Griseofulvin stabilizes microtubule dynamics, activates p53 and inhibits the proliferation of MCF-7 cells synergistically with vinblastine

International Nuclear Information System (INIS)

Rathinasamy, Krishnan; Jindal, Bhavya; Asthana, Jayant; Singh, Parminder; Balaji, Petety V; Panda, Dulal

2010-01-01

Griseofulvin, an antifungal drug, has recently been shown to inhibit proliferation of various types of cancer cells and to inhibit tumor growth in athymic mice. Due to its low toxicity, griseofulvin has drawn considerable attention for its potential use in cancer chemotherapy. This work aims to understand how griseofulvin suppresses microtubule dynamics in living cells and sought to elucidate the antimitotic and antiproliferative action of the drug. The effects of griseofulvin on the dynamics of individual microtubules in live MCF-7 cells were measured by confocal microscopy. Immunofluorescence microscopy, western blotting and flow cytometry were used to analyze the effects of griseofulvin on spindle microtubule organization, cell cycle progression and apoptosis. Further, interactions of purified tubulin with griseofulvin were studied in vitro by spectrophotometry and spectrofluorimetry. Docking analysis was performed using autodock4 and LigandFit module of Discovery Studio 2.1. Griseofulvin strongly suppressed the dynamic instability of individual microtubules in live MCF-7 cells by reducing the rate and extent of the growing and shortening phases. At or near half-maximal proliferation inhibitory concentration, griseofulvin dampened the dynamicity of microtubules in MCF-7 cells without significantly disrupting the microtubule network. Griseofulvin-induced mitotic arrest was associated with several mitotic abnormalities like misaligned chromosomes, multipolar spindles, misegregated chromosomes resulting in cells containing fragmented nuclei. These fragmented nuclei were found to contain increased concentration of p53. Using both computational and experimental approaches, we provided evidence suggesting that griseofulvin binds to tubulin in two different sites; one site overlaps with the paclitaxel binding site while the second site is located at the αβ intra-dimer interface. In combination studies, griseofulvin and vinblastine were found to exert synergistic

An agent-based model contrasts opposite effects of dynamic and stable microtubules on cleavage furrow positioning.

Science.gov (United States)

Odell, Garrett M; Foe, Victoria E

2008-11-03

From experiments by Foe and von Dassow (Foe, V.E., and G. von Dassow. 2008. J. Cell Biol. 183:457-470) and others, we infer a molecular mechanism for positioning the cleavage furrow during cytokinesis. Computer simulations reveal how this mechanism depends on quantitative motor-behavior details and explore how robustly this mechanism succeeds across a range of cell sizes. The mechanism involves the MKLP1 (kinesin-6) component of centralspindlin binding to and walking along microtubules to stimulate cortical contractility where the centralspindlin complex concentrates. The majority of astral microtubules are dynamically unstable. They bind most MKLP1 and suppress cortical Rho/myosin II activation because the tips of unstable microtubules usually depolymerize before MKLP1s reach the cortex. A subset of astral microtubules stabilizes during anaphase, becoming effective rails along which MKLP1 can actually reach the cortex. Because stabilized microtubules aim statistically at the equatorial spindle midplane, that is where centralspindlin accumulates to stimulate furrow formation.

A computational framework for cortical microtubule dynamics in realistically shaped plant cells.

Directory of Open Access Journals (Sweden)

Bandan Chakrabortty

2018-02-01

Full Text Available Plant morphogenesis is strongly dependent on the directional growth and the subsequent oriented division of individual cells. It has been shown that the plant cortical microtubule array plays a key role in controlling both these processes. This ordered structure emerges as the collective result of stochastic interactions between large numbers of dynamic microtubules. To elucidate this complex self-organization process a number of analytical and computational approaches to study the dynamics of cortical microtubules have been proposed. To date, however, these models have been restricted to two dimensional planes or geometrically simple surfaces in three dimensions, which strongly limits their applicability as plant cells display a wide variety of shapes. This limitation is even more acute, as both local as well as global geometrical features of cells are expected to influence the overall organization of the array. Here we describe a framework for efficiently simulating microtubule dynamics on triangulated approximations of arbitrary three dimensional surfaces. This allows the study of microtubule array organization on realistic cell surfaces obtained by segmentation of microscopic images. We validate the framework against expected or known results for the spherical and cubical geometry. We then use it to systematically study the individual contributions of global geometry, cell-edge induced catastrophes and cell-face induced stability to array organization in a cuboidal geometry. Finally, we apply our framework to analyze the highly non-trivial geometry of leaf pavement cells of Arabidopsis thaliana, Nicotiana benthamiana and Hedera helix. We show that our simulations can predict multiple features of the microtubule array structure in these cells, revealing, among others, strong constraints on the orientation of division planes.

A computational framework for cortical microtubule dynamics in realistically shaped plant cells

KAUST Repository

Chakrabortty, Bandan; Blilou, Ikram; Scheres, Ben; Mulder, Bela M.

2018-01-01

Plant morphogenesis is strongly dependent on the directional growth and the subsequent oriented division of individual cells. It has been shown that the plant cortical microtubule array plays a key role in controlling both these processes. This ordered structure emerges as the collective result of stochastic interactions between large numbers of dynamic microtubules. To elucidate this complex self-organization process a number of analytical and computational approaches to study the dynamics of cortical microtubules have been proposed. To date, however, these models have been restricted to two dimensional planes or geometrically simple surfaces in three dimensions, which strongly limits their applicability as plant cells display a wide variety of shapes. This limitation is even more acute, as both local as well as global geometrical features of cells are expected to influence the overall organization of the array. Here we describe a framework for efficiently simulating microtubule dynamics on triangulated approximations of arbitrary three dimensional surfaces. This allows the study of microtubule array organization on realistic cell surfaces obtained by segmentation of microscopic images. We validate the framework against expected or known results for the spherical and cubical geometry. We then use it to systematically study the individual contributions of global geometry, cell-edge induced catastrophes and cell-face induced stability to array organization in a cuboidal geometry. Finally, we apply our framework to analyze the highly non-trivial geometry of leaf pavement cells of Arabidopsis thaliana, Nicotiana benthamiana and Hedera helix. We show that our simulations can predict multiple features of the microtubule array structure in these cells, revealing, among others, strong constraints on the orientation of division planes.

A computational framework for cortical microtubule dynamics in realistically shaped plant cells

KAUST Repository

Chakrabortty, Bandan

2018-02-02

Plant morphogenesis is strongly dependent on the directional growth and the subsequent oriented division of individual cells. It has been shown that the plant cortical microtubule array plays a key role in controlling both these processes. This ordered structure emerges as the collective result of stochastic interactions between large numbers of dynamic microtubules. To elucidate this complex self-organization process a number of analytical and computational approaches to study the dynamics of cortical microtubules have been proposed. To date, however, these models have been restricted to two dimensional planes or geometrically simple surfaces in three dimensions, which strongly limits their applicability as plant cells display a wide variety of shapes. This limitation is even more acute, as both local as well as global geometrical features of cells are expected to influence the overall organization of the array. Here we describe a framework for efficiently simulating microtubule dynamics on triangulated approximations of arbitrary three dimensional surfaces. This allows the study of microtubule array organization on realistic cell surfaces obtained by segmentation of microscopic images. We validate the framework against expected or known results for the spherical and cubical geometry. We then use it to systematically study the individual contributions of global geometry, cell-edge induced catastrophes and cell-face induced stability to array organization in a cuboidal geometry. Finally, we apply our framework to analyze the highly non-trivial geometry of leaf pavement cells of Arabidopsis thaliana, Nicotiana benthamiana and Hedera helix. We show that our simulations can predict multiple features of the microtubule array structure in these cells, revealing, among others, strong constraints on the orientation of division planes.

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

Directory of Open Access Journals (Sweden)

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.

Microtubules become more dynamic but not shorter during preprophase band formation: A possible "search-and-capture" mechanism for microtubule translocation

NARCIS (Netherlands)

Vos, J.W.; Dogterom, M.; Emons, A.M.C.

2004-01-01

The dynamic behavior of the microtubule cytoskeleton plays a crucial role in cellular organization, but the physical mechanisms underlying microtubule (re)organization in plant cells are poorly understood. We investigated microtubule dynamics in tobacco BY-2 suspension cells during interphase and

Biallelic Mutations in TBCD, Encoding the Tubulin Folding Cofactor D, Perturb Microtubule Dynamics and Cause Early-Onset Encephalopathy.

Science.gov (United States)

Flex, Elisabetta; Niceta, Marcello; Cecchetti, Serena; Thiffault, Isabelle; Au, Margaret G; Capuano, Alessandro; Piermarini, Emanuela; Ivanova, Anna A; Francis, Joshua W; Chillemi, Giovanni; Chandramouli, Balasubramanian; Carpentieri, Giovanna; Haaxma, Charlotte A; Ciolfi, Andrea; Pizzi, Simone; Douglas, Ganka V; Levine, Kara; Sferra, Antonella; Dentici, Maria Lisa; Pfundt, Rolph R; Le Pichon, Jean-Baptiste; Farrow, Emily; Baas, Frank; Piemonte, Fiorella; Dallapiccola, Bruno; Graham, John M; Saunders, Carol J; Bertini, Enrico; Kahn, Richard A; Koolen, David A; Tartaglia, Marco

2016-10-06

Microtubules are dynamic cytoskeletal elements coordinating and supporting a variety of neuronal processes, including cell division, migration, polarity, intracellular trafficking, and signal transduction. Mutations in genes encoding tubulins and microtubule-associated proteins are known to cause neurodevelopmental and neurodegenerative disorders. Growing evidence suggests that altered microtubule dynamics may also underlie or contribute to neurodevelopmental disorders and neurodegeneration. We report that biallelic mutations in TBCD, encoding one of the five co-chaperones required for assembly and disassembly of the αβ-tubulin heterodimer, the structural unit of microtubules, cause a disease with neurodevelopmental and neurodegenerative features characterized by early-onset cortical atrophy, secondary hypomyelination, microcephaly, thin corpus callosum, developmental delay, intellectual disability, seizures, optic atrophy, and spastic quadriplegia. Molecular dynamics simulations predicted long-range and/or local structural perturbations associated with the disease-causing mutations. Biochemical analyses documented variably reduced levels of TBCD, indicating relative instability of mutant proteins, and defective β-tubulin binding in a subset of the tested mutants. Reduced or defective TBCD function resulted in decreased soluble α/β-tubulin levels and accelerated microtubule polymerization in fibroblasts from affected subjects, demonstrating an overall shift toward a more rapidly growing and stable microtubule population. These cells displayed an aberrant mitotic spindle with disorganized, tangle-shaped microtubules and reduced aster formation, which however did not alter appreciably the rate of cell proliferation. Our findings establish that defective TBCD function underlies a recognizable encephalopathy and drives accelerated microtubule polymerization and enhanced microtubule stability, underscoring an additional cause of altered microtubule dynamics with

The Drosophila Microtubule-Associated Protein Mars Stabilizes Mitotic Spindles by Crosslinking Microtubules through Its N-Terminal Region

DEFF Research Database (Denmark)

Zhang, Gang; Beati, Hamze; Nilsson, Jakob

2013-01-01

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...

Learning-induced and stathmin-dependent changes in microtubule stability are critical for memory and disrupted in ageing

Science.gov (United States)

Uchida, Shusaku; Martel, Guillaume; Pavlowsky, Alice; Takizawa, Shuichi; Hevi, Charles; Watanabe, Yoshifumi; Kandel, Eric R.; Alarcon, Juan Marcos; Shumyatsky, Gleb P.

2014-01-01

Changes in the stability of microtubules regulate many biological processes, but their role in memory remains unclear. Here we show that learning causes biphasic changes in the microtubule-associated network in the hippocampus. In the early phase, stathmin is dephosphorylated, enhancing its microtubule-destabilizing activity by promoting stathmin-tubulin binding, whereas in the late phase these processes are reversed leading to an increase in microtubule/KIF5-mediated localization of the GluA2 subunit of AMPA receptors at synaptic sites. A microtubule stabilizer paclitaxel decreases or increases memory when applied at the early or late phases, respectively. Stathmin mutations disrupt changes in microtubule stability, GluA2 localization, synaptic plasticity and memory. Aged wild-type mice show impairments in stathmin levels, changes in microtubule stability, and GluA2 localization. Blocking GluA2 endocytosis rescues memory deficits in stathmin mutant and aged wild-type mice. These findings demonstrate a role for microtubules in memory in young adult and aged individuals. PMID:25007915

Four-stranded mini microtubules formed by Prosthecobacter BtubAB show dynamic instability.

Science.gov (United States)

Deng, Xian; Fink, Gero; Bharat, Tanmay A M; He, Shaoda; Kureisaite-Ciziene, Danguole; Löwe, Jan

2017-07-18

Microtubules, the dynamic, yet stiff hollow tubes built from αβ-tubulin protein heterodimers, are thought to be present only in eukaryotic cells. Here, we report a 3.6-Å helical reconstruction electron cryomicroscopy structure of four-stranded mini microtubules formed by bacterial tubulin-like Prosthecobacter dejongeii BtubAB proteins. Despite their much smaller diameter, mini microtubules share many key structural features with eukaryotic microtubules, such as an M-loop, alternating subunits, and a seam that breaks overall helical symmetry. Using in vitro total internal reflection fluorescence microscopy, we show that bacterial mini microtubules treadmill and display dynamic instability, another hallmark of eukaryotic microtubules. The third protein in the btub gene cluster, BtubC, previously known as "bacterial kinesin light chain," binds along protofilaments every 8 nm, inhibits BtubAB mini microtubule catastrophe, and increases rescue. Our work reveals that some bacteria contain regulated and dynamic cytomotive microtubule systems that were once thought to be only useful in much larger and sophisticated eukaryotic cells.

Nonlinear dynamics of C-terminal tails in cellular microtubules

Science.gov (United States)

Sekulic, Dalibor L.; Sataric, Bogdan M.; Zdravkovic, Slobodan; Bugay, Aleksandr N.; Sataric, Miljko V.

2016-07-01

The mechanical and electrical properties, and information processing capabilities of microtubules are the permanent subject of interest for carrying out experiments in vitro and in silico, as well as for theoretical attempts to elucidate the underlying processes. In this paper, we developed a new model of the mechano-electrical waves elicited in the rows of very flexible C-terminal tails which decorate the outer surface of each microtubule. The fact that C-terminal tails play very diverse roles in many cellular functions, such as recruitment of motor proteins and microtubule-associated proteins, motivated us to consider their collective dynamics as the source of localized waves aimed for communication between microtubule and associated proteins. Our approach is based on the ferroelectric liquid crystal model and it leads to the effective asymmetric double-well potential which brings about the conditions for the appearance of kink-waves conducted by intrinsic electric fields embedded in microtubules. These kinks can serve as the signals for control and regulation of intracellular traffic along microtubules performed by processive motions of motor proteins, primarly from kinesin and dynein families. On the other hand, they can be precursors for initiation of dynamical instability of microtubules by recruiting the proper proteins responsible for the depolymerization process.

Mechanics of microtubules: effects of protofilament orientation.

Science.gov (United States)

Donhauser, Zachary J; Jobs, William B; Binka, Edem C

2010-09-08

Microtubules are hollow cylindrical polymers of the protein tubulin that play a number of important dynamic and structural roles in eukaryotic cells. Both in vivo and in vitro microtubules can exist in several possible configurations, differing in the number of protofilaments, helical rise of tubulin dimers, and protofilament skew angle with respect to the main tube axis. Here, finite element modeling is applied to examine the mechanical response of several known microtubule types when subjected to radial deformation. The data presented here provide an important insight into microtubule stiffness and reveal that protofilament orientation does not affect radial stiffness. Rather, stiffness is primarily dependent on the effective Young's modulus of the polymerized material and the effective radius of the microtubule. These results are also directly correlated to atomic force microscopy nanoindentation measurements to allow a more detailed interpretation of previous experiments. When combined with experimental data that show a significant difference between microtubules stabilized with a slowly hydrolyzable GTP analog and microtubules stabilized with paclitaxel, the finite element data suggest that paclitaxel increases the overall radial flexibility of the microtubule wall. Copyright 2010 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Nonlinear dynamics of C–terminal tails in cellular microtubules

Energy Technology Data Exchange (ETDEWEB)

Sekulic, Dalibor L., E-mail: dalsek@uns.ac.rs; Sataric, Bogdan M.; Sataric, Miljko V. [University of Novi Sad, Faculty of Technical Sciences, Novi Sad (Serbia); Zdravkovic, Slobodan [University of Belgrade, Institute of Nuclear Sciences Vinca, Belgrade (Serbia); Bugay, Aleksandr N. [Laboratory of Radiation Biology, Joint Institute for Nuclear Research, Dubna (Russian Federation)

2016-07-15

The mechanical and electrical properties, and information processing capabilities of microtubules are the permanent subject of interest for carrying out experiments in vitro and in silico, as well as for theoretical attempts to elucidate the underlying processes. In this paper, we developed a new model of the mechano–electrical waves elicited in the rows of very flexible C–terminal tails which decorate the outer surface of each microtubule. The fact that C–terminal tails play very diverse roles in many cellular functions, such as recruitment of motor proteins and microtubule–associated proteins, motivated us to consider their collective dynamics as the source of localized waves aimed for communication between microtubule and associated proteins. Our approach is based on the ferroelectric liquid crystal model and it leads to the effective asymmetric double-well potential which brings about the conditions for the appearance of kink–waves conducted by intrinsic electric fields embedded in microtubules. These kinks can serve as the signals for control and regulation of intracellular traffic along microtubules performed by processive motions of motor proteins, primarly from kinesin and dynein families. On the other hand, they can be precursors for initiation of dynamical instability of microtubules by recruiting the proper proteins responsible for the depolymerization process.

Nonlinear dynamics of C–terminal tails in cellular microtubules

International Nuclear Information System (INIS)

Sekulic, Dalibor L.; Sataric, Bogdan M.; Sataric, Miljko V.; Zdravkovic, Slobodan; Bugay, Aleksandr N.

2016-01-01

The mechanical and electrical properties, and information processing capabilities of microtubules are the permanent subject of interest for carrying out experiments in vitro and in silico, as well as for theoretical attempts to elucidate the underlying processes. In this paper, we developed a new model of the mechano–electrical waves elicited in the rows of very flexible C–terminal tails which decorate the outer surface of each microtubule. The fact that C–terminal tails play very diverse roles in many cellular functions, such as recruitment of motor proteins and microtubule–associated proteins, motivated us to consider their collective dynamics as the source of localized waves aimed for communication between microtubule and associated proteins. Our approach is based on the ferroelectric liquid crystal model and it leads to the effective asymmetric double-well potential which brings about the conditions for the appearance of kink–waves conducted by intrinsic electric fields embedded in microtubules. These kinks can serve as the signals for control and regulation of intracellular traffic along microtubules performed by processive motions of motor proteins, primarly from kinesin and dynein families. On the other hand, they can be precursors for initiation of dynamical instability of microtubules by recruiting the proper proteins responsible for the depolymerization process.

The engine of microtubule dynamics comes into focus.

Science.gov (United States)

Mitchison, T J

2014-05-22

In this issue, Alushin et al. report high-resolution structures of three states of the microtubule lattice: GTP-bound, which is stable to depolymerization; unstable GDP-bound; and stable Taxol and GDP-bound. By comparing these structures at near-atomic resolution, they are able to propose a detailed model for how GTP hydrolysis destabilizes the microtubule and thus powers dynamic instability and chromosome movement. Destabilization of cytoskeleton filaments by nucleotide hydrolysis is an important general principle in cell dynamics, and this work represents a major step forward on a problem with a long history. Copyright © 2014 Elsevier Inc. All rights reserved.

Microtubule-Organizing Centers.

Science.gov (United States)

Wu, Jingchao; Akhmanova, Anna

2017-10-06

The organization of microtubule networks is crucial for controlling chromosome segregation during cell division, for positioning and transport of different organelles, and for cell polarity and morphogenesis. The geometry of microtubule arrays strongly depends on the localization and activity of the sites where microtubules are nucleated and where their minus ends are anchored. Such sites are often clustered into structures known as microtubule-organizing centers, which include the centrosomes in animals and spindle pole bodies in fungi. In addition, other microtubules, as well as membrane compartments such as the cell nucleus, the Golgi apparatus, and the cell cortex, can nucleate, stabilize, and tether microtubule minus ends. These activities depend on microtubule-nucleating factors, such as γ-tubulin-containing complexes and their activators and receptors, and microtubule minus end-stabilizing proteins with their binding partners. Here, we provide an overview of the current knowledge on how such factors work together to control microtubule organization in different systems.

Microtubule catastrophe and rescue.

Science.gov (United States)

Gardner, Melissa K; Zanic, Marija; Howard, Jonathon

2013-02-01

Microtubules are long cylindrical polymers composed of tubulin subunits. In cells, microtubules play an essential role in architecture and motility. For example, microtubules give shape to cells, serve as intracellular transport tracks, and act as key elements in important cellular structures such as axonemes and mitotic spindles. To accomplish these varied functions, networks of microtubules in cells are very dynamic, continuously remodeling through stochastic length fluctuations at the ends of individual microtubules. The dynamic behavior at the end of an individual microtubule is termed 'dynamic instability'. This behavior manifests itself by periods of persistent microtubule growth interrupted by occasional switching to rapid shrinkage (called microtubule 'catastrophe'), and then by switching back from shrinkage to growth (called microtubule 'rescue'). In this review, we summarize recent findings which provide new insights into the mechanisms of microtubule catastrophe and rescue, and discuss the impact of these findings in regards to the role of microtubule dynamics inside of cells. Copyright © 2012 Elsevier Ltd. All rights reserved.

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...

Complementary activities of TPX2 and chTOG constitute an efficient importin-regulated microtubule nucleation module.

Science.gov (United States)

Roostalu, Johanna; Cade, Nicholas I; Surrey, Thomas

2015-11-01

Spindle assembly and function require precise control of microtubule nucleation and dynamics. The chromatin-driven spindle assembly pathway exerts such control locally in the vicinity of chromosomes. One of the key targets of this pathway is TPX2. The molecular mechanism of how TPX2 stimulates microtubule nucleation is not understood. Using microscopy-based dynamic in vitro reconstitution assays with purified proteins, we find that human TPX2 directly stabilizes growing microtubule ends and stimulates microtubule nucleation by stabilizing early microtubule nucleation intermediates. Human microtubule polymerase chTOG (XMAP215/Msps/Stu2p/Dis1/Alp14 homologue) only weakly promotes nucleation, but acts synergistically with TPX2. Hence, a combination of distinct and complementary activities is sufficient for efficient microtubule formation in vitro. Importins control the efficiency of the microtubule nucleation by selectively blocking the interaction of TPX2 with microtubule nucleation intermediates. This in vitro reconstitution reveals the molecular mechanism of regulated microtubule formation by a minimal nucleation module essential for chromatin-dependent microtubule nucleation in cells.

Stabilization of microtubules by inorganic phosphate and its structural analogues, the fluoride complexes of aluminum and beryllium

International Nuclear Information System (INIS)

Carlier, M.F.; Didry, D.; Melki, R.; Chabre, M.; Pantaloni, D.

1988-01-01

In order to elucidate how the elementary reactions of GTP cleavage and subsequent inorganic phosphate (P/sub i/) release, which accompany microtubule assembly, regulate microtubule dynamics, the effect of P/sub i/ and of its structural analogues AlF 4 - and BeF 3 - on the stability of GDP-microtubules has been investigated. Inorganic phosphate binds to microtubules with a low affinity (K/sub D/ = 25 mM) and slows down the rate of GDP-subunit dissociation by about 2 orders of magnitude. AlF 4 - and BeF 3 - exhibit phosphate-like effects with 1000-fold higher affinity. Evidence has been obtained for direct binding of BeF 3 - to microtubules with a stoichiometry of 1 mol of BeF 3 - per mole of GDP-subunit and an equilibrium dissociation constant of 12-15 μM. AlF 4 - and P/sub i/ compete for this site. Phosphate analogues abolish oscillatory polymerization kinetics and slow down microtubule turnover at steady state. In view of these results, the authors propose that P/sub i/ and its structural analogues bind to the site of the γ-phosphate of GTP in the E site and reconstitute a GDP-P/sub i/-microtubule, from which tubulin subunits dissociate very slowly. They therefore understand that, following GTP cleavage on microtubules, P/sub i/ release in the medium is accompanied by a structural change resulting in a large destabilization of the polymer. A cap of slowly dissociating GDP-P/sub i/-subunits prevents depolymerization of the microtubule GDP-core at steady state. The similarity with the actin system is studied

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.

Cep169, a Novel Microtubule Plus-End-Tracking Centrosomal Protein, Binds to CDK5RAP2 and Regulates Microtubule Stability.

Directory of Open Access Journals (Sweden)

Yusuke Mori

Full Text Available The centrosomal protein, CDK5RAP2, is a microcephaly protein that regulates centrosomal maturation by recruitment of a γ-tubulin ring complex (γ-TuRC onto centrosomes. In this report, we identified a novel human centrosomal protein, Cep169, as a binding partner of CDK5RAP2, a member of microtubule plus-end-tracking proteins (+TIPs. Cep169 interacts directly with CDK5RAP2 through CM1, an evolutionarily conserved domain, and colocalizes at the pericentriolar matrix (PCM around centrioles with CDK5RAP2. In addition, Cep169 interacts with EB1 through SxIP-motif responsible for EB1 binding, and colocalizes with CDK5RAP2 at the microtubule plus-end. EB1-binding-deficient Cep169 abolishes EB1 interaction and microtubule plus-end attachment, indicating Cep169 as a novel member of +TIPs. We further show that ectopic expression of either Cep169 or CDK5RAP2 induces microtubule bundling and acetylation in U2OS cells, and depletion of Cep169 induces microtubule depolymerization in HeLa cells, although Cep169 is not required for assembly of γ-tubulin onto centrosome by CDK5RAP2. These results show that Cep169 targets microtubule tips and regulates stability of microtubules with CDK5RAP2.

Microtubules Nonlinear Models Dynamics Investigations through the exp(−Φ(ξ-Expansion Method Implementation

Directory of Open Access Journals (Sweden)

Nur Alam

2016-02-01

Full Text Available In this research article, we present exact solutions with parameters for two nonlinear model partial differential equations(PDEs describing microtubules, by implementing the exp(−Φ(ξ-Expansion Method. The considered models, describing highly nonlinear dynamics of microtubules, can be reduced to nonlinear ordinary differential equations. While the first PDE describes the longitudinal model of nonlinear dynamics of microtubules, the second one describes the nonlinear model of dynamics of radial dislocations in microtubules. The acquired solutions are then graphically presented, and their distinct properties are enumerated in respect to the corresponding dynamic behavior of the microtubules they model. Various patterns, including but not limited to regular, singular kink-like, as well as periodicity exhibiting ones, are detected. Being the method of choice herein, the exp(−Φ(ξ-Expansion Method not disappointing in the least, is found and declared highly efficient.

Brassinosteroids regulate pavement cell growth by mediating BIN2-induced microtubule stabilization.

Science.gov (United States)

Liu, Xiaolei; Yang, Qin; Wang, Yuan; Wang, Linhai; Fu, Ying; Wang, Xuelu

2018-02-23

Brassinosteroids (BRs), a group of plant steroid hormones, play important roles in regulating plant development. The cytoskeleton also affects key developmental processes and a deficiency in BR biosynthesis or signaling leads to abnormal phenotypes similar to those of microtubule-defective mutants. However, how BRs regulate microtubule and cell morphology remains unknown. Here, using liquid chromatography-tandem mass spectrometry, we identified tubulin proteins that interact with Arabidopsis BRASSINOSTEROID INSENSITIVE2 (BIN2), a negative regulator of BR responses in plants. In vitro and in vivo pull-down assays confirmed that BIN2 interacts with tubulin proteins. High-speed co-sedimentation assays demonstrated that BIN2 also binds microtubules. The Arabidopsis genome also encodes two BIN2 homologs, BIN2-LIKE 1 (BIL1) and BIL2, which function redundantly with BIN2. In the bin2-3 bil1 bil2 triple mutant, cortical microtubules were more sensitive to treatment with the microtubule-disrupting drug oryzalin than in wild-type, whereas in the BIN2 gain-of-function mutant bin2-1, cortical microtubules were insensitive to oryzalin treatment. These results provide important insight into how BR regulates plant pavement cell and leaf growth by mediating the stabilization of microtubules by BIN2.

Model for the orientational ordering of the plant microtubule cortical array

Science.gov (United States)

Hawkins, Rhoda J.; Tindemans, Simon H.; Mulder, Bela M.

2010-07-01

The plant microtubule cortical array is a striking feature of all growing plant cells. It consists of a more or less homogeneously distributed array of highly aligned microtubules connected to the inner side of the plasma membrane and oriented transversely to the cell growth axis. Here, we formulate a continuum model to describe the origin of orientational order in such confined arrays of dynamical microtubules. The model is based on recent experimental observations that show that a growing cortical microtubule can interact through angle dependent collisions with pre-existing microtubules that can lead either to co-alignment of the growth, retraction through catastrophe induction or crossing over the encountered microtubule. We identify a single control parameter, which is fully determined by the nucleation rate and intrinsic dynamics of individual microtubules. We solve the model analytically in the stationary isotropic phase, discuss the limits of stability of this isotropic phase, and explicitly solve for the ordered stationary states in a simplified version of the model.

Mutations in Human Tubulin Proximal to the Kinesin-Binding Site Alter Dynamic Instability at Microtubule Plus- and Minus-Ends

Energy Technology Data Exchange (ETDEWEB)

Ti, Shih-Chieh; Pamula, Melissa C.; Howes, Stuart C.; Duellberg, Christian; Cade, Nicholas I.; Kleiner, Ralph E.; Forth, Scott; Surrey, Thomas; Nogales, Eva; Kapoor, Tarun M.

2016-04-01

The assembly of microtubule-based cellular structures depends on regulated tubulin polymerization and directional transport. In this research, we have purified and characterized tubulin heterodimers that have human β-tubulin isotype III (TUBB3), as well as heterodimers with one of two β-tubulin mutations (D417H or R262H). Both point mutations are proximal to the kinesin-binding site and have been linked to an ocular motility disorder in humans. Compared to wild-type, microtubules with these mutations have decreased catastrophe frequencies and increased average lifetimes of plus- and minus-end-stabilizing caps. Importantly, the D417H mutation does not alter microtubule lattice structure or Mal3 binding to growing filaments. Instead, this mutation reduces the affinity of tubulin for TOG domains and colchicine, suggesting that the distribution of tubulin heterodimer conformations is changed. Together, our findings reveal how residues on the surface of microtubules, distal from the GTP-hydrolysis site and inter-subunit contacts, can alter polymerization dynamics at the plus- and minus-ends of microtubules.

Effect of radiation on microtubule structure in cancer cells

International Nuclear Information System (INIS)

Tripath, Shambhoo Sharan; Panda, Dulal; Jayakumar, S.; Maikho, Thoh; Sandur, Santosh Kumar

2017-01-01

Microtubules (MT) are dynamic structural cellular components. In proliferating cells, they are essential components in cell division through the formation of the mitotic spindle. Radiotherapy is an integral part of cancer treatment for most of the solid cancers. Scanty data exists in the literature related to how ionizing radiation affects microtubule reorganization in tumor cells. In the present study, breast cancer cell line (MCF-7 cells) was exposed to different doses of radiation (2-10Gy). Cells were cultured for 24 h, fixed and stained with antitubulin antibody and subjected to immunofluorescence microscopy. In another experiment, cells were subjected to cold treatment for 5 min or 30 min for studying the disassembly of microtubules after 24 h of irradiation. Further, these cells were incubated at 37°C for 20 min for studying the reassembly of microtubules. Acetylation of microtubule was also examined after exposure of cells to radiation. Experiments were also performed by combining radiation with low concentration of CXI-Benzo 84 (MT destabilizing agent 1 and 2.5 uM). Exposure of MCF-7 cells to radiation lead to destabilization of microtubules. Interestingly, destabilization of microtubule was faster upon cold treatment in irradiated group as compared to control group. These cells failed to re-stabilize at 37°C. Radiation also reduced the acetylation level of microtubule. Combination treatment of CXI-Benzo 84 with radiation exhibited additive effect in terms of depolymerization of MT. Our results suggest that ionizing radiation indeed modulates microtubule dynamics. (author)

Ibuprofen regulation of microtubule dynamics in cystic fibrosis epithelial cells.

Science.gov (United States)

Rymut, Sharon M; Kampman, Claire M; Corey, Deborah A; Endres, Tori; Cotton, Calvin U; Kelley, Thomas J

2016-08-01

High-dose ibuprofen, an effective anti-inflammatory therapy for the treatment of cystic fibrosis (CF), has been shown to preserve lung function in a pediatric population. Despite its efficacy, few patients receive ibuprofen treatment due to potential renal and gastrointestinal toxicity. The mechanism of ibuprofen efficacy is also unclear. We have previously demonstrated that CF microtubules are slower to reform after depolymerization compared with respective wild-type controls. Slower microtubule dynamics in CF cells are responsible for impaired intracellular transport and are related to inflammatory signaling. Here, it is identified that high-dose ibuprofen treatment in both CF cell models and primary CF nasal epithelial cells restores microtubule reformation rates to wild-type levels, as well as induce extension of microtubules to the cell periphery. Ibuprofen treatment also restores microtubule-dependent intracellular transport monitored by measuring intracellular cholesterol transport. These effects are specific to ibuprofen as other cyclooxygenase inhibitors have no effect on these measures. Effects of ibuprofen are mimicked by stimulation of AMPK and blocked by the AMPK inhibitor compound C. We conclude that high-dose ibuprofen treatment enhances microtubule formation in CF cells likely through an AMPK-related pathway. These findings define a potential mechanism to explain the efficacy of ibuprofen therapy in CF. Copyright © 2016 the American Physiological Society.

Moonlighting microtubule-associated proteins: regulatory functions by day and pathological functions at night.

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Oláh, J; Tőkési, N; Lehotzky, A; Orosz, F; Ovádi, J

2013-11-01

The sensing, integrating, and coordinating features of the eukaryotic cells are achieved by the complex ultrastructural arrays and multifarious functions of the cytoskeletal network. Cytoskeleton comprises fibrous protein networks of microtubules, actin, and intermediate filaments. These filamentous polymer structures are highly dynamic and undergo constant and rapid reorganization during cellular processes. The microtubular system plays a crucial role in the brain, as it is involved in an enormous number of cellular events including cell differentiation and pathological inclusion formation. These multifarious functions of microtubules can be achieved by their decoration with proteins/enzymes that exert specific effects on the dynamics and organization of the cytoskeleton and mediate distinct functions due to their moonlighting features. This mini-review focuses on two aspects of the microtubule cytoskeleton. On the one hand, we describe the heteroassociation of tubulin/microtubules with metabolic enzymes, which in addition to their catalytic activities stabilize microtubule structures via their cross-linking functions. On the other hand, we focus on the recently identified moonlighting tubulin polymerization promoting protein, TPPP/p25. TPPP/p25 is a microtubule-associated protein and it displays distinct physiological or pathological (aberrant) functions; thus it is a prototype of Neomorphic Moonlighting Proteins. The expression of TPPP/p25 is finely controlled in the human brain; this protein is indispensable for the development of projections of oligodendrocytes that are responsible for the ensheathment of axons. The nonphysiological, higher or lower TPPP/p25 level leads to distinct CNS diseases. Mechanisms contributing to the control of microtubule stability and dynamics by metabolic enzymes and TPPP/p25 will be discussed. Copyright © 2013 Wiley Periodicals, Inc.

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

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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.

Statistical mechanics provides novel insights into microtubule stability and mechanism of shrinkage.

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Ishutesh Jain

2015-02-01

Full Text Available Microtubules are nano-machines that grow and shrink stochastically, making use of the coupling between chemical kinetics and mechanics of its constituent protofilaments (PFs. We investigate the stability and shrinkage of microtubules taking into account inter-protofilament interactions and bending interactions of intrinsically curved PFs. Computing the free energy as a function of PF tip position, we show that the competition between curvature energy, inter-PF interaction energy and entropy leads to a rich landscape with a series of minima that repeat over a length-scale determined by the intrinsic curvature. Computing Langevin dynamics of the tip through the landscape and accounting for depolymerization, we calculate the average unzippering and shrinkage velocities of GDP protofilaments and compare them with the experimentally known results. Our analysis predicts that the strength of the inter-PF interaction (E(s(m has to be comparable to the strength of the curvature energy (E(b(m such that E(s(m - E(b(m ≈ 1kBT, and questions the prevalent notion that unzippering results from the domination of bending energy of curved GDP PFs. Our work demonstrates how the shape of the free energy landscape is crucial in explaining the mechanism of MT shrinkage where the unzippered PFs will fluctuate in a set of partially peeled off states and subunit dissociation will reduce the length.

NAD+ and SIRT3 control microtubule dynamics and reduce susceptibility to antimicrotubule agents

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Harkcom, William T.; Ghosh, Ananda K.; Sung, Matthew S.; Matov, Alexandre; Brown, Kevin D.; Giannakakou, Paraskevi; Jaffrey, Samie R.

2014-01-01

Nicotinamide adenine dinucleotide (NAD+) is an endogenous enzyme cofactor and cosubstrate that has effects on diverse cellular and physiologic processes, including reactive oxygen species generation, mitochondrial function, apoptosis, and axonal degeneration. A major goal is to identify the NAD+-regulated cellular pathways that may mediate these effects. Here we show that the dynamic assembly and disassembly of microtubules is markedly altered by NAD+. Furthermore, we show that the disassembly of microtubule polymers elicited by microtubule depolymerizing agents is blocked by increasing intracellular NAD+ levels. We find that these effects of NAD+ are mediated by the activation of the mitochondrial sirtuin sirtuin-3 (SIRT3). Overexpression of SIRT3 prevents microtubule disassembly and apoptosis elicited by antimicrotubule agents and knockdown of SIRT3 prevents the protective effects of NAD+ on microtubule polymers. Taken together, these data demonstrate that NAD+ and SIRT3 regulate microtubule polymerization and the efficacy of antimicrotubule agents. PMID:24889606

Plasma membrane factor XIIIA transglutaminase activity regulates osteoblast matrix secretion and deposition by affecting microtubule dynamics.

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Hadil F Al-Jallad

2011-01-01

Full Text Available Transglutaminase activity, arising potentially from transglutaminase 2 (TG2 and Factor XIIIA (FXIIIA, has been linked to osteoblast differentiation where it is required for type I collagen and fibronectin matrix deposition. In this study we have used an irreversible TG-inhibitor to 'block -and-track' enzyme(s targeted during osteoblast differentiation. We show that the irreversible TG-inhibitor is highly potent in inhibiting osteoblast differentiation and mineralization and reduces secretion of both fibronectin and type I collagen and their release from the cell surface. Tracking of the dansyl probe by Western blotting and immunofluorescence microscopy demonstrated that the inhibitor targets plasma membrane-associated FXIIIA. TG2 appears not to contribute to crosslinking activity on the osteoblast surface. Inhibition of FXIIIA with NC9 resulted in defective secretory vesicle delivery to the plasma membrane which was attributable to a disorganized microtubule network and decreased microtubule association with the plasma membrane. NC9 inhibition of FXIIIA resulted in destabilization of microtubules as assessed by cellular Glu-tubulin levels. Furthermore, NC9 blocked modification of Glu-tubulin into 150 kDa high-molecular weight Glu-tubulin form which was specifically localized to the plasma membrane. FXIIIA enzyme and its crosslinking activity were colocalized with plasma membrane-associated tubulin, and thus, it appears that FXIIIA crosslinking activity is directed towards stabilizing the interaction of microtubules with the plasma membrane. Our work provides the first mechanistic cues as to how transglutaminase activity could affect protein secretion and matrix deposition in osteoblasts and suggests a novel function for plasma membrane FXIIIA in microtubule dynamics.

EWSR1 regulates mitosis by dynamically influencing microtubule acetylation.

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Wang, Yi-Long; Chen, Hui; Zhan, Yi-Qun; Yin, Rong-Hua; Li, Chang-Yan; Ge, Chang-Hui; Yu, Miao; Yang, Xiao-Ming

2016-08-17

EWSR1, participating in transcription and splicing, has been identified as a translocation partner for various transcription factors, resulting in translocation, which in turn plays crucial roles in tumorigenesis. Recent studies have investigated the role of EWSR1 in mitosis. However, the effect of EWSR1 on mitosis is poorly understood. Here, we observed that depletion of EWSR1 resulted in cell cycle arrest in the mitotic phase, mainly due to an increase in the time from nuclear envelope breakdown to metaphase, resulting in a high percentage of unaligned chromosomes and multipolar spindles. We also demonstrated that EWSR1 is a spindle-associated protein that interacts with α-tubulin during mitosis. EWSR1 depletion increased the cold-sensitivity of spindle microtubules, and decreased the rate of spindle assembly. EWSR1 regulated the level of microtubule acetylation in the mitotic spindle; microtubule acetylation was rescued in EWSR1-depleted mitotic cells following suppression of HDAC6 activity by its specific inhibitor or siRNA treatment. In summary, these results suggest that EWSR1 regulates the acetylation of microtubules in a cell cycle-dependent manner through its dynamic location on spindle MTs, and may be a novel regulator for mitosis progress independent of its translocation.

A mutation of the fission yeast EB1 overcomes negative regulation by phosphorylation and stabilizes microtubules

International Nuclear Information System (INIS)

Iimori, Makoto; Ozaki, Kanako; Chikashige, Yuji; Habu, Toshiyuki; Hiraoka, Yasushi; Maki, Takahisa; Hayashi, Ikuko; Obuse, Chikashi; Matsumoto, Tomohiro

2012-01-01

Mal3 is a fission yeast homolog of EB1, a plus-end tracking protein (+ TIP). We have generated a mutation (89R) replacing glutamine with arginine in the calponin homology (CH) domain of Mal3. Analysis of the 89R mutant in vitro has revealed that the mutation confers a higher affinity to microtubules and enhances the intrinsic activity to promote the microtubule-assembly. The mutant Mal3 is no longer a + TIP, but binds strongly the microtubule lattice. Live cell imaging has revealed that while the wild type Mal3 proteins dissociate from the tip of the growing microtubules before the onset of shrinkage, the mutant Mal3 proteins persist on microtubules and reduces a rate of shrinkage after a longer pausing period. Consequently, the mutant Mal3 proteins cause abnormal elongation of microtubules composing the spindle and aster. Mal3 is phosphorylated at a cluster of serine/threonine residues in the linker connecting the CH and EB1-like C-terminal motif domains. The phosphorylation occurs in a microtubule-dependent manner and reduces the affinity of Mal3 to microtubules. We propose that because the 89R mutation is resistant to the effect of phosphorylation, it can associate persistently with microtubules and confers a stronger stability of microtubules likely by reinforcing the cylindrical structure. -- Highlights: ► We characterize a mutation (mal3-89R) in fission yeast homolog of EB1. ► The mutation enhances the activity to assemble microtubules. ► Mal3 is phosphorylated in a microtubule-dependent manner. ► The phosphorylation negatively regulates the Mal3 activity.

A mutation of the fission yeast EB1 overcomes negative regulation by phosphorylation and stabilizes microtubules

Energy Technology Data Exchange (ETDEWEB)

Iimori, Makoto; Ozaki, Kanako [Graduate School of Biostudies, Kyoto University, Kitashirakawa-Oiwake cho, Sakyo ku, Kyoto, 606-8502 (Japan); Chikashige, Yuji [Kobe Advanced ICT Research Center, National Institute of Information and Communications Technology, Kobe, 651-2492 (Japan); Habu, Toshiyuki [Graduate School of Biostudies, Kyoto University, Kitashirakawa-Oiwake cho, Sakyo ku, Kyoto, 606-8502 (Japan); Radiation Biology Center, Kyoto University, Yoshida-Konoe cho, Sakyo ku, Kyoto, 606-8501 (Japan); Hiraoka, Yasushi [Kobe Advanced ICT Research Center, National Institute of Information and Communications Technology, Kobe, 651-2492 (Japan); Graduate School of Frontier Biosciences, Osaka University, 1-3 Yamadaoka, Suita, 565-0871 (Japan); Maki, Takahisa; Hayashi, Ikuko [Graduate School of Nanobioscience, Yokohama City University, Tsurumi, Yokohama, 230-0045 (Japan); Obuse, Chikashi [Graduate School of Life Science, Hokkaido University, Sapporo 001-0021 (Japan); Matsumoto, Tomohiro, E-mail: tmatsumo@house.rbc.kyoto-u.ac.jp [Graduate School of Biostudies, Kyoto University, Kitashirakawa-Oiwake cho, Sakyo ku, Kyoto, 606-8502 (Japan); Radiation Biology Center, Kyoto University, Yoshida-Konoe cho, Sakyo ku, Kyoto, 606-8501 (Japan)

2012-02-01

Mal3 is a fission yeast homolog of EB1, a plus-end tracking protein (+ TIP). We have generated a mutation (89R) replacing glutamine with arginine in the calponin homology (CH) domain of Mal3. Analysis of the 89R mutant in vitro has revealed that the mutation confers a higher affinity to microtubules and enhances the intrinsic activity to promote the microtubule-assembly. The mutant Mal3 is no longer a + TIP, but binds strongly the microtubule lattice. Live cell imaging has revealed that while the wild type Mal3 proteins dissociate from the tip of the growing microtubules before the onset of shrinkage, the mutant Mal3 proteins persist on microtubules and reduces a rate of shrinkage after a longer pausing period. Consequently, the mutant Mal3 proteins cause abnormal elongation of microtubules composing the spindle and aster. Mal3 is phosphorylated at a cluster of serine/threonine residues in the linker connecting the CH and EB1-like C-terminal motif domains. The phosphorylation occurs in a microtubule-dependent manner and reduces the affinity of Mal3 to microtubules. We propose that because the 89R mutation is resistant to the effect of phosphorylation, it can associate persistently with microtubules and confers a stronger stability of microtubules likely by reinforcing the cylindrical structure. -- Highlights: Black-Right-Pointing-Pointer We characterize a mutation (mal3-89R) in fission yeast homolog of EB1. Black-Right-Pointing-Pointer The mutation enhances the activity to assemble microtubules. Black-Right-Pointing-Pointer Mal3 is phosphorylated in a microtubule-dependent manner. Black-Right-Pointing-Pointer The phosphorylation negatively regulates the Mal3 activity.

Microtubule Catastrophe and Rescue

OpenAIRE

Gardner, Melissa K.; Zanic, Marija; Howard, Jonathon

2012-01-01

Microtubules are long cylindrical polymers composed of tubulin subunits. In cells, microtubules play an essential role in architecture and motility. For example, microtubules give shape to cells, serve as intracellular transport tracks, and act as key elements in important cellular structures such as axonemes and mitotic spindles. To accomplish these varied functions, networks of microtubules in cells are very dynamic, continuously remodeling through stochastic length fluctuations at the ends...

TBCD links centriologenesis, spindle microtubule dynamics, and midbody abscission in human cells.

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Mónica López Fanarraga

2010-01-01

Full Text Available Microtubule-organizing centers recruit alpha- and beta-tubulin polypeptides for microtubule nucleation. Tubulin synthesis is complex, requiring five specific cofactors, designated tubulin cofactors (TBCs A-E, which contribute to various aspects of microtubule dynamics in vivo. Here, we show that tubulin cofactor D (TBCD is concentrated at the centrosome and midbody, where it participates in centriologenesis, spindle organization, and cell abscission. TBCD exhibits a cell-cycle-specific pattern, localizing on the daughter centriole at G1 and on procentrioles by S, and disappearing from older centrioles at telophase as the protein is recruited to the midbody. Our data show that TBCD overexpression results in microtubule release from the centrosome and G1 arrest, whereas its depletion produces mitotic aberrations and incomplete microtubule retraction at the midbody during cytokinesis. TBCD is recruited to the centriole replication site at the onset of the centrosome duplication cycle. A role in centriologenesis is further supported in differentiating ciliated cells, where TBCD is organized into "centriolar rosettes". These data suggest that TBCD participates in both canonical and de novo centriolar assembly pathways.

Integrins Regulate Apical Constriction via Microtubule Stabilization in the Drosophila Eye Disc Epithelium

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Vilaiwan M. Fernandes

2014-12-01

Full Text Available During morphogenesis, extracellular signals trigger actomyosin contractility in subpopulations of cells to coordinate changes in cell shape. To illuminate the link between signaling-mediated tissue patterning and cytoskeletal remodeling, we study the progression of the morphogenetic furrow (MF, the wave of apical constriction that traverses the Drosophila eye imaginal disc preceding photoreceptor neurogenesis. Apical constriction depends on actomyosin contractility downstream of the Hedgehog (Hh and bone morphogenetic protein (BMP pathways. We identify a role for integrin adhesion receptors in MF progression. We show that Hh and BMP regulate integrin expression, the loss of which disrupts apical constriction and slows furrow progression; conversely, elevated integrins accelerate furrow progression. We present evidence that integrins regulate MF progression by promoting microtubule stabilization, since reducing microtubule stability rescues integrin-mediated furrow acceleration. Thus, integrins act as a genetic link between tissue-level signaling events and morphological change at the cellular level, leading to morphogenesis and neurogenesis in the eye.

Deficiency of RITA results in multiple mitotic defects by affecting microtubule dynamics.

Science.gov (United States)

Steinhäuser, K; Klöble, P; Kreis, N-N; Ritter, A; Friemel, A; Roth, S; Reichel, J M; Michaelis, J; Rieger, M A; Louwen, F; Oswald, F; Yuan, J

2017-04-01

Deregulation of mitotic microtubule (MT) dynamics results in defective spindle assembly and chromosome missegregation, leading further to chromosome instability, a hallmark of tumor cells. RBP-J interacting and tubulin-associated protein (RITA) has been identified as a negative regulator of the Notch signaling pathway. Intriguingly, deregulated RITA is involved in primary hepatocellular carcinoma and other malignant entities. We were interested in the potential molecular mechanisms behind its involvement. We show here that RITA binds to tubulin and localizes to various mitotic MT structures. RITA coats MTs and affects their structures in vitro as well as in vivo. Tumor cell lines deficient of RITA display increased acetylated α-tubulin, enhanced MT stability and reduced MT dynamics, accompanied by multiple mitotic defects, including chromosome misalignment and segregation errors. Re-expression of wild-type RITA, but not RITA Δtub ineffectively binding to tubulin, restores the phenotypes, suggesting that the role of RITA in MT modulation is mediated via its interaction with tubulin. Mechanistically, RITA interacts with tubulin/histone deacetylase 6 (HDAC6) and its suppression decreases the binding of the deacetylase HDAC6 to tubulin/MTs. Furthermore, the mitotic defects and increased MT stability are also observed in RITA -/- mouse embryonic fibroblasts. RITA has thus a novel role in modulating MT dynamics and its deregulation results in erroneous chromosome segregation, one of the major reasons for chromosome instability in tumor cells.

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

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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.

Epothilones as lead structures for the synthesis-based discovery of new chemotypes for microtubule stabilization.

Science.gov (United States)

Feyen, Fabian; Cachoux, Frédéric; Gertsch, Jürg; Wartmann, Markus; Altmann, Karl-Heinz

2008-01-01

Epothilones are macrocyclic bacterial natural products with potent microtubule-stabilizing and antiproliferative activity. They have served as successful lead structures for the development of several clinical candidates for anticancer therapy. However, the structural diversity of this group of clinical compounds is rather limited, as their structures show little divergence from the original natural product leads. Our own research has explored the question of whether epothilones can serve as a basis for the development of new structural scaffolds, or chemotypes, for microtubule stabilization that might serve as a basis for the discovery of new generations of anticancer drugs. We have elaborated a series of epothilone-derived macrolactones whose overall structural features significantly deviate from those of the natural epothilone scaffold and thus define new structural families of microtubule-stabilizing agents. Key elements of our hypermodification strategy are the change of the natural epoxide geometry from cis to trans, the incorporation of a conformationally constrained side chain, the removal of the C3-hydroxyl group, and the replacement of C12 with nitrogen. So far, this approach has yielded analogs 30 and 40 that are the most advanced, the most rigorously modified, structures, both of which are potent antiproliferative agents with low nanomolar activity against several human cancer cell lines in vitro. The synthesis was achieved through a macrolactone-based strategy or a high-yielding RCM reaction. The 12-aza-epothilone ("azathilone" 40) may be considered a "non-natural" natural product that still retains most of the overall structural characteristics of a true natural product but is structurally unique, because it lies outside of the general scope of Nature's biosynthetic machinery for polyketide synthesis. Like natural epothilones, both 30 and 40 promote tubulin polymerization in vitro and at the cellular level induce cell cycle arrest in mitosis. These

Microtubule bundling plays a role in ethylene-mediated cortical microtubule reorientation in etiolated Arabidopsis hypocotyls.

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Ma, Qianqian; Sun, Jingbo; Mao, Tonglin

2016-05-15

The gaseous hormone ethylene is known to regulate plant growth under etiolated conditions (the 'triple response'). Although organization of cortical microtubules is essential for cell elongation, the underlying mechanisms that regulate microtubule organization by hormone signaling, including ethylene, are ambiguous. In the present study, we demonstrate that ethylene signaling participates in regulation of cortical microtubule reorientation. In particular, regulation of microtubule bundling is important for this process in etiolated hypocotyls. Time-lapse analysis indicated that selective stabilization of microtubule-bundling structures formed in various arrays is related to ethylene-mediated microtubule orientation. Bundling events and bundle growth lifetimes were significantly increased in oblique and longitudinal arrays, but decreased in transverse arrays in wild-type cells in response to ethylene. However, the effects of ethylene on microtubule bundling were partially suppressed in a microtubule-bundling protein WDL5 knockout mutant (wdl5-1). This study suggests that modulation of microtubule bundles that have formed in certain orientations plays a role in reorienting microtubule arrays in response to ethylene-mediated etiolated hypocotyl cell elongation. © 2016. Published by The Company of Biologists Ltd.

GDP-to-GTP exchange on the microtubule end can contribute to the frequency of catastrophe.

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Piedra, Felipe-Andrés; Kim, Tae; Garza, Emily S; Geyer, Elisabeth A; Burns, Alexander; Ye, Xuecheng; Rice, Luke M

2016-11-07

Microtubules are dynamic polymers of αβ-tubulin that have essential roles in chromosome segregation and organization of the cytoplasm. Catastrophe-the switch from growing to shrinking-occurs when a microtubule loses its stabilizing GTP cap. Recent evidence indicates that the nucleotide on the microtubule end controls how tightly an incoming subunit will be bound (trans-acting GTP), but most current models do not incorporate this information. We implemented trans-acting GTP into a computational model for microtubule dynamics. In simulations, growing microtubules often exposed terminal GDP-bound subunits without undergoing catastrophe. Transient GDP exposure on the growing plus end slowed elongation by reducing the number of favorable binding sites on the microtubule end. Slower elongation led to erosion of the GTP cap and an increase in the frequency of catastrophe. Allowing GDP-to-GTP exchange on terminal subunits in simulations mitigated these effects. Using mutant αβ-tubulin or modified GTP, we showed experimentally that a more readily exchangeable nucleotide led to less frequent catastrophe. Current models for microtubule dynamics do not account for GDP-to-GTP exchange on the growing microtubule end, so our findings provide a new way of thinking about the molecular events that initiate catastrophe. © 2016 Piedra et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

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

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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.

YB-1 promotes microtubule assembly in vitro through interaction with tubulin and microtubules

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Baconnais Sonia

2008-09-01

Full Text Available Abstract Background YB-1 is a major regulator of gene expression in eukaryotic cells. In addition to its role in transcription, YB-1 plays a key role in translation and stabilization of mRNAs. Results We show here that YB-1 interacts with tubulin and microtubules and stimulates microtubule assembly in vitro. High resolution imaging via electron and atomic force microscopy revealed that microtubules assembled in the presence of YB-1 exhibited a normal single wall ultrastructure and indicated that YB-1 most probably coats the outer microtubule wall. Furthermore, we found that YB-1 also promotes the assembly of MAPs-tubulin and subtilisin-treated tubulin. Finally, we demonstrated that tubulin interferes with RNA:YB-1 complexes. Conclusion These results suggest that YB-1 may regulate microtubule assembly in vivo and that its interaction with tubulin may contribute to the control of mRNA translation.

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

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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.

Biallelic Mutations in TBCD, Encoding the Tubulin Folding Cofactor D, Perturb Microtubule Dynamics and Cause Early-Onset Encephalopathy

NARCIS (Netherlands)

Flex, Elisabetta; Niceta, Marcello; Cecchetti, Serena; Thiffault, Isabelle; Au, Margaret G.; Capuano, Alessandro; Piermarini, Emanuela; Ivanova, Anna A.; Francis, Joshua W.; Chillemi, Giovanni; Chandramouli, Balasubramanian; Carpentieri, Giovanna; Haaxma, Charlotte A.; Ciolfi, Andrea; Pizzi, Simone; Douglas, Ganka V.; Levine, Kara; Sferra, Antonella; Dentici, Maria Lisa; Pfundt, Rolph R.; Le Pichon, Jean-Baptiste; Farrow, Emily; Baas, Frank; Piemonte, Fiorella; Dallapiccola, Bruno; Graham, John M.; Saunders, Carol J.; Bertini, Enrico; Kahn, Richard A.; Koolen, David A.; Tartaglia, Marco

2016-01-01

Microtubules are dynamic cytoskeletal elements coordinating and supporting a variety of neuronal processes, including cell division, migration, polarity, intracellular trafficking, and signal transduction. Mutations in genes encoding tubulins and microtubule-associated proteins are known to cause

Multiscale modeling and simulation of microtubule-motor-protein assemblies.

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

2015-01-01

Microtubules and motor proteins self-organize into biologically important assemblies including the mitotic spindle and the centrosomal microtubule array. Outside of cells, microtubule-motor mixtures can form novel active liquid-crystalline materials driven out of equilibrium by adenosine triphosphate-consuming motor proteins. Microscopic motor activity causes polarity-dependent interactions between motor proteins and microtubules, but how these interactions yield larger-scale dynamical behavior such as complex flows and defect dynamics is not well understood. We develop a multiscale theory for microtubule-motor systems in which Brownian dynamics simulations of polar microtubules driven by motors are used to study microscopic organization and stresses created by motor-mediated microtubule interactions. We identify polarity-sorting and crosslink tether relaxation as two polar-specific sources of active destabilizing stress. We then develop a continuum Doi-Onsager model that captures polarity sorting and the hydrodynamic flows generated by these polar-specific active stresses. In simulations of active nematic flows on immersed surfaces, the active stresses drive turbulent flow dynamics and continuous generation and annihilation of disclination defects. The dynamics follow from two instabilities, and accounting for the immersed nature of the experiment yields unambiguous characteristic length and time scales. When turning off the hydrodynamics in the Doi-Onsager model, we capture formation of polar lanes as observed in the Brownian dynamics simulation.

Multiscale modeling and simulation of microtubule-motor-protein assemblies

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

2015-12-01

Microtubules and motor proteins self-organize into biologically important assemblies including the mitotic spindle and the centrosomal microtubule array. Outside of cells, microtubule-motor mixtures can form novel active liquid-crystalline materials driven out of equilibrium by adenosine triphosphate-consuming motor proteins. Microscopic motor activity causes polarity-dependent interactions between motor proteins and microtubules, but how these interactions yield larger-scale dynamical behavior such as complex flows and defect dynamics is not well understood. We develop a multiscale theory for microtubule-motor systems in which Brownian dynamics simulations of polar microtubules driven by motors are used to study microscopic organization and stresses created by motor-mediated microtubule interactions. We identify polarity-sorting and crosslink tether relaxation as two polar-specific sources of active destabilizing stress. We then develop a continuum Doi-Onsager model that captures polarity sorting and the hydrodynamic flows generated by these polar-specific active stresses. In simulations of active nematic flows on immersed surfaces, the active stresses drive turbulent flow dynamics and continuous generation and annihilation of disclination defects. The dynamics follow from two instabilities, and accounting for the immersed nature of the experiment yields unambiguous characteristic length and time scales. When turning off the hydrodynamics in the Doi-Onsager model, we capture formation of polar lanes as observed in the Brownian dynamics simulation.

Microtubules Growth Rate Alteration in Human Endothelial Cells

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Irina B. Alieva

2010-01-01

Full Text Available To understand how microtubules contribute to the dynamic reorganization of the endothelial cell (EC cytoskeleton, we established an EC model expressing EB3-GFP, a protein that marks microtubule plus-ends. Using this model, we were able to measure microtubule growth rate at the centrosome region and near the cell periphery of a single human EC and in the EC monolayer. We demonstrate that the majority of microtubules in EC are dynamic, the growth rate of their plus-ends is highest in the internal cytoplasm, in the region of the centrosome. Growth rate of microtubule plus-ends decreases from the cell center toward the periphery. Our data suggest the existing mechanism(s of local regulation of microtubule plus-ends growth in EC. Microtubule growth rate in the internal cytoplasm of EC in the monolayer is lower than that of single EC suggesting the regulatory effect of cell-cell contacts. Centrosomal microtubule growth rate distribution in single EC indicated the presence of two subpopulations of microtubules with “normal” (similar to those in monolayer EC and “fast” (three times as much growth rates. Our results indicate functional interactions between cell-cell contacts and microtubules.

Assembly and control of large microtubule complexes

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Korolev, Kirill; Ishihara, Keisuke; Mitchison, Timothy

Motility, division, and other cellular processes require rapid assembly and disassembly of microtubule structures. We report a new mechanism for the formation of asters, radial microtubule complexes found in very large cells. The standard model of aster growth assumes elongation of a fixed number of microtubules originating from the centrosomes. However, aster morphology in this model does not scale with cell size, and we found evidence for microtubule nucleation away from centrosomes. By combining polymerization dynamics and auto-catalytic nucleation of microtubules, we developed a new biophysical model of aster growth. The model predicts an explosive transition from an aster with a steady-state radius to one that expands as a travelling wave. At the transition, microtubule density increases continuously, but aster growth rate discontinuously jumps to a nonzero value. We tested our model with biochemical perturbations in egg extract and confirmed main theoretical predictions including the jump in the growth rate. Our results show that asters can grow even though individual microtubules are short and unstable. The dynamic balance between microtubule collapse and nucleation could be a general framework for the assembly and control of large microtubule complexes. NIH GM39565; Simons Foundation 409704; Honjo International 486 Scholarship Foundation.

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

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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

SDF1 Reduces Interneuron Leading Process Branching through Dual Regulation of Actin and Microtubules

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Lysko, Daniel E.; Putt, Mary

2014-01-01

Normal cerebral cortical function requires a highly ordered balance between projection neurons and interneurons. During development these two neuronal populations migrate from distinct progenitor zones to form the cerebral cortex, with interneurons originating in the more distant ganglionic eminences. Moreover, deficits in interneurons have been linked to a variety of neurodevelopmental disorders underscoring the importance of understanding interneuron development and function. We, and others, have identified SDF1 signaling as one important modulator of interneuron migration speed and leading process branching behavior in mice, although how SDF1 signaling impacts these behaviors remains unknown. We previously found SDF1 inhibited leading process branching while increasing the rate of migration. We have now mechanistically linked SDF1 modulation of leading process branching behavior to a dual regulation of both actin and microtubule organization. We find SDF1 consolidates actin at the leading process tip by de-repressing calpain protease and increasing proteolysis of branched-actin-supporting cortactin. Additionally, SDF1 stabilizes the microtubule array in the leading process through activation of the microtubule-associated protein doublecortin (DCX). DCX stabilizes the microtubule array by bundling microtubules within the leading process, reducing branching. These data provide mechanistic insight into the regulation of interneuron leading process dynamics during neuronal migration in mice and provides insight into how cortactin and DCX, a known human neuronal migration disorder gene, participate in this process. PMID:24695713

SDF1 reduces interneuron leading process branching through dual regulation of actin and microtubules.

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Lysko, Daniel E; Putt, Mary; Golden, Jeffrey A

2014-04-02

Normal cerebral cortical function requires a highly ordered balance between projection neurons and interneurons. During development these two neuronal populations migrate from distinct progenitor zones to form the cerebral cortex, with interneurons originating in the more distant ganglionic eminences. Moreover, deficits in interneurons have been linked to a variety of neurodevelopmental disorders underscoring the importance of understanding interneuron development and function. We, and others, have identified SDF1 signaling as one important modulator of interneuron migration speed and leading process branching behavior in mice, although how SDF1 signaling impacts these behaviors remains unknown. We previously found SDF1 inhibited leading process branching while increasing the rate of migration. We have now mechanistically linked SDF1 modulation of leading process branching behavior to a dual regulation of both actin and microtubule organization. We find SDF1 consolidates actin at the leading process tip by de-repressing calpain protease and increasing proteolysis of branched-actin-supporting cortactin. Additionally, SDF1 stabilizes the microtubule array in the leading process through activation of the microtubule-associated protein doublecortin (DCX). DCX stabilizes the microtubule array by bundling microtubules within the leading process, reducing branching. These data provide mechanistic insight into the regulation of interneuron leading process dynamics during neuronal migration in mice and provides insight into how cortactin and DCX, a known human neuronal migration disorder gene, participate in this process.

Daple Coordinates Planar Polarized Microtubule Dynamics in Ependymal Cells and Contributes to Hydrocephalus

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Maki Takagishi

2017-07-01

Full Text Available Motile cilia in ependymal cells, which line the cerebral ventricles, exhibit a coordinated beating motion that drives directional cerebrospinal fluid (CSF flow and guides neuroblast migration. At the apical cortex of these multi-ciliated cells, asymmetric localization of planar cell polarity (PCP proteins is required for the planar polarization of microtubule dynamics, which coordinates cilia orientation. Daple is a disheveled-associating protein that controls the non-canonical Wnt signaling pathway and cell motility. Here, we show that Daple-deficient mice present hydrocephalus and their ependymal cilia lack coordinated orientation. Daple regulates microtubule dynamics at the anterior side of ependymal cells, which in turn orients the cilial basal bodies required for the directional cerebrospinal fluid flow. These results demonstrate an important role for Daple in planar polarity in motile cilia and provide a framework for understanding the mechanisms and functions of planar polarization in the ependymal cells.

Biological Information Processing in Single Microtubules

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2014-03-05

generated by synchronized oscillations of microtubules, centrosomes and chromosomes regulate the dynamics of mitosis and meiosis, Yue Zhao and Qimin...of frequency by a single microtubule. Green arrows depict the peaks that appear in absorption and disappear in transmission. Purple arrows show

Interaction of the Tobacco mosaic virus movement protein with microtubules during the cell cycle in tobacco BY-2 cells.

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Boutant, Emmanuel; Fitterer, Chantal; Ritzenthaler, Christophe; Heinlein, Manfred

2009-10-01

Cell-to-cell movement of Tobacco mosaic virus (TMV) involves the interaction of virus-encoded 30-kDa movement protein (MP) with microtubules. In cells behind the infection front that accumulate high levels of MP, this activity is reflected by the formation of stabilized MP/microtubule complexes. The ability of MP to bind along and stabilize microtubules is conserved upon expression in mammalian cells. In mammalian cells, the protein also leads to inhibition of mitosis and cell division through a microtubule-independent process correlated with the loss of centrosomal gamma-tubulin and of centrosomal microtubule-nucleation activity. Since MP has the capacity to interact with plant factors involved in microtubule nucleation and dynamics, we used inducible expression in BY-2 cells to test whether MP expression inhibits mitosis and cell division also in plants. We demonstrate that MP:GFP associates with all plant microtubule arrays and, unlike in mammalian cells, does not interfere with mitosis. Thus, MP function and the interaction of MP with factors of the cytoskeleton do not entail an inhibition of mitosis in plants. We also report that the protein targets primary plasmodesmata in BY-2 cells immediately upon or during cytokinesis and that the accumulation of MP in plasmodesmata occurs in the presence of inhibitors of the cytoskeleton and the secretory pathway.

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

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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.

Combing and self-assembly phenomena in dry films of Taxol-stabilized microtubules

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Rose Franck

2007-01-01

Full Text Available AbstractMicrotubules are filamentous proteins that act as a substrate for the translocation of motor proteins. As such, they may be envisioned as a scaffold for the self-assembly of functional materials and devices. Physisorption, self-assembly and combing are here investigated as a potential prelude to microtubule-templated self-assembly. Dense films of self-assembled microtubules were successfully produced, as well as patterns of both dendritic and non-dendritic bundles of microtubules. They are presented in the present paper and the mechanism of their formation is discussed.

Active Erk Regulates Microtubule Stability in H-ras-Transformed Cells

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Rene E. Harrison

2001-01-01

Full Text Available Increasing evidence suggests that activated erk regulates cell functions, at least in part, by mechanisms that do not require gene transcription. Here we show that the map kinase, erk, decorates microtubules (MTs and mitotic spindles in both parental and mutant active rastransfected 10T1 /2 fibroblasts and MCF10A breast epithelial cells. Approximately 20% of total cellular erk decorated MTs in both cell lines. A greater proportion of activated erk was associated with MTs in the presence of mutant active H-ras than in parental cells. Activation of erk by the ras pathway coincided with a decrease in the stability of MT, as detected by a stability marker. The MKK1 inhibitor, PD98059 and transfection of a dominant negative MKK1 blocked ras-induced instability of MTs but did not modify the association of erk with MTs or affect MT stability of the parental cells. These results indicate that the subset of active erk kinase that associates with MTs contributes to their instability in the presence of a mutant active ras. The MT-associated subset of active erk likely contributes to the enhanced invasive and proliferative abilities of cells containing mutant active H-ras.

Capture of microtubule plus-ends at the actin cortex promotes axophilic neuronal migration by enhancing microtubule tension in the leading process.

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Hutchins, B Ian; Wray, Susan

2014-01-01

Microtubules are a critical part of neuronal polarity and leading process extension, thus microtubule movement plays an important role in neuronal migration. However, the dynamics of microtubules during the forward movement of the nucleus into the leading process (nucleokinesis) is unclear and may be dependent on the cell type and mode of migration used. In particular, little is known about cytoskeletal changes during axophilic migration, commonly used in anteroposterior neuronal migration. We recently showed that leading process actin flow in migrating GnRH neurons is controlled by a signaling cascade involving IP3 receptors, CaMKK, AMPK, and RhoA. In the present study, microtubule dynamics were examined in GnRH neurons. Failure of the migration of these cells leads to the neuroendocrine disorder Kallmann Syndrome. Microtubules translocated forward along the leading process shaft during migration, but reversed direction and moved toward the nucleus when migration stalled. Blocking calcium release through IP3 receptors halted migration and induced the same reversal of microtubule translocation, while blocking cortical actin flow prevented microtubules from translocating toward the distal leading process. Super-resolution imaging revealed that microtubule plus-end tips are captured at the actin cortex through calcium-dependent mechanisms. This work shows that cortical actin flow draws the microtubule network forward through calcium-dependent capture in order to promote nucleokinesis, revealing a novel mechanism engaged by migrating neurons to facilitate movement.

Explaining the Microtubule Energy Balance: Contributions Due to Dipole Moments, Charges, van der Waals and Solvation Energy

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Ahmed Taha Ayoub

2017-09-01

Full Text Available Microtubules are the main components of mitotic spindles, and are the pillars of the cellular cytoskeleton. They perform most of their cellular functions by virtue of their unique dynamic instability processes which alternate between polymerization and depolymerization phases. This in turn is driven by a precise balance between attraction and repulsion forces between the constituents of microtubules (MTs—tubulin dimers. Therefore, it is critically important to know what contributions result in a balance of the interaction energy among tubulin dimers that make up microtubules and what interactions may tip this balance toward or away from a stable polymerized state of tubulin. In this paper, we calculate the dipole–dipole interaction energy between tubulin dimers in a microtubule as part of the various contributions to the energy balance. We also compare the remaining contributions to the interaction energies between tubulin dimers and establish a balance between stabilizing and destabilizing components, including the van der Waals, electrostatic, and solvent-accessible surface area energies. The energy balance shows that the GTP-capped tip of the seam at the plus end of microtubules is stabilized only by − 9 kcal/mol, which can be completely reversed by the hydrolysis of a single GTP molecule, which releases + 14 kcal/mol and destabilizes the seam by an excess of + 5 kcal/mol. This triggers the breakdown of microtubules and initiates a disassembly phase which is aptly called a catastrophe.

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

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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.

Microtubules self-repair in response to mechanical stress

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Schaedel, Laura; John, Karin; Gaillard, Jérémie; Nachury, Maxence V.; Blanchoin, Laurent; Théry, Manuel

2015-11-01

Microtubules--which define the shape of axons, cilia and flagella, and provide tracks for intracellular transport--can be highly bent by intracellular forces, and microtubule structure and stiffness are thought to be affected by physical constraints. Yet how microtubules tolerate the vast forces exerted on them remains unknown. Here, by using a microfluidic device, we show that microtubule stiffness decreases incrementally with each cycle of bending and release. Similar to other cases of material fatigue, the concentration of mechanical stresses on pre-existing defects in the microtubule lattice is responsible for the generation of more extensive damage, which further decreases microtubule stiffness. Strikingly, damaged microtubules were able to incorporate new tubulin dimers into their lattice and recover their initial stiffness. Our findings demonstrate that microtubules are ductile materials with self-healing properties, that their dynamics does not exclusively occur at their ends, and that their lattice plasticity enables the microtubules' adaptation to mechanical stresses.

Microtubules: A network for solitary waves

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Zdravković Slobodan

2017-01-01

Full Text Available In the present paper we deal with nonlinear dynamics of microtubules. The structure and role of microtubules in cells are explained as well as one of models explaining their dynamics. Solutions of the crucial nonlinear differential equation depend on used mathematical methods. Two commonly used procedures, continuum and semi-discrete approximations, are explained. These solutions are solitary waves usually called as kink solitons, breathers and bell-type solitons. [Project of the Serbian Ministry of Education, Science and Technological Development, Grant no. III45010

Melanophores for microtubule dynamics and motility assays.

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Ikeda, Kazuho; Semenova, Irina; Zhapparova, Olga; Rodionov, Vladimir

2010-01-01

Microtubules (MTs) are cytoskeletal structures essential for cell division, locomotion, intracellular transport, and spatial organization of the cytoplasm. In most interphase cells, MTs are organized into a polarized radial array with minus-ends clustered at the centrosome and plus-ends extended to the cell periphery. This array directs transport of organelles driven by MT-based motor proteins that specifically move either to plus- or to minus-ends. Along with using MTs as tracks for cargo, motor proteins can organize MTs into a radial array in the absence of the centrosome. Transport of organelles and motor-dependent radial organization of MTs require MT dynamics, continuous addition and loss of tubulin subunits at minus- and plus-ends. A unique experimental system for studying the role of MT dynamics in these processes is the melanophore, which provides a useful tool for imaging of both dynamic MTs and moving membrane organelles. Melanophores are filled with pigment granules that are synchronously transported by motor proteins in response to hormonal stimuli. The flat shape of the cell and the radial organization of MTs facilitate imaging of dynamic MT plus-ends and monitoring of their interaction with membrane organelles. Microsurgically produced cytoplasmic fragments of melanophores are used to study the centrosome-independent rearrangement of MTs into a radial array. Here we describe the experimental approaches to study the role of MT dynamics in intracellular transport and centrosome-independent MT organization in melanophores. We focus on the preparation of cell cultures, microsurgery and microinjection, fluorescence labeling, and live imaging of MTs. 2010 Elsevier Inc. All rights reserved.

Development of other microtubule-stabilizer families: the epothilones and their derivatives.

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Brogdon, Cynthia F; Lee, Francis Y; Canetta, Renzo M

2014-05-01

Chemotherapy is the mainstay of treatment for numerous cancer types, but resistance to chemotherapy remains a major clinical issue and is one of the driving influences underlying the development of new anticancer medications. One of the most important classes of chemotherapy agents is the taxanes, which target the cytoskeleton and spindle apparatus of tumor cells by binding to the microtubules, thereby disrupting key cellular mechanisms, including mitosis. Taxane resistance, however, limits treatment options and creates a major challenge for clinicians. Ongoing research has identified several newer classes of microtubule-targeting chemotherapies that may retain activity despite clinical resistance to taxanes. Among these classes, the epothilones have been studied most extensively in the clinical setting. Like taxanes, epothilones stabilize microtubulin turnover, and they have properties favoring their development as anticancer agents. The most clinically advanced epothilone analog is ixabepilone, which is currently the only approved epothilone derivative. Ixabepilone is indicated for the treatment of metastatic or locally advanced breast cancer in combination with capecitabine after failure of an anthracycline and a taxane, or as monotherapy after failure of an anthracycline, a taxane, and capecitabine. In phase II and III trials, ixabepilone showed efficacy in several patient subgroups and in various stages of breast cancer. Common adverse reactions include peripheral sensory neuropathy and asthenia. This paper will discuss the preclinical and clinical development of epothilones and their derivatives across a variety of cancer types.

C-terminal region of MAP7 domain containing protein 3 (MAP7D3 promotes microtubule polymerization by binding at the C-terminal tail of tubulin.

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Saroj Yadav

Full Text Available MAP7 domain containing protein 3 (MAP7D3, a newly identified microtubule associated protein, has been shown to promote microtubule assembly and stability. Its microtubule binding region has been reported to consist of two coiled coil motifs located at the N-terminus. It possesses a MAP7 domain near the C-terminus and belongs to the microtubule associated protein 7 (MAP7 family. The MAP7 domain of MAP7 protein has been shown to bind to kinesin-1; however, the role of MAP7 domain in MAP7D3 remains unknown. Based on the bioinformatics analysis of MAP7D3, we hypothesized that the MAP7 domain of MAP7D3 may have microtubule binding activity. Indeed, we found that MAP7 domain of MAP7D3 bound to microtubules as well as enhanced the assembly of microtubules in vitro. Interestingly, a longer fragment MDCT that contained the MAP7 domain (MD with the C-terminal tail (CT of the protein promoted microtubule polymerization to a greater extent than MD and CT individually. MDCT stabilized microtubules against dilution induced disassembly. MDCT bound to reconstituted microtubules with an apparent dissociation constant of 3.0 ± 0.5 µM. An immunostaining experiment showed that MDCT localized along the length of the preassembled microtubules. Competition experiments with tau indicated that MDCT shares its binding site on microtubules with tau. Further, we present evidence indicating that MDCT binds to the C-terminal tail of tubulin. In addition, MDCT could bind to tubulin in HeLa cell extract. Here, we report a microtubule binding region in the C-terminal region of MAP7D3 that may have a role in regulating microtubule assembly dynamics.

Dictyoceratidan poisons: Defined mark on microtubule-tubulin dynamics.

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Gnanambal K, Mary Elizabeth; Lakshmipathy, Shailaja Vommi

2016-03-01

Tubulin/microtubule assembly and disassembly is characterized as one of the chief processes during cell growth and division. Hence drugs those perturb these process are considered to be effective in killing fast multiplying cancer cells. There is a collection of natural compounds which disturb microtubule/tubulin dis/assemblage and there have been a lot of efforts concerted in the marine realm too, to surveying such killer molecules. Close to half the natural compounds shooting out from marine invertebrates are generally with no traceable definite mechanisms of action though may be tough anti-cancerous hits at nanogram levels, hence fatefully those discoveries conclude therein without a capacity of translation from laboratory to pharmacy. Astoundingly at least 50% of natural compounds which have definite mechanisms of action causing disorders in tubulin/microtubule kinetics have an isolation history from sponges belonging to the Phylum: Porifera. Poriferans have always been a wonder worker to treat cancers with a choice of, yet precise targets on cancerous tissues. There is a specific order: Dictyoceratida within this Phylum which has contributed to yielding at least 50% of effective compounds possessing this unique mechanism of action mentioned above. However, not much notice is driven to Dictyoceratidans alongside the order: Demospongiae thus dictating the need to know its select microtubule/tubulin irritants since the unearthing of avarol in the year 1974 till date. Hence this review selectively pinpoints all the compounds, noteworthy derivatives and analogs stemming from order: Dictyoceratida focusing on the past, present and future. Copyright © 2016 Elsevier Inc. All rights reserved.

Effects of the KIF2C neck peptide on microtubules: lateral disintegration of microtubules and β-structure formation.

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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. © 2013 The Authors Journal compilation © 2013 FEBS.

Buckling analysis of orthotropic protein microtubules under axial and radial compression based on couple stress theory.

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Beni, Yaghoub Tadi; Zeverdejani, M Karimi; Mehralian, Fahimeh

2017-10-01

Protein microtubules (MTs) are one of the important intercellular components and have a vital role in the stability and strength of the cells. Due to applied external loads, protein microtubules may be involved buckling phenomenon. Due to impact of protein microtubules in cell reactions, it is important to determine their critical buckling load. Considering nature of protein microtubules, various parameters are effective on microtubules buckling. The small size of microtubules and also lack of uniformity of MTs properties in different directions caused the necessity of accuracy in the analysis of these bio-structure. In fact, microtubules must be considered as a size dependent cylinder, which behave as an orthotropic material. Hence, in the present work using first-order shear deformation model (FSDT), the buckling equations of anisotropic MTs are derived based on new modified couple stress theory (NMCST). After solving the stability equations, the influences of various parameters are measured on the MTs critical buckling load. Copyright © 2017 Elsevier Inc. All rights reserved.

Stabilization of Microtubule-Unbound Tau via Tau Phosphorylation at Ser262/356 by Par-1/MARK Contributes to Augmentation of AD-Related Phosphorylation and Aβ42-Induced Tau Toxicity.

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Kanae Ando

2016-03-01

Full Text Available Abnormal accumulation of the microtubule-interacting protein tau is associated with neurodegenerative diseases including Alzheimer's disease (AD. β-amyloid (Aβ lies upstream of abnormal tau behavior, including detachment from microtubules, phosphorylation at several disease-specific sites, and self-aggregation into toxic tau species in AD brains. To prevent the cascade of events leading to neurodegeneration in AD, it is essential to elucidate the mechanisms underlying the initial events of tau mismetabolism. Currently, however, these mechanisms remain unclear. In this study, using transgenic Drosophila co-expressing human tau and Aβ, we found that tau phosphorylation at AD-related Ser262/356 stabilized microtubule-unbound tau in the early phase of tau mismetabolism, leading to neurodegeneration. Aβ increased the level of tau detached from microtubules, independent of the phosphorylation status at GSK3-targeted SP/TP sites. Such mislocalized tau proteins, especially the less phosphorylated species, were stabilized by phosphorylation at Ser262/356 via PAR-1/MARK. Levels of Ser262 phosphorylation were increased by Aβ42, and blocking this stabilization of tau suppressed Aβ42-mediated augmentation of tau toxicity and an increase in the levels of tau phosphorylation at the SP/TP site Thr231, suggesting that this process may be involved in AD pathogenesis. In contrast to PAR-1/MARK, blocking tau phosphorylation at SP/TP sites by knockdown of Sgg/GSK3 did not reduce tau levels, suppress tau mislocalization to the cytosol, or diminish Aβ-mediated augmentation of tau toxicity. These results suggest that stabilization of microtubule-unbound tau by phosphorylation at Ser262/356 via the PAR-1/MARK may act in the initial steps of tau mismetabolism in AD pathogenesis, and that such tau species may represent a potential therapeutic target for AD.

Quantitative Analysis of Tau-Microtubule Interaction Using FRET

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Isabelle L. Di Maïo

2014-08-01

Full Text Available The interaction between the microtubule associated protein, tau and the microtubules is investigated. A fluorescence resonance energy transfer (FRET assay was used to determine the distance separating tau to the microtubule wall, as well as the binding parameters of the interaction. By using microtubules stabilized with Flutax-2 as donor and tau labeled with rhodamine as acceptor, a donor-to-acceptor distance of 54 ± 1 Å was found. A molecular model is proposed in which Flutax-2 is directly accessible to tau-rhodamine molecules for energy transfer. By titration, we calculated the stoichiometric dissociation constant to be equal to 1.0 ± 0.5 µM. The influence of the C-terminal tails of αβ-tubulin on the tau-microtubule interaction is presented once a procedure to form homogeneous solution of cleaved tubulin has been determined. The results indicate that the C-terminal tails of α- and β-tubulin by electrostatic effects and of recruitment seem to be involved in the binding mechanism of tau.

Centromere Protein (CENP)-W Interacts with Heterogeneous Nuclear Ribonucleoprotein (hnRNP) U and May Contribute to Kinetochore-Microtubule Attachment in Mitotic Cells

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Chun, Younghwa; Kim, Raehyung; Lee, Soojin

2016-01-01

Background Recent studies have shown that heterogeneous nuclear ribonucleoprotein U (hnRNP U), a component of the hnRNP complex, contributes to stabilize the kinetochore-microtubule interaction during mitosis. CENP-W was identified as an inner centromere component that plays crucial roles in the formation of a functional kinetochore complex. Results We report that hnRNP U interacts with CENP-W, and the interaction between hnRNP U and CENP-W mutually increased each other’s protein stability by inhibiting the proteasome-mediated degradation. Further, their co-localization was observed chiefly in the nuclear matrix region and at the microtubule-kinetochore interface during interphase and mitosis, respectively. Both microtubule-stabilizing and microtubule-destabilizing agents significantly decreased the protein stability of CENP-W. Furthermore, loss of microtubules and defects in microtubule organization were observed in CENP-W-depleted cells. Conclusion Our data imply that CENP-W plays an important role in the attachment and interaction between microtubules and kinetochore during mitosis. PMID:26881882

Actin and microtubule networks contribute differently to cell response for small and large strains

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Kubitschke, H.; Schnauss, J.; Nnetu, K. D.; Warmt, E.; Stange, R.; Kaes, J.

2017-09-01

Cytoskeletal filaments provide cells with mechanical stability and organization. The main key players are actin filaments and microtubules governing a cell’s response to mechanical stimuli. We investigated the specific influences of these crucial components by deforming MCF-7 epithelial cells at small (≤5% deformation) and large strains (>5% deformation). To understand specific contributions of actin filaments and microtubules, we systematically studied cellular responses after treatment with cytoskeleton influencing drugs. Quantification with the microfluidic optical stretcher allowed capturing the relative deformation and relaxation of cells under different conditions. We separated distinctive deformational and relaxational contributions to cell mechanics for actin and microtubule networks for two orders of magnitude of drug dosages. Disrupting actin filaments via latrunculin A, for instance, revealed a strain-independent softening. Stabilizing these filaments by treatment with jasplakinolide yielded cell softening for small strains but showed no significant change at large strains. In contrast, cells treated with nocodazole to disrupt microtubules displayed a softening at large strains but remained unchanged at small strains. Stabilizing microtubules within the cells via paclitaxel revealed no significant changes for deformations at small strains, but concentration-dependent impact at large strains. This suggests that for suspended cells, the actin cortex is probed at small strains, while at larger strains; the whole cell is probed with a significant contribution from the microtubules.

Cyclin G2 is a centrosome-associated nucleocytoplasmic shuttling protein that influences microtubule stability and induces a p53-dependent cell cycle arrest

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Arachchige Don, Aruni S.; Dallapiazza, Robert F.; Bennin, David A.; Brake, Tiffany; Cowan, Colleen E.; Horne, Mary C.

2006-01-01

Cyclin G2 is an atypical cyclin that associates with active protein phosphatase 2A. Cyclin G2 gene expression correlates with cell cycle inhibition; it is significantly upregulated in response to DNA damage and diverse growth inhibitory stimuli, but repressed by mitogenic signals. Ectopic expression of cyclin G2 promotes cell cycle arrest, cyclin dependent kinase 2 inhibition and the formation of aberrant nuclei [Bennin, D. A., Don, A. S., Brake, T., McKenzie, J. L., Rosenbaum, H., Ortiz, L., DePaoli-Roach, A. A., and Horne, M. C. (2002). Cyclin G2 associates with protein phosphatase 2A catalytic and regulatory B' subunits in active complexes and induces nuclear aberrations and a G 1 /S-phase cell cycle arrest. J Biol Chem 277, 27449-67]. Here we report that endogenous cyclin G2 copurifies with centrosomes and microtubules (MT) and that ectopic G2 expression alters microtubule stability. We find exogenous and endogenous cyclin G2 present at microtubule organizing centers (MTOCs) where it colocalizes with centrosomal markers in a variety of cell lines. We previously reported that cyclin G2 forms complexes with active protein phosphatase 2A (PP2A) and colocalizes with PP2A in a detergent-resistant compartment. We now show that cyclin G2 and PP2A colocalize at MTOCs in transfected cells and that the endogenous proteins copurify with isolated centrosomes. Displacement of the endogenous centrosomal scaffolding protein AKAP450 that anchors PP2A at the centrosome resulted in the depletion of centrosomal cyclin G2. We find that ectopic expression of cyclin G2 induces microtubule bundling and resistance to depolymerization, inhibition of polymer regrowth from MTOCs and a p53-dependent cell cycle arrest. Furthermore, we determined that a 100 amino acid carboxy-terminal region of cyclin G2 is sufficient to both direct GFP localization to centrosomes and induce cell cycle inhibition. Colocalization of endogenous cyclin G2 with only one of two GFP-centrin-tagged centrioles, the

Microtubule-targeting drugs rescue axonal swellings in cortical neurons from spastin knockout mice

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Coralie Fassier

2013-01-01

Mutations in SPG4, encoding the microtubule-severing protein spastin, are responsible for the most frequent form of hereditary spastic paraplegia (HSP, a heterogeneous group of genetic diseases characterized by degeneration of the corticospinal tracts. We previously reported that mice harboring a deletion in Spg4, generating a premature stop codon, develop progressive axonal degeneration characterized by focal axonal swellings associated with impaired axonal transport. To further characterize the molecular and cellular mechanisms underlying this mutant phenotype, we have assessed microtubule dynamics and axonal transport in primary cultures of cortical neurons from spastin-mutant mice. We show an early and marked impairment of microtubule dynamics all along the axons of spastin-deficient cortical neurons, which is likely to be responsible for the occurrence of axonal swellings and cargo stalling. Our analysis also reveals that a modulation of microtubule dynamics by microtubule-targeting drugs rescues the mutant phenotype of cortical neurons. Together, these results contribute to a better understanding of the pathogenesis of SPG4-linked HSP and ascertain the influence of microtubule-targeted drugs on the early axonal phenotype in a mouse model of the disease.

A structural model for microtubule minus-end recognition and protection by CAMSAP proteins

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Atherton, Joseph; Jiang, Kai; Stangier, Marcel M.; Luo, Yanzhang; Hua, Shasha; Houben, Klaartje; Van Hooff, Jolien J.E.; Joseph, Agnel Praveen; Scarabelli, Guido; Grant, Barry J.; Roberts, Anthony J.; Topf, Maya; Steinmetz, Michel O.; Baldus, Marc; Moores, Carolyn A.; Akhmanova, Anna

2017-01-01

CAMSAP and Patronin family members regulate microtubule minus-end stability and localization and thus organize noncentrosomal microtubule networks, which are essential for cell division, polarization and differentiation. Here, we found that the CAMSAP C-terminal CKK domain is widely present among

Insight into microtubule destabilization mechanism of 3,4,5-trimethoxyphenyl indanone derivatives using molecular dynamics simulation and conformational modes analysis

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Tripathi, Shubhandra; Srivastava, Gaurava; Singh, Aastha; Prakasham, A. P.; Negi, Arvind S.; Sharma, Ashok

2018-03-01

Colchicine site inhibitors are microtubule destabilizers having promising role in cancer therapeutics. In the current study, four such indanone derivatives (t1, t9, t14 and t17) with 3,4,5-trimethoxyphenyl fragment (ring A) and showing significant microtubule destabilization property have been explored. The interaction mechanism and conformational modes triggered by binding of these indanone derivatives and combretastatin at colchicine binding site (CBS) of αβ-tubulin dimer were studied using molecular dynamics (MD) simulation, principle component analysis and free energy landscape analysis. In the MD results, t1 showed binding similar to colchicine interacting in the deep hydrophobic core at the CBS. While t9, t14 and t17 showed binding conformation similar to combretastatin, with ring A superficially binding at the CBS. Results demonstrated that ring A played a vital role in binding via hydrophobic interactions and got anchored between the S8 and S9 sheets, H8 helix and T7 loop at the CBS. Conformational modes study revealed that twisting and bending conformational motions (as found in the apo system) were nearly absent in the ligand bound systems. Absence of twisting motion might causes loss of lateral contacts in microtubule, thus promoting microtubule destabilization. This study provides detailed account of microtubule destabilization mechanism by indanone ligands and combretastatin, and would be helpful for designing microtubule destabilizers with higher activity.

Luminal localization of α-tubulin K40 acetylation by cryo-EM analysis of fab-labeled microtubules.

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Virupakshi Soppina

Full Text Available The αβ-tubulin subunits of microtubules can undergo a variety of evolutionarily-conserved post-translational modifications (PTMs that provide functional specialization to subsets of cellular microtubules. Acetylation of α-tubulin residue Lysine-40 (K40 has been correlated with increased microtubule stability, intracellular transport, and ciliary assembly, yet a mechanistic understanding of how acetylation influences these events is lacking. Using the anti-acetylated tubulin antibody 6-11B-1 and electron cryo-microscopy, we demonstrate that the K40 acetylation site is located inside the microtubule lumen and thus cannot directly influence events on the microtubule surface, including kinesin-1 binding. Surprisingly, the monoclonal 6-11B-1 antibody recognizes both acetylated and deacetylated microtubules. These results suggest that acetylation induces structural changes in the K40-containing loop that could have important functional consequences on microtubule stability, bending, and subunit interactions. This work has important implications for acetylation and deacetylation reaction mechanisms as well as for interpreting experiments based on 6-11B-1 labeling.

Arabidopsis FH1 Formin Affects Cotyledon Pavement Cell Shape by Modulating Cytoskeleton Dynamics.

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Rosero, Amparo; Oulehlová, Denisa; Stillerová, Lenka; Schiebertová, Petra; Grunt, Michal; Žárský, Viktor; Cvrčková, Fatima

2016-03-01

Plant cell morphogenesis involves concerted rearrangements of microtubules and actin microfilaments. We previously reported that FH1, the main Arabidopsis thaliana housekeeping Class I membrane-anchored formin, contributes to actin dynamics and microtubule stability in rhizodermis cells. Here we examine the effects of mutations affecting FH1 (At3g25500) on cell morphogenesis and above-ground organ development in seedlings, as well as on cytoskeletal organization and dynamics, using a combination of confocal and variable angle epifluorescence microscopy with a pharmacological approach. Homozygous fh1 mutants exhibited cotyledon epinasty and had larger cotyledon pavement cells with more pronounced lobes than the wild type. The pavement cell shape alterations were enhanced by expression of the fluorescent microtubule marker GFP-microtubule-associated protein 4 (MAP4). Mutant cotyledon pavement cells exhibited reduced density and increased stability of microfilament bundles, as well as enhanced dynamics of microtubules. Analogous results were also obtained upon treatments with the formin inhibitor SMIFH2 (small molecule inhibitor of formin homology 2 domains). Pavement cell shape in wild-type (wt) and fh1 plants in some situations exhibited a differential response towards anti-cytoskeletal drugs, especially the microtubule disruptor oryzalin. Our observations indicate that FH1 participates in the control of microtubule dynamics, possibly via its effects on actin, subsequently influencing cell morphogenesis and macroscopic organ development. © The Author 2016. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Hypothesis: NDL proteins function in stress responses by regulating microtubule organization.

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Khatri, Nisha; Mudgil, Yashwanti

2015-01-01

N-MYC DOWNREGULATED-LIKE proteins (NDL), members of the alpha/beta hydrolase superfamily were recently rediscovered as interactors of G-protein signaling in Arabidopsis thaliana. Although the precise molecular function of NDL proteins is still elusive, in animals these proteins play protective role in hypoxia and expression is induced by hypoxia and nickel, indicating role in stress. Homology of NDL1 with animal counterpart N-MYC DOWNREGULATED GENE (NDRG) suggests similar functions in animals and plants. It is well established that stress responses leads to the microtubule depolymerization and reorganization which is crucial for stress tolerance. NDRG is a microtubule-associated protein which mediates the microtubule organization in animals by causing acetylation and increases the stability of α-tubulin. As NDL1 is highly homologous to NDRG, involvement of NDL1 in the microtubule organization during plant stress can also be expected. Discovery of interaction of NDL with protein kinesin light chain- related 1, enodomembrane family protein 70, syntaxin-23, tubulin alpha-2 chain, as a part of G protein interactome initiative encourages us to postulate microtubule stabilizing functions for NDL family in plants. Our search for NDL interactors in G protein interactome also predicts the role of NDL proteins in abiotic stress tolerance management. Based on published report in animals and predicted interacting partners for NDL in G protein interactome lead us to hypothesize involvement of NDL in the microtubule organization during abiotic stress management in plants.

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

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Duncan, Jason E.; Lytle, Nikki K.; Zuniga, Alfredo; Goldstein, Lawrence S. B.

2013-01-01

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. PMID:23840848

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

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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.

Noscapine alters microtubule dynamics in living cells and inhibits the progression of melanoma.

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Landen, Jaren W; Lang, Roland; McMahon, Steve J; Rusan, Nasser M; Yvon, Anne-Marie; Adams, Ashley W; Sorcinelli, Mia D; Campbell, Ross; Bonaccorsi, Paola; Ansel, John C; Archer, David R; Wadsworth, Patricia; Armstrong, Cheryl A; Joshi, Harish C

2002-07-15

Cellular microtubules, polymers of tubulin, alternate relentlessly between phases of growth and shortening. We now show that noscapine, a tubulin-binding agent, increases the time that cellular microtubules spend idle in a paused state. As a result, most mammalian cell types observed arrest in mitosis in the presence of noscapine. We demonstrate that noscapine-treated murine melanoma B16LS9 cells do not arrest in mitosis but rather become polyploid followed by cell death, whereas primary melanocytes reversibly arrest in mitosis and resume a normal cell cycle after noscapine removal. Furthermore, in a syngeneic murine model of established s.c. melanoma, noscapine treatment resulted in an 85% inhibition of tumor volume on day 17 when delivered by gavage compared with untreated animals (P microtubule dynamics, with no detected toxicity to the host. Consequently, noscapine could be a valuable chemotherapeutic agent, alone or in combination, for the treatment of advanced melanoma.

Microtubule and Cell Contact Dependency of ER-bound PTP1B Localization in Growth Cones

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Fuentes, Federico

2009-01-01

PTP1B is an ER-bound protein tyrosine phosphatase implied in the regulation of cell adhesion. Here we investigated mechanisms involved in the positioning and dynamics of PTP1B in axonal growth cones and evaluated the role of this enzyme in axons. In growth cones, PTP1B consistently localizes in the central domain, and occasionally at the peripheral region and filopodia. Live imaging of GFP-PTP1B reveals dynamic excursions of fingerlike processes within the peripheral region and filopodia. PTP1B and GFP-PTP1B colocalize with ER markers and coalign with microtubules at the peripheral region and redistribute to the base of the growth cone after treatment with nocodazole, a condition that is reversible. Growth cone contact with cellular targets is accompanied by invasion of PTP1B and stable microtubules in the peripheral region aligned with the contact axis. Functional impairment of PTP1B causes retardation of axon elongation, as well as reduction of growth cone filopodia lifetime and Src activity. Our results highlight the role of microtubules and cell contacts in the positioning of ER-bound PTP1B to the peripheral region of growth cones, which may be required for the positive role of PTP1B in axon elongation, filopodia stabilization, and Src activity. PMID:19158394

Structural differences between yeast and mammalian microtubules revealed by cryo-EM

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Howes, Stuart C. [Univ. of California, Berkeley, CA (United States). Biophysics Graduate Group; Geyer, Elisabeth A. [Univ. of Texas Southwestern Medical Center, Dallas, TX (United States). Dept. of Biophysics; Univ. of Texas Southwestern Medical Center, Dallas, TX (United States). Dept. of Biochemistry; LaFrance, Benjamin [Univ. of California, Berkeley, CA (United States). Molecular and Cell Biology Graduate Program; Zhang, Rui [Univ. of California, Berkeley, CA (United States). Howard Hughes Medical Inst.; Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Molecular Biophysics and Integrated Bioimaging Division; Kellogg, Elizabeth H. [Univ. of California, Berkeley, CA (United States). Howard Hughes Medical Inst.; Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Molecular Biophysics and Integrated Bioimaging Division; Westermann, Stefan [Univ. of Duisburg-Essen, Essen (Germany). Dept. of Molecular Genetics, Center for Medical Biotechnology; Rice, Luke M. [Univ. of Texas Southwestern Medical Center, Dallas, TX (United States). Dept. of Biophysics; Univ. of Texas Southwestern Medical Center, Dallas, TX (United States). Dept. of Biochemistry; Nogales, Eva [Univ. of California, Berkeley, CA (United States). Howard Hughes Medical Inst.; Univ. of California, Berkeley, CA (United States). Dept. of Molecular Biology and California Inst. for Quantitative Biosciences; Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Molecular Biophysics and Integrated Bioimaging Division

2017-06-26

Microtubules are polymers of αβ-tubulin heterodimers essential for all eukaryotes. Despite sequence conservation, there are significant structural differences between microtubules assembled in vitro from mammalian or budding yeast tubulin. Yeast MTs were not observed to undergo compaction at the interdimer interface as seen for mammalian microtubules upon GTP hydrolysis. Lack of compaction might reflect slower GTP hydrolysis or a different degree of allosteric coupling in the lattice. The microtubule plus end–tracking protein Bim1 binds yeast microtubules both between αβ-tubulin heterodimers, as seen for other organisms, and within tubulin dimers, but binds mammalian tubulin only at interdimer contacts. At the concentrations used in cryo-electron microscopy, Bim1 causes the compaction of yeast microtubules and induces their rapid disassembly. In conclusion, our studies demonstrate structural differences between yeast and mammalian microtubules that likely underlie their differing polymerization dynamics. These differences may reflect adaptations to the demands of different cell size or range of physiological growth temperatures.

On the nature and shape of tubulin trails: implications on microtubule self-organization.

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Glade, Nicolas

2012-06-01

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 macroscopic level. In the present work, by simulating the diffusion of tubulin in microtubule solutions at the microscopic scale, we measure the shape and intensity of tubulin trails and discuss about the assumption of microtubule self-organization due to the production of chemical trails by disassembling microtubules. We show that the tubulin trails produced by individual microtubules or small microtubule arrays are very weak and not elongated even at very high reactive rates. Although the variations of concentration due to such trails are not significant compared to natural fluctuations of the concentration of tubuline in the chemical environment, the study shows that heterogeneities of biochemical composition can form due to microtubule disassembly. They could become significant when produced by numerous microtubule ends located in the same place. Their possible formation could play a role in certain conditions of reaction. In particular, it gives a mesoscopic

TgICMAP1 is a novel microtubule binding protein in Toxoplasma gondii.

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Aoife T Heaslip

Full Text Available The microtubule cytoskeleton provides essential structural support for all eukaryotic cells and can be assembled into various higher order structures that perform drastically different functions. Understanding how microtubule-containing assemblies are built in a spatially and temporally controlled manner is therefore fundamental to understanding cell physiology. Toxoplasma gondii, a protozoan parasite, contains at least five distinct tubulin-containing structures, the spindle pole, centrioles, cortical microtubules, the conoid, and the intra-conoid microtubules. How these five structurally and functionally distinct sets of tubulin containing structures are constructed and maintained in the same cell is an intriguing problem. Previously, we performed a proteomic analysis of the T. gondii apical complex, a cytoskeletal complex located at the apical end of the parasite that is composed of the conoid, three ring-like structures, and the two short intra-conoid microtubules. Here we report the characterization of one of the proteins identified in that analysis, TgICMAP1. We show that TgICMAP1 is a novel microtubule binding protein that can directly bind to microtubules in vitro and stabilizes microtubules when ectopically expressed in mammalian cells. Interestingly, in T. gondii, TgICMAP1 preferentially binds to the intra-conoid microtubules, providing us the first molecular tool to investigate the intra-conoid microtubule assembly process during daughter construction.

Interplay between microtubule bundling and sorting factors ensures acentriolar spindle stability during C. elegans oocyte meiosis.

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Timothy J Mullen

2017-09-01

Full Text Available In many species, oocyte meiosis is carried out in the absence of centrioles. As a result, microtubule organization, spindle assembly, and chromosome segregation proceed by unique mechanisms. Here, we report insights into the principles underlying this specialized form of cell division, through studies of C. elegans KLP-15 and KLP-16, two highly homologous members of the kinesin-14 family of minus-end-directed kinesins. These proteins localize to the acentriolar oocyte spindle and promote microtubule bundling during spindle assembly; following KLP-15/16 depletion, microtubule bundles form but then collapse into a disorganized array. Surprisingly, despite this defect we found that during anaphase, microtubules are able to reorganize into a bundled array that facilitates chromosome segregation. This phenotype therefore enabled us to identify factors promoting microtubule organization during anaphase, whose contributions are normally undetectable in wild-type worms; we found that SPD-1 (PRC1 bundles microtubules and KLP-18 (kinesin-12 likely sorts those bundles into a functional orientation capable of mediating chromosome segregation. Therefore, our studies have revealed an interplay between distinct mechanisms that together promote spindle formation and chromosome segregation in the absence of structural cues such as centrioles.

Expansion and Polarity Sorting in Microtubule-Dynein Bundles(WHAT IS LIFE? THE NEXT 100 YEARS OF YUKAWA'S DREAM)

OpenAIRE

Assaf, ZEMEL; Alex, MOGILNER; Department of Neurobiology, Physiology and Behavior, University of California; Department of Neurobiology, Physiology and Behavior, University of California

2008-01-01

Interactions of multiple molecular motors with dynamic polymers, such as actin and microtubules, form the basis for many processes in the cell cytoskeleton. One example is the active 'sorting' of microtubule bundles by dynein molecular motors into aster-like arrays of microtubules; in these bundles dynein motors cross-link and slide neighboring microtubules apart. A number of models have been suggested to quantify the active dynamics of cross-linked bundles of polar filaments. In the case of ...

The growth speed of microtubules with XMAP215-coated beads coupled to their ends is increased by tensile force

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Trushko, Anastasiya; Schäffer, Erik; Howard, Jonathon

2013-01-01

The generation of pulling and pushing forces is one of the important functions of microtubules, which are dynamic and polarized structures. The ends of dynamic microtubules are able to form relatively stable links to cellular structures, so that when a microtubule grows it can exert a pushing force and when it shrinks it can exert a pulling force. Microtubule growth and shrinkage are tightly regulated by microtubule-associated proteins (MAPs) that bind to microtubule ends. Given their localization, MAPs may be exposed to compressive and tensile forces. The effect of such forces on MAP function, however, is poorly understood. Here we show that beads coated with the microtubule polymerizing protein XMAP215, the Xenopus homolog of Dis1 and chTOG, are able to link stably to the plus ends of microtubules, even when the ends are growing or shrinking; at growing ends, the beads increase the polymerization rate. Using optical tweezers, we found that tensile force further increased the microtubule polymerization rate. These results show that physical forces can regulate the activity of MAPs. Furthermore, our results show that XMAP215 can be used as a handle to sense and mechanically manipulate the dynamics of the microtubule tip. PMID:23964126

Dynamic release of nuclear RanGTP triggers TPX2-dependent microtubule assembly during the apoptotic execution phase.

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Moss, David K; Wilde, Andrew; Lane, Jon D

2009-03-01

During apoptosis, the interphase microtubule network is dismantled then later replaced by a novel, non-centrosomal microtubule array. These microtubules assist in the peripheral redistribution of nuclear fragments in the apoptotic cell; however, the regulation of apoptotic microtubule assembly is not understood. Here, we demonstrate that microtubule assembly depends upon the release of nuclear RanGTP into the apoptotic cytoplasm because this process is blocked in apoptotic cells overexpressing dominant-negative GDP-locked Ran (T24N). Actin-myosin-II contractility provides the impetus for Ran release and, consequently, microtubule assembly is blocked in blebbistatin- and Y27632-treated apoptotic cells. Importantly, the spindle-assembly factor TPX2 (targeting protein for Xklp2), colocalises with apoptotic microtubules, and siRNA silencing of TPX2, but not of the microtubule motors Mklp1 and Kid, abrogates apoptotic microtubule assembly. These data provide a molecular explanation for the assembly of the apoptotic microtubule network, and suggest important similarities with the process of RanGTP- and TPX2-mediated mitotic spindle formation.

Interactive domains in the molecular chaperone human alphaB crystallin modulate microtubule assembly and disassembly.

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Joy G Ghosh

2007-06-01

Full Text Available Small heat shock proteins regulate microtubule assembly during cell proliferation and in response to stress through interactions that are poorly understood.Novel functions for five interactive sequences in the small heat shock protein and molecular chaperone, human alphaB crystallin, were investigated in the assembly/disassembly of microtubules and aggregation of tubulin using synthetic peptides and mutants of human alphaB crystallin.The interactive sequence (113FISREFHR(120 exposed on the surface of alphaB crystallin decreased microtubule assembly by approximately 45%. In contrast, the interactive sequences, (131LTITSSLSSDGV(142 and (156ERTIPITRE(164, corresponding to the beta8 strand and the C-terminal extension respectively, which are involved in complex formation, increased microtubule assembly by approximately 34-45%. The alphaB crystallin peptides, (113FISREFHR(120 and (156ERTIPITRE(164, inhibited microtubule disassembly by approximately 26-36%, and the peptides (113FISREFHR(120 and (131LTITSSLSSDGV(142 decreased the thermal aggregation of tubulin by approximately 42-44%. The (131LTITSSLSSDGV(142 and (156ERTIPITRE(164 peptides were more effective than the widely used anti-cancer drug, Paclitaxel, in modulating tubulinmicrotubule dynamics. Mutagenesis of these interactive sequences in wt human alphaB crystallin confirmed the effects of the alphaB crystallin peptides on microtubule assembly/disassembly and tubulin aggregation. The regulation of microtubule assembly by alphaB crystallin varied over a narrow range of concentrations. The assembly of microtubules was maximal at alphaB crystallin to tubulin molar ratios between 1:4 and 2:1, while molar ratios >2:1 inhibited microtubule assembly.Interactive sequences on the surface of human alphaB crystallin collectively modulate microtubule assembly through a dynamic subunit exchange mechanism that depends on the concentration and ratio of alphaB crystallin to tubulin. These are the first

EML proteins in microtubule regulation and human disease.

Science.gov (United States)

Fry, Andrew M; O'Regan, Laura; Montgomery, Jessica; Adib, Rozita; Bayliss, Richard

2016-10-15

The EMLs are a conserved family of microtubule-associated proteins (MAPs). The founding member was discovered in sea urchins as a 77-kDa polypeptide that co-purified with microtubules. This protein, termed EMAP for echinoderm MAP, was the major non-tubulin component present in purified microtubule preparations made from unfertilized sea urchin eggs [J. Cell Sci. (1993) 104: , 445-450; J. Cell Sci. (1987) 87: (Pt 1), 71-84]. Orthologues of EMAP were subsequently identified in other echinoderms, such as starfish and sand dollar, and then in more distant eukaryotes, including flies, worms and vertebrates, where the name of ELP or EML (both for EMAP-like protein) has been adopted [BMC Dev. Biol. (2008) 8: , 110; Dev. Genes Evol. (2000) 210: , 2-10]. The common property of these proteins is their ability to decorate microtubules. However, whether they are associated with particular microtubule populations or exercise specific functions in different microtubule-dependent processes remains unknown. Furthermore, although there is limited evidence that they regulate microtubule dynamics, the biochemical mechanisms of their molecular activity have yet to be explored. Nevertheless, interest in these proteins has grown substantially because of the identification of EML mutations in neuronal disorders and oncogenic fusions in human cancers. Here, we summarize our current knowledge of the expression, localization and structure of what is proving to be an interesting and important class of MAPs. We also speculate about their function in microtubule regulation and highlight how the studies of EMLs in human diseases may open up novel avenues for patient therapy. © 2016 The Author(s); published by Portland Press Limited on behalf of the Biochemical Society.

Proteomics of cancer cell lines resistant to microtubule-stabilizing agents

DEFF Research Database (Denmark)

Albrethsen, Jakob; Angeletti, Ruth H; Horwitz, Susan Band

2014-01-01

Despite the clinical success of microtubule-interacting agents (MIA), a significant challenge for oncologists is the inability to predict the response of individual patients with cancer to these drugs. In the present study, six cell lines were compared by 2D DIGE proteomics to investigate cellula...

The dynamic interplay of plasma membrane domains and cortical microtubules in secondary cell wall patterning

Directory of Open Access Journals (Sweden)

Yoshihisa eOda

2013-12-01

Full Text Available Patterning of the cellulosic cell wall underlies the shape and function of plant cells. The cortical microtubule array plays a central role in the regulation of cell wall patterns. However, the regulatory mechanisms by which secondary cell wall patterns are established through cortical microtubules remain to be fully determined. Our recent study in xylem vessel cells revealed that a mutual inhibitory interaction between cortical microtubules and distinct plasma membrane domains leads to distinctive patterning in secondary cell walls. Our research revealed that the recycling of active and inactive ROP proteins by a specific GAP and GEF pair establishes distinct de novo plasma membrane domains. Active ROP recruits a plant-specific microtubule-associated protein, MIDD1, which mediates the mutual interaction between cortical microtubules and plasma membrane domains. In this mini review, we summarize recent research regarding secondary wall patterning, with a focus on the emerging interplay between plasma membrane domains and cortical microtubules through MIDD1 and ROP.

Katanin: A Sword Cutting Microtubules for Cellular, Developmental, and Physiological Purposes

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Ivan Luptovčiak

2017-11-01

Full Text Available KATANIN is a well-studied microtubule severing protein affecting microtubule organization and dynamic properties in higher plants. By regulating mitotic and cytokinetic and cortical microtubule arrays it is involved in the progression of cell division and cell division plane orientation. KATANIN is also involved in cell elongation and morphogenesis during plant growth. In this way KATANIN plays critical roles in diverse plant developmental processes including the development of pollen, embryo, seed, meristem, root, hypocotyl, cotyledon, leaf, shoot, and silique. KATANIN-dependent microtubule regulation seems to be under the control of plant hormones. This minireview provides an overview on available KATANIN mutants and discusses advances in our understanding of KATANIN biological roles in plants.

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.

Tubulin cofactor B regulates microtubule densities during microglia transition to the reactive states

International Nuclear Information System (INIS)

Fanarraga, M.L.; Villegas, J.C.; Carranza, G.; Castano, R.; Zabala, J.C.

2009-01-01

Microglia are highly dynamic cells of the CNS that continuously survey the welfare of the neural parenchyma and play key roles modulating neurogenesis and neuronal cell death. In response to injury or pathogen invasion parenchymal microglia transforms into a more active cell that proliferates, migrates and behaves as a macrophage. The acquisition of these extra skills implicates enormous modifications of the microtubule and actin cytoskeletons. Here we show that tubulin cofactor B (TBCB), which has been found to contribute to various aspects of microtubule dynamics in vivo, is also implicated in microglial cytoskeletal changes. We find that TBCB is upregulated in post-lesion reactive parenchymal microglia/macrophages, in interferon treated BV-2 microglial cells, and in neonate amoeboid microglia where the microtubule densities are remarkably low. Our data demonstrate that upon TBCB downregulation both, after microglia differentiation to the ramified phenotype in vivo and in vitro, or after TBCB gene silencing, microtubule densities are restored in these cells. Taken together these observations support the view that TBCB functions as a microtubule density regulator in microglia during activation, and provide an insight into the understanding of the complex mechanisms controlling microtubule reorganization during microglial transition between the amoeboid, ramified, and reactive phenotypes

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.

Influence of carbon nanotubes on the buckling of microtubule bundles in viscoelastic cytoplasm using nonlocal strain gradient theory

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A. Farajpour

Full Text Available Carbon nanotubes are a new class of microtubule-stabilizing agents since they interact with protein microtubules in living cells, interfering with cell division and inducing apoptosis. In the present work, a modified beam model is developed to investigate the effect of carbon nanotubes on the buckling of microtubule bundles in living cell. A realistic interaction model is employed using recent experimental data on the carbon nanotube-stabilized microtubules. Small scale and surface effects are taken into account applying the nonlocal strain gradient theory and surface elasticity theory. Pasternak model is used to describe the normal and shearing effects of enclosing filament matrix on the buckling behavior of the system. An exact solution is obtained for the buckling growth rates of the mixed bundle in viscoelastic surrounding cytoplasm. The present results are compared with those reported in the open literature for single microtubules and an excellent agreement is found. Finally, the effects of different parameters such as the size, chirality, position and surface energy of carbon nanotubes on the buckling growth rates of microtubule bundles are studied. It is found that the buckling growth rate may increase or decrease by adding carbon nanotubes, depending on the diameter and chirality of carbon nanotubes. Keywords: Microtubules, Carbon nanotubes, Buckling, Size effects

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 i...... to analyze the self-assembly of microtubules from tubulin are general, and many other reactions and processes may be studied as inverse problems with these methods when enough experimental data are available....

Mto2 multisite phosphorylation inactivates non-spindle microtubule nucleation complexes during mitosis

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Borek, Weronika E.; Groocock, Lynda M.; Samejima, Itaru; Zou, Juan; de Lima Alves, Flavia; Rappsilber, Juri; Sawin, Kenneth E.

2015-01-01

Microtubule nucleation is highly regulated during the eukaryotic cell cycle, but the underlying molecular mechanisms are largely unknown. During mitosis in fission yeast Schizosaccharomyces pombe, cytoplasmic microtubule nucleation ceases simultaneously with intranuclear mitotic spindle assembly. Cytoplasmic nucleation depends on the Mto1/2 complex, which binds and activates the γ-tubulin complex and also recruits the γ-tubulin complex to both centrosomal (spindle pole body) and non-centrosomal sites. Here we show that the Mto1/2 complex disassembles during mitosis, coincident with hyperphosphorylation of Mto2 protein. By mapping and mutating multiple Mto2 phosphorylation sites, we generate mto2-phosphomutant strains with enhanced Mto1/2 complex stability, interaction with the γ-tubulin complex and microtubule nucleation activity. A mutant with 24 phosphorylation sites mutated to alanine, mto2[24A], retains interphase-like behaviour even in mitotic cells. This provides a molecular-level understanding of how phosphorylation ‘switches off' microtubule nucleation complexes during the cell cycle and, more broadly, illuminates mechanisms regulating non-centrosomal microtubule nucleation. PMID:26243668

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

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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.

Centriole triplet microtubules are required for stable centriole formation and inheritance in human cells.

Science.gov (United States)

Wang, Jennifer T; Kong, Dong; Hoerner, Christian R; Loncarek, Jadranka; Stearns, Tim

2017-09-14

Centrioles are composed of long-lived microtubules arranged in nine triplets. However, the contribution of triplet microtubules to mammalian centriole formation and stability is unknown. Little is known of the mechanism of triplet microtubule formation, but experiments in unicellular eukaryotes indicate that delta-tubulin and epsilon-tubulin, two less-studied tubulin family members, are required. Here, we report that centrioles in delta-tubulin and epsilon-tubulin null mutant human cells lack triplet microtubules and fail to undergo centriole maturation. These aberrant centrioles are formed de novo each cell cycle, but are unstable and do not persist to the next cell cycle, leading to a futile cycle of centriole formation and disintegration. Disintegration can be suppressed by paclitaxel treatment. Delta-tubulin and epsilon-tubulin physically interact, indicating that these tubulins act together to maintain triplet microtubules and that these are necessary for inheritance of centrioles from one cell cycle to the next.

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....

Taking directions: the role of microtubule-bound nucleation in the self-organization of the plant cortical array

International Nuclear Information System (INIS)

Deinum, Eva E; Tindemans, Simon H; Mulder, Bela M

2011-01-01

The highly aligned cortical microtubule array of interphase plant cells is a key regulator of anisotropic cell expansion. Recent computational and analytical work has shown that the non-equilibrium self-organization of this structure can be understood on the basis of experimentally observed collisional interactions between dynamic microtubules attached to the plasma membrane. Most of these approaches assumed that new microtubules are homogeneously and isotropically nucleated on the cortical surface. Experimental evidence, however, shows that nucleation mostly occurs from other microtubules and under specific relative angles. Here, we investigate the impact of directed microtubule-bound nucleations on the alignment process using computer simulations. The results show that microtubule-bound nucleations can increase the degree of alignment achieved, decrease the timescale of the ordering process and widen the regime of dynamic parameters for which the system can self-organize. We establish that the major determinant of this effect is the degree of co-alignment of the nucleations with the parent microtubule. The specific role of sideways branching nucleations appears to allow stronger alignment while maintaining a measure of overall spatial homogeneity. Finally, we investigate the suggestion that observed persistent rotation of microtubule domains can be explained through a handedness bias in microtubule-bound nucleations, showing that this is possible only for an extreme bias and over a limited range of parameters

Taking directions: the role of microtubule-bound nucleation in the self-organization of the plant cortical array

Science.gov (United States)

Deinum, Eva E.; Tindemans, Simon H.; Mulder, Bela M.

2011-10-01

The highly aligned cortical microtubule array of interphase plant cells is a key regulator of anisotropic cell expansion. Recent computational and analytical work has shown that the non-equilibrium self-organization of this structure can be understood on the basis of experimentally observed collisional interactions between dynamic microtubules attached to the plasma membrane. Most of these approaches assumed that new microtubules are homogeneously and isotropically nucleated on the cortical surface. Experimental evidence, however, shows that nucleation mostly occurs from other microtubules and under specific relative angles. Here, we investigate the impact of directed microtubule-bound nucleations on the alignment process using computer simulations. The results show that microtubule-bound nucleations can increase the degree of alignment achieved, decrease the timescale of the ordering process and widen the regime of dynamic parameters for which the system can self-organize. We establish that the major determinant of this effect is the degree of co-alignment of the nucleations with the parent microtubule. The specific role of sideways branching nucleations appears to allow stronger alignment while maintaining a measure of overall spatial homogeneity. Finally, we investigate the suggestion that observed persistent rotation of microtubule domains can be explained through a handedness bias in microtubule-bound nucleations, showing that this is possible only for an extreme bias and over a limited range of parameters.

Direct evidence for GTP and GDP-Pi intermediates in microtubule assembly

International Nuclear Information System (INIS)

Melki, R.; Carlier, M.F.; Pantaloni, D.

1990-01-01

Identification of the kinetic intermediates in GTP hydrolysis on microtubules and characterization of their assembly properties is essential in understanding microtubule dynamics. By using an improved glass filter assay that selectively traps microtubules with a dead time of 2 s and monitoring taxol-induced rapid assembly of microtubules from [γ- 32 P, 3 H]GTP-tubulin 1:1 complex, direct evidence has been obtained for GTP- and GDP-P i -microtubule transient states in the early stages of the polymerization process. A simple kinetic analysis of GTP hydrolysis on microtubules within two sequential pseudo-first-order processes led to apparent first-order rate constants of 0.065 s -1 for the cleavage of the γ-phosphate and 0.02 s -1 for the liberation of P i , assuming a simple random model. Apparent rate constants for GTP hydrolysis and P i release were independent of the composition of the buffer used to polymerize tubulin. The significance of these values with respect to those derived from previous studies from this and other laboratories and the possibility of a vectorial model for GTP hydrolysis are discussed

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.

Synthesis and biological evaluation of structurally simplified noscapine analogues as microtubule binding agents

Czech Academy of Sciences Publication Activity Database

Ghaly, P.E.; Churchill, C.D.M.; Abou El-Magd, R.M.; Hájková, Zuzana; Dráber, Pavel; West, F.G.; Tuszyński, J.A.

2017-01-01

Roč. 95, č. 6 (2017), s. 649-655 ISSN 0008-4042 R&D Projects: GA ČR GA15-22194S Institutional support: RVO:68378050 Keywords : noscapine * microtubule * tubulin * cytotoxicity * microtubule dynamics * docking Subject RIV: EB - Genetics ; Molecular Biology OBOR OECD: Biochemistry and molecular biology Impact factor: 1.080, year: 2016

Centriole triplet microtubules are required for stable centriole formation and inheritance in human cells

Science.gov (United States)

Wang, Jennifer T; Kong, Dong; Hoerner, Christian R; Loncarek, Jadranka

2017-01-01

Centrioles are composed of long-lived microtubules arranged in nine triplets. However, the contribution of triplet microtubules to mammalian centriole formation and stability is unknown. Little is known of the mechanism of triplet microtubule formation, but experiments in unicellular eukaryotes indicate that delta-tubulin and epsilon-tubulin, two less-studied tubulin family members, are required. Here, we report that centrioles in delta-tubulin and epsilon-tubulin null mutant human cells lack triplet microtubules and fail to undergo centriole maturation. These aberrant centrioles are formed de novo each cell cycle, but are unstable and do not persist to the next cell cycle, leading to a futile cycle of centriole formation and disintegration. Disintegration can be suppressed by paclitaxel treatment. Delta-tubulin and epsilon-tubulin physically interact, indicating that these tubulins act together to maintain triplet microtubules and that these are necessary for inheritance of centrioles from one cell cycle to the next. PMID:28906251

Microtubule Regulation of Kv7 Channels Orchestrates cAMP-Mediated Vasorelaxations in Rat Arterial Smooth Muscle

DEFF Research Database (Denmark)

Lindman, Johanna; Khammy, Makhala M; Lundegaard, Pia R

2018-01-01

Microtubules can regulate GPCR (G protein-coupled receptor) signaling in various cell types. In vascular smooth muscle, activation of the β-adrenoceptor leads to production of cAMP to mediate a vasorelaxation. Little is known about the role of microtubules in smooth muscle, and given the importance...... of renal and mesenteric arteries that the microtubule stabilizer, paclitaxel, prevented. Sharp microelectrode experiments showed that colchicine treatment caused increased hyperpolarization of mesenteric artery segments in response to isoprenaline. Application of the Kv7 channel blocker, XE991, attenuated...

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.

Microtubule dynamics of the centrosome-like polar organizers from the basal land plant Marchantia polymorpha.

Science.gov (United States)

Buschmann, Henrik; Holtmannspötter, Michael; Borchers, Agnes; O'Donoghue, Martin-Timothy; Zachgo, Sabine

2016-02-01

The liverwort Marchantia employs both modern and ancestral devices during cell division: it forms preprophase bands and in addition it shows centrosome-like polar organizers. We investigated whether polar organizers and preprophase bands cooperate to set up the division plane. To this end, two novel green fluorescent protein-based microtubule markers for dividing cells of Marchantia were developed. Cells of the apical notch formed polar organizers first and subsequently assembled preprophase bands. Polar organizers were formed de novo from multiple mobile microtubule foci localizing to the nuclear envelope. The foci then became concentrated by bipolar aggregation. We determined the comet production rate of polar organizers and show that microtubule plus ends of astral microtubules polymerize faster than those found on cortical microtubules. Importantly, it was observed that conditions increasing polar organizer numbers interfere with preprophase band formation. The data show that polar organizers have much in common with centrosomes, but that they also have specialized features. The results suggest that polar organizers contribute to preprophase band formation and in this way are involved in controlling the division plane. Our analyses of the basal land plant Marchantia shed new light on the evolution of plant cell division. © 2015 The Authors. New Phytologist © 2015 New Phytologist Trust.

Microtubule nucleation and organization in dendrites

Science.gov (United States)

Delandre, Caroline; Amikura, Reiko; Moore, Adrian W.

2016-01-01

ABSTRACT Dendrite branching is an essential process for building complex nervous systems. It determines the number, distribution and integration of inputs into a neuron, and is regulated to create the diverse dendrite arbor branching patterns characteristic of different neuron types. The microtubule cytoskeleton is critical to provide structure and exert force during dendrite branching. It also supports the functional requirements of dendrites, reflected by differential microtubule architectural organization between neuron types, illustrated here for sensory neurons. Both anterograde and retrograde microtubule polymerization occur within growing dendrites, and recent studies indicate that branching is enhanced by anterograde microtubule polymerization events in nascent branches. The polarities of microtubule polymerization events are regulated by the position and orientation of microtubule nucleation events in the dendrite arbor. Golgi outposts are a primary microtubule nucleation center in dendrites and share common nucleation machinery with the centrosome. In addition, pre-existing dendrite microtubules may act as nucleation sites. We discuss how balancing the activities of distinct nucleation machineries within the growing dendrite can alter microtubule polymerization polarity and dendrite branching, and how regulating this balance can generate neuron type-specific morphologies. PMID:27097122

Manipulation and quantification of microtubule lattice integrity

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Taylor A. Reid

2017-08-01

Full Text Available Microtubules are structural polymers that participate in a wide range of cellular functions. The addition and loss of tubulin subunits allows the microtubule to grow and shorten, as well as to develop and repair defects and gaps in its cylindrical lattice. These lattice defects act to modulate the interactions of microtubules with molecular motors and other microtubule-associated proteins. Therefore, tools to control and measure microtubule lattice structure will be invaluable for developing a quantitative understanding of how the structural state of the microtubule lattice may regulate its interactions with other proteins. In this work, we manipulated the lattice integrity of in vitro microtubules to create pools of microtubules with common nucleotide states, but with variations in structural states. We then developed a series of novel semi-automated analysis tools for both fluorescence and electron microscopy experiments to quantify the type and severity of alterations in microtubule lattice integrity. These techniques will enable new investigations that explore the role of microtubule lattice structure in interactions with microtubule-associated proteins.

Wood cell-wall structure requires local 2D-microtubule disassembly by a novel plasma membrane-anchored protein.

Science.gov (United States)

Oda, Yoshihisa; Iida, Yuki; Kondo, Yuki; Fukuda, Hiroo

2010-07-13

Plant cells have evolved cortical microtubules, in a two-dimensional space beneath the plasma membrane, that regulate patterning of cellulose deposition. Although recent studies have revealed that several microtubule-associated proteins facilitate self-organization of transverse cortical microtubules, it is still unknown how diverse patterns of cortical microtubules are organized in different xylem cells, which are the major components of wood. Using our newly established in vitro xylem cell differentiation system, we found that a novel microtubule end-tracking protein, microtubule depletion domain 1 (MIDD1), was anchored to distinct plasma membrane domains and promoted local microtubule disassembly, resulting in pits on xylem cell walls. The introduction of RNA interference for MIDD1 resulted in the failure of local microtubule depletion and the formation of secondary walls without pits. Conversely, the overexpression of MIDD1 reduced microtubule density. MIDD1 has two coiled-coil domains for the binding to microtubules and for the anchorage to plasma membrane domains, respectively. Combination of the two coils caused end tracking of microtubules during shrinkage and suppressed their rescue events. Our results indicate that MIDD1 integrates spatial information in the plasma membrane with cortical microtubule dynamics for determining xylem cell wall pattern. Copyright 2010 Elsevier Ltd. All rights reserved.

Quantitative cell polarity imaging defines leader-to-follower transitions during collective migration and the key role of microtubule-dependent adherens junction formation.

Science.gov (United States)

Revenu, Céline; Streichan, Sebastian; Donà, Erika; Lecaudey, Virginie; Hufnagel, Lars; Gilmour, Darren

2014-03-01

The directed migration of cell collectives drives the formation of complex organ systems. A characteristic feature of many migrating collectives is a 'tissue-scale' polarity, whereby 'leader' cells at the edge of the tissue guide trailing 'followers' that become assembled into polarised epithelial tissues en route. Here, we combine quantitative imaging and perturbation approaches to investigate epithelial cell state transitions during collective migration and organogenesis, using the zebrafish lateral line primordium as an in vivo model. A readout of three-dimensional cell polarity, based on centrosomal-nucleus axes, allows the transition from migrating leaders to assembled followers to be quantitatively resolved for the first time in vivo. Using live reporters and a novel fluorescent protein timer approach, we investigate changes in cell-cell adhesion underlying this transition by monitoring cadherin receptor localisation and stability. This reveals that while cadherin 2 is expressed across the entire tissue, functional apical junctions are first assembled in the transition zone and become progressively more stable across the leader-follower axis of the tissue. Perturbation experiments demonstrate that the formation of these apical adherens junctions requires dynamic microtubules. However, once stabilised, adherens junction maintenance is microtubule independent. Combined, these data identify a mechanism for regulating leader-to-follower transitions within migrating collectives, based on the relocation and stabilisation of cadherins, and reveal a key role for dynamic microtubules in this process.

Microtubules as mechanical force sensors.

Science.gov (United States)

Karafyllidis, Ioannis G; Lagoudas, Dimitris C

2007-03-01

Microtubules are polymers of tubulin subunits (dimers) arranged on a hexagonal lattice. Each tubulin dimer comprises two monomers, the alpha-tubulin and beta-tubulin, and can be found in two states. In the first state a mobile negative charge is located into the alpha-tubulin monomer and in the second into the beta-tubulin monomer. Each tubulin dimer is modeled as an electrical dipole coupled to its neighbors by electrostatic forces. The location of the mobile charge in each dimer depends on the location of the charges in the dimer's neighborhood. Mechanical forces that act on the microtubule affect the distances between the dimers and alter the electrostatic potential. Changes in this potential affect the mobile negative charge location in each dimer and the charge distribution in the microtubule. The net effect is that mechanical forces affect the charge distribution in microtubules. We propose to exploit this effect and use microtubules as mechanical force sensors. We model each dimer as a two-state quantum system and, following the quantum computation paradigm, we use discrete quantum random walk on the hexagonal microtubule lattice to determine the charge distribution. Different forces applied on the microtubule are modeled as different coin biases leading to different probability distributions of the quantum walker location, which are directly connected to different charge distributions. Simulation results show that there is a strong indication that microtubules can be used as mechanical force sensors and that they can also detect the force directions and magnitudes.

Microtubule array reorientation in response to hormones does not involve changes in microtubule nucleation modes at the periclinal cell surface

Science.gov (United States)

Atkinson, Samantha; Kirik, Angela; Kirik, Viktor

2014-01-01

Aligned microtubule arrays spatially organize cell division, trafficking, and determine the direction of cell expansion in plant cells. In response to changes in environmental and developmental signals, cells reorganize their microtubule arrays into new configurations. Here, we tested the role of microtubule nucleation during hormone-induced microtubule array reorientation. We have found that in the process of microtubule array reorientation the ratios between branching, parallel, and de-novo nucleations remained constant, suggesting that the microtubule reorientation mechanism does not involve changes in nucleation modes. In the ton2/fass mutant, which has reduced microtubule branching nucleation frequency and decreased nucleation activity of the γ-tubulin complexes, microtubule arrays were able to reorient. Presented data suggest that reorientation of microtubules into transverse arrays in response to hormones does not involve changes in microtubule nucleation at the periclinal cell surface PMID:25135522

Identification and characterization of SSE15206, a microtubule depolymerizing agent that overcomes multidrug resistance

KAUST Repository

Manzoor, Safia

2018-02-13

Microtubules are highly dynamic structures that form spindle fibres during mitosis and are one of the most validated cancer targets. The success of drugs targeting microtubules, however, is often limited by the development of multidrug resistance. Here we describe the discovery and characterization of SSE15206, a pyrazolinethioamide derivative [3-phenyl-5-(3,4,5-trimethoxyphenyl)-4,5-dihydro-1H-pyrazole-1-carbothioamide] that has potent antiproliferative activities in cancer cell lines of different origins and overcomes resistance to microtubule-targeting agents. Treatment of cells with SSE15206 causes aberrant mitosis resulting in G2/M arrest due to incomplete spindle formation, a phenotype often associated with drugs that interfere with microtubule dynamics. SSE15206 inhibits microtubule polymerization both in biochemical and cellular assays by binding to colchicine site in tubulin as shown by docking and competition studies. Prolonged treatment of cells with the compound results in apoptotic cell death [increased Poly (ADP-ribose) polymerase cleavage and Annexin V/PI staining] accompanied by p53 induction. More importantly, we demonstrate that SSE15206 is able to overcome resistance to chemotherapeutic drugs in different cancer cell lines including multidrug-resistant KB-V1 and A2780-Pac-Res cell lines overexpressing MDR-1, making it a promising hit for the lead optimization studies to target multidrug resistance.

Drosophila homologue of Diaphanous 1 (DIAPH1) controls the metastatic potential of colon cancer cells by regulating microtubule-dependent adhesion.

Science.gov (United States)

Lin, Yuan-Na; Bhuwania, Ridhirama; Gromova, Kira; Failla, Antonio Virgilio; Lange, Tobias; Riecken, Kristoffer; Linder, Stefan; Kneussel, Matthias; Izbicki, Jakob R; Windhorst, Sabine

2015-07-30

Drosophila homologue of Diaphanous 1 (DIAPH1) regulates actin polymerization and microtubule (MT) stabilization upon stimulation with lysophosphatidic acid (LPA). Recently, we showed strongly reduced lung metastasis of DIAPH1-depleted colon cancer cells but we found accumulations of DIAPH1-depleted cells in bone marrow. Here, we analyzed possible organ- or tissue-specific metastasis of DIAPH1-depleted HCT-116 cells. Our data confirmed that depletion of DIAPH1 strongly inhibited lung metastasis and revealed that, in contrast to control cells, DIAPH1-depleted cells did not form metastases in further organs. Detailed mechanistic analysis on cells that were not stimulated with LPA to activate the cytoskeleton-modulating activity of DIAPH1, revealed that even under basal conditions DIAPH1 was essential for cellular adhesion to collagen. In non-stimulated cells DIAPH1 did not control actin dynamics but, interestingly, was essential for stabilization of microtubules (MTs). Additionally, DIAPH1 controlled directed vesicle trafficking and with this, local clustering of the adhesion protein integrin-β1 at the plasma membrane. Therefore, we conclude that under non-stimulating conditions DIAPH1 controls cellular adhesion by stabilizing MTs required for local clustering of integrin-β1 at the plasma membrane. Thus, blockade of DIAPH1-tubulin interaction may be a promising approach to inhibit one of the earliest steps in the metastatic cascade of colon cancer.

Modulation of microtubule assembly by the HIV-1 Tat protein is strongly dependent on zinc binding to Tat

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Muller Sylviane

2008-07-01

Full Text Available Abstract Background During HIV-1 infection, the Tat protein plays a key role by transactivating the transcription of the HIV-1 proviral DNA. In addition, Tat induces apoptosis of non-infected T lymphocytes, leading to a massive loss of immune competence. This apoptosis is notably mediated by the interaction of Tat with microtubules, which are dynamic components essential for cell structure and division. Tat binds two Zn2+ ions through its conserved cysteine-rich region in vitro, but the role of zinc in the structure and properties of Tat is still controversial. Results To investigate the role of zinc, we first characterized Tat apo- and holo-forms by fluorescence correlation spectroscopy and time-resolved fluorescence spectroscopy. Both of the Tat forms are monomeric and poorly folded but differ by local conformational changes in the vicinity of the cysteine-rich region. The interaction of the two Tat forms with tubulin dimers and microtubules was monitored by analytical ultracentrifugation, turbidity measurements and electron microscopy. At 20°C, both of the Tat forms bind tubulin dimers, but only the holo-Tat was found to form discrete complexes. At 37°C, both forms promoted the nucleation and increased the elongation rates of tubulin assembly. However, only the holo-Tat increased the amount of microtubules, decreased the tubulin critical concentration, and stabilized the microtubules. In contrast, apo-Tat induced a large amount of tubulin aggregates. Conclusion Our data suggest that holo-Tat corresponds to the active form, responsible for the Tat-mediated apoptosis.

Stability of dynamical systems

CERN Document Server

Liao, Xiaoxin; Yu, P 0

2007-01-01

The main purpose of developing stability theory is to examine dynamic responses of a system to disturbances as the time approaches infinity. It has been and still is the object of intense investigations due to its intrinsic interest and its relevance to all practical systems in engineering, finance, natural science and social science. This monograph provides some state-of-the-art expositions of major advances in fundamental stability theories and methods for dynamic systems of ODE and DDE types and in limit cycle, normal form and Hopf bifurcation control of nonlinear dynamic systems.ʺ Presents

Synchronous Oscillations in Microtubule Polymerization

Science.gov (United States)

Carlier, M. F.; Melki, R.; Pantaloni, D.; Hill, T. L.; Chen, Y.

1987-08-01

Under conditions where microtubule nucleation and growth are fast (i.e., high magnesium ion and tubulin concentrations and absence of glycerol), microtubule assembly in vitro exhibits an oscillatory regime preceding the establishment of steady state. The amplitude of the oscillations can represent >50% of the maximum turbidity change and oscillations persist for up to 20 periods of 80 s each. Oscillations are accompanied by extensive length redistribution of microtubules. Preliminary work suggests that the oscillatory kinetics can be simulated using a model in which many microtubules undergo synchronous transitions between growing and rapidly depolymerizing phases, complicated by the kinetically limiting rate of nucleotide exchange on free tubulin.

Kindlin1 regulates microtubule function to ensure normal mitosis.

Science.gov (United States)

Patel, Hitesh; Stavrou, Ifigeneia; Shrestha, Roshan L; Draviam, Viji; Frame, Margaret C; Brunton, Valerie G

2016-08-01

Loss of Kindlin 1 (Kin1) results in the skin blistering disorder Kindler Syndrome (KS), whose symptoms also include skin atrophy and reduced keratinocyte proliferation. Kin1 binds to integrins to modulate their activation and more recently it has been shown to regulate mitotic spindles and cell survival in a Plk1-dependent manner. Here we report that short-term Kin1 deletion in mouse skin results in impaired mitosis, which is associated with reduced acetylated tubulin (ac-tub) levels and cell proliferation. In cells, impaired mitosis and reduced ac-tub levels are also accompanied by reduced microtubule stability, all of which are rescued by HDAC6 inhibition. The ability of Kin1 to regulate HDAC6-dependent cellular ac-tub levels is dependent on its phosphorylation by Plk1. Taken together, these data define a novel role for Kin1 in microtubule acetylation and stability and offer a mechanistic insight into how certain KS phenotypes, such as skin atrophy and reduced cell proliferation, arise. © The Author (2016). Published by Oxford University Press on behalf of Journal of Molecular Cell Biology, IBCB, SIBS, CAS.

Near-atomic model of microtubule-tau interactions.

Science.gov (United States)

Kellogg, Elizabeth H; Hejab, Nisreen M A; Poepsel, Simon; Downing, Kenneth H; DiMaio, Frank; Nogales, Eva

2018-06-15

Tau is a developmentally regulated axonal protein that stabilizes and bundles microtubules (MTs). Its hyperphosphorylation is thought to cause detachment from MTs and subsequent aggregation into fibrils implicated in Alzheimer's disease. It is unclear which tau residues are crucial for tau-MT interactions, where tau binds on MTs, and how it stabilizes them. We used cryo-electron microscopy to visualize different tau constructs on MTs and computational approaches to generate atomic models of tau-tubulin interactions. The conserved tubulin-binding repeats within tau adopt similar extended structures along the crest of the protofilament, stabilizing the interface between tubulin dimers. Our structures explain the effect of phosphorylation on MT affinity and lead to a model of tau repeats binding in tandem along protofilaments, tethering together tubulin dimers and stabilizing polymerization interfaces. Copyright © 2018 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

TONNEAU2/FASS Regulates the Geometry of Microtubule Nucleation and Cortical Array Organization in Interphase Arabidopsis Cells[C][W

Science.gov (United States)

Kirik, Angela; Ehrhardt, David W.; Kirik, Viktor

2012-01-01

Organization of microtubules into ordered arrays involves spatial and temporal regulation of microtubule nucleation. Here, we show that acentrosomal microtubule nucleation in plant cells involves a previously unknown regulatory step that determines the geometry of microtubule nucleation. Dynamic imaging of interphase cortical microtubules revealed that the ratio of branching to in-bundle microtubule nucleation on cortical microtubules is regulated by the Arabidopsis thaliana B′′ subunit of protein phosphatase 2A, which is encoded by the TONNEAU2/FASS (TON2) gene. The probability of nucleation from γ-tubulin complexes localized at the cell cortex was not affected by a loss of TON2 function, suggesting a specific role of TON2 in regulating the nucleation geometry. Both loss of TON2 function and ectopic targeting of TON2 to the plasma membrane resulted in defects in cell shape, suggesting the importance of TON2-mediated regulation of the microtubule cytoskeleton in cell morphogenesis. Loss of TON2 function also resulted in an inability for cortical arrays to reorient in response to light stimulus, suggesting an essential role for TON2 and microtubule branching nucleation in reorganization of microtubule arrays. Our data establish TON2 as a regulator of interphase microtubule nucleation and provide experimental evidence for a novel regulatory step in the process of microtubule-dependent nucleation. PMID:22395485

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. Copyright © 2015 Elsevier Ltd. All rights reserved.

A novel mechanism important for the alignment of microtubules.

Science.gov (United States)

Wightman, Raymond; Turner, Simon R

2008-04-01

Using a live-cell imaging approach to study individual micro-tubules, we have compared microtubule behavior between net-like and aligned cortical arrays. In contrast to previous studies, a steep angled collision between the growing end of a microtubule and a preexisting microtubule was found to favor crossover. Frequencies of microtubule crossovers, bundling and catastrophes are similar regardless of whether the cell exhibited a net-like or aligned microtubule array. In the predominantly aligned array of petiole cells, severing occurs at the sites of microtubule crossovers and serves to remove unaligned microtubules and to increase microtubule density. Severing was observed to be rare in net-like arrays. Microtubule severing is carried out by the katanin enzyme. In this addendum, we present new insights into the possible mechanism of crossing over and preliminary data looking at organization of the array in a katanin mutant.

Dynamic microtubule organization and mitochondrial transport are regulated by distinct Kinesin-1 pathways

Directory of Open Access Journals (Sweden)

Anna Melkov

2015-12-01

Full Text Available The microtubule (MT plus-end motor kinesin heavy chain (Khc is well known for its role in long distance cargo transport. Recent evidence showed that Khc is also required for the organization of the cellular MT network by mediating MT sliding. We found that mutations in Khc and the gene of its adaptor protein, kinesin light chain (Klc resulted in identical bristle morphology defects, with the upper part of the bristle being thinner and flatter than normal and failing to taper towards the bristle tip. We demonstrate that bristle mitochondria transport requires Khc but not Klc as a competing force to dynein heavy chain (Dhc. Surprisingly, we demonstrate for the first time that Dhc is the primary motor for both anterograde and retrograde fast mitochondria transport. We found that the upper part of Khc and Klc mutant bristles lacked stable MTs. When following dynamic MT polymerization via the use of GFP-tagged end-binding protein 1 (EB1, it was noted that at Khc and Klc mutant bristle tips, dynamic MTs significantly deviated from the bristle parallel growth axis, relative to wild-type bristles. We also observed that GFP-EB1 failed to concentrate as a focus at the tip of Khc and Klc mutant bristles. We propose that the failure of bristle tapering is due to defects in directing dynamic MTs at the growing tip. Thus, we reveal a new function for Khc and Klc in directing dynamic MTs during polarized cell growth. Moreover, we also demonstrate a novel mode of coordination in mitochondrial transport between Khc and Dhc.

A ROP2-RIC1 pathway fine-tunes microtubule reorganization for salt tolerance in Arabidopsis.

Science.gov (United States)

Li, Changjiang; Lu, Hanmei; Li, Wei; Yuan, Ming; Fu, Ying

2017-07-01

The reorganization of microtubules induced by salt stress is required for Arabidopsis survival under high salinity conditions. RIC1 is an effector of Rho-related GTPase from plants (ROPs) and a known microtubule-associated protein. In this study, we demonstrated that RIC1 expression decreased with long-term NaCl treatment, and ric1-1 seedlings exhibited a higher survival rate under salt stress. We found that RIC1 reduced the frequency of microtubule transition from shortening to growing status and knockout of RIC1 improved the reassembly of depolymerized microtubules caused by either oryzalin treatment or salt stress. Further investigation showed that constitutively active ROP2 promoted the reassembly of microtubules and the survival of seedlings under salt stress. A rop2-1 ric1-1 double mutant rescued the salt-sensitive phenotype of rop2-1, indicating that ROP2 functions in salt tolerance through RIC1. Although ROP2 did not regulate RIC1 expression upon salt stress, a quick but mild increase of ROP2 activity was induced, led to reduction of RIC1 on microtubules. Collectively, our study reveals an ROP2-RIC1 pathway that fine-tunes microtubule dynamics in response to salt stress in Arabidopsis. This finding not only reveals a new regulatory mechanism for microtubule reorganization under salt stress but also the importance of ROP signalling for salinity tolerance. © 2017 John Wiley & Sons Ltd.

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

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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.

Dynamic Stability Experiment of Maglev Systems,

Science.gov (United States)

1995-04-01

This report summarizes the research performed on maglev vehicle dynamic stability at Argonne National Laboratory during the past few years. It also... maglev system, it is important to consider this phenomenon in the development of all maglev systems. This report presents dynamic stability experiments...on maglev systems and compares their numerical simulation with predictions calculated by a nonlinear dynamic computer code. Instabilities of an

Magnolol Inhibits the Growth of Non-Small Cell Lung Cancer via Inhibiting Microtubule Polymerization

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Jia Shen

2017-07-01

Full Text Available Background: The tubulin/microtubule system, which is an integral component of the cytoskeleton, plays an essential role in mitosis. Targeting mitotic progression by disturbing microtubule dynamics is a rational strategy for cancer treatment. Methods: Microtubule polymerization assay was performed to examine the effect of Magnolol (a novel natural phenolic compound isolated from Magnolia obovata on cellular microtubule polymerization in human non-small cell lung cancer (NSCLC cells. Cell cycle analysis, mitotic index assay, cell proliferation assay, colony formation assay, western blotting analysis of cell cycle regulators, Annexin V-FITC/PI staining, and live/dead viability staining were carried out to investigate the Magnolol’s inhibitory effect on proliferation and viability of NSCLS cells in vitro. Xenograft model of human A549 NSCLC tumor was used to determine the Magnolol’s efficacy in vivo. Results: Magnolol treatment effectively inhibited cell proliferation and colony formation of NSCLC cells. Further study proved that Magnolol induced the mitotic phase arrest and inhibited G2/M progression in a dose-dependent manner, which were mechanistically associated with expression alteration of a series of cell cycle regulators. Furthermore, Magnolol treatment disrupted the cellular microtubule organization via inhibiting the polymerization of microtubule. We also found treatment with NSCLC cells with Magnolol resulted in apoptosis activation through a p53-independent pathway, and autophgy induction via down-regulation of the Akt/mTOR pathway. Finally, Magnolol treatment significantly suppressed the NSCLC tumor growth in mouse xenograft model in vivo. Conclusion: These findings identify Magnolol as a promising candidate with anti-microtubule polymerization activity for NSCLC treatment.

Glucose regulated proteins 78 and 75 bind to the receptor for hyaluronan mediated motility in interphase microtubules

International Nuclear Information System (INIS)

Kuwabara, Hiroko; Yoneda, Masahiko; Hayasaki, Hana; Nakamura, Toshiya; Mori, Hiroshi

2006-01-01

The receptor for hyaluronan mediated motility (RHAMM), which is a hyaluronan-binding protein, is a centrosomal and microtubal protein. Here, we have identified two RHAMM-binding proteins, glucose regulated protein (GRP) 78 and GRP75, using co-immunoprecipitation analysis. These two proteins directly bound to glutathione-S-transferase-RHAMM fusion proteins. By double immunostaining, GRP78 and GRP75 colocalized with RHAMM in interphase microtubules, but were separated in mitotic spindles. Prevention of microtubule polymerization by TN-16 and vincristine sulfate induced RHAMM overexpression without a significant change in GRP78/75. Taken together, GRP78/75 and RHAMM complexes may stabilize microtubules in the interphase, associated with a downregulation of RHAMM. These results reveal a new biochemical activity of RHAMM

Centriolar CPAP/SAS-4 Imparts Slow Processive Microtubule Growth

NARCIS (Netherlands)

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

2016-01-01

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

Ionic wave propagation and collision in an excitable circuit model of microtubules

Science.gov (United States)

Guemkam Ghomsi, P.; Tameh Berinyoh, J. T.; Moukam Kakmeni, F. M.

2018-02-01

In this paper, we report the propensity to excitability of the internal structure of cellular microtubules, modelled as a relatively large one-dimensional spatial array of electrical units with nonlinear resistive features. We propose a model mimicking the dynamics of a large set of such intracellular dynamical entities as an excitable medium. We show that the behavior of such lattices can be described by a complex Ginzburg-Landau equation, which admits several wave solutions, including the plane waves paradigm. A stability analysis of the plane waves solutions of our dynamical system is conducted both analytically and numerically. It is observed that perturbed plane waves will always evolve toward promoting the generation of localized periodic waves trains. These modes include both stationary and travelling spatial excitations. They encompass, on one hand, localized structures such as solitary waves embracing bright solitons, dark solitons, and bisolitonic impulses with head-on collisions phenomena, and on the other hand, the appearance of both spatially homogeneous and spatially inhomogeneous stationary patterns. This ability exhibited by our array of proteinic elements to display several states of excitability exposes their stunning biological and physical complexity and is of high relevance in the description of the developmental and informative processes occurring on the subcellular scale.

Unifying dynamical and structural stability of equilibria

Science.gov (United States)

Arnoldi, Jean-François; Haegeman, Bart

2016-09-01

We exhibit a fundamental relationship between measures of dynamical and structural stability of linear dynamical systems-e.g. linearized models in the vicinity of equilibria. We show that dynamical stability, quantified via the response to external perturbations (i.e. perturbation of dynamical variables), coincides with the minimal internal perturbation (i.e. perturbations of interactions between variables) able to render the system unstable. First, by reformulating a result of control theory, we explain that harmonic external perturbations reflect the spectral sensitivity of the Jacobian matrix at the equilibrium, with respect to constant changes of its coefficients. However, for this equivalence to hold, imaginary changes of the Jacobian's coefficients have to be allowed. The connection with dynamical stability is thus lost for real dynamical systems. We show that this issue can be avoided, thus recovering the fundamental link between dynamical and structural stability, by considering stochastic noise as external and internal perturbations. More precisely, we demonstrate that a linear system's response to white-noise perturbations directly reflects the intensity of internal white-noise disturbance that it can accommodate before becoming stochastically unstable.

Beam Stability and Nonlinear Dynamics. Proceedings

International Nuclear Information System (INIS)

Parsa, Z.

1997-01-01

These proceedings represent papers presented at the Beam Stability and Nonlinear Dynamics symposium held in Santa Barbara in December 1996. The symposium was sponsored by the National Science Foundation as part of the United States long term accelerator research. The focus of this symposium was on nonlinear dynamics and beam stability. The topics included single-particle and many-particle dynamics, and stability in large circular accelerators such as the Large Hadron Collider(LHC). Other subjects covered were spin dynamics, nonlinear aberration correction, collective effects in the LHC, sawtooth instability and Landau damping in the presence of strong nonlinearity. There were presentations concerning plasma physics including the effect of beam echo. There are 17 papers altogether in these proceedings and 8 of them have been abstracted for the Energy Science and Technology database

Stability of molecular dynamics simulations of classical systems

DEFF Research Database (Denmark)

Toxværd, Søren

2012-01-01

The existence of a shadow Hamiltonian for discrete classical dynamics, obtained by an asymptotic expansion for a discrete symplectic algorithm, is employed to determine the limit of stability for molecular dynamics (MD) simulations with respect to the time-increment h of the discrete dynamics....... The investigation is based on the stability of the shadow energy, obtained by including the first term in the asymptotic expansion, and on the exact solution of discrete dynamics for a single harmonic mode. The exact solution of discrete dynamics for a harmonic potential with frequency ω gives a criterion...... for the limit of stability h ⩽ 2/ω. Simulations of the Lennard-Jones system and the viscous Kob-Andersen system show that one can use the limit of stability of the shadow energy or the stability criterion for a harmonic mode on the spectrum of instantaneous frequencies to determine the limit of stability of MD...

Cabazitaxel-induced stabilization of microtubules enhances radiosensitivity in ovarian cancer cells

Directory of Open Access Journals (Sweden)

Charles eKunos

2013-09-01

Full Text Available Background: Up to 40% of women with ovarian cancer have short disease-free intervals due to molecular mechanisms of chemotherapy resistance. New therapeutic strategies are sought. Ovarian cancers are sensitive to radiochemotherapy. The taxane cabazitaxel (XRP6258, Jevtana promotes tubulin assembly and stabilizes microtubules against depolymerization in cells, acting similarly in mechanism to paclitaxel. Here, sequences of cabazitaxel-radiation co-administration are tested for drug-alone cytotoxicity and optimal radiosensitization.Methods: SKOV3, OVCAR3, and TOV-112D ovarian cancer cells were administered cabazitaxel 24 h before (first, 18 h before (second, together (third, or 24 h after (fourth a single radiation dose, and then, investigated by clonogenic assay and flow cytometric assays. Radiation dose-cell survival data were fitted by two-stage multivariate analyses of variance. High content flow cytometry partitioned cabazitaxel effects into G2-phase versus M-phase events by DNA content, cyclin A2, and phospho-S10-histone H3 (PHH3. Paclitaxel served as a comparator. Findings: Cabazitaxel cytotoxicity and radiosensitization were dose dependent. Cabazitaxel added 24 h before radiation was the most lethal schedule. DNA content measurements by flow cytometry showed that cabazitaxel-treated cells accumulated in the radiosensitive G2/M 4C DNA complement compartment. Cytometry also showed that surviving cabazitaxel-induced cell cycle arrested cells resolve the arrest by entering 4C or by 8C DNA complement cell cycles.Interpretation: The radiosensitizing effect of cabazitaxel was schedule dependent, due to cell cycle redistribution, and best when cabazitaxel was given 24 h before radiation. Clinical trials of administering both cabazitaxel and radiation should be explored in women with chemoresistant ovarian cancer. Funding: Case Comprehensive Cancer Center and Sanofi-Aventis

Finding the Cell Center by a Balance of Dynein and Myosin Pulling and Microtubule Pushing: A Computational Study

Science.gov (United States)

Zhu, Jie; Burakov, Anton; Rodionov, Vladimir

2010-01-01

The centrosome position in many types of interphase cells is actively maintained in the cell center. Our previous work indicated that the centrosome is kept at the center by pulling force generated by dynein and actin flow produced by myosin contraction and that an unidentified factor that depends on microtubule dynamics destabilizes position of the centrosome. Here, we use modeling to simulate the centrosome positioning based on the idea that the balance of three forces—dyneins pulling along microtubule length, myosin-powered centripetal drag, and microtubules pushing on organelles—is responsible for the centrosome displacement. By comparing numerical predictions with centrosome behavior in wild-type and perturbed interphase cells, we rule out several plausible hypotheses about the nature of the microtubule-based force. We conclude that strong dynein- and weaker myosin-generated forces pull the microtubules inward competing with microtubule plus-ends pushing the microtubule aster outward and that the balance of these forces positions the centrosome at the cell center. The model also predicts that kinesin action could be another outward-pushing force. Simulations demonstrate that the force-balance centering mechanism is robust yet versatile. We use the experimental observations to reverse engineer the characteristic forces and centrosome mobility. PMID:20980619

Structural stability of nonlinear population dynamics.

Science.gov (United States)

Cenci, Simone; Saavedra, Serguei

2018-01-01

In population dynamics, the concept of structural stability has been used to quantify the tolerance of a system to environmental perturbations. Yet, measuring the structural stability of nonlinear dynamical systems remains a challenging task. Focusing on the classic Lotka-Volterra dynamics, because of the linearity of the functional response, it has been possible to measure the conditions compatible with a structurally stable system. However, the functional response of biological communities is not always well approximated by deterministic linear functions. Thus, it is unclear the extent to which this linear approach can be generalized to other population dynamics models. Here, we show that the same approach used to investigate the classic Lotka-Volterra dynamics, which is called the structural approach, can be applied to a much larger class of nonlinear models. This class covers a large number of nonlinear functional responses that have been intensively investigated both theoretically and experimentally. We also investigate the applicability of the structural approach to stochastic dynamical systems and we provide a measure of structural stability for finite populations. Overall, we show that the structural approach can provide reliable and tractable information about the qualitative behavior of many nonlinear dynamical systems.

Structural stability of nonlinear population dynamics

Science.gov (United States)

Cenci, Simone; Saavedra, Serguei

2018-01-01

In population dynamics, the concept of structural stability has been used to quantify the tolerance of a system to environmental perturbations. Yet, measuring the structural stability of nonlinear dynamical systems remains a challenging task. Focusing on the classic Lotka-Volterra dynamics, because of the linearity of the functional response, it has been possible to measure the conditions compatible with a structurally stable system. However, the functional response of biological communities is not always well approximated by deterministic linear functions. Thus, it is unclear the extent to which this linear approach can be generalized to other population dynamics models. Here, we show that the same approach used to investigate the classic Lotka-Volterra dynamics, which is called the structural approach, can be applied to a much larger class of nonlinear models. This class covers a large number of nonlinear functional responses that have been intensively investigated both theoretically and experimentally. We also investigate the applicability of the structural approach to stochastic dynamical systems and we provide a measure of structural stability for finite populations. Overall, we show that the structural approach can provide reliable and tractable information about the qualitative behavior of many nonlinear dynamical systems.

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...

Cellular effects of the microtubule-targeting agent peloruside A in hypoxia-conditioned colorectal carcinoma cells.

Science.gov (United States)

Řehulka, Jiří; Annadurai, Narendran; Frydrych, Ivo; Znojek, Pawel; Džubák, Petr; Northcote, Peter; Miller, John H; Hajdúch, Marián; Das, Viswanath

2017-07-01

Hypoxia is a prominent feature of solid tumors, dramatically remodeling microtubule structures and cellular pathways and contributing to paclitaxel resistance. Peloruside A (PLA), a microtubule-targeting agent, has shown promising anti-tumor effects in preclinical studies. Although it has a similar mode of action to paclitaxel, it binds to a distinct site on β-tubulin that differs from the classical taxane site. In this study, we examined the unexplored effects of PLA in hypoxia-conditioned colorectal HCT116 cancer cells. Cytotoxicity of PLA was determined by cell proliferation assay. The effects of a pre-exposure to hypoxia on PLA-induced cell cycle alterations and apoptosis were examined by flow cytometry, time-lapse imaging, and western blot analysis of selected markers. The hypoxia effect on stabilization of microtubules by PLA was monitored by an intracellular tubulin polymerization assay. Our findings show that the cytotoxicity of PLA is not altered in hypoxia-conditioned cells compared to paclitaxel and vincristine. Furthermore, hypoxia does not alter PLA-induced microtubule stabilization nor the multinucleation of cells. PLA causes cyclin B1 and G2/M accumulation followed by apoptosis. The cellular and molecular effects of PLA have been determined in normoxic conditions, but there are no reports of PLA effects in hypoxic cells. Our findings reveal that hypoxia preconditioning does not alter the sensitivity of HCT116 to PLA. These data report on the cellular and molecular effects of PLA in hypoxia-conditioned cells for the first time, and will encourage further exploration of PLA as a promising anti-tumor agent. Copyright © 2017 Elsevier B.V. All rights reserved.

Producing Conditional Mutants for Studying Plant Microtubule Function

Energy Technology Data Exchange (ETDEWEB)

Richard Cyr

2009-09-29

The cytoskeleton, and in particular its microtubule component, participates in several processes that directly affect growth and development in higher plants. Normal cytoskeletal function requires the precise and orderly arrangement of microtubules into several cell cycle and developmentally specific arrays. One of these, the cortical array, is notable for its role in directing the deposition of cellulose (the most prominent polymer in the biosphere). An understanding of how these arrays form, and the molecular interactions that contribute to their function, is incomplete. To gain a better understanding of how microtubules work, we have been working to characterize mutants in critical cytoskeletal genes. This characterization is being carried out at the subcellular level using vital microtubule gene constructs. In the last year of funding colleagues have discovered that gamma-tubulin complexes form along the lengths of cortical microtubules where they act to spawn new microtubules at a characteristic 40 deg angle. This finding complements nicely the finding from our lab (which was funded by the DOE) showing that microtubule encounters are angle dependent; high angles encounters results in catastrophic collisions while low angle encounters result in favorable zippering. The finding of a 40 deg spawn of new microtubules from extant microtubule, together with aforementioned rules of encounters, insures favorable co-alignment in the array. I was invited to write a New and Views essay on this topic and a PDF is attached (News and Views policy does not permit funding acknowledgments and so I was not allowed to acknowledge support from the DOE).

Ferritin associates with marginal band microtubules

International Nuclear Information System (INIS)

Infante, Anthony A.; Infante, Dzintra; Chan, M.-C.; How, P.-C.; Kutschera, Waltraud; Linhartova, Irena; Muellner, Ernst W.; Wiche, Gerhard; Propst, Friedrich

2007-01-01

We characterized chicken erythrocyte and human platelet ferritin by biochemical studies and immunofluorescence. Erythrocyte ferritin was found to be a homopolymer of H-ferritin subunits, resistant to proteinase K digestion, heat stable, and contained iron. In mature chicken erythrocytes and human platelets, ferritin was localized at the marginal band, a ring-shaped peripheral microtubule bundle, and displayed properties of bona fide microtubule-associated proteins such as tau. Red blood cell ferritin association with the marginal band was confirmed by temperature-induced disassembly-reassembly of microtubules. During erythrocyte differentiation, ferritin co-localized with coalescing microtubules during marginal band formation. In addition, ferritin was found in the nuclei of mature erythrocytes, but was not detectable in those of bone marrow erythrocyte precursors. These results suggest that ferritin has a function in marginal band formation and possibly in protection of the marginal band from damaging effects of reactive oxygen species by sequestering iron in the mature erythrocyte. Moreover, our data suggest that ferritin and syncolin, a previously identified erythrocyte microtubule-associated protein, are identical. Nuclear ferritin might contribute to transcriptional silencing or, alternatively, constitute a ferritin reservoir

Inhibition of glycogen synthase kinase-3 reduces extension of the axonal leading process by destabilizing microtubules in cerebellar granule neurons.

Science.gov (United States)

Inami, Yoshihiro; Omura, Mitsuru; Kubota, Kenta; Konishi, Yoshiyuki

2018-07-01

Recent studies have uncovered various molecules that play key roles in neuronal morphogenesis. Nevertheless, the mechanisms underlying the neuron-type-dependent regulation of morphogenesis remain unknown. We have previously reported that inhibition of glycogen synthase kinase-3 (GSK3) markedly reduced axonal length of cerebellar granule neurons (CGNs) in a neuron-type-dependent manner. In the present study, we investigated the mechanisms by which the growth of CGN axons was severely suppressed upon GSK3 inhibition. Using time-lapse imaging of cultured CGNs at early morphogenesis, we found that extension of the leading process was severely inhibited by the pharmacological inhibition of GSK3. The rate of somal migration was also reduced with a GSK3 inhibitor in dissociated culture as well as in microexplant culture. In addition, CGNs ectopically expressed with a catalytically inactive mutant of GSK3 exhibited a migration defect in vivo. In axonal leading processes of CGNs, detyrosination and acetylation of α-tubulin, which are known to correlate with microtubule stability, were decreased by GSK3 inhibition. A photoconversion analysis found that inhibition of GSK3 increases the turnover of microtubules. Furthermore, in the presence of paclitaxel, a microtubule-stabilizing reagent, inhibition of GSK3 recovered the axonal leading process extension that was reduced by paclitaxel. Our results suggest that GSK3 supports the extension of axonal processes by stabilizing microtubules, contrary to its function in other neuron-types, lending mechanical insight into neuron-type-dependent morphological regulation. Copyright © 2018 Elsevier B.V. All rights reserved.

A new kymogram-based method reveals unexpected effects of marker protein expression and spatial anisotropy of cytoskeletal dynamics in plant cell cortex

Czech Academy of Sciences Publication Activity Database

Cvrčková, F.; Oulehlová, Denisa

2017-01-01

Roč. 13, MAR 27 (2017), č. článku 19. ISSN 1746-4811 Institutional support: RVO:61389030 Keywords : green fluorescent protein * cortical microtubule arrays * actin stochastic dynamics * arabidopsis-thaliana * epidermal-cells * quantitative-analysis * binding domain * gfp-tubulin * f-actin * microscopy * Actin * Microtubules * Lifeact * Variable angle fluorescence microscopy * Spinning disc confocal microscopy * Kymogram * Structure stability * Lateral mobility * Anisotropy * FH1 (At3g25500) Subject RIV: EB - Genetics ; Molecular Biology OBOR OECD: Cell biology Impact factor: 3.510, year: 2016

Auxin-dependent microtubule responses and seedling development are affected in a rice mutant resistant to EPC

International Nuclear Information System (INIS)

Nick, P.; Yatou, O.; Furuya, M.; Lambert, A.M.

1994-01-01

Mutants in rice (Oryza sativa L. cv. japonica) were used to study the role of the cytoskeleton in signal-dependent morphogenesis. Mutants obtained by gamma ray irradiation were selected that failed to show inhibition of coleoptile elongation by the anti microtubular drug ethyl-N-phenylcarbamate (EPC). The mutation EPC-Resistant 31 (ER31), isolated from such a screen, caused lethality in putatively homozygous embryos. Heterozygotes exhibited drug resistance, impaired development of crown roots, and characteristic changes in the pattern of cell elongation: cell elongation was enhanced in mesocotyls and leaf sheaths, but inhibited in coleoptiles. The orientation of cortical microtubules changed correspondingly: for etiolated seedlings, compared with the wild-type, they were more transverse with respect to the long cell axis in mesocotyls and leaf sheaths, but more longitudinal in coleoptiles. In mutant coleoptiles, in contrast to wild-type, microtubules did not reorient in response to auxin, and their response to microtubule-eliminating and microtubule-stabilizing drugs was conspicuously reduced. In contrast, they responded normally to other stimuli such as gibberellins or red light. Auxin sensitivity as assayed by the dose-response for callus induction did not show any significant differences between wild-type and mutant. The mutant phenotype is interpreted in terms of an interrupted link between auxin-triggered signal transduction and microtubule reorientation. (author)

Novel mitochondrial extensions provide evidence for a link between microtubule-directed movement and mitochondrial fission

International Nuclear Information System (INIS)

Bowes, Timothy; Gupta, Radhey S.

2008-01-01

Mitochondrial dynamics play an important role in a large number of cellular processes. Previously, we reported that treatment of mammalian cells with the cysteine-alkylators, N-ethylmaleimide and ethacrynic acid, induced rapid mitochondrial fusion forming a large reticulum approximately 30 min after treatment. Here, we further investigated this phenomenon using a number of techniques including live-cell confocal microscopy. In live cells, drug-induced fusion coincided with a cessation of fast mitochondrial movement which was dependent on microtubules. During this loss of movement, thin mitochondrial tubules extending from mitochondria were also observed, which we refer to as 'mitochondrial extensions'. The formation of these mitochondrial extensions, which were not observed in untreated cells, depended on microtubules and was abolished by pretreatment with nocodazole. In this study, we provide evidence that these extensions result from of a block in mitochondrial fission combined with continued application of motile force by microtubule-dependent motor complexes. Our observations strongly suggest the existence of a link between microtubule-based mitochondrial trafficking and mitochondrial fission

Duplication in the microtubule-actin cross-linking factor 1 gene causes a novel neuromuscular condition

DEFF Research Database (Denmark)

Jørgensen, Louise H; Mosbech, Mai-Britt; Færgeman, Nils J

2014-01-01

Spectrins and plakins are important communicators linking cytoskeletal components to each other and to cellular junctions. Microtubule-actin cross-linking factor 1 (MACF1) belongs to the spectraplakin family and is involved in control of microtubule dynamics. Complete knock out of MACF1 in mice...... muscles and diminished motor skills, with heterogeneous presentation among the affected family members. To corroborate these findings we used RNA interference to knock down the VAB-10 locus containing the MACF1 homologue in C. elegans, and we could show that this also causes movement disturbances...

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.

Lateral motion and bending of microtubules studied with a new single-filament tracking routine in living cells.

Science.gov (United States)

Pallavicini, Carla; Levi, Valeria; Wetzler, Diana E; Angiolini, Juan F; Benseñor, Lorena; Despósito, Marcelo A; Bruno, Luciana

2014-06-17

The cytoskeleton is involved in numerous cellular processes such as migration, division, and contraction and provides the tracks for transport driven by molecular motors. Therefore, it is very important to quantify the mechanical behavior of the cytoskeletal filaments to get a better insight into cell mechanics and organization. It has been demonstrated that relevant mechanical properties of microtubules can be extracted from the analysis of their motion and shape fluctuations. However, tracking individual filaments in living cells is extremely complex due, for example, to the high and heterogeneous background. We introduce a believed new tracking algorithm that allows recovering the coordinates of fluorescent microtubules with ∼9 nm precision in in vitro conditions. To illustrate potential applications of this algorithm, we studied the curvature distributions of fluorescent microtubules in living cells. By performing a Fourier analysis of the microtubule shapes, we found that the curvatures followed a thermal-like distribution as previously reported with an effective persistence length of ∼20 μm, a value significantly smaller than that measured in vitro. We also verified that the microtubule-associated protein XTP or the depolymerization of the actin network do not affect this value; however, the disruption of intermediate filaments decreased the persistence length. Also, we recovered trajectories of microtubule segments in actin or intermediate filament-depleted cells, and observed a significant increase of their motion with respect to untreated cells showing that these filaments contribute to the overall organization of the microtubule network. Moreover, the analysis of trajectories of microtubule segments in untreated cells showed that these filaments presented a slower but more directional motion in the cortex with respect to the perinuclear region, and suggests that the tracking routine would allow mapping the microtubule dynamical organization in cells

Microtubules are organized independently of the centrosome in Drosophila neurons

Directory of Open Access Journals (Sweden)

Nguyen Michelle M

2011-12-01

Full Text Available Abstract Background The best-studied arrangement of microtubules is that organized by the centrosome, a cloud of microtubule nucleating and anchoring proteins is clustered around centrioles. However, noncentrosomal microtubule arrays are common in many differentiated cells, including neurons. Although microtubules are not anchored at neuronal centrosomes, it remains unclear whether the centrosome plays a role in organizing neuronal microtubules. We use Drosophila as a model system to determine whether centrosomal microtubule nucleation is important in mature neurons. Results In developing and mature neurons, centrioles were not surrounded by the core nucleation protein γ-tubulin. This suggests that the centrioles do not organize functional centrosomes in Drosophila neurons in vivo. Consistent with this idea, centriole position was not correlated with a specific region of the cell body in neurons, and growing microtubules did not cluster around the centriole, even after axon severing when the number of growing plus ends is dramatically increased. To determine whether the centrosome was required for microtubule organization in mature neurons, we used two approaches. First, we used DSas-4 centriole duplication mutants. In these mutants, centrioles were present in many larval sensory neurons, but they were not fully functional. Despite reduced centriole function, microtubule orientation was normal in axons and dendrites. Second, we used laser ablation to eliminate the centriole, and again found that microtubule polarity in axons and dendrites was normal, even 3 days after treatment. Conclusion We conclude that the centrosome is not a major site of microtubule nucleation in Drosophila neurons, and is not required for maintenance of neuronal microtubule organization in these cells.

Local Dynamic Stability Associated with Load Carrying

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Jian Liu

2013-03-01

Conclusion: Current study confirmed the sensitivity of local dynamic stability measure in load carrying situation. It was concluded that load carrying tasks were associated with declined local dynamic stability, which may result in increased risk of fall accident. This finding has implications in preventing fall accidents associated with occupational load carrying.

Effects of microtubule mechanics on hydrolysis and catastrophes

International Nuclear Information System (INIS)

Müller, N; Kierfeld, J

2014-01-01

We introduce a model for microtubule (MT) mechanics containing lateral bonds between dimers in neighboring protofilaments, bending rigidity of dimers, and repulsive interactions between protofilaments modeling steric constraints to investigate the influence of mechanical forces on hydrolysis and catastrophes. We use the allosteric dimer model, where tubulin dimers are characterized by an equilibrium bending angle, which changes from 0 ∘ to 22 ∘ by hydrolysis of a dimer. This also affects the lateral interaction and bending energies and, thus, the mechanical equilibrium state of the MT. As hydrolysis gives rise to conformational changes in dimers, mechanical forces also influence the hydrolysis rates by mechanical energy changes modulating the hydrolysis rate. The interaction via the MT mechanics then gives rise to correlation effects in the hydrolysis dynamics, which have not been taken into account before. Assuming a dominant influence of mechanical energies on hydrolysis rates, we investigate the most probable hydrolysis pathways both for vectorial and random hydrolysis. Investigating the stability with respect to lateral bond rupture, we identify initiation configurations for catastrophes along the hydrolysis pathways and values for a lateral bond rupture force. If we allow for rupturing of lateral bonds between dimers in neighboring protofilaments above this threshold force, our model exhibits avalanche-like catastrophe events. (papers)

Role of CAP350 in centriolar tubule stability and centriole assembly.

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Mikael Le Clech

Full Text Available BACKGROUND: Centrioles are microtubule-based cylindrical structures composed of nine triplet tubules and are required for the formation of the centrosome, flagella and cilia. Despite theirs importance, centriole biogenesis is poorly understood. Centrosome duplication is initiated at the G1/S transition by the sequential recruitment of a set of conserved proteins under the control of the kinase Plk4. Subsequently, the procentriole is assembled by the polymerization of centriolar tubules via an unknown mechanism involving several tubulin paralogs. METHODOLOGY/PRINCIPAL FINDINGS: Here, we developed a cellular assay to study centrosome duplication and procentriole stability based on its sensitivity to the microtubule-depolymerizing drug nocodazole. By using RNA interference experiments, we show that the stability of growing procentrioles is regulated by the microtubule-stabilizing protein CAP350, independently of hSAS-6 and CPAP which initiate procentriole growth. Furthermore, our analysis reveals the critical role of centriolar tubule stability for an efficient procentriole growth. CONCLUSIONS/SIGNIFICANCE: CAP350 belongs to a new class of proteins which associate and stabilize centriolar tubules to control centriole duplication.

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.

Changes in microtubule-associated protein tau during peripheral nerve injury and regeneration

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Guang-bin Zha

2016-01-01

Full Text Available Tau, a primary component of microtubule-associated protein, promotes microtubule assembly and/or disassembly and maintains the stability of the microtubule structure. Although the importance of tau in neurodegenerative diseases has been well demonstrated, whether tau is involved in peripheral nerve regeneration remains unknown. In the current study, we obtained sciatic nerve tissue from adult rats 0, 1, 4, 7, and 14 days after sciatic nerve crush and examined tau mRNA and protein expression levels and the location of tau in the sciatic nerve following peripheral nerve injury. The results from our quantitative reverse transcription polymerase chain reaction analysis showed that compared with the uninjured control sciatic nerve, mRNA expression levels for both tau and tau tubulin kinase 1, a serine/threonine kinase that regulates tau phosphorylation, were decreased following peripheral nerve injury. Our western blot assay results suggested that the protein expression levels of tau and phosphorylated tau initially decreased 1 day post nerve injury but then gradually increased. The results of our immunohistochemical labeling showed that the location of tau protein was not altered by nerve injury. Thus, these results showed that the expression of tau was changed following sciatic nerve crush, suggesting that tau may be involved in peripheral nerve repair and regeneration.

Phosphorylation of the yeast γ-tubulin Tub4 regulates microtubule function

DEFF Research Database (Denmark)

Lin, Tien-chen; Gombos, Linda; Neuner, Annett

2011-01-01

The yeast ¿-tubulin Tub4 is assembled with Spc97 and Spc98 into the small Tub4 complex. The Tub4 complex binds via the receptor proteins Spc72 and Spc110 to the spindle pole body (SPB), the functional equivalent of the mammalian centrosome, where the Tub4 complex organizes cytoplasmic and nuclear...... microtubules. Little is known about the regulation of the Tub4 complex. Here, we isolated the Tub4 complex with the bound receptors from yeast cells. Analysis of the purified Tub4 complex by mass spectrometry identified more than 50 phosphorylation sites in Spc72, Spc97, Spc98, Spc110 and Tub4. To examine...... the functional relevance of the phosphorylation sites, phospho-mimicking and non-phosphorylatable mutations in Tub4, Spc97 and Spc98 were analyzed. Three phosphorylation sites in Tub4 were found to be critical for Tub4 stability and microtubule organization. One of the sites is highly conserved in ¿-tubulins...

Image-based compound profiling reveals a dual inhibitor of tyrosine kinase and microtubule polymerization.

Science.gov (United States)

Tanabe, Kenji

2016-04-27

Small-molecule compounds are widely used as biological research tools and therapeutic drugs. Therefore, uncovering novel targets of these compounds should provide insights that are valuable in both basic and clinical studies. I developed a method for image-based compound profiling by quantitating the effects of compounds on signal transduction and vesicle trafficking of epidermal growth factor receptor (EGFR). Using six signal transduction molecules and two markers of vesicle trafficking, 570 image features were obtained and subjected to multivariate analysis. Fourteen compounds that affected EGFR or its pathways were classified into four clusters, based on their phenotypic features. Surprisingly, one EGFR inhibitor (CAS 879127-07-8) was classified into the same cluster as nocodazole, a microtubule depolymerizer. In fact, this compound directly depolymerized microtubules. These results indicate that CAS 879127-07-8 could be used as a chemical probe to investigate both the EGFR pathway and microtubule dynamics. The image-based multivariate analysis developed herein has potential as a powerful tool for discovering unexpected drug properties.

The Kinesin Adaptor Calsyntenin-1 Organizes Microtubule Polarity and Regulates Dynamics during Sensory Axon Arbor Development

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Mary C. Halloran

2017-04-01

Full Text Available Axon growth and branching, and development of neuronal polarity are critically dependent on proper organization and dynamics of the microtubule (MT cytoskeleton. MTs must organize with correct polarity for delivery of diverse cargos to appropriate subcellular locations, yet the molecular mechanisms regulating MT polarity remain poorly understood. Moreover, how an actively branching axon reorganizes MTs to direct their plus ends distally at branch points is unknown. We used high-speed, in vivo imaging of polymerizing MT plus ends to characterize MT dynamics in developing sensory axon arbors in zebrafish embryos. We find that axonal MTs are highly dynamic throughout development, and that the peripheral and central axons of sensory neurons show differences in MT behaviors. Furthermore, we show that Calsyntenin-1 (Clstn-1, a kinesin adaptor required for sensory axon branching, also regulates MT polarity in developing axon arbors. In wild type neurons the vast majority of MTs are directed in the correct plus-end-distal orientation from early stages of development. Loss of Clstn-1 causes an increase in MTs polymerizing in the retrograde direction. These misoriented MTs most often are found near growth cones and branch points, suggesting Clstn-1 is particularly important for organizing MT polarity at these locations. Together, our results suggest that Clstn-1, in addition to regulating kinesin-mediated cargo transport, also organizes the underlying MT highway during axon arbor development.

CLASP2 interacts with p120-catenin and governs microtubule dynamics at adherens junctions

DEFF Research Database (Denmark)

Shahbazi, Marta N; Megias, Diego; Epifano, Carolina

2013-01-01

Classical cadherins and their connections with microtubules (MTs) are emerging as important determinants of cell adhesion. However, the functional relevance of such interactions and the molecular players that contribute to tissue architecture are still emerging. In this paper, we report that the ...

Dynamic postural stability in blind athletes using the biodex stability system.

Science.gov (United States)

Aydoğ, E; Aydoğ, S T; Cakci, A; Doral, M N

2006-05-01

Three systems affect the upright standing posture in humans - visual, vestibular, and somatosensory. It is well known that the visually impaired individuals have bad postural balance. On the other hand, it is a well documented fact that some sports can improve postural balance. Therefore, it is aimed in this study to evaluate the dynamic postural stability in goal-ball athletes. Twenty blind goal-ball players, 20 sighted and 20 sedentary blind controls were evaluated using the Biodex Stability System. Three adaptation trials and three test evaluations (a 20-second balance test at a platform stability of 8) were applied to the blind people, and to the sighted with eyes open and closed. Dynamic postural stability was measured on the basis of three indices: overall, anteroposterior, and mediolateral. Means of each test score were calculated. The tests results were compared for the blind athletes, sighted (with eyes open and closed) subjects, and sedentary blind people. There were significant differences between the results of the blind people and the sighted subjects with regards to all of the three indices. Although the stability of goal-ball players was better than sedentary blinds', only ML index values were statistically different (4.47 +/- 1.24 in the goal-ball players; 6.46 +/- 3.42 in the sedentary blind, p = 0.04). Dynamic postural stability was demonstrated to be affected by vision; and it was found that blind people playing goal-ball 1 - 2 days per week have higher ML stability than the sedentary sighted people.

Dynamics and regulation at the tip : a high resolution view on microtubele assembly

NARCIS (Netherlands)

Munteanu, Laura

2008-01-01

Microtubules are highly dynamic protein polymers that and are essential for intracellular organization and fundamental processes like transport and cell division. In cells, a wide family of microtubule-associated proteins (MAPs) tightly regulates microtubule dynamics. The work presented in this

Katanin localization requires triplet microtubules in Chlamydomonas reinhardtii.

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Jessica M Esparza

Full Text Available Centrioles and basal bodies are essential for a variety of cellular processes that include the recruitment of proteins to these structures for both centrosomal and ciliary function. This recruitment is compromised when centriole/basal body assembly is defective. Mutations that cause basal body assembly defects confer supersensitivity to Taxol. These include bld2, bld10, bld12, uni3, vfl1, vfl2, and vfl3. Flagellar motility mutants do not confer sensitivity with the exception of mutations in the p60 (pf19 and p80 (pf15 subunits of the microtubule severing protein katanin. We have identified additional pf15 and bld2 (ε-tubulin alleles in screens for Taxol sensitivity. Null pf15 and bld2 alleles are viable and are not essential genes in Chlamydomonas. Analysis of double mutant strains with the pf15-3 and bld2-6 null alleles suggests that basal bodies in Chlamydomonas may recruit additional proteins beyond katanin that affect spindle microtubule stability. The bld2-5 allele is a hypomorphic allele and its phenotype is modulated by nutritional cues. Basal bodies in bld2-5 cells are missing proximal ends. The basal body mutants show aberrant localization of an epitope-tagged p80 subunit of katanin. Unlike IFT proteins, katanin p80 does not localize to the transition fibers of the basal bodies based on an analysis of the uni1 mutant as well as the lack of colocalization of katanin p80 with IFT74. We suggest that the triplet microtubules are likely to play a key role in katanin p80 recruitment to the basal body of Chlamydomonas rather than the transition fibers that are needed for IFT localization.

Effect of Footwear on Dynamic Stability during Single-leg Jump Landings.

Science.gov (United States)

Bowser, Bradley J; Rose, William C; McGrath, Robert; Salerno, Jilian; Wallace, Joshua; Davis, Irene S

2017-06-01

Barefoot and minimal footwear running has led to greater interest in the biomechanical effects of different types of footwear. The effect of running footwear on dynamic stability is not well understood. The purpose of this study was to compare dynamic stability and impact loading across 3 footwear conditions; barefoot, minimal footwear and standard running shoes. 25 injury free runners (21 male, 4 female) completed 5 single-leg jump landings in each footwear condition. Dynamic stability was assessed using the dynamic postural stability index and its directional components (mediolateral, anteroposterior, vertical). Peak vertical ground reaction force and vertical loadrates were also compared across footwear conditions. Dynamic stability was dependent on footwear type for all stability indices (ANOVA, pfootwear for the anteroposterior stability index (pfootwear (p≤0.05). Dynamic stability, peak vertical force, and average loadrates during single-leg jump landings appear to be affected by footwear type. The results suggest greater dynamic stability and lower impact loading when landing barefoot or in minimal footwear. © Georg Thieme Verlag KG Stuttgart · New York.

Branching microtubule nucleation in Xenopus egg extracts mediated by augmin and TPX2

Science.gov (United States)

Petry, Sabine; Groen, Aaron C.; Ishihara, Keisuke; Mitchison, Timothy J.; Vale, Ronald D.

2013-01-01

Summary The microtubules that comprise mitotic spindles in animal cells are nucleated at centrosomes and by spindle assembly factors that are activated in the vicinity of chromatin. Indirect evidence also has suggested that microtubules might be nucleated from pre-existing microtubules throughout the spindle, but this process has not been observed directly. Here, we demonstrate microtubule nucleation from the sides of existing microtubules in meiotic Xenopus egg extracts. Daughter microtubules grow at a low branch angle and with the same polarity as mother filaments. Branching microtubule nucleation requires gamma-tubulin and augmin and is stimulated by GTP-bound Ran and its effector TPX2, factors previously implicated in chromatin-stimulated nucleation. Because of the rapid amplification of microtubule numbers and the preservation of microtubule polarity, microtubule-dependent microtubule nucleation is well suited for spindle assembly and maintenance. PMID:23415226

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.

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

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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.

Dynamic Stability of Maglev Systems,

Science.gov (United States)

1992-04-01

AD-A259 178 ANL-92/21 Materials and Components Dynamic Stability of Technology Division Materials and Components Maglev Systems Technology Division...of Maglev Systems Y. Cai, S. S. Chen, and T. M. Mulcahy Materials and Components Technology Division D. M. Rote Center for Transportation Research...of Maglev System with L-Shaped Guideway ......................................... 6 3 Stability of M aglev System s

Microtubule self-organisation by reaction-diffusion processes causes collective transport and organisation of cellular particles

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Demongeot Jacques

2004-06-01

Full Text Available Abstract Background The transport of intra-cellular particles by microtubules is a major biological function. Under appropriate in vitro conditions, microtubule preparations behave as a 'complex' system and show 'emergent' phenomena. In particular, they form dissipative structures that self-organise over macroscopic distances by a combination of reaction and diffusion. Results Here, we show that self-organisation also gives rise to a collective transport of colloidal particles along a specific direction. Particles, such as polystyrene beads, chromosomes, nuclei, and vesicles are carried at speeds of several microns per minute. The process also results in the macroscopic self-organisation of these particles. After self-organisation is completed, they show the same pattern of organisation as the microtubules. Numerical simulations of a population of growing and shrinking microtubules, incorporating experimentally realistic reaction dynamics, predict self-organisation. They forecast that during self-organisation, macroscopic parallel arrays of oriented microtubules form which cross the reaction space in successive waves. Such travelling waves are capable of transporting colloidal particles. The fact that in the simulations, the aligned arrays move along the same direction and at the same speed as the particles move, suggest that this process forms the underlying mechanism for the observed transport properties. Conclusions This process constitutes a novel physical chemical mechanism by which chemical energy is converted into collective transport of colloidal particles along a given direction. Self-organisation of this type provides a new mechanism by which intra cellular particles such as chromosomes and vesicles can be displaced and simultaneously organised by microtubules. It is plausible that processes of this type occur in vivo.

Microtubule self-organisation by reaction-diffusion processes causes collective transport and organisation of cellular particles

Science.gov (United States)

Glade, Nicolas; Demongeot, Jacques; Tabony, James

2004-01-01

Background The transport of intra-cellular particles by microtubules is a major biological function. Under appropriate in vitro conditions, microtubule preparations behave as a 'complex' system and show 'emergent' phenomena. In particular, they form dissipative structures that self-organise over macroscopic distances by a combination of reaction and diffusion. Results Here, we show that self-organisation also gives rise to a collective transport of colloidal particles along a specific direction. Particles, such as polystyrene beads, chromosomes, nuclei, and vesicles are carried at speeds of several microns per minute. The process also results in the macroscopic self-organisation of these particles. After self-organisation is completed, they show the same pattern of organisation as the microtubules. Numerical simulations of a population of growing and shrinking microtubules, incorporating experimentally realistic reaction dynamics, predict self-organisation. They forecast that during self-organisation, macroscopic parallel arrays of oriented microtubules form which cross the reaction space in successive waves. Such travelling waves are capable of transporting colloidal particles. The fact that in the simulations, the aligned arrays move along the same direction and at the same speed as the particles move, suggest that this process forms the underlying mechanism for the observed transport properties. Conclusions This process constitutes a novel physical chemical mechanism by which chemical energy is converted into collective transport of colloidal particles along a given direction. Self-organisation of this type provides a new mechanism by which intra cellular particles such as chromosomes and vesicles can be displaced and simultaneously organised by microtubules. It is plausible that processes of this type occur in vivo. PMID:15176973

Microtubule plus end-tracking proteins play critical roles in directional growth of hyphae by regulating the dynamics of cytoplasmic microtubules in Aspergillus nidulans.

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Zeng, Cui J Tracy; Kim, Hye-Ryun; Vargas Arispuro, Irasema; Kim, Jung-Mi; Huang, An-Chi; Liu, Bo

2014-11-01

Cytoplasmic microtubules (MTs) serve as a rate-limiting factor for hyphal tip growth in the filamentous fungus Aspergillus nidulans. We hypothesized that this function depended on the MT plus end-tracking proteins (+TIPs) including the EB1 family protein EBA that decorated the MT plus ends undergoing polymerization. The ebAΔ mutation reduced colony growth and the mutant hyphae appeared in an undulating pattern instead of exhibiting unidirectional growth in the control. These phenotypes were enhanced by a mutation in another +TIP gene clipA. EBA was required for plus end-tracking of CLIPA, the Kinesin-7 motor KipA, and the XMAP215 homologue AlpA. In addition, cytoplasmic dynein also depended on EBA to track on most polymerizing MT plus ends, but not for its conspicuous appearance at the MT ends near the hyphal apex. The loss of EBA reduced the number of cytoplasmic MTs and prolonged dwelling times for MTs after reaching the hyphal apex. Finally, we found that colonies were formed in the absence of EBA, CLIPA, and NUDA together, suggesting that they were dispensable for fundamental functions of MTs. This study provided a comprehensive delineation of the relationship among different +TIPs and their contributions to MT dynamics and unidirectional hyphal expansion in filamentous fungi. © 2014 John Wiley & Sons Ltd.

Vault mobility depends in part on microtubules and vaults can be recruited to the nuclear envelope

International Nuclear Information System (INIS)

Zon, Arend van; Mossink, Marieke H.; Houtsmuller, Adriaan B.; Schoester, Martijn; Scheffer, George L.; Scheper, Rik J.; Sonneveld, Pieter; Wiemer, Erik A.C.

2006-01-01

Vaults are ribonucleoproteins that may function in intracellular transport processes. We investigated the intracellular distribution and dynamics of vaults in non-small cell lung cancer cells in which vaults are labeled with the green fluorescent protein. Immunofluorescence experiments showed that vaults are dispersed throughout the cytoplasm; a small fraction is found in close proximity to microtubules. Immunoprecipitation experiments corroborated these results showing co-precipitation of MVP and β-tubulin. Using quantitative fluorescence-recovery after photobleaching (FRAP), we demonstrated that vault mobility over longer distances in part depends on intact microtubules; vaults moving slower when microtubules are depolymerized by nocodazole. Biochemical fractionation indicated a small fraction of MVP associated with the nucleus, however, no GFP-tagged vaults could be observed inside the nucleus. We observed an accumulation of vaults at the nuclear envelope upon treatment of cells with the protein synthesis inhibitor cycloheximide. Analysis of nucleo-cytoplasmic transport using a fluorescent substrate containing a classical NLS and NES expressed in MVP +/+ and MVP -/- mouse embryonic fibroblasts indicated no differences in nuclear import/export kinetics, suggesting no role for vaults in these processes. We hypothesize that a subset of vaults moves directionally via microtubules, possibly towards the nucleus

Reliability Analysis of Dynamic Stability in Waves

DEFF Research Database (Denmark)

Søborg, Anders Veldt

2004-01-01

exhibit sufficient characteristics with respect to slope at zero heel (GM value), maximum leverarm, positive range of stability and area below the leverarm curve. The rule-based requirements to calm water leverarm curves are entirely based on experience obtained from vessels in operation and recorded......The assessment of a ship's intact stability is traditionally based on a semi-empirical deterministic concept that evaluates the characteristics of ship's calm water restoring leverarm curves. Today the ship is considered safe with respect to dynamic stability if its calm water leverarm curves...... accidents in the past. The rules therefore only leaves little room for evaluation and improvement of safety of a ship's dynamic stability. A few studies have evaluated the probability of ship stability loss in waves using Monte Carlo simulations. However, since this probability may be in the order of 10...

Microtubule protein ADP-ribosylation in vitro leads to assembly inhibition and rapid depolymerization

Energy Technology Data Exchange (ETDEWEB)

Scaife, R.M. (Fred Hutchinson Cancer Research Center, Seattle, WA (United States)); Wilson, L. (Univ. of California, Santa Barbara (United States)); Purich, D.L. (Univ. of Florida, Gainesville (United States))

1992-01-14

Bovine brain microtubule protein, containing both tubulin and microtubule-associated proteins, undergoes ADP-ribosylation in the presence of ({sup 14}C)NAD{sup +} and a turkey erythrocyte mono-ADP-ribosyltransferase in vitro. The modification reaction could be demonstrated in crude brain tissue extracts where selective ADP-ribosylation of both the {alpha} and {beta} chains of tubulin and of the high molecular weight microtubule-associated protein MAP-2 occurred. In experiments with purified microtubule protein, tubulin dimer, the high molecular weight microtubule-associated protein MAP-2, and another high molecular weight microtubule-associated protein which may be a MAP-1 species were heavily labeled. Tubulin and MAP-2 incorporated ({sup 14}C)ADP-ribose to an average extent of approximately 2.4 and 30 mol of ADP-ribose/mol of protein, respectively. Assembly of microtubule protein into microtubules in vitro was inhibited by ADP-ribosylation, and incubation of assembled steady-state microtubules with ADP-ribosyltransferase and NAD{sup +} resulted in rapid depolymerization of the microtubules. Thus, the eukaryotic enzyme can ADP-ribosylate tubulin and microtubule-associated proteins to much greater extents than previously observed with cholera and pertussis toxins, and the modification can significantly modulate microtubule assembly and disassembly.

Dynamic stability and bifurcation analysis in fractional thermodynamics

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Béda, Péter B.

2018-02-01

In mechanics, viscoelasticity was the first field of applications in studying geomaterials. Further possibilities arise in spatial non-locality. Non-local materials were already studied in the 1960s by several authors as a part of continuum mechanics and are still in focus of interest because of the rising importance of materials with internal micro- and nano-structure. When material instability gained more interest, non-local behavior appeared in a different aspect. The problem was concerned to numerical analysis, because then instability zones exhibited singular properties for local constitutive equations. In dynamic stability analysis, mathematical aspects of non-locality were studied by using the theory of dynamic systems. There the basic set of equations describing the behavior of continua was transformed to an abstract dynamic system consisting of differential operators acting on the perturbation field variables. Such functions should satisfy homogeneous boundary conditions and act as indicators of stability of a selected state of the body under consideration. Dynamic systems approach results in conditions for cases, when the differential operators have critical eigenvalues of zero real parts (dynamic stability or instability conditions). When the critical eigenvalues have non-trivial eigenspace, the way of loss of stability is classified as a typical (or generic) bifurcation. Our experiences show that material non-locality and the generic nature of bifurcation at instability are connected, and the basic functions of the non-trivial eigenspace can be used to determine internal length quantities of non-local mechanics. Fractional calculus is already successfully used in thermo-elasticity. In the paper, non-locality is introduced via fractional strain into the constitutive relations of various conventional types. Then, by defining dynamic systems, stability and bifurcation are studied for states of thermo-mechanical solids. Stability conditions and genericity

KIF7 Controls the Proliferation of Cells of the Respiratory Airway through Distinct Microtubule Dependent Mechanisms.

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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.

Solar Dynamic Power System Stability Analysis and Control

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Momoh, James A.; Wang, Yanchun

1996-01-01

The objective of this research is to conduct dynamic analysis, control design, and control performance test of solar power system. Solar power system consists of generation system and distribution network system. A bench mark system is used in this research, which includes a generator with excitation system and governor, an ac/dc converter, six DDCU's and forty-eight loads. A detailed model is used for modeling generator. Excitation system is represented by a third order model. DDCU is represented by a seventh order system. The load is modeled by the combination of constant power and constant impedance. Eigen-analysis and eigen-sensitivity analysis are used for system dynamic analysis. The effects of excitation system, governor, ac/dc converter control, and the type of load on system stability are discussed. In order to improve system transient stability, nonlinear ac/dc converter control is introduced. The direct linearization method is used for control design. The dynamic analysis results show that these controls affect system stability in different ways. The parameter coordination of controllers are recommended based on the dynamic analysis. It is concluded from the present studies that system stability is improved by the coordination of control parameters and the nonlinear ac/dc converter control stabilize system oscillation caused by the load change and system fault efficiently.

Dynamic stabilization of imploding liquid metal liner

International Nuclear Information System (INIS)

Itoh, Yasuyuki; Fujiie, Yoichi

1979-01-01

The rotational stabilization has been proposed against the Rayleigh-Taylor instability of the imploding liquid metal liner. In this paper, the discussion is made on the possibility of the dynamic stabilization by applying the oscillating azimuthal magnetic field in addition to the axial field. In contrast to the rotational stabilization, the required (field) energy for this stabilization is also used for the liner driving or the plasma confinement. In the analysis, the liner subjected to the acceleration is assumed to be infinitely long, at rest and have the situation at the start of the implosion or turnaround. At turnaround, the existence of the plasma is taken into account. The perturbed motion of the liner is discussed with a linear stability analysis. Results are as follows: (1) The dynamic stabilization at the start of the implosion is possible if the distance from the conducting wall to the liner outer surface is comparable with or less than the liner thickness. (2) At turnaround, the stability is improved with decreasing the ratio of the plasma radius to that of the liner inner surface however the kink mode (m = 1) cannot be suppressed. (author)

Oscillatory fluid flow influences primary cilia and microtubule mechanics.

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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. Copyright © 2014 Wiley Periodicals, Inc.

ANALYSIS AND OPTIMISATION OF DYNAMIC STABILITY OF MOBILE WORKING MACHINES

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Peter BIGOŠ

2014-09-01

Full Text Available This paper describes an investigation of the dynamic stability, which is specified for the mobile working machines. There are presented the basic theoretical principles of the stability theory together with an introduction of two illustrative examples of the dynamic stability analysis.

Microtubules move the nucleus to quiescence.

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Laporte, Damien; Sagot, Isabelle

2014-01-01

The nucleus is a cellular compartment that hosts several macro-molecular machines displaying a highly complex spatial organization. This tight architectural orchestration determines not only DNA replication and repair but also regulates gene expression. In budding yeast microtubules play a key role in structuring the nucleus since they condition the Rabl arrangement in G1 and chromosome partitioning during mitosis through their attachment to centromeres via the kinetochore proteins. Recently, we have shown that upon quiescence entry, intranuclear microtubules emanating from the spindle pole body elongate to form a highly stable bundle that spans the entire nucleus. Here, we examine some molecular mechanisms that may underlie the formation of this structure. As the intranuclear microtubule bundle causes a profound re-organization of the yeast nucleus and is required for cell survival during quiescence, we discuss the possibility that the assembly of such a structure participates in quiescence establishment.

Quantitative analysis of microtubule orientation in interdigitated leaf pavement cells.

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Akita, Kae; Higaki, Takumi; Kutsuna, Natsumaro; Hasezawa, Seiichiro

2015-01-01

Leaf pavement cells are shaped like a jigsaw puzzle in most dicotyledon species. Molecular genetic studies have identified several genes required for pavement cells morphogenesis and proposed that microtubules play crucial roles in the interdigitation of pavement cells. In this study, we performed quantitative analysis of cortical microtubule orientation in leaf pavement cells in Arabidopsis thaliana. We captured confocal images of cortical microtubules in cotyledon leaf epidermis expressing GFP-tubulinβ and quantitatively evaluated the microtubule orientations relative to the pavement cell growth axis using original image processing techniques. Our results showed that microtubules kept parallel orientations to the growth axis during pavement cell growth. In addition, we showed that immersion treatment of seed cotyledons in solutions containing tubulin polymerization and depolymerization inhibitors decreased pavement cell complexity. Treatment with oryzalin and colchicine inhibited the symmetric division of guard mother cells.

Linking cortical microtubule attachment and exocytosis [version 1; referees: 2 approved

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Ivar Noordstra

2017-04-01

Full Text Available Exocytosis is a fundamental cellular process whereby secreted molecules are packaged into vesicles that move along cytoskeletal filaments and fuse with the plasma membrane. To function optimally, cells are strongly dependent on precisely controlled delivery of exocytotic cargo. In mammalian cells, microtubules serve as major tracks for vesicle transport by motor proteins, and thus microtubule organization is important for targeted delivery of secretory carriers. Over the years, multiple microtubule-associated and cortical proteins have been discovered that facilitate the interaction between the microtubule plus ends and the cell cortex. In this review, we focus on mammalian protein complexes that have been shown to participate in both cortical microtubule capture and exocytosis, thereby regulating the spatial organization of secretion. These complexes include microtubule plus-end tracking proteins, scaffolding factors, actin-binding proteins, and components of vesicle docking machinery, which together allow efficient coordination of cargo transport and release.

Lysosomes are associated with microtubules and not with intermediate filaments in cultured fibroblasts.

OpenAIRE

Collot, M; Louvard, D; Singer, S J

1984-01-01

Double immunofluorescent labeling experiments for lysosomes and either microtubules or vimentin intermediate filaments in cultured well-spread fibroblasts show a remarkable degree of superposition of the lysosomes and the microtubules. Under two different sets of conditions where the microtubules and intermediate filaments are well segregated from one another, the lysosomes remain codistributed with the microtubules. It is suggested that this specific association of lysosomes with microtubule...

Microtubule reorganization in tobacco BY-2 cells stably expressing GFP-MBD

Science.gov (United States)

Granger, C. L.; Cyr, R. J.

2000-01-01

Microtubule organization plays an important role in plant morphogenesis; however, little is known about how microtubule arrays transit from one organized state to another. The use of a genetically incorporated fluorescent marker would allow long-term observation of microtubule behavior in living cells. Here, we have characterized a Nicotiana tabacum L. cv. Bright Yellow 2 (BY-2) cell line that had been stably transformed with a gfp-mbd construct previously demonstrated to label microtubules (J. Marc et al., 1998, Plant Cell 10: 1927-1939). Fluorescence levels were low, but interphase and mitotic microtubule arrays, as well as the transitions between these arrays, could be observed in individual gfp-mbd-transformed cells. By comparing several attributes of transformed and untransformed cells it was concluded that the transgenic cells are not adversely affected by low-level expression of the transgene and that these cells will serve as a useful and accurate model system for observing microtubule reorganization in vivo. Indeed, some initial observations were made that are consistent with the involvement of motor proteins in the transition between the spindle and phragmoplast arrays. Our observations also support the role of the perinuclear region in nucleating microtubules at the end of cell division with a progressive shift of these microtubules and/or nucleating activity to the cortex to form the interphase cortical array.

Sensitivity of docetaxel-resistant MCF-7 breast cancer cells to microtubule-destabilizing agents including vinca alkaloids and colchicine-site binding agents.

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Richard C Wang

Full Text Available One of the main reasons for disease recurrence in the curative breast cancer treatment setting is the development of drug resistance. Microtubule targeted agents (MTAs are among the most commonly used drugs for the treatment of breaset cancer and therefore overcoming taxane resistance is of primary clinical importance. Our group has previously demonstrated that the microtubule dynamics of docetaxel-resistant MCF-7TXT cells are insensitivity to docetaxel due to the distinct expression profiles of β-tubulin isotypes in addition to the high expression of p-glycoprotein (ABCB1. In the present investigation we examined whether taxane-resistant breast cancer cells are more sensitive to microtubule destabilizing agents including vinca alkaloids and colchicine-site binding agents (CSBAs than the non-resistant cells.Two isogenic MCF-7 breast cancer cell lines were selected for resistance to docetaxel (MCF-7TXT and the wild type parental cell line (MCF-7CC to examine if taxane-resistant breast cancer cells are sensitive to microtubule-destabilizing agents including vinca alkaloids and CSBAs. Cytotoxicity assays, immunoblotting, indirect immunofluorescence and live imaging were used to study drug resistance, apoptosis, mitotic arrest, microtubule formation, and microtubule dynamics.MCF-7TXT cells were demonstrated to be cross resistant to vinca alkaloids, but were more sensitive to treatment with colchicine compared to parental non-resistant MCF-7CC cells. Cytotoxicity assays indicated that the IC50 of MCF-7TXT cell to vinorelbine and vinblastine was more than 6 and 3 times higher, respectively, than that of MCF-7CC cells. By contrast, the IC50 of MCF-7TXT cell for colchincine was 4 times lower than that of MCF-7CC cells. Indirect immunofluorescence showed that all MTAs induced the disorganization of microtubules and the chromatin morphology and interestingly each with a unique pattern. In terms of microtubule and chromain morphology, MCF-7TXT cells were

Microtubule's conformational cap

DEFF Research Database (Denmark)

Flyvbjerg, H.

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...

The centrosomal linker and microtubules provide dual levels of spatial coordination of centrosomes.

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Marko Panic

2015-05-01

Full Text Available The centrosome is the principal microtubule organizing center in most animal cells. It consists of a pair of centrioles surrounded by pericentriolar material. The centrosome, like DNA, duplicates exactly once per cell cycle. During interphase duplicated centrosomes remain closely linked by a proteinaceous linker. This centrosomal linker is composed of rootletin filaments that are anchored to the centrioles via the protein C-Nap1. At the onset of mitosis the linker is dissolved by Nek2A kinase to support the formation of the bipolar mitotic spindle. The importance of the centrosomal linker for cell function during interphase awaits characterization. Here we assessed the phenotype of human RPE1 C-Nap1 knockout (KO cells. The absence of the linker led to a modest increase in the average centrosome separation from 1 to 2.5 μm. This small impact on the degree of separation is indicative of a second level of spatial organization of centrosomes. Microtubule depolymerisation or stabilization in C-Nap1 KO cells dramatically increased the inter-centrosomal separation (> 8 μm. Thus, microtubules position centrosomes relatively close to one another in the absence of linker function. C-Nap1 KO cells had a Golgi organization defect with a two-fold expansion of the area occupied by the Golgi. When the centrosomes of C-Nap1 KO cells showed considerable separation, two spatially distinct Golgi stacks could be observed. Furthermore, migration of C-Nap1 KO cells was slower than their wild type RPE1 counterparts. These data show that the spatial organization of centrosomes is modulated by a combination of centrosomal cohesion and microtubule forces. Furthermore a modest increase in centrosome separation has major impact on Golgi organization and cell migration.

The nucleation of microtubules in Aspergillus nidulans germlings

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Cristina de Andrade-Monteiro

1999-09-01

Full Text Available Microtubules are filaments composed of dimers of alpha- and beta-tubulins, which have a variety of functions in living cells. In fungi, the spindle pole bodies usually have been considered to be microtubule-organizing centers. We used the antimicrotubule drug Benomyl in block/release experiments to depolymerize and repolymerize microtubules in Aspergillus nidulans germlings to learn more about the microtubule nucleation process in this filamentous fungus. Twenty seconds after release from Benomyl short microtubules were formed from several bright (immunofluorescent dots distributed along the germlings, suggesting that microtubule nucleation is randomly distributed in A. nidulans germlings. Since nuclear movement is dependent on microtubules in A. nidulans we analyzed whether mutants defective in nuclear distribution along the growing hyphae (nud mutants have some obvious microtubule defect. Cytoplasmic, astral and spindle microtubules were present and appeared to be normal in all nud mutants. However, significant changes in the percentage of short versus long mitotic spindles were observed in nud mutants. This suggests that some of the nuclei of nud mutants do not reach the late stage of cell division at normal temperatures.Microtúbulos são filamentos compostos por dímeros das tubulinas a e b e têm uma variedade de funções nas células vivas. Em fungos, os corpúsculos polares dos fusos são geralmente considerados os centros organizadores dos microtúbulos. Com o objetivo de contribuir para uma melhor compreensão dos processos de nucleação dos microtúbulos no fungo filamentoso A. nidulans, nós utilizamos a droga antimicrotúbulo Benomil em experimentos de bloqueio e liberação para depolimerizar e repolimerizar os microtúbulos. Após 20 segundos de reincubação em meio sem Benomil, pequenos microtúbulos foram formados a partir de pontos distribuídos pela célula, sugerindo que os pontos de nucleação de microtúbulos s

Curcumin alters the cytoskeleton and microtubule organization on trophozoites of Giardia lamblia.

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Gutiérrez-Gutiérrez, Filiberto; Palomo-Ligas, Lissethe; Hernández-Hernández, José Manuel; Pérez-Rangel, Armando; Aguayo-Ortiz, Rodrigo; Hernández-Campos, Alicia; Castillo, Rafael; González-Pozos, Sirenia; Cortés-Zárate, Rafael; Ramírez-Herrera, Mario Alberto; Mendoza-Magaña, María Luisa; Castillo-Romero, Araceli

2017-08-01

Giardia lamblia is a worldwide protozoan responsible for a significant number of intestinal infections. There are several drugs for the treatment of giardiasis, but they often cause side effects. Curcumin, a component of turmeric, has antigiardial activity; however, the molecular target and mechanism of antiproliferative activity are not clear. The effects of curcumin on cellular microtubules have been widely investigated. Since tubulin is the most abundant protein in the cytoskeleton of Giardia, to elucidate whether curcumin has activity against the microtubules of this parasite, we treated trophozoites with curcumin and the cells were analyzed by scanning electron microscopy and confocal microscopy. Curcumin inhibited Giardia proliferation and adhesion in a time-concentration-dependent mode. The higher inhibitory concentrations of curcumin (3 and 15μM) disrupted the cytoskeletal structures of trophozoites; the damage was evident on the ventral disk, flagella and in the caudal region, also the membrane was affected. The immunofluorescence images showed altered distribution of tubulin staining on ventral disk and flagella. Additionally, we found that curcumin caused a clear reduction of tubulin expression. By docking analysis and molecular dynamics we showed that curcumin has a high probability to bind at the interface of the tubulin dimer close to the vinblastine binding site. All the data presented indicate that curcumin may inhibit Giardia proliferation by perturbing microtubules. Copyright © 2017. Published by Elsevier B.V.

Calcium-dependent depletion zones in the cortical microtubule array coincide with sites of, but do not regulate, wall ingrowth papillae deposition in epidermal transfer cells

Science.gov (United States)

Zhang, Hui-ming; Talbot, Mark J.; McCurdy, David W.; Patrick, John W.; Offler, Christina E.

2015-01-01

Trans-differentiation to a transfer-cell morphology is characterized by the localized deposition of wall ingrowth papillae that protrude into the cytosol. Whether the cortical microtubule array directs wall ingrowth papillae formation was investigated using a Vicia faba cotyledon culture system in which their adaxial epidermal cells were spontaneously induced to trans-differentiate to transfer cells. During deposition of wall ingrowth papillae, the aligned cortical microtubule arrays in precursor epidermal cells were reorganized into a randomized array characterized by circular depletion zones. Concurrence of the temporal appearance, spatial pattern, and size of depletion zones and wall ingrowth papillae was consistent with each papilla occupying a depletion zone. Surprisingly, microtubules appeared not to regulate construction of wall ingrowth papillae, as neither depolymerization nor stabilization of cortical microtubules changed their deposition pattern or morphology. Moreover, the size and spatial pattern of depletion zones was unaltered when the formation of wall ingrowth papillae was blocked by inhibiting cellulose biosynthesis. In contrast, the depletion zones were absent when the cytosolic calcium plumes, responsible for directing wall ingrowth papillae formation, were blocked or dissipated. Thus, we conclude that the depletion zones within the cortical microtubule array result from localized depolymerization of microtubules initiated by elevated cytosolic Ca2+ levels at loci where wall ingrowth papillae are deposited. The physiological significance of the depletion zones as a mechanism to accommodate the construction of wall ingrowth papillae without compromising maintenance of the plasma membrane–microtubule inter-relationship is discussed. PMID:26136268

Measurement of in vitro microtubule polymerization by turbidity and fluorescence.

Science.gov (United States)

Mirigian, Matthew; Mukherjee, Kamalika; Bane, Susan L; Sackett, Dan L

2013-01-01

Tubulin polymerization may be conveniently monitored by the increase in turbidity (optical density, or OD) or by the increase in fluorescence intensity of diamidino-phenylindole. The resulting data can be a quantitative measure of microtubule (MT) assembly, but some care is needed in interpretation, especially of OD data. Buffer formulations used for the assembly reaction significantly influence the polymerization, both by altering the critical concentration for polymerization and by altering the exact polymer produced-for example, by increasing the production of sheet polymers in addition to MT. Both the turbidity and the fluorescence methods are useful for demonstrating the effect of MT-stabilizing or -destabilizing additives. 2013 Published by Elsevier Inc.

Dynamic stability under sudden loads

International Nuclear Information System (INIS)

Simitses, G.J.

1998-01-01

The concept of dynamic stability of elastic structures subjected to sudden (step) loads is discussed. The various criteria and related methodologies for estimating critical conditions are presented with the emphasis on their similarities and differences. These are demonstrated by employing a simple mechanical model. Several structural configurations are analyzed, for demonstration purposes, with the intention of comparing critical dynamic loads to critical static loads. These configurations include shallow arches and shallow spherical caps, two bar frames, and imperfect cylindrical shells of metallic as well as laminated composite construction. In the demonstration examples, the effect of static pre loading on the dynamic critical load is presented

A Novel Plasma Membrane-Anchored Protein Regulates Xylem Cell-Wall Deposition through Microtubule-Dependent Lateral Inhibition of Rho GTPase Domains.

Science.gov (United States)

Sugiyama, Yuki; Wakazaki, Mayumi; Toyooka, Kiminori; Fukuda, Hiroo; Oda, Yoshihisa

2017-08-21

Spatial control of cell-wall deposition is essential for determining plant cell shape [1]. Rho-type GTPases, together with the cortical cytoskeleton, play central roles in regulating cell-wall patterning [2]. In metaxylem vessel cells, which are the major components of xylem tissues, active ROP11 Rho GTPases form oval plasma membrane domains that locally disrupt cortical microtubules, thereby directing the formation of oval pits in secondary cell walls [3-5]. However, the regulatory mechanism that determines the planar shape of active Rho of Plants (ROP) domains is still unknown. Here we show that IQD13 associates with cortical microtubules and the plasma membrane to laterally restrict the localization of ROP GTPase domains, thereby directing the formation of oval secondary cell-wall pits. Loss and overexpression of IQD13 led to the formation of abnormally round and narrow secondary cell-wall pits, respectively. Ectopically expressed IQD13 increased the presence of parallel cortical microtubules by promoting microtubule rescue. A reconstructive approach revealed that IQD13 confines the area of active ROP domains within the lattice of the cortical microtubules, causing narrow ROP domains to form. This activity required the interaction of IQD13 with the plasma membrane. These findings suggest that IQD13 positively regulates microtubule dynamics as well as their linkage to the plasma membrane, which synergistically confines the area of active ROP domains, leading to the formation of oval secondary cell-wall pits. This finding sheds light on the role of microtubule-plasma membrane linkage as a lateral fence that determines the planar shape of Rho GTPase domains. Copyright © 2017 Elsevier Ltd. All rights reserved.

Chlorpyrifos, chlorpyrifos-oxon, and diisopropylfluorophosphate inhibit kinesin-dependent microtubule motility

International Nuclear Information System (INIS)

Gearhart, Debra A.; Sickles, Dale W.; Buccafusco, Jerry J.; Prendergast, Mark A.; Terry, Alvin V.

2007-01-01

Diisopropylfluorophosphate, originally developed as a chemical warfare agent, is structurally similar to nerve agents, and chlorpyrifos has extensive worldwide use as an agricultural pesticide. While inhibition of cholinesterases underlies the acute toxicity of these organophosphates, we previously reported impaired axonal transport in the sciatic nerves from rats treated chronically with subthreshold doses of chlorpyrifos. Those data indicate that chlorpyrifos (and/or its active metabolite, chlorpyrifos-oxon) might directly affect the function of kinesin and/or microtubules-the principal proteins that mediate anterograde axonal transport. The current report describes in vitro assays to assess the concentration-dependent effects of chlorpyrifos (0-10 μM), chlorpyrifos-oxon (0-10 μM), and diisopropylfluorophosphate (0-0.59 nM) on kinesin-dependent microtubule motility. Preincubating bovine brain microtubules with the organophosphates did not alter kinesin-mediated microtubule motility. In contrast, preincubation of bovine brain kinesin with diisopropylfluorophosphate, chlorpyrifos, or chlorpyrifos-oxon produced a concentration-dependent increase in the number of locomoting microtubules that detached from the kinesin-coated glass cover slip. Our data suggest that the organophosphates-chlorpyrifos-oxon, chlorpyrifos, and diisopropylfluorophosphate-directly affect kinesin, thereby disrupting kinesin-dependent transport on microtubules. Kinesin-dependent movement of vesicles, organelles, and other cellular components along microtubules is fundamental to the organization of all eukaryotic cells, especially in neurons where organelles and proteins synthesized in the cell body must move down long axons to pre-synaptic sites in nerve terminals. We postulate that disruption of kinesin-dependent intracellular transport could account for some of the long-term effects of organophosphates on the peripheral and central nervous system

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

2016-01-01

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. PMID:27512034

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.

Dynamic postural stability during advancing pregnancy.

Science.gov (United States)

McCrory, J L; Chambers, A J; Daftary, A; Redfern, M S

2010-08-26

Pregnant women are at an increased risk of experiencing a fall. Numerous anatomical, physiological, and hormonal alterations occur during pregnancy, but the influence of these factors on dynamic postural stability has not been explored. The purpose of this study was to examine dynamic postural stability in pregnant women during their second and third trimesters as well as in a group of non-pregnant control women. Eighty-one women (41 pregnant, 40 controls) participated stood on a force plate that translated anteroposteriorly at small, medium, and large magnitudes. Reaction time and center of pressure (COP) movement during the translations were analyzed. Trimester, perturbation direction, and perturbation magnitude were the independent variables in a mixed-model analysis of variance on each of the following dependent variables: reaction time, initial sway, total sway, and sway velocity. Reaction time to the perturbation was not significantly different between the groups. Initial sway, total sway, and sway velocity were significantly less during the third trimester than during the second trimester and when compared to the non-pregnant controls (Ppostural stability. 2010 Elsevier Ltd. All rights reserved.

Cellular effects of curcumin on Plasmodium falciparum include disruption of microtubules.

Directory of Open Access Journals (Sweden)

Rimi Chakrabarti

Full Text Available Curcumin has been widely investigated for its myriad cellular effects resulting in reduced proliferation of various eukaryotic cells including cancer cells and the human malaria parasite Plasmodium falciparum. Studies with human cancer cell lines HT-29, Caco-2, and MCF-7 suggest that curcumin can bind to tubulin and induce alterations in microtubule structure. Based on this finding, we investigated whether curcumin has any effect on P. falciparum microtubules, considering that mammalian and parasite tubulin are 83% identical. IC50 of curcumin was found to be 5 µM as compared to 20 µM reported before. Immunofluorescence images of parasites treated with 5 or 20 µM curcumin showed a concentration-dependent effect on parasite microtubules resulting in diffuse staining contrasting with the discrete hemispindles and subpellicular microtubules observed in untreated parasites. The effect on P. falciparum microtubules was evident only in the second cycle for both concentrations tested. This diffuse pattern of tubulin fluorescence in curcumin treated parasites was similar to the effect of a microtubule destabilizing drug vinblastine on P. falciparum. Molecular docking predicted the binding site of curcumin at the interface of alpha and beta tubulin, similar to another destabilizing drug colchicine. Data from predicted drug binding is supported by results from drug combination assays showing antagonistic interactions between curcumin and colchicine, sharing a similar binding site, and additive/synergistic interactions of curcumin with paclitaxel and vinblastine, having different binding sites. This evidence suggests that cellular effects of curcumin are at least, in part, due to its perturbing effect on P. falciparum microtubules. The action of curcumin, both direct and indirect, on P. falciparum microtubules is discussed.

Autocatalytic microtubule nucleation determines the size and mass of Xenopus laevis egg extract spindles.

Science.gov (United States)

Decker, Franziska; Oriola, David; Dalton, Benjamin; Brugués, Jan

2018-01-11

Regulation of size and growth is a fundamental problem in biology. A prominent example is the formation of the mitotic spindle, where protein concentration gradients around chromosomes are thought to regulate spindle growth by controlling microtubule nucleation. Previous evidence suggests that microtubules nucleate throughout the spindle structure. However, the mechanisms underlying microtubule nucleation and its spatial regulation are still unclear. Here, we developed an assay based on laser ablation to directly probe microtubule nucleation events in Xenopus laevis egg extracts. Combining this method with theory and quantitative microscopy, we show that the size of a spindle is controlled by autocatalytic growth of microtubules, driven by microtubule-stimulated microtubule nucleation. The autocatalytic activity of this nucleation system is spatially regulated by the limiting amounts of active microtubule nucleators, which decrease with distance from the chromosomes. This mechanism provides an upper limit to spindle size even when resources are not limiting. © 2018, Decker et al.

GIT1/βPIX signaling proteins and PAK1 kinase regulate microtubule nucleation.

Science.gov (United States)

Černohorská, Markéta; Sulimenko, Vadym; Hájková, Zuzana; Sulimenko, Tetyana; Sládková, Vladimíra; Vinopal, Stanislav; Dráberová, Eduarda; Dráber, Pavel

2016-06-01

Microtubule nucleation from γ-tubulin complexes, located at the centrosome, is an essential step in the formation of the microtubule cytoskeleton. However, the signaling mechanisms that regulate microtubule nucleation in interphase cells are largely unknown. In this study, we report that γ-tubulin is in complexes containing G protein-coupled receptor kinase-interacting protein 1 (GIT1), p21-activated kinase interacting exchange factor (βPIX), and p21 protein (Cdc42/Rac)-activated kinase 1 (PAK1) in various cell lines. Immunofluorescence microscopy revealed association of GIT1, βPIX and activated PAK1 with centrosomes. Microtubule regrowth experiments showed that depletion of βPIX stimulated microtubule nucleation, while depletion of GIT1 or PAK1 resulted in decreased nucleation in the interphase cells. These data were confirmed for GIT1 and βPIX by phenotypic rescue experiments, and counting of new microtubules emanating from centrosomes during the microtubule regrowth. The importance of PAK1 for microtubule nucleation was corroborated by the inhibition of its kinase activity with IPA-3 inhibitor. GIT1 with PAK1 thus represent positive regulators, and βPIX is a negative regulator of microtubule nucleation from the interphase centrosomes. The regulatory roles of GIT1, βPIX and PAK1 in microtubule nucleation correlated with recruitment of γ-tubulin to the centrosome. Furthermore, in vitro kinase assays showed that GIT1 and βPIX, but not γ-tubulin, serve as substrates for PAK1. Finally, direct interaction of γ-tubulin with the C-terminal domain of βPIX and the N-terminal domain of GIT1, which targets this protein to the centrosome, was determined by pull-down experiments. We propose that GIT1/βPIX signaling proteins with PAK1 kinase represent a novel regulatory mechanism of microtubule nucleation in interphase cells. Copyright © 2016 Elsevier B.V. All rights reserved.

Calcium-dependent depletion zones in the cortical microtubule array coincide with sites of, but do not regulate, wall ingrowth papillae deposition in epidermal transfer cells.

Science.gov (United States)

Zhang, Hui-ming; Talbot, Mark J; McCurdy, David W; Patrick, John W; Offler, Christina E

2015-09-01

Trans-differentiation to a transfer-cell morphology is characterized by the localized deposition of wall ingrowth papillae that protrude into the cytosol. Whether the cortical microtubule array directs wall ingrowth papillae formation was investigated using a Vicia faba cotyledon culture system in which their adaxial epidermal cells were spontaneously induced to trans-differentiate to transfer cells. During deposition of wall ingrowth papillae, the aligned cortical microtubule arrays in precursor epidermal cells were reorganized into a randomized array characterized by circular depletion zones. Concurrence of the temporal appearance, spatial pattern, and size of depletion zones and wall ingrowth papillae was consistent with each papilla occupying a depletion zone. Surprisingly, microtubules appeared not to regulate construction of wall ingrowth papillae, as neither depolymerization nor stabilization of cortical microtubules changed their deposition pattern or morphology. Moreover, the size and spatial pattern of depletion zones was unaltered when the formation of wall ingrowth papillae was blocked by inhibiting cellulose biosynthesis. In contrast, the depletion zones were absent when the cytosolic calcium plumes, responsible for directing wall ingrowth papillae formation, were blocked or dissipated. Thus, we conclude that the depletion zones within the cortical microtubule array result from localized depolymerization of microtubules initiated by elevated cytosolic Ca(2+) levels at loci where wall ingrowth papillae are deposited. The physiological significance of the depletion zones as a mechanism to accommodate the construction of wall ingrowth papillae without compromising maintenance of the plasma membrane-microtubule inter-relationship is discussed. © The Author 2015. Published by Oxford University Press on behalf of the Society for Experimental Biology.

Measuring and modeling polymer concentration profiles near spindle boundaries argues that spindle microtubules regulate their own nucleation

Science.gov (United States)

Kaye, Bryan; Stiehl, Olivia; Foster, Peter J.; Shelley, Michael J.; Needleman, Daniel J.; Fürthauer, Sebastian

2018-05-01

Spindles are self-organized microtubule-based structures that segregate chromosomes during cell division. The mass of the spindle is controlled by the balance between microtubule turnover and nucleation. The mechanisms that control the spatial regulation of microtubule nucleation remain poorly understood. While previous work found that microtubule nucleators bind to pre-existing microtubules in the spindle, it is still unclear whether this binding regulates the activity of those nucleators. Here we use a combination of experiments and mathematical modeling to investigate this issue. We measured the concentration of microtubules and soluble tubulin in and around the spindle. We found a very sharp decay in the concentration of microtubules at the spindle interface. This is inconsistent with a model in which the activity of nucleators is independent of their association with microtubules but consistent with a model in which microtubule nucleators are only active when bound to pre-existing microtubules. This argues that the activity of microtubule nucleators is greatly enhanced when bound to pre-existing microtubules. Thus, microtubule nucleators are both localized and activated by the microtubules they generate.

Dynamic-Stability Characteristics of Premixed Methane Oxy-Combustion

KAUST Repository

Shroll, Andrew P.; Shanbhogue, Santosh J.; Ghoniem, Ahmed F.

2012-01-01

This work explores the dynamic stability characteristics of premixed CH 4/O 2/CO 2 mixtures in a 50 kW swirl stabilized combustor. In all cases, the methane-oxygen mixture is stoichiometric, with different dilution levels of carbon dioxide used

Rho-associated coiled-coil kinase (ROCK) protein controls microtubule dynamics in a novel signaling pathway that regulates cell migration.

Science.gov (United States)

Schofield, Alice V; Steel, Rohan; Bernard, Ora

2012-12-21

The two members of the Rho-associated coiled-coil kinase (ROCK1 and 2) family are established regulators of actin dynamics that are involved in the regulation of the cell cycle as well as cell motility and invasion. Here, we discovered a novel signaling pathway whereby ROCK regulates microtubule (MT) acetylation via phosphorylation of the tubulin polymerization promoting protein 1 (TPPP1/p25). We show that ROCK phosphorylation of TPPP1 inhibits the interaction between TPPP1 and histone deacetylase 6 (HDAC6), which in turn results in increased HDAC6 activity followed by a decrease in MT acetylation. As a consequence, we show that TPPP1 phosphorylation by ROCK increases cell migration and invasion via modulation of cellular acetyl MT levels. We establish here that the ROCK-TPPP1-HDAC6 signaling pathway is important for the regulation of cell migration and invasion.

The free energy profile of tubulin straight-bent conformational changes, with implications for microtubule assembly and drug discovery.

Directory of Open Access Journals (Sweden)

Lili X Peng

2014-02-01

Full Text Available αβ-tubulin dimers need to convert between a 'bent' conformation observed for free dimers in solution and a 'straight' conformation required for incorporation into the microtubule lattice. Here, we investigate the free energy landscape of αβ-tubulin using molecular dynamics simulations, emphasizing implications for models of assembly, and modulation of the conformational landscape by colchicine, a tubulin-binding drug that inhibits microtubule polymerization. Specifically, we performed molecular dynamics, potential-of-mean force simulations to obtain the free energy profile for unpolymerized GDP-bound tubulin as a function of the ∼12° intradimer rotation differentiating the straight and bent conformers. Our results predict that the unassembled GDP-tubulin heterodimer exists in a continuum of conformations ranging between straight and bent, but, in agreement with existing structural data, suggests that an intermediate bent state has a lower free energy (by ∼1 kcal/mol and thus dominates in solution. In agreement with predictions of the lattice model of microtubule assembly, lateral binding of two αβ-tubulins strongly shifts the conformational equilibrium towards the straight state, which is then ∼1 kcal/mol lower in free energy than the bent state. Finally, calculations of colchicine binding to a single αβ-tubulin dimer strongly shifts the equilibrium toward the bent states, and disfavors the straight state to the extent that it is no longer thermodynamically populated.

Aggregation of SND1 in Stress Granules is Associated with the Microtubule Cytoskeleton During Heat Shock Stimulus.

Science.gov (United States)

Shao, Jie; Gao, Fei; Zhang, Bingbing; Zhao, Meng; Zhou, Yunli; He, Jinyan; Ren, Li; Yao, Zhi; Yang, Jie; Su, Chao; Gao, Xingjie

2017-12-01

Stress granules (SGs) are dynamic dense structures in the cytoplasm that form in response to a variety of environmental stress stimuli. Staphylococcal nuclease and Tudor domain containing 1 (SND1) is a type of RNA-binding protein and has been identified as a transcriptional co-activator. Our previous studies have shown that SND1 is a component of the stress granule, which forms under stress conditions. Here, we observed that SND1 granules were often surrounded by ɑ-tubulin-microtubules in 45°C-treated HeLa cells at 15 min or colocalized with microtubules at 30 or 45 min. Furthermore, Nocodazole-mediated microtubule depolymerization could significantly affect the efficient recruitment of SND1 proteins to the SGs during heat shock stress. In addition, the 45°C heat shock mediated the enhancement of eIF2α phosphorylation, which was not affected by treatment with Nocodazole, an agent that disrupts the cytoskeleton. The intact microtubule cytoskeletal tracks are important for the efficient assembly of SND1 granules under heat shock stress and may facilitate SND1 shuttling between cytoplasmic RNA foci. Anat Rec, 300:2192-2199, 2017. © 2017 The Authors The Anatomical Record published by Wiley Periodicals, Inc. on behalf of American Association of Anatomists. Copyright © 2017 The Authors The Anatomical Record published by Wiley Periodicals, Inc. on behalf of American Association of Anatomists.

A direct interaction between leucine-rich repeat kinase 2 and specific β-tubulin isoforms regulates tubulin acetylation.

Science.gov (United States)

Law, Bernard M H; Spain, Victoria A; Leinster, Veronica H L; Chia, Ruth; Beilina, Alexandra; Cho, Hyun J; Taymans, Jean-Marc; Urban, Mary K; Sancho, Rosa M; Blanca Ramírez, Marian; Biskup, Saskia; Baekelandt, Veerle; Cai, Huaibin; Cookson, Mark R; Berwick, Daniel C; Harvey, Kirsten

2014-01-10

Mutations in LRRK2, encoding the multifunctional protein leucine-rich repeat kinase 2 (LRRK2), are a common cause of Parkinson disease. LRRK2 has been suggested to influence the cytoskeleton as LRRK2 mutants reduce neurite outgrowth and cause an accumulation of hyperphosphorylated Tau. This might cause alterations in the dynamic instability of microtubules suggested to contribute to the pathogenesis of Parkinson disease. Here, we describe a direct interaction between LRRK2 and β-tubulin. This interaction is conferred by the LRRK2 Roc domain and is disrupted by the familial R1441G mutation and artificial Roc domain mutations that mimic autophosphorylation. LRRK2 selectively interacts with three β-tubulin isoforms: TUBB, TUBB4, and TUBB6, one of which (TUBB4) is mutated in the movement disorder dystonia type 4 (DYT4). Binding specificity is determined by lysine 362 and alanine 364 of β-tubulin. Molecular modeling was used to map the interaction surface to the luminal face of microtubule protofibrils in close proximity to the lysine 40 acetylation site in α-tubulin. This location is predicted to be poorly accessible within mature stabilized microtubules, but exposed in dynamic microtubule populations. Consistent with this finding, endogenous LRRK2 displays a preferential localization to dynamic microtubules within growth cones, rather than adjacent axonal microtubule bundles. This interaction is functionally relevant to microtubule dynamics, as mouse embryonic fibroblasts derived from LRRK2 knock-out mice display increased microtubule acetylation. Taken together, our data shed light on the nature of the LRRK2-tubulin interaction, and indicate that alterations in microtubule stability caused by changes in LRRK2 might contribute to the pathogenesis of Parkinson disease.

Does dynamic stability govern propulsive force generation in human walking?

Science.gov (United States)

Browne, Michael G; Franz, Jason R

2017-11-01

Before succumbing to slower speeds, older adults may walk with a diminished push-off to prioritize stability over mobility. However, direct evidence for trade-offs between push-off intensity and balance control in human walking, independent of changes in speed, has remained elusive. As a critical first step, we conducted two experiments to investigate: (i) the independent effects of walking speed and propulsive force ( F P ) generation on dynamic stability in young adults, and (ii) the extent to which young adults prioritize dynamic stability in selecting their preferred combination of walking speed and F P generation. Subjects walked on a force-measuring treadmill across a range of speeds as well as at constant speeds while modulating their F P according to a visual biofeedback paradigm based on real-time force measurements. In contrast to improvements when walking slower, walking with a diminished push-off worsened dynamic stability by up to 32%. Rather, we find that young adults adopt an F P at their preferred walking speed that maximizes dynamic stability. One implication of these findings is that the onset of a diminished push-off in old age may independently contribute to poorer balance control and precipitate slower walking speeds.

Activation of Ran GTPase by a Legionella effector promotes microtubule polymerization, pathogen vacuole motility and infection.

Directory of Open Access Journals (Sweden)

Eva Rothmeier

2013-09-01

Full Text Available The causative agent of Legionnaires' disease, Legionella pneumophila, uses the Icm/Dot type IV secretion system (T4SS to form in phagocytes a distinct "Legionella-containing vacuole" (LCV, which intercepts endosomal and secretory vesicle trafficking. Proteomics revealed the presence of the small GTPase Ran and its effector RanBP1 on purified LCVs. Here we validate that Ran and RanBP1 localize to LCVs and promote intracellular growth of L. pneumophila. Moreover, the L. pneumophila protein LegG1, which contains putative RCC1 Ran guanine nucleotide exchange factor (GEF domains, accumulates on LCVs in an Icm/Dot-dependent manner. L. pneumophila wild-type bacteria, but not strains lacking LegG1 or a functional Icm/Dot T4SS, activate Ran on LCVs, while purified LegG1 produces active Ran(GTP in cell lysates. L. pneumophila lacking legG1 is compromised for intracellular growth in macrophages and amoebae, yet is as cytotoxic as the wild-type strain. A downstream effect of LegG1 is to stabilize microtubules, as revealed by conventional and stimulated emission depletion (STED fluorescence microscopy, subcellular fractionation and Western blot, or by microbial microinjection through the T3SS of a Yersinia strain lacking endogenous effectors. Real-time fluorescence imaging indicates that LCVs harboring wild-type L. pneumophila rapidly move along microtubules, while LCVs harboring ΔlegG1 mutant bacteria are stalled. Together, our results demonstrate that Ran activation and RanBP1 promote LCV formation, and the Icm/Dot substrate LegG1 functions as a bacterial Ran activator, which localizes to LCVs and promotes microtubule stabilization, LCV motility as well as intracellular replication of L. pneumophila.

Activation of Ran GTPase by a Legionella Effector Promotes Microtubule Polymerization, Pathogen Vacuole Motility and Infection

Science.gov (United States)

Rothmeier, Eva; Pfaffinger, Gudrun; Hoffmann, Christine; Harrison, Christopher F.; Grabmayr, Heinrich; Repnik, Urska; Hannemann, Mandy; Wölke, Stefan; Bausch, Andreas; Griffiths, Gareth; Müller-Taubenberger, Annette; Itzen, Aymelt; Hilbi, Hubert

2013-01-01

The causative agent of Legionnaires' disease, Legionella pneumophila, uses the Icm/Dot type IV secretion system (T4SS) to form in phagocytes a distinct “Legionella-containing vacuole” (LCV), which intercepts endosomal and secretory vesicle trafficking. Proteomics revealed the presence of the small GTPase Ran and its effector RanBP1 on purified LCVs. Here we validate that Ran and RanBP1 localize to LCVs and promote intracellular growth of L. pneumophila. Moreover, the L. pneumophila protein LegG1, which contains putative RCC1 Ran guanine nucleotide exchange factor (GEF) domains, accumulates on LCVs in an Icm/Dot-dependent manner. L. pneumophila wild-type bacteria, but not strains lacking LegG1 or a functional Icm/Dot T4SS, activate Ran on LCVs, while purified LegG1 produces active Ran(GTP) in cell lysates. L. pneumophila lacking legG1 is compromised for intracellular growth in macrophages and amoebae, yet is as cytotoxic as the wild-type strain. A downstream effect of LegG1 is to stabilize microtubules, as revealed by conventional and stimulated emission depletion (STED) fluorescence microscopy, subcellular fractionation and Western blot, or by microbial microinjection through the T3SS of a Yersinia strain lacking endogenous effectors. Real-time fluorescence imaging indicates that LCVs harboring wild-type L. pneumophila rapidly move along microtubules, while LCVs harboring ΔlegG1 mutant bacteria are stalled. Together, our results demonstrate that Ran activation and RanBP1 promote LCV formation, and the Icm/Dot substrate LegG1 functions as a bacterial Ran activator, which localizes to LCVs and promotes microtubule stabilization, LCV motility as well as intracellular replication of L. pneumophila. PMID:24068924

On atmospheric stability in the dynamic wake meandering model

DEFF Research Database (Denmark)

Keck, Rolf-Erik; de Mare, Martin Tobias; Churchfield, Matthew J.

2014-01-01

The present study investigates a new approach for capturing the effects of atmospheric stability on wind turbine wake evolution and wake meandering by using the dynamic wake meandering model. The most notable impact of atmospheric stability on the wind is the changes in length and velocity scales...... spectra and applied to the dynamic wake meandering model to capture the correct wake meandering behaviour. The ambient turbulence in all stability classes is generated using the Mann turbulence model, where the effects of non-neutral atmospheric stability are approximated by the selection of input...... in the computational domain. The changes in the turbulent length scales due to the various atmospheric stability states impact the wake meandering characteristics and thus the power generation by the individual turbines. The proposed method is compared with results from both large-eddy simulation coupled...

Short Linear Sequence Motif LxxPTPh Targets Diverse Proteins to Growing Microtubule Ends

NARCIS (Netherlands)

Kumar, Anil; Manatschal, Cristina; Rai, Ankit; Grigoriev, Ilya; Degen, Miriam Steiner; Jaussi, Rolf; Kretzschmar, Ines; Prota, Andrea E; Volkmer, Rudolf; Kammerer, Richard A.; Akhmanova, Anna; Steinmetz, Michel O.

2017-01-01

Microtubule plus-end tracking proteins (+TIPs) are involved in virtually all microtubule-based processes. End-binding (EB) proteins are considered master regulators of +TIP interaction networks, since they autonomously track growing microtubule ends and recruit a plethora of proteins to this

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.

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.

Template-free electrosynthesis of aligned poly(p-phenylene) microtubules

Institute of Scientific and Technical Information of China (English)

无

2003-01-01

Poly(p-phenylene) (PPP) microtubules with diameters of 0.2-0.8μm and lengths of～10 (m have been synthesized by direct oxidation of benzene in the mixed electrolyte of boron trifluoride diethyl etherate (BFEE) and trifluoroacetic acid (TFA) (BFEE:TFA= 2:1, by volume), containing a certain amount of sodium dodecylbenzene- sulfonate (SDBS) as surfactant. The microtubules were grown vertically on the working electrode surface. The tubular morphology has been confirmed by scanning and transmission electron microscopies and the chain structure of the skin of the tubules has been characterized by Raman spectroscopy. The electrode property, monomer/surfactant molar ratio and the value of applied potential have strong effects on the morphology of the microtubules.

Collective behavior of minus-ended motors in mitotic microtubule asters gliding toward DNA

International Nuclear Information System (INIS)

Athale, Chaitanya A; Dinarina, Ana; Nedelec, Francois; Karsenti, Eric

2014-01-01

Microtubules (MTs) nucleated by centrosomes form star-shaped structures referred to as asters. Aster motility and dynamics is vital for genome stability, cell division, polarization and differentiation. Asters move either toward the cell center or away from it. Here, we focus on the centering mechanism in a membrane independent system of Xenopus cytoplasmic egg extracts. Using live microscopy and single particle tracking, we find that asters move toward chromatinized DNA structures. The velocity and directionality profiles suggest a random-walk with drift directed toward DNA. We have developed a theoretical model that can explain this movement as a result of a gradient of MT length dynamics and MT gliding on immobilized dynein motors. In simulations, the antagonistic action of the motor species on the radial array of MTs leads to a tug-of-war purely due to geometric considerations and aster motility resembles a directed random-walk. Additionally, our model predicts that aster velocities do not change greatly with varying initial distance from DNA. The movement of asymmetric asters becomes increasingly super-diffusive with increasing motor density, but for symmetric asters it becomes less super-diffusive. The transition of symmetric asters from superdiffusive to diffusive mobility is the result of number fluctuations in bound motors in the tug-of-war. Overall, our model is in good agreement with experimental data in Xenopus cytoplasmic extracts and predicts novel features of the collective effects of motor-MT interactions. (paper)

Navier-Stokes Predictions of Dynamic Stability Derivatives: Evaluation of Steady-State Methods

National Research Council Canada - National Science Library

DeSpirito, James; Silton, Sidra I; Weinacht, Paul

2008-01-01

The prediction of the dynamic stability derivatives-roll-damping, Magnus, and pitch-damping moments-were evaluated for three spin-stabilized projectiles using steady-state computational fluid dynamic (CFD) calculations...

Arabidopsis Microtubule-Associated Protein MAP65-3 Cross-Links Antiparallel Microtubules toward Their Plus Ends in the Phragmoplast via Its Distinct C-Terminal Microtubule Binding Domain[W

Science.gov (United States)

Ho, Chin-Min Kimmy; Lee, Yuh-Ru Julie; Kiyama, Lindsay D.; Dinesh-Kumar, Savithramma P.; Liu, Bo

2012-01-01

Plant cytokinesis is brought about by the phragmoplast, which contains an antiparallel microtubule (MT) array. The MT-associated protein MAP65-3 acts as an MT-bundling factor that specifically cross-links antiparallel MTs near their plus ends. MAP65 family proteins contain an N-terminal dimerization domain and C-terminal MT interaction domain. Compared with other MAP65 isoforms, MAP65-3 contains an extended C terminus. A MT binding site was discovered in the region between amino acids 496 and 588 and found to be essential for the organization of phragmoplast MTs. The frequent cytokinetic failure caused by loss of MAP65-3 was not rescued by ectopic expression of MAP65-1 under the control of the MAP65-3 promoter, indicating nonoverlapping functions between the two isoforms. In the presence of MAP65-3, however, ectopic MAP65-1 appeared in the phragmoplast midline. We show that MAP65-1 could acquire the function of MAP65-3 when the C terminus of MAP65-3, which contains the MT binding site, was grafted to it. Our results also show that MAP65-1 and MAP65-3 may share redundant functions in MT stabilization. Such a stabilization effect was likely brought about by MT binding and bundling. We conclude that MAP65-3 contains a distinct C-terminal MT binding site with a specific role in cross-linking antiparallel MTs toward their plus ends in the phragmoplast. PMID:22570443

Dynamical stability in fluid-structure interaction

International Nuclear Information System (INIS)

Planchard, J.; Thomas, B.

1991-01-01

The aim of the paper is to investigate the dynamical stability of a group of elastic tubes placed in a cross-flow which obeys to the Navier-Stokes equations. The stability of this coupled system is deduced from the study of a quadratic eigenvalue problem arising in the linearized equations. The instability occurs when the real part of one of the eigenvalues becomes positive; the steady state is then replaced by a time-periodic state which is stable (Hopf bifurcation phenomenon). Some numerical methods for solving the quadratic eigenvalue problem are described [fr

Dynamic postural stability differences between male and female players with and without ankle sprain

NARCIS (Netherlands)

Dallinga, Joan; van der Does, Henrike; Benjaminse, Anne; Lemmink, Koen

2015-01-01

Objectives: The strategy for dynamic postural stability might be different for male and female players. Additionally, dynamic and challenging tasks are recommended to measure differences in postural stability between injured and non-injured players. Therefore, the dynamic stability index (DSI) was

Spatiotemporal relationships between growth and microtubule orientation as revealed in living root cells of Arabidopsis thaliana transformed with green-fluorescent-protein gene construct GFP-MBD

Science.gov (United States)

Granger, C. L.; Cyr, R. J.

2001-01-01

Arabidopsis thaliana plants were transformed with GFP-MBD (J. Marc et al., Plant Cell 10: 1927-1939, 1998) under the control of a constitutive (35S) or copper-inducible promoter. GFP-specific fluorescence distributions, levels, and persistence were determined and found to vary with age, tissue type, transgenic line, and individual plant. With the exception of an increased frequency of abnormal roots of 35S GFP-MBD plants grown on kanamycin-containing media, expression of GFP-MBD does not appear to affect plant phenotype. The number of leaves, branches, bolts, and siliques as well as overall height, leaf size, and seed set are similar between wild-type and transgenic plants as is the rate of root growth. Thus, we conclude that the transgenic plants can serve as a living model system in which the dynamic behavior of microtubules can be visualized. Confocal microscopy was used to simultaneously monitor growth and microtubule behavior within individual cells as they passed through the elongation zone of the Arabidopsis root. Generally, microtubules reoriented from transverse to oblique or longitudinal orientations as growth declined. Microtubule reorientation initiated at the ends of the cell did not necessarily occur simultaneously in adjacent neighboring cells and did not involve complete disintegration and repolymerization of microtubule arrays. Although growth rates correlated with microtubule reorientation, the two processes were not tightly coupled in terms of their temporal relationships, suggesting that other factor(s) may be involved in regulating both events. Additionally, microtubule orientation was more defined in cells whose growth was accelerating and less stringent in cells whose growth was decelerating, indicating that microtubule-orienting factor(s) may be sensitive to growth acceleration, rather than growth per se.

Simultaneous 3D tracking of passive tracers and microtubule bundles in an active gel

Science.gov (United States)

Fan, Yi; Breuer, Kenneth S.; Fluids Team

Kinesin-driven microtubule bundles generate a spontaneous flow in unconfined geometries. They exhibit properties of active matter, including the emergence of collective motion, reduction of apparent viscosity and consumption of local energy. Here we present results from 3D tracking of passive tracers (using Airy rings and 3D scanning) synchronized with 3D measurement of the microtubule bundles motion. This technique is applied to measure viscosity variation and collective flow in a confined geometry with particular attention paid to the self-pumping system recently reported by Wu et al. (2016). Results show that the viscosity in an equilibrium microtubule network is around half that of the isotropic unbundled microtubule solution. Cross-correlations of the active microtubule network and passive tracers define a neighborhood around microtubule bundles in which passive tracers are effectively transported. MRSEC NSF.

Self-Sustained Oscillatory Sliding Movement of Doublet Microtubules and Flagellar Bend Formation.

Directory of Open Access Journals (Sweden)

Sumio Ishijima

Full Text Available It is well established that the basis for flagellar and ciliary movements is ATP-dependent sliding between adjacent doublet microtubules. However, the mechanism for converting microtubule sliding into flagellar and ciliary movements has long remained unresolved. The author has developed new sperm models that use bull spermatozoa divested of their plasma membrane and midpiece mitochondrial sheath by Triton X-100 and dithiothreitol. These models enable the observation of both the oscillatory sliding movement of activated doublet microtubules and flagellar bend formation in the presence of ATP. A long fiber of doublet microtubules extruded by synchronous sliding of the sperm flagella and a short fiber of doublet microtubules extruded by metachronal sliding exhibited spontaneous oscillatory movements and constructed a one beat cycle of flagellar bending by alternately actuating. The small sliding displacement generated by metachronal sliding formed helical bends, whereas the large displacement by synchronous sliding formed planar bends. Therefore, the resultant waveform is a half-funnel shape, which is similar to ciliary movements.

Potential mechanisms of resistance to microtubule inhibitors.

Science.gov (United States)

Kavallaris, Maria; Annereau, Jean-Philippe; Barret, Jean-Marc

2008-06-01

Antimitotic drugs targeting the microtubules, such as the taxanes and vinca alkaloids, are widely used in the treatment of neoplastic diseases. Development of drug resistance over time, however, limits the efficacy of these agents and poses a clinical challenge to long-term improvement of patient outcomes. Understanding the mechanism(s) of drug resistance becomes paramount to allowing for alternative, if not improved, therapeutic options that might circumvent this challenge. Vinflunine, a novel microtubule inhibitor, has shown superior preclinical antitumor activity, and displays a different pattern of resistance, compared with other agents in the vinca alkaloid class.

Identification of interphase functions for the NIMA kinase involving microtubules and the ESCRT pathway.

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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

Emerging roles for microtubules in angiosperm pollen tube growth highlight new research cues

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Alessandra eMoscatelli

2015-02-01

Full Text Available In plants, actin filaments have an important role in organelle movement and cytoplasmic streaming. Otherwise microtubules have a role in restricting organelles to specific areas of the cell and in maintaining organelle morphology. In somatic plant cells, microtubules also participate in cell division and morphogenesis, allowing cells to take their definitive shape in order to perform specific functions. In the latter case, microtubules influence assembly of the cell wall, controlling the delivery of enzymes involved in cellulose synthesis and of wall modulation material to the proper sites.In angiosperm pollen tubes, organelle movement is generally attributed to the acto-myosin system, the main role of which is in distributing organelles in the cytoplasm and in carrying secretory vesicles to the apex for polarized growth. Recent data on membrane trafficking suggests a role of microtubules in fine delivery and repositioning of vesicles to sustain pollen tube growth. This review examines the role of microtubules in secretion and endocytosis, highlighting new research cues regarding cell wall construction and pollen tube-pistil crosstalk, that help unravel the role of microtubules in polarized growth.

Electrodynamic effects on microtubules

Czech Academy of Sciences Publication Activity Database

Kučera, Ondřej; Havelka, Daniel; Deriu, M.A.; Cifra, Michal

2015-01-01

Roč. 44, Jul (2015), s. 169-169 ISSN 0175-7571. [10th European-Biophysical-Societies-Association (EBSA) European Biophysics Congress. 18.07.2015-22.07.2015, Dresden] R&D Projects: GA ČR(CZ) GA15-17102S Institutional support: RVO:67985882 Keywords : Microtubules * Electric al polarity Subject RIV: JA - Electronics ; Optoelectronics, Electric al Engineering

Feeding cells induced by phytoparasitic nematodes require γ-tubulin ring complex for microtubule reorganization.

Directory of Open Access Journals (Sweden)

Mohamed Youssef Banora

2011-12-01

Full Text Available Reorganization of the microtubule network is important for the fast isodiametric expansion of giant-feeding cells induced by root-knot nematodes. The efficiency of microtubule reorganization depends on the nucleation of new microtubules, their elongation rate and activity of microtubule severing factors. New microtubules in plants are nucleated by cytoplasmic or microtubule-bound γ-tubulin ring complexes. Here we investigate the requirement of γ-tubulin complexes for giant feeding cells development using the interaction between Arabidopsis and Meloidogyne spp. as a model system. Immunocytochemical analyses demonstrate that γ-tubulin localizes to both cortical cytoplasm and mitotic microtubule arrays of the giant cells where it can associate with microtubules. The transcripts of two Arabidopsis γ-tubulin (TUBG1 and TUBG2 and two γ-tubulin complex proteins genes (GCP3 and GCP4 are upregulated in galls. Electron microscopy demonstrates association of GCP3 and γ-tubulin as part of a complex in the cytoplasm of giant cells. Knockout of either or both γ-tubulin genes results in the gene dose-dependent alteration of the morphology of feeding site and failure of nematode life cycle completion. We conclude that the γ-tubulin complex is essential for the control of microtubular network remodelling in the course of initiation and development of giant-feeding cells, and for the successful reproduction of nematodes in their plant hosts.

Dynamic voltage stability constrained congestion management framework for deregulated electricity markets

International Nuclear Information System (INIS)

Amjady, Nima; Hakimi, Mahmood

2012-01-01

Highlights: ► A new congestion management method for electricity markets is proposed. ► The proposed method includes dynamic models of generators and loads. ► Dynamic voltage stability limits are properly modeled in the proposed method. ► The proposed method is compared with several other congestion management methods. ► It leads to a more robust power system with a lower congestion management cost. - Abstract: Congestion management is an important part of power system operation in today deregulated electricity markets. However, congestion management is traditionally performed based on static analysis tools, while these tools may not correctly capture dynamic voltage stability limits of a power system. In this paper, a new congestion management framework considering dynamic voltage stability boundary of power system is proposed. For this purpose, precise dynamic modeling of power system equipment, including generators and loads, is incorporated into the proposed congestion management framework. The proposed method alleviates congestion with a lower congestion management cost and more dynamic voltage stability margin, resulting in a more robust power system, compared with the previous congestion management methods. The validity of proposed congestion management framework is studied based on the New England 39-bus power system. The obtained results confirm the validity of the developed approach.

On the dynamics of turbulent transport near marginal stability

International Nuclear Information System (INIS)

Diamond, P.H.; Hahm, T.S.

1995-03-01

A general methodology for describing the dynamics of transport near marginal stability is formulated. Marginal stability is a special case of the more general phenomenon of self-organized criticality. Simple, one field models of the dynamics of tokamak plasma self-organized criticality have been constructed, and include relevant features such as sheared mean flow and transport bifurcations. In such models, slow mode (i.e. large scale, low frequency transport events) correlation times determine the behavior of transport dynamics near marginal stability. To illustrate this, impulse response scaling exponents (z) and turbulent diffusivities (D) have been calculated for the minimal (Burgers) and sheared flow models. For the minimal model, z = 1 (indicating ballastic propagation) and D ∼(S 0 2 ) 1/3 , where S 0 2 is the noise strength. With an identically structured noise spectrum and flow with shearing rate exceeding the ambient decorrelation rate for the largest scale transport events, diffusion is recovered with z = 2 and D ∼ (S 0 2 ) 3/5 . This indicates a qualitative change in the dynamics, as well as a reduction in losses. These results are consistent with recent findings from ρ scaling scans. Several tokamak transport experiments are suggested

Obesity May Not Induce Dynamic Stability Disadvantage during Overground Walking among Young Adults.

Science.gov (United States)

Liu, Zhong-Qi; Yang, Feng

2017-01-01

Obesity has been related to postural instability during static standing. It remains unknown how obesity influences stability during dynamic movements like gait. The primary aim of this study was to investigate the effects of obesity on dynamic gait stability control in young adults during gait. Forty-four young adults (21 normal-weight and 23 obese) participated in this study. Participants walked five times at their self-selected gait speeds on a linear walkway. Their full-body kinematics were gathered by a motion capture system. Compared with normal-weight group, individuals with obesity walked more slowly with a shorter but wider step. People with obesity also spent an elongated double stance phase than those with normal weight. A reduced gait speed decreases the body center of mass's velocity relative to the base of support, leading to a reduction in dynamic stability. On the other hand, a shortened step in accompanying with a less backward-leaning trunk has the potential to bring the center of mass closer to the base of support, resulting in an increase in dynamic stability. As the result of these adaptive changes to the gait pattern, dynamic gait stability among people with obesity did not significantly differ from the one among people with normal weight. Obesity seems to not be inducing dynamic stability disadvantage in young adults during level overground walking. These findings could provide insight into the mechanisms of stability control among people affected by obesity during dynamic locomotion.

Protein friction limits diffusive and directed movements of kinesin motors on microtubules.

Science.gov (United States)

Bormuth, Volker; Varga, Vladimir; Howard, Jonathon; Schäffer, Erik

2009-08-14

Friction limits the operation of macroscopic engines and is critical to the performance of micromechanical devices. We report measurements of friction in a biological nanomachine. Using optical tweezers, we characterized the frictional drag force of individual kinesin-8 motor proteins interacting with their microtubule tracks. At low speeds and with no energy source, the frictional drag was related to the diffusion coefficient by the Einstein relation. At higher speeds, the frictional drag force increased nonlinearly, consistent with the motor jumping 8 nanometers between adjacent tubulin dimers along the microtubule, and was asymmetric, reflecting the structural polarity of the microtubule. We argue that these frictional forces arise from breaking bonds between the motor domains and the microtubule, and they limit the speed and efficiency of kinesin.

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

Directory of Open Access Journals (Sweden)

Qun Zhang

2015-12-01

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

Vehicle lateral dynamics stabilization using active suspension

Directory of Open Access Journals (Sweden)

Drobný V.

2008-12-01

Full Text Available The paper deals with the investigation of active nonlinear suspension control in order to stabilize the lateral vehicle motion in similar way as systems like ESP do. The lateral stabilization of vehicle based on braking forces can be alternatively provided by the different setting of suspension forces. The basis of this control is the nonlinear property of the tyres. The vehicle has at least four wheels and it gives one or more redundant vertical forces that can be used for the different distribution of vertical suspension forces in such a way that resulting lateral and/or longitudinal forces create the required correction moment for lateral dynamic vehicle stabilization.

CENTROSOMES AND MICROTUBULES DURING MEIOSIS IN THE MUSHROOM BOLETUS RUBINELLUS

Science.gov (United States)

McLaughlin, David J.

1971-01-01

The double centrosome in the basidium of Boletus rubinellus has been observed in three planes with the electron microscope at interphase preceding nuclear fusion, at prophase I, and at interphase I. It is composed of two components connected by a band-shaped middle part. At anaphase I a single, enlarged centrosome is found at the spindle pole, which is attached to the cell membrane. Microtubules mainly oriented parallel to the longitudinal axis of the basidium are present at prefusion, prophase I and interphase I. Cytoplasmic microtubules are absent when the spindle is present. The relationship of the centrosome in B. rubinellus to that in other organisms and the role of the cytoplasmic microtubules are discussed. PMID:4329156

Fission yeast cells undergo nuclear division in the absence of spindle microtubules.

Directory of Open Access Journals (Sweden)

Stefania Castagnetti

2010-10-01

Full Text Available Mitosis in eukaryotic cells employs spindle microtubules to drive accurate chromosome segregation at cell division. Cells lacking spindle microtubules arrest in mitosis due to a spindle checkpoint that delays mitotic progression until all chromosomes have achieved stable bipolar attachment to spindle microtubules. In fission yeast, mitosis occurs within an intact nuclear membrane with the mitotic spindle elongating between the spindle pole bodies. We show here that in fission yeast interference with mitotic spindle formation delays mitosis only briefly and cells proceed to an unusual nuclear division process we term nuclear fission, during which cells perform some chromosome segregation and efficiently enter S-phase of the next cell cycle. Nuclear fission is blocked if spindle pole body maturation or sister chromatid separation cannot take place or if actin polymerization is inhibited. We suggest that this process exhibits vestiges of a primitive nuclear division process independent of spindle microtubules, possibly reflecting an evolutionary intermediate state between bacterial and Archeal chromosome segregation where the nucleoid divides without a spindle and a microtubule spindle-based eukaryotic mitosis.

Effect of soil stabilized by cement on dynamic response of machine foundations

Directory of Open Access Journals (Sweden)

Al-Wakel Saad

2018-01-01

Full Text Available Machine foundations require significant attention from designers. The main goal of the design of machine foundation is to limit the amplitude displacement and not disturb the people who work near the machine. In some cases, if the design of machine foundations does not satisfy the acceptable value of the dynamic response (such as maximum amplitude of displacement, the stabilization of soil under the machine foundation may be used to decrease the amplitude of displacement. This paper outlines effect of stabilized soil under the foundation by cement on the displacement response of machine foundations. Three-dimensional analyses by using finite element method are carried out to investigate the effect of depth of stabilized layer with different percentage of cement content on the dynamic response of the machine foundation. In addition, the effect of area stabilized by cement material on the dynamic response of machine foundation is investigated. The results shown that, the dynamic response of machine foundations generally decreases with increasing the depth of soil layer stabilized with cement. A significant decrease in the displacement of machine foundations is occurred for the stabilized soil layer with a depth of two times of the width of foundation, and the optimum percentage of cement for stabilizing is 6%.

Prion protein inhibits microtubule assembly by inducing tubulin oligomerization

International Nuclear Information System (INIS)

Nieznanski, Krzysztof; Podlubnaya, Zoya A.; Nieznanska, Hanna

2006-01-01

A growing body of evidence points to an association of prion protein (PrP) with microtubular cytoskeleton. Recently, direct binding of PrP to tubulin has also been found. In this work, using standard light scattering measurements, sedimentation experiments, and electron microscopy, we show for First time the effect of a direct interaction between these proteins on tubulin polymerization. We demonstrate that full-length recombinant PrP induces a rapid increase in the turbidity of tubulin diluted below the critical concentration for microtubule assembly. This effect requires magnesium ions and is weakened by NaCl. Moreover, the PrP-induced light scattering structures of tubulin are cold-stable. In preparations of diluted tubulin incubated with PrP, electron microscopy revealed the presence of ∼50 nm disc-shaped structures not reported so far. These unique tubulin oligomers may form large aggregates. The effect of PrP is more pronounced under the conditions promoting microtubule formation. In these tubulin samples, PrP induces formation of the above oligomers associated with short protofilaments and sheets of protofilaments into aggregates. Noticeably, this is accompanied by a significant reduction of the number and length of microtubules. Hence, we postulate that prion protein may act as an inhibitor of microtubule assembly by inducing formation of stable tubulin oligomers

Dynamical behavior and Jacobi stability analysis of wound strings

Science.gov (United States)

Lake, Matthew J.; Harko, Tiberiu

2016-06-01

We numerically solve the equations of motion (EOM) for two models of circular cosmic string loops with windings in a simply connected internal space. Since the windings cannot be topologically stabilized, stability must be achieved (if at all) dynamically. As toy models for realistic compactifications, we consider windings on a small section of mathbb {R}^2, which is valid as an approximation to any simply connected internal manifold if the winding radius is sufficiently small, and windings on an S^2 of constant radius mathcal {R}. We then use Kosambi-Cartan-Chern (KCC) theory to analyze the Jacobi stability of the string equations and determine bounds on the physical parameters that ensure dynamical stability of the windings. We find that, for the same initial conditions, the curvature and topology of the internal space have nontrivial effects on the microscopic behavior of the string in the higher dimensions, but that the macroscopic behavior is remarkably insensitive to the details of the motion in the compact space. This suggests that higher-dimensional signatures may be extremely difficult to detect in the effective (3+1)-dimensional dynamics of strings compactified on an internal space, even if configurations with nontrivial windings persist over long time periods.

Dynamical behavior and Jacobi stability analysis of wound strings

Energy Technology Data Exchange (ETDEWEB)

Lake, Matthew J. [Naresuan University, The Institute for Fundamental Study, ' ' The Tah Poe Academia Institute' ' , Phitsanulok (Thailand); Thailand Center of Excellence in Physics, Ministry of Education, Bangkok (Thailand); Harko, Tiberiu [Babes-Bolyai University, Department of Physics, Cluj-Napoca (Romania); University College London, Department of Mathematics, London (United Kingdom)

2016-06-15

We numerically solve the equations of motion (EOM) for two models of circular cosmic string loops with windings in a simply connected internal space. Since the windings cannot be topologically stabilized, stability must be achieved (if at all) dynamically. As toy models for realistic compactifications, we consider windings on a small section of R{sup 2}, which is valid as an approximation to any simply connected internal manifold if the winding radius is sufficiently small, and windings on an S{sup 2} of constant radius R. We then use Kosambi-Cartan-Chern (KCC) theory to analyze the Jacobi stability of the string equations and determine bounds on the physical parameters that ensure dynamical stability of the windings. We find that, for the same initial conditions, the curvature and topology of the internal space have nontrivial effects on the microscopic behavior of the string in the higher dimensions, but that the macroscopic behavior is remarkably insensitive to the details of the motion in the compact space. This suggests that higher-dimensional signatures may be extremely difficult to detect in the effective (3+1)-dimensional dynamics of strings compactified on an internal space, even if configurations with nontrivial windings persist over long time periods. (orig.)

Dynamic stabilization of the imploding-shell Rayleigh-Taylor instability

International Nuclear Information System (INIS)

Boris, J.P.

1977-01-01

A method for dynamic stabilization of the Rayleigh-Taylor (R-T) instability on the surface of an imploding fusion pellet is discussed. The driving laser beams are modulated in intensity so the ablation layer is subject to a rapidly and strongly oscillating acceleration. A substantial band of the Rayleigh-Taylor instability spectrum can be stabilized by this oscillation even though the time average acceleration vector lies in the destabilizing direction. By adjusting the frequency, structure, and amplitude of the modulation, the band of dynamically stabilized modes can be made to include the most unstable and dangerous modes. Thus considerably higher aspect ratio shells (i.e., thinner shells) could implode successfully than had been previously considered stable enough. Both theory and numerical simulations support this conclusion for the case of laser-driven pellet implosions. Similar modulation via transverse beam oscillations or parallel bunching should also work to stabilize the most dangerous surface Rayleigh-Taylor modes in relativistic electron-, ion- and heavy ion-pellet fusion schemes. (U.K.)

Dynamic stability of self-similar solutions for a plasma pinch

International Nuclear Information System (INIS)

Ma, Sifeng.

1988-01-01

Linear Magnetohydrodynamic (MHD) stability theory is applied to a class of self-similar solutions which describe implosion, expansion and oscillation of an infinitely conducting plasma column. The equations of perturbation are derived in the Lagrangian coordinate system. Numerical procedures via the finite-element method are formulated, and general aspects of dynamic stability are discussed, The dynamic stability of the column when it is oscillatory is studied in detail using the Floquet theory, and the characteristic exponent is calculated numerically. A-pinch configuration is examined. It is found that self-similar oscillations in general destabilize the continua in the MHD spectrum, and parametric instability results

Effects of ultraviolet radiation on microtubule organisation and morphogenesis in plants

International Nuclear Information System (INIS)

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, whereby a highly embryogenic protoplast fraction could be enriched. 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, which was related to the absence of an microtubule network. 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 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

Fluid mechanics in dentinal microtubules provides mechanistic insights into the difference between hot and cold dental pain.

Science.gov (United States)

Lin, Min; Luo, Zheng Yuan; Bai, Bo Feng; Xu, Feng; Lu, Tian Jian

2011-03-23

Dental thermal pain is a significant health problem in daily life and dentistry. There is a long-standing question regarding the phenomenon that cold stimulation evokes sharper and more shooting pain sensations than hot stimulation. This phenomenon, however, outlives the well-known hydrodynamic theory used to explain dental thermal pain mechanism. Here, we present a mathematical model based on the hypothesis that hot or cold stimulation-induced different directions of dentinal fluid flow and the corresponding odontoblast movements in dentinal microtubules contribute to different dental pain responses. We coupled a computational fluid dynamics model, describing the fluid mechanics in dentinal microtubules, with a modified Hodgkin-Huxley model, describing the discharge behavior of intradental neuron. The simulated results agreed well with existing experimental measurements. We thence demonstrated theoretically that intradental mechano-sensitive nociceptors are not "equally sensitive" to inward (into the pulp) and outward (away from the pulp) fluid flows, providing mechanistic insights into the difference between hot and cold dental pain. The model developed here could enable better diagnosis in endodontics which requires an understanding of pulpal histology, neurology and physiology, as well as their dynamic response to the thermal stimulation used in dental practices.

Dynamic stability of a seaplane in takeoff

CSIR Research Space (South Africa)

Dala, L

2015-01-01

Full Text Available This research is based on the investigation into the dynamic stability associated with seaplanes during take-off. Various forces acting on a hydroplaning hull form have been empirically defined. Such empirical data have shown that under a certain...

The effects of 60Co γ-ray irradiation on cytoplasmic microtubules of mouse macrophages and lymphocytes

International Nuclear Information System (INIS)

Li Qianqian; Mao Zijun; Yin Zhiwei; Hu Yumin

1989-05-01

The effects of 60 Co γ-ray irradiation on cytoplasmic microtubules of mouse macrophages and lymphocytes were investigated by immunofluorescence microscopy and scanning electron microscope. The results indicated. (1) microtubule organization of the irradiated cells remarkably differed from that of the control since the doses over 4 Gy; (2) 144 hours after irradiation the alterations of microtubules have been shown to be basically r epaired ; (3) the cytoplasmic microtubules and centrioles disappeared under transmission electron microscope, the membranes irradiated and microvilli showed changes under scanning electron microscope too. From these observations and those of other workers who studied the radiation effect on extracted microtubule proteins in vitro, the authors support that 60 Co γ-ray irradiation can inhabits cytoplasmic microtubule assembling

Tau protein and adult hippocampal neurogenesis

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Almudena eFuster-Matanzo

2012-07-01

Full Text Available Tau protein is a microtubule associated protein found in the axonal compartment that stabilizes neuronal microtubules under normal physiological conditions. Tau metabolism has attracted much attention because of its role in neurodegenerative disorders called tauopathies, mainly Alzheimer disease. Here, we review recent findings suggesting that axonal outgrowth in subgranular zone during adult hippocampal neurogenesis requires a dynamic microtubule network and tau protein facilitates to maintain that dynamic cytoskeleton. Those functions are carried out in part by tau isoform with only three microtubule-binding domains (without exon 10 and by presence of hypherphosphorylated tau forms. Thus, tau is a good marker and a valuable tool to study new axons in adult neurogenesis.

Combined analytical and numerical approaches in Dynamic Stability analyses of engineering systems

Science.gov (United States)

Náprstek, Jiří

2015-03-01

Dynamic Stability is a widely studied area that has attracted many researchers from various disciplines. Although Dynamic Stability is usually associated with mechanics, theoretical physics or other natural and technical disciplines, it is also relevant to social, economic, and philosophical areas of our lives. Therefore, it is useful to occasionally highlight the general aspects of this amazing area, to present some relevant examples and to evaluate its position among the various branches of Rational Mechanics. From this perspective, the aim of this study is to present a brief review concerning the Dynamic Stability problem, its basic definitions and principles, important phenomena, research motivations and applications in engineering. The relationships with relevant systems that are prone to stability loss (encountered in other areas such as physics, other natural sciences and engineering) are also noted. The theoretical background, which is applicable to many disciplines, is presented. In this paper, the most frequently used Dynamic Stability analysis methods are presented in relation to individual dynamic systems that are widely discussed in various engineering branches. In particular, the Lyapunov function and exponent procedures, Routh-Hurwitz, Liénard, and other theorems are outlined together with demonstrations. The possibilities for analytical and numerical procedures are mentioned together with possible feedback from experimental research and testing. The strengths and shortcomings of these approaches are evaluated together with examples of their effective complementing of each other. The systems that are widely encountered in engineering are presented in the form of mathematical models. The analyses of their Dynamic Stability and post-critical behaviour are also presented. The stability limits, bifurcation points, quasi-periodic response processes and chaotic regimes are discussed. The limit cycle existence and stability are examined together with their

Calcium regulates ATP-sensitive microtubule binding by Chlamydomonas outer arm dynein.

Science.gov (United States)

Sakato, Miho; King, Stephen M

2003-10-31

The Chlamydomonas outer dynein arm contains three distinct heavy chains (alpha, beta, and gamma) that exhibit different motor properties. The LC4 protein, which binds 1-2 Ca2+ with KCa = 3 x 10-5 m, is associated with the gamma heavy chain and has been proposed to act as a sensor to regulate dynein motor function in response to alterations in intraflagellar Ca2+ levels. Here we genetically dissect the outer arm to yield subparticles containing different motor unit combinations and assess the microtubule-binding properties of these complexes both prior to and following preincubation with tubulin and ATP, which was used to inhibit ATP-insensitive (structural) microtubule binding. We observed that the alpha heavy chain exhibits a dominant Ca2+-independent ATP-sensitive MT binding activity in vitro that is inhibited by attachment of tubulin to the structural microtubule-binding domain. Furthermore, we show that ATP-sensitive microtubule binding by a dynein subparticle containing only the beta and gamma heavy chains does not occur at Ca2+ concentrations below pCa 6 but is maximally activated above pCa 5. This activity was not observed in mutant dyneins containing small deletions in the microtubule-binding region of the beta heavy chain or in dyneins that lack both the alpha heavy chain and the motor domain of the beta heavy chain. These findings strongly suggest that Ca2+ binding directly to a component of the dynein complex regulates ATP-sensitive interactions between the beta heavy chain and microtubules and lead to a model for how individual motor units are controlled within the outer dynein arm.

Optical properties of template synthesized nanowalled ZnS microtubules

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Kumar, Rajesh; Chakarvarti, S. K.

2007-12-01

Electrodeposition is a versatile technique combining low processing cost with ambient conditions that can be used to prepare metallic, polymeric and semiconducting nano/micro structures. In the present work, track-etch membranes (TEMs) of makrofol (KG) have been used as templates for synthesis of ZnS nanowalled microtubules using electrodeposition technique. The morphology of the microtubules was characterized by scanning electron microscopy. Size effects on the band gap of tubules have also been studied by UV-visible spectrophotometer.

Regulation of microtubule nucleation mediated by gamma-tubulin complexes

Czech Academy of Sciences Publication Activity Database

Sulimenko, Vadym; Hájková, Zuzana; Klebanovych, Anastasiya; Dráber, Pavel

2017-01-01

Roč. 254, č. 3 (2017), s. 1187-1199 ISSN 0033-183X R&D Projects: GA MŠk(CZ) LD13015 Institutional support: RVO:68378050 Keywords : mitotic spindle formation * ring complex * fission yeast * organizing centers * protein complex * golgi-complex * cell-cycle * pole body * augmin * centrosome * Centrosomes * Microtubule nucleation * Microtubule-organizing centers * Non-centrosomal nucleation sites * Spindle pole bodies * gamma-Tubulin complexes Subject RIV: EB - Genetics ; Molecular Biology OBOR OECD: Cell biology Impact factor: 2.870, year: 2016

Dinitroaniline herbicide resistance and the microtubule cytoskeleton.

Science.gov (United States)

Anthony; Hussey

1999-03-01

Dinitroaniline herbicides have been used for pre-emergence weed control for the past 25 years in cotton, soybean, wheat and oilseed crops. Considering their long persistence and extensive use, resistance to dinitroanilines is fairly rare. However, the most widespread dinitroaniline-resistant weeds, the highly resistant (R) and the intermediate (I) biotypes of the invasive goosegrass Eleusine indica, are now infesting more than 1000 cotton fields in the southern states of the USA. The molecular basis of this resistance has been identified, and found to be a point mutation in a major microtubule cytoskeletal protein, alpha-tubulin. These studies have served both to explain the establishment of resistance and to reveal fundamental properties of tubulin gene expression and microtubule structure.

An analytical method for solving exact solutions of a nonlinear evolution equation describing the dynamics of ionic currents along microtubules

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Md. Nur Alam

2017-11-01

Full Text Available In this article, a variety of solitary wave solutions are observed for microtubules (MTs. We approach the problem by treating the solutions as nonlinear RLC transmission lines and then find exact solutions of Nonlinear Evolution Equations (NLEEs involving parameters of special interest in nanobiosciences and biophysics. We determine hyperbolic, trigonometric, rational and exponential function solutions and obtain soliton-like pulse solutions for these equations. A comparative study against other methods demonstrates the validity of the technique that we developed and demonstrates that our method provides additional solutions. Finally, using suitable parameter values, we plot 2D and 3D graphics of the exact solutions that we observed using our method. Keywords: Analytical method, Exact solutions, Nonlinear evolution equations (NLEEs of microtubules, Nonlinear RLC transmission lines

Msd1/SSX2IP-dependent microtubule anchorage ensures spindle orientation and primary cilia formation

OpenAIRE

Hori, Akiko; Ikebe, Chiho; Tada, Masazumi; Toda, Takashi

2014-01-01

Anchoring microtubules to the centrosome is critical for cell geometry and polarity, yet the molecular mechanism remains unknown. Here we show that the conserved human Msd1/SSX2IP is required for microtubule anchoring. hMsd1/SSX2IP is delivered to the centrosome in a centriolar satellite-dependent manner and binds the microtubule-nucleator ?-tubulin complex. hMsd1/SSX2IP depletion leads to disorganised interphase microtubules and misoriented mitotic spindles with reduced length and intensity....

Deciphering the imprint of topology on nonlinear dynamical network stability

International Nuclear Information System (INIS)

Nitzbon, J; Schultz, P; Heitzig, J; Kurths, J; Hellmann, F

2017-01-01

Coupled oscillator networks show complex interrelations between topological characteristics of the network and the nonlinear stability of single nodes with respect to large but realistic perturbations. We extend previous results on these relations by incorporating sampling-based measures of the transient behaviour of the system, its survivability, as well as its asymptotic behaviour, its basin stability. By combining basin stability and survivability we uncover novel, previously unknown asymptotic states with solitary, desynchronized oscillators which are rotating with a frequency different from their natural one. They occur almost exclusively after perturbations at nodes with specific topological properties. More generally we confirm and significantly refine the results on the distinguished role tree-shaped appendices play for nonlinear stability. We find a topological classification scheme for nodes located in such appendices, that exactly separates them according to their stability properties, thus establishing a strong link between topology and dynamics. Hence, the results can be used for the identification of vulnerable nodes in power grids or other coupled oscillator networks. From this classification we can derive general design principles for resilient power grids. We find that striving for homogeneous network topologies facilitates a better performance in terms of nonlinear dynamical network stability. While the employed second-order Kuramoto-like model is parametrised to be representative for power grids, we expect these insights to transfer to other critical infrastructure systems or complex network dynamics appearing in various other fields. (paper)

ATPase Cycle of the Nonmotile Kinesin NOD Allows Microtubule End Tracking and Drives Chromosome Movement

Energy Technology Data Exchange (ETDEWEB)

Cochran, J.; Sindelar, C; Mulko, N; Collins, K; Kong, S; Hawley, R; Kull, F

2009-01-01

Segregation of nonexchange chromosomes during Drosophila melanogaster meiosis requires the proper function of NOD, a nonmotile kinesin-10. We have determined the X-ray crystal structure of the NOD catalytic domain in the ADP- and AMPPNP-bound states. These structures reveal an alternate conformation of the microtubule binding region as well as a nucleotide-sensitive relay of hydrogen bonds at the active site. Additionally, a cryo-electron microscopy reconstruction of the nucleotide-free microtubule-NOD complex shows an atypical binding orientation. Thermodynamic studies show that NOD binds tightly to microtubules in the nucleotide-free state, yet other nucleotide states, including AMPPNP, are weakened. Our pre-steady-state kinetic analysis demonstrates that NOD interaction with microtubules occurs slowly with weak activation of ADP product release. Upon rapid substrate binding, NOD detaches from the microtubule prior to the rate-limiting step of ATP hydrolysis, which is also atypical for a kinesin. We propose a model for NOD's microtubule plus-end tracking that drives chromosome movement.

Dynamics and Stability of Permanent-Magnet Synchronous Motor

OpenAIRE

He, Ren; Han, Qingzhen

2017-01-01

The aim of this article is to explore the dynamic characteristics and stability of the permanent-magnet synchronous motor (PMSM). PMSM equilibrium local stability condition and Hopf  bifurcation condition, pitchfork bifurcation condition, and fold bifurcation condition have been derived by using the Routh-Hurwitz criterion and the bifurcation theory, respectively. Bifurcation curves of the equilibrium with single and double parameters are obtained by continuation method. Numerical simulations...

On the origins of the universal dynamics of endogenous granules in mammalian cells.

Science.gov (United States)

Vanapalli, Siva A; Li, Yixuan; Mugele, Frieder; Duits, Michel H G

2009-12-01

Endogenous granules (EGs) that consist of lipid droplets and mitochondria have been commonly used to assess intracellular mechanical properties via multiple particle tracking microrheology (MPTM). Despite their widespread use, the nature of interaction of EGs with the cytoskeletal network and the type of forces driving their dynamics--both of which are crucial for the interpretation of the results from MPTM technique--are yet to be resolved. In this report, we study the dynamics of endogenous granules in mammalian cells using particle tracking methods. We find that the ensemble dynamics of EGs is diffusive in three types of mammalian cells (endothelial cells, smooth muscle cells and fibroblasts), thereby suggesting an apparent universality in their dynamical behavior. Moreover, in a given cell, the amplitude of the mean-squared displacement for EGs is an order of magnitude larger than that of injected particles. This observation along with results from ATP depletion and temperature intervention studies suggests that cytoskeletal active forces drive the dynamics of EGs. To elucidate the dynamical origin of the diffusive-like nonthermal motion, we consider three active force generation mechanisms--molecular motor transport, actomyosin contractility and microtubule polymerization forces. We test these mechanisms using pharmacological interventions. Experimental evidence and model calculations suggest that EGs are intimately linked to microtubules and that microtubule polymerization forces drive their dynamics. Thus, endogenous granules could serve as non-invasive probes for microtubule network dynamics in mammalian cells.

Standard representation and unified stability analysis for dynamic artificial neural network models.

Science.gov (United States)

Kim, Kwang-Ki K; Patrón, Ernesto Ríos; Braatz, Richard D

2018-02-01

An overview is provided of dynamic artificial neural network models (DANNs) for nonlinear dynamical system identification and control problems, and convex stability conditions are proposed that are less conservative than past results. The three most popular classes of dynamic artificial neural network models are described, with their mathematical representations and architectures followed by transformations based on their block diagrams that are convenient for stability and performance analyses. Classes of nonlinear dynamical systems that are universally approximated by such models are characterized, which include rigorous upper bounds on the approximation errors. A unified framework and linear matrix inequality-based stability conditions are described for different classes of dynamic artificial neural network models that take additional information into account such as local slope restrictions and whether the nonlinearities within the DANNs are odd. A theoretical example shows reduced conservatism obtained by the conditions. Copyright © 2017. Published by Elsevier Ltd.

Disruption of microtubule network rescues aberrant actin comets in dynamin2-depleted cells.

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Yuji Henmi

Full Text Available A large GTPase dynamin, which is required for endocytic vesicle formation, regulates the actin cytoskeleton through its interaction with cortactin. Dynamin2 mutants impair the formation of actin comets, which are induced by Listeria monocytogenes or phosphatidylinositol-4-phosphate 5-kinase. However, the role of dynamin2 in the regulation of the actin comet is still unclear. Here we show that aberrant actin comets in dynamin2-depleted cells were rescued by disrupting of microtubule networks. Depletion of dynamin2, but not cortactin, significantly reduced the length and the speed of actin comets induced by Listeria. This implies that dynamin2 may regulate the actin comet in a cortactin-independent manner. As dynamin regulates microtubules, we investigated whether perturbation of microtubules would rescue actin comet formation in dynamin2-depleted cells. Treatment with taxol or colchicine created a microtubule-free space in the cytoplasm, and made no difference between control and dynamin2 siRNA cells. This suggests that the alteration of microtubules by dynamin2 depletion reduced the length and the speed of the actin comet.

Microtubular stability affects pVHL-mediated regulation of HIF-1alpha via the p38/MAPK pathway in hypoxic cardiomyocytes.

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Miao Teng

Full Text Available BACKGROUND: Our previous research found that structural changes of the microtubule network influence glycolysis in cardiomyocytes by regulating the hypoxia-inducible factor (HIF-1α during the early stages of hypoxia. However, little is known about the underlying regulatory mechanism of the changes of HIF-1α caused by microtubule network alternation. The von Hippel-Lindau tumor suppressor protein (pVHL, as a ubiquitin ligase, is best understood as a negative regulator of HIF-1α. METHODOLOGY/PRINCIPAL FINDINGS: In primary rat cardiomyocytes and H9c2 cardiac cells, microtubule-stabilization was achieved by pretreating with paclitaxel or transfection of microtubule-associated protein 4 (MAP4 overexpression plasmids and microtubule-depolymerization was achieved by pretreating with colchicine or transfection of MAP4 siRNA before hypoxia treatment. Recombinant adenovirus vectors for overexpressing pVHL or silencing of pVHL expression were constructed and transfected in primary rat cardiomyocytes and H9c2 cells. With different microtubule-stabilizing and -depolymerizing treaments, we demonstrated that the protein levels of HIF-1α were down-regulated through overexpression of pVHL and were up-regulated through knockdown of pVHL in hypoxic cardiomyocytes. Importantly, microtubular structure breakdown activated p38/MAPK pathway, accompanied with the upregulation of pVHL. In coincidence, we found that SB203580, a p38/MAPK inhibitor decreased pVHL while MKK6 (Glu overexpression increased pVHL in the microtubule network altered-hypoxic cardiomyocytes and H9c2 cells. CONCLUSIONS/SIGNIFICANCE: This study suggests that pVHL plays an important role in the regulation of HIF-1α caused by the changes of microtubular structure and the p38/MAPK pathway participates in the process of pVHL change following microtubule network alteration in hypoxic cardiomyocytes.

Stability of nanofluids: Molecular dynamic approach and experimental study

International Nuclear Information System (INIS)

Farzaneh, H.; Behzadmehr, A.; Yaghoubi, M.; Samimi, A.; Sarvari, S.M.H.

2016-01-01

Highlights: • Nanofluid stability is investigated and discussed. • A molecular dynamic approach, considering different forces on the nanoparticles, is adopted. • Stability diagrams are presented for different thermo-fluid conditions. • An experimental investigation is carried out to confirm the theoretical approach. - Abstract: Nanofluids as volumetric absorbent in solar energy conversion devices or as working fluid in different heat exchangers have been proposed by various researchers. However, dispersion stability of nanofluids is an important issue that must be well addressed before any industrial applications. Conditions such as severe temperature gradient, high temperature of heat transfer fluid, nanoparticle mean diameters and types of nanoparticles and base fluid are among the most effective parameters on the stability of nanofluid. A molecular dynamic approach, considering kinetic energy of nanoparticles and DLVO potential energy between nanoparticles, is adopted to study the nanofluid stability for different nanofluids at different working conditions. Different forces such as Brownian, thermophoresis, drag and DLVO are considered to introduce the stability diagrams. The latter presents the conditions for which a nanofluid can be stable. In addition an experimental investigation is carried out to find a stable nanofluid and to show the validity of the theoretical approach. There is a good agreement between the experimental and theoretical results that confirms the validity of our theoretical approach.

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

Molecular dynamics study of thermodynamic stability and dynamics of [Li(glyme)]+ complex in lithium-glyme solvate ionic liquids

Science.gov (United States)

Shinoda, Wataru; Hatanaka, Yuta; Hirakawa, Masashi; Okazaki, Susumu; Tsuzuki, Seiji; Ueno, Kazuhide; Watanabe, Masayoshi

2018-05-01

Equimolar mixtures of glymes and organic lithium salts are known to produce solvate ionic liquids, in which the stability of the [Li(glyme)]+ complex plays an important role in determining the ionic dynamics. Since these mixtures have attractive physicochemical properties for application as electrolytes, it is important to understand the dependence of the stability of the [Li(glyme)]+ complex on the ion dynamics. A series of microsecond molecular dynamics simulations has been conducted to investigate the dynamic properties of these solvate ionic liquids. Successful solvate ionic liquids with high stability of the [Li(glyme)]+ complex have been shown to have enhanced ion dynamics. Li-glyme pair exchange rarely occurs: its characteristic time is longer than that of ion diffusion by one or two orders of magnitude. Li-glyme pair exchange most likely occurs through cluster formation involving multiple [Li(glyme)]+ pairs. In this process, multiple exchanges likely take place in a concerted manner without the production of energetically unfavorable free glyme or free Li+ ions.

On D-brane dynamics and moduli stabilization

Science.gov (United States)

Kitazawa, Noriaki

2017-09-01

We discuss the effect of the dynamics of D-branes on moduli stabilization in type IIB string theory compactifications, with reference to a concrete toy model of T6/Z 3 orientifold compactification with fractional D3-branes and anti-D3-branes at orbifold fixed points. The resulting attractive forces between anti-D3-branes and D3-branes, together with the repulsive forces between anti-D3-branes and O3-planes, can affect the stability of the compact space. There are no complex structure moduli in T6/Z 3 orientifold, which should thus capture some generic features of more general settings where all complex structure moduli are stabilized by three-form fluxes. The simultaneous presence of branes and anti-branes brings along the breaking of supersymmetry. Non-BPS combinations of this type are typical of “brane supersymmetry breaking” and are a necessary ingredient in the KKLT scenario for stabilizing the remaining Kähler moduli. The conclusion of our analysis is that, while mutual D-brane interactions sometimes help Kähler moduli stabilization, this is not always the case.

Cep192 controls the balance of centrosome and non-centrosomal microtubules during interphase.

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Brian P O'Rourke

Full Text Available Cep192 is a centrosomal protein that contributes to the formation and function of the mitotic spindle in mammalian cells. Cep192's mitotic activities stem largely from its role in the recruitment to the centrosome of numerous additional proteins such as gamma-tubulin and Pericentrin. Here, we examine Cep192's function in interphase cells. Our data indicate that, as in mitosis, Cep192 stimulates the nucleation of centrosomal microtubules thereby regulating the morphology of interphase microtubule arrays. Interestingly, however, cells lacking Cep192 remain capable of generating normal levels of MTs as the loss of centrosomal microtubules is augmented by MT nucleation from other sites, most notably the Golgi apparatus. The depletion of Cep192 results in a significant decrease in the level of centrosome-associated gamma-tubulin, likely explaining its impact on centrosome microtubule nucleation. However, in stark contrast to mitosis, Cep192 appears to maintain an antagonistic relationship with Pericentrin at interphase centrosomes. Interphase cells depleted of Cep192 display significantly higher levels of centrosome-associated Pericentrin while overexpression of Cep192 reduces the levels of centrosomal Pericentrin. Conversely, depletion of Pericentrin results in elevated levels of centrosomal Cep192 and enhances microtubule nucleation at centrosomes, at least during interphase. Finally, we show that depletion of Cep192 negatively impacts cell motility and alters normal cell polarization. Our current working hypothesis is that the microtubule nucleating capacity of the interphase centrosome is determined by an antagonistic balance of Cep192, which promotes nucleation, and Pericentrin, which inhibits nucleation. This in turn determines the relative abundance of centrosomal and non-centrosomal microtubules that tune cell movement and shape.

Mathematical modeling of alignment dynamics in active motor-filament systems

Science.gov (United States)

Swaminathan, Sumanth

The formation of the cytoskeleton, via motor-mediated microtubule self-organization, is an important subject of study in the biological sciences as well as in nonequilibrium, soft matter physics. Accurate modeling of the dynamics is a formidable task as it involves intrinsic nonlinearities, structural anisotropies, nonequilibrium processes, and a broad window of time scales, length scales, and densities. In this thesis, we study the ordering dynamics and pattern formations arising from motor-mediated microtubule self-organization in dilute and semi-dilute filament solutions. In the dilute case, we use a probabilistic model in which microtubules interact through motor induced, inelastic binary collisions. This model shows that initially disordered filament solutions exhibit an ordering transition resulting in the emergence of well aligned rod bundles. We study the existence and dynamic interaction of microtubule bundles analytically and numerically. Our results show a long term attraction and coalescing of bundles indicating a clear coarsening in the system; microtubule bundles concentrate into fewer orientations on a slow logarithmic time scale. In the semi-dilute case, multiple motors can bind a filament to several others and, for a critical motor density, induce a transition to an ordered state with a nonzero mean orientation. We develop a spatially homogeneous, mean-field theory that explicitly accounts for motor forcing and thermal fluctuations which enter into the model as multiplicative and additive noises respectively. Our model further incorporates a force-dependent detachment rate of motors, which in turn affects the mean and the fluctuations of the net force acting on a filament. We demonstrate that the transition to the oriented state changes from second order to first order when the force-dependent detachment becomes important. In our final analysis, we add complex spatial inhomogeneities to our mean field theory. The revised model consists of a system

The neurosteroid pregnenolone reverts microtubule derangement induced by the loss of a functional CDKL5-IQGAP1 complex.

Science.gov (United States)

Barbiero, Isabella; Peroni, Diana; Tramarin, Marco; Chandola, Chetan; Rusconi, Laura; Landsberger, Nicoletta; Kilstrup-Nielsen, Charlotte

2017-09-15

CDKL5 is a protein kinase that plays a key role for neuronal functions as testified by the onset of complex neuronal dysfunctions in patients with genetic lesions in CDKL5. Here we identify a novel interactor of CDKL5, IQGAP1, a fundamental regulator of cell migration and polarity. In accordance with a functional role of this interaction, depletion of CDKL5 impairs cell migration and impedes the localization of IQGAP1 at the leading edge. Moreover, we demonstrate that CDKL5 is required for IQGAP1 to form a functional complex with its effectors, Rac1 and the microtubule plus end tracking protein CLIP170. These defects eventually impact on the microtubule association of CLIP170, thus deranging their dynamics. CLIP170 is a cellular target of the neurosteroid pregnenolone; by blocking CLIP170 in its active conformation, pregnenolone is capable of restoring the microtubule association of CLIP170 in CDKL5 deficient cells and rescuing morphological defects in neurons devoid of CDKL5. These findings provide novel insights into CDKL5 functions and pave the way for target-specific therapeutic strategies for individuals affected with CDKL5-disorder. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

p27Kip1 Modulates Axonal Transport by Regulating α-Tubulin Acetyltransferase 1 Stability

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Giovanni Morelli

2018-05-01

Full Text Available Summary: The protein p27Kip1 plays roles that extend beyond cell-cycle regulation during cerebral cortex development, such as the regulation of neuronal migration and neurite branching via signaling pathways that converge on the actin and microtubule cytoskeletons. Microtubule-dependent transport is essential for the maturation of neurons and the establishment of neuronal connectivity though synapse formation and maintenance. Here, we show that p27Kip1 controls the transport of vesicles and organelles along the axon of mice cortical projection neurons in vitro. Moreover, suppression of the p27Kip1 ortholog, dacapo, in Drosophila melanogaster disrupts axonal transport in vivo, leading to the reduction of locomotor activity in third instar larvae and adult flies. At the molecular level, p27Kip1 stabilizes the α-tubulin acetyltransferase 1, thereby promoting the acetylation of microtubules, a post-translational modification required for proper axonal transport. : Morelli et al. report that p27Kip1/Dacapo modulates the acetylation of microtubules in axons via stabilization of ATAT1, the main α-tubulin acetyltransferase. Its conditional loss leads to the reduction of bidirectional axonal transport of vesicles and mitochondria in vitro in mice and in vivo in Drosophila. Keywords: p27Kip1, dacapo, acetylation, axonal transport, ATAT1, alpha-tubulin, HDAC6, Drosophila, mouse, cerebral cortex

KIAA0100 Modulates Cancer Cell Aggression Behavior of MDA-MB-231 through Microtubule and Heat Shock Proteins

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Zhenyu Zhong

2018-06-01

Full Text Available The KIAA0100 gene was identified in the human immature myeloid cell line cDNA library. Recent studies have shown that its expression is elevated in breast cancer and associated with more aggressive cancer types as well as poor outcomes. However, its cellular and molecular function is yet to be understood. Here we show that silencing KIAA0100 by siRNA in the breast cancer cell line MDA-MB-231 significantly reduced the cancer cells’ aggressive behavior, including cell aggregation, reattachment, cell metastasis and invasion. Most importantly, silencing the expression of KIAA0100 particularly sensitized the quiescent cancer cells in suspension culture to anoikis. Immunoprecipitation, mass spectrometry and immunofluorescence analysis revealed that KIAA0100 may play multiple roles in the cancer cells, including stabilizing microtubule structure as a microtubule binding protein, and contributing to MDA-MB-231 cells Anoikis resistance by the interaction with stress protein HSPA1A. Our study also implies that the interaction between KIAA0100 and HSPA1A may be targeted for new drug development to specifically induce anoikis cell death in the cancer cell.

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

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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.

Stability Analysis on Sparsely Encoded Associative Memory with Short-Term Synaptic Dynamics

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Xu, Muyuan; Katori, Yuichi; Aihara, Kazuyuki

This study investigates the stability of sparsely encoded associative memory in a network composed of stochastic neurons. The incorporation of short-term synaptic dynamics significantly changes the stability with respect to synaptic properties. Various states including static and oscillatory states are found in the network dynamics. Specifically, the sparseness of memory patterns raises the problem of spurious states. A mean field model is used to analyze the detailed structure in the stability and show that the performance of memory retrieval is recovered by appropriate feedback.

GTSE1 is a microtubule plus-end tracking protein that regulates EB1-dependent cell migration.

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Massimilano Scolz

Full Text Available The regulation of cell migration is a highly complex process that is often compromised when cancer cells become metastatic. The microtubule cytoskeleton is necessary for cell migration, but how microtubules and microtubule-associated proteins regulate multiple pathways promoting cell migration remains unclear. Microtubule plus-end binding proteins (+TIPs are emerging as important players in many cellular functions, including cell migration. Here we identify a +TIP, GTSE1, that promotes cell migration. GTSE1 accumulates at growing microtubule plus ends through interaction with the EB1+TIP. The EB1-dependent +TIP activity of GTSE1 is required for cell migration, as well as for microtubule-dependent disassembly of focal adhesions. GTSE1 protein levels determine the migratory capacity of both nontransformed and breast cancer cell lines. In breast cancers, increased GTSE1 expression correlates with invasive potential, tumor stage, and time to distant metastasis, suggesting that misregulation of GTSE1 expression could be associated with increased invasive potential.

Clostridium difficile toxin CDT induces formation of microtubule-based protrusions and increases adherence of bacteria.

Directory of Open Access Journals (Sweden)

Carsten Schwan

2009-10-01

Full Text Available Clostridium difficile causes antibiotic-associated diarrhea and pseudomembranous colitis by production of the Rho GTPase-glucosylating toxins A and B. Recently emerging hypervirulent Clostridium difficile strains additionally produce the binary ADP-ribosyltransferase toxin CDT (Clostridium difficile transferase, which ADP-ribosylates actin and inhibits actin polymerization. Thus far, the role of CDT as a virulence factor is not understood. Here we report by using time-lapse- and immunofluorescence microscopy that CDT and other binary actin-ADP-ribosylating toxins, including Clostridium botulinum C2 toxin and Clostridium perfringens iota toxin, induce redistribution of microtubules and formation of long (up to >150 microm microtubule-based protrusions at the surface of intestinal epithelial cells. The toxins increase the length of decoration of microtubule plus-ends by EB1/3, CLIP-170 and CLIP-115 proteins and cause redistribution of the capture proteins CLASP2 and ACF7 from microtubules at the cell cortex into the cell interior. The CDT-induced microtubule protrusions form a dense meshwork at the cell surface, which wrap and embed bacterial cells, thereby largely increasing the adherence of Clostridia. The study describes a novel type of microtubule structure caused by less efficient microtubule capture and offers a new perspective for the pathogenetic role of CDT and other binary actin-ADP-ribosylating toxins in host-pathogen interactions.

Recent Advances in Heliogyro Solar Sail Structural Dynamics, Stability, and Control Research

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Wilkie, W. Keats; Warren, Jerry E.; Horta, Lucas G.; Lyle, Karen H.; Juang, Jer-Nan; Gibbs, S. Chad; Dowell, Earl H.; Guerrant, Daniel V.; Lawrence, Dale

2015-01-01

Results from recent NASA sponsored research on the structural dynamics, stability, and control characteristics of heliogyro solar sails are summarized. Specific areas under investigation include coupled nonlinear finite element analysis of heliogyro membrane blade with solar radiation pressure effects, system identification of spinning membrane structures, and solarelastic stability analysis of heliogyro solar sails, including stability during blade deployment. Recent results from terrestrial 1-g blade dynamics and control experiments on "rope ladder" membrane blade analogs, and small-scale in vacuo system identification experiments with hanging and spinning high-aspect ratio membranes will also be presented. A low-cost, rideshare payload heliogyro technology demonstration mission concept is used as a mission context for these heliogyro structural dynamics and solarelasticity investigations, and is also described. Blade torsional dynamic response and control are also shown to be significantly improved through the use of edge stiffening structural features or inclusion of modest tip masses to increase centrifugal stiffening of the blade structure. An output-only system identification procedure suitable for on-orbit blade dynamics investigations is also developed and validated using ground tests of spinning sub-scale heliogyro blade models. Overall, analytical and experimental investigations to date indicate no intractable stability or control issues for the heliogyro solar sail concept.

GSK3 controls axon growth via CLASP-mediated regulation of growth cone microtubules

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Hur, Eun-Mi; Saijilafu; Lee, Byoung Dae; Kim, Seong-Jin; Xu, Wen-Lin; Zhou, Feng-Quan

2011-01-01

Suppression of glycogen synthase kinase 3 (GSK3) activity in neurons yields pleiotropic outcomes, causing both axon growth promotion and inhibition. Previous studies have suggested that specific GSK3 substrates, such as adenomatous polyposis coli (APC) and collapsin response mediator protein 2 (CRMP2), support axon growth by regulating the stability of axonal microtubules (MTs), but the substrate(s) and mechanisms conveying axon growth inhibition remain elusive. Here we show that CLIP (cytoplasmic linker protein)-associated protein (CLASP), originally identified as a MT plus end-binding protein, displays both plus end-binding and lattice-binding activities in nerve growth cones, and reveal that the two MT-binding activities regulate axon growth in an opposing manner: The lattice-binding activity mediates axon growth inhibition induced by suppression of GSK3 activity via preventing MT protrusion into the growth cone periphery, whereas the plus end-binding property supports axon extension via stabilizing the growing ends of axonal MTs. We propose a model in which CLASP transduces GSK3 activity levels to differentially control axon growth by coordinating the stability and configuration of growth cone MTs. PMID:21937714

HSPB1 facilitates the formation of non-centrosomal microtubules.

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Leonardo Almeida-Souza

Full Text Available The remodeling capacity of microtubules (MT is essential for their proper function. In mammals, MTs are predominantly formed at the centrosome, but can also originate from non-centrosomal sites, a process that is still poorly understood. We here show that the small heat shock protein HSPB1 plays a role in the control of non-centrosomal MT formation. The HSPB1 expression level regulates the balance between centrosomal and non-centrosomal MTs. The HSPB1 protein can be detected specifically at sites of de novo forming non-centrosomal MTs, while it is absent from the centrosomes. In addition, we show that HSPB1 binds preferentially to the lattice of newly formed MTs in vitro, suggesting that its function occurs by stabilizing MT seeds. Our findings open new avenues for the understanding of the role of HSPB1 in the development, maintenance and protection of cells with specialized non-centrosomal MT arrays.

Attitude Dynamics and Stability of a Simple Solar Photon Thruster

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Anna D. Guerman

2013-01-01

Full Text Available This paper is dedicated to the development of a model of the attitude dynamics for a nonideal Simple Solar Photon Thruster (SSPT and to the analysis of sailcraft motions with respect to their centre of mass. Derivation of the expressions for force and torque due to solar radiation that is valid for the case, when there is a misalignment of the SSPT axis with the sun direction, is followed by study of sailcraft dynamics and stability properties. Analysis of stability shows that an ideally reflecting sail is unstable, while for a sailcraft with nonideal collector, the symmetry axis is stable with respect to the Sun direction for large variety of system parameters. The motion around symmetry axis is always unstable and requires an active stabilizer.

Disassembly of microtubules and inhibition of neurite outgrowth, neuroblastoma cell proliferation, and MAP kinase tyrosine dephosphorylation by dibenzyl trisulphide.

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Rösner, H; Williams, L A; Jung, A; Kraus, W

2001-08-22

Dibenzyl trisulphide (DTS), a main lipophilic compound in Petiveria alliacea L. (Phytolaccaceae), was identified as one of the active immunomodulatory compounds in extracts of the plant. To learn more about its biological activities and molecular mechanisms, we conducted one-dimensional NMR interaction studies with bovine serum albumin (BSA) and tested DTS and related compounds in two well-established neuronal cell-and-tissue culture systems. We found that DTS preferentially binds to an aromatic region of BSA which is rich in tyrosyl residues. In SH-SY5Y neuroblastoma cells, DTS attenuates the dephosphorylation of tyrosyl residues of MAP kinase (erk1/erk2). In the same neuroblastoma cell line and in Wistar 38 human lung fibroblasts, DTS causes a reversible disassembly of microtubules, but it did not affect actin dynamics. Probably due to the disruption of the microtubule dynamics, DTS also inhibits neuroblastoma cell proliferation and neurite outgrowth from spinal cord explants. Related dibenzyl compounds with none, one, or two sulphur atoms were found to be significantly less effective. These data confirmed that the natural compound DTS has a diverse spectrum of biological properties, including cytostatic and neurotoxic actions in addition to immunomodulatory activities.

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.

Learning and Understanding System Stability Using Illustrative Dynamic Texture Examples

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Liu, Huaping; Xiao, Wei; Zhao, Hongyan; Sun, Fuchun

2014-01-01

System stability is a basic concept in courses on dynamic system analysis and control for undergraduate students with computer science backgrounds. Typically, this was taught using a simple simulation example of an inverted pendulum. Unfortunately, many difficult issues arise in the learning and understanding of the concepts of stability,…

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

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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.

TCTEX1D4, a novel protein phosphatase 1 interactor: connecting the phosphatase to the microtubule network

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Korrodi-Gregório, Luís; Vieira, Sandra I.; Esteves, Sara L. C.; Silva, Joana V.; Freitas, Maria João; Brauns, Ann-Kristin; Luers, Georg; Abrantes, Joana; Esteves, Pedro J.; da Cruz e Silva, Odete A. B.; Fardilha, Margarida; da Cruz e Silva, Edgar F.

2013-01-01

Summary Reversible phosphorylation plays an important role as a mechanism of intracellular control in eukaryotes. PPP1, a major eukaryotic Ser/Thr-protein phosphatase, acquires its specificity by interacting with different protein regulators, also known as PPP1 interacting proteins (PIPs). In the present work we characterized a physiologically relevant PIP in testis. Using a yeast two-hybrid screen with a human testis cDNA library, we identified a novel PIP of PPP1CC2 isoform, the T-complex testis expressed protein 1 domain containing 4 (TCTEX1D4) that has recently been described as a Tctex1 dynein light chain family member. The overlay assays confirm that TCTEX1D4 interacts with the different spliced isoforms of PPP1CC. Also, the binding domain occurs in the N-terminus, where a consensus PPP1 binding motif (PPP1BM) RVSF is present. The distribution of TCTEX1D4 in testis suggests its involvement in distinct functions, such as TGFβ signaling at the blood–testis barrier and acrosome cap formation. Immunofluorescence in human ejaculated sperm shows that TCTEX1D4 is present in the flagellum and in the acrosome region of the head. Moreover, TCTEX1D4 and PPP1 co-localize in the microtubule organizing center (MTOC) and microtubules in cell cultures. Importantly, the TCTEX1D4 PPP1BM seems to be relevant for complex formation, for PPP1 retention in the MTOC and movement along microtubules. These novel results open new avenues to possible roles of this dynein, together with PPP1. In essence TCTEX1D4/PPP1C complex appears to be involved in microtubule dynamics, sperm motility, acrosome reaction and in the regulation of the blood–testis barrier. PMID:23789093

Probing Conformational Dynamics of Tau Protein by Hydrogen/Deuterium Exchange Mass Spectrometry

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Huang, Richard Y.-C.; Iacob, Roxana E.; Sankaranarayanan, Sethu; Yang, Ling; Ahlijanian, Michael; Tao, Li; Tymiak, Adrienne A.; Chen, Guodong

2018-01-01

Fibrillization of the microtubule-associated protein tau has been recognized as one of the signature pathologies of the nervous system in Alzheimer's disease, progressive supranuclear palsy, and other tauopathies. The conformational transition of tau in the fibrillization process, tau monomer to soluble aggregates to fibrils in particular, remains unclear. Here we report on the use of hydrogen/deuterium exchange mass spectrometry (HDX-MS) in combination with other biochemical approaches, including Thioflavin S fluorescence measurements, enzyme-linked immunosorbent assay (ELISA), and Western blotting to understand the heparin-induced tau's fibrillization. HDX-MS studies including anti-tau antibody epitope mapping experiments provided molecular level details of the full-length tau's conformational dynamics and its regional solvent accessibility upon soluble aggregates formation. The results demonstrate that R3 region in the full-length tau's microtubule binding repeat region (MTBR) is stabilized in the aggregation process, leaving both N and C terminal regions to be solvent exposed in the soluble aggregates and fibrils. The findings also illustrate the practical utility of orthogonal analytical methodologies for the characterization of protein higher order structure. [Figure not available: see fulltext.

Cdk1-cyclin B1-mediated phosphorylation of tumor-associated microtubule-associated protein/cytoskeleton-associated protein 2 in mitosis.

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Hong, Kyung Uk; Kim, Hyun-Jun; Kim, Hyo-Sil; Seong, Yeon-Sun; Hong, Kyeong-Man; Bae, Chang-Dae; Park, Joobae

2009-06-12

During mitosis, establishment of structurally and functionally sound bipolar spindles is necessary for maintaining the fidelity of chromosome segregation. Tumor-associated microtubule-associated protein (TMAP), also known as cytoskeleton-associated protein 2 (CKAP2), is a mitotic spindle-associated protein whose level is frequently up-regulated in various malignancies. Previous reports have suggested that TMAP is a potential regulator of mitotic spindle assembly and dynamics and that it is required for chromosome segregation to occur properly. So far, there have been no reports on how its mitosis-related functions are regulated. Here, we report that TMAP is hyper-phosphorylated at the C terminus specifically during mitosis. At least four different residues (Thr-578, Thr-596, Thr-622, and Ser-627) were responsible for the mitosis-specific phosphorylation of TMAP. Among these, Thr-622 was specifically phosphorylated by Cdk1-cyclin B1 both in vitro and in vivo. Interestingly, compared with the wild type, a phosphorylation-deficient mutant form of TMAP, in which Thr-622 had been replaced with an alanine (T622A), induced a significant increase in the frequency of metaphase cells with abnormal bipolar spindles, which often displayed disorganized, asymmetrical, or narrow and elongated morphologies. Formation of these abnormal bipolar spindles subsequently resulted in misalignment of metaphase chromosomes and ultimately caused a delay in the entry into anaphase. Moreover, such defects resulting from the T622A mutation were associated with a decrease in the rate of protein turnover at spindle microtubules. These findings suggest that Cdk1-cyclin B1-mediated phosphorylation of TMAP is important for and contributes to proper regulation of microtubule dynamics and establishment of functional bipolar spindles during mitosis.

Cdk1-Cyclin B1-mediated Phosphorylation of Tumor-associated Microtubule-associated Protein/Cytoskeleton-associated Protein 2 in Mitosis*

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Uk Hong, Kyung; Kim, Hyun-Jun; Kim, Hyo-Sil; Seong, Yeon-Sun; Hong, Kyeong-Man; Bae, Chang-Dae; Park, Joobae

2009-01-01

During mitosis, establishment of structurally and functionally sound bipolar spindles is necessary for maintaining the fidelity of chromosome segregation. Tumor-associated microtubule-associated protein (TMAP), also known as cytoskeleton-associated protein 2 (CKAP2), is a mitotic spindle-associated protein whose level is frequently up-regulated in various malignancies. Previous reports have suggested that TMAP is a potential regulator of mitotic spindle assembly and dynamics and that it is required for chromosome segregation to occur properly. So far, there have been no reports on how its mitosis-related functions are regulated. Here, we report that TMAP is hyper-phosphorylated at the C terminus specifically during mitosis. At least four different residues (Thr-578, Thr-596, Thr-622, and Ser-627) were responsible for the mitosis-specific phosphorylation of TMAP. Among these, Thr-622 was specifically phosphorylated by Cdk1-cyclin B1 both in vitro and in vivo. Interestingly, compared with the wild type, a phosphorylation-deficient mutant form of TMAP, in which Thr-622 had been replaced with an alanine (T622A), induced a significant increase in the frequency of metaphase cells with abnormal bipolar spindles, which often displayed disorganized, asymmetrical, or narrow and elongated morphologies. Formation of these abnormal bipolar spindles subsequently resulted in misalignment of metaphase chromosomes and ultimately caused a delay in the entry into anaphase. Moreover, such defects resulting from the T622A mutation were associated with a decrease in the rate of protein turnover at spindle microtubules. These findings suggest that Cdk1-cyclin B1-mediated phosphorylation of TMAP is important for and contributes to proper regulation of microtubule dynamics and establishment of functional bipolar spindles during mitosis. PMID:19369249

The C-terminal region of A-kinase anchor protein 350 (AKAP350A) enables formation of microtubule-nucleation centers and interacts with pericentriolar proteins.

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Kolobova, Elena; Roland, Joseph T; Lapierre, Lynne A; Williams, Janice A; Mason, Twila A; Goldenring, James R

2017-12-15

Microtubules in animal cells assemble (nucleate) from both the centrosome and the cis-Golgi cisternae. A-kinase anchor protein 350 kDa (AKAP350A, also called AKAP450/CG-NAP/AKAP9) is a large scaffolding protein located at both the centrosome and Golgi apparatus. Previous findings have suggested that AKAP350 is important for microtubule dynamics at both locations, but how this scaffolding protein assembles microtubule nucleation machinery is unclear. Here, we found that overexpression of the C-terminal third of AKAP350A, enhanced GFP-AKAP350A(2691-3907), induces the formation of multiple microtubule-nucleation centers (MTNCs). Nevertheless, these induced MTNCs lacked "true" centriole proteins, such as Cep135. Mapping analysis with AKAP350A truncations demonstrated that AKAP350A contains discrete regions responsible for promoting or inhibiting the formation of multiple MTNCs. Moreover, GFP-AKAP350A(2691-3907) recruited several pericentriolar proteins to MTNCs, including γ-tubulin, pericentrin, Cep68, Cep170, and Cdk5RAP2. Proteomic analysis indicated that Cdk5RAP2 and Cep170 both interact with the microtubule nucleation-promoting region of AKAP350A, whereas Cep68 interacts with the distal C-terminal AKAP350A region. Yeast two-hybrid assays established a direct interaction of Cep170 with AKAP350A. Super-resolution and deconvolution microscopy analyses were performed to define the association of AKAP350A with centrosomes, and these studies disclosed that AKAP350A spans the bridge between centrioles, co-localizing with rootletin and Cep68 in the linker region. siRNA-mediated depletion of AKAP350A caused displacement of both Cep68 and Cep170 from the centrosome. These results suggest that AKAP350A acts as a scaffold for factors involved in microtubule nucleation at the centrosome and coordinates the assembly of protein complexes associating with the intercentriolar bridge.

Tubulation of class II MHC compartments is microtubule dependent and involves multiple endolysosomal membrane proteins in primary dendritic cells.

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Vyas, Jatin M; Kim, You-Me; Artavanis-Tsakonas, Katerina; Love, J Christopher; Van der Veen, Annemarthe G; Ploegh, Hidde L

2007-06-01

Immature dendritic cells (DCs) capture exogenous Ags in the periphery for eventual processing in endolysosomes. Upon maturation by TLR agonists, DCs deliver peptide-loaded class II MHC molecules from these compartments to the cell surface via long tubular structures (endolysosomal tubules). The nature and rules that govern the movement of these DC compartments are unknown. In this study, we demonstrate that the tubules contain multiple proteins including the class II MHC molecules and LAMP1, a lysosomal resident protein, as well as CD63 and CD82, members of the tetraspanin family. Endolysosomal tubules can be stained with acidotropic dyes, indicating that they are extensions of lysosomes. However, the proper trafficking of class II MHC molecules themselves is not necessary for endolysosomal tubule formation. DCs lacking MyD88 can also form endolysosomal tubules, demonstrating that MyD88-dependent TLR activation is not necessary for the formation of this compartment. Endolysosomal tubules in DCs exhibit dynamic and saltatory movement, including bidirectional travel. Measured velocities are consistent with motor-based movement along microtubules. Indeed, nocodazole causes the collapse of endolysosomal tubules. In addition to its association with microtubules, endolysosomal tubules follow the plus ends of microtubules as visualized in primary DCs expressing end binding protein 1 (EB1)-enhanced GFP.

Reassessing the roles of PIN proteins and anticlinal microtubules during pavement cell morphogenesis

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Belteton, Samuel; Sawchuk, Megan G.; Donohoe, Bryon S.; Scarpella, Enrico; Szymanski, Daniel B.

2017-11-30

The leaf epidermis is a biomechanical shell that influences the size and shape of the organ. Its morphogenesis is a multiscale process in which nanometer-scale cytoskeletal protein complexes, individual cells, and groups of cells pattern growth and define macroscopic leaf traits. Interdigitated growth of neighboring cells is an evolutionarily conserved developmental strategy. Understanding how signaling pathways and cytoskeletal proteins pattern cell walls during this form of tissue morphogenesis is an important research challenge. The cellular and molecular control of a lobed cell morphology is currently thought to involve PIN-FORMED (PIN)-type plasma membrane efflux carriers that generate subcellular auxin gradients. Auxin gradients were proposed to function across cell boundaries to encode stable offset patterns of cortical microtubules and actin filaments between adjacent cells. Many models suggest that long-lived microtubules along the anticlinal cell wall generate local cell wall heterogeneities that restrict local growth and specify the timing and location of lobe formation. Here we used Arabidopsis reverse genetics and multivariate long-term time-lapse imaging to test current cell shape control models. We found that neither PIN proteins nor microtubules along the anticlinal wall predict the patterns of lobe formation. In fields of lobing cells, anticlinal microtubules are not correlated with cell shape and are unstable at the time scales of cell expansion. Our analyses indicate that anticlinal microtubules have multiple functions in pavement cells, and that lobe initiation is likely controlled by complex interactions among cell geometry, cell wall stress patterns, and transient microtubule networks that span the anticlinal and periclinal walls.

Trait diversity promotes stability of community dynamics

DEFF Research Database (Denmark)

Zhang, Lai; Thygesen, Uffe Høgsbro; Knudsen, Kim

2013-01-01

body size. The dynamic properties of the models are described by a stability analysis of equilibrium solutions and by the non-equilibrium dynamics. We find that the introduction of trait diversity expands the set of parameters for which the equilibrium is stable and, if the community is unstable, makes....... The analysis is performed by comparing the properties of two size spectrum models. The first model considers all individuals as belonging to the same “average” species, i.e., without a description of diversity. The second model introduces diversity by further considering individuals by a trait, here asymptotic...

Direct incorporation of guanosine 5'-diphosphate into microtubules without guanosine 5'-triphosphate hydrolysis

International Nuclear Information System (INIS)

Hamel, E.; Batra, J.K.; Lin, C.M.

1986-01-01

Using highly purified calf brain tubulin bearing [8- 14 C]guanosine 5'-diphosphate (GDP) in the exchangeable nucleotide site and heat-treated microtubule-associated proteins, the authors have found that a significant proportion of exchangeable-site GDP in microtubules can be incorporated directly during guanosine 5'-triphosphate (GTP) dependent polymerization of tubulin, without an initial exchange of GDP for GTP and subsequent GTP hydrolysis during assembly. The precise amount of GDP incorporated directly into microtubules is highly dependent on specific reaction conditions, being favored by high tubulin concentrations, low GTP and Mg 2+ concentrations, and exogenous GDP in the reaction mixture. Minimum effects were observed with changes in reaction pH or temperature, changes in concentration of microtubule-associated proteins, alteration of the sulfonate buffer, or the presence of a calcium chelator in the reaction mixture. Under conditions most favorable for direct GDP incorporation, about one-third of the GDP in microtubules is incorporated directly (without GTP hydrolysis) and two-thirds is incorporated hydrolytically (as a consequence of GTP hydrolysis). Direct incorporation of GDP occurs in a constant proportion throughout elongation, and the amount of direct incorporation probably reflects the rapid equilibration of GDP and GTP at the exchangeable site that occurs before the onset of assembly

Assessment of stability during gait in patients with spinal deformity-A preliminary analysis using the dynamic stability margin.

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Simon, Anne-Laure; Lugade, Vipul; Bernhardt, Kathie; Larson, A Noelle; Kaufman, Kenton

2017-06-01

Daily living activities are dynamic, requiring spinal motion through space. Current assessment of spinal deformities is based on static measurements from full-spine standing radiographs. Tools to assess dynamic stability during gait might be useful to enhance the standard evaluation. The aim of this study was to evaluate gait dynamic imbalance in patients with spinal deformity using the dynamic stability margin (DSM). Twelve normal subjects and 17 patients with spinal deformity were prospectively recruited. A kinematic 3D gait analysis was performed for the control group (CG) and the spinal deformity group (SDG). The DSM (distance between the extrapolated center of mass and the base of support) and time-distance parameters were calculated for the right and left side during gait. The relationship between DSM and step length was assessed using three variables: gait stability, symmetry, and consistency. Variables' accuracy was validated by a discriminant analysis. Patients with spinal deformity exhibited gait instability according to the DSM (0.25m versus 0.31m) with decreased velocity (1.1ms -1 versus 1.3ms -1 ) and decreased step length (0.32m versus 0.38m). According to the discriminant analysis, gait stability was the more accurate variable (area under the curve AUC=0.98) followed by gait symmetry and consistency. However, gait consistency showed 100% of specificity, sensitivity, and accuracy of precision. The DSM showed that patients with spinal malalignment exhibit decreased gait stability, symmetry, and consistency besides gait time-distance parameter changes. Additional work is required to determine how to apply the DSM for preoperative and postoperative spinal deformity management. Copyright © 2017. Published by Elsevier B.V.

Dynamics and Stability of Permanent-Magnet Synchronous Motor

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Ren He

2017-01-01

Full Text Available The aim of this article is to explore the dynamic characteristics and stability of the permanent-magnet synchronous motor (PMSM. PMSM equilibrium local stability condition and Hopf  bifurcation condition, pitchfork bifurcation condition, and fold bifurcation condition have been derived by using the Routh-Hurwitz criterion and the bifurcation theory, respectively. Bifurcation curves of the equilibrium with single and double parameters are obtained by continuation method. Numerical simulations not only confirm the theoretical analysis results but also show one kind of codimension-two-bifurcation points of the equilibrium. PMSM, with or without external load, can exhibit rich dynamic behaviors in different parameters regions. It is shown that if unstable equilibrium appears in the parameters regions, the PMSM may not be able to work stably. To ensure the PMSMs work stably, the inherent parameters should be designed in the region which has only one stable equilibrium.

CEP295 interacts with microtubules and is required for centriole elongation.

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Chang, Ching-Wen; Hsu, Wen-Bin; Tsai, Jhih-Jie; Tang, Chieh-Ju C; Tang, Tang K

2016-07-01

Centriole duplication is a tightly ordered process during which procentrioles are assembled in G1-S and elongate during S and G2. Here, we show that human CEP295 (Drosophila Ana1) is not essential for initial cartwheel assembly, but is required to build distal half centrioles during S and G2. Using super-resolution and immunogold electron microscopy, we demonstrate that CEP295 is recruited to the proximal end of procentrioles in early S phase, when it is also localized at the centriolar microtubule wall that surrounds the human SAS6 cartwheel hub. Interestingly, depletion of CEP295 not only inhibits the recruitments of POC5 and POC1B to the distal half centrioles in G2, resulting in shorter centrioles, it also blocks the post-translational modification of centriolar microtubules (e.g. acetylation and glutamylation). Importantly, our results indicate that CEP295 directly interacts with microtubules, and that excess CEP295 could induce the assembly of overly long centrioles. Furthermore, exogenous expression of the N-terminal domain of CEP295 exerts a dominant-negative effect on centriole elongation. Collectively, these findings suggest that CEP295 is essential for building the distal half centrioles and for post-translational modification of centriolar microtubules. © 2016. Published by The Company of Biologists Ltd.

Challenges and opportunities in the high-resolution cryo-EM visualization of microtubules and their binding partners.

Science.gov (United States)

Nogales, Eva; Kellogg, Elizabeth H

2017-10-01

As non-crystallizable polymers, microtubules have been the target of cryo-electron microscopy (cryo-EM) studies since the technique was first established. Over the years, image processing strategies have been developed that take care of the unique, pseudo-helical symmetry of the microtubule. With recent progress in data quality and data processing, cryo-EM reconstructions are now reaching resolutions that allow the generation of atomic models of microtubules and the factors that bind them. These include cellular partners that contribute to microtubule cellular functions, or small ligands that interfere with those functions in the treatment of cancer. The stage is set to generate a family portrait for all identified microtubule interacting proteins and to use cryo-EM as a drug development tool in the targeting of tubulin. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

Stability in dynamical systems I

International Nuclear Information System (INIS)

Courant, E.D.; Ruth, R.D.; Weng, W.T.

1984-08-01

We have reviewed some of the basic techniques which can be used to analyze stability in nonlinear dynamical systems, particularly in circular particle accelerators. We have concentrated on one-dimensional systems in the examples in order to simply illustrate the general techniques. We began with a review of Hamiltonian dynamics and canonical transformations. We then reviewed linear equations with periodic coefficients using the basic techniques from accelerator theory. To handle nonlinear terms we developed a canonical perturbation theory. From this we calculated invariants and the amplitude dependence of the frequency. This led us to resonances. We studied the cubic resonance in detail by using a rotating coordinate system in phase space. We then considered a general isolated nonlinear resonance. In this case we calculated the width of the resonance and estimated the spacing of resonances in order to use the Chirikov criterion to restrict the validity of the analysis. Finally the resonance equation was reduced to the pendulum equation, and we examined the motion on a separatrix. This brought us to the beginnings of stochastic behavior in the neighborhood of the separatrix. It is this complex behavior in the neighborhood of the separatrix which causes the perturbation theory used here to diverge in many cases. In spite of this the methods developed here have been and are used quite successfully to study nonlinear effects in nearly integrable systems. When used with caution and in conjunction with numerical work they give tremendous insight into the nature of the phase space structure and the stability of nonlinear differential equations. 14 references

Dynamical stability of the holographic system with two competing orders

Energy Technology Data Exchange (ETDEWEB)

Du, Yiqiang [School of Physics, University of Chinese Academy of Sciences,Beijing 100049 (China); Lan, Shan-Quan [Department of Physics, Beijing Normal University,Beijing 100875 (China); Tian, Yu [School of Physics, University of Chinese Academy of Sciences,Beijing 100049 (China); State Key Laboratory of Theoretical Physics,Institute of Theoretical Physics, Chinese Academy of Science,Beijing 100190 (China); Zhang, Hongbao [Department of Physics, Beijing Normal University,Beijing 100875 (China); Theoretische Natuurkunde, Vrije Universiteit Brussel and The International Solvay Institutes,Pleinlaan 2, B-1050 Brussels (Belgium)

2016-01-04

We investigate the dynamical stability of the holographic system with two order parameters, which exhibits competition and coexistence of condensations. In the linear regime, we have developed the gauge dependent formalism to calculate the quasi-normal modes by gauge fixing, which turns out be considerably convenient. Furthermore, by giving different Gaussian wave packets as perturbations at the initial time, we numerically evolve the full nonlinear system until it arrives at the final equilibrium state. Our results show that the dynamical stability is consistent with the thermodynamical stability. Interestingly, the dynamical evolution, as well as the quasi-normal modes, shows that the relaxation time of this model is generically much longer than the simplest holographic system. We also find that the late time behavior can be well captured by the lowest lying quasi-normal modes except for the non-vanishing order towards the single ordered phase. To our knowledge, this exception is the first counter example to the general belief that the late time behavior towards a final stable state can be captured by the lowest lying quasi-normal modes. In particular, a double relation is found for this exception in certain cases.

The Emerging Role of the Cytoskeleton in Chromosome Dynamics

Directory of Open Access Journals (Sweden)

Maya Spichal

2017-05-01

Full Text Available Chromosomes underlie a dynamic organization that fulfills functional roles in processes like transcription, DNA repair, nuclear envelope stability, and cell division. Chromosome dynamics depend on chromosome structure and cannot freely diffuse. Furthermore, chromosomes interact closely with their surrounding nuclear environment, which further constrains chromosome dynamics. Recently, several studies enlighten that cytoskeletal proteins regulate dynamic chromosome organization. Cytoskeletal polymers that include actin filaments, microtubules and intermediate filaments can connect to the nuclear envelope via Linker of the Nucleoskeleton and Cytoskeleton (LINC complexes and transfer forces onto chromosomes inside the nucleus. Monomers of these cytoplasmic polymers and related proteins can also enter the nucleus and play different roles in the interior of the nucleus than they do in the cytoplasm. Nuclear cytoskeletal proteins can act as chromatin remodelers alone or in complexes with other nuclear proteins. They can also act as transcription factors. Many of these mechanisms have been conserved during evolution, indicating that the cytoskeletal regulation of chromosome dynamics is an essential process. In this review, we discuss the different influences of cytoskeletal proteins on chromosome dynamics by focusing on the well-studied model organism budding yeast.

Ase1p Organizes Antiparallel Microtubule Arrays during Interphase and Mitosis in Fission YeastV⃞

OpenAIRE

Loïodice, Isabelle; Staub, Jayme; Setty, Thanuja Gangi; Nguyen, Nam-Phuong T.; Paoletti, Anne; Tran, P. T.

2005-01-01

Proper microtubule organization is essential for cellular processes such as organelle positioning during interphase and spindle formation during mitosis. The fission yeast Schizosaccharomyces pombe presents a good model for understanding microtubule organization. We identify fission yeast ase1p, a member of the conserved ASE1/PRC1/MAP65 family of microtubule bundling proteins, which functions in organizing the spindle midzone during mitosis. Using fluorescence live cell imaging, we show that ...

Handbook of electrical power system dynamics modeling, stability, and control

CERN Document Server

Eremia, Mircea

2013-01-01

Complete guidance for understanding electrical power system dynamics and blackouts This handbook offers a comprehensive and up-to-date treatment of power system dynamics. Addressing the full range of topics, from the fundamentals to the latest technologies in modeling, stability, and control, Handbook of Electrical Power System Dynamics provides engineers with hands-on guidance for understanding the phenomena leading to blackouts so they can design the most appropriate solutions for a cost-effective and reliable operation. Focusing on system dynamics, the book details

Reassessing the Roles of PIN Proteins and Anticlinal Microtubules during Pavement Cell Morphogenesis.

Science.gov (United States)

Belteton, Samuel A; Sawchuk, Megan G; Donohoe, Bryon S; Scarpella, Enrico; Szymanski, Daniel B

2018-01-01

The leaf epidermis is a biomechanical shell that influences the size and shape of the organ. Its morphogenesis is a multiscale process in which nanometer-scale cytoskeletal protein complexes, individual cells, and groups of cells pattern growth and define macroscopic leaf traits. Interdigitated growth of neighboring cells is an evolutionarily conserved developmental strategy. Understanding how signaling pathways and cytoskeletal proteins pattern cell walls during this form of tissue morphogenesis is an important research challenge. The cellular and molecular control of a lobed cell morphology is currently thought to involve PIN-FORMED (PIN)-type plasma membrane efflux carriers that generate subcellular auxin gradients. Auxin gradients were proposed to function across cell boundaries to encode stable offset patterns of cortical microtubules and actin filaments between adjacent cells. Many models suggest that long-lived microtubules along the anticlinal cell wall generate local cell wall heterogeneities that restrict local growth and specify the timing and location of lobe formation. Here, we used Arabidopsis ( Arabidopsis thaliana ) reverse genetics and multivariate long-term time-lapse imaging to test current cell shape control models. We found that neither PIN proteins nor long-lived microtubules along the anticlinal wall predict the patterns of lobe formation. In fields of lobing cells, anticlinal microtubules are not correlated with cell shape and are unstable at the time scales of cell expansion. Our analyses indicate that anticlinal microtubules have multiple functions in pavement cells and that lobe initiation is likely controlled by complex interactions among cell geometry, cell wall stress patterns, and transient microtubule networks that span the anticlinal and periclinal walls. © 2018 American Society of Plant Biologists. All Rights Reserved.

Adaptive Dynamic Programming for Control Algorithms and Stability

CERN Document Server

Zhang, Huaguang; Luo, Yanhong; Wang, Ding

2013-01-01

There are many methods of stable controller design for nonlinear systems. In seeking to go beyond the minimum requirement of stability, Adaptive Dynamic Programming for Control approaches the challenging topic of optimal control for nonlinear systems using the tools of  adaptive dynamic programming (ADP). The range of systems treated is extensive; affine, switched, singularly perturbed and time-delay nonlinear systems are discussed as are the uses of neural networks and techniques of value and policy iteration. The text features three main aspects of ADP in which the methods proposed for stabilization and for tracking and games benefit from the incorporation of optimal control methods: • infinite-horizon control for which the difficulty of solving partial differential Hamilton–Jacobi–Bellman equations directly is overcome, and  proof provided that the iterative value function updating sequence converges to the infimum of all the value functions obtained by admissible control law sequences; • finite-...

Tubulation of Class II MHC Compartments Is Microtubule Dependent and Involves Multiple Endolysosomal Membrane Proteins in Primary Dendritic Cells1

Science.gov (United States)

Vyas, Jatin M.; Kim, You-Me; Artavanis-Tsakonas, Katerina; Love, J. Christopher; Van der Veen, Annemarthe G.; Ploegh, Hidde L.

2009-01-01

Immature dendritic cells (DCs) capture exogenous Ags in the periphery for eventual processing in endolysosomes. Upon maturation by TLR agonists, DCs deliver peptide-loaded class II MHC molecules from these compartments to the cell surface via long tubular structures (endolysosomal tubules). The nature and rules that govern the movement of these DC compartments are unknown. In this study, we demonstrate that the tubules contain multiple proteins including the class II MHC molecules and LAMP1, a lysosomal resident protein, as well as CD63 and CD82, members of the tetraspanin family. Endolysosomal tubules can be stained with acidotropic dyes, indicating that they are extensions of lysosomes. However, the proper trafficking of class II MHC molecules themselves is not necessary for endolysosomal tubule formation. DCs lacking MyD88 can also form endolysosomal tubules, demonstrating that MyD88-dependent TLR activation is not necessary for the formation of this compartment. Endolysosomal tubules in DCs exhibit dynamic and saltatory movement, including bidirectional travel. Measured velocities are consistent with motor-based movement along microtubules. Indeed, nocodazole causes the collapse of endolysosomal tubules. In addition to its association with microtubules, endolysosomal tubules follow the plus ends of microtubules as visualized in primary DCs expressing end binding protein 1 (EB1)-enhanced GFP. PMID:17513769

Cdk1-Cyclin B1-mediated Phosphorylation of Tumor-associated Microtubule-associated Protein/Cytoskeleton-associated Protein 2 in Mitosis*

OpenAIRE

Uk Hong, Kyung; Kim, Hyun-Jun; Kim, Hyo-Sil; Seong, Yeon-Sun; Hong, Kyeong-Man; Bae, Chang-Dae; Park, Joobae

2009-01-01

During mitosis, establishment of structurally and functionally sound bipolar spindles is necessary for maintaining the fidelity of chromosome segregation. Tumor-associated microtubule-associated protein (TMAP), also known as cytoskeleton-associated protein 2 (CKAP2), is a mitotic spindle-associated protein whose level is frequently up-regulated in various malignancies. Previous reports have suggested that TMAP is a potential regulator of mitotic spindle assembly and dynamics and that it is re...

Microtubule-Mediated Inositol Lipid Signaling Plays Critical Roles in Regulation of Blebbing.

Directory of Open Access Journals (Sweden)

Tatsuroh Sugiyama

Full Text Available Cells migrate by extending pseudopods such as lamellipodia and blebs. Although the signals leading to lamellipodia extension have been extensively investigated, those for bleb extension remain unclear. Here, we investigated signals for blebbing in Dictyostelium cells using a newly developed assay to induce blebbing. When cells were cut into two pieces with a microneedle, the anucleate fragments vigorously extended blebs. This assay enabled us to induce blebbing reproducibly, and analyses of knockout mutants and specific inhibitors identified candidate molecules that regulate blebbing. Blebs were also induced in anucleate fragments of leukocytes, indicating that this assay is generally applicable to animal cells. After cutting, microtubules in the anucleate fragments promptly depolymerized, followed by the extension of blebs. Furthermore, when intact cells were treated with a microtubule inhibitor, they frequently extended blebs. The depolymerization of microtubules induced the delocalization of inositol lipid phosphatidylinositol 3,4,5-trisphosphate from the cell membrane. PI3 kinase-null cells frequently extended blebs, whereas PTEN-null cells extended fewer blebs. From these observations, we propose a model in which microtubules play a critical role in bleb regulation via inositol lipid metabolism.

Designing Inter-Organisational Collectivities for Dynamic Fit: Stability, maneuvrability and Application in Disaster Relief Endeavours

Science.gov (United States)

2011-01-01

changed consumer preferences . Hence, static stability limits initial performance deviation (e.g., maintaining desired airplane altitude, maintaining...by changed consumer preferences . Hence, dynamic stability limits the duration of performance deviation (e.g., maintaining desired airplane altitude...altitude from wind gust. Initial resistance to deviation in profit level from change in consumer preferences . Dynamic stability Quickness of a

Dynamic trunk stabilization: a conceptual back injury prevention program for volleyball athletes.

Science.gov (United States)

Smith, Chad E; Nyland, John; Caudill, Paul; Brosky, Joseph; Caborn, David N M

2008-11-01

The sport of volleyball creates considerable dynamic trunk stability demands. Back injury occurs all too frequently in volleyball, particularly among female athletes. The purpose of this clinical commentary is to review functional anatomy, muscle coactivation strategies, assessment of trunk muscle performance, and the characteristics of effective exercises for the trunk or core. From this information, a conceptual progressive 3-phase volleyball-specific training program is presented to improve dynamic trunk stability and to potentially reduce the incidence of back injury among volleyball athletes. Phase 1 addresses low-velocity motor control, kinesthetic awareness, and endurance, with the clinician providing cues to teach achievement of biomechanically neutral spine alignment. Phase 2 focuses on progressively higher velocity dynamic multiplanar endurance, coordination, and strength-power challenges integrating upper and lower extremity movements, while maintaining neutral spine alignment. Phase 3 integrates volleyball-specific skill simulations by breaking down composite movement patterns into their component parts, with differing dynamic trunk stability requirements, while maintaining neutral spine alignment. Prospective research is needed to validate the efficacy of this program.

Microtubule-Targeting Agents Enter the Central Nervous System (CNS): Double-edged Swords for Treating CNS Injury and Disease.

Science.gov (United States)

Hur, Eun-Mi; Lee, Byoung Dae

2014-12-01

Microtubules have been among the most successful targets in anticancer therapy and a large number of microtubule-targeting agents (MTAs) are in various stages of clinical development for the treatment of several malignancies. Given that injury and diseases in the central nervous system (CNS) are accompanied by acute or chronic disruption of the structural integrity of neurons and that microtubules provide structural support for the nervous system at cellular and intracellular levels, microtubules are emerging as potential therapeutic targets for treating CNS disorders. It has been postulated that exogenous application of MTAs might prevent the breakdown or degradation of microtubules after injury or during neurodegeneration, which will thereby aid in preserving the structural integrity and function of the nervous system. Here we review recent evidence that supports this notion and also discuss potential risks of targeting microtubules as a therapy for treating nerve injury and neurodegenerative diseases.

Microtubule-Targeting Agents Enter the Central Nervous System (CNS: Double-edged Swords for Treating CNS Injury and Disease

Directory of Open Access Journals (Sweden)

Eun-Mi Hur

2014-12-01

Full Text Available Microtubules have been among the most successful targets in anticancer therapy and a large number of microtubule-targeting agents (MTAs are in various stages of clinical development for the treatment of several malignancies. Given that injury and diseases in the central nervous system (CNS are accompanied by acute or chronic disruption of the structural integrity of neurons and that microtubules provide structural support for the nervous system at cellular and intracellular levels, microtubules are emerging as potential therapeutic targets for treating CNS disorders. It has been postulated that exogenous application of MTAs might prevent the breakdown or degradation of microtubules after injury or during neurodegeneration, which will thereby aid in preserving the structural integrity and function of the nervous system. Here we review recent evidence that supports this notion and also discuss potential risks of targeting microtubules as a therapy for treating nerve injury and neurodegenerative diseases.

Fast Dynamic Simulation-Based Small Signal Stability Assessment and Control

Energy Technology Data Exchange (ETDEWEB)

Acharya, Naresh [General Electric Company, Fairfield, CT (United States); Baone, Chaitanya [General Electric Company, Fairfield, CT (United States); Veda, Santosh [General Electric Company, Fairfield, CT (United States); Dai, Jing [General Electric Company, Fairfield, CT (United States); Chaudhuri, Nilanjan [General Electric Company, Fairfield, CT (United States); Leonardi, Bruno [General Electric Company, Fairfield, CT (United States); Sanches-Gasca, Juan [General Electric Company, Fairfield, CT (United States); Diao, Ruisheng [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Wu, Di [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Huang, Zhenyu [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Zhang, Yu [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Jin, Shuangshuang [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Zheng, Bin [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Chen, Yousu [Pacific Northwest National Lab. (PNNL), Richland, WA (United States)

2014-12-31

Power grid planning and operation decisions are made based on simulation of the dynamic behavior of the system. Enabling substantial energy savings while increasing the reliability of the aging North American power grid through improved utilization of existing transmission assets hinges on the adoption of wide-area measurement systems (WAMS) for power system stabilization. However, adoption of WAMS alone will not suffice if the power system is to reach its full entitlement in stability and reliability. It is necessary to enhance predictability with "faster than real-time" dynamic simulations that will enable the dynamic stability margins, proactive real-time control, and improve grid resiliency to fast time-scale phenomena such as cascading network failures. Present-day dynamic simulations are performed only during offline planning studies, considering only worst case conditions such as summer peak, winter peak days, etc. With widespread deployment of renewable generation, controllable loads, energy storage devices and plug-in hybrid electric vehicles expected in the near future and greater integration of cyber infrastructure (communications, computation and control), monitoring and controlling the dynamic performance of the grid in real-time would become increasingly important. The state-of-the-art dynamic simulation tools have limited computational speed and are not suitable for real-time applications, given the large set of contingency conditions to be evaluated. These tools are optimized for best performance of single-processor computers, but the simulation is still several times slower than real-time due to its computational complexity. With recent significant advances in numerical methods and computational hardware, the expectations have been rising towards more efficient and faster techniques to be implemented in power system simulators. This is a natural expectation, given that the core solution algorithms of most commercial simulators were developed

Molecular architecture of axonemal microtubule doublets revealedby cryo-electron tomography

Energy Technology Data Exchange (ETDEWEB)

Sui, Haixin; Downing, Kenneth H.

2006-05-22

The axoneme, which forms the core of eukaryotic flagella and cilia, is one of the largest macromolecular machines with a structure that is largely conserved from protists to mammals. Microtubule doublets are structural components of axonemes containing a number of proteins besides tubulin, and are usually found in arrays of nine doublets arranged around two singlet microtubules. Coordinated sliding of adjacent doublets, which involves a host of other proteins in the axoneme, produces periodic beating movements of the axoneme. We have obtained a 3D density map of intact microtubule doublets using cryo-electron tomography and image averaging. Our map, with a resolution of about 3 nm, provides insights into locations of particular proteins within the doublets and the structural features of the doublets that define their mechanical properties. We identify likely candidates for several of these non-tubulin components of the doublets. This work offers novel insight on how tubulin protofilaments and accessory proteins attach together to form the doublets and provides a structural basis for understanding doublet function in axonemes.

Hexavalent chromium-induced differential disruption of cortical microtubules in some Fabaceae species is correlated with acetylation of α-tubulin.

Science.gov (United States)

Eleftheriou, Eleftherios P; Adamakis, Ioannis-Dimosthenis S; Michalopoulou, Vasiliki A

2016-03-01

The effects of hexavalent chromium [Cr(VI)] on the cortical microtubules (MTs) of five species of the Fabaceae family (Vicia faba, Pisum sativum, Vigna sinensis, Vigna angularis, and Medicago sativa) were investigated by confocal laser scanning microscopy after immunolocalization of total tubulin with conventional immunofluorescence techniques and of acetylated α-tubulin with the specific 6-11B-1 monoclonal antibody. Moreover, total α-tubulin and acetylated α-tubulin were quantified by Western immunoblotting and scanning densitometry. Results showed the universality of Cr(VI) detrimental effects to cortical MTs, which proved to be a sensitive and reliable subcellular marker for monitoring Cr(VI) toxicity in plant cells. However, a species-specific response was recorded, and a correlation of MT disturbance with the acetylation status of α-tubulin was demonstrated. In V. faba, MTs were depolymerized at the gain of cytoplasmic tubulin background and displayed low α-tubulin acetylation, while in P. sativum, V. sinensis, V. angularis, and M. sativa, MTs became bundled and changed orientation from perpendicular to oblique or longitudinal. Bundled MTs were highly acetylated as determined by both immunofluorescence and Western immunoblotting. Tubulin acetylation in P. sativum and M. sativa preceded MT bundling; in V. sinensis it followed MT derangement, while in V. angularis the two phenomena coincided. Total α-tubulin remained constant in all treatments. Should acetylation be an indicator of MT stabilization, it is deduced that bundled MTs became stabilized, lost their dynamic properties, and were rendered inactive. Results of this report allow the conclusion that Cr(VI) toxicity disrupts MTs and deranges the MT-mediated functions either by depolymerizing or stabilizing them.

Dynamic Stability of Pipe Conveying Fluid with Crack and Attached Masses

International Nuclear Information System (INIS)

Ahn, Tae Soo; Yoon, Han Ik; Son, In Soo; Ahn, Sung Jin

2007-01-01

In this paper, the dynamic stability of a cracked simply supported pipe conveying fluid with an attached mass is investigated. Also, the effect of attached masses on the dynamic stability of a simply supported pipe conveying fluid is presented for the different positions and depth of the crack. Based on the Euler-Bernoulli beam theory, the equation of motion can be constructed by the energy expressions using extended Hamilton's principle. The crack section is represented by a local flexibility matrix connecting two undamaged pipe segments. The crack is assumed to be in the first mode of a fracture and to be always opened during the vibrations. Finally, the critical flow velocities and stability maps of the pipe conveying fluid are obtained by changing the attached masses and crack severity

Differentiable dynamical systems an introduction to structural stability and hyperbolicity

CERN Document Server

Wen, Lan

2016-01-01

This is a graduate text in differentiable dynamical systems. It focuses on structural stability and hyperbolicity, a topic that is central to the field. Starting with the basic concepts of dynamical systems, analyzing the historic systems of the Smale horseshoe, Anosov toral automorphisms, and the solenoid attractor, the book develops the hyperbolic theory first for hyperbolic fixed points and then for general hyperbolic sets. The problems of stable manifolds, structural stability, and shadowing property are investigated, which lead to a highlight of the book, the \\Omega-stability theorem of Smale. While the content is rather standard, a key objective of the book is to present a thorough treatment for some tough material that has remained an obstacle to teaching and learning the subject matter. The treatment is straightforward and hence could be particularly suitable for self-study. Selected solutions are available electronically for instructors only. Please send email to textbooks@ams.org for more informatio...

Stabilization of computational procedures for constrained dynamical systems

Science.gov (United States)

Park, K. C.; Chiou, J. C.

1988-01-01

A new stabilization method of treating constraints in multibody dynamical systems is presented. By tailoring a penalty form of the constraint equations, the method achieves stabilization without artificial damping and yields a companion matrix differential equation for the constraint forces; hence, the constraint forces are obtained by integrating the companion differential equation for the constraint forces in time. A principal feature of the method is that the errors committed in each constraint condition decay with its corresponding characteristic time scale associated with its constraint force. Numerical experiments indicate that the method yields a marked improvement over existing techniques.

Local Dynamic Stability Assessment of Motion Impaired Elderly Using Electronic Textile Pants.

Science.gov (United States)

Liu, Jian; Lockhart, Thurmon E; Jones, Mark; Martin, Tom

2008-10-01

A clear association has been demonstrated between gait stability and falls in the elderly. Integration of wearable computing and human dynamic stability measures into home automation systems may help differentiate fall-prone individuals in a residential environment. The objective of the current study was to evaluate the capability of a pair of electronic textile (e-textile) pants system to assess local dynamic stability and to differentiate motion-impaired elderly from their healthy counterparts. A pair of e-textile pants comprised of numerous e-TAGs at locations corresponding to lower extremity joints was developed to collect acceleration, angular velocity and piezoelectric data. Four motion-impaired elderly together with nine healthy individuals (both young and old) participated in treadmill walking with a motion capture system simultaneously collecting kinematic data. Local dynamic stability, characterized by maximum Lyapunov exponent, was computed based on vertical acceleration and angular velocity at lower extremity joints for the measurements from both e-textile and motion capture systems. Results indicated that the motion-impaired elderly had significantly higher maximum Lyapunov exponents (computed from vertical acceleration data) than healthy individuals at the right ankle and hip joints. In addition, maximum Lyapunov exponents assessed by the motion capture system were found to be significantly higher than those assessed by the e-textile system. Despite the difference between these measurement techniques, attaching accelerometers at the ankle and hip joints was shown to be an effective sensor configuration. It was concluded that the e-textile pants system, via dynamic stability assessment, has the potential to identify motion-impaired elderly.

Investigation of biomechanical behavior of lumbar vertebral segments with dynamic stabilization device using finite element approach

Science.gov (United States)

Deoghare, Ashish B.; Kashyap, Siddharth; Padole, Pramod M.

2013-03-01

Degenerative disc disease is a major source of lower back pain and significantly alters the biomechanics of the lumbar spine. Dynamic stabilization device is a remedial technique which uses flexible materials to stabilize the affected lumbar region while preserving the natural anatomy of the spine. The main objective of this research work is to investigate the stiffness variation of dynamic stabilization device under various loading conditions under compression, axial rotation and flexion. Three dimensional model of the two segment lumbar spine is developed using computed tomography (CT) scan images. The lumbar structure developed is analyzed in ANSYS workbench. Two types of dynamic stabilization are considered: one with stabilizing device as pedicle instrumentation and second with stabilization device inserted around the inter-vertebral disc. Analysis suggests that proper positioning of the dynamic stabilization device is of paramount significance prior to the surgery. Inserting the device in the posterior region indicates the adverse effects as it shows increase in the deformation of the inter-vertebral disc. Analysis executed by positioning stabilizing device around the inter-vertebral disc yields better result for various stiffness values under compression and other loadings. [Figure not available: see fulltext.

Dynamic large eddy simulation: Stability via realizability

Science.gov (United States)

Mokhtarpoor, Reza; Heinz, Stefan

2017-10-01

The concept of dynamic large eddy simulation (LES) is highly attractive: such methods can dynamically adjust to changing flow conditions, which is known to be highly beneficial. For example, this avoids the use of empirical, case dependent approximations (like damping functions). Ideally, dynamic LES should be local in physical space (without involving artificial clipping parameters), and it should be stable for a wide range of simulation time steps, Reynolds numbers, and numerical schemes. These properties are not trivial, but dynamic LES suffers from such problems over decades. We address these questions by performing dynamic LES of periodic hill flow including separation at a high Reynolds number Re = 37 000. For the case considered, the main result of our studies is that it is possible to design LES that has the desired properties. It requires physical consistency: a PDF-realizable and stress-realizable LES model, which requires the inclusion of the turbulent kinetic energy in the LES calculation. LES models that do not honor such physical consistency can become unstable. We do not find support for the previous assumption that long-term correlations of negative dynamic model parameters are responsible for instability. Instead, we concluded that instability is caused by the stable spatial organization of significant unphysical states, which are represented by wall-type gradient streaks of the standard deviation of the dynamic model parameter. The applicability of our realizability stabilization to other dynamic models (including the dynamic Smagorinsky model) is discussed.

Cognitive-motor dual-task interference modulates mediolateral dynamic stability during gait in post-stroke individuals.

Science.gov (United States)

Tisserand, R; Armand, S; Allali, G; Schnider, A; Baillieul, S

2018-04-01

Gait asymmetry and dynamic balance impairments observed in post-stroke individuals increase their risk of fall. Moreover, walking while performing a cognitive task (i.e. dual-task) disturbs the control of balance in post-stroke individuals. Here we investigated the mediolateral dynamic stability in twenty-two community-dwelling participants (12 post-strokes and 10 healthy controls) while walking in single-task (normal gait) and four different dual-tasks (cognitive-motor interference). Positions of the extrapolated center of mass and mediolateral widths of both margin of stability and base of support were extracted from 35 marker trajectories. Post-stroke participants presented larger margin of stability and base of support than controls during single-task (both p dual-task was found between groups. In post-stroke participants, dual-task induced slight modification of the mediolateral stability strategy, as the margin of stability was not different between the two limbs at foot-strike, and significantly reduced the performance in every cognitive task. Post-stroke participants increased their dynamic stability in the frontal plane in single-task by extending their base of support and mainly relying on their non-paretic limb. Under cognitive-motor interference (dual-task), post-stroke participants prioritized dynamic stability over cognitive performance to ensure a safe locomotion. Thus, rehabilitation programs should consider both dynamic balance and dual-task training, even at a chronic delay following stroke, to reduce the risk of fall in post-stroke individuals. Copyright © 2018 Elsevier B.V. All rights reserved.

The validity and reliability of a dynamic neuromuscular stabilization-heel sliding test for core stability.

Science.gov (United States)

Cha, Young Joo; Lee, Jae Jin; Kim, Do Hyun; You, Joshua Sung H

2017-10-23

Core stabilization plays an important role in the regulation of postural stability. To overcome shortcomings associated with pain and severe core instability during conventional core stabilization tests, we recently developed the dynamic neuromuscular stabilization-based heel sliding (DNS-HS) test. The purpose of this study was to establish the criterion validity and test-retest reliability of the novel DNS-HS test. Twenty young adults with core instability completed both the bilateral straight leg lowering test (BSLLT) and DNS-HS test for the criterion validity study and repeated the DNS-HS test for the test-retest reliability study. Criterion validity was determined by comparing hip joint angle data that were obtained from BSLLT and DNS-HS measures. The test-retest reliability was determined by comparing hip joint angle data. Criterion validity was (ICC2,3) = 0.700 (preliability was (ICC3,3) = 0.953 (pvalidity data demonstrated a good relationship between the gold standard BSLLT and DNS-HS core stability measures. Test-retest reliability data suggests that DNS-HS core stability was a reliable test for core stability. Clinically, the DNS-HS test is useful to objectively quantify core instability and allow early detection and evaluation.

Chromatin dynamics in genome stability

DEFF Research Database (Denmark)

Nair, Nidhi; Shoaib, Muhammad; Sørensen, Claus Storgaard

2017-01-01

Genomic DNA is compacted into chromatin through packaging with histone and non-histone proteins. Importantly, DNA accessibility is dynamically regulated to ensure genome stability. This is exemplified in the response to DNA damage where chromatin relaxation near genomic lesions serves to promote...... access of relevant enzymes to specific DNA regions for signaling and repair. Furthermore, recent data highlight genome maintenance roles of chromatin through the regulation of endogenous DNA-templated processes including transcription and replication. Here, we review research that shows the importance...... of chromatin structure regulation in maintaining genome integrity by multiple mechanisms including facilitating DNA repair and directly suppressing endogenous DNA damage....

Guidelines for Computing Longitudinal Dynamic Stability Characteristics of a Subsonic Transport

Science.gov (United States)

Thompson, Joseph R.; Frank, Neal T.; Murphy, Patrick C.

2010-01-01

A systematic study is presented to guide the selection of a numerical solution strategy for URANS computation of a subsonic transport configuration undergoing simulated forced oscillation about its pitch axis. Forced oscillation is central to the prevalent wind tunnel methodology for quantifying aircraft dynamic stability derivatives from force and moment coefficients, which is the ultimate goal for the computational simulations. Extensive computations are performed that lead in key insights of the critical numerical parameters affecting solution convergence. A preliminary linear harmonic analysis is included to demonstrate the potential of extracting dynamic stability derivatives from computational solutions.

Comparison of dynamic postural stability scores between athletes with and without chronic ankle instability during lateral jump landing.

Science.gov (United States)

Shiravi, Zeinab; Shadmehr, Azadeh; Moghadam, Saeed Talebian; Moghadam, Behrouz Attarbashi

2017-01-01

Many ankle injuries occur while participating in sports that require jumping and landing such as basketball, volleyball and soccer. Most recent studies have investigated dynamic postural stability of patients with chronic ankle instability after landing from a forward jump. The present study aimed to investigate the dynamic postural stability of the athletes who suffer from chronic ankle sprain while landing from a lateral jump. Twelve athletes with self-reported unilateral chronic ankle instability (4 females and 8 males) and 12 matched controls (3 females and 9 males) voluntarily participated in the study. Dynamic postural stability index and its directional indices were measured while performing lateral jump landing test. No differences were found between athletes with and without chronic ankle instability during our landing protocol by means of the dynamic postural stability index and its directional indices. Findings showed that in each group, medial/lateral stability index is significantly higher than anterior/posterior and vertical stability indexes. Findings showed that dynamic postural stability was not significantly different between the two groups. Future studies should examine chronic ankle instability patients with more severe disabilities and expose them to more challenging dynamic balance conditions to further explore postural stability. IIIa.

JMJD5 (Jumonji Domain-containing 5) Associates with Spindle Microtubules and Is Required for Proper Mitosis.

Science.gov (United States)

He, Zhimin; Wu, Junyu; Su, Xiaonan; Zhang, Ye; Pan, Lixia; Wei, Huimin; Fang, Qiang; Li, Haitao; Wang, Da-Liang; Sun, Fang-Lin

2016-02-26

Precise mitotic spindle assembly is a guarantee of proper chromosome segregation during mitosis. Chromosome instability caused by disturbed mitosis is one of the major features of various types of cancer. JMJD5 has been reported to be involved in epigenetic regulation of gene expression in the nucleus, but little is known about its function in mitotic process. Here we report the unexpected localization and function of JMJD5 in mitotic progression. JMJD5 partially accumulates on mitotic spindles during mitosis, and depletion of JMJD5 results in significant mitotic arrest, spindle assembly defects, and sustained activation of the spindle assembly checkpoint (SAC). Inactivating SAC can efficiently reverse the mitotic arrest caused by JMJD5 depletion. Moreover, JMJD5 is found to interact with tubulin proteins and associate with microtubules during mitosis. JMJD5-depleted cells show a significant reduction of α-tubulin acetylation level on mitotic spindles and fail to generate enough interkinetochore tension to satisfy the SAC. Further, JMJD5 depletion also increases the susceptibility of HeLa cells to the antimicrotubule agent. Taken together, these results suggest that JMJD5 plays an important role in regulating mitotic progression, probably by modulating the stability of spindle microtubules. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Coupling of kinesin ATP turnover to translocation and microtubule regulation: one engine, many machines.

Science.gov (United States)

Friel, Claire T; Howard, Jonathon

2012-12-01

The cycle of ATP turnover is integral to the action of motor proteins. Here we discuss how variation in this cycle leads to variation of function observed amongst members of the kinesin superfamily of microtubule associated motor proteins. Variation in the ATP turnover cycle among superfamily members can tune the characteristic kinesin motor to one of the range of microtubule-based functions performed by kinesins. The speed at which ATP is hydrolysed affects the speed of translocation. The ratio of rate constants of ATP turnover in relation to association and dissociation from the microtubule influence the processivity of translocation. Variation in the rate-limiting step of the cycle can reverse the way in which the motor domain interacts with the microtubule producing non-motile kinesins. Because the ATP turnover cycle is not fully understood for the majority of kinesins, much work remains to show how the kinesin engine functions in such a wide variety of molecular machines.

Stability theory for dynamic equations on time scales

CERN Document Server

Martynyuk, Anatoly A

2016-01-01

This monograph is a first in the world to present three approaches for stability analysis of solutions of dynamic equations. The first approach is based on the application of dynamic integral inequalities and the fundamental matrix of solutions of linear approximation of dynamic equations. The second is based on the generalization of the direct Lyapunovs method for equations on time scales, using scalar, vector and matrix-valued auxiliary functions. The third approach is the application of auxiliary functions (scalar, vector, or matrix-valued ones) in combination with differential dynamic inequalities. This is an alternative comparison method, developed for time continuous and time discrete systems. In recent decades, automatic control theory in the study of air- and spacecraft dynamics and in other areas of modern applied mathematics has encountered problems in the analysis of the behavior of solutions of time continuous-discrete linear and/or nonlinear equations of perturbed motion. In the book “Men of Ma...

3-Amino-thieno[2,3-b]pyridines as microtubule-destabilising agents: Molecular modelling and biological evaluation in the sea urchin embryo and human cancer cells.

Science.gov (United States)

Eurtivong, Chatchakorn; Semenov, Victor; Semenova, Marina; Konyushkin, Leonid; Atamanenko, Olga; Reynisson, Jóhannes; Kiselyov, Alex

2017-01-15

A series of 3-amino-thieno[2,3-b]pyridines was prepared and tested in a phenotypic sea urchin embryo assay to identify potent and specific molecules that affect tubulin dynamics. The most active compounds featured a tricyclic core ring system with a fused cycloheptyl or cyclohexyl substituent and unsubstituted or alkyl-substituted phenyl moiety tethered via a carboxamide. Low nano-molar potency was observed in the sea urchin embryos for the most active compounds (1-5) suggestive of a microtubule-destabilising effect. The molecular modelling studies indicated that the tubulin colchicine site is inhibited, which often leads to microtubule-destabilisation in line with the sea urchin embryo results. Finally, the identified hits displayed a robust growth inhibition (GI 50 of 50-250nM) of multidrug-resistant melanoma MDA-MB-435 and breast MDA-MB-468 human cancer cell lines. This work demonstrates that for the thieno[2,3-b]pyridines the most effective mechanism of action is microtubule-destabilisation initiated by binding to the colchicine pocket. Copyright © 2016 Elsevier Ltd. All rights reserved.

Dynamical stability of slip-stacking particles

Energy Technology Data Exchange (ETDEWEB)

Eldred, Jeffrey; Zwaska, Robert

2014-09-01

We study the stability of particles in slip-stacking configuration, used to nearly double proton beam intensity at Fermilab. We introduce universal area factors to calculate the available phase space area for any set of beam parameters without individual simulation. We find perturbative solutions for stable particle trajectories. We establish Booster beam quality requirements to achieve 97% slip-stacking efficiency. We show that slip-stacking dynamics directly correspond to the driven pendulum and to the system of two standing-wave traps moving with respect to each other.

GIT1 enhances neurite outgrowth by stimulating microtubule assembly

Directory of Open Access Journals (Sweden)

Yi-sheng Li

2016-01-01

Full Text Available GIT1, a G-protein-coupled receptor kinase interacting protein, has been reported to be involved in neurite outgrowth. However, the neurobiological functions of the protein remain unclear. In this study, we found that GIT1 was highly expressed in the nervous system, and its expression was maintained throughout all stages of neuritogenesis in the brain. In primary cultured mouse hippocampal neurons from GIT1 knockout mice, there was a significant reduction in total neurite length per neuron, as well as in the average length of axon-like structures, which could not be prevented by nerve growth factor treatment. Overexpression of GIT1 significantly promoted axon growth and fully rescued the axon outgrowth defect in the primary hippocampal neuron cultures from GIT1 knockout mice. The GIT1 N terminal region, including the ADP ribosylation factor-GTPase activating protein domain, the ankyrin domains and the Spa2 homology domain, were sufficient to enhance axonal extension. Importantly, GIT1 bound to many tubulin proteins and microtubule-associated proteins, and it accelerated microtubule assembly in vitro. Collectively, our findings suggest that GIT1 promotes neurite outgrowth, at least partially by stimulating microtubule assembly. This study provides new insight into the cellular and molecular pathogenesis of GIT1-associated neurological diseases.

BWR stability using a reducing dynamical model

International Nuclear Information System (INIS)

Ballestrin Bolea, J. M.; Blazquez Martinez, J. B.

1990-01-01

BWR stability can be treated with reduced order dynamical models. When the parameters of the model came from dynamical models. When the parameters of the model came from experimental data, the predictions are accurate. In this work an alternative derivation for the void fraction equation is made, but remarking the physical structure of the parameters. As the poles of power/reactivity transfer function are related with the parameters, the measurement of the poles by other techniques such as noise analysis will lead to the parameters, but the system of equations is non-linear. Simple parametric calculation of decay ratio are performed, showing why BWRs become unstable when they are operated at low flow and high power. (Author)

Dynamic Characteristics Analysis and Stabilization of PV-Based Multiple Microgrid Clusters

DEFF Research Database (Denmark)

Zhao, Zhuoli; Yang, Ping; Wang, Yuewu

2018-01-01

-based multiple microgrid clusters. A detailed small-signal model for PV-based microgrid clusters considering local adaptive dynamic droop control mechanism of the voltage-source PV system is developed. The complete dynamic model is then used to access and compare the dynamic characteristics of the single...... microgrid and interconnected microgrids. In order to enhance system stability of the PV microgrid clusters, a tie-line flow and stabilization strategy is proposed to suppress the introduced interarea and local oscillations. Robustly selecting of the key control parameters is transformed to a multiobjective......As the penetration of PV generation increases, there is a growing operational demand on PV systems to participate in microgrid frequency regulation. It is expected that future distribution systems will consist of multiple microgrid clusters. However, interconnecting PV microgrids may lead to system...

Tracking of plus-ends reveals microtubule functional diversity in different cell types

Science.gov (United States)

Shaebani, M. Reza; Pasula, Aravind; Ott, Albrecht; Santen, Ludger

2016-07-01

Many cellular processes are tightly connected to the dynamics of microtubules (MTs). While in neuronal axons MTs mainly regulate intracellular trafficking, they participate in cytoskeleton reorganization in many other eukaryotic cells, enabling the cell to efficiently adapt to changes in the environment. We show that the functional differences of MTs in different cell types and regions is reflected in the dynamic properties of MT tips. Using plus-end tracking proteins EB1 to monitor growing MT plus-ends, we show that MT dynamics and life cycle in axons of human neurons significantly differ from that of fibroblast cells. The density of plus-ends, as well as the rescue and catastrophe frequencies increase while the growth rate decreases toward the fibroblast cell margin. This results in a rather stable filamentous network structure and maintains the connection between nucleus and membrane. In contrast, plus-ends are uniformly distributed along the axons and exhibit diverse polymerization run times and spatially homogeneous rescue and catastrophe frequencies, leading to MT segments of various lengths. The probability distributions of the excursion length of polymerization and the MT length both follow nearly exponential tails, in agreement with the analytical predictions of a two-state model of MT dynamics.

The Effect of the Loading on Dynamic Stability and Scapular Asymmetry

Directory of Open Access Journals (Sweden)

Mohammad Hassan Azarsa

2014-03-01

Full Text Available Background: Scapular stabilization and neuromuscular control provide an important parameter to characterize shoulder function during dynamic activities. Many studies have confirmed the effect of the loading on scapular position and scapulohumeral rhythm. Therefore, the evaluation of stabilizer muscles involvement in scapular asymmetry may assist in the development of clinical examination and rehabilitation program. The aim of this study was to evaluate the effect of loading on dynamic stability and scapular asymmetry in basketball players. Methods: Thirty healthy male basketball players aged between 20 to 31 years old were tested. The linear distance between scapular inferior angle and T7 spinous process was measured using a caliper in 90 degrees of unloaded scaption and with 1, 2 and 4 kg loading. The difference of distances of two sides in the above 4 positions was analyzed. Results: The amount of distances difference in two sides with 1 kg loading was minimal (9.36 mm. This difference increased to 10.19 mm and 12.22 mm, with increasing the loading to 2 and 4 kg respectively; although the 4 positions of the test did not show significant differences in distances difference (P>0.05. Conclusion: This study shows that dynamic stability of the scapula is dependent on the role of muscles, so that with increasing load on the muscles, the scapular asymmetry is more pronounced.

Use efficiency of dynamic stabilizer in the post-repair period of railways in Ukraine

Directory of Open Access Journals (Sweden)

O.V. Gubar

2013-08-01

Full Text Available Purpose. To analyze the interaction forces of the track and rolling stock at application of dynamic stabilizers, as well as the determination of the area, on which the track stabilization is necessary not only according to the technical indices but also according to the economic ones. Methodology.To achieve the research purpose the methods of analysis of material flows, peculiar to the places that need applications of dynamic stabilizers on railway transport tracks are used. Findings. Researches of relatively economic efficiency of dynamic stabilizers application, which were conducted by scientists during the last years, do not spread to comparison of cost of works on the track stabilizing and losses of railway for period of running. During the running period the trains move with a limited speed that causes corresponding financial losses. Speed limitation result in the both the motion time increase and in some occasions in the heavy consumption of fuel and energy resources. The more intensive track discord and expenses increase for its maintenance are observed in the sections of braking and acceleration immediately in front of and after the areas of limitation. The methodology relative to the estimation of economic efficiency of dynamic stabilizers application after completion of track repairs for the areas of Ukrainian railways with different operational conditions was developed. This methodology includes the losses calculation of railroad, which are predefined by extra motion time, heavy consumption of fuel and energy resources and charges for current maintenance. Originality. The methodology of cost effectiveness evaluating of the dynamic stabilizers application in the post-repair period was developed. Such an approach would allow one to take the rational decisions taking into account the features of the track sections, which were reconstructed. Practical value. The obtained results will rationally assign the works on the dynamic track

Three-dimensional fine structure of the organization of microtubules in neurite varicosities by ultra-high voltage electron microscope tomography.

Science.gov (United States)

Nishida, Tomoki; Yoshimura, Ryoichi; Endo, Yasuhisa

2017-09-01

Neurite varicosities are highly specialized compartments that are involved in neurotransmitter/ neuromodulator release and provide a physiological platform for neural functions. However, it remains unclear how microtubule organization contributes to the form of varicosity. Here, we examine the three-dimensional structure of microtubules in varicosities of a differentiated PC12 neural cell line using ultra-high voltage electron microscope tomography. Three-dimensional imaging showed that a part of the varicosities contained an accumulation of organelles that were separated from parallel microtubule arrays. Further detailed analysis using serial sections and whole-mount tomography revealed microtubules running in a spindle shape of swelling in some other types of varicosities. These electron tomographic results showed that the structural diversity and heterogeneity of microtubule organization supported the form of varicosities, suggesting that a different distribution pattern of microtubules in varicosities is crucial to the regulation of varicosities development.

Radio galaxies radiation transfer, dynamics, stability and evolution of a synchrotron plasmon

CERN Document Server

Pacholczyk, A G

1977-01-01

Radio Galaxies: Radiation Transfer, Dynamics, Stability and Evolution of a Synchrotron Plasmon deals with the physics of a region in space containing magnetic field and thermal and relativistic particles (a plasmon). The synchrotron emission and absorption of this region are discussed, along with the properties of its spectrum; its linear and circular polarization; transfer of radiation through such a region; its dynamics and expansion; and interaction with external medium.Comprised of eight chapters, this volume explores the stability, turbulence, and acceleration of particles in a synchrotro

Beam stability & nonlinear dynamics. Formal report

Energy Technology Data Exchange (ETDEWEB)

Parsa, Z. [ed.

1996-12-31

his Report includes copies of transparencies and notes from the presentations made at the Symposium on Beam Stability and Nonlinear Dynamics, December 3-5, 1996 at the Institute for Theoretical Physics, University of California, Santa Barbara California, that was made available by the authors. Editing, reduction and changes to the authors contributions were made only to fulfill the printing and publication requirements. We would like to take this opportunity and thank the speakers for their informative presentations and for providing copies of their transparencies and notes for inclusion in this Report.

Algorithm for Stabilizing a POD-Based Dynamical System

Science.gov (United States)

Kalb, Virginia L.

2010-01-01

This algorithm provides a new way to improve the accuracy and asymptotic behavior of a low-dimensional system based on the proper orthogonal decomposition (POD). Given a data set representing the evolution of a system of partial differential equations (PDEs), such as the Navier-Stokes equations for incompressible flow, one may obtain a low-dimensional model in the form of ordinary differential equations (ODEs) that should model the dynamics of the flow. Temporal sampling of the direct numerical simulation of the PDEs produces a spatial time series. The POD extracts the temporal and spatial eigenfunctions of this data set. Truncated to retain only the most energetic modes followed by Galerkin projection of these modes onto the PDEs obtains a dynamical system of ordinary differential equations for the time-dependent behavior of the flow. In practice, the steps leading to this system of ODEs entail numerically computing first-order derivatives of the mean data field and the eigenfunctions, and the computation of many inner products. This is far from a perfect process, and often results in the lack of long-term stability of the system and incorrect asymptotic behavior of the model. This algorithm describes a new stabilization method that utilizes the temporal eigenfunctions to derive correction terms for the coefficients of the dynamical system to significantly reduce these errors.

Dynamic stability of an aerodynamically efficient motorcycle

Science.gov (United States)

Sharma, Amrit; Limebeer, David J. N.

2012-08-01

Motorcycles exhibit two potentially dangerous oscillatory modes known as 'wobble' and 'weave'. The former is reminiscent of supermarket castor shimmy, while the latter is a low frequency 'fish-tailing' motion that involves a combination of rolling, yawing, steering and side-slipping motions. These unwanted dynamic features, which can occur when two-wheeled vehicles are operated at speed, have been studied extensively. The aim of this paper is to use mathematical analysis to identify important stability trends in the on-going design of a novel aerodynamically efficient motorcycle known as the ECOSSE Spirit ES1. A mathematical model of the ES1 is developed using a multi-body dynamics software package called VehicleSim [Anon, VehicleSim Lisp Reference Manual Version 1.0, Mechanical Simulation Corporation, 2008. Available at http://www.carsim.com]. This high-fidelity motorcycle model includes realistic tyre-road contact geometry, a comprehensive tyre model, tyre relaxation and a flexible frame. A parameter set representative of a modern high-performance machine and rider is used. Local stability is investigated via the eigenvalues of the linearised models that are associated with equilibrium points of interest. A comprehensive study of the effects of frame flexibilities, acceleration, aerodynamics and tyre variations is presented, and an optimal passive steering compensator is derived. It is shown that the traditional steering damper cannot be used to stabilise the ES1 over its entire operating speed range. A simple passive compensator, involving an inerter is proposed. Flexibility can be introduced deliberately into various chassis components to change the stability characteristics of the vehicle; the implications of this idea are studied.

Sync in Complex Dynamical Networks: Stability, Evolution, Control, and Application

OpenAIRE

Li, Xiang

2005-01-01

In the past few years, the discoveries of small-world and scale-free properties of many natural and artificial complex networks have stimulated significant advances in better understanding the relationship between the topology and the collective dynamics of complex networks. This paper reports recent progresses in the literature of synchronization of complex dynamical networks including stability criteria, network synchronizability and uniform synchronous criticality in different topologies, ...

Certain Actions from the Functional Movement Screen Do Not Provide an Indication of Dynamic Stability

Science.gov (United States)

Lockie, Robert G.; Callaghan, Samuel J.; Jordan, Corrin A.; Luczo, Tawni M.; Jeffriess, Matthew D.; Jalilvand, Farzad; Schultz, Adrian B.

2015-01-01

Dynamic stability is an essential physical component for team sport athletes. Certain Functional Movement Screen (FMS) exercises (deep squat; left- and right-leg hurdle step; left- and right-leg in-line lunge [ILL]; left- and right-leg active straight-leg raise; and trunk stability push-up [TSPU]) have been suggested as providing an indication of dynamic stability. No research has investigated relationships between these screens and an established test of dynamic stability such as the modified Star Excursion Balance Test (mSEBT), which measures lower-limb reach distance in posteromedial, medial, and anteromedial directions, in team sport athletes. Forty-one male and female team sport athletes completed the screens and the mSEBT. Participants were split into high-, intermediate-, and low-performing groups according to the mean of the excursions when both the left and right legs were used for the mSEBT stance. Any between-group differences in the screens and mSEBT were determined via a one-way analysis of variance with Bonferroni post hoc adjustment (p in any of the screens, and only two positive correlations between the screens and the mSEBT (TSPU and right stance leg posteromedial excursion, r = 0.37; left-leg ILL and left stance leg posteromedial excursion, r = 0.46). The mSEBT clearly indicated participants with different dynamic stability capabilities. In contrast to the mSEBT, the selected FMS exercises investigated in this study have a limited capacity to identify dynamic stability in team sport athletes. PMID:26557187

Dynamic Stabilization of Metal Oxide–Water Interfaces

Energy Technology Data Exchange (ETDEWEB)

McBriarty, Martin E.; von Rudorff, Guido Falk; Stubbs, Joanne; Eng, Peter; Blumberger, Jochen; Rosso, Kevin M.

2017-02-08

Metal oxide growth, dissolution, and redox reactivity depend on the structure and dynamics at the interface with aqueous solution. We present the most definitive analysis to date of the hydrated naturally abundant r-cut (11$\\bar{0}$2) termination of the iron oxide hematite (α-Fe₂O₃). In situ synchrotron X-ray scattering analysis reveals a ridged lateral arrangement of adsorbed water molecules hydrogen bonded to terminal aquo groups. Large-scale hybrid-functional density functional theory-based molecular dynamics (DFT-MD) simulations show how this structure is dynamically stabilized by picosecond exchange between aquo groups and adsorbed water, even under nominally dry conditions. Surface pKa prediction based on bond valence analysis suggests that water exchange may influence the proton transfer reactions associated with acid/base reactivity at the interface. Our findings rectify inconsistencies between existing models and may be extended to resolving more complex electrochemical phenomena at metal oxide-water interfaces.

Reassessing the Roles of PIN Proteins and Anticlinal Microtubules during Pavement Cell Morphogenesis1[OPEN

Science.gov (United States)

Sawchuk, Megan G.; Scarpella, Enrico

2018-01-01

The leaf epidermis is a biomechanical shell that influences the size and shape of the organ. Its morphogenesis is a multiscale process in which nanometer-scale cytoskeletal protein complexes, individual cells, and groups of cells pattern growth and define macroscopic leaf traits. Interdigitated growth of neighboring cells is an evolutionarily conserved developmental strategy. Understanding how signaling pathways and cytoskeletal proteins pattern cell walls during this form of tissue morphogenesis is an important research challenge. The cellular and molecular control of a lobed cell morphology is currently thought to involve PIN-FORMED (PIN)-type plasma membrane efflux carriers that generate subcellular auxin gradients. Auxin gradients were proposed to function across cell boundaries to encode stable offset patterns of cortical microtubules and actin filaments between adjacent cells. Many models suggest that long-lived microtubules along the anticlinal cell wall generate local cell wall heterogeneities that restrict local growth and specify the timing and location of lobe formation. Here, we used Arabidopsis (Arabidopsis thaliana) reverse genetics and multivariate long-term time-lapse imaging to test current cell shape control models. We found that neither PIN proteins nor long-lived microtubules along the anticlinal wall predict the patterns of lobe formation. In fields of lobing cells, anticlinal microtubules are not correlated with cell shape and are unstable at the time scales of cell expansion. Our analyses indicate that anticlinal microtubules have multiple functions in pavement cells and that lobe initiation is likely controlled by complex interactions among cell geometry, cell wall stress patterns, and transient microtubule networks that span the anticlinal and periclinal walls. PMID:29192026

Topics in Modeling of Cochlear Dynamics: Computation, Response and Stability Analysis

Science.gov (United States)

Filo, Maurice G.

This thesis touches upon several topics in cochlear modeling. Throughout the literature, mathematical models of the cochlea vary according to the degree of biological realism to be incorporated. This thesis casts the cochlear model as a continuous space-time dynamical system using operator language. This framework encompasses a wider class of cochlear models and makes the dynamics more transparent and easier to analyze before applying any numerical method to discretize space. In fact, several numerical methods are investigated to study the computational efficiency of the finite dimensional realizations in space. Furthermore, we study the effects of the active gain perturbations on the stability of the linearized dynamics. The stability analysis is used to explain possible mechanisms underlying spontaneous otoacoustic emissions and tinnitus. Dynamic Mode Decomposition (DMD) is introduced as a useful tool to analyze the response of nonlinear cochlear models. Cochlear response features are illustrated using DMD which has the advantage of explicitly revealing the spatial modes of vibrations occurring in the Basilar Membrane (BM). Finally, we address the dynamic estimation problem of BM vibrations using Extended Kalman Filters (EKF). Due to the limitations of noninvasive sensing schemes, such algorithms are inevitable to estimate the dynamic behavior of a living cochlea.

Cationic membranes complexed with oppositely charged microtubules: hierarchical self-assembly leading to bio-nanotubes

International Nuclear Information System (INIS)

Raviv, Uri; Needleman, Daniel J; Safinya, Cyrus R

2006-01-01

The self-assembly of microtubules and charged membranes has been studied, using x-ray diffraction and electron microscopy. Polyelectrolyte lipid complexes usually form structures templated by the lipid phase, when the polyelectrolyte curvature is much larger than the membrane spontaneous curvature. When the polyelectrolyte curvature approaches the membrane spontaneous curvature, as in microtubules, two types of new structures emerge. Depending on the conditions, vesicles either adsorb onto the microtubule, forming a 'beads on a rod' structure, or coat the microtubule, which now forms the template. Tubulin oligomers then coat the external lipid layer, forming a lipid protein nanotube. The tubulin oligomer coverage at the external layer is determined by the membrane charge density. The energy barrier between the beads on a rod and the lipid-protein nanotube states depends on the membrane bending rigidity and membrane charge density. By controlling the lipid/tubulin stoichiometry we can switch between lipid-protein nanotubes with open ends to lipid-protein nanotubes with closed end with lipid cups. This forms the basis for controlled drug encapsulation and release

Katanin spiral and ring structures shed light on power stroke for microtubule severing

Energy Technology Data Exchange (ETDEWEB)

Zehr, Elena; Szyk, Agnieszka; Piszczek, Grzegorz; Szczesna, Ewa; Zuo, Xiaobing; Roll-Mecak, Antonina

2017-08-07

Microtubule-severing enzymes katanin, spastin and fidgetin are AAA ATPases critical for the biogenesis and maintenance of complex microtubule arrays in axons, spindles and cilia. Because of a lack of 3D structures, their mechanism has remained poorly understood. We report the first X-ray structure of the monomeric AAA katanin module and cryo-EM reconstructions of the hexamer in two conformations. These reveal an unexpected asymmetric arrangement of the AAA domains mediated by structural elements unique to severing enzymes and critical for their function. Our reconstructions show that katanin cycles between open spiral and closed ring conformations, depending on the ATP occupancy of a gating protomer that tenses or relaxes inter-protomer interfaces. Cycling of the hexamer between these conformations would provide the power stroke for microtubule severing.

On Stabilizing the Variance of Dynamic Functional Brain Connectivity Time Series.

Science.gov (United States)

Thompson, William Hedley; Fransson, Peter

2016-12-01

Assessment of dynamic functional brain connectivity based on functional magnetic resonance imaging (fMRI) data is an increasingly popular strategy to investigate temporal dynamics of the brain's large-scale network architecture. Current practice when deriving connectivity estimates over time is to use the Fisher transformation, which aims to stabilize the variance of correlation values that fluctuate around varying true correlation values. It is, however, unclear how well the stabilization of signal variance performed by the Fisher transformation works for each connectivity time series, when the true correlation is assumed to be fluctuating. This is of importance because many subsequent analyses either assume or perform better when the time series have stable variance or adheres to an approximate Gaussian distribution. In this article, using simulations and analysis of resting-state fMRI data, we analyze the effect of applying different variance stabilization strategies on connectivity time series. We focus our investigation on the Fisher transformation, the Box-Cox (BC) transformation and an approach that combines both transformations. Our results show that, if the intention of stabilizing the variance is to use metrics on the time series, where stable variance or a Gaussian distribution is desired (e.g., clustering), the Fisher transformation is not optimal and may even skew connectivity time series away from being Gaussian. Furthermore, we show that the suboptimal performance of the Fisher transformation can be substantially improved by including an additional BC transformation after the dynamic functional connectivity time series has been Fisher transformed.

Antagonism between the dynein and Ndc80 complexes at kinetochores controls the stability of kinetochore-microtubule attachments during mitosis.

Science.gov (United States)

Amin, Mohammed A; McKenney, Richard J; Varma, Dileep

2018-04-20

Chromosome alignment and segregation during mitosis require kinetochore-microtubule (kMT) attachments that are mediated by the molecular motor dynein and the kMT-binding complex Ndc80. The Rod-ZW10-Zwilch (RZZ) complex is central to this coordination as it has an important role in dynein recruitment and has recently been reported to have a key function in the regulation of stable kMT attachments in Caenorhabditis elegans besides its role in activating the spindle assembly checkpoint (SAC). However, the mechanism by which these protein complexes control kMT attachments to drive chromosome motility during early mitosis is still unclear. Here, using in vitro total internal reflection fluorescence microscopy, we observed that higher concentrations of Ndc80 inhibited dynein binding to MTs, providing evidence that Ndc80 and dynein antagonize each other's function. High-resolution microscopy and siRNA-mediated functional disruption revealed that severe defects in chromosome alignment induced by depletion of dynein or the dynein adapter Spindly are rescued by codepletion of the RZZ component Rod in human cells. Interestingly, rescue of the chromosome alignment defects was independent of Rod function in SAC activation and was accompanied by a remarkable restoration of stable kMT attachments. Furthermore, the chromosome alignment rescue depended on the plus-end-directed motility of centromere protein E (CENP-E) because cells codepleted of CENP-E, Rod, and dynein could not establish stable kMT attachments or align their chromosomes properly. Our findings support the idea that dynein may control the function of the Ndc80 complex in stabilizing kMT attachments directly by interfering with Ndc80-MT binding or indirectly by controlling the Rod-mediated inhibition of Ndc80. © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

Iron oxide nanoparticles induce human microvascular endothelial cell permeability through reactive oxygen species production and microtubule remodeling

Directory of Open Access Journals (Sweden)

Shi Xianglin

2009-01-01

Full Text Available Abstract Background Engineered iron nanoparticles are being explored for the development of biomedical applications and many other industry purposes. However, to date little is known concerning the precise mechanisms of translocation of iron nanoparticles into targeted tissues and organs from blood circulation, as well as the underlying implications of potential harmful health effects in human. Results The confocal microscopy imaging analysis demonstrates that exposure to engineered iron nanoparticles induces an increase in cell permeability in human microvascular endothelial cells. Our studies further reveal iron nanoparticles enhance the permeability through the production of reactive oxygen species (ROS and the stabilization of microtubules. We also showed Akt/GSK-3β signaling pathways are involved in iron nanoparticle-induced cell permeability. The inhibition of ROS demonstrate ROS play a major role in regulating Akt/GSK-3β – mediated cell permeability upon iron nanoparticle exposure. These results provide new insights into the bioreactivity of engineered iron nanoparticles which can inform potential applications in medical imaging or drug delivery. Conclusion Our results indicate that exposure to iron nanoparticles induces an increase in endothelial cell permeability through ROS oxidative stress-modulated microtubule remodeling. The findings from this study provide new understandings on the effects of nanoparticles on vascular transport of macromolecules and drugs.

Research on Dynamics and Stability in the Stairs-Climbing of a Tracked Mobile Robot

Directory of Open Access Journals (Sweden)

Weijun Tao

2012-10-01

Full Text Available Aiming at the functional requirement of climbing up the stairs, the dynamics and stability during a tracked mobile robot's climbing of stairs is studied. First, from the analysis of its cross-country performance, the mechanical structure of the tracked mobile robot is designed and the hardware composition of its control system is given. Second, based on the analysis to its stairs-climbing process, the dynamical model of stairs-climbing is established by using the classical mechanics method. Next, the stability conditions for its stairs-climbing are determined and an evaluation method of its stairs-climbing stability is proposed, based on a mechanics analysis on the robot's backwards tumbling during the stairs-climbing process. Through simulation and experiments, the effectiveness of the dynamical model and the stability evaluation method of the tracked mobile robot in stairs-climbing is verified, which can provide design and analysis foundations for the tracked mobile robots' stairs-climbing.

Stability and chaotic dynamics of a perturbed rate gyro

International Nuclear Information System (INIS)

Chen, H.-H.

2006-01-01

An analysis of stability and chaotic dynamics is presented by a single-axis rate gyro subjected to linear feedback control loops. This rate gyro is supposed to be mounted on a space vehicle which undergoes an uncertain angular velocity ω Z (t) around its spin axis and simultaneously acceleration ω-bar X (t) occurs with respect to the output axis. The necessary and sufficient conditions of stability and degeneracy conditions for the autonomous case, whose vehicle undergoes a steady rotation, were provided by Routh-Hurwitz theory. The stability of the nonlinear nonautonomous system was investigated by Liapunov stability and instability theorems. As the electrical time constant is much smaller than the mechanical time constant, the singularly perturbed system can be obtained by the singular perturbation theory. The Liapunov stability of this system by studying the reduced and boundary-layer systems was also analyzed. Using the Melinikov technique, we can give criteria for the existence of chaos in the gyro motion when the vehicle undergoes perturbed harmonic rotation about its spin and output axes

Dynamics of actin cables in polarized growth of the filamentous fungus Aspergillus nidulans

Directory of Open Access Journals (Sweden)

Anna eBergs

2016-05-01

Full Text Available Highly polarized growth of filamentous fungi requires a continuous supply of proteins and lipids to the hyphal tip. This transport is managed by vesicle trafficking via the actin and microtubule cytoskeletons and their associated motor proteins. Particularly, actin cables originating from the hyphal tip are essential for hyphal growth. Although specific marker proteins to visualize actin cables have been developed in filamentous fungi, the exact organization and dynamics of actin cables has remained elusive. Here we visualized actin cables using tropomyosin (TpmA and Lifeact fused to fluorescent proteins in Aspergillus nidulans and studied the dynamics and regulation. GFP tagged TpmA visualized dynamic actin cables formed from the hyphal tip with cycles of elongation and shrinkage. The elongation and shrinkage rates of actin cables were similar and approximately 0.6 μm/s. Comparison of actin markers revealed that high concentrations of Lifeact reduced actin dynamics. Simultaneous visualization of actin cables and microtubules suggests temporally and spatially coordinated polymerization and depolymerization between the two cytoskeletons. Our results provide new insights into the molecular mechanism of ordered polarized growth regulated by actin cables and microtubules.

Constraints on dynamic stability during forward, backward and lateral locomotion in skilled football players.

Science.gov (United States)

Mehdizadeh, Sina; Arshi, Ahmed Reza; Davids, Keith

2016-01-01

The aim of this study was to investigate effects of speed and plane of motion on stability during locomotion in skilled football players. Ten male national-level football players participated in this study to run forward, backward and in lateral directions on a treadmill at 80%, 100% and 120% of their preferred running speeds. The coordinate data of passive reflective markers attached to body segments were recorded using motion capture systems. Time series data obtained from the ankle marker were used for further analyses. The largest finite-time Lyapunov exponent and maximum Floquet multiplier were adopted to quantify local and orbital dynamic stabilities, respectively. Results showed that speed did not significantly change local and orbital dynamic stabilities in any of running patterns. However, both local and orbital dynamic stability were significantly higher in the secondary plane of progression. Data revealed that in running, unlike walking, stability in the direction perpendicular to the direction of running is significantly higher, implying that less active control is required in the secondary plane of progression. The results of this study could be useful in sports training and rehabilitation programmes where development of fundamental exercise programmes that challenge both speed and the ability to maintain stability might produce a tangible enhancement of athletic skill level.

Effect of arm swing strategy on local dynamic stability of human gait.

Science.gov (United States)

Punt, Michiel; Bruijn, Sjoerd M; Wittink, Harriet; van Dieën, Jaap H

2015-02-01

Falling causes long term disability and can even lead to death. Most falls occur during gait. Therefore improving gait stability might be beneficial for people at risk of falling. Recently arm swing has been shown to influence gait stability. However at present it remains unknown which mode of arm swing creates the most stable gait. To examine how different modes of arm swing affect gait stability. Ten healthy young male subjects volunteered for this study. All subjects walked with four different arm swing instructions at seven different gait speeds. The Xsens motion capture suit was used to capture gait kinematics. Basic gait parameters, variability and stability measures were calculated. We found an increased stability in the medio-lateral direction with excessive arm swing in comparison to normal arm swing at all gait speeds. Moreover, excessive arm swing increased stability in the anterior-posterior and vertical direction at low gait speeds. Ipsilateral and inphase arm swing did not differ compared to a normal arm swing. Excessive arm swing is a promising gait manipulation to improve local dynamic stability. For excessive arm swing in the ML direction there appears to be converging evidence. The effect of excessive arm swing on more clinically relevant groups like the more fall prone elderly or stroke survivors is worth further investigating. Excessive arm swing significantly increases local dynamic stability of human gait. Copyright © 2014 Elsevier B.V. All rights reserved.

Effect of speed on local dynamic stability of locomotion under different task constraints in running.

Science.gov (United States)

Mehdizadeh, Sina; Arshi, Ahmed Reza; Davids, Keith

2014-01-01

A number of studies have investigated effects of speed on local dynamic stability of walking, although this relationship has been rarely investigated under changing task constraints, such as during forward and backward running. To rectify this gap in the literature, the aim of this study was to investigate the effect of running speed on local dynamic stability of forward and backward running on a treadmill. Fifteen healthy male participants took part in this study. Participants ran in forward and backward directions at speeds of 80%, 100% and 120% of their preferred running speed. The three-dimensional motion of a C7 marker was recorded using a motion capture system. Local dynamic stability of the marker was quantified using short- and long-term largest finite-time Lyapunov exponents (LyE). Results showed that short-term largest finite-time LyE values increased with participant speed meaning that local dynamic stability decreased with increasing speed. Long-term largest finite-time LyEs, however, remained unaffected as speed increased. Results of this study indicated that, as in walking, slow running is more stable than fast running. These findings improve understanding of how stability is regulated when constraints on the speed of movements is altered. Implications for the design of rehabilitation or sport practice programmes suggest how task constraints could be manipulated to facilitate adaptations in locomotion stability during athletic training.

Microtubule organization in three-dimensional confined geometries: Evaluating the role of elasticity through a combined in vitro and modeling approach

NARCIS (Netherlands)

Cosentino Lagomarsino, M.; Tanase, C.; Vos, J.W.; Emons, A.M.C.; Mulder, B.; Dogterom, M.

2007-01-01

Microtubules or microtubule bundles in cells often grow longer than the size of the cell, which causes their shape and organization to adapt to constraints imposed by the cell geometry. We test the reciprocal role of elasticity and confinement in the organization of growing microtubules in a

Transition from shod to barefoot alters dynamic stability during running.

Science.gov (United States)

Ekizos, Antonis; Santuz, Alessandro; Arampatzis, Adamantios

2017-07-01

Barefoot running recently received increased attention, with controversial results regarding its effects on injury risk and performance. Numerous studies examined the kinetic and kinematic changes between the shod and the barefoot condition. Intrinsic parameters such as the local dynamic stability could provide new insight regarding neuromuscular control when immediately transitioning from one running condition to the other. We investigated the local dynamic stability during the change from shod to barefoot running. We further measured biomechanical parameters to examine the mechanisms governing this transition. Twenty habitually shod, young and healthy participants ran on a pressure plate-equipped treadmill and alternated between shod and barefoot running. We calculated the largest Lyapunov exponents as a measure of errors in the control of the movement. Biomechanical parameters were also collected. Local dynamic stability decreased significantly (d=0.41; 2.1%) during barefoot running indicating worse control over the movement. We measured higher cadence (d=0.35; 2.2%) and total flight time (d=0.58; 19%), lower total contact time (d=0.58; -5%), total vertical displacement (d=0.39; -4%), and vertical impulse (d=1.32; 11%) over the two minutes when running barefoot. The strike index changed significantly (d=1.29; 237%) towards the front of the foot. Immediate transition from shod to the barefoot condition resulted in an increased instability and indicates a worst control over the movement. The increased instability was associated with biomechanical changes (i.e. foot strike patterns) of the participants in the barefoot condition. Possible reasons why this instability arises, might be traced in the stance phase and particularly in the push-off. The decreased stability might affect injury risk and performance. Copyright © 2017 Elsevier B.V. All rights reserved.

Interplay Between Energy-Market Dynamics and Physical Stability of a Smart Power Grid

Science.gov (United States)

Picozzi, Sergio; Mammoli, Andrea; Sorrentino, Francesco

2013-03-01

A smart power grid is being envisioned for the future which, among other features, should enable users to play the dual role of consumers as well as producers and traders of energy, thanks to emerging renewable energy production and energy storage technologies. As a complex dynamical system, any power grid is subject to physical instabilities. With existing grids, such instabilities tend to be caused by natural disasters, human errors, or weather-related peaks in demand. In this work we analyze the impact, upon the stability of a smart grid, of the energy-market dynamics arising from users' ability to buy from and sell energy to other users. The stability analysis of the resulting dynamical system is performed assuming different proposed models for this market of the future, and the corresponding stability regions in parameter space are identified. We test our theoretical findings by comparing them with data collected from some existing prototype systems.

Identification of a lysosome membrane protein which could mediate ATP-dependent stable association of lysosomes to microtubules

International Nuclear Information System (INIS)

Mithieux, G.; Rousset, B.

1989-01-01

We have previously reported that purified thyroid lysosomes bind to reconstituted microtubules to form stable complexes, a process which is inhibited by ATP. Among detergent-solubilized lysosomal membrane protein, we identified a 50-kDa molecular component which binds to preassembled microtubules. The binding of this polypeptide to microtubules was decreased in the presence of ATP. We purified this 50-kDa protein by affinity chromatography on immobilized ATP. The 50-kDa protein bound to the ATP column was eluted by 1 mM ATP. The purified protein, labeled with 125I, exhibited the ability of interacting with microtubules. The binding process was inhibited by increasing concentrations of ATP, the half-maximal inhibitory effect being obtained at an ATP concentration of 0.35 mM. The interaction of the 50-kDa protein with microtubules is a saturable phenomenon since the binding of the 125I-labeled 50-kDa protein was inhibited by unlabeled solubilized lysosomal membrane protein containing the 50-kDa polypeptide but not by the same protein fraction from which the 50-kDa polypeptide had been removed by the ATP affinity chromatography procedure. The 50-kDa protein has the property to bind to pure tubulin coupled to an insoluble matrix. The 50-kDa protein was eluted from the tubulin affinity column by ATP. These findings support the conclusion that a protein inserted into the lysosomal membrane is able to bind directly to microtubules in a process which can be regulated by ATP. We propose that this protein could account for the association of lysosomes to microtubules demonstrated both in vitro and in intact cells

The Comprehensive Biomechanics and Load-Sharing of Semirigid PEEK and Semirigid Posterior Dynamic Stabilization Systems

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D. K. Sengupta

2013-01-01

Full Text Available Alternatives to conventional rigid fusion have been proposed for several conditions related to degenerative disc disease when nonoperative treatment has failed. Semirigid fixation, in the form of dynamic stabilization or PEEK rods, is expected to provide compression under loading as well as an intermediate level of stabilization. This study systematically examines both the load-sharing characteristics and kinematics of these two devices compared to the standard of internal rigid fixators. Load-sharing was studied by using digital pressure films inserted between an artificially machined disc and two loading fixtures. Rigid rods, PEEK rods, and the dynamic stabilization system were inserted posteriorly for stabilization. The kinematics were quantified on ten, human, cadaver lumbosacral spines (L3-S1 which were tested under a pure bending moment, in flexion-extension, lateral bending, and axial rotation. The magnitude of load transmission through the anterior column was significantly greater with the dynamic device compared to PEEK rods and rigid rods. The contact pressures were distributed more uniformly, throughout the disc with the dynamic stabilization devices, and had smaller maximum point-loading (pressures on any particular point within the disc. Kinematically, the motion was reduced by both semirigid devices similarly in all directions, with slight rigidity imparted by a lateral interbody device.

Capu and Spire Assemble a Cytoplasmic Actin~Mesh that Maintains Microtubule Organization in the Drosophila Oocyte

DEFF Research Database (Denmark)

Dahlgaard, K.; Raposo, A.A.S.F.; Niccoli, T.

2007-01-01

Mutants in the actin nucleators Cappuccino and Spire disrupt the polarized microtubule network in the Drosophila oocyte that defines the anterior-posterior axis, suggesting that microtubule organization depends on actin. Here, we show that Cappuccino and Spire organize an isotropic mesh of actin...

The NFL-TBS.40-63 anti-glioblastoma peptide disrupts microtubule and mitochondrial networks in the T98G glioma cell line.

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Romain Rivalin

Full Text Available Despite aggressive therapies, including combinations of surgery, radiotherapy and chemotherapy, glioblastoma remains a highly aggressive brain cancer with the worst prognosis of any central nervous system disease. We have previously identified a neurofilament-derived cell-penetrating peptide, NFL-TBS.40-63, that specifically enters by endocytosis in glioblastoma cells, where it induces microtubule destruction and inhibits cell proliferation. Here, we explore the impact of NFL-TBS.40-63 peptide on the mitochondrial network and its functions by using global cell respiration, quantitative PCR analysis of the main actors directing mitochondrial biogenesis, western blot analysis of the oxidative phosphorylation (OXPHOS subunits and confocal microscopy. We show that the internalized peptide disturbs mitochondrial and microtubule networks, interferes with mitochondrial dynamics and induces a rapid depletion of global cell respiration. This effect may be related to reduced expression of the NRF-1 transcription factor and of specific miRNAs, which may impact mitochondrial biogenesis, in regard to default mitochondrial mobility.

Stability of power systems coupled with market dynamics

Science.gov (United States)

Meng, Jianping

This Ph.D. thesis presented here spans two relatively independent topics. The first part, Chapter 2 is self-contained, and is dedicated to studies of new algorithms for power system state estimation. The second part, encompassing the remaining chapters, is dedicated to stability analysis of power system coupled with market dynamics. The first part of this thesis presents improved Newton's methods employing efficient vectorized calculations of higher order derivatives in power system state estimation problems. The improved algorithms are proposed based on an exact Newton's method using the second order terms. By efficiently computing an exact gain matrix, combined with a special optimal multiplier method, the new algorithms show more reliable convergence compared with the existing methods of normal equations, orthogonal decomposition, and Hachtel's sparse tableau. Our methods are able to handle ill-conditioned problems, yet show minimal penalty in computational cost for well-conditioned cases. These claims are illustrated through the standard IEEE 118 and 300 bus test examples. The second part of the thesis focuses on stability analysis of market/power systems. The work presented is motivated by an emerging problem. As the frequency of market based dispatch updates increases, there will inevitably be interaction between the dynamics of markets determining the generator dispatch commands, and the physical response of generators and network interconnections, necessitating the development of stability analysis for such coupled systems. We begin with numeric tests using different market models, with detailed machine/exciter/turbine/governor dynamics, in the New England 39 bus test system. A progression of modeling refinements are introduced, including such non-ideal effects as time delays. Electricity market parameter identification algorithms are also studied based on real time data from the PJM electricity market. Finally our power market model is augmented by optimal

The effect of walking speed on local dynamic stability is sensitive to calculation methods

DEFF Research Database (Denmark)

Stenum, Jan; Bruijn, Sjoerd M; Jensen, Bente Rona

2014-01-01

Local dynamic stability has been assessed by the short-term local divergence exponent (λS), which quantifies the average rate of logarithmic divergence of infinitesimally close trajectories in state space. Both increased and decreased local dynamic stability at faster walking speeds have been...... reported. This might pertain to methodological differences in calculating λS. Therefore, the aim was to test if different calculation methods would induce different effects of walking speed on local dynamic stability. Ten young healthy participants walked on a treadmill at five speeds (60%, 80%, 100%, 120......% and 140% of preferred walking speed) for 3min each, while upper body accelerations in three directions were sampled. From these time-series, λS was calculated by three different methods using: (a) a fixed time interval and expressed as logarithmic divergence per stride-time (λS-a), (b) a fixed number...

Certain Actions from the Functional Movement Screen Do Not Provide an Indication of Dynamic Stability

Directory of Open Access Journals (Sweden)

Lockie Robert G.

2015-09-01

Full Text Available Dynamic stability is an essential physical component for team sport athletes. Certain Functional Movement Screen (FMS exercises (deep squat; left- and right-leg hurdle step; left- and right-leg in-line lunge [ILL]; left- and right-leg active straight-leg raise; and trunk stability push-up [TSPU] have been suggested as providing an indication of dynamic stability. No research has investigated relationships between these screens and an established test of dynamic stability such as the modified Star Excursion Balance Test (mSEBT, which measures lower-limb reach distance in posteromedial, medial, and anteromedial directions, in team sport athletes. Forty-one male and female team sport athletes completed the screens and the mSEBT. Participants were split into high-, intermediate-, and low-performing groups according to the mean of the excursions when both the left and right legs were used for the mSEBT stance. Any between-group differences in the screens and mSEBT were determined via a one-way analysis of variance with Bonferroni post hoc adjustment (p < 0.05. Data was pooled for a correlation analysis (p < 0.05. There were no between-group differences in any of the screens, and only two positive correlations between the screens and the mSEBT (TSPU and right stance leg posteromedial excursion, r = 0.37; left-leg ILL and left stance leg posteromedial excursion, r = 0.46. The mSEBT clearly indicated participants with different dynamic stability capabilities. In contrast to the mSEBT, the selected FMS exercises investigated in this study have a limited capacity to identify dynamic stability in team sport athletes.

and its allicin on microtubule and cancer cell lines

African Journals Online (AJOL)

STORAGESEVER

2009-10-05

Oct 5, 2009 ... microtubule protein polymer that treated by A. hirtifolium. (A), and allicin (B) in .... with a chromogenic thiol: reaction of 4-mercaptopyridine with ... transformed tumor growth in vivo by diallyl disulfide is associated with inhibition ...

Does usnic acid affect microtubules in human cancer cells? O ácido úsnico pode afetar microtúbulos em células cancerosas humanas?

Directory of Open Access Journals (Sweden)

MA. O'Neill

2010-08-01

Full Text Available Usnic acid, a lichen metabolite, is known to exert antimitotic and antiproliferative activities against normal and malignant human cells. Many chemotherapy agents exert their activities by blocking cell cycle progression, inducing cell death through apoptosis. Microtubules, protein structure involved in the segregation of chromosomes during mitosis, serve as chemotherapeutical targets due to their key role in cellular division as well as apoptosis. The aim of this work was to investigate whether usnic acid affects the formation and/or stabilisation of microtubules by visualising microtubules and determining mitotic indices after treatment. The breast cancer cell line MCF7 and the lung cancer cell line H1299 were treated with usnic acid 29 µM for 24 hours and two positive controls: vincristine (which prevents the formation of microtubules or taxol (which stabilizes microtubules. Treatment of MCF7 and H1299 cells with usnic acid did not result in any morphological changes in microtubules or increase in the mitotic index. These results suggest that the antineoplastic activity of usnic acid is not related to alterations in the formation and/or stabilisation of microtubules.O ácido úsnico, um metabólito de liquens, é conhecido por sua atividade antimitótica e antiproliferativa em células humanas normais e malignas. Muitos quimioterápicos exercem suas atividades bloqueando a progressão do ciclo celular e induzindo morte celular por apoptose. Os microtúbulos, estruturas protéicas envolvidas na segregação dos cromossomos durante a mitose, servem como alvo quimioterapêutico devido ao seu importante papel tanto na divisão celular quanto nos mecanismos de morte celular por apoptose. O objetivo deste trabalho foi investigar se o ácido úsnico afeta a formação e/ou estabilização dos microtúbulos, a partir da visualização de microtúbulos e determinação de índices mitóticos após o tratamento. Células de câncer de mama MCF7 e de c

Effects of an attention demanding task on dynamic stability during treadmill walking

Directory of Open Access Journals (Sweden)

Troy Karen L

2008-04-01

Full Text Available Abstract Background People exhibit increased difficulty balancing when they perform secondary attention-distracting tasks while walking. However, a previous study by Grabiner and Troy (J. Neuroengineering Rehabil., 2005 found that young healthy subjects performing a concurrent Stroop task while walking on a motorized treadmill exhibited decreased step width variability. However, measures of variability do not directly quantify how a system responds to perturbations. This study re-analyzed data from Grabiner and Troy 2005 to determine if performing the concurrent Stroop task directly affected the dynamic stability of walking in these same subjects. Methods Thirteen healthy volunteers walked on a motorized treadmill at their self-selected constant speed for 10 minutes both while performing the Stroop test and during undisturbed walking. This Stroop test consisted of projecting images of the name of one color, printed in text of a different color, onto a wall and asking subjects to verbally identify the color of the text. Three-dimensional motions of a marker attached to the base of the neck (C5/T1 were recorded. Marker velocities were calculated over 3 equal intervals of 200 sec each in each direction. Mean variability was calculated for each time series as the average standard deviation across all strides. Both "local" and "orbital" dynamic stability were quantified for each time series using previously established methods. These measures directly quantify how quickly small perturbations grow or decay, either continuously in real time (local or discretely from one cycle to the next (orbital. Differences between Stroop and Control trials were evaluated using a 2-factor repeated measures ANOVA. Results Mean variability of trunk movements was significantly reduced during the Stroop tests compared to normal walking. Conversely, local and orbital stability results were mixed: some measures showed slight increases, while others showed slight decreases

PWL approximation of nonlinear dynamical systems, part I: structural stability

International Nuclear Information System (INIS)

Storace, M; De Feo, O

2005-01-01

This paper and its companion address the problem of the approximation/identification of nonlinear dynamical systems depending on parameters, with a view to their circuit implementation. The proposed method is based on a piecewise-linear approximation technique. In particular, this paper describes the approximation method and applies it to some particularly significant dynamical systems (topological normal forms). The structural stability of the PWL approximations of such systems is investigated through a bifurcation analysis (via continuation methods)

The plant formin AtFH4 interacts with both actin and microtubules, and contains a newly identified microtubule-binding domain

Czech Academy of Sciences Publication Activity Database

Deeks, M.J.; Fendrych, Matyáš; Smertenko, A.; Bell, K.S.; Oparka, K.; Cvrčková, F.; Žárský, Viktor; Hussey, P.J.

2010-01-01

Roč. 123, č. 8 (2010), s. 1209-1215 ISSN 0021-9533 R&D Projects: GA MŠk(CZ) LC06004; GA ČR GAP305/10/0433 Institutional research plan: CEZ:AV0Z50380511 Keywords : Actin regulating proteins * Membrane * Microtubule Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 6.290, year: 2010

Power system dynamics and stability with synchrophasor measurement and power system toolbox

CERN Document Server

Sauer, Peter W; Chow, Joe H

2017-01-01

This new edition addresses the needs of dynamic modeling and simulation relevant to power system planning, design, and operation, including a systematic derivation of synchronous machine dynamic models together with speed and voltage control subsystems. Reduced-order modeling based on integral manifolds is used as a firm basis for understanding the derivations and limitations of lower-order dynamic models. Following these developments, a multi-machine model interconnected through the transmission network is formulated and simulated using numerical simulation methods. Energy function methods are discussed for direct evaluation of stability. Small-signal analysis is used for determining the electromechanical modes and mode-shapes, and for power system stabilizer design. Time-synchronized high-sampling-rate phasor measurement units (PMUs) to monitor power system disturbances ave been implemented throughout North America and many other countries. In this second edition, new chapters on synchrophasor measurement ...

Dynamic Stability Analysis of Autonomous Medium-Voltage Mixed-Source Microgrid

DEFF Research Database (Denmark)

Zhao, Zhuoli; Yang, Ping; Guerrero, Josep M.

2015-01-01

-space model of the autonomous MV mixed-source microgrid containing diesel generator set (DGS), grid-supporting battery energy storage system (BESS), squirrel cage induction generator (SCIG) wind turbine and network is developed. Sensitivity analysis is carried out to reveal the dynamic stability margin...

Biophysics of filament length regulation by molecular motors

International Nuclear Information System (INIS)

Kuan, Hui-Shun; Betterton, M D

2013-01-01

Regulating physical size is an essential problem that biological organisms must solve from the subcellular to the organismal scales, but it is not well understood what physical principles and mechanisms organisms use to sense and regulate their size. Any biophysical size-regulation scheme operates in a noisy environment and must be robust to other cellular dynamics and fluctuations. This work develops theory of filament length regulation inspired by recent experiments on kinesin-8 motor proteins, which move with directional bias on microtubule filaments and alter microtubule dynamics. Purified kinesin-8 motors can depolymerize chemically-stabilized microtubules. In the length-dependent depolymerization model, the rate of depolymerization tends to increase with filament length, because long filaments accumulate more motors at their tips and therefore shorten more quickly. When balanced with a constant filament growth rate, this mechanism can lead to a fixed polymer length. However, the mechanism by which kinesin-8 motors affect the length of dynamic microtubules in cells is less clear. We study the more biologically realistic problem of microtubule dynamic instability modulated by a motor-dependent increase in the filament catastrophe frequency. This leads to a significant decrease in the mean filament length and a narrowing of the filament length distribution. The results improve our understanding of the biophysics of length regulation in cells. (paper)

Mapping the structural and dynamical features of kinesin motor domains.

Directory of Open Access Journals (Sweden)

Guido Scarabelli

Full Text Available Kinesin motor proteins drive intracellular transport by coupling ATP hydrolysis to conformational changes that mediate directed movement along microtubules. Characterizing these distinct conformations and their interconversion mechanism is essential to determining an atomic-level model of kinesin action. Here we report a comprehensive principal component analysis of 114 experimental structures along with the results of conventional and accelerated molecular dynamics simulations that together map the structural dynamics of the kinesin motor domain. All experimental structures were found to reside in one of three distinct conformational clusters (ATP-like, ADP-like and Eg5 inhibitor-bound. These groups differ in the orientation of key functional elements, most notably the microtubule binding α4-α5, loop8 subdomain and α2b-β4-β6-β7 motor domain tip. Group membership was found not to correlate with the nature of the bound nucleotide in a given structure. However, groupings were coincident with distinct neck-linker orientations. Accelerated molecular dynamics simulations of ATP, ADP and nucleotide free Eg5 indicate that all three nucleotide states could sample the major crystallographically observed conformations. Differences in the dynamic coupling of distal sites were also evident. In multiple ATP bound simulations, the neck-linker, loop8 and the α4-α5 subdomain display correlated motions that are absent in ADP bound simulations. Further dissection of these couplings provides evidence for a network of dynamic communication between the active site, microtubule-binding interface and neck-linker via loop7 and loop13. Additional simulations indicate that the mutations G325A and G326A in loop13 reduce the flexibility of these regions and disrupt their couplings. Our combined results indicate that the reported ATP and ADP-like conformations of kinesin are intrinsically accessible regardless of nucleotide state and support a model where neck

Dynamic remedial action scheme using online transient stability analysis

Science.gov (United States)

Shrestha, Arun

Economic pressure and environmental factors have forced the modern power systems to operate closer to their stability limits. However, maintaining transient stability is a fundamental requirement for the operation of interconnected power systems. In North America, power systems are planned and operated to withstand the loss of any single or multiple elements without violating North American Electric Reliability Corporation (NERC) system performance criteria. For a contingency resulting in the loss of multiple elements (Category C), emergency transient stability controls may be necessary to stabilize the power system. Emergency control is designed to sense abnormal conditions and subsequently take pre-determined remedial actions to prevent instability. Commonly known as either Remedial Action Schemes (RAS) or as Special/System Protection Schemes (SPS), these emergency control approaches have been extensively adopted by utilities. RAS are designed to address specific problems, e.g. to increase power transfer, to provide reactive support, to address generator instability, to limit thermal overloads, etc. Possible remedial actions include generator tripping, load shedding, capacitor and reactor switching, static VAR control, etc. Among various RAS types, generation shedding is the most effective and widely used emergency control means for maintaining system stability. In this dissertation, an optimal power flow (OPF)-based generation-shedding RAS is proposed. This scheme uses online transient stability calculation and generator cost function to determine appropriate remedial actions. For transient stability calculation, SIngle Machine Equivalent (SIME) technique is used, which reduces the multimachine power system model to a One-Machine Infinite Bus (OMIB) equivalent and identifies critical machines. Unlike conventional RAS, which are designed using offline simulations, online stability calculations make the proposed RAS dynamic and adapting to any power system

The nucleoporin MEL-28 promotes RanGTP-dependent γ-tubulin recruitment and microtubule nucleation in mitotic spindle formation.

Science.gov (United States)

Yokoyama, Hideki; Koch, Birgit; Walczak, Rudolf; Ciray-Duygu, Fulya; González-Sánchez, Juan Carlos; Devos, Damien P; Mattaj, Iain W; Gruss, Oliver J

2014-01-01

The GTP-bound form of the Ran GTPase (RanGTP), produced around chromosomes, drives nuclear envelope and nuclear pore complex (NPC) re-assembly after mitosis. The nucleoporin MEL-28/ELYS binds chromatin in a RanGTP-regulated manner and acts to seed NPC assembly. Here we show that, upon mitotic NPC disassembly, MEL-28 dissociates from chromatin and re-localizes to spindle microtubules and kinetochores. MEL-28 directly binds microtubules in a RanGTP-regulated way via its C-terminal chromatin-binding domain. Using Xenopus egg extracts, we demonstrate that MEL-28 is essential for RanGTP-dependent microtubule nucleation and spindle assembly, independent of its function in NPC assembly. Specifically, MEL-28 interacts with the γ-tubulin ring complex and recruits it to microtubule nucleation sites. Our data identify MEL-28 as a RanGTP target that functions throughout the cell cycle. Its cell cycle-dependent binding to chromatin or microtubules discriminates MEL-28 functions in interphase and mitosis, and ensures that spindle assembly occurs only after NPC breakdown.

The zinc dyshomeostasis hypothesis of Alzheimer's disease.

Directory of Open Access Journals (Sweden)

Travis J A Craddock

Full Text Available Alzheimer's disease (AD is the most common form of dementia in the elderly. Hallmark AD neuropathology includes extracellular amyloid plaques composed largely of the amyloid-β protein (Aβ, intracellular neurofibrillary tangles (NFTs composed of hyper-phosphorylated microtubule-associated protein tau (MAP-tau, and microtubule destabilization. Early-onset autosomal dominant AD genes are associated with excessive Aβ accumulation, however cognitive impairment best correlates with NFTs and disrupted microtubules. The mechanisms linking Aβ and NFT pathologies in AD are unknown. Here, we propose that sequestration of zinc by Aβ-amyloid deposits (Aβ oligomers and plaques not only drives Aβ aggregation, but also disrupts zinc homeostasis in zinc-enriched brain regions important for memory and vulnerable to AD pathology, resulting in intra-neuronal zinc levels, which are either too low, or excessively high. To evaluate this hypothesis, we 1 used molecular modeling of zinc binding to the microtubule component protein tubulin, identifying specific, high-affinity zinc binding sites that influence side-to-side tubulin interaction, the sensitive link in microtubule polymerization and stability. We also 2 performed kinetic modeling showing zinc distribution in extra-neuronal Aβ deposits can reduce intra-neuronal zinc binding to microtubules, destabilizing microtubules. Finally, we 3 used metallomic imaging mass spectrometry (MIMS to show anatomically-localized and age-dependent zinc dyshomeostasis in specific brain regions of Tg2576 transgenic, mice, a model for AD. We found excess zinc in brain regions associated with memory processing and NFT pathology. Overall, we present a theoretical framework and support for a new theory of AD linking extra-neuronal Aβ amyloid to intra-neuronal NFTs and cognitive dysfunction. The connection, we propose, is based on β-amyloid-induced alterations in zinc ion concentration inside neurons affecting stability of

The zinc dyshomeostasis hypothesis of Alzheimer's disease.

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Craddock, Travis J A; Tuszynski, Jack A; Chopra, Deepak; Casey, Noel; Goldstein, Lee E; Hameroff, Stuart R; Tanzi, Rudolph E

2012-01-01

Alzheimer's disease (AD) is the most common form of dementia in the elderly. Hallmark AD neuropathology includes extracellular amyloid plaques composed largely of the amyloid-β protein (Aβ), intracellular neurofibrillary tangles (NFTs) composed of hyper-phosphorylated microtubule-associated protein tau (MAP-tau), and microtubule destabilization. Early-onset autosomal dominant AD genes are associated with excessive Aβ accumulation, however cognitive impairment best correlates with NFTs and disrupted microtubules. The mechanisms linking Aβ and NFT pathologies in AD are unknown. Here, we propose that sequestration of zinc by Aβ-amyloid deposits (Aβ oligomers and plaques) not only drives Aβ aggregation, but also disrupts zinc homeostasis in zinc-enriched brain regions important for memory and vulnerable to AD pathology, resulting in intra-neuronal zinc levels, which are either too low, or excessively high. To evaluate this hypothesis, we 1) used molecular modeling of zinc binding to the microtubule component protein tubulin, identifying specific, high-affinity zinc binding sites that influence side-to-side tubulin interaction, the sensitive link in microtubule polymerization and stability. We also 2) performed kinetic modeling showing zinc distribution in extra-neuronal Aβ deposits can reduce intra-neuronal zinc binding to microtubules, destabilizing microtubules. Finally, we 3) used metallomic imaging mass spectrometry (MIMS) to show anatomically-localized and age-dependent zinc dyshomeostasis in specific brain regions of Tg2576 transgenic, mice, a model for AD. We found excess zinc in brain regions associated with memory processing and NFT pathology. Overall, we present a theoretical framework and support for a new theory of AD linking extra-neuronal Aβ amyloid to intra-neuronal NFTs and cognitive dysfunction. The connection, we propose, is based on β-amyloid-induced alterations in zinc ion concentration inside neurons affecting stability of polymerized

Static and dynamic stability of pneumatic vibration isolators and systems of isolators

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Ryaboy, Vyacheslav M.

2014-01-01

Pneumatic vibration isolation is the most widespread effective method for creating vibration-free environments that are vital for precise experiments and manufacturing operations in optoelectronics, life sciences, microelectronics, nanotechnology and other areas. The modeling and design principles of a dual-chamber pneumatic vibration isolator, basically established a few decades ago, continue to attract attention of researchers. On the other hand, behavior of systems of such isolators was never explained in the literature in sufficient detail. This paper covers a range of questions essential for understanding the mechanics of pneumatic isolation systems from both design and application perspectives. The theory and a model of a single standalone isolator are presented in concise form necessary for subsequent analysis. Then the dynamics of a system of isolators supporting a payload is considered with main attention directed to two aspects of their behavior: first, the static stability of payloads with high positions of the center of gravity; second, dynamic stability of the feedback system formed by mechanical leveling valves. The direct method of calculating the maximum stable position of the center of gravity is presented and illustrated by three-dimensional stability domains; analytic formulas are given that delineate these domains. A numerical method for feedback stability analysis of self-leveling valve systems is given, and the results are compared with the analytical estimates for a single isolator. The relation between the static and dynamic phenomena is discussed.

Chromosome Bridges Maintain Kinetochore-Microtubule Attachment throughout Mitosis and Rarely Break during Anaphase.

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Pampalona, Judit; Roscioli, Emanuele; Silkworth, William T; Bowden, Brent; Genescà, Anna; Tusell, Laura; Cimini, Daniela

2016-01-01

Accurate chromosome segregation during cell division is essential to maintain genome stability, and chromosome segregation errors are causally linked to genetic disorders and cancer. An anaphase chromosome bridge is a particular chromosome segregation error observed in cells that enter mitosis with fused chromosomes/sister chromatids. The widely accepted Breakage/Fusion/Bridge cycle model proposes that anaphase chromosome bridges break during mitosis to generate chromosome ends that will fuse during the following cell cycle, thus forming new bridges that will break, and so on. However, various studies have also shown a link between chromosome bridges and aneuploidy and/or polyploidy. In this study, we investigated the behavior and properties of chromosome bridges during mitosis, with the idea to gain insight into the potential mechanism underlying chromosome bridge-induced aneuploidy. We find that only a small number of chromosome bridges break during anaphase, whereas the rest persist through mitosis into the subsequent cell cycle. We also find that the microtubule bundles (k-fibers) bound to bridge kinetochores are not prone to breakage/detachment, thus supporting the conclusion that k-fiber detachment is not the cause of chromosome bridge-induced aneuploidy. Instead, our data suggest that while the microtubules bound to the kinetochores of normally segregating chromosomes shorten substantially during anaphase, the k-fibers bound to bridge kinetochores shorten only slightly, and may even lengthen, during anaphase. This causes some of the bridge kinetochores/chromosomes to lag behind in a position that is proximal to the cell/spindle equator and may cause the bridged chromosomes to be segregated into the same daughter nucleus or to form a micronucleus.

Nonlinear Slewing Spacecraft Control Based on Exergy, Power Flow, and Static and Dynamic Stability

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Robinett, Rush D.; Wilson, David G.

2009-10-01

This paper presents a new nonlinear control methodology for slewing spacecraft, which provides both necessary and sufficient conditions for stability by identifying the stability boundaries, rigid body modes, and limit cycles. Conservative Hamiltonian system concepts, which are equivalent to static stability of airplanes, are used to find and deal with the static stability boundaries: rigid body modes. The application of exergy and entropy thermodynamic concepts to the work-rate principle provides a natural partitioning through the second law of thermodynamics of power flows into exergy generator, dissipator, and storage for Hamiltonian systems that is employed to find the dynamic stability boundaries: limit cycles. This partitioning process enables the control system designer to directly evaluate and enhance the stability and performance of the system by balancing the power flowing into versus the power dissipated within the system subject to the Hamiltonian surface (power storage). Relationships are developed between exergy, power flow, static and dynamic stability, and Lyapunov analysis. The methodology is demonstrated with two illustrative examples: (1) a nonlinear oscillator with sinusoidal damping and (2) a multi-input-multi-output three-axis slewing spacecraft that employs proportional-integral-derivative tracking control with numerical simulation results.

EFFECT OF SHOULDER SIDE PACK ON DYNAMIC POSTURAL STABILITY IN YOUNG HEALTHY FEMALE

Directory of Open Access Journals (Sweden)

Enas ELsayed Mohamed Abutaleb

2016-06-01

Full Text Available Background: Control of balance is a complex motor skill that involves integration of sensory inputs and the planning and execution of flexible movement patterns. Carrying side packs is famous in our society especially shoulder side packs. Most students carry shoulder side packs and they don't care about the way to carry them to be more balanced. The purpose of the study is to investigate the effect of carrying shoulder side pack on dynamic postural stability and to determine the best way of carrying a shoulder side pack either on the dominant side or non-dominant side that doesn’t affect dynamic postural stability in young healthy female. Methods: Sixty female volunteers aged from 18 to 25 years old participated in the study. Biodex balance system was used to measure the dynamic postural stability in three different occasions (without carrying a shoulder side pack, with carrying a shoulder side pack on the dominant side, and on the non-dominant side with a rest period in between. Results: Repeated measure analysis of variance (ANOVA followed by Bonferroni post hoc test were used to compare dynamic posture balance without carrying and during carrying a shoulder side pack on dominant and non-dominant sides. Analysis revealed that overall, anteroposterior and mediolateral stability indexes reduced significantly (P<0.0001 when carrying shoulder side pack on dominant side in comparison with when carrying shoulder side pack on non-dominant side and without carrying bag. Conclusion: It was concluded that carrying a shoulder side pack on the non-dominant side didn't disturb the postural stability when compared to carrying on the dominant side so, we recommend the students to carry shoulder side packs on the non-dominant side.

Dynamic stabilization of disruption precursors in tokamak

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Maoquan, Wang; Jianshan, Mao; Yuan, Pan [Academia Sinica, Hefei, AH (China). Inst. of Plasma Physics

1994-12-01

A method for dynamic stabilization of the disruption precursors in tokamak is proposed, that is a controlled ac current induced and added to the equilibrium current. The ac currents applied can be a sine alternative current with a relevant frequency, or a pulsed current with a suitable pulsed width {tau} and or a discontinuous pulsed current whose width {tau} is very shorter than the intervals between pulses, and or a `sawtooth` pulsed current with the time of ramp phase of the sawtooth is very much shorter than the sawtooth descending time, the ratio of them can be {<=}10{sup -3}. The physical model of the ac current drive is analyzed in detail. The suppression role of the ac current on the MHD perturbations was analyzed in theory and proved numerically. It is indicated that the ac current can make the discontinuous derivative, {Delta}`, more favorable for the tearing mode stabilities, and so, as long as the parameters of the applied ac currents are selected suitably, the MHD perturbations can be suppressed effectively, the perturbations will be in the zero-growing state, the profile of the plasma current and temperature remain in the initial states and not variate basically, the tokamak be in the stabilized operation state. (8 figs.).

Vacuolar and cytoskeletal dynamics during elicitor-induced programmed cell death in tobacco BY-2 cells.

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Higaki, Takumi; Kadota, Yasuhiro; Goh, Tatsuaki; Hayashi, Teruyuki; Kutsuna, Natsumaro; Sano, Toshio; Hasezawa, Seiichiro; Kuchitsu, Kazuyuki

2008-09-01

Responses of plant cells to environmental stresses often involve morphological changes, differentiation and redistribution of various organelles and cytoskeletal network. Tobacco BY-2 cells provide excellent model system for in vivo imaging of these intracellular events. Treatment of the cell cycle-synchronized BY-2 cells with a proteinaceous oomycete elicitor, cryptogein, induces highly synchronous programmed cell death (PCD) and provide a model system to characterize vacuolar and cytoskeletal dynamics during the PCD. Sequential observation revealed dynamic reorganization of the vacuole and actin microfilaments during the execution of the PCD. We further characterized the effects cryptogein on mitotic microtubule organization in cell cycle-synchronized cells. Cryptogein treatment at S phase inhibited formation of the preprophase band, a cortical microtubule band that predicts the cell division site. Cortical microtubules kept their random orientation till their disruption that gradually occurred during the execution of the PCD twelve hours after the cryptogein treatment. Possible molecular mechanisms and physiological roles of the dynamic behavior of the organelles and cytoskeletal network in the pathogenic signal-induced PCD are discussed.

Beam stability ampersand nonlinear dynamics. Formal report