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 Cohesins Determine the Attachment Manner of Kinetochores to Spindle Microtubules at Meiosis I in Fission Yeast

2003 ◽  
Vol 23 (11) ◽  
pp. 3965-3973 ◽  
Author(s):  
Shihori Yokobayashi ◽  
Masayuki Yamamoto ◽  
Yoshinori Watanabe
Keyword(s):  
Fission Yeast ◽  
Chromosome Segregation ◽  
Sister Chromatid ◽  
Specific Requirement ◽  
Spindle Poles ◽  
Chromatid Cohesion ◽  
Spindle Microtubules ◽  
Yeast Meiosis ◽  
Meiosis I ◽  
Random Division

ABSTRACT During mitosis, sister kinetochores attach to microtubules that extend to opposite spindle poles (bipolar attachment) and pull the chromatids apart at anaphase (equational segregation). A multisubunit complex called cohesin, including Rad21/Scc1, plays a crucial role in sister chromatid cohesion and equational segregation at mitosis. Meiosis I differs from mitosis in having a reductional pattern of chromosome segregation, in which sister kinetochores are attached to the same spindle (monopolar attachment). During meiosis, Rad21/Scc1 is largely replaced by its meiotic counterpart, Rec8. If Rec8 is inactivated in fission yeast, meiosis I is shifted from reductional to equational division. However, the reason rec8Δ cells undergo equational rather than random division has not been clarified; therefore, it has been unclear whether equational segregation is due to a loss of cohesin in general or to a loss of a specific requirement for Rec8. We report here that the equational segregation at meiosis I depends on substitutive Rad21, which relocates to the centromeres if Rec8 is absent. Moreover, we demonstrate that even if sufficient amounts of Rad21 are transferred to the centromeres at meiosis I, thereby establishing cohesion at the centromeres, rec8Δ cells never recover monopolar attachment but instead secure bipolar attachment. Thus, Rec8 and Rad21 define monopolar and bipolar attachment, respectively, at meiosis I. We conclude that cohesin is a crucial determinant of the attachment manner of kinetochores to the spindle microtubules at meiosis I in fission yeast.

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 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 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 CENP-32 is required to maintain centrosomal dominance in bipolar spindle assembly

2015 ◽  
Vol 26 (7) ◽  
pp. 1225-1237 ◽  
Author(s):  
Shinya Ohta ◽  
Laura Wood ◽  
Iyo Toramoto ◽  
Ken-Ichi Yagyu ◽  
Tatsuo Fukagawa ◽  
...  
Keyword(s):  
Chromosome Segregation ◽  
Additional Data ◽  
Weak Interactions ◽  
Metaphase Plate ◽  
Spindle Pole ◽  
Animal Cells ◽  
Central Spindle ◽  
Spindle Poles ◽  
Spindle Microtubules ◽  
Bipolar Spindles

Centrosomes nucleate spindle formation, direct spindle pole positioning, and are important for proper chromosome segregation during mitosis in most animal cells. We previously reported that centromere protein 32 (CENP-32) is required for centrosome association with spindle poles during metaphase. In this study, we show that CENP-32 depletion seems to release centrosomes from bipolar spindles whose assembly they had previously initiated. Remarkably, the resulting anastral spindles function normally, aligning the chromosomes to a metaphase plate and entering anaphase without detectable interference from the free centrosomes, which appear to behave as free asters in these cells. The free asters, which contain reduced but significant levels of CDK5RAP2, show weak interactions with spindle microtubules but do not seem to make productive attachments to kinetochores. Thus CENP-32 appears to be required for centrosomes to integrate into a fully functional spindle that not only nucleates astral microtubules, but also is able to nucleate and bind to kinetochore and central spindle microtubules. Additional data suggest that NuMA tethers microtubules at the anastral spindle poles and that augmin is required for centrosome detachment after CENP-32 depletion, possibly due to an imbalance of forces within the spindle.

scholarly journals Pivot-and-bond model explains microtubule bundle formation

2017 ◽  
Author(s):  
Marcel Prelogović ◽  
Lora Winters ◽  
Ana Milas ◽  
Iva M. Tolić ◽  
Nenad Pavin
Keyword(s):  
Fission Yeast ◽  
Physical Mechanism ◽  
Spindle Pole ◽  
Yeast Cells ◽  
Cross Linking ◽  
Stable Bundle ◽  
Wild Type ◽  
Microtubule Bundle ◽  
Angular Movement ◽  
Bond Model

ABSTRACTDuring mitosis, bundles of microtubules form a spindle, but the physical mechanism of bundle formation is still not known. Here we show that random angular movement of microtubules around the spindle pole and forces exerted by passive cross-linking proteins are sufficient for the formation of stable microtubule bundles. We test these predictions by experiments in wild-type and ase1Δ fission yeast cells. In conclusion, the angular motion drives the alignment of microtubules, which in turn allows the cross-linking proteins to connect the microtubules into a stable bundle.

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

2016 ◽  
Vol 6 (1) ◽  
Author(s):  
Gheorghe Cojoc ◽  
Ana-Maria Florescu ◽  
Alexander Krull ◽  
Anna H. Klemm ◽  
Nenad Pavin ◽  
...  
Keyword(s):  
Cell Division ◽  
Fission Yeast ◽  
General Feature ◽  
Protein Complexes ◽  
Spindle Pole ◽  
Single Object ◽  
Homologous Chromosomes ◽  
Angular Movement ◽  
Spindle Microtubules ◽  
Meiosis I

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

scholarly journals Interaction between Poly(ADP-ribose) and NuMA Contributes to Mitotic Spindle Pole Assembly

2009 ◽  
Vol 20 (21) ◽  
pp. 4575-4585 ◽  
Author(s):  
Paul Chang ◽  
Margaret Coughlin ◽  
Timothy J. Mitchison
Keyword(s):  
Binding Proteins ◽  
Magnetic Beads ◽  
Spindle Pole ◽  
Covalent Modification ◽  
Mitotic Spindles ◽  
Spindle Poles ◽  
Tankyrase 1 ◽  
Mitotic Spindle Pole

Poly(ADP-ribose) (pADPr), made by PARP-5a/tankyrase-1, localizes to the poles of mitotic spindles and is required for bipolar spindle assembly, but its molecular function in the spindle is poorly understood. To investigate this, we localized pADPr at spindle poles by immuno-EM. We then developed a concentrated mitotic lysate system from HeLa cells to probe spindle pole assembly in vitro. Microtubule asters assembled in response to centrosomes and Ran-GTP in this system. Magnetic beads coated with pADPr, extended from PARP-5a, also triggered aster assembly, suggesting a functional role of the pADPr in spindle pole assembly. We found that PARP-5a is much more active in mitosis than interphase. We used mitotic PARP-5a, self-modified with pADPr chains, to capture mitosis-specific pADPr-binding proteins. Candidate binding proteins included the spindle pole protein NuMA previously shown to bind to PARP-5a directly. The rod domain of NuMA, expressed in bacteria, bound directly to pADPr. We propose that pADPr provides a dynamic cross-linking function at spindle poles by extending from covalent modification sites on PARP-5a and NuMA and binding noncovalently to NuMA and that this function helps promote assembly of exactly two poles.

scholarly journals Amino acids whose intracellular levels change most during aging alter chronological lifespan of fission yeast

2020 ◽  
Author(s):  
Charalampos Rallis ◽  
Michael Mülleder ◽  
Graeme Smith ◽  
Yan Zi Au ◽  
Markus Ralser ◽  
...  
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Fission Yeast ◽  
Yeast Cells ◽  
Total Amino Acid ◽  
Wild Type ◽  
Chronological Lifespan ◽  
Biomarkers Of Aging ◽  
Concentration Changes ◽  
Amino Acid Levels

AbstractAmino acid deprivation or supplementation can affect cellular and organismal lifespan, but we know little about the role of concentration changes in free, intracellular amino acids during aging. Here, we determine free amino-acid levels during chronological aging of non-dividing fission yeast cells. We compare wild-type with long-lived mutant cells that lack the Pka1 protein of the protein kinase A signalling pathway. In wild-type cells, total amino-acid levels decrease during aging, but much less so in pka1 mutants. Two amino acids strongly change as a function of age: glutamine decreases, especially in wild-type cells, while aspartate increases, especially in pka1 mutants. Supplementation of glutamine is sufficient to extend the chronological lifespan of wild-type but not of pka1Δ cells. Supplementation of aspartate, on the other hand, shortens the lifespan of pka1Δ but not of wild-type cells. Our results raise the possibility that certain amino acids are biomarkers of aging, and their concentrations during aging can promote or limit cellular lifespan.

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