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Science (New York, N.Y.) Dec 1995The relative locations of several chromosomes within wheel-shaped prometaphase chromosome rosettes of human fibroblasts and HeLa cells were determined with fluorescence...
The relative locations of several chromosomes within wheel-shaped prometaphase chromosome rosettes of human fibroblasts and HeLa cells were determined with fluorescence hybridization. Homologs were consistently positioned on opposite sides of the rosette, which suggests that chromosomes are separated into two haploid sets, each derived from one parent. The relative locations of chromosomes on the rosette were mapped by dual hybridizations. The data suggest that the chromosome orders within the two haploid sets are antiparallel. This chromosome arrangement in human cells appears to be both independent of cell type- and species-specific and may influence chromosome topology throughout the cell cycle.
Topics: Cells, Cultured; Chromosomes; DNA Probes; Fibroblasts; HeLa Cells; Humans; In Situ Hybridization, Fluorescence; Metaphase
PubMed: 8525379
DOI: 10.1126/science.270.5243.1831 -
Science (New York, N.Y.) Nov 2009Two critical stages of mammalian oocyte regulation are prophase I arrest, which is important for sustaining the oocyte pool, and the progression through meiosis I (MI)...
Two critical stages of mammalian oocyte regulation are prophase I arrest, which is important for sustaining the oocyte pool, and the progression through meiosis I (MI) to produce fertilizable eggs. We have found that the spindle assembly checkpoint protein BubR1 regulates both stages in mouse oocytes. We show that oocytes depleted of BubR1 cannot sustain prophase I arrest and readily undergo germinal vesicle breakdown, a marker for reentry into MI. BubR1-depleted oocytes then arrest before completing MI, marked by failure of polar body extrusion. Both meiotic defects in BubR1-depleted oocytes are due to reduced activity of the master regulator known as the anaphase-promoting complex (APC), brought about through diminished levels of the APC coactivator Cdh1.
Topics: Anaphase-Promoting Complex-Cyclosome; Animals; Carrier Proteins; Cdc20 Proteins; Cdh1 Proteins; Cell Cycle Proteins; Cyclin B1; Female; Gene Silencing; Meiosis; Meiotic Prophase I; Mice; Oocytes; Prometaphase; Protein Serine-Threonine Kinases; Securin; Ubiquitin-Protein Ligase Complexes
PubMed: 19965510
DOI: 10.1126/science.1175326 -
The Journal of Cell Biology Aug 2010The ubiquitin ligase anaphase-promoting complex/cyclosome (APC/C) is activated at prometaphase by mitotic phosphorylation and binding of its activator, Cdc20. This...
The ubiquitin ligase anaphase-promoting complex/cyclosome (APC/C) is activated at prometaphase by mitotic phosphorylation and binding of its activator, Cdc20. This initiates cyclin A degradation, whereas cyclin B1 is stabilized by the spindle checkpoint. Upon checkpoint release, the RXXL destruction box (D box) was proposed to direct cyclin B1 to core APC/C or Cdc20. In this study, we report that endogenous cyclin B1-Cdk1 is recruited to checkpoint-inhibited, phosphorylated APC/C in prometaphase independently of Cdc20 or the cyclin B1 D box. Like cyclin A, cyclin B1 binds the APC/C by the Cdk cofactor Cks and the APC3 subunit. Prior binding to APC/C(Cdc20) makes cyclin B1 a better APC/C substrate in metaphase, driving mitotic exit and cytokinesis. We conclude that in prometaphase, the phosphorylated APC/C can recruit both cyclin A and cyclin B1 in a Cks-dependent manner. This suggests that the spindle checkpoint blocks D box recognition of APC/C-bound cyclin B1, whereas distinctive complexes between the N terminus of cyclin A and Cdc20 evade checkpoint control.
Topics: Anaphase-Promoting Complex-Cyclosome; Animals; CDC2 Protein Kinase; CDC2-CDC28 Kinases; Carrier Proteins; Cdc20 Proteins; Cell Cycle Proteins; Cell Line; Cyclin A; Cyclin B1; Cyclin-Dependent Kinases; Humans; Mitosis; Neoplasm Proteins; Nocodazole; Phosphorylation; Prometaphase; Protein Binding; Protein Kinase Inhibitors; Purines; RNA Interference; Recombinant Fusion Proteins; Roscovitine; Securin; Tubulin Modulators; Ubiquitin; Ubiquitin-Protein Ligase Complexes
PubMed: 20733055
DOI: 10.1083/jcb.200912084 -
BMC Research Notes Apr 2009NuMA is a protein that has been previously shown to play a role in focusing microtubules at the mitotic spindle poles. However, most previous work relies on experimental...
BACKGROUND
NuMA is a protein that has been previously shown to play a role in focusing microtubules at the mitotic spindle poles. However, most previous work relies on experimental methods that might cause dominant side effects on spindle formation, such as microinjection of antibodies, overexpression of mutant protein, or immunodepletion of NuMA-containing protein complexes.
FINDINGS
To circumvent these technical problems, we performed siRNA experiments in which we depleted the majority of NuMA in human cultured cells. Depleted mitotic cells show a prolonged duration of prometaphase, with spindle pole defects and with unattached, unaligned chromosomes.
CONCLUSION
Our data confirm that NuMA is important for spindle pole formation, and for cohesion of centrosome-derived microtubules with the bulk of spindle microtubules. Our findings of NuMA-dependent defects in chromosome alignment suggest that NuMA is involved in stabilizing kinetochore fibres.
PubMed: 19400937
DOI: 10.1186/1756-0500-2-64 -
PloS One 2017Mitotic prophase chromosome condensation plays an essential role in nuclear division being therefore regulated by highly conserved mechanisms. However, degrees of...
Mitotic prophase chromosome condensation plays an essential role in nuclear division being therefore regulated by highly conserved mechanisms. However, degrees of chromatin condensation in prophase-prometaphase cells may vary along the chromosomes resulting in specific condensation patterns. We examined different condensation patterns (CPs) of prophase and prometaphase chromosomes and investigated their relationship with genome size and distribution of histone H4 acetylated at lysine 5 (H4K5ac) in 17 plant species. Our results showed that most species with small genomes (2C < 5 pg) (Arachis pusilla, Bixa orellana, Costus spiralis, Eleutherine bulbosa, Indigofera campestris, Phaseolus lunatus, P. vulgaris, Poncirus trifoliata, and Solanum lycopersicum) displayed prophase chromosomes with late condensing terminal regions that were highly enriched in H4K5ac, and early condensing regions with apparently non-acetylated proximal chromatin. The species with large genomes (Allium cepa, Callisia repens, Araucaria angustifolia and Nothoscordum pulchellum) displayed uniformly condensed and acetylated prophase/prometaphase chromosomes. Three species with small genomes (Eleocharis geniculata, Rhynchospora pubera, and R. tenuis) displayed CP and H4K5ac labeling patterns similar to species with large genomes, whereas a forth species (Emilia sonchifolia) exhibited a gradual chromosome labeling, being more acetylated in the terminal regions and less acetylated in the proximal ones. The nucleolus organizer chromatin was the only chromosomal region that in prometaphase or metaphase could be hyperacetylated, hypoacetylated or non-acetylated, depending on the species. Our data indicate that the CP of a plant chromosome complement is influenced but not exclusively determined by nuclear and chromosomal DNA contents, whereas the CP of individual chromosomes is clearly correlated with H4K5ac distribution.
Topics: Acetylation; Chromatin Assembly and Disassembly; Chromosomes, Plant; Genome Size; Genome, Plant; Heterochromatin; Histones; Karyotyping; Plant Proteins; Plants; Prometaphase; Prophase; Protein Processing, Post-Translational; Species Specificity
PubMed: 28854212
DOI: 10.1371/journal.pone.0183341 -
Human Genetics Dec 1978We have constructed ideograms of human prometaphase chromosomes from synchronized and from standard 72-h lymphocyte cultures. G banding was achieved by a trypsin-Giemsa...
We have constructed ideograms of human prometaphase chromosomes from synchronized and from standard 72-h lymphocyte cultures. G banding was achieved by a trypsin-Giemsa (or Wright's stain) method. In addition to light (white) and dark (black) bands, we have distinguished three different shades of grey. This distinction is essential for proper identification of the increasing number of bands displayed by high-resolution chromosomes. The relative amount of chromatin in each category of staining intensity has been calculated and expressed as 'light value.' The ideograms represent the maximal number of bands discernible with some consistency on prometaphase chromosomes, i.e., 721 euchromatic and 62 'variable' heterochromatic or heteromorphic bands. The ideograms are based on measurements. On selected printed copies of each chromosome derived from different cells and different individuals, the relative width of each band was measured in relation to the length of the respective chromosome arm. The measurements per chromosome were averaged and used for construction of the ideograms. The distance of each border between bands or sub-bands from the centromere has been calculated on a relative scale, with positions 0 at the centromere and 1.0 at the p terminus of q terminus. The numbering system for bands and sub-bands follows the Paris Conference (1971) recommendations.
Topics: Cell Cycle; Cells, Cultured; Chromosome Banding; Chromosomes, Human; Humans; Lymphocytes; Prophase
PubMed: 738718
DOI: 10.1007/BF00286957 -
Methods in Enzymology 2018Our conventional understanding of the process of DNA replication is that it occurs in the S-phase of the cell division cycle. However, during investigations into the... (Review)
Review
Our conventional understanding of the process of DNA replication is that it occurs in the S-phase of the cell division cycle. However, during investigations into the mechanism by which common fragile sites (CFSs) drive genome instability, we observed that some DNA synthesis was still occurring in early mitosis at these loci. This curious phenomenon of mitotic DNA synthesis (which we now term "MiDAS") appears to be a form of break-induced DNA replication (BIR), a DNA repair process based on homologous recombination that has been characterized in detail only in lower eukaryotes. During MiDAS, it is proposed that parts of the human genome that are not fully replicated when cells enter mitotic prophase complete their replicative cycle at that point. To date, the loci that most depend upon this process are those whose replication can be affected by oncogene-induced DNA replication stress (RS), most notably, CFSs. From our studies, it is clear that the successful completion of MiDAS at CFSs can minimize chromosome missegregation and nondisjunction. Nevertheless, it is still not clear which loci that can undergo MiDAS, whether MiDAS is associated with mutations or genome rearrangements, or whether MiDAS really is a form of BIR. In this review, we describe methods for detecting MiDAS both in prometaphase cells and directly on isolated metaphase chromosomes. In addition, we have included methods for combining MiDAS detection either with immunofluorescence (IF) detection of proteins that are recruited to the MiDAS loci, or with fluorescence in situ hybridization using probes that target specific genomic loci.
Topics: Cell Line; DNA; Genetic Techniques; Humans; Metaphase; Mitosis; Prometaphase; Recombinational DNA Repair
PubMed: 29523241
DOI: 10.1016/bs.mie.2017.11.025 -
Clinical Genetics Sep 1985Tables of prometaphase chromosome segment sizes (750 band stage) are presented in units of percentage of haploid autosome length designating the distances from interband...
Tables of prometaphase chromosome segment sizes (750 band stage) are presented in units of percentage of haploid autosome length designating the distances from interband borders to telomeres. The Tables utilize ISCN (1981) nomenclature.
Topics: Chromosome Banding; Chromosomes, Human; Haploidy; Humans; Metaphase; Prophase
PubMed: 4064359
DOI: 10.1111/j.1399-0004.1985.tb00389.x -
Developmental Cell Apr 2013The functions of the Ndc80/Hec1 subunit of the highly conserved Ndc80 kinetochore complex are normally restricted to M phase when it exerts a pivotal kinetochore-based...
The functions of the Ndc80/Hec1 subunit of the highly conserved Ndc80 kinetochore complex are normally restricted to M phase when it exerts a pivotal kinetochore-based role. Here, we find that in mouse oocytes, depletion of Hec1 severely compromises the G2-M transition because of impaired activation of cyclin-dependent kinase 1 (Cdk1). Unexpectedly, impaired M phase entry is due to instability of the Cdk1-activating subunit, cyclin B2, which cannot be covered by cyclin B1. Hec1 protects cyclin B2 from destruction by the Cdh1-activated anaphase-promoting complex (APC(Cdh1)) and remains important for cyclin B2 stabilization during early M phase, required for the initial stages of acentrosomal spindle assembly. By late M phase, however, Hec1 and cyclin B2 become uncoupled, and although Hec1 remains stable, APC(Cdc20) triggers cyclin B2 destruction. These data identify another dimension to Hec1 function centered on M phase entry and early prometaphase progression and challenge the view that cyclin B2 is completely dispensable in mammals.
Topics: Anaphase-Promoting Complex-Cyclosome; Animals; Blotting, Western; Carrier Proteins; Cdh1 Proteins; Cell Cycle Proteins; Chromosomes, Mammalian; Cyclin B2; Female; Kinetochores; Meiosis; Metaphase; Mice; Microtubule-Associated Proteins; Oocytes; Prometaphase; Prophase; Protein Stability; Proteolysis; Securin; Spindle Apparatus; Time Factors; Ubiquitin-Protein Ligase Complexes
PubMed: 23541922
DOI: 10.1016/j.devcel.2013.02.008 -
Journal of Cell Science Nov 1983Individual spindle fibres in prometaphase spermatocytes of the cricket, Neocurtilla hexadactyla, were irradiated with an ultraviolet microbeam. The stretched...
Individual spindle fibres in prometaphase spermatocytes of the cricket, Neocurtilla hexadactyla, were irradiated with an ultraviolet microbeam. The stretched heteromorphic bivalent (X2Y) contracted to about 75% of its pre-irradiation length after irradiation of either of its two oppositely directed spindle fibres (i.e., the X2 or Y spindle fibre). The X2Y bivalent also contracted after irradiation of the connection between the kinetochores of the univalent X1 chromosome and the Y chromosome but it did not contract after irradiation of autosomal spindle fibres or of the spindle fibre of the X1 univalent sex chromosome. The spindles sometimes shrank after irradiation, but contraction of the X2Y bivalent was independent of spindle shrinkage. The data strongly suggest that the oppositely directed forces on a bivalent are not independent. One reason is that the X2Y contractions were asymmetrical: during contraction one kinetochore remained fixed in position while the other kinetochore (facing the opposite pole) moved towards the equator. In 75% of the cases the stationary kinetochore was associated with the irradiated spindle fibre. Thus, we suggest that the irradiation of a spindle fibre produces a state analogous to rigor in the irradiated spindle fibre and, because of effects on the control system, produces relaxation of tension in the oppositely directed non-irradiated spindle fibre, so that the kinetochore associated with the non-irradiated spindle fibre moves towards the equator. These experiments have identified a control system that coordinates force production to opposite poles.
Topics: Animals; Chromosomes; Dose-Response Relationship, Radiation; Male; Metaphase; Orthoptera; Spermatocytes; Spermatozoa; Spindle Apparatus; Ultraviolet Rays
PubMed: 6662864
DOI: 10.1242/jcs.64.1.69