scholarly journals How Essential Kinesin-5 Becomes Non-Essential: Force Balance and Microtubule Dynamics Matter

2020 ◽  
Author(s):  
Masashi Yukawa ◽  
Yasuhiro Teratani ◽  
Takashi Toda
Keyword(s):  
Spindle Assembly ◽  
Microtubule Dynamics ◽  
Force Balance ◽  
Fine Tuning ◽  
Mitotic Spindles ◽  
Clinical Implementation ◽  
Bipolar Spindle ◽  
Microtubule Associated Proteins ◽  
Chromatid Cohesion ◽  
Associated Proteins

The bipolar mitotic spindle drives accurate chromosome segregation by capturing the kinetochore and pulling each set of sister chromatids to the opposite poles. In this review, we describe recent findings on the multiple pathways leading to bipolar spindle formation in fission yeast and discuss these results from a broader perspective. Roles of four mitotic kinesins (Kinesin-5, Kinesin-6, Kinesin-12 and Kinesin-14) in spindle assembly are depicted, and how a group of microtubule-associated proteins, sister chromatid cohesion and the kinetochore collaborates with these motors is shown. We have paid special attention to the molecular pathways that render otherwise essential Kinesin-5 to become non-essential: how cells build bipolar mitotic spindles without the need for Kinesin-5 and where the alternate forces come from are considered. We highlight the force balance for bipolar spindle assembly and explain how outward and inward forces are generated by various ways, in which the proper fine-tuning of microtubule dynamics plays a crucial role. Overall, these new pathways have illuminated remarkable plasticity and adaptability of spindle mechanics. Kinesin molecules are regarded as prospective targets for cancer chemotherapy and many specific inhibitors have been developed. However, several hurdles have arisen against their clinical implementation. This review provides insight into possible strategies to overcome these challenges.

scholarly journals How Essential Kinesin-5 Becomes Non-Essential in Fission Yeast: Force Balance and Microtubule Dynamics Matter

Cells ◽  
2020 ◽  
Vol 9 (5) ◽  
pp. 1154 ◽  
Author(s):  
Masashi Yukawa ◽  
Yasuhiro Teratani ◽  
Takashi Toda
Keyword(s):  
Fission Yeast ◽  
Spindle Assembly ◽  
Microtubule Dynamics ◽  
Force Balance ◽  
Fine Tuning ◽  
Mitotic Spindles ◽  
Clinical Implementation ◽  
Bipolar Spindle ◽  
Microtubule Associated Proteins ◽  
Associated Proteins

The bipolar mitotic spindle drives accurate chromosome segregation by capturing the kinetochore and pulling each set of sister chromatids to the opposite poles. In this review, we describe recent findings on the multiple pathways leading to bipolar spindle formation in fission yeast and discuss these results from a broader perspective. The roles of three mitotic kinesins (Kinesin-5, Kinesin-6 and Kinesin-14) in spindle assembly are depicted, and how a group of microtubule-associated proteins, sister chromatid cohesion and the kinetochore collaborate with these motors is shown. We have paid special attention to the molecular pathways that render otherwise essential Kinesin-5 to become non-essential: how cells build bipolar mitotic spindles without the need for Kinesin-5 and where the alternate forces come from are considered. We highlight the force balance for bipolar spindle assembly and explain how outward and inward forces are generated by various ways, in which the proper fine-tuning of microtubule dynamics plays a crucial role. Overall, these new pathways have illuminated the remarkable plasticity and adaptability of spindle mechanics. Kinesin molecules are regarded as prospective targets for cancer chemotherapy and many specific inhibitors have been developed. However, several hurdles have arisen against their clinical implementation. This review provides insight into possible strategies to overcome these challenges.

scholarly journals Stu2p binds tubulin and undergoes an open-to-closed conformational change

2006 ◽  
Vol 172 (7) ◽  
pp. 1009-1022 ◽  
Author(s):  
Jawdat Al-Bassam ◽  
Mark van Breugel ◽  
Stephen C. Harrison ◽  
Anthony Hyman
Keyword(s):  
Conformational Change ◽  
Conformational Transition ◽  
Budding Yeast ◽  
Microtubule Dynamics ◽  
Open Structure ◽  
Microtubule Associated Proteins ◽  
Associated Proteins ◽  
The Common

Stu2p from budding yeast belongs to the conserved Dis1/XMAP215 family of microtubule-associated proteins (MAPs). The common feature of proteins in this family is the presence of HEAT repeat–containing TOG domains near the NH2 terminus. We have investigated the functions of the two TOG domains of Stu2p in vivo and in vitro. Our data suggest that Stu2p regulates microtubule dynamics through two separate activities. First, Stu2p binds to a single free tubulin heterodimer through its first TOG domain. A large conformational transition in homodimeric Stu2p from an open structure to a closed one accompanies the capture of a single free tubulin heterodimer. Second, Stu2p has the capacity to associate directly with microtubule ends, at least in part, through its second TOG domain. These two properties lead to the stabilization of microtubules in vivo, perhaps by the loading of tubulin dimers at microtubule ends. We suggest that this mechanism of microtubule regulation is a conserved feature of the Dis1/XMAP215 family of MAPs.

scholarly journals The impact of chromosomes and centrosomes on spindle assembly as observed in living cells.

1995 ◽  
Vol 129 (5) ◽  
pp. 1287-1300 ◽  
Author(s):  
D Zhang ◽  
R B Nicklas
Keyword(s):  
Spindle Assembly ◽  
Living Cells ◽  
Microtubule Dynamics ◽  
Specific Site ◽  
Polarization Microscopy ◽  
Spindle Microtubule ◽  
Single Chromosome ◽  
Bipolar Spindle ◽  
The Impact

We analyzed the role that chromosomes, kinetochores, and centrosomes play in spindle assembly in living grasshopper spermatocytes by reconstructing spindles lacking certain components. We used video-enhanced, polarization microscopy to distinguish the effect of each component on spindle microtubule dynamics and we discovered that both chromosomes and centrosomes make potent and very different contributions to the organization of the spindle. Remarkably, the position of a single chromosome can markedly affect the distribution of microtubules within a spindle or even alter the fate of spindle assembly. In an experimentally constructed spindle having only one chromosome, moving the chromosome to one of the two poles induces a dramatic assembly of microtubules at the nearer pole and a concomitant disassembly at the farther pole. So long as a spindle carries a single chromosome it will persist normally. A spindle will also persist even when all chromosomes are detached and then removed from the cell. If, however, a single chromosome remains in the cell but is detached from the spindle and kept in the cytoplasm, the spindle disassembles. One might expect the effect of chromosomes on spindle assembly to relate to a property of a specific site on each chromosome, perhaps the kinetochore. We have ruled out that possibility by showing that it is the size of chromosomes rather than the number of kinetochores that matters. Although chromosomes affect spindle assembly, they cannot organize a spindle in the absence of centrosomes. In contrast, centrosomes can organize a functional bipolar spindle in the absence of chromosomes. If both centrosomes and chromosomes are removed from the cell, the spindle quickly disappears.

scholarly journals The microtubule lattice and plus-end association of Drosophila Mini spindles is spatially regulated to fine-tune microtubule dynamics

2011 ◽  
Vol 22 (22) ◽  
pp. 4343-4361 ◽  
Author(s):  
Joshua D. Currie ◽  
Shannon Stewman ◽  
Gregory Schimizzi ◽  
Kevin C. Slep ◽  
Ao Ma ◽  
...  
Keyword(s):  
Dynamic Instability ◽  
Function Analysis ◽  
Microtubule Dynamics ◽  
Cell Periphery ◽  
Cell Interior ◽  
Microtubule Associated Proteins ◽  
Contact Sites ◽  
Associated Proteins ◽  
Fine Tune

Individual microtubules (MTs) exhibit dynamic instability, a behavior in which they cycle between phases of growth and shrinkage while the total amount of MT polymer remains constant. Dynamic instability is promoted by the conserved XMAP215/Dis1 family of microtubule-associated proteins (MAPs). In this study, we conducted an in vivo structure–function analysis of the Drosophila homologue Mini spindles (Msps). Msps exhibits EB1-dependent and spatially regulated MT localization, targeting to microtubule plus ends in the cell interior and decorating the lattice of growing and shrinking microtubules in the cell periphery. RNA interference rescue experiments revealed that the NH2-terminal four TOG domains of Msps function as paired units and were sufficient to promote microtubule dynamics and EB1 comet formation. We also identified TOG5 and novel inter-TOG linker motifs that are required for targeting Msps to the microtubule lattice. These novel microtubule contact sites are necessary for the interplay between the conserved TOG domains and inter-TOG MT binding that underlies the ability of Msps to promote MT dynamic instability.

scholarly journals Two XMAP215/TOG microtubule polymerases, Alp14 and Dis1, play non-exchangeable, distinct roles in microtubule organisation in fission yeast

2019 ◽  
Author(s):  
Masashi Yukawa ◽  
Tomoki Kawakami ◽  
Corinne Pinder ◽  
Takashi Toda
Keyword(s):  
Fission Yeast ◽  
Chromosome Segregation ◽  
Genome Stability ◽  
Spindle Assembly ◽  
Mitotic Progression ◽  
Spindle Microtubule ◽  
Microtubule Associated Proteins ◽  
Spatial Regulation ◽  
Associated Proteins ◽  
Microtubule Formation

AbstractProper bipolar spindle assembly underlies accurate chromosome segregation. A cohort of microtubule-associated proteins orchestrates spindle microtubule formation in a spatiotemporally coordinated manner. Among them, the conserved XMAP215/TOG family of microtubule polymerase plays a central role in spindle assembly. In fission yeast, two XMAP215/TOG members, Alp14 and Dis1, share essential roles in cell viability; however how these two proteins functionally collaborate remains undetermined. Here we show the functional interplay and specification of Alp14 and Dis1. Creation of new mutant alleles of alp14, which display temperature sensitivity in the absence of Dis1, enabled us to conduct detailed analyses of a double mutant. We have found that simultaneous inactivation of Alp14 and Dis1 results in early mitotic arrest with very short, fragile spindles. Intriguingly, these cells often undergo spindle collapse, leading to a lethal “cut” phenotype. By implementing an artificial targetting system, we have shown that Alp14 and Dis1 are not functionally exchangeable and as such are not merely redundant paralogues. Intriguingly, while Alp14 promotes microtubule nucleation, Dis1 does not. Our results uncover that the intrinsic specification, not the spatial regulation, between Alp14 and Dis1 underlies the collaborative actions of these two XMAP215/TOG members in mitotic progression, spindle integrity and genome stability.

scholarly journals Ensconsin/Map7 promotes microtubule growth and centrosome separation in Drosophila neural stem cells

2014 ◽  
Vol 204 (7) ◽  
pp. 1111-1121 ◽  
Author(s):  
Emmanuel Gallaud ◽  
Renaud Caous ◽  
Aude Pascal ◽  
Franck Bazile ◽  
Jean-Philippe Gagné ◽  
...  
Keyword(s):  
Mitotic Spindle ◽  
Spindle Assembly ◽  
Microtubule Associated Proteins ◽  
Microtubule Polymerization ◽  
Sister Chromatids ◽  
Centrosome Separation ◽  
Associated Proteins ◽  
Microtubule Growth ◽  
Mitotic Spindle Assembly

The mitotic spindle is crucial to achieve segregation of sister chromatids. To identify new mitotic spindle assembly regulators, we isolated 855 microtubule-associated proteins (MAPs) from Drosophila melanogaster mitotic or interphasic embryos. Using RNAi, we screened 96 poorly characterized genes in the Drosophila central nervous system to establish their possible role during spindle assembly. We found that Ensconsin/MAP7 mutant neuroblasts display shorter metaphase spindles, a defect caused by a reduced microtubule polymerization rate and enhanced by centrosome ablation. In agreement with a direct effect in regulating spindle length, Ensconsin overexpression triggered an increase in spindle length in S2 cells, whereas purified Ensconsin stimulated microtubule polymerization in vitro. Interestingly, ensc-null mutant flies also display defective centrosome separation and positioning during interphase, a phenotype also detected in kinesin-1 mutants. Collectively, our results suggest that Ensconsin cooperates with its binding partner Kinesin-1 during interphase to trigger centrosome separation. In addition, Ensconsin promotes microtubule polymerization during mitosis to control spindle length independent of Kinesin-1.

A role for microtubule dynamics in phagosome movement

1998 ◽  
Vol 111 (3) ◽  
pp. 303-312 ◽  
Author(s):  
A. Blocker ◽  
G. Griffiths ◽  
J.C. Olivo ◽  
A.A. Hyman ◽  
F.F. Severin
Keyword(s):  
Microtubule Dynamics ◽  
Strong Preference ◽  
Cell Periphery ◽  
Perinuclear Region ◽  
Cell Interior ◽  
Microtubule Associated Proteins ◽  
Microtubule Motors ◽  
Associated Proteins

We have shown previously that intracellular phagosome movement requires microtubules. Here we provide evidence that within cells phagosomes display two different kinds of microtubule-based movements in approximately equal proportions. The first type occurs predominantly in the cell periphery, often shortly after the phagosome is formed, and at speeds below 0.1 microm/second. The second is faster (0.2-1.5 micron/second) and occurs mainly after phagosomes have reached the cell interior. Treating cells with nanomolar concentrations of taxol or nocodazole alters microtubule dynamics without affecting either total polymer mass or microtubule organisation. Such treatments slow the accumulation of phagosomes in the perinuclear region and reduce the number of slow movements by up to 50% without affecting the frequency of fast movements. This suggests that a proportion of slow movements are mediated by microtubule dynamics while fast movements are powered by microtubule motors. In macrophages, interphase microtubules radiate from the microtubule organising centre with their plus-end towards the cell periphery. To understand the behaviour of ‘early’ phagosomes at the cell periphery we investigated their ability to bind microtubule plus-ends in vitro. We show that early phagosomes have a strong preference for microtubule plus-ends, whereas ‘late’ phagosomes do not, and that plus-end affinity requires the presence of microtubule-associated proteins within cytosol. We suggest that phagosomes can bind to the plus-ends of dynamic microtubules and move by following their shrinkage or growth.

scholarly journals Cyclin B interaction with microtubule-associated protein 4 (MAP4) targets p34cdc2 kinase to microtubules and is a potential regulator of M-phase microtubule dynamics.

1995 ◽  
Vol 128 (5) ◽  
pp. 849-862 ◽  
Author(s):  
K Ookata ◽  
S Hisanaga ◽  
J C Bulinski ◽  
H Murofushi ◽  
H Aizawa ◽  
...  
Keyword(s):  
Microtubule Dynamics ◽  
Cyclin B ◽  
Microtubule Associated Proteins ◽  
Mitotic Kinase ◽  
Culture Cells ◽  
Tissue Culture Cells ◽  
Detergent Extraction ◽  
M Phase ◽  
Associated Proteins ◽  
Free Microtubules

We previously demonstrated (Ookata et al., 1992, 1993) that the p34cdc2/cyclin B complex associates with microtubules in the mitotic spindle and premeiotic aster in starfish oocytes, and that microtubule-associated proteins (MAPs) might be responsible for this interaction. In this study, we have investigated the mechanism by which p34cdc2 kinase associates with the microtubule cytoskeleton in primate tissue culture cells whose major MAP is known to be MAP4. Double staining of primate cells with anti-cyclin B and anti-MAP4 antibodies demonstrated these two antigens were colocalized on microtubules and copartitioned following two treatments that altered MAP4 distribution. Detergent extraction before fixation removed cyclin B as well as MAP4 from the microtubules. Depolymerization of some of the cellular microtubules with nocodazole preferentially retained the microtubule localization of both cyclin B and MAP4. The association of p34cdc2/cyclin B kinase with microtubules was also shown biochemically to be mediated by MAP4. Cosedimentation of purified p34cdc2/cyclin B with purified microtubule proteins containing MAP4, but not with MAP-free microtubules, as well as binding of MAP4 to GST-cyclin B fusion proteins, demonstrated an interaction between cyclin B and MAP4. Using recombinant MAP4 fragments, we demonstrated that the Pro-rich C-terminal region of MAP4 is sufficient to mediate the cyclin B-MAP4 interaction. Since p34cdc2/cyclin B physically associated with MAP4, we examined the ability of the kinase complex to phosphorylate MAP4. Incubation of a ternary complex of p34cdc2, cyclin B, and the COOH-terminal domain of MAP4, PA4, with ATP resulted in intracomplex phosphorylation of PA4. Finally, we tested the effects of MAP4 phosphorylation on microtubule dynamics. Phosphorylation of MAP4 by p34cdc2 kinase did not prevent its binding to microtubules, but abolished its microtubule stabilizing activity. Thus, the cyclin B/MAP4 interaction we have described may be important in targeting the mitotic kinase to appropriate cytoskeletal substrates, for the regulation of spindle assembly and dynamics.

Differences in the regulation of microtubule dynamics by microtubule-associated proteins MAP1B and MAP2

1996 ◽  
Vol 35 (2) ◽  
pp. 134-146 ◽  
Author(s):  
André Vandecandelaere ◽  
Barbara Pedrotti ◽  
Michelle A. Utton ◽  
Rosy A. Calvert ◽  
Peter M. Bayley
Keyword(s):  
Microtubule Dynamics ◽  
Microtubule Associated Proteins ◽  
Associated Proteins
Sign in / Sign up

Export Citation Format

Share Document