scholarly journals Regional variation of microtubule flux reveals microtubule organization in the metaphase meiotic spindle

2008 ◽  
Vol 182 (4) ◽  
pp. 631-639 ◽  
Author(s):  
Ge Yang ◽  
Lisa A. Cameron ◽  
Paul S. Maddox ◽  
Edward D. Salmon ◽  
Gaudenz Danuser
Keyword(s):  
Chromosome Segregation ◽  
Regional Variation ◽  
Spatial Organization ◽  
Meiotic Spindle ◽  
Microtubule Organization ◽  
Spindle Poles ◽  
Egg Extract ◽  
Spindle Dynamics ◽  
Meiotic Spindles ◽  
Uniform Polarity

Continuous poleward movement of tubulin is a hallmark of metaphase spindle dynamics in higher eukaryotic cells and is essential for stable spindle architecture and reliable chromosome segregation. We use quantitative fluorescent speckle microscopy to map with high resolution the spatial organization of microtubule flux in Xenopus laevis egg extract meiotic spindles. We find that the flux velocity decreases near spindle poles by ∼20%. The regional variation is independent of functional kinetochores and centrosomes and is suppressed by inhibition of dynein/dynactin, kinesin-5, or both. Statistical analysis reveals that tubulin flows in two distinct velocity modes. We propose an association of these modes with two architecturally distinct yet spatially overlapping and dynamically cross-linked arrays of microtubules: focused polar microtubule arrays of a uniform polarity and slower flux velocities are interconnected by a dense barrel-like microtubule array of antiparallel polarities and faster flux velocities.

scholarly journals The kinesin Eg5 drives poleward microtubule flux in Xenopus laevis egg extract spindles

2004 ◽  
Vol 167 (5) ◽  
pp. 813-818 ◽  
Author(s):  
David T. Miyamoto ◽  
Zachary E. Perlman ◽  
Kendra S. Burbank ◽  
Aaron C. Groen ◽  
Timothy J. Mitchison
Keyword(s):  
Xenopus Laevis ◽  
Chromosome Segregation ◽  
Spindle Poles ◽  
Egg Extract ◽  
Constant Rate ◽  
Quantitative Image ◽  
Meiotic Spindles ◽  
Kinesin Eg5 ◽  
Poleward Flux

Although mitotic and meiotic spindles maintain a steady-state length during metaphase, their antiparallel microtubules slide toward spindle poles at a constant rate. This “poleward flux” of microtubules occurs in many organisms and may provide part of the force for chromosome segregation. We use quantitative image analysis to examine the role of the kinesin Eg5 in poleward flux in metaphase Xenopus laevis egg extract spindles. Pharmacological inhibition of Eg5 results in a dose–responsive slowing of flux, and biochemical depletion of Eg5 significantly decreases the flux rate. Our results suggest that ensembles of nonprocessive Eg5 motors drive flux in metaphase Xenopus extract spindles.

scholarly journals Central-spindle microtubules are strongly coupled to chromosomes during both anaphase A and anaphase B

2019 ◽  
Vol 30 (19) ◽  
pp. 2503-2514 ◽  
Author(s):  
Che-Hang Yu ◽  
Stefanie Redemann ◽  
Hai-Yin Wu ◽  
Robert Kiewisz ◽  
Tae Yeon Yoo ◽  
...  
Keyword(s):  
Chromosome Segregation ◽  
Tomographic Reconstruction ◽  
Mitotic Spindles ◽  
Strongly Coupled ◽  
Central Spindle ◽  
Spindle Poles ◽  
Meiotic Spindles ◽  
Spindle Microtubules ◽  
Anaphase B ◽  
Chromosome Motion

Spindle microtubules, whose dynamics vary over time and at different locations, cooperatively drive chromosome segregation. Measurements of microtubule dynamics and spindle ultrastructure can provide insight into the behaviors of microtubules, helping elucidate the mechanism of chromosome segregation. Much work has focused on the dynamics and organization of kinetochore microtubules, that is, on the region between chromosomes and poles. In comparison, microtubules in the central-spindle region, between segregating chromosomes, have been less thoroughly characterized. Here, we report measurements of the movement of central-spindle microtubules during chromosome segregation in human mitotic spindles and Caenorhabditis elegans mitotic and female meiotic spindles. We found that these central-spindle microtubules slide apart at the same speed as chromosomes, even as chromosomes move toward spindle poles. In these systems, damaging central-spindle microtubules by laser ablation caused an immediate and complete cessation of chromosome motion, suggesting a strong coupling between central-spindle microtubules and chromosomes. Electron tomographic reconstruction revealed that the analyzed anaphase spindles all contain microtubules with both ends between segregating chromosomes. Our results provide new dynamical, functional, and ultrastructural characterizations of central-spindle microtubules during chromosome segregation in diverse spindles and suggest that central-spindle microtubules and chromosomes are strongly coupled in anaphase.

scholarly journals Central spindle microtubules are strongly coupled to chromosomes during both anaphase A and anaphase B

2019 ◽  
Author(s):  
Che-Hang Yu ◽  
Stefanie Redemann ◽  
Hai-Yin Wu ◽  
Robert Kiewisz ◽  
Tae Yeon Yoo ◽  
...  
Keyword(s):  
Chromosome Segregation ◽  
Tomographic Reconstruction ◽  
Mitotic Spindles ◽  
Strongly Coupled ◽  
Central Spindle ◽  
Spindle Poles ◽  
Meiotic Spindles ◽  
Spindle Microtubules ◽  
Anaphase B ◽  
Chromosome Motion

AbstractSpindle microtubules, whose dynamics vary over time and at different locations, cooperatively drive chromosome segregation. Measurements of microtubule dynamics and spindle ultrastructure can provide insight into the behaviors of microtubules, helping elucidate the mechanism of chromosome segregation. Much work has focused on the dynamics and organization of kinetochore microtubules, i.e. on the region between chromosomes and poles. In comparison, microtubules in the central spindle region, between segregating chromosomes, have been less thoroughly characterized. Here, we report measurements of the movement of central spindle microtubules during chromosome segregation in human mitotic spindles, and Caenorhabditis elegans mitotic and female meiotic spindles. We found that these central spindle microtubules slide apart at the same speed as chromosomes, even as chromosomes move towards spindle poles. In these systems, damaging central spindle microtubules by laser ablation caused an immediate and complete cessation of chromosome motion, suggesting a strong coupling between central spindle microtubules and chromosomes. Electron tomographic reconstruction revealed that the analyzed anaphase spindles all contain microtubules with both ends between segregating chromosomes. Our results provide new dynamical, functional, and ultrastructural characterizations of central spindle microtubules during chromosome segregation in diverse spindles, and suggest that central spindle microtubules and chromosomes are strongly coupled in anaphase.

scholarly journals Chromosome structural anomalies due to aberrant spindle forces exerted at gene editing sites in meiosis

2018 ◽  
Vol 217 (10) ◽  
pp. 3416-3430 ◽  
Author(s):  
Marion Manil-Ségalen ◽  
Małgorzata Łuksza ◽  
Joanne Kanaan ◽  
Véronique Marthiens ◽  
Simon I.R. Lane ◽  
...  
Keyword(s):  
Chromosome Segregation ◽  
Gene Editing ◽  
Spindle Assembly ◽  
Chromosome Breakage ◽  
Microtubule Organizing Center ◽  
Spindle Poles ◽  
Organizing Center ◽  
Meiotic Spindles ◽  
The Impact ◽  
Structural Anomalies

Mouse female meiotic spindles assemble from acentriolar microtubule-organizing centers (aMTOCs) that fragment into discrete foci. These are further sorted and clustered to form spindle poles, thus providing balanced forces for faithful chromosome segregation. To assess the impact of aMTOC biogenesis on spindle assembly, we genetically induced their precocious fragmentation in mouse oocytes using conditional overexpression of Plk4, a master microtubule-organizing center regulator. Excessive microtubule nucleation from these fragmented aMTOCs accelerated spindle assembly dynamics. Prematurely formed spindles promoted the breakage of three different fragilized bivalents, generated by the presence of recombined Lox P sites. Reducing the density of microtubules significantly diminished the extent of chromosome breakage. Thus, improper spindle forces can lead to widely described yet unexplained chromosomal structural anomalies with disruptive consequences on the ability of the gamete to transmit an uncorrupted genome.

scholarly journals Katanin controls mitotic and meiotic spindle length

2006 ◽  
Vol 175 (6) ◽  
pp. 881-891 ◽  
Author(s):  
Karen McNally ◽  
Anjon Audhya ◽  
Karen Oegema ◽  
Francis J. McNally
Keyword(s):  
Chromosome Segregation ◽  
Eukaryotic Cell ◽  
Meiotic Spindle ◽  
Mitotic Spindles ◽  
Wild Type ◽  
Female Meiosis ◽  
C Elegans ◽  
Microtubule Severing ◽  
Spindle Poles ◽  
Type C

Accurate control of spindle length is a conserved feature of eukaryotic cell division. Lengthening of mitotic spindles contributes to chromosome segregation and cytokinesis during mitosis in animals and fungi. In contrast, spindle shortening may contribute to conservation of egg cytoplasm during female meiosis. Katanin is a microtubule-severing enzyme that is concentrated at mitotic and meiotic spindle poles in animals. We show that inhibition of katanin slows the rate of spindle shortening in nocodazole-treated mammalian fibroblasts and in untreated Caenorhabditis elegans meiotic embryos. Wild-type C. elegans meiotic spindle shortening proceeds through an early katanin-independent phase marked by increasing microtubule density and a second, katanin-dependent phase that occurs after microtubule density stops increasing. In addition, double-mutant analysis indicated that γ-tubulin–dependent nucleation and microtubule severing may provide redundant mechanisms for increasing microtubule number during the early stages of meiotic spindle assembly.

scholarly journals Male meiotic spindle features that efficiently segregate paired and lagging chromosomes

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Gunar Fabig ◽  
Robert Kiewisz ◽  
Norbert Lindow ◽  
James A Powers ◽  
Vanessa Cota ◽  
...  
Keyword(s):  
Chromosome Segregation ◽  
Electron Tomography ◽  
Male Meiosis ◽  
Meiotic Spindle ◽  
Sperm Production ◽  
C Elegans ◽  
Unpaired Chromosome ◽  
Meiotic Spindles ◽  
Anaphase B ◽  
Distance Change

Chromosome segregation during male meiosis is tailored to rapidly generate multitudes of sperm. Little is known about mechanisms that efficiently partition chromosomes to produce sperm. Using live imaging and tomographic reconstructions of spermatocyte meiotic spindles in Caenorhabditis elegans, we find the lagging X chromosome, a distinctive feature of anaphase I in C. elegans males, is due to lack of chromosome pairing. The unpaired chromosome remains tethered to centrosomes by lengthening kinetochore microtubules, which are under tension, suggesting that a ‘tug of war’ reliably resolves lagging. We find spermatocytes exhibit simultaneous pole-to-chromosome shortening (anaphase A) and pole-to-pole elongation (anaphase B). Electron tomography unexpectedly revealed spermatocyte anaphase A does not stem solely from kinetochore microtubule shortening. Instead, movement of autosomes is largely driven by distance change between chromosomes, microtubules, and centrosomes upon tension release during anaphase. Overall, we define novel features that segregate both lagging and paired chromosomes for optimal sperm production.

scholarly journals Mechanical heterogeneity and roles of parallel microtubule arrays in governing meiotic spindle length

2018 ◽  
Author(s):  
Jun Takagi ◽  
Yuta Shimamoto
Keyword(s):  
Spindle Pole ◽  
Spindle Motor ◽  
Meiotic Spindle ◽  
Functional Stability ◽  
Mechanical Heterogeneity ◽  
Spindle Poles ◽  
Egg Extract ◽  
Complex Architecture ◽  
Segregation Of Chromosomes

AbstractMetaphase spindles are arrays of microtubules whose architecture provides the mechanism for regulated force generation required for proper segregation of chromosomes during cell division. Whereas long-standing models are based on continuous antiparallel microtubule arrays connecting two spindle poles and overlapping at the equator, spindles typically possess a more complex architecture with randomly arranged short filaments. How these heterogeneous multifilament arrays generate and respond to forces has been mysterious, as it has not been possible to directly measure and perturb spindle force while observing relevant filament motility. Here, we combined microneedle-based quantitative micromanipulation with high-resolution microtubule tracking of Xenopus egg extract spindles to simultaneously examine the force and individual filament motility in situ. We found that the microtubule arrays at the middle of the spindle half are considerably weak and fluid-like, being more adaptable to perturbing forces as compared to those near the pole and the equator. We also found that a force altering spindle length induces filament translocation nearer the spindle pole, where parallel microtubules predominate, while maintaining equatorial antiparallel filaments. Molecular perturbations suggested that the distinct mechanical heterogeneity of the spindle emerges from activities of kinesin-5 and dynein, two key spindle motor proteins. Together, our data establish a link between spindle architecture and mechanics, and highlight the importance of parallel microtubule arrays in maintaining its structural and functional stability.

scholarly journals Katanin maintains meiotic metaphase chromosome alignment and spindle structure in vivo and has multiple effects on microtubules in vitro

2014 ◽  
Vol 25 (7) ◽  
pp. 1037-1049 ◽  
Author(s):  
Karen McNally ◽  
Evan Berg ◽  
Daniel B. Cortes ◽  
Veronica Hernandez ◽  
Paul E. Mains ◽  
...  
Keyword(s):  
Point Mutations ◽  
Metaphase Plate ◽  
Meiotic Spindle ◽  
Loss Of Function ◽  
Microtubule Organization ◽  
Bipolar Structure ◽  
Microtubule Severing ◽  
Meiotic Spindles

Assembly of Caenorhabditis elegans female meiotic spindles requires both MEI-1 and MEI-2 subunits of the microtubule-severing ATPase katanin. Strong loss-of-function mutants assemble apolar intersecting microtubule arrays, whereas weaker mutants assemble bipolar meiotic spindles that are longer than wild type. To determine whether katanin is also required for spindle maintenance, we monitored metaphase I spindles after a fast-acting mei-1(ts) mutant was shifted to a nonpermissive temperature. Within 4 min of temperature shift, bivalents moved off the metaphase plate, and microtubule bundles within the spindle lengthened and developed a high degree of curvature. Spindles eventually lost bipolar structure. Immunofluorescence of embryos fixed at increasing temperature indicated that MEI-1 was lost from spindle microtubules before loss of ASPM-1, indicating that MEI-1 and ASPM-1 act independently at spindle poles. We quantified the microtubule-severing activity of purified MEI-1/MEI-2 complexes corresponding to six different point mutations and found a linear relationship between microtubule disassembly rate and meiotic spindle length. Previous work showed that katanin is required for severing at points where two microtubules intersect in vivo. We show that purified MEI-1/MEI-2 complexes preferentially sever at intersections between two microtubules and directly bundle microtubules in vitro. These activities could promote parallel/antiparallel microtubule organization in meiotic spindles.

scholarly journals TPX2 levels modulate meiotic spindle size and architecture in Xenopus egg extracts

2014 ◽  
Vol 206 (3) ◽  
pp. 385-393 ◽  
Author(s):  
Kara J. Helmke ◽  
Rebecca Heald
Keyword(s):  
Parallel Architecture ◽  
Cell Types ◽  
Meiotic Spindle ◽  
Spindle Poles ◽  
Frog Species ◽  
Two Factors ◽  
Egg Extracts ◽  
Xenopus Egg ◽  
Meiotic Spindles ◽  
Xenopus Egg Extracts

The spindle segregates chromosomes in dividing eukaryotic cells, and its assembly pathway and morphology vary across organisms and cell types. We investigated mechanisms underlying differences between meiotic spindles formed in egg extracts of two frog species. Small Xenopus tropicalis spindles resisted inhibition of two factors essential for assembly of the larger Xenopus laevis spindles: RanGTP, which functions in chromatin-driven spindle assembly, and the kinesin-5 motor Eg5, which drives antiparallel microtubule (MT) sliding. This suggested a role for the MT-associated protein TPX2 (targeting factor for Xenopus kinesin-like protein 2), which is regulated by Ran and binds Eg5. Indeed, TPX2 was threefold more abundant in X. tropicalis extracts, and elevated TPX2 levels in X. laevis extracts reduced spindle length and sensitivity to Ran and Eg5 inhibition. Higher TPX2 levels recruited Eg5 to the poles, where MT density increased. We propose that TPX2 levels modulate spindle architecture through Eg5, partitioning MTs between a tiled, antiparallel array that promotes spindle expansion and a cross-linked, parallel architecture that concentrates MTs at spindle poles.

Knockdown of UCHL5IP causes abnormalities in γ-tubulin localisation, spindle organisation and chromosome alignment in mouse oocyte meiotic maturation

2013 ◽  
Vol 25 (3) ◽  
pp. 495 ◽  
Author(s):  
Ya-Peng Wang ◽  
Shu-Tao Qi ◽  
Yanchang Wei ◽  
Zhao-Jia Ge ◽  
Lei Chen ◽  
...  
Keyword(s):  
Chromosome Segregation ◽  
Oocyte Maturation ◽  
Protein Level ◽  
Mouse Oocyte ◽  
Meiotic Spindle ◽  
Meiotic Maturation ◽  
Spindle Formation ◽  
Spindle Poles ◽  
Chromosome Alignment ◽  
Oocyte Meiotic Maturation

UCHL5IP is one of the subunits of the haus complex, which is important for microtubule generation, spindle bipolarity and accurate chromosome segregation in Drosophila and human mitotic cells. In this study, the expression and localisation of UCHL5IP were explored, as well as its functions in mouse oocyte meiotic maturation. The results showed that the UCHL5IP protein level was consistent during oocyte maturation and it was localised to the meiotic spindle in MI and MII stages. Knockdown of UCHL5IP led to spindle defects, chromosome misalignment and disruption of γ-tubulin localisation in the spindle poles. These results suggest that UCHL5IP plays critical roles in spindle formation during mouse oocyte meiotic maturation.

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