scholarly journals Role of Polo Kinase and Mid1p in Determining the Site of Cell Division in Fission Yeast

1998 ◽  
Vol 143 (6) ◽  
pp. 1603-1616 ◽  
Author(s):  
Jürg Bähler ◽  
Alexander B. Steever ◽  
Sally Wheatley ◽  
Yu-li Wang ◽  
John R. Pringle ◽  
...  
Keyword(s):  
Fission Yeast ◽  
Spindle Pole ◽  
Temperature Sensitive ◽  
Kinase Gene ◽  
Division Plane ◽  
Daughter Cells ◽  
Polo Kinase ◽  
Spindle Pole Bodies ◽  
Reduced Mobility ◽  
Mitotic Cells

The fission yeast Schizosaccharomyces pombe divides symmetrically using a medial F-actin– based contractile ring to produce equal-sized daughter cells. Mutants defective in two previously described genes, mid1 and pom1, frequently divide asymmetrically. Here we present the identification of three new temperature-sensitive mutants defective in localization of the division plane. All three mutants have mutations in the polo kinase gene, plo1, and show defects very similar to those of mid1 mutants in both the placement and organization of the medial ring. In both cases, ring formation is frequently initiated near the cell poles, indicating that Mid1p and Plo1p function in recruiting medial ring components to the cell center. It has been reported previously that during mitosis Mid1p becomes hyperphosphorylated and relocates from the nucleus to a medial ring. Here we show that Mid1p first forms a diffuse cortical band during spindle formation and then coalesces into a ring before anaphase. Plo1p is required for Mid1p to exit the nucleus and form a ring, and Pom1p is required for proper placement of the Mid1p ring. Upon overexpression of Plo1p, Mid1p exits the nucleus prematurely and displays a reduced mobility on gels similar to that of the hyperphosphorylated form observed previously in mitotic cells. Genetic and two-hybrid analyses suggest that Plo1p and Mid1p act in a common pathway distinct from that involving Pom1p. Plo1p localizes to the spindle pole bodies and spindles of mitotic cells and also to the medial ring at the time of its formation. Taken together, the data indicate that Plo1p plays a role in the positioning of division sites by regulating Mid1p. Given its previously known functions in mitosis and the timing of cytokinesis, Plo1p is thus implicated as a key molecule in the spatial and temporal coordination of cytokinesis with mitosis.

scholarly journals The Meiosis-Specific Sid2p-related Protein Slk1p Regulates Forespore Membrane Assembly in Fission Yeast

2008 ◽  
Vol 19 (9) ◽  
pp. 3676-3690 ◽  
Author(s):  
Hongyan Yan ◽  
Wanzhong Ge ◽  
Ting Gang Chew ◽  
Jeng Yeong Chow ◽  
Dannel McCollum ◽  
...  
Keyword(s):  
Fission Yeast ◽  
Essential Role ◽  
Spindle Pole ◽  
Dependent Manner ◽  
Related Protein ◽  
Membrane Assembly ◽  
Daughter Cells ◽  
Spindle Pole Bodies ◽  
Septation Initiation Network ◽  
Secretory Apparatus

Cytokinesis in all organisms involves the creation of membranous barriers that demarcate individual daughter cells. In fission yeast, a signaling module termed the septation initiation network (SIN) plays an essential role in the assembly of new membranes and cell wall during cytokinesis. In this study, we have characterized Slk1p, a protein-kinase related to the SIN component Sid2p. Slk1p is expressed specifically during meiosis and localizes to the spindle pole bodies (SPBs) during meiosis I and II in a SIN-dependent manner. Slk1p also localizes to the forespore membrane during sporulation. Cells lacking Slk1p display defects associated with sporulation, leading frequently to the formation of asci with smaller and/or fewer spores. The ability of slk1Δ cells to sporulate, albeit inefficiently, is fully abolished upon compromise of function of Sid2p, suggesting that Slk1p and Sid2p play overlapping roles in sporulation. Interestingly, increased expression of the syntaxin Psy1p rescues the sporulation defect of sid2-250 slk1Δ. Thus, it is likely that Slk1p and Sid2p play a role in forespore membrane assembly by facilitating recruitment of components of the secretory apparatus, such as Psy1p, to allow membrane expansion. These studies thereby provide a novel link between the SIN and vesicle trafficking during cytokinesis.

scholarly journals The V260I Mutation in Fission Yeast α-Tubulin Atb2 Affects Microtubule Dynamics and EB1-Mal3 Localization and Activates the Bub1 Branch of the Spindle Checkpoint

2006 ◽  
Vol 17 (3) ◽  
pp. 1421-1435 ◽  
Author(s):  
Kazuhide Asakawa ◽  
Kazunori Kume ◽  
Muneyoshi Kanai ◽  
Tetsuya Goshima ◽  
Kohji Miyahara ◽  
...  
Keyword(s):  
Fission Yeast ◽  
Spindle Checkpoint ◽  
Spindle Pole ◽  
Vital Role ◽  
Microtubule Dynamics ◽  
High Rate ◽  
Single Amino Acid ◽  
Temperature Sensitive ◽  
Spindle Pole Bodies ◽  
Synthetically Lethal

We have identified a novel temperature-sensitive mutant of fission yeast α-tubulin Atb2 (atb2-983) that contains a single amino acid substitution (V260I). Atb2-983 is incorporated into the microtubules, and their overall structures are not altered noticeably, but microtubule dynamics is compromised during interphase. atb2-983 displays a high rate of chromosome missegregation and is synthetically lethal with deletions in a subset of spindle checkpoint genes including bub1, bub3, and mph1, but not with mad1, mad2, and mad3. During early mitosis in this mutant, Bub1, but not Mad2, remains for a prolonged period in the kinetochores that are situated in proximity to one of the two SPBs (spindle pole bodies). High dosage mal3+, encoding EB1 homologue, rescues atb2-983, suggesting that Mal3 function is compromised. Consistently, Mal3 localization and binding between Mal3 and Atb2-983 are impaired significantly, and a mal3 single mutant, such as atb2-983, displays prolonged Bub1 kinetochore localization. Furthermore in atb2-983 back-and-forth centromere oscillation during prometaphase is abolished. Intriguingly, this oscillation still occurs in the mal3 mutant, indicating that there is another defect independent of Mal3. These results show that microtubule dynamics is important for coordinated execution of mitotic events, in which Mal3 plays a vital role.

scholarly journals The Spindle Pole Bodies Facilitate Nuclear Envelope Division during Closed Mitosis in Fission Yeast

PLoS Biology ◽  
2007 ◽  
Vol 5 (7) ◽  
pp. e170 ◽  
Author(s):  
Liling Zheng ◽  
Cindi Schwartz ◽  
Valentin Magidson ◽  
Alexey Khodjakov ◽  
Snezhana Oliferenko
Keyword(s):  
Nuclear Envelope ◽  
Fission Yeast ◽  
Spindle Pole ◽  
Spindle Pole Bodies

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

1999 ◽  
Vol 112 (14) ◽  
pp. 2313-2321 ◽  
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.

scholarly journals A microtubule polymerase cooperates with the kinesin-6 motor and a microtubule cross-linker to promote bipolar spindle assembly in the absence of kinesin-5 and kinesin-14 in fission yeast

2017 ◽  
Vol 28 (25) ◽  
pp. 3647-3659 ◽  
Author(s):  
Masashi Yukawa ◽  
Tomoki Kawakami ◽  
Masaki Okazaki ◽  
Kazunori Kume ◽  
Ngang Heok Tang ◽  
...  
Keyword(s):  
Fission Yeast ◽  
Chromosome Segregation ◽  
Spindle Pole Body ◽  
Spindle Assembly ◽  
Spindle Pole ◽  
Temperature Sensitive ◽  
Bipolar Spindle ◽  
Pole Body ◽  
Cross Linker ◽  
Spindle Elongation

Accurate chromosome segregation relies on the bipolar mitotic spindle. In many eukaryotes, spindle formation is driven by the plus-end–directed motor kinesin-5 that generates outward force to establish spindle bipolarity. Its inhibition leads to the emergence of monopolar spindles with mitotic arrest. Intriguingly, simultaneous inactivation of the minus-end–directed motor kinesin-14 restores spindle bipolarity in many systems. Here we show that in fission yeast, three independent pathways contribute to spindle bipolarity in the absence of kinesin-5/Cut7 and kinesin-14/Pkl1. One is kinesin-6/Klp9 that engages with spindle elongation once short bipolar spindles assemble. Klp9 also ensures the medial positioning of anaphase spindles to prevent unequal chromosome segregation. Another is the Alp7/TACC-Alp14/TOG microtubule polymerase complex. Temperature-sensitive alp7cut7pkl1 mutants are arrested with either monopolar or very short spindles. Forced targeting of Alp14 to the spindle pole body is sufficient to render alp7cut7pkl1 triply deleted cells viable and promote spindle assembly, indicating that Alp14-mediated microtubule polymerization from the nuclear face of the spindle pole body could generate outward force in place of Cut7 during early mitosis. The third pathway involves the Ase1/PRC1 microtubule cross-linker that stabilizes antiparallel microtubules. Our study, therefore, unveils multifaceted interplay among kinesin-dependent and -independent pathways leading to mitotic bipolar spindle assembly.

γ-Tubulin complex-mediated anchoring of spindle microtubules to spindle-pole bodies requires Msd1 in fission yeast

2007 ◽  
Vol 9 (6) ◽  
pp. 646-653 ◽  
Author(s):  
Mika Toya ◽  
Masamitsu Sato ◽  
Uta Haselmann ◽  
Kazuhide Asakawa ◽  
Damian Brunner ◽  
...  
Keyword(s):  
Fission Yeast ◽  
Spindle Pole ◽  
Spindle Pole Bodies ◽  
Spindle Microtubules

scholarly journals Synergistic role of fission yeast Alp16GCP6 and Mzt1MOZART1 in γ-tubulin complex recruitment to mitotic spindle pole bodies and spindle assembly

2016 ◽  
Vol 27 (11) ◽  
pp. 1753-1763 ◽  
Author(s):  
Hirohisa Masuda ◽  
Takashi Toda
Keyword(s):  
Fission Yeast ◽  
Mitotic Spindle ◽  
Spindle Assembly Checkpoint ◽  
Synthetic Lethality ◽  
Spindle Assembly ◽  
Spindle Pole ◽  
Microtubule Assembly ◽  
Spindle Microtubule ◽  
Spindle Pole Bodies ◽  
Mitotic Spindle Pole

In fission yeast, γ-tubulin ring complex (γTuRC)–specific components Gfh1GCP4, Mod21GCP5, and Alp16GCP6 are nonessential for cell growth. Of these deletion mutants, only alp16Δ shows synthetic lethality with temperature-sensitive mutants of Mzt1MOZART1, a component of the γTuRC required for recruitment of the complex to microtubule-organizing centers. γ-Tubulin small complex levels at mitotic spindle pole bodies (SPBs, the centrosome equivalent in fungi) and microtubule levels for preanaphase spindles are significantly reduced in alp16Δ cells but not in gfh1Δ or mod21Δ cells. Furthermore, alp16Δ cells often form monopolar spindles and frequently lose a minichromosome when the spindle assembly checkpoint is inactivated. Alp16GCP6 promotes Mzt1-dependent γTuRC recruitment to mitotic SPBs and enhances spindle microtubule assembly in a manner dependent on its expression levels. Gfh1GCP4 and Mod21GCP5 are not required for Alp16GCP6-dependent γTuRC recruitment. Mzt1 has an additional role in the activation of the γTuRC for spindle microtubule assembly. The ratio of Mzt1 to γTuRC levels for preanaphase spindles is higher than at other stages of the cell cycle. Mzt1 overproduction enhances spindle microtubule assembly without affecting γTuRC levels at mitotic SPBs. We propose that Alp16GCP6 and Mzt1 act synergistically for efficient bipolar spindle assembly to ensure faithful chromosome segregation.

scholarly journals A Fourth Component of the Fission Yeast γ-Tubulin Complex, Alp16, Is Required for Cytoplasmic Microtubule Integrity and Becomes Indispensable When γ-Tubulin Function Is Compromised

2002 ◽  
Vol 13 (7) ◽  
pp. 2360-2373 ◽  
Author(s):  
Akiko Fujita ◽  
Leah Vardy ◽  
Miguel Angel Garcia ◽  
Takashi Toda
Keyword(s):  
Fission Yeast ◽  
Spindle Pole Body ◽  
Spindle Pole ◽  
Temperature Sensitive ◽  
Microtubule Organizing Center ◽  
Cytoplasmic Microtubule ◽  
Multicopy Suppressor ◽  
Large Complex ◽  
Synthetically Lethal ◽  
And Function

γ-Tubulin functions as a multiprotein complex, called the γ-tubulin complex (γ-TuC), and composes the microtubule organizing center (MTOC). Fission yeast Alp4 and Alp6 are homologues of two conserved γ-TuC proteins, hGCP2 and hGCP3, respectively. We isolated a novel gene, alp16 + , as a multicopy suppressor of temperature-sensitive alp6-719mutants. alp16 + encodes a 759-amino-acid protein with two conserved regions found in all other members of γ-TuC components. In addition, Alp16 contains an additional motif, which shows homology to hGCP6/Xgrip210. Gene disruption shows that alp16 + is not essential for cell viability. However, alp16 deletion displays abnormally long cytoplasmic microtubules, which curve around the cell tip. Furthermore, alp16-deleted mutants are hypersensitive to microtubule-depolymerizing drugs and synthetically lethal with either temperature-sensitive alp4-225,alp4-1891, or alp6-719 mutants. Overproduction of Alp16 is lethal, with defective phenotypes very similar to loss of Alp4 or Alp6. Alp16 localizes to the spindle pole body throughout the cell cycle and to the equatorial MTOC at postanaphase. Alp16 coimmunoprecipitates with γ-tubulin and cosediments with the γ-TuC in a large complex (>20 S). Alp16 is, however, not required for the formation of this large complex. We discuss evolutional conservation and divergence of structure and function of the γ-TuC between yeast and higher eukaryotes.

scholarly journals Protein Kinase C and the stress response pathways are required for kinetochore homeostasis

2016 ◽  
Author(s):  
Elena Ledesma-Fernández ◽  
Eva Herrero ◽  
Guðjón Ólafsson ◽  
Peter H Thorpe
Keyword(s):  
Stress Response ◽  
Spindle Assembly Checkpoint ◽  
Essential Gene ◽  
Spindle Pole ◽  
Temperature Sensitive ◽  
Gene Deletions ◽  
Spindle Pole Bodies ◽  
Genome Wide ◽  
Osmotic Stabilizer ◽  
A Genome

AbstractKinetochores serve both a structural role linking chromosomes to the mitotic spindle and a regulatory role, controlling the timing of mitosis via the spindle assembly checkpoint. To identify proteins that regulate the kinetochore we used a genome-wide fluorescence microscopy approach. We combined an array of mutants of either non-essential gene deletions or essential temperature-sensitive alleles with fluorescently tagged spindle pole bodies (centrosome) and outer kinetochores. Quantitative and qualitative analysis revealed mutants that affect the levels and distribution of kinetochores respectively. These mutants are enriched for those involved in mRNA processing, chromatin organization, DNA replication/repair and mitosis. Our data show that the Pkc1 kinase maintains the kinetochore focus via its ability to prevent cell stress and this phenotype is rescued by an osmotic stabilizer. These data support the notion that kinetochore and microtubule homeostasis are perturbed by the stress response pathways. Hence this observation provides a candidate mechanism for extracellular stress leading to chromosome segregation defects.

Chromatin behaviour during the mitotic cell cycle of Saccharomyces cerevisiae

1977 ◽  
Vol 24 (1) ◽  
pp. 81-93
Author(s):  
C.N. Gordon
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Nuclear Division ◽  
Mitotic Cell ◽  
Spindle Pole ◽  
Chromosomal Distribution ◽  
Yeast Saccharomyces Cerevisiae ◽  
Thin Sections ◽  
Daughter Cells ◽  
Spindle Pole Bodies ◽  
Direct Attachment

Chromatin behaviour during the cell division cycle of the yeast Saccharomyces cerevisiae has been investigated in cells which have been depleted of 90% of their RNA by digestion with ribonuclease. Removal of large amounts of RNA from the yeast nucleus before treatment of the cells with heavy metal fixatives and stains permits chromatin to be visualized with extreme clarity in thin sections of cells processed for electron microscopy by conventional procedures. Spindle pole bodies were also visualized by this treatment, although the associated microtubules were not. Chromatin is dispersed during interphase and occupies the non-nucleolar region of the nucleus which is known to be Feulgen-positive from light microscopy. Because spindle microtubules are not visualized, direct attachment of microtubules to chromatin fibrils could not be verified. However, chromatin was not attached directly to the spindle pole bodies and kinetochore differentiations were not observed in the nucleoplasm. During nuclear division chromatin remains dispersed and does not condense into discrete chromatids. As the nucleus expands into the bud, chromosomal distribution to the daughter cells is thought to result from the separation of the poles of the spindle apparatus with attached chromatin fibrils. However, that such distribution is occurring as the nucleus elongates is not obvious until an advanced stage of nuclear division is reached and partition of the nucleus is nearly complete. Thus, no aggregation of chromatin into metaphase or anaphase plates occurs and the appearance of chromatin during mitosis is essentially the same as in interphase. These observations indicate that the marked changes in the topological structure of chromatin which characterize mitosis in the higher eukaryotes do not occur in S. cerevisiae.

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