scholarly journals Tektin 2 is required for central spindle microtubule organization and the completion of cytokinesis

2008 ◽  
Vol 181 (4) ◽  
pp. 595-603 ◽  
Author(s):  
Thomas M. Durcan ◽  
Elizabeth S. Halpin ◽  
Trisha Rao ◽  
Nicholas S. Collins ◽  
Emily K. Tribble ◽  
...  
Keyword(s):  
Cell Formation ◽  
Binucleate Cell ◽  
Aurora B ◽  
Microtubule Organization ◽  
Spindle Microtubule ◽  
Central Spindle ◽  
Daughter Cells ◽  
Spindle Poles ◽  
Spindle Midzone ◽  
Interfering Rna

During anaphase, the nonkinetochore microtubules in the spindle midzone become compacted into the central spindle, a structure which is required to both initiate and complete cytokinesis. We show that Tektin 2 (Tek2) associates with the spindle poles throughout mitosis, organizes the spindle midzone microtubules during anaphase, and assembles into the midbody matrix surrounding the compacted midzone microtubules during cytokinesis. Tek2 small interfering RNA (siRNA) disrupts central spindle organization and proper localization of MKLP1, PRC1, and Aurora B to the midzone and prevents the formation of a midbody matrix. Video microscopy revealed that loss of Tek2 results in binucleate cell formation by aberrant fusion of daughter cells after cytokinesis. Although a myosin II inhibitor, blebbistatin, prevents actin-myosin contractility, the microtubules of the central spindle are compacted. Strikingly, Tek2 siRNA abolishes this actin-myosin–independent midzone microtubule compaction. Thus, Tek2-dependent organization of the central spindle during anaphase is essential for proper midbody formation and the segregation of daughter cells after cytokinesis.

scholarly journals Cep57 Protein Is Required for Cytokinesis by Facilitating Central Spindle Microtubule Organization

2013 ◽  
Vol 288 (20) ◽  
pp. 14384-14390 ◽  
Author(s):  
Runsheng He ◽  
Qixi Wu ◽  
Haining Zhou ◽  
Ning Huang ◽  
Jianguo Chen ◽  
...  
Keyword(s):  
Genetic Material ◽  
Microtubule Assembly ◽  
Aurora B ◽  
Microtubule Organization ◽  
Spindle Microtubule ◽  
Central Spindle ◽  
Daughter Cells ◽  
Binuclear Cells ◽  
A Cell ◽  
Centrosome Protein

Cytokinesis is the final stage of cell division in which the cytoplasm of a cell is divided into two daughter cells after the segregation of genetic material, and the central spindle and midbody are considered to be the essential structures required for the initiation and completion of cytokinesis. Here, we determined that the centrosome protein Cep57, which is localized to the central spindle and midbody, acts as a spindle organizer and is required for cytokinesis. Depletion of Cep57 disrupted microtubule assembly of the central spindle and further led to abnormal midbody localization of MKLP1, Plk1, and Aurora B, which resulted in cytokinesis failure and the formation of binuclear cells. Furthermore, we found that Cep57 directly recruited Tektin 1 to the midbody matrix to regulate microtubule organization. Thus, our data reveal that Cep57 is essential for cytokinesis via regulation of central spindle assembly and formation of the midbody.

scholarly journals Leukemia-associated RhoGEF (LARG) is a novel RhoGEF in cytokinesis and required for the proper completion of abscission

2013 ◽  
Vol 24 (18) ◽  
pp. 2785-2794 ◽  
Author(s):  
Matthew K. Martz ◽  
Elda Grabocka ◽  
Neil Beeharry ◽  
Timothy J. Yen ◽  
Philip B. Wedegaertner
Keyword(s):  
Small Interfering Rna ◽  
Live Cell Imaging ◽  
Cell Imaging ◽  
Aurora B ◽  
Intercellular Bridge ◽  
Concerted Effort ◽  
Central Spindle ◽  
Daughter Cells ◽  
Spindle Poles ◽  
Interfering Rna

Proper completion of mitosis requires the concerted effort of multiple RhoGEFs. Here we show that leukemia-associated RhoGEF (LARG), a RhoA-specific RGS-RhoGEF, is required for abscission, the final stage of cytokinesis, in which the intercellular membrane is cleaved between daughter cells. LARG colocalizes with α-tubulin at the spindle poles before localizing to the central spindle. During cytokinesis, LARG is condensed in the midbody, where it colocalizes with RhoA. HeLa cells depleted of LARG display apoptosis during cytokinesis with unresolved intercellular bridges, and rescue experiments show that expression of small interfering RNA–resistant LARG prevents this apoptosis. Moreover, live cell imaging of LARG-depleted cells reveals greatly delayed fission kinetics in abscission in which a population of cells with persistent bridges undergoes apoptosis; however, the delayed fission kinetics is rescued by Aurora-B inhibition. The formation of a Flemming body and thinning of microtubules in the intercellular bridge of cells depleted of LARG is consistent with a defect in late cytokinesis, just before the abscission event. In contrast to studies of other RhoGEFs, particularly Ect2 and GEF-H1, LARG depletion does not result in cytokinetic furrow regression nor does it affect internal mitotic timing. These results show that LARG is a novel and temporally distinct RhoGEF required for completion of abscission.

scholarly journals Annexin 11 is required for midbody formation and completion of the terminal phase of cytokinesis

2004 ◽  
Vol 165 (6) ◽  
pp. 813-822 ◽  
Author(s):  
Alejandra Tomas ◽  
Clare Futter ◽  
Stephen E. Moss
Keyword(s):  
Cell Cycle Progression ◽  
Time Lapse ◽  
Terminal Phase ◽  
Aurora B ◽  
Cycle Progression ◽  
Daughter Cells ◽  
Time Lapse Microscopy ◽  
Spindle Poles ◽  
Spindle Midzone ◽  
Late Telophase

Annexins are Ca2+-binding, membrane-fusogenic proteins with diverse but poorly understood functions. Here, we show that during cell cycle progression annexin 11 translocates from the nucleus to the spindle poles in metaphase and to the spindle midzone in anaphase. Annexin 11 is recruited to the midbody in late telophase, where it forms part of the detergent-resistant matrix that also contains CHO1. To investigate the significance of these observations, we used RNA interference to deplete cells of annexin 11. A combination of confocal and video time-lapse microscopy revealed that cells lacking annexin 11 fail to establish a functional midbody. Instead, daughter cells remain connected by intercellular bridges that contain bundled microtubules and cytoplasmic organelles but exclude normal midbody components such as MKLP1 and Aurora B. Annexin 11–depleted cells failed to complete cytokinesis and died by apoptosis. These findings demonstrate an essential role for annexin 11 in the terminal phase of cytokinesis.

scholarly journals A Small C-Terminal Sequence of Aurora B Is Responsible for Localization and Function

2005 ◽  
Vol 16 (1) ◽  
pp. 292-305 ◽  
Author(s):  
Laetitia Scrittori ◽  
Dimitrios A. Skoufias ◽  
Fabienne Hans ◽  
Véronique Gerson ◽  
Paolo Sassone-Corsi ◽  
...  
Keyword(s):  
Aurora B ◽  
Aurora A ◽  
Terminal Sequence ◽  
Function Test ◽  
Spindle Midzone ◽  
Spindle Microtubules ◽  
And Function ◽  
Interfering Rna ◽  
Sequence Requirements

Aurora B, a protein kinase required in mitosis, localizes to inner centromeres at metaphase and the spindle midzone in anaphase and is required for proper chromosome segregation and cytokinesis. Aurora A, a paralogue of Aurora B, localizes instead to centrosomes and spindle microtubules. Except for distinct N termini, Aurora B and Aurora A have highly similar sequences. We have combined small interfering RNA (siRNA) ablation of Aurora B with overexpression of truncation mutants to investigate the role of Aurora B sequence in its function. Reintroduction of Aurora B during siRNA treatment restored its localization and function. This permitted a restoration of function test to determine the sequence requirements for Aurora B targeting and function. Using this rescue protocol, neither N-terminal truncation of Aurora B unique sequence nor substitution with Aurora A N-terminal sequence affected Aurora B localization or function. Truncation of unique Aurora B C-terminal sequence from terminal residue 344 to residue 333 was without effect, but truncation to 326 abolished localization and function. Deletion of residues 326-333 completely abolished localization and blocked cells at prometaphase, establishing this sequence as critical to Aurora B function. Our findings thus establish a small sequence as essential for the distinct localization and function of Aurora B.

scholarly journals Aurora B and Kif2A control microtubule length for assembly of a functional central spindle during anaphase

2013 ◽  
Vol 202 (4) ◽  
pp. 623-636 ◽  
Author(s):  
Ryota Uehara ◽  
Yuki Tsukada ◽  
Tomoko Kamasaki ◽  
Ina Poser ◽  
Kinya Yoda ◽  
...  
Keyword(s):  
Experimental Data ◽  
Steady State ◽  
Cell Division ◽  
Self Organization ◽  
Aurora B ◽  
Spindle Formation ◽  
Central Spindle ◽  
Model Simulations ◽  
Spindle Midzone ◽  
Organization Mechanism

The central spindle is built during anaphase by coupling antiparallel microtubules (MTs) at a central overlap zone, which provides a signaling scaffold for the regulation of cytokinesis. The mechanisms underlying central spindle morphogenesis are still poorly understood. In this paper, we show that the MT depolymerase Kif2A controls the length and alignment of central spindle MTs through depolymerization at their minus ends. The distribution of Kif2A was limited to the distal ends of the central spindle through Aurora B–dependent phosphorylation and exclusion from the spindle midzone. Overactivation or inhibition of Kif2A affected interchromosomal MT length and disorganized the central spindle, resulting in uncoordinated cell division. Experimental data and model simulations suggest that the steady-state length of the central spindle and its symmetric position between segregating chromosomes are predominantly determined by the Aurora B activity gradient. On the basis of these results, we propose a robust self-organization mechanism for central spindle formation.

scholarly journals Ablation of PRC1 by Small Interfering RNA Demonstrates that Cytokinetic Abscission Requires a Central Spindle Bundle in Mammalian Cells, whereas Completion of Furrowing Does Not

2005 ◽  
Vol 16 (3) ◽  
pp. 1043-1055 ◽  
Author(s):  
Cristiana Mollinari ◽  
Jean-Philippe Kleman ◽  
Yasmina Saoudi ◽  
Sandra A. Jablonski ◽  
Julien Perard ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Time Lapse ◽  
Cell Cortex ◽  
Central Spindle ◽  
Spatial Regulation ◽  
Daughter Cells ◽  
Cell Cleavage ◽  
Temporal And Spatial ◽  
Interfering Rna ◽  
Passenger Proteins

The temporal and spatial regulation of cytokinesis requires an interaction between the anaphase mitotic spindle and the cell cortex. However, the relative roles of the spindle asters or the central spindle bundle are not clear in mammalian cells. The central spindle normally serves as a platform to localize key regulators of cell cleavage, including passenger proteins. Using time-lapse and immunofluorescence analysis, we have addressed the consequences of eliminating the central spindle by ablation of PRC1, a microtubule bundling protein that is critical to the formation of the central spindle. Without a central spindle, the asters guide the equatorial cortical accumulation of anillin and actin, and of the passenger proteins, which organize into a subcortical ring in anaphase. Furrowing goes to completion, but abscission to create two daughter cells fails. We conclude the central spindle bundle is required for abscission but not for furrowing in mammalian cells.

scholarly journals Kinesin-like Protein CHO1 Is Required for the Formation of Midbody Matrix and the Completion of Cytokinesis in Mammalian Cells

2002 ◽  
Vol 13 (6) ◽  
pp. 1832-1845 ◽  
Author(s):  
Jurgita Matuliene ◽  
Ryoko Kuriyama
Keyword(s):  
Binding Site ◽  
Mammalian Cells ◽  
Cytoplasmic Bridge ◽  
Central Spindle ◽  
Atp Binding Site ◽  
Daughter Cells ◽  
Dividing Cells ◽  
Spindle Midzone ◽  
Segregation Of Chromosomes

CHO1 is a mammalian kinesin-like motor protein of the MKLP1 subfamily. It associates with the spindle midzone during anaphase and concentrates to a midbody matrix during cytokinesis. CHO1 was originally implicated in karyokinesis, but the invertebrate homologues of CHO1 were shown to function in the midzone formation and cytokinesis. To analyze the role of the protein in mammalian cells, we mutated the ATP-binding site of CHO1 and expressed it in CHO cells. Mutant protein (CHO1F′) was able to interact with microtubules via ATP-independent microtubule-binding site(s) but failed to accumulate at the midline of the central spindle and affected the localization of endogenous CHO1. Although the segregation of chromosomes, the bundling of midzone microtubules, and the initiation of cytokinesis proceeded normally in CHO1F′-expressing cells, the completion of cytokinesis was inhibited. Daughter cells were frequently entering interphase while connected by a microtubule-containing cytoplasmic bridge from which the dense midbody matrix was missing. Depletion of endogenous CHO1 via RNA-mediated interference also affected the formation of midbody matrix in dividing cells, caused the disorganization of midzone microtubules, and resulted in abortive cytokinesis. Thus, CHO1 may not be required for karyokinesis, but it is essential for the proper midzone/midbody formation and cytokinesis in mammalian cells.

scholarly journals The Drosophila Protein Asp Is Involved in Microtubule Organization during Spindle Formation and Cytokinesis

2001 ◽  
Vol 153 (4) ◽  
pp. 637-648 ◽  
Author(s):  
James G. Wakefield ◽  
Silvia Bonaccorsi ◽  
Maurizio Gatti
Keyword(s):  
Colchicine Treatment ◽  
Spindle Pole ◽  
Mitotic Spindles ◽  
Microtubule Nucleation ◽  
Phenotypic Analysis ◽  
Microtubule Organization ◽  
Central Spindle ◽  
Spindle Poles ◽  
Drosophila Protein

Abnormal spindle (Asp) is a 220-kD microtubule-associated protein from Drosophila that has been suggested to be involved in microtubule nucleation from the centrosome. Here, we show that Asp is enriched at the poles of meiotic and mitotic spindles and localizes to the minus ends of central spindle microtubules. Localization to these structures is independent of a functional centrosome. Moreover, colchicine treatment disrupts Asp localization to the centrosome, indicating that Asp is not an integral centrosomal protein. In both meiotic and mitotic divisions of asp mutants, microtubule nucleation occurs from the centrosome, and γ-tubulin localizes correctly. However, spindle pole focusing and organization are severely affected. By examining cells that carry mutations both in asp and in asterless, a gene required for centrosome function, we have determined the role of Asp in the absence of centrosomes. Phenotypic analysis of these double mutants shows that Asp is required for the aggregation of microtubules into focused spindle poles, reinforcing the conclusion that its function at the spindle poles is independent of any putative role in microtubule nucleation. Our data also suggest that Asp has a role in the formation of the central spindle. The inability of asp mutants to correctly organize the central spindle leads to disruption of the contractile ring machinery and failure in cytokinesis.

scholarly journals Aurora A promotes chromosome congression by activating the condensin-dependent pool of KIF4A

2019 ◽  
Vol 219 (2) ◽  
Author(s):  
Elena Poser ◽  
Renaud Caous ◽  
Ulrike Gruneberg ◽  
Francis A. Barr
Keyword(s):  
Metaphase Plate ◽  
Aurora Kinase ◽  
Chromosome Condensation ◽  
Aurora B ◽  
Aurora A ◽  
Specific Point ◽  
Aurora Kinases ◽  
Central Spindle ◽  
Spindle Poles ◽  
Chromosome Congression

Aurora kinases create phosphorylation gradients within the spindle during prometaphase and anaphase, thereby locally regulating factors that promote spindle organization, chromosome condensation and movement, and cytokinesis. We show that one such factor is the kinesin KIF4A, which is present along the chromosome axes throughout mitosis and the central spindle in anaphase. These two pools of KIF4A depend on condensin I and PRC1, respectively. Previous work has shown KIF4A is activated by Aurora B at the anaphase central spindle. However, whether or not chromosome-associated KIF4A bound to condensin I is regulated by Aurora kinases remain unclear. To determine the roles of the two different pools of KIF4A, we generated specific point mutants that are unable to interact with either condensin I or PRC1 or are deficient for Aurora kinase regulation. By analyzing these mutants, we show that Aurora A phosphorylates the condensin I–dependent pool of KIF4A and thus actively promotes chromosome congression from the spindle poles to the metaphase plate.

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