scholarly journals Characterization and immunocytochemical distribution of calmodulin in higher plant endosperm cells: localization in the mitotic apparatus.

1985 ◽  
Vol 101 (2) ◽  
pp. 488-499 ◽  
Author(s):  
M Vantard ◽  
A M Lambert ◽  
J De Mey ◽  
P Picquot ◽  
L J Van Eldik
Keyword(s):  
Animal Cell ◽  
Regulatory Mechanisms ◽  
Present Observation ◽  
Chromosome Movement ◽  
Animal Cells ◽  
Higher Plant ◽  
Mitotic Apparatus ◽  
Immunogold Staining ◽  
Staining Methods ◽  
Spindle Development

In this study we have examined the immunocytochemical distribution of calmodulin during mitosis of higher plant endosperm cells. Spindle development in these cells occurs without centrioles. Instead, asterlike microtubule converging centers appear to be involved in establishing spindle polarity. By indirect immunofluorescence and immunogold staining methods with anti-calmodulin antibodies, we found endosperm calmodulin to be associated with the mitotic apparatus, particularly with asterlike and/or polar microtubule converging centers and kinetochore microtubules, in an immunocytochemical pattern distinct from that of tubulin. In addition, endosperm calmodulin and calcium showed analogous distribution profiles during mitosis. Previous reports have demonstrated that calmodulin is associated with the mitotic apparatus in animal cells. The present observation that calmodulin is also associated with the mitotic apparatus in acentriolar, higher plant endosperm cells suggests that some of the regulatory mechanisms involved in spindle formation, microtubule disassembly, and chromosome movement in plant cells may be similar to those in animal cells. However, unlike animal cell calmodulin, endosperm calmodulin appears to associate with kinetochore microtubules throughout mitosis, which suggests a specialized role for higher plant calmodulin in the regulation of kinetochore microtubule dynamics.

scholarly journals Chromosome Movement in Mitosis Requires Microtubule Anchorage at Spindle Poles

2001 ◽  
Vol 152 (3) ◽  
pp. 425-434 ◽  
Author(s):  
Michael B. Gordon ◽  
Louisa Howard ◽  
Duane A. Compton
Keyword(s):  
Electron Microscopy ◽  
Spindle Pole ◽  
Chromosome Movement ◽  
Human Homologue ◽  
Animal Cells ◽  
Mitotic Apparatus ◽  
Spindle Poles ◽  
Microtubule Attachment ◽  
Anaphase Onset ◽  
Immunofluorescence And Electron Microscopy

Anchorage of microtubule minus ends at spindle poles has been proposed to bear the load of poleward forces exerted by kinetochore-associated motors so that chromosomes move toward the poles rather than the poles toward the chromosomes. To test this hypothesis, we monitored chromosome movement during mitosis after perturbation of nuclear mitotic apparatus protein (NuMA) and the human homologue of the KIN C motor family (HSET), two noncentrosomal proteins involved in spindle pole organization in animal cells. Perturbation of NuMA alone disrupts spindle pole organization and delays anaphase onset, but does not alter the velocity of oscillatory chromosome movement in prometaphase. Perturbation of HSET alone increases the duration of prometaphase, but does not alter the velocity of chromosome movement in prometaphase or anaphase. In contrast, simultaneous perturbation of both HSET and NuMA severely suppresses directed chromosome movement in prometaphase. Chromosomes coalesce near the center of these cells on bi-oriented spindles that lack organized poles. Immunofluorescence and electron microscopy verify microtubule attachment to sister kinetochores, but this attachment fails to generate proper tension across sister kinetochores. These results demonstrate that anchorage of microtubule minus ends at spindle poles mediated by overlapping mechanisms involving both NuMA and HSET is essential for chromosome movement during mitosis.

scholarly journals A permeabilized cell model for studying cell division: a comparison of anaphase chromosome movement and cleavage furrow constriction in lysed PtK1 cells.

1981 ◽  
Vol 88 (3) ◽  
pp. 618-629 ◽  
Author(s):  
W Z Cande ◽  
K McDonald ◽  
R L Meeusen
Keyword(s):  
Cell Model ◽  
Chromosome Movement ◽  
Microtubule Depolymerization ◽  
Mitotic Apparatus ◽  
Permeabilized Cell ◽  
Culture Cells ◽  
Brij 58 ◽  
Myosin Subfragment ◽  
Myosin Subfragment 1 ◽  
Chromosome Movements

After lysis in a Brij 58-polyethylene glycol medium, PtK1 cells are permeable to small molecules, such as erythrosin B, and to proteins, such as rhodamine-labeled FAB, myosin subfragment-1, and tubulin. Holes are present in the plasma membrane, and the mitochondria are swollen and distorted, but other membrane-bounded organelles of the lysed cell model are not noticeably altered. After lysis, the mitotic apparatus is functional; chromosomes move poleward and the spindle elongates. Cells lysed while in cytokinesis will continue to divide for several minutes. Addition of crude tubulin extracts, MAP-free tubulin, or taxol to the lysis medium retards anaphase chromosome movements but does not affect cleavage. On the other hand, N-ethylmaleimide-modified myosin subfragment-1, phalloidin, and cytochalasin B inhibit cleavage but have no effect on anaphase chromosome movements under identical lysis conditions. These results suggest that actomyosin plays no functional role in anaphase chromosome movement in mammalian tissue culture cells and that microtubule depolymerization is a rate-limiting step for chromosome-to-pole movements.

Regulation of Mitosis

1969 ◽  
Vol 5 (3) ◽  
pp. 745-755
Author(s):  
W. T. JACKSON
Keyword(s):  
Mitotic Spindle ◽  
Time Lapse ◽  
Polarization Microscopy ◽  
Animal Cells ◽  
Higher Plant ◽  
Competitive Antagonism ◽  
Dividing Cells ◽  
Spindle Microtubules ◽  
Giant Nuclei

Earlier studies on the effects of the herbicide isopropyl N-phenylcarbamate (IPC) on mitosis revealed blocked metaphases, multinucleate cells, giant nuclei and an increase in number of partly contracted chromosomes. It was assumed that IPC, like colchicine, was causing these effects by disruption of the spindle apparatus by destroying the spindle microtubules. The animal hormone melatonin causes an increase in birefringence of the mitotic spindle in animal cells, presumably by increasing the number of microtubules. We have studied the effects of IPC, melatonin, and combinations of the two on mitosis in dividing endosperm cells of the African blood lily (Haemanthus katherinae Baker) in vivo by phase-contrast and polarization microscopy. Both qualitative and quantitative data are presented. Interpretation of these results has been aided materially by a time-lapse cinemicrographic analysis of dividing cells subjected to 1 and 10 p.p.m. IPC (unpublished) and by an accompanying fine-structural analysis of untreated and IPC-treated cells. Mitosis was disrupted by 0.01-10 p.p.m. IPC, the severity of the effect depending on both concentration and stage of mitosis of the cell at the time of treatment. Concentrations of IPC that caused cessation of chromosome movement also caused loss of birefringence of the mitotic spindle. Melatonin increased birefringence of the mitotic spindle in these plant cells and partly nullified the adverse effects of IPC. The results of this study demonstrate that the herbicide IPC, under our conditions, causes disruption of mitosis and loss of birefringence of the spindle. And it has been established that an animal hormone is capable of increasing the birefringence, and presumably the number of microtubules, of the mitotic spindle in dividing endosperm cells of a higher plant. Although melatonin is capable of partly nullifying the effects of IPC, a competitive antagonism is not postulated.

scholarly journals Microtubule dynamics and chromosome motion visualized in living anaphase cells.

1988 ◽  
Vol 106 (4) ◽  
pp. 1185-1192 ◽  
Author(s):  
G J Gorbsky ◽  
P J Sammak ◽  
G G Borisy
Keyword(s):  
Chromosome Segregation ◽  
Digital Imaging ◽  
Living Cells ◽  
Microtubule Dynamics ◽  
Chromosome Movement ◽  
Animal Cells ◽  
Digital Imaging Microscopy ◽  
Opposite Pole ◽  
Anaphase B ◽  
Chromosome Motion

Chromosome segregation in most animal cells is brought about through two events: the movement of the chromosomes to the poles (anaphase A) and the movement of the poles away from each other (anaphase B). Essential to an understanding of the mechanism of mitosis is information on the relative movements of components of the spindle and identification of sites of subunit loss from shortening microtubules. Through use of tubulin derivatized with X-rhodamine, photobleaching, and digital imaging microscopy of living cells, we directly determined the relative movements of poles, chromosomes, and a marked domain on kinetochore fibers during anaphase. During chromosome movement and pole-pole separation, the marked domain did not move significantly with respect to the near pole. Therefore, the kinetochore microtubules were shortened by the loss of subunits at the kinetochore, although a small amount of subunit loss elsewhere was not excluded. In anaphase A, chromosomes moved on kinetochore microtubules that remained stationary with respect to the near pole. In anaphase B, the kinetochore fiber microtubules accompanied the near pole in its movement away from the opposite pole. These results eliminate models of anaphase in which microtubules are thought to be traction elements that are drawn to and depolymerized at the pole. Our results are compatible with models of anaphase in which the kinetochore fiber microtubules remain anchored at the pole and in which microtubule dynamics are centered at the kinetochore.

scholarly journals Synthesis of DEAE-Soybean Starch Microspheres for Adhere Animal Cell Culture

2017 ◽  
Vol 9 (8) ◽  
pp. 91
Author(s):  
Lei Zhang ◽  
Mingsheng Li ◽  
Zhongren Ma ◽  
Yuping Feng
Keyword(s):  
Cell Culture ◽  
Mdck Cells ◽  
Animal Cell ◽  
Low Cost ◽  
Animal Cell Culture ◽  
Cross Linking ◽  
Animal Cells ◽  
Inoculation Density ◽  
Animal Cell Cultures ◽  
Adhere Cell

The present study outlines the synthesis of a new microcarrier for anchorage-dependent animal cell cultures. The new microcarriers were synthesized from the cross-linking soybean starch microspheres followed by modification with 2-diethylaminoethyl (DEAE). Furthermore, 5 g/100 mL of wet microspheres DEAE-soybean starch microspheres were applied in the adhere cell culture, with an inoculation density 2.0 × 105 cells/mL of BHK-21, Marc-145, and MDCK cells. The cells were shown to grow well in the DEAE-soybean starch microcarrier, with BHK-21 cells showing a higher cell density after 144 h (2.5 × 106 cells/mL) compared to cells grown on the commercial product Cytodex 1 (2.2 × 106 cells/mL). These starch microcarriers have a potential application in anchorage-dependent animal cells culture, due to its low cost and its simple process.

The regulation of mitotic spindle function

1988 ◽  
Vol 66 (6) ◽  
pp. 490-514 ◽  
Author(s):  
Stephen M. Wolniak
Keyword(s):  
Morphological Changes ◽  
Cytosolic Calcium ◽  
Spindle Pole ◽  
Chromosome Movement ◽  
Mitotic Apparatus ◽  
Physiological Regulation ◽  
New Findings ◽  
Controlling Elements ◽  
Spindle Elongation ◽  
Spindle Function

The process of mitosis includes a series of morphological changes in the cell in which the directional movements of chromosomes are the most prominent. The presence of a microtubular array, known as the spindle or mitotic apparatus, provides at least a scaffold upon which these movements take place. The precise mechanism for chromosome movement remains obscure, but new findings suggest that the kinetochore may play a key role in chromosome movement toward the spindle pole, and that sliding interactions between or among adjacent microtubules may provide the mechanochemical basis for spindle elongation. The physiological regulation of the anaphase motors and of spindle operation either before or after anaphase remains equally elusive. Elicitors that may serve as controlling elements in spindle function include shifts in cytosolic calcium activity and perhaps the activation or inactivation of protein kinases, which in turn produce changes in the state of phosphorylation of specific spindle components.

scholarly journals Molecular structure of phospholipase D and regulatory mechanisms of its activity in plant and animal cells

2012 ◽  
Vol 77 (1) ◽  
pp. 1-14 ◽  
Author(s):  
Y. S. Kolesnikov ◽  
K. P. Nokhrina ◽  
S. V. Kretynin ◽  
I. D. Volotovski ◽  
J. Martinec ◽  
...  
Keyword(s):  
Molecular Structure ◽  
Phospholipase D ◽  
Regulatory Mechanisms ◽  
Animal Cells

Demethylation of genes in animal cells

1990 ◽  
Vol 326 (1235) ◽  
pp. 241-251 ◽  
Keyword(s):  
Tissue Culture ◽  
Germ Line ◽  
Animal Cell ◽  
Gene Activation ◽  
Housekeeping Gene ◽  
Cell Types ◽  
Embryonic Cells ◽  
Animal Cells ◽  
Developmentally Regulated ◽  
I Gene

Tissue-specific animal cell genes are usually fully methylated in the germ line and become demethylated in those cell types in which they are expressed. To investigate this process, we inserted a methylated IgG K gene into fibroblasts and lymphocytes at various stages of development. The results show that this gene undergoes demethylation only in the mature lymphocytes and therefore suggest that the ability to demethylate a gene is developmentally regulated. These studies were supported by similar experiments using the rat Insulin I gene, and in this case it appears that the cis -acting elements that control demethylation may be different from those responsible for gene activation. The ability to demethylate the housekeeping gene APRT is also under developmental control, because this occurs only in embryonic cells, both in tissue culture and in transgenic mice.

scholarly journals Conservation of the pentanucleotide motif at the top of the yellow fever virus 17D 3′ stem–loop structure is not required for replication

2007 ◽  
Vol 88 (6) ◽  
pp. 1738-1747 ◽  
Author(s):  
Patrícia A. G. C. Silva ◽  
Richard Molenkamp ◽  
Tim J. Dalebout ◽  
Nathalie Charlier ◽  
Johan H. Neyts ◽  
...  
Keyword(s):  
Yellow Fever ◽  
Molecular Mechanisms ◽  
Yellow Fever Virus ◽  
Animal Cell ◽  
Fever Virus ◽  
Loop Structure ◽  
Animal Cells ◽  
Pn Sequence ◽  
Stem Loop ◽  
Stem Loop Structure

The pentanucleotide (PN) sequence 5′-CACAG-3′ at the top of the 3′ stem–loop structure of the flavivirus genome is well conserved in the arthropod-borne viruses but is more variable in flaviviruses with no known vector. In this study, the sequence requirements of the PN motif for yellow fever virus 17D (YFV) replication were determined. In general, individual mutations at either the second, third or fourth positions were tolerated and resulted in replication-competent virus. Mutations at the fifth position were lethal. Base pairing of the nucleotide at the first position of the PN motif and a nucleotide four positions downstream of the PN (ninth position) was a major determinant for replication. Despite the fact that the majority of the PN mutants were able to replicate efficiently, they were outcompeted by parental YFV-17D virus following repeated passages in double-infected cell cultures. Surprisingly, some of the virus mutants at the first and/or the ninth position that maintained the possibility of forming a base pair were found to have a similar fitness to YFV-17D under these conditions. Overall, these experiments suggest that YFV is less dependent on sequence conservation of the PN motif for replication in animal cells than West Nile virus. However, in animal cell culture, YFV has a preference for the wt CACAG PN sequence. The molecular mechanisms behind this preference remain to be elucidated.

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