scholarly journals Condensin Regulates the Stiffness of Vertebrate Centromeres

2009 ◽  
Vol 20 (9) ◽  
pp. 2371-2380 ◽  
Author(s):  
Susana A. Ribeiro ◽  
Jesse C. Gatlin ◽  
Yimin Dong ◽  
Ajit Joglekar ◽  
Lisa Cameron ◽  
...  
Keyword(s):  
Mitotic Spindle ◽  
Metaphase Plate ◽  
Centromeric Chromatin ◽  
Anaphase Onset ◽  
Sister Kinetochore ◽  
Pulling Forces ◽  
Important Regulator ◽  
Chromatin Folding ◽  
And Function ◽  
Rest Length

When chromosomes are aligned and bioriented at metaphase, the elastic stretch of centromeric chromatin opposes pulling forces exerted on sister kinetochores by the mitotic spindle. Here we show that condensin ATPase activity is an important regulator of centromere stiffness and function. Condensin depletion decreases the stiffness of centromeric chromatin by 50% when pulling forces are applied to kinetochores. However, condensin is dispensable for the normal level of compaction (rest length) of centromeres, which probably depends on other factors that control higher-order chromatin folding. Kinetochores also do not require condensin for their structure or motility. Loss of stiffness caused by condensin-depletion produces abnormal uncoordinated sister kinetochore movements, leads to an increase in Mad2(+) kinetochores near the metaphase plate and delays anaphase onset.

scholarly journals Merotelic kinetochores in mammalian tissue cells

2005 ◽  
Vol 360 (1455) ◽  
pp. 553-568 ◽  
Author(s):  
E.D Salmon ◽  
D Cimini ◽  
L.A Cameron ◽  
J.G DeLuca
Keyword(s):  
Mitotic Spindle ◽  
Binding Sites ◽  
Metaphase Plate ◽  
Mammalian Tissue ◽  
Anaphase Onset ◽  
Sister Chromatids ◽  
Pulling Forces ◽  
Tissue Cells ◽  
Early Mitosis

Merotelic kinetochore attachment is a major source of aneuploidy in mammalian tissue cells in culture. Mammalian kinetochores typically have binding sites for about 20–25 kinetochore microtubules. In prometaphase, kinetochores become merotelic if they attach to microtubules from opposite poles rather than to just one pole as normally occurs. Merotelic attachments support chromosome bi-orientation and alignment near the metaphase plate and they are not detected by the mitotic spindle checkpoint. At anaphase onset, sister chromatids separate, but a chromatid with a merotelic kinetochore may not be segregated correctly, and may lag near the spindle equator because of pulling forces toward opposite poles, or move in the direction of the wrong pole. Correction mechanisms are important for preventing segregation errors. There are probably more than 100 times as many PtK1 tissue cells with merotelic kinetochores in early mitosis, and about 16 times as many entering anaphase as the 1% of cells with lagging chromosomes seen in late anaphase. The role of spindle mechanics and potential functions of the Ndc80/Nuf2 protein complex at the kinetochore/microtubule interface is discussed for two correction mechanisms: one that functions before anaphase to reduce the number of kinetochore microtubules to the wrong pole, and one that functions after anaphase onset to move merotelic kinetochores based on the ratio of kinetochore microtubules to the correct versus incorrect pole.

Determinants of Drosophila zw10 protein localization and function

1994 ◽  
Vol 107 (4) ◽  
pp. 785-798 ◽  
Author(s):  
B.C. Williams ◽  
M.L. Goldberg
Keyword(s):  
Chromosome Segregation ◽  
Sister Chromatid ◽  
Protein Localization ◽  
Metaphase Plate ◽  
Obvious Effect ◽  
Anaphase Onset ◽  
Protein Functions ◽  
Kinetochore Region ◽  
And Function ◽  
Feedback Pathway

We have examined several issues concerning how the Drosophila l(1)zw10 gene product functions to ensure proper chromosome segregation. (a) We have found that in zw10 mutant embryos and larval neuroblasts, absence of the zw10 protein has no obvious effect on either the congression of chromosomes to the metaphase plate or the morphology of the metaphase spindle, although many aberrations are observed subsequently in anaphase. This suggests that activity of the zw10 protein becomes essential at anaphase onset, a time at which the zw10 protein is redistributed to the kinetochore region of the chromosomes. (b) The zw10 protein appears to bind to kinetochores in mitotically arrested cells, eventually accumulating to high levels within the chromosome mass. Our results imply that zw10 may act as part of a novel feedback pathway that normally renders sister chromatid separation dependent upon spindle integrity. (c) The localization of zw10 protein is altered by two mitotic mutations, rough deal and abnormal anaphase resolution, that specifically disrupt anaphase. These findings indicate that the zw10 protein functions as part of a multicomponent mechanism ensuring proper chromosome segregation at the beginning of anaphase.

scholarly journals The ESCRT protein Chmp4c regulates mitotic spindle checkpoint signaling

2018 ◽  
Vol 217 (3) ◽  
pp. 861-876 ◽  
Author(s):  
Eleni Petsalaki ◽  
Maria Dandoulaki ◽  
George Zachos
Keyword(s):  
Mitotic Spindle ◽  
Spindle Checkpoint ◽  
Metaphase Plate ◽  
Membrane Remodeling ◽  
Mitotic Progression ◽  
Mitotic Spindle Checkpoint ◽  
Checkpoint Signaling ◽  
Checkpoint Proteins ◽  
Endosomal Sorting ◽  
Anaphase Onset

The mitotic spindle checkpoint delays anaphase onset in the presence of unattached kinetochores, and efficient checkpoint signaling requires kinetochore localization of the Rod–ZW10–Zwilch (RZZ) complex. In the present study, we show that human Chmp4c, a protein involved in membrane remodeling, localizes to kinetochores in prometaphase but is reduced in chromosomes aligned at the metaphase plate. Chmp4c promotes stable kinetochore–microtubule attachments and is required for proper mitotic progression, faithful chromosome alignment, and segregation. Depletion of Chmp4c diminishes localization of RZZ and Mad1-Mad2 checkpoint proteins to prometaphase kinetochores and impairs mitotic arrest when microtubules are depolymerized by nocodazole. Furthermore, Chmp4c binds to ZW10 through a small C-terminal region, and constitutive Chmp4c kinetochore targeting causes a ZW10-dependent checkpoint metaphase arrest. In addition, Chmp4c spindle functions do not require endosomal sorting complex required for transport–dependent membrane remodeling. These results show that Chmp4c regulates the mitotic spindle checkpoint by promoting localization of the RZZ complex to unattached kinetochores.

scholarly journals Checkpoint Protein BubR1 Acts Synergistically with Mad2 to Inhibit Anaphase-promoting Complex

2002 ◽  
Vol 13 (3) ◽  
pp. 755-766 ◽  
Author(s):  
Guowei Fang
Keyword(s):  
Ubiquitin Ligase ◽  
Mitotic Spindle ◽  
Spindle Assembly Checkpoint ◽  
Kinase Activity ◽  
Metaphase Plate ◽  
Anaphase Promoting Complex ◽  
Checkpoint Control ◽  
Sister Chromatids ◽  
Direct Binding ◽  
And Function

The spindle assembly checkpoint monitors the attachment of kinetochores to the mitotic spindle and the tension exerted on kinetochores by microtubules and delays the onset of anaphase until all the chromosomes are aligned at the metaphase plate. The target of the checkpoint control is the anaphase-promoting complex (APC)/cyclosome, a ubiquitin ligase whose activation by Cdc20 is required for separation of sister chromatids. In response to activation of the checkpoint, Mad2 binds to and inhibits Cdc20-APC. I show herein that in checkpoint-arrested cells, human Cdc20 forms two separate, inactive complexes, a lower affinity complex with Mad2 and a higher affinity complex with BubR1. Purified BubR1 binds to recombinant Cdc20 and this interaction is direct. Binding of BubR1 to Cdc20 inhibits activation of APC and this inhibition is independent of its kinase activity. Quantitative analysis indicates that BubR1 is 12-fold more potent than Mad2 as an inhibitor of Cdc20. Although at high protein concentrations BubR1 and Mad2 each is sufficient to inhibit Cdc20, BubR1 and Mad2 mutually promote each other's binding to Cdc20 and function synergistically at physiological concentrations to quantitatively inhibit Cdc20-APC. Thus, BubR1 and Mad2 act cooperatively to prevent premature separation of sister chromatids by directly inhibiting APC.

scholarly journals Kinetochore motility after severing between sister centromeres using laser microsurgery: evidence that kinetochore directional instability and position is regulated by tension

1995 ◽  
Vol 108 (7) ◽  
pp. 2537-2548 ◽  
Author(s):  
R.V. Skibbens ◽  
C.L. Rieder ◽  
E.D. Salmon
Keyword(s):  
Laser Ablation ◽  
Constant Velocity ◽  
Somatic Cells ◽  
Metaphase Plate ◽  
Fundamental Property ◽  
The Other ◽  
Centromeric Chromatin ◽  
Laser Microbeam ◽  
Differential Movement ◽  
Sister Kinetochore

During mitosis in vertebrate somatic cells, the single attached kinetochore on a mono-oriented chromosome exhibits directional instability: abruptly and independently switching between constant velocity poleward and away from the pole motility states. When the non-attached sister becomes attached to the spindle (chromosome bi-orientation), the motility of the sister kinetochores becomes highly coordinated, one moving poleward while the other moves away from the pole, allowing chromosomes to congress to the spindle equator. In our kinetochore-tensiometer model, we hypothesized that this coordinated behavior is regulated by tension across the centromere produced by kinetochore movement relative to the sister kinetochore and bulk of the chromosome arms. To test this model, we severed or severely weakened the centromeric chromatin between sister kinetochores on bi-oriented newt lung cell chromosomes with a laser microbeam. This procedure converted a pair of tightly linked sister kinetochores into two mono-oriented single kinetochore-chromatin fragments that were tethered to their chromosome arms by thin compliant chromatin strands. These single kinetochore-chromatin fragments moved substantial distances off the metaphase plate, stretching their chromatin strands, before the durations of poleward and away from the pole movement again became similar. In contrast, the severed arms remained at or moved closer to the spindle equator. The poleward and away from the pole velocities of single kinetochore-chromatin fragments in prometaphase were typical of velocities exhibited by sister kinetochores on intact chromosomes from prometaphase through midanaphase A. However, severing the chromatin between sister kinetochores uncoupled the normally coordinated motility of sister kinetochores. Laser ablation also uncoupled the motilities of the single kinetochore fragments from the bulk of the arms. These results reveal that kinetochore directional instability is a fundamental property of the kinetochore and that the motilities of sister kinetochores are coordinated during congression by a stiff centromere linkage. We conclude that kinetochores act as tensiometers that sense centromere tension generated by differential movement of sister kinetochores and their chromosome arms to control switching between constant velocity P and AP motility states.

scholarly journals Merotelic Kinetochore Orientation Is a Major Mechanism of Aneuploidy in Mitotic Mammalian Tissue Cells

2001 ◽  
Vol 153 (3) ◽  
pp. 517-528 ◽  
Author(s):  
Daniela Cimini ◽  
Bonnie Howell ◽  
Paul Maddox ◽  
Alexey Khodjakov ◽  
Francesca Degrassi ◽  
...  
Keyword(s):  
Mitotic Spindle ◽  
Cultured Cells ◽  
Spindle Checkpoint ◽  
Mitotic Spindle Checkpoint ◽  
Major Mechanism ◽  
Confocal Immunofluorescence Microscopy ◽  
Anaphase Onset ◽  
Pulling Forces ◽  
Lagging Chromosome ◽  
Single Chromatid

In mitotic cells, an error in chromosome segregation occurs when a chromosome is left near the spindle equator after anaphase onset (lagging chromosome). In PtK1 cells, we found 1.16% of untreated anaphase cells exhibiting lagging chromosomes at the spindle equator, and this percentage was enhanced to 17.55% after a mitotic block with 2 μM nocodazole. A lagging chromosome seen during anaphase in control or nocodazole-treated cells was found by confocal immunofluorescence microscopy to be a single chromatid with its kinetochore attached to kinetochore microtubule bundles extending toward opposite poles. This merotelic orientation was verified by electron microscopy. The single kinetochores of lagging chromosomes in anaphase were stretched laterally (1.2–5.6-fold) in the directions of their kinetochore microtubules, indicating that they were not able to achieve anaphase poleward movement because of pulling forces toward opposite poles. They also had inactivated mitotic spindle checkpoint activities since they did not label with either Mad2 or 3F3/2 antibodies. Thus, for mammalian cultured cells, kinetochore merotelic orientation is a major mechanism of aneuploidy not detected by the mitotic spindle checkpoint. The expanded and curved crescent morphology exhibited by kinetochores during nocodazole treatment may promote the high incidence of kinetochore merotelic orientation that occurs after nocodazole washout.

scholarly journals Chromosome bi-orientation on the mitotic spindle

2005 ◽  
Vol 360 (1455) ◽  
pp. 581-589 ◽  
Author(s):  
Tomoyuki U Tanaka
Keyword(s):  
Protein Kinase ◽  
Chromosome Segregation ◽  
Mitotic Spindle ◽  
Budding Yeast ◽  
Aurora B ◽  
Cohesin Complex ◽  
Spindle Poles ◽  
Anaphase Onset ◽  
Sister Kinetochore ◽  
Dependent Mechanism

For proper chromosome segregation, sister kinetochores must attach to microtubules extending from opposite spindle poles prior to anaphase onset. This state is called sister kinetochore bi-orientation or chromosome bi-orientation. The mechanism ensuring chromosome bi-orientation lies at the heart of chromosome segregation, but is still poorly understood. Recent evidence suggests that mal-oriented kinetochore-to-pole connections are corrected in a tension-dependent mechanism. The cohesin complex and the Ipl1/Aurora B protein kinase seem to be key regulators for this correction. In this article, I discuss how cells ensure sister kinetochore bi-orientation for all chromosomes, mainly focusing on our recent findings in budding yeast.

scholarly journals Automated mitotic spindle tracking suggests a link between spindle dynamics, spindle orientation, and anaphase onset in epithelial cells

2017 ◽  
Vol 28 (6) ◽  
pp. 746-759 ◽  
Author(s):  
Matthew E. Larson ◽  
William M. Bement
Keyword(s):  
Mitotic Spindle ◽  
Spindle Positioning ◽  
Tissue Organization ◽  
Anaphase Onset ◽  
Spindle Dynamics ◽  
And Function ◽  
Spindle Position ◽  
The Relationship ◽  
Rotational Oscillations ◽  
Final Orientation

Proper spindle positioning at anaphase onset is essential for normal tissue organization and function. Here we develop automated spindle-tracking software and apply it to characterize mitotic spindle dynamics in the Xenopus laevis embryonic epithelium. We find that metaphase spindles first undergo a sustained rotation that brings them on-axis with their final orientation. This sustained rotation is followed by a set of striking stereotyped rotational oscillations that bring the spindle into near contact with the cortex and then move it rapidly away from the cortex. These oscillations begin to subside soon before anaphase onset. Metrics extracted from the automatically tracked spindles indicate that final spindle position is determined largely by cell morphology and that spindles consistently center themselves in the XY-plane before anaphase onset. Finally, analysis of the relationship between spindle oscillations and spindle position relative to the cortex reveals an association between cortical contact and anaphase onset. We conclude that metaphase spindles in epithelia engage in a stereotyped “dance,” that this dance culminates in proper spindle positioning and orientation, and that completion of the dance is linked to anaphase onset.

scholarly journals DNA Topoisomerase II Is a Determinant of the Tensile Properties of Yeast Centromeric Chromatin and the Tension Checkpoint

2008 ◽  
Vol 19 (10) ◽  
pp. 4421-4433 ◽  
Author(s):  
Tariq H. Warsi ◽  
Michelle S. Navarro ◽  
Jeff Bachant
Keyword(s):  
Error Correction ◽  
Tensile Properties ◽  
Mitotic Spindle ◽  
Topoisomerase Ii ◽  
Dna Topoisomerase Ii ◽  
Mechanical Tension ◽  
Dna Topoisomerase ◽  
Centromeric Chromatin ◽  
Sister Chromatids ◽  
Sister Kinetochore

Centromeric (CEN) chromatin is placed under mechanical tension and stretches as kinetochores biorient on the mitotic spindle. This deformation could conceivably provide a readout of biorientation to error correction mechanisms that monitor kinetochore–spindle interactions, but whether CEN chromatin acts in a tensiometer capacity is unresolved. Here, we report observations linking yeast Topoisomerase II (Top2) to both CEN mechanics and assessment of interkinetochore tension. First, in top2-4 and sumoylation-resistant top2-SNM mutants CEN chromatin stretches extensively during biorientation, resulting in increased sister kinetochore separation and preanaphase spindle extension. Our data indicate increased CEN stretching corresponds with alterations to CEN topology induced in response to tension. Second, Top2 potentiates aspects of the tension checkpoint. Mutations affecting the Mtw1 kinetochore protein activate Ipl1 kinase to detach kinetochores and induce spindle checkpoint arrest. In mtw1top2-4 and mtw1top2-SNM mutants, however, kinetochores are resistant to detachment and checkpoint arrest is attenuated. For top2-SNM cells, CEN stretching and checkpoint attenuation occur even in the absence of catenation linking sister chromatids. In sum, Top2 seems to play a novel role in CEN compaction that is distinct from decatenation. Perturbations to this function may allow weakened kinetochores to stretch CENs in a manner that mimics tension or evades Ipl1 surveillance.

scholarly journals The astrin–kinastrin/SKAP complex localizes to microtubule plus ends and facilitates chromosome alignment

2011 ◽  
Vol 192 (6) ◽  
pp. 959-968 ◽  
Author(s):  
Anja K. Dunsch ◽  
Emily Linnane ◽  
Francis A. Barr ◽  
Ulrike Gruneberg
Keyword(s):  
Light Chain ◽  
Mitotic Spindle ◽  
Metaphase Plate ◽  
Dynein Light Chain ◽  
Interacting Protein ◽  
Anaphase Onset ◽  
Chromatid Cohesion ◽  
Interaction Partners ◽  
Chromosome Alignment ◽  
Correct Alignment

Astrin is a mitotic spindle–associated protein required for the correct alignment of all chromosomes at the metaphase plate. Astrin depletion delays chromosome alignment and causes the loss of normal spindle architecture and sister chromatid cohesion before anaphase onset. Here we describe an astrin complex containing kinastrin/SKAP, a novel kinetochore and mitotic spindle protein, and three minor interaction partners: dynein light chain, Plk1, and Sgo2. Kinastrin is the major astrin-interacting protein in mitotic cells, and is required for astrin targeting to microtubule plus ends proximal to the plus tip tracking protein EB1. Cells overexpressing or depleted of kinastrin mislocalize astrin and show the same mitotic defects as astrin-depleted cells. Importantly, astrin fails to localize to and track microtubule plus ends in cells depleted of or overexpressing kinastrin. These findings suggest that microtubule plus end targeting of astrin is required for normal spindle architecture and chromosome alignment, and that perturbations of this pathway result in delayed mitosis and nonphysiological separase activation.

Sign in / Sign up

Export Citation Format

Share Document