein (P-gp) transports a variety of chemically dissimilar amphipathic compounds including anticancer drugs. Although mechanisms of P-gp drug transport are widely studied, the pathways involving its internalization are poorly understood. The present study is aimed at elucidating the pathways involved in degradation of cell surface P-gp. The fate of P-gp at the cell surface was determined by biotinylating cell surface proteins followed by flow cytometry and Western blotting. Our data shows that the half-life of endogenously expressed P-gp is 26.7±1.1 h in human colorectal cancer HCT-15 cells. Treatment of cells with Bafilomycin A1 (BafA1) a vacuolar H+ ATPase inhibitor increased the half-life of P-gp at the cell surface to 36.1±0.5 h. Interestingly, treatment with the proteasomal inhibitors MG132, MG115 or lactacystin alone did not alter the half-life of the protein. When cells were treated with both lysosomal and proteasomal inhibitors (BafA1 and MG132), the half-life was further prolonged to 39-50 h. Functional assays done with rhodamine 123 or calcein-AM, fluorescent substrates of P-gp, indicated that the transport function of P-gp was not affected by either biotinylation or treatment with BafA1 or proteasomal inhibitors. Immunofluorescence studies done with the antibody against lysosomal marker LAMP1 and the P-gp-specific antibody UIC2 in permeabilized cells indicated that intracellular P-gp is primarily localized in the lysosomal compartment. Our results suggest that the lysosomal degradation system could be targeted to increase the sensitivity of P-gp- expressing cancer cells towards chemotherapeutic drugs.

  7. Modeling the Excess Cell Surface Stored in a Complex Morphology of Bleb-Like Protrusions (United States)

    Wessler, Timothy; Yang, Xiaofeng; Chen, Alex; Roach, Nathan; Elston, Timothy C.; Wang, Qi; Jacobson, Ken; Forest, M. Gregory


    Cells transition from spread to rounded morphologies in diverse physiological contexts including mitosis and mesenchymal-to-amoeboid transitions. When these drastic shape changes occur rapidly, cell volume and surface area are approximately conserved. Consequently, the rounded cells are suddenly presented with a several-fold excess of cell surface whose area far exceeds that of a smooth sphere enclosing the cell volume. This excess is stored in a population of bleb-like protrusions (BLiPs), whose size distribution is shown by electron micrographs to be skewed. We introduce three complementary models of rounded cell morphologies with a prescribed excess surface area. A 2D Hamiltonian model provides a mechanistic description of how discrete attachment points between the cell surface and cortex together with surface bending energy can generate a morphology that satisfies a prescribed excess area and BLiP number density. A 3D random seed-and-growth model simulates efficient packing of BLiPs over a primary rounded shape, demonstrating a pathway for skewed BLiP size distributions that recapitulate 3D morphologies. Finally, a phase field model (2D and 3D) posits energy-based constitutive laws for the cell membrane, nematic F-actin cortex, interior cytosol, and external aqueous medium. The cell surface is equipped with a spontaneous curvature function, a proxy for the cell surface-cortex couple, that is a priori unknown, which the model “learns” from the thin section transmission electron micrograph image (2D) or the “seed and growth” model image (3D). Converged phase field simulations predict self-consistent amplitudes and spatial localization of pressure and stress throughout the cell for any posited stationary morphology target and cell compartment constitutive properties. The models form a general framework for future studies of cell morphological dynamics in a variety of biological contexts. PMID:27015526

  8. Modeling the Excess Cell Surface Stored in a Complex Morphology of Bleb-Like Protrusions.

    Directory of Open Access Journals (Sweden)

    Maryna Kapustina


    Full Text Available Cells transition from spread to rounded morphologies in diverse physiological contexts including mitosis and mesenchymal-to-amoeboid transitions. When these drastic shape changes occur rapidly, cell volume and surface area are approximately conserved. Consequently, the rounded cells are suddenly presented with a several-fold excess of cell surface whose area far exceeds that of a smooth sphere enclosing the cell volume. This excess is stored in a population of bleb-like protrusions (BLiPs, whose size distribution is shown by electron micrographs to be skewed. We introduce three complementary models of rounded cell morphologies with a prescribed excess surface area. A 2D Hamiltonian model provides a mechanistic description of how discrete attachment points between the cell surface and cortex together with surface bending energy can generate a morphology that satisfies a prescribed excess area and BLiP number density. A 3D random seed-and-growth model simulates efficient packing of BLiPs over a primary rounded shape, demonstrating a pathway for skewed BLiP size distributions that recapitulate 3D morphologies. Finally, a phase field model (2D and 3D posits energy-based constitutive laws for the cell membrane, nematic F-actin cortex, interior cytosol, and external aqueous medium. The cell surface is equipped with a spontaneous curvature function, a proxy for the cell surface-cortex couple, that is a priori unknown, which the model "learns" from the thin section transmission electron micrograph image (2D or the "seed and growth" model image (3D. Converged phase field simulations predict self-consistent amplitudes and spatial localization of pressure and stress throughout the cell for any posited stationary morphology target and cell compartment constitutive properties. The models form a general framework for future studies of cell morphological dynamics in a variety of biological contexts.

  9. Proteomic analysis of cell surface-associated proteins from probiotic Lactobacillus plantarum

    DEFF Research Database (Denmark)

    Beck, Hans Christian; Madsen, Søren M; Glenting, Jacob;


    In the present study, we used a proteomic approach to identify surface-associated proteins from the probiotic bacterium Lactobacillus plantarum 299v. Proteins were extracted from the cell surface using a mild wash in phosphate buffer and analysed by sodium dodecyl sulphate-polyacrylamide gel...... of probiotics in the gastrointestinal tract. The results provide the basis for future studies on the molecular mechanisms of probiotics....

  10. Mesorhizobium loti Produces nodPQ-Dependent Sulfated Cell Surface Polysaccharides▿


    Townsend, Guy E.; Forsberg, Lennart S.; Keating, David H.


    Leguminous plants and bacteria from the family Rhizobiaceae form a symbiotic relationship, which culminates in novel plant structures called root nodules. The indeterminate symbiosis that forms between Sinorhizobium meliloti and alfalfa requires biosynthesis of Nod factor, a β-1,4-linked lipochitooligosaccharide that contains an essential 6-O-sulfate modification. S. meliloti also produces sulfated cell surface polysaccharides, such as lipopolysaccharide (LPS). The physiological function of s...

  11. Cell-surface protein-protein interaction analysis with time-resolved FRET and snap-tag technologies: application to G protein-coupled receptor oligomerization. (United States)

    Comps-Agrar, Laëtitia; Maurel, Damien; Rondard, Philippe; Pin, Jean-Philippe; Trinquet, Eric; Prézeau, Laurent


    G protein-coupled receptors (GPCRs) are key players in cell-cell communication, the dysregulation of which has often deleterious effects leading to pathologies such as psychiatric and neurological diseases. Consequently, GPCRs represent excellent drug targets, and as such are the object of intense research in drug discovery for therapeutic application. Recently, the GPCR field has been revolutionized by the demonstration that GPCRs are part of large protein complexes that control their pharmacology, activity, and signaling. Moreover, in these complexes, one GPCR can either associate with itself, forming homodimers or homooligomers, or with other receptor types, forming heterodimeric or heterooligomeric receptor entities that display new receptor features. These features include alterations in ligand cooperativity and selectivity, the activation of novel signaling pathways, and novel processes of desensitization. Thus, it has become necessary to identify GPCR-associated protein complexes of interest at the cell surface, and to determine the state of oligomerization of these receptors and their interactions with their partner proteins. This is essential to understand the function of GPCRs in their native environment, as well as ways to either modulate or control receptor activity with appropriate pharmacological tools, and to develop new therapeutic strategies. This requires the development of technologies to precisely address protein-protein interactions between oligomers at the cell surface. In collaboration with Cisbio Bioassay, we have developed such a technology, which combines TR-FRET detection with a new labeling method called SnapTag. This technology has allowed us to address the oligomeric state of many GPCRs.

  12. Measurement of interaction force between RGD-peptide and Hela cell surface by optical tweezers

    Institute of Scientific and Technical Information of China (English)

    Mincheng Zhong; Guosheng Xue; Jinhua Zhou; Ziqiang Wang; Yinmei Li


    Since RGD peptides (R:arginine; G:glycine; D:aspartic acid) are found to promote cell adhesion,they are modified at numerous materials surface for medical applications such as drug delivery and regenerative medicine.Peptide-cell surface interactions play a key role in the above applications.In this letter,we study the adhesion force between the RGD-coated bead and Hela cell surface by optical tweezes.The adhesion is dominated by the binding of α5β1 and RGD-peptide with higher adhesion probability and stronger adhesion strength compared with the adhesion of bare bead and cell surface.The binding force for a single α5β1-GRGDSP pair is determined to be 16.8 pN at a loading rate of 1.5 nN/s.The unstressed off-rate is 1.65 × 10-2 s-1 and the distance of transition state for the rigid binding model is 3.0 nm.

  13. Yeast cell surface display for lipase whole cell catalyst and its applications

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Yun; Zhang, Rui; Lian, Zhongshuai; Wang, Shihui; Wright, Aaron T.


    The cell surface display technique allows for the expression of target proteins or peptides on the microbial cell surface by fusing an appropriate protein as an anchoring motif. Yeast display systems, such as Pichia pastoris, Yarowia lipolytica and Saccharomyces cerevisiae, are ideal, alternative and extensive display systems with the advantage of simple genetic manipulation and post-translational modification of expressed heterologous proteins. Engineered yeasts show high performance characteristics and variant utilizations. Herein, we comprehensively summarize the variant factors affecting lipase whole cell catalyst activity and display efficiency, including the structure and size of target proteins, screening anchor proteins, type and chain length of linkers, and the appropriate matching rules among the above-mentioned display units. Furthermore, we also address novel approaches to enhance stability and activity of recombinant lipases, such as VHb gene co-expression, multi-enzyme co-display technique, and the micro-environmental interference and self-assembly techniques. Finally, we represent the variety of applications of whole cell surface displayed lipases on yeast cells in non-aqueous phases, including synthesis of esters, PUFA enrichment, resolution of chiral drugs, organic synthesis and biofuels. We demonstrate that the lipase surface display technique is a powerful tool for functionalizing yeasts to serve as whole cell catalysts, and increasing interest is providing an impetus for broad application of this technique.

  14. Role of TI-VAMP and CD82 in EGFR cell-surface dynamics and signaling. (United States)

    Danglot, Lydia; Chaineau, Mathilde; Dahan, Maxime; Gendron, Marie-Claude; Boggetto, Nicole; Perez, Franck; Galli, Thierry


    The v-SNARE TI-VAMP (VAMP7) mediates exocytosis during neuritogenesis, phagocytosis and lysosomal secretion. It localizes to endosomes and lysosomes but also to the trans-Golgi network. Here we show that depletion of TI-VAMP enhances the endocytosis of activated EGF receptor (EGFR) without affecting constitutive endocytosis of EGFR, or transferrin uptake. This increased EGFR internalization is mainly clathrin dependent. Searching for defects in EGFR regulators, we found that TI-VAMP depletion reduces the cell surface amount of CD82, a tetraspanin known to control EGFR localization in microdomains. We further show that TI-VAMP is required for secretion from the Golgi apparatus to the cell surface, and that TI-VAMP-positive vesicles transport CD82. Quantum dots video-microscopy indicates that depletion of TI-VAMP, or its cargo CD82, restrains EGFR diffusion and the area explored by EGFR at the cell surface. Both depletions also impair MAPK signaling and enhance endocytosis of activated EGFR by increased recruitment of AP-2. These results highlight the role of TI-VAMP in the secretory pathway of a tetraspanin, and support a model in which CD82 allows EGFR entry in microdomains that control its clathrin-dependent endocytosis and signaling.

  15. Brown spider venom toxins interact with cell surface and are endocytosed by rabbit endothelial cells. (United States)

    Nowatzki, Jenifer; de Sene, Reginaldo Vieira; Paludo, Katia Sabrina; Veiga, Silvio Sanches; Oliver, Constance; Jamur, Maria Célia; Nader, Helena Bonciani; Trindade, Edvaldo S; Franco, Célia Regina C


    Bites from the Loxosceles genus (brown spiders) cause severe clinical symptoms, including dermonecrotic injury, hemorrhage, hemolysis, platelet aggregation and renal failure. Histological findings of dermonecrotic lesions in animals exposed to Loxosceles intermedia venom show numerous vascular alterations. Study of the hemorrhagic consequences of the venom in endothelial cells has demonstrated that the degeneration of blood vessels results not only from degradation of the extracellular matrix molecule or massive leukocyte infiltration, but also from a direct and primary activity of the venom on endothelial cells. Exposure of an endothelial cell line in vitro to L. intermedia venom induce morphological alterations, such as cell retraction and disadhesion to the extracellular matrix. The aim of the present study was to investigate the interaction between the venom toxins and the endothelial cell surface and their possible internalization, in order to illuminate the information about the deleterious effect triggered by venom. After treating endothelial cells with venom toxins, we observed that the venom interacts with cell surface. Venom treatment also can cause a reduction of cell surface glycoconjugates. When cells were permeabilized, it was possible to verify that some venom toxins were internalized by the endothelial cells. The venom internalization involves endocytic vesicles and the venom was detected in the lysosomes. However, no damage to lysosomal integrity was observed, suggesting that the cytotoxic effect evoked by L. intermedia venom on endothelial cells is not mediated by venom internalization.

  16. Selective radiolabeling of cell surface proteins to a high specific activity

    Energy Technology Data Exchange (ETDEWEB)

    Thompson, J.A.; Lau, A.L.; Cunningham, D.D.


    A procedure was developed for selective radiolabeling of membrane proteins on cells to higher specific activities than possible with available techniques. Cell surface amino groups were derivatized with /sup 125/I-(hydroxyphenyl)propionyl groups via /sup 125/I-sulfosuccinimidyl (hydroxyphenyl)propionate (/sup 125/II-sulfo-SHPP). This reagent preferentially labeled membrane proteins exposed at the cell surface of erythrocytes as assessed by the degree of radiolabel incorporation into erythrocyte ghost proteins and hemoglobin. Comparison with the lactoperoxidase-(/sup 125/I)iodide labeling technique revealed that /sup 125/I-sulfo-SHPP labeled cell surface proteins to a much higher specific activity and hemoglobin to a much lower specific activity. Additionally, this reagent was used for selective radiolabeling of membrane proteins on the cytoplasmic face of the plasma membrane by blocking exofacial amino groups with uniodinated sulfo-SHPP, lysing the cells, and then incubating them with /sup 125/I-sulfo-SHPP. Exclusive labeling of either side of the plasma membrane was demonstrated by the labeling of some marker proteins with well-defined spacial orientations on erythroctyes. Transmembrane proteins such as the epidermal growth factor receptor on cultured cells could also be labeled differentially from either side of the plasma membrane.

  17. Giant sacral meningeal diverticulum containing a thickened filum with lipoma in an adult with spinal cord tethering. Case report and review of the literature. (United States)

    Feigenbaum, Frank


    An unusual case of a patient with a giant intrasacral meningeal diverticulum and spinal cord tethering with a thickened filum is presented. Instead of being empty as is typical, the meningeal diverticulum in this case contained a segment of the thickened tethering filum, which entered from the thecal sac through an ostium. A search of the literature revealed no prior description of a meningeal diverticulum containing a portion of tethered filum or any other structure. Only 2 previous cases of intrasacral meningeal cyst and spinal cord tethering with a thickened filum were found, both in the non-English literature.

  18. Modulation of cell surface transport and lipid raft localization by the cytoplasmic tail of the influenza virus hemagglutinin. (United States)

    Scolari, Silvia; Imkeller, Katharina; Jolmes, Fabian; Veit, Michael; Herrmann, Andreas; Schwarzer, Roland


    Viral glycoproteins are highly variable in their primary structure, but on the other hand feature a high functional conservation to fulfil their versatile tasks during the pathogenic life cycle. Typically, all protein domains are optimized in that indispensable functions can be assigned to small conserved motifs or even individual amino acids. The cytoplasmic tail of many viral spike proteins, although of particular relevance for the virus biology, is often only insufficiently characterized. Hemagglutinin (HA), the receptor-binding protein of the influenza virus comprises a short cytoplasmic tail of 13 amino acids that exhibits three highly conserved palmitoylation sites. However, the particular importance of these modifications and the tail in general for intracellular trafficking and lateral membrane organization remains elusive. In this study, we generated HA core proteins consisting of transmembrane domain, cytoplasmic tail and a minor part of the ectodomain, tagged with a yellow fluorescent protein. Different mutation and truncation variants of these chimeric proteins were investigated using confocal microscopy, to characterize the role of cytoplasmic tail and palmitoylation for the intracellular trafficking to plasma membrane and Golgi apparatus. In addition, we assessed raft partitioning of the variants by Foerster resonance energy transfer with an established raft marker. We revealed a substantial influence of the cytoplasmic tail length on the intracellular distribution and surface exposure of the proteins. A complete removal of the tail hampers a physiological trafficking of the protein, whereas a partial truncation can be compensated by cytoplasmic palmitoylations. Plasma membrane raft partitioning on the other hand was found to imperatively require palmitoylations, and the cysteine at position 551 turned out to be of most relevance. Our data shed further light on the tight interconnection between cytoplasmic elements and intracellular trafficking and suggest a function of HA palmitoylations in both lateral sorting and anterograde trafficking of the glycoprotein.

  19. Cell surface glycan alterations in epithelial mesenchymal transition process of Huh7 hepatocellular carcinoma cell.

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    Shan Li

    Full Text Available BACKGROUND AND OBJECTIVE: Due to recurrence and metastasis, the mortality of Hepatocellular carcinoma (HCC is high. It is well known that the epithelial mesenchymal transition (EMT and glycan of cell surface glycoproteins play pivotal roles in tumor metastasis. The goal of this study was to identify HCC metastasis related differential glycan pattern and their enzymatic basis using a HGF induced EMT model. METHODOLOGY: HGF was used to induce HCC EMT model. Lectin microarray was used to detect the expression of cell surface glycan and the difference was validated by lectin blot and fluorescence cell lectin-immunochemistry. The mRNA expression levels of glycotransferases were determined by qRT-PCR. RESULTS: After HGF treatment, the Huh7 cell lost epithelial characteristics and obtained mesenchymal markers. These changes demonstrated that HGF could induce a typical cell model of EMT. Lectin microarray analysis identified a decreased affinity in seven lectins ACL, BPL, JAC, MPL, PHA-E, SNA, and SBA to the glycan of cell surface glycoproteins. This implied that glycan containing T/Tn-antigen, NA2 and bisecting GlcNAc, Siaα2-6Gal/GalNAc, terminal α or βGalNAc structures were reduced. The binding ability of thirteen lectins, AAL, LCA, LTL, ConA, NML, NPL, DBA, HAL, PTL II, WFL, ECL, GSL II and PHA-L to glycan were elevated, and a definite indication that glycan containing terminal αFuc and ± Sia-Le, core fucose, α-man, gal-β(α GalNAc, β1,6 GlcNAc branching and tetraantennary complex oligosaccharides structures were increased. These results were further validated by lectin blot and fluorescence cell lectin-immunochemistry. Furthermore, the mRNA expression level of Mgat3 decreased while that of Mgat5, FucT8 and β3GalT5 increased. Therefore, cell surface glycan alterations in the EMT process may coincide with the expression of glycosyltransferase. CONCLUSIONS: The findings of this study systematically clarify the alterations of cell surface

  20. Discovery of dual-action membrane-anchored modulators of incretin receptors.

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    Jean-Philippe Fortin

    Full Text Available BACKGROUND: The glucose-dependent insulinotropic polypeptide (GIP and the glucagon-like peptide-1 (GLP-1 receptors are considered complementary therapeutic targets for type 2 diabetes. Using recombinant membrane-tethered ligand (MTL technology, the present study focused on defining optimized modulators of these receptors, as well as exploring how local anchoring influences soluble peptide function. METHODOLOGY/PRINCIPAL FINDINGS: Serial substitution of residue 7 in membrane-tethered GIP (tGIP led to a wide range of activities at the GIP receptor, with [G(7]tGIP showing enhanced efficacy compared to the wild type construct. In contrast, introduction of G(7 into the related ligands, tGLP-1 and tethered exendin-4 (tEXE4, did not affect signaling at the cognate GLP-1 receptor. Both soluble and tethered GIP and GLP-1 were selective activators of their respective receptors. Although soluble EXE4 is highly selective for the GLP-1 receptor, unexpectedly, tethered EXE4 was found to be a potent activator of both the GLP-1 and GIP receptors. Diverging from the pharmacological properties of soluble and tethered GIP, the newly identified GIP-R agonists, (i.e. [G(7]tGIP and tEXE4 failed to trigger cognate receptor endocytosis. In an attempt to recapitulate the dual agonism observed with tEXE4, we conjugated soluble EXE4 to a lipid moiety. Not only did this soluble peptide activate both the GLP-1 and GIP receptors but, when added to receptor expressing cells, the activity persists despite serial washes. CONCLUSIONS: These findings suggest that conversion of a recombinant MTL to a soluble membrane anchored equivalent offers a means to prolong ligand function, as well as to design agonists that can simultaneously act on more than one therapeutic target.

  1. An AFM-based pit-measuring method for indirect measurements of cell-surface membrane vesicles. (United States)

    Zhang, Xiaojun; Chen, Yuan; Chen, Yong


    Circulating membrane vesicles, which are shed from many cell types, have multiple functions and have been correlated with many diseases. Although circulating membrane vesicles have been extensively characterized, the status of cell-surface membrane vesicles prior to their release is less understood due to the lack of effective measurement methods. Recently, as a powerful, micro- or nano-scale imaging tool, atomic force microscopy (AFM) has been applied in measuring circulating membrane vesicles. However, it seems very difficult for AFM to directly image/identify and measure cell-bound membrane vesicles due to the similarity of surface morphology between membrane vesicles and cell surfaces. Therefore, until now no AFM studies on cell-surface membrane vesicles have been reported. In this study, we found that air drying can induce the transformation of most cell-surface membrane vesicles into pits that are more readily detectable by AFM. Based on this, we developed an AFM-based pit-measuring method and, for the first time, used AFM to indirectly measure cell-surface membrane vesicles on cultured endothelial cells. Using this approach, we observed and quantitatively measured at least two populations of cell-surface membrane vesicles, a nanoscale population (AFM-based pit-measuring method is potentially useful for studying cell-surface membrane vesicles and for investigating the mechanisms of membrane vesicle formation/release.

  2. Membrane tethering by the atlastin GTPase depends on GTP hydrolysis but not on forming the cross-over configuration. (United States)

    Saini, Simran G; Liu, Chuang; Zhang, Peijun; Lee, Tina H


    The membrane-anchored atlastin GTPase couples nucleotide hydrolysis to the catalysis of homotypic membrane fusion to form a branched endoplasmic reticulum network. Trans dimerization between atlastins anchored in opposing membranes, accompanied by a cross-over conformational change, is thought to draw the membranes together for fusion. Previous studies on the conformational coupling of atlastin to its GTP hydrolysis cycle have been carried out largely on atlastins lacking a membrane anchor. Consequently, whether fusion involves a discrete tethering step and, if so, the potential role of GTP hydrolysis and cross-over in tethering remain unknown. In this study, we used membrane-anchored atlastins in assays that separate tethering from fusion to dissect the requirements for each. We found that tethering depended on GTP hydrolysis, but, unlike fusion, it did not depend on cross-over. Thus GTP hydrolysis initiates stable head-domain contact in trans to tether opposing membranes, whereas cross-over formation plays a more pivotal role in powering the lipid rearrangements for fusion.

  3. Role of Double-Strand Break End-Tethering during Gene Conversion in Saccharomyces cerevisiae.

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    Suvi Jain


    Full Text Available Correct repair of DNA double-strand breaks (DSBs is critical for maintaining genome stability. Whereas gene conversion (GC-mediated repair is mostly error-free, repair by break-induced replication (BIR is associated with non-reciprocal translocations and loss of heterozygosity. We have previously shown that a Recombination Execution Checkpoint (REC mediates this competition by preventing the BIR pathway from acting on DSBs that can be repaired by GC. Here, we asked if the REC can also determine whether the ends that are engaged in a GC-compatible configuration belong to the same break, since repair involving ends from different breaks will produce potentially deleterious translocations. We report that the kinetics of repair are markedly delayed when the two DSB ends that participate in GC belong to different DSBs (termed Trans compared to the case when both DSB ends come from the same break (Cis. However, repair in Trans still occurs by GC rather than BIR, and the overall efficiency of repair is comparable. Hence, the REC is not sensitive to the "origin" of the DSB ends. When the homologous ends for GC are in Trans, the delay in repair appears to reflect their tethering to sequences on the other side of the DSB that themselves recombine with other genomic locations with which they share sequence homology. These data support previous observations that the two ends of a DSB are usually tethered to each other and that this tethering facilitates both ends encountering the same donor sequence. We also found that the presence of homeologous/repetitive sequences in the vicinity of a DSB can distract the DSB end from finding its bona fide homologous donor, and that inhibition of GC by such homeologous sequences is markedly increased upon deleting Sgs1 but not Msh6.

  4. Tethered Balloon Operations at ARM AMF3 Site at Oliktok Point, AK (United States)

    Dexheimer, D.; Lucero, D. A.; Helsel, F.; Hardesty, J.; Ivey, M.


    Oliktok Point has been the home of the Atmospheric Radiation Measurement Program's (ARM) third ARM Mobile Facility, or AMF3, since October 2013. The AMF3 is operated through Sandia National Laboratories and hosts instrumentation collecting continuous measurements of clouds, aerosols, precipitation, energy, and other meteorological variables. The Arctic region is warming more quickly than any other region due to climate change and Arctic sea ice is declining to record lows. Sparsity of atmospheric data from the Arctic leads to uncertainty in process comprehension, and atmospheric general circulation models (AGCM) are understood to underestimate low cloud presence in the Arctic. Increased vertical resolution of meteorological properties and cloud measurements will improve process understanding and help AGCMs better characterize Arctic clouds. SNL is developing a tethered balloon system capable of regular operation at AMF3 in order to provide increased vertical resolution atmospheric data. The tethered balloon can be operated within clouds at altitudes up to 7,000' AGL within DOE's R-2204 restricted area. Pressure, relative humidity, temperature, wind speed, and wind direction are recorded at multiple altitudes along the tether. These data were validated against stationary met tower data in Albuquerque, NM. The altitudes of the sensors were determined by GPS and calculated using a line counter and clinometer and compared. Wireless wetness sensors and supercooled liquid water content sensors have also been deployed and their data has been compared with other sensors. This presentation will provide an overview of the balloons, sensors, and test flights flown, and will provide a preliminary look at data from sensor validation campaigns and test flights.

  5. Compact, planar, translational piezoelectric bimorph actuator with Archimedes’ spiral actuating tethers (United States)

    Yang, Chenye; Liu, Sanwei; Xie, Xin; Livermore, Carol


    The design, analytical modelling, finite element analysis (FEA), and experimental characterization of a microelectromechanical system (MEMS) out-of-plane (vertical) translational piezoelectric lead-zirconate-titanate (PZT) bimorph actuator supported on Archimedes’ spiral tethers are presented. Three types of bimorph actuators with different electrode patterns (with spiral tethers half actuated, fully actuated with uniform polarity, or fully actuated with reversed polarity) are designed and modelled. The two actuators with the highest predicted performance (half actuated and fully actuated with uniform polarity) are implemented and characterized. Both designs are fabricated by commercial processes and are compatible with integration into more complex MEMS systems. Analytical modelling and FEA are used to analyze and predict the actuators’ displacements and blocking forces. Experimental measurements of the deflections and blocking forces of actuators with full uniform actuation and half actuation validate the design. At an applied voltage of 110 V, the out-of-plane deflections of the actuators with half actuation and full uniform actuation are measured at about 17 µm and 29 µm respectively, in good agreement with analytical predictions of 17.3 µm and 34.2 µm and FEA predictions of 17.1 µm and 25.8 µm. The blocking force for devices with half-actuated tethers is predicted to be 12 mN (analytical) and 10 mN (FEA), close to the experimental value of 9 mN. The blocking force for devices with full uniform actuation is predicted to be 23 mN (analytical) and 17 mN (FEA), as compared with 15 mN in experiments.

  6. Tethered capsule OCT endomicroscopy: from bench to bedside at the primary care office (Conference Presentation) (United States)

    Gora, Michalina J.; Simmons, Leigh H.; Tiernan, Aubrey R.; Grant, Catriona N.; Soomro, Amna R.; Walker Corkery, Elizabeth S.; Rosenberg, Mireille; Metlay, Joshua P.; Tearney, Guillermo J.


    We have developed a swallowable tethered capsule OCT endomicroscopy (TCE) device that acquires microscopic images of the entire esophagus in unsedated subjects in a quick and comfortable procedure. To test its capabilities of TCE to become a population-based screening device, we conducted a clinical feasibility study in the primary care office. The swept-source OCT imaging system (1310nm central wavelength, 40kHz A-line rate, 10um axial resolution) together with the tethered capsule catheter (11x25mm capsule attached to a flexible tether) were transferred to the PCP office where unsedated patients scheduled for non-urgent PCP visits swallowed the capsule and microscopic OCT images of the entire esophagus were collected. After the whole length of the esophagus was imaged, the catheter was disinfected for reuse. Twenty subjects were enrolled in the study, including nine female and eleven male. All TCE procedures were performed by a nurse and lasted in average 5:42 ± 1:54 min. High-resolution images of the esophagus were obtained in all seventeen subjects that swallowed the capsule. Our clinical experience in this cohort, subject feedback, image quality, and technological adaptations for efficient utilization in this setting will be presented. The ease and simplicity of the procedure combined with high quality of the images demonstrate the potential for this technology to become a population-based screening device. Technology limitations and future development guided by findings from this initial experience will be discussed with the goal of effectively translating TCE to the outpatient primary care setting.

  7. Specific Measurement of Tethered Running Kinetics and its Relationship to Repeated Sprint Ability

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    Sousa Filipe


    Full Text Available Repeated sprint ability has been widely studied by researchers, however, analysis of the relationship between most kinetic variables and the effect of fatigue is still an ongoing process. To search for the best biomechanical parameter to evaluate repeated sprint ability, several kinetic variables were measured in a tethered field running test and compared regarding their sensitivity to fatigue and correlation with time trials in a free running condition. Nine male sprint runners (best average times: 100 m = 10.45 ± 0.07 s; 200 m = 21.36 ± 0.17 s; 400 m = 47.35 ± 1.09 s completed two test sessions on a synthetic track. Each session consisted of six 35 m sprints interspersed by 10 s rest under tethered field running or free running conditions. Force, power, work, an impulse and a rate of force development were all directly measured using the sensors of a new tethered running apparatus, and a one-way ANOVA with Scheffé post-hoc test used to verify differences between sprints (p < 0.05. Pearson product-moment correlation measured the relationship between mechanical variables and free running performance. A total impulse, the rate of force development and maximum force did not show significant differences for most sprints. These three variables presented low to moderate correlations with free running performance (r between 0.01 and -0.35. Maximum and mean power presented the strongest correlations with free running performance (r = -0.71 and -0.76, respectively; p < 0.001, followed by mean force (r = -0.61; p < 0.001 and total work (r = -0.50; p < 0.001. It was concluded that under a severe work-to-rest ratio condition, power variables were better suited to evaluating repeated sprint ability than the other studied variables.

  8. Numerical Modeling of a Spar Platform Tethered by a Mooring Cable

    Institute of Scientific and Technical Information of China (English)

    ZHU Xiangqian; YOO Wan-Suk


    Virtual simulation is an economical and efficient method in mechanical system design. Numerical modeling of a spar platform, tethered by a mooring cable with a spherical joint is developed for the dynamic simulation of the floating structure in ocean. The geometry modeling of the spar is created using finite element methods. The submerged part of the spar bears the buoyancy, hydrodynamic drag force, and effect of the added mass and Froude-Krylov force. Strip theory is used to sum up the forces acting on the elements. The geometry modeling of the cable is established based on the lumped-mass-and-spring modeling through which the cable is divided into 10 elements. A new element-fixed local frame is used, which is created by the element orientation vector and relative velocity of the fluid, to express the loads acting on the cable. The bottom of the cable is fixed on the seabed by spring forces, while the top of the cable is connected to the bottom of the spar platform by a modified spherical joint. This system suffers the propagating wave and current in the X-direction and the linear wave theory is applied for setting of the propagating wave. Based on the numerical modeling, the displacement-load relationships are analyzed, and the simulation results of the numerical modeling are compared with those by the commercial simulation code, ProteusDS. The comparison indicates that the numerical modeling of the spar platform tethered by a mooring cable is well developed, which provides an instruction for the optimization of a floating structure tethered by a mooring cable system.

  9. Lipid Raft-Mediated Membrane Tethering and Delivery of Hydrophobic Cargos from Liquid Crystal-Based Nanocarriers. (United States)

    Nag, Okhil K; Naciri, Jawad; Oh, Eunkeu; Spillmann, Christopher M; Delehanty, James B


    A main goal of bionanotechnology and nanoparticle (NP)-mediated drug delivery (NMDD) continues to be the development of novel biomaterials that can controllably modulate the activity of the NP-associated therapeutic cargo. One of the desired subcellular locations for targeted delivery in NMDD is the plasma membrane. However, the controlled delivery of hydrophobic cargos to the membrane bilayer poses significant challenges including cargo precipitation and lack of specificity. Here, we employ a liquid crystal NP (LCNP)-based delivery system for the controlled partitioning of a model dye cargo from within the NP core into the plasma membrane bilayer. During synthesis of the NPs, the water-insoluble model dye cargo, 3,3'-dioctadecyloxacarbocyanine perchlorate (DiO), was efficiently incorporated into the hydrophobic LCNP core as confirmed by multiple spectroscopic analyses. Conjugation of a PEGylated cholesterol derivative to the NP surface (DiO-LCNP-PEG-Chol) facilitated the localization of the dye-loaded NPs to lipid raft microdomains in the plasma membrane in HEK 293T/17 cell. Analysis of DiO cellular internalization kinetics revealed that when delivered as a LCNP-PEG-Chol NP, the half-life of DiO membrane residence time (30 min) was twice that of free DiO (DiO(free)) (15 min) delivered from bulk solution. Time-resolved laser scanning confocal microscopy was employed to visualize the passive efflux of DiO from the LCNP core and its insertion into the plasma membrane bilayer as confirmed by Förster resonance energy transfer (FRET) imaging. Finally, the delivery of DiO as a LCNP-PEG-Chol complex resulted in the attenuation of its cytotoxicity; the NP form of DiO exhibited ∼30-40% less toxicity compared to DiO(free). Our data demonstrate the utility of the LCNP platform as an efficient vehicle for the combined membrane-targeted delivery and physicochemical modulation of molecular cargos using lipid raft-mediated tethering.

  10. Complex phase behavior of a fluid in slits with semipermeable walls modified with tethered chains. (United States)

    Borówko, M; Patrykiejew, A; Rżysko, W; Sokołowski, S; Ilnytskyi, J


    We study the phase behavior of a two-component fluid in a pore with the walls modified by tethered chains. The walls are completely permeable for one component of the fluid and completely impenetrable for the second component. The fluid is perfectly mixed in a bulk phase. We have found that depending on the details of the model the fluid undergoes capillary condensation inside the pore and wetting and layering transitions at the outer walls. Moreover, we have found transitions connected with the change of symmetry of the distribution of chains and fluid inside the pore.

  11. A modular phosphate tether-mediated divergent strategy to complex polyols

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    Paul R. Hanson


    Full Text Available An efficient and divergent synthesis of polyol subunits utilizing a phosphate tether-mediated, one-pot, sequential RCM/CM/reduction process is reported. A modular, 3-component coupling strategy has been developed, in which, simple “order of addition” of a pair of olefinic-alcohol components to a pseudo-C2-symmetric phosphoryl chloride, coupled with the RCM/CM/reduction protocol, yields five polyol fragments. Each of the product polyols bears a central 1,3-anti-diol subunit with differential olefinic geometries at the periphery.

  12. Use of Tethered Enzymes as a Platform Technology for Rapid Analyte Detection.

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    Roy Cohen

    Full Text Available Rapid diagnosis for time-sensitive illnesses such as stroke, cardiac arrest, and septic shock is essential for successful treatment. Much attention has therefore focused on new strategies for rapid and objective diagnosis, such as Point-of-Care Tests (PoCT for blood biomarkers. Here we use a biomimicry-based approach to demonstrate a new diagnostic platform, based on enzymes tethered to nanoparticles (NPs. As proof of principle, we use oriented immobilization of pyruvate kinase (PK and luciferase (Luc on silica NPs to achieve rapid and sensitive detection of neuron-specific enolase (NSE, a clinically relevant biomarker for multiple diseases ranging from acute brain injuries to lung cancer. We hypothesize that an approach capitalizing on the speed and catalytic nature of enzymatic reactions would enable fast and sensitive biomarker detection, suitable for PoCT devices.We performed in-vitro, animal model, and human subject studies. First, the efficiency of coupled enzyme activities when tethered to NPs versus when in solution was tested, demonstrating a highly sensitive and rapid detection of physiological and pathological concentrations of NSE. Next, in rat stroke models the enzyme-based assay was able in minutes to show a statistically significant increase in NSE levels in samples taken 1 hour before and 0, 1, 3 and 6 hours after occlusion of the distal middle cerebral artery. Finally, using the tethered enzyme assay for detection of NSE in samples from 20 geriatric human patients, we show that our data match well (r = 0.815 with the current gold standard for biomarker detection, ELISA-with a major difference being that we achieve detection in 10 minutes as opposed to the several hours required for traditional ELISA.Oriented enzyme immobilization conferred more efficient coupled activity, and thus higher assay sensitivity, than non-tethered enzymes. Together, our findings provide proof of concept for using oriented immobilization of active

  13. Cost-effective synthesis of amine-tethered porous materials for carbon capture. (United States)

    Lu, Weigang; Bosch, Mathieu; Yuan, Daqiang; Zhou, Hong-Cai


    A truly cost-effective strategy for the synthesis of amine-tethered porous polymer networks (PPNs) has been developed. A network containing diethylenetriamine (PPN-125-DETA) exhibits a high working capacity comparable to current state-of-art technology (30 % monoethanolamine solutions), yet it requires only one third as much energy for regeneration. It has also been demonstrated to retain over 90 % capacity after 50 adsorption-desorption cycles of CO2 in a temperature-swing adsorption process. The results suggest that PPN-125-DETA is a very promising new material for carbon capture from flue gas streams.

  14. Investigation of EM Emissions by the Electrodynamic Tether, Inclusive of an Observational Program (EMET) (United States)

    Estes, Robert D.


    Our TSS-1/R investigation, which we shall refer to as EMET in this report, was an integral part of the effort by the TSS-1/R Investigators' Working Group (IWG) to come to an understanding of the complex interaction between the tethered satellite system and the ionosphere. All of the space-borne experiments were designed to collect data relevant to the local interaction. Only the ground- based experiments, EMET and its Italian counterpart Observations on the Earth's Surface of Electromagnetic Emissions (OESEE), held out any hope of characterizing the long range effects of the interaction. This was to be done by detecting electromagnetic waves generated by the system in the ionosphere, assuming the signal reached the Earth's surface with sufficient amplitude. As the type of plasma waves excited to carry charge away from the charge-exchange regions of the system at each end of the tether is one of the theoretical points about which there is greatest disagreement, a definitive identification of tether-generated waves could mark significant progress in the so-called current closure problem of electrodynamic tethers. Dr. Mario Grossi of the Smithsonian Astrophysical Observatory (SAO) initiated the investigation, and his experience in the field of ULF-ELF waves and their detection was invaluable throughout its course. Rice University had the responsibility of setting up the EMET ULF-VLF ground stations under a subcontract from SAO. Principal Investigator (PI) for the Rice effort was Prof. William E. Gordon, who was primary observer at the Arecibo Observatory during TSS-LR. Dr. Steve Noble handled major day-to-day operations, training, and planning for the ground-based measurements. Dr. James McCoy of NASA JSC, a member of the Mona/Arecibo team, was pilot for the numerous flights ferrying personnel and equipment between Puerto Rico and Mona Island. Final responsibility for the measurements rested with SAO, and the activities of field personnel and SAO investigators were

  15. Efficient display of active lipase LipB52 with a Pichia pastoris cell surface display system and comparison with the LipB52 displayed on Saccharomyces cerevisiae cell surface

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    Ma Yushu


    Full Text Available Abstract Background For industrial bioconversion processes, the utilization of surface-displayed lipase in the form of whole-cell biocatalysts is more advantageous, because the enzymes are displayed on the cell surface spontaneously, regarded as immobilized enzymes. Results Two Pichia pastoris cell surface display vectors based on the flocculation functional domain of FLO with its own secretion signal sequence or the α-factor secretion signal sequence were constructed respectively. The lipase gene lipB52 fused with the FLO gene was successfully transformed into Pichia pastoris KM71. The lipase LipB52 was expressed under the control of the AOX1 promoter and displayed on Pichia pastoris KM71 cell surface with the two Pichia pastoris cell surface display vectors. Localization of the displayed LipB52 on the cell surface was confirmed by the confocal laser scanning microscopy (CLSM. The LipB52 displayed on the Pichia pastoris cell surface exhibited activity toward p-nitrophenol ester with carbon chain length ranging from C10 to C18, and the optimum substrate was p-nitrophenol-caprate (C10, which was consistent with it displayed on the Saccharomyces cerevisiae EBY100 cell surface. The hydrolysis activity of lipase LipB52 displayed on Pichia pastoris KM71-pLHJ047 and KM71-pLHJ048 cell surface reached 94 and 91 U/g dry cell, respectively. The optimum temperature of the displayed lipases was 40°C at pH8.0, they retained over 90% activity after incubation at 60°C for 2 hours at pH 7.0, and still retained 85% activity after incubation for 3 hours. Conclusion The LipB52 displayed on the Pichia pastoris cell surface exhibited better stability than the lipase LipB52 displayed on Saccharomyces cerevisiae cell surface. The displayed lipases exhibited similar transesterification activity. But the Pichia pastoris dry cell weight per liter (DCW/L ferment culture was about 5 times than Saccharomyces cerevisiae, the lipase displayed on Pichia pastoris are more

  16. Cell-surface area codes: mobile-element related gene switches generate precise and heritable cell-surface displays of address molecules that are used for constructing embryos. (United States)

    Dreyer, W J; Roman-Dreyer, J


    We present an updated area code hypothesis supporting the proposal that cell surface display of seven-transmembrane olfactory receptors, protocadherins and other cell surface receptors provide codes that enable cells to find their correct partners as they sculpture embryos. The genetic mechanisms that program the expression of such displays have been largely unknown until very recently. However, increasing evidence now suggests that precise developmental control of the expression of these genes during embryogenesis is achieved in part by permanent and heritable changes in DNA. Using the developing immune system as a model, we discuss two different types of developmentally programmed genetic switches, each of which relies on recombination mechanisms related to mobile elements. We review new evidence suggesting the involvement of mobile element related switch mechanisms in the generation of protocadherin molecules, and their possible involvement in the control of expressions of olfactory receptors. As both recombinase and reverse transcriptase mechanisms play a role in the switching of the immunoglobulin genes, we searched the databases of expressed sequence tags (dbEST) for expression of related genes in other tissues. We present data revealing that transposases and reverse transcriptases are widely expressed in most tissues. We also searched these databases for expression of env (envelope) gene products, stimulated by provocative results suggesting that these molecules might function as cellular address receptors. We found that env genes are also expressed in large numbers in normal human tissues. One must assume that these three different types of mobile-element-related messenger RNA molecules (transposases, reverse transcriptases, and env proteins) are expressed for use in functions of value in the various tissues and have been preserved in the genome because of their selective advantages. We conclude that it is possible that many specific cell lineage decisions

  17. Tannerella forsythia strains display different cell-surface nonulosonic acids: biosynthetic pathway characterization and first insight into biological implications. (United States)

    Friedrich, Valentin; Janesch, Bettina; Windwarder, Markus; Maresch, Daniel; Braun, Matthias L; Megson, Zoë A; Vinogradov, Evgeny; Goneau, Marie-France; Sharma, Ashu; Altmann, Friedrich; Messner, Paul; Schoenhofen, Ian C; Schäffer, Christina


    Tannerella forsythia is an anaerobic, Gram-negative periodontal pathogen. A unique O-linked oligosaccharide decorates the bacterium's cell surface proteins and was shown to modulate the host immune response. In our study, we investigated the biosynthesis of the nonulosonic acid (NulO) present at the terminal position of this glycan. A bioinformatic analysis of T. forsythia genomes revealed a gene locus for the synthesis of pseudaminic acid (Pse) in the type strain ATCC 43037 while strains FDC 92A2 and UB4 possess a locus for the synthesis of legionaminic acid (Leg) instead. In contrast to the NulO in ATCC 43037, which has been previously identified as a Pse derivative (5-N-acetimidoyl-7-N-glyceroyl-3,5,7,9-tetradeoxy-l-glycero-l-manno-NulO), glycan analysis of strain UB4 performed in this study indicated a 350-Da, possibly N-glycolyl Leg (3,5,7,9-tetradeoxy-d-glycero-d-galacto-NulO) derivative with unknown C5,7 N-acyl moieties. We have expressed, purified and characterized enzymes of both NulO pathways to confirm these genes' functions. Using capillary electrophoresis (CE), CE-mass spectrometry and NMR spectroscopy, our studies revealed that Pse biosynthesis in ATCC 43037 essentially follows the UDP-sugar route described in Helicobacter pylori, while the pathway in strain FDC 92A2 corresponds to Leg biosynthesis in Campylobacter jejuni involving GDP-sugar intermediates. To demonstrate that the NulO biosynthesis enzymes are functional in vivo, we created knockout mutants resulting in glycans lacking the respective NulO. Compared to the wild-type strains, the mutants exhibited significantly reduced biofilm formation on mucin-coated surfaces, suggestive of their involvement in host-pathogen interactions or host survival. This study contributes to understanding possible biological roles of bacterial NulOs.

  18. Structural Characterization of Tip20p and Dsl1p, Subunits of the Dsl1p Vesicle Tethering Complex

    Energy Technology Data Exchange (ETDEWEB)

    Tripathi, A.; Ren, Y; Jeffrey, P; Hughson, F


    Multisubunit tethering complexes are essential for intracellular trafficking and have been proposed to mediate the initial interaction between vesicles and the membranes with which they fuse. Here we report initial structural characterization of the Dsl1p complex, whose three subunits are essential for trafficking from the Golgi apparatus to the endoplasmic reticulum (ER). Crystal structures reveal that two of the three subunits, Tip20p and Dsl1p, resemble known subunits of the exocyst complex, establishing a structural connection among several multisubunit tethering complexes and implying that many of their subunits are derived from a common progenitor. We show, moreover, that Tip20p and Dsl1p interact directly via N-terminal alpha-helices. Finally, we establish that different Dsl1p complex subunits bind independently to different ER SNARE proteins. Our results map out two alternative protein-interaction networks capable of tethering COPI-coated vesicles, via the Dsl1p complex, to ER membranes.

  19. Structure of the Membrane-tethering GRASP Domain Reveals a Unique PDZ Ligand Interaction That Mediates Golgi Biogenesis

    Energy Technology Data Exchange (ETDEWEB)

    S Truschel; D Sengupta; A Foote; A Heroux; M Macbeth; A Linstedt


    Biogenesis of the ribbon-like membrane network of the mammalian Golgi requires membrane tethering by the conserved GRASP domain in GRASP65 and GRASP55, yet the tethering mechanism is not fully understood. Here, we report the crystal structure of the GRASP55 GRASP domain, which revealed an unusual arrangement of two tandem PDZ folds that more closely resemble prokaryotic PDZ domains. Biochemical and functional data indicated that the interaction between the ligand-binding pocket of PDZ1 and an internal ligand on PDZ2 mediates the GRASP self-interaction, and structural analyses suggest that this occurs via a unique mode of internal PDZ ligand recognition. Our data uncover the structural basis for ligand specificity and provide insight into the mechanism of GRASP-dependent membrane tethering of analogous Golgi cisternae.

  20. Structure of the Membrane-tethering GRASP Domain Reveals a Unique PDZ Ligand Interaction That Mediates Golgi Biogenesis

    Energy Technology Data Exchange (ETDEWEB)

    Truschel, S.T.; Heroux, A.; Sengupta, D.; Foote, A.; Macbeth, M. R.; Linstedt, A. D.


    Biogenesis of the ribbon-like membrane network of the mammalian Golgi requires membrane tethering by the conserved GRASP domain in GRASP65 and GRASP55, yet the tethering mechanism is not fully understood. Here, we report the crystal structure of the GRASP55 GRASP domain, which revealed an unusual arrangement of two tandem PDZ folds that more closely resemble prokaryotic PDZ domains. Biochemical and functional data indicated that the interaction between the ligand-binding pocket of PDZ1 and an internal ligand on PDZ2 mediates the GRASP self-interaction, and structural analyses suggest that this occurs via a unique mode of internal PDZ ligand recognition. Our data uncover the structural basis for ligand specificity and provide insight into the mechanism of GRASP-dependent membrane tethering of analogous Golgi cisternae.

  1. Cell surface alpha 2,6 sialylation affects adhesion of breast carcinoma cells. (United States)

    Lin, Shaoqiang; Kemmner, Wolfgang; Grigull, Sabine; Schlag, Peter M


    Tumor-associated alterations of cell surface glycosylation play a crucial role in the adhesion and metastasis of carcinoma cells. The aim of this study was to examine the effect of alpha 2,6-sialylation on the adhesion properties of breast carcinoma cells. To this end mammary carcinoma cells, MDA-MB-435, were sense-transfected with sialyltransferase ST6Gal-I cDNA or antisense-transfected with a part of the ST6Gal-I sequence. Sense transfectants showed an enhanced ST6Gal-I mRNA expression and enzyme activity and an increased binding of the lectin Sambucus nigra agglutinin (SNA), specific for alpha 2,6-linked sialic acid. Transfection with ST6Gal-I in the antisense direction resulted in less enzyme activity and SNA reactivity. A sense-transfected clone carrying increased amounts of alpha 2,6-linked sialic acid adhered preferentially to collagen IV and showed reduced cell-cell adhesion and enhanced invasion capacity. In contrast, antisense transfection led to less collagen IV adhesion but enhanced homotypic cell-cell adhesion. In another approach, inhibition of ST6Gal-I enzyme activity by application of soluble antisense-oligodeoxynucleotides was studied. Antisense treatment resulted in reduced ST6 mRNA expression and cell surface 2,6-sialylation and significantly decreased collagen IV adhesion. Our results suggest that cell surface alpha 2,6-sialylation contributes to cell-cell and cell-extracellular matrix adhesion of tumor cells. Inhibition of sialytransferase ST6Gal-I by antisense-oligodeoxynucleotides might be a way to reduce the metastatic capacity of carcinoma cells.

  2. Aged mice have increased inflammatory monocyte concentration and altered expression of cell-surface functional receptors

    Indian Academy of Sciences (India)

    Kelley Strohacker; Whitney L Breslin; Katie C Carpenter; Brian K McFarlin


    The expression of monocyte cell-surface receptors represents one index of immune dysfunction, which is common with aging. Although mouse models of aging are prevalent, monocyte subset assessment is rare. Our purpose was to compare cell receptor expression on classic (CD115+/Gr-1high) and non-classic (CD115+/Gr-1low) monocytes from 80- or 20-week-old CD-1 mice. Three-colour flow cytometry was used to determine the concentration of monocyte subsets and their respective cell-surface expression of TLR2, TLR4, CD80, CD86, MHC II and CD54. These receptors were selected because they have been previously associated with altered monocyte function. Data were analysed with independent -tests; significance was set at < 0.05. Old mice had a greater concentration of both classic (258%, =0.003) and non-classic (70%, =0.026) monocytes. The classic : non-classic monocyte ratio doubled in old as compared with that in young mice (=0.006), indicating a pro-inflammatory shift. TLR4 ($\\downarrow$27%, =0.001) and CD80 ($\\downarrow$37%, =0.004) were decreased on classic monocytes from old as compared with those from young mice. TLR2 ($\\uparrow$24%, =0.002) and MHCII ($\\downarrow$21%, =0.026) were altered on non-classic monocytes from old as compared with those from young mice. The increased classic : non-classic monocyte ratio combined with changes in the cell-surface receptor expression on both monocyte subsets is indicative of immune dysfunction, which may increase age-associated disease risk.

  3. Modification of cell surface properties of Pseudomonas alcaligenes S22 during hydrocarbon biodegradation. (United States)

    Kaczorek, Ewa; Moszyńska, Sylwia; Olszanowski, Andrzej


    Biodegradation of water insoluble hydrocarbons can be significantly increased by the addition of natural surfactants one. Very promising option is the use of saponins. The obtained results indicated that in this system, after 21 days, 92% biodegradation of diesel oil could be achieved using Pseudomonas alcaligenes. No positive effect on the biodegradation process was observed using synthetic surfactant Triton X-100. The kind of carbon source influences the cell surface properties of microorganisms. Modification of the surface cell could be observed by control of the sedimentation profile. This analytical method is a new approach in microbiological analysis.

  4. Analysis of cell surface alterations in Legionella pneumophila cells treated with human apolipoprotein E. (United States)

    Palusinska-Szysz, Marta; Zdybicka-Barabas, Agnieszka; Cytryńska, Małgorzata; Wdowiak-Wróbel, Sylwia; Chmiel, Elżbieta; Gruszecki, Wiesław I


    Binding of human apolipoprotein E (apoE) to Legionella pneumophila lipopolysaccharide was analysed at the molecular level by Fourier-transform infrared spectroscopy, thereby providing biophysical evidence for apoE-L. pneumophila lipopolysaccharide interaction. Atomic force microscopy imaging of apoE-exposed L. pneumophila cells revealed alterations in the bacterial cell surface topography and nanomechanical properties in comparison with control bacteria. The changes induced by apoE binding to lipopolysaccharide on the surface of L. pneumophila cells may participate in: (1) impeding the penetration of host cells by the bacteria; (2) suppression of pathogen intracellular growth and eventually; and (3) inhibition of the development of infection.


    Institute of Scientific and Technical Information of China (English)

    WANG Xiao-mei; SHAN Jun; CHEN Zhuo-huai


    Aim: To investigate glycoconjugate changes on the cell surface of proliferative lesions and neoplasms of mice lungs at various stages of tumorigenesis, the relation between progressive development of mouse pulmonary tumors and expression of cell surface saccharide. Materials and methods: Thirty - one male A/J strain mice at 5 weeks of age were treated intraperitoneally with a single injection of 20 - methylcholanthrene (20 - MC), 292 pulmonary lesions including 31 hyperplasias, 145 alveolar adenomas, 61 papillary adenomas, 55 papillary adenocarcinomas and their combined type were obtained. The binding affinities of cells in normal respiratory epithelia and in proliferative lesions to four peroxidases - conjugated lectins, Maclura pomifera agglutinin (MPA), Arachis hypogea agglutinin (PNA), Ricinus communis agglutinin (RCA), and wheat germ agglutinin (WGA) were examined. Results: Cells of hyperplasia and alveolar adenoma showed fairly strong affinity to all the four lectins. However, part of papillary adenoma cells and greater part of papillary adenocarcinoma cells lost their binding affinity to MPA, PNA, and RCA, but not to WGA. The bindings of MPA, PNA and RNA were detected predominently on the luminal surfaces of benign tumors but not on the luminal surfaces of malignant tumors. WGA might bind to varied types of benign and malignant tumors. Pretreated with neuraminidase, the lesions enhanced the staining intensity for the four lectins, the binding sites of WGA to malignant tumor cells were numerous. A distinct difference in lectin binding affinity between hyperplasia / alveolar adenoma/papillary adenoma and papillary adenocarcinoma was clearly shown( x2 = 46.89, P < 0.01, x2 = 36.77, P < 0.01 and x2 = 52.87, P < 0.01 ) in this experiment. The complex glycoconjugates on the cell surface of malignant and benign lesions during the development of pulmonary tumor were changed,malignant tumor cells differed from the surface of benign tumor cells, the levels of

  6. Regulation of Kv2.1 K+ conductance by cell surface channel density


    Fox, Philip D.; Loftus, Rob J.; Tamkun, Michael M.


    The Kv2.1 voltage-gated K+ channel is found both freely diffusing over the plasma membrane and concentrated in micron-sized clusters localized to the soma, proximal dendrites and axon initial segment of hippocampal neurons. In transfected HEK cells, Kv2.1 channels within cluster microdomains are non-conducting. Using TIRF microscopy the number of GFP-tagged Kv2.1 channels on the HEK cell surface was compared to K+ channel conductance measured by whole-cell voltage-clamp of the same cell. This...

  7. Inhibition of experimental ascending urinary tract infection by an epithelial cell-surface receptor analogue (United States)

    Edén, C. Svanborg; Freter, R.; Hagberg, L.; Hull, R.; Hull, S.; Leffler, H.; Schoolnik, G.


    It has been shown that the establishment of urinary tract infection by Escherichia coli is dependent on attachment of the bacteria to epithelial cells1-4. The attachment involves specific epithelial cell receptors, which have been characterized as glycolipids5-10. Reversible binding to cell-surface mannosides may also be important4,11-13. This suggests an approach to the treatment of infections-that of blocking bacterial attachment with cell membrane receptor analogues. Using E. coli mutants lacking one or other of the two binding specificities (glycolipid and mannose), we show here that glycolipid analogues can block in vitro adhesion and in vivo urinary tract infection.

  8. Keynote Paper: Cell-Surface Adhesive Interactions in Microchannels and Microvessels

    CERN Document Server

    King, M R


    Adhesive interactions between white blood cells and the interior surface of the blood vessels they contact is important in inflammation and in the progression of heart disease. Parallel-plate microchannels have been useful in characterizing the strength of these interactions, in conditions that are much simplified over the complex environment these cells experience in the body. Recent computational and experimental work by several laboratories have attempted to bridge this gap between behavior observed in flow chamber experiments, and cell-surface interactions observed in the microvessels of anesthetized animals.

  9. Glycoprotein mucin molecular brush on cancer cell surface acting as mechanical barrier against drug delivery (United States)

    Wang, Xin; Shah, Aalok A.; Campbell, Robert B.; Wan, Kai-tak


    Uptake of cytotoxic drugs by typical tumor cells is limited by the dense dendritic network of oligosaccharide mucin chains that forms a mechanical barrier. Atomic force microscopy is used to directly measure the force needed to pierce the mucin layer to reach the cell surface. Measurements are analyzed by de Gennes' steric reptation theory. Multidrug resistant ovarian tumor cells shows significantly larger penetration load compared to the wide type. A pool of pancreatic, lung, colorectal, and breast cells are also characterized. The chemotherapeutic agent, benzyl-α-GalNac, for inhibiting glycosylation is shown to be effective in reducing the mechanical barrier.

  10. Shuttling of the autoantigen La between nucleus and cell surface after uv irradiation of human keratinocytes

    Energy Technology Data Exchange (ETDEWEB)

    Bachmann, M.; Chang, S.; Slor, H.; Kukulies, J.; Mueller, W.E. (Universitaet, Mainz (Germany, F.R.))


    During the past years we have established that the nuclear autoantigen La shuttles between the nucleus and the cytoplasm in tumor cells after inhibition of transcription or virus infection. We reinvestigated this shuttling using primary human keratinocytes from both healthy donors and patients with xeroderma pigmentosum. Ultraviolet irradiation resulted in both an inhibition of transcription and a translocation of La protein from the nucleus to the cytoplasm. After a prolonged inhibition of transcription La protein relocated into the nucleus and assembled with nuclear storage regions. The uv-induced shuttling included a translocation to the cell surface, where La protein colocalized with epidermal growth factor receptors.

  11. -Regular Modules

    Directory of Open Access Journals (Sweden)

    Areej M. Abduldaim


    Full Text Available We introduced and studied -regular modules as a generalization of -regular rings to modules as well as regular modules (in the sense of Fieldhouse. An -module is called -regular if for each and , there exist and a positive integer such that . The notion of -pure submodules was introduced to generalize pure submodules and proved that an -module is -regular if and only if every submodule of is -pure iff   is a -regular -module for each maximal ideal of . Many characterizations and properties of -regular modules were given. An -module is -regular iff is a -regular ring for each iff is a -regular ring for finitely generated module . If is a -regular module, then .

  12. CD133 protein N-glycosylation processing contributes to cell surface recognition of the primitive cell marker AC133 epitope. (United States)

    Mak, Anthony B; Blakely, Kim M; Williams, Rashida A; Penttilä, Pier-Andrée; Shukalyuk, Andrey I; Osman, Khan T; Kasimer, Dahlia; Ketela, Troy; Moffat, Jason


    The AC133 epitope expressed on the CD133 glycoprotein has been widely used as a cell surface marker of numerous stem cell and cancer stem cell types. It has been recently proposed that posttranslational modification and regulation of CD133 may govern cell surface AC133 recognition. Therefore, we performed a large scale pooled RNA interference (RNAi) screen to identify genes involved in cell surface AC133 expression. Gene hits could be validated at a rate of 70.5% in a secondary assay using an orthogonal RNAi system, demonstrating that our primary RNAi screen served as a powerful genetic screening approach. Within the list of hits from the primary screen, genes involved in N-glycan biosynthesis were significantly enriched as determined by Ingenuity Canonical Pathway analyses. Indeed, inhibiting biosynthesis of the N-glycan precursor using the small molecule tunicamycin or inhibiting its transfer to CD133 by generating N-glycan-deficient CD133 mutants resulted in undetectable cell surface AC133. Among the screen hits involved in N-glycosylation were genes involved in complex N-glycan processing, including the poorly characterized MGAT4C, which we demonstrate to be a positive regulator of cell surface AC133 expression. Our study identifies a set of genes involved in CD133 N-glycosylation as a direct contributing factor to cell surface AC133 recognition and provides biochemical evidence for the function and structure of CD133 N-glycans.

  13. Measuring graft stability in a tethered polyelectrolyte film by X-ray and neutron reflectivity (United States)

    Dimitriou, Michael; Galvin, Casey; Satija, Sushil; Genzer, Jan


    The instability of tethered polymer films in mild conditions has recently brought into question the limits of use of such layers in certain technologies, such as anti-fouling coatings. In order to better understand the process of chain degrafting in a polymer brush, we have used X-ray reflectivity (XR) and neutron reflectivity (NR) to examine tethered layers of poly(2-dimethylaminoethyl methacrylate) (PDMAEMA). Exposing an ~ 30 nm thick film of PDMAEMA brushes grafted on flat silica-coated substrates to a range of relative humidities (RH) resulted in reproducible thickness changes as measured by XR, illustrating the need of ambient solvent to induce degrafting. The thickness change showed non-linear behavior, increasing rapidly above ~ 70% RH and swelling to ~ 230% of its original thickness at ~ 99% RH. In order to better understand the apparent diffusive process of vapor into the brush, we have exposed brushes to isotopically labeled vapors. Using XR and NR, we examined the extent of modification in scattering length density within the brush using box and gradient models, and discuss the apparent entropic and enthalpic forces at play. We also conducted in situ aqueous measurements of similar samples to comprehend the degrafting process of a polymer brush. Through an appropriate choice of model, we detect variations in grafting density as a function of incubation time.

  14. Anti-cancer activity of novel dibenzo[b,f]azepine tethered isoxazoline derivatives

    Directory of Open Access Journals (Sweden)

    Sadashiva Maralinganadoddi


    Full Text Available Background Dibenzoazepine (DB derivatives are important and valuable compounds in medicinal chemistry. The synthesis and chemotherapeutic properties of naturally occurring DBs and different heterocyclic moiety tethered DBs are reported. Herein, we report the DB-fused hybrid structure that containing isoxazolines (DBIs and their anti-cancer activity, which could throw light on the structural and functional features of new molecules. Results and Conclusion The synthesis and characterization of novel ring DB tethered isoxazoline derivatives (DBIs were carried out. After the detailed structural characterization using 2D-NMR experiments, the compounds were identified as 5-substituted isoxazolines. The effect of newly synthesized DBIs against the invasion of murine osteosarcoma (LM8G7 cells was studied. Among the tested molecules, compound 4g (5-[−3-(4-chlorophenyl-4,5-dihydroisoxazol-5-yl-methyl]-5 H-dibenzo[b,f]azepine, was found to inhibit the invasion of LM8G7 cells strongly, when compared to other structurally related compounds. Cumulatively, the compound 4g inhibited the invasion MDA-MB-231 cells completely at 10 μM. In addition to anti-invasion property the compound 4g also inhibited the migration of LM8G7 and human ovarian cancer cells (OVSAHO dose-dependently. Compound 4g inhibited the proliferation of LM8G7, OVSAHO, human breast cancer cells (MCF-7 and human melphalan-resistant multiple myeloma (RPMI8226-LR5 cells that are comparable to cisplatin and suramin.

  15. Cholesterol-Induced Buckling in Physisorbed Polymer-Tethered Lipid Monolayers

    Directory of Open Access Journals (Sweden)

    Christoph A. Naumann


    Full Text Available The influence of cholesterol concentration on the formation of buckling structures is studied in a physisorbed polymer-tethered lipid monolayer system using epifluorescence microscopy (EPI and atomic force microscopy (AFM. The monolayer system, built using the Langmuir-Blodgett (LB technique, consists of 3 mol % poly(ethylene glycol (PEG lipopolymers and various concentrations of the phospholipid, 1-stearoyl-2-oleoyl-sn-glycero-3-phosphocholine (SOPC, and cholesterol (CHOL. In the absence of CHOL, AFM micrographs show only occasional buckling structures, which is caused by the presence of the lipopolymers in the monolayer. In contrast, a gradual increase of CHOL concentration in the range of 0–40 mol % leads to fascinating film stress relaxation phenomena in the form of enhanced membrane buckling. Buckling structures are moderately deficient in CHOL, but do not cause any notable phospholipid-lipopolymer phase separation. Our experiments demonstrate that membrane buckling in physisorbed polymer-tethered membranes can be controlled through CHOL-mediated adjustment of membrane elastic properties. They further show that CHOL may have a notable impact on molecular confinement in the presence of crowding agents, such as lipopolymers. Our results are significant, because they offer an intriguing prospective on the role of CHOL on the material properties in complex membrane architecture.

  16. Evidence for Solar Tether-cutting Magnetic Reconnection from Coronal Field Extrapolations

    CERN Document Server

    Liu, Chang; Lee, Jeongwoo; Wiegelmann, Thomas; Moore, Ronald L; Wang, Haimin


    Magnetic reconnection is one of the primary mechanisms for triggering solar eruptive events, but direct observation of its rapid process has been of challenge. In this Letter we present, using a nonlinear force-free field (NLFFF) extrapolation technique, a visualization of field line connectivity changes resulting from tether-cutting reconnection over about 30 minutes during the 2011 February 13 M6.6 flare in NOAA AR 11158. Evidence for the tether-cutting reconnection was first collected through multiwavelength observations and then by the analysis of the field lines traced from positions of four conspicuous flare 1700 A footpoints observed at the event onset. Right before the flare, the four footpoints are located very close to the regions of local maxima of magnetic twist index. Especially, the field lines from the inner two footpoints form two strongly twisted flux bundles (up to ~1.2 turns), which shear past each other and reach out close to the outer two footpoints, respectively. Immediately after the fl...

  17. Binary Mixed Homopolymer Brushes Tethered to Cellulose Nanocrystals: A Step Towards Compatibilized Polyester Blends. (United States)

    Mincheva, Rosica; Jasmani, Latifah; Josse, Thomas; Paint, Yoann; Raquez, Jena-Marie; Gerbaux, Pascal; Eyley, Samuel; Thielemans, Wim; Dubois, Philippe


    This article reports on the successful preparation and characterization of cellulose nanocrystals (CNCs) surface-modified with polylactide (PLA) and poly(butylene succinate) (PBS) binary mixed homopolymer brushes. Their synthesis was designed as a three-step procedure combining polyester synthesis and surface-modification of CNCs with simultaneous polyester grafting via a heterogeneous copper(I)-catalyzed azide-alkyne cycloaddition reaction. For comparison, single homopolymer brushes tethered to CNCs (PLLA-g-CNC and PBSBDEMPAM-g-CNC) were obtained applying the same procedure. The hairy nanoparticles were characterized in terms of chemical composition and thermal properties. Spectroscopic analyses suggested "rippled" microphase separation of both immiscible homopolyesters in the mixed brushes, while others showed impeded homopolyester crystallization after surface-grafting. A synergistic relationship between the polyesters and CNCs was also suggested, i.e., the polyester grafting increases the CNC thermal resistance, while CNC presence imparts char formation. The as-obtained binary homopolymer brushes tethered to nanoparticles makes these surface-modified cellulosic nanomaterials attractive as compatibilization/reinforcement agents for PLA/PBS blends.

  18. A Simple Flight Mill for the Study of Tethered Flight in Insects. (United States)

    Attisano, Alfredo; Murphy, James T; Vickers, Andrew; Moore, Patricia J


    Flight in insects can be long-range migratory flights, intermediate-range dispersal flights, or short-range host-seeking flights. Previous studies have shown that flight mills are valuable tools for the experimental study of insect flight behavior, allowing researchers to examine how factors such as age, host plants, or population source can influence an insects' propensity to disperse. Flight mills allow researchers to measure components of flight such as speed and distance flown. Lack of detailed information about how to build such a device can make their construction appear to be prohibitively complex. We present a simple and relatively inexpensive flight mill for the study of tethered flight in insects. Experimental insects can be tethered with non-toxic adhesives and revolve around an axis by means of a very low friction magnetic bearing. The mill is designed for the study of flight in controlled conditions as it can be used inside an incubator or environmental chamber. The strongest points are the very simple electronic circuitry, the design that allows sixteen insects to fly simultaneously allowing the collection and analysis of a large number of samples in a short time and the potential to use the device in a very limited workspace. This design is extremely flexible, and we have adjusted the mill to accommodate different species of insects of various sizes.

  19. Critical reappraisal confirms that Mitofusin 2 is an endoplasmic reticulum–mitochondria tether (United States)

    Naon, Deborah; Zaninello, Marta; Giacomello, Marta; Varanita, Tatiana; Grespi, Francesca; Lakshminaranayan, Sowmya; Serafini, Annalisa; Semenzato, Martina; Herkenne, Stephanie; Hernández-Alvarez, Maria Isabel; Zorzano, Antonio; De Stefani, Diego; Dorn, Gerald W.; Scorrano, Luca


    The discovery of the multiple roles of mitochondria–endoplasmic reticulum (ER) juxtaposition in cell biology often relied upon the exploitation of Mitofusin (Mfn) 2 as an ER–mitochondria tether. However, this established Mfn2 function was recently questioned, calling for a critical re-evaluation of Mfn2’s role in ER–mitochondria cross-talk. Electron microscopy and fluorescence-based probes of organelle proximity confirmed that ER–mitochondria juxtaposition was reduced by constitutive or acute Mfn2 deletion. Functionally, mitochondrial uptake of Ca2+ released from the ER was reduced following acute Mfn2 ablation, as well as in Mfn2−/− cells overexpressing the mitochondrial calcium uniporter. Mitochondrial Ca2+ uptake rate and extent were normal in isolated Mfn2−/− liver mitochondria, consistent with the finding that acute or chronic Mfn2 ablation or overexpression did not alter mitochondrial calcium uniporter complex component levels. Hence, Mfn2 stands as a bona fide ER–mitochondria tether whose ablation decreases interorganellar juxtaposition and communication. PMID:27647893

  20. CO2 capture and conversion with a multifunctional polyethyleneimine-tethered iminophosphine iridium catalyst/adsorbent. (United States)

    McNamara, Nicholas D; Hicks, Jason C


    Tunable, multifunctional materials able to capture CO2 and subsequently catalyze its conversion to formic acid were synthesized by the modification of branched polyethyleneimine (PEI) with an iminophosphine ligand coordinated to an Ir precatalyst. The molecular weight of the PEI backbone was an important component for material stability and catalytic activity, which were inversely related. The amine functionalities on PEI served three roles: 1) primary amines were used to tether the ligand and precatalyst, 2) amines were used to capture CO2 , and 3) amines served as a base for formate stabilization during catalysis. Ligand studies on imine and phosphine based ligands showed that a bidentate iminophosphine ligand resulted in the highest catalytic activity. X-ray photoelectron spectroscopy revealed that an increase in Ir 4f binding energy led to an increase in catalytic activity, which suggests that the electronics of the metal center play a significant role in catalysis. Catalyst loading studies revealed that there is a critical balance between free amines and ligand-metal sites that must be reached to optimize catalytic activity. Thus, it was found that the CO2 capture and conversion abilities of these materials could be optimized for reaction conditions by tuning the structure of the PEI-tethered materials.

  1. Fractional order sliding mode control for tethered satellite deployment with disturbances (United States)

    Kang, Junjie; Zhu, Zheng H.; Wang, Wei; Li, Aijun; Wang, Changqing


    This paper proposes a fractional order sliding mode control for the deployment of tethered space systems with the consideration of uncertainty of external disturbances and unmodeled system dynamics. The proposed fractional order sliding mode control consists of two sub-sliding manifolds that are defined separately for the actuated and unactuated states. This, in turn, generates a control scheme to make all states move toward to the desired states. The stability analysis of the proposed control law indicates not only all states converge to the desired states at equilibrium but also disturbances caused by the uncertainty can be suppressed satisfactorily. Parametric studies are conducted to investigate the influences of fractional order and sub-sliding manifold of unactuated states on the performance of the proposed control law. The performance is compared with the sliding mode, PD and fractional order PD control laws for a baseline scenario of tether deployment. The proposed control law performs better than others in the settling time and the maximum pitch angle control in the presence of unwanted disturbances. Effectiveness and robustness of the proposed control law are demonstrated by computer simulations.

  2. Tethered Satellites as Enabling Platforms for an Operational Space Weather Monitoring System (United States)

    Krause, L. Habash; Gilchrist, B. E.; Bilen, S.; Owens, J.; Voronka, N.; Furhop, K.


    Space weather nowcasting and forecasting models require assimilation of near-real time (NRT) space environment data to improve the precision and accuracy of operational products. Typically, these models begin with a climatological model to provide "most probable distributions" of environmental parameters as a function of time and space. The process of NRT data assimilation gently pulls the climate model closer toward the observed state (e.g. via Kalman smoothing) for nowcasting, and forecasting is achieved through a set of iterative physics-based forward-prediction calculations. The issue of required space weather observatories to meet the spatial and temporal requirements of these models is a complex one, and we do not address that with this poster. Instead, we present some examples of how tethered satellites can be used to address the shortfalls in our ability to measure critical environmental parameters necessary to drive these space weather models. Examples include very long baseline electric field measurements, magnetized ionospheric conductivity measurements, and the ability to separate temporal from spatial irregularities in environmental parameters. Tethered satellite functional requirements will be presented for each space weather parameter considered in this study.

  3. Comparison of standard, prone and cine MRI in the evaluation of tethered cord

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    Singh, Sukhjinder [Cohen Children' s Medical Center, Department of Radiology, New Hyde Park, NY (United States); Kline-Fath, Beth; Racadio, Judy M. [Cincinnati Children' s Hospital Medical Center, Department of Radiology, Cincinnati, OH (United States); Bierbrauer, Karin [Cincinnati Children' s Hospital Medical Center, Department of Neurosurgery, Cincinnati, OH (United States); Salisbury, Shelia; Macaluso, Maurizio [Cincinnati Children' s Hospital Medical Center, Division of Biostatistics and Epidemiology, Cincinnati, OH (United States); Jackson, Elizabeth C. [Cincinnati Children' s Hospital Medical Center, Department of Pediatrics, Division of Nephrology, Cincinnati, OH (United States); Egelhoff, John C. [Cincinnati Children' s Hospital Medical Center, Department of Radiology, Cincinnati, OH (United States); Phoenix Children' s Hospital, Department of Radiology, Phoenix, AZ (United States)


    Tethered cord syndrome (TCS) is defined by abnormal traction on the spinal cord that confines its movement. Surgical cord release usually stops neurological deterioration; therefore, early and accurate neuroradiological diagnosis is important. Supine MRI is the imaging modality of choice, but prone MRI and cine MRI can demonstrate cord movement. We compared the diagnostic accuracies of standard MRI, prone MRI and cine MRI in patients with clinical suspicion of TCS and evaluated inter-reader reliability for MR imaging. Children who underwent MRI for suspicion of TCS were retrospectively identified. Supine, prone and cine MRI studies were re-read by two pediatric neuroradiologists. Conus level, filum appearance and cord movement were documented. Thirteen of 49 children had tethered cord documented at surgery. Conus level had the highest diagnostic accuracy (sensitivity 69-77%, specificity 94%, positive predictive value 82-83%, negative predictive value 89-92%, correct diagnosis 88-90%) and highest between-reader concordance (98%). Prone and cine MRI did not add to the accuracy of the supine imaging. Conus level provides the highest diagnostic accuracy and inter-reader reliability in TCS. Until a larger series is evaluated, it remains questionable whether prone or cine MRI provides enough additional diagnostic information to warrant routine use. (orig.)

  4. Relative Contribution of Arms and Legs in 30 s Fully Tethered Front Crawl Swimming

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    Pedro G. Morouço


    Full Text Available The relative contribution of arm stroke and leg kicking to maximal fully tethered front crawl swimming performance remains to be solved. Twenty-three national level young swimmers (12 male and 11 female randomly performed 3 bouts of 30 s fully tethered swimming (using the whole body, only the arm stroke, and only the leg kicking. A load-cell system permitted the continuous measurement of the exerted forces, and swimming velocity was calculated from the time taken to complete a 50 m front crawl swim. As expected, with no restrictions swimmers were able to exert higher forces than that using only their arm stroke or leg kicking. Estimated relative contributions of arm stroke and leg kicking were 70.3% versus 29.7% for males and 66.6% versus 33.4% for females, with 15.6% and 13.1% force deficits, respectively. To obtain higher velocities, male swimmers are highly dependent on the maximum forces they can exert with the arm stroke (r=0.77, P<0.01, whereas female swimmers swimming velocity is more related to whole-body mean forces (r=0.81, P<0.01. The obtained results point that leg kicking plays an important role over short duration high intensity bouts and that the used methodology may be useful to identify strength and/or coordination flaws.

  5. ATP-driven Rad50 conformations regulate DNA tethering, end resection, and ATM checkpoint signaling. (United States)

    Deshpande, Rajashree A; Williams, Gareth J; Limbo, Oliver; Williams, R Scott; Kuhnlein, Jeff; Lee, Ji-Hoon; Classen, Scott; Guenther, Grant; Russell, Paul; Tainer, John A; Paull, Tanya T


    The Mre11-Rad50 complex is highly conserved, yet the mechanisms by which Rad50 ATP-driven states regulate the sensing, processing and signaling of DNA double-strand breaks are largely unknown. Here we design structure-based mutations in Pyrococcus furiosus Rad50 to alter protein core plasticity and residues undergoing ATP-driven movements within the catalytic domains. With this strategy we identify Rad50 separation-of-function mutants that either promote or destabilize the ATP-bound state. Crystal structures, X-ray scattering, biochemical assays, and functional analyses of mutant PfRad50 complexes show that the ATP-induced 'closed' conformation promotes DNA end binding and end tethering, while hydrolysis-induced opening is essential for DNA resection. Reducing the stability of the ATP-bound state impairs DNA repair and Tel1 (ATM) checkpoint signaling in Schizosaccharomyces pombe, double-strand break resection in Saccharomyces cerevisiae, and ATM activation by human Mre11-Rad50-Nbs1 in vitro, supporting the generality of the P. furiosus Rad50 structure-based mutational analyses. These collective results suggest that ATP-dependent Rad50 conformations switch the Mre11-Rad50 complex between DNA tethering, ATM signaling, and 5' strand resection, revealing molecular mechanisms regulating responses to DNA double-strand breaks.

  6. A high throughput method for quantification of cell surface bound and internalized chitosan nanoparticles. (United States)

    Tammam, Salma N; Azzazy, Hassan M E; Lamprecht, Alf


    Chitosan has become a popular polymer for drug delivery. It's hydro solubility and mild formulation conditions have made it an attractive polymer for macromolecular delivery. Accurate quantification of internalized chitosan nanoparticles (NPs) is imperative for fair assessment of the nano-formulation where it is important to determine the exact amount of drug actually being delivered into the cell, especially for macromolecular drugs where cellular entry is limited by molecule size and/or charge. The preferential affinity of wheat germ agglutinin tagged with fluorescein isothiocyanate (WGA-FITC) to chitosan is exploited in the development of a simple and rapid method for the differentiation between and quantification of cell surface bound and internalized chitosan NPs. The percentage of cell surface bound NPs could be easily determined and corrected NP uptake could be calculated accordingly. The developed method is applicable in several cell lines and has successfully been tested with NPs with different sizes (25 and 150nm) and with very low NP concentrations (20μg/mL). The method will allow for the correct evaluation of chitosan NP uptake and could be further used to evaluate chitosan based nanomedicine and provide guidelines on how to modify NPs for enhanced internalization, and improved drug delivery.

  7. Construction of a Pichia pastoris cell-surface display system using Flo1p anchor system. (United States)

    Tanino, Takanori; Fukuda, Hideki; Kondo, Akihiko


    A Pichia pastoris cell-surface display system was constructed using a Flo1p anchor system, which was developed in Saccharomyces cerevisiae. The lipase from Rhizopus oryzae with a pro sequence (ProROL) was used as the model protein and was genetically fused to the anchor consisting of amino acids 1-1099 of Flo1p (FS anchor). The resulting fusion protein FSProROL was expressed under the control of the alcohol oxidase 1 promoter (pAOX1). The fluorescence microscopy of immunolabeled P. pastoris cells revealed that ProROL was displayed on the cell surface, and Western blot analysis revealed that the fusion protein FSProROL was noncovalently attached to the cell wall and highly glycosylated. The lipase activity of P. pastoris cells was affected by the methanol concentration for the induction phase. Surprisingly, the activity of lipase displayed on the cells incubated at 60 degrees C was not only stable but also increased to about 6.5 times the initial value after 4 h incubation.

  8. Electrophysical characteristics of Azospirillum brasilense Sp245 during interaction with antibodies to various cell surface epitopes. (United States)

    Guliy, Olga I; Matora, Larisa Y; Burygin, Gennady L; Dykman, Lev A; Ostudin, Nikolai A; Bunin, Viktor D; Ignatov, Vladimir V; Ignatov, Oleg V


    This work was undertaken to examine the electrooptical characteristics of cells of Azospirillum brasilense Sp245 during their interaction with antibodies developed to various cell surface epitopes. We used the dependences of the cell suspension optical density changes induced by electroorientation on the orienting field frequency (740, 1000, 1450, 2000, and 2800kHz). Cell interactions with homologous strain-specific antibodies to the A. brasilense Sp245 O antigen and with homologous antibodies to whole bacterial cells brought about considerable changes in the electrooptical properties of the bacterial suspension. When genus-specific antibodies to the flagellin of the Azospirillum sheathed flagellum and antibodies to the serologically distinct O antigen of A. brasilense Sp7 were included in the A. brasilense Sp245 suspension, the changes caused in the electrooptical signal were slight and had values close to those for the above changes. These findings agree well with the immunochemical characteristics of the Azospirillum O antigens and with the data on the topographical distribution of the Azospirillum major cell surface antigens. The obtained results can serve as a basis for the development of a rapid test for the intraspecies detection of microorganisms.

  9. Growth condition-dependent cell surface proteome analysis of Enterococcus faecium. (United States)

    Sinnige, Jan C; de Been, Mark; Zhou, Miaomiao; Bonten, Marc J M; Willems, Rob J L; Top, Janetta


    The last 30 years Enterococcus faecium has become an important nosocomial pathogen in hospitals worldwide. The aim of this study was to obtain insight in the cell surface proteome of E. faecium when grown in laboratory and clinically relevant conditions. Enterococcus faecium E1162, a clinical blood stream isolate, was grown until mid-log phase in brain heart infusion medium (BHI) with, or without 0.02% bile salts, Tryptic Soy Broth with 1% glucose (TSBg) and urine, and its cell surface was "shaved" using immobilized trypsin. Peptides were identified using MS/MS. Mapping against the translated E1162 whole genome sequence identified 67 proteins that were differentially detected in different conditions. In urine, 14 proteins were significantly more and nine proteins less abundant relative to the other conditions. Growth in BHI-bile and TSBg, revealed four and six proteins, respectively, which were uniquely present in these conditions while two proteins were uniquely present in both conditions. Thus, proteolytic shaving of E. faecium cells identified differentially surface exposed proteins in different growth conditions. These proteins are of special interest as they provide more insight in the adaptive mechanisms and may serve as targets for the development of novel therapeutics against this multi-resistant emerging pathogen. All MS data have been deposited in the ProteomeXchange with identifier PXD002497 (

  10. Dynamics of putative raft-associated proteins at the cell surface. (United States)

    Kenworthy, Anne K; Nichols, Benjamin J; Remmert, Catha L; Hendrix, Glenn M; Kumar, Mukesh; Zimmerberg, Joshua; Lippincott-Schwartz, Jennifer


    Lipid rafts are conceptualized as membrane microdomains enriched in cholesterol and glycosphingolipid that serve as platforms for protein segregation and signaling. The properties of these domains in vivo are unclear. Here, we use fluorescence recovery after photobleaching to test if raft association affects a protein's ability to laterally diffuse large distances across the cell surface. The diffusion coefficients (D) of several types of putative raft and nonraft proteins were systematically measured under steady-state conditions and in response to raft perturbations. Raft proteins diffused freely over large distances (> 4 microm), exhibiting Ds that varied 10-fold. This finding indicates that raft proteins do not undergo long-range diffusion as part of discrete, stable raft domains. Perturbations reported to affect lipid rafts in model membrane systems or by biochemical fractionation (cholesterol depletion, decreased temperature, and cholesterol loading) had similar effects on the diffusional mobility of raft and nonraft proteins. Thus, raft association is not the dominant factor in determining long-range protein mobility at the cell surface.

  11. Differential carbohydrate binding and cell surface glycosylation of human cancer cell lines. (United States)

    Arndt, Nadia X; Tiralongo, Joe; Madge, Paul D; von Itzstein, Mark; Day, Christopher J


    Currently there is only a modest level knowledge of the glycosylation status of immortalised cell lines that are commonly used in cancer biology as well as their binding affinities to different glycan structures. Through use of glycan and lectin microarray technology, this study has endeavoured to define the different bindings of cell surface carbohydrate structures to glycan-binding lectins. The screening of breast cancer MDA-MB435 cells, cervical cancer HeLa cells and colon cancer Caco-2, HCT116 and HCT116-FM6 cells was conducted to determine their differential bindings to a variety of glycan and lectin structures printed on the array slides. An inverse relationship between the number of glycan structures recognised and the variety of cell surface glycosylation was observed. Of the cell lines tested, it was found that four bound to sialylated structures in initial screening. Secondary screening in the presence of a neuraminidase inhibitor (4-deoxy-4-guanidino-Neu5Ac2en) significantly reduced sialic acid binding. The array technology has proven to be useful in determining the glycosylation signatures of various cell-lines as well as their glycan binding preferences. The findings of this study provide the groundwork for further investigation into the numerous glycan-lectin interactions that are exhibited by immortalised cell lines.

  12. Influence of cell surface characteristics on adhesion of Saccharomyces cerevisiae to the biomaterial hydroxylapatite. (United States)

    White, Jane S; Walker, Graeme M


    The influence of the physicochemical properties of biomaterials on microbial cell adhesion is well known, with the extent of adhesion depending on hydrophobicity, surface charge, specific functional groups and acid-base properties. Regarding yeasts, the effect of cell surfaces is often overlooked, despite the fact that generalisations may not be made between closely related strains. The current investigation compared adhesion of three industrially relevant strains of Saccharomyces cerevisiae (M-type, NCYC 1681 and ALY, strains used in production of Scotch whisky, ale and lager, respectively) to the biomaterial hydroxylapatite (HAP). Adhesion of the whisky yeast was greatest, followed by the ale strain, while adhesion of the lager strain was approximately 10-times less. According to microbial adhesion to solvents (MATS) analysis, the ale strain was hydrophobic while the whisky and lager strains were moderately hydrophilic. This contrasted with analyses of water contact angles where all strains were characterised as hydrophilic. All yeast strains were electron donating, with low electron accepting potential, as indicated by both surface energy and MATS analysis. Overall, there was a linear correlation between adhesion to HAP and the overall surface free energy of the yeasts. This is the first time that the relationship between yeast cell surface energy and adherence to a biomaterial has been described.

  13. Pooled protein immunization for identification of cell surface antigens in Streptococcus sanguinis.

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    Xiuchun Ge

    Full Text Available BACKGROUND: Available bacterial genomes provide opportunities for screening vaccines by reverse vaccinology. Efficient identification of surface antigens is required to reduce time and animal cost in this technology. We developed an approach to identify surface antigens rapidly in Streptococcus sanguinis, a common infective endocarditis causative species. METHODS AND FINDINGS: We applied bioinformatics for antigen prediction and pooled antigens for immunization. Forty-seven surface-exposed proteins including 28 lipoproteins and 19 cell wall-anchored proteins were chosen based on computer algorithms and comparative genomic analyses. Eight proteins among these candidates and 2 other proteins were pooled together to immunize rabbits. The antiserum reacted strongly with each protein and with S. sanguinis whole cells. Affinity chromatography was used to purify the antibodies to 9 of the antigen pool components. Competitive ELISA and FACS results indicated that these 9 proteins were exposed on S. sanguinis cell surfaces. The purified antibodies had demonstrable opsonic activity. CONCLUSIONS: The results indicate that immunization with pooled proteins, in combination with affinity purification, and comprehensive immunological assays may facilitate cell surface antigen identification to combat infectious diseases.

  14. Identification of Cell Surface Targets through Meta-analysis of Microarray Data

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    Henry Haeberle


    Full Text Available High-resolution image guidance for resection of residual tumor cells would enable more precise and complete excision for more effective treatment of cancers, such as medulloblastoma, the most common pediatric brain cancer. Numerous studies have shown that brain tumor patient outcomes correlate with the precision of resection. To enable guided resection with molecular specificity and cellular resolution, molecular probes that effectively delineate brain tumor boundaries are essential. Therefore, we developed a bioinformatics approach to analyze micro-array datasets for the identification of transcripts that encode candidate cell surface biomarkers that are highly enriched in medulloblastoma. The results identified 380 genes with greater than a two-fold increase in the expression in the medulloblastoma compared with that in the normal cerebellum. To enrich for targets with accessibility for extracellular molecular probes, we further refined this list by filtering it with gene ontology to identify genes with protein localization on, or within, the plasma membrane. To validate this meta-analysis, the top 10 candidates were evaluated with immunohistochemistry. We identified two targets, fibrillin 2 and EphA3, which specifically stain medulloblastoma. These results demonstrate a novel bioinformatics approach that successfully identified cell surface and extracellular candidate markers enriched in medulloblastoma versus adjacent cerebellum. These two proteins are high-value targets for the development of tumor-specific probes in medulloblastoma. This bioinformatics method has broad utility for the identification of accessible molecular targets in a variety of cancers and will enable probe development for guided resection.

  15. Structural insights into alginate binding by bacterial cell-surface protein. (United States)

    Temtrirath, Kanate; Murata, Kousaku; Hashimoto, Wataru


    A gram-negative Sphingomonas sp. strain A1 inducibly forms a mouth-like pit on the cell surface in the presence of alginate and directly incorporates polymers into the cytoplasm via the pit and ABC transporter. Among the bacterial proteins involved in import of alginate, a cell-surface EfeO-like Algp7 shows an ability to bind alginate, suggesting its contribution to accumulate alginate in the pit. Here, we show identification of its positively charged cluster involved in alginate binding using X-ray crystallography, docking simulation, and site-directed mutagenesis. The tertiary structure of Algp7 was determined at a high resolution (1.99Å) by molecular replacement, although no alginates were included in the structure. Thus, an in silico model of Algp7/oligoalginate was constructed by docking simulation using atomic coordinates of Algp7 and alginate oligosaccharides, where some charged residues were found to be potential candidates for alginate binding. Site-directed mutagenesis was conducted and five purified mutants K68A, K69A, E194A, N221A, and K68A/K69A were subjected to a binding assay. UV absorption difference spectroscopy along with differential scanning fluorimetry analysis indicated that K68A/K69A exhibited a significant reduction in binding affinity with alginate than wild-type Algp7. Based on these data, Lys68/Lys69 residues of Algp7 probably play an important role in binding alginate.

  16. Regulation of collagen fibrillogenesis by cell-surface expression of kinase dead DDR2. (United States)

    Blissett, Angela R; Garbellini, Derek; Calomeni, Edward P; Mihai, Cosmin; Elton, Terry S; Agarwal, Gunjan


    The assembly of collagen fibers, the major component of the extracellular matrix (ECM), governs a variety of physiological processes. Collagen fibrillogenesis is a tightly controlled process in which several factors, including collagen binding proteins, have a crucial role. Discoidin domain receptors (DDR1 and DDR2) are receptor tyrosine kinases that bind to and are phosphorylated upon collagen binding. The phosphorylation of DDRs is known to activate matrix metalloproteases, which in turn cleave the ECM. In our earlier studies, we established a novel mechanism of collagen regulation by DDRs; that is, the extracellular domain (ECD) of DDR2, when used as a purified, soluble protein, inhibits collagen fibrillogenesis in-vitro. To extend this novel observation, the current study investigates how the DDR2-ECD, when expressed as a membrane-anchored, cell-surface protein, affects collagen fibrillogenesis by cells. We generated a mouse osteoblast cell line that stably expresses a kinase-deficient form of DDR2, termed DDR2/-KD, on its cell surface. Transmission electron microscopy, fluorescence microscopy, and hydroxyproline assays demonstrated that the expression of DDR2/-KD reduced the rate and abundance of collagen deposition and induced significant morphological changes in the resulting fibers. Taken together, our observations extend the functional roles that DDR2 and possibly other membrane-anchored, collagen-binding proteins can play in the regulation of cell adhesion, migration, proliferation and in the remodeling of the extracellular matrix.

  17. Assembly and cell surface expression of TAP-independent, chloroquine-sensitive and interferon-gamma-inducible class I MHC complexes in transformed fibroblast cell lines are regulated by tapasin. (United States)

    Fromm, Sharon Vigodman; Duady-Ben Yaakov, Shirly; Schechter, Chana; Ehrlich, Rachel


    Antigen processing and presentation by class I MHC molecules generally require assembly with peptide epitopes generated by the proteasome and transported into the ER by the transporters associated with antigen presentation (TAP). Recently, TAP-independent pathways supporting class I MHC-mediated presentation of exogenous antigens, as well as of endogenously synthesized viral antigens, were described. We now characterize a TAP-independent pathway that is operative in both TAP1- and TAP2-deficient Adenovirus (Ad)-transformed fibroblast cell lines. To the best of our knowledge, this is the first time that the existence of such a pathway has been described in non-infected cells that do not belong to the hematopoietic lineage. We show that this pathway is proteasome-independent and chloroquine-sensitive. Cell surface expression of these TAP-independent class I complexes is modulated by tapasin levels and is enhanced by IFN-gamma. The data imply that IFN-gamma increases the relative level of TAP-independent high affinity class I complexes that exit the ER on their way to the cell surface and to vacuolar compartments where peptide cleavage/exchange might take place before recycling to the cell surface. Since both TAP and tapasin expression are altered in numerous tumors and in virus-infected cells, TAP-independent class I complexes may be a valuable target source for immune responses.

  18. Cell surface profiling using high-throughput flow cytometry: a platform for biomarker discovery and analysis of cellular heterogeneity.

    Directory of Open Access Journals (Sweden)

    Craig A Gedye

    Full Text Available Cell surface proteins have a wide range of biological functions, and are often used as lineage-specific markers. Antibodies that recognize cell surface antigens are widely used as research tools, diagnostic markers, and even therapeutic agents. The ability to obtain broad cell surface protein profiles would thus be of great value in a wide range of fields. There are however currently few available methods for high-throughput analysis of large numbers of cell surface proteins. We describe here a high-throughput flow cytometry (HT-FC platform for rapid analysis of 363 cell surface antigens. Here we demonstrate that HT-FC provides reproducible results, and use the platform to identify cell surface antigens that are influenced by common cell preparation methods. We show that multiple populations within complex samples such as primary tumors can be simultaneously analyzed by co-staining of cells with lineage-specific antibodies, allowing unprecedented depth of analysis of heterogeneous cell populations. Furthermore, standard informatics methods can be used to visualize, cluster and downsample HT-FC data to reveal novel signatures and biomarkers. We show that the cell surface profile provides sufficient molecular information to classify samples from different cancers and tissue types into biologically relevant clusters using unsupervised hierarchical clustering. Finally, we describe the identification of a candidate lineage marker and its subsequent validation. In summary, HT-FC combines the advantages of a high-throughput screen with a detection method that is sensitive, quantitative, highly reproducible, and allows in-depth analysis of heterogeneous samples. The use of commercially available antibodies means that high quality reagents are immediately available for follow-up studies. HT-FC has a wide range of applications, including biomarker discovery, molecular classification of cancers, or identification of novel lineage specific or stem cell

  19. Knowledge discovery of cell-cell and cell-surface interactions (United States)

    Su, Jing

    High-throughput cell culture is an emerging technology that shows promise as a tool for research in tissue engineering, drug discovery, and medical diagnostics. An important, but overlooked, challenge is the integration of experimental methods with information processing suitable for handling large databases of cell-cell and cell-substrate interactions. In this work the traditional global descriptions of cell behaviors and surface characteristics was shown insufficient for investigating short-distance cell-to-cell and cell-to-surface interactions. Traditional summary metrics cannot distinguish information of cell near neighborhood from the average, global features, thus often is not suitable for studying distance-sensitive cell behaviors. The problem of traditional summary metrics was addressed by introducing individual-cell based local metrics that emphasize cell local environment. An individual-cell based local data analysis method was established. Contact inhibition of cell proliferation was used as a benchmark for the effectiveness of the local metrics and the method. Where global, summary metrics were unsuccessful, the local metrics successfully and quantitatively distinguished the contact inhibition effects of MC3T3-E1 cells on PLGA, PCL, and TCPS surfaces. In order to test the new metrics and analysis method in detail, a model of cell contact inhibition was proposed. Monte Carlo simulation was performed for validating the individual-cell based local data analysis method as well as the cell model itself. The simulation results well matched with the experimental observations. The parameters used in the cell model provided new descriptions of both cell behaviors and surface characteristics. Based on the viewpoint of individual cells, the local metrics and local data analysis method were extended to the investigation of cell-surface interactions, and a new high-throughput screening and knowledge discovery method on combinatorial libraries, local cell

  20. Computational analysis of the tether-pulling experiment to probe plasma membrane-cytoskeleton interaction in cells (United States)

    Schumacher, Kristopher R.; Popel, Aleksander S.; Anvari, Bahman; Brownell, William E.; Spector, Alexander A.


    Tethers are thin membrane tubes that can be formed when relatively small and localized forces are applied to cellular membranes and lipid bilayers. Tether pulling experiments have been used to better understand the fine membrane properties. These include the interaction between the plasma membrane and the underlying cytoskeleton, which is an important factor affecting membrane mechanics. We use a computational method aimed at the interpretation and design of tether pulling experiments in cells with a strong membrane-cytoskeleton attachment. In our model, we take into account the detailed information in the topology of bonds connecting the plasma membrane and the cytoskeleton. We compute the force-dependent piecewise membrane deflection and bending as well as modes of stored energy in three major regions of the system: body of the tether, membrane-cytoskeleton attachment zone, and the transition zone between the two. We apply our method to three cells: cochlear outer hair cells (OHCs), human embryonic kidney (HEK) cells, and Chinese hamster ovary (CHO) cells. OHCs have a special system of pillars connecting the membrane and the cytoskeleton, and HEK and CHO cells have the membrane-cytoskeleton adhesion arrangement via bonds (e.g., PIP2), which is common to many other cells. We also present a validation of our model by using experimental data on CHO and HEK cells. The proposed method can be an effective tool in the analyses of experiments to probe the properties of cellular membranes.

  1. Pyrolysis of Phenethyl Phenyl Ether Tethered in Mesoporous Silica. Effects of Confinement and Surface Spacer Molecules on Product Selectivity

    Energy Technology Data Exchange (ETDEWEB)

    Kidder, Michelle [ORNL; Chaffe, Alan [Monash University, Australia; Nguyen, M [Monash University, Australia; Buchanan III, A C [ORNL


    There has been expanding interest in exploring porous metal oxides as a confining environment for organic molecules resulting in altered chemical and physical properties including chemical transformations. In this paper, we examine the pyrolysis behavior of phenethyl phenyl ether (PPE) confined in mesoporous silica by covalent tethers to the pore walls as a function of tether density and the presence of co-tethered surface spacer molecules of varying structure (biphenyl, naphthyl, octyl, and hexadecyl). The PPE pyrolysis product selectivity, which is determined by two competitive free-radical pathways cycling through the two aliphatic radical intermediates (PhCH CH2OPh and PhCH2CH OPh), is shown to be dramatically different from that measured in the liquid phase as well as for PPE tethered to the exterior surface of nonporous silica nanoparticles. Tailoring the pore surface with spacer molecules further alters the selectivity such that the PPE reaction channel involving a molecular rearrangement (O-C phenyl shift in PhCH2CH OPh), which accounts for 25 % of the products in the liquid phase, can be virtually eliminated under pore confinement conditions. The origin of this change in selectivity is discussed in the context of steric constraints on the rearrangement path, confinement effects, pore surface curvature, and hydrogen bonding of PPE with residual surface silanols supplemented by nitrogen physisorption data and molecular dynamics simulations.

  2. Zirconocene and Si-tethered diynes: a happy match directed toward organometallic chemistry and organic synthesis. (United States)

    Zhang, Wen-Xiong; Zhang, Shaoguang; Xi, Zhenfeng


    Characterizing reactive organometallic intermediates is critical for understanding the mechanistic aspects of metal-mediated organic reactions. Moreover, the isolation of reactive organometallic intermediates can often result in the ability to design new synthetic methods. In this Account, we outline synthetic methods that we developed for a variety of diverse Zr/Si organo-bimetallic compounds and Si/N heteroatom-organic compounds through the detailed study of zirconacyclobutene-silacyclobutene fused compounds. Two basic components are involved in this chemistry. The first is the Si-tethered diyne, which owes its rich reactive palette to the combination of the Si-C bond and the C≡C triple bond. The second is the low-valent zirconocene species Cp(2)Zr(II), which has proven very useful in organic synthesis. The reaction of these two components affords the zirconacyclobutene-silacyclobutene fused compound, which is the key reactive Zr/Si organo-bimetallic intermediate discussed here. We discuss the three types of reactions that have been developed for the zirconacyclobutene-silacyclobutene fused intermediate. The reaction with nitriles (the C≡N triple bond) is introduced in the first section. In this one-pot reaction, up to four different components can be combined: the Si-tethered diyne can be reacted with three identical nitriles, with differing nitriles, or with a nitrile and other unsaturated organic substrates such as formamides, isocyanides, acid chlorides, aldehydes, carbodiimides, and azides. Several unexpected multiring, fused Zr/Si organo-bimetallic intermediates were isolated and characterized. A wide variety of N-heterocycles, such as 5-azaindole, pyrrole, and pyrroloazepine derivatives, were obtained. We then discuss the reaction with alkynes (the C≡C triple bond). A consecutive skeletal rearrangement, differing from that observed in the reactions with nitriles, takes place in this reaction. Finally, we discuss the reaction with the C═X substrates

  3. Characterization of Cell Surface and EPS Remodeling of Azospirillum brasilense Chemotaxis-like 1 Signal Transduction Pathway mutants by Atomic Force Microscopy

    Energy Technology Data Exchange (ETDEWEB)

    Billings, Amanda N [ORNL; Siuti, Piro [ORNL; Bible, Amber [University of Tennessee, Knoxville (UTK); Alexandre, Gladys [University of Tennessee, Knoxville (UTK); Retterer, Scott T [ORNL; Doktycz, Mitchel John [ORNL; Morrell-Falvey, Jennifer L [ORNL


    To compete in complex microbial communities, bacteria must quickly sense environmental changes and adjust cellular functions for optimal growth. Chemotaxis-like signal transduction pathways are implicated in the modulation of multiple cellular responses, including motility, EPS production, and cell-to-cell interactions. Recently, the Che1 chemotaxis-like pathway from Azospirillum brasilense was shown to modulate flocculation. In A. brasilense, cell surface properties, including EPS production, are thought to play a direct role in promoting flocculation. Using atomic force microscopy (AFM), we have detected distinct changes in the surface morphology of flocculating A. brasilense Che1 mutant strains that are absent in the wild type strain. Whereas the wild type strain produces a smooth mucosal extracellular matrix, the flocculating Che1 mutant strains produce distinctive extracellular fibril structures. Further analyses using flocculation inhibition and lectin-binding assays suggest that the composition of EPS components in the extracellular matrix differs between the cheA1 and cheY1 mutants, despite an apparent similarity in the macroscopic floc structures. Collectively, these data indicate that mutations in the Che1 pathway that result in increased flocculation are correlated with distinctive changes in the extracellular matrix structure produced by the mutants, including likely changes in the EPS structure and/or composition.

  4. Spp1, a member of the Set1 Complex, promotes meiotic DSB formation in promoters by tethering histone H3K4 methylation sites to chromosome axes. (United States)

    Sommermeyer, Vérane; Béneut, Claire; Chaplais, Emmanuel; Serrentino, Maria Elisabetta; Borde, Valérie


    Meiotic chromosomes are organized into arrays of loops that are anchored to the chromosome axis structure. Programmed DNA double-strand breaks (DSBs) that initiate meiotic recombination, catalyzed by Spo11 and accessory DSB proteins, form in loop sequences in promoters, whereas the DSB proteins are located on chromosome axes. Mechanisms bridging these two chromosomal regions for DSB formation have remained elusive. Here we show that Spp1, a conserved member of the histone H3K4 methyltransferase Set1 complex, is required for normal levels of DSB formation and is associated with chromosome axes during meiosis, where it physically interacts with the Mer2 DSB protein. The PHD finger module of Spp1, which reads H3K4 methylation close to promoters, promotes DSB formation by tethering these regions to chromosome axes and activating cleavage by the DSB proteins. This paper provides the molecular mechanism linking DSB sequences to chromosome axes and explains why H3K4 methylation is important for meiotic recombination.

  5. Cell-surface expression of Hsp70 on hematopoietic cancer cells after inhibition of HDAC activity

    DEFF Research Database (Denmark)

    Jensen, Helle; Andresen, Lars; Hansen, Karen Aagaard

    , membrane-bound Hsp70 can stimulate antigen presenting cells (APCs) to release proinflammatory cytokines and can provide a target structure for NK cell-mediated lysis. Human cancer cells frequently express Hsp70 on their cell surface, whereas the corresponding normal tissues do not. In addition, several...... clinically applied reagents, such as alkyl-lysophospholipides, chemotherapeutic agents, and anti-inflammatory reagents, have been found to enhance Hsp70 surface expression on cancer cells. We have found that inhibition of histone deacetylase (HDAC) activity leads to surface expression of Hsp70 on various...... hematopoietic cancer cells, an occurance that was not observed on naïve or activated peripheral blood cells. HDAC-inhibitor mediated Hsp70 surface expression was confined to the apoptotic Annexin V positive cells and blocked by inhibition of apoptosis. Other chemotherapeutic inducers of apoptosis...

  6. Fluid dynamics and noise in bacterial cell-cell and cell-surface scattering

    CERN Document Server

    Drescher, Knut; Cisneros, Luis H; Ganguly, Sujoy; Goldstein, Raymond E; 10.1073/pnas.1019079108


    Bacterial processes ranging from gene expression to motility and biofilm formation are constantly challenged by internal and external noise. While the importance of stochastic fluctuations has been appreciated for chemotaxis, it is currently believed that deterministic long-range fluid dynamical effects govern cell-cell and cell-surface scattering - the elementary events that lead to swarming and collective swimming in active suspensions and to the formation of biofilms. Here, we report the first direct measurements of the bacterial flow field generated by individual swimming Escherichia coli both far from and near to a solid surface. These experiments allowed us to examine the relative importance of fluid dynamics and rotational diffusion for bacteria. For cell-cell interactions it is shown that thermal and intrinsic stochasticity drown the effects of long-range fluid dynamics, implying that physical interactions between bacteria are determined by steric collisions and near-field lubrication forces. This dom...

  7. Restoration of proper trafficking to the cell surface for membrane proteins harboring cysteine mutations.

    Directory of Open Access Journals (Sweden)

    Angelica Lopez-Rodriguez

    Full Text Available A common phenotype for many genetic diseases is that the cell is unable to deliver full-length membrane proteins to the cell surface. For some forms of autism, hereditary spherocytosis and color blindness, the culprits are single point mutations to cysteine. We have studied two inheritable cysteine mutants of cyclic nucleotide-gated channels that produce achromatopsia, a common form of severe color blindness. By taking advantage of the reactivity of cysteine's sulfhydryl group, we modified these mutants with chemical reagents that attach moieties with similar chemistries to the wild-type amino acids' side chains. We show that these modifications restored proper delivery to the cell membrane. Once there, the channels exhibited normal functional properties. This strategy might provide a unique opportunity to assess the chemical nature of membrane protein traffic problems.

  8. Topical retinoic acid changes the epidermal cell surface glycosylation pattern towards that of a mucosal epithelium

    DEFF Research Database (Denmark)

    Griffiths, C E; Dabelsteen, Erik; Voorhees, J J


    for carbohydrate synthesis, are influenced by retinoids. Thus, we investigated whether epidermal cell surface glycosylation is altered in skin treated with topical RA, and contrasted it with changes induced by topical SLS. Skin biopsies were obtained from seven normal volunteers who had been treated, on three......-treated epidermis was not significantly different from that observed after vehicle treatment. Thus, RA treatment converts normal stratified epithelium towards the phenotype of mucosal epithelium with a decrease in T antigen and a concomitant increase in Ley. These changes are not observed following treatment......Topical all-trans retinoic acid (RA) produces a number of epidermal changes which are indistinguishable from those observed following treatment with a local irritant, namely sodium lauryl sulphate (SLS). This observation has led to criticism that the efficacy of RA in disorders such as photoageing...

  9. Signaling at the cell surface in the circulatory and ventilatory systems

    CERN Document Server

    Thiriet, Marc


    The volumes in this authoritative series present a multidisciplinary approach to modeling and simulation of flows in the cardiovascular and ventilatory systems, especially multiscale modeling and coupled simulations. The cardiovascular and respiratory systems are tightly coupled, as their primary function is to supply oxygen to and remove carbon dioxide from the body's cells. Because physiological conduits have deformable and reactive walls, macroscopic flow behavior and prediction must be coupled to nano- and microscopic events in a corrector scheme of regulated mechanisms when the vessel lumen caliber varies markedly. Therefore, investigation of flows of blood and air in physiological conduits requires an understanding of the biology, chemistry, and physics of these systems together with the mathematical tools to describe their functioning. Volume 3 is devoted to the set of mediators of the cell surface, especially ion and molecular carriers and catalytic receptors that, once liganded and activated, initiat...

  10. Detection of the plant parasite Cuscuta reflexa by a tomato cell surface receptor. (United States)

    Hegenauer, Volker; Fürst, Ursula; Kaiser, Bettina; Smoker, Matthew; Zipfel, Cyril; Felix, Georg; Stahl, Mark; Albert, Markus


    Parasitic plants are a constraint on agriculture worldwide. Cuscuta reflexa is a stem holoparasite that infests most dicotyledonous plants. One exception is tomato, which is resistant to C. reflexa We discovered that tomato responds to a small peptide factor occurring in Cuscuta spp. with immune responses typically activated after perception of microbe-associated molecular patterns. We identified the cell surface receptor-like protein CUSCUTA RECEPTOR 1 (CuRe1) as essential for the perception of this parasite-associated molecular pattern. CuRe1 is sufficient to confer responsiveness to the Cuscuta factor and increased resistance to parasitic C. reflexa when heterologously expressed in otherwise susceptible host plants. Our findings reveal that plants recognize parasitic plants in a manner similar to perception of microbial pathogens.

  11. Effects of sodium on cell surface and intracellular TH-naloxone binding sites

    Energy Technology Data Exchange (ETDEWEB)

    Pollack, A.E.; Wooten, G.F.


    The binding of the opiate antagonist TH-naloxone was examined in rat whole brain homogenates and in crude subcellular fractions of these homogenates (nuclear, synaptosomal, and mitochondrial fractions) using buffers that approximated intra- (low sodium concentration) and extracellular (high sodium concentration) fluids. Saturation studies showed a two-fold decrease in the dissociation constant (Kd) in all subcellular fractions examined in extracellular buffer compared to intracellular buffer. In contrast, there was no significant effect of the buffers on the Bmax. Thus, TH-naloxone did not distinguish between binding sites present on cell surface and intracellular tissues in these two buffers. These results show that the sodium effect of opiate antagonist binding is probably not a function of altered selection of intra- and extracellular binding sites. 17 references, 2 tables.

  12. Display of wasp venom allergens on the cell surface of Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Borodina, Irina; Jensen, B. M.; Søndergaard, Ib;


    Background: Yeast surface display is a technique, where the proteins of interest are expressed as fusions with yeast surface proteins and thus remain attached to the yeast cell wall after expression. Our purpose was to study whether allergens expressed on the cell surface of baker's yeast...... Saccharomyces cerevisiae preserve their native allergenic properties and whether the yeast native surface glycoproteins interfere with IgE binding. We chose to use the major allergens from the common wasp Vespula vulgaris venom: phospholipase A1, hyaluronidase and antigen 5 as the model. Results: The proteins...... were expressed on the surface as fusions with a-agglutinin complex protein AGA2. The expression was confirmed by fluorescent cytometry (FACS) after staining the cells with antibody against a C-tag attached to the C-terminal end of the allergens. Phospholipase A1 and hyaluronidase retained...

  13. Improved accuracy of cell surface shaving proteomics in Staphylococcus aureus using a false-positive control

    DEFF Research Database (Denmark)

    Solis, Nestor; Larsen, Martin Røssel; Cordwell, Stuart J


    Proteolytic treatment of intact bacterial cells is an ideal means for identifying surface-exposed peptide epitopes and has potential for the discovery of novel vaccine targets. Cell stability during such treatment, however, may become compromised and result in the release of intracellular proteins...... that complicate the final analysis. Staphylococcus aureus is a major human pathogen, causing community and hospital-acquired infections, and is a serious healthcare concern due to the increasing prevalence of multiple antibiotic resistances amongst clinical isolates. We employed a cell surface "shaving" technique...... lysis and were removed from the trypsin-shaved data set. We identified 42 predicted S. aureus COL surface proteins from 260 surface-exposed peptides. Trypsin and proteinase-K digests were highly complementary with ten proteins identified by both, 16 specific to proteinase-K treatment, 13 specific...

  14. Altered expression of epithelial cell surface glycoconjugates and intermediate filaments at the margins of mucosal wounds

    DEFF Research Database (Denmark)

    Dabelsteen, Erik; Grøn, B; Mandel, U


    Alterations in cell to cell adhesion are necessary to enable the type of cell movements that are associated with epithelial wound healing and malignant invasion. Several studies of transformed cells have related epithelial cell movement to changes in the cell surface expression of the carbohydrate......-T antigen. The changes induced by wounding in the expression of collagen IV, laminin gamma2-chain (laminin-5), and laminin alpha5-chain were similar to those found in skin wounds and served to define the region of epithelial movement. This region was found to show a marked increase in staining for both...... epithelium, a pattern of expression similar to K16, which was also strongly upregulated in both the outgrowth and the adjacent nonwounded epithelium. These findings provide further support for an influence of such carbohydrate structures on the migratory behavior of epithelial cells....

  15. Flow cytometry detection of planktonic cells with polycyclic aromatic hydrocarbons sorbed to cell surfaces

    KAUST Repository

    Cerezo, Maria I.


    Polycyclic aromatic hydrocarbons are very important components of oil pollution. These pollutants tend to sorb to cell surfaces, exerting toxic effects on organisms. Our study developed a flow cytometric method for the detection of PAHs sorbed to phytoplankton by exploiting their spectral characteristics. We discriminated between cells with PAHs from cells free of PAHs. Clear discrimination was observed with flow cytometer provided with 375 or 405nm lasers in addition to the standard 488nm laser necessary to identify phytoplankton. Using this method, we measured the relationship between the percentages of phytoplankton organisms with PAHs, with the decrease in the growth rate. Moreover, the development of this method could be extended to facilitate the study of PAHs impact on cell cultures from a large variety of organisms.

  16. The effects of a realistic hollow cathode plasma contactor model on the simulation of bare electrodynamic tether systems (United States)

    Blash, Derek M.

    The region known as Low-Earth Orbit (LEO) has become populated with artificial satellites and space debris since humanities initial venture into the region. This has turned LEO into a hazardous region. Since LEO is very valuable to many different countries, there has been a push to prevent further buildup and talk of even deorbiting spent satellites and debris already in LEO. One of the more attractive concepts available for deorbiting debris and spent satellites is a Bare Electrodynamic Tether (BET). A BET is a propellantless propulsion technique in which two objects are joined together by a thin conducting material. When these tethered objects are placed in LEO, the tether sweeps across the magnetic field lines of the Earth and induces an electromotive force (emf) along the tether. Current from the space plasma is collected on the bare tether under the action of the induced emf, and this current interacts with the Earth's magnetic field to create a drag force that can be used to deorbit spent satellites and space debris. A Plasma Contactor (PC) is used to close the electrical circuit between the BET and the ionospheric plasma. The PC requires a voltage and, depending on the device, a gas flow to emit electrons through a plasma bridge to the ionospheric plasma. The PC also can require a plasma discharge electrode and a heater to condition the PC for operation. These parameters as well as the PC performance are required to build an accurate simulation of a PC and, therefore, a BET deorbiting system. This thesis focuses on the development, validation, and implementation of a simulation tool to model the effects of a realistic hollow cathode PC system model on a BET deorbit system.

  17. Cell-Surface Receptors Transactivation Mediated by G Protein-Coupled Receptors

    Directory of Open Access Journals (Sweden)

    Fabio Cattaneo


    Full Text Available G protein-coupled receptors (GPCRs are seven transmembrane-spanning proteins belonging to a large family of cell-surface receptors involved in many intracellular signaling cascades. Despite GPCRs lack intrinsic tyrosine kinase activity, tyrosine phosphorylation of a tyrosine kinase receptor (RTK occurs in response to binding of specific agonists of several such receptors, triggering intracellular mitogenic cascades. This suggests that the notion that GPCRs are associated with the regulation of post-mitotic cell functions is no longer believable. Crosstalk between GPCR and RTK may occur by different molecular mechanism such as the activation of metalloproteases, which can induce the metalloprotease-dependent release of RTK ligands, or in a ligand-independent manner involving membrane associated non-receptor tyrosine kinases, such as c-Src. Reactive oxygen species (ROS are also implicated as signaling intermediates in RTKs transactivation. Intracellular concentration of ROS increases transiently in cells stimulated with GPCR agonists and their deliberated and regulated generation is mainly catalyzed by enzymes that belong to nicotinamide adenine dinucleotide phosphate (NADPH oxidase family. Oxidation and/or reduction of cysteine sulfhydryl groups of phosphatases tightly controls the activity of RTKs and ROS-mediated inhibition of cellular phosphatases results in an equilibrium shift from the non-phosphorylated to the phosphorylated state of RTKs. Many GPCR agonists activate phospholipase C, which catalyze the hydrolysis of phosphatidylinositol 4,5-bis-phosphate to produce inositol 1,4,5-triphosphate and diacylglicerol. The consequent mobilization of Ca2+ from endoplasmic reticulum leads to the activation of protein kinase C (PKC isoforms. PKCα mediates feedback inhibition of RTK transactivation during GPCR stimulation. Recent data have expanded the coverage of transactivation to include Serine/Threonine kinase receptors and Toll-like receptors

  18. Cell surface characteristics enable encrustation-free survival of neutrophilic iron-oxidizing bacteria (United States)

    Saini, G.; Chan, C. S.


    Microbial growth in mineralizing environments depends on the cells' ability to evade surface precipitation. Cell-mineral interactions may be required for metabolism, but if unmoderated, cells could become encrusted, which would limit diffusion of nutrients and waste across cell walls. A combination of cell surface charge and hydrophobicity could enable the survival of microbes in such environments by inhibiting mineral attachment. To investigate this mechanism, we characterized the surfaces of two neutrophilic iron-oxidizing bacteria (FeOB): Mariprofundus ferrooxydans, a Zetaproteobacterium from Fe(II)-rich submarine hydrothermal vents and a Betaproteobacterium Gallionellales strain R-1, recently isolated from a ferrous groundwater seep. Both bacteria produce iron oxyhydroxides, yet successfully escape surface encrustation while inhabiting milieu where iron minerals are also produced by abiotic processes. SEM-EDX and TEM-EELS analyses of cultured bacteria revealed no iron on the cell surfaces. Zeta potential measurements showed that these bacteria have very small negative surface charge (0 to -4 mV) over a pH range of 4-9, indicating near-neutrally charged surfaces. Water contact angle measurements and thermodynamic calculations demonstrate that both bacteria and abiotically-formed Fe oxhydroxides are hydrophilic. Extended-DLVO calculations showed that hydrophilic repulsion between cells and minerals dominates over electrostatic and Lifshitz-van der Waals interactions. This leads to overall repulsion between microbes and minerals, thus preventing surface encrustation. Low surface charge and hydrophilicity (determined by microbial adhesion to hydrocarbon assay) were common features for both live and azide-inhibited cells, which shows that surface characteristics do not depend on active metabolism. It is remarkable that these two phylogenetically-distant bacteria from different environments employ similar adaptations to prevent surface mineralization. Our results

  19. Role of calcium in growth inhibition induced by a novel cell surface sialoglycopeptide (United States)

    Betz, N. A.; Westhoff, B. A.; Johnson, T. C.; Spooner, B. S. (Principal Investigator)


    Our laboratory has purified an 18 kDa cell surface sialoglycopeptide growth inhibitor (CeReS-18) from intact bovine cerebral cortex cells. Evidence presented here demonstrates that sensitivity to CeReS-18-induced growth inhibition in BALB-c 3T3 cells is influenced by calcium, such that a decrease in the calcium concentration in the growth medium results in an increase in sensitivity to CeReS-18. Calcium did not alter CeReS-18 binding to its cell surface receptor and CeReS-18 does not bind calcium directly. Addition of calcium, but not magnesium, to CeReS-18-inhibited 3T3 cells results in reentry into the cell cycle. A greater than 3-hour exposure to increased calcium is required for escape from CeReS-18-induced growth inhibition. The calcium ionophore ionomycin could partially mimic the effect of increasing extracellular calcium, but thapsigargin was ineffective in inducing escape from growth inhibition. Increasing extracellular calcium 10-fold resulted in an approximately 7-fold increase in total cell-associated 45Ca+2, while free intracellular calcium only increased approximately 30%. However, addition of CeReS-18 did not affect total cell-associated calcium or the increase in total cell-associated calcium observed with an increase in extracellular calcium. Serum addition induced mobilization of intracellular calcium and influx across the plasma membrane in 3T3 cells, and pretreatment of 3T3 cells with CeReS-18 appeared to inhibit these calcium mobilization events. These results suggest that a calcium-sensitive step exists in the recovery from CeReS-18-induced growth inhibition. CeReS-18 may inhibit cell proliferation through a novel mechanism involving altering the intracellular calcium mobilization/regulation necessary for cell cycle progression.

  20. Sponges with covalently tethered amines for high-efficiency carbon capture

    KAUST Repository

    Qi, Genggeng


    © 2014 Macmillan Publishers Limited. All rights reserved. Adsorption using solid amine sorbents is an attractive emerging technology for energy-efficient carbon capture. Current syntheses for solid amine sorbents mainly based on physical impregnation or grafting-to methods (for example, aminosilane-grafting) lead to limited sorbent performance in terms of stability and working capacity, respectively. Here we report a family of solid amine sorbents using a grafting-from synthesis approach and synthesized by cationic polymerization of oxazolines on mesoporous silica. The sorbent with high amount of covalently tethered amines shows fast adsorption rate, high amine efficiency and sorbent capacity well exceeding the highest value reported to date for lowerature carbon dioxide sorbents under simulated flue gas conditions. The demonstrated efficiency of the new amine-immobilization chemistry may open up new avenues in the development of advanced carbon dioxide sorbents, as well as other nitrogen-functionalized systems.

  1. Dissecting functions of the conserved oligomeric Golgi tethering complex using a cell-free assay. (United States)

    Cottam, Nathanael P; Wilson, Katherine M; Ng, Bobby G; Körner, Christian; Freeze, Hudson H; Ungar, Daniel


    Vesicle transport sorts proteins between compartments and is thereby responsible for generating the non-uniform protein distribution along the eukaryotic secretory and endocytic pathways. The mechanistic details of specific vesicle targeting are not yet well characterized at the molecular level. We have developed a cell-free assay that reconstitutes vesicle targeting utilizing the recycling of resident enzymes within the Golgi apparatus. The assay has physiological properties, and could be used to show that the two lobes of the conserved oligomeric Golgi tethering complex play antagonistic roles in trans-Golgi vesicle targeting. Moreover, we can show that the assay is sensitive to several different congenital defects that disrupt Golgi function and therefore cause glycosylation disorders. Consequently, this assay will allow mechanistic insight into the targeting step of vesicle transport at the Golgi, and could also be useful for characterizing some novel cases of congenital glycosylation disorders.

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

    Hansen, Scott D; Mullins, R Dyche


    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.

  3. Adaptive sliding mode controller based on super-twist observer for tethered satellite system (United States)

    Keshtkar, Sajjad; Poznyak, Alexander


    In this work, the sliding mode control based on the super-twist observer is presented. The parameters of the controller as well as the observer are admitted to be time-varying and depending on available current measurements. In view of that, the considered controller is referred to as an adaptive one. It is shown that the deviations of the generated state estimates from real state values together with a distance of the closed-loop system trajectories to a desired sliding surface reach a μ-zone around the origin in finite time. The application of the suggested controller is illustrated for the orientation of a tethered satellite system in a required position.

  4. Optimization of motion control laws for tether crawler or elevator systems (United States)

    Swenson, Frank R.; Von Tiesenhausen, Georg


    Based on the proposal of a motion control law by Lorenzini (1987), a method is developed for optimizing motion control laws for tether crawler or elevator systems in terms of the performance measures of travel time, the smoothness of acceleration and deceleration, and the maximum values of velocity and acceleration. The Lorenzini motion control law, based on powers of the hyperbolic tangent function, is modified by the addition of a constant-velocity section, and this modified function is then optimized by parameter selections to minimize the peak acceleration value for a selected travel time or to minimize travel time for the selected peak values of velocity and acceleration. It is shown that the addition of a constant-velocity segment permits further optimization of the motion control law performance.

  5. Acoustic detection and localization from a tethered aerostat during the NATO TG-53 test (United States)

    Reiff, C.; Scanlon, M.; Noble, J.


    Acoustic sensors mounted to a tethered aerostat detect and localize transient signals from mortars, artillery, C-4, propane cannon, and small arms fire. Significant enhancements to soldier lethality and survivability can be gained when using the aerostat array to detect, localize, and cue an aerial imager to a weapon's launch site, or use the aerostat's instantaneous position and orientation to calculate a vector solution to the ground coordinates of the launch site for threat neutralization. The prototype aerostat-mounted array was tested at Yuma Proving Grounds (YPG) as part of the NATO TG-53 signature collection exercise. Acoustic wave form data was collected simultaneously with aerostat and ground-based sensor arrays for comparing wind noise, signal to noise related parameters, and atmospheric effects on propagation to an elevated array. A test description and summary of localization accuracy will be presented for various altitudes, ranges to target, and under differing meteorological conditions.

  6. Diagnosis and surgical treatment of terminal syringomyelia within spinal cord combined with tethered cord syndrome

    Directory of Open Access Journals (Sweden)

    Jing-cheng XIE


    Full Text Available Objective To summarize the clinical manifestations, imaging characteristics and experience of surgical treatment of spinal cord terminal syringomyelia with tethered cord syndrome (TCS.  Methods and Results Clinical data of 10 patients with spinal cord syringomyelia combined with TCS surgically treated under microscope from January 1999 to March 2014 in our hospital were retrospectively analyzed. There were 3 males and 7 females with average age of 15.06 years old (ranged from 2 to 35 years old. The course of disease ranged from 3 months to 20 years (average 42.17 months. Among those patients, one patient presented hydromyelia, 8 patients suffered from meningeal cyst within the sacral canal, and one patient were concurrent with sacral dermal sinus. The weakness of lower extremities, especially distal limbs, was the main clinical manifestation. Five patients were accompanied with bowel and bladder dysfunction and 5 patients with sensory disturbance below the level of syringomyelia, especially hypesthesia. Preoperative MRI showed conus medullaris disappeared at the end of spinal cord, and there was fluid signal in the lower spinal cord with hypo-intensity signal in T1WI and hyper-intensity signal in T2WI without enhancement. All patients underwent surgical procedures. Under microscope, filum terminale was cut off, drainage was performed, meningeal cyst within the sacral canal was removed, and tethered cord was released. The success rate of operations was 100%. The duration of surgery ranged from 1.52 to 3.07 h (average 2.15 h, with average intraoperative blood loss 220 ml (ranged from 100 to 410 ml. The tethering filum had been totally resected and histological examination showed typical filum tissue in all cases. No operative complication was found. Visual Analogue Scale (VAS score was decreased, and the lower limbs weakness as well as bowel and bladder dysfunction was gradually relieved after operation. The period of follow-up was ranged from 6

  7. Linear Viscoelasticity of Spherical SiO 2 Nanoparticle-Tethered Poly(butyl acrylate) Hybrids

    KAUST Repository

    Goel, Vivek


    The melt state linear viscoelastic properties of spherical silica nanoparticles with grafted poly(n-butyl acrylate) chains of varying molecular weight were probed using linear small amplitude dynamic oscillatory measurements and complementary linear stress relaxation measurements. While the pure silica-tethered-polymer hybrids with no added homopolymer exhibit solid-like response, addition of matched molecular weight free matrix homopolymer chains to this hybrid, at low concentrations of added homopolymer, maintains the solid-like response with a lowered modulus that can be factored into a silica concentration dependence and a molecular weight dependence. While the silica concentration dependence of the modulus is strong, the dependence on molecular weight is weak. On the other hand, increasing the amount of added homopolymer changes the viscoelastic response to that of a liquid with a relaxation time that scales exponentially with hybrid concentration. © 2010 American Chemical Society.

  8. Tethered capsule endomicroscopy: from bench to bedside at a primary care practice (United States)

    Gora, Michalina J.; Simmons, Leigh H.; Quénéhervé, Lucille; Grant, Catriona N.; Carruth, Robert W.; Lu, Weina; Tiernan, Aubrey; Dong, Jing; Walker-Corkery, Beth; Soomro, Amna; Rosenberg, Mireille; Metlay, Joshua P.; Tearney, Guillermo J.


    Due to the relatively high cost and inconvenience of upper endoscopic biopsy and the rising incidence of esophageal adenocarcinoma, there is currently a need for an improved method for screening for Barrett's esophagus. Ideally, such a test would be applied in the primary care setting and patients referred to endoscopy if the result is suspicious for Barrett's. Tethered capsule endomicroscopy (TCE) is a recently developed technology that rapidly acquires microscopic images of the entire esophagus in unsedated subjects. Here, we present our first experience with clinical translation and feasibility of TCE in a primary care practice. The acceptance of the TCE device by the primary care clinical staff and patients shows the potential of this device to be useful as a screening tool for a broader population.

  9. Method to maintain artificial gravity during transfer maneuvers for tethered spacecraft (United States)

    Martin, Kaela M.; Landau, Damon F.; Longuski, James M.


    Artificial gravity has long been proposed to limit the harmful effects of the micro-gravity environment on human crews during mission to Mars. A tethered spacecraft spinning at 4 rpm (to avoid motion sickness) provides an attractive configuration. However, if the spacecraft is required to spin down for impulsive maneuvers and then spin up for interplanetary travel, the propellant cost may be unacceptably high. This paper proposes a maneuver that is performed while the spacecraft is spinning thus avoiding additional spin-down and spin-up maneuvers. A control law is provided to achieve the required ΔV while maintaining spin rate. A hypothetical human mission from Earth to Mars is analyzed using the new maneuver which, in this example, may save over 700 kg of propellant.

  10. Numerical simulations of the electrodynamic interactions between the Tethered-Satellite-System and space plasma (United States)

    Vashi, Bharat I.


    The first Tethered-Satellite-System (TSS-1), scheduled for a flight in late 1992, is expected to provide relevant information related to the concept of generating an emf in a 20-km-long (or longer) conducting wire. This paper presents numerical simulations of the electrodynamic interactions between the TSS system and space plasma, using a 2D and 3D models of the system. The 2D case code simulates the motion of a long cylinder past a plasma, which is composed of electrons and H(+) ions. The system is solved by allowing the plasma to flow past the cylinder with an imposed magnetic field. The more complex 3D case is considered to study the dynamics in great detail. Results of 2D simulation show that the interaction of a satellite with plasma flowing perpendicularly to the magnetic field results in an enhancement in the current collection.

  11. Cisplatin-tethered gold nanospheres for multimodal chemo-radiotherapy of glioblastoma (United States)

    Setua, Sonali; Ouberai, Myriam; Piccirillo, Sara G.; Watts, Colin; Welland, Mark


    Glioblastoma multiforme (GBM) remains the most aggressive and challenging brain tumour to treat. We report the first successful chemo-radiotherapy on patient derived treatment resistant GBM cells using a cisplatin-tethered gold nanosphere. After intracellular uptake, the nanosphere effects DNA damage which initiates caspase-mediated apoptosis in those cells. In the presence of radiation, both gold and platinum of cisplatin, serve as high atomic number radiosensitizers leading to the emission of ionizing photoelectrons and Auger electrons. This resulted in enhanced synergy between cisplatin and radiotherapy mediated cytotoxicity, and photo/Auger electron mediated radiosensitisation leading to complete ablation of the tumour cells in an in vitro model system. This study demonstrates the potential of designed nanoparticles to target aggressive cancers in the patient derived cell lines providing a platform to move towards treatment strategies.Glioblastoma multiforme (GBM) remains the most aggressive and challenging brain tumour to treat. We report the first successful chemo-radiotherapy on patient derived treatment resistant GBM cells using a cisplatin-tethered gold nanosphere. After intracellular uptake, the nanosphere effects DNA damage which initiates caspase-mediated apoptosis in those cells. In the presence of radiation, both gold and platinum of cisplatin, serve as high atomic number radiosensitizers leading to the emission of ionizing photoelectrons and Auger electrons. This resulted in enhanced synergy between cisplatin and radiotherapy mediated cytotoxicity, and photo/Auger electron mediated radiosensitisation leading to complete ablation of the tumour cells in an in vitro model system. This study demonstrates the potential of designed nanoparticles to target aggressive cancers in the patient derived cell lines providing a platform to move towards treatment strategies. Electronic supplementary information (ESI) available: Additional figures. See DOI: 10.1039/c

  12. Fluorosilicone multi-block copolymers tethering quaternary ammonium salt groups for antimicrobial purpose

    Energy Technology Data Exchange (ETDEWEB)

    Zhou, Fang; Qin, Xiaoshuai; Li, Yancai; Ren, Lixia; Zhao, Yunhui, E-mail:; Yuan, Xiaoyan


    Highlights: • QAS-containing fluorosilicone multi-block copolymers were synthesized. • The block length of PHFBMA in the copolymers was tailored via RAFT polymerization. • Surface roughness of the copolymers decreased with the increased PHFBMA content. • A certain length of PHFBMA block enhanced C−N{sup +} percentage on the surface. - Abstract: Symmetrically structured fluorosilicone multi-block copolymers containing poly(2-(dimethylamino)ethyl methacrylate) (PDMAEMA) and poly(hexafluorobutyl methacrylate) (PHFBMA) were sequentially synthesized via reversible addition–fragmentation chain transfer polymerization, using a polydimethylsiloxane (PDMS) chain transfer agent with dithiocarbonate groups at both ends. Then, the CBABC-type block copolymers were quaternized with n-octyliodide to tether quaternary ammonium salt (QAS) groups in the PDMAEMA blocks for the antimicrobial use. The obtained fluorosilicone copolymers showed clear variations in the C-N{sup +} composition and surface morphology on their films depending on the content of the PHFBMA blocks, which were characterized by X-ray photoelectron spectroscopy and atomic force microscopy, respectively. The results indicated that the symmetrical CBABC structure favored PDMS and QAS tethered blocks migrating to the film surface. With the mass percentage of the PHFBMA increased from 0 to 32.5%, the surface roughness of the copolymer film decreased gradually with a tendency to form a smooth surface. Owing to the surface properties, fluorosilicone multi-block copolymers containing a certain amount of PHFBMA with higher C-N{sup +} content and relatively smooth morphology demonstrated obvious antimicrobial activity against Gram-positive bacteria, Bacillus subtilis and Gram-negative bacteria, Escherichia coli. The functionalized multi-block copolymers based on fluorosilicone and QAS groups would have potential applications in antimicrobial coatings.

  13. Tethered bilayer lipid membranes studied by simultaneous attenuated total reflectance infrared spectroscopy and electrochemical impedance spectroscopy (United States)

    Erbe, Andreas; Bushby, Richard J.; Evans, Stephen D.; Jeuken, Lars J. C.


    The formation of tethered lipid bilayer membranes (tBLMs) from unilamelar vesicles of egg yolk phosphatidylcholine (EggPC) on mixed self–assembled monolayers (SAMs) from varying ratios of 6-mercaptohexanol and EO3Cholesteryl on gold has been monitored by simultaneous attenuated total reflectance fourier transform infrared (ATR–FTIR) spectroscopy and electrochemical impedance spectroscopy (EIS). The influence of the lipid orientation (and hence the anisotropy) of lipids on a gold film on the dichroic ratio was studied by simulations of spectra with a matrix method for anisotropic layers. It is shown that for certain tilt angles of the dielectric tensor of the adsorbed anisotropic layer dispersive and negative absorption bands are possible. The experimental data indicates that the structure of the assemblies obtained varies with varying SAM composition. On SAMs with a high content of EO3Cholesteryl, tBLMs with reduced fluidity are formed. For SAMs with high content of 6-mercaptohexanol, the results are consistent with the adsorption of flattened vesicles, while spherical vesicles have been found in a small range of surface compositions. The kinetics of the adsorption process is consistent with the assumption of spherical vesicles as long–living intermediates for surfaces of high 6-mercaptohexanol content. No long–living spherical vesicles have been detected for surfaces with large fraction of EO3Cholesteryl tethers. The observed differences between the surfaces suggest that for the formation of tBLMs (unlike supported BLMs) no critical surface coverage of vesicles is needed prior to lipid bilayer formation. PMID:17388505

  14. Curcumin induced nanoscale CD44 molecular redistribution and antigen-antibody interaction on HepG2 cell surface

    Energy Technology Data Exchange (ETDEWEB)

    Wang Mu [Department of Chemistry, Jinan University, 601 Huangpu Road West, Tianhe District, Guangzhou 510632 (China); Ruan Yuxia [Department of Ophthalmology, The First Affiliated Hospital, Jinan University, 601 Huangpu Road West, Tianhe District, Guangzhou 510632 (China); Xing Xiaobo; Chen Qian; Peng, Yuan [Department of Chemistry, Jinan University, 601 Huangpu Road West, Tianhe District, Guangzhou 510632 (China); Cai Jiye, E-mail: [Department of Chemistry, Jinan University, 601 Huangpu Road West, Tianhe District, Guangzhou 510632 (China)


    Graphical abstract: Highlights: > In this study, we investigate the changes of CD44 expression and distribution on HepG2 cells after curcumin treatment. > We find curcumin is able to change the morphology and ultrastructure of HepG2 cells. > Curcumin can reduce the expression of CD44 molecules and induce the nanoscale molecular redistribution on cell surface. > The binding force between CD44-modified AFM tip and the HepG2 cell surface decreases after curcumin-treatment. - Abstract: The cell surface glycoprotein CD44 was implicated in the progression, metastasis and apoptosis of certain human tumors. In this study, we used atomic force microscope (AFM) to monitor the effect of curcumin on human hepatocellular carcinoma (HepG2) cell surface nanoscale structure. High-resolution imaging revealed that cell morphology and ultrastructure changed a lot after being treated with curcumin. The membrane average roughness increased (10.88 {+-} 4.62 nm to 129.70 {+-} 43.72 nm) and the expression of CD44 decreased (99.79 {+-} 0.16% to 75.14 {+-} 8.37%). Laser scanning confocal microscope (LSCM) imaging showed that CD44 molecules were located on the cell membrane. The florescence intensity in control group was weaker than that in curcumin treated cells. Most of the binding forces between CD44 antibodies and untreated HepG2 cell membrane were around 120-220 pN. After being incubated with curcumin, the major forces focused on 70-150 pN (10 {mu}M curcumin-treated) and 50-120 pN (20 {mu}M curcumin-treated). These results suggested that, as result of nanoscale molecular redistribution, changes of the cell surface were in response to external treatment of curcumin. The combination of AFM and LSCM could be a powerful method to detect the distribution of cell surface molecules and interactions between molecules and their ligands.

  15. Decreases in CD31 and CD47 levels on the cell surface during etoposide-induced Jurkat cell apoptosis. (United States)

    Azuma, Yutaro; Nakagawa, Hideaki; Dote, Kanae; Higai, Koji; Matsumoto, Kojiro


    Engulfment of apoptotic cells is regulated by 'eat me' and 'don't eat me' signals on the cell surface. Alterations to the 'eat me' signals have been well described; however, very little is known about the 'don't eat me' signals on the cell surface during apoptosis. In the present study, apoptosis of Jurkat cells was induced by treatment with topoisomerase II inhibitor etoposide, and then the CD31 and CD47 levels on the apoptotic cell surface and in microparticles were estimated by flow cytometry and immunoblotting methods in the presence of caspase, metalloproteinase, and Rho-associated coiled-coil containing protein kinase 1 (ROCK1) inhibitors. The CD31 and CD47 levels on the cell surface of apoptotic Jurkat cells had decreased after treatment with etoposide. These decreases in CD31 and CD47 levels on the apoptotic cell surface were almost completely suppressed by the caspase 3 inhibitor, Ac-DEVD-CHO, and partially suppressed by caspase 8 (Ac-IETD-CHO) and caspase 9 (Ac-LEHE-CHO) inhibitors but not by the metalloproteinase inhibitors GM6001 and TAPI-0. Microparticle counts in culture supernatants were higher during etoposide-induced apoptosis. The ROCK1 inhibitor, Y27632, suppressed blebbing formation and microparticle release. Moreover, flow cytometry and immunoblotting revealed CD31 and CD47 in the microparticles. These results indicate that CD31 and CD47 were released by the apoptotic Jurkat cells into the culture supernatant in microparticles, but not in soluble forms, resulting in decreased levels on the apoptotic cell surface.

  16. Scanning the cell surface proteome of cancer cells and identification of metastasis-associated proteins using a subtractive immunization strategy

    DEFF Research Database (Denmark)

    Rasmussen, Nicolaj; Ditzel, Henrik J


    characterization of the identified proteins. The strategy is based on subtractive immunization of mice, and we used the two isogenic cell lines, NM-2C5 and M-4A4, derived from the MDA-MB-435 cancer cell line, as a model system. Although the two cell lines are equally tumorigenic, only M-4A4 has metastatic...... of the immune system itself to scan the cell surface proteome for differentially expressed proteins. The subtractive immunization strategy should be broadly applicable as a quantitative and comparative proteomic approach for screening the cell surface and also allow generation of mAbs to study these cell...

  17. Cell-surface alterations in class IIa bacteriocin-resistant Listeria monocytogenes strains. (United States)

    Vadyvaloo, Viveka; Arous, Safia; Gravesen, Anne; Héchard, Yann; Chauhan-Haubrock, Ramola; Hastings, John W; Rautenbach, Marina


    Strains of the food-borne pathogen Listeria monocytogenes, showing either intermediate or high-level resistance to class IIa bacteriocins, were investigated to determine characteristics that correlated with their sensitivity levels. Two intermediate and one highly resistant spontaneous mutant of L. monocytogenes B73, a highly resistant mutant of L. monocytogenes 412, and a highly resistant, defined (mptA) mutant of L. monocytogenes EGDe were compared with their respective wild-type strains in order to investigate the contribution of different factors to resistance. Decreased mannose-specific phosphotransferase system gene expression (mptA, EIIAB(Man) component) was implicated in all levels of resistance, confirming previous studies by the authors' group. However, a clear correlation between d-alanine content in teichoic acid (TA), in particular the alanine : phosphorus ratio, and a more positive cell surface, as determined by cytochrome c binding, were found for the highly resistant strains. Furthermore, two of the three highly resistant strains showed a significant increase in sensitivity towards d-cycloserine (DCS). However, real-time PCR of the dltA (d-alanine esterification), and dal and ddlA genes (peptidoglycan biosynthesis) showed no change in transcriptional levels. The link between DCS sensitivity and increased d-alanine esterification of TA may be that DCS competes with alanine for transport via the alanine transporter. A possible tendency towards increased lysinylation of membrane phospholipid in the highly resistant strains was also found. A previous study reported that cell membranes of all the resistant strains, including the intermediate resistant strains, contained more unsaturated phosphatidylglycerol, which is an indication of a more fluid cell membrane. The results of that study correlate with the possible lysinylation, decreased mptA expression, d-alanine esterification of TA and more positive cell surface charge found in this study for

  18. The involvement of proteoglycans in the human plasma prekallikrein interaction with the cell surface.

    Directory of Open Access Journals (Sweden)

    Camila Lopes Veronez

    Full Text Available INTRODUCTION: The aim of this work was to evaluate the role of human plasma prekallikrein assembly and processing in cells and to determine whether proteoglycans, along with high molecular weight kininogen (H-kininogen, influence this interaction. METHODS: We used the endothelial cell line ECV304 and the epithelial cell lines CHO-K1 (wild type and CHO-745 (deficient in proteoglycans. Prekallikrein endocytosis was studied using confocal microscopy, and prekallikrein cleavage/activation was determined by immunoblotting using an antibody directed to the prekallikrein sequence C364TTKTSTR371 and an antibody directed to the entire H-kininogen molecule. RESULTS: At 37°C, prekallikrein endocytosis was assessed in the absence and presence of exogenously applied H-kininogen and found to be 1,418.4±0.010 and 1,070.3±0.001 pixels/cell, respectively, for ECV304 and 1,319.1±0.003 and 631.3±0.001 pixels/cell, respectively, for CHO-K1. No prekallikrein internalization was observed in CHO-745 in either condition. Prekallikrein colocalized with LysoTracker in the absence and presence of exogenous H-kininogen at levels of 76.0% and 88.5%, respectively, for ECV304 and at levels of 40.7% and 57.0%, respectively, for CHO-K1. After assembly on the cell surface, a plasma kallikrein fragment of 53 kDa was predominant in the incubation buffer of all the cell lines studied, indicating specific proteolysis; plasma kallikrein fragments of 48-44 kDa and 34-32 kDa were also detected in the incubation buffer, indicating non-specific cleavage. Bradykinin free H-kininogen internalization was not detected in CHO-K1 or CHO-745 cells at 37°C. CONCLUSION: The prekallikrein interaction with the cell surface is temperature-dependent and independent of exogenously applied H-kininogen, which results in prekallikrein endocytosis promoted by proteoglycans. Prekallikrein proteolysis/activation is influenced by H-kininogen/glycosaminoglycans assembly and controls plasma kallikrein

  19. [Treatment of Cystic Fibrosis with CFTR Modulators]. (United States)

    Tümmler, B


    Personalized medicine promises that medical decisions, practices and products are tailored to the individual patient. Cystic fibrosis, an inherited disorder of chloride and bicarbonate transport in exocrine glands, is the first successful example of customized drug development for mutation-specific therapy. There are two classes of CFTR modulators: potentiators that increase the activity of CFTR at the cell surface, and correctors that either promote the read-through of nonsense mutations or facilitate the translation, folding, maturation and trafficking of mutant CFTR to the cell surface. The potentiator ivacaftor and the corrector lumacaftor are approved in Germany for the treatment of people with cystic fibrosis who carry a gating mutation such as p.Gly551Asp or who are homozygous for the most common mutation p.Phe508del, respectively. This report provides an overview of the basic defect in cystic fibrosis, the population genetics of CFTR mutations in Germany and the bioassays to assess CFTR function in humans together with the major achievements of preclinical research and clinical trials to bring CFTR modulators to the clinic. Some practical information on the use of ivacaftor and lumacaftor in daily practice and an update on pitfalls, challenges and novel strategies of bench-to-bedside development of CFTR modulators are also provided.

  20. Effects of elaidic acid, a predominant industrial trans fatty acid, on bacterial growth and cell surface hydrophobicity of lactobacilli. (United States)

    Wu, Qinglong; Shah, Nagendra P


    The consumption of trans fatty acids (TFAs) increases the risk of cardiovascular diseases and coronary heart disease in human, and there are no effective ways to remove TFAs after consumption. The aim of this study was to investigate the effects of elaidic acid on bacterial growth, cell surface hydrophobicity of lactobacilli, and metabolism of elaidic acid by lactobacilli. Lactobacilli were inoculated in MRS broth containing 0, 100, 200, and 500 mg/L of elaidic acid. Viable cell counts of lactobacilli were enumerated, concentrations of elaidic acid were determined, and cell surface hydrophobicity of lactobacilli was measured. The results showed that the growth of lactobacilli was significantly inhibited by 500 mg/L of elaidic acid, however, a cell count of 8.50 log10 CFU/mL was still reached for tested lactobacilli after 24-h incubation. In particular, a reduction of elaidic acid was found for tested lactobacilli after 24-h incubation as compared to its initial concentration of 200 mg/L. However, cell surface hydrophobicity showed no correlations with the metabolism of elaidic acid by lactobacilli. Moreover, elaidic acid was able to influence cell surface hydrophobicity, and the decrease in hydrophobicity was more obvious in Lactobacillus paracasei and Lactobacillus casei compared with that in other tested lactobacilli. This study suggests that elaidic acid could change physiochemical surface properties of lactobacilli and the lactobacilli have the potential to reduce TFAs.

  1. Synthesis and application of water-soluble, photoswitchable cyanine dyes for bioorthogonal labeling of cell-surface carbohydrates. (United States)

    Mertsch, Alexander; Letschert, Sebastian; Memmel, Elisabeth; Sauer, Markus; Seibel, Jürgen


    The synthesis of cyanine dyes addressing absorption wavelengths at 550 and 648 nm is reported. Alkyne functionalized dyes were used for bioorthogonal click reactions by labeling of metabolically incorporated sugar-azides on the surface of living neuroblastoma cells, which were applied to direct stochastic optical reconstruction microscopy (dSTORM) for the visualization of cell-surface glycans in the nm-range.

  2. Regulation of NKG2D-ligand cell surface expression by intracellular calcium after HDAC-inhibitor treatment

    DEFF Research Database (Denmark)

    Jensen, Helle; Hagemann-Jensen, Michael Henrik; Lauridsen, Felicia Kathrine Bratt;


    -cells. We further show that secretion and cell surface binding of the calcium-regulating protein galectin-1 is enhanced upon HDAC-inhibitor treatment of melanoma cells. However, binding of galectin-1 to cell surface glycoproteins was not critical for constitutive or HDAC-inhibitor induced MICA/B and ULBP2......In this study we demonstrate that histone deacetylase (HDAC)-inhibitor mediated cell surface expression of the structural different NKG2D-ligands MICA/B and ULBP2 is calcium-dependent. Treatment with the calcium chelator EGTA inhibited constitutive as well as HDAC-inhibitor induced MICA/B and ULBP2...... cell surface expression on melanoma cells and Jurkat T-cells. A NKG2D-dependent cytolytic assay and staining with a recombinant NKG2D-Fc fusion protein showed that calcium chelation impaired the functional ability of NKG2D-ligands induced by HDAC-inhibitor treatment. The HDAC-inhibitor induced cell...

  3. Effects of DNP on the cell surface properties of marine bacteria and its implication for adhesion to surfaces

    Digital Repository Service at National Institute of Oceanography (India)

    Jain, A.; Nishad, K.K.; Bhosle, N.B.

    was evaluated. Interestingly, after DNP treatment there was a significant decrease in the hydrophobicity of both hydrophobic (p = 0.01) and hydrophilic (p = 0.05) cultures, whereas no significant change was observed in the cell surface charge and EPS production...

  4. Kinesin-5/Eg5 is important for transport of CARTS from the trans-Golgi network to the cell surface (United States)

    Villeneuve, Julien; van Galen, Josse; Cruz-Garcia, David; Tagaya, Mitsuo


    Here we report that the kinesin-5 motor Klp61F, which is known for its role in bipolar spindle formation in mitosis, is required for protein transport from the Golgi complex to the cell surface in Drosophila S2 cells. Disrupting the function of its mammalian orthologue, Eg5, in HeLa cells inhibited secretion of a protein called pancreatic adenocarcinoma up-regulated factor (PAUF) but, surprisingly, not the trafficking of vesicular stomatitis virus G protein (VSV-G) to the cell surface. We have previously reported that PAUF is transported from the trans-Golgi network (TGN) to the cell surface in specific carriers called CARTS that exclude VSV-G. Inhibition of Eg5 function did not affect the biogenesis of CARTS; however, their migration was delayed and they accumulated near the Golgi complex. Altogether, our findings reveal a surprising new role of Eg5 in nonmitotic cells in the facilitation of the transport of specific carriers, CARTS, from the TGN to the cell surface. PMID:23857769

  5. One Step Quick Detection of Cancer Cell Surface Marker by Integrated NiFe-based Magnetic Biosensing Cell Cultural Chip

    Institute of Scientific and Technical Information of China (English)

    Chenchen Bao; Lei Chen; Tao Wang; Chong Lei; Furong Tian; Daxiang Cui; Yong Zhou


    RGD peptides has been used to detect cell surface integrin and direct clinical effective therapeutic drug selection. Herein we report that a quick one step detection of cell surface marker that was realized by a specially designed NiFe-based magnetic biosensing cell chip combined with functionalized magnetic nanoparti-cles. Magnetic nanoparticles with 20-30 nm in diameter were prepared by coprecipitation and modified with RGD-4C, and the resultant RGD-functionalized magnetic nanoparticles were used for targeting cancer cells cul-tured on the NiFe-based magnetic biosensing chip and distinguish the amount of cell surface receptor-integrin. Cell lines such as Calu3, Hela, A549, CaFbr, HEK293 and HUVEC exhibiting different integrin expression were chosen as test samples. Calu3, Hela, HEK293 and HUVEC cells were successfully identified. This approach has advantages in the qualitative screening test. Compared with traditional method, it is fast, sensitive, low cost, easy-operative, and needs very little human intervention. The novel method has great potential in applications such as fast clinical cell surface marker detection, and diagnosis of early cancer, and can be easily extended to other biomedical applications based on molecular recognition.

  6. Cell-surface changes in cadmium-resistant Euglena: Studies using lectin-binding techniques and flow cytometry

    Energy Technology Data Exchange (ETDEWEB)

    Bonaly, J.; Brochiero, E. [Faculte de Pharmacie, Chatenay-Malabry (France)


    Most in vitro studies on contaminants focus on the short-term effects of pollutants on cells, without regard to long-term effects and the ability of cells or microorganisms to develop a specific resistance to a pollutant. Cadmium is ubiquitous environmental contaminant. This heavy metal enters the aquatic environment mainly through vapor emissions and fallout during smelting operations. Diverse mechanisms of algal resistance to toxic metals are known. Among these, the most general mechanism is the development of metal-binding proteins. In cadmium-resistant unicellular Euglena gracilis Z algae cells, the metal did not appear to be sequestered on soluble metal-binding ligands. Previous experiments have shown that resistance development is related to a diminution of cadmium penetration into cells, implicating cell surface or membrane alteration. This research investigates the mechanisms of development of cadmium resistance in Euglena cells at the cell-surface level. Sugar chains of glycoproteins and glycolipids are a predominant feature of the surface of cells. Moreover, the cell-response to environmental changes is often orchestrated through surface macromolecules such as glycoproteins. In this study, we applied this lectin method to investigate surface carbohydrate expression during and after resistance development. Our interest was twofold: (1) to learn more about the carbohydrate composition of the cell-surface of Euglena; and (2) to determine whether transition from wild cells to Cd-resistant cells changes the expression of cell-surface carbohydrates. 13 refs., 2 figs., 1 tab.

  7. Effects of rhamnolipids on cell surface hydrophobicity of PAH degrading bacteria and the biodegradation of phenanthrene. (United States)

    Zhao, Zhenyong; Selvam, Ammaiyappan; Wong, Jonathan Woon-Chung


    The effects of rhamnolipids produced by Pseudomonas aeruginosa ATCC9027 on the cell surface hydrophobicity (CSH) and the biodegradation of phenanthrene by two thermophilic bacteria, Bacillus subtilis BUM and P. aeruginosa P-CG3, and mixed inoculation of these two strains were investigated. Rhamnolipids significantly reduced the CSH of the hydrophobic BUM and resulted in a noticeable lag period in the biodegradation. However, they significantly increased the CSH and enhanced the biodegradation for the hydrophilic P-CG3. In the absence of rhamnolipids, a mixed inoculation of BUM and P-CG3 removed 82.2% of phenanthrene within 30 days and the major contributor of the biodegradation was BUM (rapid degrader) while the growth of P-CG3 (slow degrader) was suppressed. Addition of rhamnolipids promoted the surfactant-mediated-uptake of phenanthrene by P-CG3 but inhibited the uptake through direct contact by BUM. This resulted in the domination of P-CG3 during the initial stage of biodegradation and enhanced the biodegradation to 92.7%.

  8. Facile Discovery of Cell-Surface Protein Targets of Cancer Cell Aptamers. (United States)

    Bing, Tao; Shangguan, Dihua; Wang, Yinsheng


    Cancer biomarker discovery constitutes a frontier in cancer research. In recent years, cell-binding aptamers have become useful molecular probes for biomarker discovery. However, there are few successful examples, and the critical barrier resides in the identification of the cell-surface protein targets for the aptamers, where only a limited number of aptamer targets have been identified so far. Herein, we developed a universal SILAC-based quantitative proteomic method for target discovery of cell-binding aptamers. The method allowed for distinguishing specific aptamer-binding proteins from nonspecific proteins based on abundance ratios of proteins bound to aptamer-carrying bait and control bait. In addition, we employed fluorescently labeled aptamers for monitoring and optimizing the binding conditions. We were able to identify and validate selectin L and integrin α4 as the protein targets for two previously reported aptamers, Sgc-3b and Sgc-4e, respectively. This strategy should be generally applicable for the discovery of protein targets for other cell-binding aptamers, which will promote the applications of these aptamers.

  9. Cell surface phenotype of cytolytic T lymphocyte precursors in aged nude mice. (United States)

    Maryanski, J L; MacDonald, H R; Sordat, B; Cerottini, J C


    The cell surface phenotype of cytolytic T lymphocyte precursors (CTL-P) in congenitally athymic C57BL/6 nu/nu mice has been investigated. CTL-P were detected and quantitated in a limited dilution mixed leukocyte microculture assay system supplemented with interleukin 2. Minimal estimates of the frequency of CTL-P among nylon wool passed (NWP) nude spleen cells were obtained following elimination of Thy-1-bearing or Lyt-2-bearing cells with monoclonal antibodies plus complement. Alternatively, NWP spleen cells bearing Thy-1 or Lyt-2 were positively selected on a cell sorter and assayed for CTL-P frequency. Both positive and negative selection techniques demonstrated that essentially all (greater than 98%) CTL-P in NWP nude spleen expressed Thy-1 and that the majority (80-90%) expressed Lyt-2. In control NWP spleen cells from normal C57BL/6 mice, greater than 98% of CTL-P were positive for both Thy-1 and Lyt-2. These data demonstrate that most functional alloreactive CTL-P developing in the apparent absence of thymic influence already express both Thy-1 and Lyt-2 prior to exposure to antigen.

  10. Enhanced Growth Inhibition of Osteosarcoma by Cytotoxic Polymerized Liposomal Nanoparticles Targeting the Alcam Cell Surface Receptor

    Directory of Open Access Journals (Sweden)

    Noah Federman


    Full Text Available Osteosarcoma is the most common primary malignancy of bone in children, adolescents, and adults. Despite extensive surgery and adjuvant aggressive high-dose systemic chemotherapy with potentially severe bystander side effects, cure is attainable in about 70% of patients with localized disease and only 20%–30% of those patients with metastatic disease. Targeted therapies clearly are warranted in improving our treatment of this adolescent killer. However, a lack of osteosarcoma-associated/specific markers has hindered development of targeted therapeutics. We describe a novel osteosarcoma-associated cell surface antigen, ALCAM. We, then, create an engineered anti-ALCAM-hybrid polymerized liposomal nanoparticle immunoconjugate (α-AL-HPLN to specifically target osteosarcoma cells and deliver a cytotoxic chemotherapeutic agent, doxorubicin. We have demonstrated that α-AL-HPLNs have significantly enhanced cytotoxicity over untargeted HPLNs and over a conventional liposomal doxorubicin formulation. In this way, α-AL-HPLNs are a promising new strategy to specifically deliver cytotoxic agents in osteosarcoma.

  11. A Gravity-Responsive Time-Keeping Protein of the Plant and Animal Cell Surface (United States)

    Morre, D. James


    The hypothesis under investigation was that a ubiquinol (NADH) oxidase protein of the cell surface with protein disulfide-thiol interchange activity (= NOX protein) is a plant and animal time-keeping ultradian (period of less than 24 h) driver of both cell enlargement and the biological clock that responds to gravity. Despite considerable work in a large number of laboratories spanning several decades, this is, to my knowledge, our work is the first demonstration of a time-keeping biochemical reaction that is both gravity-responsive and growth-related and that has been shown to determine circadian periodicity. As such, the NOX protein may represent both the long-sought biological gravity receptor and the core oscillator of the cellular biological clock. Completed studies have resulted in 12 publications and two issued NASA-owned patents of the clock activity. The gravity response and autoentrainment were characterized in cultured mammalian cells and in two plant systems together with entrainment by light and small molecules (melatonin). The molecular basis of the oscillatory behavior was investigated using spectroscopic methods (Fourier transform infrared and circular dichroism) and high resolution electron microscopy. We have also applied these findings to an understanding of the response to hypergravity. Statistical methods for analysis of time series phenomena were developed (Foster et al., 2003).

  12. Bioadsorption of Rare Earth Elements through Cell Surface Display of Lanthanide Binding Tags. (United States)

    Park, Dan M; Reed, David W; Yung, Mimi C; Eslamimanesh, Ali; Lencka, Malgorzata M; Anderko, Andrzej; Fujita, Yoshiko; Riman, Richard E; Navrotsky, Alexandra; Jiao, Yongqin


    With the increasing demand for rare earth elements (REEs) in many emerging clean energy technologies, there is an urgent need for the development of new approaches for efficient REE extraction and recovery. As a step toward this goal, we genetically engineered the aerobic bacterium Caulobacter crescentus for REE adsorption through high-density cell surface display of lanthanide binding tags (LBTs) on its S-layer. The LBT-displayed strains exhibited enhanced adsorption of REEs compared to cells lacking LBT, high specificity for REEs, and an adsorption preference for REEs with small atomic radii. Adsorbed Tb(3+) could be effectively recovered using citrate, consistent with thermodynamic speciation calculations that predicted strong complexation of Tb(3+) by citrate. No reduction in Tb(3+) adsorption capacity was observed following citrate elution, enabling consecutive adsorption/desorption cycles. The LBT-displayed strain was effective for extracting REEs from the acid leachate of core samples collected at a prospective rare earth mine. Our collective results demonstrate a rapid, efficient, and reversible process for REE adsorption with potential industrial application for REE enrichment and separation.

  13. Modeling multivalent ligand-receptor interactions with steric constraints on configurations of cell surface receptor aggregates

    Energy Technology Data Exchange (ETDEWEB)

    Monine, Michael [Los Alamos National Laboratory; Posner, Richard [TRANSLATION GENOMICS RESAEARCH INSTITUTE; Savage, Paul [BYU; Faeder, James [UNIV OF PITTSBURGH; Hlavacek, William S [UNM


    Signal transduction generally involves multivalent protein-protein interactions, which can produce various protein complexes and post-translational modifications. The reaction networks that characterize these interactions tend to be so large as to challenge conventional simulation procedures. To address this challenge, a kinetic Monte Carlo (KMC) method has been developed that can take advantage of a model specification in terms of reaction rules for molecular interactions. A set of rules implicitly defines the reactions that can occur as a result of the interactions represented by the rules. With the rule-based KMC method, explicit generation of the underlying chemical reaction network implied by rules is avoided. Here, we apply and extend this method to characterize the interactions of a trivalent ligand with a bivalent cell-surface receptor. This system is also studied experimentally. We consider the following kinetic models: an equivalent-site model, an extension of this model, which takes into account steric constraints on the configurations of receptor aggregates, and finally, a model that accounts for cyclic receptor aggregates. Simulation results for the equivalent-site model are consistent with an equilibrium continuum model. Using these models, we investigate the effects of steric constraints and the formation of cyclic aggregates on the kinetics and equilibria of small and large aggregate formation and the percolation phase transition that occurs in this system.

  14. Scaffold Free Bio-orthogonal Assembly of 3-Dimensional Cardiac Tissue via Cell Surface Engineering (United States)

    Rogozhnikov, Dmitry; O’Brien, Paul J.; Elahipanah, Sina; Yousaf, Muhammad N.


    There has been tremendous interest in constructing in vitro cardiac tissue for a range of fundamental studies of cardiac development and disease and as a commercial system to evaluate therapeutic drug discovery prioritization and toxicity. Although there has been progress towards studying 2-dimensional cardiac function in vitro, there remain challenging obstacles to generate rapid and efficient scaffold-free 3-dimensional multiple cell type co-culture cardiac tissue models. Herein, we develop a programmed rapid self-assembly strategy to induce specific and stable cell-cell contacts among multiple cell types found in heart tissue to generate 3D tissues through cell-surface engineering based on liposome delivery and fusion to display bio-orthogonal functional groups from cell membranes. We generate, for the first time, a scaffold free and stable self assembled 3 cell line co-culture 3D cardiac tissue model by assembling cardiomyocytes, endothelial cells and cardiac fibroblast cells via a rapid inter-cell click ligation process. We compare and analyze the function of the 3D cardiac tissue chips with 2D co-culture monolayers by assessing cardiac specific markers, electromechanical cell coupling, beating rates and evaluating drug toxicity.

  15. Neisseria lactamica selectively induces mitogenic proliferation of the naive B cell pool via cell surface Ig. (United States)

    Vaughan, Andrew T; Brackenbury, Louise S; Massari, Paola; Davenport, Victoria; Gorringe, Andrew; Heyderman, Robert S; Williams, Neil A


    Neisseria lactamica is a commensal bacteria that colonizes the human upper respiratory tract mucosa during early childhood. In contrast to the closely related opportunistic pathogen Neisseria meningitidis, there is an absence of adaptive cell-mediated immunity to N. lactamica during the peak age of carriage. Instead, outer membrane vesicles derived from N. lactamica mediate a B cell-dependent proliferative response in mucosal mononuclear cells that is associated with the production of polyclonal IgM. We demonstrate in this study that this is a mitogenic human B cell response that occurs independently of T cell help and any other accessory cell population. The ability to drive B cell proliferation is a highly conserved property and is present in N. lactamica strains derived from diverse clonal complexes. CFSE staining of purified human tonsillar B cells demonstrated that naive IgD(+) and CD27(-) B cells are selectively induced to proliferate by outer membrane vesicles, including the innate CD5(+) subset. Neither purified lipooligosaccharide nor PorB from N. lactamica is likely to be responsible for this activity. Prior treatment of B cells with pronase to remove cell-surface Ig or treatment with BCR-specific Abs abrogated the proliferative response to N. lactamica outer membrane vesicles, suggesting that this mitogenic response is dependent upon the BCR.

  16. Class I major histocompatibility proteins as cell surface receptors for simian virus 40. (United States)

    Atwood, W J; Norkin, L C


    Class I major histocompatibility complex proteins appear to be the major cell surface receptors for simian virus 40 (SV40), as implied by the following observations. Adsorption of SV40 to LLC-MK2 rhesus monkey kidney cells specifically inhibited binding of a monoclonal antibody (MAb) against class I human lymphocyte antigen (HLA) proteins. Conversely, pretreatment of LLC-MK2 cells with anti-HLA MAbs inhibited infection by SV40. The ability of anti-HLA to inhibit infection was greatly reduced when the order of addition of the anti-HLA and the virus was reversed. Infection was also inhibited by preincubating SV40 with purified soluble class I protein. Finally, human lymphoblastoid cells of the Daudi line, which do not express class I major histocompatibility complex proteins, were infected at relatively low levels with SV40 virions. In a control experiment, we found that pretreatment of cells with a MAb specific for the leukocytic-function-associated antigen LFA-3 actually enhanced infection. This finding may also support the premise that class I major histocompatibility complex proteins are receptors for SV40.

  17. Interference with ubiquitination in CFTR modifies stability of core glycosylated and cell surface pools. (United States)

    Lee, Seakwoo; Henderson, Mark J; Schiffhauer, Eric; Despanie, Jordan; Henry, Katherine; Kang, Po Wei; Walker, Douglas; McClure, Michelle L; Wilson, Landon; Sorscher, Eric J; Zeitlin, Pamela L


    It is recognized that both wild-type and mutant CFTR proteins undergo ubiquitination at multiple lysines in the proteins and in one or more subcellular locations. We hypothesized that ubiquitin is added to specific sites in wild-type CFTR to stabilize it and at other sites to signal for proteolysis. Mass spectrometric analysis of wild-type CFTR identified ubiquitinated lysines 68, 710, 716, 1041, and 1080. We demonstrate that the ubiquitinated K710, K716, and K1041 residues stabilize wild-type CFTR, protecting it from proteolysis. The polyubiquitin linkage is predominantly K63. N-tail mutants, K14R and K68R, lead to increased mature band CCFTR, which can be augmented by proteasomal (but not lysosomal) inhibition, allowing trafficking to the surface. The amount of CFTR in the K1041R mutant was drastically reduced and consisted of bands A/B, suggesting that the site in transmembrane 10 (TM10) was critical to further processing beyond the proteasome. The K1218R mutant increases total and cell surface CFTR, which is further accumulated by proteasomal and lysosomal inhibition. Thus, ubiquitination at residue 1218 may destabilize wild-type CFTR in both the endoplasmic reticulum (ER) and recycling pools. Small molecules targeting the region of residue 1218 to block ubiquitination or to preserving structure at residues 710 to 716 might be protein sparing for some forms of cystic fibrosis.

  18. Recruitment of Factor H to the Streptococcus suis Cell Surface is Multifactorial

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    David Roy


    Full Text Available Streptococcus suis is an important bacterial swine pathogen and a zoonotic agent. Recently, two surface proteins of S. suis, Fhb and Fhbp, have been described for their capacity to bind factor H—a soluble complement regulatory protein that protects host cells from complement-mediated damages. Results obtained in this study showed an important role of host factor H in the adhesion of S. suis to epithelial and endothelial cells. Both Fhb and Fhbp play, to a certain extent, a role in such increased factor H-dependent adhesion. The capsular polysaccharide (CPS of S. suis, independently of the presence of its sialic acid moiety, was also shown to be involved in the recruitment of factor H. However, a triple mutant lacking Fhb, Fhbp and CPS was still able to recruit factor H resulting in the degradation of C3b in the presence of factor I. In the presence of complement factors, the double mutant lacking Fhb and Fhbp was similarly phagocytosed by human macrophages and killed by pig blood when compared to the wild-type strain. In conclusion, this study suggests that recruitment of factor H to the S. suis cell surface is multifactorial and redundant.

  19. Cell surface and transcriptional characterization of human adipose-derived adherent stromal (hADAS) cells. (United States)

    Katz, Adam J; Tholpady, Ashok; Tholpady, Sunil S; Shang, Hulan; Ogle, Roy C


    Adult human subcutaneous adipose tissue contains cells with intriguing multilineage developmental plasticity, much like marrow-derived mesenchymal stem cells. Putative stem or progenitor cells from fat have been given many different names in the literature, reflecting an early and evolving consensus regarding their phenotypic characterization. The study reported here used microarrays to evaluate over 170 genes relating to angiogenesis and extracellular matrix in undifferentiated, early-passage human adipose-derived adherent stromal (hADAS) cells isolated from three separate donors. The hADAS populations unanimously transcribed 66% of the screened genes, and 83% were transcribed by at least two of the three populations. The most highly transcribed genes relate to functional groupings such as cell adhesion, matrix proteins, growth factors and receptors, and proteases. The transcriptome of hADAS cells demonstrated by this work reveals many similarities to published profiles of bone marrow mesenchymal stem cells (MSCs). In addition, flow analysis of over 24 hADAS cell surface proteins (n = 7 donors) both confirms and expands on the existing literature and reveals strong intergroup correlation, despite an inconsistent nomenclature and the lack of standardized protocols for cell isolation and culture. Finally, based on flow analysis and reverse transcription polymerase chain reaction studies, our results suggest that hADAS cells do not express several proteins that are implicated as markers of "stemness" in other stem cell populations, including telomerase, CD133, and the membrane transporter ABCG2.

  20. Recruitment of Factor H to the Streptococcus suis Cell Surface is Multifactorial. (United States)

    Roy, David; Grenier, Daniel; Segura, Mariela; Mathieu-Denoncourt, Annabelle; Gottschalk, Marcelo


    Streptococcus suis is an important bacterial swine pathogen and a zoonotic agent. Recently, two surface proteins of S. suis, Fhb and Fhbp, have been described for their capacity to bind factor H-a soluble complement regulatory protein that protects host cells from complement-mediated damages. Results obtained in this study showed an important role of host factor H in the adhesion of S. suis to epithelial and endothelial cells. Both Fhb and Fhbp play, to a certain extent, a role in such increased factor H-dependent adhesion. The capsular polysaccharide (CPS) of S. suis, independently of the presence of its sialic acid moiety, was also shown to be involved in the recruitment of factor H. However, a triple mutant lacking Fhb, Fhbp and CPS was still able to recruit factor H resulting in the degradation of C3b in the presence of factor I. In the presence of complement factors, the double mutant lacking Fhb and Fhbp was similarly phagocytosed by human macrophages and killed by pig blood when compared to the wild-type strain. In conclusion, this study suggests that recruitment of factor H to the S. suis cell surface is multifactorial and redundant.