scholarly journals The contribution of αβ-tubulin curvature to microtubule dynamics

2014 ◽  
Vol 207 (3) ◽  
pp. 323-334 ◽  
Author(s):  
Gary J. Brouhard ◽  
Luke M. Rice
Keyword(s):  
Cell Division ◽  
Mitotic Spindle ◽  
Fundamental Property ◽  
Microtubule Dynamics ◽  
Cellular Structures ◽  
Microtubule Associated Proteins ◽  
Associated Proteins ◽  
Microtubule Growth ◽  
Microtubule Networks ◽  
And Control

Microtubules are dynamic polymers of αβ-tubulin that form diverse cellular structures, such as the mitotic spindle for cell division, the backbone of neurons, and axonemes. To control the architecture of microtubule networks, microtubule-associated proteins (MAPs) and motor proteins regulate microtubule growth, shrinkage, and the transitions between these states. Recent evidence shows that many MAPs exert their effects by selectively binding to distinct conformations of polymerized or unpolymerized αβ-tubulin. The ability of αβ-tubulin to adopt distinct conformations contributes to the intrinsic polymerization dynamics of microtubules. αβ-Tubulin conformation is a fundamental property that MAPs monitor and control to build proper microtubule networks.

scholarly journals The mitotic spindle protein CKAP2 potently increases formation and stability of microtubules

eLife ◽  
2022 ◽  
Vol 11 ◽  
Author(s):  
Thomas S McAlear ◽  
Susanne Bechstedt
Keyword(s):  
Cell Division ◽  
Growth Factor ◽  
Mitotic Spindle ◽  
Apparent Rate Constant ◽  
Microtubule Dynamics ◽  
Proliferation Marker ◽  
Microtubule Growth ◽  
Large Rearrangements ◽  
And Function

Cells increase microtubule dynamics to make large rearrangements to their microtubule cytoskeleton during cell division. Changes in microtubule dynamics are essential for the formation and function of the mitotic spindle, and misregulation can lead to aneuploidy and cancer. Using in vitro reconstitution assays we show that the mitotic spindle protein Cytoskeleton-Associated Protein 2 (CKAP2) has a strong effect on nucleation of microtubules by lowering the critical tubulin concentration 100-fold. CKAP2 increases the apparent rate constant ka of microtubule growth by 50-fold and increases microtubule growth rates. In addition, CKAP2 strongly suppresses catastrophes. Our results identify CKAP2 as the most potent microtubule growth factor to date. These finding help explain CKAP2's role as an important spindle protein, proliferation marker, and oncogene.

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.

scholarly journals Human CLASP2 specifically regulates microtubule catastrophe and rescue

2018 ◽  
Vol 29 (10) ◽  
pp. 1168-1177 ◽  
Author(s):  
Elizabeth J. Lawrence ◽  
Göker Arpag˘ ◽  
Stephen R. Norris ◽  
Marija Zanic
Keyword(s):  
Growth Rates ◽  
Molecular Mechanisms ◽  
Direct Interaction ◽  
Microtubule Dynamics ◽  
Microtubule Associated Proteins ◽  
Cellular Processes ◽  
Associated Proteins ◽  
Microtubule Growth ◽  
Protein Components

Cytoplasmic linker-associated proteins (CLASPs) are microtubule-associated proteins essential for microtubule regulation in many cellular processes. However, the molecular mechanisms underlying CLASP activity are not understood. Here, we use purified protein components and total internal reflection fluorescence microscopy to investigate the effects of human CLASP2 on microtubule dynamics in vitro. We demonstrate that CLASP2 suppresses microtubule catastrophe and promotes rescue without affecting the rates of microtubule growth or shrinkage. Strikingly, when CLASP2 is combined with EB1, a known binding partner, the effects on microtubule dynamics are strongly enhanced. We show that synergy between CLASP2 and EB1 is dependent on a direct interaction, since a truncated EB1 protein that lacks the CLASP2-binding domain does not enhance CLASP2 activity. Further, we find that EB1 targets CLASP2 to microtubules and increases the dwell time of CLASP2 at microtubule tips. Although the temporally averaged microtubule growth rates are unaffected by CLASP2, we find that microtubules grown with CLASP2 display greater variability in growth rates. Our results provide insight into the regulation of microtubule dynamics by CLASP proteins and highlight the importance of the functional interplay between regulatory proteins at dynamic microtubule ends.

scholarly journals Stu2p, the budding yeast member of the conserved Dis1/XMAP215 family of microtubule-associated proteins is a plus end–binding microtubule destabilizer

2003 ◽  
Vol 161 (2) ◽  
pp. 359-369 ◽  
Author(s):  
Mark van Breugel ◽  
David Drechsel ◽  
Anthony Hyman
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Division ◽  
Quantitative Analysis ◽  
Family Member ◽  
Budding Yeast ◽  
Microtubule Dynamics ◽  
Biochemical Activity ◽  
Microtubule Associated Proteins ◽  
Associated Proteins

The Dis1/XMAP215 family of microtubule-associated proteins conserved from yeast to mammals is essential for cell division. XMAP215, the Xenopus member of this family, has been shown to stabilize microtubules in vitro, but other members of this family have not been biochemically characterized. Here we investigate the properties of the Saccharomyces cerevisiae homologue Stu2p in vitro. Surprisingly, Stu2p is a microtubule destabilizer that binds preferentially to microtubule plus ends. Quantitative analysis of microtubule dynamics suggests that Stu2p induces microtubule catastrophes by sterically interfering with tubulin addition to microtubule ends. These results reveal both a new biochemical activity for a Dis1/XMAP215 family member and a novel mechanism for microtubule destabilization.

scholarly journals Compartimentalized control of Cdk1 drives mitotic spindle assembly

2021 ◽  
Author(s):  
Angela Flavia Serpico ◽  
Francesco Febbraro ◽  
Caterina Pisauro ◽  
Domenico Grieco
Keyword(s):  
Mitotic Spindle ◽  
Kinase Activity ◽  
Spindle Assembly ◽  
Microtubule Associated Proteins ◽  
Associated Proteins ◽  
Microtubule Growth ◽  
Mitotic Spindle Assembly ◽  
Spindle Microtubules ◽  
Growth Promoting ◽  
Fraction Bound

During cell division, dramatic microtubular rearrangements driven by cyclin B-cdk1 (Cdk1) kinase activity mark mitosis onset leading to interphase cytoskeleton dissolution and mitotic spindle assembly. Once activated by Cdc25, that reverses inhibitory phosphorylation operated by Wee1/Myt1, Cdk1 clears the cytoplasm from microtubules by inhibiting microtubule associated proteins (MAPs) with microtubule growth-promoting properties. Nevertheless, some of these MAPs are required for spindle assembly, creating quite a conundrum. We show here that a Cdk1 fraction bound to spindle structures escaped Cdc25 action and remained inhibited by phosphorylation (i-Cdk1) in mitotic human cells. Loss or restoration of i-Cdk1 inhibited or promoted spindle assembly, respectively. Furthermore, polymerizing spindle microtubules fostered i-Cdk1 by aggregating with Wee1 and excluding Cdc25. Our data reveal that spindle assembly relies on compartimentalized control of Cdk1 activity.

Filaments and membranes in the mitotic apparatus

1983 ◽  
Vol 41 ◽  
pp. 544-547
Author(s):  
Kent McDonald
Keyword(s):  
Intermediate Filaments ◽  
Mitotic Spindle ◽  
Mitotic Apparatus ◽  
Light Microscope Level ◽  
Microtubule Associated Proteins ◽  
Recent Developments ◽  
Associated Proteins ◽  
Force Generator ◽  
Spindle Function ◽  
Antibody Technology

At the light microscope level the recent developments and interest in antibody technology have permitted the localization of certain non-microtubule proteins within the mitotic spindle, e.g., calmodulin, actin, intermediate filaments, protein kinases and various microtubule associated proteins. Also, the use of fluorescent probes like chlorotetracycline suggest the presence of membranes in the spindle. Localization of non-microtubule structures in the spindle at the EM level has been less rewarding. Some mitosis researchers, e.g., Rarer, have maintained that actin is involved in mitosis movements though the bulk of evidence argues against this interpretation. Others suggest that a microtrabecular network such as found in chromatophore granule movement might be a possible force generator but there is little evidence for or against this view. At the level of regulation of spindle function, Harris and more recently Hepler have argued for the importance of studying spindle membranes. Hepler also believes that membranes might play a structural or mechanical role in moving chromosomes.

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 KIF18A's neck linker permits navigation of microtubule-bound obstacles within the mitotic spindle

2019 ◽  
Vol 2 (1) ◽  
pp. e201800169 ◽  
Author(s):  
Heidi LH Malaby ◽  
Dominique V Lessard ◽  
Christopher L Berger ◽  
Jason Stumpff
Keyword(s):  
Single Molecule ◽  
Mitotic Spindle ◽  
Binding Proteins ◽  
Mitotic Chromosome ◽  
Chromosome Movement ◽  
Wild Type ◽  
Microtubule Associated Proteins ◽  
Chromosome Alignment ◽  
Associated Proteins ◽  
Fiber Dynamics

KIF18A (kinesin-8) is required for mammalian mitotic chromosome alignment. KIF18A confines chromosome movement to the mitotic spindle equator by accumulating at the plus-ends of kinetochore microtubule bundles (K-fibers), where it functions to suppress K-fiber dynamics. It is not understood how the motor accumulates at K-fiber plus-ends, a difficult feat requiring the motor to navigate protein dense microtubule tracks. Our data indicate that KIF18A's relatively long neck linker is required for the motor's accumulation at K-fiber plus-ends. Shorter neck linker (sNL) variants of KIF18A display a deficiency in accumulation at the ends of K-fibers at the center of the spindle. Depletion of K-fiber–binding proteins reduces the KIF18A sNL localization defect, whereas their overexpression reduces wild-type KIF18A's ability to accumulate on this same K-fiber subset. Furthermore, single-molecule assays indicate that KIF18A sNL motors are less proficient in navigating microtubules coated with microtubule-associated proteins. Taken together, these results support a model in which KIF18A's neck linker length permits efficient navigation of obstacles to reach K-fiber ends during mitosis.

Regulation of Microtubule Assembly: The View From The End

2000 ◽  
Vol 6 (S2) ◽  
pp. 80-81
Author(s):  
L. Cassimeris ◽  
C. Spittle ◽  
M. Kratzer
Keyword(s):  
Chromosome Segregation ◽  
Mitotic Spindle ◽  
Dynamic Instability ◽  
Intrinsic Property ◽  
Microtubule Dynamics ◽  
Microtubule Assembly ◽  
Chromosome Movement ◽  
Spindle Formation ◽  
Associated Proteins

The mitotic spindle is responsible for chromosome movement during mitosis. It is composed of a dynamic array of microtubules and associated proteins whose assembly and constant turnover are required for both spindle formation and chromosome movement. Because microtubule assembly and turnover are necessary for chromosome segregation, we are studying how cells regulate microtubule dynamics. Microtubules are polarized polymers composed of tubulin subunits; they assemble by a process of dynamic instability where individual microtubules exist in persistent phases of elongation or rapid shortening with abrupt transitions between these two states. The switch from elongation to shortening is termed catastrophe, and the switch from shortening to elongation, rescue. Although dynamic instability is an intrinsic property of the tubulin subunits, cells use associated proteins to both speed elongation (∼ 10 fold) and regulate transitions.The only protein isolated to date capable of promoting fast polymerization consistent with rates in vivo is XMAP215, a 215 kD protein from Xenopus eggs.

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