scholarly journals Dynamic bonds and polar ejection force distribution explain kinetochore oscillations in PtK1 cells

2013 ◽  
Vol 201 (4) ◽  
pp. 577-593 ◽  
Author(s):  
Gul Civelekoglu-Scholey ◽  
Bin He ◽  
Muyao Shen ◽  
Xiaohu Wan ◽  
Emanuele Roscioli ◽  
...  
Keyword(s):  
Metaphase Plate ◽  
Quantitative Model ◽  
Force Distribution ◽  
Mitotic Chromosomes ◽  
Chromosome Behavior ◽  
Sister Chromatids ◽  
Ejection Force ◽  
Sister Kinetochore ◽  
Spindle Microtubules ◽  
Key Features

Duplicated mitotic chromosomes aligned at the metaphase plate maintain dynamic attachments to spindle microtubules via their kinetochores, and multiple motor and nonmotor proteins cooperate to regulate their behavior. Depending on the system, sister chromatids may display either of two distinct behaviors, namely (1) the presence or (2) the absence of oscillations about the metaphase plate. Significantly, in PtK1 cells, in which chromosome behavior appears to be dependent on the position along the metaphase plate, both types of behavior are observed within the same spindle, but how and why these distinct behaviors are manifested is unclear. Here, we developed a new quantitative model to describe metaphase chromosome dynamics via kinetochore–microtubule interactions mediated by nonmotor viscoelastic linkages. Our model reproduces all the key features of metaphase sister kinetochore dynamics in PtK1 cells and suggests that differences in the distribution of polar ejection forces at the periphery and in the middle of PtK1 cell spindles underlie the observed dichotomy of chromosome behavior.

Phosphorylated proteins are involved in sister-chromatid arm cohesion during meiosis I

1999 ◽  
Vol 112 (17) ◽  
pp. 2957-2969 ◽  
Author(s):  
J.A. Suja ◽  
C. Antonio ◽  
A. Debec ◽  
J.S. Rufas
Keyword(s):  
Sister Chromatid ◽  
Chromosome Structure ◽  
Metaphase Plate ◽  
Mitotic Chromosomes ◽  
Phosphorylated Proteins ◽  
Sister Chromatids ◽  
Sequential Release ◽  
Sister Kinetochore ◽  
Meiosis I ◽  
Metaphase I

Sister-chromatid arm cohesion is lost during the metaphase I/anaphase I transition to allow homologue separation. To obtain needed information on this process we have analysed in grasshopper bivalents the sequential release of arm cohesion in relation to the behaviour of chromatid axes. Results show that sister axes are associated during early metaphase I but separate during late metaphase I leading to a concomitant change of chromosome structure that implies the loss of sister-kinetochore cohesion. Afterwards, homologues initiate their separation asynchronously depending on their size, and number and position of chiasmata. In all bivalents thin chromatin strands at the telomeres appeared as the last point of contact between sister chromatids. Additionally, we have analysed the participation of phosphoproteins recognised by the MPM-2 monoclonal antibody against mitotic phosphoproteins in arm cohesion in bivalents and two different kinds of univalents. Results show the absence of MPM-2 phosphoproteins at the interchromatid domain in mitotic chromosomes and meiotic univalents, but their presence in metaphase I bivalents. These phosphoproteins are lost at the onset of anaphase I. Taken together, these data have prompted us to propose a ‘working’ model for the release of arm cohesion during meiosis I. The model suggests that MPM-2 phosphoproteins may act as cohesive proteins associating sister axes. Their modification, once all bivalents are correctly aligned at the metaphase plate, would trigger a change of chromosome structure and the sequential release of sister-kinetochore, arm, and telomere cohesions.

Proteins of the inner and outer centromere of mitotic chromosomes

Genome ◽  
1989 ◽  
Vol 31 (2) ◽  
pp. 541-552 ◽  
Author(s):  
William C. Earnshaw ◽  
Carol A. Cooke
Keyword(s):  
Negative Charge ◽  
Dna Binding Protein ◽  
Mitotic Chromosomes ◽  
Sister Chromatids ◽  
Region I ◽  
Spindle Microtubules ◽  
Region Ii ◽  
Chromosome Scaffold

We have used immunocytochemistry and molecular cloning methods to identify and characterize structural polypeptides of the centromere. These studies permit us to resolve two distinct regions: the inner and outer centromere, (i) Components of the outer centromere: autoantibodies from certain patients with rheumatic disease identify a family of three immunologically related polypeptides that we have designated CENP-A (17 kDa), CENP-B (80 kDa), and CENP-C (140 kDa). CENP-B has been cloned and sequenced. DNA sequence analysis indicates that this polypeptide possesses two large regions with extraordinary concentrations of acidic residues (region I: 61 residues with 79% glu + asp; region II: 31 residues with 87% glu + asp). Despite this concentration of negative charge, immunocytochemical experiments suggest that CENP-B may be a DNA binding protein. In these experiments, the levels of CENP-B are seen to vary reproducibly from chromosome to chromosome. The role of CENP-B in vivo is unknown. However, it is unlikely to bind directly to the spindle microtubules since it is found at an inactive centromere that apparently does not attach to the spindle. (ii) Components of the inner centromere: we have injected mice with the whole chromosome scaffold fraction to elicit production of monoclonal antibodies. One such antibody identifies two structurally related polypeptides (the INCENP antigens, 135 and 155 kDa) that are preferentially located between the sister chromatids at the centromere. The INCENP antigens undergo dramatic movements from the chromosomes to the central spindle during mitosis. They are ultimately sequestered in the midbody and discarded. Several lines of evidence suggest that the INCENP polypeptides may be involved in the regulation of sister chromatid separation at the metaphase–anaphase transition.Key words: mitosis, centromere, CENP antigens, INCENP antigens, kinetochore, disjunction.

scholarly journals Direct evaluation of cohesin-mediated sister kinetochore associations at meiosis I in fission yeast

2021 ◽  
Author(s):  
Masashi Nambu ◽  
Atsuki Kishikawa ◽  
Takatomi Yamada ◽  
Kento Ichikawa ◽  
Yunosuke Kira ◽  
...  
Keyword(s):  
Fission Yeast ◽  
Chromosome Segregation ◽  
Mitotic Chromosomes ◽  
Direct Evaluation ◽  
Homologous Chromosomes ◽  
Sister Chromatids ◽  
Pheromone Signaling ◽  
Sister Kinetochore ◽  
Novel Method ◽  
Meiosis I

Kinetochores drive chromosome segregation by mediating chromosome interactions with the spindle. In higher eukaryotes, sister kinetochores are separately positioned on opposite sides of sister centromeres during mitosis, but associate with each other during meiosis I. Kinetochore association facilitates the attachment of sister chromatids to the same pole, enabling the segregation of homologous chromosomes toward opposite poles. In the fission yeast, Schizosaccharomyces pombe, Rec8-containing meiotic cohesin is suggested to establish kinetochore associations by mediating cohesion of the centromere cores. However, cohesin-mediated kinetochore associations on intact chromosomes have never been demonstrated directly. Here, we describe a novel method for the direct evaluation of kinetochore associations on intact chromosomes in live S. pombe cells, and demonstrate that sister kinetochores and the centromere cores are positioned separately on mitotic chromosomes but associate with each other on meiosis I chromosomes. Furthermore, we demonstrate that kinetochore association depends on meiotic cohesin and the cohesin regulators, Moa1 and Mrc1, and requires mating-pheromone signaling for its establishment. These results confirm cohesin-mediated kinetochore association and its regulatory mechanisms, along with the usefulness of the developed method for its analysis.

scholarly journals Methylation of PLK1 by SET7/9 ensures accurate kinetochore–microtubule dynamics

2019 ◽  
Vol 12 (6) ◽  
pp. 462-476 ◽  
Author(s):  
Ruoying Yu ◽  
Huihui Wu ◽  
Hazrat Ismail ◽  
Shihao Du ◽  
Jun Cao ◽  
...  
Keyword(s):  
Chromosome Segregation ◽  
Kinase Activity ◽  
Genomic Stability ◽  
Lysine Methylation ◽  
Mitotic Chromosomes ◽  
Methyltransferase Activity ◽  
Sister Chromatids ◽  
Spindle Microtubules ◽  
Chemical Inhibition ◽  
Early Mitosis

Abstract Faithful segregation of mitotic chromosomes requires bi-orientation of sister chromatids, which relies on the sensing of correct attachments between spindle microtubules and kinetochores. Although the mechanisms underlying PLK1 activation have been extensively studied, the regulatory mechanisms that couple PLK1 activity to accurate chromosome segregation are not well understood. In particular, PLK1 is implicated in stabilizing kinetochore–microtubule attachments, but how kinetochore PLK1 activity is regulated to avoid hyperstabilized kinetochore–microtubules in mitosis remains elusive. Here, we show that kinetochore PLK1 kinase activity is modulated by SET7/9 via lysine methylation during early mitosis. The SET7/9-elicited dimethylation occurs at the Lys191 of PLK1, which tunes down its activity by limiting ATP utilization. Overexpression of the non-methylatable PLK1 mutant or chemical inhibition of SET7/9 methyltransferase activity resulted in mitotic arrest due to destabilized kinetochore–microtubule attachments. These data suggest that kinetochore PLK1 is essential for stable kinetochore–microtubule attachments and methylation by SET7/9 promotes dynamic kinetochore–microtubule attachments for accurate error correction. Our findings define a novel homeostatic regulation at the kinetochore that integrates protein phosphorylation and methylation with accurate chromosome segregation for maintenance of genomic stability.

Confocal analysis of chromosome behavior in wheat × maize zygotes

Genome ◽  
2004 ◽  
Vol 47 (1) ◽  
pp. 199-205 ◽  
Author(s):  
Keiichi Mochida ◽  
Hisashi Tsujimoto ◽  
Tetsuo Sasakuma
Keyword(s):  
Metaphase Plate ◽  
Chromosome Elimination ◽  
Chromosome Movement ◽  
Hybridization Method ◽  
Chromosome Behavior ◽  
Whole Mount ◽  
Hybrid Embryo ◽  
Spindle Microtubules ◽  
Spindle Structure

Herein, we profile the first embryonic mitosis in a hybrid of wheat and maize by using a whole-mount genomic in situ hybridization method and immunofluorescence staining with a tubulin-specific antibody. We have successfully captured the dynamics of each set of parental chromosomes in the first zygotic division of the hybrid embryo 24-28 h after crossing. During the first zygotic metaphase, although both sets of parental chromosomes congressed into the equatorial plate of the zygote, the maize chromosomes tended to lag in comparison with the wheat chromosomes. During anaphase, each parental chromosome separated into its sister chromosomes; however, some of the maize chromosomes lagged around the metaphase plate as segregants. The maize sister chromosomes that did move toward the pole showed delayed and asymmetric movement as compared with the wheat ones. Immunological staining of tubulin revealed a bipolar spindle structure in the first zygotic metaphase. The kinetochores of the maize chromosomes that lagged around the metaphase plate did not attach to the spindle microtubules. These results suggest that factors on the kinetochores of maize chromosomes that are required to control chromosome movement are deficient in the zygotic cell cycle.Key words: whole-mount, GISH, chromosome elimination, hybrid embryogenesis.

scholarly journals Identification of a novel nucleolar protein complex required for mitotic chromosome segregation through centromeric accumulation of Aurora B

2020 ◽  
Vol 48 (12) ◽  
pp. 6583-6596
Author(s):  
Akiko Fujimura ◽  
Yuki Hayashi ◽  
Kazashi Kato ◽  
Yuichiro Kogure ◽  
Mutsuro Kameyama ◽  
...  
Keyword(s):  
Chromosome Segregation ◽  
Protein Complex ◽  
Metaphase Plate ◽  
Nucleolar Protein ◽  
Aurora B ◽  
Mitotic Progression ◽  
Mitotic Chromosomes ◽  
Nucleolar Proteins ◽  
Chromatid Cohesion ◽  
Wd Repeat

Abstract The nucleolus is a membrane-less nuclear structure that disassembles when cells undergo mitosis. During mitosis, nucleolar factors are thus released from the nucleolus and dynamically change their subcellular localization; however, their functions remain largely uncharacterised. Here, we found that a nucleolar factor called nucleolar protein 11 (NOL11) forms a protein complex with two tryptophan-aspartic acid (WD) repeat proteins named WD-repeat protein 43 (WDR43) and Cirhin in mitotic cells. This complex, referred to here as the NWC (NOL11-WDR43-Cirhin) complex, exists in nucleoli during interphase and translocates to the periphery of mitotic chromosomes, i.e., perichromosomal regions. During mitotic progression, both the congression of chromosomes to the metaphase plate and sister chromatid cohesion are impaired in the absence of the NWC complex, as it is required for the centromeric enrichment of Aurora B and the associating phosphorylation of histone H3 at threonine 3. These results reveal the characteristics of a novel protein complex consisting of nucleolar proteins, which is required for regulating kinetochores and centromeres to ensure faithful chromosome segregation.

scholarly journals Genes Involved in Sister Chromatid Separation and Segregation in the Budding Yeast Saccharomyces cerevisiae

Genetics ◽  
2001 ◽  
Vol 159 (2) ◽  
pp. 453-470
Author(s):  
Sue Biggins ◽  
Needhi Bhalla ◽  
Amy Chang ◽  
Dana L Smith ◽  
Andrew W Murray
Keyword(s):  
Chromosome Segregation ◽  
Sister Chromatid ◽  
Mitotic Spindle ◽  
Budding Yeast ◽  
Temperature Sensitive ◽  
Chromosome Behavior ◽  
Yeast Saccharomyces Cerevisiae ◽  
Sister Chromatids ◽  
Marked Chromosome ◽  
Sister Chromatid Separation

Abstract Accurate chromosome segregation requires the precise coordination of events during the cell cycle. Replicated sister chromatids are held together while they are properly attached to and aligned by the mitotic spindle at metaphase. At anaphase, the links between sisters must be promptly dissolved to allow the mitotic spindle to rapidly separate them to opposite poles. To isolate genes involved in chromosome behavior during mitosis, we microscopically screened a temperature-sensitive collection of budding yeast mutants that contain a GFP-marked chromosome. Nine LOC (loss of cohesion) complementation groups that do not segregate sister chromatids at anaphase were identified. We cloned the corresponding genes and performed secondary tests to determine their function in chromosome behavior. We determined that three LOC genes, PDS1, ESP1, and YCS4, are required for sister chromatid separation and three other LOC genes, CSE4, IPL1, and SMT3, are required for chromosome segregation. We isolated alleles of two genes involved in splicing, PRP16 and PRP19, which impair α-tubulin synthesis thus preventing spindle assembly, as well as an allele of CDC7 that is defective in DNA replication. We also report an initial characterization of phenotypes associated with the SMT3/SUMO gene and the isolation of WSS1, a high-copy smt3 suppressor.

The influence of fixation on the morphology of mitotic chromosomes

1986 ◽  
Vol 28 (4) ◽  
pp. 536-539 ◽  
Author(s):  
Axel J. J. Dietrich
Keyword(s):  
Acetic Acid ◽  
Chemical Composition ◽  
Strong Influence ◽  
Chromosome Structure ◽  
Human Lymphocytes ◽  
Chromosome Morphology ◽  
Mitotic Chromosomes ◽  
Specific Chemical ◽  
C Banding ◽  
Sister Chromatids

It is well known that there is a strong influence of fixation, i.e., acetic methanol versus formaldehyde, on the chromosome morphology at stages of the first meiotic division. In this study the influence of both these types of fixation on the morphology of mitotic chromosomes was examined in human lymphocytes. After methanol – acetic acid (3:1) fixation, the chromosomes show the "classical" condensed shape in which it is not always possible to recognize the two sister chromatids. These chromosomes are accessible to the conventional G-, R-, and C-banding techniques. After formaldehyde fixation at a relatively high pH, the chromosomes are thinner and longer (two to six times) when compared with chromosomes following methanol – acetic acid fixation. They show a scaffold-like morphology, sometimes with a halo of thin material around it. In all cases the two sister chromatids could be recognized. This chromosome structure could be easily stained with silver, Giemsa, 4,6-diamino-2-phenyl-indole (DAPI), and fluorescein isocyanate isomere 1 (FITC). The results obtained following these stainings gave no indication to any specific chemical composition of a probable central scaffold. The scaffold-like structures were not accessible to G-, R-, or C-banding techniques. The only effect observed following these banding techniques was the disappearance of the halo of thin material around the central scaffold-like structure.Key words: chromosome structure, fixation influence, human lymphocytes.

scholarly journals The right place at the right time: Aurora B kinase localization to centromeres and kinetochores

2020 ◽  
Vol 64 (2) ◽  
pp. 299-311 ◽  
Author(s):  
Amanda J. Broad ◽  
Jennifer G. DeLuca
Keyword(s):  
Spindle Assembly Checkpoint ◽  
Protein Complexes ◽  
Centromeric Heterochromatin ◽  
Aurora B ◽  
Mitotic Chromosomes ◽  
Large Protein ◽  
Aurora B Kinase ◽  
Centromeric Protein ◽  
Spindle Microtubules ◽  
The Right

Abstract The fidelity of chromosome segregation during mitosis is intimately linked to the function of kinetochores, which are large protein complexes assembled at sites of centromeric heterochromatin on mitotic chromosomes. These key “orchestrators” of mitosis physically connect chromosomes to spindle microtubules and transduce forces through these connections to congress chromosomes and silence the spindle assembly checkpoint. Kinetochore-microtubule attachments are highly regulated to ensure that incorrect attachments are not prematurely stabilized, but instead released and corrected. The kinase activity of the centromeric protein Aurora B is required for kinetochore-microtubule destabilization during mitosis, but how the kinase acts on outer kinetochore substrates to selectively destabilize immature and erroneous attachments remains debated. Here, we review recent literature that sheds light on how Aurora B kinase is recruited to both centromeres and kinetochores and discuss possible mechanisms for how kinase interactions with substrates at distinct regions of mitotic chromosomes are regulated.

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.

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