scholarly journals The Msd1–Wdr8–Pkl1 complex anchors microtubule minus ends to fission yeast spindle pole bodies

2015 ◽  
Vol 209 (4) ◽  
pp. 549-562 ◽  
Author(s):  
Masashi Yukawa ◽  
Chiho Ikebe ◽  
Takashi Toda
Keyword(s):  
Fission Yeast ◽  
Critical Role ◽  
Spindle Pole Body ◽  
Spindle Pole ◽  
Microtubule Organizing Center ◽  
Spatiotemporal Regulation ◽  
Spindle Pole Bodies ◽  
Organizing Center ◽  
Spindle Microtubules ◽  
Insight Into

The minus ends of spindle microtubules are anchored to a microtubule-organizing center. The conserved Msd1/SSX2IP proteins are localized to the spindle pole body (SPB) and the centrosome in fission yeast and humans, respectively, and play a critical role in microtubule anchoring. In this paper, we show that fission yeast Msd1 forms a ternary complex with another conserved protein, Wdr8, and the minus end–directed Pkl1/kinesin-14. Individual deletion mutants displayed the identical spindle-protrusion phenotypes. Msd1 and Wdr8 were delivered by Pkl1 to mitotic SPBs, where Pkl1 was tethered through Msd1–Wdr8. The spindle-anchoring defect imposed by msd1/wdr8/pkl1 deletions was suppressed by a mutation of the plus end–directed Cut7/kinesin-5, which was shown to be mutual. Intriguingly, Pkl1 motor activity was not required for its anchoring role once targeted to the SPB. Therefore, spindle anchoring through Msd1–Wdr8–Pkl1 is crucial for balancing the Cut7/kinesin-5–mediated outward force at the SPB. Our analysis provides mechanistic insight into the spatiotemporal regulation of two opposing kinesins to ensure mitotic spindle bipolarity.

scholarly journals Fission yeast MOZART1/Mzt1 is an essential γ-tubulin complex component required for complex recruitment to the microtubule organizing center, but not its assembly

2013 ◽  
Vol 24 (18) ◽  
pp. 2894-2906 ◽  
Author(s):  
Hirohisa Masuda ◽  
Risa Mori ◽  
Masashi Yukawa ◽  
Takashi Toda
Keyword(s):  
Fission Yeast ◽  
Spindle Pole Body ◽  
Spindle Pole ◽  
Temperature Sensitive ◽  
Microtubule Organizing Center ◽  
Microtubule Organization ◽  
Unique Role ◽  
Pole Body ◽  
Organizing Center ◽  
Core Components

γ-Tubulin plays a universal role in microtubule nucleation from microtubule organizing centers (MTOCs) such as the animal centrosome and fungal spindle pole body (SPB). γ-Tubulin functions as a multiprotein complex called the γ-tubulin complex (γ-TuC), consisting of GCP1–6 (GCP1 is γ-tubulin). In fungi and flies, it has been shown that GCP1–3 are core components, as they are indispensable for γ-TuC complex assembly and cell division, whereas the other three GCPs are not. Recently a novel conserved component, MOZART1, was identified in humans and plants, but its precise functions remain to be determined. In this paper, we characterize the fission yeast homologue Mzt1, showing that it is essential for cell viability. Mzt1 is present in approximately equal stoichiometry with Alp4/GCP2 and localizes to all the MTOCs, including the SPB and interphase and equatorial MTOCs. Temperature-sensitive mzt1 mutants display varying degrees of compromised microtubule organization, exhibiting multiple defects during both interphase and mitosis. Mzt1 is required for γ-TuC recruitment, but not sufficient to localize to the SPB, which depends on γ-TuC integrity. Intriguingly, the core γ-TuC assembles in the absence of Mzt1. Mzt1 therefore plays a unique role within the γ-TuC components in attachment of this complex to the major MTOC site.

γ-Tubulin and the fungal microtubule cytoskeleton

1995 ◽  
Vol 73 (S1) ◽  
pp. 352-358 ◽  
Author(s):  
Berl R. Oakley
Keyword(s):  
Cell Cycle ◽  
Mitotic Spindle ◽  
Spindle Pole Body ◽  
Spindle Pole ◽  
Microtubule Organizing Center ◽  
Pole Body ◽  
Spindle Pole Bodies ◽  
Key Words Tubulin ◽  
Organizing Center ◽  
Chromosomal Condensation

γ-Tubulin is present in phylogenetically diverse eukaryotes. It is a component of microtubule organizing centers such as the spindle pole bodies of fungi. In Aspergillus nidulans and Schizosaccharomyces pombe, it is essential for nuclear division, and, thus, for viability. In A. nidulans, nuclei carrying a γ-tubulin disruption can be maintained in heterokaryons, and the phenotypes caused by the disruption can be determined in uninucleate spores produced by the heterokaryons. Experiments with heterokaryons created in strains with mutations that allow synchronization of the cell cycle reveal that γ-tubulin is not required for the transition from the G1 phase of the cell cycle through S phase to G2, nor for the entry into mitosis as judged by chromosomal condensation. It is, however, required for the formation of the mitotic spindle and for the successful completion of mitosis. Staining with the MPM-2 monoclonal antibody reveals that spindle pole body replication occurs in the absence of functional γ-tubulin. Finally, human γ-tubulin functions in fission yeast, and this indicates that γ-tubulin has similar functions in widely divergent organisms. Key words: tubulin, microtubule, spindle pole body, microtubule organizing center.

Genetic interactions between CDC31 and KAR1, two genes required for duplication of the microtubule organizing center in Saccharomyces cerevisiae.

Genetics ◽  
1994 ◽  
Vol 137 (2) ◽  
pp. 407-422 ◽  
Author(s):  
E A Vallen ◽  
W Ho ◽  
M Winey ◽  
M D Rose
Keyword(s):  
Copy Number ◽  
Gene Dosage ◽  
Spindle Pole Body ◽  
Direct Interaction ◽  
Spindle Pole ◽  
High Copy Number ◽  
Temperature Sensitive ◽  
Microtubule Organizing Center ◽  
Recessive Mutations ◽  
Organizing Center

Abstract KAR1 encodes an essential component of the yeast spindle pole body (SPB) that is required for karyogamy and SPB duplication. A temperature-sensitive mutation, kar1-delta 17, mapped to a region required for SPB duplication and for localization to the SPB. To identify interacting SPB proteins, we isolated 13 dominant mutations and 3 high copy number plasmids that suppressed the temperature sensitivity of kar1-delta 17. Eleven extragenic suppressor mutations mapped to two linkage groups, DSK1 and DSK2. The extragenic suppressors were specific for SPB duplication and did not suppress karyogamy-defective alleles. The major class, DSK1, consisted of mutations in CDC31. CDC31 is required for SPB duplication and encodes a calmodulin-like protein that is most closely related to caltractin/centrin, a protein associated with the Chlamydomonas basal body. The high copy number suppressor plasmids contained the wild-type CDC31 gene. One CDC31 suppressor allele conferred a temperature-sensitive defect in SPB duplication, which was counter-suppressed by recessive mutations in KAR1. In spite of the evidence for a direct interaction, the strongest CDC31 alleles, as well as both DSK2 alleles, suppressed a complete deletion of KAR1. However, the CDC31 alleles also made the cell supersensitive to KAR1 gene dosage, arguing against a simple bypass mechanism of suppression. We propose a model in which Kar1p helps localize Cdc31p to the SPB and that Cdc31p then initiates SPB duplication via interaction with a downstream effector.

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 Mobility, Microtubule Nucleation and Structure of Microtubule-organizing Centers in Multinucleated Hyphae ofAshbya gossypii

2010 ◽  
Vol 21 (1) ◽  
pp. 18-28 ◽  
Author(s):  
Claudia Lang ◽  
Sandrine Grava ◽  
Tineke van den Hoorn ◽  
Rhonda Trimble ◽  
Peter Philippsen ◽  
...  
Keyword(s):  
Spindle Pole Body ◽  
Hyphal Growth ◽  
Spindle Pole ◽  
Microtubule Organizing Center ◽  
Microtubule Nucleation ◽  
Cytoplasmic Microtubule ◽  
Organizing Center ◽  
Cytoplasmic Side ◽  
Independent Mode

We investigated the migration of multiple nuclei in hyphae of the filamentous fungus Ashbya gossypii. Three types of cytoplasmic microtubule (cMT)-dependent nuclear movements were characterized using live cell imaging: short-range oscillations (up to 4.5 μm/min), rotations (up to 180° in 30 s), and long-range nuclear bypassing (up to 9 μm/min). These movements were superimposed on a cMT-independent mode of nuclear migration, cotransport with the cytoplasmic stream. This latter mode is sufficient to support wild-type-like hyphal growth speeds. cMT-dependent nuclear movements were led by a nuclear-associated microtubule-organizing center, the spindle pole body (SPB), which is the sole site of microtubule nucleation in A. gossypii. Analysis of A. gossypii SPBs by electron microscopy revealed an overall laminar structure similar to the budding yeast SPB but with distinct differences at the cytoplasmic side. Up to six perpendicular and tangential cMTs emanated from a more spherical outer plaque. The perpendicular and tangential cMTs most likely correspond to short, often cortex-associated cMTs and to long, hyphal growth-axis–oriented cMTs, respectively, seen by in vivo imaging. Each SPB nucleates its own array of cMTs, and the lack of overlapping cMT arrays between neighboring nuclei explains the autonomous nuclear oscillations and bypassing observed in A. gossypii hyphae.

scholarly journals The role of Aspergillus nidulans polo-like kinase PlkA in microtubule-organizing center control

2021 ◽  
Author(s):  
Xiaolei Gao ◽  
Saturnino Herrero ◽  
Valentin Wernet ◽  
Sylvia Erhardt ◽  
Oliver Valerius ◽  
...  
Keyword(s):  
Aspergillus Nidulans ◽  
Cell Types ◽  
Spindle Pole ◽  
Receptor Protein ◽  
Animal Cells ◽  
Microtubule Organizing Center ◽  
Spindle Pole Bodies ◽  
Organizing Center ◽  
Ring Complex ◽  
Core Components

Centrosomes are important microtubule-organizing centers (MTOC) in animal cells. In addition, non-centrosomal MTOCs (ncMTOCs) were described in many cell types. Functional analogs of centrosomes in fungi are the spindle pole bodies (SPBs). In Aspergillus nidulans additional MTOCs were discovered at septa (sMTOC). Although the core components are conserved in both MTOCs, their composition and organization are different and dynamic. Here, we show that the polo-like kinase PlkA binds the γ-tubulin ring complex (γ-TuRC) receptor protein ApsB and contributes to targeting ApsB to both MTOCs. PlkA coordinates SPB outer plaque with sMTOC activities. PlkA kinase activity was required for astral MT formation involving ApsB recruitment. PlkA also interacted with the γ-TuRC inner plaque receptor protein PcpA. Mitosis was delayed without PlkA, and the PlkA protein was required for proper mitotic spindle morphology, although this function was independent of its catalytic activity. Our results suggest polo-like kinase as a regulator of MTOC activities and as a scaffolding unit through interaction with γ-tubulin ring complex receptors.

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 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 Kinetochore capture by spindle microtubules: why fission yeast may prefer pivoting to search-and-capture

2019 ◽  
Author(s):  
Indrani Nayak ◽  
Dibyendu Das ◽  
Amitabha Nandi
Keyword(s):  
Fission Yeast ◽  
Cell Biology ◽  
Rotational Diffusion ◽  
Spindle Pole Body ◽  
Experimental Studies ◽  
Spindle Pole ◽  
Capture Process ◽  
Geometrical Constraints ◽  
The Common ◽  
Spindle Microtubules

The mechanism by which microtubules find kinetochores during spindle formation is a key question in cell biology. Previous experimental studies have shown that although search-and-capture of kinetochores by dynamic microtubules is a dominant mechanism in many organisms, several other capture mechanisms are also possible. One such mechanism reported in Schizosaccharomyces pombe shows that microtubules can exhibit a prolonged pause between growth and shrinkage. During the pause, the microtubules pivoted at the spindle pole body search for the kinetochores by performing an angular diffusion. Is the latter mechanism purely accidental, or could there be any physical advantage underlying its selection? To compare the efficiency of these two mechanisms, we numerically study distinct models and compute the timescales of kinetochore capture as a function of microtubule number N. We find that the capture timescales have non-trivial dependences on microtubule number, and one mechanism may be preferred over the other depending on this number. While for small N (as in fission yeast), the typical capture times due to rotational diffusion are lesser than those for search-and-capture, the situation is reversed beyond a certain N. The capture times for rotational diffusion tend to saturate due to geometrical constraints, while those for search-and-capture reduce monotonically with increasing N making it physically more efficient. The results provide a rationale for the common occurrence of classic search-and-capture process in many eukaryotes which have few hundreds of dynamic microtubules, as well as justify exceptions in cells with fewer microtubules.

scholarly journals Tts1, the fission yeast homologue of the TMEM33 family, functions in NE remodeling during mitosis

2014 ◽  
Vol 25 (19) ◽  
pp. 2970-2983 ◽  
Author(s):  
Dan Zhang ◽  
Snezhana Oliferenko
Keyword(s):  
Fission Yeast ◽  
Structural Features ◽  
Spindle Pole ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
C Terminus ◽  
Spindle Pole Bodies ◽  
Evolutionarily Conserved ◽  
Pore Complexes ◽  
Insight Into

The fission yeast Schizosaccharomyces pombe undergoes “closed” mitosis in which the nuclear envelope (NE) stays intact throughout chromosome segregation. Here we show that Tts1, the fission yeast TMEM33 protein that was previously implicated in organizing the peripheral endoplasmic reticulum (ER), also functions in remodeling the NE during mitosis. Tts1 promotes insertion of spindle pole bodies (SPBs) in the NE at the onset of mitosis and modulates distribution of the nuclear pore complexes (NPCs) during mitotic NE expansion. Structural features that drive partitioning of Tts1 to the high-curvature ER domains are crucial for both aspects of its function. An amphipathic helix located at the C-terminus of Tts1 is important for ER shaping and modulating the mitotic NPC distribution. Of interest, the evolutionarily conserved residues at the luminal interface of the third transmembrane region function specifically in promoting SPB-NE insertion. Our data illuminate cellular requirements for remodeling the NE during “closed” nuclear division and provide insight into the structure and functions of the eukaryotic TMEM33 family.

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