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Molecular Biology of the Cell Aug 2011In many eukaryotes, disruption of the spindle checkpoint protein Mad2 results in an increase in meiosis I nondisjunction, suggesting that Mad2 has a conserved role in...
In many eukaryotes, disruption of the spindle checkpoint protein Mad2 results in an increase in meiosis I nondisjunction, suggesting that Mad2 has a conserved role in ensuring faithful chromosome segregation in meiosis. To characterize the meiotic function of Mad2, we analyzed individual budding yeast cells undergoing meiosis. We find that Mad2 sets the duration of meiosis I by regulating the activity of APC(Cdc20). In the absence of Mad2, most cells undergo both meiotic divisions, but securin, a substrate of the APC/C, is degraded prematurely, and prometaphase I/metaphase I is accelerated. Some mad2Δ cells have a misregulation of meiotic cell cycle events and undergo a single aberrant division in which sister chromatids separate. In these cells, both APC(Cdc20) and APC(Ama1) are prematurely active, and meiosis I and meiosis II events occur in a single meiotic division. We show that Mad2 indirectly regulates APC(Ama1) activity by decreasing APC(Cdc20) activity. We propose that Mad2 is an important meiotic cell cycle regulator that ensures the timely degradation of APC/C substrates and the proper orchestration of the meiotic divisions.
Topics: Anaphase-Promoting Complex-Cyclosome; Cdc20 Proteins; Cell Cycle Proteins; Cell Nucleolus; Chromosomal Proteins, Non-Histone; Gene Knockout Techniques; Kinetochores; M Phase Cell Cycle Checkpoints; Mad2 Proteins; Meiosis; Metaphase; Nuclear Proteins; Prometaphase; Protein Tyrosine Phosphatases; Recombinant Fusion Proteins; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Spores; Time-Lapse Imaging; Ubiquitin-Protein Ligase Complexes
PubMed: 21697504
DOI: 10.1091/mbc.E11-04-0378 -
Cytogenetics and Cell Genetics 1998High resolution RBA-banded early prometaphase chromosomes of cattle (Bos taurus L.) and goat (Capra hircus L.), from thymidine synchronized lymphocyte cultures, are... (Comparative Study)
Comparative Study
High resolution RBA-banded early prometaphase chromosomes of cattle (Bos taurus L.) and goat (Capra hircus L.), from thymidine synchronized lymphocyte cultures, are compared at a level of 700 bands per haploid genome, with the purpose of detecting the extent of banding homologies between the two species and improving the resolution level of the ISCNDA (1989) standardized RBA-banded karyotypes. The results demonstrate that, at this level of resolution, at least 10 autosomes can be fully homologized between the two species, namely chromosomes 11, 16, 17, 18, 20, 21, 22, 23, 24 and 26, whereas the remaining autosomes show variations in some regions which require further investigations on more elongated chromosomes. Such investigations should also involve the use of GTG, GBG and RBG-banding techniques. These variations concern basically the relative distance of the bands from the centromeres or telomeres, appearance of subbands, and clustering of some positive bands.
Topics: Animals; Cattle; Cells, Cultured; Chromosome Banding; Chromosome Mapping; Goats; In Situ Hybridization, Fluorescence; Karyotyping; Lymphocytes; Male; Metaphase
PubMed: 9925950
DOI: 10.1159/000015148 -
Cancer Research Mar 2009The Aurora (Ipl) kinase family plays important roles in the regulation of mitosis and tumorigenesis. The tumor suppressor RASSF1A controls mitotic progression by...
The Aurora (Ipl) kinase family plays important roles in the regulation of mitosis and tumorigenesis. The tumor suppressor RASSF1A controls mitotic progression by regulating anaphase-promoting complex (APC)-Cdc20 activity and microtubule stability, but the mechanism by which this action is regulated has not been previously established. Here, we show that Aurora A and B associate with and phosphorylate RASSF1A on serine 203 in vivo at different times and in different subcellular compartments during mitosis. Notably, both depletion of Aurora A by RNA interference and expression of a nonphosphorylatable RASSF1A (S203A) mutant gene led to a marked delay in prometaphase progression. This is likely because of the failure of RASSF1A to dissociate from Cdc20, constitutive inhibition of APC-Cdc20, and accumulation of mitotic cyclins. In contrast, the delay in prometaphase progression caused by Aurora A depletion was largely normalized by phosphomimetic RASSF1A (S203D). Finally, RASSF1A phosphorylation on serine 203 was up-regulated in Aurora A-overexpressing human tumors. These findings indicate that Aurora A plays a critical role in RASSF1A-APC-Cdc20 regulatory mechanisms that control normal prometaphase progression and that are involved in tumorigenesis. [Cancer Res 2009;69(6):2314-23.
Topics: Aurora Kinases; Cdc20 Proteins; Cell Cycle Proteins; Cell Division; G2 Phase; HeLa Cells; Humans; Lung Neoplasms; Mitosis; Mutagenesis, Site-Directed; Phosphorylation; Prometaphase; Protein Serine-Threonine Kinases; RNA, Small Interfering; Subcellular Fractions; Tumor Suppressor Proteins; Up-Regulation
PubMed: 19276349
DOI: 10.1158/0008-5472.CAN-08-3984 -
Journal of Medical Genetics Jul 1988High resolution prometaphase chromosome banding has allowed the detection of discrete chromosome aberrations which escaped earlier metaphase examinations. Consistent... (Review)
Review
High resolution prometaphase chromosome banding has allowed the detection of discrete chromosome aberrations which escaped earlier metaphase examinations. Consistent tiny deletions have been detected in some well established malformation syndromes: an interstitial deletion in 15q11/12 in the majority of patients with the Prader-Willi syndrome and in a minority of patients with the Angelman (happy puppet) syndrome; a terminal deletion of 17p13.3 in most patients examined with the Miller-Dieker syndrome; an interstitial deletion of 8q23.3/24.1 in a large majority of patients with the Giedion-Langer syndrome; an interstitial deletion of 11p13 in virtually all patients with the WAGR (Wilms' tumour-aniridia-gonadoblastoma-retardation) syndrome; and an interstitial deletion in 22q11 in about one third of patients with the DiGeorge sequence. In addition, a combination of chromosome prometaphase banding and DNA marker studies has allowed the localisation of the genes for retinoblastoma and for Wilms' tumour and the clarification of both the autosomal recessive nature of the mutation and the possible somatic mutations by which the normal allele can be lost in retina and kidney cells. After a number of X linked genes had been mapped, discrete deletions in the X chromosome were detected by prometaphase banding with specific attention paid to the sites of the gene(s) in males who had from one to up to four different X linked disorders plus mental retardation. Furthermore, the detection of balanced translocations in probands with disorders caused by autosomal dominant or X linked genes has allowed a better insight into the localisation of these genes. In some females with X linked disorders, balanced X; autosomal translocations have allowed the localisation of X linked genes at the breakpoint on the X chromosome. Balanced autosome; autosome translocations segregating with autosomal dominant conditions have provided some clues to the gene location of these conditions. In two conditions, Greig cephalopolysyndactyly and dominant aniridia, two translocation families with one common breakpoint have allowed quite a confident location of the genes at the common breakpoint at 7p13 and 11p13, respectively.
Topics: Chromosome Aberrations; Chromosome Banding; Chromosome Deletion; Chromosome Disorders; Chromosome Mapping; Genetic Markers; Humans; Karyotyping; Sex Chromosome Aberrations; Syndrome; Translocation, Genetic
PubMed: 3050093
DOI: 10.1136/jmg.25.7.454 -
Biochimica Et Biophysica Acta Sep 2008Aneuploidy, an abnormal number of chromosomes, is a trait shared by most solid tumors. Chromosomal instability (CIN) manifested as aneuploidy might promote tumorigenesis... (Review)
Review
Aneuploidy, an abnormal number of chromosomes, is a trait shared by most solid tumors. Chromosomal instability (CIN) manifested as aneuploidy might promote tumorigenesis and cause increased resistance to anti-cancer therapies. The mitotic checkpoint or spindle assembly checkpoint is a major signaling pathway involved in the prevention of CIN. We review current knowledge on the contribution of misregulation of mitotic checkpoint proteins to tumor formation and will address to what extent this contribution is due to chromosome segregation errors directly. We propose that both checkpoint and non-checkpoint functions of these proteins contribute to the wide array of oncogenic phenotypes seen upon their misregulation.
Topics: Aneuploidy; Animals; Cdc20 Proteins; Cell Cycle Proteins; Cell Survival; Chromosomal Instability; Chromosome Segregation; Humans; Mad2 Proteins; Mice; Mitosis; Models, Animal; Neoplasms; Nuclear Proteins; Prometaphase; Protein Serine-Threonine Kinases; Saccharomyces cerevisiae Proteins
PubMed: 18472014
DOI: 10.1016/j.bbcan.2008.04.001 -
Journal of Visualized Experiments : JoVE Dec 2017In humans, chromosome segregation errors in oocytes are responsible for the majority of miscarriages and birth defects. Moreover, as women age, their risk of conceiving...
In humans, chromosome segregation errors in oocytes are responsible for the majority of miscarriages and birth defects. Moreover, as women age, their risk of conceiving an aneuploid fetus increases dramatically and this phenomenon is known as the maternal age effect. One requirement for accurate chromosome segregation during the meiotic divisions is maintenance of sister chromatid cohesion during the extended prophase period that oocytes experience. Cytological evidence in both humans and model organisms suggests that meiotic cohesion deteriorates during the aging process. In addition, segregation errors in human oocytes are most prevalent during meiosis I, consistent with premature loss of arm cohesion. The use of model organisms is critical for unraveling the mechanisms that underlie age-dependent loss of cohesion. Drosophila melanogaster offers several advantages for studying the regulation of meiotic cohesion in oocytes. However, until recently, only genetic tests were available to assay for loss of arm cohesion in oocytes of different genotypes or under different experimental conditions. Here, a detailed protocol is provided for using fluorescence in situ hybridization (FISH) to directly visualize defects in arm cohesion in prometaphase I and metaphase I arrested Drosophila oocytes. By generating a FISH probe that hybridizes to the distal arm of the X chromosome and collecting confocal Z stacks, a researcher can visualize the number of individual FISH signals in three dimensions and determine whether sister chromatid arms are separated. The procedure outlined makes it possible to quantify arm cohesion defects in hundreds of Drosophila oocytes. As such, this method provides an important tool for investigating the mechanisms that contribute to cohesion maintenance as well as the factors that lead to its demise during the aging process.
Topics: Animals; Drosophila; Female; Humans; In Situ Hybridization, Fluorescence; Metaphase; Oocytes; Prometaphase
PubMed: 29286418
DOI: 10.3791/56802 -
Human Genetics 1982A method is described for the analysis of chromosomes in prophase and early metaphase. It involves culturing the lymphocytes in medium RPMI-1640, supplemented with 10%...
A method is described for the analysis of chromosomes in prophase and early metaphase. It involves culturing the lymphocytes in medium RPMI-1640, supplemented with 10% autologous plasma instead of fetal bovine serum. Living cells are treated with actinomycin D and colcemid for 1 h prior to harvest and harvested early at 65 h of incubation, using a hypotonic solution formulated by Ohnuki (1968). The method has been tested on several hundred clinical samples on a routine basis. On average, 30% of the dividing cells were in prometaphase.
Topics: Azure Stains; Cell Division; Chromosome Banding; Chromosomes, Human; Colchicine; Culture Media; Dactinomycin; Humans; Hypertonic Solutions; Lymphocytes; Metaphase; Plasma; Prophase
PubMed: 7106769
DOI: 10.1007/BF00569213 -
Biochemical Pharmacology Apr 2015The present study sought to determine the correlation between 2-methoxyestradiol (2-MeO-E2)-induced cell cycle arrest and 2-MeO-E2-induced apoptosis. Exposure of Jurkat...
The present study sought to determine the correlation between 2-methoxyestradiol (2-MeO-E2)-induced cell cycle arrest and 2-MeO-E2-induced apoptosis. Exposure of Jurkat T cell clone (JT/Neo) to 2-MeO-E2 (0.5-1.0 μM) caused G2/M arrest, Bak activation, Δψm loss, caspase-9 and -3 activation, PARP cleavage, intracellular ROS accumulation, and apoptotic DNA fragmentation, whereas none of these events except for G2/M arrest were induced in Jurkat T cells overexpressing Bcl-2 (JT/Bcl-2). Under these conditions, Cdk1 phosphorylation at Thr-161 and dephosphorylation at Tyr-15, up-regulation of cyclin B1 expression, histone H1 phosphorylation, Cdc25C phosphorylation at Thr-48, Bcl-2 phosphorylation at Thr-56 and Ser-70, Mcl-1 phosphorylation at Ser-159/Thr-163, and Bim phosphorylation were detected irrespective of Bcl-2 overexpression. Concomitant treatment of JT/Neo cells with 2-MeO-E2 and the G1/S blocking agent aphidicolin resulted in G1/S arrest and abrogation of all apoptotic events, including Cdk1 activation, phosphorylation of Bcl-2, Mcl-1 and Bim, and ROS accumulation. The 2-MeO-E2-induced phosphorylation of Bcl-2 family proteins and mitochondrial apoptotic events were suppressed by a Cdk1 inhibitor, but not by an Aurora A kinase (AURKA), Aurora B kinase (AURKB), JNK, or p38 MAPK inhibitor. Immunofluorescence microscopic analysis revealed that 2-MeO-E2-induced mitotic arrest was caused by mitotic spindle network impairment and prometaphase arrest. Whereas 10-20 μM 2-MeO-E2 reduced the proportion of intracellular polymeric tubulin to monomeric tubulin, 0.5-5.0 μM 2-MeO-E2 increased it. These results demonstrate that the apoptogenic effect of 2-MeO-E2 (0.5-1.0 μM) was attributable to mitotic spindle defect-mediated prometaphase arrest, Cdk1 activation, phosphorylation of Bcl-2, Mcl-1, and Bim, and activation of Bak and mitochondria-dependent caspase cascade.
Topics: 2-Methoxyestradiol; Antineoplastic Agents; Apoptosis; Caspase 3; Enzyme Activation; Estradiol; G1 Phase Cell Cycle Checkpoints; G2 Phase Cell Cycle Checkpoints; Humans; Jurkat Cells; Microtubules; Mitochondria; Mitosis; Phosphorylation; Prometaphase; Proto-Oncogene Proteins c-bcl-2; Spindle Apparatus
PubMed: 25732194
DOI: 10.1016/j.bcp.2015.02.011 -
Chromosoma 1975Chromosome and granule movements in meiotic prophase and prometaphase have been studied by time-lapse cinemicrography in live spermatocytes of the house cricket, Acheta...
Chromosome and granule movements in meiotic prophase and prometaphase have been studied by time-lapse cinemicrography in live spermatocytes of the house cricket, Acheta domesticus. Chromosome movements in prophase cells, up to one hour or more before breakdown of the nuclear envelope, are described. These movements are frequent but saltatory; are based mostly at chromosome ends but also at kinetochores; occur in very intimate association with the inside of the nuclear envelope; are directed towards and away from the extranuclear centres (centrioles); tend weakly to accumulate bivalents round the two centres and reach a velocity of 0.65 micron/sec. Saltatory movements in granules associated with extranuclear asters are remarkably similar to basic characteristics to the intranuclear chromosome movements. Surprisingly, the chromosome movements (and those granules) are reversably blocked by colcemid (but not lumi-colcemid), and yet occur in the apparent absence of an intranuclear envelope. However, kinetochore movements in very early prometaphase are similar in velocity and other respects to prophase movements; later prometaphase movements are clearly slower, and those of anaphase very much slower still. -The prophase movements suggest a two component model for motion: a non-microtubule, linear force producer together withrotubules with a skeletal, orientational role. Arguably, both these components are also necessary for chromosome movements in prometaphase and anaphase.
Topics: Animals; Cell Nucleus; Chromosomes; Cytoplasmic Granules; Demecolcine; Male; Meiosis; Micromanipulation; Microtubules; Models, Biological; Movement; Orthoptera; Photomicrography; Spermatozoa; Statistics as Topic; Time Factors
PubMed: 1132283
DOI: 10.1007/BF00285133 -
Cell Motility and the Cytoskeleton 1992Injection of CREST antikinetochore antiserum (AKA) containing antibodies to the kinetochore into living prometaphase PtK2 cells decreased chromosome velocity to near...
Injection of CREST antikinetochore antiserum (AKA) containing antibodies to the kinetochore into living prometaphase PtK2 cells decreased chromosome velocity to near zero. Injection of either phosphate-buffered saline or CREST antiserum without antikinetochore antibodies (antikinetochore negative: AKN) had no effect on prometaphase oscillations. AKA antiserum injected into anaphase cells at the beginning of chromatid separation had no effect on anaphase chromosome velocity, spindle elongation, or cytokinesis. Visible binding of antikinetochore antibodies in prometaphase cells at room temperature occurred between 5 and 15 minutes after injection. Anaphase cells injected at the beginning of chromatid separation had bound antibody at the end of anaphase. AKA antiserum recognizes in Western blots proteins associated with the primary constriction: CENP-B, -C, and -D, as reported by other workers. The control antiserum, AKN, does not recognize these proteins. These results imply that the antigens recognized by CREST antibodies are important for chromosome movement. Whether or not these antigens are themselves motor molecules cannot be addressed by the present data. In addition, the results suggest that these antigens are not involved in an important way in anaphase movement.
Topics: Anaphase; Animals; Autoantibodies; Autoimmune Diseases; Cell Division; Cell Line, Transformed; Centromere; Chromosomes; Fluorescent Antibody Technique; Macropodidae; Metaphase; Scleroderma, Systemic; Spindle Apparatus
PubMed: 1451187
DOI: 10.1002/cm.970230208