Higher-order structure of human mitotic chromosomes.

Bak, A. Leth; Zeuthen, Jesper; Crick, Francis

doi:10.1073/pnas.74.4.1595

Cited by 131 publications

(59 citation statements)

References 9 publications

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“…Even in unicellular eukaryotic organisms, such as yeast, interphase chromatin DNA is already compacted ~1000-fold in the nucleus, relative to the length of the naked DNA duplex. Forming properly condensed chromosomes upon entry into mitosis at prophase requires a further compaction, by severalfold, of interphase chromatin DNA (Bak et al, 1977;Earnshaw, 1988;Kireeva et al, 2004). In the fission yeast Schizosaccharomyces pombe, which has a small genome, chromatin condenses ~fivefold upon mitotic entry, judging from fluorescence in situ hybridization (FISH) images of chromosome arms (Saka et al, 1994).…”

Section: Introductionmentioning

confidence: 99%

Condensin phosphorylated by the Aurora-B-like kinase Ark1 is continuously required until telophase in a mode distinct from Top2

Nakazawa

Mehrotra

Ebe

et al. 2011

Journal of Cell Science

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SummaryCondensin is a conserved protein complex that functions in chromosome condensation and segregation. It has not been previously unequivocally determined whether condensin is required throughout mitosis. Here, we examined whether Schizosaccharomyces pombe condensin continuously acts on chromosomes during mitosis and compared its role with that of DNA topoisomerase II (Top2). Using double mutants containing a temperature-sensitive allele of the condensin SMC2 subunit cut14 (cut14-208) or of top2, together with the cold-sensitive nda3-KM311 mutation (in -tubulin), temperature-shift experiments were performed. These experiments allowed inactivation of condensin or Top2 at various stages throughout mitosis, even after late anaphase. The results established that mitotic chromosomes require condensin and Top2 throughout mitosis, even in telophase. We then showed that the Cnd2 subunit of condensin (also known as Barren) is the target subunit of Aurora-B-like kinase Ark1 and that Ark1-mediated phosphorylation of Cnd2 occurred throughout mitosis. The phosphorylation sites in Cnd2 were determined by mass spectrometry, and alanine and glutamate residue replacement mutant constructs for these sites were constructed. Alanine substitution mutants of Cnd2, which mimic the unphosphorylated protein, exhibited broad mitotic defects, including at telophase, and overexpression of these constructs caused a severe dominantnegative effect. By contrast, glutamate substitution mutants, which mimic the phosphorylated protein, alleviated the segregation defect in Ark1-inhibited cells. In telophase, the condensin subunits in cut14-208 mutant accumulated in lumps that contained telomeric DNA and proteins that failed to segregate. Condensin might thus serve to keep the segregated chromosomes apart during telophase.

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Section: Introductionmentioning

confidence: 99%

Condensin phosphorylated by the Aurora-B-like kinase Ark1 is continuously required until telophase in a mode distinct from Top2

Nakazawa

Mehrotra

Ebe

et al. 2011

Journal of Cell Science

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“…Chromosome condensation is an enigmatic phenomenon in mitosis (Bak et al 1977). Its occurrence is an essential step before subsequent sister chromatid separation in anaphase (for review, see Murray 1998).…”

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confidence: 99%

Fission yeast condensin complex: essential roles of non-SMC subunits for condensation and Cdc2 phosphorylation of Cut3/SMC4

Sutani¹,

Yuasa²,

Tomonaga³

et al. 1999

Genes & Development

246

280

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The condensin complex in frog extracts, containing two SMC (structural maintenance of chromosomes) and three non-SMC subunits, promotes mitotic chromosome condensation, and its supercoiling activity increases during mitosis by Cdc2 phosphorylation. Here, we report that fission yeast has the same five-member condensin complex, each of which is essential for mitotic condensation. The condensin complex was purified and the subunits were identified by microsequencing. Cnd1, Cnd2, and Cnd3, three non-SMC subunits showing a high degree of sequence conservation to frog subunits, are essential for viability, and their gene disruption leads to a phenotype indistinguishable from that observed in cut3-477 and cut14-208, known mutations in SMC4 and SMC2-like subunits. Condensin subunits tagged with GFP were observed to alter dramatically their localization during the cell cycle, enriched in the nucleus during mitosis, but cytoplasmic during other stages. This stage-specific alteration in localization requires mitosis-specific phosphorylation of the T19 Cdc2 site in Cut3. The T19 site is phosphorylated in vitro by Cdc2 kinase and shows the maximal phosphorylation in metaphase in vivo. Its alanine substitution mutant fails to suppress the temperature-sensitive phenotype of cut3-477, and shows deficiency in condensation, probably because Cut3 T19A remains cytoplasmic. Therefore, direct Cdc2 phosphorylation of fission yeast condensin may facilitate its nuclear accumulation during mitosis.

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“…BAK et al (1) suggested that the degree of compaction of DNA in human metaphase chromosomes is about 8,400.…”

Section: Resultsmentioning

confidence: 99%

Polytene Chromosomes Isolated From Nuclei of Tokunagayusurika Akamushi (Diptera, Chironomidae): Structural Transformation Caused by Salt, Detergent Polyanions and Enzymes

Yanagida

Masuda

1980

Dev Growth Differ

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Polytene chromosomes of Tokunagayusurika akamushi (Chironomidae) were isolated from nuclei of the salivary glands. The isolated chromosomes retained their normal morphology, revealing bands by phase contrast microscopy.Extensive structural transformations were observed when the isolated chromosomes were immersed in solutions of high salt concentration, polyanions (dextran sulfate and heparin), the ionic detergent SDS (0.01 % or more) or enzymes. The morphological changes could be described in terms of combinations of the following characteristics : (1) reversible or irreversible changes, (2) expansion or no expansion, (3) disappearance of bands, and (4) appearance of new bands. The structural features of the transformed chromosomes are described.Studies on polytene chromosomes at the molecular level have been hampered by the technical difficulty of isolating intact chromosomes in sufficient quantity. Various techniques have been employed to break the nuclear membranes, but few can be applied in biochemical investigations, either because the procedures cause destruction of the chromosomes or because insufficient material is available. Recently SEDAT and MAN~JELIDIS ( 14) reported the detailed fine structure of the polytene chromosomes of Drosophila free from nuclear membranes. They employed Buffer A (3) in which the chromosome structure appeared to be preserved, and removed the nuclear membranes by addition of 2-4 M urea. This paper reports that the procedure of SEDAT and MANUELIDIS could be applied for isolation of polytene chromosomes from the salivary gland nuclei of Tokunagayusurika akamushi. Characteristic chromomeres, or bands, in the isolated chromosomes could be seen by phasg contrast microscopy without staining. Their structural stability of the chromosomes was investigated, and the extensive structural changes caused by salt, detergent, polyanions and enzyme are described. MATERIALS AND METHODS Preparation of polytene chromosomesTokunagayusurika akamuslii (1 2), also called Orthocladius akamushi, is a common chironomid midge in Japan. This species is unique in morphology and development, and was therefore classified into a new genus, Tokunagayusurika, by SASA (12). The larvae of this species seem to be suitable material to use in large scale preparation of polytene chromosomes; the red colored larvae, which are as large as the largest chironomid Chironomus plumosus, are used as bait in fishing in Japan, and cultivated larvae

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Higher-order structure of human mitotic chromosomes.

Cited by 131 publications

References 9 publications

Condensin phosphorylated by the Aurora-B-like kinase Ark1 is continuously required until telophase in a mode distinct from Top2

Condensin phosphorylated by the Aurora-B-like kinase Ark1 is continuously required until telophase in a mode distinct from Top2

Fission yeast condensin complex: essential roles of non-SMC subunits for condensation and Cdc2 phosphorylation of Cut3/SMC4

Polytene Chromosomes Isolated From Nuclei of Tokunagayusurika Akamushi (Diptera, Chironomidae): Structural Transformation Caused by Salt, Detergent Polyanions and Enzymes

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