scholarly journals Nek2A kinase stimulates centrosome disjunction and is required for formation of bipolar mitotic spindles

2003 ◽  
Vol 14 (7) ◽  
pp. 2876-2889 ◽  
Author(s):  
Alison J. Faragher ◽  
Andrew M. Fry
Keyword(s):  
Chromosome Segregation ◽  
Direct Evidence ◽  
Coiled Coil ◽  
Mitotic Spindles ◽  
Wild Type ◽  
Spindle Poles ◽  
Mother And Daughter ◽  
Correct Execution ◽  
A Cell ◽  
Daughter Centriole

Nek2A is a cell cycle-regulated kinase of the never in mitosis A (NIMA) family that is highly enriched at the centrosome. One model for Nek2A function proposes that it regulates cohesion between the mother and daughter centriole through phosphorylation of C-Nap1, a large coiled-coil protein that localizes to centriolar ends. Phosphorylation of C-Nap1 at the G2/M transition may trigger its displacement from centrioles, promoting their separation and subsequent bipolar spindle formation. To test this model, we generated tetracycline-inducible cell lines overexpressing wild-type and kinase-dead versions of Nek2A. Live cell imaging revealed that active Nek2A stimulates the sustained splitting of interphase centrioles indicative of loss of cohesion. However, this splitting is accompanied by only a partial reduction in centriolar C-Nap1. Strikingly, induction of kinase-dead Nek2A led to formation of monopolar spindles with unseparated spindle poles that lack C-Nap1. Furthermore, kinase-dead Nek2A interfered with chromosome segregation and cytokinesis and led to an overall change in the DNA content of the cell population. These results provide the first direct evidence in human cells that Nek2A function is required for the correct execution of mitosis, most likely through promotion of centrosome disjunction. However, they suggest that loss of centriole cohesion and C-Nap1 displacement may be distinct mitotic events.

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 Mitotic Chromosome Biorientation in Fission Yeast Is Enhanced by Dynein and a Minus-end–directed, Kinesin-like Protein

2007 ◽  
Vol 18 (6) ◽  
pp. 2216-2225 ◽  
Author(s):  
Ekaterina L. Grishchuk ◽  
Ilia S. Spiridonov ◽  
J. Richard McIntosh
Keyword(s):  
Fission Yeast ◽  
Chromosome Segregation ◽  
Mitotic Chromosome ◽  
Spindle Pole ◽  
Ultrastructural Analysis ◽  
Yeast Cells ◽  
Mitotic Spindles ◽  
Wild Type ◽  
Spindle Poles ◽  
Spindle Microtubules

Chromosome biorientation, the attachment of sister kinetochores to sister spindle poles, is vitally important for accurate chromosome segregation. We have studied this process by following the congression of pole-proximal kinetochores and their subsequent anaphase segregation in fission yeast cells that carry deletions in any or all of this organism's minus end–directed, microtubule-dependent motors: two related kinesin 14s (Pkl1p and Klp2p) and dynein. None of these deletions abolished biorientation, but fewer chromosomes segregated normally without Pkl1p, and to a lesser degree without dynein, than in wild-type cells. In the absence of Pkl1p, which normally localizes to the spindle and its poles, the checkpoint that monitors chromosome biorientation was defective, leading to frequent precocious anaphase. Ultrastructural analysis of mutant mitotic spindles suggests that Pkl1p contributes to error-free biorientation by promoting normal spindle pole organization, whereas dynein helps to anchor a focused bundle of spindle microtubules at the pole.

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 Mars promotes dTACC dephosphorylation on mitotic spindles to ensure spindle stability

2008 ◽  
Vol 182 (1) ◽  
pp. 27-33 ◽  
Author(s):  
Shengjiang Tan ◽  
Ekaterina Lyulcheva ◽  
Jon Dean ◽  
Daimark Bennett
Keyword(s):  
Chromosome Segregation ◽  
Coiled Coil ◽  
Protein Phosphatase 1 ◽  
Mitotic Spindles ◽  
Developmental Context ◽  
Microtubule Associated Proteins ◽  
Associated Proteins ◽  
Spindle Stability ◽  
Map Function

Microtubule-associated proteins (MAPs) ensure the fidelity of chromosome segregation by controlling microtubule (MT) dynamics and mitotic spindle stability. However, many aspects of MAP function and regulation are poorly understood in a developmental context. We show that mars, which encodes a Drosophila melanogaster member of the hepatoma up-regulated protein family of MAPs, is essential for MT stabilization during early embryogenesis. As well as associating with spindle MTs in vivo, Mars binds directly to protein phosphatase 1 (PP1) and coimmunoprecipitates from embryo extracts with minispindles and Drosophila transforming acidic coiled-coil (dTACC), two MAPs that function as spindle assembly factors. Disruption of binding to PP1 or loss of mars function results in elevated levels of phosphorylated dTACC on spindles. A nonphosphorylatable form of dTACC is capable of rescuing the lethality of mars mutants. We propose that Mars mediates spatially controlled dephosphorylation of dTACC, which is critical for spindle stabilization.

CDP1, a Novel Saccharomyces cerevisiae Gene Required for Proper Nuclear Division and Chromosome Segregation

Genetics ◽  
1996 ◽  
Vol 144 (4) ◽  
pp. 1387-1397 ◽  
Author(s):  
Pamela K Foreman ◽  
Ronald W Davis
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Chromosome Segregation ◽  
Nuclear Division ◽  
Immunofluorescent Staining ◽  
Temperature Sensitive ◽  
Permissive Temperature ◽  
Mitotic Spindles ◽  
Wild Type ◽  
New Gene ◽  
Mutant Cells

To identify new gene products involved in chromosome segregation, we isolated Saccharomyces cerevisiae mutants that require centromere binding factor I (Cbf1p) for viability. One Cbf1p-dependent mutant (denoted cdp1-1) was selected for further analysis. The CDP1 gene encodes a novel 125-kD protein that is notably similar to previously identified mouse, human and Caenorhabditis elegans proteins. CDP1Δ and cdp1-1 mutant cells were temperature sensitive for growth. At the permissive temperature, cdp1-1 and cdp1Δ cells lost chromosomes at a frequencies ∼20-fold and ∼110-fold higher than wild-type cells, respectively. These mutants also displayed unusually long and numerous bundles of cytoplasmic microtubules as revealed by immunofluorescent staining. In addition, we occasionally observed improperly oriented mitotic spindles, residing entirely within one of the cells. Presumably as a result of undergoing nuclear division with improperly oriented spindles, a large percentage of cdp1 cells had accumulated multiple nuclei. While cdp1 mutant cells were hypersensitive to the microtubule-disrupting compound thiabendazole, they showed increased resistance to the closely related compound benomyl relative to wild-type cells. Taken together, these results suggest that Cdp1p plays a role in governing tubulin dynamics within the cell and may interact directly with microtubules or tubulin.

scholarly journals Tension on kinetochore substrates is insufficient to prevent Aurora-triggered detachment

2018 ◽  
Author(s):  
Anna K. de Regt ◽  
Charles L. Asbury ◽  
Sue Biggins
Keyword(s):  
Chromosome Segregation ◽  
Direct Evidence ◽  
Underlying Mechanism ◽  
Aurora B ◽  
Trial And Error ◽  
Spindle Poles ◽  
Pulling Forces ◽  
Low Tension

Introduction / AbstractChromosome segregation requires large macromolecular structures called kinetochores to attach dynamic microtubules from opposite spindle poles1,2. Attachments are made iteratively, through a trial-and-error process, and proper attachments come under tension from the pulling forces of microtubules3,4. However, if sister kinetochores bind microtubules from the same pole1,2, these defective attachments lack tension and must be destabilized to give another chance for proper attachments to form. This vital error correction process requires Aurora B kinase, which phosphorylates kinetochores lacking tension to reduce their affinity for microtubules5-11. An unresolved question is how Aurora B distinguishes the level of tension on kinetochores. There are conflicting reports on the underlying mechanism12-16, owing in part to the difficulties of manipulating kinetochore tension in vivo and distinguishing kinase from opposing phosphatase activity. To address these issues, we have reconstituted Aurora B-triggered kinetochore detachment in an in vitro optical trapping-based flow assay. Here, we test an outstanding model by determining whether kinetochore tension is sufficient to prevent kinase-triggered detachments. Strikingly, Aurora B detaches kinetochores from microtubules under both high and low tension, providing direct evidence that the kinase does not distinguish correct versus incorrect attachments by recognizing tension-dependent changes in the conformation of its kinetochore substrates.

scholarly journals The centrosomin protein is required for centrosome assembly and function during cleavage in Drosophila

Development ◽  
1999 ◽  
Vol 126 (13) ◽  
pp. 2829-2839 ◽  
Author(s):  
T.L. Megraw ◽  
K. Li ◽  
L.R. Kao ◽  
T.C. Kaufman
Keyword(s):  
Chromosome Segregation ◽  
Late Stage ◽  
Mitotic Spindles ◽  
Cleavage Stage ◽  
Centrosomal Protein ◽  
Spindle Poles ◽  
Gamma Tubulin ◽  
And Function ◽  
Giant Nuclei ◽  
Centrosomal Proteins

Centrosomin is a 150 kDa centrosomal protein of Drosophila melanogaster. To study the function of Centrosomin in the centrosome, we have recovered mutations that are viable but male and female sterile (cnnmfs). We have shown that these alleles (1, 2, 3, 7, 8 and hk21) induce a maternal effect on early embryogenesis and result in the accumulation of low or undetectable levels of Centrosomin in the centrosomes of cleavage stage embryos. Hemizygous cnn females produce embryos that show dramatic defects in chromosome segregation and spindle organization during the syncytial cleavage divisions. In these embryos the syncytial divisions proceed as far as the twelfth cycle, and embryos fail to cellularize. Aberrant divisions and nuclear fusions occur in the early cycles of the nuclear divisions, and become more prominent at later stages. Giant nuclei are seen in late stage embryos. The spindles that form in mutant embryos exhibit multiple anomalies. There is a high occurrence of apparently linked spindles that share poles, indicating that Centrosomin is required for the proper spacing and separation of mitotic spindles within the syncytium. Spindle poles in the mutants contain little or no detectable amounts of the centrosomal proteins CP60, CP190 and (gamma)-tubulin and late stage embryos often do not have astral microtubules at their spindle poles. Spindle morphology and centrosomal composition suggest that the primary cause of these division defects in mutant embryos is centrosomal malfunction. These results suggest that Centrosomin is required for the assembly and function of centrosomes during the syncytial cleavage divisions.

scholarly journals A Kinesin Mutant with an Atypical Bipolar Spindle Undergoes Normal Mitosis

2003 ◽  
Vol 14 (4) ◽  
pp. 1717-1726 ◽  
Author(s):  
A. I. Marcus ◽  
W. Li ◽  
H. Ma ◽  
R. J. Cyr
Keyword(s):  
Chromosome Segregation ◽  
Plant Cells ◽  
Spindle Assembly ◽  
Wild Type Allele ◽  
Wild Type ◽  
Bipolar Spindle ◽  
Type Allele ◽  
Spindle Poles ◽  
Normal Mitosis ◽  
Arabidopsis Cells

Motor proteins have been implicated in various aspects of mitosis, including spindle assembly and chromosome segregation. Here, we show that acentrosomal Arabidopsis cells that are mutant for the kinesin, ATK1, lack microtubule accumulation at the predicted spindle poles during prophase and have reduced spindle bipolarity during prometaphase. Nonetheless, all abnormalities are rectified by anaphase and chromosome segregation appears normal. We conclude that ATK1 is required for normal microtubule accumulation at the spindle poles during prophase and possibly functions in spindle assembly during prometaphase. Because aberrant spindle morphology in these mutants is resolved by anaphase, we postulate that mitotic plant cells contain an error-correcting mechanism. Moreover, ATK1 function seems to be dosage-dependent, because cells containing one wild-type allele take significantly longer to proceed to anaphase as compared with cells containing two wild-type alleles.

Polar organization of gamma-tubulin in acentriolar mitotic spindles of Drosophila melanogaster cells

1995 ◽  
Vol 108 (7) ◽  
pp. 2645-2653 ◽  
Author(s):  
A. Debec ◽  
C. Detraves ◽  
C. Montmory ◽  
G. Geraud ◽  
M. Wright
Keyword(s):  
Spatial Organization ◽  
Polyclonal Antibodies ◽  
Spindle Pole ◽  
Dependent Manner ◽  
Mitotic Spindles ◽  
Wild Type ◽  
A Cell ◽  
Gamma Tubulin ◽  
Cycle Dependent ◽  
Carboxy Terminal

The spindle pole localization of gamma-tubulin was compared in wild type and acentriolar cultured Drosophila cells using polyclonal antibodies specifically raised against the carboxy terminal amino acid sequence of Drosophila gamma-tubulin-1 (-KSEDSRSVTSAGS). During interphase, gamma-tubulin was present in the centrosome of wild type cells and accumulated around this organelle in a cell cycle dependent manner. In contrast, no such structure was observed in acentriolar cells. Wild type mitoses were homogeneously composed of biconical spindles, with two centrosome-associated gamma-tubulin spots at the poles. The mitotic apparatuses observed in the acentriolar cells were heterogeneous; multipolar mitoses, bipolar mitoses with a barrel-shaped spindle and bipolar mitoses with biconical spindles were observed. In acentriolar cells, gamma-tubulin accumulation at mitotic poles was dependent on spindle microtubule integrity. Most acentriolar spindles presented a dispersed gamma-tubulin labeling at the poles. Only well polarized and biconical acentriolar spindles showed a strong gamma-tubulin polar spot. Finally, acentriolar mitotic poles were not organized around true centrosomes. In contrast to wild type cells, in acentriolar cells the Bx63 centrosome-associated antigen was absent and the gamma-tubulin containing material dispersed readily following microtubule disassembly. These observations confirm that gamma-tubulin plays an essential role in the nucleation of microtubules even in the absence of mitotic polar organelles. In addition the data suggest that the mechanisms involved in the bipolarization of wild type and acentriolar mitoses are different, and that centrioles play a role in the spatial organization of the nucleating material containing gamma-tubulin.

Sign in / Sign up

Export Citation Format

Share Document