Mitosis, arrest

As a whole, the results presented above indicate that low concentrations of Vinca alkaloids, probably similar to therapeutic concentrations, have an antiproliferative activity that is due to inhibition of mitotic spindle function by changing the dynamics of microtubules rather than by depolymerizing them. A growing body of evidence seems to indicate that, specially in cancer cells, where mitosis regulation is already disrupted, the suppression of microtubule dynamics with mitosis arrest induces a signaling cascade leading to cell death by apoptosis (a type of programmed cell death) [172-174]. 

DCPA inhibits the growth of grass species by dismpting the mitotic sequence, probably at entry (190). DCPA influences spindle formation and function (181) and causes root-tip swelling (182) and britde shoot tissue (191). It has been reported that DCPA, like colchicine and vinblastine, attests mitosis at prometaphase and is associated with formation of polymorphic nuclei after mitotic arrest (192). Pronamide also inhibits root growth by dismpting the mitotic sequence in a manner similar to the effect of colchicine and the dinitroanilines (193,194). Cinmethylin and bensuhde prevent mitotic entry by unknown mechanisms (194). 

Vinca alkaloids are derived from the Madagascar periwinkle plant, Catharanthus roseus. The main alkaloids are vincristine, vinblastine and vindesine. Vinca alkaloids are cell-cycle-specific agents and block cells in mitosis. This cellular activity is due to their ability to bind specifically to tubulin and to block the ability of the protein to polymerize into microtubules. This prevents spindle formation in mitosing cells and causes arrest at metaphase. Vinca alkaloids also inhibit other cellular activities that involve microtubules, such as leukocyte phagocytosis and chemotaxis as well as axonal transport in neurons. Side effects of the vinca alkaloids such as their neurotoxicity may be due to disruption of these functions. 

Another drug is taxol, which is extracted from the bark of the Pacific yew tree, Taxus brevijolia. Unlike colchicine and the vinca alkaloids, taxol binds tightly to microtubules and stabilizes them against depolymerization by Ca. It also enhances the rate and yield of microtubule assembly, thereby decreasing the amount of soluble tubulin in the cytosol pool. Again, the overall effect of taxol is to arrest dividing cells in mitosis. Taxol is used in cancer chemotherapy. 

Despite the complexity of the experiments and the enormous data manipulation necessary, complex biological pathways, as well as new drug targets are being identified by this method. Examples include screens for compounds that arrest cells in mitosis, that block cell migration, and that block the secretory pathway [50], or assays with primary T cells from PLP TCR transgenic mice for their inhibitory activity on the proliferation and secretion of proinflammatory cytokines in PLP-reactive T cells [51], and identification of small-molecule inhibitors of histone acetyltransferase activity [52]. 

Eehner We haven t done these experiments. But we have looked in string mutants where cells arrest in G2 before mitosis 14 and in cyclinAl cyclinB double mutants where cells arrest in G2 before mitosis 15. The timing of dacapo expression in the epidermis appears to be normal in these mutants. 

The Xenopus system has proven instrumental in determining the mechanism controlling exit from mitosis at the metaphase/anaphase transition. Studies in this area have relied heavily on extracts prepared from fully mature oocytes/ unfertilized eggs that are arrested at metaphase of the second meiotic division. Upon Ca2+ addition, anaphase is initiated and the extract enters the first embryonic cell cycle to replicate DNA. The activity responsible for metaphase arrest was discovered by Masui at the same time as MPF (Masui Markert 1971), and given the name cytostatic factor (CSF). CSF has never been purified... 

Parthenogenetic one-cell embryos were fused very soon after an activating treatment with parthenogenetic one-cell embryos entering first mitosis. We expected that hybrids obtained in such experiments would either arrest in M-... 

Kalab P, Kubiak JZ, Verlhac MH, Colledge WH, Maro B 1996 Activation of p90Kk during meiotic maturation and first mitosis in mouse oocytes and eggs MAP kinase-independent and dependent activation. Development 122 1957—1964 Kubiak JZ, Weber M, dePennart H, Winston N, Maro B 1993 The metaphase II arrest in mouse oocytes is controlled through microtubule-dependent destruction of cyclin B in the presence of CSF. EMBO J 12 3773-3778... 

Despite close scrutiny of infected cell nuclei, there is no reported evidence of passage into mitosis. Isolated infected cell nuclei had an approximate 4N content of DNA (Jasmer, 1993). Therefore, T. spiralis infection evidently induces terminally differentiated skeletal muscle cells to re-enter the cell cycle and chronically arrest in G2/M. 

Observations are made in metaphase cells arrested with a spindle inhibitor such as colchicine or colcemid to accumulate cells in a metaphase-like stage of mitosis (c-metaphase) before hypotonic treatment to enlarge cells and fixation with alcohol-acetic acid solution. Cells are then dispersed on to microscope slides and stained and slides are randomized, coded and analyzed for chromosome aberrations with high-power light microscopy. Details of the procedure are given in Dean and Danford (1984) and Preston et al. (1981, 1987). The UKEMS guidelines (Scott et al., 1990) recommend that all tests be repeated regardless of the outcome of the first test and... 

The protein content is measured by biochemical methods, such as methods of Lowry or Bradford [14,18,19,21,23], now usually by commercially available kits. The DNA content can be measured by flow cytometry, and the chromosomes can be counted directly in cells after their arrest in the metaphase of the mitosis by Colcemide [21]. 

Quinn, C.M., Kagedal, K., Terman, A., et al. (2004) Induction of fibroblast apohpoprotein E expression during apoptosis, starvation-induced growth arrest and mitosis. Biochem. J., 378, 753-761. 

Stoler, S., Keith, K.C., Curnick, K.E., and Fitzgerald-Hayes, M. (1995) A mutation in CSE4, an essential gene encoding a novel chromatin-associated protein in yeast, causes chromosome nondisjunction and cell cycle arrest at mitosis. Gene Dev. 9, 573-586. 

Kanoh K, Kohno S, Katada J, Takahashi J, Uno 1. (1999) (-)-Phenylahistin arrests cells in mitosis by inhibiting tnbnlin polymerization. J Antibiot 52 134-141. 

Dividing cells in culture exposed to vinblastine or vincristine are arrested from further growth during mitosis (12,13). In fact, the antimitotic effects of this class of compounds is ubiquitous. These effects are observed at relatively low concentrations (<1 iM), and are reversible when drug is removed from the media prior to lysis of the arrested cells. The concentration of drug required to elicit an antimitotic effect is usually comparable to that required to produce a cytotoxic effect in the same cell type (14,15). Originally, this type of analysis was exceedingly laborious, but the introduction of laser- and computer-based fluoresence activated cell sorters (FACS) has rendered this type of analysis routine. Nevertheless, a cytotoxic, non-cell cycle-specific bisindole alkaloid has yet to be discovered. 

Vincristine and vinblastine are generally considered to act specifically on the metaphase portion of the mitotic (M) stage of the cell cycle as a consequence of perturbations of the structure and function of tubulin. A characteristic action of the drugs is production of mitotic arrest in which the tJercentage of cells in mitosis in a given population of cells will rise from a few percent to 50% and more after treatment with a drug such as vinblastine. There are reports, however, that these drugs can interfere with other phases of the cell cycle in ways not clearly related to interference with tubulin function (5). 

The contractile proteins of the spindle apparatus must draw apart the replicated chromosomes before the cell can divide. This process is prevented by the so-called spindle poisons (see also colchicine, p. 316) that arrest mitosis at metaphase by disrupting the assembly of microtubules into spindle threads. The vinca alkaloids, vincristine and vinblastine (from the periwinkle plant. Vinca rosea) exert such a cell-cycle-specific effect. Damage to the nervous system is a predicted adverse effect arising from injury to microtubule-operated axonal transport mechanisms. 

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