Cell cycle-dependent specific positioning and clustering of centromeres and telomeres in fission yeast.

Fluorescence in situ hybridization (FISH) shows that fission yeast centromeres and telomeres make up specific spatial arrangements in the nucleus. Their positioning and clustering are cell cycle regulated. In G2, centromeres cluster adjacent to the spindle pole body (SPB), while in mitosis, their association with each other and with the SPB is disrupted. Similarly, telomeres cluster at the nuclear periphery in G2 and their associations are disrupted in mitosis. Mitotic centromeres interact with the spindle. They remain undivided until the spindle reaches a critical length, then separate and move towards the poles. This demonstrated, for the first time, that anaphase A occurs in fission yeast. The mode of anaphase A and B is similar to that of higher eukaryotes. In nda3 and cut7 mutants defective in tubulin of a kinesin-related motor, cells are blocked in early stages of mitosis due to the absence of the spindle, and centromeres dissociate but remain close to the SPB, whereas in a metaphase-arrested nuc2 mutant, they reside at the middle of the spindle. FISH is therefore a powerful tool for analyzing mitotic chromosome movement and disjunction using various mutants. Surprisingly, in top2 defective in DNA topoisomerase II, while most chromatid DNAs remain undivided, sister centromeres are separated. Significance of this finding is discussed. In contrast, most chromatid DNAs are separated but telomeric DNAs are not in cut1 mutant. In cut1, the dependence of SPB duplication on the completion of mitosis is abolished. In crm1 mutant cells defective in higher-order chromosome organization, the interphase arrangements of centromeres and telomeres are disrupted.

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Asymmetry of the spindle pole bodies and spg1p GAP segregation during mitosis in fission yeast

Journal of Cell Science ◽

10.1242/jcs.112.14.2313 ◽

1999 ◽

Vol 112 (14) ◽

pp. 2313-2321 ◽

Cited By ~ 22

Author(s):

L. Cerutti ◽

V. Simanis

Keyword(s):

Cell Cycle ◽

Fission Yeast ◽

Spindle Pole Body ◽

Mitotic Cell ◽

Spindle Pole ◽

The Other ◽

Septum Formation ◽

Spindle Pole Bodies ◽

Interphase Cells ◽

Early Mitosis

In the fission yeast Schizosaccharomyces pombe, the onset of septum formation is induced by a signal transduction network involving several protein kinases and a GTPase switch. One of the roles of the spg1p GTPase is to localise the cdc7p protein kinase to the poles of the mitotic spindle, from where the onset of septation is thought to be signalled at the end of mitosis. Immunofluorescence studies have shown that cdc7p is located on both spindle pole bodies early in mitosis, but only on one during the later stages of anaphase. This is mediated by inactivation of spg1p on one pole before the other. The GAP for spg1p is a complex of two proteins, cdc16p and byr4p. Localisation of cdc16p and byr4p by indirect immunofluorescence during the mitotic cell cycle showed that both proteins are present on the spindle pole body in interphase cells. During mitosis, byr4p is seen first on both poles of the spindle, then on only one. This occurs prior to cdc7p becoming asymmetric. In contrast, the signal due to cdc16p decreases to a low level during early mitosis, before being seen strongly on the same pole as byr4p. Double staining indicates that this is the opposite pole to that which retains cdc7p in late anaphase. Examination of the effect of inactivating cdc16p at various stages of the cell cycle suggests that cdc16p, together with cdc2p plays a role in restraining septum formation during interphase. The asymmetric inactivation of spg1p is mediated by recruitment of the cdc16p-byr4p GAP to one of the poles of the spindle before the other, and the asymmetry of the spindle pole bodies may be established early during mitosis. Moreover, the spindle pole bodies appear to be non-equivalent even after division has been completed.

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Cdk1 promotes cytokinesis in fission yeast through activation of the septation initiation network

Molecular Biology of the Cell ◽

10.1091/mbc.e14-04-0936 ◽

2014 ◽

Vol 25 (15) ◽

pp. 2250-2259 ◽

Cited By ~ 6

Author(s):

Nicole Rachfall ◽

Alyssa E. Johnson ◽

Sapna Mehta ◽

Jun-Song Chen ◽

Kathleen L. Gould

Keyword(s):

Cell Cycle ◽

Spindle Pole Body ◽

Complete Removal ◽

Spindle Pole ◽

Cyclin Dependent Kinase ◽

Gtpase Activating Protein ◽

Pole Body ◽

Kinase Cascade ◽

Septation Initiation Network ◽

Cycle Dependent

In Schizosaccharomyces pombe, late mitotic events are coordinated with cytokinesis by the septation initiation network (SIN), an essential spindle pole body (SPB)–associated kinase cascade, which controls the formation, maintenance, and constriction of the cytokinetic ring. It is not fully understood how SIN initiation is temporally regulated, but it depends on the activation of the GTPase Spg1, which is inhibited during interphase by the essential bipartite GTPase-activating protein Byr4-Cdc16. Cells are particularly sensitive to the modulation of Byr4, which undergoes cell cycle–dependent phosphorylation presumed to regulate its function. Polo-like kinase, which promotes SIN activation, is partially responsible for Byr4 phosphorylation. Here we show that Byr4 is also controlled by cyclin-dependent kinase (Cdk1)–mediated phosphorylation. A Cdk1 nonphosphorylatable Byr4 phosphomutant displays severe cell division defects, including the formation of elongated, multinucleate cells, failure to maintain the cytokinetic ring, and compromised SPB association of the SIN kinase Cdc7. Our analyses show that Cdk1-mediated phosphoregulation of Byr4 facilitates complete removal of Byr4 from metaphase SPBs in concert with Plo1, revealing an unexpected role for Cdk1 in promoting cytokinesis through activation of the SIN pathway.

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Cytoplasmic microtubular system implicated in de novo formation of a Rabl-like orientation of chromosomes in fission yeast

Journal of Cell Science ◽

10.1242/jcs.114.13.2427 ◽

2001 ◽

Vol 114 (13) ◽

pp. 2427-2435 ◽

Cited By ~ 2

Author(s):

Bunshiro Goto ◽

Koei Okazaki ◽

Osami Niwa

Keyword(s):

Fission Yeast ◽

De Novo ◽

Nuclear Periphery ◽

Spindle Pole Body ◽

Spindle Pole ◽

Limited Area ◽

Cytoplasmic Microtubule ◽

Random Arrangement ◽

Triplex Structure ◽

Microtubular System

Chromosomes are not packed randomly in the nucleus. The Rabl orientation is an example of the non-random arrangement of chromosomes, centromeres are grouped in a limited area near the nuclear periphery and telomeres are located apart from centromeres. This orientation is established during mitosis and maintained through subsequent interphase in a range of species. We report that a Rabl-like configuration can be formed de novo without a preceding mitosis during the transition from the sexual phase to the vegetative phase of the life cycle in fission yeast. In this process, each of the dispersed centromeres is often associated with a novel Sad1-containing body that is contacting a cytoplasmic microtubule laterally (Sad1 is a component of the spindle pole body (SPB)). The Sad1-containing body was colocalized with other known SPB components, Kms1 and Spo15 but not with Cut12, indicating that it represents a novel SPB-related complex. The existence of the triplex structure (centromere-microtubule-Sad1 body) suggests that the clustering of centromeres is controlled by a cytoplasmic microtubular system. Accordingly, when microtubules are destabilized, clustering is markedly reduced.

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Spc98p Directs the Yeast γ-Tubulin Complex into the Nucleus and Is Subject to Cell Cycle-dependent Phosphorylation on the Nuclear Side of the Spindle Pole Body

Molecular Biology of the Cell ◽

10.1091/mbc.9.4.775 ◽

1998 ◽

Vol 9 (4) ◽

pp. 775-793 ◽

Cited By ~ 63

Author(s):

Gislene Pereira ◽

Michael Knop ◽

Elmar Schiebel

Keyword(s):

Cell Cycle ◽

Spindle Pole Body ◽

Mitotic Checkpoint ◽

Spindle Pole ◽

Nuclear Localization Sequence ◽

Checkpoint Control ◽

Yeast Saccharomyces Cerevisiae ◽

Pole Body ◽

Cycle Dependent ◽

Cytoplasmic Side

In the yeast Saccharomyces cerevisiae, microtubules are organized by the spindle pole body (SPB), which is embedded in the nuclear envelope. Microtubule organization requires the γ-tubulin complex containing the γ-tubulin Tub4p, Spc98p, and Spc97p. The Tub4p complex is associated with cytoplasmic and nuclear substructures of the SPB, which organize the cytoplasmic and nuclear microtubules. Here we present evidence that the Tub4p complex assembles in the cytoplasm and then either binds to the cytoplasmic side of the SPB or is imported into the nucleus followed by binding to the nuclear side of the SPB. Nuclear import of the Tub4p complex is mediated by the essential nuclear localization sequence of Spc98p. Our studies also indicate that Spc98p in the Tub4p complex is phosphorylated at the nuclear, but not at the cytoplasmic, side of the SPB. This phosphorylation is cell cycle dependent and occurs after SPB duplication and nucleation of microtubules by the new SPB and therefore may have a role in mitotic spindle function. In addition, activation of the mitotic checkpoint stimulates Spc98p phosphorylation. The kinase Mps1p, which functions in SPB duplication and mitotic checkpoint control, seems to be involved in Spc98p phosphorylation. Our results also suggest that the nuclear and cytoplasmic Tub4p complexes are regulated differently.

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Quantifying Tubulin Concentration and Microtubule Number Throughout the Fission Yeast Cell Cycle

Biomolecules ◽

10.3390/biom9030086 ◽

2019 ◽

Vol 9 (3) ◽

pp. 86 ◽

Cited By ~ 4

Author(s):

Isabelle Loiodice ◽

Marcel Janson ◽

Penny Tavormina ◽

Sebastien Schaub ◽

Divya Bhatt ◽

...

Keyword(s):

Cell Cycle ◽

Fission Yeast ◽

Genetic Model ◽

Spindle Pole Body ◽

Model Organism ◽

Spindle Pole ◽

Living Cells ◽

Fluorescent Imaging ◽

Mother And Daughter ◽

In The Wild

The fission yeast Schizosaccharomyces pombe serves as a good genetic model organism for the molecular dissection of the microtubule (MT) cytoskeleton. However, analysis of the number and distribution of individual MTs throughout the cell cycle, particularly during mitosis, in living cells is still lacking, making quantitative modelling imprecise. We use quantitative fluorescent imaging and analysis to measure the changes in tubulin concentration and MT number and distribution throughout the cell cycle at a single MT resolution in living cells. In the wild-type cell, both mother and daughter spindle pole body (SPB) nucleate a maximum of 23 ± 6 MTs at the onset of mitosis, which decreases to a minimum of 4 ± 1 MTs at spindle break down. Interphase MT bundles, astral MT bundles, and the post anaphase array (PAA) microtubules are composed primarily of 1 ± 1 individual MT along their lengths. We measure the cellular concentration of αβ-tubulin subunits to be ~5 µM throughout the cell cycle, of which one-third is in polymer form during interphase and one-quarter is in polymer form during mitosis. This analysis provides a definitive characterization of αβ-tubulin concentration and MT number and distribution in fission yeast and establishes a foundation for future quantitative comparison of mutants defective in MTs.

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Regulation of spindle pole body assembly and cytokinesis by the centrin-binding protein Sfi1 in fission yeast

Molecular Biology of the Cell ◽

10.1091/mbc.e13-11-0699 ◽

2014 ◽

Vol 25 (18) ◽

pp. 2735-2749 ◽

Cited By ~ 22

Author(s):

I-Ju Lee ◽

Ning Wang ◽

Wen Hu ◽

Kersey Schott ◽

Jürg Bähler ◽

...

Keyword(s):

Cell Cycle ◽

Cell Division ◽

Fission Yeast ◽

Budding Yeast ◽

Binding Protein ◽

Spindle Pole Body ◽

Spindle Pole ◽

Centrosome Duplication ◽

Pole Body ◽

The Individual

Centrosomes play critical roles in the cell division cycle and ciliogenesis. Sfi1 is a centrin-binding protein conserved from yeast to humans. Budding yeast Sfi1 is essential for the initiation of spindle pole body (SPB; yeast centrosome) duplication. However, the recruitment and partitioning of Sfi1 to centrosomal structures have never been fully investigated in any organism, and the presumed importance of the conserved tryptophans in the internal repeats of Sfi1 remains untested. Here we report that in fission yeast, instead of doubling abruptly at the initiation of SPB duplication and remaining at a constant level thereafter, Sfi1 is gradually recruited to SPBs throughout the cell cycle. Like an sfi1Δ mutant, a Trp-to-Arg mutant (sfi1-M46) forms monopolar spindles and exhibits mitosis and cytokinesis defects. Sfi1-M46 protein associates preferentially with one of the two daughter SPBs during mitosis, resulting in a failure of new SPB assembly in the SPB receiving insufficient Sfi1. Although all five conserved tryptophans tested are involved in Sfi1 partitioning, the importance of the individual repeats in Sfi1 differs. In summary, our results reveal a link between the conserved tryptophans and Sfi1 partitioning and suggest a revision of the model for SPB assembly.

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The Phosphorylation State of the 110 kDa Component of the Yeast Spindle Pole Body Shows Cell Cycle Dependent Regulation

Biochemical and Biophysical Research Communications ◽

10.1006/bbrc.1996.0728 ◽

1996 ◽

Vol 222 (2) ◽

pp. 236-242 ◽

Cited By ~ 11

Author(s):

Douglas A. Stirling ◽

Michael J.R. Stark

Keyword(s):

Cell Cycle ◽

Spindle Pole Body ◽

Spindle Pole ◽

Phosphorylation State ◽

Pole Body ◽

Cycle Dependent

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In vivo localisation of fission yeast cyclin-dependent kinase cdc2p and cyclin B cdc13p during mitosis and meiosis

Journal of Cell Science ◽

10.1242/jcs.114.14.2627 ◽

2001 ◽

Vol 114 (14) ◽

pp. 2627-2640 ◽

Cited By ~ 1

Author(s):

Anabelle Decottignies ◽

Patrick Zarzov ◽

Paul Nurse

Keyword(s):

Fission Yeast ◽

Fluorescent Protein ◽

Yellow Fluorescent Protein ◽

Nuclear Periphery ◽

Spindle Pole Body ◽

Spindle Pole ◽

Cyclin Dependent Kinase ◽

Spindle Pole Bodies ◽

Mitosis And Meiosis

We investigated the in vivo localisation of fission yeast cyclin-dependent kinase cdc2p during mitosis and meiosis. Fusion to yellow fluorescent protein (YFP) revealed that cdc2-YFP is present in the cytoplasm at all stages of the cell cycle. Nuclear cdc2-YFP fluorescence oscillates with that of cdc13-YFP cyclin. At G1/S, at least one of cdc13p, cig1p or cig2p B-type cyclins is required for the accumulation of cdc2-YFP into the nucleus. Cdc2-YFP and cdc13-YFP are highly enriched on the spindle pole body of cells in late G2 or arrested at S phase. Both accumulate on the spindle pole bodies and the spindle in prophase and metaphase independently of the microtubule-associated protein dis1p. In anaphase, the cdc2p/cdc13p complex leaves the spindle prior to sister chromatid separation, and cdc13-YFP is enriched at the nuclear periphery before fluorescence disappears. If cdc13p cannot be recognized by the anaphase-promoting complex, cdc2-YFP and cdc13-YFP remain associated with the spindle. In mating cells, cdc2-YFP enters the nucleus as soon as the cells undergo fusion. During karyogamy and meiotic prophase, cdc2-YFP is highly enriched on the centromeres. In meiosis I, association of cdc2-YFP with the spindle and the spindle pole bodies shows differences to mitotic cells, suggesting different mechanisms of spindle formation. This study suggests that changes in cdc2p localisation are important for both mitosis and meiosis regulation.

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Cell Cycle-Dependent Control and Roles of DNA Topoisomerase II

Genes ◽

10.3390/genes10110859 ◽

2019 ◽

Vol 10 (11) ◽

pp. 859 ◽

Cited By ~ 14

Author(s):

Joyce H. Lee ◽

James M. Berger

Keyword(s):

Cell Cycle ◽

Cell Cycle Progression ◽

Topoisomerase Ii ◽

Type Ii ◽

Dna Topoisomerase Ii ◽

Dna Topoisomerase ◽

Cycle Progression ◽

Double Stranded Dna ◽

Topo Ii ◽

Cycle Dependent

Type II topoisomerases are ubiquitous enzymes in all branches of life that can alter DNA superhelicity and unlink double-stranded DNA segments during processes such as replication and transcription. In cells, type II topoisomerases are particularly useful for their ability to disentangle newly-replicated sister chromosomes. Growing lines of evidence indicate that eukaryotic topoisomerase II (topo II) activity is monitored and regulated throughout the cell cycle. Here, we discuss the various roles of topo II throughout the cell cycle, as well as mechanisms that have been found to govern and/or respond to topo II function and dysfunction. Knowledge of how topo II activity is controlled during cell cycle progression is important for understanding how its misregulation can contribute to genetic instability and how modulatory pathways may be exploited to advance chemotherapeutic development.

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