-
ELife Aug 2019According to the prevailing 'clock' model, chromosome decondensation and nuclear envelope reformation when cells exit mitosis are byproducts of Cdk1 inactivation at the...
According to the prevailing 'clock' model, chromosome decondensation and nuclear envelope reformation when cells exit mitosis are byproducts of Cdk1 inactivation at the metaphase-anaphase transition, controlled by the spindle assembly checkpoint. However, mitotic exit was recently shown to be a function of chromosome separation during anaphase, assisted by a midzone Aurora B phosphorylation gradient - the 'ruler' model. Here we found that Cdk1 remains active during anaphase due to ongoing APC/C- and APC/C-mediated degradation of B-type Cyclins in and human cells. Failure to degrade B-type Cyclins during anaphase prevented mitotic exit in a Cdk1-dependent manner. Cyclin B1-Cdk1 localized at the spindle midzone in an Aurora B-dependent manner, with incompletely separated chromosomes showing the highest Cdk1 activity. Slowing down anaphase chromosome motion delayed Cyclin B1 degradation and mitotic exit in an Aurora B-dependent manner. Thus, a crosstalk between molecular 'rulers' and 'clocks' licenses mitotic exit only after proper chromosome separation.
Topics: Anaphase; Animals; Aurora Kinase B; CDC2 Protein Kinase; Cell Line; Cyclin B1; Drosophila; Drosophila Proteins; Humans; Proteolysis; Spatio-Temporal Analysis
PubMed: 31424385
DOI: 10.7554/eLife.47646 -
Communications Biology Jan 2023Profilin 1-encoded by PFN1-is a small actin-binding protein with a tumour suppressive role in various adenocarcinomas and pagetic osteosarcomas. However, its...
Profilin 1-encoded by PFN1-is a small actin-binding protein with a tumour suppressive role in various adenocarcinomas and pagetic osteosarcomas. However, its contribution to tumour development is not fully understood. Using fix and live cell imaging, we report that Profilin 1 inactivation results in multiple mitotic defects, manifested prominently by anaphase bridges, multipolar spindles, misaligned and lagging chromosomes, and cytokinesis failures. Accordingly, next-generation sequencing technologies highlighted that Profilin 1 knock-out cells display extensive copy-number alterations, which are associated with complex genome rearrangements and chromothripsis events in primary pagetic osteosarcomas with Profilin 1 inactivation. Mechanistically, we show that Profilin 1 is recruited to the spindle midzone at anaphase, and its deficiency reduces the supply of actin filaments to the cleavage furrow during cytokinesis. The mitotic defects are also observed in mouse embryonic fibroblasts and mesenchymal cells deriving from a newly generated knock-in mouse model harbouring a Pfn1 loss-of-function mutation. Furthermore, nuclear atypia is also detected in histological sections of mutant femurs. Thus, our results indicate that Profilin 1 has a role in regulating cell division, and its inactivation triggers mitotic defects, one of the major mechanisms through which tumour cells acquire chromosomal instability.
Topics: Animals; Humans; Mice; Anaphase; Cytokinesis; Fibroblasts; Genomic Instability; Mitosis; Profilins; Osteosarcoma
PubMed: 36599901
DOI: 10.1038/s42003-022-04392-8 -
Molecular Biology of the Cell Sep 2019Spindle microtubules, whose dynamics vary over time and at different locations, cooperatively drive chromosome segregation. Measurements of microtubule dynamics and...
Spindle microtubules, whose dynamics vary over time and at different locations, cooperatively drive chromosome segregation. Measurements of microtubule dynamics and spindle ultrastructure can provide insight into the behaviors of microtubules, helping elucidate the mechanism of chromosome segregation. Much work has focused on the dynamics and organization of kinetochore microtubules, that is, on the region between chromosomes and poles. In comparison, microtubules in the central-spindle region, between segregating chromosomes, have been less thoroughly characterized. Here, we report measurements of the movement of central-spindle microtubules during chromosome segregation in human mitotic spindles and mitotic and female meiotic spindles. We found that these central-spindle microtubules slide apart at the same speed as chromosomes, even as chromosomes move toward spindle poles. In these systems, damaging central-spindle microtubules by laser ablation caused an immediate and complete cessation of chromosome motion, suggesting a strong coupling between central-spindle microtubules and chromosomes. Electron tomographic reconstruction revealed that the analyzed anaphase spindles all contain microtubules with both ends between segregating chromosomes. Our results provide new dynamical, functional, and ultrastructural characterizations of central-spindle microtubules during chromosome segregation in diverse spindles and suggest that central-spindle microtubules and chromosomes are strongly coupled in anaphase.
Topics: Anaphase; Animals; Caenorhabditis elegans; Caenorhabditis elegans Proteins; Cell Line, Tumor; Chromosome Segregation; Chromosomes; Humans; Kinetochores; Meiosis; Microtubules; Spindle Apparatus; Spindle Poles
PubMed: 31339442
DOI: 10.1091/mbc.E19-01-0074 -
PLoS Genetics Nov 2020In mitosis and meiosis, chromosome segregation is triggered by the Anaphase-Promoting Complex/Cyclosome (APC/C), a multi-subunit ubiquitin ligase that targets proteins...
In mitosis and meiosis, chromosome segregation is triggered by the Anaphase-Promoting Complex/Cyclosome (APC/C), a multi-subunit ubiquitin ligase that targets proteins for degradation, leading to the separation of chromatids. APC/C activation requires phosphorylation of its APC3 and APC1 subunits, which allows the APC/C to bind its co-activator Cdc20. The identity of the kinase(s) responsible for APC/C activation in vivo is unclear. Cyclin B3 (CycB3) is an activator of the Cyclin-Dependent Kinase 1 (Cdk1) that is required for meiotic anaphase in flies, worms and vertebrates. It has been hypothesized that CycB3-Cdk1 may be responsible for APC/C activation in meiosis but this remains to be determined. Using Drosophila, we found that mutations in CycB3 genetically enhance mutations in tws, which encodes the B55 regulatory subunit of Protein Phosphatase 2A (PP2A) known to promote mitotic exit. Females heterozygous for CycB3 and tws loss-of-function alleles lay embryos that arrest in mitotic metaphase in a maternal effect, indicating that CycB3 promotes anaphase in mitosis in addition to meiosis. This metaphase arrest is not due to the Spindle Assembly Checkpoint (SAC) because mutation of mad2 that inactivates the SAC does not rescue the development of embryos from CycB3-/+, tws-/+ females. Moreover, we found that CycB3 promotes APC/C activity and anaphase in cells in culture. We show that CycB3 physically associates with the APC/C, is required for phosphorylation of APC3, and promotes APC/C association with its Cdc20 co-activators Fizzy and Cortex. Our results strongly suggest that CycB3-Cdk1 directly activates the APC/C to promote anaphase in both meiosis and mitosis.
Topics: Anaphase; Anaphase-Promoting Complex-Cyclosome; Animals; Animals, Genetically Modified; Apc3 Subunit, Anaphase-Promoting Complex-Cyclosome; CDC2 Protein Kinase; Cdc20 Proteins; Cell Line; Chromosome Segregation; Cyclin B; Drosophila Proteins; Drosophila melanogaster; Female; Loss of Function Mutation; Mad2 Proteins; Male; Metaphase; Models, Animal; Mutagenesis; Phosphoprotein Phosphatases; Phosphorylation
PubMed: 33137813
DOI: 10.1371/journal.pgen.1009184 -
Biophysical Journal Apr 2015Elongation of the mitotic spindle during anaphase B contributes to chromosome segregation in many cells. Here, we quantitatively test the ability of two models for...
Elongation of the mitotic spindle during anaphase B contributes to chromosome segregation in many cells. Here, we quantitatively test the ability of two models for spindle length control to describe the dynamics of anaphase B spindle elongation using experimental data from Drosophila embryos. In the slide-and-flux-or-elongate (SAFE) model, kinesin-5 motors persistently slide apart antiparallel interpolar microtubules (ipMTs). During pre-anaphase B, this outward sliding of ipMTs is balanced by depolymerization of their minus ends at the poles, producing poleward flux, while the spindle maintains a constant length. Following cyclin B degradation, ipMT depolymerization ceases so the sliding ipMTs can push the poles apart. The competing slide-and-cluster (SAC) model proposes that MTs nucleated at the equator are slid outward by the cooperative actions of the bipolar kinesin-5 and a minus-end-directed motor, which then pulls the sliding MTs inward and clusters them at the poles. In assessing both models, we assume that kinesin-5 preferentially cross-links and slides apart antiparallel MTs while the MT plus ends exhibit dynamic instability. However, in the SAC model, minus-end-directed motors bind the minus ends of MTs as cargo and transport them poleward along adjacent, parallel MT tracks, whereas in the SAFE model, all MT minus ends that reach the pole are depolymerized by kinesin-13. Remarkably, the results show that within a narrow range of MT dynamic instability parameters, both models can reproduce the steady-state length and dynamics of pre-anaphase B spindles and the rate of anaphase B spindle elongation. However, only the SAFE model reproduces the change in MT dynamics observed experimentally at anaphase B onset. Thus, although both models explain many features of anaphase B in this system, our quantitative evaluation of experimental data regarding several different aspects of spindle dynamics suggests that the SAFE model provides a better fit.
Topics: Anaphase; Animals; Drosophila; Kinesins; Microtubules; Models, Biological; Spindle Apparatus
PubMed: 25902440
DOI: 10.1016/j.bpj.2015.03.018 -
Seminars in Cell & Developmental Biology May 2010Kinetochores have been proposed to play multiple roles in mitotic chromosome alignment, including initial microtubule (MT) capture, monitoring MT attachments,... (Review)
Review
Kinetochores have been proposed to play multiple roles in mitotic chromosome alignment, including initial microtubule (MT) capture, monitoring MT attachments, prometaphase and anaphase chromosome movement and tension generation at metaphase. In addition, kinetochores are essential components of the spindle assembly checkpoint (SAC), and couple chromosome alignment with SAC silencing at metaphase. Although the molecular details of these activities remain under investigation, cytoplasmic dynein has been implicated in several aspects of MT and SAC regulation. Recent work clarifies the contribution of dynein to MT interactions and to events that drive anaphase onset. This review summarizes these studies and provides new models for dynein function.
Topics: Anaphase; Animals; Aspergillus; Cytoplasm; Dyneins; Gene Silencing; Humans; Kinetochores; Metaphase; Microtubules; Mitosis; Models, Biological; Phosphorylation
PubMed: 20045078
DOI: 10.1016/j.semcdb.2009.12.015 -
Open Biology Mar 2013The correct assembly and timely disassembly of the mitotic spindle is crucial for the propagation of the genome during cell division. Aurora kinases play a central role... (Review)
Review
The correct assembly and timely disassembly of the mitotic spindle is crucial for the propagation of the genome during cell division. Aurora kinases play a central role in orchestrating bipolar spindle establishment, chromosome alignment and segregation. In most eukaryotes, ranging from amoebas to humans, Aurora activity appears to be required both at the spindle pole and the kinetochore, and these activities are often split between two different Aurora paralogues, termed Aurora A and B. Polar and equatorial functions of Aurora kinases have generally been considered separately, with Aurora A being mostly involved in centrosome dynamics, whereas Aurora B coordinates kinetochore attachment and cytokinesis. However, double inactivation of both Aurora A and B results in a dramatic synergy that abolishes chromosome segregation. This suggests that these two activities jointly coordinate mitotic progression. Accordingly, recent evidence suggests that Aurora A and B work together in both spindle assembly in metaphase and disassembly in anaphase. Here, we provide an outlook on these shared functions of the Auroras, discuss the evolution of this family of mitotic kinases and speculate why Aurora kinase activity may be required at both ends of the spindle microtubules.
Topics: Anaphase; Aurora Kinases; Centrosome; Evolution, Molecular; Humans; Microtubules; Spindle Apparatus
PubMed: 23516109
DOI: 10.1098/rsob.120185 -
BioEssays : News and Reviews in... Jul 2017At metaphase in mitotic cells, pulling forces at the kinetochore-microtubule interface create tension by stretching the centromeric chromatin between oppositely oriented... (Review)
Review
At metaphase in mitotic cells, pulling forces at the kinetochore-microtubule interface create tension by stretching the centromeric chromatin between oppositely oriented sister kinetochores. This tension is important for stabilizing the end-on kinetochore microtubule attachment required for proper bi-orientation of sister chromosomes as well as for satisfaction of the Spindle Assembly Checkpoint and entry into anaphase. How force is coupled by proteins to kinetochore microtubules and resisted by centromere stretch is becoming better understood as many of the proteins involved have been identified. Recent application of genetically encoded fluorescent tension sensors within the mechanical linkage between the centromere and kinetochore microtubules are beginning to reveal - from live cell assays - protein specific contributions that are functionally important.
Topics: Anaphase; Animals; Chromatin; Chromosome Segregation; Humans; Kinetochores; Microtubules; Mitosis; Spindle Apparatus
PubMed: 28582586
DOI: 10.1002/bies.201600216 -
Trends in Biochemical Sciences Jun 2006Proteolysis mediated by the ubiquitin-proteasome system is a crucial regulatory mechanism in signal transduction cascades of temporal cellular processes such as cell... (Review)
Review
Proteolysis mediated by the ubiquitin-proteasome system is a crucial regulatory mechanism in signal transduction cascades of temporal cellular processes such as cell division. Two principal subtypes of modular ubiquitin ligase, the anaphase-promoting complex or cyclosome (APC/C) and the Skp1/Cullin-1/F-box protein complex, have emerged as essential regulators of key events in the cell cycle. The importance of these ligases is best illustrated by their roles in the checkpoint and repair pathways or in response to multiple stresses, where they affect activation of the M-phase-promoting factor or proper formation and/or maintenance of the mitotic spindle. Recent studies have considerably improved our understanding of the function of the concerted action of the phosphorylation and ubiquitin or SUMO systems in the regulation of the stability and activity of key components of the mitotic checkpoint.
Topics: Anaphase; Animals; Cell Division; Humans; Phosphorylation; Proteasome Endopeptidase Complex; Protein Processing, Post-Translational; SUMO-1 Protein; Signal Transduction; Ubiquitin
PubMed: 16647857
DOI: 10.1016/j.tibs.2006.04.001 -
Communications Biology Dec 2021Mitotic divisions achieve equal re-partition of chromosomes into daughter cells. In their recent work in , Sen, Harrison et al. propose that the risk of mis-segregation...
Mitotic divisions achieve equal re-partition of chromosomes into daughter cells. In their recent work in , Sen, Harrison et al. propose that the risk of mis-segregation in human mitotic cells is higher than previously thought and identify the existence of an early-anaphase correction mechanism. The study documents kinetochore dynamics in unprecedented detail, providing a detailed look at the events preceding loss of correct chromosomal numericity and genomic stability.
Topics: Anaphase; Chromosome Segregation; Humans; Kinetochores; Mitosis
PubMed: 34903809
DOI: 10.1038/s42003-021-02933-1