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Sample records for affect microtubule dynamics

  1. Dynamics of Microtubule Instabilities

    CERN Document Server

    Antal, T; Redner, S

    2007-01-01

    We investigate the dynamics of an idealized model of microtubule growth that evolves by: (i) attachment of guanosine triphosphate (GTP) at rate lambda, (ii) conversion of GTP to guanosine diphosphate (GDP) at rate 1, and (iii) detachment of GDP at rate mu. As a function of these rates, a microtubule can grow steadily or its length can fluctuate wildly. For mu=0, we find the exact tubule and GTP cap length distributions, and power-law length distributions of GTP and GDP islands. For mu=infinity, we argue that the time between catastrophes, where the microtubule shrinks to zero length, scales as exp(lambda). We also find the phase boundary between a growing and shrinking microtubule.

  2. Aging of dynamically stabilized microtubules

    CERN Document Server

    Ebbinghaus, M

    2009-01-01

    The microtubule network, an important part of the cytoskeleton, is constantly remodeled by alternating phases of growth and shrinkage of individual filaments. Plus-end tracking proteins (+TIPs) interact with the microtubule and in many cases alter its dynamics. While it is established that the prototypal CLIP-170 enhances microtubule stability by increasing rescues, the plus-end tracking mechanism is still under debate. We present a model for microtubule dynamics in which a rescue factor is dynamically added to the filament while growing. As a consequence, the filament shows aging behavior which should be experimentally accessible and thus allow one to exclude some hypothesized models of the inclusion of rescue factors at the microtubule plus end. Additionally, we show the strong influence of the cell geometry on the quantitative results.

  3. Catechol-o-methyltransferase expression and 2-methoxyestradiol affect microtubule dynamics and modify steroid receptor signaling in leiomyoma cells.

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    Salama A Salama

    Full Text Available CONTEXT: Development of optimal medicinal treatments of uterine leiomyomas represents a significant challenge. 2-Methoxyestradiol (2ME is an endogenous estrogen metabolite formed by sequential action of CYP450s and catechol-O-methyltransferase (COMT. Our previous study demonstrated that 2ME is a potent antiproliferative, proapoptotic, antiangiogenic, and collagen synthesis inhibitor in human leiomyomas cells (huLM. OBJECTIVES: Our objectives were to investigate whether COMT expression, by the virtue of 2ME formation, affects the growth of huLM, and to explore the cellular and molecular mechanisms whereby COMT expression or treatment with 2ME affect these cells. RESULTS: Our data demonstrated that E(2-induced proliferation was less pronounced in cells over-expressing COMT or treated with 2ME (500 nM. This effect on cell proliferation was associated with microtubules stabilization and diminution of estrogen receptor alpha (ERalpha and progesterone receptor (PR transcriptional activities, due to shifts in their subcellular localization and sequestration in the cytoplasm. In addition, COMT over expression or treatment with 2ME reduced the expression of hypoxia-inducible factor -1alpha (HIF-1 alpha and the basal level as well as TNF-alpha-induced aromatase (CYP19 expression. CONCLUSIONS: COMT over expression or treatment with 2ME stabilize microtubules, ameliorates E(2-induced proliferation, inhibits ERalpha and PR signaling, and reduces HIF-1 alpha and CYP19 expression in human uterine leiomyoma cells. Thus, microtubules are a candidate target for treatment of uterine leiomyomas. In addition, the naturally occurring microtubule-targeting agent 2ME represents a potential new therapeutic for uterine leiomyomas.

  4. Visualization of microtubule growth in living platelets reveals a dynamic marginal band with multiple microtubules

    NARCIS (Netherlands)

    S. Patel-Hett (Sunita); J.L. Richardson (Jennifer); H. Schulze (Harald); K. Drabek (Ksenija); N.A. Isaac (Natasha); K. Hoffmeister (Karin); R.A. Shivdasani (Ramesh); J.C. Bulinski (J. Chloë); N.J. Galjart (Niels); J.H. Hartwig (John); J. Italiano (Joseph)

    2008-01-01

    textabstractThe marginal band of microtubules maintains the discoid shape of resting blood platelets. Although studies of platelet microtubule coil structure conclude that it is composed of a single microtubule, no investigations of its dynamics exist. In contrast to previous studies, permeabilized

  5. Dynamics and regulation of plant interphase microtubules: a comparative view.

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    Hashimoto, Takashi

    2003-12-01

    Microtubule and actin cytoskeletons are fundamental to a variety of cellular activities within eukaryotic organisms. Extensive information on the dynamics and functions of microtubules, as well as on their regulatory proteins, have been revealed in fungi and animals, and corresponding pictures are now slowly emerging in plants. During interphase, plant cells contain highly dynamic cortical microtubules that organize into ordered arrays, which are apparently regulated by distinct groups of microtubule regulators. Comparison with fungal and animal microtubules highlights both conserved and unique mechanisms for the regulation of the microtubule cytoskeleton in plants.

  6. Microtubules: dynamically unstable stochastic phase-switching polymers

    Science.gov (United States)

    Zakharov, P. N.; Arzhanik, V. K.; Ulyanov, E. V.; Gudimchuk, N. B.; Ataullakhanov, F. I.

    2016-08-01

    One of the simplest molecular motors, a biological microtubule, is reviewed as an example of a highly nonequilibrium molecular machine capable of stochastic transitions between slow growth and rapid disassembly phases. Basic properties of microtubules are described, and various approaches to simulating their dynamics, from statistical chemical kinetics models to molecular dynamics models using the Metropolis Monte Carlo and Brownian dynamics methods, are outlined.

  7. Deceivingly dynamic: Learning-dependent changes in stathmin and microtubules.

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    Uchida, Shusaku; Shumyatsky, Gleb P

    2015-10-01

    Microtubules, one of the major cytoskeletal structures, were previously considered stable and only indirectly involved in synaptic structure and function in mature neurons. However, recent evidence demonstrates that microtubules are dynamic and have an important role in synaptic structure, synaptic plasticity, and memory. In particular, learning induces changes in microtubule turnover and stability, and pharmacological manipulation of microtubule dynamics alters synaptic plasticity and long-term memory. These learning-induced changes in microtubules are controlled by the phosphoprotein stathmin, whose only known cellular activity is to negatively regulate microtubule formation. During the first eight hours following learning, changes in the phosphorylation of stathmin go through two phases causing biphasic shifts in microtubules stability/instability. These shifts, in turn, regulate memory formation by controlling in the second phase synaptic transport of the GluA2 subunit of AMPA receptors. Improper regulation of stathmin and microtubule dynamics has been observed in aged animals and in patients with Alzheimer's disease and depression. Thus, recent work on stathmin and microtubules has identified new molecular players in the early stages of memory encoding.

  8. The Role of Molecular Microtubule Motors and the Microtubule Cytoskeleton in Stress Granule Dynamics

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    Kristen M. Bartoli

    2011-01-01

    Full Text Available Stress granules (SGs are cytoplasmic foci that appear in cells exposed to stress-induced translational inhibition. SGs function as a triage center, where mRNAs are sorted for storage, degradation, and translation reinitiation. The underlying mechanisms of SGs dynamics are still being characterized, although many key players have been identified. The main components of SGs are stalled 48S preinitiation complexes. To date, many other proteins have also been found to localize in SGs and are hypothesized to function in SG dynamics. Most recently, the microtubule cytoskeleton and associated motor proteins have been demonstrated to function in SG dynamics. In this paper, we will discuss current literature examining the function of microtubules and the molecular microtubule motors in SG assembly, coalescence, movement, composition, organization, and disassembly.

  9. Calcium-independent disruption of microtubule dynamics by nanosecond pulsed electric fields in U87 human glioblastoma cells.

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    Carr, Lynn; Bardet, Sylvia M; Burke, Ryan C; Arnaud-Cormos, Delia; Leveque, Philippe; O'Connor, Rodney P

    2017-01-24

    High powered, nanosecond duration, pulsed electric fields (nsPEF) cause cell death by a mechanism that is not fully understood and have been proposed as a targeted cancer therapy. Numerous chemotherapeutics work by disrupting microtubules. As microtubules are affected by electrical fields, this study looks at the possibility of disrupting them electrically with nsPEF. Human glioblastoma cells (U87-MG) treated with 100, 10 ns, 44 kV/cm pulses at a frequency of 10 Hz showed a breakdown of their interphase microtubule network that was accompanied by a reduction in the number of growing microtubules. This effect is temporally linked to loss of mitochondrial membrane potential and independent of cellular swelling and calcium influx, two factors that disrupt microtubule growth dynamics. Super-resolution microscopy revealed microtubule buckling and breaking as a result of nsPEF application, suggesting that nsPEF may act directly on microtubules.

  10. Calcium-independent disruption of microtubule dynamics by nanosecond pulsed electric fields in U87 human glioblastoma cells

    Science.gov (United States)

    Carr, Lynn; Bardet, Sylvia M.; Burke, Ryan C.; Arnaud-Cormos, Delia; Leveque, Philippe; O’Connor, Rodney P.

    2017-01-01

    High powered, nanosecond duration, pulsed electric fields (nsPEF) cause cell death by a mechanism that is not fully understood and have been proposed as a targeted cancer therapy. Numerous chemotherapeutics work by disrupting microtubules. As microtubules are affected by electrical fields, this study looks at the possibility of disrupting them electrically with nsPEF. Human glioblastoma cells (U87-MG) treated with 100, 10 ns, 44 kV/cm pulses at a frequency of 10 Hz showed a breakdown of their interphase microtubule network that was accompanied by a reduction in the number of growing microtubules. This effect is temporally linked to loss of mitochondrial membrane potential and independent of cellular swelling and calcium influx, two factors that disrupt microtubule growth dynamics. Super-resolution microscopy revealed microtubule buckling and breaking as a result of nsPEF application, suggesting that nsPEF may act directly on microtubules. PMID:28117459

  11. The kinesin-13 KLP10A motor regulates oocyte spindle length and affects EB1 binding without altering microtubule growth rates

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    Kevin K. Do

    2014-06-01

    Full Text Available Kinesin-13 motors are unusual in that they do not walk along microtubules, but instead diffuse to the ends, where they remove tubulin dimers, regulating microtubule dynamics. Here we show that Drosophila kinesin-13 klp10A regulates oocyte meiosis I spindle length and is haplo-insufficient – KLP10A, reduced by RNAi or a loss-of-function P element insertion mutant, results in elongated and mispositioned oocyte spindles, and abnormal cortical microtubule asters and aggregates. KLP10A knockdown by RNAi does not significantly affect microtubule growth rates in oocyte spindles, but, unexpectedly, EB1 binding and unbinding are slowed, suggesting a previously unobserved role for kinesin-13 in mediating EB1 binding interactions with microtubules. Kinesin-13 may regulate spindle length both by disassembling subunits from microtubule ends and facilitating EB1 binding to plus ends. We also observe an increased number of paused microtubules in klp10A RNAi knockdown spindles, consistent with a reduced frequency of microtubule catastrophes. Overall, our findings indicate that reduced kinesin-13 decreases microtubule disassembly rates and affects EB1 interactions with microtubules, rather than altering microtubule growth rates, causing spindles to elongate and abnormal cortical microtubule asters and aggregates to form.

  12. Integrated modeling methodology for microtubule dynamics and Taxol kinetics with experimentally identifiable parameters.

    Science.gov (United States)

    Zhao, He; Sokhansanj, Bahrad A

    2007-10-01

    Microtubule dynamics play a critical role in cell function and stress response, modulating mitosis, morphology, signaling, and transport. Drugs such as paclitaxel (Taxol) can impact tubulin polymerization and affect microtubule dynamics. While theoretical methods have been previously proposed to simulate microtubule dynamics, we develop a methodology here that can be used to compare model predictions with experimental data. Our model is a hybrid of (1) a simple two-state stochastic formulation of tubulin polymerization kinetics and (2) an equilibrium approximation for the chemical kinetics of Taxol drug binding to microtubule ends. Model parameters are biologically realistic, with values taken directly from experimental measurements. Model validation is conducted against published experimental data comparing optical measurements of microtubule dynamics in cultured cells under normal and Taxol-treated conditions. To compare model predictions with experimental data requires applying a "windowing" strategy on the spatiotemporal resolution of the simulation. From a biological perspective, this is consistent with interpreting the microtubule "pause" phenomenon as at least partially an artifact of spatiotemporal resolution limits on experimental measurement.

  13. Microtubule dynamics in the peripheral nervous system: A matter of balance.

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    Almeida-Souza, Leonardo; Timmerman, Vincent; Janssens, Sophie

    2011-11-01

    The special architecture of neurons in the peripheral nervous system, with axons extending for long distances, represents a major challenge for the intracellular transport system. Two recent studies show that mutations in the small heat shock protein HSPB1, which cause an axonal type of Charcot-Marie-Tooth (CMT) neuropathy, affect microtubule dynamics and impede axonal transport. Intriguingly, while at presymptomatic age the neurons in the mutant HSPB1 mouse show a hyperstable microtubule network, at postsymptomatic age, the microtubule network completely lost its stability as reflected by a marked decrease in tubulin acetylation levels. We here propose a model explaining the role of microtubule stabilization and tubulin acetylation in the pathogenesis of HSPB1 mutations.

  14. Modeling the Effects of Drug Binding on the Dynamic Instability of Microtubules

    CERN Document Server

    Hinow, Peter; Lopus, Manu; Jordan, Mary Ann; Tuszynski, Jack A

    2010-01-01

    We propose a stochastic model that accounts for the growth, catastrophe and rescue processes of steady state microtubules assembled from MAP-free tubulin. Both experimentally and theoretically we study the perturbation of microtubule dynamic instability by S-methyl-D-DM1, a synthetic derivative of the microtubule-targeted agent maytansine and a potential anticancer agent. We find that to be an effective suppressor of microtubule dynamics a drug must primarily suppress the loss of GDP tubulin from the microtubule tip.

  15. Nonlinear Dynamics of Dipoles in Microtubules: Pseudo-Spin Model

    CERN Document Server

    Nesterov, Alexander I; Berman, Gennady P; Mavromatos, Nick E

    2016-01-01

    We perform a theoretical study of the dynamics of the electric field excitations in a microtubule by taking into consideration the realistic cylindrical geometry, dipole-dipole interactions of the tubulin-based protein heterodimers, the radial electric field produced by the solvent, and a possible degeneracy of energy states of individual heterodimers. The consideration is done in the frames of the classical pseudo-spin model. We derive the system of nonlinear dynamical ordinary differential equations of motion for interacting dipoles, and the continuum version of these equations. We obtain the solutions of these equations in the form of snoidal waves, solitons, kinks, and localized spikes. Our results will help to a better understanding of the functional properties of microtubules including the motor protein dynamics and the information transfer processes. Our considerations are based on classical dynamics. Some speculations on the role of possible quantum effects are also made.

  16. Nonlinear dynamics of dipoles in microtubules: Pseudospin model.

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    Nesterov, Alexander I; Ramírez, Mónica F; Berman, Gennady P; Mavromatos, Nick E

    2016-06-01

    We perform a theoretical study of the dynamics of the electric field excitations in a microtubule by taking into consideration the realistic cylindrical geometry, dipole-dipole interactions of the tubulin-based protein heterodimers, the radial electric field produced by the solvent, and a possible degeneracy of energy states of individual heterodimers. The consideration is done in the frame of the classical pseudospin model. We derive the system of nonlinear dynamical partial differential equations of motion for interacting dipoles and the continuum version of these equations. We obtain the solutions of these equations in the form of snoidal waves, solitons, kinks, and localized spikes. Our results will help to achieve a better understanding of the functional properties of microtubules including the motor protein dynamics and the information transfer processes. Our considerations are based on classical dynamics. Some speculations on the role of possible quantum effects are also made.

  17. Microtubule Dynamicity Is More Important than Stability in Memory Formation: an In Vivo Study.

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    Atarod, Deyhim; Eskandari-Sedighi, Ghazaleh; Pazhoohi, Farid; Karimian, Seyed Morteza; Khajeloo, Mojtaba; Riazi, Gholam Hossein

    2015-06-01

    It has been shown that microtubule (MT) activity and dynamics can have huge impacts on synaptic plasticity and memory formation. This is mainly due to various functions of MTs in neurons; MTs are involved in dendritic spine formation, axonal transportation, neuronal polarity, and receptor trafficking. Recent studies from our group and other labs have suggested the possible role of brain MT dynamicity and activity in memory; however, there is a need for more detailed studies regarding this aspect. In this study, we have tried to evaluate the importance of microtubule dynamicity rather than stability in memory formation in vivo. In order to investigate the role of MT stability in memory formation, we treated mice with paclitaxel-a classic microtubule-stabilizing agent. We then studied the behavior of treated animals using Morris water maze (MWM) test. To measure the effect of injected paclitaxel on MT polymerization kinetics, we conducted polymerization assays on brain extracts of the same paclitaxel-treated animals. Our results show that paclitaxel treatment affects animals' memory in a negative way and treated animals behave poorly in MWM compared to control group. In addition, our kinetics studies show that MT stability is significantly increased in brain extracts from paclitaxel-treated mice, but MT dynamics is reduced. Thus, we suggest that dynamicity is a very important feature of MT protein structures, and regarding memory formation, dynamicity is more important than stability and high activity.

  18. TOG Proteins Are Spatially Regulated by Rac-GSK3β to Control Interphase Microtubule Dynamics.

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    Kathryn P Trogden

    Full Text Available Microtubules are regulated by a diverse set of proteins that localize to microtubule plus ends (+TIPs where they regulate dynamic instability and mediate interactions with the cell cortex, actin filaments, and organelles. Although individual +TIPs have been studied in depth and we understand their basic contributions to microtubule dynamics, there is a growing body of evidence that these proteins exhibit cross-talk and likely function to collectively integrate microtubule behavior and upstream signaling pathways. In this study, we have identified a novel protein-protein interaction between the XMAP215 homologue in Drosophila, Mini spindles (Msps, and the CLASP homologue, Orbit. These proteins have been shown to promote and suppress microtubule dynamics, respectively. We show that microtubule dynamics are regionally controlled in cells by Rac acting to suppress GSK3β in the peripheral lamellae/lamellipodium. Phosphorylation of Orbit by GSK3β triggers a relocalization of Msps from the microtubule plus end to the lattice. Mutation of the Msps-Orbit binding site revealed that this interaction is required for regulating microtubule dynamic instability in the cell periphery. Based on our findings, we propose that Msps is a novel Rac effector that acts, in partnership with Orbit, to regionally regulate microtubule dynamics.

  19. Taxol differentially modulates the dynamics of microtubules assembled from unfractionated and purified beta-tubulin isotypes.

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    Derry, W B; Wilson, L; Khan, I A; Luduena, R F; Jordan, M A

    1997-03-25

    Substoichiometric binding of taxol to tubulin in microtubules potently suppresses microtubule dynamics, which appears to be the most sensitive antiproliferative mechanism of taxol. To determine whether the beta-tubulin isotype composition of a microtubule can modulate sensitivity to taxol, we measured the effects of substoichiometric ratios of taxol bound to tubulin in microtubules on the dynamics of microtubules composed of purified alphabeta(II)-, alphabeta(III)-, or alphabeta(IV)-tubulin isotypes and compared the results with the effects of taxol on microtubules assembled from unfractionated tubulin. Substoichiometric ratios of bound taxol in microtubules assembled from purified beta-tubulin isotypes or unfractionated tubulin potently suppressed the shortening rates and the lengths shortened per shortening event. Correlation of the suppression of the shortening rate with the stoichiometry of bound taxol revealed that microtubules composed of purified alphabeta(II)-, alphabeta(III)-, and alphabeta(IV)-tubulin were, respectively, 1.6-, 7.4-, and 7.2-fold less sensitive to the effects of bound taxol than microtubules assembled from unfractionated tubulin. These results indicate that taxol differentially modulates microtubule dynamics depending upon the beta-tubulin isotype composition. The results are consistent with recent studies correlating taxol resistance in tumor cells with increased levels of beta(III0- and beta(IV)-tubulin expression and suggest that altered cellular expression of beta-tubulin isotypes can be an important mechanism by which tumor cells develop resistance to taxol.

  20. Enhanced dynamic instability of microtubules in a ROS free inert environment.

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    Islam, Md Sirajul; Kabir, Arif Md Rashedul; Inoue, Daisuke; Sada, Kazuki; Kakugo, Akira

    2016-04-01

    Reactive oxygen species (ROS), one of the regulators in various biological processes, have recently been suspected to modulate microtubule (MT) dynamics in cells. However due to complicated cellular environment and unavailability of any in vitro investigation, no detail is understood yet. Here, by performing simple in vitro investigations, we have unveiled the effect of ROS on MT dynamics. By studying dynamic instability of MTs in a ROS free environment and comparing with that in the presence of ROS, we disclosed that MTs showed enhanced dynamics in the ROS free environment. All the parameters that define dynamic instability of MTs e.g., growth and shrinkage rates, rescue and catastrophe frequencies were significantly affected by the presence of ROS. This work clearly reveals the role of ROS in modulating MT dynamics in vitro, and would be a great help in understanding the role of ROS in regulation of MT dynamics in cells.

  1. Mechanism of dynamic reorientation of cortical microtubules due to mechanical stress

    CERN Document Server

    Muratov, Alexander

    2015-01-01

    Directional growth caused by gravitropism and corresponding bending of plant cells has been explored since 19th century, however, many aspects of mechanisms underlying the perception of gravity at the molecular level are still not well known. Perception of gravity in root and shoot gravitropisms is usually attributed to gravisensitive cells, called statocytes, which exploit sedimentation of macroscopic and heavy organelles, amyloplasts, to sense the direction of gravity. Gravity stimulus is then transduced into distal elongation zone, which is several mm far from statocytes, where it causes stretching. It is suggested that gravity stimulus is conveyed by gradients in auxin flux. We propose a theoretical model that may explain how concentration gradients and/or stretching may indirectly affect the global orientation of cortical microtubules, attached to the cell membrane and induce their dynamic reorientation perpendicular to the gradients. In turn, oriented microtubules arrays direct the growth and orientatio...

  2. Stathmin regulates microtubule dynamics and microtubule organizing center polarization in activated T cells.

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    Filbert, Erin L; Le Borgne, Marie; Lin, Joseph; Heuser, John E; Shaw, Andrey S

    2012-06-01

    Polarization of T cells involves reorientation of the microtubule organizing center (MTOC). Because activated ERK is localized at the immunological synapse, we investigated its role by showing that ERK activation is important for MTOC polarization. Suspecting that ERK phosphorylates a regulator of microtubules, we next focused on stathmin, a known ERK substrate. Our work indicates that during T cell activation, ERK is recruited to the synapse, allowing it to phosphorylate stathmin molecules near the immunological synapse. Supporting an important role of stathmin phosphorylation in T cell activation, we showed that T cell activation results in increased microtubule growth rate dependent on the presence of stathmin. The significance of this finding was demonstrated by results showing that CTLs from stathmin(-/-) mice displayed defective MTOC polarization and defective target cell cytolysis. These data implicate stathmin as a regulator of the microtubule network during T cell activation.

  3. Contributions of microtubule dynamic instability and rotational diffusion to kinetochore capture

    CERN Document Server

    Blackwell, Robert; Edelmaier, Christopher; Gergely, Zachary R; Flynn, Patrick J; Montes, Salvador; Crapo, Ammon; Doostan, Alireza; McIntosh, J Richard; Glaser, Matthew A; Betterton, Meredith D

    2016-01-01

    Microtubule dynamic instability allows search and capture of kinetochores during spindle formation, an important process for accurate chromosome segregation during cell division. Recent work has found that microtubule rotational diffusion about minus-end attachment points contributes to kinetochore capture in fission yeast, but the relative contributions of dynamic instability and rotational diffusion are not well understood. We have developed a biophysical model of kinetochore capture in small fission-yeast nuclei using hybrid Brownian dynamics/kinetic Monte Carlo simulation techniques. With this model, we have studied the importance of dynamic instability and microtubule rotational diffusion for kinetochore capture, both to the lateral surface of a microtubule and at or near its end. Over a range of biologically relevant parameters, microtubule rotational diffusion decreased capture time, but made a relatively small contribution compared to dynamic instability. At most, rotational diffusion reduced capture ...

  4. Thymoquinone disrupts the microtubule dynamics in fission yeast Schizosaccharomyces pombe

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    Nusrat Masood

    2016-11-01

    Full Text Available Mad2 deletion strain of Schizosaccharomyces pombe was found to be sensitive to thymoquinone, a signature molecule present in Nigella sativa in a dose-dependent manner. Mad2 protein is an indispensable part of mitotic spindle checkpoint complex and is required for the cell cycle arrest in response to the spindle defects. Although the expression of α tubulin was not affected in thymoquinone treated cells, but the expression of β-tubulin was reduced. Further, the absence of microtubule in thymoquinone treated cells suggests its involvement in tubulin polymerization. Molecular docking studies revealed that thymoquinone specifically binds to β-tubulin near the Taxotere binding site of Tub1 (Tubulin α-β dimer. These studies additionally showed that thymoquinone interacts with the residues present in chain B, which is an inherent part of Mad2 protein of mitotic checkpoint complex (MCC. We concluded that the thymoquinone disrupts the microtubule polymerization that leads to the requirement of spindle checkpoint protein for the cell survival.

  5. Cep70 promotes microtubule assembly in vitro by increasing microtubule elongation

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    Xingjuan Shi; Jun Wang; Yunfan Yang; Yuan Ren; Jun Zhou; Dengwen Li

    2012-01-01

    Microtubules are dynamic cytoskeletal polymers present in all eukaryotic cells,In animal cells,they are organized by the centrosome,the major microtubule-organizing center.Many centrosomal proteins act coordinately to modulate microtubule assembly and organization.Our previous work has shown that Cep70,a novel centrosomal protein regulates microtubule assembly and organization in mammalian cells.However,the molecular details remain to be investigated,in this study,we investigated the molecular mechanism of how Cep70 regulates microtubule assembly using purified proteins.Our data showed that Cep70 increased the microtubule length without affecting the microtubule number in the purified system.These results demonstrate that Cep70 could directly regulate microtubule assembly by promoting microtubule elongation instead of microtubule nucleation.

  6. A plus-end raft to control microtubule dynamics and function.

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    Galjart, Niels; Perez, Franck

    2003-02-01

    Cells require a properly oriented and organised microtubule array to transmit positional information. Recent data have revealed a heterogeneous population of microtubule-binding proteins that accumulates mainly at distal ends of polymerising microtubules. Two mechanisms may account for this concentration: transient immobilisation, which involves association of proteins with growing ends, followed by release more proximally; and deposition at ends via a molecular motor. As with lipid rafts, protein concentration at distal ends may allow a cascade of interactions in the restricted area of a microtubule plus end. This may, in turn, control the dynamic behaviour of this cytoskeletal network and its anchoring to other structures.

  7. Paired arrangement of kinetochores together with microtubule pivoting and dynamics drive kinetochore capture in meiosis I.

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    Cojoc, Gheorghe; Florescu, Ana-Maria; Krull, Alexander; Klemm, Anna H; Pavin, Nenad; Jülicher, Frank; Tolić, Iva M

    2016-01-01

    Kinetochores are protein complexes on the chromosomes, whose function as linkers between spindle microtubules and chromosomes is crucial for proper cell division. The mechanisms that facilitate kinetochore capture by microtubules are still unclear. In the present study, we combine experiments and theory to explore the mechanisms of kinetochore capture at the onset of meiosis I in fission yeast. We show that kinetochores on homologous chromosomes move together, microtubules are dynamic and pivot around the spindle pole, and the average capture time is 3-4 minutes. Our theory describes paired kinetochores on homologous chromosomes as a single object, as well as angular movement of microtubules and their dynamics. For the experimentally measured parameters, the model reproduces the measured capture kinetics and shows that the paired configuration of kinetochores accelerates capture, whereas microtubule pivoting and dynamics have a smaller contribution. Kinetochore pairing may be a general feature that increases capture efficiency in meiotic cells.

  8. Halogenated auxins affect microtubules and root elongation in Lactuca sativa

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    Zhang, N.; Hasenstein, K. H.

    2000-01-01

    We studied the effect of 4,4,4-trifluoro-3-(indole-3-)butyric acid (TFIBA), a recently described root growth stimulator, and 5,6-dichloro-indole-3-acetic acid (DCIAA) on growth and microtubule (MT) organization in roots of Lactuca sativa L. DCIAA and indole-3-butyric acid (IBA) inhibited root elongation and depolymerized MTs in the cortex of the elongation zone, inhibited the elongation of stele cells, and promoted xylem maturation. Both auxins caused the plane of cell division to shift from anticlinal to periclinal. In contrast, TFIBA (100 micromolar) promoted elongation of primary roots by 40% and stimulated the elongation of lateral roots, even in the presence of IBA, the microtubular inhibitors oryzalin and taxol, or the auxin transport inhibitor naphthylphthalamic acid. However, TFIBA inhibited the formation of lateral root primordia. Immunostaining showed that TFIBA stabilized MTs orientation perpendicular to the root axis, doubled the cortical cell length, but delayed xylem maturation. The data indicate that the auxin-induced inhibition of elongation and swelling of roots results from reoriented phragmoplasts, the destabilization of MTs in elongating cells, and promotion of vessel formation. In contrast, TFIBA induced promotion of root elongation by enhancing cell length, prolonging transverse MT orientation, delaying cell and xylem maturation.

  9. Microtubule dynamics control HGF-induced lung endothelial barrier enhancement.

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    Xinyong Tian

    Full Text Available Microtubules (MT play a vital role in many cellular functions, but their role in peripheral actin cytoskeletal dynamics which is essential for control of endothelial barrier and monolayer integrity is less understood. We have previously described the enhancement of lung endothelial cell (EC barrier by hepatocyte growth factor (HGF which was associated with Rac1-mediated remodeling of actin cytoskeleton. This study investigated involvement of MT-dependent mechanisms in the HGF-induced enhancement of EC barrier. HGF-induced Rac1 activation was accompanied by phosphorylation of stathmin, a regulator of MT dynamics. HGF also stimulated MT peripheral growth monitored by time lapse imaging and tracking analysis of EB-1-decorated MT growing tips, and increased the pool of acetylated tubulin. These effects were abolished by EC pretreatment with HGF receptor inhibitor, downregulation of Rac1 pathway, or by expression of a stathmin-S63A phosphorylation deficient mutant. Expression of stathmin-S63A abolished the HGF protective effects against thrombin-induced activation of RhoA cascade, permeability increase, and EC barrier dysfunction. These results demonstrate a novel MT-dependent mechanism of HGF-induced EC barrier regulation via Rac1/PAK1/stathmin-dependent control of MT dynamics.

  10. Microtubules as key coordinators of nuclear envelope and endoplasmic reticulum dynamics during mitosis.

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    Schlaitz, Anne-Lore

    2014-07-01

    During mitosis, cells comprehensively restructure their interior to promote the faithful inheritance of DNA and cytoplasmic contents. In metazoans, this restructuring entails disassembly of the nuclear envelope, redistribution of its components into the endoplasmic reticulum (ER) and eventually nuclear envelope reassembly around the segregated chromosomes. The microtubule cytoskeleton has recently emerged as a critical regulator of mitotic nuclear envelope and ER dynamics. Microtubules and associated molecular motors tear open the nuclear envelope in prophase and remove nuclear envelope remnants from chromatin. Additionally, two distinct mechanisms of microtubule-based regulation of ER dynamics operate later in mitosis. First, association of the ER with microtubules is reduced, preventing invasion of ER into the spindle area, and second, organelle membrane is actively cleared from metaphase chromosomes. However, we are only beginning to understand the role of microtubules in shaping and distributing ER and other organelles during mitosis.

  11. Dynamic microtubules regulate dendritic spine morphology and synaptic plasticity

    NARCIS (Netherlands)

    Jaworski, J.; Kapitein, L.C.; Montenegro Gouveia, S.; Dortland, B.R.; Wulf, P.S.; Grigoriev, I.; Camera, P.; Spangler, S.A.; Di Stefano, P.; Demmers, J.; Krugers, H.; Defilippi, P.; Akhmanova, A.; Hoogenraad, C.C.

    2009-01-01

    Dendritic spines are the major sites of excitatory synaptic input, and their morphological changes have been linked to learning and memory processes. Here, we report that growing microtubule plus ends decorated by the microtubule tip-tracking protein EB3 enter spines and can modulate spine morpholog

  12. Microtubule assembly affects bone mass by regulating both osteoblast and osteoclast functions: stathmin deficiency produces an osteopenic phenotype in mice.

    Science.gov (United States)

    Liu, Hongbin; Zhang, Rongrong; Ko, Seon-Yle; Oyajobi, Babatunde O; Papasian, Christopher J; Deng, Hong-Wen; Zhang, Shujun; Zhao, Ming

    2011-09-01

    Cytoskeleton microtubules regulate various cell signaling pathways that are involved in bone cell function. We recently reported that inhibition of microtubule assembly by microtubule-targeting drugs stimulates osteoblast differentiation and bone formation. To further elucidate the role of microtubules in bone homeostasis, we characterized the skeletal phenotype of mice null for stathmin, an endogenous protein that inhibits microtubule assembly. In vivo micro-computed tomography (µCT) and histology revealed that stathmin deficiency results in a significant reduction of bone mass in adult mice concurrent with decreased osteoblast and increased osteoclast numbers in bone tissues. Phenotypic analyses of primary calvarial cells and bone marrow cells showed that stathmin deficiency inhibited osteoblast differentiation and induced osteoclast formation. In vitro overexpression studies showed that increased stathmin levels enhanced osteogenic differentiation of preosteoblast MC3T3-E1 cells and mouse bone marrow-derived cells and attenuated osteoclast formation from osteoclast precursor Raw264.7 cells and bone marrow cells. Results of immunofluorescent studies indicated that overexpression of stathmin disrupted radial microtubule filaments, whereas deficiency of stathmin stabilized the microtubule network structure in these bone cells. In addition, microtubule-targeting drugs that inhibit microtubule assembly and induce osteoblast differentiation lost these effects in the absence of stathmin. Collectively, these results suggest that stathmin, which alters microtubule dynamics, plays an essential role in maintenance of postnatal bone mass by regulating both osteoblast and osteoclast functions in bone. \\

  13. Microtubule-based nanomaterials: Exploiting nature's dynamic biopolymers.

    Science.gov (United States)

    Bachand, George D; Spoerke, Erik D; Stevens, Mark J

    2015-06-01

    For more than a decade now, biomolecular systems have served as an inspiration for the development of synthetic nanomaterials and systems that are capable of reproducing many of unique and emergent behaviors of living systems. One intriguing element of such systems may be found in a specialized class of proteins known as biomolecular motors that are capable of performing useful work across multiple length scales through the efficient conversion of chemical energy. Microtubule (MT) filaments may be considered within this context as their dynamic assembly and disassembly dissipate energy, and perform work within the cell. MTs are one of three cytoskeletal filaments in eukaryotic cells, and play critical roles in a range of cellular processes including mitosis and vesicular trafficking. Based on their function, physical attributes, and unique dynamics, MTs also serve as a powerful archetype of a supramolecular filament that underlies and drives multiscale emergent behaviors. In this review, we briefly summarize recent efforts to generate hybrid and composite nanomaterials using MTs as biomolecular scaffolds, as well as computational and synthetic approaches to develop synthetic one-dimensional nanostructures that display the enviable attributes of the natural filaments.

  14. Microtubule Dynamics and Oscillating State for Mitotic Spindle

    CERN Document Server

    Rashid-Shomali, Safura

    2010-01-01

    We present a physical mechanism that can cause the mitotic spindle to oscillate. The driving force for this mechanism emerges from the polymerization of astral microtubules interacting with the cell cortex. We show that Brownian ratchet model for growing microtubules reaching the cell cortex, mediate an effective mass to the spindle body and therefore force it to oscillate. We compare the predictions of this mechanism with the previous mechanisms which were based on the effects of motor proteins. Finally we combine the effects of microtubules polymerization and motor proteins, and present the detailed phase diagram for possible oscillating states.

  15. Dietary flavonoid fisetin binds to β-tubulin and disrupts microtubule dynamics in prostate cancer cells.

    Science.gov (United States)

    Mukhtar, Eiman; Adhami, Vaqar Mustafa; Sechi, Mario; Mukhtar, Hasan

    2015-10-28

    Microtubule targeting based therapies have revolutionized cancer treatment; however, resistance and side effects remain a major limitation. Therefore, novel strategies that can overcome these limitations are urgently needed. We made a novel discovery that fisetin, a hydroxyflavone, is a microtubule stabilizing agent. Fisetin binds to tubulin and stabilizes microtubules with binding characteristics far superior than paclitaxel. Surface plasmon resonance and computational docking studies suggested that fisetin binds to β-tubulin with superior affinity compared to paclitaxel. Fisetin treatment of human prostate cancer cells resulted in robust up-regulation of microtubule associated proteins (MAP)-2 and -4. In addition, fisetin treated cells were enriched in α-tubulin acetylation, an indication of stabilization of microtubules. Fisetin significantly inhibited PCa cell proliferation, migration, and invasion. Nudc, a protein associated with microtubule motor dynein/dynactin complex that regulates microtubule dynamics, was inhibited with fisetin treatment. Further, fisetin treatment of a P-glycoprotein overexpressing multidrug-resistant cancer cell line NCI/ADR-RES inhibited the viability and colony formation. Our results offer in vitro proof-of-concept for fisetin as a microtubule targeting agent. We suggest that fisetin could be developed as an adjuvant for treatment of prostate and other cancer types.

  16. Hoxb1b controls oriented cell division, cell shape and microtubule dynamics in neural tube morphogenesis.

    Science.gov (United States)

    Zigman, Mihaela; Laumann-Lipp, Nico; Titus, Tom; Postlethwait, John; Moens, Cecilia B

    2014-02-01

    Hox genes are classically ascribed to function in patterning the anterior-posterior axis of bilaterian animals; however, their role in directing molecular mechanisms underlying morphogenesis at the cellular level remains largely unstudied. We unveil a non-classical role for the zebrafish hoxb1b gene, which shares ancestral functions with mammalian Hoxa1, in controlling progenitor cell shape and oriented cell division during zebrafish anterior hindbrain neural tube morphogenesis. This is likely distinct from its role in cell fate acquisition and segment boundary formation. We show that, without affecting major components of apico-basal or planar cell polarity, Hoxb1b regulates mitotic spindle rotation during the oriented neural keel symmetric mitoses that are required for normal neural tube lumen formation in the zebrafish. This function correlates with a non-cell-autonomous requirement for Hoxb1b in regulating microtubule plus-end dynamics in progenitor cells in interphase. We propose that Hox genes can influence global tissue morphogenesis by control of microtubule dynamics in individual cells in vivo.

  17. The molecular dynamics of crawling migration in microtubule-disrupted keratocytes.

    Science.gov (United States)

    Nakashima, Hitomi; Okimura, Chika; Iwadate, Yoshiaki

    2015-01-01

    Cell-crawling migration plays an essential role in complex biological phenomena. It is now generally believed that many processes essential to such migration are regulated by microtubules in many cells, including fibroblasts and neurons. However, keratocytes treated with nocodazole, which is an inhibitor of microtubule polymerization - and even keratocyte fragments that contain no microtubules - migrate at the same velocity and with the same directionality as normal keratocytes. In this study, we discovered that not only these migration properties, but also the molecular dynamics that regulate such properties, such as the retrograde flow rate of actin filaments, distributions of vinculin and myosin II, and traction forces, are also the same in nocodazole-treated keratocytes as those in untreated keratocytes. These results suggest that microtubules are not in fact required for crawling migration of keratocytes, either in terms of migrating properties or of intracellular molecular dynamics.

  18. Dynamics of microtubule asters in microfabricated chambers: The role of catastrophes

    Science.gov (United States)

    Faivre-Moskalenko, Cendrine; Dogterom, Marileen

    2002-01-01

    Recent in vivo as well as in vitro experiments have indicated that microtubule pushing alone is sufficient to position a microtubule-organizing center within a cell. Here, we investigate the effect of catastrophes on the dynamics of microtubule asters within microfabricated chambers that mimic the confining geometry of living cells. The use of a glass bead as the microtubule-organizing center allows us to manipulate the aster by using optical tweezers. In the case in which microtubules preexist, we show that because of microtubule buckling, repositioning almost never occurs after relocation with the optical tweezers, although initial microtubule growth always leads the aster to the geometrical center of the chamber. When a catastrophe promoter is added, we find instead that the aster is able to efficiently explore the chamber geometry even after being relocated with the optical tweezers. As predicted by theoretical calculations, the results of our in vitro experiments clearly demonstrate the need for catastrophes for proper positioning in a confining geometry. These findings correlate with recent observations of nuclear positioning in fission yeast cells. PMID:12486218

  19. Aurora B suppresses microtubule dynamics and limits central spindle size by locally activating KIF4A

    Science.gov (United States)

    Nunes Bastos, Ricardo; Gandhi, Sapan R.; Baron, Ryan D.; Gruneberg, Ulrike; Nigg, Erich A.

    2013-01-01

    Anaphase central spindle formation is controlled by the microtubule-stabilizing factor PRC1 and the kinesin KIF4A. We show that an MKlp2-dependent pool of Aurora B at the central spindle, rather than global Aurora B activity, regulates KIF4A accumulation at the central spindle. KIF4A phosphorylation by Aurora B stimulates the maximal microtubule-dependent ATPase activity of KIF4A and promotes its interaction with PRC1. In the presence of phosphorylated KIF4A, microtubules grew more slowly and showed long pauses in growth, resulting in the generation of shorter PRC1-stabilized microtubule overlaps in vitro. Cells expressing only mutant forms of KIF4A lacking the Aurora B phosphorylation site overextended the anaphase central spindle, demonstrating that this regulation is crucial for microtubule length control in vivo. Aurora B therefore ensures that suppression of microtubule dynamic instability by KIF4A is restricted to a specific subset of microtubules and thereby contributes to central spindle size control in anaphase. PMID:23940115

  20. Push or Pull? -- Cryo-Electron Microscopy of Microtubule's Dynamic Instability and Its Roles in the Kinetochore

    Science.gov (United States)

    Wang, Hong-Wei

    2009-03-01

    Microtubule is a biopolymer made up of alpha-beta-tubulin heterodimers. The tubulin dimers assemble head-to-tail as protofilaments and about 13 protofilaments interact laterally to form a hollow cylindrical structure which is the microtubule. As the major cytoskeleton in all eukaryotic cells, microtubules have the intrinsic property to switch stochastically between growth and shrinkage phases, a phenomenon termed as their dynamic instability. Microtubule's dynamic instability is closely related to the types of nucleotide (GTP or GDP) that binds to the beta-tubulin. We have biochemically trapped two types of assembly states of tubulin with GTP or GDP bound representing the polymerizing and depolymerizing ends of microtubules respectively. Using cryo-electron microscopy, we have elucidated the structures of these intermediate assemblies, showing that tubulin protofilaments demonstrate various curvatures and form different types of lateral interactions depending on the nucleotide states of tubulin and the temperature. Our work indicates that during the microtubule's dynamic cycle, tubulin undergoes various assembly states. These states, different from the straight microtubule, lend the highly dynamic and complicated behavior of microtubules. Our study of microtubule's interaction with certain kinetochore complexes suggests that the intermediate assemblies are responsible for specific mechanical forces that are required during the mitosis or meiosis. Our discoveries strongly suggest that a microtubule is a molecular machine rather than a simple cellular scaffold.

  1. Autoinhibition of TBCB regulates EB1-mediated microtubule dynamics.

    Science.gov (United States)

    Carranza, Gerardo; Castaño, Raquel; Fanarraga, Mónica L; Villegas, Juan Carlos; Gonçalves, João; Soares, Helena; Avila, Jesus; Marenchino, Marco; Campos-Olivas, Ramón; Montoya, Guillermo; Zabala, Juan Carlos

    2013-01-01

    Tubulin cofactors (TBCs) participate in the folding, dimerization, and dissociation pathways of the tubulin dimer. Among them, TBCB and TBCE are two CAP-Gly domain-containing proteins that together efficiently interact with and dissociate the tubulin dimer. In the study reported here we showed that TBCB localizes at spindle and midzone microtubules during mitosis. Furthermore, the motif DEI/M-COO(-) present in TBCB, which is similar to the EEY/F-COO(-) element characteristic of EB proteins, CLIP-170, and α-tubulin, is required for TBCE-TBCB heterodimer formation and thus for tubulin dimer dissociation. This motif is responsible for TBCB autoinhibition, and our analysis suggests that TBCB is a monomer in solution. Mutants of TBCB lacking this motif are derepressed and induce microtubule depolymerization through an interaction with EB1 associated with microtubule tips. TBCB is also able to bind to the chaperonin complex CCT containing α-tubulin, suggesting that it could escort tubulin to facilitate its folding and dimerization, recycling or degradation.

  2. A polarised population of dynamic microtubules mediates homeostatic length control in animal cells.

    Directory of Open Access Journals (Sweden)

    Remigio Picone

    Full Text Available Because physical form and function are intimately linked, mechanisms that maintain cell shape and size within strict limits are likely to be important for a wide variety of biological processes. However, while intrinsic controls have been found to contribute to the relatively well-defined shape of bacteria and yeast cells, the extent to which individual cells from a multicellular animal control their plastic form remains unclear. Here, using micropatterned lines to limit cell extension to one dimension, we show that cells spread to a characteristic steady-state length that is independent of cell size, pattern width, and cortical actin. Instead, homeostatic length control on lines depends on a population of dynamic microtubules that lead during cell extension, and that are aligned along the long cell axis as the result of interactions of microtubule plus ends with the lateral cell cortex. Similarly, during the development of the zebrafish neural tube, elongated neuroepithelial cells maintain a relatively well-defined length that is independent of cell size but dependent upon oriented microtubules. A simple, quantitative model of cellular extension driven by microtubules recapitulates cell elongation on lines, the steady-state distribution of microtubules, and cell length homeostasis, and predicts the effects of microtubule inhibitors on cell length. Together this experimental and theoretical analysis suggests that microtubule dynamics impose unexpected limits on cell geometry that enable cells to regulate their length. Since cells are the building blocks and architects of tissue morphogenesis, such intrinsically defined limits may be important for development and homeostasis in multicellular organisms.

  3. Automated screening of microtubule growth dynamics identifies MARK2 as a regulator of leading edge microtubules downstream of Rac1 in migrating cells.

    Directory of Open Access Journals (Sweden)

    Yukako Nishimura

    Full Text Available Polarized microtubule (MT growth in the leading edge is critical to directed cell migration, and is mediated by Rac1 GTPase. To find downstream targets of Rac1 that affect MT assembly dynamics, we performed an RNAi screen of 23 MT binding and regulatory factors and identified RNAi treatments that suppressed changes in MT dynamics induced by constitutively activated Rac1. By analyzing fluorescent EB3 dynamics with automated tracking, we found that RNAi treatments targeting p150(glued, APC2, spastin, EB1, Op18, or MARK2 blocked Rac1-mediated MT growth in lamellipodia. MARK2 was the only protein whose RNAi targeting additionally suppressed Rac1 effects on MT orientation in lamellipodia, and thus became the focus of further study. We show that GFP-MARK2 rescued effects of MARK2 depletion on MT growth lifetime and orientation, and GFP-MARK2 localized in lamellipodia in a Rac1-activity-dependent manner. In a wound-edge motility assay, MARK2-depleted cells failed to polarize their centrosomes or exhibit oriented MT growth in the leading edge, and displayed defects in directional cell migration. Thus, automated image analysis of MT assembly dynamics identified MARK2 as a target regulated downstream of Rac1 that promotes oriented MT growth in the leading edge to mediate directed cell migration.

  4. Label-Free Imaging of Single Microtubule Dynamics Using Spatial Light Interference Microscopy.

    Science.gov (United States)

    Kandel, Mikhail E; Teng, Kai Wen; Selvin, Paul R; Popescu, Gabriel

    2017-01-24

    Due to their diameter, of only 24 nm, single microtubules are extremely challenging to image without the use of extrinsic contrast agents. As a result, fluorescence tagging is the common method to visualize their motility. However, such investigation is limited by photobleaching and phototoxicity. We experimentally demonstrate the capability of combining label-free spatial light interference microscopy (SLIM) with numerical processing for imaging single microtubules in a gliding assay. SLIM combines four different intensity images to obtain the optical path length map associated with the sample. Because of the use of broadband fields, the sensitivity to path length is better than 1 nm without (temporal) averaging and better than 0.1 nm upon averaging. Our results indicate that SLIM can image the dynamics of microtubules in a full field of view, of 200 × 200 μm(2), over many hours. Modeling the microtubule transport via the diffusion-advection equation, we found that the dispersion relation yields the standard deviation of the velocity distribution, without the need for tracking individual tubes. Interestingly, during a 2 h window, the microtubules begin to decelerate, at 100 pm/s(2) over a 20 min period. Thus, SLIM is likely to serve as a useful tool for understanding molecular motor activity, especially over large time scales, where fluorescence methods are of limited utility.

  5. Alterations in ovarian cancer cell adhesion drive taxol resistance by increasing microtubule dynamics in a FAK-dependent manner.

    Science.gov (United States)

    McGrail, Daniel J; Khambhati, Niti N; Qi, Mark X; Patel, Krishan S; Ravikumar, Nithin; Brandenburg, Chandler P; Dawson, Michelle R

    2015-04-17

    Chemorefractory ovarian cancer patients show extremely poor prognosis. Microtubule-stabilizing Taxol (paclitaxel) is a first-line treatment against ovarian cancer. Despite the close interplay between microtubules and cell adhesion, it remains unknown if chemoresistance alters the way cells adhere to their extracellular environment, a process critical for cancer metastasis. To investigate this, we isolated Taxol-resistant populations of OVCAR3 and SKOV3 ovarian cancer cell lines. Though Taxol-resistant cells neither effluxed more drug nor gained resistance to other chemotherapeutics, they did display increased microtubule dynamics. These changes in microtubule dynamics coincided with faster attachment rates and decreased adhesion strength, which correlated with increased surface β1-integrin expression and decreased focal adhesion formation, respectively. Adhesion strength correlated best with Taxol-sensitivity, and was found to be independent of microtubule polymerization but dependent on focal adhesion kinase (FAK), which was up-regulated in Taxol-resistant cells. FAK inhibition also decreased microtubule dynamics to equal levels in both populations, indicating alterations in adhesive signaling are up-stream of microtubule dynamics. Taken together, this work demonstrates that Taxol-resistance dramatically alters how ovarian cancer cells adhere to their extracellular environment causing down-stream increases in microtubule dynamics, providing a therapeutic target that may improve prognosis by not only recovering drug sensitivity, but also decreasing metastasis.

  6. Tank binding kinase 1 is a centrosome-associated kinase necessary for microtubule dynamics and mitosis.

    Science.gov (United States)

    Pillai, Smitha; Nguyen, Jonathan; Johnson, Joseph; Haura, Eric; Coppola, Domenico; Chellappan, Srikumar

    2015-12-10

    TANK Binding Kinase 1 (TBK1) is a non-canonical IκB kinase that contributes to KRAS-driven lung cancer. Here we report that TBK1 plays essential roles in mammalian cell division. Specifically, levels of active phospho-TBK1 increase during mitosis and localize to centrosomes, mitotic spindles and midbody, and selective inhibition or silencing of TBK1 triggers defects in spindle assembly and prevents mitotic progression. TBK1 binds to the centrosomal protein CEP170 and to the mitotic apparatus protein NuMA, and both CEP170 and NuMA are TBK1 substrates. Further, TBK1 is necessary for CEP170 centrosomal localization and binding to the microtubule depolymerase Kif2b, and for NuMA binding to dynein. Finally, selective disruption of the TBK1-CEP170 complex augments microtubule stability and triggers defects in mitosis, suggesting that TBK1 functions as a mitotic kinase necessary for microtubule dynamics and mitosis.

  7. KATNAL1 regulation of sertoli cell microtubule dynamics is essential for spermiogenesis and male fertility.

    Directory of Open Access Journals (Sweden)

    Lee B Smith

    Full Text Available Spermatogenesis is a complex process reliant upon interactions between germ cells (GC and supporting somatic cells. Testicular Sertoli cells (SC support GCs during maturation through physical attachment, the provision of nutrients, and protection from immunological attack. This role is facilitated by an active cytoskeleton of parallel microtubule arrays that permit transport of nutrients to GCs, as well as translocation of spermatids through the seminiferous epithelium during maturation. It is well established that chemical perturbation of SC microtubule remodelling leads to premature GC exfoliation demonstrating that microtubule remodelling is an essential component of male fertility, yet the genes responsible for this process remain unknown. Using a random ENU mutagenesis approach, we have identified a novel mouse line displaying male-specific infertility, due to a point mutation in the highly conserved ATPase domain of the novel KATANIN p60-related microtubule severing protein Katanin p60 subunit A-like1 (KATNAL1. We demonstrate that Katnal1 is expressed in testicular Sertoli cells (SC from 15.5 days post-coitum (dpc and that, consistent with chemical disruption models, loss of function of KATNAL1 leads to male-specific infertility through disruption of SC microtubule dynamics and premature exfoliation of spermatids from the seminiferous epithelium. The identification of KATNAL1 as an essential regulator of male fertility provides a significant novel entry point into advancing our understanding of how SC microtubule dynamics promotes male fertility. Such information will have resonance both for future treatment of male fertility and the development of non-hormonal male contraceptives.

  8. Synergistic suppression of microtubule dynamics by discodermolide and paclitaxel in non-small cell lung carcinoma cells.

    Science.gov (United States)

    Honore, Stéphane; Kamath, Kathy; Braguer, Diane; Horwitz, Susan Band; Wilson, Leslie; Briand, Claudette; Jordan, Mary Ann

    2004-07-15

    Discodermolide is a new microtubule-targeted antimitotic drug in Phase I clinical trials that, like paclitaxel, stabilizes microtubule dynamics and enhances microtubule polymer mass in vitro and in cells. Despite their apparently similar binding sites on microtubules, discodermolide acts synergistically with paclitaxel to inhibit proliferation of A549 human lung cancer cells (L. Martello et al., Clin. Cancer Res., 6: 1978-1987, 2000). To understand their synergy, we examined the effects of the two drugs singly and in combination in A549 cells and found that, surprisingly, their antiproliferative synergy is related to their ability to synergistically inhibit microtubule dynamic instability and mitosis. The combination of discodermolide and paclitaxel at their antiproliferative IC(50)s (7 nm for discodermolide and 2 nm for paclitaxel) altered all of the parameters of dynamic instability synergistically except the time-based rescue frequency. For example, together the drugs inhibited overall microtubule dynamicity by 71%, but each drug individually inhibited dynamicity by only 24%, giving a combination index (CI) of 0.23. Discodermolide and paclitaxel also synergistically blocked cell cycle progression at G(2)-M (41, 9.6, and 16% for both drugs together, for discodermolide alone, and for paclitaxel alone, respectively; CI = 0.59), and they synergistically enhanced apoptosis (CI = 0.85). Microtubules are unique receptors for drugs. The results suggest that ligands that bind to large numbers of binding sites on an individual microtubule can interact in a poorly understood manner to synergistically suppress microtubule dynamic instability and inhibit both mitosis and cell proliferation, with important consequences for combination clinical therapy with microtubule-targeted drugs.

  9. LARG and mDia1 link Galpha12/13 to cell polarity and microtubule dynamics.

    Science.gov (United States)

    Goulimari, Polyxeni; Knieling, Helga; Engel, Ulrike; Grosse, Robert

    2008-01-01

    Regulation of cell polarity is a process observed in all cells. During directed migration, cells orientate their microtubule cytoskeleton and the microtubule-organizing-center (MTOC), which involves integrins and downstream Cdc42 and glycogen synthase kinase-3beta activity. However, the contribution of G protein-coupled receptor signal transduction for MTOC polarity is less well understood. Here, we report that the heterotrimeric Galpha(12) and Galpha(13) proteins are necessary for MTOC polarity and microtubule dynamics based on studies using Galpha(12/13)-deficient mouse embryonic fibroblasts. Cell polarization involves the Galpha(12/13)-interacting leukemia-associated RhoGEF (LARG) and the actin-nucleating diaphanous formin mDia1. Interestingly, LARG associates with pericentrin and localizes to the MTOC and along microtubule tracks. We propose that Galpha(12/13) proteins exert essential functions linking extracellular signals to microtubule dynamics and cell polarity via RhoGEF and formin activity.

  10. Erucin, the major isothiocyanate in arugula (Eruca sativa, inhibits proliferation of MCF7 tumor cells by suppressing microtubule dynamics.

    Directory of Open Access Journals (Sweden)

    Olga Azarenko

    Full Text Available Consumption of cruciferous vegetables is associated with reduced risk of various types of cancer. Isothiocyanates including sulforaphane and erucin are believed to be responsible for this activity. Erucin [1-isothiocyanato-4-(methylthiobutane], which is metabolically and structurally related to sulforaphane, is present in large quantities in arugula (Eruca sativa, Mill., kohlrabi and Chinese cabbage. However, its cancer preventive mechanisms remain poorly understood. We found that erucin inhibits proliferation of MCF7 breast cancer cells (IC50 = 28 µM in parallel with cell cycle arrest at mitosis (IC50 = 13 µM and apoptosis, by a mechanism consistent with impairment of microtubule dynamics. Concentrations of 5-15 µM erucin suppressed the dynamic instability of microtubules during interphase in the cells. Most dynamic instability parameters were inhibited, including the rates and extents of growing and shortening, the switching frequencies between growing and shortening, and the overall dynamicity. Much higher erucin concentrations were required to reduce the microtubule polymer mass. In addition, erucin suppressed dynamic instability of microtubules reassembled from purified tubulin in similar fashion. The effects of erucin on microtubule dynamics, like those of sulforaphane, are similar qualitatively to those of much more powerful clinically-used microtubule-targeting anticancer drugs, including taxanes and the vinca alkaloids. The results suggest that suppression of microtubule dynamics by erucin and the resulting impairment of critically important microtubule-dependent cell functions such as mitosis, cell migration and microtubule-based transport may be important in its cancer preventive activities.

  11. Erucin, the major isothiocyanate in arugula (Eruca sativa), inhibits proliferation of MCF7 tumor cells by suppressing microtubule dynamics.

    Science.gov (United States)

    Azarenko, Olga; Jordan, Mary Ann; Wilson, Leslie

    2014-01-01

    Consumption of cruciferous vegetables is associated with reduced risk of various types of cancer. Isothiocyanates including sulforaphane and erucin are believed to be responsible for this activity. Erucin [1-isothiocyanato-4-(methylthio)butane], which is metabolically and structurally related to sulforaphane, is present in large quantities in arugula (Eruca sativa, Mill.), kohlrabi and Chinese cabbage. However, its cancer preventive mechanisms remain poorly understood. We found that erucin inhibits proliferation of MCF7 breast cancer cells (IC50 = 28 µM) in parallel with cell cycle arrest at mitosis (IC50 = 13 µM) and apoptosis, by a mechanism consistent with impairment of microtubule dynamics. Concentrations of 5-15 µM erucin suppressed the dynamic instability of microtubules during interphase in the cells. Most dynamic instability parameters were inhibited, including the rates and extents of growing and shortening, the switching frequencies between growing and shortening, and the overall dynamicity. Much higher erucin concentrations were required to reduce the microtubule polymer mass. In addition, erucin suppressed dynamic instability of microtubules reassembled from purified tubulin in similar fashion. The effects of erucin on microtubule dynamics, like those of sulforaphane, are similar qualitatively to those of much more powerful clinically-used microtubule-targeting anticancer drugs, including taxanes and the vinca alkaloids. The results suggest that suppression of microtubule dynamics by erucin and the resulting impairment of critically important microtubule-dependent cell functions such as mitosis, cell migration and microtubule-based transport may be important in its cancer preventive activities.

  12. AKAP9, a Regulator of Microtubule Dynamics, Contributes to Blood-Testis Barrier Function.

    Science.gov (United States)

    Venkatesh, Deepak; Mruk, Dolores; Herter, Jan M; Cullere, Xavier; Chojnacka, Katarzyna; Cheng, C Yan; Mayadas, Tanya N

    2016-02-01

    The blood-testis barrier (BTB), formed between adjacent Sertoli cells, undergoes extensive remodeling to facilitate the transport of preleptotene spermatocytes across the barrier from the basal to apical compartments of the seminiferous tubules for further development and maturation into spermatozoa. The actin cytoskeleton serves unique structural and supporting roles in this process, but little is known about the role of microtubules and their regulators during BTB restructuring. The large isoform of the cAMP-responsive scaffold protein AKAP9 regulates microtubule dynamics and nucleation at the Golgi. We found that conditional deletion of Akap9 in mice after the initial formation of the BTB at puberty leads to infertility. Akap9 deletion results in marked alterations in the organization of microtubules in Sertoli cells and a loss of barrier integrity despite a relatively intact, albeit more apically localized F-actin and BTB tight junctional proteins. These changes are accompanied by a loss of haploid spermatids due to impeded meiosis. The barrier, however, progressively reseals in older Akap9 null mice, which correlates with a reduction in germ cell apoptosis and a greater incidence of meiosis. However, spermiogenesis remains defective, suggesting additional roles for AKAP9 in this process. Together, our data suggest that AKAP9 and, by inference, the regulation of the microtubule network are critical for BTB function and subsequent germ cell development during spermatogenesis.

  13. PAR-1/MARK: a kinase essential for maintaining the dynamic state of microtubules.

    Science.gov (United States)

    Hayashi, Kenji; Suzuki, Atsushi; Ohno, Shigeo

    2012-01-01

    The serine/threonine kinase, PAR-1, is an essential component of the evolutionary-conserved polarity-regulating system, PAR-aPKC system, which plays indispensable roles in establishing asymmetric protein distributions and cell polarity in various biological contexts (Suzuki, A. and Ohno, S. (2006). J. Cell Sci., 119: 979-987; Matenia, D. and Mandelkow, E.M. (2009). Trends Biochem. Sci., 34: 332-342). PAR-1 is also known as MARK, which phosphorylates classical microtubule-associated proteins (MAPs) and detaches MAPs from microtubules (Matenia, D. and Mandelkow, E.M. (2009). Trends Biochem. Sci., 34: 332-342). This MARK activity of PAR-1 suggests its role in microtubule (MT) dynamics, but surprisingly, only few studies have been carried out to address this issue. Here, we summarize our recent study on live imaging analysis of MT dynamics in PAR-1b-depleted cells, which clearly demonstrated the positive role of PAR-1b in maintaining MT dynamics (Hayashi, K., Suzuki, A., Hirai, S., Kurihara, Y., Hoogenraad, C.C., and Ohno, S. (2011). J. Neurosci., 31: 12094-12103). Importantly, our results further revealed the novel physiological function of PAR-1b in maintaining dendritic spine morphology in mature neurons.

  14. Kinesin-12 motors cooperate to suppress microtubule catastrophes and drive the formation of parallel microtubule bundles.

    Science.gov (United States)

    Drechsler, Hauke; McAinsh, Andrew D

    2016-03-22

    Human Kinesin-12 (hKif15) plays a crucial role in assembly and maintenance of the mitotic spindle. These functions of hKif15 are partially redundant with Kinesin-5 (Eg5), which can cross-link and drive the extensile sliding of antiparallel microtubules. Although both motors are known to be tetramers, the functional properties of hKif15 are less well understood. Here we reveal how single or multiple Kif15 motors can cross-link, transport, and focus the plus-ends of intersecting microtubules. During transport, Kif15 motors step simultaneously along both microtubules with relative microtubule transport driven by a velocity differential between motor domain pairs. Remarkably, this differential is affected by the underlying intersection geometry: the differential is low on parallel and extreme on antiparallel microtubules where one motor domain pair becomes immobile. As a result, when intersecting microtubules are antiparallel, canonical transport of one microtubule along the other is allowed because one motor is firmly attached to one microtubule while it is stepping on the other. When intersecting microtubules are parallel, however, Kif15 motors can drive (biased) parallel sliding because the motor simultaneously steps on both microtubules that it cross-links. These microtubule rearrangements will focus microtubule plus-ends and finally lead to the formation of parallel bundles. At the same time, Kif15 motors cooperate to suppress catastrophe events at polymerizing microtubule plus-ends, raising the possibility that Kif15 motors may synchronize the dynamics of bundles that they have assembled. Thus, Kif15 is adapted to operate on parallel microtubule substrates, a property that clearly distinguishes it from the other tetrameric spindle motor, Eg5.

  15. Regulation of Kif15 localization and motility by the C-terminus of TPX2 and microtubule dynamics

    Science.gov (United States)

    Mann, Barbara J.; Balchand, Sai K.; Wadsworth, Patricia

    2017-01-01

    Mitotic motor proteins generate force to establish and maintain spindle bipolarity, but how they are temporally and spatially regulated in vivo is unclear. Prior work demonstrated that a microtubule-associated protein, TPX2, targets kinesin-5 and kinesin-12 motors to spindle microtubules. The C-terminal domain of TPX2 contributes to the localization and motility of the kinesin-5, Eg5, but it is not known whether this domain regulates kinesin-12, Kif15. We found that the C-terminal domain of TPX2 contributes to the localization of Kif15 to spindle microtubules in cells and suppresses motor walking in vitro. Kif15 and Eg5 are partially redundant motors, and overexpressed Kif15 can drive spindle formation in the absence of Eg5 activity. Kif15-dependent bipolar spindle formation in vivo requires the C-terminal domain of TPX2. In the spindle, fluorescent puncta of GFP-Kif15 move toward the equatorial region at a rate equivalent to microtubule growth. Reduction of microtubule growth with paclitaxel suppresses GFP-Kif15 motility, demonstrating that dynamic microtubules contribute to Kif15 behavior. Our results show that the C-terminal region of TPX2 regulates Kif15 in vitro, contributes to motor localization in cells, and is required for Kif15 force generation in vivo and further reveal that dynamic microtubules contribute to Kif15 behavior in vivo. PMID:27852894

  16. Mathematical modeling of the intracellular protein dynamics: the importance of active transport along microtubules.

    Science.gov (United States)

    Szymańska, Zuzanna; Parisot, Martin; Lachowicz, Mirosław

    2014-12-21

    In this paper we propose a mathematical model of protein and mRNA transport inside a cell. The spatio-temporal model takes into account the active transport along microtubules in the cytoplasm as well as diffusion and is able to reproduce the oscillatory changes in protein concentration observed in many experimental data. In the model the protein and the mRNA interact with each other that allows us to classify the model as a simple gene regulatory network. The proposed model is generic and may be adapted to specific signaling pathways. On the basis of numerical simulations, we formulate a new hypothesis that the oscillatory dynamics is allowed by the mRNA active transport along microtubules from the nucleus to distant locations.

  17. The Salmonella effector SseJ disrupts microtubule dynamics when ectopically expressed in normal rat kidney cells

    Science.gov (United States)

    Raines, Sally A.; Hodgkinson, Michael R.; Dowle, Adam A.

    2017-01-01

    Salmonella effector protein SseJ is secreted by Salmonella into the host cell cytoplasm where it can then modify host cell processes. Whilst host cell small GTPase RhoA has previously been shown to activate the acyl-transferase activity of SseJ we show here an un-described effect of SseJ protein production upon microtubule dynamism. SseJ prevents microtubule collapse and this is independent of SseJ’s acyl-transferase activity. We speculate that the effects of SseJ on microtubules would be mediated via its known interactions with the small GTPases of the Rho family. PMID:28235057

  18. ATX-2, the C. elegans Ortholog of Human Ataxin-2, Regulates Centrosome Size and Microtubule Dynamics.

    Directory of Open Access Journals (Sweden)

    Michael D Stubenvoll

    2016-09-01

    Full Text Available Centrosomes are critical sites for orchestrating microtubule dynamics, and exhibit dynamic changes in size during the cell cycle. As cells progress to mitosis, centrosomes recruit more microtubules (MT to form mitotic bipolar spindles that ensure proper chromosome segregation. We report a new role for ATX-2, a C. elegans ortholog of Human Ataxin-2, in regulating centrosome size and MT dynamics. ATX-2, an RNA-binding protein, forms a complex with SZY-20 in an RNA-independent fashion. Depleting ATX-2 results in embryonic lethality and cytokinesis failure, and restores centrosome duplication to zyg-1 mutants. In this pathway, SZY-20 promotes ATX-2 abundance, which inversely correlates with centrosome size. Centrosomes depleted of ATX-2 exhibit elevated levels of centrosome factors (ZYG-1, SPD-5, γ-Tubulin, increasing MT nucleating activity but impeding MT growth. We show that ATX-2 influences MT behavior through γ-Tubulin at the centrosome. Our data suggest that RNA-binding proteins play an active role in controlling MT dynamics and provide insight into the control of proper centrosome size and MT dynamics.

  19. Persistence Length of Stable Microtubules

    Science.gov (United States)

    Hawkins, Taviare; Mirigian, Matthew; Yasar, M. Selcuk; Ross, Jennifer

    2011-03-01

    Microtubules are a vital component of the cytoskeleton. As the most rigid of the cytoskeleton filaments, they give shape and support to the cell. They are also essential for intracellular traffic by providing the roadways onto which organelles are transported, and they are required to reorganize during cellular division. To perform its function in the cell, the microtubule must be rigid yet dynamic. We are interested in how the mechanical properties of stable microtubules change over time. Some ``stable'' microtubules of the cell are recycled after days, such as in the axons of neurons or the cilia and flagella. We measured the persistence length of freely fluctuating taxol-stabilized microtubules over the span of a week and analyzed them via Fourier decomposition. As measured on a daily basis, the persistence length is independent of the contour length. Although measured over the span of the week, the accuracy of the measurement and the persistence length varies. We also studied how fluorescently-labeling the microtubule affects the persistence length and observed that a higher labeling ratio corresponded to greater flexibility. National Science Foundation Grant No: 0928540 to JLR.

  20. Visualization and analysis of microtubule dynamics using dual color-coded display of plus-end labels.

    Directory of Open Access Journals (Sweden)

    Amy K Garrison

    Full Text Available Investigating spatial and temporal control of microtubule dynamics in live cells is critical to understanding cell morphogenesis in development and disease. Tracking fluorescently labeled plus-end-tracking proteins over time has become a widely used method to study microtubule assembly. Here, we report a complementary approach that uses only two images of these labels to visualize and analyze microtubule dynamics at any given time. Using a simple color-coding scheme, labeled plus-ends from two sequential images are pseudocolored with different colors and then merged to display color-coded ends. Based on object recognition algorithms, these colored ends can be identified and segregated into dynamic groups corresponding to four events, including growth, rescue, catastrophe, and pause. Further analysis yields not only their spatial distribution throughout the cell but also provides measurements such as growth rate and direction for each labeled end. We have validated the method by comparing our results with ground-truth data derived from manual analysis as well as with data obtained using the tracking method. In addition, we have confirmed color-coded representation of different dynamic events by analyzing their history and fate. Finally, we have demonstrated the use of the method to investigate microtubule assembly in cells and provided guidance in selecting optimal image acquisition conditions. Thus, this simple computer vision method offers a unique and quantitative approach to study spatial regulation of microtubule dynamics in cells.

  1. Suppression of microtubule dynamics by discodermolide by a novel mechanism is associated with mitotic arrest and inhibition of tumor cell proliferation.

    Science.gov (United States)

    Honore, Stéphane; Kamath, Kathy; Braguer, Diane; Wilson, Leslie; Briand, Claudette; Jordan, Mary Ann

    2003-12-01

    Discodermolide is a new microtubule-targeted drug in Phase I clinical trials that inhibits tumor growth and induces G(2)-M cell cycle arrest. It is effective against paclitaxel-resistant cell lines and acts synergistically in combination with paclitaxel. Suppression of microtubule dynamics by microtubule-targeted drugs has been hypothesized to be responsible for their ability to inhibit mitotic progression and cell proliferation. To determine whether discodermolide blocks mitosis by an effect on microtubule dynamics, we analyzed the effects of discodermolide on microtubule dynamics in living A549 human lung cancer cells during interphase at concentrations that block mitosis and inhibit cell proliferation. We found that discodermolide (7-166 nM) significantly suppressed microtubule dynamic instability. At the IC(50) for proliferation (7 nM discodermolide, 72 h), overall dynamicity was reduced by 23%. The principal parameters of dynamic instability suppressed by discodermolide were the microtubule shortening rate and length shortened. In addition, discodermolide markedly increased the frequency of rescued catastrophes. At the discodermolide concentration that resulted in 50% of maximal mitotic block (83 nM, 20 h), most microtubules were completely non-dynamic, no anaphases occurred, and all spindles were abnormal. The dynamicity of the remaining dynamic microtubules was reduced by 62%. The results indicate that a principal mechanism of inhibition of cell proliferation and mitotic block by discodermolide is suppression of microtubule dynamics. Importantly, the results indicate significant additional stabilizing effects of discodermolide on microtubule dynamics as compared with those of paclitaxel that may in turn reflect differences in their binding sites and their effects on tubulin conformation.

  2. External electric field effects on the mechanical properties of the αβ-tubulin dimer of microtubules: a molecular dynamics study.

    Science.gov (United States)

    Saeidi, H R; Lohrasebi, A; Mahnam, K

    2014-08-01

    The mechanical properties of the αβ-tubulin dimer of microtubules was modeled by using the molecular dynamics (MD) simulation method. The effect on the mechanical properties of the dimer of the existence and nonexistence of an applied electric field, either constant or periodic, was studied. Since there are charged or polar groups in the dimer structure, the electric field can interact with the dimer. The elastic constant and Young's modulus of the dimer were decreased when the dimer was exposed to a constant electric field of 0.03 V/nm. Furthermore, applying an oscillating electric field in the 1 GHz range to the dimer increased the elastic constant and Young's modulus of the dimer. These parameters were related to dimer rigidity and, consequently, in this frequency range, the application of electric fields may affect the function of microtubules.

  3. Dietary flavonoid fisetin binds to β-tubulin and disrupts microtubule dynamics in prostate cancer cells

    OpenAIRE

    Mukhtar, Eiman; Adhami, Vaqar Mustafa; Sechi, Mario; Mukhtar, Hasan

    2015-01-01

    Microtubule targeting based therapies have revolutionized cancer treatment; however, resistance and side effects remain a major limitation. Therefore, novel strategies that can overcome these limitations are urgently needed. We made a novel discovery that fisetin, a hydroxyflavone, is a microtubule stabilizing agent. Fisetin binds to tubulin and stabilizes microtubules with binding characteristics far superior than paclitaxel. Surface plasmon resonance and computational docking studies sugges...

  4. Microtubule-associated protein-4 controls nanovesicle dynamics and T cell activation.

    Science.gov (United States)

    Bustos-Morán, Eugenio; Blas-Rus, Noelia; Martin-Cófreces, Noa Beatriz; Sánchez-Madrid, Francisco

    2017-04-01

    The immune synapse (IS) is a specialized structure formed at the contact area between T lymphocytes and antigen-presenting cells (APCs) that is essential for the adaptive immune response. Proper T cell activation requires its polarization towards the APC, which is highly dependent on the tubulin cytoskeleton. Microtubule-associated protein-4 (MAP4) is a microtubule (MT)-stabilizing protein that controls MTs in physiological processes, such as cell division, migration, vesicular transport or primary cilia formation. In this study, we assessed the role of MAP4 in T cell activation. MAP4 decorates the pericentrosomal area and MTs of the T cell, and it is involved in MT detyrosination and stable assembly in response to T cell activation. In addition, MAP4 prompts the timely translocation of the MT-organizing center (MTOC) towards the IS and the dynamics of signaling nanovesicles that sustains T cell activation. However, MAP4 acts as a negative regulator of other T cell activation-related signals, including diacylglycerol (DAG) production and IL2 secretion. Our data indicate that MAP4 acts as a checkpoint molecule that balances positive and negative hallmarks of T cell activation.

  5. Human TUBB3 mutations perturb microtubule dynamics, kinesin interactions, and axon guidance

    Science.gov (United States)

    Tischfield, Max A.; Baris, Hagit N.; Wu, Chen; Rudolph, Guenther; Van Maldergem, Lionel; He, Wei; Chan, Wai-Man; Andrews, Caroline; Demer, Joseph L.; Robertson, Richard L.; Mackey, David A.; Ruddle, Jonathan B.; Bird, Thomas D.; Gottlob, Irene; Pieh, Christina; Traboulsi, Elias I.; Pomeroy, Scott L.; Hunter, David G.; Soul, Janet S.; Newlin, Anna; Sabol, Louise J.; Doherty, Edward J.; de Uzcátegui, Clara E.; de Uzcátegui, Nicolas; Collins, Mary Louise Z.; Sener, Emin C.; Wabbels, Bettina; Hellebrand, Heide; Meitinger, Thomas; de Berardinis, Teresa; Magli, Adriano; Schiavi, Costantino; Pastore-Trossello, Marco; Koc, Feray; Wong, Agnes M.; Levin, Alex V.; Geraghty, Michael T.; Descartes, Maria; Flaherty, Maree; Jamieson, Robyn V.; Møller, H. U.; Meuthen, Ingo; Callen, David F.; Kerwin, Janet; Lindsay, Susan; Meindl, Alfons; Gupta, Mohan L.; Pellman, David; Engle, Elizabeth C.

    2011-01-01

    We report that eight heterozygous missense mutations in TUBB3, encoding the neuron-specific β-tubulin isotype III, result in a spectrum of human nervous system disorders we now call the TUBB3 syndromes. Each mutation causes the ocular motility disorder CFEOM3, whereas some also result in intellectual and behavioral impairments, facial paralysis, and/or later-onset axonal sensorimotor polyneuropathy. Neuroimaging reveals a spectrum of abnormalities including hypoplasia of oculomotor nerves, and dysgenesis of the corpus callosum, anterior commissure, and corticospinal tracts. A knock-in disease mouse model reveals axon guidance defects without evidence of cortical cell migration abnormalities. We show the disease-associated mutations can impair tubulin heterodimer formation in vitro, although folded mutant heterodimers can still polymerize into microtubules. Modeling each mutation in yeast tubulin demonstrates that all alter dynamic instability whereas a subset disrupts the interaction of microtubules with kinesin motors. These findings demonstrate normal TUBB3 is required for axon guidance and maintenance in mammals. PMID:20074521

  6. Analysis of dynamic instability of steady-state microtubules in vitro by video-enhanced differential interference contrast microscopy with an appendix by Emin Oroudjev.

    Science.gov (United States)

    Yenjerla, Mythili; Lopus, Manu; Wilson, Leslie

    2010-01-01

    Microtubules are major constituents of the cytoskeleton which display dynamic properties. They exhibit dynamic instability which is defined as the stochastic switching between growing and shortening at microtubule ends. Dynamic instability plays an important role in diverse cellular functions including cell migration and mitosis. Many successful antimitotic drugs and microtubule-associated proteins (MAPs) are known to modulate microtubule dynamics, and it is important to analyze the in vitro dynamic instability of microtubules to study the mechanism of action of microtubule-targeted therapeutics and MAPs. In this chapter, we describe a method to analyze the in vitro dynamic instability of microtubules at steady state using video-enhanced differential contrast (VE-DIC) microscopy in detail. In this method, microtubules are assembled to steady state at 30 degrees C with MAP-free tubulin in a slide chamber in the presence of GTP, using sea urchin axonemes as nucleating seeds. Images of microtubules are enhanced and recorded in real time by a video camera and an image processor connected to a DIC microscope which is maintained at 30 degrees C. We use two software programs to track and analyze the growing and shortening of plus or minus ends of microtubules in the real-time images recorded using VE-DIC. In this chapter, we describe the instructions to use the tracking software Real Time Measurement II (RTM II) program. The instructions to use the analysis software Microtubule Life History Analysis Procedures (MT-LHAP) in Igor Pro software have been described in detail in an appendix (Oroudjev, 2010) following this chapter.

  7. Dynamic assembly of polymer nanotube networks via kinesin powered microtubule filaments

    Science.gov (United States)

    Paxton, Walter F.; Bouxsein, Nathan F.; Henderson, Ian M.; Gomez, Andrew; Bachand, George D.

    2015-06-01

    We describe for the first time how biological nanomotors may be used to actively self-assemble mesoscale networks composed of diblock copolymer nanotubes. The collective force generated by multiple kinesin nanomotors acting on a microtubule filament is large enough to overcome the energy barrier required to extract nanotubes from polymer vesicles comprised of poly(ethylene oxide-b-butadiene) in spite of the higher force requirements relative to extracting nanotubes from lipid vesicles. Nevertheless, large-scale polymer networks were dynamically assembled by the motors. These networks displayed enhanced robustness, persisting more than 24 h post-assembly (compared to 4-5 h for corresponding lipid networks). The transport of materials in and on the polymer membranes differs substantially from the transport on analogous lipid networks. Specifically, our data suggest that polymer mobility in nanotubular structures is considerably different from planar or 3D structures, and is stunted by 1D confinement of the polymer subunits. Moreover, quantum dots adsorbed onto polymer nanotubes are completely immobile, which is related to this 1D confinement effect and is in stark contrast to the highly fluid transport observed on lipid tubules.We describe for the first time how biological nanomotors may be used to actively self-assemble mesoscale networks composed of diblock copolymer nanotubes. The collective force generated by multiple kinesin nanomotors acting on a microtubule filament is large enough to overcome the energy barrier required to extract nanotubes from polymer vesicles comprised of poly(ethylene oxide-b-butadiene) in spite of the higher force requirements relative to extracting nanotubes from lipid vesicles. Nevertheless, large-scale polymer networks were dynamically assembled by the motors. These networks displayed enhanced robustness, persisting more than 24 h post-assembly (compared to 4-5 h for corresponding lipid networks). The transport of materials in and on

  8. An antitubulin agent BCFMT inhibits proliferation of cancer cells and induces cell death by inhibiting microtubule dynamics.

    Directory of Open Access Journals (Sweden)

    Ankit Rai

    Full Text Available Using cell based screening assay, we identified a novel anti-tubulin agent (Z-5-((5-(4-bromo-3-chlorophenylfuran-2-ylmethylene-2-thioxothiazolidin-4-one (BCFMT that inhibited proliferation of human cervical carcinoma (HeLa (IC(50, 7.2 ± 1.8 µM, human breast adenocarcinoma (MCF-7 (IC(50, 10.0 ± 0.5 µM, highly metastatic breast adenocarcinoma (MDA-MB-231 (IC(50, 6.0 ± 1 µM, cisplatin-resistant human ovarian carcinoma (A2780-cis (IC(50, 5.8 ± 0.3 µM and multi-drug resistant mouse mammary tumor (EMT6/AR1 (IC(50, 6.5 ± 1 µM cells. Using several complimentary strategies, BCFMT was found to inhibit cancer cell proliferation at G2/M phase of the cell cycle apparently by targeting microtubules. In addition, BCFMT strongly suppressed the dynamics of individual microtubules in live MCF-7 cells. At its half maximal proliferation inhibitory concentration (10 µM, BCFMT reduced the rates of growing and shortening phases of microtubules in MCF-7 cells by 37 and 40%, respectively. Further, it increased the time microtubules spent in the pause (neither growing nor shortening detectably state by 135% and reduced the dynamicity (dimer exchange per unit time of microtubules by 70%. In vitro, BCFMT bound to tubulin with a dissociation constant of 8.3 ± 1.8 µM, inhibited tubulin assembly and suppressed GTPase activity of microtubules. BCFMT competitively inhibited the binding of BODIPY FL-vinblastine to tubulin with an inhibitory concentration (K(i of 5.2 ± 1.5 µM suggesting that it binds to tubulin at the vinblastine site. In cultured cells, BCFMT-treatment depolymerized interphase microtubules, perturbed the spindle organization and accumulated checkpoint proteins (BubR1 and Mad2 at the kinetochores. BCFMT-treated MCF-7 cells showed enhanced nuclear accumulation of p53 and its downstream p21, which consequently activated apoptosis in these cells. The results suggested that BCFMT inhibits proliferation of several types of cancer cells including drug

  9. CLIP-170 facilitates the formation of kinetochore-microtubule attachments.

    Science.gov (United States)

    Tanenbaum, Marvin E; Galjart, Niels; van Vugt, Marcel A T M; Medema, René H

    2006-01-11

    CLIP-170 is a microtubule 'plus end tracking' protein involved in several microtubule-dependent processes in interphase. At the onset of mitosis, CLIP-170 localizes to kinetochores, but at metaphase, it is no longer detectable at kinetochores. Although RNA interference (RNAi) experiments have suggested an essential role for CLIP-170 during mitosis, the molecular function of CLIP-170 in mitosis has not yet been revealed. Here, we used a combination of high-resolution microscopy and RNAi-mediated depletion to study the function of CLIP-170 in mitosis. We found that CLIP-170 dynamically localizes to the outer most part of unattached kinetochores and to the ends of growing microtubules. In addition, we provide evidence that a pool of CLIP-170 is transported along kinetochore-microtubules by the dynein/dynactin complex. Interference with CLIP-170 expression results in defective chromosome congression and diminished kinetochore-microtubule attachments, but does not detectibly affect microtubule dynamics or kinetochore-microtubule stability. Taken together, our results indicate that CLIP-170 facilitates the formation of kinetochore-microtubule attachments, possibly through direct capture of microtubules at the kinetochore.

  10. Microtubules are Essential for Mitochondrial Dynamics-Fission, Fusion, and Motility- in Dictyostelium discoideum.

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    Laken C. Woods

    2016-03-01

    Full Text Available Mitochondrial function is dependent upon mitochondrial structure which is in turn dependent upon mitochondrial dynamics, including fission, fusion, and motility. Here we examined the relationship between mitochondrial dynamics and the cytoskeleton in Dictyostelium discoideum. Using time-lapse analysis, we quantified mitochondrial fission, fusion, and motility in the presence of cytoskeleton disrupting pharmaceuticals and the absence of the potential mitochondria-cytoskeleton linker protein, CluA. Our results indicate that microtubules are essential for mitochondrial movement, as well as fission and fusion; actin plays a less significant role, perhaps selecting the mitochondria for transport. We also suggest that CluA is not a linker protein but plays an unidentified role in mitochondrial fission and fusion. The significance of our work is to gain further insight into the role the cytoskeleton plays in mitochondrial dynamics and function. By better understanding these processes we can better appreciate the underlying mitochondrial contributions to many neurological disorders characterized by altered mitochondrial dynamics, structure, and/or function.

  11. Microtubule dynamics from mating through the first zygotic division in the budding yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Maddox, P; Chin, E; Mallavarapu, A; Yeh, E; Salmon, E D; Bloom, K

    1999-03-08

    We have used time-lapse digital imaging microscopy to examine cytoplasmic astral microtubules (Mts) and spindle dynamics during the mating pathway in budding yeast Saccharomyces cerevisiae. Mating begins when two cells of opposite mating type come into proximity. The cells arrest in the G1 phase of the cell cycle and grow a projection towards one another forming a shmoo projection. Imaging of microtubule dynamics with green fluorescent protein (GFP) fusions to dynein or tubulin revealed that the nucleus and spindle pole body (SPB) became oriented and tethered to the shmoo tip by a Mt-dependent search and capture mechanism. Dynamically unstable astral Mts were captured at the shmoo tip forming a bundle of three or four astral Mts. This bundle changed length as the tethered nucleus and SPB oscillated toward and away from the shmoo tip at growth and shortening velocities typical of free plus end astral Mts (approximately 0.5 micrometer/min). Fluorescent fiduciary marks in Mt bundles showed that Mt growth and shortening occurred primarily at the shmoo tip, not the SPB. This indicates that Mt plus end assembly/disassembly was coupled to pushing and pulling of the nucleus. Upon cell fusion, a fluorescent bar of Mts was formed between the two shmoo tip bundles, which slowly shortened (0.23 +/- 0.07 micrometer/min) as the two nuclei and their SPBs came together and fused (karyogamy). Bud emergence occurred adjacent to the fused SPB approximately 30 min after SPB fusion. During the first mitosis, the SPBs separated as the spindle elongated at a constant velocity (0.75 micrometer/min) into the zygotic bud. There was no indication of a temporal delay at the 2-micrometer stage of spindle morphogenesis or a lag in Mt nucleation by replicated SPBs as occurs in vegetative mitosis implying a lack of normal checkpoints. Thus, the shmoo tip appears to be a new model system for studying Mt plus end dynamic attachments and much like higher eukaryotes, the first mitosis after haploid

  12. FoxO regulates microtubule dynamics and polarity to promote dendrite branching in Drosophila sensory neurons.

    Science.gov (United States)

    Sears, James C; Broihier, Heather T

    2016-10-01

    The size and shape of dendrite arbors are defining features of neurons and critical determinants of neuronal function. The molecular mechanisms establishing arborization patterns during development are not well understood, though properly regulated microtubule (MT) dynamics and polarity are essential. We previously found that FoxO regulates axonal MTs, raising the question of whether it also regulates dendritic MTs and morphology. Here we demonstrate that FoxO promotes dendrite branching in all classes of Drosophila dendritic arborization (da) neurons. FoxO is required both for initiating growth of new branches and for maintaining existing branches. To elucidate FoxO function, we characterized MT organization in both foxO null and overexpressing neurons. We find that FoxO directs MT organization and dynamics in dendrites. Moreover, it is both necessary and sufficient for anterograde MT polymerization, which is known to promote dendrite branching. Lastly, FoxO promotes proper larval nociception, indicating a functional consequence of impaired da neuron morphology in foxO mutants. Together, our results indicate that FoxO regulates dendrite structure and function and suggest that FoxO-mediated pathways control MT dynamics and polarity.

  13. The Dynamic Pollen Tube Cytoskeleton: Live Cell Studies Using Actin-Binding and Microtubule-Binding Reporter Proteins

    Institute of Scientific and Technical Information of China (English)

    Alice Y. Cheung; Qiao-hong Duan; Silvia Santos Costa; Barend H.J.de Graaf; Veronica S.Di Stilio; Jose Feijo; Hen-Ming Wu

    2008-01-01

    Pollen tubes elongate within the pistil to transport sperm cells to the embryo sac for fertilization.Growth occurs exclusively at the tube apex,rendering pollen tube elongation a most dramatic polar cell growth process.A hall-mark pollen tube feature is its cytoskeleton,which comprises elaborately organized and dynamic actin microfilaments and microtubules.Pollen tube growth is dependent on the actin cytoskeleton;its organization and regulation have been exalined extensively by various approaches.including fluorescent protein labeled actin-binding proteins in live cell studies.Using the previously described GFP-NtADF1 and GFP-LIADF1, and a new actin reporter protein NtPLIM2b-GFP,we re-affirm that the predominant actin structures in elongating tobacco and lily pollen tubes are long,streaming actin cables along the pollen tube shank,and a subapical structure comprising shorter actin cables.The subapical collection of actin microfilaments undergoes dynamic changes,giving rise to the appearance of structures that range from basket-or funnel-shaped,mesh-like to a subtle ring.NtPLIM2b-GFP is used in combination with a guanine nucleotide exchange factor for the Rho GTPases,AtROP-GEF1,to illustrate the use of these actin reporter proteins to explore the linkage between the polar cell growth process and its actin cytoskeleton.Contrary to the actin cytoskeleton,microtubules appear not to play a direct role in supporting the polar cell growth process in angiosperm pollen tubes.Using a microtubule reporter protein based on the microtubule end-binding protein from Arabidopsis AtEB1,GFP-AtEB1,we show that the extensive microtubule network in elongating pollen tubes displays varying degrees of dynamics.These reporter proteins provide versatile tools to explore the functional connection between major structural and signaling components of the polar pollen tube growth process.

  14. The Gas2 family protein Pigs is a microtubule +TIP that affects cytoskeleton organisation.

    Science.gov (United States)

    Girdler, Gemma C; Applewhite, Derek A; Perry, Wick M G; Rogers, Stephen L; Röper, Katja

    2016-01-01

    Coordination between different cytoskeletal systems is crucial for many cell biological functions, including cell migration and mitosis, and also plays an important role during tissue morphogenesis. Proteins of the class of cytoskeletal crosslinkers, or cytolinkers, have the ability to interact with more than one cytoskeletal system at a time and are prime candidates to mediate any coordination. One such class comprises the Gas2-like proteins, combining a conserved calponin-homology-type actin-binding domain and a Gas2 domain predicted to bind microtubules (MTs). This domain combination is also found in spectraplakins, huge cytolinkers that play important roles in many tissues in both invertebrates and vertebrates. Here, we dissect the ability of the single Drosophila Gas2-like protein Pigs to interact with both actin and MT cytoskeletons, both in vitro and in vivo, and illustrate complex regulatory interactions that determine the localisation of Pigs to and its effects on the cytoskeleton.

  15. plusTipTracker: Quantitative image analysis software for the measurement of microtubule dynamics.

    Science.gov (United States)

    Applegate, Kathryn T; Besson, Sebastien; Matov, Alexandre; Bagonis, Maria H; Jaqaman, Khuloud; Danuser, Gaudenz

    2011-11-01

    Here we introduce plusTipTracker, a Matlab-based open source software package that combines automated tracking, data analysis, and visualization tools for movies of fluorescently-labeled microtubule (MT) plus end binding proteins (+TIPs). Although +TIPs mark only phases of MT growth, the plusTipTracker software allows inference of additional MT dynamics, including phases of pause and shrinkage, by linking collinear, sequential growth tracks. The algorithm underlying the reconstruction of full MT trajectories relies on the spatially and temporally global tracking framework described in Jaqaman et al. (2008). Post-processing of track populations yields a wealth of quantitative phenotypic information about MT network architecture that can be explored using several visualization modalities and bioinformatics tools included in plusTipTracker. Graphical user interfaces enable novice Matlab users to track thousands of MTs in minutes. In this paper, we describe the algorithms used by plusTipTracker and show how the package can be used to study regional differences in the relative proportion of MT subpopulations within a single cell. The strategy of grouping +TIP growth tracks for the analysis of MT dynamics has been introduced before (Matov et al., 2010). The numerical methods and analytical functionality incorporated in plusTipTracker substantially advance this previous work in terms of flexibility and robustness. To illustrate the enhanced performance of the new software we thus compare computer-assembled +TIP-marked trajectories to manually-traced MT trajectories from the same movie used in Matov et al. (2010).

  16. Dynamics of centrosome translocation and microtubule organization in neocortical neurons during distinct modes of polarization.

    Science.gov (United States)

    Sakakibara, Akira; Sato, Toshiyuki; Ando, Ryota; Noguchi, Namiko; Masaoka, Makoto; Miyata, Takaki

    2014-05-01

    Neuronal migration and process formation require cytoskeletal organization and remodeling. Recent studies suggest that centrosome translocation is involved in initial axon outgrowth, while the role of centrosomal positioning is not clear. Here, we examine relations between centrosomal positioning, axonogenesis, and microtubule (MT) polarization in multipolar and bipolar neocortical neurons. We monitored dynamic movements of centrosomes and MT plus ends in migratory neurons in embryonic mouse cerebral slices. In locomoting bipolar neurons, the centrosome oriented toward the pia-directed leading process. Bipolar neurons displayed dense MT plus end dynamics in leading processes, while trailing processes showed clear bidirectional MTs. In migrating multipolar neurons, new processes emerged irrespective of centrosome localization, followed by centrosome reorientations toward the dominant process. Anterograde movements of MT plus ends occurred in growing processes and retrograde movements were observed after retraction of the distal tip. In multipolar neurons, axon formed by tangential extension of a dominant process and the centrosome oriented toward the growing axon, while in locomoting neurons, an axon formed opposite to the direction of migration and the centrosome localized to the base of the leading process. Our data suggest that MT organization may alter centrosomal localization and that centrosomal positioning does not necessarily direct process formation.

  17. Dynamic Synchronization of Teacher-Students Affection in Affective Instruction

    Science.gov (United States)

    Zhang, Wenhai; Lu, Jiamei

    2011-01-01

    Based on Bower's affective network theory, the article links the dynamic analysis of affective factors in affective instruction, and presents affective instruction strategic of dynamic synchronization between teacher and students to implement the best ideal mood that promotes students' cognition and affection together. In the process of teaching,…

  18. Non-critical string theory formulation of microtubule dynamics and quantum aspects of brain function

    CERN Document Server

    Mavromatos, Nikolaos E

    1995-01-01

    Microtubule (MT) networks, subneural paracrystalline cytosceletal structures, seem to play a fundamental role in the neurons. We cast here the complicated MT dynamics in the form of a 1+1-dimensional non-critical string theory, thus enabling us to provide a consistent quantum treatment of MTs, including enviromental {\\em friction} effects. We suggest, thus, that the MTs are the microsites, in the brain, for the emergence of stable, macroscopic quantum coherent states, identifiable with the {\\em preconscious states}. Quantum space-time effects, as described by non-critical string theory, trigger then an {\\em organized collapse} of the coherent states down to a specific or {\\em conscious state}. The whole process we estimate to take {\\cal O}(1\\,{\\rm sec}), in excellent agreement with a plethora of experimental/observational findings. The {\\em microscopic arrow of time}, endemic in non-critical string theory, and apparent here in the self-collapse process, provides a satisfactory and simple resolution to the age...

  19. Microtubule Destabilizer KIF2A Undergoes Distinct Site-Specific Phosphorylation Cascades that Differentially Affect Neuronal Morphogenesis

    Directory of Open Access Journals (Sweden)

    Tadayuki Ogawa

    2015-09-01

    Full Text Available Neurons exhibit dynamic structural changes in response to extracellular stimuli. Microtubules (MTs provide rapid and dramatic cytoskeletal changes within the structural framework. However, the molecular mechanisms and signaling networks underlying MT dynamics remain unknown. Here, we have applied a comprehensive and quantitative phospho-analysis of the MT destabilizer KIF2A to elucidate the regulatory mechanisms of MT dynamics within neurons in response to extracellular signals. Interestingly, we identified two different sets of KIF2A phosphorylation profiles that accelerate (A-type and brake (B-type the MT depolymerization activity of KIF2A. Brain-derived neurotrophic factor (BDNF stimulates PAK1 and CDK5 kinases, which decrease the MT depolymerizing activity of KIF2A through B-type phosphorylation, resulting in enhanced outgrowth of neural processes. In contrast, lysophosphatidic acid (LPA induces ROCK2 kinase, which suppresses neurite outgrowth from round cells via A-type phosphorylation. We propose that these two mutually exclusive forms of KIF2A phosphorylation differentially regulate neuronal morphogenesis during development.

  20. Electric fields generated by synchronized oscillations of microtubules, centrosomes and chromosomes regulate the dynamics of mitosis and meiosis

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    Zhao Yue

    2012-07-01

    Full Text Available Abstract Super-macromolecular complexes play many important roles in eukaryotic cells. Classical structural biological studies focus on their complicated molecular structures, physical interactions and biochemical modifications. Recent advances concerning intracellular electric fields generated by cell organelles and super-macromolecular complexes shed new light on the mechanisms that govern the dynamics of mitosis and meiosis. In this review we synthesize this knowledge to provide an integrated theoretical model of these cellular events. We suggest that the electric fields generated by synchronized oscillation of microtubules, centrosomes, and chromatin fibers facilitate several events during mitosis and meiosis, including centrosome trafficking, chromosome congression in mitosis and synapsis between homologous chromosomes in meiosis. These intracellular electric fields are generated under energy excitation through the synchronized electric oscillations of the dipolar structures of microtubules, centrosomes and chromosomes, three of the super-macromolecular complexes within an animal cell.

  1. Electric fields generated by synchronized oscillations of microtubules, centrosomes and chromosomes regulate the dynamics of mitosis and meiosis.

    Science.gov (United States)

    Zhao, Yue; Zhan, Qimin

    2012-07-02

    Super-macromolecular complexes play many important roles in eukaryotic cells. Classical structural biological studies focus on their complicated molecular structures, physical interactions and biochemical modifications. Recent advances concerning intracellular electric fields generated by cell organelles and super-macromolecular complexes shed new light on the mechanisms that govern the dynamics of mitosis and meiosis. In this review we synthesize this knowledge to provide an integrated theoretical model of these cellular events. We suggest that the electric fields generated by synchronized oscillation of microtubules, centrosomes, and chromatin fibers facilitate several events during mitosis and meiosis, including centrosome trafficking, chromosome congression in mitosis and synapsis between homologous chromosomes in meiosis. These intracellular electric fields are generated under energy excitation through the synchronized electric oscillations of the dipolar structures of microtubules, centrosomes and chromosomes, three of the super-macromolecular complexes within an animal cell.

  2. Coordination of microtubule and microfilament dynamics by Drosophila Rho1, Spire and Cappuccino.

    Science.gov (United States)

    Rosales-Nieves, Alicia E; Johndrow, James E; Keller, Lani C; Magie, Craig R; Pinto-Santini, Delia M; Parkhurst, Susan M

    2006-04-01

    The actin-nucleation factors Spire and Cappuccino (Capu) regulate the onset of ooplasmic streaming in Drosophila melanogaster. Although this streaming event is microtubule-based, actin assembly is required for its timing. It is not understood how the interaction of microtubules and microfilaments is mediated in this context. Here, we demonstrate that Capu and Spire have microtubule and microfilament crosslinking activity. The spire locus encodes several distinct protein isoforms (SpireA, SpireC and SpireD). SpireD was recently shown to nucleate actin, but the activity of the other isoforms has not been addressed. We find that SpireD does not have crosslinking activity, whereas SpireC is a potent crosslinker. We show that SpireD binds to Capu and inhibits F-actin/microtubule crosslinking, and activated Rho1 abolishes this inhibition, establishing a mechanistic basis for the regulation of Capu and Spire activity. We propose that Rho1, cappuccino and spire are elements of a conserved developmental cassette that is capable of directly mediating crosstalk between microtubules and microfilaments.

  3. Escape from Mitotic Arrest: An Unexpected Connection Between Microtubule Dynamics and Epigenetic Regulation of Centromeric Chromatin in Schizosaccharomyces pombe.

    Science.gov (United States)

    George, Anuja A; Walworth, Nancy C

    2015-12-01

    Accurate chromosome segregation is necessary to ensure genomic integrity. Segregation depends on the proper functioning of the centromere, kinetochore, and mitotic spindle microtubules and is monitored by the spindle assembly checkpoint (SAC). In the fission yeast Schizosaccharomyces pombe, defects in Dis1, a microtubule-associated protein that influences microtubule dynamics, lead to mitotic arrest as a result of an active SAC and consequent failure to grow at low temperature. In a mutant dis1 background (dis1-288), loss of function of Msc1, a fission yeast homolog of the KDM5 family of proteins, suppresses the growth defect and promotes normal mitosis. Genetic analysis implicates a histone deacetylase (HDAC)-linked pathway in suppression because HDAC mutants clr6-1, clr3∆, and sir2∆, though not hos2∆, also promote normal mitosis in the dis1-288 mutant. Suppression of the dis phenotype through loss of msc1 function requires the spindle checkpoint protein Mad2 and is limited by the presence of the heterochromatin-associated HP1 protein homolog Swi6. We speculate that alterations in histone acetylation promote a centromeric chromatin environment that compensates for compromised dis1 function by allowing for successful kinetochore-microtubule interactions that can satisfy the SAC. In cells arrested in mitosis by mutation of dis1, loss of function of epigenetic determinants such as Msc1 or specific HDACs can promote cell survival. Because the KDM5 family of proteins has been implicated in human cancers, an appreciation of the potential role of this family of proteins in chromosome segregation is warranted.

  4. In vitro reconstitution of dynamic microtubules interacting with actin filament networks

    NARCIS (Netherlands)

    Preciado Lopez, M.; Huber, F.; Grigoriev, Ilya; Steinmetz, M.O.; Akhmanova, Anna; Dogterom, M.; Koenderink, G.H.

    2014-01-01

    Interactions between microtubules and actin filaments (F-actin) are essential for eukaryotic cell migration, polarization, growth, and division. Although the importance of these interactions has been long recognized, the inherent complexity of the cell interior hampers a detailed mechanistic study o

  5. Dynamic behavior of GFP-CLIP-170 reveals fast protein turnover on microtubule plus ends.

    NARCIS (Netherlands)

    K.A. Drägestein (Katharina Asja); W.A. van Cappellen (Gert); J.A.J. van Haren (Jeffrey); G.D. Tsibidis (George); A.S. Akhmanova (Anna); T.A. Knoch (Tobias); F.G. Grosveld (Frank); N.J. Galjart (Niels)

    2008-01-01

    textabstractMicrotubule (MT) plus end – tracking proteins (+TIPs) specifi cally recognize the ends of growing MTs. +TIPs are involved in diverse cellular processes such as cell division, cell migration, and cell polarity. Although +TIP tracking is important for these processes, the mechanisms underl

  6. Theory of dynamic force spectroscopy for kinetochore-microtubule attachments: rupture force distribution

    CERN Document Server

    Ghanti, Dipanwita

    2016-01-01

    Application of pulling force, under force-clamp conditions, to kinetochore-microtubule attachments {\\it in-vitro} revealed a catch-bond-like behavior. In an earlier paper ({\\it Sharma et al. Phys. Biol. (2014)} the physical origin of this apparently counter-intuitive phenomenon was traced to the nature of the force-dependence of the (de-)polymerization kinetics of the microtubules. In this brief communication that work is extended to situations where the external forced is ramped up till the attachment gets ruptured. In spite of the fundamental differences in the underlying mechanisms, the trend of variation of the rupture force distribution observed in our model kinetochore-microtubule attachment with the increasing loading rate is qualitatively similar to that displayed by the catch bonds formed in some other ligand-receptor systems. Our theoretical predictions can be tested experimentally by a straightforward modification of the protocol for controlling the force in the optical trap set up that was used in...

  7. Potent antiproliferative cembrenoids accumulate in tobacco upon infection with Rhodococcus fascians and trigger unusual microtubule dynamics in human glioblastoma cells.

    Directory of Open Access Journals (Sweden)

    Aminata P Nacoulma

    Full Text Available AIMS: Though plant metabolic changes are known to occur during interactions with bacteria, these were rarely challenged for pharmacologically active compounds suitable for further drug development. Here, the occurrence of specific chemicals with antiproliferative activity against human cancer cell lines was evidenced in hyperplasia (leafy galls induced when plants interact with particular phytopathogens, such as the Actinomycete Rhodococcus fascians. METHODS: We examined leafy galls fraction F3.1.1 on cell proliferation, cell division and cytoskeletal disorganization of human cancer cell lines using time-lapse videomicroscopy imaging, combined with flow cytometry and immunofluorescence analysis. We determined the F3.1.1-fraction composition by gas chromatography coupled to mass spectrometry. RESULTS: The leafy galls induced on tobacco by R. fascians yielded fraction F3.1.1 which inhibited proliferation of glioblastoma U373 cells with an IC50 of 4.5 µg/mL, F.3.1.1 was shown to increase cell division duration, cause nuclear morphological deformations and cell enlargement, and, at higher concentrations, karyokinesis defects leading to polyploidization and apoptosis. F3.1.1 consisted of a mixture of isomers belonging to the cembrenoids. The cellular defects induced by F3.1.1 were caused by a peculiar cytoskeletal disorganization, with the occurrence of fragmented tubulin and strongly organized microtubule aggregates within the same cell. Colchicine, paclitaxel, and cembrene also affected U373 cell proliferation and karyokinesis, but the induced microtubule rearrangement was very different from that provoked by F3.1.1. Altogether our data indicate that the cembrenoid isomers in F3.1.1 have a unique mode of action and are able to simultaneously modulate microtubule polymerization and stability.

  8. The acetylenic tricyclic bis(cyano enone), TBE-31, targets microtubule dynamics and cell polarity in migrating cells.

    Science.gov (United States)

    Chan, Eddie; Saito, Akira; Honda, Tadashi; Di Guglielmo, Gianni M

    2016-04-01

    Cell migration is dependent on the microtubule network for structural support as well as for the proper delivery and positioning of polarity proteins at the leading edge of migrating cells. Identification of drugs that target cytoskeletal-dependent cell migration and protein transport in polarized migrating cells is important in understanding the cell biology of normal and tumor cells and can lead to new therapeutic targets in disease processes. Here, we show that the tricyclic compound TBE-31 directly binds to tubulin and interferes with microtubule dynamics, as assessed by end binding 1 (EB1) live cell imaging. Interestingly, this interference is independent of in vitro tubulin polymerization. Using immunofluorescence microscopy, we also observed that TBE-31 interferes with the polarity of migratory cells. The polarity proteins Rac1, IQGAP and Tiam1 were localized at the leading edge of DMSO-treated migrating cell, but were observed to be in multiple protrusions around the cell periphery of TBE-31-treated cells. Finally, we observed that TBE-31 inhibits the migration of Rat2 fibroblasts with an IC50 of 0.75 μM. Taken together, our results suggest that the inhibition of cell migration by TBE-31 may result from the improper maintenance of cell polarity of migrating cells.

  9. Atomic-resolution structure of the CAP-Gly domain of dynactin on polymeric microtubules determined by magic angle spinning NMR spectroscopy.

    Science.gov (United States)

    Yan, Si; Guo, Changmiao; Hou, Guangjin; Zhang, Huilan; Lu, Xingyu; Williams, John Charles; Polenova, Tatyana

    2015-11-24

    Microtubules and their associated proteins perform a broad array of essential physiological functions, including mitosis, polarization and differentiation, cell migration, and vesicle and organelle transport. As such, they have been extensively studied at multiple levels of resolution (e.g., from structural biology to cell biology). Despite these efforts, there remain significant gaps in our knowledge concerning how microtubule-binding proteins bind to microtubules, how dynamics connect different conformational states, and how these interactions and dynamics affect cellular processes. Structures of microtubule-associated proteins assembled on polymeric microtubules are not known at atomic resolution. Here, we report a structure of the cytoskeleton-associated protein glycine-rich (CAP-Gly) domain of dynactin motor on polymeric microtubules, solved by magic angle spinning NMR spectroscopy. We present the intermolecular interface of CAP-Gly with microtubules, derived by recording direct dipolar contacts between CAP-Gly and tubulin using double rotational echo double resonance (dREDOR)-filtered experiments. Our results indicate that the structure adopted by CAP-Gly varies, particularly around its loop regions, permitting its interaction with multiple binding partners and with the microtubules. To our knowledge, this study reports the first atomic-resolution structure of a microtubule-associated protein on polymeric microtubules. Our approach lays the foundation for atomic-resolution structural analysis of other microtubule-associated motors.

  10. Cell cycle-dependent microtubule-based dynamic transport of cytoplasmic dynein in mammalian cells.

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    Takuya Kobayashi

    Full Text Available BACKGROUND: Cytoplasmic dynein complex is a large multi-subunit microtubule (MT-associated molecular motor involved in various cellular functions including organelle positioning, vesicle transport and cell division. However, regulatory mechanism of the cell-cycle dependent distribution of dynein has not fully been understood. METHODOLOGY/PRINCIPAL FINDINGS: Here we report live-cell imaging of cytoplasmic dynein in HeLa cells, by expressing multifunctional green fluorescent protein (mfGFP-tagged 74-kDa intermediate chain (IC74. IC74-mfGFP was successfully incorporated into functional dynein complex. In interphase, dynein moved bi-directionally along with MTs, which might carry cargos such as transport vesicles. A substantial fraction of dynein moved toward cell periphery together with EB1, a member of MT plus end-tracking proteins (+TIPs, suggesting +TIPs-mediated transport of dynein. In late-interphase and prophase, dynein was localized at the centrosomes and the radial MT array. In prometaphase and metaphase, dynein was localized at spindle MTs where it frequently moved from spindle poles toward chromosomes or cell cortex. +TIPs may be involved in the transport of spindle dyneins. Possible kinetochore and cortical dyneins were also observed. CONCLUSIONS AND SIGNIFICANCE: These findings suggest that cytoplasmic dynein is transported to the site of action in preparation for the following cellular events, primarily by the MT-based transport. The MT-based transport may have greater advantage than simple diffusion of soluble dynein in rapid and efficient transport of the limited concentration of the protein.

  11. A pericentrin-related protein homolog in Aspergillus nidulans plays important roles in nucleus positioning and cell polarity by affecting microtubule organization.

    Science.gov (United States)

    Chen, Peiying; Gao, Rongsui; Chen, Shaochun; Pu, Li; Li, Pin; Huang, Ying; Lu, Ling

    2012-12-01

    Pericentrin is a large coiled-coil protein in mammalian centrosomes that serves as a multifunctional scaffold for anchoring numerous proteins. Recent studies have linked numerous human disorders with mutated or elevated levels of pericentrin, suggesting unrecognized contributions of pericentrin-related proteins to the development of these disorders. In this study, we characterized AnPcpA, a putative homolog of pericentrin-related protein in the model filamentous fungus Aspergillus nidulans, and found that it is essential for conidial germination and hyphal development. Compared to the hyphal apex localization pattern of calmodulin (CaM), which has been identified as an interactive partner of the pericentrin homolog, GFP-AnPcpA fluorescence dots are associated mainly with nuclei, while the accumulation of CaM at the hyphal apex depends on the function of AnPcpA. In addition, the depletion of AnPcpA by an inducible alcA promoter repression results in severe growth defects and abnormal nuclear segregation. Most interestingly, in mature hyphal cells, knockdown of pericentrin was able to significantly induce changes in cell shape and cytoskeletal remodeling; it resulted in some enlarged compartments with condensed nuclei and anucleate small compartments as well. Moreover, defects in AnPcpA significantly disrupted the microtubule organization and nucleation, suggesting that AnPcpA may affect nucleus positioning by influencing microtubule organization.

  12. Dynamic Musical Communication of Core Affect

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    Nicole eFlaig

    2014-03-01

    Full Text Available Is there something special about the way music communicates feelings? Theorists since Meyer (1956 have attempted to explain how music could stimulate varied and subtle affective experiences by violating learned expectancies, or by mimicking other forms of social interaction. Our proposal is that music speaks to the brain in its own language; it need not imitate any other form of communication. We review recent theoretical and empirical literature, which suggests that all conscious processes consist of dynamic neural events, produced by spatially dispersed processes in the physical brain. Intentional thought and affective experience arise as dynamical aspects of neural events taking place in multiple brain areas simultaneously. At any given moment, this content comprises a unified scene that is integrated into a dynamic core through synchrony of neuronal oscillations. We propose that 1 neurodynamic synchrony with musical stimuli gives rise to musical qualia including tonal and temporal expectancies, and that 2 music-synchronous responses couple into core neurodynamics, enabling music to directly modulate core affect. Expressive music performance, for example, may recruit rhythm-synchronous neural responses to support affective communication. We suggest that the dynamic relationship between musical expression and the experience of affect presents a unique opportunity for the study of emotional experience. This may help elucidate the neural mechanisms underlying arousal and valence, and offer a new approach to exploring the complex dynamics of the how and why of emotional experience.

  13. Dynamic musical communication of core affect.

    Science.gov (United States)

    Flaig, Nicole K; Large, Edward W

    2014-01-01

    Is there something special about the way music communicates feelings? Theorists since Meyer (1956) have attempted to explain how music could stimulate varied and subtle affective experiences by violating learned expectancies, or by mimicking other forms of social interaction. Our proposal is that music speaks to the brain in its own language; it need not imitate any other form of communication. We review recent theoretical and empirical literature, which suggests that all conscious processes consist of dynamic neural events, produced by spatially dispersed processes in the physical brain. Intentional thought and affective experience arise as dynamical aspects of neural events taking place in multiple brain areas simultaneously. At any given moment, this content comprises a unified "scene" that is integrated into a dynamic core through synchrony of neuronal oscillations. We propose that (1) neurodynamic synchrony with musical stimuli gives rise to musical qualia including tonal and temporal expectancies, and that (2) music-synchronous responses couple into core neurodynamics, enabling music to directly modulate core affect. Expressive music performance, for example, may recruit rhythm-synchronous neural responses to support affective communication. We suggest that the dynamic relationship between musical expression and the experience of affect presents a unique opportunity for the study of emotional experience. This may help elucidate the neural mechanisms underlying arousal and valence, and offer a new approach to exploring the complex dynamics of the how and why of emotional experience.

  14. Models for microtubule cargo transport coupling the Langevin equation to stochastic stepping motor dynamics: Caring about fluctuations.

    Science.gov (United States)

    Bouzat, Sebastián

    2016-01-01

    One-dimensional models coupling a Langevin equation for the cargo position to stochastic stepping dynamics for the motors constitute a relevant framework for analyzing multiple-motor microtubule transport. In this work we explore the consistence of these models focusing on the effects of the thermal noise. We study how to define consistent stepping and detachment rates for the motors as functions of the local forces acting on them in such a way that the cargo velocity and run-time match previously specified functions of the external load, which are set on the base of experimental results. We show that due to the influence of the thermal fluctuations this is not a trivial problem, even for the single-motor case. As a solution, we propose a motor stepping dynamics which considers memory on the motor force. This model leads to better results for single-motor transport than the approaches previously considered in the literature. Moreover, it gives a much better prediction for the stall force of the two-motor case, highly compatible with the experimental findings. We also analyze the fast fluctuations of the cargo position and the influence of the viscosity, comparing the proposed model to the standard one, and we show how the differences on the single-motor dynamics propagate to the multiple motor situations. Finally, we find that the one-dimensional character of the models impede an appropriate description of the fast fluctuations of the cargo position at small loads. We show how this problem can be solved by considering two-dimensional models.

  15. MVL-PLA2, a snake venom phospholipase A2, inhibits angiogenesis through an increase in microtubule dynamics and disorganization of focal adhesions.

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    Amine Bazaa

    Full Text Available Integrins are essential protagonists of the complex multi-step process of angiogenesis that has now become a major target for the development of anticancer therapies. We recently reported and characterized that MVL-PLA2, a novel phospholipase A2 from Macrovipera lebetina venom, exhibited anti-integrin activity. In this study, we show that MVL-PLA2 also displays potent anti-angiogenic properties. This phospholipase A2 inhibited adhesion and migration of human microvascular-endothelial cells (HMEC-1 in a dose-dependent manner without being cytotoxic. Using Matrigel and chick chorioallantoic membrane assays, we demonstrated that MVL-PLA2, as well as its catalytically inactivated form, significantly inhibited angiogenesis both in vitro and in vivo. We have also found that the actin cytoskeleton and the distribution of alphav beta3 integrin, a critical regulator of angiogenesis and a major component of focal adhesions, were disturbed after MVL-PLA2 treatment. In order to further investigate the mechanism of action of this protein on endothelial cells, we analyzed the dynamic instability behavior of microtubules in living endothelial cells. Interestingly, we showed that MVL-PLA2 significantly increased microtubule dynamicity in HMEC-1 cells by 40%. We propose that the enhancement of microtubule dynamics may explain the alterations in the formation of focal adhesions, leading to inhibition of cell adhesion and migration.

  16. Mammalian diaphanous-related formin 1 regulates GSK3β-dependent microtubule dynamics required for T cell migratory polarization.

    Directory of Open Access Journals (Sweden)

    Baoxia Dong

    Full Text Available The mammalian diaphanous-related formin (mDia1, a Rho-regulated cytoskeletal modulator, has been shown to promote T lymphocyte chemotaxis and interaction with antigen presenting cells, but the mechanisms underpinning mDia1 roles in these processes have not been defined. Here we show that mDia1(-/- T cells exhibit impaired lymphocyte function-associated antigen 1 (LFA-1-mediated T cell adhesion, migration and in vivo trafficking. These defects are associated with impaired microtubule (MT polarization and stabilization, altered MT dynamics and reduced peripheral clustering of the MT plus-end-protein, adenomatous polyposis coli (APC in migrating T cells following LFA-1-engagement. Loss of mDia1 also leads to impaired inducible inactivation of the glycogen synthase kinase (GSK 3β as well as hyperphosphorylation and reduced levels of APC in migrating T cells. These findings identify essential roles for the mDia1 formin in modulating GSK3β-dependent MT contributions to induction of T-cell polarity, adhesion and motility.

  17. Mammalian diaphanous-related formin 1 regulates GSK3β-dependent microtubule dynamics required for T cell migratory polarization.

    Science.gov (United States)

    Dong, Baoxia; Zhang, Steven S; Gao, Wen; Su, Haichun; Chen, Jun; Jin, Fuzi; Bhargava, Ajay; Chen, Xiequn; Jorgensen, Lars; Alberts, Arthur S; Zhang, Jinyi; Siminovitch, Katherine A

    2013-01-01

    The mammalian diaphanous-related formin (mDia1), a Rho-regulated cytoskeletal modulator, has been shown to promote T lymphocyte chemotaxis and interaction with antigen presenting cells, but the mechanisms underpinning mDia1 roles in these processes have not been defined. Here we show that mDia1(-/-) T cells exhibit impaired lymphocyte function-associated antigen 1 (LFA-1)-mediated T cell adhesion, migration and in vivo trafficking. These defects are associated with impaired microtubule (MT) polarization and stabilization, altered MT dynamics and reduced peripheral clustering of the MT plus-end-protein, adenomatous polyposis coli (APC) in migrating T cells following LFA-1-engagement. Loss of mDia1 also leads to impaired inducible inactivation of the glycogen synthase kinase (GSK) 3β as well as hyperphosphorylation and reduced levels of APC in migrating T cells. These findings identify essential roles for the mDia1 formin in modulating GSK3β-dependent MT contributions to induction of T-cell polarity, adhesion and motility.

  18. Recruitment of EB1, a Master Regulator of Microtubule Dynamics, to the Surface of the Theileria annulata Schizont

    KAUST Repository

    Woods, Kerry L.

    2013-05-09

    The apicomplexan parasite Theileria annulata transforms infected host cells, inducing uncontrolled proliferation and clonal expansion of the parasitized cell population. Shortly after sporozoite entry into the target cell, the surrounding host cell membrane is dissolved and an array of host cell microtubules (MTs) surrounds the parasite, which develops into the transforming schizont. The latter does not egress to invade and transform other cells. Instead, it remains tethered to host cell MTs and, during mitosis and cytokinesis, engages the cell\\'s astral and central spindle MTs to secure its distribution between the two daughter cells. The molecular mechanism by which the schizont recruits and stabilizes host cell MTs is not known. MT minus ends are mostly anchored in the MT organizing center, while the plus ends explore the cellular space, switching constantly between phases of growth and shrinkage (called dynamic instability). Assuming the plus ends of growing MTs provide the first point of contact with the parasite, we focused on the complex protein machinery associated with these structures. We now report how the schizont recruits end-binding protein 1 (EB1), a central component of the MT plus end protein interaction network and key regulator of host cell MT dynamics. Using a range of in vitro experiments, we demonstrate that T. annulata p104, a polymorphic antigen expressed on the schizont surface, functions as a genuine EB1-binding protein and can recruit EB1 in the absence of any other parasite proteins. Binding strictly depends on a consensus SxIP motif located in a highly disordered C-terminal region of p104. We further show that parasite interaction with host cell EB1 is cell cycle regulated. This is the first description of a pathogen-encoded protein to interact with EB1 via a bona-fide SxIP motif. Our findings provide important new insight into the mode of interaction between Theileria and the host cell cytoskeleton. 2013 Woods et al.

  19. Recruitment of EB1, a master regulator of microtubule dynamics, to the surface of the Theileria annulata schizont.

    Directory of Open Access Journals (Sweden)

    Kerry L Woods

    2013-05-01

    Full Text Available The apicomplexan parasite Theileria annulata transforms infected host cells, inducing uncontrolled proliferation and clonal expansion of the parasitized cell population. Shortly after sporozoite entry into the target cell, the surrounding host cell membrane is dissolved and an array of host cell microtubules (MTs surrounds the parasite, which develops into the transforming schizont. The latter does not egress to invade and transform other cells. Instead, it remains tethered to host cell MTs and, during mitosis and cytokinesis, engages the cell's astral and central spindle MTs to secure its distribution between the two daughter cells. The molecular mechanism by which the schizont recruits and stabilizes host cell MTs is not known. MT minus ends are mostly anchored in the MT organizing center, while the plus ends explore the cellular space, switching constantly between phases of growth and shrinkage (called dynamic instability. Assuming the plus ends of growing MTs provide the first point of contact with the parasite, we focused on the complex protein machinery associated with these structures. We now report how the schizont recruits end-binding protein 1 (EB1, a central component of the MT plus end protein interaction network and key regulator of host cell MT dynamics. Using a range of in vitro experiments, we demonstrate that T. annulata p104, a polymorphic antigen expressed on the schizont surface, functions as a genuine EB1-binding protein and can recruit EB1 in the absence of any other parasite proteins. Binding strictly depends on a consensus SxIP motif located in a highly disordered C-terminal region of p104. We further show that parasite interaction with host cell EB1 is cell cycle regulated. This is the first description of a pathogen-encoded protein to interact with EB1 via a bona-fide SxIP motif. Our findings provide important new insight into the mode of interaction between Theileria and the host cell cytoskeleton.

  20. SAS-4 is recruited to a dynamic structure in newly forming centrioles that is stabilized by the gamma-tubulin-mediated addition of centriolar microtubules.

    Science.gov (United States)

    Dammermann, Alexander; Maddox, Paul S; Desai, Arshad; Oegema, Karen

    2008-02-25

    Centrioles are surrounded by pericentriolar material (PCM), which is proposed to promote new centriole assembly by concentrating gamma-tubulin. Here, we quantitatively monitor new centriole assembly in living Caenorhabditis elegans embryos, focusing on the conserved components SAS-4 and SAS-6. We show that SAS-4 and SAS-6 are coordinately recruited to the site of new centriole assembly and reach their maximum levels during S phase. Centriolar SAS-6 is subsequently reduced by a mechanism intrinsic to the early assembly pathway that does not require progression into mitosis. Centriolar SAS-4 remains in dynamic equilibrium with the cytoplasmic pool until late prophase, when it is stably incorporated in a step that requires gamma-tubulin and microtubule assembly. These results indicate that gamma-tubulin in the PCM stabilizes the nascent daughter centriole by promoting microtubule addition to its outer wall. Such a mechanism may help restrict new centriole assembly to the vicinity of preexisting parent centrioles that recruit PCM.

  1. Structural investigations into the binding mode of novel neolignans Cmp10 and Cmp19 microtubule stabilizers by in silico molecular docking, molecular dynamics, and binding free energy calculations.

    Science.gov (United States)

    Tripathi, Shubhandra; Kumar, Akhil; Kumar, B Sathish; Negi, Arvind S; Sharma, Ashok

    2016-06-01

    Microtubule stabilizers provide an important mode of treatment via mitotic cell arrest of cancer cells. Recently, we reported two novel neolignans derivatives Cmp10 and Cmp19 showing anticancer activity and working as microtubule stabilizers at micromolar concentrations. In this study, we have explored the binding site, mode of binding, and stabilization by two novel microtubule stabilizers Cmp10 and Cmp19 using in silico molecular docking, molecular dynamics (MD) simulation, and binding free energy calculations. Molecular docking studies were performed to explore the β-tubulin binding site of Cmp10 and Cmp19. Further, MD simulations were used to probe the β-tubulin stabilization mechanism by Cmp10 and Cmp19. Binding affinity was also compared for Cmp10 and Cmp19 using binding free energy calculations. Our docking results revealed that both the compounds bind at Ptxl binding site in β-tubulin. MD simulation studies showed that Cmp10 and Cmp19 binding stabilizes M-loop (Phe272-Val288) residues of β-tubulin and prevent its dynamics, leading to a better packing between α and β subunits from adjacent tubulin dimers. In addition, His229, Ser280 and Gln281, and Arg278, Thr276, and Ser232 were found to be the key amino acid residues forming H-bonds with Cmp10 and Cmp19, respectively. Consequently, binding free energy calculations indicated that Cmp10 (-113.655 kJ/mol) had better binding compared to Cmp19 (-95.216 kJ/mol). This study provides useful insight for better understanding of the binding mechanism of Cmp10 and Cmp19 and will be helpful in designing novel microtubule stabilizers.

  2. Long astral microtubules and RACK-1 stabilize polarity domains during maintenance phase in Caenorhabditis elegans embryos.

    Directory of Open Access Journals (Sweden)

    Erkang Ai

    Full Text Available Cell polarity is a very well conserved process important for cell differentiation, cell migration, and embryonic development. After the establishment of distinct cortical domains, polarity cues have to be stabilized and maintained within a fluid and dynamic membrane to achieve proper cell asymmetry. Microtubules have long been thought to deliver the signals required to polarize a cell. While previous studies suggest that microtubules play a key role in the establishment of polarity, the requirement of microtubules during maintenance phase remains unclear. In this study, we show that depletion of Caenorhabditis elegans RACK-1, which leads to short astral microtubules during prometaphase, specifically affects maintenance of cortical PAR domains and Dynamin localization. We then investigated the consequence of knocking down other factors that also abolish astral microtubule elongation during polarity maintenance phase. We found a correlation between short astral microtubules and the instability of PAR-6 and PAR-2 domains during maintenance phase. Our data support a necessary role for astral microtubules in the maintenance phase of cell polarity.

  3. Crowding of molecular motors determines microtubule depolymerization

    CERN Document Server

    Reese, Louis; Frey, Erwin

    2011-01-01

    Assembly and disassembly dynamics of microtubules (MTs) is tightly controlled by MT associated proteins. Here, we investigate how plus-end-directed depolymerases of the kinesin-8 family regulate MT depolymerization dynamics. Employing an individual-based model, we reproduce experimental findings. Moreover, crowding is identified as the key regulatory mechanism of depolymerization dynamics. Our analysis gives two qualitatively distinct regimes. For motor densities above a particular threshold, a macroscopic traffic jam emerges at the plus-end and the MT dynamics become independent of the motor concentration. Below this threshold, microscopic traffic jams at the tip arise which cancel out the effect of the depolymerization kinetics such that the depolymerization speed is solely determined by the motor density. Because this density changes over the MT length, length-dependent regulation is possible. Remarkably, motor cooperativity does not affect the depolymerization speed but only the end-residence time of depo...

  4. Modeling microtubule oscillations

    DEFF Research Database (Denmark)

    Jobs, E.; Wolf, D.E.; Flyvbjerg, H.

    1997-01-01

    Synchronization of molecular reactions in a macroscopic volume may cause the volume's physical properties to change dynamically and thus reveal much about the reactions. As an example, experimental time series for so-called microtubule oscillations are analyzed in terms of a minimal model...... for this complex polymerization-depolymerization cycle. The model reproduces well the qualitatively different time series that result from different experimental conditions, and illuminates the role and importance of individual processes in the cycle. Simple experiments are suggested that can further test...... and define the model and the polymer's reaction cycle....

  5. The use of compressive sensing and peak detection in the reconstruction of microtubules length time series in the process of dynamic instability.

    Science.gov (United States)

    Mahrooghy, Majid; Yarahmadian, Shantia; Menon, Vineetha; Rezania, Vahid; Tuszynski, Jack A

    2015-10-01

    Microtubules (MTs) are intra-cellular cylindrical protein filaments. They exhibit a unique phenomenon of stochastic growth and shrinkage, called dynamic instability. In this paper, we introduce a theoretical framework for applying Compressive Sensing (CS) to the sampled data of the microtubule length in the process of dynamic instability. To reduce data density and reconstruct the original signal with relatively low sampling rates, we have applied CS to experimental MT lament length time series modeled as a Dichotomous Markov Noise (DMN). The results show that using CS along with the wavelet transform significantly reduces the recovery errors comparing in the absence of wavelet transform, especially in the low and the medium sampling rates. In a sampling rate ranging from 0.2 to 0.5, the Root-Mean-Squared Error (RMSE) decreases by approximately 3 times and between 0.5 and 1, RMSE is small. We also apply a peak detection technique to the wavelet coefficients to detect and closely approximate the growth and shrinkage of MTs for computing the essential dynamic instability parameters, i.e., transition frequencies and specially growth and shrinkage rates. The results show that using compressed sensing along with the peak detection technique and wavelet transform in sampling rates reduces the recovery errors for the parameters.

  6. Microtubule heterogeneity of Ornithogalum umbellatum ovary epidermal cells: non-stable cortical microtubules and stable lipotubuloid microtubules

    Directory of Open Access Journals (Sweden)

    Maria Kwiatkowska

    2011-07-01

    Full Text Available Lipotubuloids, structures containing lipid bodies and microtubules, are described in ovary epidermalcells of Ornithogalum umbellatum. Microtubules of lipotubuloids can be fixed in electron microscope fixativecontaining only buffered OsO4 or in glutaraldehyde with OsO4 post-fixation, or in a mixture of OsO4 and glutaraldehyde[1]. None of these substances fixes cortical microtubules of ovary epidermis of this plant which ischaracterized by dynamic longitudinal growth. However, cortical microtubules can be fixed with cold methanolaccording immunocytological methods with the use of b-tubulin antibodies and fluorescein. The existence ofcortical microtubules has also been evidenced by EM observations solely after the use of taxol, microtubulestabilizer, and fixation in a glutaraldehyde/OsO4 mixture. These microtubules mostly lie transversely, sometimesobliquely, and rarely parallel to the cell axis. Staining, using Ruthenium Red and silver hexamine, has revealedthat lipotubuloid microtubules surface is covered with polysaccharides. The presumption has been made thatthe presence of a polysaccharide layer enhances the stability of lipotubuloid microtubules.

  7. Physical Basis of Large Microtubule Aster Growth

    CERN Document Server

    Ishihara, Keisuke; Mitchison, Timothy J

    2016-01-01

    Microtubule asters - radial arrays of microtubules organized by centrosomes - play a fundamental role in the spatial coordination of animal cells. The standard model of aster growth assumes a fixed number of microtubules originating from the centrosomes. However, aster morphology in this model does not scale with cell size, and we recently found evidence for non-centrosomal microtubule nucleation. Here, we combine autocatalytic nucleation and polymerization dynamics to develop a biophysical model of aster growth. Our model predicts that asters expand as traveling waves and recapitulates all major aspects of aster growth. As the nucleation rate increases, the model predicts an explosive transition from stationary to growing asters with a discontinuous jump of the growth velocity to a nonzero value. Experiments in frog egg extract confirm the main theoretical predictions. Our results suggest that asters observed in large frog and amphibian eggs are a meshwork of short, unstable microtubules maintained by autoca...

  8. Effect of microtubule-associated protein tau in dynamics of single-headed motor proteins KIF1A

    CERN Document Server

    Sparacino, J; Lamberti, P W

    2013-01-01

    Intracellular transport based on molecular motors and its regulation are crucial to the functioning of cells. Filamentary tracks of the cells are abundantly decorated with non-motile microtubule-associated proteins, such as tau. Motivated by experiments on kinesin-tau interactions [Dixit et al. Science 319, 1086 (2008)] we developed a stochastic model of interacting single-headed motor proteins KIF1A that also takes into account the interactions between motor proteins and tau molecules. Our model reproduce experimental observations and predicts significant effects of tau on bound time and run length which suggest an important role of tau in regulation of kinesin-based transport.

  9. Discodermolide interferes with the binding of tau protein to microtubules.

    Science.gov (United States)

    Kar, Santwana; Florence, Gordon J; Paterson, Ian; Amos, Linda A

    2003-03-27

    We investigated whether discodermolide, a novel antimitotic agent, affects the binding to microtubules of tau protein repeat motifs. Like taxol, the new drug reduces the proportion of tau that pellets with microtubules. Despite their differing structures, discodermolide, taxol and tau repeats all bind to a site on beta-tubulin that lies within the microtubule lumen and is crucial in controlling microtubule assembly. Low concentrations of tau still bind strongly to the outer surfaces of preformed microtubules when the acidic C-terminal regions of at least six tubulin dimers are available for interaction with each tau molecule; otherwise binding is very weak.

  10. Pharmacologic reductions of total tau levels; implications for the role of microtubule dynamics in regulating tau expression

    Directory of Open Access Journals (Sweden)

    Dickey Chad A

    2006-07-01

    Full Text Available Abstract The microtubule-associated protein tau (MAPT is a pathological component of several neurodegenerative diseases and clinical dementias. Here, we have investigated the effects of a series of commercially available FDA-approved compounds and natural products on total tau protein levels using a cell-based approach that allows for the rapid and efficient measurement of changes in protein expression. Results The compounds that reduced tau largely fell within 3 functional categories with the largest percentage being microtubule regulators. Several of these candidates were validated in both a human neuroglioma and a human neuroblastoma cell line. While these drugs lead to a rapid reduction in tau protein levels, a selective decrease in MAPT mRNA expression was also observed. Conclusion These findings suggest that the identified compounds that reduce tau levels may act either through direct effects on the MAPT promoter itself or by altering a feedback transcriptional mechanism regulating MAPT transcription. This is particularly interesting in light of recent evidence suggesting that MAPT 5' UTR mutations in late-onset PD and PSP cases alter the expression of tau mRNA. In fact, one of the compounds we identified, rotenone, has been used extensively to model PD in rodents. These observations may provide key insights into the mechanism of tau turnover within the neuron while also providing the first evidence that selectively reducing tau protein levels may be possible using compounds that are FDA-approved for other uses.

  11. Anomalous motor mediated cargo transport in microtubule networks

    Science.gov (United States)

    Vandal, Steven; Macveigh-Fierro, Daniel; Shen, Zhiyuan; Lemoi, Kyle; Vidali, Luis; Ross, Jennifer; Tuzel, Erkan

    Cargo transport is an important biological mechanism by which cells locomote, self-organize, and actively transport organelles. This transport is mediated by the cytoskeletal network and molecular motors; however, it is not known how network self-organization and dynamics affect these transport processes. In order to develop a mechanistic understanding of cargo transport, we use a coarse-grained Brownian dynamics model that incorporates the dynamics of these networks, as well as experimentally determined motor properties. We will test these models with two experimental systems: (1) in vitro microtubule networks with kinesin-1 motors, and quantum dot cargos on recreated microtubule networks, and (2) an excellent model organism, the moss Physcomitrella patens, in which chloroplasts are transported via the microtubule network by means of kinesin-like proteins. Phenomenological network characterizations are made, both in vivo and in vitro, and cargo motility is characterized using Mean Squared Displacement (MSD) measurements. Our simulations shed light on the role of network density and motor properties on the observed transport behavior, and improve our understanding of cargo transport in cells.

  12. Wave->Diffusion Transition in Microtubules

    OpenAIRE

    2005-01-01

    In this paper the heat transport in microtubules (MT) is investigated. When the dimension of the structure is of the order of the de Broglie wave length the transport phenomena must be analyzed within quantum mechanics. In this paper we developed the Dirac type thermal equation for MT .The solution of the equation-the temperature fields for electrons can be wave type or diffusion type depending on the dynamics of the scattering. Key words: Microtubules ultrashort laser pulses, Dirac thermal e...

  13. Dynamic artificial neural networks with affective systems.

    Science.gov (United States)

    Schuman, Catherine D; Birdwell, J Douglas

    2013-01-01

    Artificial neural networks (ANNs) are processors that are trained to perform particular tasks. We couple a computational ANN with a simulated affective system in order to explore the interaction between the two. In particular, we design a simple affective system that adjusts the threshold values in the neurons of our ANN. The aim of this paper is to demonstrate that this simple affective system can control the firing rate of the ensemble of neurons in the ANN, as well as to explore the coupling between the affective system and the processes of long term potentiation (LTP) and long term depression (LTD), and the effect of the parameters of the affective system on its performance. We apply our networks with affective systems to a simple pole balancing example and briefly discuss the effect of affective systems on network performance.

  14. Microtubules, Tubulins and Associated Proteins.

    Science.gov (United States)

    Raxworthy, Michael J.

    1988-01-01

    Reviews much of what is known about microtubules, which are biopolymers consisting predominantly of subunits of the globular protein, tubulin. Describes the functions of microtubules, their structure and assembly, microtube associated proteins, and microtubule-disrupting agents. (TW)

  15. Microtubule remodeling mediates the inhibition of store-operated calcium entry (SOCE) during mitosis in COS-7 cells.

    Science.gov (United States)

    Russa, Afadhali Denis; Ishikita, Naoyuki; Masu, Kazuki; Akutsu, Hitomi; Saino, Tomoyuki; Satoh, Yoh-ichi

    2008-12-01

    Regulation of the intracellular calcium ion concentration ([Ca(2+)](i)) is critical, because calcium signaling controls diverse and vital cellular processes such as secretion, proliferation, division, gene transcription, and apoptosis. Store-operated calcium entry (SOCE) is the main mechanism through which non-excitable cells replenish and thus maintain this delicate balance. There is limited evidence which indicates that SOCE may be inhibited during mitosis, and the mechanisms leading to the presumed inhibition has not been elucidated. In the present study, we examined and compared the [Ca(2+)](i) dynamics of COS-7 cells in mitotic and non-mitotic phases with special reference paid to SOCE. Laser scanning confocal microscopy to monitor [Ca(2+)](i) dynamics revealed that SOCE was progressively inhibited in mitosis and became virtually absent during the metaphase. We used various cytoskeletal modifying drugs and immunofluorescence to assess the contribution of microtubule and actin filaments in SOCE signaling. Nocodazole treatment caused microtubule reorganization and retraction from the cell periphery that mimicked the natural mitotic microtubule remodeling that was also accompanied by SOCE inhibition. Short exposure to paclitaxel, a microtubule-stabilizing drug, bolstered SOCE, whereas long exposure resulted in microtubule disruption and SOCE inhibition. Actin-modifying drugs did not affect SOCE. These findings indicate that mitotic microtubule remodeling plays a significant role in the inhibition of SOCE during mitosis.

  16. Microtubule stabilization reduces scarring and causes axon regeneration after spinal cord injury

    NARCIS (Netherlands)

    F. Hellal (Farida); A. Hurtado (Andres); J. Ruschel (Jörg); K.C. Flynn (Kevin); C.J. Laskowski (Claudia); M. Umlauf (Martina); L.C. Kapitein (Lukas); D. Strikis (Dinara); V. Lemmon (Vance); J. Bixby (John); C.C. Hoogenraad (Casper); F. Bradke (Frank)

    2011-01-01

    textabstractHypertrophic scarring and poor intrinsic axon growth capacity constitute major obstacles for spinal cord repair. These processes are tightly regulated by microtubule dynamics. Here, moderate microtubule stabilization decreased scar formation after spinal cord injury in rodents through va

  17. How Resource Phenology Affects Consumer Population Dynamics.

    Science.gov (United States)

    Bewick, Sharon; Cantrell, R Stephen; Cosner, Chris; Fagan, William F

    2016-02-01

    Climate change drives uneven phenology shifts across taxa, and this can result in changes to the phenological match between interacting species. Shifts in the relative phenology of partner species are well documented, but few studies have addressed the effects of such changes on population dynamics. To explore this, we develop a phenologically explicit model describing consumer-resource interactions. Focusing on scenarios for univoltine insects, we show how changes in resource phenology can be reinterpreted as transformations in the year-to-year recursion relationships defining consumer population dynamics. This perspective provides a straightforward path for interpreting the long-term population consequences of phenology change. Specifically, by relating the outcome of phenological shifts to species traits governing recursion relationships (e.g., consumer fecundity or competitive scenario), we demonstrate how changes in relative phenology can force systems into different dynamical regimes, with major implications for resource management, conservation, and other areas of applied dynamics.

  18. Effects of silver ions (Ag+) on contractile ring function and microtubule dynamics during first cleavage in Ilyanassa obsoleta

    Science.gov (United States)

    Conrad, A. H.; Stephens, A. P.; Paulsen, A. Q.; Schwarting, S. S.; Conrad, G. W.; Spooner, B. S. (Principal Investigator)

    1994-01-01

    The terminal phase of cell division involves tight constriction of the cleavage furrow contractile ring, stabilization/elongation of the intercellular bridge, and final separation of the daughter cells. At first cleavage, the fertilized eggs of the mollusk, Ilyanassa obsoleta, form two contractile rings at right angles to each other in the same cytoplasm that constrict to tight necks and partition the egg into a trefoil shape. The cleavage furrow contractile ring (CF) normally constricts around many midbody microtubules (MTs) and results in cleavage; the polar lobe constriction contractile ring (PLC) normally constricts around very few MTs and subsequently relaxes without cleavage. In the presence of Ag+ ions, the PLC 1) begins MT-dependent rapid constriction sooner than controls, 2) encircles more MTs than control egg PLCs, 3) elongates much more than control PLCs, and 4) remains tightly constricted and effectively cleaves the polar lobe from the egg. If Ag(+)-incubated eggs are returned to normal seawater at trefoil, tubulin fluorescence disappears from the PLC neck and the neck relaxes. If nocodazole, a drug that depolymerizes MTs, is added to Ag(+)-incubated eggs during early PLC constriction, the PLC is not stabilized and eventually relaxes. However, if nocodazole is added to Ag(+)-incubated eggs at trefoil, tubulin fluorescence disappears from the PLC neck but the neck remains constricted. These results suggest that Ag+ accelerates and gradually stabilizes the PLC constriction by a mechanism that is initially MT-dependent, but that progressively becomes MT-independent.

  19. Measurement of Binding Force between Microtubule-Associated Protein and Microtubule

    Institute of Scientific and Technical Information of China (English)

    XU Chun-Hua; GUO Hong-Lian; QU E; LI Zhao-Lin; YUAN Ming; CHENG Bing-Ying; ZHANG Dao-Zhong

    2007-01-01

    Microtubule-associated proteins (MAPs) are important proteins in cells. They can regulate the organization,dynamics and function of microtubules. We measure the binding force between microtubule and a new plant MAP, i.e. AtMAP65-1, by dual-optical tweezers. The force is obtained to be 14.6±3.5 pN from the data statistics and analysis. This force measurement is helpful to understand the function and mechanism of MAPs from the mechanical point of view and lays the groundwork for future measurements of the mechanical properties of other biological macro-molecules.

  20. Fission yeast mtr1p regulates interphase microtubule cortical dwell-time

    Directory of Open Access Journals (Sweden)

    Frédérique Carlier-Grynkorn

    2014-06-01

    Full Text Available The microtubule cytoskeleton plays important roles in cell polarity, motility and division. Microtubules inherently undergo dynamic instability, stochastically switching between phases of growth and shrinkage. In cells, some microtubule-associated proteins (MAPs and molecular motors can further modulate microtubule dynamics. We present here the fission yeast mtr1+, a new regulator of microtubule dynamics that appears to be not a MAP or a motor. mtr1-deletion (mtr1Δ primarily results in longer microtubule dwell-time at the cell tip cortex, suggesting that mtr1p acts directly or indirectly as a destabilizer of microtubules. mtr1p is antagonistic to mal3p, the ortholog of mammalian EB1, which stabilizes microtubules. mal3Δ results in short microtubules, but can be partially rescued by mtr1Δ, as the double mutant mal3Δ mtr1Δ exhibits longer microtubules than mal3Δ single mutant. By sequence homology, mtr1p is predicted to be a component of the ribosomal quality control complex. Intriguingly, deletion of a predicted ribosomal gene, rps1801, also resulted in longer microtubule dwell-time similar to mtr1Δ. The double-mutant mal3Δ rps1801Δ also exhibits longer microtubules than mal3Δ single mutant alone. Our study suggests a possible involvement of mtr1p and the ribosome complex in modulating microtubule dynamics.

  1. The dual specificity phosphatase Cdc14B bundles and stabilizes microtubules

    Energy Technology Data Exchange (ETDEWEB)

    Plumley, Hyekyung [ORNL; Liu, Yie [ORNL; Gomez, Marla V [ORNL; Wang, Yisong [ORNL

    2005-01-01

    The Cdc14 dual-specificity phosphatases regulate key events in the eukaryotic cell cycle. However, little is known about the function of mammalian CDC14B family members. Here, we demonstrate that subcellular localization of CDC14B protein is cell cycle regulated. CDC14B can bind, bundle, and stabilize microtubules in vitro independently of its catalytic activity. Basic amino acid residues within the nucleolar targeting domain are important for both retaining CDC14B in the nucleolus and preventing microtubule bundling. Overexpression of CDC14B resulted in the formation of cytoplasmic CDC14B and microtubule bundles in interphase cells. These microtubule bundles were resistant to microtubule depolymerization reagents and enriched in acetylated -tubulin. Expression of cytoplasmic forms of CDC14B impaired microtubule nucleation from the microtubule organization center. CDC14B is thus a novel microtubule-bundling and -stabilizing protein, whose regulated subcellular localization may help modulate spindle and microtubule dynamics in mitosis.

  2. In vitro assembly of plant tubulin in the absence of microtubule-stabilizing reagents

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    The assembly of microtubules is essential for physiological functions of microtubules. Addition of microtubule-stabilizing reagents or microtubule "seeds" is usually necessary for plant tubulin assembly in vitro, which hinders the investigation of plant microtubule dynamics. In the present note, highly purified plant tubulins have been obtained from lily pollen, a non-microtubule-stabilizing reagent or microtubule "seed" system for plant tubulin assembly has been established and the analysis of plant tubulin assembly performed. Experiment results showed that purified tubulin polymerized in vitro, and a typical microtubule structure was observed with electron microscopy. The kinetics curve of tubulin assembly exhibited typical "parabola". The presence of taxol significantly altered the character of plant tubulin assembly, including that abnormal microtubules were assembled and the critical concentration for plant tubulin assembly was decreased exceedingly from 3 mg/mL in the absence of taxol to 0.043 mg/mL in the presence of taxol.

  3. Microtubules Are Essential for Guard-Cell Function in Vicia and Arabidopsis

    Institute of Scientific and Technical Information of China (English)

    William Eisinger; David Ehrhardt; Winslow Briggs

    2012-01-01

    Radially arranged cortical microtubules are a prominent feature of guard cells.Guard cells expressing GFPtubulin showed consistent changes in the appearance of microtubules when stomata opened or closed.Guard cells showed fewer microtubule structures as stomata closed,whether induced by transfer to darkness,ABA,hydrogen peroxide,or sodium hydrogen carbonate.Guard cells kept in the dark (closed stomata) showed increases in microtubule structures and stomatal aperture on light treatment.GFP-EB1,marking microtubule growing plus ends,showed no change in number of plus ends or velocity of assembly on stomatal closure.Since the number of growing plus ends and the rate of plus-end growth did not change when microtubule structure numbers declined,microtubule instability and/or rearrangement must be responsible for the apparent loss of microtubules.Guard cells with closed stomata showed more cytosolic GFP-fluorescence than those with open stomata as cortical microtubules became disassembled,although with a large net loss in total fluorescence.Microtubule-targeted drugs blocked guard-cell function in Vicia and Arabidopsis.Oryzalin disrupted guard-cell microtubules and prevented stomatal opening and taxol stabilized guard-cell microtubules and delayed stomatal closure.Gas exchange measurements indicated that the transgenes for fluorescent-labeled proteins did not disrupt normal stomatal function.These dynamic changes in guard-cell microtubules combined with our inhibitor studies provide evidence for an active role of microtubules in guard-cell function.

  4. Microtubule stability and MAPI B upregulation control neuritogenesis in CAD cells

    Institute of Scientific and Technical Information of China (English)

    Wen LI; Jin-tang XIA; Yue FENG

    2006-01-01

    Aim: To study the role of microtubule dynamics and microtubule associated protein 1B (MAP1B) in regulation of the neurite extension in CAD catecholaminergic neuronal cell line. Methods: The neuritogenesis of the CAD cells was abolished by inhibiting microtubule polymerization with nocodazole and by blocking microtubule depolymerization with taxol. MAP1B and tubulin protein expression was detected by Western blot. Immunofluorescent staining of tubulins was observed by fluorescent and confocal microscopy. Results: Microtubule dynamics was essential for CAD neurite extension. Dosage analysis revealed that neurite extension was much more sensitive to nocodazole than to taxol, suggesting a functional requirement for highly active microtubule assembly. A remarkable upregulation of MAP1B protein was detected during neurite extension accompanied with increased microtubule stability. Conclusion: Upregulation of MAP1B leads to the stabilization of newly formed microtubules in the developing neurites, which in turn promotes neurite extension.

  5. Alignment of Microtubule Imagery

    CERN Document Server

    Yu, Feiyang; Bakker, Erwin M

    2011-01-01

    This work discusses preliminary work aimed at simulating and visualizing the growth process of a tiny structure inside the cell---the microtubule. Difficulty of recording the process lies in the fact that the tissue preparation method for electronic microscopes is highly destructive to live cells. Here in this paper, our approach is to take pictures of microtubules at different time slots and then appropriately combine these images into a coherent video. Experimental results are given on real data.

  6. Studies of dynamical processes affecting global climate

    Energy Technology Data Exchange (ETDEWEB)

    Keller, C.; Cooper, D.; Eichinger, W. [and others

    1998-12-31

    This is the final report of a three-year, Laboratory Directed Research and Development project at the Los Alamos National Laboratory (LANL). The main objective was, by a combined theoretical and observational approach, to develop improved models of dynamic processes in the oceans and atmosphere and to incorporate them into large climate codes, chiefly in four main areas: numerical physics, chemistry, water vapor, and ocean-atmosphere interactions. Main areas of investigation included studies of: cloud parameterizations for global climate codes, Lidar and the planetary boundary layer, chemistry, climate variability using coupled ocean-atmospheric models, and numerical physical methods. This project employed a unique approach that included participation of a number of University of California faculty, postdoctoral fellows and graduate students who collaborated with Los Alamos research staff on specific tasks, thus greatly enhancing the research output. Overall accomplishments during the sensing of the atmospheric planetary were: (1) first two- and three-dimensional remote sensing of the atmospheric planetary boundary layer using Lidars, (2) modeling of 20-year cycle in both pressure and sea surface temperatures in North Pacific, (3) modeling of low frequency internal variability, (4) addition of aerosols to stratosphere to simulate Pinatubo effect on ozone, (5) development of fast, comprehensive chemistry in the troposphere for urban pollution studies, (6) new prognostic cloud parameterization in global atmospheric code remedied problems with North Pacific atmospheric circulation and excessive equatorial precipitation, (7) development of a unique aerosol analysis technique, the aerosol time-of-flight mass spectrometer (ATOFMS), which allows real-time analysis of the size and chemical composition of individual aerosol particles, and (8) numerical physics applying Approximate Inertial Manifolds to ocean circulation. 14 refs., 6 figs.

  7. Micro-Level Affect Dynamics in Psychopathology Viewed From Complex Dynamical System Theory

    NARCIS (Netherlands)

    Wichers, M.; Wigman, J. T. W.; Myin-Germeys, I.

    2015-01-01

    This article discusses the role of moment-to-moment affect dynamics in mental disorder and aims to integrate recent literature on this topic in the context of complex dynamical system theory. First, we will review the relevance of temporal and contextual aspects of affect dynamics in relation to psy

  8. Phospholipase D activation correlates with microtubule reorganization in living plant cells

    NARCIS (Netherlands)

    P.B. Dhonukshe; A.M. Laxalt; J. Goedhart; Th.W.J. Gadella; T. Munnik

    2003-01-01

    A phospholipase D (PLD) was shown recently to decorate microtubules in plant cells. Therefore, we used tobacco BY-2 cells expressing the microtubule reporter GFP-MAP4 to test whether PLD activation affects the organization of plant microtubules. Within 30 min of adding n-butanol, a potent activator

  9. Biological Information Processing in Single Microtubules

    Science.gov (United States)

    2014-03-05

    the time for reviewing instructions, searching existing data sources, gathering and maintaining the data needed, and completing and reviewing the...generated by synchronized oscillations of microtubules, centrosomes and chromosomes regulate the dynamics of mitosis and meiosis , Yue Zhao and Qimin

  10. S. pombe kinesins-8 promote both nucleation and catastrophe of microtubules.

    Directory of Open Access Journals (Sweden)

    Muriel Erent

    Full Text Available The kinesins-8 were originally thought to be microtubule depolymerases, but are now emerging as more versatile catalysts of microtubule dynamics. We show here that S. pombe Klp5-436 and Klp6-440 are non-processive plus-end-directed motors whose in vitro velocities on S. pombe microtubules at 7 and 23 nm s(-1 are too slow to keep pace with the growing tips of dynamic interphase microtubules in living S. pombe. In vitro, Klp5 and 6 dimers exhibit a hitherto-undescribed combination of strong enhancement of microtubule nucleation with no effect on growth rate or catastrophe frequency. By contrast in vivo, both Klp5 and Klp6 promote microtubule catastrophe at cell ends whilst Klp6 also increases the number of interphase microtubule arrays (IMAs. Our data support a model in which Klp5/6 bind tightly to free tubulin heterodimers, strongly promoting the nucleation of new microtubules, and then continue to land as a tubulin-motor complex on the tips of growing microtubules, with the motors then dissociating after a few seconds residence on the lattice. In vivo, we predict that only at cell ends, when growing microtubule tips become lodged and their growth slows down, will Klp5/6 motor activity succeed in tracking growing microtubule tips. This mechanism would allow Klp5/6 to detect the arrival of microtubule tips at cells ends and to amplify the intrinsic tendency for microtubules to catastrophise in compression at cell ends. Our evidence identifies Klp5 and 6 as spatial regulators of microtubule dynamics that enhance both microtubule nucleation at the cell centre and microtubule catastrophe at the cell ends.

  11. Taxol crystals can masquerade as stabilized microtubules.

    Directory of Open Access Journals (Sweden)

    Margit Foss

    Full Text Available Taxol is a potent anti-mitotic drug used in chemotherapy, angioplastic stents, and cell biology research. By binding and stabilizing microtubules, Taxol inhibits their dynamics, crucial for cell division, motility, and survival. The drug has also been reported to induce formation of asters and bundles composed of stabilized microtubules. Surprisingly, at commonly used concentrations, Taxol forms crystals that rapidly bind fluorescent tubulin subunits, generating structures with an uncanny resemblance to microtubule asters and bundles. Kinetic and topological considerations suggest that tubulin subunits, rather than microtubules, bind the crystals. This sequestration of tubulin from the subunit pool would be expected to shift the equilibrium of free to polymerized tubulin to disfavor assembly. Our results imply that some previously reported Taxol-induced asters or bundles could include or be composed of tubulin-decorated Taxol crystals. Thus, reevaluation of certain morphological, chemical, and physical properties of Taxol-treated microtubules may be necessary. Moreover, our findings suggest a novel mechanism for chemotherapy-induced cytotoxicity in non-dividing cells, with far-reaching medical implications.

  12. Do prokaryotes contain microtubules?

    Science.gov (United States)

    Bermudes, D.; Hinkle, G.; Margulis, L.

    1994-01-01

    In eukaryotic cells, microtubules are 24-nm-diameter tubular structures composed of a class of conserved proteins called tubulin. They are involved in numerous cell functions including ciliary motility, nerve cell elongation, pigment migration, centrosome formation, and chromosome movement. Although cytoplasmic tubules and fibers have been observed in bacteria, some with diameters similar to those of eukaryotes, no homologies to eukaryotic microtubules have been established. Certain groups of bacteria including azotobacters, cyanobacteria, enteric bacteria, and spirochetes have been frequently observed to possess microtubule-like structures, and others, including archaebacteria, have been shown to be sensitive to drugs that inhibit the polymerization of microtubules. Although little biochemical or molecular biological information is available, the differences observed among these prokaryotic structures suggest that their composition generally differs among themselves as well as from that of eukaryotes. We review the distribution of cytoplasmic tubules in prokaryotes, even though, in all cases, their functions remain unknown. At least some tend to occur in cells that are large, elongate, and motile, suggesting that they may be involved in cytoskeletal functions, intracellular motility, or transport activities comparable to those performed by eukaryotic microtubules. In Escherichia coli, the FtsZ protein is associated with the formation of a ring in the division zone between the newly forming offspring cells. Like tubulin, FtsZ is a GTPase and shares with tubulin a 7-amino-acid motif, making it a promising candidate in which to seek the origin of tubulins.

  13. Pseudomonas aeruginosa exotoxin Y-mediated tau hyperphosphorylation impairs microtubule assembly in pulmonary microvascular endothelial cells.

    Directory of Open Access Journals (Sweden)

    Ron Balczon

    Full Text Available Pseudomonas aeruginosa uses a type III secretion system to introduce the adenylyl and guanylyl cyclase exotoxin Y (ExoY into the cytoplasm of endothelial cells. ExoY induces Tau hyperphosphorylation and insolubility, microtubule breakdown, barrier disruption and edema, although the mechanism(s responsible for microtubule breakdown remain poorly understood. Here we investigated both microtubule behavior and centrosome activity to test the hypothesis that ExoY disrupts microtubule dynamics. Fluorescence microscopy determined that infected pulmonary microvascular endothelial cells contained fewer microtubules than control cells, and further studies demonstrated that the microtubule-associated protein Tau was hyperphosphorylated following infection and dissociated from microtubules. Disassembly/reassembly studies determined that microtubule assembly was disrupted in infected cells, with no detectable effects on either microtubule disassembly or microtubule nucleation by centrosomes. This effect of ExoY on microtubules was abolished when the cAMP-dependent kinase phosphorylation site (Ser-214 on Tau was mutated to a non-phosphorylatable form. These studies identify Tau in microvascular endothelial cells as the target of ExoY in control of microtubule architecture following pulmonary infection by Pseudomonas aeruginosa and demonstrate that phosphorylation of tau following infection decreases microtubule assembly.

  14. Kinetochore-Dependent Microtubule Rescue Ensures Their Efficient and Sustained Interactions in Early Mitosis

    Science.gov (United States)

    Gandhi, Sapan R.; Gierliński, Marek; Mino, Akihisa; Tanaka, Kozo; Kitamura, Etsushi; Clayton, Lesley; Tanaka, Tomoyuki U.

    2011-01-01

    Summary How kinetochores regulate microtubule dynamics to ensure proper kinetochore-microtubule interactions is unknown. Here, we studied this during early mitosis in Saccharomyces cerevisiae. When a microtubule shrinks and its plus end reaches a kinetochore bound to its lateral surface, the microtubule end attempts to tether the kinetochore. This process often fails and, responding to this failure, microtubule rescue (conversion from shrinkage to growth) occurs, preventing kinetochore detachment from the microtubule end. This rescue is promoted by Stu2 transfer (ortholog of vertebrate XMAP215/ch-TOG) from the kinetochore to the microtubule end. Meanwhile, microtubule rescue distal to the kinetochore is also promoted by Stu2, which is transported by a kinesin-8 motor Kip3 along the microtubule from the kinetochore. Microtubule extension following rescue facilitates interaction with other widely scattered kinetochores, diminishing long delays in collecting the complete set of kinetochores by microtubules. Thus, kinetochore-dependent microtubule rescue ensures efficient and sustained kinetochore-microtubule interactions in early mitosis. PMID:22075150

  15. CYLD Regulates Noscapine Activity in Acute Lymphoblastic Leukemia via a Microtubule-Dependent Mechanism.

    Science.gov (United States)

    Yang, Yunfan; Ran, Jie; Sun, Lei; Sun, Xiaodong; Luo, Youguang; Yan, Bing; Tala; Liu, Min; Li, Dengwen; Zhang, Lei; Bao, Gang; Zhou, Jun

    2015-01-01

    Noscapine is an orally administrable drug used worldwide for cough suppression and has recently been demonstrated to disrupt microtubule dynamics and possess anticancer activity. However, the molecular mechanisms regulating noscapine activity remain poorly defined. Here we demonstrate that cylindromatosis (CYLD), a microtubule-associated tumor suppressor protein, modulates the activity of noscapine both in cell lines and in primary cells of acute lymphoblastic leukemia (ALL). Flow cytometry and immunofluorescence microscopy reveal that CYLD increases the ability of noscapine to induce mitotic arrest and apoptosis. Examination of cellular microtubules as well as in vitro assembled microtubules shows that CYLD enhances the effect of noscapine on microtubule polymerization. Microtubule cosedimentation and fluorescence titration assays further reveal that CYLD interacts with microtubule outer surface and promotes noscapine binding to microtubules. These findings thus demonstrate CYLD as a critical regulator of noscapine activity and have important implications for ALL treatment.

  16. Microtubule doublets are double-track railways for intraflagellar transport trains.

    Science.gov (United States)

    Stepanek, Ludek; Pigino, Gaia

    2016-05-06

    The cilium is a large macromolecular machine that is vital for motility, signaling, and sensing in most eukaryotic cells. Its conserved core structure, the axoneme, contains nine microtubule doublets, each comprising a full A-microtubule and an incomplete B-microtubule. However, thus far, the function of this doublet geometry has not been understood. We developed a time-resolved correlative fluorescence and three-dimensional electron microscopy approach to investigate the dynamics of intraflagellar transport (IFT) trains, which carry ciliary building blocks along microtubules during the assembly and disassembly of the cilium. Using this method, we showed that each microtubule doublet is used as a bidirectional double-track railway: Anterograde IFT trains move along B-microtubules, and retrograde trains move along A-microtubules. Thus, the microtubule doublet geometry provides direction-specific rails to coordinate bidirectional transport of ciliary components.

  17. Multiscale Polar Theory of Microtubule and Motor-Protein Assemblies

    Science.gov (United States)

    Gao, Tong; Blackwell, Robert; Glaser, Matthew A.; Betterton, M. D.; Shelley, Michael J.

    2015-01-01

    Microtubules and motor proteins are building blocks of self-organized subcellular biological structures such as the mitotic spindle and the centrosomal microtubule array. These same ingredients can form new “bioactive” liquid-crystalline fluids that are intrinsically out of equilibrium and which display complex flows and defect dynamics. It is not yet well understood how microscopic activity, which involves polarity-dependent interactions between motor proteins and microtubules, yields such larger-scale dynamical structures. In our multiscale theory, Brownian dynamics simulations of polar microtubule ensembles driven by cross-linking motors allow us to study microscopic organization and stresses. Polarity sorting and cross-link relaxation emerge as two polar-specific sources of active destabilizing stress. On larger length scales, our continuum Doi-Onsager theory captures the hydrodynamic flows generated by polarity-dependent active stresses. The results connect local polar structure to flow structures and defect dynamics. PMID:25679909

  18. The ability to induce microtubule acetylation is a general feature of formin proteins.

    Directory of Open Access Journals (Sweden)

    Susan F Thurston

    Full Text Available Cytoplasmic microtubules exist as distinct dynamic and stable populations within the cell. Stable microtubules direct and maintain cell polarity and it is thought that their stabilization is dependent on coordinative organization between the microtubule network and the actin cytoskeleton. A growing body of work suggests that some members of the formin family of actin remodeling proteins also regulate microtubule organization and stability. For example, we showed previously that expression of the novel formin INF1 is sufficient to induce microtubule stabilization and tubulin acetylation, but not tubulin detyrosination. An important issue with respect to the relationship between formins and microtubules is the determination of which formin domains mediate microtubule stabilization. INF1 has a distinct microtubule-binding domain at its C-terminus and the endogenous INF1 protein is associated with the microtubule network. Surprisingly, the INF1 microtubule-binding domain is not essential for INF1-induced microtubule acetylation. We show here that expression of the isolated FH1 + FH2 functional unit of INF1 is sufficient to induce microtubule acetylation independent of the INF1 microtubule-binding domain. It is not yet clear whether or not microtubule stabilization is a general property of all mammalian formins; therefore we expressed constitutively active derivatives of thirteen of the fifteen mammalian formin proteins in HeLa and NIH3T3 cells and measured their effects on stress fiber formation, MT organization and MT acetylation. We found that expression of the FH1 + FH2 unit of the majority of mammalian formins is sufficient to induce microtubule acetylation. Our results suggest that the regulation of microtubule acetylation is likely a general formin activity and that the FH2 should be thought of as a dual-function domain capable of regulating both actin and microtubule networks.

  19. Relations between affective music and speech: Evidence from dynamics of affective piano performance and speech production

    Directory of Open Access Journals (Sweden)

    Xiaoluan eLiu

    2015-07-01

    Full Text Available This study compares affective piano performance with speech production from the perspective of dynamics: unlike previous research, this study uses finger force and articulatory effort as indexes reflecting the dynamics of affective piano performance and speech production respectively. Moreover, for the first time physical constraints such as piano fingerings and speech articulatory distance are included due to their potential contribution to different patterns of dynamics. A piano performance experiment and speech production experiment were conducted in four emotions: anger, fear, happiness and sadness. The results show that in both piano performance and speech production, anger and happiness generally have high dynamics while sadness has the lowest dynamics, with fear in the middle. Fingerings interact with fear in the piano experiment and articulatory distance interacts with anger in the speech experiment, i.e., large physical constraints produce significantly higher dynamics than small physical constraints in piano performance under the condition of fear and in speech production under the condition of anger. Using production experiments, this study firstly supports previous perception studies on relations between affective music and speech. Moreover, this is the first study to show quantitative evidence for the importance of considering motor aspects such as dynamics in comparing music performance and speech production in which motor mechanisms play a crucial role.

  20. Relations between affective music and speech: evidence from dynamics of affective piano performance and speech production.

    Science.gov (United States)

    Liu, Xiaoluan; Xu, Yi

    2015-01-01

    This study compares affective piano performance with speech production from the perspective of dynamics: unlike previous research, this study uses finger force and articulatory effort as indexes reflecting the dynamics of affective piano performance and speech production respectively. Moreover, for the first time physical constraints such as piano fingerings and speech articulatory constraints are included due to their potential contribution to different patterns of dynamics. A piano performance experiment and speech production experiment were conducted in four emotions: anger, fear, happiness and sadness. The results show that in both piano performance and speech production, anger and happiness generally have high dynamics while sadness has the lowest dynamics. Fingerings interact with fear in the piano experiment and articulatory constraints interact with anger in the speech experiment, i.e., large physical constraints produce significantly higher dynamics than small physical constraints in piano performance under the condition of fear and in speech production under the condition of anger. Using production experiments, this study firstly supports previous perception studies on relations between affective music and speech. Moreover, this is the first study to show quantitative evidence for the importance of considering motor aspects such as dynamics in comparing music performance and speech production in which motor mechanisms play a crucial role.

  1. Depolymerizing kinesins Kip3 and MCAK shape cellular microtubule architecture by differential control of catastrophe.

    Science.gov (United States)

    Gardner, Melissa K; Zanic, Marija; Gell, Christopher; Bormuth, Volker; Howard, Jonathon

    2011-11-23

    Microtubules are dynamic filaments whose ends alternate between periods of slow growth and rapid shortening as they explore intracellular space and move organelles. A key question is how regulatory proteins modulate catastrophe, the conversion from growth to shortening. To study this process, we reconstituted microtubule dynamics in the absence and presence of the kinesin-8 Kip3 and the kinesin-13 MCAK. Surprisingly, we found that, even in the absence of the kinesins, the microtubule catastrophe frequency depends on the age of the microtubule, indicating that catastrophe is a multistep process. Kip3 slowed microtubule growth in a length-dependent manner and increased the rate of aging. In contrast, MCAK eliminated the aging process. Thus, both kinesins are catastrophe factors; Kip3 mediates fine control of microtubule length by narrowing the distribution of maximum lengths prior to catastrophe, whereas MCAK promotes rapid restructuring of the microtubule cytoskeleton by making catastrophe a first-order random process.

  2. A novel role for aquaporin-5 in enhancing microtubule organization and stability.

    Directory of Open Access Journals (Sweden)

    Venkataramana K Sidhaye

    Full Text Available Aquaporin-5 (AQP5 is a water-specific channel located on the apical surface of airway epithelial cells. In addition to regulating transcellular water permeability, AQP5 can regulate paracellular permeability, though the mechanisms by which this occurs have not been determined. Microtubules also regulate paracellular permeability. Here, we report that AQP5 promotes microtubule assembly and helps maintain the assembled microtubule steady state levels with slower turnover dynamics in cells. Specifically, reduced levels of AQP5 correlated with lower levels of assembled microtubules and decreased paracellular permeability. In contrast, overexpression of AQP5 increased assembly of microtubules, with evidence of increased MT stability, and promoted the formation of long straight microtubules in the apical domain of the epithelial cells. These findings indicate that AQP5-mediated regulation of microtubule dynamics modulates airway epithelial barrier properties and epithelial function.

  3. PACSIN1, a Tau-interacting protein, regulates axonal elongation and branching by facilitating microtubule instability.

    Science.gov (United States)

    Liu, Yingying; Lv, Kaosheng; Li, Zenglong; Yu, Albert C H; Chen, Jianguo; Teng, Junlin

    2012-11-16

    Tau is a major member of the neuronal microtubule-associated proteins. It promotes tubulin assembly and stabilizes axonal microtubules. Previous studies have demonstrated that Tau forms cross-bridges between microtubules, with some particles located on cross-bridges, suggesting that some proteins interact with Tau and might be involved in regulating Tau-related microtubule dynamics. This study reports that PACSIN1 interacts with Tau in axon. PACSIN1 blockade results in impaired axonal elongation and a higher number of primary axonal branches in mouse dorsal root ganglia neurons, which is induced by increasing the binding ability of Tau to microtubules. In PACSIN1-blocked dorsal root ganglia neurons, a greater amount of Tau is inclined to accumulate in the central domain of growth cones, and it promotes the stability of the microtubule network. Taken together, these results suggest that PACSIN1 is an important Tau binding partner in regulating microtubule dynamics and forming axonal plasticity.

  4. Microtubule's conformational cap

    DEFF Research Database (Denmark)

    Chretien, D.; Janosi, I.; Taveau, J.C.

    1999-01-01

    The molecular mechanisms that allow elongation of the unstable microtubule lattice remain unclear. It is usually thought that the GDP-liganded tubulin lattice is capped by a small layer of GTP- or GDP-P(i)-liganded molecules, the so called "GTP-cap". Here, we point-out that the elastic properties...

  5. Dynamic modelling of processes in rivers affected by precipitation runoff

    DEFF Research Database (Denmark)

    Jacobsen, Judith L.

    1997-01-01

    In this thesis, models for the dynamics of oxygen and organic matter in receiving waters (such as rivers and creeks), which are affected by rain, are developed. A time series analysis framework is used, but presented with special emphasis on continuous time state space models. Also, the concept...

  6. Dynamics of learner affective development in early FLL

    Directory of Open Access Journals (Sweden)

    Jelena Mihaljević Djigunović

    2012-10-01

    Full Text Available Affective learner factors were first considered as a cause of success in language learning. This was followed by a change in approach and recently authors (e.g., Edelenbos, Johnstone, & Kubanek, 2006 have considered them an important outcome, especially in early foreign language learning (FLL. Current research into affective learner factors in early FLL tries to catch the developmental aspects too, and studies are emerging that take a contextual view as well. This paper describes a study on affective characteristics of young FL learners that combines the developmental and contextual perspectives. Using the case study methodology the author analyses the affective profiles of three young learners of English as a foreign language who were followed for 4 years. The analyses are done taking into account their immediate language learning environment, home support, out-of-school exposure to English and language achievement. The findings suggest that affective learner factors contribute to the dynamic complexity of early FLL.

  7. Network Diversity and Affect Dynamics: The Role of Personality Traits

    Science.gov (United States)

    Alshamsi, Aamena; Pianesi, Fabio; Lepri, Bruno; Pentland, Alex; Rahwan, Iyad

    2016-01-01

    People divide their time unequally among their social contacts due to time constraints and varying strength of relationships. It was found that high diversity of social communication, dividing time more evenly among social contacts, is correlated with economic well-being both at macro and micro levels. Besides economic well-being, it is not clear how the diversity of social communication is also associated with the two components of individuals’ subjective well-being, positive and negative affect. Specifically, positive affect and negative affect are two independent dimensions representing the experience (feeling) of emotions. In this paper, we investigate the relationship between the daily diversity of social communication and dynamic affect states that people experience in their daily lives. We collected two high-resolution datasets that capture affect scores via daily experience sampling surveys and social interaction through wearable sensing technologies: sociometric badges for face-to-face interaction and smart phones for mobile phone calls. We found that communication diversity correlates with desirable affect states–e.g. an increase in the positive affect state or a decrease in the negative affect state–for some personality types, but correlates with undesirable affect states for others. For example, diversity in phone calls is experienced as good by introverts, but bad by extroverts; diversity in face-to-face interaction is experienced as good by people who tend to be positive by nature (trait) but bad for people who tend to be not positive by nature. More broadly, the moderating effect of personality type on the relationship between diversity and affect was detected without any knowledge of the type of social tie or the content of communication. This provides further support for the power of unobtrusive sensing in understanding social dynamics, and in measuring the effect of potential interventions designed to improve well-being. PMID:27035904

  8. Network Diversity and Affect Dynamics: The Role of Personality Traits.

    Science.gov (United States)

    Alshamsi, Aamena; Pianesi, Fabio; Lepri, Bruno; Pentland, Alex; Rahwan, Iyad

    2016-01-01

    People divide their time unequally among their social contacts due to time constraints and varying strength of relationships. It was found that high diversity of social communication, dividing time more evenly among social contacts, is correlated with economic well-being both at macro and micro levels. Besides economic well-being, it is not clear how the diversity of social communication is also associated with the two components of individuals' subjective well-being, positive and negative affect. Specifically, positive affect and negative affect are two independent dimensions representing the experience (feeling) of emotions. In this paper, we investigate the relationship between the daily diversity of social communication and dynamic affect states that people experience in their daily lives. We collected two high-resolution datasets that capture affect scores via daily experience sampling surveys and social interaction through wearable sensing technologies: sociometric badges for face-to-face interaction and smart phones for mobile phone calls. We found that communication diversity correlates with desirable affect states--e.g. an increase in the positive affect state or a decrease in the negative affect state--for some personality types, but correlates with undesirable affect states for others. For example, diversity in phone calls is experienced as good by introverts, but bad by extroverts; diversity in face-to-face interaction is experienced as good by people who tend to be positive by nature (trait) but bad for people who tend to be not positive by nature. More broadly, the moderating effect of personality type on the relationship between diversity and affect was detected without any knowledge of the type of social tie or the content of communication. This provides further support for the power of unobtrusive sensing in understanding social dynamics, and in measuring the effect of potential interventions designed to improve well-being.

  9. Network Diversity and Affect Dynamics: The Role of Personality Traits.

    Directory of Open Access Journals (Sweden)

    Aamena Alshamsi

    Full Text Available People divide their time unequally among their social contacts due to time constraints and varying strength of relationships. It was found that high diversity of social communication, dividing time more evenly among social contacts, is correlated with economic well-being both at macro and micro levels. Besides economic well-being, it is not clear how the diversity of social communication is also associated with the two components of individuals' subjective well-being, positive and negative affect. Specifically, positive affect and negative affect are two independent dimensions representing the experience (feeling of emotions. In this paper, we investigate the relationship between the daily diversity of social communication and dynamic affect states that people experience in their daily lives. We collected two high-resolution datasets that capture affect scores via daily experience sampling surveys and social interaction through wearable sensing technologies: sociometric badges for face-to-face interaction and smart phones for mobile phone calls. We found that communication diversity correlates with desirable affect states--e.g. an increase in the positive affect state or a decrease in the negative affect state--for some personality types, but correlates with undesirable affect states for others. For example, diversity in phone calls is experienced as good by introverts, but bad by extroverts; diversity in face-to-face interaction is experienced as good by people who tend to be positive by nature (trait but bad for people who tend to be not positive by nature. More broadly, the moderating effect of personality type on the relationship between diversity and affect was detected without any knowledge of the type of social tie or the content of communication. This provides further support for the power of unobtrusive sensing in understanding social dynamics, and in measuring the effect of potential interventions designed to improve well-being.

  10. The Microtubule Regulatory Protein Stathmin Is Required to Maintain the Integrity of Axonal Microtubules in Drosophila.

    Directory of Open Access Journals (Sweden)

    Jason E Duncan

    Full Text Available Axonal transport, a form of long-distance, bi-directional intracellular transport that occurs between the cell body and synaptic terminal, is critical in maintaining the function and viability of neurons. We have identified a requirement for the stathmin (stai gene in the maintenance of axonal microtubules and regulation of axonal transport in Drosophila. The stai gene encodes a cytosolic phosphoprotein that regulates microtubule dynamics by partitioning tubulin dimers between pools of soluble tubulin and polymerized microtubules, and by directly binding to microtubules and promoting depolymerization. Analysis of stai function in Drosophila, which has a single stai gene, circumvents potential complications with studies performed in vertebrate systems in which mutant phenotypes may be compensated by genetic redundancy of other members of the stai gene family. This has allowed us to identify an essential function for stai in the maintenance of the integrity of axonal microtubules. In addition to the severe disruption in the abundance and architecture of microtubules in the axons of stai mutant Drosophila, we also observe additional neurological phenotypes associated with loss of stai function including a posterior paralysis and tail-flip phenotype in third instar larvae, aberrant accumulation of transported membranous organelles in stai deficient axons, a progressive bang-sensitive response to mechanical stimulation reminiscent of the class of Drosophila mutants used to model human epileptic seizures, and a reduced adult lifespan. Reductions in the levels of Kinesin-1, the primary anterograde motor in axonal transport, enhance these phenotypes. Collectively, our results indicate that stai has an important role in neuronal function, likely through the maintenance of microtubule integrity in the axons of nerves of the peripheral nervous system necessary to support and sustain long-distance axonal transport.

  11. Acentrosomal Microtubule Assembly in Mitosis: The Where, When, and How.

    Science.gov (United States)

    Meunier, Sylvain; Vernos, Isabelle

    2016-02-01

    In mitosis the cell assembles the bipolar spindle, a microtubule (MT)-based apparatus that segregates the duplicated chromosomes into two daughter cells. Most animal cells enter mitosis with duplicated centrosomes that provide an active source of dynamic MTs. However, it is now established that spindle assembly relies on the nucleation of acentrosomal MTs occurring around the chromosomes after nuclear envelope breakdown, and on pre-existing microtubules. Where chromosome-dependent MT nucleation occurs, when MT amplification takes place and how the two pathways function are still key questions that generate some controversies. We reconcile the data and present an integrated model accounting for acentrosomal microtubule assembly in the dividing cell.

  12. Sliding of microtubules by a team of dynein motors: Understanding the effect of spatial distribution of motor tails and mutual exclusion of motor heads on microtubules

    Science.gov (United States)

    Singh, Hanumant Pratap; Takshak, Anjneya; Mall, Utkarsh; Kunwar, Ambarish

    2016-06-01

    Molecular motors are natural nanomachines that use the free energy released from ATP hydrolysis to generate mechanical forces. Cytoplasmic dynein motors often work collectively as a team to drive important processes such as axonal growth, proplatelet formation and mitosis, as forces generated by single motors are insufficient. A large team of dynein motors is used to slide cytoskeletal microtubules with respect to one another during the process of proplatelet formation and axonal growth. These motors attach to a cargo microtubule via their tail domains, undergo the process of detachment and reattachment of their head domains on another track microtubule, while sliding the cargo microtubule along the track. Traditional continuum/mean-field approaches used in the past are not ideal for studying the sliding mechanism of microtubules, as they ignore spatial and temporal fluctuations due to different possible distributions of motor tails on cargo filament, as well as binding/unbinding of motors from their track. Therefore, these models cannot be used to address important questions such as how the distribution of motor tails on microtubules, or how the mutual exclusion of motor heads on microtubule tracks affects the sliding velocity of cargo microtubule. To answer these, here we use a computational stochastic model where we model each dynein motor explicitly. In our model, we use both random as well as uniform distributions of dynein motors on cargo microtubule, as well as mutual exclusion of motors on microtubule tracks. We find that sliding velocities are least affected by the distribution of motor tails on microtubules, whereas they are greatly affected by mutual exclusion of motor heads on microtubule tracks. We also find that sliding velocity depends on the length of cargo microtubule if mutual exclusion among motor heads is considered.

  13. Centrosome and microtubule instability in aging Drosophila cells

    Science.gov (United States)

    Schatten, H.; Chakrabarti, A.; Hedrick, J.

    1999-01-01

    Several cytoskeletal changes are associated with aging which includes alterations in muscle structure leading to muscular atrophy, and weakening of the microtubule network which affects cellular secretion and maintenance of cell shape. Weakening of the microtubule network during meiosis in aging oocytes can result in aneuploidy or trisomic zygotes with increasing maternal age. Imbalances of cytoskeletal organization can lead to disease such as Alzheimer's, muscular disorders, and cancer. Because many cytoskeletal diseases are related to age we investigated the effects of aging on microtubule organization in cell cultures of the Drosophila cell model system (Schneider S-1 and Kc23 cell lines). This cell model is increasingly being used as an alternative system to mammalian cell cultures. Drosophila cells are amenable to genetic manipulations and can be used to identify and manipulate genes which are involved in the aging processes. Immunofluorescence, scanning, and transmission electron microscopy were employed for the analysis of microtubule organizing centers (centrosomes) and microtubules at various times after subculturing cells in fresh medium. Our results reveal that centrosomes and the microtubule network becomes significantly affected in aging cells after 5 days of subculture. At 5-14 days of subculture, 1% abnormal out of 3% mitoses were noted which were clearly distinguishable from freshly subcultured control cells in which 3% of cells undergo normal mitosis with bipolar configurations. Microtubules are also affected in the midbody during cell division. The midbody in aging cells becomes up to 10 times longer when compared with midbodies in freshly subcultured cells. During interphase, microtubules are often disrupted and disorganized, which may indicate improper function related to transport of cell organelles along microtubules. These results are likely to help explain some cytoskeletal disorders and diseases related to aging.

  14. Studying neuronal microtubule organization and microtubule-associated proteins using single molecule localization microscopy

    NARCIS (Netherlands)

    Chazeau, Anaël; Katrukha, Eugene A; Hoogenraad, Casper C; Kapitein, Lukas C

    2016-01-01

    The formation and maintenance of highly polarized neurons critically depends on the proper organization of the microtubule (MT) cytoskeleton. In axons, MTs are uniformly oriented with their plus-end pointing outward whereas in mature dendrites MTs have mixed orientations. MT organization and dynamic

  15. Interactive affective sharing versus non-interactive affective sharing in work groups : Comparative effects of group affect on work group performance and dynamics

    NARCIS (Netherlands)

    Klep, Annefloor; Wisse, Barbara; Van Der Flier, Henk

    2011-01-01

    This study explores whether the dynamic path to group affect, which is characterized by interactive affective sharing processes, yields different effects on task performance and group dynamics than the static path to group affect, which arises from non-interactive affective sharing. The results of o

  16. The microtubule plus-end-tracking protein CLIP-170 associates with the spermatid manchette and is essential for spermatogenesis.

    NARCIS (Netherlands)

    A.S. Akhmanova (Anna); A.L. Mausset-Bonnefont (Anne-Laure); W.A. van Cappellen (Gert); N. Keijzer (Nanda); C.C. Hoogenraad (Casper); T. Stepanova (Tatiana); K. Drabek (Ksenija); J. van der Wees (Jacqueline); M. Mommaas (Mieke); J. Onderwater (Jos); H. van der Meulen (Hans); M.E. Tanenbaum (Marvin); R.H. Medema (Rene); J.W. Hoogerbrugge (Jos); J.T.M. Vreeburg (Jan); E.J. Uringa; J.A. Grootegoed (Anton); F.G. Grosveld (Frank); N.J. Galjart (Niels)

    2005-01-01

    textabstractCLIP-170 is a microtubule "plus-end-tracking protein" implicated in the control of microtubule dynamics, dynactin localization, and the linking of endosomes to microtubules. To investigate the function of mouse CLIP-170, we generated CLIP-170 knockout and GFP-CLIP-170 knock-in alleles. R

  17. The microtubule plus-end-tracking protein CLIP-170 associates with the spermatid manchette and is essential for spermatogenesis

    NARCIS (Netherlands)

    Akhmanova, A.S.; Mausset-Bonnefont, A.-L.; Cappellen, W. van; Keijzer, N.; Hoogenraad, C.C.; Stepanova, T.; Drabek, K.; Wees, J. van der; Mommaas, M.; Onderwater, J.; Meulen, H. van der; Tanenbaum, M.E.; Medema, R.H.; Hoogerbrugge, J.; Vreeburg, J.; Uringa, E.-J.; Grootegoed, J.A.; Grosveld, F.; Galjart, N.

    2005-01-01

    CLIP-170 is a microtubule "plus-end-tracking protein" implicated in the control of microtubule dynamics, dynactin localization, and the linking of endosomes to microtubules. To investigate the function of mouse CLIP-170, we generated CLIP-170 knockout and GFP-CLIP-170 knock-in alleles. Residual CLIP

  18. Electrostatically biased binding of kinesin to microtubules.

    Directory of Open Access Journals (Sweden)

    Barry J Grant

    2011-11-01

    Full Text Available The minimum motor domain of kinesin-1 is a single head. Recent evidence suggests that such minimal motor domains generate force by a biased binding mechanism, in which they preferentially select binding sites on the microtubule that lie ahead in the progress direction of the motor. A specific molecular mechanism for biased binding has, however, so far been lacking. Here we use atomistic Brownian dynamics simulations combined with experimental mutagenesis to show that incoming kinesin heads undergo electrostatically guided diffusion-to-capture by microtubules, and that this produces directionally biased binding. Kinesin-1 heads are initially rotated by the electrostatic field so that their tubulin-binding sites face inwards, and then steered towards a plus-endwards binding site. In tethered kinesin dimers, this bias is amplified. A 3-residue sequence (RAK in kinesin helix alpha-6 is predicted to be important for electrostatic guidance. Real-world mutagenesis of this sequence powerfully influences kinesin-driven microtubule sliding, with one mutant producing a 5-fold acceleration over wild type. We conclude that electrostatic interactions play an important role in the kinesin stepping mechanism, by biasing the diffusional association of kinesin with microtubules.

  19. Globally visualizing the microtubule-dependent transport behaviors of influenza virus in live cells.

    Science.gov (United States)

    Liu, Shu-Lin; Zhang, Li-Juan; Wang, Zhi-Gang; Zhang, Zhi-Ling; Wu, Qiu-Mei; Sun, En-Ze; Shi, Yun-Bo; Pang, Dai-Wen

    2014-04-15

    Understanding the microtubule-dependent behaviors of viruses in live cells is very meaningful for revealing the mechanisms of virus infection and endocytosis. Herein, we used a quantum dots-based single-particle tracking technique to dynamically and globally visualize the microtubule-dependent transport behaviors of influenza virus in live cells. We found that the intersection configuration of microtubules can interfere with the transport behaviors of the virus in live cells, which lead to the changing and long-time pausing of the transport behavior of viruses. Our results revealed that most of the viruses moved along straight microtubules rapidly and unidirectionally from the cell periphery to the microtubule organizing center (MTOC) near the bottom of the cell, and the viruses were confined in the grid of microtubules near the top of the cell and at the MTOC near the bottom of the cell. These results provided deep insights into the influence of entire microtubule geometry on the virus infection.

  20. Tubulin bond energies and microtubule biomechanics determined from nanoindentation in silico

    CERN Document Server

    Kononova, Olga; Theisen, Kelly E; Marx, Kenneth A; Dima, Ruxandra I; Ataullakhanov, Fazly I; Grishchuk, Ekaterina L; Barsegov, Valeri

    2015-01-01

    Microtubules, the primary components of the chromosome segregation machinery, are stabilized by longitudinal and lateral non-covalent bonds between the tubulin subunits. However, the thermodynamics of these bonds and the microtubule physico-chemical properties are poorly understood. Here, we explore the biomechanics of microtubule polymers using multiscale computational modeling and nanoindentations in silico of a contiguous microtubule fragment. A close match between the simulated and experimental force-deformation spectra enabled us to correlate the microtubule biomechanics with dynamic structural transitions at the nanoscale. Our mechanical testing revealed that the compressed MT behaves as a system of rigid elements interconnected through a network of lateral and longitudinal elastic bonds. The initial regime of continuous elastic deformation of the microtubule is followed by the transition regime, during which the microtubule lattice undergoes discrete structural changes, which include first the reversib...

  1. A Chimeric Cetuximab-Functionalized Corona as a Potent Delivery System for Microtubule-Destabilizing Nanocomplexes to Hepatocellular Carcinoma Cells: A Focus on EGFR and Tubulin Intracellular Dynamics.

    Science.gov (United States)

    Poojari, Radhika; Kini, Sudarshan; Srivastava, Rohit; Panda, Dulal

    2015-11-01

    In this study, we have developed microtubule destabilizing agents combretastatin A4 (CA4) or 2-methoxyestradiol (2ME) encapsulated poly(d,l-lactide-co-glycolide)-b-poly(ethylene glycol) (PLGA-b-PEG) nanocomplexes for targeted delivery to human hepatocellular carcinoma (HCC) cells. An epidermal growth factor receptor (EGFR) is known to be overexpressed in HCC cells. Therefore, the targeting moiety cetuximab (Cet), an anti-EGFR chimeric monoclonal antibody, is functionalized on the surface of these diblock copolymeric coronas. Cetuximab is associated with the extracellular domain of the EGFR; therefore, the uptake of the cetuximab conjugated nanocomplexes occurred efficiently in EGFR overexpressing HCC cells indicating potent internalization of the complex. The cetuximab targeted-PLGA-b-PEG nanocomplexes encapsulating CA4 or 2ME strongly inhibited phospho-EGFR expression, depolymerized microtubules, produced spindle abnormalities, stalled mitosis, and induced apoptosis in Huh7 cells compared to the free drugs, CA4 or 2ME. Further, the combinatorial strategy of targeted nanocomplexes, Cet-PLGA-b-PEG-CA4 NP and Cet-PLGA-b-PEG-2ME NP, significantly reduced the migration of Huh7 cells, and markedly enhanced the anticancer effects of the microtubule-targeted drugs in Huh7 cells compared to the free drugs, CA4 or 2ME. The results indicated that EGFR receptor-mediated internalization via cetuximab facilitated enhanced uptake of the nanocomplexes leading to potent anticancer efficacy in Huh7 cells. Cetuximab-functionalized PLGA-b-PEG nanocomplexes possess a strong potential for the targeted delivery of CA4 or 2ME in EGFR overexpressed HCC cells, and the strategy may be useful for selectively targeting microtubules in these cells.

  2. Sulfo-SMCC Prevents Annealing of Taxol-Stabilized Microtubules In Vitro

    CERN Document Server

    Prabhune, Meenakshi; Schmidt, Christoph F

    2015-01-01

    Microtubule structure and functions have been widely studied in vitro and in cells. Research has shown that cysteines on tubulin play a crucial role in the polymerization of microtubules. Here, we show that blocking sulfhydryl groups of cysteines in taxol-stabilized polymerized microtubules with a commonly used chemical crosslinker prevents temporal end-to-end annealing of microtubules in vitro. This can dramatically affect the length distribution of the microtubules. The crosslinker sulfosuccinimidyl 4-(N-maleimidomethyl)cyclohexane-1-carboxylate, sulfo-SMCC, consists of a maleimide and a N-hydroxysuccinimide ester group to bind to sulfhydryl groups and primary amines, respectively. Interestingly, addition of a maleimide dye alone does not show the same prevention of annealing in stabilized microtubules. This study shows that the sulfhydryl groups of cysteines of tubulin that are vital for the polymerization are also important for the subsequent annealing of microtubules.

  3. A thermodynamic model of microtubule assembly and disassembly.

    Directory of Open Access Journals (Sweden)

    Bernard M A G Piette

    Full Text Available Microtubules are self-assembling polymers whose dynamics are essential for the normal function of cellular processes including chromosome separation and cytokinesis. Therefore understanding what factors effect microtubule growth is fundamental to our understanding of the control of microtubule based processes. An important factor that determines the status of a microtubule, whether it is growing or shrinking, is the length of the GTP tubulin microtubule cap. Here, we derive a Monte Carlo model of the assembly and disassembly of microtubules. We use thermodynamic laws to reduce the number of parameters of our model and, in particular, we take into account the contribution of water to the entropy of the system. We fit all parameters of the model from published experimental data using the GTP tubulin dimer attachment rate and the lateral and longitudinal binding energies of GTP and GDP tubulin dimers at both ends. Also we calculate and incorporate the GTP hydrolysis rate. We have applied our model and can mimic published experimental data, which formerly suggested a single layer GTP tubulin dimer microtubule cap, to show that these data demonstrate that the GTP cap can fluctuate and can be several microns long.

  4. A thermodynamic model of microtubule assembly and disassembly.

    Science.gov (United States)

    Piette, Bernard M A G; Liu, Junli; Peeters, Kasper; Smertenko, Andrei; Hawkins, Timothy; Deeks, Michael; Quinlan, Roy; Zakrzewski, Wojciech J; Hussey, Patrick J

    2009-08-11

    Microtubules are self-assembling polymers whose dynamics are essential for the normal function of cellular processes including chromosome separation and cytokinesis. Therefore understanding what factors effect microtubule growth is fundamental to our understanding of the control of microtubule based processes. An important factor that determines the status of a microtubule, whether it is growing or shrinking, is the length of the GTP tubulin microtubule cap. Here, we derive a Monte Carlo model of the assembly and disassembly of microtubules. We use thermodynamic laws to reduce the number of parameters of our model and, in particular, we take into account the contribution of water to the entropy of the system. We fit all parameters of the model from published experimental data using the GTP tubulin dimer attachment rate and the lateral and longitudinal binding energies of GTP and GDP tubulin dimers at both ends. Also we calculate and incorporate the GTP hydrolysis rate. We have applied our model and can mimic published experimental data, which formerly suggested a single layer GTP tubulin dimer microtubule cap, to show that these data demonstrate that the GTP cap can fluctuate and can be several microns long.

  5. Alendronate affects calcium dynamics in cardiomyocytes in vitro.

    Science.gov (United States)

    Kemeny-Suss, Naomi; Kasneci, Amanda; Rivas, Daniel; Afilalo, Jonathan; Komarova, Svetlana V; Chalifour, Lorraine E; Duque, Gustavo

    2009-01-01

    Therapy with bisphosphonates, including alendronate (ALN), is considered a safe and effective treatment for osteoporosis. However, recent studies have reported an unexpected increase in serious atrial fibrillation (AF) in patients treated with bisphosphonates. The mechanism that explains this side effect remains unknown. Since AF is associated with an altered sarcoendoplasmic reticulum calcium load, we studied how ALN affects cardiomyocyte calcium homeostasis and protein isoprenylation in vitro. Acute and long-term (48h) treatment of atrial and ventricular cardiomyocytes with ALN (10(-8)-10(-6)M) was performed. Changes in calcium dynamics were determined by both fluorescence measurement of cytosolic free Ca(2+) concentration and western blot analysis of calcium-regulating proteins. Finally, effect of ALN on protein farnesylation was also identified. In both atrial and ventricular cardiomyocytes, ALN treatment delayed and diminished calcium responses to caffeine. Only in atrial cells, long-term exposure to ALN-induced transitory calcium oscillations and led to the development of oscillatory component in calcium responses to caffeine. Changes in calcium dynamics were accompanied by changes in expression of proteins controlling sarcoendoplasmic reticulum calcium. In contrast, ALN minimally affected protein isoprenylation in these cells. In summary, treatment of atrial cardiomyocytes with ALN-induced abnormalities in calcium dynamics consistent with induction of a self-stimulatory, pacemaker-like behavior, which may contribute to the development of cardiac side effects associated with these drugs.

  6. Analysis of Soybean Microtubule Persistence Length; New Evidence on the Correlation between Structural Composition and Mechanical Properties

    Science.gov (United States)

    Shojania Feizabadi, Mitra; Winton, Carly; Barrientos, Jimmy

    2012-02-01

    Recent studies on microtubules composed of different β tubulin isotypes indicate their different functionality in terms of their dynamical behavior or the mechanism of their interaction with chemotherapeutic drugs. Along these lines, the result of our recent study measuring the rigidity of neural and non-neural samples of microtubules with different β tubulin isotype compositions suggests that the distinguished mechanical properties of microtubules, such as rigidity, may also be associated with the different distribution of their β tubulin isotypes. In our current study, we have reported the persistence length of a single soybean microtubule. This plant microtubule has a structural composition different from that of mammalian microtubules. Under the same experimental methods of measurement, the soybean microtubules showed a different persistence length as compared to the value of the persistence length that we estimated in the study of both single Bovine Brain and MCF7 microtubules.

  7. Insights from an erroneous kinetochore-microtubule attachment state.

    Science.gov (United States)

    Cane, Stuart; McGilvray, Philip T; Maresca, Thomas J

    2013-01-01

    Faithful distribution of the genome requires that sister kinetochores, which assemble on each chromatid during cell division, interact with dynamic microtubules from opposite spindle poles in a configuration called chromosome biorientation. Biorientation produces tension that increases the affinity of kinetochores for microtubules via ill-defined mechanisms. Non-bioriented kinetochore-microtubule (kt-MT) interactions are prevalent but short-lived due to an error correction pathway that reduces the affinity of kinetochores for microtubules. Interestingly, incorrect kt-MT interactions can be stabilized by experimentally applying force to misoriented chromosomes. Here, a live-cell force assay is utilized to characterize the molecular composition of a common type of improper kt-MT attachment. Our force-related studies are also discussed in the context of current models for tension-dependent stabilization of kt-MT interactions.

  8. Mechanical Models of Microtubule Bundle Collapse in Alzheimer's Disease

    Science.gov (United States)

    Sendek, Austin; Singh, Rajiv; Cox, Daniel

    2013-03-01

    Amyloid-beta aggregates initiate Alzheimer's disease, and downstream trigger degradation of tau proteins that act as microtubule bundle stabilizers and mechanical spacers. Currently it is unclear which of tau cutting by proteases, tau phosphorylation, or tau aggregation are responsible for cytoskeleton degradation., We construct a percolation simulation of the microtubule bundle using a molecular spring model for the taus and including depletion force attraction between microtubules and membrane/actin cytoskeletal surface tension. The simulation uses a fictive molecular dynamics to model the motion of the individual microtubules within the bundle as a result of random tau removal, and calculates the elastic modulus of the bundle as the tau concentration falls. We link the tau removal steps to kinetic tau steps in various models of tau degradation. Supported by US NSF Grant DMR 1207624

  9. Mechanical modulation of cardiac microtubules.

    Science.gov (United States)

    White, Ed

    2011-07-01

    Microtubules are a major component of the cardiac myocyte cytoskeleton. Interventions that alter it may influence cardiac mechanical and electrical activity by disrupting the trafficking of proteins to and from the surface membrane by molecular motors such as dynein, which use microtubules as tracks to step along. Free tubulin dimers may transfer GTP to the α-subunits of G-proteins, thus an increase in free tubulin could increase the activity of G-proteins; evidence for and against such a role exists. There is more general agreement that microtubules act as compression-resisting structures within myocytes, influencing visco-elasticity of myocytes and increasing resistance to shortening when proliferated and resisting deformation from longitudinal shear stress. In response to pressure overload, there can be post-translational modifications resulting in more stable microtubules and an increase in microtubule density. This is accompanied by contractile dysfunction of myocytes which can be reversed by microtubule disruption. There are reports of mechanically induced changes in electrical activity that are dependent upon microtubules, but at present, a consensus is lacking on whether disruption or proliferation would be beneficial in the prevention of arrhythmias. Microtubules certainly play a role in the response of cardiac myocytes to mechanical stimulation, the exact nature and significance of this role is still to be fully determined.

  10. Stabilizing versus destabilizing the microtubules: a double-edge sword for an effective cancer treatment option?

    Science.gov (United States)

    Fanale, Daniele; Bronte, Giuseppe; Passiglia, Francesco; Calò, Valentina; Castiglia, Marta; Di Piazza, Florinda; Barraco, Nadia; Cangemi, Antonina; Catarella, Maria Teresa; Insalaco, Lavinia; Listì, Angela; Maragliano, Rossella; Massihnia, Daniela; Perez, Alessandro; Toia, Francesca; Cicero, Giuseppe; Bazan, Viviana

    2015-01-01

    Microtubules are dynamic and structural cellular components involved in several cell functions, including cell shape, motility, and intracellular trafficking. In proliferating cells, they are essential components in the division process through the formation of the mitotic spindle. As a result of these functions, tubulin and microtubules are targets for anticancer agents. Microtubule-targeting agents can be divided into two groups: microtubule-stabilizing, and microtubule-destabilizing agents. The former bind to the tubulin polymer and stabilize microtubules, while the latter bind to the tubulin dimers and destabilize microtubules. Alteration of tubulin-microtubule equilibrium determines the disruption of the mitotic spindle, halting the cell cycle at the metaphase-anaphase transition and, eventually, resulting in cell death. Clinical application of earlier microtubule inhibitors, however, unfortunately showed several limits, such as neurological and bone marrow toxicity and the emergence of drug-resistant tumor cells. Here we review several natural and synthetic microtubule-targeting agents, which showed antitumor activity and increased efficacy in comparison to traditional drugs in various preclinical and clinical studies. Cryptophycins, combretastatins, ombrabulin, soblidotin, D-24851, epothilones and discodermolide were used in clinical trials. Some of them showed antiangiogenic and antivascular activity and others showed the ability to overcome multidrug resistance, supporting their possible use in chemotherapy.

  11. Stabilizing versus Destabilizing the Microtubules: A Double-Edge Sword for an Effective Cancer Treatment Option?

    Science.gov (United States)

    Fanale, Daniele; Bronte, Giuseppe; Passiglia, Francesco; Calò, Valentina; Castiglia, Marta; Di Piazza, Florinda; Barraco, Nadia; Cangemi, Antonina; Catarella, Maria Teresa; Insalaco, Lavinia; Listì, Angela; Maragliano, Rossella; Massihnia, Daniela; Perez, Alessandro; Toia, Francesca; Cicero, Giuseppe; Bazan, Viviana

    2015-01-01

    Microtubules are dynamic and structural cellular components involved in several cell functions, including cell shape, motility, and intracellular trafficking. In proliferating cells, they are essential components in the division process through the formation of the mitotic spindle. As a result of these functions, tubulin and microtubules are targets for anticancer agents. Microtubule-targeting agents can be divided into two groups: microtubule-stabilizing, and microtubule-destabilizing agents. The former bind to the tubulin polymer and stabilize microtubules, while the latter bind to the tubulin dimers and destabilize microtubules. Alteration of tubulin-microtubule equilibrium determines the disruption of the mitotic spindle, halting the cell cycle at the metaphase-anaphase transition and, eventually, resulting in cell death. Clinical application of earlier microtubule inhibitors, however, unfortunately showed several limits, such as neurological and bone marrow toxicity and the emergence of drug-resistant tumor cells. Here we review several natural and synthetic microtubule-targeting agents, which showed antitumor activity and increased efficacy in comparison to traditional drugs in various preclinical and clinical studies. Cryptophycins, combretastatins, ombrabulin, soblidotin, D-24851, epothilones and discodermolide were used in clinical trials. Some of them showed antiangiogenic and antivascular activity and others showed the ability to overcome multidrug resistance, supporting their possible use in chemotherapy. PMID:26484003

  12. Stabilizing versus Destabilizing the Microtubules: A Double-Edge Sword for an Effective Cancer Treatment Option?

    Directory of Open Access Journals (Sweden)

    Daniele Fanale

    2015-01-01

    Full Text Available Microtubules are dynamic and structural cellular components involved in several cell functions, including cell shape, motility, and intracellular trafficking. In proliferating cells, they are essential components in the division process through the formation of the mitotic spindle. As a result of these functions, tubulin and microtubules are targets for anticancer agents. Microtubule-targeting agents can be divided into two groups: microtubule-stabilizing, and microtubule-destabilizing agents. The former bind to the tubulin polymer and stabilize microtubules, while the latter bind to the tubulin dimers and destabilize microtubules. Alteration of tubulin-microtubule equilibrium determines the disruption of the mitotic spindle, halting the cell cycle at the metaphase-anaphase transition and, eventually, resulting in cell death. Clinical application of earlier microtubule inhibitors, however, unfortunately showed several limits, such as neurological and bone marrow toxicity and the emergence of drug-resistant tumor cells. Here we review several natural and synthetic microtubule-targeting agents, which showed antitumor activity and increased efficacy in comparison to traditional drugs in various preclinical and clinical studies. Cryptophycins, combretastatins, ombrabulin, soblidotin, D-24851, epothilones and discodermolide were used in clinical trials. Some of them showed antiangiogenic and antivascular activity and others showed the ability to overcome multidrug resistance, supporting their possible use in chemotherapy.

  13. Xenopus oocyte wound healing as a model system for analysis of microtubule-actin interactions.

    Science.gov (United States)

    Zhang, Tong; Mandato, Craig A

    2007-01-01

    Microtubule-actin interactions are fundamental to many cellular processes such as cytokinesis and cellular locomotion. Investigating the mechanism of microtubule-actin interactions is the key to understand the cellular morphogenesis and related pathological processes. The abundance and highly dynamic nature of microtubules and F-actin raise a serious challenge when trying to distinguish between the real and fortuitous interactions within a cell. Xenopus oocyte wound model represents an ideal system to study microtubule-actin interactions as well as microtubule-dependent control of the actin polymerization. Here, we describe a series of cytoskeleton specific treatments in Xenopus oocyte wound healing experiments and use confocal fluorescence microscopy to analyze fixed oocytes to examine microtubule-actin interactions.

  14. Polyamine sharing between tubulin dimers favours microtubule nucleation and elongation via facilitated diffusion.

    Directory of Open Access Journals (Sweden)

    Alain Mechulam

    2009-01-01

    Full Text Available We suggest for the first time that the action of multivalent cations on microtubule dynamics can result from facilitated diffusion of GTP-tubulin to the microtubule ends. Facilitated diffusion can promote microtubule assembly, because, upon encountering a growing nucleus or the microtubule wall, random GTP-tubulin sliding on their surfaces will increase the probability of association to the target sites (nucleation sites or MT ends. This is an original explanation for understanding the apparent discrepancy between the high rate of microtubule elongation and the low rate of tubulin association at the microtubule ends in the viscous cytoplasm. The mechanism of facilitated diffusion requires an attraction force between two tubulins, which can result from the sharing of multivalent counterions. Natural polyamines (putrescine, spermidine, and spermine are present in all living cells and are potent agents to trigger tubulin self-attraction. By using an analytical model, we analyze the implication of facilitated diffusion mediated by polyamines on nucleation and elongation of microtubules. In vitro experiments using pure tubulin indicate that the promotion of microtubule assembly by polyamines is typical of facilitated diffusion. The results presented here show that polyamines can be of particular importance for the regulation of the microtubule network in vivo and provide the basis for further investigations into the effects of facilitated diffusion on cytoskeleton dynamics.

  15. The model of fungal population dynamics affected by nystatin

    Science.gov (United States)

    Voychuk, Sergei I.; Gromozova, Elena N.; Sadovskiy, Mikhail G.

    Fungal diseases are acute problems of the up-to-day medicine. Significant increase of resistance of microorganisms to the medically used antibiotics and a lack of new effective drugs follows in a growth of dosage of existing chemicals to solve the problem. Quite often such approach results in side effects on humans. Detailed study of fungi-antibiotic dynamics can identify new mechanisms and bring new ideas to overcome the microbial resistance with a lower dosage of antibiotics. In this study, the dynamics of the microbial population under antibiotic treatment was investigated. The effects of nystatin on the population of Saccharomyces cerevisiae yeasts were used as a model system. Nystatin effects were investigated both in liquid and solid media by viability tests. Dependence of nystatin action on osmotic gradient was evaluated in NaCl solutions. Influences of glucose and yeast extract were additionally analyzed. A "stepwise" pattern of the cell death caused by nystatin was the most intriguing. This pattern manifested in periodical changes of the stages of cell death against stages of resistance to the antibiotic. The mathematical model was proposed to describe cell-antibiotic interactions and nystatin viability effects in the liquid medium. The model implies that antibiotic ability to cause a cells death is significantly affected by the intracellular compounds, which came out of cells after their osmotic barriers were damaged

  16. Kinesin-1 Translocation along Human Breast Cancer Cell Microtubules in Vitro

    Science.gov (United States)

    Shojania Feizabadi, Mitra; Jun, Yonggun

    2015-03-01

    A principle approach to better understand intra-cellular microtubule based transport is to study such it in vitro. Such in vitro examinations have predominantly used microtubules polymerized from bovine brain tubulin, but motor function can also in principle be affected by the specific tubulin isotypes present in different cells. The human breast cancer cells carry different beta tubulin isotype distribution. However, it is entirely unknown whether transport along the microtubules is different in these cells. In this work we have characterized, for the first time, the translocation specifications of kinesin-1 along human breast cancer cell microtubules polymerized in vitro. We found that as compared with the translocation along bovine brain microtubules, kinesin-1 shows a fifty percent shorter processive run length and slightly slower velocity under similar experimental conditions. These first time results support the regulatory role of tubulin isotypes in regards to motor protein translocations, and quantify the translocation specifications of kinesin-1 along microtubules of human breast cancer cells.

  17. Termination of Protofilament Elongation by Eribulin Induces Lattice Defects that Promote Microtubule Catastrophes

    NARCIS (Netherlands)

    Doodhi, Harinath; Prota, Andrea E; Rodríguez-García, Ruddi; Xiao, Hui; Custar, Daniel W; Bargsten, Katja; Katrukha, Eugene A; Hilbert, Manuel; Hua, Shasha; Jiang, Kai; Grigoriev, Ilya; Yang, Chia-Ping H; Cox, David; Horwitz, Susan Band; Kapitein, Lukas C; Akhmanova, Anna; Steinmetz, Michel O

    2016-01-01

    Microtubules are dynamic polymers built of tubulin dimers that attach in a head-to-tail fashion to form protofilaments, which further associate laterally to form a tube. Asynchronous elongation of individual protofilaments can potentially lead to an altered microtubule-end structure that promotes su

  18. The octarepeat region of hamster PrP (PrP51-91) enhances the formation of microtubule and antagonize Cu~(2+)-induced microtubule-disrupting activity

    Institute of Scientific and Technical Information of China (English)

    Xiaoli Li; Chenfang Dong; Song Shi; Guirong Wang; Yuan Li; Xin Wang; Qi Shi; Chan Tian; Ruimin Zhou; Chen Gao; Xiaoping Dong

    2009-01-01

    Prion protein (PrP) is considered to associate with microtubule and its major component, tubulin. In the present study, octarepeat region of PrP (PrP51-91) was expressed in prokaryotic-expressing system. Using GST pull-down assay and co-immunoprecipitation, the mol-ecular interaction between PrP51-91 and tubulin was observed. Our data also demonstrated that PrP51-91 could efficiently stimulate microtubule assembly in vitro, indicating a potential effect of PrP on microtu-bule dynamics. Moreover, PrP51-91 was confirmed to be able to antagonize Cu~(2+)-induced microtubule-disrupt-ing activity in vivo, partially protecting against Cu~(2+) intoxication to culture cells and stabilize cellular micro-tubule structure. The association of the octarepeat region of PrP with tubulin may further provide insight into the biological function of PrP in the neurons.

  19. Characterization of affective states by pupillary dynamics and autonomic correlates.

    Science.gov (United States)

    Onorati, Francesco; Barbieri, Riccardo; Mauri, Maurizio; Russo, Vincenzo; Mainardi, Luca

    2013-01-01

    With the recent advent of new recording devices and an easier access to signal processing tools, researchers are increasingly exploring and studying the Pupil Dilation (PD) signal. Recently, numerous studies pointed out the relations between PD dynamics and psychophysiological states. Although it is well known that PD is controlled by the Autonomic Nervous System (ANS), and ANS responses are related to emotional events/stimuli, the relationship between emotional states and PD is still an open issue. The aim of this study is to define the statistical properties of the PD signal, to understand its relation with ANS correlates such as Heart Rate Variability (HRV) and respiration (RESP), and to explore if PD could provide information for the evaluation of the psychophysiological response of ANS to affective triggering events. ECG, RESP, and PD data from 13 normal subjects were recorded during a memory recall paradigm, and processed with spectral and cross-spectral analysis. Our results demonstrate that variability indices extracted from fast PD oscillations, not observable through standard cardiorespiratory identification in the frequency domain, would be able to discern psychophysiological responses elicited by basic emotional stimuli. A strong linear coupling was found between the variables, due to the influence of RESP on both PD and HRV within the High Frequency (HF) band, from 0.15 to 0.45 Hz. Most importantly, our results point at PD features as possible candidates for characterizing basic emotional stimuli.

  20. Myomegalin is necessary for the formation of centrosomal and Golgi-derived microtubules

    Directory of Open Access Journals (Sweden)

    Régine Roubin

    2012-12-01

    The generation of cellular microtubules is initiated at specific sites such as the centrosome and the Golgi apparatus that contain nucleation complexes rich in γ-tubulin. The microtubule growing plus-ends are stabilized by plus-end tracking proteins (+TIPs, mainly EB1 and associated proteins. Myomegalin was identified as a centrosome/Golgi protein associated with cyclic nucleotide phosphodiesterase. We show here that Myomegalin exists as several isoforms. We characterize two of them. One isoform, CM-MMG, harbors a conserved domain (CM1, recently described as a nucleation activator, and is related to a family of γ-tubulin binding proteins, which includes Drosophila centrosomin. It localizes at the centrosome and at the cis-Golgi in an AKAP450-dependent manner. It recruits γ-tubulin nucleating complexes and promotes microtubule nucleation. The second isoform, EB-MMG, is devoid of CM1 domain and has a unique N-terminus with potential EB1-binding sites. It localizes at the cis-Golgi and can localize to microtubule plus-ends. EB-MMG binds EB1 and affects its loading on microtubules and microtubule growth. Depletion of Myomegalin by small interfering RNA delays microtubule growth from the centrosome and Golgi apparatus, and decreases directional migration of RPE1 cells. In conclusion, the Myomegalin gene encodes different isoforms that regulate microtubules. At least two of these have different roles, demonstrating a previously unknown mechanism to control microtubules in vertebrate cells.

  1. Inhibition of kinesin-5 improves regeneration of injured axons by a novel microtubule-based mechanism

    Institute of Scientific and Technical Information of China (English)

    Peter W. Baas; Andrew J. Matamoros

    2015-01-01

    Microtubules have been identiifed as a powerful target for augmenting regeneration of injured adult axons in the central nervous system. Drugs that stabilize microtubules have shown some promise, but there are concerns that abnormally stabilizing microtubules may have only limited beneifts for regeneration, while at the same time may be detrimental to the normal work that microtubules perform for the axon. Kinesin-5 (also called kif11 or Eg5), a molecular motor protein best known for its crucial role in mitosis, acts as a brake on microtubule movements by other motor proteins in the axon. Drugs that inhibit kinesin-5, originally developed to treat cancer, result in greater mobility of microtubules in the axon and an overall shift in the forces on the microtubule array. As a result, the axon grows faster, retracts less, and more readily enters environments that are inhibitory to axonal regeneration. Thus, drugs that inhibit kinesin-5 offer a novel microtubule-based means to boost axonal regeneration without the concerns that ac-company abnormal stabilization of the microtubule array. Even so, inhibiting kinesin-5 is not without its own caveats, such as potential problems with navigation of the regenerating axon to its target, as well as morphological effects on dendrites that could affect learning and memory if the drugs reach the brain.

  2. Inhibition of kinesin-5 improves regeneration of injured axons by a novel microtubule-based mechanism

    Directory of Open Access Journals (Sweden)

    Peter W Baas

    2015-01-01

    Full Text Available Microtubules have been identified as a powerful target for augmenting regeneration of injured adult axons in the central nervous system. Drugs that stabilize microtubules have shown some promise, but there are concerns that abnormally stabilizing microtubules may have only limited benefits for regeneration, while at the same time may be detrimental to the normal work that microtubules perform for the axon. Kinesin-5 (also called kif11 or Eg5, a molecular motor protein best known for its crucial role in mitosis, acts as a brake on microtubule movements by other motor proteins in the axon. Drugs that inhibit kinesin-5, originally developed to treat cancer, result in greater mobility of microtubules in the axon and an overall shift in the forces on the microtubule array. As a result, the axon grows faster, retracts less, and more readily enters environments that are inhibitory to axonal regeneration. Thus, drugs that inhibit kinesin-5 offer a novel microtubule-based means to boost axonal regeneration without the concerns that accompany abnormal stabilization of the microtubule array. Even so, inhibiting kinesin-5 is not without its own caveats, such as potential problems with navigation of the regenerating axon to its target, as well as morphological effects on dendrites that could affect learning and memory if the drugs reach the brain.

  3. Kinetochore-microtubule attachment is sufficient to satisfy the human spindle assembly checkpoint.

    Science.gov (United States)

    Etemad, Banafsheh; Kuijt, Timo E F; Kops, Geert J P L

    2015-12-01

    The spindle assembly checkpoint (SAC) is a genome surveillance mechanism that protects against aneuploidization. Despite profound progress on understanding mechanisms of its activation, it remains unknown what aspect of chromosome-spindle interactions is monitored by the SAC: kinetochore-microtubule attachment or the force generated by dynamic microtubules that signals stable biorientation of chromosomes? To answer this, we uncoupled these two processes by expressing a non-phosphorylatable version of the main microtubule-binding protein at kinetochores (HEC1-9A), causing stabilization of incorrect kinetochore-microtubule attachments despite persistent activity of the error-correction machinery. The SAC is fully functional in HEC1-9A-expressing cells, yet cells in which chromosomes cannot biorient but are stably attached to microtubules satisfy the SAC and exit mitosis. SAC satisfaction requires neither intra-kinetochore stretching nor dynamic microtubules. Our findings support the hypothesis that in human cells the end-on interactions of microtubules with kinetochores are sufficient to satisfy the SAC without the need for microtubule-based pulling forces.

  4. Termination of Protofilament Elongation by Eribulin Induces Lattice Defects that Promote Microtubule Catastrophes.

    Science.gov (United States)

    Doodhi, Harinath; Prota, Andrea E; Rodríguez-García, Ruddi; Xiao, Hui; Custar, Daniel W; Bargsten, Katja; Katrukha, Eugene A; Hilbert, Manuel; Hua, Shasha; Jiang, Kai; Grigoriev, Ilya; Yang, Chia-Ping H; Cox, David; Horwitz, Susan Band; Kapitein, Lukas C; Akhmanova, Anna; Steinmetz, Michel O

    2016-07-11

    Microtubules are dynamic polymers built of tubulin dimers that attach in a head-to-tail fashion to form protofilaments, which further associate laterally to form a tube. Asynchronous elongation of individual protofilaments can potentially lead to an altered microtubule-end structure that promotes sudden depolymerization, termed catastrophe [1-4]. However, how the dynamics of individual protofilaments relates to overall growth persistence has remained unclear. Here, we used the microtubule targeting anti-cancer drug Eribulin [5-7] to explore the consequences of stalled protofilament elongation on microtubule growth. Using X-ray crystallography, we first revealed that Eribulin binds to a site on β-tubulin that is required for protofilament plus-end elongation. Based on the structural information, we engineered a fluorescent Eribulin molecule. We demonstrate that single Eribulin molecules specifically interact with microtubule plus ends and are sufficient to either trigger a catastrophe or induce slow and erratic microtubule growth in the presence of EB3. Interestingly, we found that Eribulin increases the frequency of EB3 comet "splitting," transient events where a slow and erratically progressing comet is followed by a faster comet. This observation possibly reflects the "healing" of a microtubule lattice. Because EB3 comet splitting was also observed in control microtubules in the absence of any drugs, we propose that Eribulin amplifies a natural pathway toward catastrophe by promoting the arrest of protofilament elongation.

  5. Dynamics of Affective Experience and Behavior in Depressed Adolescents

    Science.gov (United States)

    Sheeber, Lisa B.; Allen, Nicholas B.; Leve, Craig; Davis, Betsy; Shortt, Joann Wu; Katz, Lynn Fainsilber

    2009-01-01

    Background: Depression is often characterized as a disorder of affect regulation. However, research focused on delineating the key dimensions of affective experience (other than valence) that are abnormal in depressive disorder has been scarce, especially in child and adolescent samples. As definitions of affect regulation center around processes…

  6. A Hierarchical Latent Stochastic Differential Equation Model for Affective Dynamics

    Science.gov (United States)

    Oravecz, Zita; Tuerlinckx, Francis; Vandekerckhove, Joachim

    2011-01-01

    In this article a continuous-time stochastic model (the Ornstein-Uhlenbeck process) is presented to model the perpetually altering states of the core affect, which is a 2-dimensional concept underlying all our affective experiences. The process model that we propose can account for the temporal changes in core affect on the latent level. The key…

  7. Microtubule affinity-regulating kinase 4: structure, function, and regulation.

    Science.gov (United States)

    Naz, Farha; Anjum, Farah; Islam, Asimul; Ahmad, Faizan; Hassan, Md Imtaiyaz

    2013-11-01

    MAP/Microtubule affinity-regulating kinase 4 (MARK4) belongs to the family of serine/threonine kinases that phosphorylate the microtubule-associated proteins (MAP) causing their detachment from the microtubules thereby increasing microtubule dynamics and facilitating cell division, cell cycle control, cell polarity determination, cell shape alterations, etc. The MARK4 gene encodes two alternatively spliced isoforms, L and S that differ in their C-terminal region. These isoforms are differentially regulated in human tissues including central nervous system. MARK4L is a 752-residue-long polypeptide that is divided into three distinct domains: (1) protein kinase domain (59-314), (2) ubiquitin-associated domain (322-369), and (3) kinase-associated domain (703-752) plus 54 residues (649-703) involved in the proper folding and function of the enzyme. In addition, residues 65-73 are considered to be the ATP-binding domain and Lys88 is considered as ATP-binding site. Asp181 has been proposed to be the active site of MARK4 that is activated by phosphorylation of Thr214 side chain. The isoform MARK4S is highly expressed in the normal brain and is presumably involved in neuronal differentiation. On the other hand, the isoform MARK4L is upregulated in hepatocarcinoma cells and gliomas suggesting its involvement in cell cycle. Several biological functions are also associated with MARK4 including microtubule bundle formation, nervous system development, and positive regulation of programmed cell death. Therefore, MARK4 is considered as the most suitable target for structure-based rational drug design. Our sequence, structure- and function-based analysis should be helpful for better understanding of mechanisms of regulation of microtubule dynamics and MARK4 associated diseases.

  8. On the Nature and Shape of Tubulin Trails: Implications on Microtubule Self-Organization

    CERN Document Server

    Glade, Nicolas

    2012-01-01

    Microtubules, major elements of the cell skeleton are, most of the time, well organized in vivo, but they can also show self-organizing behaviors in time and/or space in purified solutions in vitro. Theoretical studies and models based on the concepts of collective dynamics in complex systems, reaction-diffusion processes and emergent phenomena were proposed to explain some of these behaviors. In the particular case of microtubule spatial self-organization, it has been advanced that microtubules could behave like ants, self-organizing by 'talking to each other' by way of hypothetic (because never observed) concentrated chemical trails of tubulin that are expected to be released by their disassembling ends. Deterministic models based on this idea yielded indeed like-looking spatio-temporal self-organizing behaviors. Nevertheless the question remains of whether microscopic tubulin trails produced by individual or bundles of several microtubules are intense enough to allow microtubule self-organization at a macr...

  9. Phospholipase d activation correlates with microtubule reorganization in living plant cells.

    Science.gov (United States)

    Dhonukshe, Pankaj; Laxalt, Ana M; Goedhart, Joachim; Gadella, Theodorus W J; Munnik, Teun

    2003-11-01

    A phospholipase D (PLD) was shown recently to decorate microtubules in plant cells. Therefore, we used tobacco BY-2 cells expressing the microtubule reporter GFP-MAP4 to test whether PLD activation affects the organization of plant microtubules. Within 30 min of adding n-butanol, a potent activator of PLD, cortical microtubules were released from the plasma membrane and partially depolymerized, as visualized with four-dimensional confocal imaging. The isomers sec- and tert-butanol, which did not activate PLD, did not affect microtubule organization. The effect of treatment on PLD activation was monitored by the in vivo formation of phosphatidylbutanol, a specific reporter of PLD activity. Tobacco cells also were treated with mastoparan, xylanase, NaCl, and hypoosmotic stress as reported activators of PLD. We confirmed the reports and found that all treatments induced microtubule reorganization and PLD activation within the same time frame. PLD still was activated in microtubule-stabilized (taxol) and microtubule-depolymerized (oryzalin) situations, suggesting that PLD activation triggers microtubular reorganization and not vice versa. Exogenously applied water-soluble synthetic phosphatidic acid did not affect the microtubular cytoskeleton. Cell cycle studies revealed that n-butanol influenced not just interphase cortical microtubules but also those in the preprophase band and phragmoplast, but not those in the spindle structure. Cell growth and division were inhibited in the presence of n-butanol, whereas sec- and tert-butanol had no such effects. Using these novel insights, we propose a model for the mechanism by which PLD activation triggers microtubule reorganization in plant cells.

  10. ErbB2-dependent chemotaxis requires microtubule capture and stabilization coordinated by distinct signaling pathways.

    Directory of Open Access Journals (Sweden)

    Khedidja Benseddik

    Full Text Available Activation of the ErbB2 receptor tyrosine kinase stimulates breast cancer cell migration. Cell migration is a complex process that requires the synchronized reorganization of numerous subcellular structures including cell-to-matrix adhesions, the actin cytoskeleton and microtubules. How the multiple signaling pathways triggered by ErbB2 coordinate, in time and space, the various processes involved in cell motility, is poorly defined. We investigated the mechanism whereby ErbB2 controls microtubules and chemotaxis. We report that activation of ErbB2 increased both cell velocity and directed migration. Impairment of the Cdc42 and RhoA GTPases, but not of Rac1, prevented the chemotactic response. RhoA is a key component of the Memo/ACF7 pathway whereby ErbB2 controls microtubule capture at the leading edge. Upon Memo or ACF7 depletion, microtubules failed to reach the leading edge and cells lost their ability to follow the chemotactic gradient. Constitutive ACF7 targeting to the membrane in Memo-depleted cells reestablished directed migration. ErbB2-mediated activation of phospholipase C gamma (PLCγ also contributed to cell guidance. We further showed that PLCγ signaling, via classical protein kinases C, and Memo signaling converged towards a single pathway controlling the microtubule capture complex. Finally, inhibiting the PI3K/Akt pathway did not affect microtubule capture, but disturbed microtubule stability, which also resulted in defective chemotaxis. PI3K/Akt-dependent stabilization of microtubules involved repression of GSK3 activity on the one hand and inhibition of the microtubule destabilizing protein, Stathmin, on the other hand. Thus, ErbB2 triggers distinct and complementary pathways that tightly coordinate microtubule capture and microtubule stability to control chemotaxis.

  11. Oxidative stress decreases microtubule growth and stability in ventricular myocytes

    OpenAIRE

    Drum, BML; Yuan, C.; Li, L; Liu, Q.; Wordeman, L; Santana, LF

    2016-01-01

    © 2016 Elsevier Ltd.Microtubules (MTs) have many roles in ventricular myocytes, including structural stability, morphological integrity, and protein trafficking. However, despite their functional importance, dynamic MTs had never been visualized in living adult myocytes. Using adeno-associated viral vectors expressing the MT-associated protein plus end binding protein 3 (EB3) tagged with EGFP, we were able to perform live imaging and thus capture and quantify MT dynamics in ventricular myocyt...

  12. The microtubule aster formation and its role in nuclear envelope assembly around the sperm chromatin in Xenopus egg extracts

    Institute of Scientific and Technical Information of China (English)

    YANG Ning; CHEN Zhongcai; LU Ping; ZHANG Chuanmao; ZHAI Zhonghe; TANG Xiaowei

    2003-01-01

    Nuclear envelope is a dynamic structure in the cell cycle. At the beginning of mitosis, nuclear envelope breaks down and its components disperse into the cytoplasm. At the end of mitosis, nuclear envelope reassembles using the dispersed components. Searching for the mechanisms of the nuclear disassembly and reassembly has for a long time been one of the key projects for cell biologists. In this report we show that microtubules take a role in the nuclear envelope assembly around the sperm chromatin in Xenopus egg extracts. Microtubule cytoskeleton has been demonstrated to take roles in the transport of intracellular membranes such as Golgi and ER vesicles. We found that the nuclear envelope assembly needs functional microtubules. At the beginning of the nuclear assembly, microtubules nucleated to form a microtubule aster around the centrosome at the base of the sperm head. Using the microtubule drug colchicine to disrupt the microtubule nucleation, nuclear envelope reassembly was seriously inhibited. If the microtubules were stabilized by taxol, another microtubule drug, the nuclear envelope reassembly was also interfered, although a significantly large aster formed around the chromatin. Based on these observations, we propose that microtubules play an important role in the nuclear envelope reassembly maybe by transporting the nuclear envelope precursors to the chromatin surfaces.

  13. Dependency of microtubule-associated proteins (MAPs) for tubulin stability and assembly; use of estramustine phosphate in the study of microtubules.

    Science.gov (United States)

    Fridén, B; Wallin, M

    1991-07-10

    Microtubule-associated proteins (MAPs) were separated from tubulin with several different methods. The ability of the isolated MAPs to reinduce assembly of phosphocellulose purified tubulin differed markedly between the different methods. MAPs isolated by addition of 0.35 M NaCl to taxol-stabilized microtubules stimulated tubulin assembly most effectively, while addition of 0.6 M NaCl produced MAPs with a substantially lower ability to stimulate tubulin assembly. The second best preparation was achieved with phosphocellulose chromatographic separation of MAPs with 0.6 M NaCl elution. The addition of estramustine phosphate to microtubules reconstituted of MAPs prepared by 0.35 M NaCl or phosphocellulose chromatography, induced less disassembly than for microtubules assembled from unseparated proteins, and was almost without effect on microtubules reconstituted from MAPs prepared by taxol and 0.6 M NaCl. Estramustine phosphate binds to the tubulin binding part of the MAPs, and the results do therefore indicate that the MAPs are altered by the separation methods. Since the MAPs are regarded as highly stable molecules, one probable alteration could be aggregation of the MAPs, as also indicated by the results. The purified tubulin itself seemed not to be affected by the phosphocellulose purification, since the microtubule proteins were unchanged by the low buffer strenght used during the cromatography. However, the assembly competence after a prolonged incubation of the microtubule proteins at 4 degrees C was dependent on intact bindings between the tubulin and MAPs.

  14. Molecular mechanisms of antitumor activity of taxanes. I. Interaction of docetaxel with microtubules

    Directory of Open Access Journals (Sweden)

    Sabina Tabaczar

    2010-11-01

    Full Text Available Docetaxel (Taxotere, a new semisynthetic taxoid, is a mitotic inhibitor, widely used in monotherapy or in combination with other anticancer drugs against many types of cancer. The structure and dynamics of microtubules as the main target for docetaxel activity inside the cell and the taxane-binding site on β-tubulin are discussed. Microtubules are highly dynamic assemblies of α- and β-tubulin. They readily polymerize and depolymerize in cells and these dynamic behaviours are crucial to cell mitosis. Microtubule instability is attributed to their capability to hydrolyze GTP to GDP, which causes their depolymerization. Addition of new α-, β-tubulin heterodimer bound to GTP leads to tubulin polymerization, which increases the length of the microtubule. Docetaxel alters the polymerization dynamics of microtubules, which causes blockage of cell mitosis, and consequently induces apoptotic and non-apoptotic cell death. Docetaxel specifically acts on the S, M and G2 phases of the cell cycle. This paper reviews the current state of knowledge related to the molecular mechanisms of docetaxel action on the cell cycle and microtubule dynamics. In addition, a brief survey of the present state of research on the new generation (2nd and 3rd of taxanes is presented.

  15. Mechanism for the catastrophe-promoting activity of the microtubule destabilizer Op18/stathmin.

    Science.gov (United States)

    Gupta, Kamlesh K; Li, Chunlei; Duan, Aranda; Alberico, Emily O; Kim, Oleg V; Alber, Mark S; Goodson, Holly V

    2013-12-17

    Regulation of microtubule dynamic instability is crucial for cellular processes, ranging from mitosis to membrane transport. Stathmin (also known as oncoprotein 18/Op18) is a prominent microtubule destabilizer that acts preferentially on microtubule minus ends. Stathmin has been studied intensively because of its association with multiple types of cancer, but its mechanism of action remains controversial. Two models have been proposed. One model is that stathmin promotes microtubule catastrophe indirectly, and does so by sequestering tubulin; the other holds that stathmin alters microtubule dynamics by directly destabilizing growing microtubules. Stathmin's sequestration activity is well established, but the mechanism of any direct action is mysterious because stathmin binds to microtubules very weakly. To address these issues, we have studied interactions between stathmin and varied tubulin polymers. We show that stathmin binds tightly to Dolastatin-10 tubulin rings, which mimic curved tubulin protofilaments, and that stathmin depolymerizes stabilized protofilament-rich polymers. These observations lead us to propose that stathmin promotes catastrophe by binding to and acting upon protofilaments exposed at the tips of growing microtubules. Moreover, we suggest that stathmin's minus-end preference results from interactions between stathmin's N terminus and the surface of α-tubulin that is exposed only at the minus end. Using computational modeling of microtubule dynamics, we show that these mechanisms could account for stathmin's observed activities in vitro, but that both the direct and sequestering activities are likely to be relevant in a cellular context. Taken together, our results suggest that stathmin can promote catastrophe by direct action on protofilament structure and interactions.

  16. Sclerotium rolfsii dynamics in soil as affected by crop sequences

    NARCIS (Netherlands)

    Leoni, C.; Braak, ter C.J.F.; Gilsanz, J.C.; Dogliotti, S.; Rossing, W.A.H.; Bruggen, van A.H.C.

    2014-01-01

    Crop rotation has been used for the management of soilborne diseases for centuries, but has not often been planned based on scientific knowledge. Our objective was to generate information on Sclerotium rolfsii dynamics under different crop or intercrop activities, and design and test a research appr

  17. Aging affects attunement in perceiving length by dynamic touch

    NARCIS (Netherlands)

    Withagen, Rob; Caljouw, Simone R.

    2011-01-01

    Earlier studies have revealed age-dependent differences in perception by dynamic touch. In the present study, we examined whether the capacity to learn deteriorates with aging. Adopting an ecological approach to learning, the authors examined the process of attunement-that is, the changes in what in

  18. Microtubule-microtubule sliding by kinesin-1 is essential for normal cytoplasmic streaming in Drosophila oocytes.

    Science.gov (United States)

    Lu, Wen; Winding, Michael; Lakonishok, Margot; Wildonger, Jill; Gelfand, Vladimir I

    2016-08-23

    Cytoplasmic streaming in Drosophila oocytes is a microtubule-based bulk cytoplasmic movement. Streaming efficiently circulates and localizes mRNAs and proteins deposited by the nurse cells across the oocyte. This movement is driven by kinesin-1, a major microtubule motor. Recently, we have shown that kinesin-1 heavy chain (KHC) can transport one microtubule on another microtubule, thus driving microtubule-microtubule sliding in multiple cell types. To study the role of microtubule sliding in oocyte cytoplasmic streaming, we used a Khc mutant that is deficient in microtubule sliding but able to transport a majority of cargoes. We demonstrated that streaming is reduced by genomic replacement of wild-type Khc with this sliding-deficient mutant. Streaming can be fully rescued by wild-type KHC and partially rescued by a chimeric motor that cannot move organelles but is active in microtubule sliding. Consistent with these data, we identified two populations of microtubules in fast-streaming oocytes: a network of stable microtubules anchored to the actin cortex and free cytoplasmic microtubules that moved in the ooplasm. We further demonstrated that the reduced streaming in sliding-deficient oocytes resulted in posterior determination defects. Together, we propose that kinesin-1 slides free cytoplasmic microtubules against cortically immobilized microtubules, generating forces that contribute to cytoplasmic streaming and are essential for the refinement of posterior determinants.

  19. Space charges can significantly affect the dynamics of accelerator maps

    Science.gov (United States)

    Bountis, Tassos; Skokos, Charalampos

    2006-10-01

    Space charge effects can be very important for the dynamics of intense particle beams, as they repeatedly pass through nonlinear focusing elements, aiming to maximize the beam's luminosity properties in the storage rings of a high energy accelerator. In the case of hadron beams, whose charge distribution can be considered as “frozen” within a cylindrical core of small radius compared to the beam's dynamical aperture, analytical formulas have been recently derived [C. Benedetti, G. Turchetti, Phys. Lett. A 340 (2005) 461] for the contribution of space charges within first order Hamiltonian perturbation theory. These formulas involve distribution functions which, in general, do not lead to expressions that can be evaluated in closed form. In this Letter, we apply this theory to an example of a charge distribution, whose effect on the dynamics can be derived explicitly and in closed form, both in the case of 2-dimensional as well as 4-dimensional mapping models of hadron beams. We find that, even for very small values of the “perveance” (strength of the space charge effect) the long term stability of the dynamics changes considerably. In the flat beam case, the outer invariant “tori” surrounding the origin disappear, decreasing the size of the beam's dynamical aperture, while beyond a certain threshold the beam is almost entirely lost. Analogous results in mapping models of beams with 2-dimensional cross section demonstrate that in that case also, even for weak tune depressions, orbital diffusion is enhanced and many particles whose motion was bounded now escape to infinity, indicating that space charges can impose significant limitations on the beam's luminosity.

  20. Space charges can significantly affect the dynamics of accelerator maps

    Energy Technology Data Exchange (ETDEWEB)

    Bountis, Tassos [Department of Mathematics, University of Patras, GR-26500 Patras (Greece) and Center for Research and Applications of Nonlinear Systems (CRANS), University of Patras, GR-26500 Patras (Greece)]. E-mail: tassos50@otenet.gr; Skokos, Charalampos [Center for Research and Applications of Nonlinear Systems (CRANS), University of Patras, GR-26500 Patras (Greece)

    2006-10-09

    Space charge effects can be very important for the dynamics of intense particle beams, as they repeatedly pass through nonlinear focusing elements, aiming to maximize the beam's luminosity properties in the storage rings of a high energy accelerator. In the case of hadron beams, whose charge distribution can be considered as 'frozen' within a cylindrical core of small radius compared to the beam's dynamical aperture, analytical formulas have been recently derived [C. Benedetti, G. Turchetti, Phys. Lett. A 340 (2005) 461] for the contribution of space charges within first order Hamiltonian perturbation theory. These formulas involve distribution functions which, in general, do not lead to expressions that can be evaluated in closed form. In this Letter, we apply this theory to an example of a charge distribution, whose effect on the dynamics can be derived explicitly and in closed form, both in the case of 2-dimensional as well as 4-dimensional mapping models of hadron beams. We find that, even for very small values of the 'perveance' (strength of the space charge effect) the long term stability of the dynamics changes considerably. In the flat beam case, the outer invariant 'tori' surrounding the origin disappear, decreasing the size of the beam's dynamical aperture, while beyond a certain threshold the beam is almost entirely lost. Analogous results in mapping models of beams with 2-dimensional cross section demonstrate that in that case also, even for weak tune depressions, orbital diffusion is enhanced and many particles whose motion was bounded now escape to infinity, indicating that space charges can impose significant limitations on the beam's luminosity.

  1. A Stochastic Multiscale Model That Explains the Segregation of Axonal Microtubules and Neurofilaments in Neurological Diseases.

    Directory of Open Access Journals (Sweden)

    Chuan Xue

    2015-08-01

    Full Text Available The organization of the axonal cytoskeleton is a key determinant of the normal function of an axon, which is a long thin projection of a neuron. Under normal conditions two axonal cytoskeletal polymers, microtubules and neurofilaments, align longitudinally in axons and are interspersed in axonal cross-sections. However, in many neurotoxic and neurodegenerative disorders, microtubules and neurofilaments segregate apart from each other, with microtubules and membranous organelles clustered centrally and neurofilaments displaced to the periphery. This striking segregation precedes the abnormal and excessive neurofilament accumulation in these diseases, which in turn leads to focal axonal swellings. While neurofilament accumulation suggests an impairment of neurofilament transport along axons, the underlying mechanism of their segregation from microtubules remains poorly understood for over 30 years. To address this question, we developed a stochastic multiscale model for the cross-sectional distribution of microtubules and neurofilaments in axons. The model describes microtubules, neurofilaments and organelles as interacting particles in a 2D cross-section, and is built upon molecular processes that occur on a time scale of seconds or shorter. It incorporates the longitudinal transport of neurofilaments and organelles through this domain by allowing stochastic arrival and departure of these cargoes, and integrates the dynamic interactions of these cargoes with microtubules mediated by molecular motors. Simulations of the model demonstrate that organelles can pull nearby microtubules together, and in the absence of neurofilament transport, this mechanism gradually segregates microtubules from neurofilaments on a time scale of hours, similar to that observed in toxic neuropathies. This suggests that the microtubule-neurofilament segregation can be a consequence of the selective impairment of neurofilament transport. The model generates the

  2. Predicting when climate-driven phenotypic change affects population dynamics

    NARCIS (Netherlands)

    McLean, Nina; Lawson, C.R.; Leech, David; Van de Pol, M.

    2016-01-01

    Species' responses to climate change are variable and diverse, yet our understanding of how different responses (e.g. physiological, behavioural, demographic) relate and how they affect the parameters most relevant for conservation (e.g. population persistence) is lacking. Despite this, studies that

  3. Interaction of an Overexpressed gamma-Tubulin with Microtubules In Vivo and In Vitro.

    Science.gov (United States)

    Kofron, M; Nadezdina, E; Vassilev, A; Matuliene, J; Essner, R; Kato, J; Kuriyama, R

    1998-08-01

    gamma-Tubulin is an ubiquitous MTOC (microtubule-organizing center) component essential for the regulation of microtubule functions. A 1.8 kb cDNA coding for gamma-tubulin was isolated from CHO cells. Analysis of nucleotide sequence predicts a protein of 451 amino acids, which is over 97% identical to human and Xenopus gamma-tubulin. When CHO cells were transiently transfected with the gamma-tubulin clone, epitope-tagged full-length, as well as truncated polypeptides (amino acids 1-398 and 1-340), resulted in the formation of cytoplasmic foci of various sizes. Although one of the foci was identified as the centrosome, the rest of the dots were not associated with any other centrosomal components tested so far. The pattern of microtubule organization was not affected by induction of such gamma-tubulin-containing dots in transfected cells. In addition, the cytoplasmic foci were unable to serve as the site for microtubule regrowth in nocodazole-treated cells upon removal of the drug, suggesting that gamma-tubulin-containing foci were not involved in the activity for microtubule formation and organization. Using the monomeric form of Chlamydomonas gamma-tubulin purified from insect Sf9 cells (), interaction between gamma-tubulin and microtubules was further investigated by immunoelectron microscopy. Microtubules incubated with gamma-tubulin monomers in vitro were associated with more gold particles conjugated with gamma-tubulin than in controls where no exogenous gamma-tubulin was added. However, binding of gamma-tubulin to microtubules was not extensive and was easily lost during sample preparation. Although gamma-tubulin was detected at the minus end of microtubules several times more frequently than the plus end, the majority of gold particles were seen along the microtubule length. These results contradict the previous reports (; ), which might be ascribed to the difference in the level of protein expression in transfected cells.

  4. Affective Dynamics of Leadership: An Experimental Test of Affect Control Theory

    Science.gov (United States)

    Schroder, Tobias; Scholl, Wolfgang

    2009-01-01

    Affect Control Theory (ACT; Heise 1979, 2007) states that people control social interactions by striving to maintain culturally shared feelings about the situation. The theory is based on mathematical models of language-based impression formation. In a laboratory experiment, we tested the predictive power of a new German-language ACT model with…

  5. Dynamically Tracking Anxious Individuals' Affective Response to Valenced Information.

    Science.gov (United States)

    Fua, Karl C; Teachman, Bethany A

    2017-03-30

    Past research has shown that an individual's feelings at any given moment reflect currently experienced stimuli as well as internal representations of similar past experiences. However, anxious individuals' affective reactions to streams of interrelated valenced information (vs. reactions to static stimuli that are arguably less ecologically valid) are rarely tracked. The present study provided a first examination of the newly developed Tracking Affect Ratings Over Time (TAROT) task to continuously assess anxious individuals' affective reactions to streams of information that systematically change valence. Undergraduate participants (N = 141) completed the TAROT task in which they listened to narratives containing positive, negative, and neutral physically- or socially-relevant events, and indicated how positive or negative they felt about the information they heard as each narrative unfolded. The present study provided preliminary evidence for the validity and reliability of the task. Within scenarios, participants higher (vs. lower) in anxiety showed many expected negative biases, reporting more negative mean ratings and overall summary ratings, changing their pattern of responding more quickly to negative events, and responding more negatively to neutral events. Furthermore, individuals higher (vs. lower) in anxiety tended to report more negative minimums during and after positive events, and less positive maximums after negative events. Together, findings indicate that positive events were less impactful for anxious individuals, whereas negative experiences had a particularly lasting impact on future affective responses. The TAROT task is able to efficiently capture a number of different cognitive biases, and may help clarify the mechanisms that underlie anxious individuals' biased negative processing. (PsycINFO Database Record

  6. Incorporating Affect in an Engineering Student's Epistemological Dynamics

    CERN Document Server

    Danielak, Brian A; Elby, Andrew

    2010-01-01

    Research has linked a student's affect to her epistemology (Boaler & Greeno, 2000), but those constructs often apply broadly to a discipline and/or classroom culture. Independently, an emerging line of research shows that a student in a given classroom and discipline can shift between multiple locally coherent epistemological stances (Hammer, Elby, Scherr, & Redish, 2005). Our case study of Judy, an undergraduate engineering major, begins our long-term effort at uniting these two bodies of literature.

  7. Tau Induces Cooperative Taxol Binding to Microtubules

    Science.gov (United States)

    Ross, Jennifer; Santangelo, Christian; Victoria, Makrides; Fygenson, Deborah

    2004-03-01

    Taxol and tau are two ligands which stabilize the microtubule (MT) lattice. Taxol is an anti-mitotic drug that binds β tubulin in the MT interior. Tau is a MT-associated protein that binds both α and β tubulin on the MT exterior. Both taxol and tau reduce MT dynamics and promote tubulin polymerization. Tau alone also acts as a buttress to bundle, stiffen, and space MTs. A structural study recently suggested that taxol and tau may interact by binding to the same site. Using fluorescence recovery after photobleaching, we find that tau induces taxol to bind MTs cooperatively depending on the tau concentration. We develop a model that correctly fits the data in the absence of tau and yields a measure of taxol cooperativity when tau is present.

  8. Microtubule as nanobioelectronic nonlinear circuit

    Directory of Open Access Journals (Sweden)

    Sekulić Dalibor L.

    2012-01-01

    Full Text Available In recent years, the use of biological molecules has offered exciting alternatives to conventional synthetic methods. Specific methods use various biological templates to direct the deposition and patterning of inorganic materials. Here we have established a new electrical model of microtubules as a biological nanoscale circuit based on polyelectrolyte features of cylindrical biopolymers. Our working hypothesis is that microtubules play an active role in sub-cellular computation and signaling via electronic and protonic conductivity and can thus be made useful in hybrid materials that offer novel electronic characteristics. We verify these hypotheses both computationally and analytically through a quantitative model based on the atomic resolution structures of the key functional proteins.

  9. Detailed Per-residue Energetic Analysis Explains the Driving Force for Microtubule Disassembly.

    Directory of Open Access Journals (Sweden)

    Ahmed T Ayoub

    2015-06-01

    Full Text Available Microtubules are long filamentous hollow cylinders whose surfaces form lattice structures of αβ-tubulin heterodimers. They perform multiple physiological roles in eukaryotic cells and are targets for therapeutic interventions. In our study, we carried out all-atom molecular dynamics simulations for arbitrarily long microtubules that have either GDP or GTP molecules in the E-site of β-tubulin. A detailed energy balance of the MM/GBSA inter-dimer interaction energy per residue contributing to the overall lateral and longitudinal structural stability was performed. The obtained results identified the key residues and tubulin domains according to their energetic contributions. They also identified the molecular forces that drive microtubule disassembly. At the tip of the plus end of the microtubule, the uneven distribution of longitudinal interaction energies within a protofilament generates a torque that bends tubulin outwardly with respect to the cylinder's axis causing disassembly. In the presence of GTP, this torque is opposed by lateral interactions that prevent outward curling, thus stabilizing the whole microtubule. Once GTP hydrolysis reaches the tip of the microtubule (lateral cap, lateral interactions become much weaker, allowing tubulin dimers to bend outwards, causing disassembly. The role of magnesium in the process of outward curling has also been demonstrated. This study also showed that the microtubule seam is the most energetically labile inter-dimer interface and could serve as a trigger point for disassembly. Based on a detailed balance of the energetic contributions per amino acid residue in the microtubule, numerous other analyses could be performed to give additional insights into the properties of microtubule dynamic instability.

  10. Thermodynamic and structural analysis of microtubule assembly: the role of GTP hydrolysis.

    Science.gov (United States)

    Vulevic, B; Correia, J J

    1997-03-01

    Different models have been proposed that link the tubulin heterodimer nucleotide content and the role of GTP hydrolysis with microtubule assembly and dynamics. Here we compare the thermodynamics of microtubule assembly as a function of nucleotide content by van't Hoff analysis. The thermodynamic parameters of tubulin assembly in 30-100 mM piperazine-N,N'-bis(2-ethanesulfonic acid), 1 mM MgSO4, 2 mM EGTA, pH 6.9, in the presence of a weakly hydrolyzable analog, GMPCPP, the dinucleotide analog GMPCP plus 2 M glycerol, and GTP plus 2 M glycerol were obtained together with data for taxol-GTP/GDP tubulin assembly (GMPCPP and GMPCP are the GTP and GDP nucleotide analogs where the alpha beta oxygen has been replaced by a methylene, -CH2-). All of the processes studied are characterized by a positive enthalpy, a positive entropy, and a large, negative heat capacity change. GMPCP-induced assembly has the largest negative heat capacity change and GMPCPP has the second largest, whereas GTP/2 M glycerol- and taxol-induced assembly have more positive values, respectively. A large, negative heat capacity is most consistent with the burial of water-accessible hydrophobic surface area, which gives rise to the release of bound water. The heat capacity changes observed with GTP/2 M glycerol-induced and with taxol-induced assembly are very similar, -790 +/- 190 cal/mol/k, and correspond to the burial of 3330 +/- 820 A2 of nonpolar surface area. This value is shown to be very similar to an estimate of the buried nonpolar surface in a reconstructed microtubule lattice. Polymerization data from GMPCP- and GMPCPP-induced assembly are consistent with buried nonpolar surface areas that are 3 and 6 times larger. A linear enthalpy-entropy and enthalpy-free energy plot for tubulin polymerization reactions verifies that enthalpy-entropy compensation for this system is based upon true biochemical correlation, most likely corresponding to a dominant hydrophobic effect. Entropy analysis suggests

  11. Microtubules in Dendritic Spine Development

    OpenAIRE

    2008-01-01

    It is generally believed that only the actin cytoskeleton resides in dendritic spines and controls spine morphology and plasticity. Here we report that microtubules (MTs) are present in spines and that shRNA knockdown of the MT-plus end binding protein EB3 significantly reduces spine formation. Furthermore, stabilization and inhibition of MTs by low doses of taxol and nocodazole enhance and impair spine formation elicited by BDNF, respectively. Therefore, MTs play an important role in the con...

  12. Agulhas leakage dynamics affects decadal variability in Atlantic overturning circulation.

    Science.gov (United States)

    Biastoch, A; Böning, C W; Lutjeharms, J R E

    2008-11-27

    Predicting the evolution of climate over decadal timescales requires a quantitative understanding of the dynamics that govern the meridional overturning circulation (MOC). Comprehensive ocean measurement programmes aiming to monitor MOC variations have been established in the subtropical North Atlantic (RAPID, at latitude 26.5 degrees N, and MOVE, at latitude 16 degrees N) and show strong variability on intraseasonal to interannual timescales. Observational evidence of longer-term changes in MOC transport remains scarce, owing to infrequent sampling of transoceanic sections over past decades. Inferences based on long-term sea surface temperature records, however, supported by model simulations, suggest a variability with an amplitude of +/-1.5-3 Sv (1 Sv = 10(6) m(3) s(-1)) on decadal timescales in the subtropics. Such variability has been attributed to variations of deep water formation in the sub-arctic Atlantic, particularly the renewal rate of Labrador Sea Water. Here we present results from a model simulation that suggest an additional influence on decadal MOC variability having a Southern Hemisphere origin: dynamic signals originating in the Agulhas leakage region at the southern tip of Africa. These contribute a MOC signal in the tropical and subtropical North Atlantic that is of the same order of magnitude as the northern source. A complete rationalization of observed MOC changes therefore also requires consideration of signals arriving from the south.

  13. CYLD Regulates Noscapine Activity in Acute Lymphoblastic Leukemia via a Microtubule-Dependent Mechanism

    OpenAIRE

    Yang, Yunfan; Ran, Jie; Sun, Lei; Sun, Xiaodong; Luo, Youguang; Yan, Bing; Tala,; Liu, Min; Li, Dengwen; Zhang, Lei; Bao, Gang; Zhou, Jun

    2015-01-01

    Noscapine is an orally administrable drug used worldwide for cough suppression and has recently been demonstrated to disrupt microtubule dynamics and possess anticancer activity. However, the molecular mechanisms regulating noscapine activity remain poorly defined. Here we demonstrate that cylindromatosis (CYLD), a microtubule-associated tumor suppressor protein, modulates the activity of noscapine both in cell lines and in primary cells of acute lymphoblastic leukemia (ALL). Flow cytometry a...

  14. Detailed Per-residue Energetic Analysis Explains the Driving Force for Microtubule Disassembly

    OpenAIRE

    Ayoub, Ahmed T.; Mariusz Klobukowski; Tuszynski, Jack A

    2015-01-01

    Microtubules are long filamentous hollow cylinders whose surfaces form lattice structures of αβ-tubulin heterodimers. They perform multiple physiological roles in eukaryotic cells and are targets for therapeutic interventions. In our study, we carried out all-atom molecular dynamics simulations for arbitrarily long microtubules that have either GDP or GTP molecules in the E-site of β-tubulin. A detailed energy balance of the MM/GBSA inter-dimer interaction energy per residue contributing to t...

  15. Enantioselective effects of (+)- and (-)-citronellal on animal and plant microtubules.

    Science.gov (United States)

    Altshuler, Osnat; Abu-Abied, Mohamad; Chaimovitsh, David; Shechter, Alona; Frucht, Hilla; Dudai, Nativ; Sadot, Einat

    2013-09-27

    Citronellal is a major component of Corymbia citriodora and Cymbopogon nardus essential oils. Herein it is shown that whereas (+)-citronellal (1) is an effective microtubule (MT)-disrupting compound, (-)-citronellal (2) is not. Quantitative image analysis of fibroblast cells treated with 1 showed total fluorescence associated with fibers resembling that in cells treated with the MT-disrupting agents colchicine and vinblastine; in the presence of 2, the fluorescence more closely resembled that in control cells. The distribution of tubulin in soluble and insoluble fractions in the presence of 1 also resembled that in the presence of colchicine, whereas similar tubulin distribution was obtained in the presence of 2 and in control cells. In vitro polymerization of MTs was inhibited by 1 but not 2. Measurements of MT dynamics in plant cells showed similar MT elongation and shortening rates in control and 2-treated cells, whereas in the presence of 1, much fewer and shorter MTs were observed and no elongation or shrinkage was detected. Taken together, the MT system is suggested to be able to discriminate between different enantiomers of the same compound. In addition, the activity of essential oils rich in citronellal is affected by the relative content of the two enantiomers of this monoterpenoid.

  16. Fission yeast Scp3 potentially maintains microtubule orientation through bundling.

    Directory of Open Access Journals (Sweden)

    Kanako Ozaki

    Full Text Available Microtubules play important roles in organelle transport, the maintenance of cell polarity and chromosome segregation and generally form bundles during these processes. The fission yeast gene scp3+ was identified as a multicopy suppressor of the cps3-81 mutant, which is hypersensitive to isopropyl N-3-chlorophenylcarbamate (CIPC, a poison that induces abnormal multipolar spindle formation in higher eukaryotes. In this study, we investigated the function of Scp3 along with the effect of CIPC in the fission yeast Schizosaccharomyces pombe. Microscopic observation revealed that treatment with CIPC, cps3-81 mutation and scp3+ gene deletion disturbed the orientation of microtubules in interphase cells. Overexpression of scp3+ suppressed the abnormal orientation of microtubules by promoting bundling. Functional analysis suggested that Scp3 functions independently from Ase1, a protein largely required for the bundling of the mitotic spindle. A strain lacking the ase1+ gene was more sensitive to CIPC, with the drug affecting the integrity of the mitotic spindle, indicating that CIPC has a mitotic target that has a role redundant with Ase1. These results suggested that multiple systems are independently involved to ensure microtubule orientation by bundling in fission yeast.

  17. How Predation and Landscape Fragmentation Affect Vole Population Dynamics

    DEFF Research Database (Denmark)

    Dalkvist, Trine; Sibly, Richard M.; Topping, Chris J.

    2011-01-01

    population cycles. Because these factors covary along the gradient it is difficult to distinguish their effects experimentally in the field. The distinction is here attempted using realistic agent-based modelling. Methodology/Principal Findings: By using a spatially explicit computer simulation model based...... on behavioural and ecological data from the field vole (Microtus agrestis), we generated a number of repeated time series of vole densities whose mean population size and amplitude were measured. Subsequently, these time series were subjected to statistical autoregressive modelling, to investigate the effects...... on vole population dynamics of making predators more specialised, of altering the breeding season, and increasing the level of habitat fragmentation. We found that fragmentation as well as the presence of specialist predators are necessary for the occurrence of population cycles. Habitat fragmentation...

  18. Lubricated wrinkles: imposed constraints affect the dynamics of wrinkle coarsening

    CERN Document Server

    Kodio, Ousmane; Vella, Dominic

    2016-01-01

    We study the dynamic coarsening of wrinkles in an elastic sheet that is compressed while lying on a thin layer of viscous liquid. When the ends of the sheet are instantaneously brought together by a small distance, viscous resistance initially prevents the sheet from adopting a globally buckled shape. Instead, the sheet accommodates the compression by wrinkling. Previous scaling arguments suggested that a balance between the sheet's bending stiffness and viscous effects lead to a wrinkle wavelength $\\lambda$ that increases with time $t$ according to $\\lambda\\propto t^{1/6}$. We show that taking proper account of the compression constraint leads to a logarithmic correction of this result, $\\lambda\\propto (t/\\log t)^{1/6}$. This correction is significant over experimentally observable time spans, and leads us to reassess previously published experimental data.

  19. Lubricated wrinkles: Imposed constraints affect the dynamics of wrinkle coarsening

    Science.gov (United States)

    Kodio, Ousmane; Griffiths, Ian M.; Vella, Dominic

    2017-01-01

    We study the dynamic coarsening of wrinkles in an elastic sheet that is compressed while lying on a thin layer of viscous liquid. When the ends of the sheet are instantaneously brought together by a small distance, viscous resistance initially prevents the sheet from adopting a globally buckled shape. Instead, the sheet accommodates the compression by wrinkling. Previous scaling arguments suggested that a balance between the sheet's bending stiffness and viscous effects lead to a wrinkle wavelength λ that increases with time t according to λ ∝t1 /6 . We show that taking proper account of the compression constraint leads to a logarithmic correction of this result, λ ∝(t/logt ) 1 /6 . This correction is significant over experimentally observable time spans and leads us to reassess previously published experimental data.

  20. Dynamic touch is affected in children with cerebral palsy.

    Science.gov (United States)

    Ocarino, Juliana M; Fonseca, Sergio T; Silva, Paula L P; Gonçalves, Gabriela G P; Souza, Thales R; Mancini, Marisa C

    2014-02-01

    Children with developmental disorders such as cerebral palsy have limited opportunities for effortful interactions with objects and tools. The goal of the study was to investigate whether children with cerebral palsy have deficits in their ability to perceive object length by dynamic touch when compared to typically developing children. Fourteen children with typical development and 12 children with cerebral palsy were asked to report the length of hand-held rods after wielding them out of sight. Multilevel regression models indicated that I1 (maximum principal moment of inertia) was a significant predictor of perceived length - LP (pcerebral palsy (group factor) partially explained such variance (p=.002). In addition, accuracy and reliability of the length judgments made by children with cerebral palsy were significantly lower than the typically developing children (p<.05). Theoretical and clinical implications of these results were identified and discussed.

  1. Laulimalide induces dose-dependent modulation of microtubule behaviour in the C. elegans embryo.

    Directory of Open Access Journals (Sweden)

    Megha Bajaj

    Full Text Available Laulimalide is a microtubule-binding drug that was originally isolated from marine sponges. High concentrations of laulimalide stabilize microtubules and inhibit cell division similarly to paclitaxel; however, there are important differences with respect to the nature of the specific cellular defects between these two drugs and their binding sites on the microtubule. In this study, we used Caenorhabditis elegans embryos to investigate the acute effects of laulimalide on microtubules in vivo, with a direct comparison to paclitaxel. We observed surprising dose-dependent effects for laulimalide, whereby microtubules were stabilized at concentrations above 100 nM, but destabilized at concentrations between 50 and 100 nM. Despite this behaviour at low concentrations, laulimalide acted synergistically with paclitaxel to stabilize microtubules when both drugs were used at sub-effective concentrations, consistent with observations of synergistic interactions between these two drugs in other systems. Our results indicate that laulimalide induces a concentration-dependent, biphasic change in microtubule polymer dynamics in the C. elegans embryo.

  2. Social decisions affect neural activity to perceived dynamic gaze.

    Science.gov (United States)

    Latinus, Marianne; Love, Scott A; Rossi, Alejandra; Parada, Francisco J; Huang, Lisa; Conty, Laurence; George, Nathalie; James, Karin; Puce, Aina

    2015-11-01

    Gaze direction, a cue of both social and spatial attention, is known to modulate early neural responses to faces e.g. N170. However, findings in the literature have been inconsistent, likely reflecting differences in stimulus characteristics and task requirements. Here, we investigated the effect of task on neural responses to dynamic gaze changes: away and toward transitions (resulting or not in eye contact). Subjects performed, in random order, social (away/toward them) and non-social (left/right) judgment tasks on these stimuli. Overall, in the non-social task, results showed a larger N170 to gaze aversion than gaze motion toward the observer. In the social task, however, this difference was no longer present in the right hemisphere, likely reflecting an enhanced N170 to gaze motion toward the observer. Our behavioral and event-related potential data indicate that performing social judgments enhances saliency of gaze motion toward the observer, even those that did not result in gaze contact. These data and that of previous studies suggest two modes of processing visual information: a 'default mode' that may focus on spatial information; a 'socially aware mode' that might be activated when subjects are required to make social judgments. The exact mechanism that allows switching from one mode to the other remains to be clarified.

  3. The Drosophila microtubule-associated protein mars stabilizes mitotic spindles by crosslinking microtubules through its N-terminal region.

    Directory of Open Access Journals (Sweden)

    Gang Zhang

    Full Text Available Correct segregation of genetic material relies on proper assembly and maintenance of the mitotic spindle. How the highly dynamic microtubules (MTs are maintained in stable mitotic spindles is a key question to be answered. Motor and non-motor microtubule associated proteins (MAPs have been reported to stabilize the dynamic spindle through crosslinking adjacent MTs. Mars, a novel MAP, is essential for the early development of Drosophila embryos. Previous studies showed that Mars is required for maintaining an intact mitotic spindle but did not provide a molecular mechanism for this function. Here we show that Mars is able to stabilize the mitotic spindle in vivo. Both in vivo and in vitro data reveal that the N-terminal region of Mars functions in the stabilization of the mitotic spindle by crosslinking adjacent MTs.

  4. How Fear of Future Outcomes Affects Social Dynamics

    Science.gov (United States)

    Podobnik, Boris; Jusup, Marko; Wang, Zhen; Stanley, H. Eugene

    2016-11-01

    Mutualistic relationships among the different species are ubiquitous in nature. To prevent mutualism from slipping into antagonism, a host often invokes a "carrot and stick" approach towards symbionts with a stabilizing effect on their symbiosis. In open human societies, a mutualistic relationship arises when a native insider population attracts outsiders with benevolent incentives in hope that the additional labor will improve the standard of all. A lingering question, however, is the extent to which insiders are willing to tolerate outsiders before mutualism slips into antagonism. To test the assertion by Karl Popper that unlimited tolerance leads to the demise of tolerance, we model a society under a growing incursion from the outside. Guided by their traditions of maintaining the social fabric and prizing tolerance, the insiders reduce their benevolence toward the growing subpopulation of outsiders but do not invoke punishment. This reduction of benevolence intensifies as less tolerant insiders (e.g., "radicals") openly renounce benevolence. Although more tolerant insiders maintain some level of benevolence, they may also tacitly support radicals out of fear for the future. If radicals and their tacit supporters achieve a critical majority, herd behavior ensues and the relation between the insider and outsider subpopulations turns antagonistic. To control the risk of unwanted social dynamics, we map the parameter space within which the tolerance of insiders is in balance with the assimilation of outsiders, the tolerant insiders maintain a sustainable majority, and any reduction in benevolence occurs smoothly. We also identify the circumstances that cause the relations between insiders and outsiders to collapse or that lead to the dominance of the outsiders.

  5. Form of an evolutionary tradeoff affects eco-evolutionary dynamics in a predator-prey system.

    Science.gov (United States)

    Kasada, Minoru; Yamamichi, Masato; Yoshida, Takehito

    2014-11-11

    Evolution on a time scale similar to ecological dynamics has been increasingly recognized for the last three decades. Selection mediated by ecological interactions can change heritable phenotypic variation (i.e., evolution), and evolution of traits, in turn, can affect ecological interactions. Hence, ecological and evolutionary dynamics can be tightly linked and important to predict future dynamics, but our understanding of eco-evolutionary dynamics is still in its infancy and there is a significant gap between theoretical predictions and empirical tests. Empirical studies have demonstrated that the presence of genetic variation can dramatically change ecological dynamics, whereas theoretical studies predict that eco-evolutionary dynamics depend on the details of the genetic variation, such as the form of a tradeoff among genotypes, which can be more important than the presence or absence of the genetic variation. Using a predator-prey (rotifer-algal) experimental system in laboratory microcosms, we studied how different forms of a tradeoff between prey defense and growth affect eco-evolutionary dynamics. Our experimental results show for the first time to our knowledge that different forms of the tradeoff produce remarkably divergent eco-evolutionary dynamics, including near fixation, near extinction, and coexistence of algal genotypes, with quantitatively different population dynamics. A mathematical model, parameterized from completely independent experiments, explains the observed dynamics. The results suggest that knowing the details of heritable trait variation and covariation within a population is essential for understanding how evolution and ecology will interact and what form of eco-evolutionary dynamics will result.

  6. Dissecting the nanoscale distributions and functions of microtubule-end-binding proteins EB1 and ch-TOG in interphase HeLa cells.

    Directory of Open Access Journals (Sweden)

    Satoko Nakamura

    Full Text Available Recently, the EB1 and XMAP215/TOG families of microtubule binding proteins have been demonstrated to bind autonomously to the growing plus ends of microtubules and regulate their behaviour in in vitro systems. However, their functional redundancy or difference in cells remains obscure. Here, we compared the nanoscale distributions of EB1 and ch-TOG along microtubules using high-resolution microscopy techniques, and also their roles in microtubule organisation in interphase HeLa cells. The ch-TOG accumulation sites protruded ∼100 nm from the EB1 comets. Overexpression experiments showed that ch-TOG and EB1 did not interfere with each other's localisation, confirming that they recognise distinct regions at the ends of microtubules. While both EB1 and ch-TOG showed similar effects on microtubule plus end dynamics and additively increased microtubule dynamicity, only EB1 exhibited microtubule-cell cortex attachment activity. These observations indicate that EB1 and ch-TOG regulate microtubule organisation differently via distinct regions in the plus ends of microtubules.

  7. Nitrogen Additions Affect Root Dynamics in a Boreal Forest Ecosystem

    Science.gov (United States)

    Turner, K. M.; Treseder, K. K.

    2004-12-01

    As with many ecosystems, North American boreal forests are increasingly subjected to anthropogenic nitrogen deposition. To examine potential effects on plant growth, we created nitrogen fertilization plots in three sites along an Alaskan fire chronosequence composed of forests aged 5, 17, and 80 years. Each site had been exposed to two years of nitrogen fertilization, with four control plots and four nitrogen plots per site. General observations indicate that aboveground net primary productivity appears to be nitrogen limited in each site. We hypothesized that nitrogen fertilization would positively influence root dynamics as well, with nitrogen additions resulting in an increase in standing root biomass and length. To test our hypothesis, we used a minirhizotron camera to collect sequential images of roots in the top 10 cm of soil in both nitrogen fertilized and control plots in each site. Images were collected monthly during the growing season, with a total of five sampling times between May 2003 and May 2004. We then analyzed the images with WinRhizotron root measurement software. Nitrogen fertilization had varying effects on root biomass among the three sites, with a significant site by N interaction (P = 0.039). A decrease in root biomass was observed in the 5 and 80 year old sites, dropping from 207 g/m2 to 79 g/m2 and from 230 g/m2 to 129 g/m2 for the youngest and oldest sites, respectively. In contrast, root biomass increased from 52 g/m2 to 107 g/m2 in the 17 year old site. (Values are for the top 10 cm of soil only, and likely underestimate total root stocks.) Patterns in standing root lengths diverged from those of root biomass, with a 2.5-fold overall increase under nitrogen fertilization across all sites (P = 0.004). There were no significant differences among sites in nitrogen response. Standing root biomass and length differed from one another in their responses to nitrogen fertilization because nitrogen additions decreased specific root weight (as g

  8. The temporal dynamics of cortisol and affective states in depressed and non-depressed individuals

    NARCIS (Netherlands)

    Booij, Sanne H.; Bos, Elisabeth H.; de Jonge, Peter; Oldehinkel, Albertine J.

    2016-01-01

    Objectives: Cortisol levels have been related to mood disorders at the group level, but not much is known about how cortisol relates to affective states within individuals over time. We examined the temporal dynamics of cortisol and affective states in depressed and non-depressed individuals in dail

  9. A Mechanism for Cytoplasmic Streaming: Kinesin-Driven Alignment of Microtubules and Fast Fluid Flows.

    Science.gov (United States)

    Monteith, Corey E; Brunner, Matthew E; Djagaeva, Inna; Bielecki, Anthony M; Deutsch, Joshua M; Saxton, William M

    2016-05-10

    The transport of cytoplasmic components can be profoundly affected by hydrodynamics. Cytoplasmic streaming in Drosophila oocytes offers a striking example. Forces on fluid from kinesin-1 are initially directed by a disordered meshwork of microtubules, generating minor slow cytoplasmic flows. Subsequently, to mix incoming nurse cell cytoplasm with ooplasm, a subcortical layer of microtubules forms parallel arrays that support long-range, fast flows. To analyze the streaming mechanism, we combined observations of microtubule and organelle motions with detailed mathematical modeling. In the fast state, microtubules tethered to the cortex form a thin subcortical layer and undergo correlated sinusoidal bending. Organelles moving in flows along the arrays show velocities that are slow near the cortex and fast on the inward side of the subcortical microtubule layer. Starting with fundamental physical principles suggested by qualitative hypotheses, and with published values for microtubule stiffness, kinesin velocity, and cytoplasmic viscosity, we developed a quantitative coupled hydrodynamic model for streaming. The fully detailed mathematical model and its simulations identify key variables that can shift the system between disordered (slow) and ordered (fast) states. Measurements of array curvature, wave period, and the effects of diminished kinesin velocity on flow rates, as well as prior observations on f-actin perturbation, support the model. This establishes a concrete mechanistic framework for the ooplasmic streaming process. The self-organizing fast phase is a result of viscous drag on kinesin-driven cargoes that mediates equal and opposite forces on cytoplasmic fluid and on microtubules whose minus ends are tethered to the cortex. Fluid moves toward plus ends and microtubules are forced backward toward their minus ends, resulting in buckling. Under certain conditions, the buckling microtubules self-organize into parallel bending arrays, guiding varying directions

  10. Effects of tertiary amine local anesthetics on the assembly and disassembly of brain microtubules in vitro.

    Science.gov (United States)

    Genna, J M; Coffe, G; Pudles, J

    1980-09-01

    From kinetic and electron microscopy studies on the effects of procaine, tetracaine and dibucaine on the polymerization and depolymerization of the microtubules isolated from pig and rat brains the following results were obtained. 1. Procaine or tetracaine, at the concentration range of 0.5--20 mM and of 0.5--5 mM respectively, increases the rate of tubulin polymerization (24 degrees C or 37 degrees C) and of microtubule depolymerization (4 degrees C) as a linear function of the concentration of the anesthetics, while identical amounts of microtubules are formed. In the absence of microtubule-associated proteins the polymerization of tubulin is not induced by 10 mM procaine, furthermore, the critical concentration of microtubule proteins necessary for assembly into microtubules is not affected at this concentration level of the anesthetic. This suggests that procaine affects not the nucleation, but rather the elongation process. 2. Dibucaine, from 0.5 mM to 3 mM increases the lag time of the polymerization reaction, while from 0.5 mM to 2 mM it linearly decreases both tubulin polymerization (24 degrees C) and microtubule depolymerization (4 degrees C) rates. Dibucaine, up to mM concentration, does not affect the extent of tubulin polymerization; however, above this concentration it induces the formation of amorphous aggregates. 3. Procaine or tetracaine enhances the depolymerizing effect of calcium on microtubules. The half-maximal values for the depolymerizing effect of calcium were 0.96, 0.71 and 0.51 mM for the control, in the presence of 10 mM procaine and 5 mM tetracaine respectively.

  11. Joined at the hip: kinetochores, microtubules, and spindle assembly checkpoint signaling.

    Science.gov (United States)

    Sacristan, Carlos; Kops, Geert J P L

    2015-01-01

    Error-free chromosome segregation relies on stable connections between kinetochores and spindle microtubules. The spindle assembly checkpoint (SAC) monitors such connections and relays their absence to the cell cycle machinery to delay cell division. The molecular network at kinetochores that is responsible for microtubule binding is integrated with the core components of the SAC signaling system. Molecular-mechanistic understanding of how the SAC is coupled to the kinetochore-microtubule interface has advanced significantly in recent years. The latest insights not only provide a striking view of the dynamics and regulation of SAC signaling events at the outer kinetochore but also create a framework for understanding how that signaling may be terminated when kinetochores and microtubules connect.

  12. [The role of cortical microtubules in moss protonemal cells during dehydration/rehydration cycle].

    Science.gov (United States)

    Chen, Zhi-Ling; Ouyang, Hao-Miao; Liu, Xiang-Lin; Xia, Gui-Xian

    2003-05-01

    Plant cells response to water deficit through a variety of physiological processes. In this work, we studied the function of microtubule cytoskeleton during dehydration/rehydration cycle in moss (Atrichum undulatum) protonemal cells as a model system. The morphological and cytological change of protonemal cells during dehydration and rehydration cycle were first investigated. Under normal conditions, protonemal cells showed bright green colour and appeared wet and fresh. Numerous chloroplasts distributed regularly throughout the cytoplasm in each cell. After dehydration treatment, protonemal cells lost most of their chlorophylls and turned to look yellow and dry. In addition, dehydration caused plasmolysis in these cells. Upon rehydration, the cells could recover completely from the dehydrated state. These results indicated that moss had a remarkable intrinsic ability to survive from the extreme drought stress. Microtubule, an important component of cytoskeleton, is considered to play crucial roles in the responses to some environmental stresses such as cold and light. To see if it is also involved in the drought tolerance, dynamic organization of microtubules in protonemal cells of Atrichum undulatum subjected to drought and rehydration were examined by indirect immunofluorescence combined with confocal lasersharp scanning microscopy. The cortical microtubules were arranged into a fine structure with a predominant orientation parallel to the long axis of the cells in the control cells. After dehydration, the microtubule organization was remarkablly altered and the fine microtubule structure disappeared whereas some thicker cables formed. When the cells were grown under rehydration conditions, the fine microtubule arrays reappeared. These results provided a piece of evidence that microtubules play a role in the cellular responses to drought stress in moss. Furthermore, we analyzed the effects of the microtubule-disrupting agent colchicine on the morphology recovery

  13. Movement of chromosomes with severed kinetochore microtubules.

    Science.gov (United States)

    Forer, Arthur; Johansen, Kristen M; Johansen, Jørgen

    2015-05-01

    Experiments dating from 1966 and thereafter showed that anaphase chromosomes continued to move poleward after their kinetochore microtubules were severed by ultraviolet microbeam irradiation. These observations were initially met with scepticism as they contradicted the prevailing view that kinetochore fibre microtubules pulled chromosomes to the pole. However, recent experiments using visible light laser microbeam irradiations have corroborated these earlier experiments as anaphase chromosomes again were shown to move poleward after their kinetochore microtubules were severed. Thus, multiple independent studies using different techniques have shown that chromosomes can indeed move poleward without direct microtubule connections to the pole, with only a kinetochore 'stub' of microtubules. An issue not yet settled is: what propels the disconnected chromosome? There are two not necessarily mutually exclusive proposals in the literature: (1) chromosome movement is propelled by the kinetochore stub interacting with non-kinetochore microtubules and (2) chromosome movement is propelled by a spindle matrix acting on the stub. In this review, we summarise the data indicating that chromosomes can move with severed kinetochore microtubules and we discuss proposed mechanisms for chromosome movement with severed kinetochore microtubules.

  14. Microtubules and associated molecular motors in Neurospora crassa.

    Science.gov (United States)

    Mouriño-Pérez, Rosa Reyna; Riquelme, Meritxell; Callejas-Negrete, Olga Alicia; Galván-Mendoza, José Iván

    2016-01-01

    The cytoskeleton provides structure, shape and movement to various cells. Microtubules (MTs) are tubular structures made of α and β-tubulin heterodimers organized in 13 protofilaments, forming a hollow cylinder. A vast group of MT-associated proteins determines the function, behavior and interaction of the MTs with other cellular components. Among these proteins, molecular motors such as the dynein-dynactin complex and kinesin superfamily play roles in MT organization and organelle transport. This article focuses on the MT cytoskeleton and associated molecular motors in the filamentous fungus Neurospora crassa In addition to reviewing current available information for this fungus and contrasting it with knowledge of other fungal species, we present new experimental results that support the role of dynein, dynactin and conventional kinesin in MT organization, dynamics and transport of subcellular structures (nuclei and secretory vesicles). In wild type hyphae of N. crassa, cytoplasmic MTs are arranged longitudinally along hyphae and display a helical curvature. They interlace with one another to form a network throughout the cytoplasm. N. crassa dynein and dynactin mutants have a scant and disorganized MT cytoskeleton, an erratic and reduced Spitzenkörper (Spk) and distorted hyphal morphology. In contrast, hyphae of mutants with defective conventional kinesin exhibit only minor disruptions in MT and Spk organization. Although nuclear positioning is affected in all mutants, the MT-associated motor proteins are not major contributors to nuclear movement during hyphal growth. Cytoplasmic bulk flow is the vehicle for nuclear displacement in growing hyphal regions of N. crassa Motors are involved in nuclei saltatory movements in both retrograde or anterograde direction. In the dynein and kinesin mutants, micro and macrovesicles can reach the Spk, although growth is slightly impaired and the Spk displays an erratic path. Hyphal growth requires MTs, and their associated

  15. Effects of 3-repeat tau on taxol mobility through microtubules

    Science.gov (United States)

    Park, Hyunjoo; Fygenson, Deborah; Kim, Mahn Won

    2005-03-01

    Both the anti-cancer drug taxol and the microtubule-associated protein tau suppress dynamics of microtubules (MT). We have observed taxol mobility with full-length 3-repeat tau, one of six tau isoforms, using fluorescence recovery after photobleaching (FRAP) on MTs and compare with earlier results on recombinant full-length adult 4-repeat tau. Taxol mobility becomes highly sensitive to taxol concentration in the presence of 3-repeat tau (up to 1:1 molar ratio) as it does in the presence of 4-repeat tau, but is 2 to 3 times faster at low taxol concentrations. Fitting to a mean-field binding reaction model [J.L. Ross et.al, PNAS 101:12910-5 (2004)] suggests that the presence of 3-repeat tau enhances taxol movement through pores in the MT walls.

  16. Mobility of Taxol in Microtubule Bundles

    Science.gov (United States)

    Ross, J.

    2003-06-01

    Mobility of taxol inside microtubules was investigated using fluorescence recovery after photobleaching (FRAP) on flow-aligned bundles. Bundles were made of microtubules with either GMPCPP or GTP at the exchangeable site on the tubulin dimer. Recovery times were sensitive to bundle thickness and packing, indicating that taxol molecules are able to move laterally through the bundle. The density of open binding sites along a microtubule was varied by controlling the concentration of taxol in solution for GMPCPP samples. With > 63% sites occupied, recovery times were independent of taxol concentration and, therefore, inversely proportional to the microscopic dissociation rate, k_{off}. It was found that 10*k_{off} (GMPCPP) ~ k_{off} (GTP), consistent with, but not fully accounting for, the difference in equilibrium constants for taxol on GMPCPP and GTP microtubules. With taxol along the microtubule interior is hindered by rebinding events when open sites are within ~7 nm of each other.

  17. TRESK background K(+ channel is inhibited by PAR-1/MARK microtubule affinity-regulating kinases in Xenopus oocytes.

    Directory of Open Access Journals (Sweden)

    Gabriella Braun

    Full Text Available TRESK (TWIK-related spinal cord K(+ channel, KCNK18 is a major background K(+ channel of sensory neurons. Dominant-negative mutation of TRESK is linked to familial migraine. This important two-pore domain K(+ channel is uniquely activated by calcineurin. The calcium/calmodulin-dependent protein phosphatase directly binds to the channel and activates TRESK current several-fold in Xenopus oocytes and HEK293 cells. We have recently shown that the kinase, which is responsible for the basal inhibition of the K(+ current, is sensitive to the adaptor protein 14-3-3. Therefore we have examined the effect of the 14-3-3-inhibited PAR-1/MARK, microtubule-associated-protein/microtubule affinity-regulating kinase on TRESK in the Xenopus oocyte expression system. MARK1, MARK2 and MARK3 accelerated the return of TRESK current to the resting state after the calcium-dependent activation. Several other serine-threonine kinase types, generally involved in the modulation of other ion channels, failed to influence TRESK current recovery. MARK2 phosphorylated the primary determinant of regulation, the cluster of three adjacent serine residues (S274, 276 and 279 in the intracellular loop of mouse TRESK. In contrast, serine 264, the 14-3-3-binding site of TRESK, was not phosphorylated by the kinase. Thus MARK2 selectively inhibits TRESK activity via the S274/276/279 cluster, but does not affect the direct recruitment of 14-3-3 to the channel. TRESK is the first example of an ion channel phosphorylated by the dynamically membrane-localized MARK kinases, also known as general determinants of cellular polarity. These results raise the possibility that microtubule dynamics is coupled to the regulation of excitability in the neurons, which express TRESK background potassium channel.

  18. Laminin/β1 integrin signal triggers axon formation by promoting microtubule assembly and stabilization

    Institute of Scientific and Technical Information of China (English)

    Wen-Liang Lei; Shi-Ge Xing; Cai-Yun Deng; Xiang-Chun Ju; Xing-Yu Jiang; Zhen-Ge Luo

    2012-01-01

    Axon specification during neuronal polarization is closely associated with increased microtubule stabilization in one of the neurites of unpolarized neuron,but how this increased microtubule stability is achieved is unclear.Here,we show that extracellular matrix (ECM) component laminin promotes neuronal polarization via regulating directional microtubule assembly through β1 integrin (Itgb1).Contact with laminin coated on culture substrate or polystyrene beads was sufficient for axon specification of undifferentiated neurites in cultured hippocampal neurons and cortical slices.Active Itgb1 was found to be concentrated in laminin-contacting neurites.Axon formation was promoted and abolished by enhancing and attenuating Itgbl signaling,respectively.Interestingly,laminin contact promoted plus-end microtubule assembly in a manner that required Itgbl.Moreover,stabilizing microtubules partially prevented polarization defects caused by ltgbl downregulation.Finally,genetic ablation of ltgbl in dorsal telencephalic progenitors caused deficits in axon development of cortical pyramidal neurons.Thus,laminin/Itgb1 signaling plays an instructive role in axon initiation and growth,both in vitro and in vivo,through the regulation of microtubule assembly.This study has established a linkage between an extrinsic factor and intrinsic cytoskeletai dynamics during neuronal polarization.

  19. Symmetry Breaking in an Edgeless Epithelium by Fat2-Regulated Microtubule Polarity

    Directory of Open Access Journals (Sweden)

    Dong-Yuan Chen

    2016-05-01

    Full Text Available Planar cell polarity (PCP information is a critical determinant of organ morphogenesis. While PCP in bounded epithelial sheets is increasingly well understood, how PCP is organized in tubular and acinar tissues is not. Drosophila egg chambers (follicles are an acinus-like “edgeless epithelium” and exhibit a continuous, circumferential PCP that does not depend on pathways active in bounded epithelia; this follicle PCP directs formation of an ellipsoid rather than a spherical egg. Here, we apply an imaging algorithm to “unroll” the entire 3D tissue surface and comprehensively analyze PCP onset. This approach traces chiral symmetry breaking to plus-end polarity of microtubules in the germarium, well before follicles form and rotate. PCP germarial microtubules provide chiral information that predicts the direction of whole-tissue rotation as soon as independent follicles form. Concordant microtubule polarity, but not microtubule alignment, requires the atypical cadherin Fat2, which acts at an early stage to translate plus-end bias into coordinated actin-mediated collective cell migration. Because microtubules are not required for PCP or migration after follicle rotation initiates, while dynamic actin and extracellular matrix are, polarized microtubules lie at the beginning of a handoff mechanism that passes early chiral PCP of the cytoskeleton to a supracellular planar polarized extracellular matrix and elongates the organ.

  20. Early stages of spindle formation and independence of chromosome and microtubule cycles in Haemanthus endosperm.

    Science.gov (United States)

    Smirnova, E A; Bajer, A S

    1998-01-01

    We analyzed transformation of the interphase microtubular cytoskeleton into the prophase spindle and followed the pattern of spindle axis determination. Microtubules in endosperm of the higher plant Haemanthus (Scadoxus) were stained by the immunogold and immunogold silver-enhanced methods. Basic structural units involved in spindle morphogenesis were "microtubule converging centers." We emphasized the importance of relative independence of chromosomal and microtubular cycles, and the influence of these cycles on the progress of mitosis. Cells with moderately desynchronized cycles were functional, but extreme desynchronization led to aberrant mitosis. There were three distinct phases of spindle development. The first one comprised interphase and early to mid-prophase. During this phase, the interphase microtubule meshwork radiating from the nuclear surface into the cytoplasm rearranged and formed a dense microtubule cage around the nucleus. The second phase comprised mid to late prophase, and resulted in the formation of normal (bipolar) or transitory aberrant (apolar or multipolar) prophase spindles. The third phase comprised late prophase with prometaphase. The onset of prometaphase was accompanied by a rapid association of microtubule converging centers with kinetochores. In this stage aberrant spindles transformed invariably into bipolar ones. Lateral association of a few bipolar kinetochore fibers at early prometaphase established the core of the bipolar spindle and its alignment. We concluded that (1) spindle formation is a largely independent microtubular process modified by the chromosomal/kinetochore cycle; and (2) the initial polarity of the spindle is established by microtubule converging centers, which are a functional substitute of the centrosome/MTOC. We believe that the dynamics of microtubule converging centers is an expression of microtubule self-organization driven by motor proteins as proposed by Mitchison [1992: Philos. Trans. R. Soc. Lond. B

  1. Identification of a TPX2-like microtubule-associated protein in Drosophila.

    Directory of Open Access Journals (Sweden)

    Gohta Goshima

    Full Text Available Chromosome segregation during mitosis and meiosis relies on the spindle and the functions of numerous microtubule-associated proteins (MAPs. One of the best-studied spindle MAPs is the highly conserved TPX2, which has been reported to have characteristic intracellular dynamics and molecular activities, such as nuclear localisation in interphase, poleward movement in the metaphase spindle, microtubule nucleation, microtubule stabilisation, microtubule bundling, Aurora A kinase activation, kinesin-5 binding, and kinesin-12 recruitment. This protein has been shown to be essential for spindle formation in every cell type analysed so far. However, as yet, TPX2 homologues have not been found in the Drosophila genome. In this study, I found that the Drosophila protein Ssp1/Mei-38 has significant homology to TPX2. Sequence conservation was limited to the putative spindle microtubule-associated region of TPX2, and intriguingly, D-TPX2 (Ssp1/Mei-38 lacks Aurora A- and kinesin-5-binding domains, which are highly conserved in other animal and plant species, including many insects such as ants and bees. D-TPX2 uniformly localised to kinetochore microtubule-enriched regions of the metaphase spindle in the S2 cell line, and it had microtubule binding and bundling activities in vitro. In comparison with other systems, the contribution of D-TPX2 to cell division seems to be minor; live cell imaging of microtubules and chromosomes after RNAi knockdown identified significant delay in chromosome congression in only 18% of the cells. Thus, while this conserved spindle protein is present in Drosophila, other mechanisms may largely compensate for its spindle assembly and chromosome segregation functions.

  2. Decoherence Time of a Microtubule

    CERN Document Server

    Hiramatsu, T; Sakakibara, K; Hiramatsu, Takashi; Matsui, Tetsuo; Sakakibara, Kazuhiko

    2006-01-01

    We formulate and study a quantum field theory of a microtubule, a basic element of living cells. Following the quantum theory of consciousness by Hameroff and Penrose, we let the system to make self-reductions, and measure the decoherence time $\\tau_N$ (the mean interval between two successive reductions) of a cluster consisting of more than $N$ neighboring cells (tubulins). $\\tau_N$ is interpreted as an instance of the stream of consciousness. For a sufficiently small electron hopping amplitude, $\\tau_N$ obeys an exponential law, $\\tau_N \\sim \\exp(c' N)$, and may take realistic values $\\tau_N $ \\raisebox{-0.5ex} {$\\stackrel{>}{\\sim}$} $ 10^{-2}$ sec for $N \\raisebox{-0.5ex} {$\\stackrel{>}{\\sim}$} 1100$.

  3. Differences in the dynamics of affective and cognitive processing - An ERP study.

    Science.gov (United States)

    Mueller, Christina J; Fritsch, Nathalie; Hofmann, Markus J; Kuchinke, Lars

    2017-01-15

    A controversy in emotion research concerns the question of whether affective or cognitive primacy are evident in processing affective stimuli and the factors contributing to each alternative. Using electrophysiological recordings in an adapted visual oddball paradigm allowed tracking the dynamics of affective and cognitive effects. Stimuli consisted of face pictures displaying affective expressions with rare oddballs differing from frequent stimuli in either affective expression, structure (while frequent stimuli were shown frontally these deviants were turned sideways) or they differed on both dimensions, i.e. in affective expression and structure. Results revealed a defined sequence of differences in ERP amplitudes: For stimuli deviating in their affective expression only, P1 modulations ~100ms were evident, while affective differences of structure deviants were not evident before the N170 time window. All three types of deviants differed in P300 amplitudes, indicating integration of affective and structural information. These results encompass evidence for both, cognitive and affective primacy depending on stimulus properties. Specifically affective primacy is only visible when the respective facial features can be extracted with ease. When structural differences make face processing harder, however, cognitive primacy is brought forward.

  4. Two-state mechanochemical model for microtubule growth

    CERN Document Server

    Zhang, Yunxin

    2011-01-01

    In this study, a two-state mechanochemical model is presented to describe the dynamic properties of microtubule (MT) growth in cells. The MT switches between two states, assembly state and disassembly state. In assembly state, the growth of microtubule includes two processes: GTP-tubulin binding to the tip of protofilament (PF) and conformational change of PF, during which the penultimate GTP is hydrolyzed and the first tubulin unit that curls out the MT surface is rearranged into MT surface using the energy released from GTP hydrolysis. In disassembly state, the shortening of microtubule is also described by two processes, the release of GDP-tibulin from the tip of PF and one new tubulin unit curls out from the MT surface. Switches between these two states, which are usually called rescue and catastrophe, happen stochastically with external force dependent rates. Using this two-state model with parameters obtained by fitting the recent experimental data, detailed properties of MT growth are obtained, we find...

  5. A mechanism for reorientation of cortical microtubule arrays driven by microtubule severing.

    Science.gov (United States)

    Lindeboom, Jelmer J; Nakamura, Masayoshi; Hibbel, Anneke; Shundyak, Kostya; Gutierrez, Ryan; Ketelaar, Tijs; Emons, Anne Mie C; Mulder, Bela M; Kirik, Viktor; Ehrhardt, David W

    2013-12-06

    Environmental and hormonal signals cause reorganization of microtubule arrays in higher plants, but the mechanisms driving these transitions have remained elusive. The organization of these arrays is required to direct morphogenesis. We discovered that microtubule severing by the protein katanin plays a crucial and unexpected role in the reorientation of cortical arrays, as triggered by blue light. Imaging and genetic experiments revealed that phototropin photoreceptors stimulate katanin-mediated severing specifically at microtubule intersections, leading to the generation of new microtubules at these locations. We show how this activity serves as the basis for a mechanism that amplifies microtubules orthogonal to the initial array, thereby driving array reorientation. Our observations show how severing is used constructively to build a new microtubule array.

  6. Structural comparison of the Caenorhabditis elegans and human Ndc80 complexes bound to microtubules reveals distinct binding behavior

    Science.gov (United States)

    Wilson-Kubalek, Elizabeth M.; Cheeseman, Iain M.; Milligan, Ronald A.

    2016-01-01

    During cell division, kinetochores must remain tethered to the plus ends of dynamic microtubule polymers. However, the molecular basis for robust kinetochore–microtubule interactions remains poorly understood. The conserved four-subunit Ndc80 complex plays an essential and direct role in generating dynamic kinetochore–microtubule attachments. Here we compare the binding of the Caenorhabditis elegans and human Ndc80 complexes to microtubules at high resolution using cryo–electron microscopy reconstructions. Despite the conserved roles of the Ndc80 complex in diverse organisms, we find that the attachment mode of these complexes for microtubules is distinct. The human Ndc80 complex binds every tubulin monomer along the microtubule protofilament, whereas the C. elegans Ndc80 complex binds more tightly to β-tubulin. In addition, the C. elegans Ndc80 complex tilts more toward the adjacent protofilament. These structural differences in the Ndc80 complex between different species may play significant roles in the nature of kinetochore–microtubule interactions. PMID:26941333

  7. Centriolar CPAP/SAS-4 Imparts Slow Processive Microtubule Growth.

    Science.gov (United States)

    Sharma, Ashwani; Aher, Amol; Dynes, Nicola J; Frey, Daniel; Katrukha, Eugene A; Jaussi, Rolf; Grigoriev, Ilya; Croisier, Marie; Kammerer, Richard A; Akhmanova, Anna; Gönczy, Pierre; Steinmetz, Michel O

    2016-05-23

    Centrioles are fundamental and evolutionarily conserved microtubule-based organelles whose assembly is characterized by microtubule growth rates that are orders of magnitude slower than those of cytoplasmic microtubules. Several centriolar proteins can interact with tubulin or microtubules, but how they ensure the exceptionally slow growth of centriolar microtubules has remained mysterious. Here, we bring together crystallographic, biophysical, and reconstitution assays to demonstrate that the human centriolar protein CPAP (SAS-4 in worms and flies) binds and "caps" microtubule plus ends by associating with a site of β-tubulin engaged in longitudinal tubulin-tubulin interactions. Strikingly, we uncover that CPAP activity dampens microtubule growth and stabilizes microtubules by inhibiting catastrophes and promoting rescues. We further establish that the capping function of CPAP is important to limit growth of centriolar microtubules in cells. Our results suggest that CPAP acts as a molecular lid that ensures slow assembly of centriolar microtubules and, thereby, contributes to organelle length control.

  8. The smallest active fragment of microtubule-associated protein 4 and its interaction with microtubules in phosphate buffer.

    Science.gov (United States)

    Hashi, Yurika; Nagase, Lisa; Matsushima, Kazuyuki; Kotani, Susumu

    2012-01-01

    To analyze the interaction between microtubule-associated protein (MAP) 4 and microtubules physicochemically, a MAP4 active site fragment was designed for nuclear magnetic resonance (NMR) use. The fragment was bacterially expressed and purified to homogeneity. The buffer conditions for NMR were optimized to support microtubule assembly. The fragment was found to bind to microtubules under the optimized buffer conditions.

  9. Factors affecting the haptic filled-space illusion for dynamic touch

    NARCIS (Netherlands)

    Sanders, A.F.J.; Kappers, A.M.L.

    2009-01-01

    In the haptic filled-space illusion for active dynamic touch, observers move their fingertip across an unfilled extent or an extent filled with intermediate stimulations. Previous researchers have reported lengths of filled extents to be overestimated, but the parameters affecting the strength of th

  10. Dynamic Effects of Craving and Negative Affect on Adolescent Smoking Relapse

    NARCIS (Netherlands)

    Zundert, R.M.P. van; Ferguson, S.G.; Shiffman, S.; Engels, R.C.M.E.

    2012-01-01

    Objective: This study examined dynamic effects of daily variations in craving and negative affect on the next-day risk of the first lapse and relapse among 149 adolescent daily smokers who achieved at least 24 hr of abstinence. Method: Participants completed real-time assessments of their smoking, c

  11. Environmental Factors Affecting Computer Assisted Language Learning Success: A Complex Dynamic Systems Conceptual Model

    Science.gov (United States)

    Marek, Michael W.; Wu, Wen-Chi Vivian

    2014-01-01

    This conceptual, interdisciplinary inquiry explores Complex Dynamic Systems as the concept relates to the internal and external environmental factors affecting computer assisted language learning (CALL). Based on the results obtained by de Rosnay ["World Futures: The Journal of General Evolution", 67(4/5), 304-315 (2011)], who observed…

  12. School Factors Explaining Achievement on Cognitive and Affective Outcomes : Establishing a Dynamic Model of Educational Effectiveness

    NARCIS (Netherlands)

    Creemers, Bert; Kyriakides, Leonidas

    2010-01-01

    The dynamic model of educational effectiveness defines school level factors associated with student outcomes. Emphasis is given to the two main aspects of policy, evaluation, and improvement in schools which affect quality of teaching and learning at both the level of teachers and students: a) teach

  13. Modeling affect dynamics : State-of-the-art and future challenges

    NARCIS (Netherlands)

    Hamaker, E.L.; Ceulemans, Eva; Grasman, R.P.P.P.; Tuerlinckx, Francis

    2015-01-01

    The current article aims to provide an up-to-date synopsis of available techniques to study affect dynamics using intensive longitudinal data (ILD). We do so by introducing the following eight dichotomies that help elucidate what kind of data one has, what process aspects are of interest, and what r

  14. Imperfect or Perfect Dynamic Bipolarity? The Case of Antonymous Affective Judgments

    Science.gov (United States)

    Vautier, Stephane; Steyer, Rolf; Jmel, Said; Raufaste, Eric

    2005-01-01

    How is affective change rated with positive adjectives such as good related to change rated with negative adjectives such as bad? Two nested perfect and imperfect forms of dynamic bipolarity are defined using latent change structural equation models based on tetrads of items. Perfect bipolarity means that latent change scores correlate -1.…

  15. One-parameter nonrelativistic supersymmetry for microtubules

    CERN Document Server

    Rosu, H C

    2003-01-01

    The simple supersymmetric model of Caticha [PRA 51, 4264 (1995)], as used by Rosu [PRE 55, 2038 (1997)] for microtubules, is generalized to the case of Mielnik's one-parameter nonrelativistic susy [JMP 25, 3387 (1984)

  16. The human kinesin-14 HSET tracks the tips of growing microtubules in vitro.

    Science.gov (United States)

    Braun, Marcus; Lansky, Zdenek; Bajer, Seweryn; Fink, Gero; Kasprzak, Andrzej A; Diez, Stefan

    2013-09-01

    Tip-tracking of kinesin-14 motor proteins is believed to be crucial for the assembly and maintenance of dynamic microtubule arrays. However, in contrast to other members of the kinesin-14 family, H. sapiens kinesin-14 HSET has so far never been observed to be prominently located at microtubule plus ends. Here, using an in vitro microtubule dynamics reconstitution assay we observe tip-tracking of GFP-HSET in the presence of H. sapiens EB1 (hsEB1). Tip-tracking depended on the SxIP-like motif in HSET as well as on the EB homology domain in hsEB1. D. melanogaster Ncd and S. pombe Klp2 tip-tracking reconstitution assays accompanied by kinesin-14 amino acid sequence comparisons suggest that SxIP-like motif mediated tip-tracking dependent on EB family proteins is conserved in the kinesin-14 family of molecular motors.

  17. Oxidative stress decreases microtubule growth and stability in ventricular myocytes.

    Science.gov (United States)

    Drum, Benjamin M L; Yuan, Can; Li, Lei; Liu, Qinghang; Wordeman, Linda; Santana, L Fernando

    2016-04-01

    Microtubules (MTs) have many roles in ventricular myocytes, including structural stability, morphological integrity, and protein trafficking. However, despite their functional importance, dynamic MTs had never been visualized in living adult myocytes. Using adeno-associated viral vectors expressing the MT-associated protein plus end binding protein 3 (EB3) tagged with EGFP, we were able to perform live imaging and thus capture and quantify MT dynamics in ventricular myocytes in real time under physiological conditions. Super-resolution nanoscopy revealed that EB1 associated in puncta along the length of MTs in ventricular myocytes. The vast (~80%) majority of MTs grew perpendicular to T-tubules at a rate of 0.06μm∗s(-1) and growth was preferentially (82%) confined to a single sarcomere. Microtubule catastrophe rate was lower near the Z-line than M-line. Hydrogen peroxide increased the rate of catastrophe of MTs ~7-fold, suggesting that oxidative stress destabilizes these structures in ventricular myocytes. We also quantified MT dynamics after myocardial infarction (MI), a pathological condition associated with increased production of reactive oxygen species (ROS). Our data indicate that the catastrophe rate of MTs increases following MI. This contributed to decreased transient outward K(+) currents by decreasing the surface expression of Kv4.2 and Kv4.3 channels after MI. On the basis of these data, we conclude that, under physiological conditions, MT growth is directionally biased and that increased ROS production during MI disrupts MT dynamics, decreasing K(+) channel trafficking.

  18. Motor function in interpolar microtubules during metaphase

    OpenAIRE

    Deutsch, J. M.; Lewis, Ian P.

    2013-01-01

    We analyze experimental observations of microtubules undergoing small fluctuations about a "balance point" when mixed in solution of two different kinesin motor proteins, KLP61F and Ncd. It has been proposed that the microtubule movement is due to stochastic variations in the densities of the two species of motor proteins. We test this hypothesis here by showing how it maps onto a one-dimensional random walk in a random environment. Our estimate of the amplitude of the fluctuations agrees wit...

  19. Patient affect experiencing following therapist interventions in short-term dynamic psychotherapy.

    Science.gov (United States)

    Town, Joel M; Hardy, Gillian E; McCullough, Leigh; Stride, Chris

    2012-01-01

    The aim of this research was to examine the relationship between therapist interventions and patient affect responses in Short-Term Dynamic Psychotherapy (STDP). The Affect Experiencing subscale from the Achievement of Therapeutic Objectives Scale (ATOS) was adapted to measure individual immediate affect experiencing (I-AES) responses in relation to therapist interventions coded within the preceding speaking turn, using the Psychotherapy Interaction Coding (PIC) system. A hierarchical linear modelling procedure was used to assess the change in affect experiencing and the relationship between affect experiencing and therapist interventions within and across segments of therapy. Process data was taken from six STDP cases; in total 24 hours of video-taped sessions were examined. Therapist interventions were found to account for a statistically significant amount of variance in immediate affect experiencing. Higher levels of immediate affect experiencing followed the therapist's use of Confrontation, Clarification and Support compared to Questions, Self-disclosure and Information interventions. Therapist Confrontation interventions that attempted to direct pressure towards either the visceral experience of affect or a patient's defences against feelings led to the highest levels of immediate affect experiencing. The type of therapist intervention accounts for a small but significant amount of the variation observed in a patient's immediate emotional arousal. Empirical findings support clinical theory in STDP that suggests strategic verbal responses promote the achievement of this specific therapeutic objective.

  20. TIPsy tour guides: How microtubule plus-end tracking proteins (+TIPs facilitate axon guidance

    Directory of Open Access Journals (Sweden)

    Elizabeth A Bearce

    2015-06-01

    Full Text Available The growth cone is a dynamic cytoskeletal vehicle, which drives the end of a developing axon. It serves to interpret and navigate through the complex landscape and guidance cues of the early nervous system. The growth cone’s distinctive cytoskeletal organization offers a fascinating platform to study how extracellular cues can be translated into mechanical outgrowth and turning behaviors. While many studies of cell motility highlight the importance of actin networks in signaling, adhesion, and propulsion, both seminal and emerging works in the field have highlighted a unique and necessary role for microtubules in growth cone navigation. Here, we focus on the role of singular pioneer microtubules, which extend into the growth cone periphery and are regulated by a diverse family of microtubule plus-end tracking proteins (+TIPs. These +TIPs accumulate at the dynamic ends of microtubules, where they are well-positioned to encounter and respond to key signaling events downstream of guidance receptors, catalyzing immediate changes in microtubule stability and actin cross-talk, that facilitate both axonal outgrowth and turning events.

  1. Factors affecting the dynamic response of pre-stressed anchors after transient excitation

    Institute of Scientific and Technical Information of China (English)

    Xu Huijun; Li Qingfeng

    2011-01-01

    The wide application of pre-stressed bolting technology in coal mine tunnels has made the nondestructive stress wave reflection method of determining bolting quality an important one.The effect of the support plate on the dynamic response of the pre-stressed anchor is of particular interest.A theoreticalanalysis and numerical simulations are used to identify the factors affecting the contact stress between the support plate and the rock wall.A formula allowing the calculation of contact stress is presented.Stress wave propagation through the nut,support plate,and rock wall are predicted.The dynamic response signals were measured in the field using prestressed anchors pre-tightened to different torques.The effects from the support plate on the dynamic response were recorded and the results compared to the predictions of pre-stressed anchor.This work provides a theoretical reference for the signal processing of dynamic reflected wave signals in anchor bolts.

  2. An Investigation of Factors Affecting Elementary School Students’ BMI Values Based on the System Dynamics Modeling

    Directory of Open Access Journals (Sweden)

    Tian-Syung Lan

    2014-01-01

    Full Text Available This study used system dynamics method to investigate the factors affecting elementary school students’ BMI values. The construction of the dynamic model is divided into the qualitative causal loop and the quantitative system dynamics modeling. According to the system dynamics modeling, this study consisted of research on the four dimensions: student’s personal life style, diet-relevant parenting behaviors, advocacy and implementation of school nutrition education, and students’ peer interaction. The results of this study showed that students with more adequate health concepts usually have better eating behaviors and consequently have less chance of becoming obese. In addition, this study also verified that educational attainment and socioeconomic status of parents have a positive correlation with students’ amounts of physical activity, and nutrition education has a prominent influence on changing students’ high-calorie diets.

  3. The role of microtubule-associated protein 1B in axonal growth and neuronal migration in the central nervous system

    Institute of Scientific and Technical Information of China (English)

    Maoguang Yang; Xiaoyu Yang; Minfei Wu; Peng Xia; Chunxin Wang; Peng Yan; Qi Gao; Jian Liu; Haitao Wang; Xingwei Duan

    2012-01-01

    In this review, we discuss the role of microtubule-associated protein 1B (MAP1B) and its phosphorylation in axonal development and regeneration in the central nervous system. MAP1B exhibits similar functions during axonal development and regeneration. MAP1B and phosphorylated MAP1B in neurons and axons maintain a dynamic balance between cytoskeletal components, and regulate the stability and interaction of microtubules and actin to promote axonal growth, neural connectivity and regeneration in the central nervous system.

  4. Pseudo-Spin Model for the Cytoskeletal Microtubule Surface

    Institute of Scientific and Technical Information of China (English)

    CHEN Ying; QIU Xi-Jun; LI Ru-Xin

    2004-01-01

    @@ Due to the inherent symmetry structures and the electric properties in the microtubule (MT), we treat the MT as a one-dimensional ferroelectric system and describe the nonlinear dynamics of dimer electric dipoles in one protofilament of the MT by virtue of the double-well potential. Consequently, the physical problem has been mapped onto the pseudo-spin system, and the mean-field approximation has been taken to obtain some physical results, including the expression for the phase transition temperature Tc and the estimated value of Tc (≈ 312 K).

  5. Role of Exchange Protein Activated by cAMP 1 in Regulating Rates of Microtubule Formation in Cystic Fibrosis Epithelial Cells.

    Science.gov (United States)

    Rymut, Sharon M; Ivy, Tracy; Corey, Deborah A; Cotton, Calvin U; Burgess, James D; Kelley, Thomas J

    2015-12-01

    The regulation of microtubule dynamics in cystic fibrosis (CF) epithelial cells and the consequences of reduced rates of microtubule polymerization on downstream CF cellular events, such as cholesterol accumulation, a marker of impaired intracellular transport, are explored here. It is identified that microtubules in both CF cell models and in primary CF nasal epithelial cells repolymerize at a slower rate compared with respective controls. Previous studies suggest a role for cAMP in modulating organelle transport in CF cells, implicating a role for exchange protein activated by cAMP (EPAC) 1, a regulator of microtubule elongation, as a potential mechanism. EPAC1 activity is reduced in CF cell models and in Cftr(-/-) mouse lung compared with respective non-CF controls. Stimulation of EPAC1 activity with the selective EPAC1 agonist, 8-cpt-2-O-Me-cAMP, stimulates microtubule repolymerization to wild-type rates in CF cells. EPAC1 activation also alleviates cholesterol accumulation in CF cells, suggesting a direct link between microtubule regulation and intracellular transport. To verify the relationship between transport and microtubule regulation, expression of the protein, tubulin polymerization-promoting protein, was knocked down in non-CF human tracheal (9/HTEo(-)) cells to mimic the microtubule dysregulation in CF cells. Transduced cells with short hairpin RNA targeting tubulin polymerization-promoting protein exhibit CF-like perinuclear cholesterol accumulation and other cellular manifestations of CF cells, thus supporting a role for microtubule regulation as a mechanism linking CFTR function to downstream cellular manifestation.

  6. Microtubules and Lis-1/NudE/dynein regulate invasive cell-on-cell migration in Drosophila.

    Directory of Open Access Journals (Sweden)

    Nachen Yang

    Full Text Available The environment through which cells migrate in vivo differs considerably from the in vitro environment where cell migration is often studied. In vivo many cells migrate in crowded and complex 3-dimensional tissues and may use other cells as the substratum on which they move. This includes neurons, glia and their progenitors in the brain. Here we use a Drosophila model of invasive, collective migration in a cellular environment to investigate the roles of microtubules and microtubule regulators in this type of cell movement. Border cells are of epithelial origin and have no visible microtubule organizing center (MTOC. Interestingly, microtubule plus-end growth was biased away from the leading edge. General perturbation of the microtubule cytoskeleton and analysis by live imaging showed that microtubules in both the migrating cells and the substrate cells affect movement. Also, whole-tissue and cell autonomous deletion of the microtubule regulator Stathmin had distinct effects. A screen of 67 genes encoding microtubule interacting proteins uncovered cell autonomous requirements for Lis-1, NudE and Dynein in border cell migration. Net cluster migration was decreased, with initiation of migration and formation of dominant front cell protrusion being most dramatically affected. Organization of cells within the cluster and localization of cell-cell adhesion molecules were also abnormal. Given the established role of Lis-1 in migrating neurons, this could indicate a general role of Lis-1/NudE, Dynein and microtubules, in cell-on-cell migration. Spatial regulation of cell-cell adhesion may be a common theme, consistent with observing both cell autonomous and non-autonomous requirements in both systems.

  7. Identifying Core Affect in Individuals from fMRI Responses to Dynamic Naturalistic Audiovisual Stimuli.

    Science.gov (United States)

    Kim, Jongwan; Wang, Jing; Wedell, Douglas H; Shinkareva, Svetlana V

    2016-01-01

    Recent research has demonstrated that affective states elicited by viewing pictures varying in valence and arousal are identifiable from whole brain activation patterns observed with functional magnetic resonance imaging (fMRI). Identification of affective states from more naturalistic stimuli has clinical relevance, but the feasibility of identifying these states on an individual trial basis from fMRI data elicited by dynamic multimodal stimuli is unclear. The goal of this study was to determine whether affective states can be similarly identified when participants view dynamic naturalistic audiovisual stimuli. Eleven participants viewed 5s audiovisual clips in a passive viewing task in the scanner. Valence and arousal for individual trials were identified both within and across participants based on distributed patterns of activity in areas selectively responsive to audiovisual naturalistic stimuli while controlling for lower level features of the stimuli. In addition, the brain regions identified by searchlight analyses to represent valence and arousal were consistent with previously identified regions associated with emotion processing. These findings extend previous results on the distributed representation of affect to multimodal dynamic stimuli.

  8. Identifying Core Affect in Individuals from fMRI Responses to Dynamic Naturalistic Audiovisual Stimuli

    Science.gov (United States)

    Kim, Jongwan; Wang, Jing; Wedell, Douglas H.

    2016-01-01

    Recent research has demonstrated that affective states elicited by viewing pictures varying in valence and arousal are identifiable from whole brain activation patterns observed with functional magnetic resonance imaging (fMRI). Identification of affective states from more naturalistic stimuli has clinical relevance, but the feasibility of identifying these states on an individual trial basis from fMRI data elicited by dynamic multimodal stimuli is unclear. The goal of this study was to determine whether affective states can be similarly identified when participants view dynamic naturalistic audiovisual stimuli. Eleven participants viewed 5s audiovisual clips in a passive viewing task in the scanner. Valence and arousal for individual trials were identified both within and across participants based on distributed patterns of activity in areas selectively responsive to audiovisual naturalistic stimuli while controlling for lower level features of the stimuli. In addition, the brain regions identified by searchlight analyses to represent valence and arousal were consistent with previously identified regions associated with emotion processing. These findings extend previous results on the distributed representation of affect to multimodal dynamic stimuli. PMID:27598534

  9. On the dynamic covariation between interpersonal behavior and affect: prediction from neuroticism, extraversion, and agreeableness.

    Science.gov (United States)

    Côté, S; Moskowitz, D S

    1998-10-01

    It was posited that the traits of Neuroticism, Extraversion, and Agreeableness are predictors of dynamic intraindividual processes involving interpersonal behavior and affect. Hypotheses derived from the behavioral concordance model that individuals with high scores on a trait would experience more positively valenced affect when engaging in behavior concordant with that trait than individuals with low scores on the trait were tested. Participants completed a questionnaire measure of the traits and reported on behavior and affect during interpersonal interactions using event-contingent sampling forms approximately 6 times a day for 20 days. Trait scores were related to indexes of the association between each dimension of interpersonal behavior and affect calculated for each individual. Previous findings concerning the trait of Agreeableness were replicated, and results strongly supported the behavioral concordance model for the trait of Neuroticism. Thus, at least some traits can provide information about intraindividual processes that vary over time.

  10. Rearrangements of microtubule cytoskeleton in stomatal closure of Arabidopsis induced by nitric oxide

    Institute of Scientific and Technical Information of China (English)

    ZHANG YongMei; WU ZhongYi; WANG XueChen; YU Rong

    2008-01-01

    NO (nitric oxide), known as a key signal molecule in plant, plays important roles in regulation of stomatal movement. In this study, microtubule dynamics and its possible mechanism in the NO signal pathway were investigated. The results were as follows: (ⅰ) In vivo stomatal aperture assays revealed that both vinblastine (microtubule-disrupting drug) and SNP (exogenous NO donor) prevented stomatal opening in the light, and vinblastine even could enhance the inhibitory effect of SNP, whereas taxol (a microtubule-stabilizing agent) was able to reduce this effect; (ⅱ) microtubules in the opening Arabi-dopsis guard cells expressing GFP:α-tubulin-6 (AtGFP:α-tubulin-6) were organized in parallel, straight and dense bundles, radiating from the ventral side to the dorsal side, and most of them were localized perpendicularly to the ventral wall; (ⅲ) under the same environmental conditions, treated with SNP for 30 min, the radial arrays of microtubules in guard cells began to break down, twisted partially and be-came oblique or exhibited a random pattern; (ⅳ) furthermore, the involvement of cytosolic Ca2+ in this event was tested. Stomatal aperture assays revealed that BAPTA-AM (a chelator of Ca2+) greatly sup-pressed the effect of NO on stomatal closure; however, it did not show the same function on stomatal closure induced by vinblastine. When BAPTA-AM was added to the SNP-pretreated solution, the SNP-induced disordered microtubulue cytoskeleton in guard cells underwent rearrangement in a time-dependent manner. After 30 min of treatment with BAPTA-AM, the cortical microtubules resumed the original radial distribution, almost the same as the control. All this indicates that NO may promote rearrangement of microtubule cytoskeleton via elevation of [Ca2+]cyt (free Ca2+ concentration in the cy-toplasm), finally leading to stomatal closure.

  11. [Disruption of organization of mitotic microtubules in root meristem cells of Allium cepa induced by chloral hydrate].

    Science.gov (United States)

    Smirnova, E A; Svetlitskaia, O M; Chentsov, Iu S

    2002-01-01

    Data are presented on the effect of chlorahydrate on microtubule organization in the root meristem of Allium cepa. Our studies show that an incomplete preprophase band commonly appears during G2-prophase transition, yet the major effect is the lack of perinuclear microtubules, leading to inhibition of the prophase spindle formation and transition to C-mitosis. Upon chloralhydrate treatment of metaphase cells, we found cells with chromosomes regularly aligned within the metaphase plate and differently disorganized mitotic spindles. Concurrently, C-metaphase cells with remnants of kinetochore fibers were present. In addition, normal bipolar and abnormal irregular types of chromosome segregation were detected, this representing multipolar and diffuse anaphases. The major difference between them is the presence of polar microtubules during multipolar anaphase, and their lacking during diffuse anaphase. Alternatively, microtubule clusters between segregated groups of chromosomes are typical for cells with diffuse anaphase. During bipolar anaphase, excessive aster-like microtubules emanate from the spindle poles, and in telophase accessory phragmoplasts are observed at the cell periphery. The formation of incomplete phragmoplasts was observed after normal bipolar and abnormal chromosome segregation. We conclude that chloralhydrate may affect the nuclear surface capability to initiate the growth of perinuclear microtubules, thus blocking the prophase spindle formation. It also disturbs the spatial interaction between microtubules, which is crucial for the formation and functioning of various microtubular systems (preprophase band, spindle and phragmoplast).

  12. Toward affect-inclusive models of cognitive dynamics: Coupling epistemological resources and emotions

    CERN Document Server

    Gupta, Ayush; Elby, Andrew

    2013-01-01

    Many prominent lines of research on student's reasoning and conceptual change within learning sciences and physics education research have not attended to the role of learners' affect or emotions in the dynamics of their conceptual reasoning. This is despite evidence from psychology and cognitive- and neuro- sciences that emotions are deeply integrated with cognition and documented associations in education research between emotions and academic performance. The few studies that have aimed to integrate emotions within models of learners' cognition, have mostly done so at a coarse grain size. In this manuscript, toward the long-term goal of incorporating emotions into fine-grained models of in-themoment cognitive dynamics, we present a case study of Judy, an undergraduate electrical engineering and physics major. We argue that a fine-grained aspect of Judy's affect, her annoyance at a particular kind of homework problem, stabilizes a context-dependent epistemological stance she displays, about an unbridgeable ...

  13. Dynamic cerebral autoregulatory capacity is affected early in Type 2 diabetes

    DEFF Research Database (Denmark)

    Kim, Y.S.; Immink, R.V.; Stok, W.J.;

    2008-01-01

    ) and impaired in subjects with DM+ (30+/-5 degrees ; Pblood pressure oscillations by affected dynamic cerebral autoregulation. The steady-state response of MCA V(mean) to postural change was comparable for all groups (-12+/-6% in CTRL subjects......, -15+/-6% in subjects with DM- and -15+/-7% in subjects with DM+). HbA(1c) (glycated haemoglobin) and the duration of diabetes, but not blood pressure, were determinants of transfer function phase. In conclusion, dysfunction of dynamic cerebral autoregulation in subjects with Type 2 diabetes appears......Type 2 diabetes is associated with an increased risk of endothelial dysfunction and microvascular complications with impaired autoregulation of tissue perfusion. Both microvascular disease and cardiovascular autonomic neuropathy may affect cerebral autoregulation. In the present study, we tested...

  14. Types and concentrations of metal ions affect local structure and dynamics of RNA

    Science.gov (United States)

    Wang, Jun; Xiao, Yi

    2016-10-01

    The roles that metal ions play in the structure and dynamics of RNA molecules are long-standing problems that have been studied extensively but are still not well understood. Here we show that metal ions have distributions around RNA molecules that strongly depend on the types and concentrations of the metal ions and also the electrostatic surface of the molecule. In particular, the ion distributions may not balance all the local electronegativity of the molecule. These ion distributions do not only greatly affect local structures but also lead to different local dynamics of RNA. We studied the effects of different ion solutions on the structure and dynamics of RNA by taking the pre Q1 riboswitch aptamer domain as an illustrative example and using molecular dynamics simulations. Since the local structures and dynamics of RNAs are important to their functions, our results also indicate that the selection of proper ion conditions is necessary to model them correctly, in contrast to the use of diverse ion solutions in current molecular dynamics simulations.

  15. beta-Dystroglycan modulates the interplay between actin and microtubules in human-adhered platelets.

    Science.gov (United States)

    Cerecedo, Doris; Cisneros, Bulmaro; Suárez-Sánchez, Rocío; Hernández-González, Enrique; Galván, Iván

    2008-05-01

    To maintain the continuity of an injured blood vessel, platelets change shape, secrete granule contents, adhere, aggregate, and retract in a haemostatic plug. Ordered arrays of microtubules, microfilaments, and associated proteins are responsible for these platelet responses. In full-spread platelets, microfilament bundles in association with other cytoskeleton proteins are anchored in focal contacts. Recent studies in migrating cells suggest that co-ordination and direct physical interaction of microtubules and actin network modulate adhesion development. In platelets, we have proposed a feasible association between these two cytoskeletal systems, as well as the participation of the dystrophin-associated protein complex, as part of the focal adhesion complex. The present study analysed the participation of microtubules and actin during the platelet adhesion process. Confocal microscopy, fluorescence resonance transfer energy and immunoprecipitation assays were used to provide evidence of a cross-talk between these two cytoskeletal systems. Interestingly, beta-dystroglycan was found to act as an interplay protein between actin and microtubules and an additional communication between these two cytoskeleton networks was maintained through proteins of focal adhesion complex. Altogether our data are indicative of a dynamic co-participation of actin filaments and microtubules in modulating focal contacts to achieve platelet function.

  16. The Effect of the Crocus Sativus L. Carotenoid, Crocin, on the Polymerization of Microtubules, in Vitro

    Directory of Open Access Journals (Sweden)

    Hossein Zarei Jaliani

    2013-01-01

    Full Text Available Objective(s: Crocin, as the main carotenoid of saffron, has shown anti-tumor activity both in vitro and in vivo. Crocin might interact with cellular proteins and modulate their functions, but the exact target of this carotenoid and the other compounds of the saffron have not been discovered yet. Microtubular proteins, as one of the most important proteins inside the cells, have several functions in nearly all kinds of cellular processes. The aim of this study was to investigate whether crocin affects microtubule polymerization and tubulin structure. Materials and Methods: Microtubules were extracted from sheep brains after two cycles of temperature-dependant assembly-disassembly in the polymerization buffer (PMG. Then phosphocellulose P11 column was used to prepare MAP-free tubulin. Turbidimetric assay of microtubules was performed by incubation of tubulins at 37 ºC in PIPES buffer. To investigate the intrinsic fluorescence spectra of tubulins, the emission spectra of tryptophans was monitored. To test the interaction of crocin with tubulin in more details, ANS has been used. Results: Crocin extremely affected the tubulin polymerization and structure. Ultraviolet spectroscopy indicated that crocin increased polymerization of microtubules by nearly a factor of two. Fluorescence spectroscopic data also pointed to significant conformational changes of tubulin. Conclusion: We showed that crocin increased tubulin polymerization and microtubule nucleation rate and this effect was concentration dependant. After entering cell, crocin can modulate cellular proteins and their functions. Concerning the results of this study, crocin would be able to affect several cell processes through interaction with tubulin proteins or microtubules.

  17. A study of microtubule dipole lattices

    Science.gov (United States)

    Nandi, Shubhendu

    Microtubules are cytoskeletal protein polymers orchestrating a host of important cellular functions including, but not limited to, cell support, cell division, cell motility and cell transport. In this thesis, we construct a toy-model of the microtubule lattice composed of vector Ising spins representing tubulin molecules, the building block of microtubules. Nearest-neighbor and next-to-nearest neighbor interactions are considered within an anisotropic dielectric medium. As a consequence of the helical topology, we observe that certain spin orientations render the lattice frustrated with nearest neighbor ferroelectric and next-to-nearest neighbor antiferroelectric bonds. Under these conditions, the lattice displays the remarkable property of stabilizing certain spin patterns that are robust to thermal fluctuations. We model this behavior in the framework of a generalized Ising model known as the J1 - J2 model and theoretically determine the set of stable patterns. Employing Monte-Carlo methods, we demonstrate the stability of such patterns in the microtubule lattice at human physiological temperatures. This suggests a novel biological mechanism for storing information in living organisms, whereby the tubulin spin (dipole moment) states become information bits and information gets stored in microtubules in a way that is robust to thermal fluctuations.

  18. Novel insights into mammalian embryonic neural stem cell division: focus on microtubules.

    Science.gov (United States)

    Mora-Bermúdez, Felipe; Huttner, Wieland B

    2015-12-01

    During stem cell divisions, mitotic microtubules do more than just segregate the chromosomes. They also determine whether a cell divides virtually symmetrically or asymmetrically by establishing spindle orientation and the plane of cell division. This can be decisive for the fate of the stem cell progeny. Spindle defects have been linked to neurodevelopmental disorders, yet the role of spindle orientation for mammalian neurogenesis has remained controversial. Here we explore recent advances in understanding how the microtubule cytoskeleton influences mammalian neural stem cell division. Our focus is primarily on the role of spindle microtubules in the development of the cerebral cortex. We also highlight unique characteristics in the architecture and dynamics of cortical stem cells that are tightly linked to their mode of division. These features contribute to setting these cells apart as mitotic "rule breakers," control how asymmetric a division is, and, we argue, are sufficient to determine the fate of the neural stem cell progeny in mammals.

  19. Prickle isoforms control the direction of tissue polarity by microtubule independent and dependent mechanisms

    Science.gov (United States)

    Sharp, Katherine A.; Axelrod, Jeffrey D.

    2016-01-01

    ABSTRACT Planar cell polarity signaling directs the polarization of cells within the plane of many epithelia. While these tissues exhibit asymmetric localization of a set of core module proteins, in Drosophila, more than one mechanism links the direction of core module polarization to the tissue axes. One signaling system establishes a polarity bias in the parallel, apical microtubules upon which vesicles containing core proteins traffic. Swapping expression of the differentially expressed Prickle isoforms, Prickle and Spiny-legs, reverses the direction of core module polarization. Studies in the proximal wing and the anterior abdomen indicated that this results from their differential control of microtubule polarity. Prickle and Spiny-legs also control the direction of polarization in the distal wing (D-wing) and the posterior abdomen (P-abd). We report here that this occurs without affecting microtubule polarity in these tissues. The direction of polarity in the D-wing is therefore likely determined by a novel mechanism independent of microtubule polarity. In the P-abd, Prickle and Spiny-legs interpret at least two directional cues through a microtubule-polarity-independent mechanism. PMID:26863941

  20. Prickle isoforms control the direction of tissue polarity by microtubule independent and dependent mechanisms.

    Science.gov (United States)

    Sharp, Katherine A; Axelrod, Jeffrey D

    2016-02-10

    Planar cell polarity signaling directs the polarization of cells within the plane of many epithelia. While these tissues exhibit asymmetric localization of a set of core module proteins, in Drosophila, more than one mechanism links the direction of core module polarization to the tissue axes. One signaling system establishes a polarity bias in the parallel, apical microtubules upon which vesicles containing core proteins traffic. Swapping expression of the differentially expressed Prickle isoforms, Prickle and Spiny-legs, reverses the direction of core module polarization. Studies in the proximal wing and the anterior abdomen indicated that this results from their differential control of microtubule polarity. Prickle and Spiny-legs also control the direction of polarization in the distal wing (D-wing) and the posterior abdomen (P-abd). We report here that this occurs without affecting microtubule polarity in these tissues. The direction of polarity in the D-wing is therefore likely determined by a novel mechanism independent of microtubule polarity. In the P-abd, Prickle and Spiny-legs interpret at least two directional cues through a microtubule-polarity-independent mechanism.

  1. Prickle isoforms control the direction of tissue polarity by microtubule independent and dependent mechanisms

    Directory of Open Access Journals (Sweden)

    Katherine A. Sharp

    2016-03-01

    Full Text Available Planar cell polarity signaling directs the polarization of cells within the plane of many epithelia. While these tissues exhibit asymmetric localization of a set of core module proteins, in Drosophila, more than one mechanism links the direction of core module polarization to the tissue axes. One signaling system establishes a polarity bias in the parallel, apical microtubules upon which vesicles containing core proteins traffic. Swapping expression of the differentially expressed Prickle isoforms, Prickle and Spiny-legs, reverses the direction of core module polarization. Studies in the proximal wing and the anterior abdomen indicated that this results from their differential control of microtubule polarity. Prickle and Spiny-legs also control the direction of polarization in the distal wing (D-wing and the posterior abdomen (P-abd. We report here that this occurs without affecting microtubule polarity in these tissues. The direction of polarity in the D-wing is therefore likely determined by a novel mechanism independent of microtubule polarity. In the P-abd, Prickle and Spiny-legs interpret at least two directional cues through a microtubule-polarity-independent mechanism.

  2. Aspergillus nidulans Dis1/XMAP215 protein AlpA localizes to spindle pole bodies and microtubule plus ends and contributes to growth directionality.

    Science.gov (United States)

    Enke, Cathrin; Zekert, Nadine; Veith, Daniel; Schaaf, Carolin; Konzack, Sven; Fischer, Reinhard

    2007-03-01

    The dynamics of cytoplasmic microtubules (MTs) is largely controlled by a protein complex at the MT plus end. In Schizosaccharomyces pombe and in filamentous fungi, MT plus end-associated proteins also determine growth directionality. We have characterized the Dis1/XMAP215 family protein AlpA from Aspergillus nidulans and show that it determines MT dynamics as well as hyphal morphology. Green fluorescent protein-tagged AlpA localized to MT-organizing centers (centrosomes) and to MT plus ends. The latter accumulation occurred independently of conventional kinesin or the Kip2-familiy kinesin KipA. alpA deletion strains were viable and only slightly temperature sensitive. Mitosis, nuclear migration, and nuclear positioning were not affected, but hyphae grew in curves rather than straight, which appeared to be an effect of reduced MT growth and dynamics.

  3. The dynamic interplay between appraisal and core affect in daily life

    Directory of Open Access Journals (Sweden)

    Peter eKuppens

    2012-10-01

    Full Text Available Appraisals and core affect are both considered central to the experience of emotion. In this study we examine the bidirectional relationships between these two components of emotional experience by examining how core affect changes following how people appraise events and how appraisals in turn change following how they feel in daily life. In an experience sampling study, participants recorded their core affect and appraisals of ongoing events; data were analyzed using cross-lagged multilevel modeling. Valence-appraisal relationships were found to be characterized by congruency: The same appraisals that were associated with a change in pleasure-displeasure (motivational congruency, other-agency, coping potential, and future expectancy, changed themselves as a function of pleasure-displeasure. In turn, mainly secondary appraisals of who is responsible and how one is able to cope with events were associated with changes in arousal, which itself is followed by changes in the future appraised relevance of events. These results integrate core affect and appraisal approaches to emotion by demonstrating the dynamic interplay of how appraisals are followed by changes in core affect which in turn change our basis for judging future events.

  4. How volatilities nonlocal in time affect the price dynamics in complex financial systems.

    Science.gov (United States)

    Tan, Lei; Zheng, Bo; Chen, Jun-Jie; Jiang, Xiong-Fei

    2015-01-01

    What is the dominating mechanism of the price dynamics in financial systems is of great interest to scientists. The problem whether and how volatilities affect the price movement draws much attention. Although many efforts have been made, it remains challenging. Physicists usually apply the concepts and methods in statistical physics, such as temporal correlation functions, to study financial dynamics. However, the usual volatility-return correlation function, which is local in time, typically fluctuates around zero. Here we construct dynamic observables nonlocal in time to explore the volatility-return correlation, based on the empirical data of hundreds of individual stocks and 25 stock market indices in different countries. Strikingly, the correlation is discovered to be non-zero, with an amplitude of a few percent and a duration of over two weeks. This result provides compelling evidence that past volatilities nonlocal in time affect future returns. Further, we introduce an agent-based model with a novel mechanism, that is, the asymmetric trading preference in volatile and stable markets, to understand the microscopic origin of the volatility-return correlation nonlocal in time.

  5. How volatilities nonlocal in time affect the price dynamics in complex financial systems.

    Directory of Open Access Journals (Sweden)

    Lei Tan

    Full Text Available What is the dominating mechanism of the price dynamics in financial systems is of great interest to scientists. The problem whether and how volatilities affect the price movement draws much attention. Although many efforts have been made, it remains challenging. Physicists usually apply the concepts and methods in statistical physics, such as temporal correlation functions, to study financial dynamics. However, the usual volatility-return correlation function, which is local in time, typically fluctuates around zero. Here we construct dynamic observables nonlocal in time to explore the volatility-return correlation, based on the empirical data of hundreds of individual stocks and 25 stock market indices in different countries. Strikingly, the correlation is discovered to be non-zero, with an amplitude of a few percent and a duration of over two weeks. This result provides compelling evidence that past volatilities nonlocal in time affect future returns. Further, we introduce an agent-based model with a novel mechanism, that is, the asymmetric trading preference in volatile and stable markets, to understand the microscopic origin of the volatility-return correlation nonlocal in time.

  6. Spatial and spatiotemporal variation in metapopulation structure affects population dynamics in a passively dispersing arthropod.

    Science.gov (United States)

    De Roissart, Annelies; Wang, Shaopeng; Bonte, Dries

    2015-11-01

    The spatial and temporal variation in the availability of suitable habitat within metapopulations determines colonization-extinction events, regulates local population sizes and eventually affects local population and metapopulation stability. Insights into the impact of such a spatiotemporal variation on the local population and metapopulation dynamics are principally derived from classical metapopulation theory and have not been experimentally validated. By manipulating spatial structure in artificial metapopulations of the spider mite Tetranychus urticae, we test to which degree spatial (mainland-island metapopulations) and spatiotemporal variation (classical metapopulations) in habitat availability affects the dynamics of the metapopulations relative to systems where habitat is constantly available in time and space (patchy metapopulations). Our experiment demonstrates that (i) spatial variation in habitat availability decreases variance in metapopulation size and decreases density-dependent dispersal at the metapopulation level, while (ii) spatiotemporal variation in habitat availability increases patch extinction rates, decreases local population and metapopulation sizes and decreases density dependence in population growth rates. We found dispersal to be negatively density dependent and overall low in the spatial variable mainland-island metapopulation. This demographic variation subsequently impacts local and regional population dynamics and determines patterns of metapopulation stability. Both local and metapopulation-level variabilities are minimized in mainland-island metapopulations relative to classical and patchy ones.

  7. A new cable truss support system for coal roadways affected by dynamic pressure

    Institute of Scientific and Technical Information of China (English)

    Hong Yan; Fulian He

    2012-01-01

    The support of coal roadways is seriously affected by intense dynamic pressures.This can lead to problems with large deformation of the roof and the two side walls of coal roadways.Rapid convergence of the walls and roof,a high damage rate to the bolts and cables,or even abrupt roof collapse or rib spalling can occur during the service period of these coal roadways.Analyzing the main support measures used in China leads to a proposed new cable truss supporting system.Thorough study of the entire structure shows the superiority of this design for roadways suffering under dynamic pressure.A corresponding mechanical model of the rock surrounding the cable truss system is described in this paper and formulas for calculating pre-tightening forces of the truss cable,and the minimum anchoring forces,were deduced.The new support system was applied to a typical roadway affected by intensive dynamic pressure that is located in the Xinyuan Coal Mine.The results show that the largest subsidence of the roof was 97 mm,the convergence of the two sides was less than 248 mm,and the average depth of the loose,fractured layer was only 6.12 mm.This proves that the new support system is feasible and effective.

  8. A modified dynamic evolving neural-fuzzy approach to modeling customer satisfaction for affective design.

    Science.gov (United States)

    Kwong, C K; Fung, K Y; Jiang, Huimin; Chan, K Y; Siu, Kin Wai Michael

    2013-01-01

    Affective design is an important aspect of product development to achieve a competitive edge in the marketplace. A neural-fuzzy network approach has been attempted recently to model customer satisfaction for affective design and it has been proved to be an effective one to deal with the fuzziness and non-linearity of the modeling as well as generate explicit customer satisfaction models. However, such an approach to modeling customer satisfaction has two limitations. First, it is not suitable for the modeling problems which involve a large number of inputs. Second, it cannot adapt to new data sets, given that its structure is fixed once it has been developed. In this paper, a modified dynamic evolving neural-fuzzy approach is proposed to address the above mentioned limitations. A case study on the affective design of mobile phones was conducted to illustrate the effectiveness of the proposed methodology. Validation tests were conducted and the test results indicated that: (1) the conventional Adaptive Neuro-Fuzzy Inference System (ANFIS) failed to run due to a large number of inputs; (2) the proposed dynamic neural-fuzzy model outperforms the subtractive clustering-based ANFIS model and fuzzy c-means clustering-based ANFIS model in terms of their modeling accuracy and computational effort.

  9. A shift from exploitation to interference competition with increasing density affects population and community dynamics.

    Science.gov (United States)

    Holdridge, Erica M; Cuellar-Gempeler, Catalina; terHorst, Casey P

    2016-08-01

    Intraspecific competition influences population and community dynamics and occurs via two mechanisms. Exploitative competition is an indirect effect that occurs through use of a shared resource and depends on resource availability. Interference competition occurs by obstructing access to a resource and may not depend on resource availability. Our study tested whether the strength of interference competition changes with protozoa population density. We grew experimental microcosms of protozoa and bacteria under different combinations of protozoan density and basal resource availability. We then solved a dynamic predator-prey model for parameters of the functional response using population growth rates measured in our experiment. As population density increased, competition shifted from exploitation to interference, and competition was less dependent on resource levels. Surprisingly, the effect of resources was weakest when competition was the most intense. We found that at low population densities, competition was largely exploitative and resource availability had a large effect on population growth rates, but the effect of resources was much weaker at high densities. This shift in competitive mechanism could have implications for interspecific competition, trophic interactions, community diversity, and natural selection. We also tested whether this shift in the mechanism of competition with protozoa density affected the structure of the bacterial prey community. We found that both resources and protozoa density affected the structure of the bacterial prey community, suggesting that competitive mechanism may also affect trophic interactions.

  10. Natural product Celastrol destabilizes tubulin heterodimer and facilitates mitotic cell death triggered by microtubule-targeting anti-cancer drugs.

    Directory of Open Access Journals (Sweden)

    Hakryul Jo

    Full Text Available BACKGROUND: Microtubule drugs are effective anti-cancer agents, primarily due to their ability to induce mitotic arrest and subsequent cell death. However, some cancer cells are intrinsically resistant or acquire a resistance. Lack of apoptosis following mitotic arrest is thought to contribute to drug resistance that limits the efficacy of the microtubule-targeting anti-cancer drugs. Genetic or pharmacological agents that selectively facilitate the apoptosis of mitotic arrested cells present opportunities to strengthen the therapeutic efficacy. METHODOLOGY AND PRINCIPAL FINDINGS: We report a natural product Celastrol targets tubulin and facilitates mitotic cell death caused by microtubule drugs. First, in a small molecule screening effort, we identify Celastrol as an inhibitor of neutrophil chemotaxis. Subsequent time-lapse imaging analyses reveal that inhibition of microtubule-mediated cellular processes, including cell migration and mitotic chromosome alignment, is the earliest events affected by Celastrol. Disorganization, not depolymerization, of mitotic spindles appears responsible for mitotic defects. Celastrol directly affects the biochemical properties of tubulin heterodimer in vitro and reduces its protein level in vivo. At the cellular level, Celastrol induces a synergistic apoptosis when combined with conventional microtubule-targeting drugs and manifests an efficacy toward Taxol-resistant cancer cells. Finally, by time-lapse imaging and tracking of microtubule drug-treated cells, we show that Celastrol preferentially induces apoptosis of mitotic arrested cells in a caspase-dependent manner. This selective effect is not due to inhibition of general cell survival pathways or mitotic kinases that have been shown to enhance microtubule drug-induced cell death. CONCLUSIONS AND SIGNIFICANCE: We provide evidence for new cellular pathways that, when perturbed, selectively induce the apoptosis of mitotic arrested cancer cells, identifying a

  11. Inertial forces affect fluid front displacement dynamics in a pore-throat network model.

    Science.gov (United States)

    Moebius, Franziska; Or, Dani

    2014-08-01

    The seemingly regular and continuous motion of fluid displacement fronts in porous media at the macroscopic scale is propelled by numerous (largely invisible) pore-scale abrupt interfacial jumps and pressure bursts. Fluid fronts in porous media are characterized by sharp phase discontinuities and by rapid pore-scale dynamics that underlie their motion; both attributes challenge standard continuum theories of these flow processes. Moreover, details of pore-scale dynamics affect front morphology and subsequent phase entrapment behind a front and thereby shape key macroscopic transport properties of the unsaturated zone. The study presents a pore-throat network model that focuses on quantifying interfacial dynamics and interactions along fluid displacement fronts. The porous medium is represented by a lattice of connected pore throats capable of detaining menisci and giving rise to fluid-fluid interfacial jumps (the study focuses on flow rate controlled drainage). For each meniscus along the displacement front we formulate a local inertial, capillary, viscous, and hydrostatic force balance that is then solved simultaneously for the entire front. The model enables systematic evaluation of the role of inertia and boundary conditions. Results show that while displacement patterns are affected by inertial forces mainly by invasion of throats with higher capillary resistance, phase entrapment (residual saturation) is largely unaffected by inertia, limiting inertial effects on hydrological properties behind a front. Interfacial jump velocities are often an order of magnitude larger than mean front velocity, are strongly dependent on geometrical throat dimensions, and become less predictable (more scattered) when inertia is considered. Model simulations of the distributions of capillary pressure fluctuations and waiting times between invasion events follow an exponential distribution and are in good agreement with experimental results. The modeling approach provides insights

  12. Biological Activity of 4-Substituted Methoxybenzoyl-Aryl-Thiazole (SMART): An Active Microtubule Inhibitor

    OpenAIRE

    2010-01-01

    Formation of microtubules is a dynamic process that involves polymerization and depolymerization of αβ-tubulin heterodimers. Drugs that enhance or inhibit tubulin polymerization can destroy this dynamic process, arresting cells in the G2/M phase of the cell cycle. Although drugs that target tubulin generally demonstrate cytotoxic potency in the sub-nanomolar range, resistance due to drug efflux is a common phenomenon among the antitubulin agents. We recently reported a class of 4-Substituted ...

  13. Salt—Water Dynamics in Highly Salinized Topsoil of Salt—Affected Soil During Water Infiltration

    Institute of Scientific and Technical Information of China (English)

    WANGXUE-FENG; YOUWEN-RUI; 等

    1991-01-01

    Continuous monitoring of salt and water movement in the soil profile of highly salinized topsoil under steadystate infiltration was conducted.It gives that salt and water dynamics during convection-diffusion period can be divided into three stages:1.formation of a salt peak,2.the salt peak moving downwards till the appearance of the summit of the salt peak,3.the salt peak moving further downwards with the peak value decreasing.Results show that the maximum salt peak appears at the same depth if soil texture and outflow condition are the same.Factors affecting salt and water movement and ion components in the outflow solution underinfiltration are discussed.

  14. Factors affecting the haptic filled-space illusion for dynamic touch.

    Science.gov (United States)

    Sanders, Abram F J; Kappers, Astrid M L

    2009-02-01

    In the haptic filled-space illusion for active dynamic touch, observers move their fingertip across an unfilled extent or an extent filled with intermediate stimulations. Previous researchers have reported lengths of filled extents to be overestimated, but the parameters affecting the strength of the illusion are still largely unknown. In the current research, we show that the illusion persists when intermediate stimulations do not provide information about the extent's length. In addition, the results show that the strength of the illusion increases with the number of filler elements. In contrast with earlier research, we control for movement speed differences between filled and unfilled extents. The results suggest that the strength of the illusion is independent of the overall average movement speed. Insight into factors affecting the strength of the illusion may provide a better understanding of the kinematic mechanisms underlying haptic length perception.

  15. Microtubules guide root hair tip growth

    NARCIS (Netherlands)

    Sieberer, B.; Ketelaar, M.J.; Esseling, J.J.; Emons, A.M.C.

    2005-01-01

    The ability to establish cell polarity is crucial to form and function of an individual cell. Polarity underlies critical processes during cell development, such as cell growth, cell division, cell differentiation and cell signalling. Interphase cytoplasmic microtubules in tip-growing fission yeast

  16. A role for katanin in plant cell division: microtubule organization in dividing root cells of fra2 and lue1Arabidopsis thaliana mutants.

    Science.gov (United States)

    Panteris, Emmanuel; Adamakis, Ioannis-Dimosthenis S; Voulgari, Georgia; Papadopoulou, Galini

    2011-07-01

    Severing of microtubules by katanin has proven to be crucial for cortical microtubule organization in elongating and differentiating plant cells. On the contrary, katanin is currently not considered essential during cell division in plants as it is in animals. However, defects in cell patterning have been observed in katanin mutants, implying a role for it in dividing plant cells. Therefore, microtubule organization was studied in detail by immunofluorescence in dividing root cells of fra2 and lue1 katanin mutants of Arabidopsis thaliana. In both, early preprophase bands consisted of poorly aligned microtubules, prophase spindles were multipolar, and the microtubules of expanding phragmoplasts were elongated, bended toward and connected to the surface of daughter nuclei. Accordingly, severing by katanin seems to be necessary for the proper organization of these microtubule arrays. In both fra2 and lue1, metaphase/anaphase spindles and initiating phragmoplasts exhibited typical organization. However, they were obliquely oriented more frequently than in the wild type. It is proposed that this oblique orientation may be due to prophase spindle multipolarity and results in a failure of the cell plate to follow the predetermined division plane, during cytokinesis, producing oblique cell walls in the roots of both mutants. It is therefore concluded that, like in animal cells, katanin is important for plant cell division, influencing the organization of several microtubule arrays. Moreover, failure in microtubule severing indirectly affects the orientation of the division plane.

  17. Mechanical properties of organelles driven by microtubule-dependent molecular motors in living cells.

    Directory of Open Access Journals (Sweden)

    Luciana Bruno

    Full Text Available The organization of the cytoplasm is regulated by molecular motors which transport organelles and other cargoes along cytoskeleton tracks. Melanophores have pigment organelles or melanosomes that move along microtubules toward their minus and plus end by the action of cytoplasmic dynein and kinesin-2, respectively. In this work, we used single particle tracking to characterize the mechanical properties of motor-driven organelles during transport along microtubules. We tracked organelles with high temporal and spatial resolutions and characterized their dynamics perpendicular to the cytoskeleton track. The quantitative analysis of these data showed that the dynamics is due to a spring-like interaction between melanosomes and microtubules in a viscoelastic microenvironment. A model based on a generalized Langevin equation explained these observations and predicted that the stiffness measured for the motor complex acting as a linker between organelles and microtubules is ∼ one order smaller than that determined for motor proteins in vitro. This result suggests that other biomolecules involved in the interaction between motors and organelles contribute to the mechanical properties of the motor complex. We hypothesise that the high flexibility observed for the motor linker may be required to improve the efficiency of the transport driven by multiple copies of motor molecules.

  18. TCS1, a Microtubule-Binding Protein, Interacts with KCBP/ZWICHEL to Regulate Trichome Cell Shape in Arabidopsis thaliana.

    Directory of Open Access Journals (Sweden)

    Liangliang Chen

    2016-10-01

    Full Text Available How cell shape is controlled is a fundamental question in developmental biology, but the genetic and molecular mechanisms that determine cell shape are largely unknown. Arabidopsis trichomes have been used as a good model system to investigate cell shape at the single-cell level. Here we describe the trichome cell shape 1 (tcs1 mutants with the reduced trichome branch number in Arabidopsis. TCS1 encodes a coiled-coil domain-containing protein. Pharmacological analyses and observations of microtubule dynamics show that TCS1 influences the stability of microtubules. Biochemical analyses and live-cell imaging indicate that TCS1 binds to microtubules and promotes the assembly of microtubules. Further results reveal that TCS1 physically associates with KCBP/ZWICHEL, a microtubule motor involved in the regulation of trichome branch number. Genetic analyses indicate that kcbp/zwi is epistatic to tcs1 with respect to trichome branch number. Thus, our findings define a novel genetic and molecular mechanism by which TCS1 interacts with KCBP to regulate trichome cell shape by influencing the stability of microtubules.

  19. Doublecortin Is Excluded from Growing Microtubule Ends and Recognizes the GDP-Microtubule Lattice.

    Science.gov (United States)

    Ettinger, Andreas; van Haren, Jeffrey; Ribeiro, Susana A; Wittmann, Torsten

    2016-06-20

    Many microtubule (MT) functions are mediated by a diverse class of proteins (+TIPs) at growing MT plus ends that control intracellular MT interactions and dynamics and depend on end-binding proteins (EBs) [1]. Cryoelectron microscopy has recently identified the EB binding site as the interface of four tubulin dimers that undergoes a conformational change in response to β-tubulin GTP hydrolysis [2, 3]. Doublecortin (DCX), a MT-associated protein (MAP) required for neuronal migration during cortical development [4, 5], binds to the same site as EBs [6], and recent in vitro studies proposed DCX localization to growing MT ends independent of EBs [7]. Because this conflicts with observations in neurons [8, 9] and the molecular function of DCX is not well understood, we revisited intracellular DCX dynamics at low expression levels. Here, we report that DCX is not a +TIP in cells but, on the contrary, is excluded from the EB1 domain. In addition, we find that DCX-MT interactions are highly sensitive to MT geometry. In cells, DCX binding was greatly reduced at MT segments with high local curvature. Remarkably, this geometry-dependent binding to MTs was completely reversed in the presence of taxanes, which reconciles incompatible observations in cells [9] and in vitro [10]. We propose a model explaining DCX specificity for different MT geometries based on structural changes induced by GTP hydrolysis that decreases the spacing between adjacent tubulin dimers [11]. Our data are consistent with a unique mode of MT interaction in which DCX specifically recognizes this compacted GDP-like MT lattice.

  20. Dynamical response of dark matter to galaxy evolution affects direct-detection experiments

    Science.gov (United States)

    Petersen, Michael S.; Katz, Neal; Weinberg, Martin D.

    2016-12-01

    Over a handful of rotation periods, dynamical processes in barred galaxies induce nonaxisymmetric structure in dark matter halos. Using n -body simulations of a Milky Way-like barred galaxy, we identify both a trapped dark matter component, a shadow bar, and a strong response wake in the dark matter distribution that affects the predicted dark matter detection rates for current experiments. The presence of a baryonic disk, together with well-known dynamical processes (e.g. spiral structure and bar instabilities), increases the dark matter density in the disk plane. We find that the magnitude of the combined stellar and shadow bar evolution, when isolated from the effect of the axisymmetric gravitational potential of the disk, accounts for >30 % of this overall increase in disk-plane density. This is significantly larger than that of previously claimed deviations from the standard halo model. The dark matter density and kinematic wakes driven by the Milky Way bar increase the detectability of dark matter overall, especially for the experiments with higher vmin . These astrophysical features increase the detection rate by more than a factor of 2 when compared to the standard halo model and by a factor of 10 for experiments with high minimum recoil energy thresholds. These same features increase (decrease) the annual modulation for low (high) minimum recoil energy experiments. We present physical arguments for why these dynamics are generic for barred galaxies such as the Milky Way rather than contingent on a specific galaxy model.

  1. Asymmetrical loading affects intersegmental dynamics during the swing phase of walking.

    Science.gov (United States)

    Smith, Jeremy D; Royer, Todd D; Martin, Philip E

    2013-08-01

    Much of the work related to lower extremity inertia manipulations has focused on temporal, kinematic and traditional inverse dynamics assessments during locomotion. Intersegmental dynamics is an analytical technique that provides further insights into mechanisms underlying linked-segment motion. The purpose of this study was to determine how intersegmental dynamics during the swing phase of walking are altered during asymmetrical lower extremity loading. Participants walked overground at a speed of 1.57 ms(-1) with 0, 0.5, 1.0, and 2.0 kg attached to one foot. Net, interaction, gravitational, and muscle moments were computed. Moment magnitudes at joints of the loaded leg increased systematically with increasing load, whereas unloaded leg moments were unaffected by loading. With increasing load, relative contributions of interaction moments about the knee and hip and gravitational moment about the ankle increased (i.e., 21%, 8%, and 44% increases, respectively), whereas the relative contributions of muscle moments about all three joints declined (i.e., -4%, -13%, and -8% decreases for the ankle, knee, and hip, respectively for unloaded vs. 2.0 kg). These results suggest that altered inertia properties of the limb not only affected the amount of muscular effort required to swing the leg, but also changed the nature of the interaction between segments.

  2. Effect of hexylene glycol-altered microtubule distributions on cytokinesis and polar lobe formation in fertilized eggs of Ilyanassa obsoleta

    Science.gov (United States)

    Conrad, A. H.; Stephens, A. P.; Conrad, G. W.; Spooner, B. S. (Principal Investigator)

    1994-01-01

    Some effects of gravity on early morphogenesis are correlated with microtubule locations within cells. During first cleavage in Ilyanassa obsoleta embryos, a transitory polar lobe constriction forms and then relaxes, allowing the polar lobe to merge with one daughter cell. If the polar lobe is equally divided or removed, morphogenesis is severely disrupted. To examine microtuble locations during early Ilyanassa development, eggs were fixed and stained for polymerized alpha-tubulin during first cleavage. The mitotic apparatus assembles at the animal pole. The cleavage furrow forms between the asters, constricting to a stabilized intercellular bridge encircling midbody-bound microtubules, whereas the polar lobe constriction forms below and parallel to the spindle, constricting to a transitory intercellular bridge encircling no detectable microtubules. At metaphase an alpha-tubulin epitope is distributed throughout the spindle, whereas a beta-tubulin epitope is present predominantly in the asters. Incubation in hexylene glycol, a drug that increases microtubule polymerization, during mitosis causes the polar lobe constriction to tighten around polymerized alpha-tubulin and remain stably constricted. If hexylene glycol is removed, alpha-tubulin staining disappears from the polar lobe constriction, which relaxes, whereas microtubules remain in the cleavage furrow, which remains constricted. These observations suggest that asymmetric distribution of microtubules affects early Ilyanassa cleavage patterns, and that continued presence of microtubules extending through an intercellular bridge is important for stabilization of the bridge constriction prior to completion of cytokinesis. These data provide the basis for further analysis of the role of microtubules in possible microgravity disruptions of Ilyanassa development.

  3. Molecular dynamics simulation of heat-affected zone of copper metal ablated with femtosecond laser

    Science.gov (United States)

    Hirayama, Yoichi; Obara, Minoru

    2005-03-01

    Femtosecond laser ablation of materials with high thermal conductivity is of paramount importance, because the chemical composition and properties of the area ablated with femtosecond laser are kept unchanged. The material processing by femtosecond laser can well control the heat-affected zone, compared to nanosecond laser ablation. We report on the heat-affected zone of crystalline copper (Cu) by use of femtosecond laser experimentally and theoretically. Laser ablation of Cu is investigated theoretically by two temperature model and molecular dynamics (MD) simulation. The MD simulation takes into account of electron temperature and thermal diffusion length calculated by two temperature model. The dependence of lattice temperature on time and depth is calculated by the MD simulation and two temperature model. The heat-affected zone estimated from the temperature is mainly studied and calculated to be 3 nm at 0.02 J/cm2 which is below the threshold fluence of 0.137 J/cm2. In addition, the thickness of heat-affected zone of copper crystal ablated with femtosecond Ti:sapphire laser is experimentally studied. As a result of X-ray diffraction (XRD) of the ablated surface, the surface crystallinity is partially changed into disordered structure from crystal form. The residual energy left in the metal, which is not used for ablation, will induce liquid phase, leading to the amorphous phase of the metal during resolidification. The thickness of heat-affected zone depends on laser fluence and is experimentally measured to be less than 1 μm at higher laser fluences than the ablation threshold.

  4. Can ocean acidification affect population dynamics of the barnacle Semibalanus balanoides at its southern range edge?

    Science.gov (United States)

    Findlay, Helen S; Burrows, Michael T; Kendall, Michael A; Spicer, John I; Widdicombe, Stephen

    2010-10-01

    The global ocean and atmosphere are warming. There is increasing evidence suggesting that, in addition to other environmental factors, climate change is affecting species distributions and local population dynamics. Additionally, as a consequence of the growing levels of atmospheric carbon dioxide (CO2), the oceans are taking up increasing amounts of this CO2, causing ocean pH to decrease (ocean acidification). The relative impacts of ocean acidification on population dynamics have yet to be investigated, despite many studies indicating that there will be at least a sublethal impact on many marine organisms, particularly key calcifying organisms. Using empirical data, we forced a barnacle (Semibalanus balanoides) population model to investigate the relative influence of sea surface temperature (SST) and ocean acidification on a population nearing the southern limit of its geographic distribution. Hindcast models were compared to observational data from Cellar Beach (southwestern United Kingdom). Results indicate that a declining pH trend (-0.0017 unit/yr), indicative of ocean acidification over the past 50 years, does not cause an observable impact on the population abundance relative to changes caused by fluctuations in temperature. Below the critical temperature (here T(crit) = 13.1 degrees C), pH has a more significant affect on population dynamics at this southern range edge. However, above this value, SST has the overriding influence. At lower SST, a decrease in pH (according to the National Bureau of Standards, pHNBs) from 8.2 to 7.8 can significantly decrease the population abundance. The lethal impacts of ocean acidification observed in experiments on early life stages reduce cumulative survival by approximately 25%, which again will significantly alter the population level at this southern limit. Furthermore, forecast predictions from this model suggest that combined acidification and warming cause this local population to die out 10 years earlier than

  5. Decreased summer drought affects plant productivity and soil carbon dynamics in a Mediterranean woodland

    Directory of Open Access Journals (Sweden)

    M. F. Cotrufo

    2011-09-01

    Full Text Available Precipitation patterns are expected to change in the Mediterranean region within the next decades, with projected decreases in total rainfall and increases in extreme events. We manipulated precipitation patterns in a Mediterranean woodland, dominated by Arbutus unedo L., to study the effects of changing precipitation regimes on above-ground net primary production (ANPP and soil C dynamics, specifically plant-derived C input to soil and soil respiration (SR. Experimental plots were exposed to either a 20 % reduction of throughfall or to water addition targeted at maintaining soil water content above a minimum of 10 % v/v. Treatments were compared to control plots which received ambient precipitation. Enhanced soil moisture during summer months highly stimulated annual stem primary production, litter fall, SR and net annual plant-derived C input to soil which on average increased by 130 %, 26 %, 58 % and 220 %, respectively, as compared to the control. In contrast, the 20 % reduction in throughfall (equivalent to 10 % reduction in precipitation did not significantly change soil moisture at the site, and therefore did not significantly affect ANPP or SR. We conclude that minor changes (around 10 % reduction in precipitation amount are not likely to significantly affect ANPP or soil C dynamics in Mediterranean woodlands. However, if summer rain increases, C cycling will significantly accelerate but soil C stocks are not likely to be changed in the short-term. More studies involving modelling of long-term C dynamics are needed to predict if the estimated increases in soil C input under wet conditions is going to be sustained and if labile C is being substituted to stable C, with a negative effect on long-term soil C stocks.

  6. Mastoid vibration affects dynamic postural control during gait in healthy older adults

    Science.gov (United States)

    Chien, Jung Hung; Mukherjee, Mukul; Kent, Jenny; Stergiou, Nicholas

    2017-01-01

    Vestibular disorders are difficult to diagnose early due to the lack of a systematic assessment. Our previous work has developed a reliable experimental design and the result shows promising results that vestibular sensory input while walking could be affected through mastoid vibration (MV) and changes are in the direction of motion. In the present paper, we wanted to extend this work to older adults and investigate how manipulating sensory input through mastoid vibration (MV) could affect dynamic postural control during walking. Three levels of MV (none, unilateral, and bilateral) applied via vibrating elements placed on the mastoid processes were combined with the Locomotor Sensory Organization Test (LSOT) paradigm to challenge the visual and somatosensory systems. We hypothesized that the MV would affect sway variability during walking in older adults. Our results revealed that MV significantly not only increased the amount of sway variability but also decreased the temporal structure of sway variability only in anterior-posterior direction. Importantly, the bilateral MV stimulation generally produced larger effects than the unilateral. This is an important finding that confirmed our experimental design and the results produced could guide a more reliable screening of vestibular system deterioration. PMID:28128341

  7. Mastoid vibration affects dynamic postural control during gait in healthy older adults

    Science.gov (United States)

    Chien, Jung Hung; Mukherjee, Mukul; Kent, Jenny; Stergiou, Nicholas

    2017-01-01

    Vestibular disorders are difficult to diagnose early due to the lack of a systematic assessment. Our previous work has developed a reliable experimental design and the result shows promising results that vestibular sensory input while walking could be affected through mastoid vibration (MV) and changes are in the direction of motion. In the present paper, we wanted to extend this work to older adults and investigate how manipulating sensory input through mastoid vibration (MV) could affect dynamic postural control during walking. Three levels of MV (none, unilateral, and bilateral) applied via vibrating elements placed on the mastoid processes were combined with the Locomotor Sensory Organization Test (LSOT) paradigm to challenge the visual and somatosensory systems. We hypothesized that the MV would affect sway variability during walking in older adults. Our results revealed that MV significantly not only increased the amount of sway variability but also decreased the temporal structure of sway variability only in anterior-posterior direction. Importantly, the bilateral MV stimulation generally produced larger effects than the unilateral. This is an important finding that confirmed our experimental design and the results produced could guide a more reliable screening of vestibular system deterioration.

  8. Microtubule-driven nuclear movements and linear elements as meiosis-specific characteristics of the fission yeasts Schizosaccharomyces versatilis and Schizosaccharomyces pombe.

    Science.gov (United States)

    Svoboda, A; Bähler, J; Kohli, J

    1995-11-01

    Meiotic prophase in Schizosaccharomyces pombe is characterized by striking nuclear movements and the formation of linear elements along chromosomes instead of tripartite synaptonemal complexes. We analysed the organization of nuclei and microtubules in cells of fission yeasts undergoing sexual differentiation. S. japonicus var. versatilis and S. pombe cells were studied in parallel, taking advantage of the better cytology in S. versatilis. During conjugation, microtubules were directed towards the mating projection. These microtubules seem to lead the haploid nuclei together in the zygote by interaction with the spindle pole bodies at the nuclear periphery. After karyogamy, arrays of microtubules emanating from the spindle pole body of the diploid nucleus extended to both cell poles. The same differentiated microtubule configuration was elaborated upon induction of azygotic meiosis in S. pombe. The cyclic movements of the elongated nuclei between the cell poles is reflected by a dynamic and coordinated shortening and lengthening of the two microtubule arrays. When the nucleus was at a cell end, one array was short while the other bridged the whole cell length. Experiments with inhibitors showed that microtubules are required for karyogamy and for the elongated shape and movement of nuclei during meiotic prophase. In both fission yeasts the SPBs and nucleoli are at the leading ends of the moving nuclei. Astral and cytoplasmic microtubules were also prominent during meiotic divisions and sporulation. We further show that in S. versatilis the linear elements formed during meiotic prophase are similar to those in S. pombe. Tripartite synaptonemal complexes were never detected. Taken together, these findings suggest that S. pombe and S. versatilis share basic characteristics in the organization of microtubules and the structure and behaviour of nuclei during their meiotic cell cycle. The prominent differentiations of microtubules and nuclei may be involved in the

  9. The formin DIAPH1 (mDia1) regulates megakaryocyte proplatelet formation by remodeling the actin and microtubule cytoskeletons.

    Science.gov (United States)

    Pan, Jiajia; Lordier, Larissa; Meyran, Deborah; Rameau, Philippe; Lecluse, Yann; Kitchen-Goosen, Susan; Badirou, Idinath; Mokrani, Hayat; Narumiya, Shuh; Alberts, Arthur S; Vainchenker, William; Chang, Yunhua

    2014-12-18

    Megakaryocytes are highly specialized precursor cells that produce platelets via cytoplasmic extensions called proplatelets. Proplatelet formation (PPF) requires profound changes in microtubule and actin organization. In this work, we demonstrated that DIAPH1 (mDia1), a mammalian homolog of Drosophila diaphanous that works as an effector of the small GTPase Rho, negatively regulates PPF by controlling the dynamics of the actin and microtubule cytoskeletons. Moreover, we showed that inhibition of both DIAPH1 and the Rho-associated protein kinase (Rock)/myosin pathway increased PPF via coordination of both cytoskeletons. We provide evidence that 2 major effectors of the Rho GTPase pathway (DIAPH1 and Rock/myosin II) are involved not only in Rho-mediated stress fibers assembly, but also in the regulation of microtubule stability and dynamics during PPF.

  10. How do Plants Organize Microtubules Without a Centrosome?

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    A microtubule nucleates from a γ-tubulin complex, which consists of γ-tubulin, proteins from the SPC97/SPC98 family, and the WD40 motif protein GCP-WD. We analyzed the phylogenetic relationships of the genes encoding these proteins and found that the components of this complex are widely conserved among land plants and other eukaryotes. By contrast,the interphase and mitotic arrays of microtubules in land plants differ from those in other eukaryotes. In the interphase cortical array, the majority of microtubules nucleate on existing microtubules in the absence of conspicuous microtubule organizing centers (MTOCs), such as a centrosome. During mitosis, the spindle also forms in the absence of conspicuous MTOCs. Both poles of the spindle are broad, and branched structures of microtubules called microtubule converging centers form at the poles. In this review, we hypothesize that the microtubule converging centers form via microtubuledependent microtubule nucleation, as in the case of the interphase arrays. The evolutionary insights arising from the molecular basis of the diversity in microtubule organization are discussed.

  11. Resolving bundled microtubules using anti-tubulin nanobodies.

    Science.gov (United States)

    Mikhaylova, Marina; Cloin, Bas M C; Finan, Kieran; van den Berg, Robert; Teeuw, Jalmar; Kijanka, Marta M; Sokolowski, Mikolaj; Katrukha, Eugene A; Maidorn, Manuel; Opazo, Felipe; Moutel, Sandrine; Vantard, Marylin; Perez, Frank; van Bergen en Henegouwen, Paul M P; Hoogenraad, Casper C; Ewers, Helge; Kapitein, Lukas C

    2015-08-11

    Microtubules are hollow biopolymers of 25-nm diameter and are key constituents of the cytoskeleton. In neurons, microtubules are organized differently between axons and dendrites, but their precise organization in different compartments is not completely understood. Super-resolution microscopy techniques can detect specific structures at an increased resolution, but the narrow spacing between neuronal microtubules poses challenges because most existing labelling strategies increase the effective microtubule diameter by 20-40 nm and will thereby blend neighbouring microtubules into one structure. Here we develop single-chain antibody fragments (nanobodies) against tubulin to achieve super-resolution imaging of microtubules with a decreased apparent diameter. To test the resolving power of these novel probes, we generate microtubule bundles with a known spacing of 50-70 nm and successfully resolve individual microtubules. Individual bundled microtubules can also be resolved in different mammalian cells, including hippocampal neurons, allowing novel insights into fundamental mechanisms of microtubule organization in cell- and neurobiology.

  12. Does Climate Change Mitigation Activity Affect Crude Oil Prices? Evidence from Dynamic Panel Model

    Directory of Open Access Journals (Sweden)

    Jude C. Dike

    2014-01-01

    Full Text Available This paper empirically investigates how climate change mitigation affects crude oil prices while using carbon intensity as the indicator for climate change mitigation. The relationship between crude oil prices and carbon intensity is estimated using an Arellano and Bond GMM dynamic panel model. This study undertakes a regional-level analysis because of the geographical similarities among the countries in a region. Regions considered for the study are Africa, Asia and Oceania, Central and South America, the EU, the Middle East, and North America. Results show that there is a positive relationship between crude oil prices and carbon intensity, and a 1% change in carbon intensity is expected to cause about 1.6% change in crude oil prices in the short run and 8.4% change in crude oil prices in the long run while the speed of adjustment is 19%.

  13. Does consideration of water routing affect simulated water and carbon dynamics in terrestrial ecosystems?

    Directory of Open Access Journals (Sweden)

    G. Tang

    2013-10-01

    Full Text Available The cycling of carbon in terrestrial ecosystems is closely coupled with the cycling of water. An important mechanism connecting ecological and hydrological processes in terrestrial ecosystems is lateral flow of water along landscapes. Few studies, however, have examined explicitly how consideration of water routing affects simulated water and carbon dynamics in terrestrial ecosystems. The objective of this study is to explore how consideration of water routing in a process-based hydroecological model affects simulated water and carbon dynamics. To achieve that end, we rasterized the regional hydroecological simulation systems (RHESSys and employed the rasterized RHESSys (R-RHESSys in a forested watershed. We performed and compared two contrasting simulations, one with and another without water routing. We found that R-RHESSys is able to correctly simulate major hydrological and ecological variables regardless of whether water routing is considered. When water routing was neglected, however, soil water table depth and saturation deficit were simulated to be smaller and spatially more homogeneous. As a result, evaporation, forest productivity and soil heterotrophic respiration also were simulated to be spatially more homogeneous compared to simulation with water routing. When averaged for the entire watershed, however, differences in simulated water and carbon fluxes are not significant between the two simulations. Overall, the study demonstrated that consideration of water routing enabled R-RHESSys to better capture our preconception of the spatial patterns of water table depth and saturation deficit across the watershed. Because the spatial pattern of soil moisture is fundamental to water efflux from land to the atmosphere, forest productivity and soil microbial activity, ecosystem and carbon cycle models, therefore, need to explicitly represent water routing in order to accurately quantify the magnitudes and patterns of water and carbon fluxes

  14. Motor protein accumulation on antiparallel microtubule overlaps

    CERN Document Server

    Kuan, Hui-Shun

    2015-01-01

    Biopolymers serve as one-dimensional tracks on which motor proteins move to perform their biological roles. Motor protein phenomena have inspired theoretical models of one-dimensional transport, crowding, and jamming. Experiments studying the motion of Xklp1 motors on reconstituted antiparallel microtubule overlaps demonstrated that motors recruited to the overlap walk toward the plus end of individual microtubules and frequently switch between filaments. We study a model of this system that couples the totally asymmetric simple exclusion process (TASEP) for motor motion with switches between antiparallel filaments and binding kinetics. We determine steady-state motor density profiles for fixed-length overlaps using exact and approximate solutions of the continuum differential equations and compare to kinetic Monte Carlo simulations. The center region, far from the overlap ends, has a constant motor density as one would na\\"ively expect. However, rather than following a simple binding equilibrium, the center ...

  15. The actin-microtubule cross-linking activity of Drosophila Short stop is regulated by intramolecular inhibition.

    Science.gov (United States)

    Applewhite, Derek A; Grode, Kyle D; Duncan, Mara C; Rogers, Stephen L

    2013-09-01

    Actin and microtubule dynamics must be precisely coordinated during cell migration, mitosis, and morphogenesis--much of this coordination is mediated by proteins that physically bridge the two cytoskeletal networks. We have investigated the regulation of the Drosophila actin-microtubule cross-linker Short stop (Shot), a member of the spectraplakin family. Our data suggest that Shot's cytoskeletal cross-linking activity is regulated by an intramolecular inhibitory mechanism. In its inactive conformation, Shot adopts a "closed" conformation through interactions between its NH(2)-terminal actin-binding domain and COOH-terminal EF-hand-GAS2 domain. This inactive conformation is targeted to the growing microtubule plus end by EB1. On activation, Shot binds along the microtubule through its COOH-terminal GAS2 domain and binds to actin with its NH(2)-terminal tandem CH domains. We propose that this mechanism allows Shot to rapidly cross-link dynamic microtubules in response to localized activating signals at the cell cortex.

  16. Microtubule-Targeting Therapy for Prostate Cancer

    Science.gov (United States)

    2007-02-01

    Cancer1828 Mol Cancer Ther 2005;4(12). December 2005 22. Sambrook J, Fritsch EF, Maniatis T. Molecular cloning : a laboratory manual. 2nd ed. Cold Spring...Harbor (NY): Cold Spring Harbor Laboratory; 1989. 23. Zhu XX, Kozarsky K, Strahler JR, et al. Molecular cloning of a novel human leukemia-associated...of Cancer Research, Abstract #4940, 2005. 3. Mistry, SJ, Atweh, GF. Microtubule targeting therapy: Anti-stathmin based molecular cancer

  17. Facial feedback affects valence jugdments of dynamic and static emotional expressions

    Directory of Open Access Journals (Sweden)

    Sylwia eHyniewska

    2015-03-01

    Full Text Available The ability to judge others’ emotions is required for the establishment and maintenance of smooth interactions in a community. Several lines of evidence suggest that the attribution of meaning to a face is influenced by the facial actions produced by an observer during the observation of a face. However, empirical studies testing causal relationships between observers’ facial actions and emotion judgments have reported mixed findings. This study is the first to measure emotion judgments in terms of valence and arousal dimensions while comparing dynamic versus static presentations of facial expressions. We presented pictures and videos of facial expressions of anger and happiness. Participants (N = 36 were asked to differentiate between the gender of faces by activating the corrugator supercilii muscle (brow lowering and zygomaticus major muscle (cheek raising. They were also asked to evaluate the internal states of the stimuli using the affect grid while maintaining the facial action until they finished responding. The cheek-raising condition increased the attributed valence scores compared with the brow-lowering condition. This effect of facial actions was observed for static as well as for dynamic facial expressions. These data suggest that facial feedback mechanisms contribute to the judgment of the valence of emotional facial expressions.

  18. Wolbachia-Host Interactions: Host Mating Patterns Affect Wolbachia Density Dynamics.

    Science.gov (United States)

    Zhao, Dong-Xiao; Zhang, Xiang-Fei; Chen, Da-Song; Zhang, Yan-Kai; Hong, Xiao-Yue

    2013-01-01

    Wolbachia are maternally inherited intracellular bacteria that infect a wide range of arthropods and cause an array of effects on host reproduction, fitness and mating behavior. Although our understanding of the Wolbachia-associated effects on hosts is rapidly expanding, our knowledge of the host factors that mediate Wolbachia dynamics is rudimentary. Here, we explore the interactions between Wolbachia and its host, the two-spotted spider mite Tetranychus urticae Koch. Our results indicate that Wolbachia induces strong cytoplasmic incompatibility (CI), increases host fecundity, but has no effects on the longevity of females and the mating competitiveness of males in T. urticae. Most importantly, host mating pattern was found to affect Wolbachia density dynamics during host aging. Mating of an uninfected mite of either sex with an infected mite attenuates the Wolbachia density in the infected mite. According to the results of Wolbachia localization, this finding may be associated with the tropism of Wolbachia for the reproductive tissue in adult spider mites. Our findings describe a new interaction between Wolbachia and their hosts.

  19. The Dynamical Response of Dark Matter to Galaxy Evolution Affects Direct-Detection Experiments

    CERN Document Server

    Petersen, Michael S; Weinberg, Martin D

    2016-01-01

    Over a handful of rotation periods, dynamical processes in barred galaxies induce non-axisymmetric structure in dark matter halos. Using n-body simulations of a Milky Way-like barred galaxy, we identify both a trapped dark-matter component, a shadow bar, and a strong response wake in the dark-matter distribution that affects the predicted dark-matter detection rates for current experiments. The presence of a baryonic disk together with well-known dynamical processes (e.g. spiral structure and bar instabilities) increase the dark matter density in the disk plane. We find that the magnitude of the combined stellar and shadow bar evolution, when isolated from the effect of the axisymmetric gravitational potential of the disk, accounts for >30% of this overall increase in disk-plane density. This is significantly larger that of previously claimed deviations from the standard halo model. The dark-matter density and kinematic wakes driven by the Milky Way bar increase the detectability of dark matter overall, espec...

  20. Wolbachia-Host Interactions: Host Mating Patterns Affect Wolbachia Density Dynamics.

    Directory of Open Access Journals (Sweden)

    Dong-Xiao Zhao

    Full Text Available Wolbachia are maternally inherited intracellular bacteria that infect a wide range of arthropods and cause an array of effects on host reproduction, fitness and mating behavior. Although our understanding of the Wolbachia-associated effects on hosts is rapidly expanding, our knowledge of the host factors that mediate Wolbachia dynamics is rudimentary. Here, we explore the interactions between Wolbachia and its host, the two-spotted spider mite Tetranychus urticae Koch. Our results indicate that Wolbachia induces strong cytoplasmic incompatibility (CI, increases host fecundity, but has no effects on the longevity of females and the mating competitiveness of males in T. urticae. Most importantly, host mating pattern was found to affect Wolbachia density dynamics during host aging. Mating of an uninfected mite of either sex with an infected mite attenuates the Wolbachia density in the infected mite. According to the results of Wolbachia localization, this finding may be associated with the tropism of Wolbachia for the reproductive tissue in adult spider mites. Our findings describe a new interaction between Wolbachia and their hosts.

  1. Factors affecting population dynamics of maternally transmitted endosymbionts in Bemisia tabaci.

    Directory of Open Access Journals (Sweden)

    Huipeng Pan

    Full Text Available While every individual of Bemisia tabaci (Hemiptera: Aleyrodidae harbors the primary symbiont (P-symbiont Portiera, the infection frequencies of the six secondary symbionts (S-symbionts including Hamiltonella, Arsenophonus, Cardinium, Wolbachia, Rickettsia and Fritschea vary greatly among different populations. To characterize the factors influencing the infection dynamics of the six S-symbionts in B. tabaci, gene-specific PCR were conducted to screen for the presence of the P-symbiont Portiera and the six S-symbionts in 61 (17 B and 44 Q biotypes field populations collected from different plant species and locations in China. All individuals of the 61 populations hosted the P-symbiont Portiera, but none of them harbored Arsenophonus and Fritschea. The presence and infection rates of Hamiltonella, Cardinium, Rickettsia, Wolbachia and their co-infections Rickettsia + Hamiltonella (RH, Rickettsia + Cardinium (RC, Hamiltonella + Cardinium (HC and Rickettsia + Hamiltonella + Cardinium (RHC varied significantly among the 61 field populations; and the observed variations can be explained by biotypes, sexes, host plants and geographical locations of these field populations. Taken together, at least three factors including biotype, host plant and geographical location affect the infection dynamics of S-symbionts in B. tabaci.

  2. N dynamics of Inner Mongolia typical steppe as affected by grazing

    Science.gov (United States)

    Giese, M.; Gao, Y. Z.; Brueck, H.; Butterbach-Bahl, K.

    2012-04-01

    For large areas of Inner Mongolian semi-arid grasslands, as for many regions of the Eurasian steppe belt, substantial land degradation was reported as a consequence of excessive overgrazing during the last decades. Nitrogen is considered as a key element for ecosystem functions and therefore, a comprehensive analysis of the system's N balance and cycle as affected by land-use change is of fundamental importance to maintain, improve or restore ecosystem services such as forage production, carbon sequestration and diversity conservation. In this comprehensive case study of a Chinese typical steppe, we present an in-depth analysis of N dynamics including the balance of N gains and losses, and N cycling. N pools and fluxes were simultaneously quantified on three grassland sites of contrasting grazing intensity. Our N balances indicated the ungrazed site as N sink with annual net N input of up to 3 g N m-2, mainly due to N input by dust deposition, whereas the heavily grazed site must be considered as N source with net losses of up to 1.7 g m-2. Mayor N losses occurred via dust emissions and excrement export from grazing sites, the latter as a consequence of the common practice of keeping sheep in paddocks overnight. Compared to these fluxes, gaseous N losses, export of animal products (live weight and wool) and biological N2 fixation were of minor relevance. Heavy grazing reduced pool sizes of both topsoil organic N, and above- and belowground biomass N. Furthermore, grazing reduced N fluxes with regard to N uptake, decomposition, gross microbial N turnover, and immobilization. Most N-related processes were more intensive in seasons of higher water availability indicating complex interactions between land-use intensity and climate variability. The projected increase of annual atmospheric N wet deposition and changes in rainfall pattern will likely affect the N sink-source pathways and N flux dynamics, indicating high potential impact of future N enrichment and climate

  3. Tuning microtubule-based transport via filamentous MAPs: the problem of dynein

    Science.gov (United States)

    Vershinin, Michael; Xu, Jing; Razafsky, David S.; King, Stephen J.; Gross, Steven P.

    2010-01-01

    We recently proposed that regulating the single-to-multiple motor transition was a likely strategy for regulating kinesin-based transport in vivo. Here, we use an in vitro bead assay coupled with an optical trap to investigate how this proposed regulatory mechanism affects dynein-based transport. We show that tau’s regulation of kinesin function can proceed without interfering with dynein-based transport. Surprisingly, at extremely high tau levels—where kinesin cannot bind microtubules—dynein can still contact microtubules. The difference between tau’s effects on kinesin- and dynein-based motility suggests that tau can be used to tune relative amounts of plus-end and minus-end directed transport. As in the case of kinesin, we find that the 3RS isoform of tau is a more potent inhibitor of dynein binding to microtubules. We show that this isoform-specific effect is not due to steric interference of tau’s projection domains, but rather due to tau’s interactions with the motor at the microtubule surface. Nonetheless, we do observe a modest steric interference effect of tau away from the microtubule and discuss the potential implications of this for molecular motor structure. PMID:18373727

  4. MICROTUBULE ORGANIZATION 1 regulates structure and function of microtubule arrays during mitosis and cytokinesis in the Arabidopsis root.

    Science.gov (United States)

    Kawamura, Eiko; Himmelspach, Regina; Rashbrooke, Madeleine C; Whittington, Angela T; Gale, Kevin R; Collings, David A; Wasteneys, Geoffrey O

    2006-01-01

    MICROTUBULE ORGANIZATION 1 (MOR1) is a plant member of the highly conserved MAP215/Dis1 family of microtubule-associated proteins. Prior studies with the temperature-sensitive mor1 mutants of Arabidopsis (Arabidopsis thaliana), which harbor single amino acid substitutions in an N-terminal HEAT repeat, proved that MOR1 regulates cortical microtubule organization and function. Here we demonstrate by use of live cell imaging and immunolabeling that the mor1-1 mutation generates specific defects in the microtubule arrays of dividing vegetative cells. Unlike the universal cortical microtubule disorganization in elongating mor1-1 cells, disruption of mitotic and cytokinetic microtubule arrays was not detected in all dividing cells. Nevertheless, quantitative analysis identified distinct defects in preprophase bands (PPBs), spindles, and phragmoplasts. In nearly one-half of dividing cells at the restrictive temperature of 30 degrees C, PPBs were not detected prior to spindle formation, and those that did form were often disrupted. mor1-1 spindles and phragmoplasts were short and abnormally organized and persisted for longer times than in wild-type cells. The reduced length of these arrays predicts that the component microtubule lengths are also reduced, suggesting that microtubule length is a critical determinant of spindle and phragmoplast structure, orientation, and function. Microtubule organizational defects led to aberrant chromosomal arrangements, misaligned or incomplete cell plates, and multinucleate cells. Antiserum raised against an N-terminal MOR1 sequence labeled the full length of microtubules in interphase arrays, PPBs, spindles, and phragmoplasts. Continued immunolabeling of the disorganized and short microtubules of mor1-1 at the restrictive temperature demonstrated that the mutant mor1-1(L174F) protein loses function without dissociating from microtubules, providing important insight into the mechanism by which MOR1 may regulate microtubule length.

  5. Reduced short term adaptation to robot generated dynamic environment in children affected by Cerebral Palsy

    Directory of Open Access Journals (Sweden)

    Di Rosa Giuseppe

    2011-05-01

    Full Text Available Abstract Background It is known that healthy adults can quickly adapt to a novel dynamic environment, generated by a robotic manipulandum as a structured disturbing force field. We suggest that it may be of clinical interest to evaluate to which extent this kind of motor learning capability is impaired in children affected by cerebal palsy. Methods We adapted the protocol already used with adults, which employs a velocity dependant viscous field, and compared the performance of a group of subjects affected by Cerebral Palsy (CP group, 7 subjects with a Control group of unimpaired age-matched children. The protocol included a familiarization phase (FA, during which no force was applied, a force field adaptation phase (CF, and a wash-out phase (WO in which the field was removed. During the CF phase the field was shut down in a number of randomly selected "catch" trials, which were used in order to evaluate the "learning index" for each single subject and the two groups. Lateral deviation, speed and acceleration peaks and average speed were evaluated for each trajectory; a directional analysis was performed in order to inspect the role of the limb's inertial anisotropy in the different experimental phases. Results During the FA phase the movements of the CP subjects were more curved, displaying greater and variable directional error; over the course of the CF phase both groups showed a decreasing trend in the lateral error and an after-effect at the beginning of the wash-out, but the CP group had a non significant adaptation rate and a lower learning index, suggesting that CP subjects have reduced ability to learn to compensate external force. Moreover, a directional analysis of trajectories confirms that the control group is able to better predict the force field by tuning the kinematic features of the movements along different directions in order to account for the inertial anisotropy of arm. Conclusions Spatial abnormalities in children affected

  6. Reduced short term adaptation to robot generated dynamic environment in children affected by Cerebral Palsy

    Science.gov (United States)

    2011-01-01

    Background It is known that healthy adults can quickly adapt to a novel dynamic environment, generated by a robotic manipulandum as a structured disturbing force field. We suggest that it may be of clinical interest to evaluate to which extent this kind of motor learning capability is impaired in children affected by cerebal palsy. Methods We adapted the protocol already used with adults, which employs a velocity dependant viscous field, and compared the performance of a group of subjects affected by Cerebral Palsy (CP group, 7 subjects) with a Control group of unimpaired age-matched children. The protocol included a familiarization phase (FA), during which no force was applied, a force field adaptation phase (CF), and a wash-out phase (WO) in which the field was removed. During the CF phase the field was shut down in a number of randomly selected "catch" trials, which were used in order to evaluate the "learning index" for each single subject and the two groups. Lateral deviation, speed and acceleration peaks and average speed were evaluated for each trajectory; a directional analysis was performed in order to inspect the role of the limb's inertial anisotropy in the different experimental phases. Results During the FA phase the movements of the CP subjects were more curved, displaying greater and variable directional error; over the course of the CF phase both groups showed a decreasing trend in the lateral error and an after-effect at the beginning of the wash-out, but the CP group had a non significant adaptation rate and a lower learning index, suggesting that CP subjects have reduced ability to learn to compensate external force. Moreover, a directional analysis of trajectories confirms that the control group is able to better predict the force field by tuning the kinematic features of the movements along different directions in order to account for the inertial anisotropy of arm. Conclusions Spatial abnormalities in children affected by cerebral palsy may be

  7. Cellular Samurai: katanin and the severing of microtubules.

    Science.gov (United States)

    Quarmby, L

    2000-08-01

    Recent biochemical studies of the AAA ATPase, katanin, provide a foundation for understanding how microtubules might be severed along their length. These in vitro studies are complemented by a series of recent reports of direct in vivo observation of microtubule breakage, which indicate that the in vitro phenomenon of catalysed microtubule severing is likely to be physiological. There is also new evidence that microtubule severing by katanin is important for the production of non-centrosomal microtubules in cells such as neurons and epithelial cells. Although it has been difficult to establish the role of katanin in mitosis, new genetic evidence indicates that a katanin-like protein, MEI-1, plays an essential role in meiosis in C. elegans. Finally, new proteins involved in the severing of axonemal microtubules have been discovered in the deflagellation system of Chlamydomonas.

  8. Microtubule Associated Proteins in Plants and the Processes They Manage

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    Microtubule associated proteins (MAPs) are proteins that physically bind to microtubules in eukaryotes. MAPs play important roles in regulating the polymerization and organization of microtubules and in using the ensuing microtubule arrays to carry out a variety of cellular functions. In plants, MAPs manage the construction, repositioning, and dismantling of four distinct microtubule arrays throughout the cell cycle. Three of these arrays, the cortical array, the preprophase band,and the phragmoplast, are prominent to plants and are responsible for facilitating cell wall deposition and modification,transducing signals, demarcating the plane of cell division, and forming the new cell plate during cytokinesis, This review highlights important aspects of how MAPs in plants establish and maintain microtubule arrays as well as regulate cell growth, cell division, and cellular responses to the environment.

  9. Cortical microtubule arrays are initiated from a nonrandom prepattern driven by atypical microtubule initiation.

    Science.gov (United States)

    Lindeboom, Jelmer J; Lioutas, Antonios; Deinum, Eva E; Tindemans, Simon H; Ehrhardt, David W; Emons, Anne Mie C; Vos, Jan W; Mulder, Bela M

    2013-03-01

    The ordered arrangement of cortical microtubules in growing plant cells is essential for anisotropic cell expansion and, hence, for plant morphogenesis. These arrays are dismantled when the microtubule cytoskeleton is rearranged during mitosis and reassembled following completion of cytokinesis. The reassembly of the cortical array has often been considered as initiating from a state of randomness, from which order arises at least partly through self-organizing mechanisms. However, some studies have shown evidence for ordering at early stages of array assembly. To investigate how cortical arrays are initiated in higher plant cells, we performed live-cell imaging studies of cortical array assembly in tobacco (Nicotiana tabacum) Bright Yellow-2 cells after cytokinesis and drug-induced disassembly. We found that cortical arrays in both cases did not initiate randomly but with a significant overrepresentation of microtubules at diagonal angles with respect to the cell axis, which coincides with the predominant orientation of the microtubules before their disappearance from the cell cortex in preprophase. In Arabidopsis (Arabidopsis thaliana) root cells, recovery from drug-induced disassembly was also nonrandom and correlated with the organization of the previous array, although no diagonal bias was observed in these cells. Surprisingly, during initiation, only about one-half of the new microtubules were nucleated from locations marked by green fluorescent protein-γ-tubulin complex protein2-tagged γ-nucleation complexes (γ-tubulin ring complex), therefore indicating that a large proportion of early polymers was initiated by a noncanonical mechanism not involving γ-tubulin ring complex. Simulation studies indicate that the high rate of noncanonical initiation of new microtubules has the potential to accelerate the rate of array repopulation.

  10. Drosophila Stathmin: A Microtubule-destabilizing Factor Involved in Nervous System Formation

    OpenAIRE

    Ozon, Sylvie; Guichet, Antoine; Gavet, Olivier; Roth, Siegfried; Sobel, André

    2002-01-01

    Stathmin is a ubiquitous regulatory phosphoprotein, the generic element of a family of neural phosphoproteins in vertebrates that possess the capacity to bind tubulin and interfere with microtubule dynamics. Although stathmin and the other proteins of the family have been associated with numerous cell regulations, their biological roles remain elusive, as in particular inactivation of the stathmin gene in the mouse resulted in no clear deleterious phenotype. We identified stathmin phosphoprot...

  11. The Vip1 inositol polyphosphate kinase family regulates polarized growth and modulates the microtubule cytoskeleton in fungi.

    Science.gov (United States)

    Pöhlmann, Jennifer; Risse, Carmen; Seidel, Constanze; Pohlmann, Thomas; Jakopec, Visnja; Walla, Eva; Ramrath, Pascal; Takeshita, Norio; Baumann, Sebastian; Feldbrügge, Michael; Fischer, Reinhard; Fleig, Ursula

    2014-09-01

    Microtubules (MTs) are pivotal for numerous eukaryotic processes ranging from cellular morphogenesis, chromosome segregation to intracellular transport. Execution of these tasks requires intricate regulation of MT dynamics. Here, we identify a new regulator of the Schizosaccharomyces pombe MT cytoskeleton: Asp1, a member of the highly conserved Vip1 inositol polyphosphate kinase family. Inositol pyrophosphates generated by Asp1 modulate MT dynamic parameters independent of the central +TIP EB1 and in a dose-dependent and cellular-context-dependent manner. Importantly, our analysis of the in vitro kinase activities of various S. pombe Asp1 variants demonstrated that the C-terminal phosphatase-like domain of the dual domain Vip1 protein negatively affects the inositol pyrophosphate output of the N-terminal kinase domain. These data suggest that the former domain has phosphatase activity. Remarkably, Vip1 regulation of the MT cytoskeleton is a conserved feature, as Vip1-like proteins of the filamentous ascomycete Aspergillus nidulans and the distantly related pathogenic basidiomycete Ustilago maydis also affect the MT cytoskeleton in these organisms. Consistent with the role of interphase MTs in growth zone selection/maintenance, all 3 fungal systems show aspects of aberrant cell morphogenesis. Thus, for the first time we have identified a conserved biological process for inositol pyrophosphates.

  12. The Vip1 inositol polyphosphate kinase family regulates polarized growth and modulates the microtubule cytoskeleton in fungi.

    Directory of Open Access Journals (Sweden)

    Jennifer Pöhlmann

    2014-09-01

    Full Text Available Microtubules (MTs are pivotal for numerous eukaryotic processes ranging from cellular morphogenesis, chromosome segregation to intracellular transport. Execution of these tasks requires intricate regulation of MT dynamics. Here, we identify a new regulator of the Schizosaccharomyces pombe MT cytoskeleton: Asp1, a member of the highly conserved Vip1 inositol polyphosphate kinase family. Inositol pyrophosphates generated by Asp1 modulate MT dynamic parameters independent of the central +TIP EB1 and in a dose-dependent and cellular-context-dependent manner. Importantly, our analysis of the in vitro kinase activities of various S. pombe Asp1 variants demonstrated that the C-terminal phosphatase-like domain of the dual domain Vip1 protein negatively affects the inositol pyrophosphate output of the N-terminal kinase domain. These data suggest that the former domain has phosphatase activity. Remarkably, Vip1 regulation of the MT cytoskeleton is a conserved feature, as Vip1-like proteins of the filamentous ascomycete Aspergillus nidulans and the distantly related pathogenic basidiomycete Ustilago maydis also affect the MT cytoskeleton in these organisms. Consistent with the role of interphase MTs in growth zone selection/maintenance, all 3 fungal systems show aspects of aberrant cell morphogenesis. Thus, for the first time we have identified a conserved biological process for inositol pyrophosphates.

  13. Calculation of the Electromagnetic Field Around a Microtubule

    Directory of Open Access Journals (Sweden)

    D. Havelka

    2009-01-01

    Full Text Available Microtubules are important structures in the cytoskeleton which organizes the cell. A single microtubule is composed of electrically polar structures, tubulin heterodimers, which have a strong electric dipole moment. Vibrations are expected to be generated in microtubules, thus tubulin heterodimers oscillate as electric dipoles. This gives rise to an electromagnetic field which is detected around the cells. We calculate here the electromagnetic field of microtubules if they are excited at 1 GHz. This paper includes work done for the bachelor thesis of the first author. 

  14. Population dynamics of dechlorinators and factors affecting the level and products of PCB dechlorination in sediments

    Energy Technology Data Exchange (ETDEWEB)

    Kim, J.S.; Sokol, R.C.; Liu, X.; Bethoney, C.M.; Rhee, G.Y. [State Univ. of New York and New York State Department of Health, Albany, NY (United States)

    1996-12-31

    Microbial dechlorination of polychlorinated biphenyls (PCBs) often stops although a significant number of removable chlorines remain. To determine the reason for the cessation, we investigated the limitation of organic carbon, PCB bioavailability, and inhibition by metabolic products. Enrichment with carbon sources did not induce additional chlorination, indicating the plateau was not due to depletion of organic carbon. The bioavailability was not limiting, since a subcritical micelle concentration of the surfactant, which enhanced desorption without inhibiting dechlorinating microorganisms, failed to lower the plateau. Neither was it due to accumulation of metabolites, since no additional dechlorination was detected when plateau sediments were incubated with fresh medium. Similarly, dechlorination was not inhibited in freshly spiked sediment slurries. Dechlorination ended up at the same level with nearly identical congener profiles, regardless of treatment. These results indicate that cessation of dechlorination was due to the accumulation of daughter congeners, which cannot be used as electron acceptors by microbes. To determine whether the decreasing availability affected the microorganisms, we determined the population dynamics of dechlorinators using the most probable number technique. The growth dynamics of the dechlorinators mirrored the time course of dechlorination. It started when the population increased by two orders of magnitude. Once dechlorination stopped the dechlorinating population also began to decrease. When dechlorinators were inoculated into PCB-free sediments, the population decreased over time. The decrease of the population as dechlorination ceased confirms that the diminishing availability of congeners was the reason for the incomplete dechlorination. Recent findings have shown that a second phase of dechlorination of certain congeners can occur after a long lag. 45 refs., 8 figs.

  15. Human SAS-6 C-Terminus Nucleates and Promotes Microtubule Assembly in Vitro by Binding to Microtubules.

    Science.gov (United States)

    Gupta, Hindol; Badarudeen, Binshad; George, Athira; Thomas, Geethu Emily; Gireesh, K K; Manna, Tapas K

    2015-10-20

    Centrioles are essential components of the animal centrosome and play crucial roles in the formation of cilia and flagella. They are cylindrical structures composed of nine triplet microtubules organized around a central cartwheel. Recent studies have identified spindle assembly abnormal protein SAS-6 as a critical component necessary for formation of the cartwheel. However, the molecular details of how the cartwheel participates in centriolar microtubule assembly have not been clearly understood. In this report, we show that the C-terminal tail (residues 470-657) of human SAS-6, HsSAS-6 C, the region that has been shown to extend toward the centriolar wall where the microtubule triplets are organized, nucleated and induced microtubule polymerization in vitro. The N-terminus (residues 1-166) of HsSAS-6, the domain known to be involved in formation of the central hub of the cartwheel, did not, however, exert any effect on microtubule polymerization. HsSAS-6 C bound to the microtubules and localized along the lengths of the microtubules in vitro. Microtubule pull-down and coimmunoprecipitation (Co-IP) experiments with S-phase synchronized HeLa cell lysates showed that the endogenous HsSAS-6 coprecipitated with the microtubules, and it mediated interaction with tubulin. Isothermal calorimetry titration and size exclusion chromatography showed that HsSAS-6 C bound to the αβ-tubulin dimer in vitro. The results demonstrate that HsSAS-6 possesses an intrinsic microtubule assembly promoting activity and further implicate that its outer exposed C-terminal tail may play critical roles in microtubule assembly and stabilizing microtubule attachment with the centriolar cartwheel.

  16. Interplay between kinesin-1 and cortical dynein during axonal outgrowth and microtubule organization in Drosophila neurons.

    Science.gov (United States)

    del Castillo, Urko; Winding, Michael; Lu, Wen; Gelfand, Vladimir I

    2015-12-28

    In this study, we investigated how microtubule motors organize microtubules in Drosophila neurons. We showed that, during the initial stages of axon outgrowth, microtubules display mixed polarity and minus-end-out microtubules push the tip of the axon, consistent with kinesin-1 driving outgrowth by sliding antiparallel microtubules. At later stages, the microtubule orientation in the axon switches from mixed to uniform polarity with plus-end-out. Dynein knockdown prevents this rearrangement and results in microtubules of mixed orientation in axons and accumulation of microtubule minus-ends at axon tips. Microtubule reorganization requires recruitment of dynein to the actin cortex, as actin depolymerization phenocopies dynein depletion, and direct recruitment of dynein to the membrane bypasses the actin requirement. Our results show that cortical dynein slides 'minus-end-out' microtubules from the axon, generating uniform microtubule arrays. We speculate that differences in microtubule orientation between axons and dendrites could be dictated by differential activity of cortical dynein.

  17. Heat-shock protein 70 binds microtubules and interacts with kinesin in tobacco pollen tubes.

    Science.gov (United States)

    Parrotta, Luigi; Cresti, Mauro; Cai, Giampiero

    2013-09-01

    The heat-shock proteins of 70 kDa are a family of ubiquitously expressed proteins important for protein folding. Heat-shock protein 70 assists other nascent proteins to achieve the spatial structure and ultimately helps the cell to protect against stress factors, such as heat. These proteins are localized in different cellular compartments and are associated with the cytoskeleton. We identified a heat-shock protein 70 isoform in the pollen tube of tobacco that binds to microtubules in an ATP-dependent manner. The heat-shock protein 70 was identified as part of the so-called ATP-MAP (ATP-dependent microtubule-associated protein) fraction, which also includes the 90-kDa kinesin, a mitochondria-associated motor protein. The identity of heat-shock protein 70 was validated by immunological assays and mass spectrometry. Sequence analysis showed that this heat-shock protein 70 is more similar to specific heat-shock proteins of Arabidopsis than to corresponding proteins of tobacco. Two-dimensional electrophoresis indicated that this heat-shock protein 70 isoform only is part of the ATP-MAP fraction and that is associated with the mitochondria of pollen tubes. Sedimentation assays showed that the binding of heat-shock protein 70 to microtubules is not affected by AMPPNP but it increases in the presence of the 90-kDa kinesin. Binding of heat-shock protein 70 to microtubules occurs only partially in the presence of ATP but it does not occur if, in addition to ATP, the 90-kDa kinesin is also present. Data suggest that the binding (but not the release) of heat-shock protein 70 to microtubules is facilitated by the 90-kDa kinesin.

  18. Tubers from patients with tuberous sclerosis complex are characterized by changes in microtubule biology through ROCK2 signalling.

    Science.gov (United States)

    Ferrer, Isidre; Mohan, Pooja; Chen, Helen; Castellsague, Joan; Gómez-Baldó, Laia; Carmona, Marga; García, Nadia; Aguilar, Helena; Jiang, Jihong; Skowron, Margaretha; Nellist, Mark; Ampuero, Israel; Russi, Antonio; Lázaro, Conxi; Maxwell, Christopher A; Pujana, Miguel Angel

    2014-07-01

    Most patients with tuberous sclerosis complex (TSC) develop cortical tubers that cause severe neurological disabilities. It has been suggested that defects in neuronal differentiation and/or migration underlie the appearance of tubers. However, the precise molecular alterations remain largely unknown. Here, by combining cytological and immunohistochemical analyses of tubers from nine TSC patients (four of them diagnosed with TSC2 germline mutations), we show that alteration of microtubule biology through ROCK2 signalling contributes to TSC neuropathology. All tubers showed a larger number of binucleated neurons than expected relative to control cortex. An excess of normal and altered cytokinetic figures was also commonly observed. Analysis of centrosomal markers suggested increased microtubule nucleation capacity, which was supported by the analysis of an expression dataset from cortical tubers and control cortex, and subsequently linked to under-expression of Rho-associated coiled-coil containing kinase 2 (ROCK2). Thus, augmented microtubule nucleation capacity was observed in mouse embryonic fibroblasts and human fibroblasts deficient in the Tsc2/TSC2 gene product, tuberin. Consistent with ROCK2 under-expression, microtubule acetylation was found to be increased with tuberin deficiency; this alteration was abrogated by rapamycin treatment and mimicked by HDAC6 inhibition. Together, the results of this study support the hypothesis that loss of TSC2 expression can alter microtubule organization and dynamics, which, in turn, deregulate cell division and potentially impair neuronal differentiation.

  19. Spatiotemporal dynamics of black-tailed prairie dog colonies affected by plague

    Science.gov (United States)

    Augustine, D.J.; Matchett, M.R.; Toombs, T.P.; Cully, J.F.; Johnson, T.L.; Sidle, John G.

    2008-01-01

    Black-tailed prairie dogs (Cynomys ludovicianus) are a key component of the disturbance regime in semi-arid grasslands of central North America. Many studies have compared community and ecosystem characteristics on prairie dog colonies to grasslands without prairie dogs, but little is known about landscape-scale patterns of disturbance that prairie dog colony complexes may impose on grasslands over long time periods. We examined spatiotemporal dynamics in two prairie dog colony complexes in southeastern Colorado (Comanche) and northcentral Montana (Phillips County) that have been strongly influenced by plague, and compared them to a complex unaffected by plague in northwestern Nebraska (Oglala). Both plague-affected complexes exhibited substantial spatiotemporal variability in the area occupied during a decade, in contrast to the stability of colonies in the Oglala complex. However, the plague-affected complexes differed in spatial patterns of colony movement. Colonies in the Comanche complex in shortgrass steppe shifted locations over a decade. Only 10% of the area occupied in 1995 was still occupied by prairie dogs in 2006. In 2005 and 2006 respectively, 74 and 83% of the total area of the Comanche complex occurred in locations that were not occupied in 1995, and only 1% of the complex was occupied continuously over a decade. In contrast, prairie dogs in the Phillips County complex in mixed-grass prairie and sagebrush steppe primarily recolonized previously occupied areas after plague-induced colony declines. In Phillips County, 62% of the area occupied in 1993 was also occupied by prairie dogs in 2004, and 12% of the complex was occupied continuously over a decade. Our results indicate that plague accelerates spatiotemporal movement of prairie dog colonies, and have significant implications for landscape-scale effects of prairie dog disturbance on grassland composition and productivity. These findings highlight the need to combine landscape-scale measures of

  20. Effects of a disease affecting a predator on the dynamics of a predator-prey system.

    Science.gov (United States)

    Auger, Pierre; McHich, Rachid; Chowdhury, Tanmay; Sallet, Gauthier; Tchuente, Maurice; Chattopadhyay, Joydev

    2009-06-07

    We study the effects of a disease affecting a predator on the dynamics of a predator-prey system. We couple an SIRS model applied to the predator population, to a Lotka-Volterra model. The SIRS model describes the spread of the disease in a predator population subdivided into susceptible, infected and removed individuals. The Lotka-Volterra model describes the predator-prey interactions. We consider two time scales, a fast one for the disease and a comparatively slow one for predator-prey interactions and for predator mortality. We use the classical "aggregation method" in order to obtain a reduced equivalent model. We show that there are two possible asymptotic behaviors: either the predator population dies out and the prey tends to its carrying capacity, or the predator and prey coexist. In this latter case, the predator population tends either to a "disease-free" or to a "disease-endemic" state. Moreover, the total predator density in the disease-endemic state is greater than the predator density in the "disease-free" equilibrium (DFE).

  1. A dynamic evolution model of human opinion as affected by advertising

    Science.gov (United States)

    Luo, Gui-Xun; Liu, Yun; Zeng, Qing-An; Diao, Su-Meng; Xiong, Fei

    2014-11-01

    We propose a new model to investigate the dynamics of human opinion as affected by advertising, based on the main idea of the CODA model and taking into account two practical factors: one is that the marginal influence of an additional friend will decrease with an increasing number of friends; the other is the decline of memory over time. Simulations show several significant conclusions for both advertising agencies and the general public. A small difference of advertising’s influence on individuals or advertising coverage will result in significantly different advertising effectiveness within a certain interval of value. Compared to the value of advertising’s influence on individuals, the advertising coverage plays a more important role due to the exponential decay of memory. Meanwhile, some of the obtained results are in accordance with people’s daily cognition about advertising. The real key factor in determining the success of advertising is the intensity of exchanging opinions, and people’s external actions always follow their internal opinions. Negative opinions also play an important role.

  2. Carbon and nitrogen dynamics in early stages of forest litter decomposition as affected by nitrogen addition

    Institute of Scientific and Technical Information of China (English)

    DENG Xiao-wen; LIU Ying; HAN Shi-jie

    2009-01-01

    The effects of nitrogen (N) availability and tree species on the dynamics of carbon and nitrogen at early stage of decomposition of forest litter were studied in a 13-week laboratory incubation experiment. Fresh litter samples including needle litter (Pinus koraiensis) and two types of broadleaf litters (Quercus mongolica and Tilia amurensis) were collected from a broadleaf-korean pine mixed forest in the northern slope of Changbai Mountain (China). Different doses of N (equal to 0, 30 and 50 kg·ha-1yr-1, respectively, as NH4NO3) were added to litter during the experiment period. The litter decomposition rate expressed as mass loss and respiration rate increased significantly with increasing N availability. The mass loss and cumulative CO2-C emission were higher in leaf litter compared to that in needle litter. The dissolved organic Carbon (DOC) concentrations in litter leachate varied widely between the species, but were not greatly affected by N treatments. Regardless of the N addition rate, both N treatments and species had no significant effect on dissolved organic N (DON) concentrations in litter leachate. About 52·78% of added N was retained in the litter. The percentage of N retention was positively correlated (R2=0.91, p<0.05) with the litter mass loss. This suggested that a forest floor with easily decomposed litter might have higher potential N sink strength than that with more slowly decomposed litter.

  3. How Major Depressive Disorder affects the ability to decode multimodal dynamic emotional stimuli

    Directory of Open Access Journals (Sweden)

    FILOMENA SCIBELLI

    2016-09-01

    Full Text Available Most studies investigating the processing of emotions in depressed patients reported impairments in the decoding of negative emotions. However, these studies adopted static stimuli (mostly stereotypical facial expressions corresponding to basic emotions which do not reflect the way people experience emotions in everyday life. For this reason, this work proposes to investigate the decoding of emotional expressions in patients affected by Recurrent Major Depressive Disorder (RMDDs using dynamic audio/video stimuli. RMDDs’ performance is compared with the performance of patients with Adjustment Disorder with Depressed Mood (ADs and healthy (HCs subjects. The experiments involve 27 RMDDs (16 with acute depression - RMDD-A, and 11 in a compensation phase - RMDD-C, 16 ADs and 16 HCs. The ability to decode emotional expressions is assessed through an emotion recognition task based on short audio (without video, video (without audio and audio/video clips. The results show that AD patients are significantly less accurate than HCs in decoding fear, anger, happiness, surprise and sadness. RMDD-As with acute depression are significantly less accurate than HCs in decoding happiness, sadness and surprise. Finally, no significant differences were found between HCs and RMDD-Cs in a compensation phase. The different communication channels and the types of emotion play a significant role in limiting the decoding accuracy.

  4. Microtubule binding by the formin Cappuccino and its implications for Drosophila oogenesis

    OpenAIRE

    Roth-Johnson, Elizabeth Anne

    2014-01-01

    Coordination of actin and microtubule cytoskeletal networks is required for a number of fundamental cellular processes. Formin family actin nucleators are emerging coordinators of the actin and microtubule cytoskeletons, as they can both nucleate actin filaments and bind microtubules in vitro. To gain a more detailed mechanistic understanding of formin-microtubule interactions and formin-mediated actin-microtubule crosstalk, we studied microtubule binding by Cappuccino (Capu), a formin involv...

  5. In vivo FRET imaging revealed a regulatory role of RanGTP in kinetochore-microtubule attachments via Aurora B kinase.

    Directory of Open Access Journals (Sweden)

    Yoke-Peng Lee

    Full Text Available Under the fluctuating circumstances provided by the innate dynamics of microtubules and opposing tensions resulted from microtubule-associated motors, it is vital to ensure stable kinetochore-microtubule attachments for accurate segregation. However, a comprehensive understanding of how this regulation is mechanistically achieved remains elusive. Using our newly designed live cell FRET time-lapse imaging, we found that post-metaphase RanGTP is crucial in the maintenance of stable kinetochore-microtubule attachments by regulating Aurora B kinase via the NES-bearing Mst1. More importantly, our study demonstrates that by ensuring stable alignment of metaphase chromosomes prior to segregation, RanGTP is indispensible in governing the genomic integrity and the fidelity of cell cycle progression. Our findings suggest an additional role of RanGTP beyond its known function in mitotic spindle assembly during the prometaphase-metaphase transition.

  6. Effects of mutual interaction of Laccaria laccata with Trichoderma harzianum and T. virens on the morphology of microtubules and mitochondria.

    Science.gov (United States)

    Zadworny, M; Tuszyńska, S; Samardakiewicz, S; Werner, A

    2007-01-01

    Organelles are known to respond to challenges caused by many stress factors. The morphology of the microtubular cytoskeleton and mitochondria during mutual interaction in coculture of Laccaria laccata with Trichoderma harzianum and T. virens were examined. Hyphae from the interaction region were sampled between 4 and 12 days of growth. Microtubules were labelled with a specific antibody and mitochondria with 3,3'-dihexyloxacarbocyanine iodide, and the organelles were examined microscopically. The morphology of microtubules and mitochondria were similar in all three fungi. Microtubules were arranged in long arrays parallel to the hyphal axis and mitochondria formed an interconnected network. In hyphae growing within the interaction zone, microtubules became wavy and eventually fragmented or depolymerised, and mitochondria also became fragmented. The effects were time-dependent. In general, the organelles of all three fungi were affected during the interaction, but L. laccata was affected the least and to the same extent by each of the saprotrophic fungi. The saprotrophic fungi were affected by L. laccata to a similar extent at 4 and 8 days of interaction. Our results suggest that the studied fungi antagonistically affect each other at the cellular level, although the mechanisms involved remain to be elucidated.

  7. Molecular Pathway of Microtubule Organization at the Golgi Apparatus

    NARCIS (Netherlands)

    Wu, Jingchao; de Heus, Cecilia; Liu, Qingyang; Bouchet, Benjamin P; Noordstra, Ivar; Jiang, Kai; Hua, Shasha; Martin, Maud; Yang, Chao; Grigoriev, Ilya; Katrukha, Eugene A; Altelaar, A F Maarten; Hoogenraad, Casper C; Qi, Robert Z; Klumperman, Judith; Akhmanova, Anna

    2016-01-01

    The Golgi apparatus controls the formation of non-centrosomal microtubule arrays important for Golgi organization, polarized transport, cell motility, and cell differentiation. Here, we show that CAMSAP2 stabilizes and attaches microtubule minus ends to the Golgi through a complex of AKAP450 and myo

  8. Resolving bundled microtubules using anti-tubulin nanobodies

    NARCIS (Netherlands)

    Mikhaylova, Marina; Cloin, Bas M C; Finan, Kieran; van den Berg, Robert; Teeuw, Jalmar; Kijanka, Marta M; Sokolowski, Mikolaj; Katrukha, Eugene A; Maidorn, Manuel; Opazo, Felipe; Moutel, Sandrine; Vantard, Marylin; Perez, Frank; van Bergen en Henegouwen, Paul M P; Hoogenraad, Casper C; Ewers, Helge; Kapitein, Lukas C

    2015-01-01

    Microtubules are hollow biopolymers of 25-nm diameter and are key constituents of the cytoskeleton. In neurons, microtubules are organized differently between axons and dendrites, but their precise organization in different compartments is not completely understood. Super-resolution microscopy techn

  9. Interconnections between cell wall polymers, wall mechanics, and cortical microtubules: Teasing out causes and consequences.

    Science.gov (United States)

    Xiao, Chaowen; Anderson, Charles T

    2016-09-01

    In plants, cell wall components including cellulose, hemicelluloses, and pectins interact with each other to form complex extracellular network structures that control cell growth and maintain cell shape. However, it is still not clear exactly how different wall polymers interact, how the conformations and interactions of cell wall polymers relate to wall mechanics, and how these factors impinge on intracellular structures such as the cortical microtubule cytoskeleton. Here, based on studies of Arabidopsis thaliana xxt1 xxt2 mutants, which lack detectable xyloglucan in their walls and display aberrant wall mechanics, altered cellulose patterning and biosynthesis, and reduced cortical microtubule stability, we discuss the potential relationships between cell wall biosynthesis, wall mechanics, and cytoskeletal dynamics in an effort to better understand their roles in controlling plant growth and morphogenesis.

  10. Pironetin reacts covalently with cysteine-316 of α-tubulin to destabilize microtubule

    Science.gov (United States)

    Yang, Jianhong; Wang, Yuxi; Wang, Taijing; Jiang, Jian; Botting, Catherine H.; Liu, Huanting; Chen, Qiang; Yang, Jinliang; Naismith, James H.; Zhu, Xiaofeng; Chen, Lijuan

    2016-06-01

    Molecules that alter the normal dynamics of microtubule assembly and disassembly include many anticancer drugs in clinical use. So far all such therapeutics target β-tubulin, and structural biology has explained the basis of their action and permitted design of new drugs. However, by shifting the profile of β-tubulin isoforms, cancer cells become resistant to treatment. Compounds that bind to α-tubulin are less well characterized and unexploited. The natural product pironetin is known to bind to α-tubulin and is a potent inhibitor of microtubule polymerization. Previous reports had identified that pironetin reacts with lysine-352 residue however analogues designed on this model had much lower potency, which was difficult to explain, hindering further development. We report crystallographic and mass spectrometric data that reveal that pironetin forms a covalent bond to cysteine-316 in α-tubulin via a Michael addition reaction. These data provide a basis for the rational design of α-tubulin targeting chemotherapeutics.

  11. Asymptotic Analysis of Microtubule-Based Transport by Multiple Identical Molecular Motors

    CERN Document Server

    McKinley, Scott A; Fricks, John; Kramer, Peter R

    2011-01-01

    We describe a system of stochastic differential equations (SDEs) which model the interaction between processive molecular motors, such as kinesin and dynein, and the biomolecular cargo they tow as part of microtubule-based intracellular transport. We show that the classical experimental environment fits within a parameter regime which is qualitatively distinct from conditions one expects to find in living cells. Through an asymptotic analysis of our system of SDEs, we develop a means for applying in vitro observations of the nonlinear response by motors to forces induced on the attached cargo to make analytical predictions for two parameter regimes that have thus far eluded direct experimental observation: 1) highly viscous in vivo transport and 2) dynamics when multiple identical motors are attached to the cargo and microtubule.

  12. Microtubules in Plant Cells: Strategies and Methods for Immunofluorescence, Transmission Electron Microscopy and Live Cell Imaging

    Science.gov (United States)

    Celler, Katherine; Fujita, Miki; Kawamura, Eiko; Ambrose, Chris; Herburger, Klaus; Wasteneys, Geoffrey O.

    2016-01-01

    Microtubules are required throughout plant development for a wide variety of processes, and different strategies have evolved to visualize and analyze them. This chapter provides specific methods that can be used to analyze microtubule organization and dynamic properties in plant systems and summarizes the advantages and limitations for each technique. We outline basic methods for preparing samples for immunofluorescence labelling, including an enzyme-based permeabilization method, and a freeze-shattering method, which generates microfractures in the cell wall to provide antibodies access to cells in cuticle-laden aerial organs such as leaves. We discuss current options for live cell imaging of MTs with fluorescently tagged proteins (FPs), and provide chemical fixation, high pressure freezing/freeze substitution, and post-fixation staining protocols for preserving MTs for transmission electron microscopy and tomography. PMID:26498784

  13. Role of the Number of Microtubules in Chromosome Segregation during Cell Division

    CERN Document Server

    Bertalan, Zsolt; La Porta, Caterina A M; Zapperi, Stefano

    2015-01-01

    Faithful segregation of genetic material during cell division requires alignment of chromosomes between two spindle poles and attachment of their kinetochores to each of the poles. Failure of these complex dynamical processes leads to chromosomal instability (CIN), a characteristic feature of several diseases including cancer. While a multitude of biological factors regulating chromosome congression and bi-orientation have been identified, it is still unclear how they are integrated so that coherent chromosome motion emerges from a large collection of random and deterministic processes. Here we address this issue by a three dimensional computational model of motor-driven chromosome congression and bi-orientation during mitosis. Our model reveals that successful cell division requires control of the total number of microtubules: if this number is too small bi-orientation fails, while if it is too large not all the chromosomes are able to congress. The optimal number of microtubules predicted by our model compa...

  14. HILI destabilizes microtubules by suppressing phosphorylation and Gigaxonin-mediated degradation of TBCB

    Science.gov (United States)

    Tan, Hao; Liao, Hua; Zhao, Lianfang; Lu, Yilu; Jiang, Siyuan; Tao, Dachang; Liu, Yunqiang; Ma, Yongxin

    2017-01-01

    Human PIWIL2, aka HILI, is a member of PIWI protein family and overexpresses in various tumors. However, the underlying mechanisms of HILI in tumorigenesis remain largely unknown. TBCB has a critical role in regulating microtubule dynamics and is overexpressed in many cancers. Here we report that HILI inhibits Gigaxonin-mediated TBCB ubiquitination and degradation by interacting with TBCB, promoting the binding between HSP90 and TBCB, and suppressing the interaction between Gigaxonin and TBCB. Meanwhile, HILI can also reduce phosphorylation level of TBCB induced by PAK1. Our results showed that HILI suppresses microtubule polymerization and promotes cell proliferation, migration and invasion via TBCB for the first time, revealing a novel mechanism for HILI in tumorigenesis. PMID:28393858

  15. Multimodal microtubule binding by the Ndc80 kinetochore complex.

    Science.gov (United States)

    Alushin, Gregory M; Musinipally, Vivek; Matson, Daniel; Tooley, John; Stukenberg, P Todd; Nogales, Eva

    2012-11-01

    The Ndc80 complex is a key site of kinetochore-microtubule attachment during cell division. The human complex engages microtubules with a globular 'head' formed by tandem calponin-homology domains and an 80-amino-acid unstructured 'tail' that contains sites of phosphoregulation by the Aurora B kinase. Using biochemical, cell biological and electron microscopy analyses, we dissected the roles of the tail in binding of microtubules and mediation of cooperative interactions between Ndc80 complexes. Two segments of the tail that contain Aurora B phosphorylation sites become ordered at interfaces; one with tubulin and the second with an adjacent Ndc80 head on the microtubule surface, forming interactions that are disrupted by phosphorylation. We propose a model in which Ndc80's interaction with either growing or shrinking microtubule ends can be tuned by the phosphorylation state of its tail.

  16. Tensile stress stimulates microtubule outgrowth in living cells

    Science.gov (United States)

    Kaverina, Irina; Krylyshkina, Olga; Beningo, Karen; Anderson, Kurt; Wang, Yu-Li; Small, J. Victor

    2002-01-01

    Cell motility is driven by the sum of asymmetric traction forces exerted on the substrate through adhesion foci that interface with the actin cytoskeleton. Establishment of this asymmetry involves microtubules, which exert a destabilising effect on adhesion foci via targeting events. Here, we demonstrate the existence of a mechano-sensing mechanism that signals microtubule polymerisation and guidance of the microtubules towards adhesion sites under increased stress. Stress was applied either by manipulating the body of cells moving on glass with a microneedle or by stretching a flexible substrate that cells were migrating on. We propose a model for this mechano-sensing phenomenon whereby microtubule polymerisation is stimulated and guided through the interaction of a microtubule tip complex with actin filaments under tension.

  17. Emotion expression of an affective state space; a humanoid robot displaying a dynamic emotional state during a soccer game

    NARCIS (Netherlands)

    van der Mey, A.; Smit, F; Droog, K.J.; Visser, A.

    2010-01-01

    Following a soccer game is an example where clear emotions are displayed. This example is worked out for a humanoid robot which can express emotions with body language. The emotions expressed by the robot are not just stimuli-response, but are based on an affective state which shows dynamic behavior

  18. Nitrogen storage dynamics are affected by masting events in Fagus crenata.

    Science.gov (United States)

    Han, Qingmin; Kabeya, Daisuke; Iio, Atsuhiro; Inagaki, Yoshiyuki; Kakubari, Yoshitaka

    2014-03-01

    It is generally assumed that the production of a large crop of seeds depletes stores of resources and that these take more than 1 year to replenish; this is accepted, theoretically, as the proximate mechanism of mast seeding (resource budget model). However, direct evidence of resource depletion in masting trees is very rare. Here, we trace seasonal and inter-annual variations in nitrogen (N) concentration and estimate the N storage pool of individuals after full masting of Fagus crenata in two stands. In 2005, a full masting year, the amount of N in fruit litter represented half of the N present in mature leaves in an old stand (age 190-260 years), and was about equivalent to the amount of N in mature leaves in a younger stand (age 83-84 years). Due to this additional burden, both tissue N concentration and individual N storage decreased in 2006; this was followed by significant replenishment in 2007, although a substantial N store remained even after full masting. These results indicate that internal storage may be important and that N may be the limiting factor for fruiting. In the 4 years following full masting, the old stand experienced two moderate masting events separated by 2 years, whilst trees in the younger stand did not fruit. This different fruiting behavior may be related to different "costs of reproduction" in the full masting year 2005, thus providing more evidence that N may limit fruiting. Compared to the non-fruiting stand, individuals in the fruiting stand exhibited an additional increase in N concentrations in roots early in the 2007 growing season, suggesting additional N uptake from the soil to supply resource demand. The enhanced uptake may alleviate the N storage depletion observed in the full masting year. This study suggests that masting affects N cycle dynamics in mature Fagus crenata and N may be one factor limiting fruiting.

  19. Efficient event-driven simulations shed new light on microtubule organization in the plant cortical array

    Science.gov (United States)

    Tindemans, Simon H.; Deinum, Eva E.; Lindeboom, Jelmer J.; Mulder, Bela M.

    2014-04-01

    The dynamics of the plant microtubule cytoskeleton is a paradigmatic example of the complex spatiotemporal processes characterising life at the cellular scale. This system is composed of large numbers of spatially extended particles, each endowed with its own intrinsic stochastic dynamics, and is capable of non-equilibrium self-organisation through collisional interactions of these particles. To elucidate the behaviour of such a complex system requires not only conceptual advances, but also the development of appropriate computational tools to simulate it. As the number of parameters involved is large and the behaviour is stochastic, it is essential that these simulations be fast enough to allow for an exploration of the phase space and the gathering of sufficient statistics to accurately pin down the average behaviour as well as the magnitude of fluctuations around it. Here we describe a simulation approach that meets this requirement by adopting an event-driven methodology that encompasses both the spontaneous stochastic changes in microtubule state as well as the deterministic collisions. In contrast with finite time step simulations this technique is intrinsically exact, as well as several orders of magnitude faster, which enables ordinary PC hardware to simulate systems of ˜ 10^3 microtubules on a time scale ˜ 10^{3} faster than real time. In addition we present new tools for the analysis of microtubule trajectories on curved surfaces. We illustrate the use of these methods by addressing a number of outstanding issues regarding the importance of various parameters on the transition from an isotropic to an aligned and oriented state.

  20. Efficient event-driven simulations shed new light on microtubule organisation in the plant cortical array

    Directory of Open Access Journals (Sweden)

    Simon H. Tindemans

    2014-04-01

    Full Text Available The dynamics of the plant microtubule cytoskeleton is a paradigmatic example of the complex spatiotemporal processes characterising life at the cellular scale. This system is composed of large numbers of spatially extended particles, each endowed with its own intrinsic stochastic dynamics, and is capable of non-equilibrium self-organisation through collisional interactions of these particles. To elucidate the behaviour of such a complex system requires not only conceptual advances, but also the development of appropriate computational tools to simulate it. As the number of parameters involved is large and the behaviour is stochastic, it is essential that these simulations be fast enough to allow for an exploration of the phase space and the gathering of sufficient statistics to accurately pin down the average behaviour as well as the magnitude of fluctuations around it. Here we describe a simulation approach that meets this requirement by adopting an event-driven methodology that encompasses both the spontaneous stochastic changes in microtubule state as well as the deterministic collisions. In contrast with finite time step simulations this technique is intrinsically exact, as well as several orders of magnitude faster, which enables ordinary PC hardware to simulate systems of $sim 10^3$ microtubules on a time scale $sim 10^{3}$ faster than real time. In addition we present new tools for the analysis of microtubule trajectories on curved surfaces. We illustrate the use of these methods by addressing a number of outstanding issues regarding the importance of various parameters on the transition from an isotropic to an aligned and oriented state.

  1. Examination of actin and microtubule dependent APC localisations in living mammalian cells

    Directory of Open Access Journals (Sweden)

    Adams Matthew

    2006-01-01

    Full Text Available Abstract Background The trafficking of the adenomatous polyposis coli (APC tumour suppressor protein in mammalian cells is a perennially controversial topic. Immunostaining evidence for an actin-associated APC localisation at intercellular junctions has been previously presented, though live imaging of mammalian junctional APC has not been documented. Results Using live imaging of transfected COS-7 cells we observed intercellular junction-associated pools of GFP-APC in addition to previously documented microtubule-associated GFP-APC and a variety of minor localisations. Although both microtubule and junction-associated populations could co-exist within individual cells, they differed in their subcellular location, dynamic behaviour and sensitivity to cytoskeletal poisons. GFP-APC deletion mutant analysis indicated that a protein truncated immediately after the APC armadillo repeat domain retained the ability to localise to adhesive membranes in transfected cells. Supporting this, we also observed junctional APC immunostaining in cultures of human colorectal cancer cell line that express truncated forms of APC. Conclusion Our data indicate that APC can be found in two spatially separate populations at the cell periphery and these populations can co-exist in the same cell. The first localisation is highly dynamic and associated with microtubules near free edges and in cell vertices, while the second is comparatively static and is closely associated with actin at sites of cell-cell contact. Our imaging confirms that human GFP-APC possesses many of the localisations and behaviours previously seen by live imaging of Xenopus GFP-APC. However, we report the novel finding that GFP-APC puncta can remain associated with the ends of shrinking microtubules. Deletion analysis indicated that the N-terminal region of the APC protein mediated its junctional localisation, consistent with our observation that truncated APC proteins in colon cancer cell lines are

  2. KIF7 Controls the Proliferation of Cells of the Respiratory Airway through Distinct Microtubule Dependent Mechanisms.

    Directory of Open Access Journals (Sweden)

    Garry L Coles

    2015-10-01

    Full Text Available The cell cycle must be tightly coordinated for proper control of embryonic development and for the long-term maintenance of organs such as the lung. There is emerging evidence that Kinesin family member 7 (Kif7 promotes Hedgehog (Hh signaling during embryonic development, and its misregulation contributes to diseases such as ciliopathies and cancer. Kif7 encodes a microtubule interacting protein that controls Hh signaling through regulation of microtubule dynamics within the primary cilium. However, whether Kif7 has a function in nonciliated cells remains largely unknown. The role Kif7 plays in basic cell biological processes like cell proliferation or cell cycle progression also remains to be elucidated. Here, we show that Kif7 is required for coordination of the cell cycle, and inactivation of this gene leads to increased cell proliferation in vivo and in vitro. Immunostaining and transmission electron microscopy experiments show that Kif7dda/dda mutant lungs are hyperproliferative and exhibit reduced alveolar epithelial cell differentiation. KIF7 depleted C3H10T1/2 fibroblasts and Kif7dda/dda mutant mouse embryonic fibroblasts have increased growth rates at high cellular densities, suggesting that Kif7 may function as a general regulator of cellular proliferation. We ascertained that in G1, Kif7 and microtubule dynamics regulate the expression and activity of several components of the cell cycle machinery known to control entry into S phase. Our data suggest that Kif7 may function to regulate the maintenance of the respiratory airway architecture by controlling cellular density, cell proliferation, and cycle exit through its role as a microtubule associated protein.

  3. Cap-Gly proteins at microtubule plus ends: is EB1 detyrosination involved?

    Directory of Open Access Journals (Sweden)

    Anouk Bosson

    Full Text Available Localization of CAP-Gly proteins such as CLIP170 at microtubule+ends results from their dual interaction with α-tubulin and EB1 through their C-terminal amino acids -EEY. Detyrosination (cleavage of the terminal tyrosine of α-tubulin by tubulin-carboxypeptidase abolishes CLIP170 binding. Can detyrosination affect EB1 and thus regulate the presence of CLIP170 at microtubule+ends as well? We developed specific antibodies to discriminate tyrosinated vs detyrosinated forms of EB1 and detected only tyrosinated EB1 in fibroblasts, astrocytes, and total brain tissue. Over-expressed EB1 was not detyrosinated in cells and chimeric EB1 with the eight C-terminal amino acids of α-tubulin was only barely detyrosinated. Our results indicate that detyrosination regulates CLIPs interaction with α-tubulin, but not with EB1. They highlight the specificity of carboxypeptidase toward tubulin.

  4. Cap-Gly proteins at microtubule plus ends: is EB1 detyrosination involved?

    Science.gov (United States)

    Bosson, Anouk; Soleilhac, Jean-Marc; Valiron, Odile; Job, Didier; Andrieux, Annie; Moutin, Marie-Jo

    2012-01-01

    Localization of CAP-Gly proteins such as CLIP170 at microtubule+ends results from their dual interaction with α-tubulin and EB1 through their C-terminal amino acids -EEY. Detyrosination (cleavage of the terminal tyrosine) of α-tubulin by tubulin-carboxypeptidase abolishes CLIP170 binding. Can detyrosination affect EB1 and thus regulate the presence of CLIP170 at microtubule+ends as well? We developed specific antibodies to discriminate tyrosinated vs detyrosinated forms of EB1 and detected only tyrosinated EB1 in fibroblasts, astrocytes, and total brain tissue. Over-expressed EB1 was not detyrosinated in cells and chimeric EB1 with the eight C-terminal amino acids of α-tubulin was only barely detyrosinated. Our results indicate that detyrosination regulates CLIPs interaction with α-tubulin, but not with EB1. They highlight the specificity of carboxypeptidase toward tubulin.

  5. Effects of spermine binding on Taxol-stabilized microtubules

    Science.gov (United States)

    Cheng, Shengfeng; Regmi, Chola

    Previous studies have shown that polyamines such as spermine present in cells at physiological concentrations can facilitate the polymerization of tubulins into microtubules (MTs). A recent experiment demonstrates that in the presence of high-concentration spermine, Taxol-stabilized MTs undergo a shape transformation into inverted tubulin tubules (ITTs), the outside surface of which corresponds to the inside surface of a regular MT. However, the molecular mechanism underlying the shape transformation of MTs into ITTs is unclear. We perform all atom molecular dynamics simulations on Taxol-stabilized MT sheets containing two protofilaments surrounded by spermine ions. The spermine concentration is varied from 0 to 25mM to match the range probed experimentally. We identify important spermine binding regions on the MT surface and the influence of the spermine binding on the structure and dynamics of MTs. In contrast to Taxol, our results show that spermine binding seems to decrease the flexibility of tubulin proteins, resulting in weaker tubulin-tubulin contacts and promoting the bending of protofilaments into curved protofilaments, inverted rings, and eventually inverted tubules.

  6. Kinesins have a dual function in organizing microtubules during both tip growth and cytokinesis in Physcomitrella patens.

    Science.gov (United States)

    Hiwatashi, Yuji; Sato, Yoshikatsu; Doonan, John H

    2014-03-01

    Microtubules (MTs) play a crucial role in the anisotropic deposition of cell wall material, thereby affecting the direction of growth. A wide range of tip-growing cells display highly polarized cell growth, and MTs have been implicated in regulating directionality and expansion. However, the molecular machinery underlying MT dynamics in tip-growing plant cells remains unclear. Here, we show that highly dynamic MT bundles form cyclically in the polarized expansion zone of the moss Physcomitrella patens caulonemal cells through the coalescence of growing MT plus ends. Furthermore, the plant-specific kinesins (KINID1) that are is essential for the proper MT organization at cytokinesis also regulate the turnover of the tip MT bundles as well as the directionality and rate of cell growth. The plus ends of MTs grow toward the expansion zone, and KINID1 is necessary for the stability of a single coherent focus of MTs in the center of the zone, whose formation coincides with the accumulation of KINID1. We propose that KINID-dependent MT bundling is essential for the correct directionality of growth as well as for promoting growth per se. Our findings indicate that two localized cell wall deposition processes, tip growth and cytokinesis, previously believed to be functionally and evolutionarily distinct, share common and plant-specific MT regulatory components.

  7. Dynamic Association between Negative Affect and Alcohol Lapses following Alcohol Treatment

    Science.gov (United States)

    Witkiewitz, Katie; Villarroel, Nadia Aracelliz

    2009-01-01

    Clinical research has found a strong association between negative affect and returning to alcohol use after a period of abstinence. Yet little is known about the probability of a lapse given a particular level of negative affect or whether there is a reciprocal relationship between negative affect and alcohol use across time. The goal of the…

  8. A pseudo-spin model for the wall of cytoskeletal microtubule

    Institute of Scientific and Technical Information of China (English)

    Chen Ying; Qiu Xi-Jun; Li Ru-Xin

    2005-01-01

    A pseudo-spin model is intended to describe the physical dynamics of unbound electrons in the wall of cytoskeletal microtubule (MT). Due to the inherent symmetry of the structure and the electric properties in the MT, one may treat it as a one-dimensional ferroelectric system, and describe the nonlinear dynamics of dimer electric dipoles in one protofilament of the MT by virtue of the double-well potential. Consequently, the physical problem has been mapped onto the pseudo-spin system, and the mean-field approximation has been taken to get some physical results.

  9. Communication: Microsecond dynamics of the protein and water affect electron transfer in a bacterial bc{sub 1} complex

    Energy Technology Data Exchange (ETDEWEB)

    Martin, Daniel R.; Matyushov, Dmitry V., E-mail: dmitrym@asu.edu [Department of Physics and Department of Chemistry and Biochemistry, Arizona State University, P.O. Box 871504, Tempe, Arizona 85287 (United States)

    2015-04-28

    Cross-membrane electron transport between cofactors localized in proteins of mitochondrial respiration and bacterial photosynthesis is the source of all biological energy. The statistics and dynamics of nuclear fluctuations in these protein/membrane/water heterogeneous systems are critical for their energetic efficiency. The results of 13 μs of atomistic molecular dynamics simulations of the membrane-bound bc{sub 1} bacterial complex are analyzed here. The reaction is affected by a broad spectrum of nuclear modes, with the slowest dynamics in the range of time-scales ∼0.1-1.6 μs contributing half of the reaction reorganization energy. Two reorganization energies are required to describe protein electron transfer due to dynamical arrest of protein conformations on the observation window. This mechanistic distinction allows significant lowering of activation barriers for reactions in proteins.

  10. Tripolin A, a novel small-molecule inhibitor of aurora A kinase, reveals new regulation of HURP's distribution on microtubules.

    Directory of Open Access Journals (Sweden)

    Iliana A Kesisova

    Full Text Available Mitotic regulators exhibiting gain of function in tumor cells are considered useful cancer therapeutic targets for the development of small-molecule inhibitors. The human Aurora kinases are a family of such targets. In this study, from a panel of 105 potential small-molecule inhibitors, two compounds Tripolin A and Tripolin B, inhibited Aurora A kinase activity in vitro. In human cells however, only Tripolin A acted as an Aurora A inhibitor. We combined in vitro, in vivo single cell and in silico studies to demonstrate the biological action of Tripolin A, a non-ATP competitive inhibitor. Tripolin A reduced the localization of pAurora A on spindle microtubules (MTs, affected centrosome integrity, spindle formation and length, as well as MT dynamics in interphase, consistent with Aurora A inhibition by RNAi or other specific inhibitors, such as MLN8054 or MLN8237. Interestingly, Tripolin A affected the gradient distribution towards the chromosomes, but not the MT binding of HURP (Hepatoma Up-Regulated Protein, a MT-associated protein (MAP and substrate of the Aurora A kinase. Therefore Tripolin A reveals a new way of regulating mitotic MT stabilizers through Aurora A phosphorylation. Tripolin A is predicted to bind Aurora A similarly but not identical to MLN8054, therefore it could be used to dissect pathways orchestrated by Aurora kinases as well as a scaffold for further inhibitor development.

  11. Tripolin A, a novel small-molecule inhibitor of aurora A kinase, reveals new regulation of HURP's distribution on microtubules.

    Science.gov (United States)

    Kesisova, Iliana A; Nakos, Konstantinos C; Tsolou, Avgi; Angelis, Dimitrios; Lewis, Joe; Chatzaki, Aikaterini; Agianian, Bogos; Giannis, Athanassios; Koffa, Maria D

    2013-01-01

    Mitotic regulators exhibiting gain of function in tumor cells are considered useful cancer therapeutic targets for the development of small-molecule inhibitors. The human Aurora kinases are a family of such targets. In this study, from a panel of 105 potential small-molecule inhibitors, two compounds Tripolin A and Tripolin B, inhibited Aurora A kinase activity in vitro. In human cells however, only Tripolin A acted as an Aurora A inhibitor. We combined in vitro, in vivo single cell and in silico studies to demonstrate the biological action of Tripolin A, a non-ATP competitive inhibitor. Tripolin A reduced the localization of pAurora A on spindle microtubules (MTs), affected centrosome integrity, spindle formation and length, as well as MT dynamics in interphase, consistent with Aurora A inhibition by RNAi or other specific inhibitors, such as MLN8054 or MLN8237. Interestingly, Tripolin A affected the gradient distribution towards the chromosomes, but not the MT binding of HURP (Hepatoma Up-Regulated Protein), a MT-associated protein (MAP) and substrate of the Aurora A kinase. Therefore Tripolin A reveals a new way of regulating mitotic MT stabilizers through Aurora A phosphorylation. Tripolin A is predicted to bind Aurora A similarly but not identical to MLN8054, therefore it could be used to dissect pathways orchestrated by Aurora kinases as well as a scaffold for further inhibitor development.

  12. The microtubule plus-end tracking proteins SPR1 and EB1b interact to maintain polar cell elongation and directional organ growth in Arabidopsis.

    Science.gov (United States)

    Galva, Charitha; Kirik, Viktor; Lindeboom, Jelmer J; Kaloriti, Despoina; Rancour, David M; Hussey, Patrick J; Bednarek, Sebastian Y; Ehrhardt, David W; Sedbrook, John C

    2014-11-01

    The microtubule plus-end tracking proteins (+TIPs) END BINDING1b (EB1b) and SPIRAL1 (SPR1) are required for normal cell expansion and organ growth. EB proteins are viewed as central regulators of +TIPs and cell polarity in animals; SPR1 homologs are specific to plants. To explore if EB1b and SPR1 fundamentally function together, we combined genetic, biochemical, and cell imaging approaches in Arabidopsis thaliana. We found that eb1b-2 spr1-6 double mutant roots exhibit substantially more severe polar expansion defects than either single mutant, undergoing right-looping growth and severe axial twisting instead of waving on tilted hard-agar surfaces. Protein interaction assays revealed that EB1b and SPR1 bind each other and tubulin heterodimers, which is suggestive of a microtubule loading mechanism. EB1b and SPR1 show antagonistic association with microtubules in vitro. Surprisingly, our combined analyses revealed that SPR1 can load onto microtubules and function independently of EB1 proteins, setting SPR1 apart from most studied +TIPs in animals and fungi. Moreover, we found that the severity of defects in microtubule dynamics in spr1 eb1b mutant hypocotyl cells correlated well with the severity of growth defects. These data indicate that SPR1 and EB1b have complex interactions as they load onto microtubule plus ends and direct polar cell expansion and organ growth in response to directional cues.

  13. The Microtubule Plus-End Tracking Proteins SPR1 and EB1b Interact to Maintain Polar Cell Elongation and Directional Organ Growth in Arabidopsis[W

    Science.gov (United States)

    Galva, Charitha; Kirik, Viktor; Lindeboom, Jelmer J.; Kaloriti, Despoina; Rancour, David M.; Hussey, Patrick J.; Bednarek, Sebastian Y.; Ehrhardt, David W.; Sedbrook, John C.

    2014-01-01

    The microtubule plus-end tracking proteins (+TIPs) END BINDING1b (EB1b) and SPIRAL1 (SPR1) are required for normal cell expansion and organ growth. EB proteins are viewed as central regulators of +TIPs and cell polarity in animals; SPR1 homologs are specific to plants. To explore if EB1b and SPR1 fundamentally function together, we combined genetic, biochemical, and cell imaging approaches in Arabidopsis thaliana. We found that eb1b-2 spr1-6 double mutant roots exhibit substantially more severe polar expansion defects than either single mutant, undergoing right-looping growth and severe axial twisting instead of waving on tilted hard-agar surfaces. Protein interaction assays revealed that EB1b and SPR1 bind each other and tubulin heterodimers, which is suggestive of a microtubule loading mechanism. EB1b and SPR1 show antagonistic association with microtubules in vitro. Surprisingly, our combined analyses revealed that SPR1 can load onto microtubules and function independently of EB1 proteins, setting SPR1 apart from most studied +TIPs in animals and fungi. Moreover, we found that the severity of defects in microtubule dynamics in spr1 eb1b mutant hypocotyl cells correlated well with the severity of growth defects. These data indicate that SPR1 and EB1b have complex interactions as they load onto microtubule plus ends and direct polar cell expansion and organ growth in response to directional cues. PMID:25415978

  14. Calmodulin immunolocalization to cortical microtubules is calcium independent

    Energy Technology Data Exchange (ETDEWEB)

    Fisher, D.D.; Cyr, R.J.

    1992-01-01

    Calcium affects the stability of cortical microtubules (MTs) in lysed protoplasts. This calmodulin (CaM)-mediated interaction may provide a mechanism that serves to integrate cellular behavior with MT function. To test the hypothesis that CaM associates with these MTs, monoclonal antibodies were produced against CaM, and one (designated mAb1D10), was selected for its suitability as an immunocytochemical reagent. It is shown that CaM associates with the cortical Mats of cultured carrot (Daucus carota L.) and tobacco (Nicotiana tobacum L.) cells. Inasmuch as CaM interacts with calcium and affects the behavior of these Mats, we hypothesized that calcium would alter this association. To test this, protoplasts containing taxol-stabilized Mats were lysed in the presence of various concentrations of calcium and examined for the association of Cam with cortical Mats. At 1 [mu]M calcium, many protoplasts did not have CaM in association with the cortical Mats, while at 3.6 [mu]M calcium, this association was completely abolished. The results are discussed in terms of a model in which CaM associates with Mats via two types of interactions; one calcium dependent and one independent.

  15. Calmodulin immunolocalization to cortical microtubules is calcium independent

    Energy Technology Data Exchange (ETDEWEB)

    Fisher, D.D.; Cyr, R.J.

    1992-12-31

    Calcium affects the stability of cortical microtubules (MTs) in lysed protoplasts. This calmodulin (CaM)-mediated interaction may provide a mechanism that serves to integrate cellular behavior with MT function. To test the hypothesis that CaM associates with these MTs, monoclonal antibodies were produced against CaM, and one (designated mAb1D10), was selected for its suitability as an immunocytochemical reagent. It is shown that CaM associates with the cortical Mats of cultured carrot (Daucus carota L.) and tobacco (Nicotiana tobacum L.) cells. Inasmuch as CaM interacts with calcium and affects the behavior of these Mats, we hypothesized that calcium would alter this association. To test this, protoplasts containing taxol-stabilized Mats were lysed in the presence of various concentrations of calcium and examined for the association of Cam with cortical Mats. At 1 {mu}M calcium, many protoplasts did not have CaM in association with the cortical Mats, while at 3.6 {mu}M calcium, this association was completely abolished. The results are discussed in terms of a model in which CaM associates with Mats via two types of interactions; one calcium dependent and one independent.

  16. Toxicity and interaction of titanium dioxide nanoparticles with microtubule protein

    Institute of Scientific and Technical Information of China (English)

    Zahra Naghdi Gheshlaghi; Gholam Hossein Riazi; Shahin Ahmadian; Mahmoud Ghafari; Roya Mahinpour

    2008-01-01

    Titanium dioxide (TiO2) nanoparticles (NPs) are widely used in several manufactured products. The small size of NPs facilitates their uptake into cells as well as transcytosis across epithelial cells into blood and lymph circulation to reach different sites, such as the central nervous system. Different studies have shown the risks that TiO2 NPs in the neuronal system and other organs present. As membranebound layer aggregates or single particles, TiO2 NPs can enter not only cells, but also mitochondria and nuclei.Therefore these particles can interact with cytoplasmic proteins such as microtubules (MTs). MTs are cytoskeletal proteins that are essential in eukaryotic cells for a variety of functions, such as cellular transport, cell motility and mitosis. MTs in neurons are used to transport substances such as neurotransmitters. Single TiO2 NPs in cytoplasm can interact with these proteins and affect their crucial functions in different tissues. In this study, we showed the effects of TiO2 NPs on MT polymerization and structure using ultraviolet spectrophotometer and fluorometry. The fluorescent spectroscopy showed a significant tubulin conformational change in the presence of TiO2 NPs and the ultraviolet spectroscopy results showed that TiO2 NPs affect tubulin polymerization and decrease it. The aim of this study was to find the potential risks that TiO2 NPs pose to human organs and cells.

  17. Effects of ultraviolet radiation on microtubule organisation and morphogenesis in plants

    Energy Technology Data Exchange (ETDEWEB)

    Staxen, I.

    1994-09-01

    The involvement of the cytoskeleton in the development of somatic embryos was studied in Larix x eurolepis. Protoplasts were isolated from both somatic embryo-regenerating and non-generating cultures and fractionated on a discontinuous Percoll density gradient. Protoplasts of two cell lines of Larix eurolepis, one with regenerating potential and one lacking this potential, were compared. In contrast to the non-regenerating line were a protoplast-like organisation of the cortical microtubules was maintained, re-organisation of this microtubular network occurred in the regenerable line after only three days of culture, indicating that organised growth was occurring. However, this early organisation of cortical microtubules may not always be a valid marker for regenerable and non-regenerable material. In order to investigate the effect of ultraviolet-B (UV-B, 280-320 nm) radiation on the microtubule cytoskeleton, protoplasts were isolated from leaves of Petunia hybrida and subjected to four different doses of UV-B radiation. The organisation of the microtubules and the progression of the cells through the cell cycle was observed at 0, 24, 48 and 72 h after irradiation. UV-B induced breaks in the cortical microtubules resulting in shorter fragments with increasing amounts of radiation. Also, the division of the protoplasts was delayed. Whole Petunia plants were grown in growth chambers in the presence and absence of UV-B. The plants responded to UV-B with increased rates of CO{sub 2} assimilation, a 60% increase in UV-screening compounds and the changes in the morphology of the leaves that were reflected in a 70-100% increase in leaf area and 20% decrease in leaf thickness. The microtubules of the epidermal cells was not affected by UV-B, nor was the number of epidermal cells (per unit area). The increase in leaf area in the UV-treated plants appeared due to stimulation of cell division in the leaf meristems. 111 refs, 5 figs, 2 tabs.

  18. Microtubules in the Cerebral Cortex: Role in Memory and Consciousness

    Science.gov (United States)

    Woolf, Nancy J.

    This chapter raises the question whether synaptic connections in the cerebral cortex are adequate in accounting for higher cognition, especially cognition involving multimodal processing. A recent and novel approach to brain mechanics is outlined, one that involves microtubules and microtubule-associated protein-2 (MAP2). In addition to effects on the neuronal membrane, neurotransmitters exert actions on microtubules. These neurotransmitter effects alter the MAP2 phosphorylation state and rates of microtubule polymerization and transport. It is argued that these processes are important to the physical basis of memory and consciousness. In support of this argument, MAP2 is degraded with learning in discrete cortical modules. How this relates to synaptic change related to learning is unknown. The specific proposal is advanced that learning alters microtubules in the subsynaptic zone lying beneath the synapse, and that this forms the physical basis of long-term memory storage because microtubule networks determine the synapse strength by directing contacts with actin filaments and transport of synaptic proteins. It is argued that this is more probable than memory-related physical storage in the synapse itself. Comparisons to consciousness are made and it is concluded that there is a link between microtubules, memory and consciousness.

  19. An improved quantitative analysis method for plant cortical microtubules.

    Science.gov (United States)

    Lu, Yi; Huang, Chenyang; Wang, Jia; Shang, Peng

    2014-01-01

    The arrangement of plant cortical microtubules can reflect the physiological state of cells. However, little attention has been paid to the image quantitative analysis of plant cortical microtubules so far. In this paper, Bidimensional Empirical Mode Decomposition (BEMD) algorithm was applied in the image preprocessing of the original microtubule image. And then Intrinsic Mode Function 1 (IMF1) image obtained by decomposition was selected to do the texture analysis based on Grey-Level Cooccurrence Matrix (GLCM) algorithm. Meanwhile, in order to further verify its reliability, the proposed texture analysis method was utilized to distinguish different images of Arabidopsis microtubules. The results showed that the effect of BEMD algorithm on edge preserving accompanied with noise reduction was positive, and the geometrical characteristic of the texture was obvious. Four texture parameters extracted by GLCM perfectly reflected the different arrangements between the two images of cortical microtubules. In summary, the results indicate that this method is feasible and effective for the image quantitative analysis of plant cortical microtubules. It not only provides a new quantitative approach for the comprehensive study of the role played by microtubules in cell life activities but also supplies references for other similar studies.

  20. An Improved Quantitative Analysis Method for Plant Cortical Microtubules

    Directory of Open Access Journals (Sweden)

    Yi Lu

    2014-01-01

    Full Text Available The arrangement of plant cortical microtubules can reflect the physiological state of cells. However, little attention has been paid to the image quantitative analysis of plant cortical microtubules so far. In this paper, Bidimensional Empirical Mode Decomposition (BEMD algorithm was applied in the image preprocessing of the original microtubule image. And then Intrinsic Mode Function 1 (IMF1 image obtained by decomposition was selected to do the texture analysis based on Grey-Level Cooccurrence Matrix (GLCM algorithm. Meanwhile, in order to further verify its reliability, the proposed texture analysis method was utilized to distinguish different images of Arabidopsis microtubules. The results showed that the effect of BEMD algorithm on edge preserving accompanied with noise reduction was positive, and the geometrical characteristic of the texture was obvious. Four texture parameters extracted by GLCM perfectly reflected the different arrangements between the two images of cortical microtubules. In summary, the results indicate that this method is feasible and effective for the image quantitative analysis of plant cortical microtubules. It not only provides a new quantitative approach for the comprehensive study of the role played by microtubules in cell life activities but also supplies references for other similar studies.

  1. Oscillatory fluid flow influences primary cilia and microtubule mechanics.

    Science.gov (United States)

    Espinha, Lina C; Hoey, David A; Fernandes, Paulo R; Rodrigues, Hélder C; Jacobs, Christopher R

    2014-07-01

    Many tissues are sensitive to mechanical stimuli; however, the mechanotransduction mechanism used by cells remains unknown in many cases. The primary cilium is a solitary, immotile microtubule-based extension present on nearly every mammalian cell which extends from the basal body. The cilium is a mechanosensitive organelle and has been shown to transduce fluid flow-induced shear stress in tissues, such as the kidney and bone. The majority of microtubules assemble from the mother centriole (basal body), contributing significantly to the anchoring of the primary cilium. Several studies have attempted to quantify the number of microtubules emanating from the basal body and the results vary depending on the cell type. It has also been shown that cellular response to shear stress depends on microtubular integrity. This study hypothesizes that changing the microtubule attachment of primary cilia in response to a mechanical stimulus could change primary cilia mechanics and, possibly, mechanosensitivity. Oscillatory fluid flow was applied to two different cell types and the microtubule attachment to the ciliary base was quantified. For the first time, an increase in microtubules around primary cilia both with time and shear rate in response to oscillatory fluid flow stimulation was demonstrated. Moreover, it is presented that the primary cilium is required for this loading-induced cellular response. This study has demonstrated a new role for the cilium in regulating alterations in the cytoplasmic microtubule network in response to mechanical stimulation, and therefore provides a new insight into how cilia may regulate its mechanics and thus the cells mechanosensitivity.

  2. Roles for microtubule and microfilament cytoskeletons in animal cell cytokinesis

    Institute of Scientific and Technical Information of China (English)

    CHEN Zhongcai; CAI Shang; JIANG Qing; ZHANG Chuanmao; TANG Xiaowei

    2005-01-01

    Microtubule and microfilament cytoskeletons play key roles in the whole process of cytokinesis. Although a number of hypotheses have been proposed to elucidate the mechanism of cytokinesis by microtubule and actin filament cytoskeletons, many reports are conflicting. In our study, combining the cytoskeletons drug treatments with the time-lapse video technology, we retested the key roles of microtubule and actin filament in cytokinesis. The results showed that depolymerization of microtubules by Nocodazole after the initiation of furrowing would not inhibit the furrow ingression, but obviously decrease the stiffness of daughter cells. Depolymerizing actin filaments by Cytochalasin B before metaphase would inhibit the initiation of furrowing but not chromosome segregation, resulting in the formation of binucleate cells; however, depolymerizing actin filaments during anaphase would prevent furrowing and lead to the regress of established furrow, also resulting in the formation of binucleate cells. Further, depolymerizing microtubules and actin filaments simultaneously after metaphase would cause the quick regress of the furrow and the formation of binucleate cells. From these results we propose that a successful cytokinesis requires functions and coordination of both the microtubule and actin filament cytoskeletons. Microtubule cytoskeleton may function in the positioning and initiation of cleavage furrow, and the actin filament cytoskeleton may play key roles in the initiation and ingression of the furrow.

  3. Structural insights into microtubule doublet interactions inaxonemes

    Energy Technology Data Exchange (ETDEWEB)

    Downing, Kenneth H.; Sui, Haixin

    2007-06-06

    Coordinated sliding of microtubule doublets, driven by dynein motors, produces periodic beating of the axoneme. Recent structural studies of the axoneme have used cryo-electron tomography to reveal new details of the interactions among some of the multitude of proteins that form the axoneme and regulate its movement. Connections among the several sets of dyneins, in particular, suggest ways in which their actions may be coordinated. Study of the molecular architecture of isolated doublets has provided a structural basis for understanding the doublet's mechanical properties that are related to the bending of the axoneme, and has also offered insight into its potential role in the mechanism of dynein activity regulation.

  4. Assembly of bipolar microtubule structures by passive cross-linkers and molecular motors

    Science.gov (United States)

    Johann, D.; Goswami, D.; Kruse, K.

    2016-06-01

    During cell division, sister chromatids are segregated by the mitotic spindle, a bipolar assembly of interdigitating antiparallel polar filaments called microtubules. The spindle contains the midzone, a stable region of overlapping antiparallel microtubules, that is essential for maintaining bipolarity. Although a lot is known about the molecular players involved, the mechanism underlying midzone formation and maintenance is still poorly understood. We study the interaction of polar filaments that are cross-linked by molecular motors moving directionally and by passive cross-linkers diffusing along microtubules. Using a particle-based stochastic model, we find that the interplay of motors and passive cross-linkers can generate a stable finite overlap between a pair of antiparallel polar filaments. We develop a mean-field theory to study this mechanism in detail and investigate the influence of steric interactions between motors and passive cross-linkers on the overlap dynamics. In the presence of interspecies steric interactions, passive cross-linkers mimic the behavior of molecular motors and stable finite overlaps are generated even for non-cross-linking motors. Finally, we develop a mean-field theory for a bundle of aligned polar filaments and show that they can self-organize into a spindlelike pattern. Our work suggests possible ways as to how cells can generate spindle midzones and control their extensions.

  5. Microtubule-associated proteins and tubulin interaction by isothermal titration calorimetry.

    Science.gov (United States)

    Tsvetkov, P O; Barbier, P; Breuzard, G; Peyrot, V; Devred, F

    2013-01-01

    Microtubules play an important role in a number of vital cell processes such as cell division, intracellular transport, and cell architecture. The highly dynamic structure of microtubules is tightly regulated by a number of stabilizing and destabilizing microtubule-associated proteins (MAPs), such as tau and stathmin. Because of their importance, tubulin-MAPs interactions have been extensively studied using various methods that provide researchers with complementary but sometimes contradictory thermodynamic data. Isothermal titration calorimetry (ITC) is the only direct thermodynamic method that enables a full thermodynamic characterization (stoichiometry, enthalpy, entropy of binding, and association constant) of the interaction after a single titration experiment. This method has been recently applied to study tubulin-MAPs interactions in order to bring new insights into molecular mechanisms of tubulin regulation. In this chapter, we review the technical specificity of this method and then focus on the use of ITC in the investigation of tubulin-MAPs binding. We describe technical issues which could arise during planning and carrying out the ITC experiments, in particular with fragile proteins such as tubulin. Using examples of stathmin and tau, we demonstrate how ITC can be used to gain major insights into tubulin-MAP interaction.

  6. 3D reconstruction of cortical microtubules using multi-angle total internal reflection fluorescence microscopy

    Science.gov (United States)

    Jin, Luhong; Xiu, Peng; Zhou, Xiaoxu; Fan, Jiannan; Kuang, Cuifang; Liu, Xu; Xu, Yingke

    2017-01-01

    Total internal reflection fluorescence microscopy (TIRFM) has been widely used in biomedical research to visualize cellular processes near the cell surface. In this study, a novel multi-angle ring-illuminated TIRFM system, equipped with two galvo mirrors that are on conjugate plan of a 4f optical system was developed. Multi-angle TIRFM generates images with different penetration depths through the controlled variation of the incident angle of illuminating laser. We presented a method to perform three-dimensional (3-D) reconstruction of microtubules from multi-angle TIRFM images. The performance of our method was validated in simulated microtubules with variable signal-to-noise ratios (SNR) and the axial resolution and accuracy of reconstruction were evaluated in selecting different numbers of illumination angles or in different SNR conditions. In U373 cells, we reconstructed the 3-D localization of microtubules near the cell surface with high resolution using over a hundred different illumination angles. Theoretically, the presented TIRFM setup and 3-D reconstruction method can achieve 40 nm axial resolution in experimental conditions where SNR is as low as 2, with 35 different illumination angles. Moreover, our system and reconstruction method have the potential to be used in live cells to track membrane dynamics in 3-D.

  7. Structural microtubule cap: Stability, catastrophe, rescue, and third state

    DEFF Research Database (Denmark)

    Flyvbjerg, H.; Chretien, D.; Janosi, I.M.

    2002-01-01

    Microtubules polymerize from GTP-liganded tubulin dinners, but are essentially made of GDP-liganded tubulin. We investigate the tug-of-war resulting from the fact that GDP-liganded tubulin favors a curved configuration, but is forced to remain in a straight one when part of a microtubule. We point...... of two well-established facts: protofilaments made of GDP-liganded tubulin have intrinsic curvature, and microtubules are elastic, made from material that can yield to forces, in casu its own intrinsic forces. We explore possible properties of this structural cap, and demonstrate 1) how it allows both...

  8. Regulation of localization and activity of the microtubule depolymerase MCAK.

    Science.gov (United States)

    Tanenbaum, Marvin E; Medema, René H; Akhmanova, Anna

    2011-03-01

    Mitotic Centromere Associated Kinesin (MCAK) is a potent microtubule depolymerizing and catastrophe-inducing factor, which uses the energy of ATP hydrolysis to destabilize microtubule ends. MCAK is localized to inner centromeres, kinetochores and spindle poles of mitotic cells, and is also present in the cytoplasm. Both in interphase and in mitosis, MCAK can specifically accumulate at the growing microtubule ends. Here we discuss the mechanisms, which modulate subcellular localization and activity of MCAK through the interaction with the End Binding (EB) proteins and phosphorylation.

  9. Mitochondrial inheritance is mediated by microtubules in mammalian cell division.

    Science.gov (United States)

    Lawrence, Elizabeth; Mandato, Craig

    2013-11-01

    The mitochondrial network fragments and becomes uniformly dispersed within the cytoplasm when mammalian cells enter mitosis. Such morphology and distribution of mitochondria was previously thought to facilitate the stochastic inheritance of mitochondria by daughter cells. In contrast, we recently reported that mitochondria in dividing mammalian cells are inherited by an ordered mechanism of inheritance mediated by microtubules. We showed that mitochondria are progressively enriched at the cell equator and depleted at the poles throughout division. Furthermore, the mitochondrial distribution during division is dependent on microtubules, indicating an ordered inheritance strategy. The microtubule-mediated positioning of mitochondria in dividing mammalian cells may have functional consequences for cell division and/or mitochondrial inheritance.

  10. Fluvastatin interferes with hepatitis C virus replication via microtubule bundling and a doublecortin-like kinase-mediated mechanism.

    Directory of Open Access Journals (Sweden)

    Naushad Ali

    Full Text Available Hepatitis C virus (HCV-induced alterations in lipid metabolism and cellular protein expression contribute to viral pathogenesis. The mechanism of pleiotropic actions of cholesterol-lowering drugs, statins, against HCV and multiple cancers are not well understood. We investigated effects of fluvastatin (FLV on microtubule-associated and cancer stem cell marker (CSC, doublecortin-like kinase 1 (DCLK1 during HCV-induced hepatocarcinogenesis. HCV replication models, cancer cell lines and normal human hepatocytes were used to investigate the antiviral and antitumor effects of statins. FLV treatment resulted in induction of microtubule bundling, cell-cycle arrest and alterations in cellular DCLK1 distribution in HCV-expressing hepatoma cells. These events adversely affected the survival of liver-derived tumor cells without affecting normal human hepatocytes. FLV downregulated HCV replication in cell culture where the ATP pool and cell viability were not compromised. Pravastatin did not exhibit these effects on HCV replication, microtubules and cancer cells. The levels of miR-122 that regulates liver homeostasis and provides HCV genomic stability remained at steady state whereas DCLK1 mRNA levels were considerably reduced during FLV treatment. We further demonstrated that HCV replication was increased with DCLK1 overexpression. In conclusion, unique effects of FLV on microtubules and their binding partner DCLK1 are likely to contribute to its anti-HCV and antitumor activities in addition to its known inhibitory effects on 3-hydroxy-3-methylglutary-CoA reductase (HMGCR.

  11. Visualizing and Analyzing Branching Microtubule Nucleation Using Meiotic Xenopus Egg Extracts and TIRF Microscopy

    Science.gov (United States)

    King, Matthew; Petry, Sabine

    2016-01-01

    Mitotic and meiotic spindles consist primarily of microtubules, which originate from centrosomes and within the vicinity of chromatin. Indirect evidence suggested that microtubules also originate throughout the spindle, but the high microtubule density within the spindle precludes the direct observation of this phenomenon. By using meiotic Xenopus laevis egg extract and employing total internal reflection (TIRF) microscopy, microtubule nucleation from preexisting microtubules could be demonstrated and analyzed. Branching microtubule nucleation is an ideal mechanism to assemble and maintain a mitotic spindle, because microtubule numbers are amplified while preserving their polarity. Here, we describe the assays that made these findings possible and the experiments that helped identify the key molecular players involved. PMID:27193844

  12. Opposing microtubule motors control motility, morphology and cargo segregation during ER-to-Golgi transport

    Directory of Open Access Journals (Sweden)

    Anna K. Brown

    2014-04-01

    Full Text Available We recently demonstrated that dynein and kinesin motors drive multiple aspects of endosomal function in mammalian cells. These functions include driving motility, maintaining morphology (notably through providing longitudinal tension to support vesicle fission, and driving cargo sorting. Microtubule motors drive bidirectional motility during traffic between the endoplasmic reticulum (ER and Golgi. Here, we have examined the role of microtubule motors in transport carrier motility, morphology, and domain organization during ER-to-Golgi transport. We show that, consistent with our findings for endosomal dynamics, microtubule motor function during ER-to-Golgi transport of secretory cargo is required for motility, morphology, and cargo sorting within vesicular tubular carriers en route to the Golgi. Our data are consistent with previous findings that defined roles for dynein-1, kinesin-1 (KIF5B and kinesin-2 in this trafficking step. Our high resolution tracking data identify some intriguing aspects. Depletion of kinesin-1 reduces the number of motile structures seen, which is in line with other findings relating to the role of kinesin-1 in ER export. However, those transport carriers that were produced had a much greater run length suggesting that this motor can act as a brake on anterograde motility. Kinesin-2 depletion did not significantly reduce the number of motile transport carriers but did cause a similar increase in run length. These data suggest that kinesins act as negative regulators of ER-to-Golgi transport. Depletion of dynein not only reduced the number of motile carriers formed but also caused tubulation of carriers similar to that seen for sorting nexin-coated early endosomes. Our data indicated that the previously observed anterograde–retrograde polarity of transport carriers in transit to the Golgi from the ER is maintained by microtubule motor function.

  13. Identification of interphase functions for the NIMA kinase involving microtubules and the ESCRT pathway.

    Directory of Open Access Journals (Sweden)

    Meera Govindaraghavan

    2014-03-01

    Full Text Available The Never in Mitosis A (NIMA kinase (the founding member of the Nek family of kinases has been considered a mitotic specific kinase with nuclear restricted roles in the model fungus Aspergillus nidulans. By extending to A. nidulans the results of a synthetic lethal screen performed in Saccharomyces cerevisiae using the NIMA ortholog KIN3, we identified a conserved genetic interaction between nimA and genes encoding proteins of the Endosomal Sorting Complex Required for Transport (ESCRT pathway. Absence of ESCRT pathway functions in combination with partial NIMA function causes enhanced cell growth defects, including an inability to maintain a single polarized dominant cell tip. These genetic insights suggest NIMA potentially has interphase functions in addition to its established mitotic functions at nuclei. We therefore generated endogenously GFP-tagged NIMA (NIMA-GFP which was fully functional to follow its interphase locations using live cell spinning disc 4D confocal microscopy. During interphase some NIMA-GFP locates to the tips of rapidly growing cells and, when expressed ectopically, also locates to the tips of cytoplasmic microtubules, suggestive of non-nuclear interphase functions. In support of this, perturbation of NIMA function either by ectopic overexpression or through partial inactivation results in marked cell tip growth defects with excess NIMA-GFP promoting multiple growing cell tips. Ectopic NIMA-GFP was found to locate to the plus ends of microtubules in an EB1 dependent manner, while impairing NIMA function altered the dynamic localization of EB1 and the cytoplasmic microtubule network. Together, our genetic and cell biological analyses reveal novel non-nuclear interphase functions for NIMA involving microtubules and the ESCRT pathway for normal polarized fungal cell tip growth. These insights extend the roles of NIMA both spatially and temporally and indicate that this conserved protein kinase could help integrate cell

  14. PREFACE: Cooperative dynamics Cooperative dynamics

    Science.gov (United States)

    Gov, Nir

    2011-09-01

    , the biological problems that inspire these physical studies cover fundamental processes, from cell division up to the dynamics within axons and neurons. We would like to acknowledge and thank all contributors for their submissions, which made this special issue possible in the first place. Moreover, we would like to thank the staff at IOP Publishing for helping us with the administrative aspects and for co-ordinating the refereeing process. We hope that readers will enjoy this collection of papers and that it will trigger them to further explore the endless open physics questions presented by biological systems. Cooperative dynamics contents How does the antagonism between capping and anti-capping proteins affect actin network dynamics? Longhua Hu and Garegin A Papoian The emergence of sarcomeric, graded-polarity and spindle-like patterns in bundles of short cytoskeletal polymers and two opposite molecular motorsE M Craig, S Dey and A Mogilner Model of myosin node aggregation into a contractile ring: the effect of local alignmentNikola Ojkic, Jian-Qiu Wu and Dimitrios Vavylonis Loop formation of microtubules during gliding at high densityLynn Liu, Erkan Tüzel and Jennifer L Ross Protein-coat dynamics and cluster phases in intracellular traffickingGreg Huber, Hui Wang and Ranjan Mukhopadhyay Conformational changes, diffusion and collective behavior in monomeric kinesin-based motilityKerwyn Casey Huang, Christian Vega and Ajay Gopinathan One-dimensional deterministic transport in neurons measured by dispersion-relation phase spectroscopyRu Wang, Zhuo Wang, Joe Leigh, Nahil Sobh, Larry Millet, Martha U Gillette, Alex J Levine and Gabriel Popescu

  15. Effects of the KIF2C neck peptide on microtubules: lateral disintegration of microtubules and β-structure formation.

    Science.gov (United States)

    Shimizu, Youské; Shimizu, Takashi; Nara, Masayuki; Kikumoto, Mahito; Kojima, Hiroaki; Morii, Hisayuki

    2013-04-01

    Members of the kinesin-13 sub-family, including KIF2C, depolymerize microtubules. The positive charge-rich 'neck' region extending from the N-terminus of the catalytic head is considered to be important in the depolymerization activity. Chemically synthesized peptides, covering the basic region (A182-E200), induced a sigmoidal increase in the turbidity of a microtubule suspension. The increase was suppressed by salt addition or by reduction of basicity by amino acid substitutions. Electron microscopic observations revealed ring structures surrounding the microtubules at high peptide concentrations. Using the peptide A182-D218, we also detected free thin straight filaments, probably protofilaments disintegrated from microtubules. Therefore, the neck region, even without the catalytic head domain, may induce lateral disintegration of microtubules. With microtubules lacking anion-rich C-termini as a result of subtilisin treatment, addition of the peptide induced only a moderate increase in turbidity, and rings and protofilaments were rarely detected, while aggregations, also thought to be caused by lateral disintegration, were often observed in electron micrographs. Thus, the C-termini are not crucial for the action of the peptides in lateral disintegration but contribute to structural stabilization of the protofilaments. Previous structural studies indicated that the neck region of KIF2C is flexible, but our IR analysis suggests that the cation-rich region (K190-A204) forms β-structure in the presence of microtubules, which may be of significance with regard to the action of the neck region. Therefore, the neck region of KIF2C is sufficient to cause disintegration of microtubules into protofilaments, and this may contribute to the ability of KIF2C to cause depolymerization of microtubules.

  16. Feedback Mechanism for Microtubule Length Regulation by Stathmin Gradients

    Science.gov (United States)

    Zeitz, Maria; Kierfeld, Jan

    2014-12-01

    We formulate and analyze a theoretical model for the regulation of microtubule (MT) polymerization dynamics by the signaling proteins Rac1 and stathmin. In cells, the MT growth rate is inhibited by cytosolic stathmin, which, in turn, is inactivated by Rac1. Growing MTs activate Rac1 at the cell edge, which closes a positive feedback loop. We investigate both tubulin sequestering and catastrophe promotion as mechanisms for MT growth inhibition by stathmin. For a homogeneous stathmin concentration in the absence of Rac1, we find a switch-like regulation of the MT mean length by stathmin. For constitutively active Rac1 at the cell edge, stathmin is deactivated locally, which establishes a spatial gradient of active stathmin. In this gradient, we find a stationary bimodal MT length distributions for both mechanisms of MT growth inhibition by stathmin. One subpopulation of the bimodal length distribution can be identified with fast growing and long pioneering MTs in the region near the cell edge, which have been observed experimentally. The feedback loop is closed through Rac1 activation by MTs. For tubulin sequestering by stathmin, this establishes a bistable switch with two stable states: one stable state corresponds to upregulated MT mean length and bimodal MT length distributions, i.e., pioneering MTs; the other stable state corresponds to an interrupted feedback with short MTs. Stochastic effects as well as external perturbations can trigger switching events. For catastrophe promoting stathmin we do not find bistability.

  17. Dynamic and Geological-Ecological Spatial Planning Approach in Hot Mud Volcano Affected Area in Porong-Sidoarjo

    Directory of Open Access Journals (Sweden)

    Haryo Sulistyarso

    2010-08-01

    Full Text Available By May 29t h 2006 with an average hot mud volcano volume of 100,000 m3 /per day, disasters on well kick (i.e. Lapindo Brantas Ltd. in Banjar Panji 1 drilling well have deviated the Spatial Planning of Sidoarjo’s Regency for 2003- 2013. Regional Development Concept that is aimed at developing triangle growth pole model on SIBORIAN (SIdoarjo-JaBOn-KRIaAN could not be implemented. This planning cannot be applied due to environmental imbalance to sub district of Porong that was damaged by hot mud volcano. In order to anticipate deviations of the Regional and Spatial Planning of Sidoarjo Regency for 2003-2013, a review on regional planning and dynamic implementation as well as Spatial Planning Concept based on geologicalecological condition are required, especially the regions affected by well kick disaster. The spatial analysis is based on the geological and ecological condition by using an overlay technique using several maps of hot mud volcano affected areas. In this case, dynamic implementation is formulated to the responsiblity plan that can happen at any time because of uncertain ending of the hot mud volcano eruption disaster in Porong. The hot mud volcano affected areas in the Sidoarjo’s Spatial Planning 2009-2029 have been decided as a geologic protected zone. The result of this research is scenarios of spatial planning for the affected area (short term, medium term and long term spatial planning scenarios.

  18. Delayed system response times affect immediate physiology and the dynamics of subsequent button press behavior.

    Science.gov (United States)

    Kohrs, Christin; Hrabal, David; Angenstein, Nicole; Brechmann, André

    2014-11-01

    System response time research is an important issue in human-computer interactions. Experience with technical devices and general rules of human-human interactions determine the user's expectation, and any delay in system response time may lead to immediate physiological, emotional, and behavioral consequences. We investigated such effects on a trial-by-trial basis during a human-computer interaction by measuring changes in skin conductance (SC), heart rate (HR), and the dynamics of button press responses. We found an increase in SC and a deceleration of HR for all three delayed system response times (0.5, 1, 2 s). Moreover, the data on button press dynamics was highly informative since subjects repeated a button press with more force in response to delayed system response times. Furthermore, the button press dynamics could distinguish between correct and incorrect decisions and may thus even be used to infer the uncertainty of a user's decision.

  19. Microtubule as a Transmission Line for Ionic Currents

    Institute of Scientific and Technical Information of China (English)

    ILI(C) D.I.; SATARI(C) M.V.; RALEVI(C) N.

    2009-01-01

    We establish a new model for ionic waves along microtubules based on polyelectrolyte features of cylindrical biopolymers. The nonlinear transmission line described by a nonlinear differential equation is obtained with stable kink solution pertinent to the shape of the front of accompanying potential. The localized ionic wave could be used to explain the behavior of microtubules as biomolecular transistors capable of amplifying electrical information in neurons.

  20. Microtubule modification influences cellular response to amyloid-β exposure

    Directory of Open Access Journals (Sweden)

    Nicole Shamitko-Klingensmith

    2016-05-01

    Full Text Available During the normal aging process, cytoskeletal changes such as a reduction in density or disruption of cytoskeletal components occur that can affect neuronal function. As aging is the biggest risk factor for Alzheimer's disease (AD, this study sought to determine how microtubule (MT modification influences cellular response upon exposure to β-amyloid1-42 (Aβ1-42, which is implicated in AD. The MT networks of hypothalamic GT1-7 neurons were modified by common disrupting or stabilizing drugs, and then the physical and mechanical properties of the modified neurons were determined. The MT modified neurons were then exposed to Aβ1-42 and the ability of the neurons to cope with this exposure was determined by a variety of biochemical assays. Flow cytometry studies indicated that MT disruption reduced the binding of Aβ1-42 to the plasma membrane by 45% per cell compared to neurons with stabilized or unaltered MTs. Although the cells with disrupted MTs experienced less peptide-membrane binding, they experienced similar or increased levels of cytotoxicity caused by the Aβ1-42 exposure. In contrast, MT stabilization delayed toxicity caused by Aβ1-42. These results demonstrate that MT modification significantly influences the ability of neurons to cope with toxicity induced by Aβ1-42.

  1. Transport properties of melanosomes along microtubules interpreted by a tug-of-war model with loose mechanical coupling.

    Directory of Open Access Journals (Sweden)

    Sebastián Bouzat

    Full Text Available In this work, we explored theoretically the transport of organelles driven along microtubules by molecular motors of opposed polarities using a stochastic model that considers a Langevin dynamics for the cargo, independent cargo-motor linkers and stepping motion for the motors. It has been recently proposed that the stiffness of the motor plays an important role when multiple motors collectively transport a cargo. Therefore, we considered in our model the recently reported values for the stiffness of the cargo-motor linker determined in living cells (∼0.01 pN/nm, which is significantly lower than the motor stiffness obtained in in vitro assays and used in previous studies. Our model could reproduce the multimodal velocity distributions and typical trajectory characteristics including the properties of the reversions in the overall direction of motion observed during melanosome transport along microtubules in Xenopus laevis melanophores. Moreover, we explored the contribution of the different motility states of the cargo-motor system to the different modes of the velocity distributions and could identify the microscopic mechanisms of transport leading to trajectories compatible with those observed in living cells. Finally, by changing the attachment and detachment rates, the model could reproduce the different velocity distributions observed during melanosome transport along microtubules in Xenopus laevis melanophores stimulated for aggregation and dispersion. Our analysis suggests that active tug-of-war processes with loose mechanical coupling can account for several aspects of cargo transport along microtubules in living cells.

  2. Dynamic and Static Exercises Differentially Affect Plasma Cytokine Content in Elite Endurance- and Strength-Trained Athletes and Untrained Volunteers

    Science.gov (United States)

    Kapilevich, Leonid V.; Zakharova, Anna N.; Kabachkova, Anastasia V.; Kironenko, Tatyana A.; Orlov, Sergei N.

    2017-01-01

    Extensive exercise increases the plasma content of IL-6, IL-8, IL-15, leukemia inhibitory factor (LIF), and several other cytokines via their augmented transcription in skeletal muscle cells. However, the relative impact of aerobic and resistant training interventions on cytokine production remains poorly defined. In this study, we compared effects of dynamic and static load on cytokine plasma content in elite strength- and endurance-trained athletes vs. healthy untrained volunteers. The plasma cytokine content was measured before, immediately after, and 30 min post-exercise using enzyme-linked immunosorbent assay. Pedaling on a bicycle ergometer increased IL-6 and IL-8 content in the plasma of trained athletes by about 4- and 2-fold, respectively. In contrast to dynamic load, weightlifting had negligible impact on these parameters in strength exercise-trained athletes. Unlike IL-6 and IL-8, dynamic exercise had no impact on IL-15 and LIF, whereas static load increases the content of these cytokines by ~50%. Two-fold increment of IL-8 content seen in athletes subjected to dynamic exercise was absent in untrained individuals, whereas the ~50% increase in IL-15 triggered by static load in the plasma of weightlifting athletes was not registered in the control group. Thus, our results show the distinct impact of static and dynamic exercises on cytokine content in the plasma of trained athletes. They also demonstrate that both types of exercises differentially affect cytokine content in plasma of athletes and untrained persons.

  3. Dynein prevents erroneous kinetochore-microtubule attachments in mitosis.

    Science.gov (United States)

    Barisic, Marin; Maiato, Helder

    2015-01-01

    Equal distribution of the genetic material during cell division relies on efficient congression of chromosomes to the metaphase plate. Prior to their alignment, the Dynein motor recruited to kinetochores transports a fraction of laterally-attached chromosomes along microtubules toward the spindle poles. By doing that, Dynein not only contributes to chromosome movements, but also prevents premature stabilization of end-on kinetochore-microtubule attachments. This is achieved by 2 parallel mechanisms: 1) Dynein-mediated poleward movement of chromosomes counteracts opposite polar-ejection forces (PEFs) on chromosome arms by the microtubule plus-end-directed motors chromokinesins. Otherwise, they could stabilize erroneous syntelic kinetochore-microtubule attachments and lead to the random ejection of chromosomes away from the spindle poles; and 2) By transporting chromosomes to the spindle poles, Dynein brings the former to the zone of highest Aurora A kinase activity, further destabilizing kinetochore-microtubule attachments. Thus, Dynein plays an important role in keeping chromosome segregation error-free by preventing premature stabilization of kinetochore-microtubule attachments near the spindle poles.

  4. Cortical microtubules are responsible for gravity resistance in plants.

    Science.gov (United States)

    Hoson, Takayuki; Matsumoto, Shouhei; Soga, Kouichi; Wakabayashi, Kazuyuki

    2010-06-01

    Mechanical resistance to the gravitational force is a principal gravity response in plants distinct from gravitropism. In the final step of gravity resistance, plants increase the rigidity of their cell walls. Here we discuss the role of cortical microtubules, which sustain the function of the cell wall, in gravity resistance. Hypocotyls of Arabidopsis tubulin mutants were shorter and thicker than the wild-type, and showed either left-handed or right-handed helical growth at 1 g. The degree of twisting phenotype was intensified under hypergravity conditions. Hypergravity also induces reorientation of cortical microtubules from transverse to longitudinal directions in epidermal cells. In tubulin mutants, the percentage of cells with longitudinal microtubules was high even at 1 g, and it was further increased by hypergravity. The left-handed helical growth mutants had right-handed microtubule arrays, whereas the right-handed mutant had left-handed arrays. Moreover, blockers of mechanoreceptors suppressed both the twisting phenotype and reorientation of microtubules in tubulin mutants. These results support the hypothesis that cortical microtubules play an essential role in maintenance of normal growth phenotype against the gravitational force, and suggest that mechanoreceptors are involved in signal perception in gravity resistance. Space experiments will confirm whether this view is applicable to plant resistance to 1 g gravity, as to the resistance to hypergravity.

  5. In vitro study on the alterations of brain tubulin structure and assembly affected by magnetite nanoparticles.

    Science.gov (United States)

    Dadras, Ali; Riazi, Gholam Hossein; Afrasiabi, Ali; Naghshineh, Ali; Ghalandari, Behafarid; Mokhtari, Farzad

    2013-03-01

    In recent decades, considerable efforts have been made to understand the mechanism of memory, cognition, and relevant neurodegenerative diseases in the human brain. Several studies have shown the importance of microtubule proteins in the memory mechanism and memory dysfunction. Microtubules possess dynamicity, which is essential for functions of neuronal networks. Microtubule-associated proteins, i.e., tau, play vital roles in microtubule stability. On the other hand, the ferromagnetic mineral magnetite (Fe(3)O(4)) has been detected in the normal human brain, and elevated levels of magnetite are also observed in the brains of Alzheimer's disease patients. Therefore, we propose that a relationship between microtubule organization in axons and brain magnetite nanoparticles is possible. In this study we found alterations of microtubule polymerization in the presence of increasing concentrations of magnetite through transmission electron microscopy images and a turbidimetry method. Structural changes of microtubule and tau protein, as an essential microtubule-associated protein for tubulin assembly, were detected via circular dichroism spectroscopy, intrinsic fluorescence, and 8-anilino-1-naphthalenesulfonic acid fluorometry. We predicted three possible binding sites on tau protein and one possible binding site on tubulin dimer for magnetite nanoparticles. Magnetite also causes the morphology of PC12 cells to change abnormally and cell viability to decrease. Finally, we suggest that magnetite changes microtubule dynamics and polymerization through two paths: (1) changing the secondary and tertiary structure of tubulin and (2) binding to either tubulin dimer or tau protein and preventing tau-tubulin interaction.

  6. Forces affecting employment dynamics in Drenthe; Case study in a leading rural region in the Netherlands

    NARCIS (Netherlands)

    Terluin, I.J.; Post, J.H.; Wisselink, A.J.; Overbeek, M.M.M.

    1999-01-01

    In this report the focus is on employment dynamics in Drenthe since the beginning of the 1980s. This study is part of an EU wide research project on employment development in leading and lagging rural regions of the EU. Total employment in Drenthe increased by 24,500 jobs or with over 20% in the per

  7. Lipopolysaccharide infusion enhances dynamic cerebral autoregulation without affecting cerebral oxygen vasoreactivity in healthy volunteers

    DEFF Research Database (Denmark)

    Berg, Ronan Mg; Plovsing, Ronni R; Evans, Kevin A;

    2013-01-01

    Sepsis may be associated with disturbances in cerebral oxygen transport and cerebral haemodynamic function, thus rendering the brain particularly susceptible to hypoxia. The purpose of this study was to assess the impact of isocapnic hypoxia and hyperoxia on dynamic cerebral autoregulation...... in a human-experimental model of the systemic inflammatory response during the early stages of sepsis....

  8. Forces affecting employment dynamics in Groningen; Case study in a lagging rural region in the Netherlands

    NARCIS (Netherlands)

    Terluin, I.J.; Post, J.H.; Wisselink, A.J.; Overbeek, M.M.M.

    1999-01-01

    In this report the focus is on employment dynamics in Groningen since the beginning of the 1980s. This study is part of an EU wide research project on employment development in leading and lagging rural regions of the EU. Total employment in Groningen increased by 36,000 jobs or with one quarter in

  9. The Role of Self-Efficacy, Goal, and Affect in Dynamic Motivational Self-Regulation

    Science.gov (United States)

    Seo, Myeong-gu; Ilies, Remus

    2009-01-01

    In this paper, we examined the within-person relationship between self-efficacy and performance in an Internet-based stock investment simulation in which participants engaged in a series of stock trading activities trying to achieve performance goals in response to dynamic task environments (performance feedback and stock market movements).…

  10. Mechanisms of mindfulness: The dynamics of affective adaptation during open monitoring.

    Science.gov (United States)

    Uusberg, Helen; Uusberg, Andero; Talpsep, Teri; Paaver, Marika

    2016-07-01

    Mindfulness - the nonjudgmental awareness of the present experience - is thought to facilitate affective adaptation through increased exposure to emotions and faster extinction of habitual responses. To test this framework, the amplification of the Late Positive Potential (LPP) by negative relative to neutral images was analyzed across stimulus repetitions while 37 novices performed an open monitoring mindfulness exercise. Compared to two active control conditions where attention was either diverted to a distracting task or the stimuli were attended without mindfulness instructions, open monitoring enhanced the initial LPP response to negative stimuli, indicating increased emotional exposure. Across successive repetitions, mindfulness reduced and ultimately removed the affective LPP amplification, suggesting extinction of habitual emotional reactions. This effect arose from reduced negative as well enlarged neutral LPPs. Unlike stimuli from control conditions, the images previously viewed with mindfulness instructions did not elicit affective LPP amplification during subsequent re-exposure, suggesting reconsolidation of stimulus meaning.

  11. Live Cell Imaging Reveals Structural Associations between the Actin and Microtubule Cytoskeleton in Arabidopsis [W] [OA

    Science.gov (United States)

    Sampathkumar, Arun; Lindeboom, Jelmer J.; Debolt, Seth; Gutierrez, Ryan; Ehrhardt, David W.; Ketelaar, Tijs; Persson, Staffan

    2011-01-01

    In eukaryotic cells, the actin and microtubule (MT) cytoskeletal networks are dynamic structures that organize intracellular processes and facilitate their rapid reorganization. In plant cells, actin filaments (AFs) and MTs are essential for cell growth and morphogenesis. However, dynamic interactions between these two essential components in live cells have not been explored. Here, we use spinning-disc confocal microscopy to dissect interaction and cooperation between cortical AFs and MTs in Arabidopsis thaliana, utilizing fluorescent reporter constructs for both components. Quantitative analyses revealed altered AF dynamics associated with the positions and orientations of cortical MTs. Reorganization and reassembly of the AF array was dependent on the MTs following drug-induced depolymerization, whereby short AFs initially appeared colocalized with MTs, and displayed motility along MTs. We also observed that light-induced reorganization of MTs occurred in concert with changes in AF behavior. Our results indicate dynamic interaction between the cortical actin and MT cytoskeletons in interphase plant cells. PMID:21693695

  12. The Microtubule Plus-End Tracking Protein CLASP2 Is Required for Hematopoiesis and Hematopoietic Stem Cell Maintenance

    Directory of Open Access Journals (Sweden)

    Ksenija Drabek

    2012-10-01

    Full Text Available Mammalian CLASPs are microtubule plus-end tracking proteins whose essential function as regulators of microtubule behavior has been studied mainly in cultured cells. We show here that absence of murine CLASP2 in vivo results in thrombocytopenia, progressive anemia, and pancytopenia, due to defects in megakaryopoiesis, in erythropoiesis, and in the maintenance of hematopoietic stem cell activity. Furthermore, microtubule stability and organization are affected upon attachment of Clasp2 knockout hematopoietic stem-cell-enriched populations, and these cells do not home efficiently toward their bone marrow niche. Strikingly, CLASP2-deficient hematopoietic stem cells contain severely reduced mRNA levels of c-Mpl, which encodes the thrombopoietin receptor, an essential factor for megakaryopoiesis and hematopoietic stem cell maintenance. Our data suggest that thrombopoietin signaling is impaired in Clasp2 knockout mice. We propose that the CLASP2-mediated stabilization of microtubules is required for proper attachment, homing, and maintenance of hematopoietic stem cells and that this is necessary to sustain c-Mpl transcription.

  13. The Developmental Dynamics of Children's Academic Performance and Mothers' Homework-Related Affect and Practices

    Science.gov (United States)

    Silinskas, Gintautas; Kiuru, Noona; Aunola, Kaisa; Lerkkanen, Marja-Kristiina; Nurmi, Jari-Erik

    2015-01-01

    This study investigated the longitudinal associations between children's academic performance and their mothers' affect, practices, and perceptions of their children in homework situations. The children's (n = 2,261) performance in reading and math was tested in Grade 1 and Grade 4, and the mothers (n = 1,476) filled out questionnaires on their…

  14. Temporal Dynamics of Physical Activity and Affect in Depressed and Nondepressed Individuals

    NARCIS (Netherlands)

    Stavrakakis, Nikolaos; Booij, Sanne H.; Roest, Annelieke M.; de Jonge, Peter; Oldehinkel, Albertine J.; Bos, Elisabeth H.

    2015-01-01

    Objective: The association between physical activity and affect found in longitudinal observational studies is generally small to moderate. It is unknown how this association generalizes to individuals. The aim of the present study was to investigate interindividual differences in the bidirectional

  15. Exploring Dynamical Assessments of Affect, Behavior, and Cognition and Math State Test Achievement

    Science.gov (United States)

    San Pedro, Maria Ofelia Z.; Snow, Erica L.; Baker, Ryan S.; McNamara, Danielle S.; Heffernan, Neil T.

    2015-01-01

    There is increasing evidence that fine-grained aspects of student performance and interaction within educational software are predictive of long-term learning. Machine learning models have been used to provide assessments of affect, behavior, and cognition based on analyses of system log data, estimating the probability of a student's particular…

  16. A shift from exploitation to interference competition with increasing density affects population and community dynamics

    OpenAIRE

    Holdridge, Erica M.; Cuellar‐Gempeler, Catalina; terHorst, Casey P.

    2016-01-01

    Abstract Intraspecific competition influences population and community dynamics and occurs via two mechanisms. Exploitative competition is an indirect effect that occurs through use of a shared resource and depends on resource availability. Interference competition occurs by obstructing access to a resource and may not depend on resource availability. Our study tested whether the strength of interference competition changes with protozoa population density. We grew experimental microcosms of ...

  17. Wolbachia-Host Interactions: Host Mating Patterns Affect Wolbachia Density Dynamics

    OpenAIRE

    Dong-Xiao Zhao; Xiang-Fei Zhang; Da-Song Chen; Yan-Kai Zhang; Xiao-Yue Hong

    2013-01-01

    Wolbachia are maternally inherited intracellular bacteria that infect a wide range of arthropods and cause an array of effects on host reproduction, fitness and mating behavior. Although our understanding of the Wolbachia-associated effects on hosts is rapidly expanding, our knowledge of the host factors that mediate Wolbachia dynamics is rudimentary. Here, we explore the interactions between Wolbachia and its host, the two-spotted spider mite Tetranychus urticae Koch. Our results indicate th...

  18. The Katanin Microtubule Severing Protein in Plants

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    Katanin is a heterodimeric microtubule (MT) severing protein that uses energy from ATP hydrolysis to generate internal breaks along MTs. Katanin p60, one of the two subunits, possesses ATPase and MT-bindinglsevering activities, and the p80 subunit is responsible for targeting of katanin to certain subcellular locations. In animals, katanin plays an important role in the release of MTs from their nucleation sites in the centrosome. It is also involved in severing MTs into smaller fragments which can serve as templates for further polymerization to increase MT number during meiotic and mitotic spindle assembly. Katanin homologs are present in a wide variety of plant species. The Arabidopsis katanin homolog has been shown to possess ATP-dependent MT severing activity in vitro and exhibit a punctate localization pattern at the cell cortex and the perinuclear region. Disruption of katanin functions by genetic mutations causes a delay in the disappearance of the perinuclear MT array and results in an aberrant organization of cortical MTs in elongating cells. Consequently, katanin mutations lead to defects in cell elongation, cellulose microfibril deposition, and hormonal responses. Studies of katanin in plants provide new insights into our understanding of its roles in cellular functions.

  19. Microtubules, polarity and vertebrate neural tube morphogenesis.

    Science.gov (United States)

    Cearns, Michael D; Escuin, Sarah; Alexandre, Paula; Greene, Nicholas D E; Copp, Andrew J

    2016-07-01

    Microtubules (MTs) are key cellular components, long known to participate in morphogenetic events that shape the developing embryo. However, the links between the cellular functions of MTs, their effects on cell shape and polarity, and their role in large-scale morphogenesis remain poorly understood. Here, these relationships were examined with respect to two strategies for generating the vertebrate neural tube: bending and closure of the mammalian neural plate; and cavitation of the teleost neural rod. The latter process has been compared with 'secondary' neurulation that generates the caudal spinal cord in mammals. MTs align along the apico-basal axis of the mammalian neuroepithelium early in neural tube closure, participating functionally in interkinetic nuclear migration, which indirectly impacts on cell shape. Whether MTs play other functional roles in mammalian neurulation remains unclear. In the zebrafish, MTs are important for defining the neural rod midline prior to its cavitation, both by localizing apical proteins at the tissue midline and by orienting cell division through a mirror-symmetric MT apparatus that helps to further define the medial localization of apical polarity proteins. Par proteins have been implicated in centrosome positioning in neuroepithelia as well as in the control of polarized morphogenetic movements in the neural rod. Understanding of MT functions during early nervous system development has so far been limited, partly by techniques that fail to distinguish 'cause' from 'effect'. Future developments will likely rely on novel ways to selectively impair MT function in order to investigate the roles they play.

  20. Tau neurofibrillary pathology and microtubule stability.

    Science.gov (United States)

    Michaelis, Mary L; Dobrowsky, Rick T; Li, Guibin

    2002-12-01

    We previously reported that nonomolar concentrations of Taxol and several structurally diverse microtubule (MT)-stabilizing agents significantly enhanced the survival of neurons in the presence of fibrils of amyloid beta peptide (Abeta). Pretreatment of neurons with MT-stabilizing drugs also blocked Abeta-induced activation of tau hyperphosphorylation. Although tau is a substrate for several kinases, we initially focused on cdk5, as this tau kinase has been shown to be activated in Abeta-treated neurons and Alzheimer's disease (AD) brain. In an in vitro kinase assay, Taxol inhibited activation of cdk5 by Abeta. In addition, the proposed cellular cascade in which calpain activation leads to cleavage of the cdk5 regulator, p35, to the strong kinase activator p25 was also prevented. Taxol did not directly inhibit the activity of either cdk5 or calpain, indicating that other cellular components are required for the effect of the drug on Abeta activation of tau phosphorylation. Our results suggest that drugs that interact with MTs can alter signaling events in neurons, possibly because some MTs play a role in organizing protein complexes involved in responses to Abeta. Thus the cytoskeletal network may serve as a biosensor of cellular well-being.

  1. A stage-structured, spatially explicit migration model for Myotis bats: mortality location affects system dynamics

    Science.gov (United States)

    Erickson, Richard A.; Thogmartin, Wayne E.; Russell, Robin E.; Diffendorfer, James E.; Jennifer A. Szymanski,; ,

    2014-01-01

    Bats are ecologically and economically important species because they consume insects, transport nutrients, and pollinate flowers.  Many species of bats, including those in the Myotis genus, are facing population decline and increased extinction risk.  Despite these conservation concerns, few models exist for providing insight into the population dynamics of bats in a spatially explicit context.  We developed a model for bats by considering the stage-structured colonial life history of Myotis bats with their annual migration behavior.  This model provided insight into network dynamics.  We specifically focused on two Myotis species living in the eastern United States: the Indiana bat (M. sodalis), which is a Federally listed endangered species, and the little brown bat (M. lucifugus), which is under consideration for listing as an endangered species.  We found that multiple equilibria exist for the local, migratory subpopulations even though the total population was constant.  These equilibria suggest the location and magnitude of stressors such as White-nose Syndrome, meteorological phenomena, or impacts of wind turbines on survival influence system dynamics and risk of population extirpation in difficult to predict ways.

  2. Nucleotide binding affects intrinsic dynamics and structural communication in Ras GTPases.

    Science.gov (United States)

    Fanelli, Francesca; Raimondi, Francesco

    2013-01-01

    The Ras superfamily comprises many guanine nucleotide-binding proteins (G proteins) that are essential to intracellular signal transduction. These proteins act biologically as molecular switches, which, cycling between OFF and ON states, play fundamental role in cell biology. This review article summarizes the inferences from the widest computational analyses done so far on Ras GTPases aimed at providing a comprehensive structural/dynamic view of the trans-family and family-specific functioning mechanisms. These variegated comparative analyses could infer the evolutionary and intrinsic flexibilities as well as the structural communication features in the most representative G protein families in different functional states. In spite of the low sequence similarities, the members of the Ras superfamily share the topology of the Ras-like domain, including the nucleotide binding site. GDP and GTP make very similar interactions in all GTPases and differences in their binding modes are localized around the γ-phosphate of GTP. Remarkably, such subtle local differences result in significant differences in the functional dynamics and structural communication features of the protein. In Ras GTPases, the nucleotide plays a central and active role in dictating functional dynamics, establishing the major structure network, and mediating the communication paths instrumental in function retention and specialization. Collectively, the results of these studies support the speculation that an "extended conformational selection model" that embraces a repertoire of selection and adjustment processes is likely more suitable to describe the nucleotide behavior in these important molecular switches.

  3. Nonequilibrium water dynamics in the rhizosphere: How mucilage affects water flow in soils

    Science.gov (United States)

    Kroener, Eva; Zarebanadkouki, Mohsen; Kaestner, Anders; Carminati, Andrea

    2014-08-01

    The flow of water from soil to plant roots is controlled by the properties of the narrow region of soil close to the roots, the rhizosphere. In particular, the hydraulic properties of the rhizosphere are altered by mucilage, a polymeric gel exuded by the roots. In this paper we present experimental results and a conceptual model of water flow in unsaturated soils mixed with mucilage. A central hypothesis of the model is that the different drying/wetting rate of mucilage compared to the bulk soil results in nonequilibrium relations between water content and water potential in the rhizosphere. We coupled this nonequilibrium relation with the Richards equation and obtained a constitutive equation for water flow in soil and mucilage. To test the model assumptions, we measured the water retention curve and the saturated hydraulic conductivity of sandy soil mixed with mucilage from chia seeds. Additionally, we used neutron radiography to image water content in a layer of soil mixed with mucilage during drying and wetting cycles. The radiographs demonstrated the occurrence of nonequilibrium water dynamics in the soil-mucilage mixture. The experiments were simulated by numerically solving the nonequilibrium model. Our study provides conceptual and experimental evidences that mucilage has a strong impact on soil water dynamics. During drying, mucilage maintains a greater soil water content for an extended time, while during irrigation it delays the soil rewetting. We postulate that mucilage exudation by roots attenuates plant water stress by modulating water content dynamics in the rhizosphere.

  4. Experimental protocols for and studies of the effects of surface passivation and water isotopes on the gliding speed of microtubules propelled by kinesin-1

    Science.gov (United States)

    Maloney, Roger Andrew

    This dissertation explores how the kinesin-1 and microtubule system is affected by surface passivation and water isotopes. Surface passivation was found to affect the gliding speed that microtubules exhibit in the gliding motility assay and the lengths of microtubules supported by the passivation. It was also found that gliding speeds of microtubules are very sensitive to temperature changes. Studies changing the water isotope were a first attempt to investigate if changing the solvent changed the osmotic pressure of the solution kinesin and microtubules were in. No osmotic pressure changes were observed, however, the experiments using different isotopes of water did illuminate the possibility that kinesin may be sensitive to viscosity changes in the solvent. This experiment also suggests further experiments that can be specifically designed to probe osmotic pressure changes. This thesis was also the first thesis ever, to the best of the author's knowledge, to be done in a completely open format. All information and notebook entries that are related to it, as well as the thesis itself, can be found on the website OpenWetWare. The thesis can also be found there including all the different versions that went into its editing. The philosophy and process of making data open and accessible to every one is also discussed.

  5. On the significance of microtubule flexural behavior in cytoskeletal mechanics.

    Directory of Open Access Journals (Sweden)

    Mehrdad Mehrbod

    Full Text Available Quantitative description of cell mechanics has challenged biological scientists for the past two decades. Various structural models have been attempted to analyze the structure of the cytoskeleton. One important aspect that has been largely ignored in all these modeling approaches is related to the flexural and buckling behavior of microtubular filaments. The objective of this paper is to explore the influence of this flexural and buckling behavior in cytoskeletal mechanics.In vitro the microtubules are observed to buckle in the first mode, reminiscent of a free, simply-supported beam. In vivo images of microtubules, however, indicate that the buckling mostly occurs in higher modes. This buckling mode switch takes place mostly because of the lateral support of microtubules via their connections to actin and intermediate filaments. These lateral loads are exerted throughout the microtubule length and yield a considerable bending behavior that, unless properly accounted for, would produce erroneous results in the modeling and analysis of the cytoskeletal mechanics.One of the promising attempts towards mechanical modeling of the cytoskeleton is the tensegrity model, which simplifies the complex network of cytoskeletal filaments into a combination merely of tension-bearing actin filaments and compression-bearing microtubules. Interestingly, this discrete model can qualitatively explain many experimental observations in cell mechanics. However, evidence suggests that the simplicity of this model may undermine the accuracy of its predictions, given the model's underlying assumption that "every single member bears solely either tensile or compressive behavior," i.e. neglecting the flexural behavior of the microtubule filaments. We invoke an anisotropic continuum model for microtubules and compare the bending energy stored in a single microtubule with its axial strain energy at the verge of buckling. Our results suggest that the bending energy can

  6. Gene organization, evolution and expression of the microtubule-associated protein ASAP (MAP9

    Directory of Open Access Journals (Sweden)

    Giorgi Dominique

    2008-09-01

    tissues rich in microtubules. ASAP associates to the mitotic spindle and cytoplasmic microtubules, and represents a key factor of mitosis with possible involvement in other cell cycle processes. It may have a role in spermatogenesis and also represents a potential new target for antitumoral drugs. Possible involvement in neuron dynamics also highlights ASAP as a candidate target in neurodegenerative diseases.

  7. Association between microtubules and Golgi vesicles isolated from rat parotid glands.

    Science.gov (United States)

    Coffe, G; Raymond, M N

    1990-01-01

    We report an isolation procedure of trans-Golgi vesicles (GVs) from rat parotid glands. Various organelle markers were used, particularly galactosyl transferase as a trans-Golgi marker, to test the purity of the GV fraction. A quantitative in vitro binding assay between microtubules and GVs is described. The vesicles were incubated with taxol-induced microtubules, layered between 50% and 43% sucrose cushions and subjected to centrifugation. Unlike free microtubules which were sedimented, the GV-bound microtubules co-migrated upward with GVs. Quantification of these bound microtubules was carried out by densitometric scanning of Coomassie blue-stained gels. The association between microtubules and GVs followed a saturation curve, with a plateau value of 20 micrograms of microtubule protein bound to 500 micrograms of GV fraction. The half-saturation of the GV sites was obtained with a microtubule concentration of 20 micrograms/ml. Electron microscopy of negatively stained re-floated material showed numerous microtubule-vesicle complexes. Coating of microtubules with an excess of brain microtubule-associated proteins (MAPs) abolished binding. In the absence of exogenous microtubules, we showed that the GV fraction was already interacting with a class of endogenous rat parotid microtubules. This class of colcemid and cold-stable microtubules represents 10-20% of the total tubulin content of the parotid cell.

  8. LKB1 destabilizes microtubules in myoblasts and contributes to myoblast differentiation.

    Directory of Open Access Journals (Sweden)

    Isma Mian

    Full Text Available BACKGROUND: Skeletal muscle myoblast differentiation and fusion into multinucleate myotubes is associated with dramatic cytoskeletal changes. We find that microtubules in differentiated myotubes are highly stabilized, but premature microtubule stabilization blocks differentiation. Factors responsible for microtubule destabilization in myoblasts have not been identified. FINDINGS: We find that a transient decrease in microtubule stabilization early during myoblast differentiation precedes the ultimate microtubule stabilization seen in differentiated myotubes. We report a role for the serine-threonine kinase LKB1 in both microtubule destabilization and myoblast differentiation. LKB1 overexpression reduced microtubule elongation in a Nocodazole washout assay, and LKB1 RNAi increased it, showing LKB1 destabilizes microtubule assembly in myoblasts. LKB1 levels and activity increased during myoblast differentiation, along with activation of the known LKB1 substrates AMP-activated protein kinase (AMPK and microtubule affinity regulating kinases (MARKs. LKB1 overexpression accelerated differentiation, whereas RNAi impaired it. CONCLUSIONS: Reduced microtubule stability precedes myoblast differentiation and the associated ultimate microtubule stabilization seen in myotubes. LKB1 plays a positive role in microtubule destabilization in myoblasts and in myoblast differentiation. This work suggests a model by which LKB1-induced microtubule destabilization facilitates the cytoskeletal changes required for differentiation. Transient destabilization of microtubules might be a useful strategy for enhancing and/or synchronizing myoblast differentiation.

  9. The impact of chief executive officer personality on top management team dynamics:one mechanism by which leadership affects organizational performance.

    Science.gov (United States)

    Peterson, Randall S; Smith, D Brent; Martorana, Paul V; Owens, Pamela D

    2003-10-01

    This article explores 1 mechanism by which leader personality affects organizational performance. The authors hypothesized and tested the effects of leader personality on the group dynamics of the top management team (TMT) and of TMT dynamics on organizational performance. To test their hypotheses, the authors used the group dynamics q-sort method, which is designed to permit rigorous, quantitative comparisons of data derived from qualitative sources. Results from independent observations of chief executive officer (CEO) personality and TMT dynamics for 17 CEOs supported the authors' hypothesized relationships both between CEO personality and TMT group dynamics and between TMT dynamics and organizational performance.

  10. Direct Cytoplasmic Delivery and Nuclear Targeting Delivery of HPMA-MT Conjugates in a Microtubules Dependent Fashion.

    Science.gov (United States)

    Zhong, Jiaju; Zhu, Xi; Luo, Kui; Li, Lian; Tang, Manlin; Liu, Yanxi; Zhou, Zhou; Huang, Yuan

    2016-09-06

    As the hearts of tumor cells, the nucleus is the ultimate target of many chemotherapeutic agents and genes. However, nuclear drug delivery is always hampered by multiple intracellular obstacles, such as low efficiency of lysosome escape and insufficient nuclear trafficking. Herein, an N-(2-hydroxypropyl) methacrylamide (HPMA) polymer-based drug delivery system was designed, which could achieve direct cytoplasmic delivery by a nonendocytic pathway and transport into the nucleus in a microtubules dependent fashion. A special targeting peptide (MT), derived from an endogenic parathyroid hormone-related protein, was conjugated to the polymer backbone, which could accumulate into the nucleus a by microtubule-mediated pathway. The in vitro studies found that low temperature and NaN3 could not influence the cell internalization of the conjugates. Besides, no obvious overlay of the conjugates with lysosome demonstrated that the polymer conjugates could enter the tumor cell cytoplasm by a nonendocytic pathway, thus avoiding the drug degradation in the lysosome. Furthermore, after suppression of the microtubule dynamics with microtubule stabilizing docetaxel (DTX) and destabilizing nocodazole (Noc), the nuclear accumulation of polymeric conjugates was significantly inhibited. Living cells fluorescence recovery after photobleaching study found that the nuclear import rate of conjugates was 2-fold faster compared with the DTX and Noc treated groups. These results demonstrated that the conjugates transported into the nucleus in a microtubules dependent way. Therefore, in addition to direct cytoplasmic delivery, our peptide conjugated polymeric platform could simultaneously mediate nuclear drug accumulation, which may open a new path for further intracellular genes/peptides delivery.

  11. Oleuropein-Enriched Olive Leaf Extract Affects Calcium Dynamics and Impairs Viability of Malignant Mesothelioma Cells

    Directory of Open Access Journals (Sweden)

    Carla Marchetti

    2015-01-01

    Full Text Available Malignant mesothelioma is a poor prognosis cancer in urgent need of alternative therapies. Oleuropein, the major phenolic of olive tree (Olea europaea L., is believed to have therapeutic potentials for various diseases, including tumors. We obtained an oleuropein-enriched fraction, consisting of 60% w/w oleuropein, from olive leaves, and assessed its effects on intracellular Ca2+ and cell viability in mesothelioma cells. Effects of the oleuropein-enriched fraction on Ca2+ dynamics and cell viability were studied in the REN mesothelioma cell line, using fura-2 microspectrofluorimetry and MTT assay, respectively. Fura-2-loaded cells, transiently exposed to the oleuropein-enriched fraction, showed dose-dependent transient elevations of cytosolic Ca2+ concentration (Ca2+i. Application of standard oleuropein and hydroxytyrosol, and of the inhibitor of low-voltage T-type Ca2+ channels NNC-55-0396, suggested that the effect is mainly due to oleuropein acting through its hydroxytyrosol moiety on T-type Ca2+ channels. The oleuropein-enriched fraction and standard oleuropein displayed a significant antiproliferative effect, as measured on REN cells by MTT cell viability assay, with IC50 of 22 μg/mL oleuropein. Data suggest that our oleuropein-enriched fraction from olive leaf extract could have pharmacological application in malignant mesothelioma anticancer therapy, possibly by targeting T-type Ca2+ channels and thereby dysregulating intracellular Ca2+ dynamics.

  12. Screening of mutations affecting protein stability and dynamics of FGFR1—A simulation analysis

    Directory of Open Access Journals (Sweden)

    C. George Priya Doss

    2012-12-01

    Full Text Available Single amino acid substitutions in Fibroblast Growth Factor Receptor 1 (FGFR1 destabilize protein and have been implicated in several genetic disorders like various forms of cancer, Kallamann syndrome, Pfeiffer syndrome, Jackson Weiss syndrome, etc. In order to gain functional insight into mutation caused by amino acid substitution to protein function and expression, special emphasis was laid on molecular dynamics simulation techniques in combination with in silico tools such as SIFT, PolyPhen 2.0, I-Mutant 3.0 and SNAP. It has been estimated that 68% nsSNPs were predicted to be deleterious by I-Mutant, slightly higher than SIFT (37%, PolyPhen 2.0 (61% and SNAP (58%. From the observed results, P722S mutation was found to be most deleterious by comparing results of all in silico tools. By molecular dynamics approach, we have shown that P722S mutation leads to increase in flexibility, and deviated more from the native structure which was supported by the decrease in the number of hydrogen bonds. In addition, biophysical analysis revealed a clear insight of stability loss due to P722S mutation in FGFR1 protein. Majority of mutations predicted by these in silico tools were in good concordance with the experimental results.

  13. STARDUST-U experiments on fluid-dynamic conditions affecting dust mobilization during LOVAs

    Science.gov (United States)

    Poggi, L. A.; Malizia, A.; Ciparisse, J. F.; Tieri, F.; Gelfusa, M.; Murari, A.; Del Papa, C.; Giovannangeli, I.; Gaudio, P.

    2016-07-01

    Since 2006 the Quantum Electronics and Plasma Physics (QEP) Research Group together with ENEA FusTech of Frascati have been working on dust re-suspension inside tokamaks and its potential capability to jeopardize the integrity of future fusion nuclear plants (i.e. ITER or DEMO) and to be a risk for the health of the operators. Actually, this team is working with the improved version of the "STARDUST" facility, i.e. "STARDUST-Upgrade". STARDUST-U facility has four new air inlet ports that allow the experimental replication of Loss of Vacuum Accidents (LOVAs). The experimental campaign to detect the different pressurization rates, local air velocity, temperature, have been carried out from all the ports in different accident conditions and the principal results will be analyzed and compared with the numerical simulations obtained through a CFD (Computational Fluid Dynamic) code. This preliminary thermo fluid-dynamic analysis of the accident is crucial for numerical model development and validation, and for the incoming experimental campaign of dust resuspension inside STARDUST-U due to well-defined accidents presented in this paper.

  14. Inactivation of Tor proteins affects the dynamics of endocytic proteins in early stage of endocytosis

    Indian Academy of Sciences (India)

    Brandon Tenay; Evin Kimberlin; Michelle Williams; Juliette Denise; Joshua Fakilahyel; Kyoungtae Kim

    2013-06-01

    Tor2 is an activator of the Rom2/Rho1 pathway that regulates -factor internalization. Since the recruitment of endocytic proteins such as actin-binding proteins and the amphiphysins precedes the internalization of -factor, we hypothesized that loss of Tor function leads to an alteration in the dynamics of the endocytic proteins. We report here that endocytic proteins, Abp1 and Rvs167, are less recruited to endocytic sites not only in tor2 but also tor1 mutants. Furthermore, we found that the endocytic proteins Rvs167 and Sjl2 are completely mistargeted to the cytoplasm in tor1tor2ts double mutant cells. We also demonstrate here that the efficiency of endocytic internalization or scission in all tor mutants was drastically decreased. In agreement with the Sjl2 mislocalization, we found that in tor1tor2ts double mutant cells, as well as other tor mutant cells, the overall PIP2 level was dramatically increased. Finally, the cell wall chitin content in tor2ts and tor1tor2ts mutant cells was also significantly increased. Taken together, both functional Tor proteins, Tor1 and Tor2, are essentially required for proper endocytic protein dynamics at the early stage of endocytosis.

  15. Microtubule dynamics. II. Kinetics of self-assembly

    DEFF Research Database (Denmark)

    Flyvbjerg, H.; Jobs, E.

    1997-01-01

    Inverse scattering theory describes the conditions necessary and sufficient to determine an unknown potential from known scattering data. No similar theory exists for when and how one may deduce the kinetics of an unknown chemical reaction from quantitative information about its final state and its...... various initial concentrations of monomers. Distinct mathematical properties of the kinetics of the unknown reaction pathway are found. These properties are shown to restrict the kinetics to a single model that generalizes Oosawa's classical nucleation-polymerization model. The methods used here...

  16. Drainage and leaching dynamics in a cropped hummocky soil landscape with erosion-affected pedogenesis

    Science.gov (United States)

    Gerke, Horst H.; Rieckh, Helene; Sommer, Michael

    2016-04-01

    Hummocky soil landscapes are characterized by 3D spatial patterns of soil types that result from erosion-affected pedogenesis. Due to tillage and water erosion, truncated profiles have been formed at steep and mid slopes and colluvial soils at hollows. Pedogenetic variations in soil horizons at the different hillslope positions suggested feedback effects between erosion affected soil properties, the water balances, and the crop growth and leaching rates. Water balance simulations compared uniform with hillslope position-specific crop and root growths for soils at plateau, flat mid slope, steep slope, and hollow using the Hydrus-1D program. The boundary condition data were monitored at the CarboZALF-D experimental field site, which was cropped with perennial lucerne (Medicago sativa L.) in 2013 and 2014. Crop and root growth was assumed proportional to observed leaf area index (LAI). Fluxes of dissolved organic and inorganic carbon (DOC, DIC) were obtained from simulated water fluxes and measured DOC and DIC concentrations. For the colluvic soil, the predominately upward flow led to a net input in DIC and DOC. For the truncated soils at steep slopes, a reduced crop growth caused an relative increase in drainage, suggesting an accelerated leaching, which in the long term could accelerate the soil development and more soil variations along eroding hillslopes in arable soil landscapes.

  17. Improved Dynamic Modeling of the Cascade Distillation Subsystem and Analysis of Factors Affecting Its Performance

    Science.gov (United States)

    Perry, Bruce A.; Anderson, Molly S.

    2015-01-01

    The Cascade Distillation Subsystem (CDS) is a rotary multistage distiller being developed to serve as the primary processor for wastewater recovery during long-duration space missions. The CDS could be integrated with a system similar to the International Space Station Water Processor Assembly to form a complete water recovery system for future missions. A preliminary chemical process simulation was previously developed using Aspen Custom Modeler® (ACM), but it could not simulate thermal startup and lacked detailed analysis of several key internal processes, including heat transfer between stages. This paper describes modifications to the ACM simulation of the CDS that improve its capabilities and the accuracy of its predictions. Notably, the modified version can be used to model thermal startup and predicts the total energy consumption of the CDS. The simulation has been validated for both NaC1 solution and pretreated urine feeds and no longer requires retuning when operating parameters change. The simulation was also used to predict how internal processes and operating conditions of the CDS affect its performance. In particular, it is shown that the coefficient of performance of the thermoelectric heat pump used to provide heating and cooling for the CDS is the largest factor in determining CDS efficiency. Intrastage heat transfer affects CDS performance indirectly through effects on the coefficient of performance.

  18. Environmental Factors Affecting Temporal and Spatial Dynamics of Soil Erosion in Xingguo County, South China

    Institute of Scientific and Technical Information of China (English)

    WANG Ku; SHI Xue-Zheng; YU Dong-Sheng; SHI De-Ming; CHEN Jing-Ming; XU Bin-Bin; LIANG Yin; LI De-Cheng

    2005-01-01

    By using soil erosion maps of four different time periods and a digital elevation model (DEM), in combination with the remote sensing and GIS technologies, soil erosion dynamics in Xingguo County of Jiangxi Province in South China were analyzed on both temporal and spatial scales in soils of different parent materials, altitudes and slopes. The results showed that from 1958 to 2000 severe soil erosion was coming under control with a decreasing percentage of the land under severe erosion. It was also found that the soils developed from Quaternary red clay, granite and purple shale were more susceptible to soil erosion and that areas sitting between 200 to 500 m in altitude with a slope less than 3° or between 7° to 20° where human activities were frequent remained to be zones where soil erosion was most likely to occur. These areas deserve special attention in monitoring and controlling.

  19. Large dynamic range operation of ultra-higher number MWFLs affected by MZI-SI

    Science.gov (United States)

    Narimah Aziz, Siti; Arsad, Norhana; Ashrif Abu Bakar, Ahmad; Sushita Menon, P.; Shaari, Sahbudin

    2016-11-01

    This report presents a large dynamic operation of wavelength numbers of laser lines that have been periodically filtered using an MZI-SI filter effect. A 70 nm span range for a wider periodic comb filter with 0.60 nm wavelength spacing was achieved through an advanced triple-loop ring-cavity fiber laser with a combination of MZI-SI. Almost all of the best 95 numbers of wavelengths are flattened at 6 dB of peak power fluctuation in a 52 nm range. By adjusting the rotation angles of a polarization controller (PC), the ultra-wide range multiwavelength spectrum has been shifted by 36 nm in a range from 1522.8-1558.6 nm.

  20. Melatonin affects the order, dynamics and hydration of brain membrane lipids

    Science.gov (United States)

    Akkas, Sara B.; Inci, Servet; Zorlu, Faruk; Severcan, Feride

    2007-05-01

    The brain is especially susceptible to free radical attack since it is rich in polyunsaturated fatty acids and consumes very high amounts of oxygen. Melatonin is a non-enzymatic amphiphilic antioxidant hormone that is widely used in medicine for protective and treatment purposes in cases of oxidative stress. In the present work, the effects of the clinically used dose of melatonin (a single intraperitoneal dose of 100 mg/kg) on rat brain homogenate were investigated as a function of temperature using Fourier transform infrared spectroscopy. The results showed that the lipid to protein ratio decreases in the melatonin treated brain samples. Moreover, it is revealed that melatonin disorders and decreases the dynamics of lipids and induces a strengthening in the hydrogen bonding between the functional groups of both melatonin and the polar parts of lipids and/or water at physiological temperatures.

  1. The interplay between language, gesture, and affect during communicative transition: a dynamic systems approach.

    Science.gov (United States)

    Parladé, Meaghan V; Iverson, Jana M

    2011-05-01

    From a dynamic systems perspective, transition points in development are times of increased instability, during which behavioral patterns are susceptible to temporary decoupling. This study investigated the impact of the vocabulary spurt on existing patterns of communicative coordination. Eighteen typically developing infants were videotaped at home 1 month before, at, and after the vocabulary spurt. Infants were identified as spurters if they underwent a discrete phase transition in vocabulary development (marked by an inflection point), and compared with a group of nonspurters whose word-learning rates followed a trajectory of continuous change. Relative to surrounding sessions, there were significant reductions in overall coordination of communicative behaviors and in words produced in coordination at the vocabulary spurt session for infants who experienced more dramatic vocabulary growth. In contrast, nonspurters demonstrated little change across sessions. Findings underscore the importance of transitions as opportunities for observing processes of developmental change.

  2. Small doses, big troubles: modeling growth dynamics of organisms affecting microalgal production cultures in closed photobioreactors.

    Science.gov (United States)

    Forehead, Hugh I; O'Kelly, Charles J

    2013-02-01

    The destruction of mass cultures of microalgae by biological contamination of culture medium is a pervasive and expensive problem, in industry and research. A mathematical model has been formulated that attempts to explain contaminant growth dynamics in closed photobioreactors (PBRs). The model simulates an initial growth phase without PBR dilution, followed by a production phase in which culture is intermittently removed. Contaminants can be introduced at any of these stages. The model shows how exponential growth from low initial inocula can lead to "explosive" growth in the population of contaminants, appearing days to weeks after inoculation. Principal influences are contaminant growth rate, PBR dilution rate, and the size of initial contaminant inoculum. Predictions corresponded closely with observed behavior of two contaminants, Uronema sp. and Neoparamoeba sp., found in operating PBRs. A simple, cheap and effective protocol was developed for short-term prediction of contamination in PBRs, using microscopy and archived samples.

  3. Assessing when chromosomal rearrangements affect the dynamics of speciation: implications from computer simulations.

    Science.gov (United States)

    Feder, Jeffrey L; Nosil, Patrik; Flaxman, Samuel M

    2014-01-01

    Many hypotheses have been put forth to explain the origin and spread of inversions, and their significance for speciation. Several recent genic models have proposed that inversions promote speciation with gene flow due to the adaptive significance of the genes contained within them and because of the effects inversions have on suppressing recombination. However, the consequences of inversions for the dynamics of genome wide divergence across the speciation continuum remain unclear, an issue we examine here. We review a framework for the genomics of speciation involving the congealing of the genome into alternate adaptive states representing species ("genome wide congealing"). We then place inversions in this context as examples of how genetic hitchhiking can potentially hasten genome wide congealing. Specifically, we use simulation models to (i) examine the conditions under which inversions may speed genome congealing and (ii) quantify predicted magnitudes of these effects. Effects of inversions on promoting speciation were most common and pronounced when inversions were initially fixed between populations before secondary contact and adaptation involved many genes with small fitness effects. Further work is required on the role of underdominance and epistasis between a few loci of major effect within inversions. The results highlight five important aspects of the roles of inversions in speciation: (i) the geographic context of the origins and spread of inversions, (ii) the conditions under which inversions can facilitate divergence, (iii) the magnitude of that facilitation, (iv) the extent to which the buildup of divergence is likely to be biased within vs. outside of inversions, and (v) the dynamics of the appearance and disappearance of exceptional divergence within inversions. We conclude by discussing the empirical challenges in showing that inversions play a central role in facilitating speciation with gene flow.

  4. Fluid front displacement dynamics affecting pressure fluctuations and phase entrapment in porous media

    Science.gov (United States)

    Moebius, F.; Or, D.

    2012-04-01

    Many natural and engineering processes involve motion of fluid fronts in porous media, from infiltration and drainage in hydrology to reservoir management in petroleum engineering. Macroscopically smooth and continuous motion of displacement fronts involves numerous rapid interfacial jumps and local reconfigurations. Detailed observations of displacement processes in micromodels illustrate the wide array of fluid interfacial dynamics ranging from irregular jumping-pinning motions to gradual pore scale invasions. The pressure fluctuations associated with interfacial motions reflect not only pore geometry (as traditionally hypothesized) but there is a strong influence of boundary conditions (e.g., mean drainage rate). The time scales associated with waiting time distribution of individual invasion events and decay time of inertial oscillations (following a rapid interfacial jump) provide a means for distinguishing between displacement regimes. Direct observations using high-speed camera combined with concurrent pressure signal measurements were instrumental in clarifying influences of flow rates, pore size, and gravity on burst size distribution and waiting times. We compared our results with the early experimental and theoretical study on burst size and waiting time distribution during slow drainage processes of Måløy et al. [Måløy et al., 1992]. Results provide insights on critical invasion events that exert strong influence on macroscopic phenomena such as front morphology and residual phase entrapment behind leading to hysteresis. Måløy, K. J., L. Furuberg, J. Feder, and T. Jossang (1992), Dynamics of Slow Drainage in Porous-Media, Phys Rev Lett, 68(14), 2161-2164.

  5. The bias associated with amplicon sequencing does not affect the quantitative assessment of bacterial community dynamics.

    Directory of Open Access Journals (Sweden)

    Federico M Ibarbalz

    Full Text Available The performance of two sets of primers targeting variable regions of the 16S rRNA gene V1-V3 and V4 was compared in their ability to describe changes of bacterial diversity and temporal turnover in full-scale activated sludge. Duplicate sets of high-throughput amplicon sequencing data of the two 16S rRNA regions shared a collection of core taxa that were observed across a series of twelve monthly samples, although the relative abundance of each taxon was substantially different between regions. A case in point was the changes in the relative abundance of filamentous bacteria Thiothrix, which caused a large effect on diversity indices, but only in the V1-V3 data set. Yet the relative abundance of Thiothrix in the amplicon sequencing data from both regions correlated with the estimation of its abundance determined using fluorescence in situ hybridization. In nonmetric multidimensional analysis samples were distributed along the first ordination axis according to the sequenced region rather than according to sample identities. The dynamics of microbial communities indicated that V1-V3 and the V4 regions of the 16S rRNA gene yielded comparable patterns of: 1 the changes occurring within the communities along fixed time intervals, 2 the slow turnover of activated sludge communities and 3 the rate of species replacement calculated from the taxa-time relationships. The temperature was the only operational variable that showed significant correlation with the composition of bacterial communities over time for the sets of data obtained with both pairs of primers. In conclusion, we show that despite the bias introduced by amplicon sequencing, the variable regions V1-V3 and V4 can be confidently used for the quantitative assessment of bacterial community dynamics, and provide a proper qualitative account of general taxa in the community, especially when the data are obtained over a convenient time window rather than at a single time point.

  6. Intracellular spatial localization regulated by the microtubule network.

    Directory of Open Access Journals (Sweden)

    Jing Chen

    Full Text Available The commonly recognized mechanisms for spatial regulation inside the cell are membrane-bounded compartmentalization and biochemical association with subcellular organelles. We use computational modeling to investigate another spatial regulation mechanism mediated by the microtubule network in the cell. Our results demonstrate that the mitotic spindle can impose strong sequestration and concentration effects on molecules with binding affinity for microtubules, especially dynein-directed cargoes. The model can recapitulate the essence of three experimental observations on distinct microtubule network morphologies: the sequestration of germ plasm components by the mitotic spindles in the Drosophila syncytial embryo, the asymmetric cell division initiated by the time delay in centrosome maturation in the Drosophila neuroblast, and the diffusional block between neighboring energids in the Drosophila syncytial embryo. Our model thus suggests that the cell cycle-dependent changes in the microtubule network are critical for achieving different spatial regulation effects. The microtubule network provides a spatially extensive docking platform for molecules and gives rise to a "structured cytoplasm", in contrast to a free and fluid environment.

  7. A Quantitative Method for Microtubule Analysis in Fluorescence Images.

    Science.gov (United States)

    Lan, Xiaodong; Li, Lingfei; Hu, Jiongyu; Zhang, Qiong; Dang, Yongming; Huang, Yuesheng

    2015-12-01

    Microtubule analysis is of significant value for a better understanding of normal and pathological cellular processes. Although immunofluorescence microscopic techniques have proven useful in the study of microtubules, comparative results commonly rely on a descriptive and subjective visual analysis. We developed an objective and quantitative method based on image processing and analysis of fluorescently labeled microtubular patterns in cultured cells. We used a multi-parameter approach by analyzing four quantifiable characteristics to compose our quantitative feature set. Then we interpreted specific changes in the parameters and revealed the contribution of each feature set using principal component analysis. In addition, we verified that different treatment groups could be clearly discriminated using principal components of the multi-parameter model. High predictive accuracy of four commonly used multi-classification methods confirmed our method. These results demonstrated the effectiveness and efficiency of our method in the analysis of microtubules in fluorescence images. Application of the analytical methods presented here provides information concerning the organization and modification of microtubules, and could aid in the further understanding of structural and functional aspects of microtubules under normal and pathological conditions.

  8. Microtubules contribute to maintain nucleus shape in epithelial cell monolayer

    Science.gov (United States)

    Tremblay, Dominique; Andrzejewski, Lukasz; Pelling, Andrew

    2013-03-01

    INTRODUCTION: Tissue strains can result in significant nuclear deformations and may regulate gene expression. However, the precise role of the cytoskeleton in regulating nuclear mechanics remains poorly understood. Here, we investigate the nuclear deformability of Madin-Darky canine kidney cells (MDCK) under various stretching conditions to clarify the role of the microtubules and actin network on the mechanical behavior of the nucleus. METHODS: A custom-built cell-stretching device allowing for real time imaging of MDCK nuclei was used. Cells were seeded on a silicone membrane coated with rat-tail collagen I. A nuclear stain, Hoechst-33342, was used to image nuclei during stretching. We exposed cells to a compressive and non-compressive stretching strain field of 25%. Nocodazole and cytochalasin-D were used to depolymerize the microtubules and actin network. RESULTS: Nuclei in control cells stretched more along their minor axis than major axis with a deformation of 5% and 2% respectively. This anisotropy vanished completely in microtubule-deprived cells and these cells showed a very high nuclear deformability along the minor axis when exposed to a compressive stretching strain field. CONCLUSIONS: The microtubules drive the anisotropic deformability of MDCK nuclei in a monolayer and maintain nuclear shape when exposed to compressive strain. Such intrinsic mechanical behavior indicates that microtubules are essential to maintain nuclear shape and may prevent down regulation of gene expression.

  9. The feasibility of coherent energy transfer in microtubules.

    Science.gov (United States)

    Craddock, Travis John Adrian; Friesen, Douglas; Mane, Jonathan; Hameroff, Stuart; Tuszynski, Jack A

    2014-11-06

    It was once purported that biological systems were far too 'warm and wet' to support quantum phenomena mainly owing to thermal effects disrupting quantum coherence. However, recent experimental results and theoretical analyses have shown that thermal energy may assist, rather than disrupt, quantum coherent transport, especially in the 'dry' hydrophobic interiors of biomolecules. Specifically, evidence has been accumulating for the necessary involvement of quantum coherent energy transfer between uniquely arranged chromophores in light harvesting photosynthetic complexes. The 'tubulin' subunit proteins, which comprise microtubules, also possess a distinct architecture of chromophores, namely aromatic amino acids, including tryptophan. The geometry and dipolar properties of these aromatics are similar to those found in photosynthetic units indicating that tubulin may support coherent energy transfer. Tubulin aggregated into microtubule geometric lattices may support such energy transfer, which could be important for biological signalling and communication essential to living processes. Here, we perform a computational investigation of energy transfer between chromophoric amino acids in tubulin via dipole excitations coupled to the surrounding thermal environment. We present the spatial structure and energetic properties of the tryptophan residues in the microtubule constituent protein tubulin. Plausibility arguments for the conditions favouring a quantum mechanism of signal propagation along a microtubule are provided. Overall, we find that coherent energy transfer in tubulin and microtubules is biologically feasible.

  10. Mangrove forest distributions and dynamics (19752005) of the tsunami-affected region of Asia

    Science.gov (United States)

    Giri, C.; Zhu, Z.; Tieszen, L.L.; Singh, A.; Gillette, S.; Kelmelis, J.A.

    2008-01-01

    Aim: We aimed to estimate the present extent of tsunami-affected mangrove forests and determine the rates and causes of deforestation from 1975 to 2005. Location: Our study region covers the tsunami-affected coastal areas of Indonesia, Malaysia, Thailand, Burma (Myanmar), Bangladesh, India and Sri Lanka in Asia. Methods: We interpreted time-series Landsat data using a hybrid supervised and unsupervised classification approach. Landsat data were geometrically corrected to an accuracy of plus-or-minus half a pixel, an accuracy necessary for change analysis. Each image was normalized for solar irradiance by converting digital number values to the top-of-the atmosphere reflectance. Ground truth data and existing maps and data bases were used to select training samples and also for iterative labelling. We used a post-classification change detection approach. Results: were validated with the help of local experts and/or high-resolution commercial satellite data. Results The region lost 12% of its mangrove forests from 1975 to 2005, to a present extent of c. 1,670,000 ha. Rates and causes of deforestation varied both spatially and temporally. Annual deforestation was highest in Burma (c. 1%) and lowest in Sri Lanka (0.1%). In contrast, mangrove forests in India and Bangladesh remained unchanged or gained a small percentage. Net deforestation peaked at 137,000 ha during 1990-2000, increasing from 97,000 ha during 1975-90, and declining to 14,000 ha during 2000-05. The major causes of deforestation were agricultural expansion (81%), aquaculture (12%) and urban development (2%). Main conclusions: We assessed and monitored mangrove forests in the tsunami-affected region of Asia using the historical archive of Landsat data. We also measured the rates of change and determined possible causes. The results of our study can be used to better understand the role of mangrove forests in saving lives and property from natural disasters such as the Indian Ocean tsunami, and to identify

  11. Interaction between microtubules and the Drosophila formin Cappuccino and its effect on actin assembly.

    Science.gov (United States)

    Roth-Johnson, Elizabeth A; Vizcarra, Christina L; Bois, Justin S; Quinlan, Margot E

    2014-02-14

    Formin family actin nucleators are potential coordinators of the actin and microtubule cytoskeletons, as they can both nucleate actin filaments and bind microtubules in vitro. To gain a more detailed mechanistic understanding of formin-microtubule interactions and formin-mediated actin-microtubule cross-talk, we studied microtubule binding by Cappuccino (Capu), a formin involved in regulating actin and microtubule organization during Drosophila oogenesis. We found that two distinct domains within Capu, FH2 and tail, work together to promote high-affinity microtubule binding. The tail domain appears to bind microtubules through nonspecific charge-based interactions. In contrast, distinct residues within the FH2 domain are important for microtubule binding. We also report the first visualization of a formin polymerizing actin filaments in the presence of microtubules. Interestingly, microtubules are potent inhibitors of the actin nucleation activity of Capu but appear to have little effect on Capu once it is bound to the barbed end of an elongating filament. Because Capu does not simultaneously bind microtubules and assemble actin filaments in vitro, its actin assembly and microtubule binding activities likely require spatial and/or temporal regulation within the Drosophila oocyte.

  12. Interaction between Microtubules and the Drosophila Formin Cappuccino and Its Effect on Actin Assembly*

    Science.gov (United States)

    Roth-Johnson, Elizabeth A.; Vizcarra, Christina L.; Bois, Justin S.; Quinlan, Margot E.

    2014-01-01

    Formin family actin nucleators are potential coordinators of the actin and microtubule cytoskeletons, as they can both nucleate actin filaments and bind microtubules in vitro. To gain a more detailed mechanistic understanding of formin-microtubule interactions and formin-mediated actin-microtubule cross-talk, we studied microtubule binding by Cappuccino (Capu), a formin involved in regulating actin and microtubule organization during Drosophila oogenesis. We found that two distinct domains within Capu, FH2 and tail, work together to promote high-affinity microtubule binding. The tail domain appears to bind microtubules through nonspecific charge-based interactions. In contrast, distinct residues within the FH2 domain are important for microtubule binding. We also report the first visualization of a formin polymerizing actin filaments in the presence of microtubules. Interestingly, microtubules are potent inhibitors of the actin nucleation activity of Capu but appear to have little effect on Capu once it is bound to the barbed end of an elongating filament. Because Capu does not simultaneously bind microtubules and assemble actin filaments in vitro, its actin assembly and microtubule binding activities likely require spatial and/or temporal regulation within the Drosophila oocyte. PMID:24362037

  13. Disturbance frequency and vertical distribution of seeds affect long-term population dynamics: a mechanistic seed bank model.

    Science.gov (United States)

    Eager, Eric Alan; Haridas, Chirakkal V; Pilson, Diana; Rebarber, Richard; Tenhumberg, Brigitte

    2013-08-01

    Seed banks are critically important for disturbance specialist plants because seeds of these species germinate only in disturbed soil. Disturbance and seed depth affect the survival and germination probability of seeds in the seed bank, which in turn affect population dynamics. We develop a density-dependent stochastic integral projection model to evaluate the effect of stochastic soil disturbances on plant population dynamics with an emphasis on mimicking how disturbances vertically redistribute seeds within the seed bank. We perform a simulation analysis of the effect of the frequency and mean depth of disturbances on the population's quasi-extinction probability, as well as the long-term mean and variance of the total density of seeds in the seed bank. We show that increasing the frequency of disturbances increases the long-term viability of the population, but the relationship between the mean depth of disturbance and the long-term viability of the population are not necessarily monotonic for all parameter combinations. Specifically, an increase in the probability of disturbance increases the long-term viability of the total seed bank population. However, if the probability of disturbance is too low, a shallower mean depth of disturbance can increase long-term viability, a relationship that switches as the probability of disturbance increases. However, a shallow disturbance depth is beneficial only in scenarios with low survival in the seed bank.

  14. Species sorting and patch dynamics in harlequin metacommunities affect the relative importance of environment and space.

    Science.gov (United States)

    Leibold, Mathew A; Loeuille, Nicolas

    2015-12-01

    Metacommunity theory indicates that variation in local community structure can be partitioned into components including those related to local environmental conditions vs. spatial effects and that these can be quantified using statistical methods based on variation partitioning. It has been hypothesized that joint associations of community composition with environment and space could be due to patch dynamics involving colonization-extinction processes in environmentally heterogeneous landscapes but this has yet to be theoretically shown. We develop a two-patch, type-two, species competition model in such a "harlequin" landscape (where different patches have different environments) to evaluate how composition is related to environmental and spatial effects as a function of background extinction rate. Using spatially implicit analytical models, we find that the environmental association of community composition declines with extinction rate as expected. Using spatially explicit simulation models, we further find that there is an increase in the spatial structure with extinction due to spatial patterning into clusters that are not related to environmental conditions but that this increase is limited. Natural metacommunities often show both environment and spatial determination even under conditions of relatively high isolation and these could be more easily explained by our model than alternative metacommunity models.

  15. Club position relative to the golfer's swing plane meaningfully affects swing dynamics.

    Science.gov (United States)

    MacKenzie, Sasho J

    2012-06-01

    Previous research indicates that the motion of the golf club is not planar and that the plane traced out by the club is different than that of the golfer's hands. The aim of the present study was to investigate how the position of the club, relative to the golfer's swing plane, influences the motion of the club by using a four-segment (torso, upper arm, forearm, and club), three-dimensional forward dynamics model. A genetic algorithm optimized the coordination of the model's four muscular torque generators to produce the best golf swings possible under six different conditions. The series of simulations were designed to demonstrate the effect of positioning the club above, and below, the golfer's swing plane as well as the effect of changing the steepness of the golfer's swing plane. The simulation results suggest that positioning the club below the golfer's swing plane, early in the downswing, will facilitate the squaring of the clubface for impact, while positioning the club above the plane will have the opposite effect. It was also demonstrated that changing the steepness of the golfer's swing plane by 10 degrees can have little effect on the delivery of the clubhead to the ball.

  16. Memory and obesity affect the population dynamics of asexual freshwater planarians

    Science.gov (United States)

    Dunkel, Jörn; Talbot, Jared; Schötz, Eva-Maria

    2011-04-01

    Asexual reproduction in multicellular organisms is a complex biophysical process that is not yet well understood quantitatively. Here, we report a detailed population study for the asexual freshwater planarian Schmidtea mediterranea, which can reproduce via transverse fission due to a large stem cell contingent. Our long-term observations of isolated non-interacting planarian populations reveal that the characteristic fission waiting time distributions for head and tail fragments differ significantly from each other. The stochastic fission dynamics of tail fragments exhibits non-negligible memory effects, implying that an accurate mathematical description of future data should be based on non-Markovian tree models. By comparing the effective growth of non-interacting planarian populations with those of self-interacting populations, we are able to quantify the influence of interactions between flatworms and physical conditions on the population growth. A surprising result is the non-monotonic relationship between effective population growth rate and nutrient supply: planarians exhibit a tendency to become 'obese' if the feeding frequency exceeds a critical level, resulting in a decreased reproduction activity. This suggests that these flatworms, which possess many genes homologous to those of humans, could become a new model system for studying dietary effects on reproduction and regeneration in multicellular organisms.

  17. Automated dynamic headspace/GC-MS analyses affect the repeatability of volatiles in irradiated Turkey.

    Science.gov (United States)

    Nam, Ki-Chang; Cordray, Joseph; Ahn, Dong U

    2004-03-24

    Although a dynamic headspace/gas chromatography-mass spectrometry (DH/GC-MS) method is an effective tool for determining volatiles of irradiated turkey meat, the profile of volatiles may be changeable depending upon the availability of oxygen in the sample vial and sample holding time before purge. The objective of this study was to evaluate the effects of helium flushing and sample holding time before purge on the volatiles profiles of irradiated raw and cooked turkey breast meat. Vacuum-packaged turkey breasts were irradiated at 2.5 kGy, and the volatiles of irradiated raw and cooked samples were analyzed using a DH/GC-MS with different holding times up to 280 min. The amounts of dimethyl disulfide and dimethyl trisulfide decreased as sample holding time in an autosampler (4 degrees C) before purge increased, whereas those of aldehdyes increased as holding time increased due to lipid oxidation. Helium flush of sample vials before sample loading on an autosampler retarded lipid oxidation and minimized the changes of sulfur volatiles in raw meat but was not enough to prevent oxidative changes in cooked meat. Although DH/GC-MS is a convenient method for automatic analysis of volatiles in meat samples, the number of samples that can be loaded in an autosampler at a time should be limited within the range that can permit reasonable repeatabilities for target volatile compounds.

  18. Continuous and discrete extreme climatic events affecting the dynamics of a high-arctic reindeer population.

    Science.gov (United States)

    Chan, Kung-Sik; Mysterud, Atle; Øritsland, Nils Are; Severinsen, Torbjørn; Stenseth, Nils Chr

    2005-10-01

    Climate at northern latitudes are currently changing both with regard to the mean and the temporal variability at any given site, increasing the frequency of extreme events such as cold and warm spells. Here we use a conceptually new modelling approach with two different dynamic terms of the climatic effects on a Svalbard reindeer population (the Brøggerhalvøya population) which underwent an extreme icing event ("locked pastures") with 80% reduction in population size during one winter (1993/94). One term captures the continuous and linear effect depending upon the Arctic Oscillation and another the discrete (rare) "event" process. The introduction of an "event" parameter describing the discrete extreme winter resulted in a more parsimonious model. Such an approach may be useful in strongly age-structured ungulate populations, with young and very old individuals being particularly prone to mortality factors during adverse conditions (resulting in a population structure that differs before and after extreme climatic events). A simulation study demonstrates that our approach is able to properly detect the ecological effects of such extreme climate events.

  19. Dynamic regimes of buoyancy-affected two-phase flow in unconsolidated porous media

    Science.gov (United States)

    Stöhr, M.; Khalili, A.

    2006-03-01

    The invasion and subsequent flow of a nonwetting fluid (NWF) in a three-dimensional, unconsolidated porous medium saturated with a wetting fluid of higher density and viscosity have been studied experimentally using a light-transmission technique. Distinct dynamic regimes have been found for different relative magnitudes of viscous, capillary, and gravity forces. It is shown that the ratio of viscous and hydrostatic pressure gradients can be used as a relevant dimensionless number K for the characterization of the different flow regimes. For low values of K , the invasion is characterized by the migration and fragmentation of isolated clusters of the NWF resulting from the prevalence of gravity and capillary forces. At high values of K , the dominance of viscous and gravity forces leads to an anisotropic fingerlike invasion. When the invasion stops after the breakthrough of the NWF at the open upper boundary, the invasion structure retracts under the influence of gravity and transforms into stable vertical channels. It is shown that the stability of these channels is the result of a balance between hydrostatic and viscous pressure gradients.

  20. Seasonal timing of first rain storms affects rare plant population dynamics

    Science.gov (United States)

    Levine, J.M.; McEachern, A.K.; Cowan, C.

    2011-01-01

    A major challenge in forecasting the ecological consequences of climate change is understanding the relative importance of changes to mean conditions vs. changes to discrete climatic events, such as storms, frosts, or droughts. Here we show that the first major storm of the growing season strongly influences the population dynamics of three rare and endangered annual plant species in a coastal California (USA) ecosystem. In a field experiment we used moisture barriers and water addition to manipulate the timing and temperature associated with first major rains of the season. The three focal species showed two- to fivefold variation in per capita population growth rates between the different storm treatments, comparable to variation found in a prior experiment imposing eightfold differences in season-long precipitation. Variation in germination was a major demographic driver of how two of three species responded to the first rains. For one of these species, the timing of the storm was the most critical determinant of its germination, while the other showed enhanced germination with colder storm temperatures. The role of temperature was further supported by laboratory trials showing enhanced germination in cooler treatments. Our work suggests that, because of species-specific cues for demographic transitions such as germination, changes to discrete climate events may be as, if not more, important than changes to season-long variables.

  1. Disorganization of cortical microtubules stimulates tangential expansion and reduces the uniformity of cellulose microfibril alignment among cells in the root of Arabidopsis.

    Science.gov (United States)

    Baskin, Tobias I; Beemster, Gerrit T S; Judy-March, Jan E; Marga, Françoise

    2004-08-01

    To test the role of cortical microtubules in aligning cellulose microfibrils and controlling anisotropic expansion, we exposed Arabidopsis thaliana roots to moderate levels of the microtubule inhibitor, oryzalin. After 2 d of treatment, roots grow at approximately steady state. At that time, the spatial profiles of relative expansion rate in length and diameter were quantified, and roots were cryofixed, freeze-substituted, embedded in plastic, and sectioned. The angular distribution of microtubules as a function of distance from the tip was quantified from antitubulin immunofluorescence images. In alternate sections, the overall amount of alignment among microfibrils and their mean orientation as a function of position was quantified with polarized-light microscopy. The spatial profiles of relative expansion show that the drug affects relative elongation and tangential expansion rates independently. The microtubule distributions averaged to transverse in the growth zone for all treatments, but on oryzalin the distributions became broad, indicating poorly organized arrays. At a subcellular scale, cellulose microfibrils in oryzalin-treated roots were as well aligned as in controls; however, the mean alignment direction, while consistently transverse in the controls, was increasingly variable with oryzalin concentration, meaning that microfibril orientation in one location tended to differ from that of a neighboring location. This conclusion was confirmed by direct observations of microfibrils with field-emission scanning electron microscopy. Taken together, these results suggest that cortical microtubules ensure microfibrils are aligned consistently across the organ, thereby endowing the organ with a uniform mechanical structure.

  2. Disorganization of Cortical Microtubules Stimulates Tangential Expansion and Reduces the Uniformity of Cellulose Microfibril Alignment among Cells in the Root of Arabidopsis1

    Science.gov (United States)

    Baskin, Tobias I.; Beemster, Gerrit T.S.; Judy-March, Jan E.; Marga, Françoise

    2004-01-01

    To test the role of cortical microtubules in aligning cellulose microfibrils and controlling anisotropic expansion, we exposed Arabidopsis thaliana roots to moderate levels of the microtubule inhibitor, oryzalin. After 2 d of treatment, roots grow at approximately steady state. At that time, the spatial profiles of relative expansion rate in length and diameter were quantified, and roots were cryofixed, freeze-substituted, embedded in plastic, and sectioned. The angular distribution of microtubules as a function of distance from the tip was quantified from antitubulin immunofluorescence images. In alternate sections, the overall amount of alignment among microfibrils and their mean orientation as a function of position was quantified with polarized-light microscopy. The spatial profiles of relative expansion show that the drug affects relative elongation and tangential expansion rates independently. The microtubule distributions averaged to transverse in the growth zone for all treatments, but on oryzalin the distributions became broad, indicating poorly organized arrays. At a subcellular scale, cellulose microfibrils in oryzalin-treated roots were as well aligned as in controls; however, the mean alignment direction, while consistently transverse in the controls, was increasingly variable with oryzalin concentration, meaning that microfibril orientation in one location tended to differ from that of a neighboring location. This conclusion was confirmed by direct observations of microfibrils with field-emission scanning electron microscopy. Taken together, these results suggest that cortical microtubules ensure microfibrils are aligned consistently across the organ, thereby endowing the organ with a uniform mechanical structure. PMID:15299138

  3. Quantum Computation in Brain Microtubules? Decoherence and Biological Feasibility

    CERN Document Server

    Hagan, S; Tuszynski, J A

    2000-01-01

    The Penrose-Hameroff (`Orch OR') model of quantum computation in brain microtubules has been criticized as regards the issue of environmental decoherence. A recent report by Tegmark finds that microtubules can maintain quantum coherence for only $10^{-13}$ s, far too short to be neurophysiologically relevant. Here, we critically examine the assumptions behind Tegmark's calculation and find that: 1) Tegmark's commentary is not aimed at an existing model in the literature but rather at a hybrid that replaces the superposed protein conformations of the `Orch OR' theory with a soliton in superposition along the microtubule, 2) Tegmark predicts decreasing decoherence times at lower temperature, in direct contradiction of the observed behavior of quantum states, 3) recalculation after correcting Tegmark's equation for differences between his model and the `Orch OR' model (superposition separation, charge vs. dipole, dielectric constant) lengthens the decoherence time to $10^{-5} - 10^{-4}$ s and invalidates a criti...

  4. Disruption of microtubule integrity initiates mitosis during CNS repair.

    Science.gov (United States)

    Bossing, Torsten; Barros, Claudia S; Fischer, Bettina; Russell, Steven; Shepherd, David

    2012-08-14

    Mechanisms of CNS repair have vital medical implications. We show that traumatic injury to the ventral midline of the embryonic Drosophila CNS activates cell divisions to replace lost cells. A pilot screen analyzing transcriptomes of single cells during repair pointed to downregulation of the microtubule-stabilizing GTPase mitochondrial Rho (Miro) and upregulation of the Jun transcription factor Jun-related antigen (Jra). Ectopic Miro expression can prevent midline divisions after damage, whereas Miro depletion destabilizes cortical β-tubulin and increases divisions. Disruption of cortical microtubules, either by chemical depolymerization or by overexpression of monomeric tubulin, triggers ectopic mitosis in the midline and induces Jra expression. Conversely, loss of Jra renders midline cells unable to replace damaged siblings. Our data indicate that upon injury, the integrity of the microtubule cytoskeleton controls cell division in the CNS midline, triggering extra mitosis to replace lost cells. The conservation of the identified molecules suggests that similar mechanisms may operate in vertebrates.

  5. Root cooling strongly affects diel leaf growth dynamics, water and carbohydrate relations in Ricinus communis.

    Science.gov (United States)

    Poiré, Richard; Schneider, Heike; Thorpe, Michael R; Kuhn, Arnd J; Schurr, Ulrich; Walter, Achim

    2010-03-01

    In laboratory and greenhouse experiments with potted plants, shoots and roots are exposed to temperature regimes throughout a 24 h (diel) cycle that can differ strongly from the regime under which these plants have evolved. In the field, roots are often exposed to lower temperatures than shoots. When the root-zone temperature in Ricinus communis was decreased below a threshold value, leaf growth occurred preferentially at night and was strongly inhibited during the day. Overall, leaf expansion, shoot biomass growth, root elongation and ramification decreased rapidly, carbon fluxes from shoot to root were diminished and carbohydrate contents of both root and shoot increased. Further, transpiration rate was not affected, yet hydrostatic tensions in shoot xylem increased. When root temperature was increased again, xylem tension reduced, leaf growth recovered rapidly, carbon fluxes from shoot to root increased, and carbohydrate pools were depleted. We hypothesize that the decreased uptake of water in cool roots diminishes the growth potential of the entire plant - especially diurnally, when the growing leaf loses water via transpiration. As a consequence, leaf growth and metabolite concentrations can vary enormously, depending on root-zone temperature and its heterogeneity inside pots.

  6. A compartmental model predicts that dietary potassium affects lithium dynamics in rats.

    Science.gov (United States)

    Everts, H B; Jang, H; Boston, R C; Canolty, N L

    1996-05-01

    Lithium is the treatment of choice for manic depression, but therapy often results in nephrogenic diabetes insipidus and lithium intoxication. To investigate the effects of dietary potassium on potential side effects of lithium therapy, a mathematical model was built using the modeling program SAAM (Simulation, Analysis, And Modeling). Experimental data modeled were from adult male Sprague-Dawley rats fed diets with or without lithium and one of three levels of potassium for 17 d. A five-compartment model of lithium dynamics was built that was consistent with data from rats fed a lithium-containing diet adequate in potassium. This model was then compared with data from rats fed the other two lithium-containing diets. The model predicts that both the fractional transfer coefficient and rate of transport of lithium to the serum compartment from the kidney compartment are lower in rats fed the potassium-adequate diet than in those fed the potassium-deficient diet, and even lower in those fed the potassium-supplemented diet. In addition, fractional transfer coefficients into the serum compartment from the sampled and simulated tissue compartments changed differently with time depending on the amount of dietary potassium. The model also predicts that there would be less accumulation of lithium in the kidney, sampled tissue and simulated tissue compartments with supplemental dietary potassium. The model suggests that potassium supplementation, after a 7-d delay, protects against nephrogenic diabetes insipidus and the potentially toxic accumulation of lithium by decreasing the reabsorption of lithium from the kidneys and increasing lithium efflux from the tissues.

  7. The EF loop in green proteorhodopsin affects conformation and photocycle dynamics.

    Science.gov (United States)

    Mehler, Michaela; Scholz, Frank; Ullrich, Sandra J; Mao, Jiafei; Braun, Markus; Brown, Lynda J; Brown, Richard C D; Fiedler, Sarah A; Becker-Baldus, Johanna; Wachtveitl, Josef; Glaubitz, Clemens

    2013-07-16

    The proteorhodopsin family consists of retinal proteins of marine bacterial origin with optical properties adjusted to their local environments. For green proteorhodopsin, a highly specific mutation in the EF loop, A178R, has been found to cause a surprisingly large redshift of 20 nm despite its distance from the chromophore. Here, we analyze structural and functional consequences of this EF loop mutation by time-resolved optical spectroscopy and solid-state NMR. We found that the primary photoreaction and the formation of the K-like photo intermediate is almost pH-independent and slower compared to the wild-type, whereas the decay of the K-intermediate is accelerated, suggesting structural changes within the counterion complex upon mutation. The photocycle is significantly elongated mainly due to an enlarged lifetime of late photo intermediates. Multidimensional MAS-NMR reveals mutation-induced chemical shift changes propagating from the EF loop to the chromophore binding pocket, whereas dynamic nuclear polarization-enhanced (13)C-double quantum MAS-NMR has been used to probe directly the retinylidene conformation. Our data show a modified interaction network between chromophore, Schiff base, and counterion complex explaining the altered optical and kinetic properties. In particular, the mutation-induced distorted structure in the EF loop weakens interactions, which help reorienting helix F during the reprotonation step explaining the slower photocycle. These data lead to the conclusion that the EF loop plays an important role in proton uptake from the cytoplasm but our data also reveal a clear interaction pathway between the EF loop and retinal binding pocket, which might be an evolutionary conserved communication pathway in retinal proteins.

  8. Investigating nitrate dynamics in a fine-textured soil affected by feedlot effluents

    Science.gov (United States)

    Veizaga, E. A.; Rodríguez, L.; Ocampo, C. J.

    2016-10-01

    N dynamics resulting from FL operations under heavy soils.

  9. Septins localize to microtubules during nutritional limitation in Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Vázquez de Aldana Carlos R

    2008-10-01

    Full Text Available Abstract Background In Saccharomyces cerevisiae, nutrient limitation stimulates diploid cells to undergo DNA replication and meiosis, followed by the formation of four haploid spores. Septins are a family of proteins that assemble a ring structure at the mother-daughter neck during vegetative growth, where they control cytokinesis. In sporulating cells, the septin ring disassembles and septins relocalize to the prospore membrane. Results Here, we demonstrate that nutrient limitation triggers a change in the localization of at least two vegetative septins (Cdc10 and Cdc11 from the bud neck to the microtubules. The association of Cdc10 and Cdc11 with microtubules persists into meiosis, and they are found associated with the meiotic spindle until the end of meiosis II. In addition, the meiosis-specific septin Spr28 displays similar behavior, suggesting that this is a common feature of septins. Septin association to microtubules is a consequence of the nutrient limitation signal, since it is also observed when haploid cells are incubated in sporulation medium and when haploid or diploid cells are grown in medium containing non-fermentable carbon sources. Moreover, during meiosis II, when the nascent prospore membrane is formed, septins moved from the microtubules to this membrane. Proper organization of the septins on the membrane requires the sporulation-specific septins Spr3 and Spr28. Conclusion Nutrient limitation in S. cerevisiae triggers the sporulation process, but it also induces the disassembly of the septin bud neck ring and relocalization of the septin subunits to the nucleus. Septins remain associated with microtubules during the meiotic divisions and later, during spore morphogenesis, they are detected associated to the nascent prospore membranes surrounding each nuclear lobe. Septin association to microtubules also occurs during growth in non-fermentable carbon sources.

  10. Three-dimensional reconstruction of tubulin sheets and re-investigation of microtubule surface lattice.

    Science.gov (United States)

    Schultheiss, R; Mandelkow, E

    1983-10-25

    Sheets are incomplete microtubule walls observed as polymorphic variants of microtubule assembly. Their substructure is similar to that of microtubules, as shown by two-dimensional optical and computer reconstruction. We have extended earlier studies by computing a three-dimensional reconstruction. From a re-investigation of the surface lattice it appears that the three-start helix of microtubules is right-handed rather than left-handed.

  11. In vivo and in vitro effects of the mitochondrial uncoupler FCCP on microtubules.

    OpenAIRE

    Maro, B.; Marty, M C; Bornens, M

    1982-01-01

    FCCP (carbonylcyanide-p-trifluoromethoxyphenylhydrazone), a potent uncoupler of oxidative phosphorylation, induces the complete disruption of cellular microtubules. A further analysis of this effect on BHK21 cells has shown that a decrease in the number of microtubules can be observed 15 min after adding FCCP and there is complete disruption after 60 min. Regrowth of microtubules was initiated 30 min after removal of FCCP, in marked contrast with the rapid reversion observed when microtubules...

  12. Kinesin-3 in the basidiomycete Ustilago maydis transports organelles along the entire microtubule array.

    Science.gov (United States)

    Steinberg, Gero

    2015-01-01

    The molecular motor kinesin-3 transports early endosomes along microtubules in filamentous fungi. It was reported that kinesin-3 from the ascomycete fungi Aspergillus nidulans and Neurospora crassa use a subset of post-translationally modified and more stable microtubules. Here, I show that kinesin-3 from the basidiomycete Ustilago maydis moves along all hyphal microtubules. This difference is likely due to variation in cell cycle control and associated organization of the microtubule array.

  13. A hopping mechanism for cargo transport by molecular motors in crowded microtubules

    CERN Document Server

    Goldman, Carla

    2010-01-01

    Most models designed to study the bidirectional movement of cargos as they are driven by molecular motors rely on the idea that motors of different polarities can be coordinated by external agents if arranged into a motor-cargo complex to perform the necessary work [gross04]. Although these models have provided us with important insights into these phenomena, there are still many unanswered questions regarding the mechanisms through which the movement of the complex takes place on crowded microtubules. For example (i) how does cargo-binding affect motor motility? and in connection with that - (ii) how does the presence of other motors (and also other cargos) on the microtubule affect the motility of the motor-cargo complex? We discuss these questions from a different perspective. The movement of a cargo is conceived here as a hopping process resulting from the transference of cargo between neighboring motors. In the light of this, we examine the conditions under which cargo might display bidirectional movemen...

  14. Simulation of water and nitrogen dynamics as affected by drip fertigation strategies

    Institute of Scientific and Technical Information of China (English)

    ZHANG Jian-jun; LI Jiu-sheng; ZHAO Bing-qiang; LI Yan-ting

    2015-01-01

    The aim of drip fertigation is synchronising the application of water and nutrients with crop requirements, and maintaining the proper concentration and distribution of nutrient and water in the soil. The wetting patterns and nutrient distributions under drip fertigation have been proved to be closely related to the fertigation strategies. In order to ifnd out the critical factors that affect the nutrient distribution under different drip fertigaiton strategies, a computer simulation model HYDRUS2D/3D was used to simulate the water and nitrate distribution for various fertigation strategies from a surface point source. Simulation results were compared with the observed ones from our previous studies. A 15° wedge-shaped plexiglass container was used in our experiment to represent one-twenty-fourth of the complete cylinder. The height of container is 40 cm, and the radius is 41 cm. The ammonium nitrate solution was added through a no. 7 needle connected to a Mariotte tube with a lfexible hose. The soil water content, nitrate and ammonium concentrations were measured. The comparison of simulated and observed data demonstrated that the model performed reliably. The numerical analysis for various fertigation strategies from a surface point source showed that: (1) The total amount of irrigation water, the concentration of the fertilizer solution and the amount of pure water used to lfush the pipeline after fertilizer solution application are the three critical factors inlfu-encing the distribution of water and fertilizer nitrogen in the soil. (2) The fresh water irrigation duration prior to fertigation has no obvious effect on nitrate distribution. The longer lfushing time period after fertigation resulted in nitrate accumulation closer to the wetting front. From the point of avoiding the possibility of nitrate loss from the root zone, we recommended that the lfushing time period should be as shorter as possible. (3) For a given amount of fertilizer, higher

  15. Cortical Microtubule Arrays Are Initiated from a Nonrandom Prepattern Driven by Atypical Microtubule Initiation1[W][OA

    Science.gov (United States)

    Lindeboom, Jelmer J.; Lioutas, Antonios; Deinum, Eva E.; Tindemans, Simon H.; Ehrhardt, David W.; Emons, Anne Mie C.; Vos, Jan W.; Mulder, Bela M.

    2013-01-01

    The ordered arrangement of cortical microtubules in growing plant cells is essential for anisotropic cell expansion and, hence, for plant morphogenesis. These arrays are dismantled when the microtubule cytoskeleton is rearranged during mitosis and reassembled following completion of cytokinesis. The reassembly of the cortical array has often been considered as initiating from a state of randomness, from which order arises at least partly through self-organizing mechanisms. However, some studies have shown evidence for ordering at early stages of array assembly. To investigate how cortical arrays are initiated in higher plant cells, we performed live-cell imaging studies of cortical array assembly in tobacco (Nicotiana tabacum) Bright Yellow-2 cells after cytokinesis and drug-induced disassembly. We found that cortical arrays in both cases did not initiate randomly but with a significant overrepresentation of microtubules at diagonal angles with respect to the cell axis, which coincides with the predominant orientation of the microtubules before their disappearance from the cell cortex in preprophase. In Arabidopsis (Arabidopsis thaliana) root cells, recovery from drug-induced disassembly was also nonrandom and correlated with the organization of the previous array, although no diagonal bias was observed in these cells. Surprisingly, during initiation, only about one-half of the new microtubules were nucleated from locations marked by green fluorescent protein-γ-tubulin complex protein2-tagged γ-nucleation complexes (γ-tubulin ring complex), therefore indicating that a large proportion of early polymers was initiated by a noncanonical mechanism not involving γ-tubulin ring complex. Simulation studies indicate that the high rate of noncanonical initiation of new microtubules has the potential to accelerate the rate of array repopulation. PMID:23300168

  16. The microtubule-stabilizing drug Epothilone D increases axonal sprouting following transection injury in vitro.

    Science.gov (United States)

    Brizuela, Mariana; Blizzard, Catherine A; Chuckowree, Jyoti A; Dawkins, Edgar; Gasperini, Robert J; Young, Kaylene M; Dickson, Tracey C

    2015-05-01

    Neuronal cytoskeletal alterations, in particular the loss and misalignment of microtubules, are considered a hallmark feature of the degeneration that occurs after traumatic brain injury (TBI). Therefore, microtubule-stabilizing drugs are attractive potential therapeutics for use following TBI. The best-known drug in this category is Paclitaxel, a widely used anti-cancer drug that has produced promising outcomes when employed in the treatment of various animal models of nervous system trauma. However, Paclitaxel is not ideal for the treatment of patients with TBI due to its limited blood-brain barrier (BBB) permeability. Herein we have characterized the effect of the brain penetrant microtubule-stabilizing agent Epothilone D (Epo D) on post-injury axonal sprouting in an in vitro model of CNS trauma. Epo D was found to modulate axonal sprout number in a dose dependent manner, increasing the number of axonal sprouts generated post-injury. Elevated sprouting was observed when analyzing the total population of injured neurons, as well as in selective analysis of Thy1-YFP-labeled excitatory neurons. However, we found no effect of Epo D on axonal sprout length or outgrowth speed. These findings indicate that Epo D specifically affects injury-induced axonal sprout generation, but not net growth. Our investigation demonstrates that primary cultures of cortical neurons are tolerant of Epo D exposure, and that Epo D significantly increases their regenerative response following structural injury. Therefore Epo D may be a potent therapeutic for enhancing regeneration following CNS injury. This article is part of a Special Issue entitled 'Traumatic Brain Injury'.

  17. Melt front propagation in dielectrics upon femtosecond laser irradiation: Formation dynamics of a heat-affected layer

    Science.gov (United States)

    Garcia-Lechuga, Mario; Solis, Javier; Siegel, Jan

    2016-04-01

    Several studies in dielectrics have reported the presence of a thin heat-affected layer underneath the ablation crater produced by femtosecond laser irradiation. In this work, we present a time-resolved microscopy technique that is capable of monitoring the formation dynamics of this layer and apply it to the study of a phosphate glass exposed to single pulses below the ablation threshold. A few nanoseconds after laser excitation, a melt front interface can be detected, which propagates into the bulk, gradually slowing down its speed. By means of image analysis combined with optical modeling, we are able to determine the temporal evolution of the layer thickness and its refractive index. Initially, a strong transient decrease in the refractive index is observed, which partially recovers afterwards. The layer resolidifies after approximately 1 μs after excitation, featuring a maximum thickness of several hundreds of nanometers.

  18. Dynamic modulation of inflammatory pain-related affective and sensory symptoms by optical control of amygdala metabotropic glutamate receptor 4.

    Science.gov (United States)

    Zussy, C; Gómez-Santacana, X; Rovira, X; De Bundel, D; Ferrazzo, S; Bosch, D; Asede, D; Malhaire, F; Acher, F; Giraldo, J; Valjent, E; Ehrlich, I; Ferraguti, F; Pin, J-P; Llebaria, A; Goudet, C

    2016-12-20

    Contrary to acute pain, chronic pain does not serve as a warning signal and must be considered as a disease per se. This pathology presents a sensory and psychological dimension at the origin of affective and cognitive disorders. Being largely refractory to current pharmacotherapies, identification of endogenous systems involved in persistent and chronic pain is crucial. The amygdala is a key brain region linking pain sensation with negative emotions. Here, we show that activation of a specific intrinsic neuromodulatory system within the amygdala associated with type 4 metabotropic glutamate receptors (mGlu4) abolishes sensory and affective symptoms of persistent pain such as hypersensitivity to pain, anxiety- and depression-related behaviors, and fear extinction impairment. Interestingly, neuroanatomical and synaptic analysis of the amygdala circuitry suggests that the effects of mGlu4 activation occur outside the central nucleus via modulation of multisensory thalamic inputs to lateral amygdala principal neurons and dorso-medial intercalated cells. Furthermore, we developed optogluram, a small diffusible photoswitchable positive allosteric modulator of mGlu4. This ligand allows the control of endogenous mGlu4 activity with light. Using this photopharmacological approach, we rapidly and reversibly inhibited behavioral symptoms associated with persistent pain through optical control of optogluram in the amygdala of freely behaving animals. Altogether, our data identify amygdala mGlu4 signaling as a mechanism that bypasses central sensitization processes to dynamically modulate persistent pain symptoms. Our findings help to define novel and more precise therapeutic interventions for chronic pain, and exemplify the potential of optopharmacology to study the dynamic activity of endogenous neuromodulatory mechanisms in vivo.Molecular Psychiatry advance online publication, 20 December 2016; doi:10.1038/mp.2016.223.

  19. Deciphering the Dynamics of Non-Covalent Interactions Affecting Thermal Stability of a Protein: Molecular Dynamics Study on Point Mutant of Thermus thermophilus Isopropylmalate Dehydrogenase.

    Science.gov (United States)

    Sharma, Reetu; Sastry, G Narahari

    2015-01-01

    Thermus thermophilius isopropylmalate dehydrogenase catalyzes oxidative decarboxylation and dehydrogenation of isopropylmalate. Substitution of leucine to alanine at position 172 enhances the thermal stability among the known point mutants. Exploring the dynamic properties of non-covalent interactions such as saltbridges, hydrogen bonds and hydrophobic interactions to explain thermal stability of a protein is interesting in its own right. In this study dynamic changes in the non-covalent interactions are studied to decipher the deterministic features of thermal stability of a protein considering a case study of a point mutant in Thermus thermophilus isopropylmalate dehydrogenase. A total of four molecular dynamic simulations of 0.2 μs were carried out on wild type and mutant's functional dimers at 300 K and 337 K. Higher thermal stability of the mutant as compared to wild type is revealed by root mean square deviation, root mean square fluctuations and Cα-Cα distance with an increase in temperature from 300 K to 337 K. Most of the regions of wild type fluctuate higher than the corresponding regions of mutant with an increase in temperature. Cα-Cα distance analysis suggests that long distance networks are significantly affected in wild type as compared to the mutant. Short lived contacts are higher in wild type, while long lived contacts are lost at 337 K. The mutant forms less hydrogen bonds with water as compared to wild type at 337 K. In contrast to wild type, the mutant shows significant increase in unique saltbridges, hydrogen bonds and hydrophobic contacts at 337 K. The current study indicates that there is a strong inter-dependence of thermal stability on the way in which non-covalent interactions reorganize, and it is rewarding to explore this connection in single mutant studies.

  20. Deciphering the Dynamics of Non-Covalent Interactions Affecting Thermal Stability of a Protein: Molecular Dynamics Study on Point Mutant of Thermus thermophilus Isopropylmalate Dehydrogenase.

    Directory of Open Access Journals (Sweden)

    Reetu Sharma

    Full Text Available Thermus thermophilius isopropylmalate dehydrogenase catalyzes oxidative decarboxylation and dehydrogenation of isopropylmalate. Substitution of leucine to alanine at position 172 enhances the thermal stability among the known point mutants. Exploring the dynamic properties of non-covalent interactions such as saltbridges, hydrogen bonds and hydrophobic interactions to explain thermal stability of a protein is interesting in its own right. In this study dynamic changes in the non-covalent interactions are studied to decipher the deterministic features of thermal stability of a protein considering a case study of a point mutant in Thermus thermophilus isopropylmalate dehydrogenase. A total of four molecular dynamic simulations of 0.2 μs were carried out on wild type and mutant's functional dimers at 300 K and 337 K. Higher thermal stability of the mutant as compared to wild type is revealed by root mean square deviation, root mean square fluctuations and Cα-Cα distance with an increase in temperature from 300 K to 337 K. Most of the regions of wild type fluctuate higher than the corresponding regions of mutant with an increase in temperature. Cα-Cα distance analysis suggests that long distance networks are significantly affected in wild type as compared to the mutant. Short lived contacts are higher in wild type, while long lived contacts are lost at 337 K. The mutant forms less hydrogen bonds with water as compared to wild type at 337 K. In contrast to wild type, the mutant shows significant increase in unique saltbridges, hydrogen bonds and hydrophobic contacts at 337 K. The current study indicates that there is a strong inter-dependence of thermal stability on the way in which non-covalent interactions reorganize, and it is rewarding to explore this connection in single mutant studies.

  1. How does stress affect human being—a molecular dynamic simulation study on cortisol and its glucocorticoid receptor

    Directory of Open Access Journals (Sweden)

    Dan Zhang

    2017-03-01

    Full Text Available Stress can be either positive or negative to human beings. Under stressful conditions, the mental and physical conditions of human can be affected. There exists certain relation between stress and illness. The cortisol and other glucocorticoids bind to the same receptor, which is called glucocorticoid receptor. Some evidences indicated that cortisol molecule binding to its glucocorticoid receptor was necessary for the stress response. Up to now, the structure–function relationships between cortisol molecule and its glucocorticoid receptor have not been deliberated from the atomic-level. In order to get a detailed understanding of the structure–function relationships between the cortisol molecule and glucocorticoids receptor, we have carried out molecular dynamic (MD simulations on glucocorticoid receptor (Apo system and cortisol with its glucocorticoid receptor complex (HCY system. On the basis of molecular dynamic simulations, a couple of key residues were identified, which were crucial for the binding of cortisol molecule. The results of binding free energy calculations are in good agreement with the experiment data. Our research gives clear insights from atomic-level into the structural–functional aspects of cortisol molecule and its glucocorticoid receptor, and also provides valuable information for the design of drug which can treat stress related illnesses.

  2. Microtubules accelerate the kinase activity of Aurora-B by a reduction in dimensionality.

    Science.gov (United States)

    Noujaim, Michael; Bechstedt, Susanne; Wieczorek, Michal; Brouhard, Gary J

    2014-01-01

    Aurora-B is the kinase subunit of the Chromosome Passenger Complex (CPC), a key regulator of mitotic progression that corrects improper kinetochore attachments and establishes the spindle midzone. Recent work has demonstrated that the CPC is a microtubule-associated protein complex and that microtubules are able to activate the CPC by contributing to Aurora-B auto-phosphorylation in trans. Aurora-B activation is thought to occur when the local concentration of Aurora-B is high, as occurs when Aurora-B is enriched at centromeres. It is not clear, however, whether distributed binding to large structures such as microtubules would increase the local concentration of Aurora-B. Here we show that microtubules accelerate the kinase activity of Aurora-B by a "reduction in dimensionality." We find that microtubules increase the kinase activity of Aurora-B toward microtubule-associated substrates while reducing the phosphorylation levels of substrates not associated to microtubules. Using the single molecule assay for microtubule-associated proteins, we show that a minimal CPC construct binds to microtubules and diffuses in a one-dimensional (1D) random walk. The binding of Aurora-B to microtubules is salt-dependent and requires the C-terminal tails of tubulin, indicating that the interaction is electrostatic. We show that the rate of Aurora-B auto-activation is faster with increasing concentrations of microtubules. Finally, we demonstrate that microtubules lose their ability to stimulate Aurora-B when their C-terminal tails are removed by proteolysis. We propose a model in which microtubules act as scaffolds for the enzymatic activity of Aurora-B. The scaffolding activity of microtubules enables rapid Aurora-B activation and efficient phosphorylation of microtubule-associated substrates.

  3. Acaricide treatment affects viral dynamics in Varroa destructor-infested honey bee colonies via both host physiology and mite control.

    Science.gov (United States)

    Locke, Barbara; Forsgren, Eva; Fries, Ingemar; de Miranda, Joachim R

    2012-01-01

    Honey bee (Apis mellifera) colonies are declining, and a number of stressors have been identified that affect, alone or in combination, the health of honey bees. The ectoparasitic mite Varroa destructor, honey bee viruses that are often closely associated with the mite, and pesticides used to control the mite population form a complex system of stressors that may affect honey bee health in different ways. During an acaricide treatment using Apistan (plastic strips coated with tau-fluvalinate), we analyzed the infection dynamics of deformed wing virus (DWV), sacbrood virus (SBV), and black queen cell virus (BQCV) in adult bees, mite-infested pupae, their associated Varroa mites, and uninfested pupae, comparing these to similar samples from untreated control colonies. Titers of DWV increased initially with the onset of the acaricide application and then slightly decreased progressively coinciding with the removal of the Varroa mite infestation. This initial increase in DWV titers suggests a physiological effect of tau-fluvalinate on the host's susceptibility to viral infection. DWV titers in adult bees and uninfested pupae remained higher in treated colonies than in untreated colonies. The titers of SBV and BQCV did not show any direct relationship with mite infestation and showed a variety of possible effects of the acaricide treatment. The results indicate that other factors besides Varroa mite infestation may be important to the development and maintenance of damaging DWV titers in colonies. Possible biochemical explanations for the observed synergistic effects between tau-fluvalinate and virus infections are discussed.

  4. Infestation dynamics of Brevipalpus phoenicis (Geijskes) (Acari: Tenuipalpidae) in citrus orchards as affected by edaphic and climatic variables.

    Science.gov (United States)

    Laranjeira, Francisco Ferraz; Silva, Suely Xavier de Brito; de Andrade, Eduardo Chumbinho; Almeida, Décio de Oliveira; da Silva, Tibério Santos Martins; Soares, Ana Cristina Fermino; Freitas-Astúa, Juliana

    2015-08-01

    Brevipalpus phoenicis (Geijskes) is a cosmopolitan and polyphagous mite that transmits important phytoviruses, such as coffee ringspot virus, passion fruit green spot virus and Citrus leprosis virus C. To characterise the dynamics of the probability and the rate of B. phoenicis infestation in response to edaphic and climatic factors, monthly inspections were performed in nine orchards in a citrus region of the State of Bahia, Brazil, for 35 months. Three fruits per plant were examined using a magnifying glass (10×) on 21 plants distributed along a "W"-shaped path in each orchard. Meteorological data were collected from a conventional station. To determine the correlations among the climatic variables, the data were analysed using Spearman correlations. Variables were selected by principal component analysis, and those that contributed the most to differentiate the groups were evaluated via a Mann-Whitney test. Using the quantile-quantile method, the limit values for the following climatic variables were determined: temperature (24.5 °C), photoperiod (12 h), relative humidity (83%), evapotranspiration (71 mm) and rainy days (14 days). The combination of longer days, higher temperatures, lower relative humidity levels and lower evapotranspiration increased the probability of B. phoenicis infestation, whereas successive rain events decreased that risk. Infestation rates were negatively affected by relative humidity levels above 83% and were positively affected by a decreasing available soil-water fraction and increasing insolation and photoperiod.

  5. Microtubule organization : from the centrosome to the Golgi apparatus

    NARCIS (Netherlands)

    Wu, J.

    2017-01-01

    Similar to the skeleton of a human body, every cell possesses the so-called cytoskeleton, a system of filaments that support cell shape and enable cells to divide and move. One of the major types of cytoskeletal fibers are microtubules, microscopic tubes that cells use as rails to transport their co

  6. Structure of the microtubule-binding domain of flagellar dynein.

    Science.gov (United States)

    Kato, Yusuke S; Yagi, Toshiki; Harris, Sarah A; Ohki, Shin-ya; Yura, Kei; Shimizu, Youské; Honda, Shinya; Kamiya, Ritsu; Burgess, Stan A; Tanokura, Masaru

    2014-11-04

    Flagellar dyneins are essential microtubule motors in eukaryotes, as they drive the beating motions of cilia and flagella. Unlike myosin and kinesin motors, the track binding mechanism of dyneins and the regulation between the strong and weak binding states remain obscure. Here we report the solution structure of the microtubule-binding domain of flagellar dynein-c/DHC9 (dynein-c MTBD). The structure reveals a similar overall helix-rich fold to that of the MTBD of cytoplasmic dynein (cytoplasmic MTBD), but dynein-c MTBD has an additional flap, consisting of an antiparallel b sheet. The flap is positively charged and highly flexible. Despite the structural similarity to cytoplasmic MTBD, dynein-c MTBD shows only a small change in the microtubule- binding affinity depending on the registry change of coiled coil-sliding, whereby lacks the apparent strong binding state. The surface charge distribution of dynein-c MTBD also differs from that of cytoplasmic MTBD, which suggests a difference in the microtubule-binding mechanism.

  7. Neuronal microtubule organization: from minus end to plus end

    NARCIS (Netherlands)

    Yau, K.W.

    2016-01-01

    Neurons are highly polarized cells consisting of a dendritic part and axonal part. Dendrites receive signals from other cells while axons transmit signals to other cells. In this thesis, mostly hippocampal neurons from rat embryos are used to study fundamental aspects of the microtubule organization

  8. Total synthesis of the potent microtubule-stabilizing agent (+)-discodermolide.

    Science.gov (United States)

    Harried, Scott S; Lee, Christopher P; Yang, Ge; Lee, Tony I H; Myles, David C

    2003-08-22

    The total synthesis of the potent microtubule-stabilizing, antimitotic agent (+)-discodermolide is described. The convergent synthetic strategy takes advantage of the diastereoselective alkylation of a ketone enolate to establish the key C15-C16 bond. The synthesis is amenable to preparation of gram-scale quantities of (+)-discodermolide and analogues.

  9. Direct Modulation of Microtubule Stability Contributes to Anthracene General Anesthesia

    Science.gov (United States)

    Emerson, Daniel J.; Weiser, Brian P.; Psonis, John; Liao, Zhengzheng; Taratula, Olena; Fiamengo, Ashley; Wang, Xiaozhao; Sugasawa, Keizo; Smith, Amos B.; Eckenhoff, Roderic G; Dmochowski, Ivan J.

    2013-01-01

    Recently, we identified 1-aminoanthracene as a fluorescent general anesthetic. To investigate the mechanism of action, a photoactive analogue, 1-azidoanthracene, was synthesized. Administration of 1-azidoanthrace