scholarly journals DISTRIBUTION OF LABELED CHROMATIN

1970 ◽  
Vol 47 (1) ◽  
pp. 99-106 ◽  
Author(s):  
Jean H. Priest ◽  
Robert H. Shikes
Keyword(s):  
Mammalian Cells ◽  
Tritiated Thymidine ◽  
Daughter Cells ◽  
Chromatid Segregation ◽  
Count Distribution ◽  
Random Segregation ◽  
Single Grain ◽  
Discrete Populations ◽  
Rule Out ◽  
Diploid Human Fibroblast

The question of whether distribution of chromatids to daughter cells in mitosis is a random or nonrandom process was investigated by study of the distribution of labeled chromatin in anaphase pairs at M1 and M2 after a pulse of tritiated thymidine. Diploid and tetraploid rat and diploid human fibroblast-like cells in serial monlayer culture were synchronized by two different methods to "purify" M1 and M2 anaphases: metaphase shake, and FUdR block to DNA synthesis followed by exogenous thymidine. Exposed grains of NTB-2 emulsion were counted over M1 and M2 anaphase pairs. An analysis (by pair) of diploid M2 anaphase grain counts showed two discrete populations of daughters with less and with more radioactivity. A similar analysis of diploid M1 and tetrapolid M2 anaphases showed a single grain-count distribution. These findings may support a nonrandom model of chromatid segregation for diploid mammalian cells but do not rule out random segregation until sound mathematical models are formulated for expected random grain distributions in M2 anaphases of cells with differing numbers of chromosomes.

scholarly journals Random segregation of chromatids at mitosis in Saccharomyces cerevisiae.

Genetics ◽  
1991 ◽  
Vol 127 (3) ◽  
pp. 463-473 ◽  
Author(s):  
M W Neff ◽  
D J Burke
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Sister Chromatid ◽  
Mitotic Chromosome ◽  
Sister Chromatid Exchanges ◽  
Daughter Cells ◽  
Quantitative Measurements ◽  
Chromatid Segregation ◽  
Dna Strands ◽  
Random Segregation ◽  
Chromatid Exchanges

Abstract Previous experiments suggest that mitotic chromosome segregation in some fungi is a nonrandom process in which chromatids of the same replicative age are destined for cosegregation. We have investigated the pattern of chromatid segregation in Saccharomyces cerevisiae by labeling the DNA of a strain auxotrophic for thymidine with 5-bromodeoxyuridine. The fate of DNA strands was followed qualitatively by immunofluorescence microscopy and quantitatively by microphotometry using an anti-5-bromodeoxyuridine monoclonal antibody. Chromatids of the same replicative age were distributed randomly to daughter cells at mitosis. Quantitative measurements showed that the amount of fluorescence in the daughter nuclei derived from parents with hemilabeled chromosomes diminished in intensity by one half. The concentration of 5-bromodeoxyuridine used in the experiments had little effect on the frequency of either homologous or sister chromatid exchanges. We infer that the 5-bromodeoxyuridine was distributed randomly due to mitotic segregation of chromatids and not via sister chromatid exchanges.

Topoisomerase II inhibition prevents anaphase chromatid segregation in mammalian cells independently of the generation of DNA strand breaks

1993 ◽  
Vol 105 (2) ◽  
pp. 563-569 ◽  
Author(s):  
D.J. Clarke ◽  
R.T. Johnson ◽  
C.S. Downes
Keyword(s):  
Chromosome Segregation ◽  
Mammalian Cells ◽  
Topoisomerase Ii ◽  
Dna Strand Breaks ◽  
Temperature Sensitive ◽  
Dna Topoisomerase ◽  
Strand Breaks ◽  
Temperature Sensitive Mutants ◽  
Chromatid Segregation ◽  
Dna Strand

Yeast temperature-sensitive mutants of DNA topoisomerase II are incapable of chromosome condensation and anaphase chromatid segregation. In mammalian cells, topoisomerase II inhibitors such as etoposide (VP-16-123) have similar effects. Unfortunately, conclusions drawn from work with mammalian cells have been limited by the fact that the standard inhibitors of topoisomerase II also generate DNA strand breaks, which when produced by other agents (e.g. ionizing radiation) are known to affect progression into and through mitosis. Here we show that the anti-tumour agent ICRF-193, recently identified as a topoisomerase II inhibitor operating by a non-standard mechanism, generates neither covalent complexes between topoisomerase II and DNA, nor adjacent DNA strand breaks, in mitotic HeLa. However, the drug does prevent anaphase segregation in HeLa and PtK2 cells, with effects similar to those of etoposide. We therefore conclude that topoisomerase II function is required for anaphase chromosome segregation in mammalian cells, as it is in yeast.

scholarly journals Sphingolipid Domains in the Plasma Membranes of Fibroblasts Are Not Enriched with Cholesterol

2013 ◽  
Vol 288 (23) ◽  
pp. 16855-16861 ◽  
Author(s):  
Jessica F. Frisz ◽  
Haley A. Klitzing ◽  
Kaiyan Lou ◽  
Ian D. Hutcheon ◽  
Peter K. Weber ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Plasma Membrane ◽  
High Resolution ◽  
Mammalian Cells ◽  
Plasma Membranes ◽  
Secondary Ion Mass Spectrometry ◽  
Cholesterol Depletion ◽  
Fibroblast Cells ◽  
Secondary Ion ◽  
Rule Out

The plasma membranes of mammalian cells are widely expected to contain domains that are enriched with cholesterol and sphingolipids. In this work, we have used high-resolution secondary ion mass spectrometry to directly map the distributions of isotope-labeled cholesterol and sphingolipids in the plasma membranes of intact fibroblast cells. Although acute cholesterol depletion reduced sphingolipid domain abundance, cholesterol was evenly distributed throughout the plasma membrane and was not enriched within the sphingolipid domains. Thus, we rule out favorable cholesterol-sphingolipid interactions as dictating plasma membrane organization in fibroblast cells. Because the sphingolipid domains are disrupted by drugs that depolymerize the cells actin cytoskeleton, cholesterol must instead affect the sphingolipid organization via an indirect mechanism that involves the cytoskeleton.

scholarly journals Inhibition of Chromosomal Separation Provides Insights into Cleavage Furrow Stimulation in Cultured Epithelial Cells

1998 ◽  
Vol 9 (8) ◽  
pp. 2173-2184 ◽  
Author(s):  
Sally P. Wheatley ◽  
Christopher B. O’Connell ◽  
Yu-li Wang
Keyword(s):  
Epithelial Cells ◽  
Mammalian Cells ◽  
Topoisomerase Ii ◽  
Rat Kidney ◽  
Myosin Ii ◽  
Daughter Cells ◽  
Irregular Shapes ◽  
Anaphase Onset ◽  
Cultured Epithelial Cells ◽  
Normal Rat

While astral microtubules are believed to be primarily responsible for the stimulation of cytokinesis in Echinodermembryos, it has been suggested that a signal emanating from the chromosomal region and mediated by the interzonal microtubules stimulates cytokinesis in cultured mammalian cells. To test this hypothesis, we examined cytokinesis in normal rat kidney cells treated with an inhibitor of topoisomerase II, (+)-1,2-bis(3,5-dioxopiperaz-inyl-1-yl)propane, which prevents the separation of sister chromatids and the formation of a spindle interzone. The majority of treated cells showed various degrees of abnormality in cytokinesis. Furrows frequently deviated from the equatorial plane, twisting daughter cells into irregular shapes. Some cells developed furrows in regions outside the equator or far away from the spindle. In addition, F-actin and myosin II accumulated at the lateral ingressing margins but did not form a continuous band along the equator as in control cells. Imaging of microinjected 5- (and 6-) carboxymtetramethylrhodamine-tubulin revealed that a unique set of microtubules projected out from the chromosomal vicinity upon anaphase onset. These microtubules emanated toward the lateral cortex, where they delineated sites of microtubule bundle formation, cortical ingression, and F-actin and myosin II accumulation. As centrosome integrity and astral microtubules appeared unperturbed by (+)-1,2-bis(3,5-dioxopiperaz-inyl-1-yl)propane treatment, the present observations cannot be easily explained by the conventional model involving astral microtubules. We suggest that in cultured epithelial cells the organization of the chromosomes dictates the organization of midzone microtubules, which in turn determines and maintains the cleavage activity.

scholarly journals Kinetochore-microtubule stability governs the metaphase requirement for Eg5

2014 ◽  
Vol 25 (13) ◽  
pp. 2051-2060 ◽  
Author(s):  
A. Sophia Gayek ◽  
Ryoma Ohi
Keyword(s):  
Physical Properties ◽  
Human Cell ◽  
Mitotic Spindle ◽  
Mammalian Cells ◽  
Human Cell Lines ◽  
Bipolar Spindle ◽  
Daughter Cells ◽  
Microtubule Stability ◽  
The Stability ◽  
Bipolar Spindles

The mitotic spindle is a bipolar, microtubule (MT)-based cellular machine that segregates the duplicated genome into two daughter cells. The kinesin-5 Eg5 establishes the bipolar geometry of the mitotic spindle, but previous work in mammalian cells suggested that this motor is unimportant for the maintenance of spindle bipolarity. Although it is known that Kif15, a second mitotic kinesin, enforces spindle bipolarity in the absence of Eg5, how Kif15 functions in this capacity and/or whether other biochemical or physical properties of the spindle promote its bipolarity have been poorly studied. Here we report that not all human cell lines can efficiently maintain bipolarity without Eg5, despite their expressing Kif15. We show that the stability of chromosome-attached kinetochore-MTs (K-MTs) is important for bipolar spindle maintenance without Eg5. Cells that efficiently maintain bipolar spindles without Eg5 have more stable K-MTs than those that collapse without Eg5. Consistent with this observation, artificial destabilization of K-MTs promotes spindle collapse without Eg5, whereas stabilizing K-MTs improves bipolar spindle maintenance without Eg5. Our findings suggest that either rapid K-MT turnover pulls poles inward or slow K-MT turnover allows for greater resistance to inward-directed forces.

Studies of the 'Adaptive' Repair Response in Human Lymphocytes and V79 Cells after Treatment with MNNG and MNU

1988 ◽  
Vol 7 (4) ◽  
pp. 337-341 ◽  
Author(s):  
D. Anderson ◽  
P. Fisher ◽  
P.C. Jenkinson ◽  
B.J. Phillips
Keyword(s):  
Adaptive Response ◽  
Mammalian Cells ◽  
Tritiated Thymidine ◽  
Human Lymphocytes ◽  
Alternative Methods ◽  
X Rays ◽  
V79 Cells ◽  
Repair Mechanisms ◽  
Experimental Protocols ◽  
Mutagenic Effects

In bacteria, there is evidence that a damage inducible repair response system known as the adaptive response exists since pretreatment with low doses of a simple monofunctional alkylating agent leads to a decrease in both the lethal and mutagenic effects of a subsequent challenge dose of the agent. The evidence for an analagous system in mammalian cells has proved to be inconsistent to date. The induction of chromosome repair mechanisms in human cells by low-dose radiation from tritiated thymidine has been shown to make the cells refractory to the induction of chromosome aberrations by X-rays. The present communication investigates the induction of an adaptive response in human lymphocytes from four donors and V79 cells using SCE and mutation as endpoints and MNNG and MNU for the adapting and challenging treatment. It is clear that a reproducible model of the adaptive response in human lymphocytes is difficult to establish because of the variability between different donors and different culture times. In V79 cells, assays with much larger cell numbers are required to detect a reproducible response with such small changes in mutant frequency. To demonstrate an adaptive response conclusively in mammalian cells will probably require the use of more sensitive experimental protocols and alternative methods of administration of adaptive doses of mutagen.

Random single chromatid and nonrandom double chromatid type segregation of human acrocentric chromosomes in BrdU-labeled mitoses

1984 ◽  
Vol 26 (2) ◽  
pp. 137-140 ◽  
Author(s):  
S. Chemitiganti ◽  
R. S. Verma ◽  
S. Ved Brat ◽  
H. Dosik
Keyword(s):  
Human Leukocytes ◽  
Satellite Association ◽  
Chromatid Segregation ◽  
Random Segregation ◽  
Acrocentric Chromosomes ◽  
Single Chromatid ◽  
Cis And Trans

Chromatid segregation was analyzed using satellite association of 5-bromodeoxyuridine (BrdU) differentially stained acrocentric chromosomes of human leukocytes. Data were classified into cis and trans configurations in second and third division cycles. It was found that single chromatid types have random segregation (1:1) while nonrandom segregation was noted for double chromatid types. The nonrandom segregation hypothesis of earlier investigators needs to be reexamined.Key words: chromatid segregation, BrdU, mitosis, acrocentric.

scholarly journals RENEWAL OF CELLS WITHIN TASTE BUDS

1965 ◽  
Vol 27 (2) ◽  
pp. 263-272 ◽  
Author(s):  
Lloyd M. Beidler ◽  
Ronald L. Smallman
Keyword(s):  
Epithelial Cells ◽  
Life Span ◽  
Tritiated Thymidine ◽  
Mitotic Division ◽  
Considerable Change ◽  
Chemical Stimulation ◽  
Taste Bud ◽  
Average Cell ◽  
Daughter Cells ◽  
Rat Tongue

Colchicine blocks mitotic division of the epithelial cells surrounding the taste bud of the rat tongue. Response to chemical stimulation decreases 50 per cent 3 hours after colchicine injection as measured by recording the electrical activity from the taste nerve bundle. Radioautography, using tritiated thymidine, shows that those epithelial cells surrounding the taste bud divide and that some of the daughter cells enter the taste bud and slowly move toward the center. The life span of the average cell is about 250 ± 50 hours, although some cells have a much shorter and others a much longer life span. These studies suggest that the cells within the taste bud, as well as the nerves, undergo considerable change with time. Corresponding changes in function are considered.

scholarly journals The Abscission Checkpoint: A Guardian of Chromosomal Stability

Cells ◽  
2021 ◽  
Vol 10 (12) ◽  
pp. 3350
Author(s):  
Eleni Petsalaki ◽  
George Zachos
Keyword(s):  
Mammalian Cells ◽  
Kinase Activity ◽  
Nuclear Pore ◽  
Aurora B ◽  
Intercellular Bridge ◽  
Endosomal Sorting ◽  
Daughter Cells ◽  
Chromosomal Passenger ◽  
Dividing Cells ◽  
Chromatin Bridges

The abscission checkpoint contributes to the fidelity of chromosome segregation by delaying completion of cytokinesis (abscission) when there is chromatin lagging in the intercellular bridge between dividing cells. Although additional triggers of an abscission checkpoint-delay have been described, including nuclear pore defects, replication stress or high intercellular bridge tension, this review will focus only on chromatin bridges. In the presence of such abnormal chromosomal tethers in mammalian cells, the abscission checkpoint requires proper localization and optimal kinase activity of the Chromosomal Passenger Complex (CPC)-catalytic subunit Aurora B at the midbody and culminates in the inhibition of Endosomal Sorting Complex Required for Transport-III (ESCRT-III) components at the abscission site to delay the final cut. Furthermore, cells with an active checkpoint stabilize the narrow cytoplasmic canal that connects the two daughter cells until the chromatin bridges are resolved. Unsuccessful resolution of chromatin bridges in checkpoint-deficient cells or in cells with unstable intercellular canals can lead to chromatin bridge breakage or tetraploidization by regression of the cleavage furrow. In turn, these outcomes can lead to accumulation of DNA damage, chromothripsis, generation of hypermutation clusters and chromosomal instability, which are associated with cancer formation or progression. Recently, many important questions regarding the mechanisms of the abscission checkpoint have been investigated, such as how the presence of chromatin bridges is signaled to the CPC, how Aurora B localization and kinase activity is regulated in late midbodies, the signaling pathways by which Aurora B implements the abscission delay, and how the actin cytoskeleton is remodeled to stabilize intercellular canals with DNA bridges. Here, we review recent progress toward understanding the mechanisms of the abscission checkpoint and its role in guarding genome integrity at the chromosome level, and consider its potential implications for cancer therapy.

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