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PLoS Genetics May 2018It is widely accepted in eukaryotes that the cleavage furrow only initiates after mitosis completion. In fission yeast, cytokinesis requires the synthesis of a septum...
It is widely accepted in eukaryotes that the cleavage furrow only initiates after mitosis completion. In fission yeast, cytokinesis requires the synthesis of a septum tightly coupled to cleavage furrow ingression. The current cytokinesis model establishes that simultaneous septation and furrow ingression only initiate after spindle breakage and mitosis exit. Thus, this model considers that although Cdk1 is inactivated at early-anaphase, septation onset requires the long elapsed time until mitosis completion and full activation of the Hippo-like SIN pathway. Here, we studied the precise timing of septation onset regarding mitosis by exploiting both the septum-specific detection with the fluorochrome calcofluor and the high-resolution electron microscopy during anaphase and telophase. Contrarily to the existing model, we found that both septum and cleavage furrow start to ingress at early anaphase B, long before spindle breakage, with a slow ingression rate during anaphase B, and greatly increasing after telophase onset. This shows that mitosis and cleavage furrow ingression are not concatenated but simultaneous events in fission yeast. We found that the timing of septation during early anaphase correlates with the cell size and is regulated by the corresponding levels of SIN Etd1 and Rho1. Cdk1 inactivation was directly required for timely septation in early anaphase. Strikingly the reduced SIN activity present after Cdk1 loss was enough to trigger septation by immediately inducing the medial recruitment of the SIN kinase complex Sid2-Mob1. On the other hand, septation onset did not depend on the SIN asymmetry establishment, which is considered a hallmark for SIN activation. These results recalibrate the timing of key cytokinetic events in fission yeast; and unveil a size-dependent control mechanism that synchronizes simultaneous nuclei separation with septum and cleavage furrow ingression to safeguard the proper chromosome segregation during cell division.
Topics: Anaphase; Benzenesulfonates; CDC2 Protein Kinase; Cell Cycle Proteins; Cell Nucleus; Cytokinesis; Microscopy, Electron, Transmission; Microscopy, Fluorescence; Protein Kinases; Schizosaccharomyces; Schizosaccharomyces pombe Proteins; Spindle Apparatus; Telophase; Time Factors; rho GTP-Binding Proteins
PubMed: 29813053
DOI: 10.1371/journal.pgen.1007388 -
EMBO Reports Oct 2021The isotropic metaphase actin cortex progressively polarizes as the anaphase spindle elongates during mitotic exit. This involves the loss of actomyosin cortex from...
The isotropic metaphase actin cortex progressively polarizes as the anaphase spindle elongates during mitotic exit. This involves the loss of actomyosin cortex from opposing cell poles and the accumulation of an actomyosin belt at the cell centre. Although these spatially distinct cortical remodelling events are coordinated in time, here we show that they are independent of each other. Thus, actomyosin is lost from opposing poles in anaphase cells that lack an actomyosin ring owing to centralspindlin depletion. In examining potential regulators of this process, we identify a role for Aurora B kinase in actin clearance at cell poles. Upon combining Aurora B inhibition with centralspindlin depletion, cells exiting mitosis fail to change shape and remain completely spherical. Additionally, we demonstrate a requirement for Aurora B in the clearance of cortical actin close to anaphase chromatin in cells exiting mitosis with a bipolar spindle and in monopolar cells forced to divide while flat. Altogether, these data suggest a novel role for Aurora B activity in facilitating DNA-mediated polar relaxation at anaphase, polarization of the actomyosin cortex, and cell division.
Topics: Actomyosin; Anaphase; Aurora Kinase B; Cytokinesis; Mitosis; Spindle Apparatus
PubMed: 34431205
DOI: 10.15252/embr.202152387 -
Developmental Cell Jan 2015Polyploidization is a natural process that frequently accompanies differentiation; its deregulation is linked to genomic instability and cancer. Despite its relevance,...
Polyploidization is a natural process that frequently accompanies differentiation; its deregulation is linked to genomic instability and cancer. Despite its relevance, why cells select different polyploidization mechanisms is unknown. Here we report a systematic genetic analysis of endomitosis, a process in which megakaryocytes become polyploid by entering mitosis but aborting anaphase. Whereas ablation of the APC/C cofactor Cdc20 results in mitotic arrest and severe thrombocytopenia, lack of the kinases Aurora-B, Cdk1, or Cdk2 does not affect megakaryocyte polyploidization or platelet levels. Ablation of Cdk1 forces a switch to endocycles without mitosis, whereas polyploidization in the absence of Cdk1 and Cdk2 occurs in the presence of aberrant re-replication events. Importantly, ablation of these kinases rescues the defects in Cdc20 null megakaryocytes. These findings suggest that endomitosis can be functionally replaced by alternative polyploidization mechanisms in vivo and provide the cellular basis for therapeutic approaches aimed to discriminate mitotic and polyploid cells.
Topics: Anaphase; Animals; Cdc20 Proteins; Cells, Cultured; Megakaryocytes; Mice; Mitosis; Polyploidy; Protein Serine-Threonine Kinases
PubMed: 25625205
DOI: 10.1016/j.devcel.2014.12.015 -
International Journal of Molecular... Jul 2023Plant height, petiole length, and the angle of the leaf petiole and branch angles are crucial traits determining plant architecture and yield in soybean ( L.). Here, we...
Plant height, petiole length, and the angle of the leaf petiole and branch angles are crucial traits determining plant architecture and yield in soybean ( L.). Here, we characterized a soybean mutant with super-short petioles (SSP) and enlarged petiole angles (named ) through phenotypic observation, anatomical structure analysis, and bulk sequencing analysis. To identify the gene responsible for the mutant phenotype, we established a pipeline involving bulk sequencing, variant calling, functional annotation by SnpEFF (v4.0e) software, and Integrative Genomics Viewer analysis, and we initially identified , encoding a homolog of Anaphase-promoting complex subunit 8 (APC8). Another mutant, , with a large deletion of many genes including , has super-short petioles and an enlarged petiole angle, similar to the phenotype. Characterization of the mutant together with quantitative trait locus mapping and allelic variation analysis confirmed as the gene involved in the phenotype. In , a 4 bp deletion in leads to a 380 aa truncated protein due to a premature stop codon. The dysfunction or absence of caused severe defects in morphology, anatomical structure, and physiological traits. Transcriptome analysis and weighted gene co-expression network analysis revealed multiple pathways likely involved in these phenotypes, including ubiquitin-mediated proteolysis and gibberellin-mediated pathways. Our results demonstrate that dysfunction of leads to diverse functional consequences in different tissues, indicating that this APC8 homolog plays key roles in cell differentiation and elongation in a tissue-specific manner. Deciphering the molecular control of petiole length and angle enriches our knowledge of the molecular network regulating plant architecture in soybean and should facilitate the breeding of high-yielding soybean cultivars with compact plant architecture.
Topics: Glycine max; Anaphase; Plant Breeding; Chromosome Mapping; Phenotype
PubMed: 37446203
DOI: 10.3390/ijms241311024 -
Philosophical Transactions of the Royal... Sep 1999Ubiquitin-mediated proteolysis is fundamental to cell cycle progression. In the fission yeast Schizosaccharomyces pombe, a mitotic cyclin (Cdc13), a key cell cycle... (Review)
Review
Ubiquitin-mediated proteolysis is fundamental to cell cycle progression. In the fission yeast Schizosaccharomyces pombe, a mitotic cyclin (Cdc13), a key cell cycle regulator, is degraded for exiting mitosis, while Cut2 has to be destroyed for the onset of sister chromatid separation in anaphase. Ubiquitination of these proteins requires the special destruction box (DB) sequences locating in their N-termini and the large, 20S complex called the anaphase-promoting complex or cyclosome. Here we show that cyclosome function during metaphase-anaphase progression is regulated by the protein kinase A (PKA) inactivation pathway, ubiquitination of the cyclosome subunit, and cellular localization of the target substrates. Evidence is provided that the cyclosome plays pleiotropic roles in the cell cycle: mutations in the subunit genes show a common anaphase defect, but subunit-specific phenotypes such as in G1/S or G2/M transition, septation and cytokinesis, stress response and heavy metal sensitivity, are additionally produced, suggesting that different subunits take distinct parts of complex cyclosome functions. Inactivation of PKA is important for the activation of the cyclosome for promoting anaphase, perhaps through dephosphorylation of the subunits such as Cut9 (Apc6). Cut4 (Apc1), the largest subunit, plays an essential role in the assembly and functional regulation of the cyclosome in response to cell cycle arrest and stresses. Cut4 is highly modified, probably by ubiquitination, when it is not assembled into the 20S cyclosome. Sds23 is implicated in DB-mediated ubiquitination possibly through regulating de-ubiquitination, while Cut8 is necessary for efficient proteolysis of Cdc13 and Cut2 coupled with cytokinesis. Unexpectedly, the timing of proteolysis is dependent on cellular localization of the substrate. Cdc13 enriched along the spindle disappears first, followed by decay of the nuclear signal, whereas Cut2 in the nucleus disappears first, followed by decline in the spindle signal during metaphase-anaphase progression.
Topics: Anaphase; Anaphase-Promoting Complex-Cyclosome; Apc1 Subunit, Anaphase-Promoting Complex-Cyclosome; Cyclic AMP-Dependent Protein Kinases; Fungal Proteins; Humans; Ligases; Metaphase; Mutation; Schizosaccharomyces; Substrate Specificity; Ubiquitin-Protein Ligase Complexes; Ubiquitin-Protein Ligases; Ubiquitins
PubMed: 10582241
DOI: 10.1098/rstb.1999.0499 -
Fungal Genetics and Biology : FG & B Oct 2011Mitosis in Aspergillus nidulans is very rapid, requiring less than 5 min at 37 °C in germlings (Bergen and Morris, 1983). In this time the cytoplasmic microtubules...
Mitosis in Aspergillus nidulans is very rapid, requiring less than 5 min at 37 °C in germlings (Bergen and Morris, 1983). In this time the cytoplasmic microtubules (MTs) must disassemble, the mitotic spindle assemble, function and disassemble, and cytoplasmic MTs reassemble. It follows that cytoplasmic MTs must be extremely dynamic in this period and we were interested, in particular, in examining the processes of MT disassembly in prophase and reassembly in anaphase and telophase. We observed a diploid strain that expressed GFP-α-tubulin. We used a spinning disk confocal microscope that allowed rapid image capture, which proved necessary because microtubule dynamics were extremely rapid. We found, for the first time, that microtubule severing occurs in prophase in a filamentous fungus and that catastrophe rather than nucleation limits astral microtubule growth.
Topics: Anaphase; Aspergillus nidulans; Diploidy; Haploidy; Microtubules; Mitosis; Prophase; Spindle Apparatus; Telophase
PubMed: 21807107
DOI: 10.1016/j.fgb.2011.07.003 -
Proceedings of the National Academy of... Apr 1994By using monopolar spindles artificially induced in sea urchin embryos, we examined whether or not the presence of two opposing poles was an indispensable condition for...
By using monopolar spindles artificially induced in sea urchin embryos, we examined whether or not the presence of two opposing poles was an indispensable condition for keeping chromosomes at a fixed distance from the pole at metaphase and for the anaphase chromosome movement. Chromosomes were stained with Hoechst dye 33342 and their behavior was followed in the monopolar and the control bipolar spindles. In the monopolar spindle, chromosomes were first arranged on a curved metaphase plate and then spread on a part of the imaginary surface of a sphere whose center was the monopole. The estimated chromosome-to-pole distance was similar to that of bipolar spindles at metaphase and remained fixed until chromosomes started to move toward the pole. The average duration of metaphase in the monopolar spindle was 6 times longer than that in the bipolar spindle. The poleward movement of chromosomes in the monopolar spindle was similar to the anaphase A (chromosome-to-pole movement) in the bipolar spindle with respect to the velocity, duration, distance, and synchronization of migration. These results show that even half of the normal spindle has capacities for the arrangement of chromosomes at metaphase and for the anaphase A chromosome movement. Based on these results, we were able to exclude some existing theories of metaphase, such as the one based on the balance of forces between the two poles.
Topics: Anaphase; Animals; Chromosomes; In Vitro Techniques; Metaphase; Microscopy, Fluorescence; Movement; Sea Urchins; Spindle Apparatus; Video Recording
PubMed: 8171013
DOI: 10.1073/pnas.91.9.3921 -
Nature Communications May 2018Kinetochores are multi-protein complexes that power chromosome movements by tracking microtubules plus-ends in the mitotic spindle. Human kinetochores bind up to 20...
Kinetochores are multi-protein complexes that power chromosome movements by tracking microtubules plus-ends in the mitotic spindle. Human kinetochores bind up to 20 microtubules, even though single microtubules can generate sufficient force to move chromosomes. Here, we show that high microtubule occupancy at kinetochores ensures robust chromosome segregation by providing a strong mechanical force that favours segregation of merotelic attachments during anaphase. Using low doses of the microtubules-targeting agent BAL27862 we reduce microtubule occupancy and observe that spindle morphology is unaffected and bi-oriented kinetochores can still oscillate with normal intra-kinetochore distances. Inter-kinetochore stretching is, however, dramatically reduced. The reduction in microtubule occupancy and inter-kinetochore stretching does not delay satisfaction of the spindle assembly checkpoint or induce microtubule detachment via Aurora-B kinase, which was so far thought to release microtubules from kinetochores under low stretching. Rather, partial microtubule occupancy slows down anaphase A and increases incidences of lagging chromosomes due to merotelically attached kinetochores.
Topics: Anaphase; Aurora Kinase B; Benzimidazoles; Cell Line; Chromosome Segregation; Humans; Intravital Microscopy; Kinetochores; Microscopy, Electron; Microtubules; Oxadiazoles; Spindle Apparatus
PubMed: 29795284
DOI: 10.1038/s41467-018-04427-x -
The Journal of Cell Biology Dec 2020Anaphase chromosome movement is thought to be mediated by pulling forces generated by end-on attachment of microtubules to the outer face of kinetochores. However, it...
Anaphase chromosome movement is thought to be mediated by pulling forces generated by end-on attachment of microtubules to the outer face of kinetochores. However, it has been suggested that during C. elegans female meiosis, anaphase is mediated by a kinetochore-independent pushing mechanism with microtubules only attached to the inner face of segregating chromosomes. We found that the kinetochore proteins KNL-1 and KNL-3 are required for preanaphase chromosome stretching, suggesting a role in pulling forces. In the absence of KNL-1,3, pairs of homologous chromosomes did not separate and did not move toward a spindle pole. Instead, each homolog pair moved together with the same spindle pole during anaphase B spindle elongation. Two masses of chromatin thus ended up at opposite spindle poles, giving the appearance of successful anaphase.
Topics: Anaphase; Animals; Caenorhabditis elegans; Caenorhabditis elegans Proteins; Kinetochores; Microtubule-Associated Proteins
PubMed: 33064834
DOI: 10.1083/jcb.202005179 -
PloS One 2016Accurate chromosome segregation during cell division is essential to maintain genome stability, and chromosome segregation errors are causally linked to genetic...
Accurate chromosome segregation during cell division is essential to maintain genome stability, and chromosome segregation errors are causally linked to genetic disorders and cancer. An anaphase chromosome bridge is a particular chromosome segregation error observed in cells that enter mitosis with fused chromosomes/sister chromatids. The widely accepted Breakage/Fusion/Bridge cycle model proposes that anaphase chromosome bridges break during mitosis to generate chromosome ends that will fuse during the following cell cycle, thus forming new bridges that will break, and so on. However, various studies have also shown a link between chromosome bridges and aneuploidy and/or polyploidy. In this study, we investigated the behavior and properties of chromosome bridges during mitosis, with the idea to gain insight into the potential mechanism underlying chromosome bridge-induced aneuploidy. We find that only a small number of chromosome bridges break during anaphase, whereas the rest persist through mitosis into the subsequent cell cycle. We also find that the microtubule bundles (k-fibers) bound to bridge kinetochores are not prone to breakage/detachment, thus supporting the conclusion that k-fiber detachment is not the cause of chromosome bridge-induced aneuploidy. Instead, our data suggest that while the microtubules bound to the kinetochores of normally segregating chromosomes shorten substantially during anaphase, the k-fibers bound to bridge kinetochores shorten only slightly, and may even lengthen, during anaphase. This causes some of the bridge kinetochores/chromosomes to lag behind in a position that is proximal to the cell/spindle equator and may cause the bridged chromosomes to be segregated into the same daughter nucleus or to form a micronucleus.
Topics: Anaphase; Cells, Cultured; Chromosome Segregation; Chromosomes, Human; HeLa Cells; Humans; Kinetochores; Mammary Glands, Human; Microtubules; Mitosis
PubMed: 26784746
DOI: 10.1371/journal.pone.0147420