Dissertations / Theses on the topic 'Spindle'
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Satyarebala, Chilaka. "Role of CENP-A NAC/CAD network in spindle assembly and spindle checkpoint /." Zürich : ETH, 2008. http://e-collection.ethbib.ethz.ch/show?type=diss&nr=18103.
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Rout, Michael Paul. "The identification and characterization of components of the yeast spindle pole body and spindle." Thesis, University of Cambridge, 1989. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.334226.
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Müller-Reichert, Thomas. "Spindle organization in three dimensions." Doctoral thesis, Saechsische Landesbibliothek- Staats- und Universitaetsbibliothek Dresden, 2006. http://nbn-resolving.de/urn:nbn:de:swb:14-1166107130476-22269.
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During cell division, chromosome segregation takes place on bipolar, microtubulebased spindles. Here, C. elegans is used to analyze spindle organization under both mitotic and meiotic conditions. First, the role of SAS-4 in organizing centrosome structure was analyzed. Partial depletion of SAS-4 in early embryos results in structurally defective centrioles. The study of this protein sheds light on the poorly understood role of the centrioles in dictating centrosome size. Second, the ultrastructure of wild-type mitotic spindle components was analyzed by electron tomography. This 3-D analysis reveals morphologically distinct microtubule end morphologies in the mitotic spindle pole. These results have structural implications for models of microtubule interactions with centrosomes Third, spindle assembly was studied in female meiosis. Specifically, the role of the microtubule severing complex katanin in spindle organization was analyzed. Electron tomography reveals fragmentation of spindle microtubules and suggests a novel katanin-dependent mechanism of meiotic spindle assembly. In this model, relatively long microtubules seen near the meiotic chromatin are converted into numerous short fragments, thus increasing the total number of polymers in an acentrosomal environment. Taken together, these results provide novel insights into the three-dimensional organization of microtubules during spindle assemblyDie Segregation der Chromosomen während der Zellteilung wird duch bipolare, von Microtubuli-aufgebauten Spindlen gewährleistet. In der vorliegenden Arbeit wird C. elegans zur Analyse der Spindelorganisation unter mitotischen und meiotischen Bedingungen herangezogen. Erstens wird die Rolle von SAS-4 in der Organisation von Zentrosomen untersucht. Die partielle Depletierung von SAS-4 in frühen Embryonen führt zu strukturell defekten Zentriolen und wirft somit Licht auf die wenig verstandene Rolle der Zentriolen in der Bestimmung der Zentrosomengröße. Zweitens wird die Ultrastruktur der mitotischen Spindelkomponenten im Wildtyp durch Elektronentomographie untersucht. Diese 3-D-Analyse zeigt, dass im mitotischen Spindlepol unterschiedliche Morphologien der Mikrotubulienden zu finden sind. Diese Ergebnisse haben strukturelle Implikationen für Modelle der Mikrotubuli-Zentrosomen-Interaktionen. Drittens wird der Aufbau der Spindel in der weiblichen Meiose, speziell die Rolle des Mikrotubuli-schneidenden Kataninkomplexes in der Spindelorganisation, untersucht. Die Elektronentomographie zeigt hier eine Fragmentierung der Spindelmikrotubuli. Basierend auf diesem Ergebnis wird ein neues Katanin-abhängiges Modell der Formierung der Meiosespindel entwickelt, in dem relativ lange Microtubuli in Nähe des meiotischen Chromatins in zahlreiche kurze Mikrotubuli “zerschnitten” werden. Dies erhöht die Anzahl der verfügbaren Polymere in dieser azentrosomalen Situation. Zusammenfassend bringen diese Ergebnisse neue Einsichten in die räumliche Organisation der Mikrotubuli während des Spindelaufbaus
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Wigge, Philip Anthony. "An analysis of the spindle and spindle pole body in the budding yeast Saccharomyces cerevisiae." Thesis, University of Cambridge, 2000. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.621860.
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Cavazza, Tommaso 1985. "Unravelling integrated kinetics during spindle assembly." Doctoral thesis, Universitat Pompeu Fabra, 2015. http://hdl.handle.net/10803/325428.
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To ensure faithful segregation of the genetic material, during mitosis the cell assembles the mitotic spindle, a macromolecular machine made of dynamic microtubules and associated proteins. Spindle microtubules are assembled de novo by the centrosomes and through the RanGTP pathway that is sufficient for spindle self-organization. In this thesis I used a proteomics approach to study the RanGTP pathway induced microtubules and spindle self-organization and I propose that it occurs through a multi-step mechanism. I then addressed how the centrosomal and acentrosomal microtubule assembly pathways are integrated to build the bipolar spindle. My data suggest that, in small cells, centrosome maturation defines an optimal and essential balance between these two pathways due to a competition for limiting amounts of tubulin. Interestingly in big cells, I found that centrosome maturation ensures the correct positioning of each centrosome to a spindle pole and thereby assures proper centrosome segregation to the daughter cells. Finally, I present molecular insights about the interaction of the microtubule polymerase XMAP215 with its mitotic interactor TACC3.Durant la mitosi, per assegurar que el material genètic es segrega correctament, la cèl•lula construeix el fus mitòtic, una maquinària macromolecular formada de microtúbuls dinàmics i proteïnes associades. Els microtúbuls del fus són formats de novo pels centrosomes o a través de la “RanGTP pathway”, la qual és autosuficient per promoure l'autoorganització del fus mitòtic. En aquesta tesi he utilitzat tècniques de proteòmica per estudiar com la “RanGTP pathway” indueix l'autoorganització dels microtúbuls i del fus mitòtic a través d’un procés amb múltiples passos (tal i com indiquen els resultats obtinguts). Paral•lelament he estudiat com els microtúbuls formats per ambdues vies (centrosomes i “RanGTP pathway”) es coordinen per la construcció del fus bipolar. Els meus resultats suggereixen que, en cèl•lules petites, la maduració dels centrosomes defineix un equilibri òptim i essencial entre els dos sistemes, com a conseqüència de la disponibilitat limitada de tubulina. Per altra banda, en cèl•lules grans la maduració dels centrosomes condiciona la correcta localització d’aquests als pols del fus mitòtic, assegurant-ne la segregació a les dues cèl•lules filles. Finalment, exposo els detalls moleculars de la interacció entre la polimerasa de microtúbuls XMAP215 i el seu interactor mitòtic TACC3.
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Overington, Y. H. "Aspects of hollow spindle fancy yarn." Thesis, University of Manchester, 1986. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.556105.
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Donaldson, Anne Dunlop. "The yeast spindle pole component spc42." Thesis, University of Cambridge, 1994. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.319938.
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Banks, Robert William. "Studies on the mammalian muscle spindle." Thesis, Durham University, 1994. http://etheses.dur.ac.uk/9594/.
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The subjects of these studies are the major components of the mammalian muscle spindle, which is an encapsulated proprioceptor serving to monitor skeletal muscle length and length change. Those components are: specialized intrafusal muscle fibres; sensory nerve endings that form intimate contacts with the intrafusal fibres; and motor nerve fibres by means of which the central nervous system can exercise control over the sensitivity of the spindle. My first important contribution was to establish the number of types of intrafusal fibre (1-8, 11). Their different mechanical properties help to shape the responses of the sensory endings in characteristic ways (papers 15 and 42). Detailed reconstructions of sensory endings revealed recognizable features of the primary ending that were consistently associated with the different intrafusal fibres (10, 13, 18, 20, 33). The sites of nerve impulse generation and coding are being studied in relation to the branching pattern of the sensory nerve fibres (45, 50, 55). Analysis of the innervation of individual spindles has revealed the interplay of random and deterministic factors in spindle construction (20, 36, 37, 40, 41, 44, 48, 52, 53). As yet it is unknown how the differences that exist between muscles in this respect are related to their specific roles in motor control or kinaesthesia. However, reflex activity appears to be grossly disturbed in muscles that have been reinnervated following nerve section, since functional endings may be formed in inappropriate locations (22, 25, 28- 31,34, 38, 39, 43, 46). The motor innervation of the spindle was for long controversial, especially concerning the distribution of the different functional categories of axon. I have pursued histophysiological and probabilistic approaches to this problem, about which there now appears to be a large measure of agreement in favour of my conclusions (9, 12, 19, 21, 23, 26, 27, 35, 41, 42, 44, 48, 49). Papers 1-6 in the following list are based on work that originally formed part of a thesis presented in candidature for the degree of Ph. D. in the Faculty of Medicine, University of Sheffield. For each full paper of which 1 am a co-author an estimate of my contribution to the overall effort is given as a percentage in the list.
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Yuan, Ivan. "Generation of synthetic spindle checkpoint signals." Thesis, University of Edinburgh, 2016. http://hdl.handle.net/1842/22030.
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The spindle checkpoint ensures proper chromosome segregation by monitoring kinetochore-microtubule interactions: unattached kinetochores recruit checkpoint proteins that combine to form a diffusible inhibitor which delays anaphase, thus buying cells time to fix attachment errors. Although the major checkpoint proteins were identified some 25 years ago, we have only just begun to understand how they assemble at unattached kinetochores to generate the crucial checkpoint signal. Much of this can be attributed to the difficulty associated with studying these proteins at the kinetochores, which are highly complex and thus often make clean dissection of function impossible. To circumvent this problem, a synthetic version of the spindle checkpoint was engineered on an ectopic location on a chromosome arm away from kinetochores in S. pombe. This work describes how the co-targeting of only two checkpoint components, the outer kinetochore protein Spc7 and the checkpoint kinase Mph1, was found to be sufficient to successfully generate a checkpoint-dependent metaphase arrest and how this paves the way for a clearer, more joined-up understanding of how the checkpoint works.
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Holinger, Eric P. "The spindle pole body phosphoproteome and the importance of phosphorylation on the Saccharomyces cerevisiae spindle pole body." Connect to online resource, 2008. http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&res_dat=xri:pqdiss&rft_dat=xri:pqdiss:3303825.
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Wang, Bin. "Phosphoproteome studies of human mitotic spindle proteins." Diss., lmu, 2010. http://nbn-resolving.de/urn:nbn:de:bvb:19-123191.
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Sen, Onur. "Phospho-regulation of the spindle assembly checkpoint." Thesis, University of Edinburgh, 2016. http://hdl.handle.net/1842/15873.
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Mitosis is a highly regulated process by which a cell duplicates and distributes its chromosomal DNA into two identical daughter cells equally. Equal distribution of the chromosomes is crucial for accurate propagation of genetic information. This is essential for maintaining viability and preventing genomic instability that can potentially lead to cancer. In order to avoid unequal distribution of chromosomes, cells employ a surveillance mechanism called the spindle assembly checkpoint (SAC). The SAC is an inhibitory signalling network, which delays segregation of chromosomes, until they have stably attached to spindle microtubules through their multi-protein platforms, known as kinetochores. The main target of the SAC is the anaphase promoter complex/ cyclosome (APC/C), an E3 ubiquitin ligase. Specifically APC/C and its activator Cdc20 are inhibited by the main effector of the SAC, called the mitotic checkpoint complex (MCC). The MCC consists of Cdc20, Mad2, Mad3 and Bub3 (except S. pombe) proteins, which are recruited to the unattached kinetochores to promote MCC assembly. Once the chromosomes stably attach to the spindle, the SAC is turned off, MCC disassembles, and APC/CCdc20 is released from the inhibition. Activated APC/CCdc20 then targets its two main substrates, securin and cyclin B, for proteasomal degradation, and thereby triggers anaphase onset and mitotic exit. SAC signalling involves many protein components, whose activities are essentially regulated by direct protein-protein interactions and/ or post-translational modifications. One of these major modifications is phosphorylation, which is mediated by the SAC kinases such as Aurora B, Mps1 and Bub1. A number of studies have characterised SAC related substrates of Aurora B and Mps1 kinases in several model organisms. On the other hand, Bub1 kinase activity has been thought to play a key role in chromosome bi-orientation and more of an auxiliary role in SAC activation. The aim of this study is to investigate the importance of Bub1 kinase activity for SAC response in fission yeast Schizosaccharomyces pombe (S. pombe). SAC activation assays, using various degrees of spindle perturbation, have demonstrated that Bub1 kinase activity plays an important role in SAC maintenance. In order to examine the pathways downstream of Bub1, we set out to indicate Bub1 substrates which may be involved in SAC signalling. According to studies in various species, Cdc20 appears to be a prominent candidate, whose phosphorylation by Cdk1 and Bub1 kinases has been reported to regulate its mitotic activity. To investigate whether Cdc20 is phosphorylated by Bub1 in vitro, we purified recombinant S. pombe proteins from insect cells. Subsequent kinase assays identified Cdc20 as an in vitro substrate of Bub1, and the phosphorylated sites in Cdc20 were mapped by mass spectrometry. To address if this phospho-modification is involved in SAC regulation, phosphorylation mutants of Cdc20 were analysed in terms of their abilities to activate and silence SAC in vivo. Results show that phosphorylation of Cdc20 C-terminus promotes SAC maintenance in response to spindle damage. Furthermore, the mutations mimicking Bub1-mediated phosphorylation of Cdc20 Cterminus restore the SAC defects in the absence of Bub1 kinase activity. In addition, we purified S. pombe mitotic checkpoint complex (MCC) from insect cells, and analysed the interactions between its components (Cdc20, Mad2 and Mad3) by cross-linking mass spectrometry. Crystal structure of S.pombe MCC has been determined recently, which lacks Mad3 C-terminus and flexible C-terminal tail of Cdc20. Using an MCC with full length Mad3, we identified novel interactions between the C-terminal tails Mad3 and Cdc20, which are in close proximity to the identified Cdc20 phosphorylation sites. Briefly, in this study we confirm the previously known roles of Bub1 kinase activity (chromosome bi-orientation). Moreover, we propose a new pathway (in addition to the well-established H2A pathway) mediated by Cdc20, that may be important to maintain the SAC response.
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Knezevich, Stevan Robert. "Molecular characterization of pediatric spindle cell tumors." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 2000. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape4/PQDD_0021/NQ48668.pdf.
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Plumb, Kemp. "Measuring mitotic spindle dynamics in budding yeast." Thesis, McGill University, 2010. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=86726.
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In order to carry out its life cycle and produce viable progeny through cell division, a cell must successfully coordinate and execute a number of complex processes with high fidelity, in an environment dominated by thermal noise. One important example of such a process is the assembly and positioning of the mitotic spindle prior to chromosome segregation. The mitotic spindle is a modular structure composed of two spindle pole bodies, separated in space and spanned by filamentous proteins called microtubules, along which the genetic material of the cell is held. The spindle is responsible for alignment and subsequent segregation of chromosomes into two equal parts; proper spindle positioning and timing ensure that genetic material is appropriately divided amongst mother and daughter cells.In this thesis, I describe fluorescence confocal microscopy and automated image analysis algorithms, which I have used to observe and analyze the real space dynamics of the mitotic spindle in budding yeast. The software can locate structures in three spatial dimensions and track their movement in time. By selecting fluorescent proteins which specifically label the spindle poles and cell periphery, mitotic spindle dynamics have been measured in a coordinate system relevant to the cell division. I describe how I have characterised the accuracy and precision of the algorithms by simulating fluorescence data for both spindle poles and the budding yeast cell surface.
In this thesis I also describe the construction of a microfluidic apparatus that allows for the measurement of long time-scale dynamics of individual cells and the development of a cell population. The tools developed in this thesis work will facilitate in-depth quantitative analysis of the non-equilibrium processes in living cells.
La régulation du cycle cellulaire et la prolifération de générations viables requièrent à la fois la reproductibilité et la coordination des différents processus complexes en jeu dans un environnement toutefois dominé par l'agitation thermique. Un exemple essentiel est l'assemblage et la migration du fuseau mitotique qui doivent avoir lieu correctement avant même la ségrégation des chromosomes. Le fuseau mitotique est une structure transitoire composée de deux pôles séparés par des filaments de protéines appelés les microtubules, auxquelles est rattaché le matériel génétique de la cellule. Le fuseau mitotique est notamment impliqué dans l'alignement des chromosomes, puis leur ségrégation vers des pôles opposés de la cellule ; d'où la nécessité d'un positionnement spatial précis et temporellement coordonné du fuseau mitotique pour assurer la division correcte du matériel génétique entre les cellules mère et fille.
Dans ce mémoire, je décrirai les techniques de microscopie confocale par fluorescence ainsi que les algorithmes automatisés d'analyse d'image, que j'ai mis en oeuvre pour observer et analyser la dynamique spatiale en temps réel du fuseau mitotique chez la levure bourgeonnante. Les programmes développés permettent de localiser dans l'espace tridimensionnel des structures subcellulaires et de détecter leurs déplacements au cours du temps. Le marquage par protéines fluorescentes des pôles du fuseau mitotique et de la membrane cellulaire a permis de quantifier la dynamique du fuseau mitotique dans un système de coordonnées pertinent pour la division cellulaire. Je décrirai des simulations numériques de signaux fluorescents de ces structures subcellulaires qui m'ont permis de caractériser la fiabilité et de quantifier la précision des programmes d'analyse.
Je terminerai ce mémoire par la description d'un dispositif microfluidique permettant à la fois la culture de cellules et la caractérisation de leur dynamique à l'échelle individuelle, et ce sur de longues échelles de temps. Les outils développés au cours de cette thèse et présentés dans ce mémoire offrent la possibilité d'analyses quantitatives nouvelles des processus hors équilibre en jeu chez les cellules vivantes en général.
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Kutlu, Asim. "Design and development of a lathe spindle." Thesis, KTH, Maskinkonstruktion (Inst.), 2016. http://urn.kb.se/resolve?urn=urn:nbn:se:kth:diva-187477.
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Verktygsmaskiner möjliggör tillverkning av materiekroppar med olika form. Svarvar är de vanligaste maskinerna för att bearbeta runda detaljer. Man kan säga att en svarvspindeln möjliggör hela skärprocessen och att den därför är den viktigaste komponenten i en svarv. En svarvaxel, eller en spindel, bearbetar det roterande ämnet med ett stationärt skärverktyg, och materialet tas bort vid skärverktygets kontakt med ämnet.Det här arbetet syftar till att konstruera en spindel som uppfyller en given kravspecifikation. Specifikationen innehåller prestandakrav, som rotationshastighet och kraft, geometri och dimensionskrav, som storlek och håldiameter, samt vilka komponenttyper som ska användas, som t.ex. motorn.I enlighet medprestandakravet definieras maximalt lastscenario. Med givna svarvningsparametrar beräknas skärkraften på materialet vid dess kontakt med skärverktyget. En inbyggd motor som är tillräckligt kraftfull och uppfyller hastighets- och effektkraven väljs och en preliminär konstruktion skapas med valda lager, lagerordningar och axelmaterial.Beteendet av den preliminära spindelkonstruktionen, under påverkan av skärkrafter och rotation analyseras statiskt och dynamiskt med hjälp av FEM och analytiska modeller. Inom ett givet område optimeras avståndet mellan lagren, vilka utgör spindelns stöd.Genom verifikation och optimering av den preliminära konstruktionen skapas den slutliga detaljkonstruktionen. Slutkonstruktionen innefattar alla nödvändiga detaljer och viktiga krav beaktas och balanseras, som att vara tillverkningsbar och monterbar, tätningssystem, ett system för hastighetsmätning och lägesmätning, kabelvägar och andra nödvändiga punkter. I slutänden konstrueras en spindel som uppfyller den definierade kravspecifikationen.Machine tools enable the industry to shape almost any material by a variety of methods. Lathes are one of the most common machines to cut circular parts with precision and accuracy. And the spindle of a lathe can be entitled as the most critical mechanical component which makes the cutting process possible. A lathe spindle rotates the workpiece to be cut against a stationary rigid cutting tool, therefore removing material through the contact edge. In this thesis, a spindle is aimed to be designed which complies with a set of specifications defined. These specifications consists of performance requirements such as speed and power, dimensional constraints for space and bore diameter, and component types which must be used, such as for the motor.Based on the performance requirements, a maximum loading case with cutting parameters is defined. With these cutting parameters, cutting forces acting to the material from at the contact point with the cutting tool are calculated. A built-in motor with sufficient power and speed specifications is selected based on the maximum cutting forces and speed requirements. A preliminary design is made up by selecting bearings, bearing arrangements and shaft material.With static and dynamic analysis conducted on the preliminary design through analytical models and FEM, the behavior of the spindle is investigated separately under the cutting forces and during the rotation. Within an allowable range, optimization is made on the bearing span distances which are the support locations for the spindle.Following the verification and optimization of the preliminary design, the final detail design of the spindle is made. The final design includes the design of all the necessary parts, by taking the manufacturability, assemblability, sealing design, a system for speed and position measurements, cable paths and more necessary points into account. Ultimately, a spindle which meets the requirements and specifications successfully is designed as the expected outcome of this thesis.
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Winters, Lora. "Mechanism of spindle assembly in Schizosaccharomyces pombe-." Doctoral thesis, Saechsische Landesbibliothek- Staats- und Universitaetsbibliothek Dresden, 2017. http://nbn-resolving.de/urn:nbn:de:bsz:14-qucosa-225764.
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At the onset of cell division microtubules growing from spindle pole bodies (SPB) interact with each other to form the mitotic spindle enabling proper chromosome positioning and segregation. However, the exact mechanism of microtubule dynamics and microtubule associated proteins (MAPs) underlying spindle assembly is still not well understood. We developed an in vivo method to observe spindle assembly in the fission yeast Schizosaccharomyces pombe by inducing depolymerization of already formed and grown spindles by subjecting the cells to low temperatures, followed by subsequent repolymerization at a permissive temperature. We observed that microtubules pivot, i.e., perform angular movement around the SPB in a random manner, exploring the intranuclear space. Eventually microtubules extending from opposite SPBs come into contact and establish an antiparallel connection thus reassembling the spindle. Mutant approaches revealed that deletion of ase1 and klp5 did not prevent spindle reassembly, however introduced aberrations during the spindle formation. Amazingly, cut7p showed direct colocalization with microtubule overlap during spindle reassembly. Abrogation of cut7p led to inability to form a functional spindle. Thus, cut7p is the main regulator of spindle formation in fission yeast. None of the mutant strains affected microtubule pivoting, confirming that microtubule pivoting is a random movement unrelated to MAPs.
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Wells, Allan R. "Spectral analysis of multi-spindle machining heads /." Online version of thesis, 1994. http://hdl.handle.net/1850/12019.
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Sundberg, Holly. "Analysis of the spindle pole component Spc110p /." Thesis, Connect to this title online; UW restricted, 1996. http://hdl.handle.net/1773/9225.
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Connolly, Amy. "Oocyte Meiotic Spindle Assembly in Caenorhabditis Elegans." Thesis, University of Oregon, 2014. http://hdl.handle.net/1794/18492.
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As in many organisms, Caenorhabditis elegans oocytes assemble bipolar meiotic spindles in the absence of centrosomes. While the assembly of the mitotic spindle in C. elegans has been studied in some detail, how the poles assemble in the absence of centrosomes remains poorly understood. In an ongoing screen for temperature-sensitive (TS), embryonic-lethal mutants, we have identified TS mutations in multiple genes required for oocyte meiotic spindle pole assembly. We have so far identified mutations in four genes: or1178ts in mei-1, which encodes the catalytic domain of the microtubule severing complex katanin; or447ts in klp-18, which encodes a kinesin 12 family member; or645ts in aspm-1, which encodes a microtubule scaffolding protein; and or1092ts and or1292ts in klp-7, which encode a kinesin 13/MCAK family member. By using live cell imaging of oocytes from transgenic strains expressing GFP and mCherry fusion to proteins associated with the spindle, we have found and confirmed other findings that klp-18 promotes spindle bipolarity and that MEI-1 promotes pole assembly both by severing microtubules and by recruiting ASPM-1. More recently, we have found that klp-7 is required for maintaining bipolarity in the meiotic spindle by preventing the number of poles that can form. In klp-7(-) mutants, we observed in addition to extraneous poles an excess accumulation of microtubules during Meiosis I. Futhermore, reducing klp-7 function can restore bipolarity in a klp-18(-) monopolar spindle mutant background. We also observed that disruption of the kinetochore factor KNL-1 in klp-7(-) mutants exacerbates the extra spindle pole phenotype. We suggest that in oocyte meiosis, klp-7 is required to limit microtubule accumulation and pole assembly and that it may carry out these functions in a kinetochore-dependent manner.
This dissertation includes previously published co-authored material.
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Dunsch, Anja Katrin. "Control of the mitotic spindle by dynein light chain 1 complexes." Thesis, University of Oxford, 2013. http://ora.ox.ac.uk/objects/uuid:b2fd5670-a035-42ca-aaef-78a30aeaa084.
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Robust control mechanisms ensure faithful inheritance of an intact genome through the processes of mitosis and cytokinesis. Different populations of the cytoplasmic dynein motor defined by specific dynein adaptor complexes are required for the formation of a stable bipolar mitotic spindle. This study analysed how different dynein subcomplexes contribute to spindle formation and orientation. Various dynein subpopulations were identified by mass spectrometry. I have shown that the dynein light chain 1 (DYNLL1) directly interacts with the kinetochore localised Astrin-Kinastrin complex as well as the spindle microtubule associated complex formed by CHICA and HMMR. I have characterised both complexes and identified unique functions in chromosome alignment and mitotic spindle orientation, respectively. I have found that Kinastrin (C15orf23) is the major Astrin-interacting protein in mitotic cells and is required for Astrin targeting to microtubule plus ends proximal to the plus tip tracking protein EB1. Fixed cell microscopy revealed that cells over-expressing or depleted of Kinastrin mislocalise Astrin. Additionally, depletion of the Astrin-Kinastrin complex delays chromosome alignment and causes the loss of normal spindle architecture and sister chromatid cohesion before anaphase onset (Dunsch et al., 2011). Using immunoprecipitation and microtubule binding assays, I have shown that CHICA and HMMR interact with one another, and target to the spindle by a microtubule-binding site in the amino-terminal region of HMMR. CHICA interacts with DYNLL1 by a series of conserved TQT motifs in the carboxy-terminal region. Depletion of DYNLL1, CHICA or HMMR causes a slight increase in mitotic index but has little effect on spindle formation or checkpoint function. Fixed and live cell microscopy reveal, however, that the asymmetric distribution of cor tical dynein is lost and the spindle in these cells fails to orient correctly in relation to the culture surface (Dunsch et al., 2012). These findings presented here suggest that the Astrin-Kinastrin complex is required for normal spindle architecture and chromosome alignment, and that per turbations of this pathway result in delayed mitosis and non-physiological separase activation, whereas HMMR and CHICA act as par t of a dynein-DYNLL1 complex with a specific function defining or controlling spindle orientation.
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Huňka, Radek. "Výměna nástrojů u svislého soustruhu." Master's thesis, Vysoké učení technické v Brně. Fakulta strojního inženýrství, 2012. http://www.nusl.cz/ntk/nusl-230356.
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This master's thesis deals with the problems of tool changing of vertical turning lathe. Specifically about modular system of spindle locking in a ram. The master's thesis is shows types of structural design, choosing the best design with supporting technical - economics evaluation. After this is realized series of calculation controls. These define dimensions of the mechanical parts.
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Costa, Mariana Fernandes Alves. "Molecular remodelling of the spindle architecture during metaphase arrest in oocytes." Thesis, University of Edinburgh, 2018. http://hdl.handle.net/1842/31255.
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Oocytes of most species assemble and maintain a functional bipolar spindle in the absence of centrosomes. Strikingly, after bipolar spindle formation, oocytes arrest in metaphase for several hours before fertilisation. How the dynamic spindle maintains its bipolarity during this long arrest is poorly understood. I hypothesise that the bipolar spindle is stably maintained by changes in the distribution of microtubule-associated proteins (MAPs) on the spindle during the long oocyte arrest. To test this, I generated transgenic flies expressing GFP-tagged microtubule-associated proteins (MAPs), and found that 13 out of 24 proteins change localisation between early and late oocytes. I refer to these changes in MAP localisation after establishment of bipolarity as 'spindle maturation'. In order to identify the molecular mechanisms triggering MAP relocalisation, I manipulated the kinase activity of the cell cycle regulator Cdk1 by over-expressing non-degradable cyclin A or B, the major activators of Cdk1. Their expression prevented re-localisation of distinct sets of MAPs, and disrupted spindle bipolarity and accurate chromosome segregation in oocytes. Kinesin-6 Pavarotti/MKlp1 localised strongly to the spindle equator in late oocytes, whilst nearly always absent from this region in early oocytes. The localisation of Pavarotti to the spindle equator in late oocytes was reduced when cyclin B is over-expressed in oocytes, suggesting a role for Cdk1/cyclin B complex in regulating Pavarotti localisation. Indeed, a Pavarotti/Mklp1 mutant non-phosphorylatable by Cdk1 prematurely localised to the meiotic spindle and disrupted spindle bipolarity. Moreover, removal of Pavarotti from the metaphase-I spindle by RNAi induced spindle defects in oocytes. Therefore, it is likely that the microtubule cross-linking activity of Pavarotti enhances the stability of the metaphase-I spindle during the long arrest. Consistent with this, I found that the microtubule density in the spindle equator is higher in late oocytes. Altogether, I propose that remodelling the molecular architecture of the spindle during the long oocyte arrest is important to stabilise the bipolar spindle without centrosomes.
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Rahman, Aminur Khosru. "Mathematical modelling of vibrations in spindle bearing assemblies." Thesis, Brunel University, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.264691.
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Karthikeyan, Bindu Kumar. "Tribo-dynamics of high speed precision spindle bearings." Thesis, Loughborough University, 2008. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.547399.
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Turner, Craig Robert. "Transduction and adaptation in the frog muscle spindle." Thesis, University of Cambridge, 1997. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.627207.
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Rodgers, Justin Fraser. "Classification and projection strength of muscle spindle afferents." Thesis, King's College London (University of London), 1995. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.309421.
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Pan, Qinwei. "Dynamic control of spindle motors for milling applications." Thesis, University of Sheffield, 2018. http://etheses.whiterose.ac.uk/16333/.
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This programme of research is concerned with investigating novel methods for enhancing the machining of metals in terms of increasing material removal rates while maintaining surface finish and dimensional tolerance. This thesis provides a detailed investigation of the interaction between the drive control system, the power converter and the cutting process. Specifically, the research explores the merits of high speed and precise torque modulation of the drive and its influence on the cutting tool to suppress chatter in milling operations. The coupling between the controller and power converter behaviour and the dynamic cutting conditions is the key novelty in this research.
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Brown, Austin (Austin R. ). "Axially force limited grinding spindle for robotic grinding." Thesis, Massachusetts Institute of Technology, 2018. http://hdl.handle.net/1721.1/119966.
Full textAbstract:
Thesis: S.B., Massachusetts Institute of Technology, Department of Mechanical Engineering, 2018.Cataloged from PDF version of thesis.
Includes bibliographical references (page 35).
Grinding and Polishing of small parts is often easily performed by human hands, yet is challenging to automate. The grinding and polishing process is best done using a force-control scheme, which human hands perform naturally. Heavy robotic arms, which favor a position-control scheme, are difficult to control precisely, and trajectory errors can cause excessive grinding force which leads to burning of the part or destruction of the grinding wheel. Prior art of direct force control on a large robot arm requires the end-effector to have a 6-axis dynamometer, which is unwieldy, costly, and greatly limits the speed/precision of the process. We will discuss a new type of grinding spindle which is axially compliant, allowing the position-control robot arm to be used in a force-control nature. The spindle has a disjoint force-displacement curve, effectively operating in two modes: position-control mode at first, until a critical force is exceeded, when the spindle transitions into force-mode, keeping constant grinding force on the part though a certain range of travel. This limits the amount of force which can be imparted during grinding to a safe amount. The spindle is very simple and mechanically robust. We have built this hybrid position-force control spindle and tested it. The spindle was shown to perform correctly and successfully completed the test grind.
by Austin Brown.
S.B.
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Dodson, Helen. "Analysis of the spindle assembly checkpoint in vertebrates." Thesis, University of Edinburgh, 2003. http://hdl.handle.net/1842/13671.
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I have cloned the chicken BubR1 cDNA and raised antibodies to the protein. The protein is highly conserved when compared to other vertebrate BubR1s with 55% identity and 70% similarity to human. The antibody recognises a protein of approximately 150 kDa and stains the kinetochores of chicken cells during prometaphase with the signal disappearing as they become attached to microtubules, a localisation typical of spindle checkpoint proteins. In order to further characterise the function of this protein in vertebrates, I have attempted to generate a chicken DT40 cell line conditionally null for BubR1. I have successfully targeted one allele of the BubR1 locus in DT40 cells. BubR1 (+/-) cells grow normally and have an intact checkpoint. Since I expect BubR1 to be essential, the gene must be expressed under the control of the tetracycline operator in the heterozygote cells. Attempts to target the second allele have failed thus far although the expected targeting frequency is 1/20 clones. It appears that expression of exogenous BubR1 is detrimental to these cells and they shut off expression when possible. It seems that expression levels of this essential checkpoint component are critical to avoid problems within the cell. In addition to this study in chicken cells I have cloned two other vertebrate checkpoint components from Xenopus laevis. The proteins Zw10 and Rod were initially identified in Drosophila and since no homologues have been identified in yeast these are considered as metazoan-specific components of the spindle assembly checkpoint. In order to study the checkpoint and the kinetochore, I have cloned partial Xenopus cDNA for Zw10 and Rod and raised antibodies to these proteins. Zw10 is localised to the kinetochores of metaphase Xenopus cells and along with Rod is present in a complex of approximately 11S in Xenopus egg extracts.
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Nixon, Faye Margaret. "A three-dimensional study of mitotic spindle ultrastructure." Thesis, University of Liverpool, 2016. http://livrepository.liverpool.ac.uk/3001861/.
Full textAbstract:
The mitotic spindle is the complex machinery that enables a cell to segregate its genetic material faithfully. Without fidelity in this process, genetic abnormalities may occur which, in turn, can lead to cancer. Microtubules of the mitotic spindle are bundled together to form kinetochore fibres which contribute to the movement of replicated chromosomes during cell division, helping to ensure the DNA is equally divided. Electron microscopy has long been used to observe inter-microtubule bridges within kinetochore fibres and, although they are thought to confer stability to the fibres thus ensuring faithful mitosis, their identity and function is yet to be fully established. These inter-microtubule cross-linkers were thought to be simple structures, providing a bridge directly between two microtubules. Here, three dimensional electron microscopy was used to examine these cross-linkers in unprecedented detail - revealing a complex network of microtubule connectors which we have termed 'the mesh'. To establish the functional role of the mesh, levels of a known microtubule crosslinker, the TACC3-chTOG-clathrin complex, were manipulated. In cells with too much mesh, kinetochore microtubules were found to be disorganised - forming tighter clusters within fibres and deviating from their parallel trajectories. Kinetochore fibres were also found to contain more microtubules than in control cells, and took longer to complete mitosis. These observations provide evidence that the mesh plays a functional role in the organisation within kinetochore fibres, directly impacting the ability of the cell to divide successfully. In addition to fine kinetochore fibre structure, scanning electron microscopy was optimised to examine changes to the whole spindle structure after manipulation of the mesh. This revealed changes to spindle architecture when microtubuleassociated proteins were altered, and offers insight into how cancer can be initiated when these proteins are dysregulated.
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Breuer, Manuel. "The role of HURP in acentriolar spindle assembly." Paris 6, 2010. http://www.theses.fr/2010PA066013.
Full textAbstract:
Les fuseaux acentriolaires se forment grâce à la nucléation de microtubules à partir des MTOCs. Les mécanismes qui aboutissent à l’assemblage de la région centrale du fuseau, essentielle pour la mise en place de la bipolarité, sont très peu connus. HURP (Hepatoma UpRegulated Protein) s’accumule de préférence sur les microtubules dans cette région. HURP est également une cible de RanGTP. Par ailleurs, l’accumulation est dépendante du moteur Eg5, un facteur essentiel à la mise en place de la bipolarité. Les femelles invalidées pour HURP sont stériles. Une analyse détailleé in vivo montre que HURP est requis pour l’établissement et le maintien de la bipolarité. Des mesures de croissance et de densité des microtubules par microscopie de type spinning disk démontrent un rôle crucial de HURP dans l’assemblage des microtubules à proximité des chromosomes. Je propose un modèle où HURP, stabilisateur de microtubules dans la région enrichie en MTs interpolaires, assure un échafaudage permettant la formation d’un fuseau robuste et la congression des chromosomes
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Lizarraga, Sofia B. "Regulation of spindle assembly by the Ran pathway." Available to US Hopkins community, 2003. http://wwwlib.umi.com/dissertations/dlnow/3080716.
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Parthasarathy, Anand Kumar. "Feasibility analysis of FPGA based spindle motor controller." Diss., Online access via UMI:, 2009.
Find full textAbstract:
Thesis (M.S.)--State University of New York at Binghamton, Thomas J. Watson School of Engineering and Applied Science, Department of Electrical and Computer Engineering, 2009.Includes bibliographical references.
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Sarhan, Ahmed Aly Diaa Mohammed. "Monitoring of cutting forces by the intelligent spindle." 京都大学 (Kyoto University), 2007. http://hdl.handle.net/2433/136224.
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Hannabuss, J. C. "In vitro reconstitution of the anaphase central spindle." Thesis, University College London (University of London), 2018. http://discovery.ucl.ac.uk/10044566/.
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The anaphase central spindle is the major regulatory scaffold for coordinating cytokinesis; the final stage in cell division. It is constructed from two populations of microtubules, which emanate from opposite halves of the cell, until they meet at the centre, form antiparallel overlaps, and are cross-linked by microtubule bundling proteins. This cross-linked overlapping region marks the division plane, and recruits proteins that promote the membrane cleavage furrow, and the final abscission of the two daughter cells. For robust cortical signalling, and localisation of the abscission apparatus, the cell must regulate the length and alignment of the overlapping region to create a clear narrow band. What are the molecular mechanisms underlying this regulation? To address this question, I use a novel assay, which allows me to use TIRF microscopy to study free-floating, dynamic, microtubule bundles, nucleated de novo in solution, and organised by purified recombinant proteins. By reverse engineering the anaphase central spindle in this way, I have discovered that only two human central spindle proteins, PRC1 and KIF4a, are necessary to organise dynamic microtubules into bundled structures with narrow, aligned, antiparallel overlaps. These results also suggest a novel functional role for human KIF4a, as an antiparallel sliding motor, capable of aligning microtubule overlaps in a bundle.
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Roca, Marianne. "The spindle assembly checkpoint in Phallusia mammillata embryos." Electronic Thesis or Diss., Sorbonne université, 2019. http://www.theses.fr/2019SORUS500.
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Le point de contrôle du fuseau mitotique (Spindle Assembly Checkpoint : SAC) retarde l’anaphase jusqu’à ce que tous les chromosomes soient attachés correctement aux microtubules. Le SAC permet ainsi d’éviter des erreurs de ségrégation des chromosomes aboutissant à des cellules filles aneuploïdes (i.e. avec un nombre anormal de chromosomes). L’aneuploïdie, délétère pour les cellules, peut entrainer des problèmes de développement et est observée dans les cancers. Cependant, chez certaines espèces, le SAC n’est pas efficace au cours de la phase précoce du développement embryonnaire. J’ai mis en évidence que chez l’ascidie P. mammillata, un organisme marin du groupe des chordés, le SAC devient efficace au 8ème cycle cellulaire et son efficacité augmente dans les cycles suivants. J’ai démontré qu’en partie ventrale l’identité des cellules antérieures induisait la présence d’un SAC plus efficace mais que d’autres facteurs modulaient aussi l’efficacité du SAC. J’ai étudié les mécanismes moléculaires impliqués dans les variations de l’efficacité du SAC au cours du développement. Mes expériences ont révélé la présence des composants du SAC tout au long de l’embryogenèse. Cependant, j’ai pu montrer que les protéines du SAC ne se localisent pas au niveau des kinétochores lorsque le SAC est inefficace au début du développement mais qu’elles s’y localisent bien dans l’ovocyte en méiose et dans l’embryon plus tardif, lequel se caractérise par un SAC actif. Ma thèse a permis de montrer que P. mammillata est un organisme expérimental de grand intérêt pour l’étude du SAC au cours de l’embryogenèseDuring mitosis, progression through anaphase must take place only when all chromosomes are correctly attached to spindle microtubules to avoid chromosome mis-segregation and the generation of aneuploid cells (i.e. with an abnormal chromosome number). Embryos containing aneuploid cells can exhibit developmental defects and lethality. Furthermore, cancer cells are often aneuploid. To prevent such deleterious aneuploidy, a control mechanism, the spindle assembly checkpoint (SAC), delays metaphase-anaphase transition until all chromosomes are properly attached to spindle microtubules. However, the SAC is not efficient during early development in some species. During my thesis, I analyzed the activity of the SAC during the development of the marine chordate P. mammillata. I showed that in P. mammillata embryos, the SAC becomes efficient at the 8th cell cycle and its efficiency increases progressively in the following cell cycles. Although, I demonstrated that patterning of the embryo along the anteroposterior axis influences SAC efficiency, my experiments suggest that additional parameters modulate SAC efficiency. I searched the molecular mechanisms, which control SAC efficiency during development. I collected evidence showing that SAC components are present in oocytes and all post-fertilization stages. I found that SAC proteins localize at kinetochores during meiosis and at later stages when there is an efficient SAC while they do not accumulate on unattached kinetochores in early SAC deficient embryos. My thesis work establishes P. mammillata as a valuable experimental organism to study SAC regulation during embryogenesis
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Zapletal, Jan. "Návrh vřetene obráběcího stroje." Master's thesis, Vysoké učení technické v Brně. Fakulta strojního inženýrství, 2016. http://www.nusl.cz/ntk/nusl-241859.
Full textAbstract:
This master’s thesis deals with design of spindle for small three axis milling machine. The first part of this thesis includes research in milling spindles and overview of several spindles from different manufacturers. The second part deals with calculation of necessary parameters, design of spindle and subsequent simulation verification. This thesis also includes a 3D model of spindle, assembly drawing and four manufacturing drawings.
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Žádník, Lubomír. "Návrh vřeteníku soustružnického centra." Master's thesis, Vysoké učení technické v Brně. Fakulta strojního inženýrství, 2020. http://www.nusl.cz/ntk/nusl-417727.
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The diploma thesis deals with the design of a built-in headstock of a turning center. The aim of the work is the design of a spindle for machines up to 10 kW. The spindle is driven by a two-speed gearbox and a belt drive. In the introduction, the work deals with the description of individual parts of the turning headstock. A design proposal is performed, accompanied by a number of technical calculations. The result of the work is a 3D model, drawing documentation and calculation report.
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Burian, Viktor. "Návrh vřeteníku frézovacího centra." Master's thesis, Vysoké učení technické v Brně. Fakulta strojního inženýrství, 2018. http://www.nusl.cz/ntk/nusl-382051.
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This master´s thesis deals with the design of a cartridge spindle for milling machines. The aim of the work is to design a spindle with a power of 15 kW, which allows the automatic exchange of tools with ISO interface. In the beginning, the thesis deals with the description of the individual components related to the construction of machine tools spindle. A spindle design concept is chosen, for which a number of calculations were made concerning the structure itself. Based on the proposed design, a 3D model and drawing documentation were made.
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Florian, Stefan [Verfasser]. "Bipolar spindle formation beyond Eg5: A study on antagonists and synergists of the molecular motor Eg5 during mitotic spindle formation / Stefan Florian." Konstanz : Bibliothek der Universität Konstanz, 2011. http://d-nb.info/1038383951/34.
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Stevenson, Deja Lee. "Whole-Body Vibration and Its Effects on Electromechanical Delay and Vertical Jump Performance." Diss., CLICK HERE for online access, 2005. http://contentdm.lib.byu.edu/ETD/image/etd867.pdf.
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Eliscovich, Carolina. "Spindle-Localized CPE-Mediated Translation Controls Mediotic Chromosome Segregation." Doctoral thesis, Universitat Pompeu Fabra, 2008. http://hdl.handle.net/10803/7123.
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La progresión meiótica y el desarrollo embrionario temprano están programados, en parte, por la activación tradcuccional de mRNAs maternos como lo son los que codifican para las proteinas de ciclina B1 o mos. Estos mRNAs no son traducidos al mismo tiempo ni en el mismo lugar. Por lo contrario, su traducción está especificamente regulada por elementos de poliadenilación citoplasmática (CPEs) presentes en sus 3'UTRs. Los elementos CPEs reclutan a la proteina de unión a CPE (CPE-binding protein CPEB (Colegrove-Otero et al., 2005; de Moor et al., 2005; Mendez and Richter, 2001; Richter, 2007)). Esta proteina de unión al RNA no sólo determina cuándo y en qué medida un mRNA será activado traduccionalmente por poliadenilación citoplasmática (Mendez et al., 2000a; Mendez et al., 2000b; Mendez et al., 2002) sino que también participa, junto con el represor de la traducción Maskin, en el transporte y la localización de sus mRNAs diana hacia los sitios de localización subcelular donde su traducción ocurrirá (Huang et al., 2003; Huang and Richter, 2004). Durante el desarrollo embrionario de Xenopus, CPEB se encuentra localizada en el polo animal de los oocitos y más tarde, sobre el huso mitótico y centrosomas en el embrión (Groisman et al., 2000). Se ha demostrado que embriones de Xenopus inyectados con agentes que interrumpen la traducción dependiente de poliadenilación citoplasmática, detienen la división celular y presentan estructuras mitóticas anormales (Groisman et al., 2000). En este trabajo que derivó en mi tesis doctoral, hemos demostrado que la activación traduccional localizada en el huso mitótico de mRNAs regulados por CPEB que codifican para proteinas con una conocida función en aspectos estructurales del ciclo celular como la formación del huso mitótico y la segregación cromosómica, es esencial para completar la primera división meiótica y para la correcta segregación cromosómica en oocitos de Xenopus.
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Vanneste, David. "The role of molecular motor HKLP2 in spindle assembly." Doctoral thesis, Universitat Pompeu Fabra, 2010. http://hdl.handle.net/10803/31822.
Full textAbstract:
During cell division, a macromolecular machine is assembled to segregate the two sets of chromosomes carrying the genetic material into the two daughter cells. This machine, the mitotic spindle, is a bipolar array composed of microtubules and proteins that control their dynamics and organization. Kinesins are molecular motors that can apply forces to the microtubules and arrange them in space to assemble a bipolar spindle.
Here we characterized the human kinesin Hklp2 as an important player in bipolar spindle assembly and maintenance.
Hklp2 is a kinesin that localizes to the spindle microtubules and the chromosomes in mitosis. Its localization to the microtubules depends on TPX2, a crucial factor in microtubule nucleation. Hklp2 localization to the chromosomes necessitates the presence of cell proliferation marker Ki67 whose function is currently unknown. Hklp2 helps the separation of the centrosomes during the establishment of spindle bipolarity and plays a role in the subsequent maintenance of bipolarity during metaphase. This role is probably achieved through a cross-linking of the microtubules. Interestingly, the populations of the kinesin present on the microtubules and the chromosomes have different functions.Durante la división celular, tiene lugar el ensamblaje de la maquinaria macromolecular encargada de segregar los dos conjuntos de cromosomas en las dos células hijas. Esta maquinaria, el huso mitótico, es una matriz bipolar compuesta de microtúbulos y de proteínas que controlan la dinámica y la organización. Las quinesinas son motores moleculares que pueden ejercer fuerzas sobre los microtúbulos y organizarlos en el espacio para dar lugar al huso bipolar. Este estudio aporta evidencias del papel importante de la quinesina humana Hklp2 en el ensamblaje y en la estabilidad del huso mitótico. Hklp2 es una quinesina que se localiza en los microtúbulos del huso y en los cromosomas durante la mitosis. Su localización en los microtúbulos depende de TPX2, un factor importante en la nucleación de microtúbulos. Para que Hklp2 se localice en los cromosomas, este necesita la presencia del marcador de proliferación Ki67, la función del cual se desconoce actualmente. Hklp2 participa en la separación de los centrosomas durante el establecimiento de la bipolaridad del huso mitótico y posteriormente juega un papel importante en el mantenimiento de esta bipolaridad durante la metafase. Esta función probablemente se consiga haciendo lazos entre los microtúbulos. Las poblaciones de la quinesina presente en los microtúbulos y en los cromosomas tienen funciones diferentes.
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Schlaitz, Anne-Lore. "Regulation of Mitotic Spindle Assembly in Caenorhabditis elegans Embryos." Doctoral thesis, Saechsische Landesbibliothek- Staats- und Universitaetsbibliothek Dresden, 2007. http://nbn-resolving.de/urn:nbn:de:swb:14-1181247079528-57268.
Full textAbstract:
The mitotic spindle is a bipolar microtubule-based structure that mediates proper cell division by segregating the genetic material and by positioning the cytokinesis cleavage plane. Spindle assembly is a complex process, involving the modulation of microtubule dynamics, microtubule focusing at spindle poles and the formation of stable microtubule attachments to chromosomes. The cellular events leading to spindle formation are highly regulated, and mitotic kinases have been implicated in many aspects of this process. However, little is known about their counteracting phosphatases. A screen for genes required for early embryonic cell divisions in C. elegans identified rsa-1 (for regulator of spindle assembly 1), a putative Protein Phosphatase 2A (PP2A) regulatory subunit whose silencing causes defects in spindle formation. Upon rsa-1(RNAi), spindle poles collapse onto each other and microtubule amounts are strongly reduced. My thesis work demonstrates that RSA-1 indeed functions as a PP2A regulatory subunit. RSA-1 associates with the PP2A enzyme and recruits it to centrosomes. The centrosome binding of PP2A furthermore requires the new protein RSA-2 as well as the core centrosomal protein SPD-5 and is based on a hierarchical protein-protein interaction pathway. When PP2A is lacking at centrosomes after rsa-1(RNAi), the centrosomal amounts of two critical mitotic effectors, the microtubule destabilizer KLP-7 and the kinetochore microtubule stabilizer TPXL-1, are altered. KLP-7 is increased, which may account for the reduction of microtubule outgrowth from centrosomes in rsa-1(RNAi) embryos. TPXL-1 is lost from centrosomes, which may explain why spindle poles collapse in the absence of RSA-1. TPXL-1 physically associates with RSA-1 and RSA-2, suggesting that it is a direct target of PP2A. In summary, this work defines the role of a novel PP2A complex in mitotic spindle assembly and suggests a model for how different microtubule re-organization steps might be coordinated during spindle formation.
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Erturk, Alper. "Dynamic Modeling Of Spindle-tool Assemblies In Machining Centers." Master's thesis, METU, 2006. http://etd.lib.metu.edu.tr/upload/12607236/index.pdf.
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Regenerative chatter is a well-known machining problem that results in unstable cutting process, poor surface quality, reduced material removal rate and damage on the machine tool itself. Stability lobe diagrams supply stable depth of cut –
spindle speed combinations and they can be used to avoid chatter. The main requirement for generating the stability lobe diagrams is the system dynamics information at the tool tip in the form of point frequency response function (FRF). In this work, an analytical model that uses structural coupling and modification methods for modeling the dynamics of spindle-holder-tool assemblies in order to obtain the tool point FRF is presented. The resulting FRF obtained by the model can be used in the existing analytical and numerical models for constructing the stability lobe diagrams. Timoshenko beam theory is used in the model for improved accuracy and the results are compared with those of Euler-Bernoulli beam theory. The importance of using Timoshenko beam theory in the model is pointed out, and the circumstances, under which the theory being used in the model becomes more important, are explained. The model is verified by comparing the results obtained by the model with those of a reliable finite element software for a case study. The computational superiority in using the model developed against the finite element software is also demonstrated. Then, the model is used for studying the effects of bearing and contact dynamics at the spindle-holder and holder-tool interfaces on the tool point FRF. Based on the results of the effect analysis, a new approach is suggested for the identification of bearing and interface parameters from experimental measurements, which is demonstrated on a spindle-holder-tool assembly. The model is also employed for studying the effects of design and operational parameters on the tool point FRF, from the results of which, suggestions are made regarding the design of spindles and selection of operational parameters. Finally, it is experimentally demonstrated that the stability lobe diagram of an assembly can be predicted pretty accurately by using the model proposed, and furthermore the stability lobe diagram can be modified in a predictable manner for improving chatter stability.
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Smith, Christopher. "Kinetochore dynamics and their attachment to the mitotic spindle." Thesis, University of Warwick, 2016. http://wrap.warwick.ac.uk/83227/.
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Chromosome segregation is a mechanical process that requires assembly of the mitotic spindle – a dynamic microtubule-based force-generating machine (Dumont & Mitchison, 2009). Connections between chromosomes and the spindle are mediated by kinetochores, pairs of multi-protein complexes assembled on centromeric chromatin that harness the pushing and pulling forces associated with microtubule dynamics to power chromosome movement (Rago & Cheeseman, 2013). This structure is reported to be compliant (Wan et al., 2009; Maresca & Salmon, 2009; Uchida et al., 2009; Dumont et al., 2012; Drpic et al., 2015; Etemad et al., 2015; Tauchman et al., 2015; Magidson et al., 2016), which is the basis upon which it can transduce forces, and satisfy mitotic checkpoints. However recent studies have suggested that the kinetochore is a non-compliant linkage (Suzuki et al., 2014), while others negate the requirement for intra-kinetochore stretch to allow onset of anaphase (Etemad et al., 2015; Tauchman et al., 2015; Magidson et al., 2016). It is therefore crucial that the reasons for these differences are elucidated, and a new model developed to explain regulatory pathways sensitive to intra-kinetochore structure. I have developed a novel semi-automated imaging assay to track kinetochore proteins tagged with two different coloured fluorescent proteins in 3D, coupled with a detailed assay for measurement of the required correction for 3D chromatic aberration directly from cells. I reveal using three dimensional tracking that the kinetochore forms a non-compliant linkage between centromeric chromatin and kintetochore spindle fibres. Instead the outer kinetochore layer can swivel around the inner kinetochore/centromere, which results in a reduction in intra-kinetochore distance when viewed in lower dimensions, the only 3D measurement thus far being by Etemad et al. (2015). I show that swivel provides a mechanical flexibility that enables kinetochores at the periphery of the spindle to engage microtubules. Swivel reduces as cells approach anaphase, explaining previous suggestions that the kinetochore needs to be stretched upon bi-orientation prior to anaphase onset (Maresca & Salmon, 2009; Uchida et al., 2009). Reduction in swivel suggests there is an organisational change linked to checkpoint satisfaction, and/or changes required in kinetochore mechanochemistry before sister chromatid dysjunction. I show that Cdk1 inactivation is required to suppress kinetochore-microtubule dynamics in anaphase, and therefore may mediate required changes in kinetochore state. Finally, my work opens up the possibility to map the 3D architecture of the kinetochore, and between different domains within proteins and complexes. With such an architectural understanding, it will now be possible to investigate the mechanisms of organisational changes that occur during cycles of microtubule attachment and the regulatory processes of error-correction and SAC activation/silencing.
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Tarailo, Maja. "Spindle assembly checkpoint and chromosome stability in Caenorhabditis elegans." Thesis, University of British Columbia, 2007. http://hdl.handle.net/2429/5587.
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In many species, proper chromosome segregation is accomplished with the aid of a
surveillance mechanism, the spindle assembly checkpoint. To identify the mechanisms involved
in this process, the mutants that suppress or enhance the mdf-1(gk2)/MAD1 checkpoint lethality
were characterized. The suppressors of mdf-1 (gk2) fall into two classes. The major class of
suppressors compensates for the loss of the checkpoint by delaying mitotic progression. This
class includes two known suppressors and anaphase promoting complex/cyclosome (APC/C)
components, emb-30/APC4 and fzy-1/CDC2O, and four new such (suppressors of spindle
checkpoint defect) genes. One of the new such genes was found to be an APC5-like gene not
previously identified as a component of the APC/C in C. elegans. This analysis revealed that
APC5 and APCJO genes have paralogs in the C. elegans genome.
Furthermore, a class of suppressors was identified that does not delay mitotic
progression. In mouse cells, mutations that result in defective apoptosis rescued the lethality
associated with deletion of the Mad2 gene. The suppressor mutants were analyzed for the
apoptotic response and the such-7(h1985) suppressor with normal anaphase onset was found to
abrogate the DNA damage-induced apoptosis. Despite the defective apoptotic response in this
suppressor, loss of apoptosis alone could not rescue the mdf-1 lethality in C. elegans, indicating
that other processes affected by the such-7 mutation, could account for the rescue of mdf-1(gk2)
lethality.
The upstream components required for the genome stability would be recognized as
enhancers of the mdf-1 (gk2) lethality. In yeast, 79 genes were found to result in synthetic lethal
phenotype with MAD1. Of the 21 non-essential putative C. elegans orthologs assayed, nine
enhanced mdf-1(gk2) lethality. The enhancers have a specific effect on the SAC, since six of
them enhanced the mdf-2(tm2910)/MAD2 lethality, three also enhanced the san-1 (ok1580)
lethality and none enhanced lethality in the kinetochore mutant him-1O(e15l1ts)/NUF2. In
addition two interactions, hcp-1 and bub-3, were identified in C. elegans that are not conserved
in yeast. This analysis also showed that HCP-1 and HCP-2, the two CENP-F-related proteins,
have a non redundant role, since only the hcp-1(RNAi) enhances lethality of the SAC mutants.
48
Decker, Franziska. "Mechanisms of microtubule nucleation in metaphase spindles and how they set spindle size." Doctoral thesis, 2017. https://tud.qucosa.de/id/qucosa%3A31788.
Full textAbstract:
Regulation of size and growth is a fundamental problem in biology and often closely related to functionality and fitness. A prominent example is the mitotic spindle, whose size needs to be perfectly tuned to ensure proper chromosome segregation during cell division. It is known that spindle size generally scales with cell volume, most likely as a result of limiting components. However, this relation breaks down in very large cells where spindles have a maximum size. How the size and microtubule mass are set and why spindles show an upper size limit in large cells is still not understood.
Spindles mainly consist of highly dynamic short microtubules that turn over very quickly in comparison to the lifetime of the entire structure. Thus, microtubules need to be constantly created throughout the spindle, a process called nucleation. Understanding the role of microtubule nucleation in setting the size of spindles is limited by the fact that little is known about the rate, distribution, and regulation of microtubule nucleation in these structures. This is partly due to the lack of methods to measure microtubule nucleation in spindles.
During this work, I developed an assay based on laser ablation to probe microtubule nucleation in monopolar spindles assembled in Xenopus laevis egg extract. Using this new method in combination with quantitative microscopy, I found that microtubule nucleation in these structures is spatially regulated. Furthermore, I observed that nucleation is stimulated by pre-existing microtubules leading to new microtubule growth in their physical proximity.
Combining my experimental results on nucleation with theory and further biochemical perturbations, I show that this autocatalytic nucleation mechanism is limited by the availability of active nucleators. In spindles, the amount of active nucleators decreases with distance from the chromosomes. Thus, this mechanism provides an upper limit to spindle size even when resources are not limiting.
49
Maeda, Osamu. "Expert spindle design system." Thesis, 2003. http://hdl.handle.net/2429/14534.
Full textAbstract:
Machine tool spindle is the most important mechanical component in removing metal during machining operations. The structural dynamics of the spindle are evaluated at the tool tip since it directly affects the material removal rate. Flexible spindles lead to unstable chatter vibrations, which can be avoided only by reducing the material removal rate. In addition, the spindle motor must have sufficient torque and power to overcome the cutting resistance of work materials to be machined. The spindles are currently designed based on accumulated experience, basic laws of machine design, and metal cutting mechanics. This thesis presents an expert spindle design system strategy which is based on the efficient utilization of past design experience, the laws of machine design and metal cutting dynamics. The configuration of the spindle is decided by identifying the work material, desired cutting conditions and most common tools which will be used on the machine tool. The spindle drive mechanism, drive motor, bearing types, and spindle shaft dimensions are selected based on the target application. The thesis provides a set of fuzzy design rules which lead to interactive and automatic design of spindle drive configurations. The structural dynamics of the spindle are automatically optimized by distributing the bearings along the spindle shaft. The proposed strategy is to iteratively predict the Frequency Response Function (FRF) of the spindle at the tool tip using the Finite Element Method (FEM) based on Timoshenko Beam elements. Predicted FRF of the spindle is integrated to the chatter vibration stability law which indicates whether the design would lead to chatter vibration free cutting operation at the desired speed and depth of cut. The bearing spacings are iteratively optimized without violating the design constraints of the spindle. The proposed expert system design is demonstrated by automatically designing several spindles which are found on industrial machine tools.
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Lai, Yu-An, and 賴昱安. "Automatic Identification of Sleep Spindle." Thesis, 2007. http://ndltd.ncl.edu.tw/handle/22125809941288803350.
Full textAbstract:
碩士淡江大學
電機工程學系碩士班
95
Sleep disorders has an important part in area of the psychiatric diseases. Usually, it''s also the complication of other diseases. Sleep apnea syndrome and sleep dyssomnia are common sleep diseases, which sometimes will even cause the death. Sleep spindles are transient sleep EEG waveforms. Along with K-complexes, they are the hallmarks of Stage 2 sleep. Furthermore, sleep spindles can be used to analyse sleep microstructure. The interest in sleep spindles lies not only in its representation to the onset of QS-2, but also in other interesting findings, such as those relating SS, normal aging, infant pathologies and sleeping positions. The visual pattern detection of all-night EEG recordings which have typically 1000 spindles is time consuming and tedious. Therefore, developing a fast and accurate automatic identification system of sleep spindle can reduce doctor’s labor and realize quantitative diagnosis of sleep spindle.