WorldWideScience

Sample records for cdk7 localization mitotic

  1. Analysis list: Cdk7 [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Cdk7 Pluripotent stem cell + mm9 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/target/Cd...k7.1.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/target/Cdk7.5.tsv http://dbarchive.bioscienced...bc.jp/kyushu-u/mm9/target/Cdk7.10.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/colo/Cdk7.Pluripotent_s

  2. Localization of topoisomerase II in mitotic chromosomes

    OpenAIRE

    1985-01-01

    In the preceding article we described a polyclonal antibody that recognizes cSc-1, a major polypeptide component of the chicken mitotic chromosome scaffold. This polypeptide was shown to be chicken topoisomerase II. In the experiments described in the present article we use indirect immunofluorescence and immunoelectron microscopy to examine the distribution of topoisomerase II within intact chromosomes. We also describe a simple experimental protocol that differentiates antigens that are int...

  3. Identification of a novel mitotic phosphorylation motif associated with protein localization to the mitotic apparatus

    Energy Technology Data Exchange (ETDEWEB)

    Yang, Feng; Camp, David G.; Gritsenko, Marina A.; Luo, Quanzhou; Kelly, Ryan T.; Clauss, Therese RW; Brinkley, William R.; Smith, Richard D.; Stenoien, David L.

    2007-11-16

    The chromosomal passenger complex (CPC) is a critical regulator of chromosome, cytoskeleton and membrane dynamics during mitosis. Here, we identified phosphopeptides and phosphoprotein complexes recognized by a phosphorylation specific antibody that labels the CPC using liquid chromatography coupled to mass spectrometry. A mitotic phosphorylation motif (PX{G/T/S}{L/M}[pS]P or WGL[pS]P) was identified in 11 proteins including Fzr/Cdh1 and RIC-8, two proteins with potential links to the CPC. Phosphoprotein complexes contained known CPC components INCENP, Aurora-B and TD-60, as well as SMAD2, 14-3-3 proteins, PP2A, and Cdk1, a likely kinase for this motif. Protein sequence analysis identified phosphorylation motifs in additional proteins including SMAD2, Plk3 and INCENP. Mitotic SMAD2 and Plk3 phosphorylation was confirmed using phosphorylation specific antibodies, and in the case of Plk3, phosphorylation correlates with its localization to the mitotic apparatus. A mutagenesis approach was used to show INCENP phosphorylation is required for midbody localization. These results provide evidence for a shared phosphorylation event that regulates localization of critical proteins during mitosis.

  4. Localization of latency-associated nuclear antigen (LANA) on mitotic chromosomes.

    Science.gov (United States)

    Rahayu, Retno; Ohsaki, Eriko; Omori, Hiroko; Ueda, Keiji

    2016-09-01

    In latent infection of Kaposi's sarcoma-associated herpesvirus (KSHV), viral gene expression is extremely limited and copy numbers of viral genomes remain constant. Latency-associated nuclear antigen (LANA) is known to have a role in maintaining viral genome copy numbers in growing cells. Several studies have shown that LANA is localized in particular regions on mitotic chromosomes, such as centromeres/pericentromeres. We independently examined the distinct localization of LANA on mitotic chromosomes during mitosis, using super-resolution laser confocal microscopy and correlative fluorescence microscopy-electron microscopy (FM-EM) analyses. We found that the majority of LANA were not localized at particular regions such as telomeres/peritelomeres, centromeres/pericentromeres, and cohesion sites, but at the bodies of condensed chromosomes. Thus, LANA may undergo various interactions with the host factors on the condensed chromosomes in order to tether the viral genome to mitotic chromosomes and realize faithful viral genome segregation during cell division. PMID:27254595

  5. Upregulation of CDK7 in gastric cancer cell promotes tumor cell proliferation and predicts poor prognosis.

    Science.gov (United States)

    Wang, Qiuhong; Li, Manhua; Zhang, Xunlei; Huang, Hua; Huang, Jianfei; Ke, Jing; Ding, Haifang; Xiao, Jinzhang; Shan, Xiaohang; Liu, Qingqing; Bao, Bojun; Yang, Lei

    2016-06-01

    CDK7 has been known as a component of CDK activating kinase (CAK) complex, the complex was composed of CDK7, Cyclin H and RING finger protein Mat1 that contribute to cell cycle progression by phosphorylating other CDKs. In addition, the complex is also an essential component of general transcription factor TFIIH which controls transcription via activating RNA polymerase II by serines 5 and 7 phosphorylation of the carboxyl-terminal domain (CTD) of its largest subunit. However, the role of CDK7 in the pathogenesis of gastric cancer has not been identified. Our study showed that CDK7 was significantly upregulated and positively correlated with tumor grade, infiltration depth, lymph node, Ki-67, and predicted poor prognosis in 173 gastric cancer specimens by immunohistochemistrical analyses. Furthermore, in vitro results indicated that CDK7 promoted proliferation of gastric cancer cells by CCK8, clone formation analyses and flow cytometric analyses, while CDK7 knockdown led to decreased cell proliferation. Our study will provide a theoretical basis for the study of CDK7 in gastric cancer. PMID:27155449

  6. Targeting Transcriptional Addictions In Small Cell Lung Cancer With a Covalent CDK7 Inhibitor

    OpenAIRE

    Christensen, Camilla L.; Kwiatkowski, Nicholas; Abraham, Brian J; Carretero, Julian; Al-Shahrour, Fatima; Zhang, Tinghu; Chipumuro, Edmond; Herter-Sprie, Grit S.; Akbay, Esra A; Altabef, Abigail; Zhang, Jianming; Shimamura, Takeshi; Capelletti, Marzia; Reibel, Jakob B.; Cavanaugh, Jillian

    2014-01-01

    Small cell lung cancer (SCLC) is an aggressive disease with high mortality. The identification of effective pharmacological strategies to target SCLC biology represents an urgent need. Using a high-throughput cellular screen of a diverse chemical library we observe that SCLC is sensitive to transcription-targeting drugs, and in particular to THZ1, a recent identified covalent inhibitor of cyclin-dependent kinase 7 (CDK7). We find that expression of super-enhancer associated transcription fact...

  7. Human Zwint-1 Specifies Localization of Zeste White 10 to Kinetochores and Is Essential for Mitotic Checkpoint Signaling

    Institute of Scientific and Technical Information of China (English)

    HongmeiWang; XiaoyuHu; XiaDing; ZhenDou; ZhihongYank; AndrewW.Shaw; MaikunTcng; DonW.Cleveland; MichaelL.Goldberg; LiwenNiu; XucbiaoYao

    2005-01-01

    Chromosome segregation in mitosis is orchestrated by dynamic interaction between spindle microtubules and the kinetochore, a multiprotein complex assembled onto centromeric DNA of the chromosome. Here we show that Zwint-1 is required and is sufficient for kinetochore localization of Zestc White 10 (ZW10) in HeLa cells. Zwint-1 specifies the kinetochore association of ZW10 by interacting with its N-terminal domain. Suppression of synthesis of Zwint-1 by small interfering RNA abolishes the localization of ZW10 to the kinetochore, demonstrating the requirement of Zwint-1 for ZWl0 kinetochore localization. In addition, dcplction of Zwint-1 affects no mitotic arrest but causes aberrant premature chromo. some segregation. These Zwint-l-suppressed cells dis. play chromosome bridge phenotype with sister chromatids inter-connected. Moreover, Zwint-1 is required for stable association of CENP.F and dynamitin but not BUB1 with the kinetochore. Finally, our studies showthat Zwint-1 is a new component of the mitotic check. point, as cells lacking Zwint-1 fail to arrest in mitosis when exposed to microtubule inhibitors, yielding inter. phase cells with multinuclei. As ZWl0 and Zwint.1 are absent from yeast, we reasoned that metazoans evolved an elaborate spindle checkpoint machinery to ensure faithful chromosome segregation in mitosis.

  8. Genetic inactivation of Cdk7 leads to cell cycle arrest and induces premature aging due to adult stem cell exhaustion

    OpenAIRE

    Ganuza, Miguel; Sáiz-Ladera, Cristina; Cañamero, Marta; Gómez, Gonzalo; Schneider, Ralph; Blasco, María A.; Pisano, David; Paramio, Jesús M.; Santamaría, David; Barbacid, Mariano

    2012-01-01

    Employing a conditionally inactive gene trap allele, Cdk7's function in regulating cellular proliferation by Cdk1/2-phosphorylation is convincingly dissected from alternative notions on CTD-phosphorylation of RNA Pol II. Premature aging phenotypes caused by stem cell depletion lend the necessary functional support.

  9. Isolated late local recurrences with high mitotic count and early local recurrences following breast-conserving therapy are associated with increased risk on distant metastasis

    International Nuclear Information System (INIS)

    Purpose: Local recurrence (LR) after breast-conserving therapy (BCT) is associated with an increased risk for the development of distant metastasis. We studied risk factors for distant metastasis risk (DMR) and poor prognosis within a group of patients with LR as first event. Patients and Methods: From a cohort of 1481 breast carcinomas treated with BCT in the period 1980-1994, a total of 68 pT1-3 N0-1 patients developed LR as first event. We have studied risk factors for the development of distant metastasis within this group of patients with LR. In addition to clinical factors (age at BCT and LR, mode of detection, location of LR, and treatment of LR), the histology slides of the primary and the recurrent tumor were reviewed. Immunohistochemical staining was performed for the following proteins: bcl-2, cyclin D1, E-cadherin, EGF receptor, ER, PR, Ki-67, c-erbB-2/neu, and p53. Statistical analyses were performed using conditional logistic regression. Results: At a median follow-up after LR of 5.6 years, the 5-year DMR was 53%. In univariate analysis, none of the factors of the primary tumor was found to be associated with DMR after LR. Of the recurrent tumor the following factors were found to be risk factors for high DMR after LR: interval between treatment of the primary tumor and LR at 2 years or less (relative risk, 2.38; 95% confidence interval, 1.22-4.76; p=0.008) and high mitotic count (relative risk, 2.51; 95% confidence interval, 1.03-6.15; p=0.04). All patients with noninvasive recurrent tumor were alive at the time of analysis. Patients with an interval of greater than 2 years and a recurrent tumor with high mitotic count were found to have an equally poor prognosis compared to patients with LRs detected after a short interval. Conclusion: LR after BCT is associated with higher DMR and poor prognosis. Patients with LR within 2 years after BCT are especially at high risk. Late recurrences with high mitotic count have the same poor prognosis as early

  10. The SUMO-targeted ubiquitin ligase RNF4 localizes to etoposide-exposed mitotic chromosomes: implication for a novel DNA damage response during mitosis.

    Science.gov (United States)

    Saito, Masayuki; Fujimitsu, Yuka; Sasano, Takeshi; Yoshikai, Yushi; Ban-Ishihara, Reiko; Nariai, Yuko; Urano, Takeshi; Saitoh, Hisato

    2014-04-25

    RNF4, a SUMO-targeted ubiquitin ligase (STUbL), localizes to the nucleus and functions in the DNA damage response during interphase of the cell cycle. RNF4 also exists in cells undergoing mitosis, where its regulation and function remain poorly understood. Here we showed that administration of etoposide, an anticancer DNA topoisomerase II poison, to mitotic human cervical cancer HeLa cells induced SUMO-2/3-dependent localization of RNF4 to chromosomes. The FK2 antibody signals, indicative of poly/multi-ubiquitin assembly, were detected on etoposide-exposed mitotic chromosomes, whereas the signals were negligible in cells depleted for RNF4 by RNA interference. This suggests that RNF4 functions as a STUbL in the etoposide-induced damage response during mitosis. Indeed, RNF4-depletion sensitized mitotic HeLa cells to etoposide and increased cells with micronuclei. These results indicate the importance of the RNF4-mediated STUbL pathway during mitosis for the maintenance of chromosome integrity and further implicate RNF4 as a target for topo II poison-based therapy for cancer patients. PMID:24695317

  11. Phosphorylation by PKA potentiates retinoic acid receptor α activity by means of increasing interaction with and phosphorylation by cyclin H/cdk7

    OpenAIRE

    Gaillard, Emilie; Bruck, Nathalie; Brelivet, Yann; Bour, Gaétan; Lalevée, Sébastien; Bauer, Annie; Poch, Olivier; Moras, Dino; Rochette-Egly, Cécile

    2006-01-01

    Nuclear retinoic acid receptors (RARs) work as ligand-dependent heterodimeric RAR/retinoid X receptor transcription activators, which are targets for phosphorylations. The N-terminal activation function (AF)-1 domain of RARα is phosphorylated by the cyclin-dependent kinase (cdk) 7/cyclin H complex of the general transcription factor TFIIH and the C-terminal AF-2 domain by the cAMP-dependent protein kinase A (PKA). Here, we report the identification of a molecular pathway by which phosphorylat...

  12. Triptolide Induces Cell Killing in Multidrug-Resistant Tumor Cells via CDK7/RPB1 Rather than XPB or p44.

    Science.gov (United States)

    Yi, Jun-Mei; Huan, Xia-Juan; Song, Shan-Shan; Zhou, Hu; Wang, Ying-Qing; Miao, Ze-Hong

    2016-07-01

    Multidrug resistance (MDR) is a major cause of tumor treatment failure; therefore, drugs that can avoid this outcome are urgently needed. We studied triptolide, which directly kills MDR tumor cells with a high potency and a broad spectrum of cell death. Triptolide did not inhibit P-glycoprotein (P-gp) drug efflux and reduced P-gp and MDR1 mRNA resulting from transcription inhibition. Transcription factors including c-MYC, SOX-2, OCT-4, and NANOG were not correlated with triptolide-induced cell killing, but RPB1, the largest subunit of RNA polymerase II, was critical in mediating triptolide's inhibition of MDR cells. Triptolide elicited antitumor and anti-MDR activity through a universal mechanism: by activating CDK7 by phosphorylating Thr170 in both parental and MDR cell lines and in SK-OV-3 cells. The CDK7-selective inhibitor BS-181 partially rescued cell killing induced by 72-hour treatment of triptolide, which may be due to partial rescue of RPB1 degradation. We suggest that a precise phosphorylation site on RPB1 (Ser1878) was phosphorylated by CDK7 in response to triptolide. In addition, XPB and p44, two transcription factor TFIIH subunits, did not contribute to triptolide-driven RPB1 degradation and cell killing, although XPB was reported to covalently bind to triptolide. Several clinical trials are underway to test triptolide and its analogues for treating cancer and other diseases, so our data may help expand potential clinical uses of triptolide, as well as offer a compound that overcomes tumor MDR. Future investigations into the primary molecular target(s) of triptolide responsible for RPB1 degradation may suggest novel anti-MDR target(s) for therapeutic development. Mol Cancer Ther; 15(7); 1495-503. ©2016 AACR. PMID:27197304

  13. Bacterial mitotic machineries

    DEFF Research Database (Denmark)

    Gerdes, Kenn; Møller-Jensen, Jakob; Ebersbach, Gitte;

    2004-01-01

    Here, we review recent progress that yields fundamental new insight into the molecular mechanisms behind plasmid and chromosome segregation in prokaryotic cells. In particular, we describe how prokaryotic actin homologs form mitotic machineries that segregate DNA before cell division. Thus, the P...

  14. Mitotic spindle perturbations

    NARCIS (Netherlands)

    Tame, M.A.

    2016-01-01

    Microtubules are major components of the cytoskeleton and form the bipolar spindle apparatus during mitosis. The mitotic spindle consists of highly dynamic microtubule polymers that are under constant modulation, controlled by multiple motor proteins and microtubule-associated proteins. This tight s

  15. New mitotic regulators released from chromatin

    Directory of Open Access Journals (Sweden)

    Hideki eYokoyama

    2013-12-01

    Full Text Available Faithful action of the mitotic spindle segregates duplicated chromosomes into daughter cells. Perturbations of this process result in chromosome mis-segregation, leading to chromosomal instability and cancer development. Chromosomes are not simply passengers segregated by spindle microtubules but rather play a major active role in spindle assembly. The GTP bound form of the Ran GTPase (RanGTP, produced around chromosomes, locally activates spindle assembly factors. Recent studies have uncovered that chromosomes organize mitosis beyond spindle formation. They distinctly regulate other mitotic events, such as spindle maintenance in anaphase, which is essential for chromosome segregation. Furthermore, the direct function of chromosomes is not only to produce RanGTP but, in addition, to release key mitotic regulators from chromatin. Chromatin-remodeling factors and nuclear pore complex proteins, which have established functions on chromatin in interphase, dissociate from mitotic chromatin and function in spindle assembly or maintenance. Thus, chromosomes actively organize their own segregation using chromatin-releasing mitotic regulators as well as RanGTP.

  16. Mitotic regulation by NIMA-related kinases

    Directory of Open Access Journals (Sweden)

    Blot Joelle

    2007-08-01

    Full Text Available Abstract The NIMA-related kinases represent a family of serine/threonine kinases implicated in cell cycle control. The founding member of this family, the NIMA kinase of Aspergillus nidulans, as well as the fission yeast homologue Fin1, contribute to multiple aspects of mitotic progression including the timing of mitotic entry, chromatin condensation, spindle organization and cytokinesis. Mammals contain a large family of eleven NIMA-related kinases, named Nek1 to Nek11. Of these, there is now substantial evidence that Nek2, Nek6, Nek7 and Nek9 also regulate mitotic events. At least three of these kinases, as well as NIMA and Fin1, have been localized to the microtubule organizing centre of their respective species, namely the centrosome or spindle pole body. Here, they have important functions in microtubule organization and mitotic spindle assembly. Other Nek kinases have been proposed to play microtubule-dependent roles in non-dividing cells, most notably in regulating the axonemal microtubules of cilia and flagella. In this review, we discuss the evidence that NIMA-related kinases make a significant contribution to the orchestration of mitotic progression and thereby protect cells from chromosome instability. Furthermore, we highlight their potential as novel chemotherapeutic targets.

  17. Bacterial mitotic machineries

    DEFF Research Database (Denmark)

    Gerdes, Kenn; Møller-Jensen, Jakob; Ebersbach, Gitte; Kruse, Torben; Nordström, Kurt

    2004-01-01

    Here, we review recent progress that yields fundamental new insight into the molecular mechanisms behind plasmid and chromosome segregation in prokaryotic cells. In particular, we describe how prokaryotic actin homologs form mitotic machineries that segregate DNA before cell division. Thus, the P......M protein of plasmid R1 forms F actin-like filaments that separate and move plasmid DNA from mid-cell to the cell poles. Evidence from three different laboratories indicate that the morphogenetic MreB protein may be involved in segregation of the bacterial chromosome....

  18. Mitotic chromosome condensation in vertebrates

    Energy Technology Data Exchange (ETDEWEB)

    Vagnarelli, Paola, E-mail: P.Vagnarelli@ed.ac.uk

    2012-07-15

    Work from several laboratories over the past 10-15 years has revealed that, within the interphase nucleus, chromosomes are organized into spatially distinct territories [T. Cremer, C. Cremer, Chromosome territories, nuclear architecture and gene regulation in mammalian cells, Nat. Rev. Genet. 2 (2001) 292-301 and T. Cremer, M. Cremer, S. Dietzel, S. Muller, I. Solovei, S. Fakan, Chromosome territories-a functional nuclear landscape, Curr. Opin. Cell Biol. 18 (2006) 307-316]. The overall compaction level and intranuclear location varies as a function of gene density for both entire chromosomes [J.A. Croft, J.M. Bridger, S. Boyle, P. Perry, P. Teague,W.A. Bickmore, Differences in the localization and morphology of chromosomes in the human nucleus, J. Cell Biol. 145 (1999) 1119-1131] and specific chromosomal regions [N.L. Mahy, P.E. Perry, S. Gilchrist, R.A. Baldock, W.A. Bickmore, Spatial organization of active and inactive genes and noncoding DNA within chromosome territories, J. Cell Biol. 157 (2002) 579-589] (Fig. 1A, A'). In prophase, when cyclin B activity reaches a high threshold, chromosome condensation occurs followed by Nuclear Envelope Breakdown (NEB) [1]. At this point vertebrate chromosomes appear as compact structures harboring an attachment point for the spindle microtubules physically recognizable as a primary constriction where the two sister chromatids are held together. The transition from an unshaped interphase chromosome to the highly structured mitotic chromosome (compare Figs. 1A and B) has fascinated researchers for several decades now; however a definite picture of how this process is achieved and regulated is not yet in our hands and it will require more investigation to comprehend the complete process. From a biochemical point of view a vertebrate mitotic chromosomes is composed of DNA, histone proteins (60%) and non-histone proteins (40%) [6]. I will discuss below what is known to date on the contribution of these two different classes

  19. Mitotic chromosome condensation in vertebrates

    International Nuclear Information System (INIS)

    Work from several laboratories over the past 10–15 years has revealed that, within the interphase nucleus, chromosomes are organized into spatially distinct territories [T. Cremer, C. Cremer, Chromosome territories, nuclear architecture and gene regulation in mammalian cells, Nat. Rev. Genet. 2 (2001) 292–301 and T. Cremer, M. Cremer, S. Dietzel, S. Muller, I. Solovei, S. Fakan, Chromosome territories—a functional nuclear landscape, Curr. Opin. Cell Biol. 18 (2006) 307–316]. The overall compaction level and intranuclear location varies as a function of gene density for both entire chromosomes [J.A. Croft, J.M. Bridger, S. Boyle, P. Perry, P. Teague,W.A. Bickmore, Differences in the localization and morphology of chromosomes in the human nucleus, J. Cell Biol. 145 (1999) 1119–1131] and specific chromosomal regions [N.L. Mahy, P.E. Perry, S. Gilchrist, R.A. Baldock, W.A. Bickmore, Spatial organization of active and inactive genes and noncoding DNA within chromosome territories, J. Cell Biol. 157 (2002) 579–589] (Fig. 1A, A'). In prophase, when cyclin B activity reaches a high threshold, chromosome condensation occurs followed by Nuclear Envelope Breakdown (NEB) [1]. At this point vertebrate chromosomes appear as compact structures harboring an attachment point for the spindle microtubules physically recognizable as a primary constriction where the two sister chromatids are held together. The transition from an unshaped interphase chromosome to the highly structured mitotic chromosome (compare Figs. 1A and B) has fascinated researchers for several decades now; however a definite picture of how this process is achieved and regulated is not yet in our hands and it will require more investigation to comprehend the complete process. From a biochemical point of view a vertebrate mitotic chromosomes is composed of DNA, histone proteins (60%) and non-histone proteins (40%) [6]. I will discuss below what is known to date on the contribution of these two different

  20. Phosphoproteins are components of mitotic microtubule organizing centers.

    OpenAIRE

    Vandre, D.D.; Davis, F. M.; Rao, P. N.; Borisy, G G

    1984-01-01

    Protein phosphorylation has been suggested as an important control mechanism for the events leading toward the initiation and completion of mitosis. Using a monoclonal antibody recognizing a class of phosphoproteins abundant in mitotic cells, we demonstrated the localization of a subset of these phosphoproteins to several discrete mitotic structures. Patchy immunofluorescence was present in the interphase nuclei, but a significant increase in nuclear immunofluorescence was apparent at prophas...

  1. Nuclear Chk1 prevents premature mitotic entry.

    Science.gov (United States)

    Matsuyama, Makoto; Goto, Hidemasa; Kasahara, Kousuke; Kawakami, Yoshitaka; Nakanishi, Makoto; Kiyono, Tohru; Goshima, Naoki; Inagaki, Masaki

    2011-07-01

    Chk1 inhibits the premature activation of the cyclin-B1-Cdk1. However, it remains controversial whether Chk1 inhibits Cdk1 in the centrosome or in the nucleus before the G2-M transition. In this study, we examined the specificity of the mouse monoclonal anti-Chk1 antibody DCS-310, with which the centrosome was stained. Conditional Chk1 knockout in mouse embryonic fibroblasts reduced nuclear but not centrosomal staining with DCS-310. In Chk1(+/myc) human colon adenocarcinoma (DLD-1) cells, Chk1 was detected in the nucleus but not in the centrosome using an anti-Myc antibody. Through the combination of protein array and RNAi technologies, we identified Ccdc-151 as a protein that crossreacted with DCS-310 on the centrosome. Mitotic entry was delayed by expression of the Chk1 mutant that localized in the nucleus, although forced immobilization of Chk1 to the centrosome had little impact on the timing of mitotic entry. These results suggest that nuclear but not centrosomal Chk1 contributes to correct timing of mitotic entry. PMID:21628425

  2. Human Nek7-interactor RGS2 is required for mitotic spindle organization.

    Science.gov (United States)

    de Souza, Edmarcia Elisa; Hehnly, Heidi; Perez, Arina Marina; Meirelles, Gabriela Vaz; Smetana, Juliana Helena Costa; Doxsey, Stephen; Kobarg, Jörg

    2015-01-01

    The mitotic spindle apparatus is composed of microtubule (MT) networks attached to kinetochores organized from 2 centrosomes (a.k.a. spindle poles). In addition to this central spindle apparatus, astral MTs assemble at the mitotic spindle pole and attach to the cell cortex to ensure appropriate spindle orientation. We propose that cell cycle-related kinase, Nek7, and its novel interacting protein RGS2, are involved in mitosis regulation and spindle formation. We found that RGS2 localizes to the mitotic spindle in a Nek7-dependent manner, and along with Nek7 contributes to spindle morphology and mitotic spindle pole integrity. RGS2-depletion leads to a mitotic-delay and severe defects in the chromosomes alignment and congression. Importantly, RGS2 or Nek7 depletion or even overexpression of wild-type or kinase-dead Nek7, reduced γ-tubulin from the mitotic spindle poles. In addition to causing a mitotic delay, RGS2 depletion induced mitotic spindle misorientation coinciding with astral MT-reduction. We propose that these phenotypes directly contribute to a failure in mitotic spindle alignment to the substratum. In conclusion, we suggest a molecular mechanism whereupon Nek7 and RGS2 may act cooperatively to ensure proper mitotic spindle organization. PMID:25664600

  3. Micromechanics of human mitotic chromosomes

    International Nuclear Information System (INIS)

    Eukaryote cells dramatically reorganize their long chromosomal DNAs to facilitate their physical segregation during mitosis. The internal organization of folded mitotic chromosomes remains a basic mystery of cell biology; its understanding would likely shed light on how chromosomes are separated from one another as well as into chromosome structure between cell divisions. We report biophysical experiments on single mitotic chromosomes from human cells, where we combine micromanipulation, nano-Newton-scale force measurement and biochemical treatments to study chromosome connectivity and topology. Results are in accord with previous experiments on amphibian chromosomes and support the 'chromatin network' model of mitotic chromosome structure. Prospects for studies of chromosome-organizing proteins using siRNA expression knockdowns, as well as for differential studies of chromosomes with and without mutations associated with genetic diseases, are also discussed

  4. Rab11 endosomes contribute to mitotic spindle organization and orientation.

    Science.gov (United States)

    Hehnly, Heidi; Doxsey, Stephen

    2014-03-10

    During interphase, Rab11-GTPase-containing endosomes recycle endocytic cargo. However, little is known about Rab11 endosomes in mitosis. Here, we show that Rab11 localizes to the mitotic spindle and regulates dynein-dependent endosome localization at poles. We found that mitotic recycling endosomes bind γ-TuRC components and associate with tubulin in vitro. Rab11 depletion or dominant-negative Rab11 expression disrupts astral microtubules, delays mitosis, and redistributes spindle pole proteins. Reciprocally, constitutively active Rab11 increases astral microtubules, restores γ-tubulin spindle pole localization, and generates robust spindles. This suggests a role for Rab11 activity in spindle pole maturation during mitosis. Rab11 depletion causes misorientation of the mitotic spindle and the plane of cell division. These findings suggest a molecular mechanism for the organization of astral microtubules and the mitotic spindle through Rab11-dependent control of spindle pole assembly and function. We propose that Rab11 and its associated endosomes cocontribute to these processes through retrograde transport to poles by dynein. PMID:24561039

  5. Smurf2 as a novel mitotic regulator: From the spindle assembly checkpoint to tumorigenesis

    Directory of Open Access Journals (Sweden)

    Moore Finola E

    2009-07-01

    Full Text Available Abstract The execution of the mitotic program with high fidelity is dependent upon precise spatiotemporal regulation of posttranslational protein modifications. For example, the timely polyubiquitination of critical mitotic regulators by Anaphase Promoting Complex/Cyclosome (APC/C is essential for the metaphase to anaphase transition and mitotic exit. The spindle assembly checkpoint prevents unscheduled activity of APC/C-Cdc20 in early mitosis, allowing bipolar attachment of kinetochores to mitotic spindle and facilitating equal segregation of sister chromatids. The critical effector of the spindle checkpoint, Mitotic arrest deficient 2 (Mad2, is recruited to unattached kinetochores forming a complex with other regulatory proteins to efficiently and cooperatively inhibit APC/C-Cdc20. A weakened and/or dysfunctional spindle checkpoint has been linked to the development of genomic instability in both cell culture and animal models, and evidence suggests that aberrant regulation of the spindle checkpoint plays a critical role in human carcinogenesis. Recent studies have illuminated a network of both degradative and non-degradative ubiquitination events that regulate the metaphase to anaphase transition and mitotic exit. Within this context, our recent work showed that the HECT (Homologous to E6-AP C-terminus-family E3 ligase Smurf2 (Smad specific ubiquitin regulatory factor 2, known as a negative regulator of transforming growth factor-beta (TGF-β signaling, is required for a functional spindle checkpoint by promoting the functional localization and stability of Mad2. Here we discuss putative models explaining the role of Smurf2 as a new regulator in the spindle checkpoint. The dynamic mitotic localization of Smurf2 to the centrosome and other critical mitotic structures provides implications about mitotic checkpoint control dependent on various ubiquitination events. Finally, deregulated Smurf2 activity may contribute to carcinogenesis by

  6. Radiation-induced mitotic catastrophe in PARG-deficient cells

    International Nuclear Information System (INIS)

    Poly(ADP-ribosyl)ation is a post-translational modification of proteins involved in the regulation of chromatin structure, DNA metabolism, cell division and cell death. Through the hydrolysis of poly(ADP-ribose) (PAR), Poly(ADP-ribose) glyco-hydrolase (PARG) has a crucial role in the control of life-and-death balance following DNA insult. Comprehension of PARG function has been hindered by the existence of many PARG isoforms encoded by a single gene and displaying various subcellular localizations. To gain insight into the function of PARG in response to irradiation, we constitutively and stably knocked down expression of PARG isoforms in HeLa cells. PARG depletion leading to PAR accumulation was not deleterious to undamaged cells and was in fact rather beneficial, because it protected cells from spontaneous single-strand breaks and telomeric abnormalities. By contrast, PARG-deficient cells showed increased radiosensitivity, caused by defects in the repair of single- and double-strand breaks and in mitotic spindle checkpoint, leading to alteration of progression of mitosis. Irradiated PARG-deficient cells displayed centrosome amplification leading to mitotic supernumerary spindle poles, and accumulated aberrant mitotic figures, which induced either polyploidy or cell death by mitotic catastrophe. Our results suggest that PARG could be a novel potential therapeutic target for radiotherapy. (authors)

  7. Variations in the association of papillomavirus E2 proteins with mitotic chromosomes

    OpenAIRE

    Jaquelline G de Oliveira; Colf, Leremy A.; Alison A McBride

    2006-01-01

    The E2 protein segregates episomal bovine papillomavirus (BPV) genomes to daughter cells by tethering them to mitotic chromosomes, thus ensuring equal distribution and retention of viral DNA. To date, only the BPV1 E2 protein has been shown to bind to mitotic chromosomes. We assessed the localization of 13 different animal and human E2 proteins from seven papillomavirus genera, and we show that most of them are stably bound to chromosomes throughout mitosis. Furthermore, in contrast to the ra...

  8. Brd4 Marks Select Genes on Mitotic Chromatin and Directs Postmitotic Transcription

    OpenAIRE

    Dey, Anup; Nishiyama, Akira; Karpova, Tatiana; McNally, James; Ozato, Keiko

    2009-01-01

    On entry into mitosis, many transcription factors dissociate from chromatin, resulting in global transcriptional shutdown. During mitosis, some genes are marked to ensure the inheritance of their expression in the next generation of cells. The nature of mitotic gene marking, however, has been obscure. Brd4 is a double bromodomain protein that localizes to chromosomes during mitosis and is implicated in holding mitotic memory. In interphase, Brd4 interacts with P-TEFb and functions as a global...

  9. Loading of PAX3 to Mitotic Chromosomes Is Mediated by Arginine Methylation and Associated with Waardenburg Syndrome.

    Science.gov (United States)

    Wu, Tsu-Fang; Yao, Ya-Li; Lai, I-Lu; Lai, Chien-Chen; Lin, Pei-Lun; Yang, Wen-Ming

    2015-08-14

    PAX3 is a transcription factor critical to gene regulation in mammalian development. Mutations in PAX3 are associated with Waardenburg syndrome (WS), but the mechanism of how mutant PAX3 proteins cause WS remains unclear. Here, we found that PAX3 loads on mitotic chromosomes using its homeodomain. PAX3 WS mutants with mutations in homeodomain lose the ability to bind mitotic chromosomes. Moreover, loading of PAX3 on mitotic chromosomes requires arginine methylation, which is regulated by methyltransferase PRMT5 and demethylase JMJD6. Mutant PAX3 proteins that lose mitotic chromosome localization block cell proliferation and normal development of zebrafish. These results reveal the molecular mechanism of PAX3s loading on mitotic chromosomes and the importance of this localization pattern in normal development. Our findings suggest that PAX3 WS mutants interfere with the normal functions of PAX3 in a dominant negative manner, which is important to the understanding of the pathogenesis of Waardenburg syndrome. PMID:26149688

  10. Loading of PAX3 to Mitotic Chromosomes Is Mediated by Arginine Methylation and Associated with Waardenburg Syndrome*

    Science.gov (United States)

    Wu, Tsu-Fang; Yao, Ya-Li; Lai, I-Lu; Lai, Chien-Chen; Lin, Pei-Lun; Yang, Wen-Ming

    2015-01-01

    PAX3 is a transcription factor critical to gene regulation in mammalian development. Mutations in PAX3 are associated with Waardenburg syndrome (WS), but the mechanism of how mutant PAX3 proteins cause WS remains unclear. Here, we found that PAX3 loads on mitotic chromosomes using its homeodomain. PAX3 WS mutants with mutations in homeodomain lose the ability to bind mitotic chromosomes. Moreover, loading of PAX3 on mitotic chromosomes requires arginine methylation, which is regulated by methyltransferase PRMT5 and demethylase JMJD6. Mutant PAX3 proteins that lose mitotic chromosome localization block cell proliferation and normal development of zebrafish. These results reveal the molecular mechanism of PAX3s loading on mitotic chromosomes and the importance of this localization pattern in normal development. Our findings suggest that PAX3 WS mutants interfere with the normal functions of PAX3 in a dominant negative manner, which is important to the understanding of the pathogenesis of Waardenburg syndrome. PMID:26149688

  11. Arsenic-induced promoter hypomethylation and over-expression of ERCC2 reduces DNA repair capacity in humans by non-disjunction of the ERCC2-Cdk7 complex.

    Science.gov (United States)

    Paul, Somnath; Banerjee, Nilanjana; Chatterjee, Aditi; Sau, Tanmoy J; Das, Jayanta K; Mishra, Prafulla K; Chakrabarti, Partha; Bandyopadhyay, Arun; Giri, Ashok K

    2014-04-01

    Arsenic in drinking water is of critical concern in West Bengal, India, as it results in several physiological symptoms including dermatological lesions and cancers. Impairment of the DNA repair mechanism has been associated with arsenic-induced genetic damage as well as with several cancers. ERCC2 (Excision Repair Cross-Complementing rodent repair, complementation group 2), mediates DNA-repair by interacting with Cdk-activating kinase (CAK) complex, which helps in DNA proof-reading during transcription. Arsenic metabolism alters epigenetic regulation; we tried to elucidate the regulation of ERCC2 in arsenic-exposed humans. Water, urine, nails, hair and blood samples from one hundred and fifty seven exposed and eighty eight unexposed individuals were collected. Dose dependent validation was done in vitro using HepG2 and HEK-293. Arsenic content in the biological samples was higher in the exposed individuals compared with the content in unexposed individuals (p < 0.001). Bisulfite-modified methylation specific PCR showed a significant (p < 0.0001) hypomethylation of the ERCC2 promoter in the arsenic-exposed individuals. Densitometric analysis of immunoblots showed a nearly two-fold increase in expression of ERCC2 in exposed individuals, but there was an enhanced genotoxic insult as measured by micronuclei frequency. Immuno-precipitation and western blotting revealed an increased (p < 0.001) association of Cdk7 with ERCC2 in highly arsenic exposed individuals. The decrease in CAK activity was determined by observing the intensity of Ser(392) phosphorylation in p53, in vitro, which decreased with an increase in arsenic dose. Thus we infer that arsenic biotransformation leads to promoter hypomethylation of ERCC2, which in turn inhibits the normal functioning of the CAK-complex, thus affecting DNA-repair; this effect was highest among the arsenic exposed individuals with dermatological lesions. PMID:24473091

  12. Analysis of the mitotic exit control system using locked levels of stable mitotic cyclin.

    Science.gov (United States)

    Drapkin, Benjamin J; Lu, Ying; Procko, Andrea L; Timney, Benjamin L; Cross, Frederick R

    2009-01-01

    Cyclin-dependent kinase (Cdk) both promotes mitotic entry (spindle assembly and anaphase) and inhibits mitotic exit (spindle disassembly and cytokinesis), leading to an elegant quantitative hypothesis that a single cyclin oscillation can function as a ratchet to order these events. This ratchet is at the core of a published ODE model for the yeast cell cycle. However, the ratchet model requires appropriate cyclin dose-response thresholds. Here, we test the inhibition of mitotic exit in budding yeast using graded levels of stable mitotic cyclin (Clb2). In opposition to the ratchet model, stable levels of Clb2 introduced dose-dependent delays, rather than hard thresholds, that varied by mitotic exit event. The ensuing cell cycle was highly abnormal, suggesting a novel reason for cyclin degradation. Cdc14 phosphatase antagonizes Clb2-Cdk, and Cdc14 is released from inhibitory nucleolar sequestration independently of stable Clb2. Thus, Cdc14/Clb2 balance may be the appropriate variable for mitotic regulation. Although our results are inconsistent with the aforementioned ODE model, revision of the model to allow Cdc14/Clb2 balance to control mitotic exit corrects these discrepancies, providing theoretical support for our conclusions. PMID:19920813

  13. Radmis, a novel mitotic spindle protein that functions in cell division of neural progenitors.

    Directory of Open Access Journals (Sweden)

    Takahito Yumoto

    Full Text Available Developmental dynamics of neural stem/progenitor cells (NSPCs are crucial for embryonic and adult neurogenesis, but its regulatory factors are not fully understood. By differential subtractive screening with NSPCs versus their differentiated progenies, we identified the radmis (radial fiber and mitotic spindle/ckap2l gene, a novel microtubule-associated protein (MAP enriched in NSPCs. Radmis is a putative substrate for the E3-ubiquitin ligase, anaphase promoting complex/cyclosome (APC/C, and is degraded via the KEN box. Radmis was highly expressed in regions of active neurogenesis throughout life, and its distribution was dynamically regulated during NSPC division. In embryonic and perinatal brains, radmis localized to bipolar mitotic spindles and radial fibers (basal processes of dividing NSPCs. As central nervous system development proceeded, radmis expression was lost in most brain regions, except for several neurogenic regions. In adult brain, radmis expression persisted in the mitotic spindles of both slowly-dividing stem cells and rapid amplifying progenitors. Overexpression of radmis in vitro induced hyper-stabilization of microtubules, severe defects in mitotic spindle formation, and mitotic arrest. In vivo gain-of-function using in utero electroporation revealed that radmis directed a reduction in NSPC proliferation and a concomitant increase in cell cycle exit, causing a reduction in the Tbr2-positive basal progenitor population and shrinkage of the embryonic subventricular zone. Besides, radmis loss-of-function by shRNAs induced the multipolar mitotic spindle structure, accompanied with the catastrophe of chromosome segregation including the long chromosome bridge between two separating daughter nuclei. These findings uncover the indispensable role of radmis in mitotic spindle formation and cell-cycle progression of NSPCs.

  14. The bipolar assembly domain of the mitotic motor kinesin-5

    OpenAIRE

    Acar, Seyda; Carlson, David B.; Budamagunta, Madhu S.; Yarov-Yarovoy, Vladimir; Correia, John J.; Niñonuevo, Milady R.; Jia, Weitao; Tao, Li; Leary, Julie A.; Voss, John C.; Evans, James E.; Scholey, Jonathan M.

    2013-01-01

    An outstanding unresolved question is how does the mitotic spindle utilize microtubules and mitotic motors to coordinate accurate chromosome segregation during mitosis? This process depends upon the mitotic motor, kinesin-5, whose unique bipolar architecture, with pairs of motor domains lying at opposite ends of a central rod, allows it to crosslink microtubules within the mitotic spindle and to coordinate their relative sliding during spindle assembly, maintenance and elongation. The structu...

  15. Sequential phosphorylation of GRASP65 during mitotic Golgi disassembly

    Directory of Open Access Journals (Sweden)

    Danming Tang

    2012-09-01

    GRASP65 phosphorylation during mitosis and dephosphorylation after mitosis are required for Golgi disassembly and reassembly during the cell cycle. At least eight phosphorylation sites on GRASP65 have been identified, but whether they are modified in a coordinated fashion during mitosis is so far unknown. In this study, we raised phospho-specific antibodies that recognize phosphorylated T220/T224, S277 and S376 residues of GRASP65, respectively. Biochemical analysis showed that cdc2 phosphorylates all three sites, while plk1 enhances the phosphorylation. Microscopic studies using these antibodies for double and triple labeling demonstrate sequential phosphorylation and dephosphorylation during the cell cycle. S277 and S376 are phosphorylated from late G2 phase through metaphase until telophase when the new Golgi is reassembled. T220/224 is not modified until prophase, but is highly modified from prometaphase to anaphase. In metaphase, phospho-T220/224 signal localizes on both Golgi haze and mitotic Golgi clusters that represent dispersed Golgi vesicles and Golgi remnants, respectively, while phospho-S277 and S376 labeling is more concentrated on mitotic Golgi clusters. Expression of a phosphorylation-resistant GRASP65 mutant T220A/T224A inhibited mitotic Golgi fragmentation to a much larger extent than the expression of the S277A and S376A mutants. In cytokinesis, T220/224 dephosphorylation occurs prior to that of S277, but after S376. This study provides evidence that GRASP65 is sequentially phosphorylated and dephosphorylated during mitosis at different sites to orchestrate Golgi disassembly and reassembly during cell division, with phosphorylation of the T220/224 site being most critical in the process.

  16. Integrin-linked kinase regulates interphase and mitotic microtubule dynamics.

    Directory of Open Access Journals (Sweden)

    Simin Lim

    Full Text Available Integrin-linked kinase (ILK localizes to both focal adhesions and centrosomes in distinct multiprotein complexes. Its dual function as a kinase and scaffolding protein has been well characterized at focal adhesions, where it regulates integrin-mediated cell adhesion, spreading, migration and signaling. At the centrosomes, ILK regulates mitotic spindle organization and centrosome clustering. Our previous study showed various spindle defects after ILK knockdown or inhibition that suggested alteration in microtubule dynamics. Since ILK expression is frequently elevated in many cancer types, we investigated the effects of ILK overexpression on microtubule dynamics. We show here that overexpressing ILK in HeLa cells was associated with a shorter duration of mitosis and decreased sensitivity to paclitaxel, a chemotherapeutic agent that suppresses microtubule dynamics. Measurement of interphase microtubule dynamics revealed that ILK overexpression favored microtubule depolymerization, suggesting that microtubule destabilization could be the mechanism behind the decreased sensitivity to paclitaxel, which is known to stabilize microtubules. Conversely, the use of a small molecule inhibitor selective against ILK, QLT-0267, resulted in suppressed microtubule dynamics, demonstrating a new mechanism of action for this compound. We further show that treatment of HeLa cells with QLT-0267 resulted in higher inter-centromere tension in aligned chromosomes during mitosis, slower microtubule regrowth after cold depolymerization and the presence of a more stable population of spindle microtubules. These results demonstrate that ILK regulates microtubule dynamics in both interphase and mitotic cells.

  17. Measuring mitotic spindle dynamics in budding yeast

    Science.gov (United States)

    Plumb, Kemp

    In order to carry out its life cycle and produce viable progeny through cell division, a cell must successfully coordinate and execute a number of complex processes with high fidelity, in an environment dominated by thermal noise. One important example of such a process is the assembly and positioning of the mitotic spindle prior to chromosome segregation. The mitotic spindle is a modular structure composed of two spindle pole bodies, separated in space and spanned by filamentous proteins called microtubules, along which the genetic material of the cell is held. The spindle is responsible for alignment and subsequent segregation of chromosomes into two equal parts; proper spindle positioning and timing ensure that genetic material is appropriately divided amongst mother and daughter cells. In this thesis, I describe fluorescence confocal microscopy and automated image analysis algorithms, which I have used to observe and analyze the real space dynamics of the mitotic spindle in budding yeast. The software can locate structures in three spatial dimensions and track their movement in time. By selecting fluorescent proteins which specifically label the spindle poles and cell periphery, mitotic spindle dynamics have been measured in a coordinate system relevant to the cell division. I describe how I have characterised the accuracy and precision of the algorithms by simulating fluorescence data for both spindle poles and the budding yeast cell surface. In this thesis I also describe the construction of a microfluidic apparatus that allows for the measurement of long time-scale dynamics of individual cells and the development of a cell population. The tools developed in this thesis work will facilitate in-depth quantitative analysis of the non-equilibrium processes in living cells.

  18. The Mechanics of Mitotic Cell Rounding

    OpenAIRE

    Stewart, Martin

    2012-01-01

    During mitosis, adherent animal cells undergo a drastic shape change, from essentially flat to round, in a process known as mitotic cell rounding (MCR). The aim of this thesis was to critically examine the physical and biological basis of MCR. The experimental part of this thesis employed a combined optical microscope-atomic force microscope (AFM) setup in conjunction with flat tipless cantilevers to analyze cell mechanics, shape and volume. To this end, two AFM assays were developed: the ...

  19. Cell cycle-dependent SUMO-1 conjugation to nuclear mitotic apparatus protein (NuMA)

    Energy Technology Data Exchange (ETDEWEB)

    Seo, Jae Sung; Kim, Ha Na; Kim, Sun-Jick; Bang, Jiyoung; Kim, Eun-A; Sung, Ki Sa [Department of Biological Sciences, Sungkyunkwan University, Suwon 440-746 (Korea, Republic of); Yoon, Hyun-Joo [TissueGene Inc. 9605 Medical Center Dr., Rockville, MD 20850 (United States); Yoo, Hae Yong [Department of Health Sciences and Technology, Samsung Advanced Institute for Health Sciences and Technology, Samsung Medical Center, Sungkyunkwan University, Seoul 135-710 (Korea, Republic of); Choi, Cheol Yong, E-mail: choicy@skku.ac.kr [Department of Biological Sciences, Sungkyunkwan University, Suwon 440-746 (Korea, Republic of)

    2014-01-03

    Highlights: •NuMA is modified by SUMO-1 in a cell cycle-dependent manner. •NuMA lysine 1766 is the primary target site for SUMOylation. •SUMOylation-deficient NuMA induces multiple spindle poles during mitosis. •SUMOylated NuMA induces microtubule bundling. -- Abstract: Covalent conjugation of proteins with small ubiquitin-like modifier 1 (SUMO-1) plays a critical role in a variety of cellular functions including cell cycle control, replication, and transcriptional regulation. Nuclear mitotic apparatus protein (NuMA) localizes to spindle poles during mitosis, and is an essential component in the formation and maintenance of mitotic spindle poles. Here we show that NuMA is a target for covalent conjugation to SUMO-1. We find that the lysine 1766 residue is the primary NuMA acceptor site for SUMO-1 conjugation. Interestingly, SUMO modification of endogenous NuMA occurs at the entry into mitosis and this modification is reversed after exiting from mitosis. Knockdown of Ubc9 or forced expression of SENP1 results in impairment of the localization of NuMA to mitotic spindle poles during mitosis. The SUMOylation-deficient NuMA mutant is defective in microtubule bundling, and multiple spindles are induced during mitosis. The mitosis-dependent dynamic SUMO-1 modification of NuMA might contribute to NuMA-mediated formation and maintenance of mitotic spindle poles during mitosis.

  20. A mitotic SKAP isoform regulates spindle positioning at astral microtubule plus ends.

    Science.gov (United States)

    Kern, David M; Nicholls, Peter K; Page, David C; Cheeseman, Iain M

    2016-05-01

    The Astrin/SKAP complex plays important roles in mitotic chromosome alignment and centrosome integrity, but previous work found conflicting results for SKAP function. Here, we demonstrate that SKAP is expressed as two distinct isoforms in mammals: a longer, testis-specific isoform that was used for the previous studies in mitotic cells and a novel, shorter mitotic isoform. Unlike the long isoform, short SKAP rescues SKAP depletion in mitosis and displays robust microtubule plus-end tracking, including localization to astral microtubules. Eliminating SKAP microtubule binding results in severe chromosome segregation defects. In contrast, SKAP mutants specifically defective for plus-end tracking facilitate proper chromosome segregation but display spindle positioning defects. Cells lacking SKAP plus-end tracking have reduced Clasp1 localization at microtubule plus ends and display increased lateral microtubule contacts with the cell cortex, which we propose results in unbalanced dynein-dependent cortical pulling forces. Our work reveals an unappreciated role for the Astrin/SKAP complex as an astral microtubule mediator of mitotic spindle positioning. PMID:27138257

  1. Cell cycle-dependent SUMO-1 conjugation to nuclear mitotic apparatus protein (NuMA)

    International Nuclear Information System (INIS)

    Highlights: •NuMA is modified by SUMO-1 in a cell cycle-dependent manner. •NuMA lysine 1766 is the primary target site for SUMOylation. •SUMOylation-deficient NuMA induces multiple spindle poles during mitosis. •SUMOylated NuMA induces microtubule bundling. -- Abstract: Covalent conjugation of proteins with small ubiquitin-like modifier 1 (SUMO-1) plays a critical role in a variety of cellular functions including cell cycle control, replication, and transcriptional regulation. Nuclear mitotic apparatus protein (NuMA) localizes to spindle poles during mitosis, and is an essential component in the formation and maintenance of mitotic spindle poles. Here we show that NuMA is a target for covalent conjugation to SUMO-1. We find that the lysine 1766 residue is the primary NuMA acceptor site for SUMO-1 conjugation. Interestingly, SUMO modification of endogenous NuMA occurs at the entry into mitosis and this modification is reversed after exiting from mitosis. Knockdown of Ubc9 or forced expression of SENP1 results in impairment of the localization of NuMA to mitotic spindle poles during mitosis. The SUMOylation-deficient NuMA mutant is defective in microtubule bundling, and multiple spindles are induced during mitosis. The mitosis-dependent dynamic SUMO-1 modification of NuMA might contribute to NuMA-mediated formation and maintenance of mitotic spindle poles during mitosis

  2. Mlp1 Acts as a Mitotic Scaffold to Spatially Regulate Spindle Assembly Checkpoint Proteins in Aspergillus nidulans

    OpenAIRE

    De Souza, Colin P.; Hashmi, Shahr B.; Nayak, Tania; Oakley, Berl; Osmani, Stephen A.

    2009-01-01

    During open mitosis several nuclear pore complex (NPC) proteins have mitotic specific localizations and functions. We find that the Aspergillus nidulans Mlp1 NPC protein has previously unrealized mitotic roles involving spatial regulation of spindle assembly checkpoint (SAC) proteins. In interphase, An-Mlp1 tethers the An-Mad1 and An-Mad2 SAC proteins to NPCs. During a normal mitosis, An-Mlp1, An-Mad1, and An-Mad2 localize similarly on, and around, kinetochores until telophase when they trans...

  3. A Novel Pathway that Coordinates Mitotic Exit with Spindle Position

    OpenAIRE

    Nelson, Scott A.; Cooper, John A.

    2007-01-01

    In budding yeast, the spindle position checkpoint (SPC) delays mitotic exit until the mitotic spindle moves into the neck between the mother and bud. This checkpoint works by inhibiting the mitotic exit network (MEN), a signaling cascade initiated and controlled by Tem1, a small GTPase. Tem1 is regulated by a putative guanine exchange factor, Lte1, but the function and regulation of Lte1 remains poorly understood. Here, we identify novel components of the checkpoint that operate upstream of L...

  4. Novel insights into mitotic chromosome condensation

    Science.gov (United States)

    Piskadlo, Ewa; Oliveira, Raquel A.

    2016-01-01

    The fidelity of mitosis is essential for life, and successful completion of this process relies on drastic changes in chromosome organization at the onset of nuclear division. The mechanisms that govern chromosome compaction at every cell division cycle are still far from full comprehension, yet recent studies provide novel insights into this problem, challenging classical views on mitotic chromosome assembly. Here, we briefly introduce various models for chromosome assembly and known factors involved in the condensation process (e.g. condensin complexes and topoisomerase II). We will then focus on a few selected studies that have recently brought novel insights into the mysterious way chromosomes are condensed during nuclear division.

  5. Mitotic stopwatch for the blast fungus Magnaporthe oryzae during invasion of rice cells.

    Science.gov (United States)

    Jones, Kiersun; Jenkinson, Cory B; Borges Araújo, Maíra; Zhu, Jie; Kim, Rebecca Y; Kim, Dong Won; Khang, Chang Hyun

    2016-08-01

    To study nuclear dynamics of Magnaporthe oryzae, we developed a novel mitotic reporter strain with GFP-NLS (localized in nuclei during interphase but in the cytoplasm during mitosis) and H1-tdTomato (localized in nuclei throughout the cell cycle). Time-lapse confocal microscopy of the reporter strain during host cell invasion provided several new insights into nuclear division and migration in M. oryzae: (i) mitosis lasts about 5min; (ii) mitosis is semi-closed; (iii) septal pores are closed during mitosis; and (iv) a nucleus exhibits extreme constriction (approximately from 2μm to 0.5μm), elongation (over 5μm), and long migration (over 16μm). Our observations raise new questions about mechanisms controlling the mitotic dynamics, and the answers to these questions may result in new means to prevent fungal proliferation without negatively affecting the host cell cycle. PMID:27321562

  6. The budding yeast Ipl1/Aurora protein kinase regulates mitotic spindle disassembly

    OpenAIRE

    Buvelot, Stéphanie; Tatsutani, Sean Y.; Vermaak, Danielle; Biggins, Sue

    2003-01-01

    Ipl1p is the budding yeast member of the Aurora family of protein kinases, critical regulators of genomic stability that are required for chromosome segregation, the spindle checkpoint, and cytokinesis. Using time-lapse microscopy, we found that Ipl1p also has a function in mitotic spindle disassembly that is separable from its previously identified roles. Ipl1–GFP localizes to kinetochores from G1 to metaphase, transfers to the spindle after metaphase, and accumulates at the spindle midzone ...

  7. Mitotic Catastrophe的研究进展%Progress in Mitotic Catastrophe

    Institute of Scientific and Technical Information of China (English)

    张博; 周平坤

    2007-01-01

    细胞死亡是多细胞生物生命过程中重要的生理或病理现象,可分为坏死和程序性细胞死亡,而后者根据死亡细胞的形态学和发生机制的不同又可分为凋亡、自吞噬和mitotic catastrophe,其中mitotic catastrophe是近年来才被揭示报道,是指细胞在有丝分裂过程中死亡的现象,是一种发生在细胞有丝分裂期由于异常的细胞分裂而导致的细胞死亡,它常常伴随着细胞有丝分裂检查点的异常和基因或纺锤体结构的损伤而发生.现对mitotic catastrophe及相关的调控机制进行综述.

  8. Kaposi's sarcoma herpesvirus C-terminal LANA concentrates at pericentromeric and peri-telomeric regions of a subset of mitotic chromosomes

    International Nuclear Information System (INIS)

    The Kaposi's sarcoma-associated herpesvirus (KSHV) latency-associated nuclear antigen (LANA) tethers KSHV terminal repeat (TR) DNA to mitotic chromosomes to efficiently segregate episomes to progeny nuclei. LANA contains N- and C-terminal chromosome binding regions. We now show that C-terminal LANA preferentially concentrates to paired dots at pericentromeric and peri-telomeric regions of a subset of mitotic chromosomes through residues 996-1139. Deletions within C-terminal LANA abolished both self-association and chromosome binding, consistent with a requirement for self-association to bind chromosomes. A deletion abolishing TR DNA binding did not affect chromosome targeting, indicating LANA's localization is not due to binding its recognition sequence in chromosomal DNA. LANA distributed similarly on human and non-human mitotic chromosomes. These results are consistent with C-terminal LANA interacting with a cell factor that concentrates at pericentromeric and peri-telomeric regions of mitotic chromosomes

  9. The bipolar assembly domain of the mitotic motor kinesin-5.

    Science.gov (United States)

    Acar, Seyda; Carlson, David B; Budamagunta, Madhu S; Yarov-Yarovoy, Vladimir; Correia, John J; Niñonuevo, Milady R; Jia, Weitao; Tao, Li; Leary, Julie A; Voss, John C; Evans, James E; Scholey, Jonathan M

    2013-01-01

    An outstanding unresolved question is how does the mitotic spindle utilize microtubules and mitotic motors to coordinate accurate chromosome segregation during mitosis? This process depends upon the mitotic motor, kinesin-5, whose unique bipolar architecture, with pairs of motor domains lying at opposite ends of a central rod, allows it to crosslink microtubules within the mitotic spindle and to coordinate their relative sliding during spindle assembly, maintenance and elongation. The structural basis of kinesin-5's bipolarity is, however, unknown, as protein asymmetry has so far precluded its crystallization. Here we use electron microscopy of single molecules of kinesin-5 and its subfragments, combined with hydrodynamic analysis plus mass spectrometry, circular dichroism and site-directed spin label electron paramagnetic resonance spectroscopy, to show how a staggered antiparallel coiled-coil 'BASS' (bipolar assembly) domain directs the assembly of four kinesin-5 polypeptides into bipolar minifilaments. PMID:23299893

  10. Mitotic Diversity in Homeostatic Human Interfollicular Epidermis

    Directory of Open Access Journals (Sweden)

    Katharina Nöske

    2016-01-01

    Full Text Available Despite decades of skin research, regulation of proliferation and homeostasis in human epidermis is still insufficiently understood. To address the role of mitoses in tissue regulation, we utilized human long-term skin equivalents and systematically assessed mitoses during early epidermal development and long-term epidermal regeneration. We now demonstrate four different orientations: (1 horizontal, i.e., parallel to the basement membrane (BM and suggestive of symmetric divisions; (2 oblique with an angle of 45°–70°; or (3 perpendicular, suggestive of asymmetric division. In addition, we demonstrate a fourth substantial fraction of suprabasal mitoses, many of which are committed to differentiation (Keratin K10-positive. As verified also for normal human skin, this spatial mitotic organization is part of the regulatory program of human epidermal tissue homeostasis. As a potential marker for asymmetric division, we investigated for Numb and found that it was evenly spread in almost all undifferentiated keratinocytes, but indeed asymmetrically distributed in some mitoses and particularly frequent under differentiation-repressing low-calcium conditions. Numb deletion (stable knockdown by CRISPR/Cas9, however, did not affect proliferation, neither in a three-day follow up study by life cell imaging nor during a 14-day culture period, suggesting that Numb is not essential for the general control of keratinocyte division.

  11. Regulation of mitotic spindle orientation: an integrated view.

    Science.gov (United States)

    di Pietro, Florencia; Echard, Arnaud; Morin, Xavier

    2016-08-01

    Mitotic spindle orientation is essential for cell fate decisions, epithelial maintenance, and tissue morphogenesis. In most animal cell types, the dynein motor complex is anchored at the cell cortex and exerts pulling forces on astral microtubules to position the spindle. Early studies identified the evolutionarily conserved Gαi/LGN/NuMA complex as a key regulator that polarizes cortical force generators. In recent years, a combination of genetics, biochemistry, modeling, and live imaging has contributed to decipher the mechanisms of spindle orientation. Here, we highlight the dynamic nature of the assembly of this complex and discuss the molecular regulation of its localization. Remarkably, a number of LGN-independent mechanisms were described recently, whereas NuMA remains central in most pathways involved in recruiting force generators at the cell cortex. We also describe the emerging role of the actin cortex in spindle orientation and discuss how dynamic astral microtubule formation is involved. We further give an overview on instructive external signals that control spindle orientation in tissues. Finally, we discuss the influence of cell geometry and mechanical forces on spindle orientation. PMID:27432284

  12. A Protein Interaction Map of the Mitotic Spindle

    OpenAIRE

    Wong, Jonathan; Nakajima, Yuko; Westermann, Stefan; Shang, Ching; Kang, Jung-seog; Goodner, Crystal; Houshmand, Pantea; Fields, Stanley; Chan, Clarence S.M.; Drubin, David; Barnes, Georjana; Hazbun, Tony

    2007-01-01

    The mitotic spindle consists of a complex network of proteins that segregates chromosomes in eukaryotes. To strengthen our understanding of the molecular composition, organization, and regulation of the mitotic spindle, we performed a system-wide two-hybrid screen on 94 proteins implicated in spindle function in Saccharomyces cerevisiae. We report 604 predominantly novel interactions that were detected in multiple screens, involving 303 distinct prey proteins. We uncovered a pattern of extens...

  13. A mitotic recombination map proximal to the APC locus on chromosome 5q and assessment of influences on colorectal cancer risk

    Directory of Open Access Journals (Sweden)

    Clark Susan

    2009-06-01

    Full Text Available Abstract Background Mitotic recombination is important for inactivating tumour suppressor genes by copy-neutral loss of heterozygosity (LOH. Although meiotic recombination maps are plentiful, little is known about mitotic recombination. The APC gene (chr5q21 is mutated in most colorectal tumours and its usual mode of LOH is mitotic recombination. Methods We mapped mitotic recombination boundaries ("breakpoints" between the centromere (~50 Mb and APC (~112 Mb in early colorectal tumours. Results Breakpoints were non-random, with the highest frequency between 65 Mb and 75 Mb, close to a low copy number repeat region (68–71 Mb. There were, surprisingly, few breakpoints close to APC, contrary to expectations were there constraints on tumorigenesis caused by uncovering recessive lethal alleles or if mitotic recombination were mechanistically favoured by a longer residual chromosome arm. The locations of mitotic and meiotic recombination breakpoints were correlated, suggesting that the two types of recombination are influenced by similar processes, whether mutational or selective in origin. Breakpoints were also associated with higher local G+C content. The recombination and gain/deletion breakpoint maps on 5q were not, however, associated, perhaps owing to selective constraints on APC dosage in early colorectal tumours. Since polymorphisms within the region of frequent mitotic recombination on 5q might influence the frequency of LOH, we tested the 68–71 Mb low copy number repeat and nearby tagSNPs, but no associations with colorectal cancer risk were found. Conclusion LOH on 5q is non-random, but local factors do not greatly influence the rate of LOH at APC or explain inter differential susceptibility to colorectal tumours.

  14. Evidence that phosphorylation by the mitotic kinase Cdk1 promotes ICER monoubiquitination and nuclear delocalization

    Energy Technology Data Exchange (ETDEWEB)

    Memin, Elisabeth, E-mail: molinac@mail.montclair.edu [Department of Biochemistry and Molecular Biology, University of Medicine and Dentistry of New Jersey-New Jersey Medical School, Newark, NJ 07103 (United States); Genzale, Megan [Department of Biochemistry and Molecular Biology, University of Medicine and Dentistry of New Jersey-New Jersey Medical School, Newark, NJ 07103 (United States); Crow, Marni; Molina, Carlos A. [Department of Biology and Molecular Biology, Montclair State University, Montclair, NJ, 07043 (United States)

    2011-10-15

    In contrast to normal prostatic cells, the transcriptional repressor Inducible cAMP Early Repressor (ICER) is undetected in the nuclei of prostate cancer cells. The molecular mechanisms for ICER abnormal expression in prostate cancer cells remained largely unknown. In this report data is presented demonstrating that ICER is phosphorylated by the mitotic kinase cdk1. Phosphorylation of ICER on a discrete residue targeted ICER to be monoubiquitinated. Different from unphosphorylated, phosphorylated and polyubiquitinated ICER, monoubiquitinated ICER was found to be cytosolic. Taken together, these results hinted on a mechanism for the observed abnormal subcellular localization of ICER in human prostate tumors.

  15. Evidence that phosphorylation by the mitotic kinase Cdk1 promotes ICER monoubiquitination and nuclear delocalization

    International Nuclear Information System (INIS)

    In contrast to normal prostatic cells, the transcriptional repressor Inducible cAMP Early Repressor (ICER) is undetected in the nuclei of prostate cancer cells. The molecular mechanisms for ICER abnormal expression in prostate cancer cells remained largely unknown. In this report data is presented demonstrating that ICER is phosphorylated by the mitotic kinase cdk1. Phosphorylation of ICER on a discrete residue targeted ICER to be monoubiquitinated. Different from unphosphorylated, phosphorylated and polyubiquitinated ICER, monoubiquitinated ICER was found to be cytosolic. Taken together, these results hinted on a mechanism for the observed abnormal subcellular localization of ICER in human prostate tumors.

  16. Molecular origin of mitotic aneuploidies in preimplantation embryos.

    Science.gov (United States)

    Mantikou, Eleni; Wong, Kai Mee; Repping, Sjoerd; Mastenbroek, Sebastiaan

    2012-12-01

    Mitotic errors are common in human preimplantation embryos. The occurrence of mitotic errors is highest during the first three cleavages after fertilization and as a result about three quarters of human preimplantation embryos show aneuploidies and are chromosomally mosaic at day three of development. The origin of these preimplantation mitotic aneuploidies and the molecular mechanisms involved are being discussed in this review. At later developmental stages the mitotic aneuploidy rate is lower. Mechanisms such as cell arrest, apoptosis, active correction of the aneuploidies and preferential allocation of the aneuploid cells to the extra-embryonic tissues could underlie this lower rate. Understanding the mechanisms that cause mitotic aneuploidies in human preimplantation embryos and the way human preimplantation embryos deal with these aneuploidies might lead to ways to limit the occurrence of aneuploidies, in order to ultimately increase the quality of embryos and with that the likelihood of a successful pregnancy in IVF/ICSI. This article is part of a Special Issue entitled: Molecular Genetics of Human Reproductive Failure. PMID:22771499

  17. Mitotic Exit Control as an Evolved Complex System

    Energy Technology Data Exchange (ETDEWEB)

    Bosl, W; Li, R

    2005-04-25

    The exit from mitosis is the last critical decision a cell has to make during a division cycle. A complex regulatory system has evolved to evaluate the success of mitotic events and control this decision. Whereas outstanding genetic work in yeast has led to rapid discovery of a large number of interacting genes involved in the control of mitotic exit, it has also become increasingly difficult to comprehend the logic and mechanistic features embedded in the complex molecular network. Our view is that this difficulty stems in part from the attempt to explain mitotic exit control using concepts from traditional top-down engineering design, and that exciting new results from evolutionary engineering design applied to networks and electronic circuits may lend better insights. We focus on four particularly intriguing features of the mitotic exit control system: the two-stepped release of Cdc14; the self-activating nature of Tem1 GTPase; the spatial sensor associated with the spindle pole body; and the extensive redundancy in the mitotic exit network. We attempt to examine these design features from the perspective of evolutionary design and complex system engineering.

  18. Molecular mechanisms of DNA recombination: testing mitotic and meiotic models

    International Nuclear Information System (INIS)

    A hyperhaploid n + 1 strain of Saccharomyces cerevisiae (LBL1) disomic for chromosome VII was employed to isolate hyper-rec and hypo-rec mutations affecting spontaneous mitotic gene conversion and intergenic recombination. The genotype of LBL1 permits simultaneous and independent identification of rec mutations that enhance or diminish gene conversion and those that enhance or diminish intergenic recombination. Five phenotypic groups of rec mutants were isolated following ultraviolet light mutagenesis. Rec mutations that simultaneously abolish or enhance both classes of recombinational events were detected. These results demonstrate that gene conversion and intergenic recombination are under joint genetic control in mitotic cells. Conversion-specific and intergenic recombination-specific rec mutants were also recovered. Their properties indicate that conversion and intergenic recombination are separable pheonomena dependent upon discrete REC genes. The rec mutants isolated in LBL1 provide a method to test molecular models of mitotic and meiotic recombination

  19. Identification of a BET family Bromodomain / Casein Kinase II / TAF-containing complex as a regulator of mitotic condensin function

    OpenAIRE

    Kim, Hyun-Soo; Mukhopadhyay, Rituparna; Rothbart, Scott B.; Silva, Andrea C.; Vanoosthuyse, Vincent; Radovani, Ernest; Kislinger, Thomas; Roguev, Assen; Ryan, Colm J.; Xu, Jiewei; Jahari, Harlizawati; Hardwick, Kevin G.; Greenblatt, Jack F.; Krogan, Nevan J.; Fillingham, Jeffrey S.

    2014-01-01

    Condensin is a central regulator of mitotic genome structure, with mutants showing poorly condensed chromosomes and profound segregation defects. Here we identify NCT complex, comprising the Nrc1 BET-family tandem bromodomain protein (SPAC631.02), Casein Kinase II (CKII) and several TAFs, as a regulator of condensin function. We show that NCT and condensin bind similar genomic regions, but only briefly co-localize during the periods of chromosome condensation and decondensation. This pattern ...

  20. The moyamoya disease susceptibility variant RNF213 R4810K (rs112735431) induces genomic instability by mitotic abnormality

    Energy Technology Data Exchange (ETDEWEB)

    Hitomi, Toshiaki [Department of Health and Environmental Sciences, Graduate School of Medicine, Kyoto University, Kyoto (Japan); Habu, Toshiyuki [Radiation Biology Center, Kyoto University, Kyoto (Japan); Kobayashi, Hatasu; Okuda, Hiroko; Harada, Kouji H. [Department of Health and Environmental Sciences, Graduate School of Medicine, Kyoto University, Kyoto (Japan); Osafune, Kenji [Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto (Japan); Taura, Daisuke; Sone, Masakatsu [Department of Medicine and Clinical Science, Graduate School of Medicine, Kyoto University, Kyoto (Japan); Asaka, Isao; Ameku, Tomonaga; Watanabe, Akira; Kasahara, Tomoko; Sudo, Tomomi; Shiota, Fumihiko [Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto (Japan); Hashikata, Hirokuni; Takagi, Yasushi [Department of Neurosurgery, Graduate School of Medicine, Kyoto University, Kyoto (Japan); Morito, Daisuke [Faculty of Life Sciences, Kyoto Sangyo University, Kyoto (Japan); Miyamoto, Susumu [Department of Neurosurgery, Graduate School of Medicine, Kyoto University, Kyoto (Japan); Nakao, Kazuwa [Department of Medicine and Clinical Science, Graduate School of Medicine, Kyoto University, Kyoto (Japan); Koizumi, Akio, E-mail: koizumi.akio.5v@kyoto-u.ac.jp [Department of Health and Environmental Sciences, Graduate School of Medicine, Kyoto University, Kyoto (Japan)

    2013-10-04

    Highlights: •Overexpression of RNF213 R4810K inhibited cell proliferation. •Overexpression of RNF213 R4810K had the time of mitosis 4-fold and mitotic failure. •R4810K formed a complex with MAD2 more readily than wild-type. •iPSECs from the MMD patients had elevated mitotic failure compared from the control. •RNF213 R4810K induced mitotic abnormality and increased risk of aneuploidy. -- Abstract: Moyamoya disease (MMD) is a cerebrovascular disease characterized by occlusive lesions in the Circle of Willis. The RNF213 R4810K polymorphism increases susceptibility to MMD. In the present study, we characterized phenotypes caused by overexpression of RNF213 wild type and R4810K variant in the cell cycle to investigate the mechanism of proliferation inhibition. Overexpression of RNF213 R4810K in HeLa cells inhibited cell proliferation and extended the time of mitosis 4-fold. Ablation of spindle checkpoint by depletion of mitotic arrest deficiency 2 (MAD2) did not shorten the time of mitosis. Mitotic morphology in HeLa cells revealed that MAD2 colocalized with RNF213 R4810K. Immunoprecipitation revealed an RNF213/MAD2 complex: R4810K formed a complex with MAD2 more readily than RNF213 wild-type. Desynchronized localization of MAD2 was observed more frequently during mitosis in fibroblasts from patients (n = 3, 61.0 ± 8.2%) compared with wild-type subjects (n = 6, 13.1 ± 7.7%; p < 0.01). Aneuploidy was observed more frequently in fibroblasts (p < 0.01) and induced pluripotent stem cells (iPSCs) (p < 0.03) from patients than from wild-type subjects. Vascular endothelial cells differentiated from iPSCs (iPSECs) of patients and an unaffected carrier had a longer time from prometaphase to metaphase than those from controls (p < 0.05). iPSECs from the patients and unaffected carrier had significantly increased mitotic failure rates compared with controls (p < 0.05). Thus, RNF213 R4810K induced mitotic abnormalities and increased risk of genomic instability.

  1. The moyamoya disease susceptibility variant RNF213 R4810K (rs112735431) induces genomic instability by mitotic abnormality

    International Nuclear Information System (INIS)

    Highlights: •Overexpression of RNF213 R4810K inhibited cell proliferation. •Overexpression of RNF213 R4810K had the time of mitosis 4-fold and mitotic failure. •R4810K formed a complex with MAD2 more readily than wild-type. •iPSECs from the MMD patients had elevated mitotic failure compared from the control. •RNF213 R4810K induced mitotic abnormality and increased risk of aneuploidy. -- Abstract: Moyamoya disease (MMD) is a cerebrovascular disease characterized by occlusive lesions in the Circle of Willis. The RNF213 R4810K polymorphism increases susceptibility to MMD. In the present study, we characterized phenotypes caused by overexpression of RNF213 wild type and R4810K variant in the cell cycle to investigate the mechanism of proliferation inhibition. Overexpression of RNF213 R4810K in HeLa cells inhibited cell proliferation and extended the time of mitosis 4-fold. Ablation of spindle checkpoint by depletion of mitotic arrest deficiency 2 (MAD2) did not shorten the time of mitosis. Mitotic morphology in HeLa cells revealed that MAD2 colocalized with RNF213 R4810K. Immunoprecipitation revealed an RNF213/MAD2 complex: R4810K formed a complex with MAD2 more readily than RNF213 wild-type. Desynchronized localization of MAD2 was observed more frequently during mitosis in fibroblasts from patients (n = 3, 61.0 ± 8.2%) compared with wild-type subjects (n = 6, 13.1 ± 7.7%; p < 0.01). Aneuploidy was observed more frequently in fibroblasts (p < 0.01) and induced pluripotent stem cells (iPSCs) (p < 0.03) from patients than from wild-type subjects. Vascular endothelial cells differentiated from iPSCs (iPSECs) of patients and an unaffected carrier had a longer time from prometaphase to metaphase than those from controls (p < 0.05). iPSECs from the patients and unaffected carrier had significantly increased mitotic failure rates compared with controls (p < 0.05). Thus, RNF213 R4810K induced mitotic abnormalities and increased risk of genomic instability

  2. Mitotic Spindle Disruption by Alternating Electric Fields Leads to Improper Chromosome Segregation and Mitotic Catastrophe in Cancer Cells.

    Science.gov (United States)

    Giladi, Moshe; Schneiderman, Rosa S; Voloshin, Tali; Porat, Yaara; Munster, Mijal; Blat, Roni; Sherbo, Shay; Bomzon, Zeev; Urman, Noa; Itzhaki, Aviran; Cahal, Shay; Shteingauz, Anna; Chaudhry, Aafia; Kirson, Eilon D; Weinberg, Uri; Palti, Yoram

    2015-01-01

    Tumor Treating Fields (TTFields) are low intensity, intermediate frequency, alternating electric fields. TTFields are a unique anti-mitotic treatment modality delivered in a continuous, noninvasive manner to the region of a tumor. It was previously postulated that by exerting directional forces on highly polar intracellular elements during mitosis, TTFields could disrupt the normal assembly of spindle microtubules. However there is limited evidence directly linking TTFields to an effect on microtubules. Here we report that TTFields decrease the ratio between polymerized and total tubulin, and prevent proper mitotic spindle assembly. The aberrant mitotic events induced by TTFields lead to abnormal chromosome segregation, cellular multinucleation, and caspase dependent apoptosis of daughter cells. The effect of TTFields on cell viability and clonogenic survival substantially depends upon the cell division rate. We show that by extending the duration of exposure to TTFields, slowly dividing cells can be affected to a similar extent as rapidly dividing cells. PMID:26658786

  3. Airborne urban particles (Milan winter-PM2.5) cause mitotic arrest and cell death: Effects on DNA, mitochondria, AhR binding and spindle organization

    International Nuclear Information System (INIS)

    Highlights: → PM2.5 induces mitotic arrest in BEAS-2B cells. → PM2.5 induces DNA damage and activates DNA damage response. → AhR regulated genes (Cyp1A1, Cyp1B1 and AhRR) are upregulated after PM exposure. → Mitotic spindle assembly is perturbed in PM exposed cells. - Abstract: Airborne particulate matter (PM) is considered to be an important contributor to lung diseases. In the present study we report that Milan winter-PM2.5 inhibited proliferation in human bronchial epithelial cells (BEAS-2B) by inducing mitotic arrest. The cell cycle arrest was followed by an increase in mitotic-apoptotic cells, mitotic slippage and finally an increase in 'classical' apoptotic cells. Exposure to winter-PM10 induced only a slight effect which may be due to the presence of PM2.5 in this fraction while pure combustion particles failed to disturb mitosis. Fewer cells expressing the mitosis marker phospho-histone H3 compared to cells with condensed chromosomes, suggest that PM2.5 induced premature mitosis. PM2.5 was internalized into the cells and often localized in laminar organelles, although particles without apparent plasma membrane covering were also seen. In PM-containing cells mitochondria and lysosomes were often damaged, and in mitotic cells fragmented chromosomes often appeared. PM2.5 induced DNA strands breaks and triggered a DNA-damage response characterized by increased phosphorylation of ATM, Chk2 and H2AX; as well as induced a marked increase in expression of the aryl hydrocarbon receptor (AhR)-regulated genes, CYP1A1, CYP1B1 and AhRR. Furthermore, some disturbance of the organization of microtubules was indicated. It is hypothesized that the induced mitotic arrest and following cell death was due to a premature chromosome condensation caused by a combination of DNA, mitochondrial and spindle damage.

  4. Action study of mumio preparation on mitotic index by autoradiography way

    International Nuclear Information System (INIS)

    In this chapter author made conclusion that leading of mumio preparation raise the mitotic activity and promote of rapid passing by cells mitotic cycle that lead to rapid partition and raising of quantity cells in hemopoietic organs

  5. The effects of X-rays on the mitotic activity of mouse epidermis

    Energy Technology Data Exchange (ETDEWEB)

    Knowlton, N.P. Jr.; Hempelmann, L.H.; Hoffman, J.G.

    1949-04-19

    This report describes a simplified technique of obtaining the mitotic index of mouse skin and indicates the surprising sensitivity of the mitotic activity of mouse epithelium to the effects of x-rays.

  6. Changes in distribution of nuclear matrix antigens during the mitotic cell cycle.

    Science.gov (United States)

    Chaly, N; Bladon, T; Setterfield, G; Little, J E; Kaplan, J G; Brown, D L

    1984-08-01

    We examined the distribution of nonlamin nuclear matrix antigens during the mitotic cell cycle in mouse 3T3 fibroblasts. Four monoclonal antibodies produced against isolated nuclear matrices were used to characterize antigens by the immunoblotting of isolated nuclear matrix preparations, and were used to localize the antigens by indirect immunofluorescence. For comparison, lamins and histones were localized using human autoimmune antibodies. At interphase, the monoclonal antibodies recognized non-nucleolar and nonheterochromatin nuclear components. Antibody P1 stained the nuclear periphery homogeneously, with some small invaginations toward the interior of the nucleus. Antibody I1 detected an antigen distributed as fine granules throughout the nuclear interior. Monoclonals PI1 and PI2 stained both the nuclear periphery and interior, with some characteristic differences. During mitosis, P1 and I1 were chromosome-associated, whereas PI1 and PI2 dispersed in the cytoplasm. Antibody P1 heavily stained the periphery of the chromosome mass, and we suggest that the antigen may play a role in maintaining interphase and mitotic chromosome order. With antibody I1, bright granules were distributed along the chromosomes and there was also some diffuse internal staining. The antigen to I1 may be involved in chromatin/chromosome higher-order organization throughout the cell cycle. Antibodies PI1 and PI2 were redistributed independently during prophase, and dispersed into the cytoplasm during prometaphase. Antibody PI2 also detected antigen associated with the spindle poles. PMID:6378926

  7. Cdc20 control of cell fate during prolonged mitotic arrest

    DEFF Research Database (Denmark)

    Nilsson, Jakob

    2011-01-01

    The fate of cells arrested in mitosis by antimitotic compounds is complex but is influenced by competition between pathways promoting cell death and pathways promoting mitotic exit. As components of both of these pathways are regulated by Cdc20-dependent degradation, I hypothesize that variations...

  8. THE INFLUENCE OF CAFFEINE ON MITOTIC DIVISION AT CAPSICUM ANNUUM L.

    Directory of Open Access Journals (Sweden)

    Elena Rosu

    2006-08-01

    Full Text Available The paper presents, the caffeine effects in mitotic division at Capsicum annuum L.. The treatment has determined the lessening of the mitotic index (comparative with the control variant, until mitotic division total inhibition, as well as an growth frequency of division aberation in anaphase and telophase.

  9. Mechanism of APC/CCDC20 activation by mitotic phosphorylation.

    Science.gov (United States)

    Qiao, Renping; Weissmann, Florian; Yamaguchi, Masaya; Brown, Nicholas G; VanderLinden, Ryan; Imre, Richard; Jarvis, Marc A; Brunner, Michael R; Davidson, Iain F; Litos, Gabriele; Haselbach, David; Mechtler, Karl; Stark, Holger; Schulman, Brenda A; Peters, Jan-Michael

    2016-05-10

    Chromosome segregation and mitotic exit are initiated by the 1.2-MDa ubiquitin ligase APC/C (anaphase-promoting complex/cyclosome) and its coactivator CDC20 (cell division cycle 20). To avoid chromosome missegregation, APC/C(CDC20) activation is tightly controlled. CDC20 only associates with APC/C in mitosis when APC/C has become phosphorylated and is further inhibited by a mitotic checkpoint complex until all chromosomes are bioriented on the spindle. APC/C contains 14 different types of subunits, most of which are phosphorylated in mitosis on multiple sites. However, it is unknown which of these phospho-sites enable APC/C(CDC20) activation and by which mechanism. Here we have identified 68 evolutionarily conserved mitotic phospho-sites on human APC/C bound to CDC20 and have used the biGBac technique to generate 47 APC/C mutants in which either all 68 sites or subsets of them were replaced by nonphosphorylatable or phospho-mimicking residues. The characterization of these complexes in substrate ubiquitination and degradation assays indicates that phosphorylation of an N-terminal loop region in APC1 is sufficient for binding and activation of APC/C by CDC20. Deletion of the N-terminal APC1 loop enables APC/C(CDC20) activation in the absence of mitotic phosphorylation or phospho-mimicking mutations. These results indicate that binding of CDC20 to APC/C is normally prevented by an autoinhibitory loop in APC1 and that its mitotic phosphorylation relieves this inhibition. The predicted location of the N-terminal APC1 loop implies that this loop controls interactions between the N-terminal domain of CDC20 and APC1 and APC8. These results reveal how APC/C phosphorylation enables CDC20 to bind and activate the APC/C in mitosis. PMID:27114510

  10. Impaired mitotic progression and preimplantation lethality in mice lacking OMCG1, a new evolutionarily conserved nuclear protein

    DEFF Research Database (Denmark)

    Artus, Jérôme; Vandormael-Pournin, Sandrine; Frödin, Morten; Nacerddine, Karim; Babinet, Charles; Cohen-Tannoudji, Michel

    2005-01-01

    vitro cultured Omcg1-null blastocysts exhibit a dramatic reduction in the total cell number, a high mitotic index, and the presence of abnormal mitotic figures. Importantly, we found that Omcg1 disruption results in the lengthening of M phase rather than in a mitotic block. We show that the mitotic...

  11. Mitochondrial genome regulates mitotic fidelity by maintaining centrosomal homeostasis

    OpenAIRE

    Donthamsetty, Shashikiran; Brahmbhatt, Meera; Pannu, Vaishali; Rida, Padmashree CG; Ramarathinam, Sujatha; Ogden, Angela; Cheng, Alice; Singh, Keshav K.; Aneja, Ritu

    2014-01-01

    Centrosomes direct spindle morphogenesis to assemble a bipolar mitotic apparatus to enable error-free chromosome segregation and preclude chromosomal instability (CIN). Amplified centrosomes, a hallmark of cancer cells, set the stage for CIN, which underlies malignant transformation and evolution of aggressive phenotypes. Several studies report CIN and a tumorigenic and/or aggressive transformation in mitochondrial DNA (mtDNA)-depleted cells. Although several nuclear-encoded proteins are impl...

  12. A transmembrane inner nuclear membrane protein in the mitotic spindle

    OpenAIRE

    Figueroa, Ricardo; Gudise, Santhosh; Larsson, Veronica; Hallberg, Einar

    2010-01-01

    We have recently characterized a novel transmembrane protein of the inner nuclear membrane of mammalian cells. The protein has two very interesting features. First, despite being an integral membrane protein it is able to concentrate in the membranes colocalizing with the mitotic spindle in metaphase and anaphase. Hence, the protein was named Samp1, Spindle associated membrane protein 1. Secondly, it displays a functional connection to centrosomes. This article discusses various aspects of Sa...

  13. A Genetic Map of DICTYOSTELIUM DISCOIDEUM Based on Mitotic Recombination

    OpenAIRE

    Welker, Dennis L.; Williams, Keith L.

    1982-01-01

    A genetic map of the cellular slime mold Dictyostelium discoideum is presented in which 42 loci are ordered on five of the seven linkage groups. Although most of the loci were ordered using standing mitotic crossing-over techniques in which recessive selective markers were employed, use was also made of unselected recombined haploid strains. Consistent with cytological studies in which the chromosomes appear to be acrocentric, only a single arm has been found for each of the five linkage grou...

  14. Mitotic Origins of Chromosomal Instability in Colorectal Cancer

    OpenAIRE

    Dalton, W. Brian; Yang, Vincent W.

    2007-01-01

    Mitosis is a crucial part of the cell cycle. A successful mitosis requires the proper execution of many complex cellular behaviors. Thus, there are many points at which mitosis may be disrupted. In cancer cells, chronic disruption of mitosis can lead to unequal segregation of chromosomes, a phenomenon known as chromosomal instability. A majority of colorectal tumors suffer from this instability, and recent studies have begun to reveal the specific ways in which mitotic defects promote chromos...

  15. Mitotic exit: Determining the PP2A dephosphorylation program.

    Science.gov (United States)

    Pereira, Gislene; Schiebel, Elmar

    2016-08-29

    In mitotic exit, proteins that were highly phosphorylated are sequentially targeted by the phosphatase PP2A-B55, but what underlies substrate selection is unclear. In this issue, Cundell et al. (2016. J. Cell Biol http://dx.doi.org/10.1083/jcb.201606033) identify the determinants of PP2A-B55's dephosphorylation program, thereby influencing spindle disassembly, nuclear envelope reformation, and cytokinesis. PMID:27551057

  16. Evaluation of Morphological Diversity, Mitotic Instability and Fertility in Tritipyrum

    OpenAIRE

    zolfaghar shahriari; mohammad assad; hosein hassani

    2014-01-01

    Tritipyrum is prone to mitotic instability, stiff straw and low fertility. A complete randomized design was used to evaluate morphological diversity, fertility and grain yield of 7 new synthetic Tritipyrum lines and their F1 offspring’sin a cross with bread wheat. Fertility, harvest index, biological yield, 1000-grains weight (TGW) and grain yield of Tritipyrum derived genotypes were significantly lower than bread wheat and Triticale. The morphology of Tritipyrum was found to be similar with ...

  17. Carbamazepine induces mitotic arrest in mammalian Vero cells

    International Nuclear Information System (INIS)

    We reported recently that the anticonvulsant drug carbamazepine, at supratherapeutic concentrations, exerts antiproliferative effects in mammalian Vero cells, but the underlying mechanism has not been elucidated. This motivates us to examine rigorously whether growth arrest was associated with structural changes in cellular organization during mitosis. In the present work, we found that exposure of the cells to carbamazepine led to an increase in mitotic index, mainly due to the sustained block at the metaphase/anaphase boundary, with the consequent inhibition of cell proliferation. Indirect immunofluorescence, using antibodies directed against spindle apparatus proteins, revealed that mitotic arrest was associated with formation of monopolar spindles, caused by impairment of centrosome separation. The final consequence of the spindle defects induced by carbamazepine, depended on the duration of cell cycle arrest. Following the time course of accumulation of metaphase and apoptotic cells during carbamazepine treatments, we observed a causative relationship between mitotic arrest and induction of cell death. Conversely, cells released from the block of metaphase by removal of the drug, continued to progress through mitosis and resume normal proliferation. Our results show that carbamazepine shares a common antiproliferative mechanism with spindle-targeted drugs and contribute to a better understanding of the cytostatic activity previously described in Vero cells. Additional studies are in progress to extend these initial findings that define a novel mode of action of carbamazepine in cultured mammalian cells

  18. Role of senescence and mitotic catastrophe in cancer therapy

    Directory of Open Access Journals (Sweden)

    Shukla Yogeshwer

    2010-01-01

    Full Text Available Abstract Senescence and mitotic catastrophe (MC are two distinct crucial non-apoptotic mechanisms, often triggered in cancer cells and tissues in response to anti-cancer drugs. Chemotherapeuticals and myriad other factors induce cell eradication via these routes. While senescence drives the cells to a state of quiescence, MC drives the cells towards death during the course of mitosis. The senescent phenotype distinguishes tumor cells that survived drug exposure but lost the ability to form colonies from those that recover and proliferate after treatment. Although senescent cells do not proliferate, they are metabolically active and may secrete proteins with potential tumor-promoting activities. The other anti-proliferative response of tumor cells is MC that is a form of cell death that results from abnormal mitosis and leads to the formation of interphase cells with multiple micronuclei. Different classes of cytotoxic agents induce MC, but the pathways of abnormal mitosis differ depending on the nature of the inducer and the status of cell-cycle checkpoints. In this review, we compare the two pathways and mention that they are activated to curb the growth of tumors. Altogether, we have highlighted the possibilities of the use of senescence targeting drugs, mitotic kinases and anti-mitotic agents in fabricating novel strategies in cancer control.

  19. Carbamazepine induces mitotic arrest in mammalian Vero cells

    Energy Technology Data Exchange (ETDEWEB)

    Perez Martin, J.M.; Fernandez Freire, P.; Labrador, V. [Departamento de Biologia, Facultad de Ciencias, Universidad Autonoma de Madrid, Cantoblanco, 28049 Madrid (Spain); Hazen, M.J. [Departamento de Biologia, Facultad de Ciencias, Universidad Autonoma de Madrid, Cantoblanco, 28049 Madrid (Spain)], E-mail: mariajose.hazen@uam.es

    2008-01-01

    We reported recently that the anticonvulsant drug carbamazepine, at supratherapeutic concentrations, exerts antiproliferative effects in mammalian Vero cells, but the underlying mechanism has not been elucidated. This motivates us to examine rigorously whether growth arrest was associated with structural changes in cellular organization during mitosis. In the present work, we found that exposure of the cells to carbamazepine led to an increase in mitotic index, mainly due to the sustained block at the metaphase/anaphase boundary, with the consequent inhibition of cell proliferation. Indirect immunofluorescence, using antibodies directed against spindle apparatus proteins, revealed that mitotic arrest was associated with formation of monopolar spindles, caused by impairment of centrosome separation. The final consequence of the spindle defects induced by carbamazepine, depended on the duration of cell cycle arrest. Following the time course of accumulation of metaphase and apoptotic cells during carbamazepine treatments, we observed a causative relationship between mitotic arrest and induction of cell death. Conversely, cells released from the block of metaphase by removal of the drug, continued to progress through mitosis and resume normal proliferation. Our results show that carbamazepine shares a common antiproliferative mechanism with spindle-targeted drugs and contribute to a better understanding of the cytostatic activity previously described in Vero cells. Additional studies are in progress to extend these initial findings that define a novel mode of action of carbamazepine in cultured mammalian cells.

  20. A comprehensive characterization of the nuclear microRNA repertoire of post-mitotic neurons

    Directory of Open Access Journals (Sweden)

    Sharof Abdumalikovich Khudayberdiev

    2013-11-01

    Full Text Available MicroRNAs (miRNAs are small non-coding RNAs with important functions in the development and plasticity of post-mitotic neurons. In addition to the well-described cytoplasmic function of miRNAs in post-transcriptional gene regulation, recent studies suggested that miRNAs could also be involved in transcriptional and post-transcriptional regulatory processes in the nuclei of proliferating cells. However, whether miRNAs localize to and function within the nucleus of post-mitotic neurons is unknown. Using a combination of microarray hybridization and small RNA deep sequencing, we identified a specific subset of miRNAs which are enriched in the nuclei of neurons. Nuclear enrichment of specific candidate miRNAs (miR-25 and miR-92a could be independently validated by Northern blot, quantitative real-time PCR (qRT-PCR and fluorescence in situ hybridization (FISH. By cross-comparison to published reports, we found that nuclear accumulation of miRNAs might be linked to a down-regulation of miRNA expression during in vitro development of cortical neurons. Importantly, by generating a comprehensive isomiR profile of the nuclear and cytoplasmic compartment, we found a significant overrepresentation of guanine nucleotides at the 3’ terminus of nuclear-enriched isomiRs, suggesting the presence of neuron-specific mechanisms involved in miRNA nuclear localization. In conclusion, our results provide a starting point for future studies addressing the nuclear function of specific miRNAs and the detailed mechanisms underlying subcellular localization of miRNAs in neurons and possibly other polarized cell types.

  1. Proteomics Analysis with a Nano Random Forest Approach Reveals Novel Functional Interactions Regulated by SMC Complexes on Mitotic Chromosomes.

    Science.gov (United States)

    Ohta, Shinya; Montaño-Gutierrez, Luis F; de Lima Alves, Flavia; Ogawa, Hiromi; Toramoto, Iyo; Sato, Nobuko; Morrison, Ciaran G; Takeda, Shunichi; Hudson, Damien F; Rappsilber, Juri; Earnshaw, William C

    2016-08-01

    Packaging of DNA into condensed chromosomes during mitosis is essential for the faithful segregation of the genome into daughter nuclei. Although the structure and composition of mitotic chromosomes have been studied for over 30 years, these aspects are yet to be fully elucidated. Here, we used stable isotope labeling with amino acids in cell culture to compare the proteomes of mitotic chromosomes isolated from cell lines harboring conditional knockouts of members of the condensin (SMC2, CAP-H, CAP-D3), cohesin (Scc1/Rad21), and SMC5/6 (SMC5) complexes. Our analysis revealed that these complexes associate with chromosomes independently of each other, with the SMC5/6 complex showing no significant dependence on any other chromosomal proteins during mitosis. To identify subtle relationships between chromosomal proteins, we employed a nano Random Forest (nanoRF) approach to detect protein complexes and the relationships between them. Our nanoRF results suggested that as few as 113 of 5058 detected chromosomal proteins are functionally linked to chromosome structure and segregation. Furthermore, nanoRF data revealed 23 proteins that were not previously suspected to have functional interactions with complexes playing important roles in mitosis. Subsequent small-interfering-RNA-based validation and localization tracking by green fluorescent protein-tagging highlighted novel candidates that might play significant roles in mitotic progression. PMID:27231315

  2. APC/C-Cdh1-dependent anaphase and telophase progression during mitotic slippage

    Directory of Open Access Journals (Sweden)

    Toda Kazuhiro

    2012-02-01

    Full Text Available Abstract Background The spindle assembly checkpoint (SAC inhibits anaphase progression in the presence of insufficient kinetochore-microtubule attachments, but cells can eventually override mitotic arrest by a process known as mitotic slippage or adaptation. This is a problem for cancer chemotherapy using microtubule poisons. Results Here we describe mitotic slippage in yeast bub2Δ mutant cells that are defective in the repression of precocious telophase onset (mitotic exit. Precocious activation of anaphase promoting complex/cyclosome (APC/C-Cdh1 caused mitotic slippage in the presence of nocodazole, while the SAC was still active. APC/C-Cdh1, but not APC/C-Cdc20, triggered anaphase progression (securin degradation, separase-mediated cohesin cleavage, sister-chromatid separation and chromosome missegregation, in addition to telophase onset (mitotic exit, during mitotic slippage. This demonstrates that an inhibitory system not only of APC/C-Cdc20 but also of APC/C-Cdh1 is critical for accurate chromosome segregation in the presence of insufficient kinetochore-microtubule attachments. Conclusions The sequential activation of APC/C-Cdc20 to APC/C-Cdh1 during mitosis is central to accurate mitosis. Precocious activation of APC/C-Cdh1 in metaphase (pre-anaphase causes mitotic slippage in SAC-activated cells. For the prevention of mitotic slippage, concomitant inhibition of APC/C-Cdh1 may be effective for tumor therapy with mitotic spindle poisons in humans.

  3. Mitotic Spindle Disruption by Alternating Electric Fields Leads to Improper Chromosome Segregation and Mitotic Catastrophe in Cancer Cells

    OpenAIRE

    Moshe Giladi; Schneiderman, Rosa S; Tali Voloshin; Yaara Porat; Mijal Munster; Roni Blat; Shay Sherbo; Zeev Bomzon; Noa Urman; Aviran Itzhaki; Shay Cahal; Anna Shteingauz; Aafia Chaudhry; Kirson, Eilon D.; Uri Weinberg

    2015-01-01

    Tumor Treating Fields (TTFields) are low intensity, intermediate frequency, alternating electric fields. TTFields are a unique anti-mitotic treatment modality delivered in a continuous, noninvasive manner to the region of a tumor. It was previously postulated that by exerting directional forces on highly polar intracellular elements during mitosis, TTFields could disrupt the normal assembly of spindle microtubules. However there is limited evidence directly linking TTFields to an effect on mi...

  4. Comparison of staining of mitotic figures by haematoxylin and eosin-and crystal violet stains, in oral epithelial dysplasia and squamous cell carcinoma

    Directory of Open Access Journals (Sweden)

    Ankle Madhuri

    2007-01-01

    Full Text Available Mitosis of cells gives rise to tissue integrity. Defects during mitosis bring about abnormalities. Excessive proliferation of cells due to increased mitosis is one such outcome, which is the hallmark in precancer and cancer. The localization of proliferating cells or their precursors may not be obvious and easy. Establishing an easy way to distinguish these mitotic cells will help in grading and understanding their biological potential. Although immunohistochemistry is an advanced method in use, the cost and time factor makes it less feasible for many laboratories. Selective histochemical stains like toluidine blue, giemsa and crystal violet have been used in tissues including the developing brain, neural tissue and skin. Aim of the study: 1To compare the staining of mitotic cells in haematoxylin and eosin with that in crystal violet. 2To compare the number of mitotic figures present in normal oral mucosa, epithelial dysplasia and oral squamous cell carcinoma in crystal violet-stained sections with that in H and E-stained sections. Materials and Methods: Ten tissues of normal oral mucosa and 15 tissues each of oral epithelial dysplasia seen in tobacco-associated leukoplakia and squamous cell carcinoma were studied to evaluate the selectivity of 1% crystal violet for mitotic figures. The staining was compared with standard H and E staining. Statistical analysis was done using Man-Whitney U test. Results: A statistically significant increase in the mean mitotic count was observed in crystal violet-stained sections of epithelial dysplasia as compared to the H and E-stained sections ( p = 0.0327. A similar increase in the mitotic counts was noted in crystal violet-stained sections of oral squamous cell carcinoma as compared to the H and E-stained sections.( p = 0.0443. No significant difference was found in the mitotic counts determined in dysplasia or carcinoma by either the crystal violet ( p = 0.4429 or the H and E-staining techniques ( p = 0

  5. Mitotic delay of irradiated cells and its connection with quantity of radiation injuries

    International Nuclear Information System (INIS)

    The study is dedicated to development of mathematical approach to interpret radiation-induced mitosic delay. An assumption is made that mitotic delay is conditioned by discrete injuries distributed in cells according to stochasticity of interaction of radiation and target substance. It is supposed to consider the problem on injuries nature causing mitotic delay and to use the developed method for accounting the effect of radiation-induced mitotic delay on registered chromosomal aberration yield. 10 refs.; 2 figs.; 3 tabs

  6. Cyto-3D-print to attach mitotic cells.

    Science.gov (United States)

    Castroagudin, Michelle R; Zhai, Yujia; Li, Zhi; Marnell, Michael G; Glavy, Joseph S

    2016-08-01

    The Cyto-3D-print is an adapter that adds cytospin capability to a standard centrifuge. Like standard cytospinning, Cyto-3D-print increases the surface attachment of mitotic cells while giving a higher degree of adaptability to other slide chambers than available commercial devices. The use of Cyto-3D-print is cost effective, safe, and applicable to many slide designs. It is durable enough for repeated use and made of biodegradable materials for environment-friendly disposal. PMID:26464272

  7. Microtubule Dynamics and Oscillating State for Mitotic Spindle

    CERN Document Server

    Rashid-Shomali, Safura

    2010-01-01

    We present a physical mechanism that can cause the mitotic spindle to oscillate. The driving force for this mechanism emerges from the polymerization of astral microtubules interacting with the cell cortex. We show that Brownian ratchet model for growing microtubules reaching the cell cortex, mediate an effective mass to the spindle body and therefore force it to oscillate. We compare the predictions of this mechanism with the previous mechanisms which were based on the effects of motor proteins. Finally we combine the effects of microtubules polymerization and motor proteins, and present the detailed phase diagram for possible oscillating states.

  8. Inhibition of mitotic-specific histone phophorylation by sodium arsenite

    Energy Technology Data Exchange (ETDEWEB)

    Cobo, J.M. [Universidad de Alcala de Henares, Madrid (Spain); Valdez, J.G.; Gurley, L.R. [Los Alamos National Lab., NM (United States)

    1994-10-01

    Synchronized cultures of Chinese hamster cells (line CHO) were used to measure the effects of 10{mu}M sodium arsenite on histone phosphorylation. This treatment caused cell proliferation to be temporarily arrested, after which the cells spontaneously resumed cell proliferation in a radiomimetric manner. Immediately following treatment, it was found that sodium arsenite affected only mitotic-specific HI and H3 phosphorylations. Neither interphase, nor mitotic, H2A and H4 phosphorylations were affected, nor was interphase HI Phosphorylation affected. The phosphorylation of HI was inhibited only in mitosis, reducing HI phosphorylation to 38.1% of control levels, which was the level of interphase HI phosphorylation. The phosphorylation of both H3 variants was inhibited in mitosis, the less hydrophobic H3 to 19% and the more hydrophobic H3 to 24% of control levels. These results suggest that sodium arsenite may inhibite cell proliferation by interfering with the cyclin B/p34{sup cdc2} histone kinase activity which is thought to play a key role in regulating the cell cycle. It has been proposed by our laboratory that HI and H3 phosphorylations play a role in restructuring interphase chromatin into metaphase chromosomes. Interference of this process by sodium arsenite may lead to structurally damaged chromosomes resulting in the increased cancer risks known to be produced by arsenic exposure from the environment.

  9. Comparison of staining of mitotic figures by haematoxylin and eosin-and crystal violet stains, in oral epithelial dysplasia and squamous cell carcinoma

    OpenAIRE

    Ankle Madhuri; Kale Alka; Charantimath Seema

    2007-01-01

    Mitosis of cells gives rise to tissue integrity. Defects during mitosis bring about abnormalities. Excessive proliferation of cells due to increased mitosis is one such outcome, which is the hallmark in precancer and cancer. The localization of proliferating cells or their precursors may not be obvious and easy. Establishing an easy way to distinguish these mitotic cells will help in grading and understanding their biological potential. Although immunohistochemistry is an advanced method in u...

  10. INFLUENCE OF SODIUM METABISULPHITE (E 223) ON MITOTIC DIVISION IN CALENDULA OFFICINALIS L.s

    OpenAIRE

    Romeo-Cristian Marc; Gabriela Capraru

    2008-01-01

    This paper presents the cytogenetic effects induced by sodium metabisulphite (E 223) (a food additive used as preservative) in meristematic cells of Calendula officinalis L. root tips. The treatment has determined the lessening of the mitotic index (comparative with the control variant), until mitotic division total inhibition, as well as a growth frequency of division aberration in anaphase and telophase.

  11. Genetic variation in mitotic regulatory pathway genes is associated with breast tumor grade

    DEFF Research Database (Denmark)

    Purrington, Kristen S; Slettedahl, Seth; Bolla, Manjeet K;

    2014-01-01

    Mitotic index is an important component of histologic grade and has an etiologic role in breast tumorigenesis. Several small candidate gene studies have reported associations between variation in mitotic genes and breast cancer risk. We measured associations between 2156 single nucleotide polymor...

  12. Par1b induces asymmetric inheritance of plasma membrane domains via LGN-dependent mitotic spindle orientation in proliferating hepatocytes

    NARCIS (Netherlands)

    Slim, Christiaan L; Lázaro-Diéguez, Francisco; Bijlard, Marjolein; Toussaint, Mathilda J M; de Bruin, Alain; Du, Quansheng; Müsch, Anne; van Ijzendoorn, Sven C D

    2013-01-01

    The development and maintenance of polarized epithelial tissue requires a tightly controlled orientation of mitotic cell division relative to the apical polarity axis. Hepatocytes display a unique polarized architecture. We demonstrate that mitotic hepatocytes asymmetrically segregate their apical p

  13. Cytoplasmic flows as signatures for the mechanics of mitotic positioning

    CERN Document Server

    Nazockdast, Ehssan; Needleman, Daniel; Shelley, Michael

    2015-01-01

    The proper positioning of the mitotic spindle is crucial for asymmetric cell division and generating cell diversity during development. Proper position in the single-cell embryo of Caenorhabditis elegans is achieved initially by the migration and rotation of the pronuclear complex (PNC) and its two associated centrosomal arrays of microtubules (MTs). We present here the first systematic theoretical study of how these $O(1000)$ centrosomal microtubules (MTs) interact through the immersing cytoplasm, the cell periphery and PNC, and with each other, to achieve proper position. This study is made possible through our development of a highly efficient and parallelized computational framework that accounts explicitly for long-ranged hydrodynamic interactions (HIs) between the MTs, while also capturing their flexibility, dynamic instability, and interactions with molecular motors and boundaries. First, we show through direct simulation that previous estimates of the PNC drag coefficient, based on either ignoring or ...

  14. Aurora A's functions during mitotic exit: the Guess Who game

    Directory of Open Access Journals (Sweden)

    David eReboutier

    2015-12-01

    Full Text Available Until recently, the knowledge of Aurora A kinase functions during mitosis was limited to pre-metaphase events, particularly centrosome maturation, G2/M transition, and mitotic spindle assembly. However, an involvement of Aurora A in post-metaphase events was also suspected, but not clearly demonstrated due to the technical difficulty to perform the appropriate experiments. Recent developments of both an analog specific version of Aurora A, and of small molecule inhibitors have led to the first demonstration that Aurora A is required for the early steps of cytokinesis. As in pre-metaphase, Aurora A plays diverse functions during anaphase, essentially participating in astral microtubules dynamics and central spindle assembly and functioning. The present review describes the experimental systems used to decipher new functions of Aurora A during late mitosis and situate these functions into the context of cytokinesis mechanisms.

  15. Aurora A's Functions During Mitotic Exit: The Guess Who Game.

    Science.gov (United States)

    Reboutier, David; Benaud, Christelle; Prigent, Claude

    2015-01-01

    Until recently, the knowledge of Aurora A kinase functions during mitosis was limited to pre-metaphase events, particularly centrosome maturation, G2/M transition, and mitotic spindle assembly. However, an involvement of Aurora A in post-metaphase events was also suspected, but not clearly demonstrated due to the technical difficulty to perform the appropriate experiments. Recent developments of both an analog-specific version of Aurora A and small molecule inhibitors have led to the first demonstration that Aurora A is required for the early steps of cytokinesis. As in pre-metaphase, Aurora A plays diverse functions during anaphase, essentially participating in astral microtubules dynamics and central spindle assembly and functioning. The present review describes the experimental systems used to decipher new functions of Aurora A during late mitosis and situate these functions into the context of cytokinesis mechanisms. PMID:26734572

  16. In-silico modeling of the mitotic spindle assembly checkpoint.

    Directory of Open Access Journals (Sweden)

    Bashar Ibrahim

    Full Text Available BACKGROUND: The Mitotic Spindle Assembly Checkpoint ((MSAC is an evolutionary conserved mechanism that ensures the correct segregation of chromosomes by restraining cell cycle progression from entering anaphase until all chromosomes have made proper bipolar attachments to the mitotic spindle. Its malfunction can lead to cancer. PRINCIPLE FINDINGS: We have constructed and validated for the human (MSAC mechanism an in silico dynamical model, integrating 11 proteins and complexes. The model incorporates the perspectives of three central control pathways, namely Mad1/Mad2 induced Cdc20 sequestering based on the Template Model, MCC formation, and APC inhibition. Originating from the biochemical reactions for the underlying molecular processes, non-linear ordinary differential equations for the concentrations of 11 proteins and complexes of the (MSAC are derived. Most of the kinetic constants are taken from literature, the remaining four unknown parameters are derived by an evolutionary optimization procedure for an objective function describing the dynamics of the APC:Cdc20 complex. MCC:APC dissociation is described by two alternatives, namely the "Dissociation" and the "Convey" model variants. The attachment of the kinetochore to microtubuli is simulated by a switching parameter silencing those reactions which are stopped by the attachment. For both, the Dissociation and the Convey variants, we compare two different scenarios concerning the microtubule attachment dependent control of the dissociation reaction. Our model is validated by simulation of ten perturbation experiments. CONCLUSION: Only in the controlled case, our models show (MSAC behaviour at meta- to anaphase transition in agreement with experimental observations. Our simulations revealed that for (MSAC activation, Cdc20 is not fully sequestered; instead APC is inhibited by MCC binding.

  17. Orc1 Binding to Mitotic Chromosomes Precedes Spatial Patterning during G1 Phase and Assembly of the Origin Recognition Complex in Human Cells.

    Science.gov (United States)

    Kara, Nihan; Hossain, Manzar; Prasanth, Supriya G; Stillman, Bruce

    2015-05-01

    Replication of eukaryotic chromosomes occurs once every cell division cycle in normal cells and is a tightly controlled process that ensures complete genome duplication. The origin recognition complex (ORC) plays a key role during the initiation of DNA replication. In human cells, the level of Orc1, the largest subunit of ORC, is regulated during the cell division cycle, and thus ORC is a dynamic complex. Upon S phase entry, Orc1 is ubiquitinated and targeted for destruction, with subsequent dissociation of ORC from chromosomes. Time lapse and live cell images of human cells expressing fluorescently tagged Orc1 show that Orc1 re-localizes to condensing chromatin during early mitosis and then displays different nuclear localization patterns at different times during G1 phase, remaining associated with late replicating regions of the genome in late G1 phase. The initial binding of Orc1 to mitotic chromosomes requires C-terminal amino acid sequences that are similar to mitotic chromosome-binding sequences in the transcriptional pioneer protein FOXA1. Depletion of Orc1 causes concomitant loss of the mini-chromosome maintenance (Mcm2-7) helicase proteins on chromatin. The data suggest that Orc1 acts as a nucleating center for ORC assembly and then pre-replication complex assembly by binding to mitotic chromosomes, followed by gradual removal from chromatin during the G1 phase. PMID:25784553

  18. Meiotic and Mitotic Cell Cycle Mutants Involved in Gametophyte Development in Arabidopsis

    Institute of Scientific and Technical Information of China (English)

    Jingjing Liu; Li-Jia Qu

    2008-01-01

    The alternation between diploid and haploid generations is fundamentalin the life cycles of both animals and plants.The meiotic cell cycle is common to both animals and plants gamete formation, but in animals the products of meiosis are gametes,whereas for most plants,subsequent mitotic cell cycles are needed for their formation. Clarifying the regulatory mechanisms of mitotic cell cycle progression during gametophyte development will help understanding of sexual reproduction in plants.Many mutants defective in gametophyte development and,in particular,many meiotic and mitotic cell cycle mutants in Arabidopsis male and female gametophyte development were identified through both forward and reverse genetics approaches.

  19. The essential mitotic peptidyl–prolyl isomerase Pin1 binds and regulates mitosis-specific phosphoproteins

    OpenAIRE

    Shen, Minhui; Stukenberg, P. Todd; Kirschner, Marc W; Lu, Kun Ping

    1998-01-01

    Phosphorylation of mitotic proteins on the Ser/Thr-Pro motifs has been shown to play an important role in regulating mitotic progression. Pin1 is a novel essential peptidyl–prolyl isomerase (PPIase) that inhibits entry into mitosis and is also required for proper progression through mitosis, but its substrate(s) and function(s) remain to be determined. Here we report that in both human cells and Xenopus extracts, Pin1 interacts directly with a subset of mitotic phosphoproteins on phosphorylat...

  20. The CRO-1 gene of Saccharomyces cerevisiae controls mitotic crossing over, chromosomal stability and sporulation

    International Nuclear Information System (INIS)

    The properties of a novel temperature-sensitive recombination-defective mutant of Saccharomyces cerevisiae, cro1-1 is described. The cro1-1 mutant is the first instance of a rec mutation that reduces drastically the rates of spontaneous mitotic crossing-over events but not those of gene conversional events. The cro1-1 mutation thus provides evidence that mitotic crossing-over is dependent upon gene products that are not essential for gene conversional events. The cro1-1 mutation also results in enhanced mitotic-chromosomal instability and MATa/MATα cro1-1/cro1-1 mutants are sporulation deficient. These phenotypes indicate that the CRO1 gene modulates mitotic chromosomal integrity and is essential for normal meiosis. The cro1-1 mutant possesses Holliday junction resolvase activity, hence its recombinational defect does not involve failure to execute this putative final recombinational step. 7 refs., 1 fig., 5 tabs

  1. Relationship of cyclic AMP levels to duration of radiation-induced mitotic delay

    International Nuclear Information System (INIS)

    A study has been made of the effect of several phosphodiesterase inhibitors (caffeine, theophylline, methyl-isobutyl xanthine (MIX), diethyl-amino-1-reserpine (DL-152) and 4-(-3-butoxy-4-methoxybenzyl-2-imidazolidinone) (Ro-20-1742)) on the duration of X-ray induced mitotic delay and on cyclic AMP concentration in replicate monolayer cultures of CHO cells. Only caffeine (1mM) of the methyl xanthines protected the cells against radiation-induced mitotic delay, without changing the cyclic AMP level. Elevated AMP levels and mitotic delay were demonstrated for unirradiated cells treated with MIX, Ro-20-1724 and DL-152, although the timing and extent of the delays varied for the different compounds. There was synergism between Ro-20-1724 and irradiation for lengthening the period of mitotic delay. (U.K.)

  2. Effects of 5-fluorouracil on the mitotic activity of onion root tips apical meristem

    Directory of Open Access Journals (Sweden)

    Waldemar Lechowicz

    2015-05-01

    Full Text Available The effects of various concentrations of 5-FU on the mitotic activity of onion root tips apical meristem were investigated during 24-hour incubation in 5-FU and postincubation in water. The incubation in 5-FU caused a reversible inhibition of mitotic activity, and waves of the partially synchronised mitoses were observed during the period of postincubation. The most pronounced synchronisation of mitoses was obtained after incubation in 100 mg/l. 5-FU but the mitotic index of the resumed mitotic activity amounted to only one half of the control value. 5-FU was found to cause some cytological changes in meristematic cells such as enlargement of the nucleoli, change in the interphasic nuclei structure, appearance of subchromatid and chromatid aberrations and micronuclei. The effects of 5-FU on nucleic acids and the cell division cycle ace discussed and compared with the effects of 5-FUdR.

  3. Phosphorylation and disassembly of intermediate filaments in mitotic cells

    International Nuclear Information System (INIS)

    As baby hamster kidney (BHK-21) cells enter mitosis, networks of intermediate filaments (IFs) are transformed into cytoplasmic aggregates of protofilaments. Coincident with this morphological change, the phosphate content of vimentin increases from 0.3 mol of Pi per mol of protein in interphase to 1.9 mol of Pi per mol of protein in mitosis. A similar increase in phosphate content is observed with desmin, from 0.5 mol of Pi per mol of protein to 1.5 mol of Pi per mol of protein. Fractionation of mitotic cell lysates by hydroxylapatite column chromatography reveals the presence of two IF protein kinase activities, designated as IF protein kinase I and IF protein kinase II. Comparison of two-dimensional 32P-labeled phosphopeptide maps of vimentin and desmin phosphorylated in vivo in mitosis, and in vitro using partially purified kinase fractions, reveals extensive similarity in the two sets of phosphorylation sites. Phosphorylation of in vitro polymerized IFs by IF protein kinase II induces complete disassembly as determined by negative-stain electron microscopy. The results support the idea that the disassembly of IFs in mitosis is regulated by the phosphorylation of its subunit proteins

  4. Phosphorylation and disassembly of intermediate filaments in mitotic cells

    Energy Technology Data Exchange (ETDEWEB)

    Chou, Yinghao; Rosevear, E.; Goldman, R.D. (Northwestern Univ. Medical School, Chicago, IL (USA))

    1989-03-01

    As baby hamster kidney (BHK-21) cells enter mitosis, networks of intermediate filaments (IFs) are transformed into cytoplasmic aggregates of protofilaments. Coincident with this morphological change, the phosphate content of vimentin increases from 0.3 mol of P{sub i} per mol of protein in interphase to 1.9 mol of P{sub i} per mol of protein in mitosis. A similar increase in phosphate content is observed with desmin, from 0.5 mol of P{sub i} per mol of protein to 1.5 mol of P{sub i} per mol of protein. Fractionation of mitotic cell lysates by hydroxylapatite column chromatography reveals the presence of two IF protein kinase activities, designated as IF protein kinase I and IF protein kinase II. Comparison of two-dimensional {sup 32}P-labeled phosphopeptide maps of vimentin and desmin phosphorylated in vivo in mitosis, and in vitro using partially purified kinase fractions, reveals extensive similarity in the two sets of phosphorylation sites. Phosphorylation of in vitro polymerized IFs by IF protein kinase II induces complete disassembly as determined by negative-stain electron microscopy. The results support the idea that the disassembly of IFs in mitosis is regulated by the phosphorylation of its subunit proteins.

  5. Maintaining Genome Stability in Defiance of Mitotic DNA Damage

    Science.gov (United States)

    Ferrari, Stefano; Gentili, Christian

    2016-01-01

    The implementation of decisions affecting cell viability and proliferation is based on prompt detection of the issue to be addressed, formulation and transmission of a correct set of instructions and fidelity in the execution of orders. While the first and the last are purely mechanical processes relying on the faithful functioning of single proteins or macromolecular complexes (sensors and effectors), information is the real cue, with signal amplitude, duration, and frequency ultimately determining the type of response. The cellular response to DNA damage is no exception to the rule. In this review article we focus on DNA damage responses in G2 and Mitosis. First, we set the stage describing mitosis and the machineries in charge of assembling the apparatus responsible for chromosome alignment and segregation as well as the inputs that control its function (checkpoints). Next, we examine the type of issues that a cell approaching mitosis might face, presenting the impact of post-translational modifications (PTMs) on the correct and timely functioning of pathways correcting errors or damage before chromosome segregation. We conclude this essay with a perspective on the current status of mitotic signaling pathway inhibitors and their potential use in cancer therapy. PMID:27493659

  6. Centromeric barrier disruption leads to mitotic defects in Schizosaccharomyces pombe.

    Science.gov (United States)

    Gaither, Terilyn L; Merrett, Stephanie L; Pun, Matthew J; Scott, Kristin C

    2014-04-01

    Centromeres are cis-acting chromosomal domains that direct kinetochore formation, enabling faithful chromosome segregation and preserving genome stability. The centromeres of most eukaryotic organisms are structurally complex, composed of nonoverlapping, structurally and functionally distinct chromatin subdomains, including the specialized core chromatin that underlies the kinetochore and pericentromeric heterochromatin. The genomic and epigenetic features that specify and preserve the adjacent chromatin subdomains critical to centromere identity are currently unknown. Here we demonstrate that chromatin barriers regulate this process in Schizosaccharomyces pombe. Reduced fitness and mitotic chromosome segregation defects occur in strains that carry exogenous DNA inserted at centromere 1 chromatin barriers. Abnormal phenotypes are accompanied by changes in the structural integrity of both the centromeric core chromatin domain, containing the conserved CENP-A(Cnp1) protein, and the flanking pericentric heterochromatin domain. Barrier mutant cells can revert to wild-type growth and centromere structure at a high frequency after the spontaneous excision of integrated exogenous DNA. Our results reveal a previously undemonstrated role for chromatin barriers in chromosome segregation and in the prevention of genome instability. PMID:24531725

  7. X-ray induction of mitotic and meiotic chromosome aberrations

    International Nuclear Information System (INIS)

    In 1964 six pairs of rat kangaroo (Potorous tridactylis) were obtained from Australia. The tissues of these animals were used to initiate cell lines. Since this species has a low chromosome number of six pairs, each pair with its own distinctive morphology, it is particularly favorable for cytogenetic research. In cell cultures derived from the corneal endothelial tissues of one animal there emerged a number of haploid cells. The number of haploid cells in the cultures reached as high as 20% of the total mitotic configurations. The in vitro diploid and haploid mixture cell cultures could be a resemblance or a coincidence to the mixture existence of the diploid primary spermatocytes and the haploid secondary spermatocytes (gametes) in the in vivo testicular tissues of the male animals. It would be interesting to compare reactions of the haploid and diploid cell mixture, either in the cultures or in the testes, to x-ray exposure. Two other studies involving x-ray effects on Chinese hamster oocyte maturation and meiotic chromosomes and the x-ray induction of Chinese hamster spermatocyte meiotic chromosome aberrations have been done in this laboratory. A review of these three studies involving diploid and haploid chromosomes may lead to further research in the x-ray induction of chromosome aberrations

  8. SAP-like domain in nucleolar spindle associated protein mediates mitotic chromosome loading as well as interphase chromatin interaction

    International Nuclear Information System (INIS)

    Highlights: → The SAP-like domain in NuSAP is a functional DNA-binding domain with preference for dsDNA. → This SAP-like domain is essential for chromosome loading during early mitosis. → NuSAP is highly dynamic on mitotic chromatin, as evident from photobleaching experiments. → The SAP-like domain also mediates NuSAP-chromatin interaction in interphase nucleoplasm. -- Abstract: Nucleolar spindle associated protein (NuSAP) is a microtubule-stabilizing protein that localizes to chromosome arms and chromosome-proximal microtubules during mitosis and to the nucleus, with enrichment in the nucleoli, during interphase. The critical function of NuSAP is underscored by the finding that its depletion in HeLa cells results in various mitotic defects. Moreover, NuSAP is found overexpressed in multiple cancers and its expression levels often correlate with the aggressiveness of cancer. Due to its localization on chromosome arms and combination of microtubule-stabilizing and DNA-binding properties, NuSAP takes a special place within the extensive group of spindle assembly factors. In this study, we identify a SAP-like domain that shows DNA binding in vitro with a preference for dsDNA. Deletion of the SAP-like domain abolishes chromosome arm binding of NuSAP during mitosis, but is not sufficient to abrogate its chromosome-proximal localization after anaphase onset. Fluorescence recovery after photobleaching experiments revealed the highly dynamic nature of this NuSAP-chromatin interaction during mitosis. In interphase cells, NuSAP also interacts with chromatin through its SAP-like domain, as evident from its enrichment on dense chromatin regions and intranuclear mobility, measured by fluorescence correlation spectroscopy. The obtained results are in agreement with a model where NuSAP dynamically stabilizes newly formed microtubules on mitotic chromosomes to enhance chromosome positioning without immobilizing these microtubules. Interphase NuSAP-chromatin interaction

  9. SAP-like domain in nucleolar spindle associated protein mediates mitotic chromosome loading as well as interphase chromatin interaction

    Energy Technology Data Exchange (ETDEWEB)

    Verbakel, Werner, E-mail: werner.verbakel@chem.kuleuven.be [Laboratory of Biomolecular Dynamics, Katholieke Universiteit Leuven, Celestijnenlaan 200G, Bus 2403, 3001 Heverlee (Belgium); Carmeliet, Geert, E-mail: geert.carmeliet@med.kuleuven.be [Laboratory of Experimental Medicine and Endocrinology, Katholieke Universiteit Leuven, Herestraat 49, Bus 902, 3000 Leuven (Belgium); Engelborghs, Yves, E-mail: yves.engelborghs@fys.kuleuven.be [Laboratory of Biomolecular Dynamics, Katholieke Universiteit Leuven, Celestijnenlaan 200G, Bus 2403, 3001 Heverlee (Belgium)

    2011-08-12

    Highlights: {yields} The SAP-like domain in NuSAP is a functional DNA-binding domain with preference for dsDNA. {yields} This SAP-like domain is essential for chromosome loading during early mitosis. {yields} NuSAP is highly dynamic on mitotic chromatin, as evident from photobleaching experiments. {yields} The SAP-like domain also mediates NuSAP-chromatin interaction in interphase nucleoplasm. -- Abstract: Nucleolar spindle associated protein (NuSAP) is a microtubule-stabilizing protein that localizes to chromosome arms and chromosome-proximal microtubules during mitosis and to the nucleus, with enrichment in the nucleoli, during interphase. The critical function of NuSAP is underscored by the finding that its depletion in HeLa cells results in various mitotic defects. Moreover, NuSAP is found overexpressed in multiple cancers and its expression levels often correlate with the aggressiveness of cancer. Due to its localization on chromosome arms and combination of microtubule-stabilizing and DNA-binding properties, NuSAP takes a special place within the extensive group of spindle assembly factors. In this study, we identify a SAP-like domain that shows DNA binding in vitro with a preference for dsDNA. Deletion of the SAP-like domain abolishes chromosome arm binding of NuSAP during mitosis, but is not sufficient to abrogate its chromosome-proximal localization after anaphase onset. Fluorescence recovery after photobleaching experiments revealed the highly dynamic nature of this NuSAP-chromatin interaction during mitosis. In interphase cells, NuSAP also interacts with chromatin through its SAP-like domain, as evident from its enrichment on dense chromatin regions and intranuclear mobility, measured by fluorescence correlation spectroscopy. The obtained results are in agreement with a model where NuSAP dynamically stabilizes newly formed microtubules on mitotic chromosomes to enhance chromosome positioning without immobilizing these microtubules. Interphase Nu

  10. A chemical tool box defines mitotic and interphase roles for Mps1 kinase

    OpenAIRE

    Lan, Weijie; Don W Cleveland

    2010-01-01

    In this issue, three groups (Hewitt et al. 2010. J. Cell Biol. doi:10.1083/jcb.201002133; Maciejowski et al. 2010. J. Cell Biol. doi:10.1083/jcb.201001050; Santaguida et al. 2010. J. Cell Biol. doi:10.1083/jcb.201001036) use chemical inhibitors to analyze the function of the mitotic checkpoint kinase Mps1. These studies demonstrate that Mps1 kinase activity ensures accurate chromosome segregation through its recruitment to kinetochores of mitotic checkpoint proteins, formation of interphase a...

  11. Stereospecific phosphorylation by the central mitotic kinase Cdk1-cyclin B.

    Science.gov (United States)

    Etzkorn, Felicia A; Zhao, Song

    2015-04-17

    The cis vs trans conformation, or shape, of phosphoserine-proline (pSer-Pro), a prevalent motif in cell cycle proteins, may play a significant role in regulating mitosis. We demonstrate that Cdk1-cyclin B, the central mitotic kinase, is specific for the trans conformation, not cis, of synthetic, locked Ser-Pro 11-residue peptide substrates, using LC-MSMS detection and sequencing of phosphorylated products. This substrate stereospecificity may contribute an additional level of mitotic regulation. PMID:25603287

  12. Identification of Drosophila mitotic genes by combining co-expression analysis and RNA interference.

    Directory of Open Access Journals (Sweden)

    Maria Patrizia Somma

    2008-07-01

    Full Text Available RNAi screens have, to date, identified many genes required for mitotic divisions of Drosophila tissue culture cells. However, the inventory of such genes remains incomplete. We have combined the powers of bioinformatics and RNAi technology to detect novel mitotic genes. We found that Drosophila genes involved in mitosis tend to be transcriptionally co-expressed. We thus constructed a co-expression-based list of 1,000 genes that are highly enriched in mitotic functions, and we performed RNAi for each of these genes. By limiting the number of genes to be examined, we were able to perform a very detailed phenotypic analysis of RNAi cells. We examined dsRNA-treated cells for possible abnormalities in both chromosome structure and spindle organization. This analysis allowed the identification of 142 mitotic genes, which were subdivided into 18 phenoclusters. Seventy of these genes have not previously been associated with mitotic defects; 30 of them are required for spindle assembly and/or chromosome segregation, and 40 are required to prevent spontaneous chromosome breakage. We note that the latter type of genes has never been detected in previous RNAi screens in any system. Finally, we found that RNAi against genes encoding kinetochore components or highly conserved splicing factors results in identical defects in chromosome segregation, highlighting an unanticipated role of splicing factors in centromere function. These findings indicate that our co-expression-based method for the detection of mitotic functions works remarkably well. We can foresee that elaboration of co-expression lists using genes in the same phenocluster will provide many candidate genes for small-scale RNAi screens aimed at completing the inventory of mitotic proteins.

  13. Evidence of Selection against Complex Mitotic-Origin Aneuploidy during Preimplantation Development.

    Directory of Open Access Journals (Sweden)

    Rajiv C McCoy

    2015-10-01

    Full Text Available Whole-chromosome imbalances affect over half of early human embryos and are the leading cause of pregnancy loss. While these errors frequently arise in oocyte meiosis, many such whole-chromosome abnormalities affecting cleavage-stage embryos are the result of chromosome missegregation occurring during the initial mitotic cell divisions. The first wave of zygotic genome activation at the 4-8 cell stage results in the arrest of a large proportion of embryos, the vast majority of which contain whole-chromosome abnormalities. Thus, the full spectrum of meiotic and mitotic errors can only be detected by sampling after the initial cell divisions, but prior to this selective filter. Here, we apply 24-chromosome preimplantation genetic screening (PGS to 28,052 single-cell day-3 blastomere biopsies and 18,387 multi-cell day-5 trophectoderm biopsies from 6,366 in vitro fertilization (IVF cycles. We precisely characterize the rates and patterns of whole-chromosome abnormalities at each developmental stage and distinguish errors of meiotic and mitotic origin without embryo disaggregation, based on informative chromosomal signatures. We show that mitotic errors frequently involve multiple chromosome losses that are not biased toward maternal or paternal homologs. This outcome is characteristic of spindle abnormalities and chaotic cell division detected in previous studies. In contrast to meiotic errors, our data also show that mitotic errors are not significantly associated with maternal age. PGS patients referred due to previous IVF failure had elevated rates of mitotic error, while patients referred due to recurrent pregnancy loss had elevated rates of meiotic error, controlling for maternal age. These results support the conclusion that mitotic error is the predominant mechanism contributing to pregnancy losses occurring prior to blastocyst formation. This high-resolution view of the full spectrum of whole-chromosome abnormalities affecting early embryos

  14. Arsenite-induced mitotic death involves stress response and is independent of tubulin polymerization

    International Nuclear Information System (INIS)

    Arsenite, a known mitotic disruptor, causes cell cycle arrest and cell death at anaphase. The mechanism causing mitotic arrest is highly disputed. We compared arsenite to the spindle poisons nocodazole and paclitaxel. Immunofluorescence analysis of α-tubulin in interphase cells demonstrated that, while nocodazole and paclitaxel disrupt microtubule polymerization through destabilization and hyperpolymerization, respectively, microtubules in arsenite-treated cells remain comparable to untreated cells even at supra-therapeutic concentrations. Immunofluorescence analysis of α-tubulin in mitotic cells showed spindle formation in arsenite- and paclitaxel-treated cells but not in nocodazole-treated cells. Spindle formation in arsenite-treated cells appeared irregular and multi-polar. γ-tubulin staining showed that cells treated with nocodazole and therapeutic concentrations of paclitaxel contained two centrosomes. In contrast, most arsenite-treated mitotic cells contained more than two centrosomes, similar to centrosome abnormalities induced by heat shock. Of the three drugs tested, only arsenite treatment increased expression of the inducible isoform of heat shock protein 70 (HSP70i). HSP70 and HSP90 proteins are intimately involved in centrosome regulation and mitotic spindle formation. HSP90 inhibitor 17-DMAG sensitized cells to arsenite treatment and increased arsenite-induced centrosome abnormalities. Combined treatment of 17-DMAG and arsenite resulted in a supra-additive effect on viability, mitotic arrest, and centrosome abnormalities. Thus, arsenite-induced abnormal centrosome amplification and subsequent mitotic arrest is independent of effects on tubulin polymerization and may be due to specific stresses that are protected against by HSP90 and HSP70

  15. PIASy Mediates SUMO-2/3 Conjugation of Poly(ADP-ribose) Polymerase 1 (PARP1) on Mitotic Chromosomes*

    OpenAIRE

    Ryu, Hyunju; Al-Ani, Gada; Deckert, Katelyn; Kirkpatrick, Donald; Gygi, Steven P.; Dasso, Mary; Azuma, Yoshiaki

    2010-01-01

    PIASy is a small ubiquitin-related modifier (SUMO) ligase that modifies chromosomal proteins in mitotic Xenopus egg extracts and plays an essential role in mitotic chromosome segregation. We have isolated a novel SUMO-2/3-modified mitotic chromosomal protein and identified it as poly(ADP-ribose) polymerase 1 (PARP1). PARP1 was robustly conjugated to SUMO-2/3 on mitotic chromosomes but not on interphase chromatin. PIASy promotes SUMOylation of PARP1 both in egg extracts and in vitro reconstitu...

  16. The spindle protein CHICA mediates localization of the chromokinesin Kid to the mitotic spindle

    NARCIS (Netherlands)

    Santamaria, Anna; Nagel, Susanna; Sillje, Herman H W; Nigg, Erich A

    2008-01-01

    Microtubule-based motor proteins provide essential forces for bipolar organization of spindle microtubules and chromosome movement, prerequisites of chromosome segregation during the cell cycle. Here, we describe the functional characterization of a novel spindle protein, termed "CHICA," that was or

  17. Localization of gamma-tubulin in interphase and mitotic cells of a unicellular eukaryote, Giardia intestinalis

    Czech Academy of Sciences Publication Activity Database

    Nohýnková, E.; Dráber, Pavel; Reisching, J.; Kulda, J.

    2000-01-01

    Roč. 79, č. 6 (2000), s. 438-445. ISSN 0171-9335 R&D Projects: GA MZd IZ3190; GA MŠk VS96142; GA ČR GA204/98/1054 Institutional research plan: CEZ:AV0Z5052915 Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.801, year: 2000

  18. The effect of magnesium on mitotic spindle formation in Schizosaccharomyces pombe.

    Science.gov (United States)

    Uz, Gulsen; Sarikaya, Aysegul Topal

    2016-01-01

    Magnesium (Mg2+), an essential ion for cells and biological systems, is involved in a variety of cellular processes, including the formation and breakdown of microtubules. The results of a previous investigation suggested that as cells grow the intracellular Mg2+ concentration falls, thereby stimulating formation of the mitotic spindle. In the present work, we used a Mg2+-deficient Schizosaccharomyces pombe strain GA2, in which two essential membrane Mg2+ transporter genes (homologs of ALR1 and ALR2 in Saccharomyces cerevisae) were deleted, and its parental strain Sp292, to examine the extent to which low Mg2+ concentrations can affect mitotic spindle formation. The two S. pombe strains were transformed with a plasmid carrying a GFP-α2-tubulin construct to fluorescently label microtubules. Using the free Mg2+-specific fluorescent probe mag-fura-2, we confirmed that intracellular free Mg2+ levels were lower in GA2 than in the parental strain. Defects in interphase microtubule organization, a lower percentage of mitotic spindle formation and a reduced mitotic index were also observed in the GA2 strain. Although there was interphase microtubule polymerization, the lower level of mitotic spindle formation in the Mg2+-deficient strain suggested a greater requirement for Mg2+ in this phenomenon than previously thought. PMID:27560651

  19. PP1 initiates the dephosphorylation of MASTL, triggering mitotic exit and bistability in human cells

    Science.gov (United States)

    Rogers, Samuel; Fey, Dirk; McCloy, Rachael A.; Parker, Benjamin L.; Mitchell, Nicholas J.; Payne, Richard J.; Daly, Roger J.; James, David E.; Caldon, C. Elizabeth; Watkins, D. Neil; Croucher, David R.; Burgess, Andrew

    2016-01-01

    ABSTRACT Entry into mitosis is driven by the phosphorylation of thousands of substrates, under the master control of Cdk1. During entry into mitosis, Cdk1, in collaboration with MASTL kinase, represses the activity of the major mitotic protein phosphatases, PP1 and PP2A, thereby ensuring mitotic substrates remain phosphorylated. For cells to complete and exit mitosis, these phosphorylation events must be removed, and hence, phosphatase activity must be reactivated. This reactivation of phosphatase activity presumably requires the inhibition of MASTL; however, it is not currently understood what deactivates MASTL and how this is achieved. In this study, we identified that PP1 is associated with, and capable of partially dephosphorylating and deactivating, MASTL during mitotic exit. Using mathematical modelling, we were able to confirm that deactivation of MASTL is essential for mitotic exit. Furthermore, small decreases in Cdk1 activity during metaphase are sufficient to initiate the reactivation of PP1, which in turn partially deactivates MASTL to release inhibition of PP2A and, hence, create a feedback loop. This feedback loop drives complete deactivation of MASTL, ensuring a strong switch-like activation of phosphatase activity during mitotic exit. PMID:26872783

  20. A mitotic kinase scaffold depleted in testicular seminomas impacts spindle orientation in germ line stem cells.

    Science.gov (United States)

    Hehnly, Heidi; Canton, David; Bucko, Paula; Langeberg, Lorene K; Ogier, Leah; Gelman, Irwin; Santana, L Fernando; Wordeman, Linda; Scott, John D

    2015-01-01

    Correct orientation of the mitotic spindle in stem cells underlies organogenesis. Spindle abnormalities correlate with cancer progression in germ line-derived tumors. We discover a macromolecular complex between the scaffolding protein Gravin/AKAP12 and the mitotic kinases, Aurora A and Plk1, that is down regulated in human seminoma. Depletion of Gravin correlates with an increased mitotic index and disorganization of seminiferous tubules. Biochemical, super-resolution imaging, and enzymology approaches establish that this Gravin scaffold accumulates at the mother spindle pole during metaphase. Manipulating elements of the Gravin-Aurora A-Plk1 axis prompts mitotic delay and prevents appropriate assembly of astral microtubules to promote spindle misorientation. These pathological responses are conserved in seminiferous tubules from Gravin(-/-) mice where an overabundance of Oct3/4 positive germ line stem cells displays randomized orientation of mitotic spindles. Thus, we propose that Gravin-mediated recruitment of Aurora A and Plk1 to the mother (oldest) spindle pole contributes to the fidelity of symmetric cell division. PMID:26406118

  1. Heat shock protein inhibitors, 17-DMAG and KNK437, enhance arsenic trioxide-induced mitotic apoptosis

    International Nuclear Information System (INIS)

    Arsenic trioxide (ATO) has recently emerged as a promising therapeutic agent in leukemia because of its ability to induce apoptosis. However, there is no sufficient evidence to support its therapeutic use for other types of cancers. In this study, we investigated if, and how, 17-dimethylaminoethylamino-17-demethoxy-geldanamycin (17-DMAG), an antagonist of heat shock protein 90 (HSP90), and KNK437, a HSP synthesis inhibitor, potentiated the cytotoxic effect of ATO. Our results showed that cotreatment with ATO and either 17-DMAG or KNK437 significantly increased ATO-induced cell death and apoptosis. siRNA-mediated attenuation of the expression of the inducible isoform of HSP70 (HSP70i) or HSP90α/β also enhanced ATO-induced apoptosis. In addition, cotreatment with ATO and 17-DMAG or KNK437 significantly increased ATO-induced mitotic arrest and ATO-induced BUBR1 phosphorylation and PDS1 accumulation. Cotreatment also significantly increased the percentage of mitotic cells with abnormal mitotic spindles and promoted metaphase arrest as compared to ATO treatment alone. These results indicated that 17-DMAG or KNK437 may enhance ATO cytotoxicity by potentiating mitotic arrest and mitotic apoptosis possibly through increased activation of the spindle checkpoint.

  2. High frequency induction of mitotic recombination by ionizing radiation in Mlh1 null mouse cells

    International Nuclear Information System (INIS)

    Mitotic recombination in somatic cells involves crossover events between homologous autosomal chromosomes. This process can convert a cell with a heterozygous deficiency to one with a homozygous deficiency if a mutant allele is present on one of the two homologous autosomes. Thus mitotic recombination often represents the second mutational step in tumor suppressor gene inactivation. In this study we examined the frequency and spectrum of ionizing radiation (IR)-induced autosomal mutations affecting Aprt expression in a mouse kidney cell line null for the Mlh1 mismatch repair (MMR) gene. The mutant frequency results demonstrated high frequency induction of mutations by IR exposure and the spectral analysis revealed that most of this response was due to the induction of mitotic recombinational events. High frequency induction of mitotic recombination was not observed in a DNA repair-proficient cell line or in a cell line with an MMR-independent mutator phenotype. These results demonstrate that IR exposure can initiate a process leading to mitotic recombinational events and that MMR function suppresses these events from occurring

  3. Association of Mitotic Regulation Pathway Polymorphisms with Pancreatic Cancer Risk and Outcome

    Science.gov (United States)

    Couch, Fergus J.; Wang, Xianshu; Bamlet, William R.; de Andrade, Mariza; Petersen, Gloria M.; McWilliams, Robert R.

    2009-01-01

    Background Mitosis is a highly regulated process that serves to ensure the fidelity of cell division. Disruption of mitotic regulators leading to aneuploidy and polyploidy is commonly observed in cancer cells. Single nucleotide polymorphisms (SNPs) in regulators of mitosis may promote chromosome mis-segregation and influence pancreatic cancer and/or survival. Methods Thirty four SNPs, previously associated with breast cancer risk, from 33 genes involved in regulation of mitosis, were investigated for associations with pancreatic cancer risk in 1,143 Caucasian patients with pancreatic adenocarcinoma and 1,097 unaffected controls from the Mayo Clinic. Associations with survival from pancreatic cancer were also assessed using 1,030 pancreatic cancer cases with known outcome. Results Two SNPs in the APC (rs2431238) and NIN (rs10145182) loci, out of 34 examined, were significantly associated with pancreatic cancer risk (p=0.035 and p=0.038, respectively). Further analyses of individuals categorized by smoking and BMI identified several SNPs displaying significant associations (p<0.05) with pancreatic cancer risk, including APC rs2431238 in individuals with high body mass index (BMI≥30) (p=0.031) and NIN rs10145182 in ever smokers (p=0.01). In addition, survival analyses detected significant associations between SNPs in EIF3S10 and overall survival (p=0.009), SNPs from five genes and survival in resected cancer cases (p<0.05), and SNPs from two other genes (p<0.05) and survival of locally advanced cancer cases. Conclusion Common variation in genes encoding regulators of mitosis may independently influence pancreatic cancer susceptibility and survival. PMID:20056645

  4. The Effect Of PHA And SEA On Mitotic Index Of Lymphocyte Cell Of Macaca Fasciulare

    International Nuclear Information System (INIS)

    The observation of influences of PHA (phytohemagglutinin) and SEA (staphilucoccal enterotoxin A) on mitotic index of lymphocyte of Macaca Fascicularis had been done. Half milliliters of lymphocyte cells stimulated with PHA or SEA were cultured in 10 ml RPMI + 1.0 ml Fetal Bouvine Serum (FBS ) + 0.1 ml L-glutamine + 0.15 ml PHA or 0.1 ml SEA ( 0.5 μg/ml ) + 0.1 ml Colchisin on 37 degree C for 96 hours. The result demonstrated that the frequency of mitotic index stimulated with PHA was higher than that of SEA. The average of mitotic index with PHA was 18.56 %, and with SEA was 8.3 %. (author)

  5. Semaphorin-Plexin Signaling Controls Mitotic Spindle Orientation during Epithelial Morphogenesis and Repair

    DEFF Research Database (Denmark)

    Xia, Jingjing; Swiercz, Jakub M.; Bañón-Rodríguez, Inmaculada;

    2015-01-01

    Morphogenesis, homeostasis, and regeneration of epithelial tissues rely on the accurate orientation of cell divisions, which is specified by the mitotic spindle axis. To remain in the epithelial plane, symmetrically dividing epithelial cells align their mitotic spindle axis with the plane. Here, we...... show that this alignment depends on epithelial cell-cell communication via semaphorin-plexin signaling. During kidney morphogenesis and repair, renal tubular epithelial cells lacking the transmembrane receptor Plexin-B2 or its semaphorin ligands fail to correctly orient the mitotic spindle, leading to...... severe defects in epithelial architecture and function. Analyses of a series of transgenic and knockout mice indicate that Plexin-B2 controls the cell division axis by signaling through its GTPase-activating protein (GAP) domain and Cdc42. Our data uncover semaphorin-plexin signaling as a central...

  6. Mitotic cells contract actomyosin cortex and generate pressure to round against or escape epithelial confinement

    Science.gov (United States)

    Sorce, Barbara; Escobedo, Carlos; Toyoda, Yusuke; Stewart, Martin P.; Cattin, Cedric J.; Newton, Richard; Banerjee, Indranil; Stettler, Alexander; Roska, Botond; Eaton, Suzanne; Hyman, Anthony A.; Hierlemann, Andreas; Müller, Daniel J.

    2015-11-01

    Little is known about how mitotic cells round against epithelial confinement. Here, we engineer micropillar arrays that subject cells to lateral mechanical confinement similar to that experienced in epithelia. If generating sufficient force to deform the pillars, rounding epithelial (MDCK) cells can create space to divide. However, if mitotic cells cannot create sufficient space, their rounding force, which is generated by actomyosin contraction and hydrostatic pressure, pushes the cell out of confinement. After conducting mitosis in an unperturbed manner, both daughter cells return to the confinement of the pillars. Cells that cannot round against nor escape confinement cannot orient their mitotic spindles and more likely undergo apoptosis. The results highlight how spatially constrained epithelial cells prepare for mitosis: either they are strong enough to round up or they must escape. The ability to escape from confinement and reintegrate after mitosis appears to be a basic property of epithelial cells.

  7. Bistability of mitotic entry and exit switches during open mitosis in mammalian cells.

    Science.gov (United States)

    Hégarat, Nadia; Rata, Scott; Hochegger, Helfrid

    2016-07-01

    Mitotic entry and exit are switch-like transitions that are driven by the activation and inactivation of Cdk1 and mitotic cyclins. This simple on/off reaction turns out to be a complex interplay of various reversible reactions, feedback loops, and thresholds that involve both the direct regulators of Cdk1 and its counteracting phosphatases. In this review, we summarize the interplay of the major components of the system and discuss how they work together to generate robustness, bistability, and irreversibility. We propose that it may be beneficial to regard the entry and exit reactions as two separate reversible switches that are distinguished by differences in the state of phosphatase activity, mitotic proteolysis, and a dramatic rearrangement of cellular components after nuclear envelope breakdown, and discuss how the major Cdk1 activity thresholds could be determined for these transitions. PMID:27231150

  8. Mitotic recombination induced by chemical and physical agents in the yeast Saccharomyces cerevisiae

    International Nuclear Information System (INIS)

    The treatment of diploid cultures of yeast with ultraviolet light (uv), γ-rays, nitrous acid (na) and ethyl methane sulphonate (ems) results in increases in cell death, mitotic gene conversion and crossing-over. Acridine orange (ao) treatment, in contrast, was effective only in increasing the frequency of gene conversion. The individual mutagens were effective in the order uv>na>γ-rays>ao>ems. Prior treatment of yeast cultures in starvation medium produced a significant reduction in the yield of induced gene conversion. The results have been interpreted on the basis of a general model of mitotic gene conversion which involves the post-replication repair of induced lesions involving de novo DNA synthesis without genetic exchange. In contrast mitotic crossing-over appears to involve the action of a repair system independent from excision or post-replication repair which involves genetic exchange between homologous chromosomes

  9. Cell fate after mitotic arrest in different tumor cells is determined by the balance between slippage and apoptotic threshold

    Energy Technology Data Exchange (ETDEWEB)

    Galán-Malo, Patricia; Vela, Laura; Gonzalo, Oscar; Calvo-Sanjuán, Rubén; Gracia-Fleta, Lucía; Naval, Javier; Marzo, Isabel, E-mail: imarzo@unizar.es

    2012-02-01

    Microtubule poisons and other anti-mitotic drugs induce tumor death but the molecular events linking mitotic arrest to cell death are still not fully understood. We have analyzed cell fate after mitotic arrest produced by the microtubule-destabilizing drug vincristine in a panel of human tumor cell lines showing different response to vincristine. In Jurkat, RPMI 8226 and HeLa cells, apoptosis was triggered shortly after vincristine-induced mitotic arrest. However, A549 cells, which express a great amount of Bcl-x{sub L} and undetectable amounts of Bak, underwent mitotic slippage prior to cell death. However, when Bcl-x{sub L} gene was silenced in A549 cells, vincristine induced apoptosis during mitotic arrest. Another different behavior was found in MiaPaca2 cells, where vincristine caused death by mitotic catastrophe that switched to apoptosis when cyclin B1 degradation was prevented by proteasome inhibition. Overexpression of Bcl-x{sub L} or silencing Bax and Bak expression delayed the onset of apoptosis in Jurkat and RPMI 8226 cells, enabling mitotic slippage and endoreduplication. In HeLa cells, overexpression of Bcl-x{sub L} switched cell death from apoptosis to mitotic catastrophe. Mcl-1 offered limited protection to vincristine-induced cell death and Mcl-1 degradation was not essential for vincristine-induced death. All these results, taken together, indicate that the Bcl-x{sub L}/Bak ratio and the ability to degrade cyclin B1 determine cell fate after mitotic arrest in the different tumor cell types. Highlights: ► Vincristine induces cell death by apoptosis or mitotic catastrophe. ► Apoptosis-proficient cells die by apoptosis during mitosis upon vincristine treatment. ► p53wt apoptosis-deficient cells undergo apoptosis from a G1-like tetraploid state. ► p53mt apoptosis-deficient cells can survive and divide giving rise to 8N cells.

  10. BRIT1/MCPH1 is essential for mitotic and meiotic recombination DNA repair and maintaining genomic stability in mice.

    Directory of Open Access Journals (Sweden)

    Yulong Liang

    2010-01-01

    Full Text Available BRIT1 protein (also known as MCPH1 contains 3 BRCT domains which are conserved in BRCA1, BRCA2, and other important molecules involved in DNA damage signaling, DNA repair, and tumor suppression. BRIT1 mutations or aberrant expression are found in primary microcephaly patients as well as in cancer patients. Recent in vitro studies suggest that BRIT1/MCPH1 functions as a novel key regulator in the DNA damage response pathways. To investigate its physiological role and dissect the underlying mechanisms, we generated BRIT1(-/- mice and identified its essential roles in mitotic and meiotic recombination DNA repair and in maintaining genomic stability. Both BRIT1(-/- mice and mouse embryonic fibroblasts (MEFs were hypersensitive to gamma-irradiation. BRIT1(-/- MEFs and T lymphocytes exhibited severe chromatid breaks and reduced RAD51 foci formation after irradiation. Notably, BRIT1(-/- mice were infertile and meiotic homologous recombination was impaired. BRIT1-deficient spermatocytes exhibited a failure of chromosomal synapsis, and meiosis was arrested at late zygotene of prophase I accompanied by apoptosis. In mutant spermatocytes, DNA double-strand breaks (DSBs were formed, but localization of RAD51 or BRCA2 to meiotic chromosomes was severely impaired. In addition, we found that BRIT1 could bind to RAD51/BRCA2 complexes and that, in the absence of BRIT1, recruitment of RAD51 and BRCA2 to chromatin was reduced while their protein levels were not altered, indicating that BRIT1 is involved in mediating recruitment of RAD51/BRCA2 to the damage site. Collectively, our BRIT1-null mouse model demonstrates that BRIT1 is essential for maintaining genomic stability in vivo to protect the hosts from both programmed and irradiation-induced DNA damages, and its depletion causes a failure in both mitotic and meiotic recombination DNA repair via impairing RAD51/BRCA2's function and as a result leads to infertility and genomic instability in mice.

  11. Mammalian neurogenesis requires Treacle-Plk1 for precise control of spindle orientation, mitotic progression, and maintenance of neural progenitor cells.

    Directory of Open Access Journals (Sweden)

    Daisuke Sakai

    Full Text Available The cerebral cortex is a specialized region of the brain that processes cognitive, motor, somatosensory, auditory, and visual functions. Its characteristic architecture and size is dependent upon the number of neurons generated during embryogenesis and has been postulated to be governed by symmetric versus asymmetric cell divisions, which mediate the balance between progenitor cell maintenance and neuron differentiation, respectively. The mechanistic importance of spindle orientation remains controversial, hence there is considerable interest in understanding how neural progenitor cell mitosis is controlled during neurogenesis. We discovered that Treacle, which is encoded by the Tcof1 gene, is a novel centrosome- and kinetochore-associated protein that is critical for spindle fidelity and mitotic progression. Tcof1/Treacle loss-of-function disrupts spindle orientation and cell cycle progression, which perturbs the maintenance, proliferation, and localization of neural progenitors during cortical neurogenesis. Consistent with this, Tcof1(+/- mice exhibit reduced brain size as a consequence of defects in neural progenitor maintenance. We determined that Treacle elicits its effect via a direct interaction with Polo-like kinase1 (Plk1, and furthermore we discovered novel in vivo roles for Plk1 in governing mitotic progression and spindle orientation in the developing mammalian cortex. Increased asymmetric cell division, however, did not promote increased neuronal differentiation. Collectively our research has therefore identified Treacle and Plk1 as novel in vivo regulators of spindle fidelity, mitotic progression, and proliferation in the maintenance and localization of neural progenitor cells. Together, Treacle and Plk1 are critically required for proper cortical neurogenesis, which has important implications in the regulation of mammalian brain size and the pathogenesis of congenital neurodevelopmental disorders such as microcephaly.

  12. High LET radiation enhances nocodazole induced cell death in HeLa cells through mitotic catastrophe and apoptosis

    International Nuclear Information System (INIS)

    To understand how human tumor cells respond to the combined treatment with nocodazole and high linear energy transfer (LET) radiation, alterations in cell cycle, mitotic disturbances and cell death were investigated in the present study. Human cervix carcinoma HeLa cells were exposed to nocodazole for 18 h immediately followed by high LET iron ion irradiation and displayed a sequence of events leading to DNA damages, mitotic aberrations, interphase restitution and endocycle as well as cell death. A prolonged mitotic arrest more than 10 h was observed following nocodazole exposure, no matter the irradiation was present or not. The occurrence of mitotic slippage following the mitotic arrest was only drug-dependent and the irradiation did not accelerate it. The amount of polyploidy cells was increased following mitotic slippage. No detectable G2 or G1 arrest was observed in cells upon the combined treatment and the cells reentered the cell cycle still harboring unrepaired cellular damages. This premature entry caused an increase of multipolar mitotic spindles and amplification of centrosomes, which gave rise to lagging chromosomal material, failure of cytokinesis and polyploidization. These mitotic disturbances and their outcomes confirmed the incidence of mitotic catastrophe and delayed apoptotic features displayed by terminal-transferased UTP- nick end-labeling (TUNEL) method after the combined treatment. These results suggest that the addition of high-LET iron ion irradiation to nocodazole enhanced mitotic catastrophe and delayed apoptosis in HeLa cells. These might be important cell death mechanisms involved in tumor cells in response to the treatment of antimitotic drug combined with high LET radiation. (author)

  13. Influence of sodium phosphate (E 339) on mitotic division in Calendula officinalis L.

    OpenAIRE

    Romeo-Cristian Marc; Gabriela Capraru

    2008-01-01

    This paper includes the cytogenetic effects induced by sodium phosphate (E 339) food additive in meristematic cells of Calendula officinalis L. root tips. The increase of food additive concentration determined the decrease of mitotic index, while the frequency and the type of chromosome aberrations are much greater in treated variants, comparatively with control.

  14. Frequencies of mutagen-induced coincident mitotic recombination at unlinked loci in Saccharomyces cerevisiae

    Energy Technology Data Exchange (ETDEWEB)

    Freeman, Kathryn M. [Department of Biology, College of the Holy Cross, One College Street, Worcester, MA 01610-2395 (United States); Hoffmann, George R. [Department of Biology, College of the Holy Cross, One College Street, Worcester, MA 01610-2395 (United States)]. E-mail: ghoffmann@holycross.edu

    2007-03-01

    Frequencies of coincident genetic events were measured in strain D7 of Saccharomyces cerevisiae. This diploid strain permits the detection of mitotic gene conversion involving the trp5-12 and trp5-27 alleles, mitotic crossing-over and gene conversion leading to the expression of the ade2-40 and ade2-119 alleles as red and pink colonies, and reversion of the ilv1-92 allele. The three genes are on different chromosomes, and one might expect that coincident (simultaneous) genetic alterations at two loci would occur at frequencies predicted by those of the single alterations acting as independent events. Contrary to this expectation, we observed that ade2 recombinants induced by bleomycin, {beta}-propiolactone, and ultraviolet radiation occur more frequently among trp5 convertants than among total colonies. This excess among trp5 recombinants indicates that double recombinants are more common than expected for independent events. No similar enrichment was found among Ilv{sup +} revertants. The possibility of an artifact in which haploid yeasts that mimic mitotic recombinants are generated by a low frequency of cryptic meiosis has been excluded. Several hypotheses that can explain the elevated incidence of coincident mitotic recombination have been evaluated, but the cause remains uncertain. Most evidence suggests that the excess is ascribable to a subset of the population being in a recombination-prone state.

  15. A selective stain for mitotic figures, particularly in the developing brain.

    Science.gov (United States)

    Fraser, F J

    1982-07-01

    A selective stain for mitotic figures is valuable where autoradiographic counting is not required, especially in the developing brain. Most work in this field has been based on conventional nuclear stains which do not differentiate mitotic figures from resting cells by color. Hematoxylin, Feulgen, gallocyanin and Nissl methods have been used particularly. The method described uses a modified Bouin fixative, followed by hydrolysis in 1 N HCl. Mitotic figures are selectively stained using crystal violet, with nuclear fast red as the counterstain for resting cells. The method has been tested using material from postnatal and fetal sheep, guinea pig and rat. Using paraffin mounted serial sections it is applicable to all organs. The method was very successful on developing rat brain, particularly for detail and quantitative estimation in the early stages of prenatal development, which was of primary interest. Nucleated cells of the erythrocytic series, keratin and what appear to be mast cells were found to stain. When nuclear counting or cell recognition were required these did not cause any difficulty, except in prenatal liver. The highly selective method presented stains mitotic figures, in all tissue tested, an intense blue against a background of red resting cells. PMID:6183796

  16. Hsp70 protects mitotic cells against heat-induced centrosome damage and division abnormalities

    NARCIS (Netherlands)

    Hut, HMJ; Kampinga, HH; Sibon, OCM

    2005-01-01

    The effect of heat shock on centrosomes has been mainly studied in interphase cells. Centrosomes play a key role in proper segregation of DNA during mitosis. However, the direct effect and consequences of heat shock on mitotic cells and a possible cellular defense system against proteotoxic stress d

  17. Involvement of CNOT3 in mitotic progression through inhibition of MAD1 expression

    Energy Technology Data Exchange (ETDEWEB)

    Takahashi, Akinori [Division of Oncology, Department of Cancer Biology, Institute of Medical Science, University of Tokyo, Minato-ku, Tokyo 108-8639 (Japan); Kikuguchi, Chisato [Cell Signal Unit, Okinawa Institute of Science and Technology, Kunigami, Okinawa 904-0412 (Japan); Morita, Masahiro; Shimodaira, Tetsuhiro; Tokai-Nishizumi, Noriko; Yokoyama, Kazumasa; Ohsugi, Miho; Suzuki, Toru [Division of Oncology, Department of Cancer Biology, Institute of Medical Science, University of Tokyo, Minato-ku, Tokyo 108-8639 (Japan); Yamamoto, Tadashi, E-mail: tyamamot@ims.u-tokyo.ac.jp [Division of Oncology, Department of Cancer Biology, Institute of Medical Science, University of Tokyo, Minato-ku, Tokyo 108-8639 (Japan); Cell Signal Unit, Okinawa Institute of Science and Technology, Kunigami, Okinawa 904-0412 (Japan)

    2012-03-09

    Highlights: Black-Right-Pointing-Pointer CNOT3 depletion increases the mitotic index. Black-Right-Pointing-Pointer CNOT3 inhibits the expression of MAD1. Black-Right-Pointing-Pointer CNOT3 destabilizes the MAD1 mRNA. Black-Right-Pointing-Pointer MAD1 knockdown attenuates the CNOT3 depletion-induced mitotic arrest. -- Abstract: The stability of mRNA influences the dynamics of gene expression. The CCR4-NOT complex, the major deadenylase in mammalian cells, shortens the mRNA poly(A) tail and contributes to the destabilization of mRNAs. The CCR4-NOT complex plays pivotal roles in various physiological functions, including cell proliferation, apoptosis, and metabolism. Here, we show that CNOT3, a subunit of the CCR4-NOT complex, is involved in the regulation of the spindle assembly checkpoint, suggesting that the CCR4-NOT complex also plays a part in the regulation of mitosis. CNOT3 depletion increases the population of mitotic-arrested cells and specifically increases the expression of MAD1 mRNA and its protein product that plays a part in the spindle assembly checkpoint. We showed that CNOT3 depletion stabilizes the MAD1 mRNA, and that MAD1 knockdown attenuates the CNOT3 depletion-induced increase of the mitotic index. Basing on these observations, we propose that CNOT3 is involved in the regulation of the spindle assembly checkpoint through its ability to regulate the stability of MAD1 mRNA.

  18. The flavonoid eupatorin inactivates the mitotic checkpoint leading to polyploidy and apoptosis

    Energy Technology Data Exchange (ETDEWEB)

    Salmela, Anna-Leena [VTT Technical Research Centre of Finland, Medical Biotechnology, P.O. Box 106, Turku (Finland); Turku Graduate School of Biomedical Sciences, Turku (Finland); Turku Centre for Biotechnology, P.O. Box 123, University of Turku (Finland); Pouwels, Jeroen; Kukkonen-Macchi, Anu [VTT Technical Research Centre of Finland, Medical Biotechnology, P.O. Box 106, Turku (Finland); Waris, Sinikka; Toivonen, Pauliina [Turku Centre for Biotechnology, P.O. Box 123, University of Turku (Finland); Jaakkola, Kimmo [VTT Technical Research Centre of Finland, Medical Biotechnology, P.O. Box 106, Turku (Finland); Maeki-Jouppila, Jenni [VTT Technical Research Centre of Finland, Medical Biotechnology, P.O. Box 106, Turku (Finland); Turku Centre for Biotechnology, P.O. Box 123, University of Turku (Finland); Drug Discovery Graduate School, University of Turku (Finland); Kallio, Lila, E-mail: lila.kallio@vtt.fi [VTT Technical Research Centre of Finland, Medical Biotechnology, P.O. Box 106, Turku (Finland); Kallio, Marko J. [VTT Technical Research Centre of Finland, Medical Biotechnology, P.O. Box 106, Turku (Finland); Turku Centre for Biotechnology, P.O. Box 123, University of Turku (Finland); Centre of Excellence for Translational Genome-Scale Biology, P.O. Box 106, Academy of Finland (Finland)

    2012-03-10

    The spindle assembly checkpoint (SAC) is a conserved mechanism that ensures the fidelity of chromosome distribution in mitosis by preventing anaphase onset until the correct bipolar microtubule-kinetochore attachments are formed. Errors in SAC function may contribute to tumorigenesis by inducing numerical chromosome anomalies (aneuploidy). On the other hand, total disruption of SAC can lead to massive genomic imbalance followed by cell death, a phenomena that has therapeutic potency. We performed a cell-based high-throughput screen with a compound library of 2000 bioactives for novel SAC inhibitors and discovered a plant-derived phenolic compound eupatorin (3 Prime ,5-dihydroxy-4 Prime ,6,7-trimethoxyflavone) as an anti-mitotic flavonoid. The premature override of the microtubule drug-imposed mitotic arrest by eupatorin is dependent on microtubule-kinetochore attachments but not interkinetochore tension. Aurora B kinase activity, which is essential for maintenance of normal SAC signaling, is diminished by eupatorin in cells and in vitro providing a mechanistic explanation for the observed forced mitotic exit. Eupatorin likely has additional targets since eupatorin treatment of pre-mitotic cells causes spindle anomalies triggering a transient M phase delay followed by impaired cytokinesis and polyploidy. Finally, eupatorin potently induces apoptosis in multiple cancer cell lines and suppresses cancer cell proliferation in organotypic 3D cell culture model.

  19. Mitotic phosphorylation of VCIP135 blocks p97ATPase-mediated Golgi membrane fusion

    Energy Technology Data Exchange (ETDEWEB)

    Totsukawa, Go; Matsuo, Ayaka; Kubota, Ayano; Taguchi, Yuya; Kondo, Hisao, E-mail: hk228@med.kyushu-u.ac.jp

    2013-04-05

    Highlights: •VCIP135 is mitotically phosphorylated on Threonine-760 and Serine-767 by Cdc2. •Phosphorylated VCIP135 does not bind to p97ATPase. •The phosphorylation of VCIP135 inhibits p97ATPase-mediated Golgi membrane fusion. -- Abstract: In mammals, the Golgi apparatus is disassembled early mitosis and reassembled at the end of mitosis. For Golgi disassembly, membrane fusion needs to be blocked. Golgi biogenesis requires two distinct p97ATPase-mediated membrane fusion, the p97/p47 and p97/p37 pathways. We previously reported that p47 phosphorylation on Serine-140 and p37 phosphorylation on Serine-56 and Threonine-59 result in mitotic inhibition of the p97/p47 and the p97/p37 pathways, respectively [11,14]. In this study, we show another mechanism of mitotic inhibition of p97-mediated Golgi membrane fusion. We clarified that VCIP135, an essential factor in both p97 membrane fusion pathways, is phosphorylated on Threonine-760 and Serine-767 by Cdc2 at mitosis and that this phosphorylated VCIP135 does not bind to p97. An in vitro Golgi reassembly assay revealed that VCIP135(T760E, S767E), which mimics mitotic phosphorylation, caused no cisternal regrowth. Our results indicate that the phosphorylation of VCIP135 on Threonine-760 and Serine-767 inhibits p97-mediated Golgi membrane fusion at mitosis.

  20. Discrimination of bromodeoxyuridine labelled and unlabelled mitotic cells in flow cytometric bromodeoxyuridine/DNA analysis

    DEFF Research Database (Denmark)

    Jensen, P O; Larsen, J K; Christensen, I J;

    1994-01-01

    Bromodeoxyuridine (BrdUrd) labelled and unlabelled mitotic cells, respectively, can be discriminated from interphase cells using a new method, based on immunocytochemical staining of BrdUrd and flow cytometric four-parameter analysis of DNA content, BrdUrd incorporation, and forward and orthogona...

  1. The flavonoid eupatorin inactivates the mitotic checkpoint leading to polyploidy and apoptosis

    International Nuclear Information System (INIS)

    The spindle assembly checkpoint (SAC) is a conserved mechanism that ensures the fidelity of chromosome distribution in mitosis by preventing anaphase onset until the correct bipolar microtubule–kinetochore attachments are formed. Errors in SAC function may contribute to tumorigenesis by inducing numerical chromosome anomalies (aneuploidy). On the other hand, total disruption of SAC can lead to massive genomic imbalance followed by cell death, a phenomena that has therapeutic potency. We performed a cell-based high-throughput screen with a compound library of 2000 bioactives for novel SAC inhibitors and discovered a plant-derived phenolic compound eupatorin (3′,5-dihydroxy-4′,6,7-trimethoxyflavone) as an anti-mitotic flavonoid. The premature override of the microtubule drug-imposed mitotic arrest by eupatorin is dependent on microtubule–kinetochore attachments but not interkinetochore tension. Aurora B kinase activity, which is essential for maintenance of normal SAC signaling, is diminished by eupatorin in cells and in vitro providing a mechanistic explanation for the observed forced mitotic exit. Eupatorin likely has additional targets since eupatorin treatment of pre-mitotic cells causes spindle anomalies triggering a transient M phase delay followed by impaired cytokinesis and polyploidy. Finally, eupatorin potently induces apoptosis in multiple cancer cell lines and suppresses cancer cell proliferation in organotypic 3D cell culture model.

  2. Classification of mitotic figures with convolutional neural networks and seeded blob features

    Directory of Open Access Journals (Sweden)

    Christopher D Malon

    2013-01-01

    Full Text Available Background: The mitotic figure recognition contest at the 2012 International Conference on Pattern Recognition (ICPR challenges a system to identify all mitotic figures in a region of interest of hematoxylin and eosin stained tissue, using each of three scanners (Aperio, Hamamatsu, and multispectral. Methods: Our approach combines manually designed nuclear features with the learned features extracted by convolutional neural networks (CNN. The nuclear features capture color, texture, and shape information of segmented regions around a nucleus. The use of a CNN handles the variety of appearances of mitotic figures and decreases sensitivity to the manually crafted features and thresholds. Results : On the test set provided by the contest, the trained system achieves F1 scores up to 0.659 on color scanners and 0.589 on multispectral scanner. Conclusions : We demonstrate a powerful technique combining segmentation-based features with CNN, identifying the majority of mitotic figures with a fair precision. Further, we show that the approach accommodates information from the additional focal planes and spectral bands from a multi-spectral scanner without major redesign.

  3. Akt Inhibitor A-443654 Interferes with Mitotic Progression by Regulating Aurora A Kinase Expression

    Directory of Open Access Journals (Sweden)

    Xuesong Liu

    2008-08-01

    Full Text Available Both Akt and Aurora A kinase have been shown to be important targets for intervention for cancer therapy. We report here that Compound A (A-443654, a specific Akt inhibitor, interferes with mitotic progression and bipolar spindle formation. Compound A induces G2/M accumulation, defects in centrosome separation, and formation of either monopolar arrays or disorganized spindles. On the basis of gene expression array studies, we identified Aurora A as one of the genes regulated transcriptionally by Akt inhibitors including Compound A. Inhibition of the phosphatidylinositol 3-kinase (PI3K/Akt pathway, either by PI3K inhibitor LY294002 or by Compound A, dramatically inhibits the promoter activity of Aurora A, whereas the mammalian target of rapamycin inhibitor has little effect, suggesting that Akt might be responsible for up-regulating Aurora A for mitotic progression. Further analysis of the Aurora A promoter region indicates that the Ets element but not the Sp1 element is required for Compound A-sensitive transcriptional control of Aurora A. Overexpression of Aurora A in cells treated with Compound A attenuates the mitotic arrest and the defects in bipolar spindle formation induced by Akt inhibition. Our studies suggest that that Akt may promote mitotic progression through the transcriptional regulation of Aurora A.

  4. Frequencies of mutagen-induced coincident mitotic recombination at unlinked loci in Saccharomyces cerevisiae

    International Nuclear Information System (INIS)

    Frequencies of coincident genetic events were measured in strain D7 of Saccharomyces cerevisiae. This diploid strain permits the detection of mitotic gene conversion involving the trp5-12 and trp5-27 alleles, mitotic crossing-over and gene conversion leading to the expression of the ade2-40 and ade2-119 alleles as red and pink colonies, and reversion of the ilv1-92 allele. The three genes are on different chromosomes, and one might expect that coincident (simultaneous) genetic alterations at two loci would occur at frequencies predicted by those of the single alterations acting as independent events. Contrary to this expectation, we observed that ade2 recombinants induced by bleomycin, β-propiolactone, and ultraviolet radiation occur more frequently among trp5 convertants than among total colonies. This excess among trp5 recombinants indicates that double recombinants are more common than expected for independent events. No similar enrichment was found among Ilv+ revertants. The possibility of an artifact in which haploid yeasts that mimic mitotic recombinants are generated by a low frequency of cryptic meiosis has been excluded. Several hypotheses that can explain the elevated incidence of coincident mitotic recombination have been evaluated, but the cause remains uncertain. Most evidence suggests that the excess is ascribable to a subset of the population being in a recombination-prone state

  5. Direct preparation protocol to obtain mitotic chromosomes from canine mammary tumors.

    Science.gov (United States)

    Morais, C S D; Affonso, P R A M; Bitencourt, J A; Wenceslau, A A

    2015-01-01

    Currently, mammary neoplasms in female canines are a serious problem in veterinary clinics. In addition, the canine species is an excellent disease model for human oncology because of the biological and genetic similarities between the species. Cytogenetics has allowed further study of the characterization of neoplasms in canines. We hypothesized that the use of a direct preparation protocol for mitotic chromosome analysis would provide a simple and low cost protocol for use in all laboratories. The objective of this method is to display in a few hours of dividing cells just like the time of collection since cell division in tissue can be obtained. Ten female canines with the spontaneous occurrence of mammary neoplasia were used to test a pioneering direct preparation protocol to obtain mitotic chromosomes. The excised breast tumor tissue fragments were subjected to the protocol consisting of treatment with colchicine, treatment with hypotonic solution, and fixation. Mitotic chromosomes were absent in cell suspensions of only two samples among the 10 materials analyzed, based on the analysis of five blades for each preparation obtained. So, the cell suspension obtained allowed for the observation of eight tissue samples viable for cytogenetic analysis, five of which had excellent numbers of mitotic chromosomes. However, the technique was unsuccessful in producing high-quality cell suspensions because of inadequate condensation and scattering of chromosomes. While adjustments to methodological procedures are needed, this protocol represents a low cost and simplified method to study the cytogenetics of canine tumors. PMID:26782592

  6. Flow Sorting of Mitotic Chromosomes in Common Wheat (Triticum aestivum L.)

    Czech Academy of Sciences Publication Activity Database

    Vrána, Jan; Doleželová, Marie; Šimková, Hana; Čihalíková, Jarmila; Lysák, Martin; Doležel, Jaroslav

    2000-01-01

    Roč. 156, - (2000), s. 2033-2041. ISSN 0016-6731 R&D Projects: GA ČR GV521/96/K117; GA MŠk ME 376 Institutional research plan: CEZ:AV0Z5038910 Keywords : Mitotic Chromosomes * Triticum aestivum L. Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 4.687, year: 2000

  7. Suspension of Mitotic Activity in Dentate Gyrus of the Hibernating Ground Squirrel

    Directory of Open Access Journals (Sweden)

    Victor I. Popov

    2011-01-01

    Full Text Available Neurogenesis occurs in the adult mammalian hippocampus, a region of the brain important for learning and memory. Hibernation in Siberian ground squirrels provides a natural model to study mitosis as the rapid fall in body temperature in 24 h (from 35-36°C to +4–6°C permits accumulation of mitotic cells at different stages of the cell cycle. Histological methods used to study adult neurogenesis are limited largely to fixed tissue, and the mitotic state elucidated depends on the specific phase of mitosis at the time of day. However, using an immunohistochemical study of doublecortin (DCX and BrdU-labelled neurons, we demonstrate that the dentate gyrus of the ground squirrel hippocampus contains a population of immature cells which appear to possess mitotic activity. Our data suggest that doublecortin-labelled immature cells exist in a mitotic state and may represent a renewable pool for generation of new neurons within the dentate gyrus.

  8. The Drosophila Microtubule-Associated Protein Mars Stabilizes Mitotic Spindles by Crosslinking Microtubules through Its N-Terminal Region

    DEFF Research Database (Denmark)

    Zhang, Gang; Beati, Hamze; Nilsson, Jakob;

    2013-01-01

    reported to stabilize the dynamic spindle through crosslinking adjacent MTs. Mars, a novel MAP, is essential for the early development of Drosophila embryos. Previous studies showed that Mars is required for maintaining an intact mitotic spindle but did not provide a molecular mechanism for this function....... Here we show that Mars is able to stabilize the mitotic spindle in vivo. Both in vivo and in vitro data reveal that the N-terminal region of Mars functions in the stabilization of the mitotic spindle by crosslinking adjacent MTs....

  9. An in vivo model of mitotic cell death and Ras/MAPK signaling

    International Nuclear Information System (INIS)

    Full text: We have created the first and only existing tissue-model of mitotic cell death using the nematode C. elegans. We are able to measure radiation sensitivity in C. elegans by microscopically scoring the percentage of radiation-induced abnormal vulvae. We have found that these abnormalities are due to the death of the vulva cells after their third (and final) division, consistent with post-mitotic cell death. In C. elegans the Ras/MAPK signaling pathway is primarily responsible for the development of the hermaphrodite vulva, and is highly conserved to the mammalian Ras/MAPK pathway. We began by studying the effects of radiation on worm strains with mild loss-of-function (lof) mutations in components of the Ras/MAPK pathway. While the mutant strains that we studied have no abnormalities in normal vulva development, we found that all were radiosensitive, with increased radiation-induced vulval abnormalities as compared to wild-type worms. We therefore wanted to see if overexpression of the Ras/MAPK pathway would confer radioresistance in our system, so we irradiated a gain-of-function (gof) EGFR mutant worm strain. We found that this strain was radioresistant, with less radiation-induced vulval abnormalities than wild-type worms. We have concluded that the Ras/MAPK pathway protects against mitotic cell death in C. elegans. We wanted to better understand the downstream effectors of Ras/MAPK signaling that facilitate protection from mitotic cell death. Since mitotic cell death is due to DNA damage, we hypothesized that worm strains with mutations in the DNA damage response pathway should also be sensitive to mitotic cell death. We have begun analyzing worms with mutations in cell cycle checkpoint genes and DNA damage sensor genes, and have found that all of the strains tested thus far are highly radiosensitive. We plan to genetically cross gain-of-function Ras/MAPK mutants and loss-of-function checkpoint or damage response mutants, and determine the linearity of

  10. The Exon Junction Complex Controls the Efficient and Faithful Splicing of a Subset of Transcripts Involved in Mitotic Cell-Cycle Progression.

    Science.gov (United States)

    Fukumura, Kazuhiro; Wakabayashi, Shunichi; Kataoka, Naoyuki; Sakamoto, Hiroshi; Suzuki, Yutaka; Nakai, Kenta; Mayeda, Akila; Inoue, Kunio

    2016-01-01

    The exon junction complex (EJC) that is deposited onto spliced mRNAs upstream of exon-exon junctions plays important roles in multiple post-splicing gene expression events, such as mRNA export, surveillance, localization, and translation. However, a direct role for the human EJC in pre-mRNA splicing has not been fully understood. Using HeLa cells, we depleted one of the EJC core components, Y14, and the resulting transcriptome was analyzed by deep sequencing (RNA-Seq) and confirmed by RT-PCR. We found that Y14 is required for efficient and faithful splicing of a group of transcripts that is enriched in short intron-containing genes involved in mitotic cell-cycle progression. Tethering of EJC core components (Y14, eIF4AIII or MAGOH) to a model reporter pre-mRNA harboring a short intron showed that these core components are prerequisites for the splicing activation. Taken together, we conclude that the EJC core assembled on pre-mRNA is critical for efficient and faithful splicing of a specific subset of short introns in mitotic cell cycle-related genes. PMID:27490541

  11. Discrimination of bromodeoxyuridine labelled and unlabelled mitotic cells in flow cytometric bromodeoxyuridine/DNA analysis

    DEFF Research Database (Denmark)

    Jensen, P O; Larsen, J K; Christensen, I J;

    1994-01-01

    Bromodeoxyuridine (BrdUrd) labelled and unlabelled mitotic cells, respectively, can be discriminated from interphase cells using a new method, based on immunocytochemical staining of BrdUrd and flow cytometric four-parameter analysis of DNA content, BrdUrd incorporation, and forward and orthogonal...... light scatter. The method was optimized using the human leukemia cell lines HL-60 and K-562. Samples of 10(5) ethanol-fixed cells were treated with pepsin/HCl and stained as a nuclear suspension with anti-BrdUrd antibody, FITC-conjugated secondary antibody, and propidium iodide. Labelled mitoses could...... fluorescence distribution. This interpretation was supported by experiments using mitotic arrest, fluorescence activated cell sorting and microscopy, and comparison with an alternative flow cytometric method for discrimination of mitoses....

  12. Analysis of the Ceratitis capitata y chromosome using in situ hybridization to mitotic chromosomes

    International Nuclear Information System (INIS)

    In Ceratitis capitata the Y chromosome is responsible for sex-determination. We used fluorescence in situ hybridization (FISH) for cytogenetic analysis of mitotic chromosomes. FISH with the wild-type strain EgyptII and two repetitive DNA probes enabled us to differentiate between the short and the long arm of the Y chromosome and gives a much better resolution than C-banding of mitotic chromosomes. We identified the Y-chromosomal breakpoints in Y-autosome translocations using FISH. Even more complex rearrangements i.e. deletions and insertions in some translocation strains were detected by this method. A strategy for mapping the primary sex determination factor in Ceratitis capitata by FISH is presented. (author)

  13. Mitotic Exit Function of Polo-like Kinase Cdc5 Is Dependent on Sequential Activation by Cdk1.

    Science.gov (United States)

    Rodriguez-Rodriguez, Jose-Antonio; Moyano, Yolanda; Játiva, Soraya; Queralt, Ethel

    2016-05-31

    To complete mitosis, Saccharomyces cerevisiae needs to activate the mitotic phosphatase Cdc14. Two pathways contribute to Cdc14 regulation: FEAR (Cdc14 early anaphase release) and MEN (mitotic exit network). Cdc5 polo-like kinase was found to be an important mitotic exit component. However, its specific role in mitotic exit regulation and its involvement in Cdc14 release remain unclear. Here, we provide insight into the mechanism by which Cdc5 contributes to the timely release of Cdc14. Our genetic and biochemical data indicate that Cdc5 acts in parallel with MEN during anaphase. This MEN-independent Cdc5 function requires active separase and activation by Cdk1-dependent phosphorylation. Cdk1 first phosphorylates Cdc5 to activate it in early anaphase, and then, in late anaphase, further phosphorylation of Cdc5 by Cdk1 is needed to promote its MEN-related functions. PMID:27210759

  14. Human papillomavirus type 16 E7 oncoprotein engages but does not abrogate the mitotic spindle assembly checkpoint

    Energy Technology Data Exchange (ETDEWEB)

    Yu, Yueyang [Division of Infectious Diseases, Brigham and Women' s Hospital and Biological and Biomedical Sciences Program, Harvard Medical School, Boston, MA 02115 (United States); Munger, Karl, E-mail: kmunger@rics.bwh.harvard.edu [Division of Infectious Diseases, Brigham and Women' s Hospital and Biological and Biomedical Sciences Program, Harvard Medical School, Boston, MA 02115 (United States)

    2012-10-10

    The mitotic spindle assembly checkpoint (SAC) ensures faithful chromosome segregation during mitosis by censoring kinetochore-microtubule interactions. It is frequently rendered dysfunctional during carcinogenesis causing chromosome missegregation and genomic instability. There are conflicting reports whether the HPV16 E7 oncoprotein drives chromosomal instability by abolishing the SAC. Here we report that degradation of mitotic cyclins is impaired in cells with HPV16 E7 expression. RNAi-mediated depletion of Mad2 or BubR1 indicated the involvement of the SAC, suggesting that HPV16 E7 expression causes sustained SAC engagement. Mutational analyses revealed that HPV16 E7 sequences that are necessary for retinoblastoma tumor suppressor protein binding as well as sequences previously implicated in binding the nuclear and mitotic apparatus (NuMA) protein and in delocalizing dynein from the mitotic spindle contribute to SAC engagement. Importantly, however, HPV16 E7 does not markedly compromise the SAC response to microtubule poisons.

  15. Regulation of mitotic progression : Focus on Plk1 function and the novel Ska complex at kinetochores

    OpenAIRE

    Hanisch, Anja

    2007-01-01

    During mitosis the duplicated chromosomes have to be faithfully segregated into the nascent daughter cells in order to maintain genomic stability. This critical process is dependent on the rearrangement of the interphase microtubule (MT) network, resulting in the formation of a bipolar mitotic spindle. For proper chromosome segregation all chromosomes have to become connected to MTs emanating from opposite spindle poles. The MT attachment sites on the chromosomes are the kinetochores (KTs), w...

  16. The forces that center the mitotic spindle in the C. elegans embryo

    OpenAIRE

    Garzon-Coral, Carlos

    2015-01-01

    The precise positioning of the mitotic spindle to the cell center during mitosis is a fundamental process for chromosome segregation and the division plane definition. Despite its importance, the mechanism for spindle centering remains elusive. To study this mechanism, the dynamic of the microtubules was characterized at the bulk and at the cortex in the C. elegans embryo. Then, this dynamic was correlated to the centering forces of the spindle that were studied by applying calibrated magneti...

  17. Mechanism of the mitotic kinesin CENP-E in tethering kinetochores to spindle microtubules

    OpenAIRE

    Kim, Yumi

    2009-01-01

    The mitotic kinesin CENP-E is an essential kinetochore motor that directly contributes to the capture and stabilization of spindle microtubules by kinetochores. Although it has been well established that CENP-E is essential for metaphase chromosome alignment and reduction of CENP-E leads to high rates of whole chromosome missegregation in cells, its properties as a microtubule- dependent motor, the mechanism by which CENP-E contributes to the dynamic linkage between kinetochores and spindle m...

  18. c-Mos forces the mitotic cell cycle to undergo meiosis II to produce haploid gametes

    OpenAIRE

    Tachibana, Kazunori; Tanaka, Daisuke; Isobe, Tomohiro; Kishimoto, Takeo

    2000-01-01

    The meiotic cycle reduces ploidy through two consecutive M phases, meiosis I and meiosis II, without an intervening S phase. To maintain ploidy through successive generations, meiosis must be followed by mitosis after the recovery of diploidy by fertilization. However, the coordination from meiotic to mitotic cycle is still unclear. Mos, the c-mos protooncogene product, is a key regulator of meiosis in vertebrates. In contrast to the previous observation that Mos f...

  19. Costunolide causes mitotic arrest and enhances radiosensitivity in human hepatocellular carcinoma cells

    Directory of Open Access Journals (Sweden)

    Chen Chih-Jen

    2011-05-01

    Full Text Available Abstract Purpose This work aimed to investigate the effect of costunolide, a sesquiterpene lactone isolated from Michelia compressa, on cell cycle distribution and radiosensitivity of human hepatocellular carcinoma (HCC cells. Methods The assessment used in this study included: cell viability assay, cell cycle analysis by DNA histogram, expression of phosphorylated histone H3 (Ser 10 by flow cytometer, mitotic index by Liu's stain and morphological observation, mitotic spindle alignment by immunofluorescence of alpha-tubulin, expression of cell cycle-related proteins by Western blotting, and radiation survival by clonogenic assay. Results Our results show that costunolide reduced the viability of HA22T/VGH cells. It caused a rapid G2/M arrest at 4 hours shown by DNA histogram. The increase in phosphorylated histone H3 (Ser 10-positive cells and mitotic index indicates costunolide-treated cells are arrested at mitosis, not G2, phase. Immunofluorescence of alpha-tubulin for spindle formation further demonstrated these cells are halted at metaphase. Costunolide up-regulated the expression of phosphorylated Chk2 (Thr 68, phosphorylated Cdc25c (Ser 216, phosphorylated Cdk1 (Tyr 15 and cyclin B1 in HA22T/VGH cells. At optimal condition causing mitotic arrest, costunolide sensitized HA22T/VGH HCC cells to ionizing radiation with sensitizer enhancement ratio up to 1.9. Conclusions Costunolide could reduce the viability and arrest cell cycling at mitosis in hepatoma cells. Logical exploration of this mitosis-arresting activity for cancer therapeutics shows costunolide enhanced the killing effect of radiotherapy against human HCC cells.

  20. Mitotic Spindle Positioning in Saccharomyces cerevisiae Is Accomplished by Antagonistically Acting Microtubule Motor Proteins

    OpenAIRE

    Cottingham, Frank R.; Hoyt, M. Andrew

    1997-01-01

    Proper positioning of the mitotic spindle is often essential for cell division and differentiation processes. The asymmetric cell division characteristic of budding yeast, Saccharomyces cerevisiae, requires that the spindle be positioned at the mother–bud neck and oriented along the mother–bud axis. The single dynein motor encoded by the S. cerevisiae genome performs an important but nonessential spindle-positioning role. We demonstrate that kinesin-related Kip3p makes a major contribution to...

  1. The role of p53 in the response to mitotic spindle damage

    International Nuclear Information System (INIS)

    The p53 tumour suppressor protein has defined roles in G1/S and G2/M cell cycle checkpoint in response to a range of cellular stresses including DNA damage, dominant oncogene expression, hypoxia, metabolic changes and viral infection. In addition to these responses, p53 can also be activated when damage occurs to the mitotic spindle. Initially, spindle damage activates a p53-independent checkpoint which functions at the metaphase-anaphase transition and prevents cells from progressing through mitosis until the completion of spindle formation. Cells eventually escape from this block (a process termed 'mitotic slippage'), and an aberrant mitosis ensues in which sister chromatids fail to segregate properly. After a delay period, p53 responds to this mitotic failure by instituting a G1-like growth arrest, with an intact nucleus containing 4N DNA, but without the cells undergoing division. Cells lacking wild-type p53 are still able to arrest transiently at mitosis, and also fail to undergo division, underscoring that the delay in mitosis is p53-independent. However, these cells are not prevented from re-entering the cell cycle and can reduplicate their DNA unchecked, leading to polyploidy. Additionally, p53-null cells which experience spindle failure often show the appearance of micronuclei arising from poorly segregated chromosomes which have de-condensed and been enclosed in a nuclear envelope. The ability of p53 to prevent their formation suggests an additional G2 involvement which prevents nuclear breakdown prior to mitosis. The molecular mechanism by which p53 is able to sense mitotic failure is still unknown, but may be linked to the ability of p53 to regulate duplication of the centrosome, the organelle which nucleates spindle formation. (authors)

  2. Interaction of the Betapapillomavirus E2 Tethering Protein with Mitotic Chromosomes▿

    OpenAIRE

    Sekhar, Vandana; Reed, Shawna C.; Alison A McBride

    2009-01-01

    During persistent papillomavirus infection, the viral E2 protein tethers the viral genome to the host cell chromosomes, ensuring maintenance and segregation of the viral genome during cell division. However, E2 proteins from different papillomaviruses interact with distinct chromosomal regions and targets. The tethering mechanism has been best characterized for bovine papillomavirus type 1 (BPV1), where the E2 protein tethers the viral genome to mitotic chromosomes in complex with the cellula...

  3. Mitotic Kinesin-Like Protein 2 Binds and Colocalizes with Papillomavirus E2 during Mitosis▿

    OpenAIRE

    Yu, Ting; Peng, Yu-Cai; Androphy, Elliot J.

    2006-01-01

    MKlp2 is a kinesin-like motor protein of the central mitotic spindle required for completion of cytokinesis. Papillomavirus E2 is a sequence specific DNA binding protein that regulates viral transcription and replication and is responsible for partitioning viral episomes into daughter cells during cell division. We demonstrate that MKlp2 specifically associates with the E2 protein during mitosis. Using chromatin immunoprecipitation, we show viral genomes are in complex with MKlp2 only within ...

  4. Hair cell recovery in mitotically blocked cultures of the bullfrog saccule

    OpenAIRE

    Baird, Richard A.; Burton, Miriam D.; Fashena, David S.; Naeger, Rebecca A.

    2000-01-01

    Hair cells in many nonmammalian vertebrates are regenerated by the mitotic division of supporting cell progenitors and the differentiation of the resulting progeny into new hair cells and supporting cells. Recent studies have shown that nonmitotic hair cell recovery after aminoglycoside-induced damage can also occur in the vestibular organs. Using hair cell and supporting cell immunocytochemical markers, we have used confocal and electron microscopy to examine the fate...

  5. Caspase-Mediated Specific Cleavage of BubR1 Is a Determinant of Mitotic Progression

    OpenAIRE

    Kim, Mijin; Murphy, Katie; Liu, Fang; Parker, Sharon E.; Dowling, Melissa L.; Baff, Wesley; Kao, Gary D.

    2007-01-01

    The fidelity of chromosomal duplication is monitored by cell cycle checkpoints operational during mitosis. One such cell cycle delay is invoked by microtubule-targeting agents such as nocodazole or paclitaxel (Taxol) and is mediated by mitotic checkpoint proteins that include BubR1. Relatively little is known about the regulation of expression and stability of BubR1 (or other checkpoint proteins) and how these factors dictate the durability of the cell cycle delay. We report here that treatme...

  6. Mutagenesis and mitotic recombination in Aspergillus niger, an expedition from gene to genome

    OpenAIRE

    Vondervoort, Peter Jozef Ida van de

    2007-01-01

    Aspergillus niger is a cosmopolitan fungus and its spores can be found in air and soil worldwide. This saprophyte is used in food biotechnology for the production of proteins, mainly enzymes and for the production of organic acids. In the production of proteins, several problems are encountered such as repressed gene-expression, morphologic and genetic instability and undesired metabolite accumulation. To find solutions to these problems, often mutagenesis and mitotic recombination have been ...

  7. Costunolide causes mitotic arrest and enhances radiosensitivity in human hepatocellular carcinoma cells

    International Nuclear Information System (INIS)

    This work aimed to investigate the effect of costunolide, a sesquiterpene lactone isolated from Michelia compressa, on cell cycle distribution and radiosensitivity of human hepatocellular carcinoma (HCC) cells. The assessment used in this study included: cell viability assay, cell cycle analysis by DNA histogram, expression of phosphorylated histone H3 (Ser 10) by flow cytometer, mitotic index by Liu's stain and morphological observation, mitotic spindle alignment by immunofluorescence of alpha-tubulin, expression of cell cycle-related proteins by Western blotting, and radiation survival by clonogenic assay. Our results show that costunolide reduced the viability of HA22T/VGH cells. It caused a rapid G2/M arrest at 4 hours shown by DNA histogram. The increase in phosphorylated histone H3 (Ser 10)-positive cells and mitotic index indicates costunolide-treated cells are arrested at mitosis, not G2, phase. Immunofluorescence of alpha-tubulin for spindle formation further demonstrated these cells are halted at metaphase. Costunolide up-regulated the expression of phosphorylated Chk2 (Thr 68), phosphorylated Cdc25c (Ser 216), phosphorylated Cdk1 (Tyr 15) and cyclin B1 in HA22T/VGH cells. At optimal condition causing mitotic arrest, costunolide sensitized HA22T/VGH HCC cells to ionizing radiation with sensitizer enhancement ratio up to 1.9. Costunolide could reduce the viability and arrest cell cycling at mitosis in hepatoma cells. Logical exploration of this mitosis-arresting activity for cancer therapeutics shows costunolide enhanced the killing effect of radiotherapy against human HCC cells

  8. A role for Arf1 in mitotic Golgi disassembly, chromosome segregation, and cytokinesis

    OpenAIRE

    Altan-Bonnet, Nihal; Phair, Robert D.; Polishchuk, Roman S.; Weigert, Roberto; Lippincott-Schwartz, Jennifer

    2003-01-01

    In mitosis, chromosome, cytoskeleton, and organelle dynamics must be coordinated for successful cell division. Here, we present evidence for a role for Arf1, a small GTPase associated with the Golgi apparatus, in the orchestration of mitotic Golgi breakdown, chromosome segregation, and cytokinesis. We show that early in mitosis Arf1 becomes inactive and dissociates from Golgi membranes. This is followed by the dispersal of numerous Arf1-dependent peripheral Golgi proteins and subsequent Golgi...

  9. Prognostic value of mitotic index and Bcl2 expression in male breast cancer.

    OpenAIRE

    Lacle, M.M.; van der Pol, C.C.; Witkamp, A. J.; van der Wall, E.; van Diest, P.J.

    2013-01-01

    The incidence of male breast cancer (MBC) is rising. Current treatment regimens for MBC are extrapolated from female breast cancer (FBC), based on the assumption that FBC prognostic features and therapeutic targets can be extrapolated to MBC. However, there is yet little evidence that prognostic features that have been developed and established in FBC are applicable to MBC as well. In a recent study on FBC, a combination of mitotic index and Bcl2 expression proved to be of strong prognostic v...

  10. Mitotically active microglandular hyperplasia of the cervix: a case series with implications for the differential diagnosis.

    Science.gov (United States)

    Abi-Raad, Rita; Alomari, Ahmed; Hui, Pei; Buza, Natalia

    2014-09-01

    Microglandular hyperplasia (MGH) is a benign proliferation of endocervical glands with relatively uniform columnar or cuboidal nuclei, and rare to absent mitoses. Endometrial adenocarcinomas with mucinous differentiation or a microglandular pattern can closely mimic MGH, often resulting in a diagnostic dilemma in small biopsy specimens. Rare unusual morphologic features-mild to moderate nuclear atypia, solid or reticular growth pattern, hobnail and signet ring cells-have been previously reported in MGH. We present 9 cases of unusual, mitotically active-between 5 and 11 mitotic figures per 10 HPF-MGH, all of which presented as endocervical polyps and had morphologic features otherwise typical of MGH. The patients' age ranged between 35 and 56 yr, 2 patients were postmenopausal. High-risk human papillomavirus status was available in 7 patients, all of which were negative. The Ki-67 proliferation index ranged between 1% and 15%, and all cases were negative for p16, carcinoembryonic antigen, and vimentin immunostains. The clinical follow-up ranged from 3 to 76.2 mo, with a median of 40.7 mo, all patients were doing well without evidence of endocervical or endometrial malignancy. In summary, this case series documents the presence of rare cases of MGH demonstrating significant mitotic activity (up to 11/10 HPF) without a negative impact on the clinical prognosis. Mitotic activity alone should be interpreted with caution in small biopsy specimens with microglandular growth pattern. Immunohistochemical stains, especially p16, carcinoembryonic antigen, and vimentin, may be helpful-in addition to the patient's clinical history and human papillomavirus status to rule out endocervical or endometrial malignancy. PMID:25083971

  11. Drought Resistance and Mitotic Instability of Tritipyrum Compared with Triticale and Bread Wheat

    OpenAIRE

    shahriari, zolfaghar; Mohammad Taghi ASSAD; Hosein Shahsavand HASANI

    2012-01-01

    This study presents the first data on the drought resistance pattern of seven new synthetic 6x primary Tritipyrum amphiploid linesand evaluates their mitotic instability. The primary Tritipyrum lines were crossed with Iranian 6x bread wheat ‘Navid’ cultivar and theirF1 and F2 progenies were obtained. Two experiments with complete randomized design were conducted under optimum and limitedwater conditions to evaluate Tritipyrum-derived genotypes for drought resistance in greenhouse. Under optim...

  12. Quantitative phosphoproteomics reveals new roles for the protein phosphatase PP6 in mitotic cells.

    Science.gov (United States)

    Rusin, Scott F; Schlosser, Kate A; Adamo, Mark E; Kettenbach, Arminja N

    2015-10-13

    Protein phosphorylation is an important regulatory mechanism controlling mitotic progression. Protein phosphatase 6 (PP6) is an essential enzyme with conserved roles in chromosome segregation and spindle assembly from yeast to humans. We applied a baculovirus-mediated gene silencing approach to deplete HeLa cells of the catalytic subunit of PP6 (PP6c) and analyzed changes in the phosphoproteome and proteome in mitotic cells by quantitative mass spectrometry-based proteomics. We identified 408 phosphopeptides on 272 proteins that increased and 298 phosphopeptides on 220 proteins that decreased in phosphorylation upon PP6c depletion in mitotic cells. Motif analysis of the phosphorylated sites combined with bioinformatics pathway analysis revealed previously unknown PP6c-dependent regulatory pathways. Biochemical assays demonstrated that PP6c opposed casein kinase 2-dependent phosphorylation of the condensin I subunit NCAP-G, and cellular analysis showed that depletion of PP6c resulted in defects in chromosome condensation and segregation in anaphase, consistent with dysregulation of condensin I function in the absence of PP6 activity. PMID:26462736

  13. Effect of Various Doses of Nicotine on Mitotic Index in Esophageal Mucosa

    Directory of Open Access Journals (Sweden)

    S. Khajeh Jahromi

    2016-07-01

    Full Text Available Introduction & Objective: Nicotine could directly act as a cancer promoter. The purpose of this study was to evaluate effects of nicotine on mitotic index in esophagus epithelium. Materials & Methods: In the present study 30 adult male mice were used. Animals were ran-domly divided into three groups. Group A or the control group received vehicle, groups B and C received nicotine intraperitoneally at doses of 0.2 and 0.4 mg/kg once daily for 14 days, re-spectively. Evaluations were made using kI-67 immunohistochemistry and Hematoxilin& Eo-sin for proliferative activity and morphometric study on esophagus mucosa, respectively. Results: Administration of nicotine in group C, showed a significant increase (P<0.05 in KI-67 index 34.15±2.50vs. 10.41±1.4 compared with the control subjects. The other parameters such as epithelial height, lamina propria, muscular mucosa and mucosa height in nicotine- treated groups were not affected. Nicotine at dose of 0.2 mg/kg did not change the mitotic in-dex in epithelium when compared with the control group. Conclusion: This study indicates nicotine at dose of 0.4 mg/kg increases mitotic activity in basal cells in esophagus epithelium. (Sci J Hamadan Univ Med Sci 2016; 23 (2:126-133

  14. Live Cell Imaging of the Cancer-related Transcription Factor RUNX2 during Mitotic Progression

    Science.gov (United States)

    Pockwinse, Shirwin M.; Kota, Krishna P.; Quaresma, Alexandre J.C.; Imbalzano, Anthony N.; Lian, Jane B.; van Wijnen, Andre J.; Stein, Janet L.; Stein, Gary S.; Nickerson, Jeffrey A.

    2010-01-01

    The nuclear matrix bound transcription factor RUNX2 is a lineage-specific developmental regulator that is linked to cancer. We have previously shown that RUNX2 controls transcription of both RNA polymerase II genes and RNA polymerase I dependent ribosomal RNA genes. RUNX2 is epigenetically retained through mitosis on both classes of target genes in condensed chromosomes. We have used fluorescence recovery after photobleaching (FRAP) to measure the relative binding kinetics of EGFP-RUNX2 at transcription sites in the nucleus and nucleoli during interphase, as well as on mitotic chromosomes. RUNX2 becomes more strongly bound as cells go from interphase through prophase, with a doubling of the most tightly bound “immobile fraction”. RUNX2 exchange then becomes much more facile during metaphase to telophase. During interphase the less tightly bound pool of RUNX2 exchanges more slowly at nucleoli than at subnuclear foci, and the non-exchanging immobile fraction is greater in nucleoli. These results are consistent with a model in which the molecular mechanism of RUNX2 binding is different at protein-coding and ribosomal RNA genes. The binding interactions of RUNX2 change as cells go through mitosis, with binding affinity increasing as chromosomes condense and then decreasing through subsequent mitotic phases. The increased residence of RUNX2 at mitotic chromosomes may reflect its epigenetic function in “bookmarking” of target genes in cancer cells. PMID:20945391

  15. APC2 and Axin promote mitotic fidelity by facilitating centrosome separation and cytoskeletal regulation.

    Science.gov (United States)

    Poulton, John S; Mu, Frank W; Roberts, David M; Peifer, Mark

    2013-10-01

    To ensure the accurate transmission of genetic material, chromosome segregation must occur with extremely high fidelity. Segregation errors lead to chromosomal instability (CIN), with deleterious consequences. Mutations in the tumor suppressor adenomatous polyposis coli (APC) initiate most colon cancers and have also been suggested to promote disease progression through increased CIN, but the mechanistic role of APC in preventing CIN remains controversial. Using fly embryos as a model, we investigated the role of APC proteins in CIN. Our findings suggest that APC2 loss leads to increased rates of chromosome segregation error. This occurs through a cascade of events beginning with incomplete centrosome separation leading to failure to inhibit formation of ectopic cleavage furrows, which result in mitotic defects and DNA damage. We test several hypotheses related to the mechanism of action of APC2, revealing that APC2 functions at the embryonic cortex with several protein partners, including Axin, to promote mitotic fidelity. Our in vivo data demonstrate that APC2 protects genome stability by modulating mitotic fidelity through regulation of the cytoskeleton. PMID:24026117

  16. Late A-bomb effects on proliferation and mitotic inhibition of T- and B-lymphocytes

    International Nuclear Information System (INIS)

    In order to investigate late effects of ionization radiation and aging on T- and B-lymphocytes, mitotic ability of T- and B-lymphocytes in the peripheral blood of 266 A-bomb survivors was examined by determining the incorporation of [3H]-thymidine. Phytohemagglutinin (PHA) and pokeweed mitogen (PWM) were used as inducers. Furthermore, mitotic inhibition of lymphocytes induced by a lymphatic inhibitor which was in part prepared from ulex seed extracts (USE) was examined. A decreased reaction of peripheral lymphocytes to PHA was seen in men exposed to 100-199 rad; a decreased reaction to PWM was seen in women exposed to more than 200 rad. According to the age group at examination, these decreased reactions were remarkable in men aged 60 years or younger and women aged 60 years or older. Among men less than 60-year-old exposed to 100-199 rad, PWM-induced mitosis of lymphocytes tended to be inhibited remarkably by USE. These results suggest the involvement of late A-bomb effects in mitotic regulation of T- and B-lymphocytes of aged A-bomb survivors. (Namekawa, K.)

  17. Inhibition of the mitotic exit network in response to damaged telomeres.

    Directory of Open Access Journals (Sweden)

    Mauricio Valerio-Santiago

    Full Text Available When chromosomal DNA is damaged, progression through the cell cycle is halted to provide the cells with time to repair the genetic material before it is distributed between the mother and daughter cells. In Saccharomyces cerevisiae, this cell cycle arrest occurs at the G2/M transition. However, it is also necessary to restrain exit from mitosis by maintaining Bfa1-Bub2, the inhibitor of the Mitotic Exit Network (MEN, in an active state. While the role of Bfa1 and Bub2 in the inhibition of mitotic exit when the spindle is not properly aligned and the spindle position checkpoint is activated has been extensively studied, the mechanism by which these proteins prevent MEN function after DNA damage is still unclear. Here, we propose that the inhibition of the MEN is specifically required when telomeres are damaged but it is not necessary to face all types of chromosomal DNA damage, which is in agreement with previous data in mammals suggesting the existence of a putative telomere-specific DNA damage response that inhibits mitotic exit. Furthermore, we demonstrate that the mechanism of MEN inhibition when telomeres are damaged relies on the Rad53-dependent inhibition of Bfa1 phosphorylation by the Polo-like kinase Cdc5, establishing a new key role of this kinase in regulating cell cycle progression.

  18. Hair cell recovery in mitotically blocked cultures of the bullfrog saccule

    Science.gov (United States)

    Baird, R. A.; Burton, M. D.; Fashena, D. S.; Naeger, R. A.

    2000-01-01

    Hair cells in many nonmammalian vertebrates are regenerated by the mitotic division of supporting cell progenitors and the differentiation of the resulting progeny into new hair cells and supporting cells. Recent studies have shown that nonmitotic hair cell recovery after aminoglycoside-induced damage can also occur in the vestibular organs. Using hair cell and supporting cell immunocytochemical markers, we have used confocal and electron microscopy to examine the fate of damaged hair cells and the origin of immature hair cells after gentamicin treatment in mitotically blocked cultures of the bullfrog saccule. Extruding and fragmenting hair cells, which undergo apoptotic cell death, are replaced by scar formations. After losing their bundles, sublethally damaged hair cells remain in the sensory epithelium for prolonged periods, acquiring supporting cell-like morphology and immunoreactivity. These modes of damage appear to be mutually exclusive, implying that sublethally damaged hair cells repair their bundles. Transitional cells, coexpressing hair cell and supporting cell markers, are seen near scar formations created by the expansion of neighboring supporting cells. Most of these cells have morphology and immunoreactivity similar to that of sublethally damaged hair cells. Ultrastructural analysis also reveals that most immature hair cells had autophagic vacuoles, implying that they originated from damaged hair cells rather than supporting cells. Some transitional cells are supporting cells participating in scar formations. Supporting cells also decrease in number during hair cell recovery, supporting the conclusion that some supporting cells undergo phenotypic conversion into hair cells without an intervening mitotic event.

  19. Insulin growth factors regulate the mitotic cycle in cultured rat sympathetic neuroblasts

    International Nuclear Information System (INIS)

    While neuronal mitosis is uniquely restricted to early development, the underlying regulation remains to be defined. The authors have now developed a dissociated, embryonic sympathetic neuron culture system that uses fully defined medium in which cells enter the mitotic cycle. The cultured cells expressed two neuronal traits, tyrosine hydroxylase and the neuron-specific 160-kDa neurofilament subunit protein, but were devoid of glial fibrillary acidic protein, a marker for non-myelin-forming Schwann cells in ganglia. Approximately one-third of the tyrosine hydroxylase-positive cells synthesized DNA in culture, specifically incorporating [3H]thymidine into their nuclei. They used this system to define factors regulating the mitotic cycle in sympathetic neuroblasts. Members of the insulin family of growth factors, including insulin and insulin-like growth factors I and II, regulated DNA synthesis in the presumptive neuroblasts. Insulin more than doubled the proportion of tyrosine hydroxylase-positive cells entering the mitotic cycle, as indicated by autoradiography of [3H]thymidine incorporation into nuclei. Scintillation spectrometry was an even more sensitive index of DNA synthesis. In contrast, the trophic protein nerve growth factor exhibited no mitogenic effect, suggesting that the mitogenic action of insulin growth factors is highly specific. The observations are discussed in the context of the detection of insulin growth factors and receptors in the developing brain

  20. The proteolysis of mitotic cyclins in mammalian cells persists from the end of mitosis until the onset of S phase.

    OpenAIRE

    Brandeis, M.; Hunt, T

    1996-01-01

    We have studied how the cell cycle-specific oscillations of mitotic B-type cyclins are generated in mouse fibroblasts. A reporter enzyme comprising the N-terminus of a B-type cyclin fused to bacterial chloramphenicol acetyl transferase (CAT) was degraded at the end of mitosis like endogenous cyclins. Point mutations in the destruction box of this construct completely abolished its mitotic instability. When the destructible reporter was driven by the cyclin B2 promoter, CAT activity mimicked t...

  1. The SUMO protease SENP1 is required for cohesion maintenance and mitotic arrest following spindle poison treatment

    Energy Technology Data Exchange (ETDEWEB)

    Era, Saho [Fondazione IFOM, Istituto FIRC di Oncologia Molecolare, IFOM-IEO campus, Via Adamello 16, 20139 Milan (Italy); Radiation Genetics, Graduate School of Medicine, Kyoto University, Yoshida-Konoe, Sakyo-ku, Kyoto 606-8501 (Japan); Abe, Takuya; Arakawa, Hiroshi [Fondazione IFOM, Istituto FIRC di Oncologia Molecolare, IFOM-IEO campus, Via Adamello 16, 20139 Milan (Italy); Kobayashi, Shunsuke [Radiation Genetics, Graduate School of Medicine, Kyoto University, Yoshida-Konoe, Sakyo-ku, Kyoto 606-8501 (Japan); Szakal, Barnabas [Fondazione IFOM, Istituto FIRC di Oncologia Molecolare, IFOM-IEO campus, Via Adamello 16, 20139 Milan (Italy); Yoshikawa, Yusuke; Motegi, Akira; Takeda, Shunichi [Radiation Genetics, Graduate School of Medicine, Kyoto University, Yoshida-Konoe, Sakyo-ku, Kyoto 606-8501 (Japan); Branzei, Dana, E-mail: dana.branzei@ifom.eu [Fondazione IFOM, Istituto FIRC di Oncologia Molecolare, IFOM-IEO campus, Via Adamello 16, 20139 Milan (Italy)

    2012-09-28

    Highlights: Black-Right-Pointing-Pointer SENP1 knockout chicken DT40 cells are hypersensitive to spindle poisons. Black-Right-Pointing-Pointer Spindle poison treatment of SENP1{sup -/-} cells leads to increased mitotic slippage. Black-Right-Pointing-Pointer Mitotic slippage in SENP1{sup -/-} cells associates with apoptosis and endoreplication. Black-Right-Pointing-Pointer SENP1 counteracts sister chromatid separation during mitotic arrest. Black-Right-Pointing-Pointer Plk1-mediated cohesion down-regulation is involved in colcemid cytotoxicity. -- Abstract: SUMO conjugation is a reversible posttranslational modification that regulates protein function. SENP1 is one of the six SUMO-specific proteases present in vertebrate cells and its altered expression is observed in several carcinomas. To characterize SENP1 role in genome integrity, we generated Senp1 knockout chicken DT40 cells. SENP1{sup -/-} cells show normal proliferation, but are sensitive to spindle poisons. This hypersensitivity correlates with increased sister chromatid separation, mitotic slippage, and apoptosis. To test whether the cohesion defect had a causal relationship with the observed mitotic events, we restored the cohesive status of sister chromatids by introducing the TOP2{alpha}{sup +/-} mutation, which leads to increased catenation, or by inhibiting Plk1 and Aurora B kinases that promote cohesin release from chromosomes during prolonged mitotic arrest. Although TOP2{alpha} is SUMOylated during mitosis, the TOP2{alpha}{sup +/-} mutation had no obvious effect. By contrast, inhibition of Plk1 or Aurora B rescued the hypersensitivity of SENP1{sup -/-} cells to colcemid. In conclusion, we identify SENP1 as a novel factor required for mitotic arrest and cohesion maintenance during prolonged mitotic arrest induced by spindle poisons.

  2. RBPJ, the major transcriptional effector of Notch signaling, remains associated with chromatin throughout mitosis, suggesting a role in mitotic bookmarking.

    Directory of Open Access Journals (Sweden)

    Robert J Lake

    2014-03-01

    Full Text Available Mechanisms that maintain transcriptional memory through cell division are important to maintain cell identity, and sequence-specific transcription factors that remain associated with mitotic chromatin are emerging as key players in transcriptional memory propagation. Here, we show that the major transcriptional effector of Notch signaling, RBPJ, is retained on mitotic chromatin, and that this mitotic chromatin association is mediated through the direct association of RBPJ with DNA. We further demonstrate that RBPJ binds directly to nucleosomal DNA in vitro, with a preference for sites close to the entry/exit position of the nucleosomal DNA. Genome-wide analysis in the murine embryonal-carcinoma cell line F9 revealed that roughly 60% of the sites occupied by RBPJ in asynchronous cells were also occupied in mitotic cells. Among them, we found that a fraction of RBPJ occupancy sites shifted between interphase and mitosis, suggesting that RBPJ can be retained on mitotic chromatin by sliding on DNA rather than disengaging from chromatin during mitotic chromatin condensation. We propose that RBPJ can function as a mitotic bookmark, marking genes for efficient transcriptional activation or repression upon mitotic exit. Strikingly, we found that sites of RBPJ occupancy were enriched for CTCF-binding motifs in addition to RBPJ-binding motifs, and that RBPJ and CTCF interact. Given that CTCF regulates transcription and bridges long-range chromatin interactions, our results raise the intriguing hypothesis that by collaborating with CTCF, RBPJ may participate in establishing chromatin domains and/or long-range chromatin interactions that could be propagated through cell division to maintain gene expression programs.

  3. The SUMO protease SENP1 is required for cohesion maintenance and mitotic arrest following spindle poison treatment

    International Nuclear Information System (INIS)

    Highlights: ► SENP1 knockout chicken DT40 cells are hypersensitive to spindle poisons. ► Spindle poison treatment of SENP1−/− cells leads to increased mitotic slippage. ► Mitotic slippage in SENP1−/− cells associates with apoptosis and endoreplication. ► SENP1 counteracts sister chromatid separation during mitotic arrest. ► Plk1-mediated cohesion down-regulation is involved in colcemid cytotoxicity. -- Abstract: SUMO conjugation is a reversible posttranslational modification that regulates protein function. SENP1 is one of the six SUMO-specific proteases present in vertebrate cells and its altered expression is observed in several carcinomas. To characterize SENP1 role in genome integrity, we generated Senp1 knockout chicken DT40 cells. SENP1−/− cells show normal proliferation, but are sensitive to spindle poisons. This hypersensitivity correlates with increased sister chromatid separation, mitotic slippage, and apoptosis. To test whether the cohesion defect had a causal relationship with the observed mitotic events, we restored the cohesive status of sister chromatids by introducing the TOP2α+/− mutation, which leads to increased catenation, or by inhibiting Plk1 and Aurora B kinases that promote cohesin release from chromosomes during prolonged mitotic arrest. Although TOP2α is SUMOylated during mitosis, the TOP2α+/− mutation had no obvious effect. By contrast, inhibition of Plk1 or Aurora B rescued the hypersensitivity of SENP1−/− cells to colcemid. In conclusion, we identify SENP1 as a novel factor required for mitotic arrest and cohesion maintenance during prolonged mitotic arrest induced by spindle poisons.

  4. Cloning of four cycling from maize indicates that higher plants have three structurally distinct groups of mitotic cyclins

    OpenAIRE

    Renaudin, J P; Colasanti, J; RIME, Hélène; Z. Yuan; Sundaresan, V.

    1994-01-01

    While a large number of cyclins have been described in animals and yeasts, very limited information is available regarding cyclins in plants. We describe here the isolation of cDNA clones encoding four putative mitotic cyclins from maize. All four cyclins were able to induce maturation of Xenopus oocytes, demonstrating that they can act as mitotic cyclins in this system. Northern analysis showed that all four cyclins were expressed only in actively dividing tissues and organs, with a stronger...

  5. Conditional Mutations in the Mitotic Chromosome Binding Function of the Bovine Papillomavirus Type 1 E2 Protein

    OpenAIRE

    Zheng, Peng-Sheng; Brokaw, Jane; Alison A McBride

    2005-01-01

    The papillomavirus E2 protein is required for viral transcriptional regulation, DNA replication and genome segregation. We have previously shown that the E2 transactivator protein and BPV1 genomes are associated with mitotic chromosomes; E2 links the genomes to cellular chromosomes to ensure efficient segregation to daughter nuclei. The transactivation domain of the E2 protein is necessary and sufficient for association of the E2 protein with mitotic chromosomes. To determine which residues o...

  6. Promotion of mitotic catastrophe via activation of PTEN by paclitaxel with supplement of mulberry water extract in bladder cancer cells

    OpenAIRE

    Nien-Cheng Chen; Charng-Cherng Chyau; Yi-Ju Lee; Hsien-Chun Tseng; Fen-Pi Chou

    2016-01-01

    Paclitaxel is a mitotic inhibitor used in cancer chemotherapy. Mulberry fruit is rich in phenolic compounds and flavonoids and exhibits chemopreventive activities. In this study, mulberry water extract (MWE) was used as a supplement to synergize with the effects of paclitaxel in the treatment of the TSGH 8301 human bladder cancer cell line. Treatment with paclitaxel combined with MWE (paclitaxel/MWE) enhanced the cytotoxicity of paclitaxel and induced severe G2/M arrest, mitotic catastrophe a...

  7. Effects of dietary fat on metabolic and structural adpatation of mitotic and postmitotic tissues to calorie restriction in mice

    OpenAIRE

    Khraiwesh, Husam M.

    2016-01-01

    In this work it is analyzed the effect of aging and its possible prevention by calorie restriction (CR) in liver and skeletal muscle from mice. These organs were selected as models of mitotic and post-mitotic tissues, respectively. Furthermore, the effect of dietary fat under CR was also investigated. For this purpose, animals submitted to CR were separated into three CR groups with soybean oil (high in ω−6 polyunsaturated fatty acids), fish oil (with a high content of polyu...

  8. Positive feedback promotes mitotic exit via the APC/C-Cdh1-separase-Cdc14 axis in budding yeast.

    Science.gov (United States)

    Hatano, Yuhki; Naoki, Koike; Suzuki, Asuka; Ushimaru, Takashi

    2016-10-01

    The mitotic inhibitor securin is degraded via the ubiquitin ligase anaphase-promoting complex/cyclosome (APC/C)-Cdc20 after anaphase onset. This triggers activation of the mitotic protease separase and thereby sister chromatid separation. However, only a proportion of securin molecules are degraded at metaphase-anaphase transition and the remaining molecules are still present in anaphase. The roles of securin and separase in late mitosis remain elusive. Here, we show that securin still inhibits separase to repress mitotic exit in anaphase in budding yeast. APC/C-Cdh1-mediated securin degradation at telophase further liberated separase, which promotes Cdc14 release and mitotic exit. Separase executed these events via its proteolytic action and that in the Cdc14 early release (FEAR) network. Cdc14 release further activated APC/C-Cdh1 in the manner of a positive feedback loop. Thus, the positive feedback promotes mitotic exit via the APC/C-Cdh1-separase-Cdc14 axis. This study shows the importance of the two-step degradation mode of securin and the role of separase in mitotic exit. PMID:27418100

  9. Using Automated Image Analysis Algorithms to Distinguish Normal, Aberrant, and Degenerate Mitotic Figures Induced by Eg5 Inhibition.

    Science.gov (United States)

    Bigley, Alison L; Klein, Stephanie K; Davies, Barry; Williams, Leigh; Rudmann, Daniel G

    2016-07-01

    Modulation of the cell cycle may underlie the toxicologic or pharmacologic responses of a potential therapeutic agent and contributes to decisions on its preclinical and clinical safety and efficacy. The descriptive and quantitative assessment of normal, aberrant, and degenerate mitotic figures in tissue sections is an important end point characterizing the effect of xenobiotics on the cell cycle. Historically, pathologists used manual counting and special staining visualization techniques such as immunohistochemistry for quantification of normal, aberrant, and degenerate mitotic figures. We designed an automated image analysis algorithm for measuring these mitotic figures in hematoxylin and eosin (H&E)-stained sections. Algorithm validation methods used data generated from a subcutaneous human transitional cell carcinoma xenograft model in nude rats treated with the cell cycle inhibitor Eg5. In these studies, we scanned and digitized H&E-stained xenografts and applied a complex ruleset of sequential mathematical filters and shape discriminators for classification of cell populations demonstrating normal, aberrant, or degenerate mitotic figures. The resultant classification system enabled the representations of three identifiable degrees of morphological change associated with tumor differentiation and compound effects. The numbers of mitotic figure variants and mitotic indices data generated corresponded to a manual assessment by a pathologist and supported automated algorithm verification and application for both efficacy and toxicity studies. PMID:26936079

  10. Delayed cell death associated with mitotic catastrophe in γ-irradiated stem-like glioma cells

    International Nuclear Information System (INIS)

    Stem-like tumor cells are regarded as highly resistant to ionizing radiation (IR). Previous studies have focused on apoptosis early after irradiation, and the apoptosis resistance observed has been attributed to reduced DNA damage or enhanced DNA repair compared to non-stem tumor cells. Here, early and late radioresponse of patient-derived stem-like glioma cells (SLGCs) and differentiated cells directly derived from them were examined for cell death mode and the influence of stem cell-specific growth factors. Primary SLGCs were propagated in serum-free medium with the stem-cell mitogens epidermal growth factor (EGF) and fibroblast growth factor-2 (FGF-2). Differentiation was induced by serum-containing medium without EGF and FGF. Radiation sensitivity was evaluated by assessing proliferation, clonogenic survival, apoptosis, and mitotic catastrophe. DNA damage-associated γH2AX as well as p53 and p21 expression were determined by Western blots. SLGCs failed to apoptose in the first 4 days after irradiation even at high single doses up to 10 Gy, but we observed substantial cell death later than 4 days postirradiation in 3 of 6 SLGC lines treated with 5 or 10 Gy. This delayed cell death was observed in 3 of the 4 SLGC lines with nonfunctional p53, was associated with mitotic catastrophe and occurred via apoptosis. The early apoptosis resistance of the SLGCs was associated with lower γH2AX compared to differentiated cells, but we found that the stem-cell culture cytokines EGF plus FGF-2 strongly reduce γH2AX levels. Nonetheless, in two p53-deficient SLGC lines examined γIR-induced apoptosis even correlated with EGF/FGF-induced proliferation and mitotic catastrophe. In a line containing CD133-positive and -negative stem-like cells, the CD133-positive cells proliferated faster and underwent more γIR-induced mitotic catastrophe. Our results suggest the importance of delayed apoptosis, associated mitotic catastrophe, and cellular proliferation for γIR-induced death of

  11. A study of the low level radiation effect on the mitotic index of the basal cells in the buccal pouch of hamsters

    Energy Technology Data Exchange (ETDEWEB)

    Kang, Byung Cheol; You, Dong Soo [Dept. of Oral and Maxillofacial Radiology, College of Dentistry, Seoul National University, Seoul (Korea, Republic of)

    1993-08-15

    The purpose of this study was to investigate the defects of the low level irradiation on the mitotic index of the basal cells in the buccal pouch of hamsters (golden hamster: APG strain). After colchicine was administrated to the hamsters through the intraperitoneal, the low level radiation (5461 mR) was exposed in the buccal pouch of hamsters. The mitotic index of the basal cells was estimated 2 hours after irradiation. The results were as follows: 1. The mean mitotic index of the control group was 4.32. 2. The mean mitotic index of the irradiated group was 2.46. 3. T-test of data in the irradiated group showed significant difference from the mitotic endex in the control group. These results suggested the lowered mitotic index of the irradiated group resulted from the low level irradiation.

  12. Plk1 Inhibition Causes Post-Mitotic DNA Damage and Senescence in a Range of Human Tumor Cell Lines

    Science.gov (United States)

    Bowman, Doug; Shinde, Vaishali; Lasky, Kerri; Shi, Judy; Vos, Tricia; Stringer, Bradley; Amidon, Ben; D'Amore, Natalie; Hyer, Marc L.

    2014-01-01

    Plk1 is a checkpoint protein whose role spans all of mitosis and includes DNA repair, and is highly conserved in eukaryotes from yeast to man. Consistent with this wide array of functions for Plk1, the cellular consequences of Plk1 disruption are diverse, spanning delays in mitotic entry, mitotic spindle abnormalities, and transient mitotic arrest leading to mitotic slippage and failures in cytokinesis. In this work, we present the in vitro and in vivo consequences of Plk1 inhibition in cancer cells using potent, selective small-molecule Plk1 inhibitors and Plk1 genetic knock-down approaches. We demonstrate for the first time that cellular senescence is the predominant outcome of Plk1 inhibition in some cancer cell lines, whereas in other cancer cell lines the dominant outcome appears to be apoptosis, as has been reported in the literature. We also demonstrate strong induction of DNA double-strand breaks in all six lines examined (as assayed by γH2AX), which occurs either during mitotic arrest or mitotic-exit, and may be linked to the downstream induction of senescence. Taken together, our findings expand the view of Plk1 inhibition, demonstrating the occurrence of a non-apoptotic outcome in some settings. Our findings are also consistent with the possibility that mitotic arrest observed as a result of Plk1 inhibition is at least partially due to the presence of unrepaired double-strand breaks in mitosis. These novel findings may lead to alternative strategies for the development of novel therapeutic agents targeting Plk1, in the selection of biomarkers, patient populations, combination partners and dosing regimens. PMID:25365521

  13. Plk1 inhibition causes post-mitotic DNA damage and senescence in a range of human tumor cell lines.

    Science.gov (United States)

    Driscoll, Denise L; Chakravarty, Arijit; Bowman, Doug; Shinde, Vaishali; Lasky, Kerri; Shi, Judy; Vos, Tricia; Stringer, Bradley; Amidon, Ben; D'Amore, Natalie; Hyer, Marc L

    2014-01-01

    Plk1 is a checkpoint protein whose role spans all of mitosis and includes DNA repair, and is highly conserved in eukaryotes from yeast to man. Consistent with this wide array of functions for Plk1, the cellular consequences of Plk1 disruption are diverse, spanning delays in mitotic entry, mitotic spindle abnormalities, and transient mitotic arrest leading to mitotic slippage and failures in cytokinesis. In this work, we present the in vitro and in vivo consequences of Plk1 inhibition in cancer cells using potent, selective small-molecule Plk1 inhibitors and Plk1 genetic knock-down approaches. We demonstrate for the first time that cellular senescence is the predominant outcome of Plk1 inhibition in some cancer cell lines, whereas in other cancer cell lines the dominant outcome appears to be apoptosis, as has been reported in the literature. We also demonstrate strong induction of DNA double-strand breaks in all six lines examined (as assayed by γH2AX), which occurs either during mitotic arrest or mitotic-exit, and may be linked to the downstream induction of senescence. Taken together, our findings expand the view of Plk1 inhibition, demonstrating the occurrence of a non-apoptotic outcome in some settings. Our findings are also consistent with the possibility that mitotic arrest observed as a result of Plk1 inhibition is at least partially due to the presence of unrepaired double-strand breaks in mitosis. These novel findings may lead to alternative strategies for the development of novel therapeutic agents targeting Plk1, in the selection of biomarkers, patient populations, combination partners and dosing regimens. PMID:25365521

  14. The Effect of Olive Oil Mill Effluent on the Mitotic Cell Division and Total Protein Amount of the Root Tips of Triticum aestivumL.

    OpenAIRE

    Aybeke, Mehmet; OLGUN, Göksel

    2000-01-01

    In this work sitotoxic and mutagenic effects Olive Oil Mill Effluent (OOME) on the root tips of Triticum aestivumL. were investigated. In this purpose, germination rate of seeds, mitotic division abnormalities and total protein amounts were evaluated. The seeds kept in various OOME concentrastions, it was determinated that germination rate decreased, whilst mitotic abnormalities and mitotic cell division frequency increased. Especially, the increased cell division frequency was of signif...

  15. Dynamic Localization of the Human Papillomavirus Type 11 Origin Binding Protein E2 through Mitosis While in Association with the Spindle Apparatus

    OpenAIRE

    Dao, Luan D.; Duffy, Aaron; Van Tine, Brian A.; Wu, Shwu-Yuan; Chiang, Cheng-Ming; Broker, Thomas R.; Chow, Louise T.

    2006-01-01

    Papillomaviral DNA replicates as extrachromosomal plasmids in squamous epithelium. Viral DNA must segregate equitably into daughter cells to persist in dividing basal/parabasal cells. We have previously reported that the viral origin binding protein E2 of human papillomavirus types 11 (HPV-11), 16, and 18 colocalized with the mitotic spindles. In this study, we show the localization of the HPV-11 E2 protein to be dynamic. It colocalized with the mitotic spindles during prophase and metaphase....

  16. Targeting Transcriptional Addictions in Small Cell Lung Cancer with a Covalent CDK7 Inhibitor

    DEFF Research Database (Denmark)

    Christensen, Camilla L; Kwiatkowski, Nicholas; Abraham, Brian J;

    2014-01-01

    Small cell lung cancer (SCLC) is an aggressive disease with high mortality, and the identification of effective pharmacological strategies to target SCLC biology represents an urgent need. Using a high-throughput cellular screen of a diverse chemical library, we observe that SCLC is sensitive to ...

  17. Targeting transcriptional addictions in small cell lung cancer with a covalent CDK7 inhibitor.

    Science.gov (United States)

    Christensen, Camilla L; Kwiatkowski, Nicholas; Abraham, Brian J; Carretero, Julian; Al-Shahrour, Fatima; Zhang, Tinghu; Chipumuro, Edmond; Herter-Sprie, Grit S; Akbay, Esra A; Altabef, Abigail; Zhang, Jianming; Shimamura, Takeshi; Capelletti, Marzia; Reibel, Jakob B; Cavanaugh, Jillian D; Gao, Peng; Liu, Yan; Michaelsen, Signe R; Poulsen, Hans S; Aref, Amir R; Barbie, David A; Bradner, James E; George, Rani E; Gray, Nathanael S; Young, Richard A; Wong, Kwok-Kin

    2014-12-01

    Small cell lung cancer (SCLC) is an aggressive disease with high mortality, and the identification of effective pharmacological strategies to target SCLC biology represents an urgent need. Using a high-throughput cellular screen of a diverse chemical library, we observe that SCLC is sensitive to transcription-targeting drugs, in particular to THZ1, a recently identified covalent inhibitor of cyclin-dependent kinase 7. We find that expression of super-enhancer-associated transcription factor genes, including MYC family proto-oncogenes and neuroendocrine lineage-specific factors, is highly vulnerability to THZ1 treatment. We propose that downregulation of these transcription factors contributes, in part, to SCLC sensitivity to transcriptional inhibitors and that THZ1 represents a prototype drug for tailored SCLC therapy. PMID:25490451

  18. Implications of mitotic and meiotic irregularities in common beans (Phaseolus vulgaris L.).

    Science.gov (United States)

    Lima, D C; Braz, G T; Dos Reis, G B; Techio, V H; Davide, L C; de F B Abreu, A

    2016-01-01

    The common bean has great social and economic importance in Brazil and is the subject of a high number of publications, especially in the fields of genetics and breeding. Breeding programs aim to increase grain yield; however, mitosis and meiosis represent under explored research areas that have a direct impact on grain yield. Therefore, the study of cell division could be another tool available to bean geneticists and breeders. The aim of this study was to investigate irregularities occurring during the cell cycle and meiosis in common bean. The common bean cultivar used was BRSMG Talismã, which owing to its high yield and grain quality is recommended for cultivation in Brazil. We classified the interphase nuclei, estimated the mitotic and meiotic index, grain pollen viability, and percentage of abnormalities in both processes. The mitotic index was 4.1%, the interphase nucleus was non-reticulated, and 19% of dividing somatic cells showed abnormal behavior. Meiosis also presented irregularities resulting in a meiotic index of 44.6%. Viability of pollen grains was 94.3%. These results indicate that the common bean cultivar BRSMG Talismã possesses repair mechanisms that compensate for changes by producing a large number of pollen grains. Another important strategy adopted by bean plants to ensure stability is the elimination of abnormal cells by apoptosis. As the common bean cultivar BRSMG Talismã is recommended for cultivation because of its good agronomic performance, it can be concluded that mitotic and meiotic irregularities have no negative influence on its grain quality and yield. PMID:27323072

  19. Aurora kinase inhibitors: a new class of drugs targeting the regulatory mitotic system.

    Science.gov (United States)

    Pérez Fidalgo, José Alejandro; Roda, Desamparados; Roselló, Susana; Rodríguez-Braun, Edith; Cervantes, Andrés

    2009-12-01

    The present review gives a perspective on the Aurora kinase family members, their function in normal cells, their role in cancer progression as well as their potential as target for anticancer treatment. Mitosis has been an important target for anticancer therapy development, leading to some specific drugs mainly addressing Tubulines, as a key structure of the mitotic spindle. Vinca alkaloids, taxanes or epotilones are good examples of conventionally developed antimitotic agents. However, novel classes of antineoplastic drugs are being studied, targeting the regulatory system that controls functional aspects of mitosis, such as Aurora or Polo-like kinases or Kinespondin inhibitors. The specific role of the different Aurora kinase proteins as regulator enzymes of the mitotic process in normal cells is discussed. Some of the mechanisms that link Aurora overexpression with cancer are also considered. Thereafter, the clinical and preclinical development of the different Aurora kinase inhibitors is presented. This is nowadays a very active area of therapeutic research and at least, sixteen new compounds are being studied as potential antineoplastic drugs. Most of them are in a very early phase of clinical development. However, we summarized the most recently published findings related with these drugs: main characteristics, way of administration, dose limiting toxicities and recommended doses for further studies. Another important aspect in Aurora kinase inhibition is the study and validation of potential biomarkers to optimize the clinical development. Several studies included pharmacodynamic assessments in normal blood cells, skin or/and tumor biopsies. Several proposals included a higher mitotic index, a decreased number of mitosis with bipolar spindles or normal alignment of chromosomes and inhibition of histone H3 phosphorylation. Future strategies and challenges for trials with Aurora kinase inhibitors are also discussed. PMID:20045785

  20. Exit from Arsenite-Induced Mitotic Arrest Is p53 Dependent

    OpenAIRE

    McNeely, Samuel C.; Xu, Xiaogiang; Taylor, B. Frazier; Zacharias, Wolfgang; McCabe, Michael J.; States, J.Christopher

    2006-01-01

    Background Arsenic is both a human carcinogen and a chemotherapeutic agent, but the mechanism of neither arsenic-induced carcinogenesis nor tumor selective cytotoxicity is clear. Using a model cell line in which p53 expression is regulated exogenously in a tetracycline-off system (TR9-7 cells), our laboratory has shown that arsenite disrupts mitosis and that p53-deficient cells [p53(−)], in contrast to p53-expressing cells [p53(+)], display greater sensitivity to arsenite-induced mitotic arre...

  1. Study on the radioprotective effect of cystamine and mexamine during two subsequent mitotic cycles

    Energy Technology Data Exchange (ETDEWEB)

    Sidorov, V.P. (Vsesoyuznyj Nauchno-ssledovatel' skij Inst. I Khimizatsii Lesnogo Khozyajstva, Pushkino (USSR))

    The radioprotective agents were found to be effective in relation to chromosomal aberrations occuring during both the first and the second mitotic cycles. It was shown that the radioprotective effect of cystamine and mexamine is completely removed by the effect of the inhibitor of DNA synthesis, 5-aminouracil. It is suggested that the radioprotective effect of the protective agents is realized through the formation of complexes between the radioprotective agent and the genetically active loci of chromosome DNA rather than through the reduction of radiation-induced DNA lesions.

  2. Mitotic chromosome loss in a radiation-sensitive strain of the yeast Saccharomyces cerevisiae

    International Nuclear Information System (INIS)

    Cells of Saccharomyces cerevisiae with mutations in the RAD52 gene have previously been shown to be defective in meiotic and mitotic recombination, in sporulation, and in repair of radiation-induced damage to DNA. In this study we show that diploid cells homozygous for rad52 lose chromosomes at high frequencies and that these frequencies of loss can be increased dramatically by exposure of these cells to x-rays. Genetic analyses of survivors of x-ray treatment demonstrate that chromosome loss events result in the conversion of diploid cells to cells with near haploid chromosome numbers

  3. Mitotic instability of Pichia pinus diploid cells 2. Segregation induced by gamma-irradiation

    International Nuclear Information System (INIS)

    Two types of genetic damages resulted from gamma rays in diploid cells of yeasts Pichia pinus MH4: nonlethal damages resulting in increasing frequency of mitotic crossing-over and lethal damages leading to cell death or arising unstable clones have been described. Survaving irradiated cells with lethal damages (which nature is not established) originate ''grown'' colonies which instability is manifested in the increased frequency of segregation of lethal sections and aneuploid segregants; in such colonies segregants without one, two or three nonhomologous chromosomes are often found. It is concluded that losses of separate chromosomes are not those primary damages which result in radiation inactivation of cells

  4. Prolonged mitotic arrest induces a caspase-dependent DNA damage response at telomeres that determines cell survival.

    Science.gov (United States)

    Hain, Karolina O; Colin, Didier J; Rastogi, Shubhra; Allan, Lindsey A; Clarke, Paul R

    2016-01-01

    A delay in the completion of metaphase induces a stress response that inhibits further cell proliferation or induces apoptosis. This response is thought to protect against genomic instability and is important for the effects of anti-mitotic cancer drugs. Here, we show that mitotic arrest induces a caspase-dependent DNA damage response (DDR) at telomeres in non-apoptotic cells. This pathway is under the control of Mcl-1 and other Bcl-2 family proteins and requires caspase-9, caspase-3/7 and the endonuclease CAD/DFF40. The gradual caspase-dependent loss of the shelterin complex protein TRF2 from telomeres promotes a DDR that involves DNA-dependent protein kinase (DNA-PK). Suppression of mitotic telomere damage by enhanced expression of TRF2, or the inhibition of either caspase-3/7 or DNA-PK during mitotic arrest, promotes subsequent cell survival. Thus, we demonstrate that mitotic stress is characterised by the sub-apoptotic activation of a classical caspase pathway, which promotes telomere deprotection, activates DNA damage signalling, and determines cell fate in response to a prolonged delay in mitosis. PMID:27230693

  5. Moderate intensity static magnetic fields affect mitotic spindles and increase the antitumor efficacy of 5-FU and Taxol.

    Science.gov (United States)

    Luo, Yan; Ji, Xinmiao; Liu, Juanjuan; Li, Zhiyuan; Wang, Wenchao; Chen, Wei; Wang, Junfeng; Liu, Qingsong; Zhang, Xin

    2016-06-01

    Microtubules are the fundamental components in mitotic spindle, which plays essential roles in cell division. It was well known that purified microtubules could be affected by static magnetic fields (SMFs) in vitro because of the diamagnetic anisotropy of tubulin. However, whether these effects lead to cell division defects was unknown. Here we find that 1T SMFs induce abnormal mitotic spindles and increase mitotic index. Synchronization experiments show that SMFs delay cell exit from mitosis and cause mitotic arrest. These mimic the cellular effects of a microtubule-targeting drug Paclitaxel (Taxol), which is frequently used in combination with 5-Fluorouracil (5-FU) and Cisplatin in cancer treatment. Using four different human cancer cell lines, HeLa, HCT116, CNE-2Z and MCF7, we find that SMFs increase the antitumor efficacy of 5-FU or 5-FU/Taxol, but not Cisplatin, which indicates that the SMF-induced combinational effects with chemodrugs are drug-specific. Our study not only reveals the effect of SMFs on microtubules to cause abnormal mitotic spindles and delay cells exit from mitosis, but also implies the potential applications of SMFs in combination with chemotherapy drugs 5-FU or 5-FU/Taxol, but not with Cisplatin in cancer treatment. PMID:26775206

  6. Cell death, chromosome damage and mitotic delay in normal human, ataxia telangiectasia and retinoblastoma fibroblasts after X-irradiation

    International Nuclear Information System (INIS)

    We recently showed (Scott and Zampetti-Bosseler 1980) that X-ray sensitive mouse lymphoma cells sustain more chromosome damage, mitotic delay and spindle defects than X-ray resistant cells. We proposed that (a) chromosome aberrations contribute much more to lethality than spindle defects, and (b) that DNA lesions are less effectively repaired in the sensitive cells and give rise to more G2 mitotic delay and chromosome aberrations. Our present results on human fibroblasts with reported differential sensitivity to ionizing radiation (i.e. normal donors and patients with ataxia telangiectasia and retinoblastoma) support the first hypothesis since we observed a positive correlation between chromosome aberration frequencies and cell killing and no induced spindle defects. Our second hypothesis is however not substantiated since X-ray sensitive fibroblasts from the ataxia patient suffered less mitotic delay than cells from normal donors. A common lesion for mitotic delay and chromosome aberrations can still be assumed by adopting the hypothesis of Painter and Young (1981) that the defect in ataxia cells is not in repair but in a failure of DNA damage to initiate mitotic delay. In contrast to other reports, we found the retinoblastoma cells to be of normal radiation sensitivity (cell killing and aberrations). (author)

  7. Taxifolin enhances andrographolide-induced mitotic arrest and apoptosis in human prostate cancer cells via spindle assembly checkpoint activation.

    Directory of Open Access Journals (Sweden)

    Zhong Rong Zhang

    Full Text Available Andrographolide (Andro suppresses proliferation and triggers apoptosis in many types of cancer cells. Taxifolin (Taxi has been proposed to prevent cancer development similar to other dietary flavonoids. In the present study, the cytotoxic and apoptotic effects of the addition of Andro alone and Andro and Taxi together on human prostate carcinoma DU145 cells were assessed. Andro inhibited prostate cancer cell proliferation by mitotic arrest and activation of the intrinsic apoptotic pathway. Although the effect of Taxi alone on DU145 cell proliferation was not significant, the combined use of Taxi with Andro significantly potentiated the anti-proliferative effect of increased mitotic arrest and apoptosis by enhancing the cleavage of poly(ADP-ribose polymerase, and caspases-7 and -9. Andro together with Taxi enhanced microtubule polymerization in vitro, and they induced the formation of twisted and elongated spindles in the cancer cells, thus leading to mitotic arrest. In addition, we showed that depletion of MAD2, a component in the spindle assembly checkpoint (SAC, alleviated the mitotic block induced by the two compounds, suggesting that they trigger mitotic arrest by SAC activation. This study suggests that the anti-cancer activity of Andro can be significantly enhanced in combination with Taxi by disrupting microtubule dynamics and activating the SAC.

  8. Centromeric Transcription Regulates Aurora-B Localization and Activation

    Directory of Open Access Journals (Sweden)

    Michael D. Blower

    2016-05-01

    Full Text Available Centromeric transcription is widely conserved; however, it is not clear what role centromere transcription plays during mitosis. Here, I find that centromeres are transcribed in Xenopus egg extracts into a long noncoding RNA (lncRNA; cen-RNA that localizes to mitotic centromeres, chromatin, and spindles. cen-RNAs bind to the chromosomal passenger complex (CPC in vitro and in vivo. Blocking transcription or antisense inhibition of cen-RNA leads to a reduction of CPC localization to the inner centromere and misregulation of CPC component Aurora-B activation independently of known centromere recruitment pathways. Additionally, transcription is required for normal bipolar attachment of kinetochores to the mitotic spindle, consistent with a role for cen-RNA in CPC regulation. This work demonstrates that cen-RNAs promote normal kinetochore function through regulation of the localization and activation of the CPC and confirm that lncRNAs are components of the centromere.

  9. The effect of mitotic inhibitors on DNA strand size and radiation-associated break repair in Down syndrome fibroblasts

    International Nuclear Information System (INIS)

    The effect of mitotic inhibitors on formation and repair of DNA breaks was studied in cultured fibroblasts from patients with Down syndrome in order to investigate the hypothesis that the karyotyping procedure itself may play a role in the increased chromosome breakage seen in these cells after gamma radiation exposure. Using the nondenaturing elution and alkaline elution techniques to examine fibroblasts from Down syndrome patients and from controls, no specific abnormalities in Down syndrome cells could be detected after exposure to mitotic inhibitors, including rate and extent of elution of DNA from filters as well as repair of radiation-induced DNA breaks. In both normal and Down syndrome cell strains, however, exposure to mitotic inhibitors was associated with a decrease in cellular DNA strand size, suggesting the presence of drug-induced DNA strand breaks. The mechanism of increased chromosome sensitivity of Down syndrome cells to gamma radiation remains unknown. (orig.)

  10. DNA lesions induced by replication stress trigger mitotic aberration and tetraploidy development.

    Directory of Open Access Journals (Sweden)

    Yosuke Ichijima

    Full Text Available During tumorigenesis, cells acquire immortality in association with the development of genomic instability. However, it is still elusive how genomic instability spontaneously generates during the process of tumorigenesis. Here, we show that precancerous DNA lesions induced by oncogene acceleration, which induce situations identical to the initial stages of cancer development, trigger tetraploidy/aneuploidy generation in association with mitotic aberration. Although oncogene acceleration primarily induces DNA replication stress and the resulting lesions in the S phase, these lesions are carried over into the M phase and cause cytokinesis failure and genomic instability. Unlike directly induced DNA double-strand breaks, DNA replication stress-associated lesions are cryptogenic and pass through cell-cycle checkpoints due to limited and ineffective activation of checkpoint factors. Furthermore, since damaged M-phase cells still progress in mitotic steps, these cells result in chromosomal mis-segregation, cytokinesis failure and the resulting tetraploidy generation. Thus, our results reveal a process of genomic instability generation triggered by precancerous DNA replication stress.

  11. Effect of propolis on mitotic and cellular proliferation indices in human blood lymphocytes

    Energy Technology Data Exchange (ETDEWEB)

    Montoro, A.; Almonacid, M.; Villaescusa, J. [Valencia Hospital Univ. la Fe, Servicio de Proteccion Radiologica (Spain); Barquinero, J. [Barcelona Univ. Autonom, Servicio de Dosimetria Biologica, Unidad de Antropologia, Dept. de Biologia Animal, Vegetal y Ecologia, barcelona (Spain); Barrios, L. [Barcelona Univ. Autonoma, Dept. de Biologia Celular y Fisiologia. Unidad de Biologia Celular (Spain); Verdu, G. [Valencia Univ. Politecnica, Dept. de Ingenieria Quimica y Nuclear (Spain); Perez, J. [Hospital la Fe, Seccion de Radiofisica, Servicio de Radioterapia, valencia (Spain)

    2006-07-01

    The study of the frequency of chromosomal aberrations per cell is the tool used in Biological dosimetry studies. Using dose-effect calibration curve obtained in our laboratory, we can evaluate the radioprotector effect of the EEP (ethanolic extract of propolis) in cultures in vitro. Propolis is the generic name for resinous substance collected by honeybees. The results showed a reduction in chromosomal aberrations's frequency of up to 50 %. The following study consisted of analyzing human peripheral blood lymphocytes exposed to 2 Gy {gamma} rays, in presence and absence of EEP, the change in the frequency of chromosome aberrations was analysed with biological dosimetry. The protection against the formation of dicentric and ring was dose-dependent, but there seemed to be a maximum protection, i.e. a further increase in the concentration of EEP does not show additional protection. This work studies the effect of the EEP of the cellular cycle using the mitotic and cellular proliferation index, as an alternative for the screening cytostatic activity. The results indicate that the lymphocytes which were cultures in presence of EEP exhibited a significant and dependent-concentration decrease in mitotic index and proliferation kinetics. The possible mechanisms involved in the radioprotective influence of EEP are discussed. (authors)

  12. Visualization of the chromosome scaffold and intermediates of loop domain compaction in extracted mitotic cells.

    Science.gov (United States)

    Sheval, Eugene V; Polyakov, Vladimir Y

    2006-12-01

    A novel extraction protocol for cells cultured on coverslips is described. Observations of the extraction process in a perfusion chamber reveal that cells of all mitotic stages are not detached from coverslips during extraction, and all stages can be recognized using phase contrast images. We studied the extracted cell morphology and distribution of a major scaffold component - topoisomerase IIalpha, in extracted metaphase and anaphase cells. An extraction using 2M NaCl leads to destruction of chromosomes at the light microscope level. Immunogold studies demonstrate that the only residual structure observed is an axial chromosome scaffold that contains topoisomerase IIalpha. In contrast, mitotic chromosomes are swelled only partially after an extraction using dextran sulphate and heparin, and it appears that this treatment does not lead to total destruction of loop domains. In this case, the chromosome scaffold and numerous structures resembling small rosettes are revealed inside extracted cells. The rosettes observed condense after addition of Mg2+-ions and do not contain topoisomerase IIalpha suggesting that these structures correspond to intermediates of loop domain compaction. We propose a model of chromosome structure in which the loop domains are condensed into highly regular structures with rosette organization. PMID:17029868

  13. The effect of x-ray induced mitotic delay on chromosome aberration yields in human lymphocytes

    International Nuclear Information System (INIS)

    The extent to which X-ray induced mitotic delay at 150 and 400 rad influences chromosome aberration yields was examined in human peripheral blood lymphocytes. The dicentric was used as a marker and aberration yields were obtained for mixed cultures prepared from equal numbers of normal and irradiated cells. The cultures were terminated following incubation times of 36-120 h. Greater mitotic delay of the order of a few hours was observed at the higher dose. However most reduction in the numbers of lymphocytes arriving at metaphase by 48 h may be ascribed to interphase death of failure to transform. Analysis of the dicentric distributions which were expected to follow Poisson statistics indicated that cells containing dicentrics were delayed relative to irradiated but aberration-free cells. Cells with one dicentric moved more easily through the first cell cycle than cells containing two dicentrics. Following accidental partial body irradiation, selection in culture favouring the unirradiated lymphocytes does not distort the aberration yield sufficiently to warrant incubation times in excess of the standard 48-52 h

  14. Effect of propolis on mitotic and cellular proliferation indices in human blood lymphocytes

    International Nuclear Information System (INIS)

    The study of the frequency of chromosomal aberrations per cell is the tool used in Biological dosimetry studies. Using dose-effect calibration curve obtained in our laboratory, we can evaluate the radioprotector effect of the EEP (ethanolic extract of propolis) in cultures in vitro. Propolis is the generic name for resinous substance collected by honeybees. The results showed a reduction in chromosomal aberrations's frequency of up to 50 %. The following study consisted of analyzing human peripheral blood lymphocytes exposed to 2 Gy γ rays, in presence and absence of EEP, the change in the frequency of chromosome aberrations was analysed with biological dosimetry. The protection against the formation of dicentric and ring was dose-dependent, but there seemed to be a maximum protection, i.e. a further increase in the concentration of EEP does not show additional protection. This work studies the effect of the EEP of the cellular cycle using the mitotic and cellular proliferation index, as an alternative for the screening cytostatic activity. The results indicate that the lymphocytes which were cultures in presence of EEP exhibited a significant and dependent-concentration decrease in mitotic index and proliferation kinetics. The possible mechanisms involved in the radioprotective influence of EEP are discussed. (authors)

  15. The Emerging Nexus of Active DNA Demethylation and Mitochondrial Oxidative Metabolism in Post-Mitotic Neurons

    Directory of Open Access Journals (Sweden)

    Huan Meng

    2014-12-01

    Full Text Available The variable patterns of DNA methylation in mammals have been linked to a number of physiological processes, including normal embryonic development and disease pathogenesis. Active removal of DNA methylation, which potentially regulates neuronal gene expression both globally and gene specifically, has been recently implicated in neuronal plasticity, learning and memory processes. Model pathways of active DNA demethylation involve ten-eleven translocation (TET methylcytosine dioxygenases that are dependent on oxidative metabolites. In addition, reactive oxygen species (ROS and oxidizing agents generate oxidative modifications of DNA bases that can be removed by base excision repair proteins. These potentially link the two processes of active DNA demethylation and mitochondrial oxidative metabolism in post-mitotic neurons. We review the current biochemical understanding of the DNA demethylation process and discuss its potential interaction with oxidative metabolism. We then summarise the emerging roles of both processes and their interaction in neural plasticity and memory formation and the pathophysiology of neurodegeneration. Finally, possible therapeutic approaches for neurodegenerative diseases are proposed, including reprogramming therapy by global DNA demethylation and mitohormesis therapy for locus-specific DNA demethylation in post-mitotic neurons.

  16. Post-mitotic role of nucleostemin as a promoter of skeletal muscle cell differentiation

    Energy Technology Data Exchange (ETDEWEB)

    Hirai, Hiroyuki; Romanova, Liudmila; Kellner, Steven; Verma, Mayank; Rayner, Samuel [Stem Cell Institute, University of Minnesota, Room 2-216, MTRF, 2001 6th St. SE, Minneapolis, MN 55455 (United States); Asakura, Atsushi, E-mail: asakura@umn.edu [Stem Cell Institute, University of Minnesota, Room 2-216, MTRF, 2001 6th St. SE, Minneapolis, MN 55455 (United States); Kikyo, Nobuaki, E-mail: kikyo001@umn.edu [Stem Cell Institute, University of Minnesota, Room 2-216, MTRF, 2001 6th St. SE, Minneapolis, MN 55455 (United States)

    2010-01-01

    Nucleostemin (NS) is a nucleolar protein abundantly expressed in a variety of proliferating cells and undifferentiated cells. Its known functions include cell cycle regulation and the control of pre-rRNA processing. It also has been proposed that NS has an additional role in undifferentiated cells due to its downregulation during stem cell differentiation and its upregulation during tissue regeneration. Here, however, we demonstrate that skeletal muscle cell differentiation has a unique expression profile of NS in that it is continuously expressed during differentiation. NS was expressed at similar levels in non-proliferating muscle stem cells (satellite cells), rapidly proliferating precursor cells (myoblasts) and post-mitotic terminally differentiated cells (myotubes and myofibers). The sustained expression of NS during terminal differentiation is necessary to support increased protein synthesis during this process. Downregulation of NS inhibited differentiation of myoblasts to myotubes, accompanied by striking downregulation of key myogenic transcription factors, such as myogenin and MyoD. In contrast, upregulation of NS inhibited proliferation and promoted muscle differentiation in a p53-dependent manner. Our findings provide evidence that NS has an unexpected role in post-mitotic terminal differentiation. Importantly, these findings also indicate that, contrary to suggestions in the literature, the expression of NS cannot always be used as a reliable indicator for undifferentiated cells or proliferating cells.

  17. Multisite phosphorylation of Pin1-associated mitotic phosphoproteins revealed by monoclonal antibodies MPM-2 and CC-3

    Directory of Open Access Journals (Sweden)

    Vincent Michel

    2004-06-01

    Full Text Available Abstract Background The peptidyl-prolyl isomerase Pin1 recently revealed itself as a new player in the regulation of protein function by phosphorylation. Pin1 isomerizes the peptide bond of specific phosphorylated serine or threonine residues preceding proline in several proteins involved in various cellular events including mitosis, transcription, differentiation and DNA damage response. Many Pin1 substrates are antigens of the phosphodependent monoclonal antibody MPM-2, which reacts with a subset of proteins phosphorylated at the G2/M transition. Results As MPM-2 is not a general marker of mitotic phosphoproteins, and as most mitotic substrates are phosphorylated more than once, we used a different phosphodependent antibody, mAb CC-3, to identify additional mitotic phosphoproteins and eventual Pin1 substrates by combining affinity purification, MALDI-TOF mass spectrometry and immunoblotting. Most CC-3-reactive phosphoproteins appeared to be known or novel MPM-2 antigens and included the RNA-binding protein p54nrb/nmt55, the spliceosomal protein SAP155, the Ki-67 antigen, MAP-1B, DNA topoisomerases II α and β, the elongation factor hSpt5 and the largest subunit of RNA polymerase II. The CC-3 mitotic antigens were also shown to be Pin1 targets. The fine CC-3- and MPM-2-epitope mapping of the RNA polymerase II carboxy-terminal domain confirmed that the epitopes were different and could be generated in vitro by distinct kinases. Finally, the post-mitotic dephosphorylation of both CC-3 and MPM-2 antigens was prevented when cellular Pin1 activity was blocked by the selective inhibitor juglone. Conclusion These observations indicate that the mitotic phosphoproteins associated with Pin1 are phosphorylated on multiple sites, suggesting combinatorial regulation of substrate recognition and isomerization.

  18. Influence of irradiation at different stages of mitotic cycle upon production of sister chromatid exchanges in cultured Chinese hamster cells

    International Nuclear Information System (INIS)

    Frequency of sister chromatid exchanges (SCE) and microexchanges in Chinese hamster cells has been studied by means of the method of differential staining of chromatids on irradiation at different stages of the mitotic cycle. It is shown that the irradiation enhances frequency of SCE and microexchanges if it is carried out before the end of DNA replication synthesis. Comparison of frequency depenedence of radiation-induced microexchanges and SCE at different stages of the mitotic cycle results in the conclusion that the microexchanges are none other than small SCE

  19. Influence of irradiation at different stages of mitotic cycle unon production of sister chromatid exchanges in cultured chinese hamster cells

    International Nuclear Information System (INIS)

    Frequency of.sister chromatid exchanges (SCE) and microexchanges in the chinese hamster cells was investigated by means of the method of differential colouring of chromatids on irradiation at different stages of the mitotic cycle. It is shown that the irradiation increases frequency of SCE and microexchanges if it is performed before the end of the replicative DNA synthesis. Comparison of frequency dependence of radiation-induced microexchanges and SCE at different stages of the mitotic cycle permits to conclude that the microexchanges are small SCE

  20. Mitotic inhibition of ICR 2A frog cells exposed to 265-313 nm monochromatic ultraviolet wavelengths and photoreactivating light

    International Nuclear Information System (INIS)

    Exposure of ICR 2A frog cells to 265, 289, 302 or 313 nm U.V. radiation caused a decrease in the MI of the irradiated cells in a fluence-dependent fashion. Treatment of cells with PRL immediately after U.V.-irradiation resulted in a smaller decrease in the MI, demonstrating that pyrimidine dimers played a role in the mitotic inhibition induced by these U.V. wavelengths. The effect of PRL on 313 nm-irradiated cells was much smaller than for the other wavelengths tested, indicating that non-dimer photoproducts were of importance in the mitotic inhibition induced by this U.V. wavelength. (author)

  1. High throughput screening of natural products for anti-mitotic effects in MDA-MB-231 human breast carcinoma cells

    OpenAIRE

    Mazzio, E; Badisa, R; Mack, N; Deiab, S.; Soliman, KFA

    2013-01-01

    Some of the most effective anti-mitotic microtubule-binding agents, such as paclitaxel (Taxus brevifolia) were originally discovered through robust NCI botanical screenings. In this study, a high-through microarray format was utilized to screen 897 aqueous extracts of commonly used natural products (0.00015–0.5 mg/ml) relative to paclitaxel for anti-mitotic effects (independent of toxicity) on proliferation of MDA-MB-231 cells. The data obtained showed that less than 1.34 % tested showed inhi...

  2. Autoradiographic studies on the cell kinetics after the whole body X-irradiation. 3. Post-irradiation changes in the mitotic activity and mitotic cycle of the rat's brain subependymal cells

    Energy Technology Data Exchange (ETDEWEB)

    Gracheva, N.D. (Tsentral' nyj Nauchno-Issledovatel' skij Rentgeno-Radiologicheskij Inst., Leningrad (USSR))

    1982-04-01

    The study is carried out on subependymal cells of rat brain usig autoradiography with 3H-tymidine introduced 60-80 min before whole body X-ray irradiation i doses 50, 150, or 300 R. In postradiation cell system the dynamics of mitotic index Isub(M) and changes of mitotic cycle aevaluated according to the curve of labelled mitosis (CLM) are studied. A new interpretation of ''classical delay of mitosis'' is given, which points not to excessive delay in phases of mitotic cycle but to dependence on dose time from the irradiation moment before appearance of the first mitoses of survived cells as cells which were dividing at the time of death. At that, all the cells of the system have experienced one hour block independent of the dose. It is showm that the curve Isub(M) can serve as a measure of dead proliferative cells. The changes of CLM, conditioned by mitotic deathe f cells irradiated in G2-and S-phases and its other peculiarities are discussed.

  3. Effects of the rad52 gene on recombination in Saccharomyces cerevisiae. [Comparison of. gamma. -, uv-induced meiotic and spontaneous mitotic recombination

    Energy Technology Data Exchange (ETDEWEB)

    Prakash, S.; Prakash, L.; Burke, W.; Montelone, B.A.

    1979-01-01

    Effects of the rad52 mutation in Saccharomyces cerevisiae on meiotic, ..gamma..-ray-induced, uv-induced, and spontaneous mitotic recombination were studied. The rad52/rad52 diploids undergo premeiotic DNA synthesis; sporulation occurs but inviable spores are produced. Intra- and intergenic recombination during meiosis were examined in cells transferred from sporulation medium to vegetative medium at different time intervals. No intragenic recombination was observed at the hisl-1/hisl-315 and trp5-2/trp5-48 heteroalleles. Gene-centromere recombination was also not observed in rad52/rad52 diploids. No ..gamma..-ray-induced intragenic mitotic recombination is seen in rad52/rad52 diploids and uv-induced intragenic recombination is greatly reduced. However, spontaneous mitotic recombination is not similarly affected. The RAD52 gene thus functions in recombination in meiosis and in ..gamma..-ray and uv-induced mitotic recombination but not in spontaneous mitotic recombination.

  4. Acrylamide effects on kinesin-related proteins of the mitotic/meiotic spindle

    International Nuclear Information System (INIS)

    The microtubule (MT) motor protein kinesin is a vital component of cells and organs expressing acrylamide (ACR) toxicity. As a mechanism of its potential carcinogenicity, we determined whether kinesins involved in cell division are inhibited by ACR similar to neuronal kinesin [Sickles, D.W., Brady, S.T., Testino, A.R., Friedman, M.A., and Wrenn, R.A. (1996). Direct effect of the neurotoxicant acrylamide on kinesin-based microtubule motility. Journal of Neuroscience Research 46, 7-17.] Kinesin-related genes were isolated from rat testes [Navolanic, P.M., and Sperry, A.O. (2000). Identification of isoforms of a mitotic motor in mammalian spermatogenesis. Biology of Reproduction 62, 1360-1369.], their kinesin-like proteins expressed in bacteria using recombinant DNA techniques and the effects of ACR, glycidamide (GLY) and propionamide (a non-neurotoxic metabolite) on the function of two of the identified kinesin motors were tested. KIFC5A MT bundling activity, required for mitotic spindle formation, was measured in an MT-binding assay. Both ACR and GLY caused a similar concentration-dependent reduction in the binding of MT; concentrations of 100 μM ACR or GLY reduced its activity by 60%. KRP2 MT disassembling activity was assayed using the quantity of tubulin disassembled from taxol-stabilized MT. Both ACR and GLY inhibited KRP2-induced MT disassembly. GLY was substantially more potent; significant reductions of 60% were achieved by 500 μM, a comparable inhibition by ACR required a 5 mM concentration. Propionamide had no significant effect on either kinesin, except KRP2 at 10 mM. This is the first report of ACR inhibition of a mitotic/meiotic motor protein. ACR (or GLY) inhibition of kinesin may be an alternative mechanism to DNA adduction in the production of cell division defects and potential carcinogenicity. We conclude that ACR may act on multiple kinesin family members and produce toxicities in organs highly dependent on microtubule-based functions

  5. Cremophor EL stimulates mitotic recombination in uvsH//uvsH diploid strain of Aspergillus nidulans

    Directory of Open Access Journals (Sweden)

    Cleverson Busso

    2004-03-01

    Full Text Available Cremophor EL is a solubilizer and emulsifier agent used in the pharmaceutical and foodstuff industries. The solvent is the principal constituent of paclitaxel's clinical formulation vehicle. Since mitotic recombination plays a crucial role in multistep carcinogenesis, the study of the recombinagenic potential of chemical compounds is of the utmost importance. In our research genotoxicity of cremophor EL has been studied by using an uvsH//uvsH diploid strain of Aspergillus nidulans. Since it spends a great part of its cell cycle in the G2period, this fungus is a special screening system for the study of mitotic recombination induced by chemical substances. Homozygotization Indexes (HI for paba and bi markers from heterozygous B211//A837 diploid strain were determined for the evaluation of the recombinagenic effect of cremophor EL. It has been shown that cremophor EL induces increase in mitotic crossing-over events at nontoxic concentrations (0.05 and 0.075% v/v.Cremofor EL (CEL é um solubilizante e emulsificante amplamente utilizado nas indústrias farmacêuticas e de gêneros alimentícios. É o principal veículo empregado nas formulações clínicas do antineoplásico paclitaxel. Considerando-se que a recombinação mitótica desempenha importante função no processo de carcinogênese, o estudo de substâncias químicas com potencial recombinagênico assume importância crucial, no sentido de se detectar aquelas que eventualmente possam atuar como promotoras de neoplasias. A genotoxicidade do cremofor EL foi estudada no presente trabalho, utilizando-se uma linhagem diplóide uvsH//uvsH de Aspergillus nidulans. Neste fungo as células vegetativas comumente repousam no período G2 do ciclo celular, facilitando a ocorrência da recombinação mitótica. O efeito recombinagênico do CEL foi avaliado através da determinação dos Índices de Homozigotização para os marcadores nutricionais paba e bi do diplóide heterozigoto B211//A837. Os

  6. Inhibition of the Ras-ERK pathway in mitotic COS7 cells is due to the inability of EGFR/Raf to transduce EGF signaling to downstream proteins.

    Science.gov (United States)

    Shi, Huaiping; Zhang, Tianying; Yi, Yongqing; Ma, Yue

    2016-06-01

    Although previous studies have shown that Ras-ERK signaling in mitosis is closed due to the inhibition of signal transduction, the events involved in the molecular mechanisms are still unclear. In the present study, we investigated the Ras-ERK signaling pathway in mitotic COS7 cells. The results demonstrated that treatment with epidermal growth factor (EGF) failed to increase the endocytosis of EGF-EGFR (EGF receptor) complexes in mitotic COS7 cells, although a large amount of endosomes were found in asynchronous COS7 cells. Clathrin expression levels in mitotic COS7 cells were inhibited whereas caveolin expression levels in mitotic COS7 cells were almost unaffected. Y1068 and Y1086 residues of EGFR in the mitotic COS7 cells were activated. However, Grb2 and Shc in the mitotic COS7 cells did not bind to activated EGFR. Ras activity was inhibited in the mitotic COS7 cells whereas its downstream protein, Raf, was obviously phosphorylated by EGF in mitosis. Treatment with phorbol 12-myristate 13-acetate (PMA) also increased the phosphorylation levels of Raf in the mitotic COS7 cells. Nevertheless, Raf phosphorylation in mitosis was significantly inhibited by AG1478. Lastly, activation of EGF-mediated MEK and ERK in the mitotic COS7 cells was obviously inhibited. In summary, our results suggest that the Ras-ERK pathway is inhibited in mitotic COS7 cells which may be the dual result of the difficulty in the transduction of EGF signaling by EGFR or Raf to downstream proteins. PMID:27004682

  7. Proteomic profiling revealed the functional networks associated with mitotic catastrophe of HepG2 hepatoma cells induced by 6-bromine-5-hydroxy-4-methoxybenzaldehyde

    International Nuclear Information System (INIS)

    Mitotic catastrophe, a form of cell death resulting from abnormal mitosis, is a cytotoxic death pathway as well as an appealing mechanistic strategy for the development of anti-cancer drugs. In this study, 6-bromine-5-hydroxy-4-methoxybenzaldehyde was demonstrated to induce DNA double-strand break, multipolar spindles, sustain mitotic arrest and generate multinucleated cells, all of which indicate mitotic catastrophe, in human hepatoma HepG2 cells. We used proteomic profiling to identify the differentially expressed proteins underlying mitotic catastrophe. A total of 137 differentially expressed proteins (76 upregulated and 61 downregulated proteins) were identified. Some of the changed proteins have previously been associated with mitotic catastrophe, such as DNA-PKcs, FoxM1, RCC1, cyclin E, PLK1-pT210, 14-3-3σ and HSP70. Multiple isoforms of 14-3-3, heat-shock proteins and tubulin were upregulated. Analysis of functional significance revealed that the 14-3-3-mediated signaling network was the most significantly enriched for the differentially expressed proteins. The modulated proteins were found to be involved in macromolecule complex assembly, cell death, cell cycle, chromatin remodeling and DNA repair, tubulin and cytoskeletal organization. These findings revealed the overall molecular events and functional signaling networks associated with spindle disruption and mitotic catastrophe. - Graphical abstract: Display Omitted Research highlights: → 6-bromoisovanillin induced spindle disruption and sustained mitotic arrest, consequently resulted in mitotic catastrophe. → Proteomic profiling identified 137 differentially expressed proteins associated mitotic catastrophe. → The 14-3-3-mediated signaling network was the most significantly enriched for the altered proteins. → The macromolecule complex assembly, cell cycle, chromatin remodeling and DNA repair, tubulin organization were also shown involved in mitotic catastrophe.

  8. Mdb1, a fission yeast homolog of human MDC1, modulates DNA damage response and mitotic spindle function.

    Directory of Open Access Journals (Sweden)

    Yi Wei

    Full Text Available During eukaryotic DNA damage response (DDR, one of the earliest events is the phosphorylation of the C-terminal SQ motif of histone H2AX (H2A in yeasts. In human cells, phosphorylated H2AX (γH2AX is recognized by MDC1, which serves as a binding platform for the accumulation of a myriad of DDR factors on chromatin regions surrounding DNA lesions. Despite its important role in DDR, no homolog of MDC1 outside of metazoans has been described. Here, we report the characterization of Mdb1, a protein from the fission yeast Schizosaccharomyces pombe, which shares significant sequence homology with human MDC1 in their C-terminal tandem BRCT (tBRCT domains. We show that in vitro, recombinant Mdb1 protein binds a phosphorylated H2A (γH2A peptide, and the phospho-specific binding requires two conserved phospho-binding residues in the tBRCT domain of Mdb1. In vivo, Mdb1 forms nuclear foci at DNA double strand breaks (DSBs induced by the HO endonuclease and ionizing radiation (IR. IR-induced Mdb1 focus formation depends on γH2A and the phospho-binding residues of Mdb1. Deleting the mdb1 gene does not overtly affect DNA damage sensitivity in a wild type background, but alters the DNA damage sensitivity of cells lacking another γH2A binder Crb2. Overexpression of Mdb1 causes severe DNA damage sensitivity in a manner that requires the interaction between Mdb1 and γH2A. During mitosis, Mdb1 localizes to spindles and concentrates at spindle midzones at late mitosis. The spindle midzone localization of Mdb1 requires its phospho-binding residues, but is independent of γH2A. Loss of Mdb1 or mutating its phospho-binding residues makes cells more resistant to the microtubule depolymerizing drug thiabendazole. We propose that Mdb1 performs dual roles in DDR and mitotic spindle regulation.

  9. Polyglutamylated Tubulin Binding Protein C1orf96/CSAP Is Involved in Microtubule Stabilization in Mitotic Spindles

    Science.gov (United States)

    Ohta, Shinya; Hamada, Mayako; Sato, Nobuko; Toramoto, Iyo

    2015-01-01

    The centrosome-associated C1orf96/Centriole, Cilia and Spindle-Associated Protein (CSAP) targets polyglutamylated tubulin in mitotic microtubules (MTs). Loss of CSAP causes critical defects in brain development; however, it is unclear how CSAP association with MTs affects mitosis progression. In this study, we explored the molecular mechanisms of the interaction of CSAP with mitotic spindles. Loss of CSAP caused MT instability in mitotic spindles and resulted in mislocalization of Nuclear protein that associates with the Mitotic Apparatus (NuMA), with defective MT dynamics. Thus, CSAP overload in the spindles caused extensive MT stabilization and recruitment of NuMA. Moreover, MT stabilization by CSAP led to high levels of polyglutamylation on MTs. MT depolymerization by cold or nocodazole treatment was inhibited by CSAP binding. Live-cell imaging analysis suggested that CSAP-dependent MT-stabilization led to centrosome-free MT aster formation immediately upon nuclear envelope breakdown without γ-tubulin. We therefore propose that CSAP associates with MTs around centrosomes to stabilize MTs during mitosis, ensuring proper bipolar spindle formation and maintenance. PMID:26562023

  10. The transforming parasite Theileria co-opts host cell mitotic and central spindles to persist in continuously dividing cells.

    Directory of Open Access Journals (Sweden)

    Conrad von Schubert

    Full Text Available The protozoan parasite Theileria inhabits the host cell cytoplasm and possesses the unique capacity to transform the cells it infects, inducing continuous proliferation and protection against apoptosis. The transforming schizont is a multinucleated syncytium that resides free in the host cell cytoplasm and is strictly intracellular. To maintain transformation, it is crucial that this syncytium is divided over the two daughter cells at each host cell cytokinesis. This process was dissected using different cell cycle synchronization methods in combination with the targeted application of specific inhibitors. We found that Theileria schizonts associate with newly formed host cell microtubules that emanate from the spindle poles, positioning the parasite at the equatorial region of the mitotic cell where host cell chromosomes assemble during metaphase. During anaphase, the schizont interacts closely with host cell central spindle. As part of this process, the schizont recruits a host cell mitotic kinase, Polo-like kinase 1, and we established that parasite association with host cell central spindles requires Polo-like kinase 1 catalytic activity. Blocking the interaction between the schizont and astral as well as central spindle microtubules prevented parasite segregation between the daughter cells during cytokinesis. Our findings provide a striking example of how an intracellular eukaryotic pathogen that evolved ways to induce the uncontrolled proliferation of the cells it infects usurps the host cell mitotic machinery, including Polo-like kinase 1, one of the pivotal mitotic kinases, to ensure its own persistence and survival.

  11. Par1b induces asymmetric inheritance of plasma membrane domains via LGN-dependent mitotic spindle orientation in proliferating hepatocytes.

    Directory of Open Access Journals (Sweden)

    Christiaan L Slim

    2013-12-01

    Full Text Available The development and maintenance of polarized epithelial tissue requires a tightly controlled orientation of mitotic cell division relative to the apical polarity axis. Hepatocytes display a unique polarized architecture. We demonstrate that mitotic hepatocytes asymmetrically segregate their apical plasma membrane domain to the nascent daughter cells. The non-polarized nascent daughter cell can form a de novo apical domain with its new neighbor. This asymmetric segregation of apical domains is facilitated by a geometrically distinct "apicolateral" subdomain of the lateral surface present in hepatocytes. The polarity protein partitioning-defective 1/microtubule-affinity regulating kinase 2 (Par1b/MARK2 translates this positional landmark to cortical polarity by promoting the apicolateral accumulation of Leu-Gly-Asn repeat-enriched protein (LGN and the capture of nuclear mitotic apparatus protein (NuMA-positive astral microtubules to orientate the mitotic spindle. Proliferating hepatocytes thus display an asymmetric inheritance of their apical domains via a mechanism that involves Par1b and LGN, which we postulate serves the unique tissue architecture of the developing liver parenchyma.

  12. Curcumin-treated cancer cells show mitotic disturbances leading to growth arrest and induction of senescence phenotype.

    Science.gov (United States)

    Mosieniak, Grażyna; Sliwinska, Małgorzata A; Przybylska, Dorota; Grabowska, Wioleta; Sunderland, Piotr; Bielak-Zmijewska, Anna; Sikora, Ewa

    2016-05-01

    Cellular senescence is recognized as a potent anticancer mechanism that inhibits carcinogenesis. Cancer cells can also undergo senescence upon chemo- or radiotherapy. Curcumin, a natural polyphenol derived from the rhizome of Curcuma longa, shows anticancer properties both in vitro and in vivo. Previously, we have shown that treatment with curcumin leads to senescence of human cancer cells. Now we identified the molecular mechanism underlying this phenomenon. We observed a time-dependent accumulation of mitotic cells upon curcumin treatment. The time-lapse analysis proved that those cells progressed through mitosis for a significantly longer period of time. A fraction of cells managed to divide or undergo mitotic slippage and then enter the next phase of the cell cycle. Cells arrested in mitosis had an improperly formed mitotic spindle and were positive for γH2AX, which shows that they acquired DNA damage during prolonged mitosis. Moreover, the DNA damage response pathway was activated upon curcumin treatment and the components of this pathway remained upregulated while cells were undergoing senescence. Inhibition of the DNA damage response decreased the number of senescent cells. Thus, our studies revealed that the induction of cell senescence upon curcumin treatment resulted from aberrant progression through the cell cycle. Moreover, the DNA damage acquired by cancer cells, due to mitotic disturbances, activates an important molecular mechanism that determines the potential anticancer activity of curcumin. PMID:26916504

  13. The participation of elevated levels of cyclic GMP in the recovery from radiation-induced mitotic delay

    International Nuclear Information System (INIS)

    The levels of cyclic AMP and cyclic GMP have been measured in Physarum plasmodia before and after treatment with gamma-radiation, 2 mM caffeine, or combinations of the two agents compared to the length of the radiation-induced mitotic delay. Caffeine alone produces a rapid transient elevation of cyclic AMP and a slower delayed elevation of cyclic GMP. Irradiation elicits an immediate transient increase in cyclic AMP and a later cyclic GMP increase which accompanies or precedes the delayed mitosis. A composite pattern is produced by combinations of radiation and caffeine, a distinctive feature of which is an elevated level of cyclic GMP near the time of the radiation-delayed and caffeine-promoted mitosis. With pretreatment by caffeine, the least radiation-induced mitotic delay occurs when plasmodia are irradiated during the caffeine-elicited increase in cyclic GMP. The plasmodium becomes refractory to the reduction of mitotic delay by caffeine at approximately the time it becomes refractory to the further elevation of cyclic GMP by caffeine. The data support a role for cyclic AMP in the onset of and for cyclic GMP in the recovery from mitotic delay induced by ionizing radiation. (author)

  14. Draft Genome Sequences of Supercritical CO[subscript 2]-Tolerant Bacteria Bacillus subterraneus MITOT1 and Bacillus cereus MIT0214

    OpenAIRE

    Peet, Kyle C.; Thompson, Janelle R.

    2015-01-01

    We report draft genome sequences of Bacillus subterraneus MITOT1 and Bacillus cereus MIT0214 isolated through enrichment of samples from geologic sequestration sites in pressurized bioreactors containing a supercritical (sc) CO[subscript 2] headspace. Their genome sequences expand the phylogenetic range of sequenced bacilli and allow characterization of molecular mechanisms of scCO[subscript 2] tolerance.

  15. Increase in mitotic recombination in diploid cells of Aspergillus nidulans in response to ethidium bromide

    Directory of Open Access Journals (Sweden)

    Tânia C.A. Becker

    2003-01-01

    Full Text Available Ethidium bromide (EB is an intercalating inhibitor of topoisomerase II and its activities are related to chemotherapeutic drugs used in anti-cancer treatments. EB promotes several genotoxic effects in exposed cells by stabilising the DNA-enzyme complex. The recombinagenic potential of EB was evaluated in our in vivo study by the loss of heterozygosity of nutritional markers in diploid Aspergillus nidulans cells through Homozygotization Index (HI. A DNA repair mutation, uvsZ and a chromosome duplication DP (II-I were introduced in the genome of tested cells to obtain a sensitive system for the recombinagenesis detection. EB-treated diploid cells had HI values significantly greater than the control at both concentrations (4.0 x 10-3 and 5.0 x 10-3 mM. Results indicate that the intercalating agent is potentially capable of inducing mitotic crossing-over in diploid A. nidulans cells.

  16. Effect of caffeine on radiation-induced mitotic delay: delayed expression of G2 arrest

    International Nuclear Information System (INIS)

    In the presence of 5 mM caffeine, irradiated (1.5 Gy) S and G2 cells progressed to mitosis in register and without arrest in G2. Caffeine (5 mM) markedly reduced mitotic delay even after radiation doses up to 20 Gy. When caffeine was removed from irradiated (1.5 Gy) and caffeine-treated cells, a period of G2 arrest followed, similar in length to that produced by radiation alone. The arrest expressed was independent of the duration of the caffeine treatment for exposures up to 3 hr. The similarity of the response to the cited effects of caffeine on S-phase delay suggests a common basis for delay induction in S and G2 phases

  17. Radiosensitivity of chromosomes in two successive mitotic cycles of human lymphocytes

    Energy Technology Data Exchange (ETDEWEB)

    Luchnik, N.V.; Poryadkova, N.A.

    1988-11-01

    A culture of human lymphocytes was irradiated with /gamma/-quanta in a dose of 0.5 Gy with different ratios of cells in first (M1) and second (M2) mitotic cycle and the frequency of aberrations induced at stage G2 was analyzed. With increase in interval of time between the start of culturing and irradiation, total yield of aberrations increased in a regular way. However, if the M1:M2 ratio is considered, then it turns out that in M2 chromosomes are /approximately/1.5 times more sensitive than in M1: within the limits of each cycle, radiosensitivity is constant and does not depend on its duration. It was established in accordance with data of other authors that 5-bromodeoxyuridine (5-BdU) increases radiosensitivity materially.

  18. Technological exploration of BAC-FISH on mitotic chromosomes of maize

    Institute of Scientific and Technical Information of China (English)

    Yongsheng TAO; Zuxin ZHANG; Yonglin CHEN; Lijia LI; Yonglian ZHENG

    2008-01-01

    The rice BAC-DNA was used as probes and fluorescence in situ hybridization (FISH) was applied to the interphase and metaphase mitotic chromosomes of maize. To optimize the BAC-FISH technique, we respect-ively assayed the effect of several factors, including maize or rice genomic Cot DNA used as blocking reagent of DNA, washing temperatures and FAD concentration in the washing buffer and in the hybrid solution. The results show that Cot DNA of maize genome blocked the repet-itive sequence of the rice BAC-DNA when the Cot value was below 50. Meanwhile, it was necessary to adjust the Cot value according to the different probes and their ratios. Decreasing the concentration of FAD in the hybridization mixtures, adjusting the washing rate after hybridization, and most especially, blocking the rice-specific repetitive sequences of BAC-DNA could improve the positive signals of BAC-FISH.

  19. Characterization of TcCYC6 from Trypanosoma cruzi, a gene with homology to mitotic cyclins.

    Science.gov (United States)

    Di Renzo, María Agostina; Laverrière, Marc; Schenkman, Sergio; Wehrendt, Diana Patricia; Tellez-Iñón, María Teresa; Potenza, Mariana

    2016-06-01

    Trypanosoma cruzi, the etiologic agent of Chagas disease, is a protozoan parasite with a life cycle that alternates between replicative and non-replicative forms, but the components and mechanisms that regulate its cell cycle are poorly described. In higher eukaryotes, cyclins are proteins that activate cyclin-dependent kinases (CDKs), by associating with them along the different stages of the cell cycle. These cyclin-CDK complexes exert their role as major modulators of the cell cycle by phosphorylating specific substrates. For the correct progression of the cell cycle, the mechanisms that regulate the activity of cyclins and their associated CDKs are diverse and must be controlled precisely. Different types of cyclins are involved in specific phases of the eukaryotic cell cycle, preferentially activating certain CDKs. In this work, we characterized TcCYC6, a putative coding sequence of T. cruzi which encodes a protein with homology to mitotic cyclins. The overexpression of this sequence, fused to a tag of nine amino acids from influenza virus hemagglutinin (TcCYC6-HA), showed to be detrimental for the proliferation of epimastigotes in axenic culture and affected the cell cycle progression. In silico analysis revealed an N-terminal segment similar to the consensus sequence of the destruction box, a hallmark for the degradation of several mitotic cyclins. We experimentally determined that the TcCYC6-HA turnover decreased in the presence of proteasome inhibitors, suggesting that TcCYC6 degradation occurs via ubiquitin-proteasome pathway. The results obtained in this study provide first evidence that TcCYC6 expression and degradation are finely regulated in T. cruzi. PMID:26709077

  20. Ionizing radiation is a potent inducer of mitotic recombination in mouse embryonic stem cells

    International Nuclear Information System (INIS)

    Highlights: → Embryonic stem cells have a distinct mutational response to X-rays. → X-rays induce more mutations in embryonic stem cells than in somatic cells. → Mitotic recombination is more readily induced by X-rays in embryonic stem cells. → Radiation hazards may have different consequences on different types of cells. - Abstract: Maintenance of genomic integrity in embryonic cells is pivotal to proper embryogenesis, organogenesis and to the continuity of species. Cultured mouse embryonic stem cells (mESCs), a model for early embryonic cells, differ from cultured somatic cells in their capacity to remodel chromatin, in their repertoire of DNA repair enzymes, and in the regulation of cell cycle checkpoints. Using 129XC3HF1 mESCs heterozygous for Aprt, we characterized loss of Aprt heterozygosity after exposure to ionizing radiation. We report here that the frequency of loss of heterozygosity mutants in mESCs can be induced several hundred-fold by exposure to 5-10 Gy of X-rays. This induction is 50-100-fold higher than the induction reported for mouse adult or embryonic fibroblasts. The primary mechanism underlying the elevated loss of heterozygosity after irradiation is mitotic recombination, with lesser contributions from deletions and gene conversions that span Aprt. Aprt point mutations and epigenetic inactivation are very rare in mESCs compared to fibroblasts. Mouse ESCs, therefore, are distinctive in their response to ionizing radiation and studies of differentiated cells may underestimate the mutagenic effects of ionizing radiation on ESC or other stem cells. Our findings are important to understanding the biological effects of ionizing radiation on early development and carcinogenesis.

  1. Ionizing radiation is a potent inducer of mitotic recombination in mouse embryonic stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Denissova, Natalia G.; Tereshchenko, Irina V.; Cui, Eric [Department of Genetics, Rutgers University, Piscataway, 145 Bevier Rd, NJ 08854 (United States); Stambrook, Peter J. [Department of Molecular Genetics, Biochemistry and Microbiology, University of Cincinnati, Cincinnati, OH 45267 (United States); Shao, Changshun [Department of Genetics, Rutgers University, Piscataway, 145 Bevier Rd, NJ 08854 (United States); Tischfield, Jay A., E-mail: jay@biology.rutgers.edu [Department of Genetics, Rutgers University, Piscataway, 145 Bevier Rd, NJ 08854 (United States)

    2011-10-01

    Highlights: {yields} Embryonic stem cells have a distinct mutational response to X-rays. {yields} X-rays induce more mutations in embryonic stem cells than in somatic cells. {yields} Mitotic recombination is more readily induced by X-rays in embryonic stem cells. {yields} Radiation hazards may have different consequences on different types of cells. - Abstract: Maintenance of genomic integrity in embryonic cells is pivotal to proper embryogenesis, organogenesis and to the continuity of species. Cultured mouse embryonic stem cells (mESCs), a model for early embryonic cells, differ from cultured somatic cells in their capacity to remodel chromatin, in their repertoire of DNA repair enzymes, and in the regulation of cell cycle checkpoints. Using 129XC3HF1 mESCs heterozygous for Aprt, we characterized loss of Aprt heterozygosity after exposure to ionizing radiation. We report here that the frequency of loss of heterozygosity mutants in mESCs can be induced several hundred-fold by exposure to 5-10 Gy of X-rays. This induction is 50-100-fold higher than the induction reported for mouse adult or embryonic fibroblasts. The primary mechanism underlying the elevated loss of heterozygosity after irradiation is mitotic recombination, with lesser contributions from deletions and gene conversions that span Aprt. Aprt point mutations and epigenetic inactivation are very rare in mESCs compared to fibroblasts. Mouse ESCs, therefore, are distinctive in their response to ionizing radiation and studies of differentiated cells may underestimate the mutagenic effects of ionizing radiation on ESC or other stem cells. Our findings are important to understanding the biological effects of ionizing radiation on early development and carcinogenesis.

  2. Automated high-throughput quantification of mitotic spindle positioning from DIC movies of Caenorhabditis embryos.

    Directory of Open Access Journals (Sweden)

    David Cluet

    Full Text Available The mitotic spindle is a microtubule-based structure that elongates to accurately segregate chromosomes during anaphase. Its position within the cell also dictates the future cell cleavage plan, thereby determining daughter cell orientation within a tissue or cell fate adoption for polarized cells. Therefore, the mitotic spindle ensures at the same time proper cell division and developmental precision. Consequently, spindle dynamics is the matter of intensive research. Among the different cellular models that have been explored, the one-cell stage C. elegans embryo has been an essential and powerful system to dissect the molecular and biophysical basis of spindle elongation and positioning. Indeed, in this large and transparent cell, spindle poles (or centrosomes can be easily detected from simple DIC microscopy by human eyes. To perform quantitative and high-throughput analysis of spindle motion, we developed a computer program ACT for Automated-Centrosome-Tracking from DIC movies of C. elegans embryos. We therefore offer an alternative to the image acquisition and processing of transgenic lines expressing fluorescent spindle markers. Consequently, experiments on large sets of cells can be performed with a simple setup using inexpensive microscopes. Moreover, analysis of any mutant or wild-type backgrounds is accessible because laborious rounds of crosses with transgenic lines become unnecessary. Last, our program allows spindle detection in other nematode species, offering the same quality of DIC images but for which techniques of transgenesis are not accessible. Thus, our program also opens the way towards a quantitative evolutionary approach of spindle dynamics. Overall, our computer program is a unique macro for the image- and movie-processing platform ImageJ. It is user-friendly and freely available under an open-source licence. ACT allows batch-wise analysis of large sets of mitosis events. Within 2 minutes, a single movie is processed

  3. Ki-67 expression is superior to mitotic count and novel proliferation markers PHH3, MCM4 and mitosin as a prognostic factor in thick cutaneous melanoma

    International Nuclear Information System (INIS)

    Tumor cell proliferation is a predictor of survival in cutaneous melanoma. The aim of the present study was to evaluate the prognostic impact of mitotic count, Ki-67 expression and novel proliferation markers phosphohistone H3 (PHH3), minichromosome maintenance protein 4 (MCM4) and mitosin, and to compare the results with histopathological variables. 202 consecutive cases of nodular cutaneous melanoma were initially included. Mitotic count (mitosis per mm2) was assessed on H&E sections, and Ki-67 expression was estimated by immunohistochemistry on standard sections. PHH3, MCM4 and mitosin were examined by staining of tissue microarrays (TMA) sections. Increased mitotic count and elevated Ki-67 expression were strongly associated with increased tumor thickness, presence of ulceration and tumor necrosis. Furthermore, high mitotic count and elevated Ki-67 expression were also associated with Clark's level of invasion and presence of vascular invasion. High expression of PHH3 and MCM4 was correlated with high mitotic count, elevated Ki-67 expression and tumor ulceration, and increased PHH3 frequencies were associated with tumor thickness and presence of tumor necrosis. Univariate analyses showed a worse outcome in cases with elevated Ki-67 expression and high mitotic count, whereas PHH3, MCM4 and mitosin were not significant. Tumor cell proliferation by Ki-67 had significant prognostic impact by multivariate analysis. Ki-67 was a stronger and more robust prognostic indicator than mitotic count in this series of nodular melanoma. PHH3, MCM4 and mitosin did not predict patient survival

  4. Fully functional global genome repair of (6-4) photoproducts and compromised transcription-coupled repair of cyclobutane pyrimidine dimers in condensed mitotic chromatin

    Energy Technology Data Exchange (ETDEWEB)

    Komura, Jun-ichiro, E-mail: junkom@med.tohoku.ac.jp [Department of Cell Biology, Tohoku University Graduate School of Medicine, Sendai 980-8575 (Japan); Ikehata, Hironobu [Department of Cell Biology, Tohoku University Graduate School of Medicine, Sendai 980-8575 (Japan); Mori, Toshio [Radioisotope Research Center, Nara Medical University, Kashihara, Nara 634-8521 (Japan); Ono, Tetsuya [Department of Cell Biology, Tohoku University Graduate School of Medicine, Sendai 980-8575 (Japan)

    2012-03-10

    During mitosis, chromatin is highly condensed, and activities such as transcription and semiconservative replication do not occur. Consequently, the condensed condition of mitotic chromatin is assumed to inhibit DNA metabolism by impeding the access of DNA-transacting proteins. However, about 40 years ago, several researchers observed unscheduled DNA synthesis in UV-irradiated mitotic chromosomes, suggesting the presence of excision repair. We re-examined this subject by directly measuring the removal of UV-induced DNA lesions by an ELISA and by a Southern-based technique in HeLa cells arrested at mitosis. We observed that the removal of (6-4) photoproducts from the overall genome in mitotic cells was as efficient as in interphase cells. This suggests that global genome repair of (6-4) photoproducts is fully functional during mitosis, and that the DNA in mitotic chromatin is accessible to proteins involved in this mode of DNA repair. Nevertheless, not all modes of DNA repair seem fully functional during mitosis. We also observed that the removal of cyclobutane pyrimidine dimers from the dihydrofolate reductase and c-MYC genes in mitotic cells was very slow. This suggests that transcription-coupled repair of cyclobutane pyrimidine dimers is compromised or non-functional during mitosis, which is probably the consequence of mitotic transcriptional repression. -- Highlights: Black-Right-Pointing-Pointer Global genome repair of (6-4) photoproducts is fully active in mitotic cells. Black-Right-Pointing-Pointer DNA in condensed mitotic chromatin does not seem inaccessible or inert. Black-Right-Pointing-Pointer Mitotic transcriptional repression may impair transcription-coupled repair.

  5. Fully functional global genome repair of (6-4) photoproducts and compromised transcription-coupled repair of cyclobutane pyrimidine dimers in condensed mitotic chromatin

    International Nuclear Information System (INIS)

    During mitosis, chromatin is highly condensed, and activities such as transcription and semiconservative replication do not occur. Consequently, the condensed condition of mitotic chromatin is assumed to inhibit DNA metabolism by impeding the access of DNA-transacting proteins. However, about 40 years ago, several researchers observed unscheduled DNA synthesis in UV-irradiated mitotic chromosomes, suggesting the presence of excision repair. We re-examined this subject by directly measuring the removal of UV-induced DNA lesions by an ELISA and by a Southern-based technique in HeLa cells arrested at mitosis. We observed that the removal of (6-4) photoproducts from the overall genome in mitotic cells was as efficient as in interphase cells. This suggests that global genome repair of (6-4) photoproducts is fully functional during mitosis, and that the DNA in mitotic chromatin is accessible to proteins involved in this mode of DNA repair. Nevertheless, not all modes of DNA repair seem fully functional during mitosis. We also observed that the removal of cyclobutane pyrimidine dimers from the dihydrofolate reductase and c-MYC genes in mitotic cells was very slow. This suggests that transcription-coupled repair of cyclobutane pyrimidine dimers is compromised or non-functional during mitosis, which is probably the consequence of mitotic transcriptional repression. -- Highlights: ► Global genome repair of (6-4) photoproducts is fully active in mitotic cells. ► DNA in condensed mitotic chromatin does not seem inaccessible or inert. ► Mitotic transcriptional repression may impair transcription-coupled repair.

  6. Immunohistochemical localization of epidermal growth factor in rat and man

    DEFF Research Database (Denmark)

    Poulsen, Steen Seier; Nexø, Ebba

    1986-01-01

    Epidermal growth factor (EGF) is a peptide which stimulates cell mitotic activity and differentiation, has a cytoprotective effect on the gastroduodenal mucosa, and inhibits gastric acid secretion. The immunohistochemical localization of EGF in the Brunner's glands and the submandibular glands is...... well documented. The localization of EGF in other tissues is still unclarified. In the present study, the immunohistochemical localization of EGF in tissues from rat, man and a 20 week human fetus were investigated. In man and rat, immunoreaction was found in the submandibular glands, the serous glands...

  7. Flow cytometric analysis of mitotic cycle perturbation by chemical carcinogens in cultured epithelial cells. [Effects of benzo(a)pyrene-diol-epoxide on mitotic cycle of cultural mouse liver epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Pearlman, A.L.

    1978-08-01

    A system for kinetic analysis of mitotic cycle perturbation by various agents was developed and applied to the study of the mitotic cycle effects and dependency of the chemical carcinogen benzo(a)pyrene-diolepoxide, DE, upon a mouse lever epithelial cell line, NMuLi. The study suggests that the targets of DE action are not confined to DNA alone but may include cytoplasmic structures as well. DE was found to affect cells located in virtually every phase of the mitotic cycle, with cells that were actively synthesizing DNA showing the strongest response. However, the resulting perturbations were not confined to S-phase alone. DE slowed traversal through S-phase by about 40% regardless of the cycle phase of the cells exposed to it, and slowed traversal through G/sub 2/M by about 50%. When added to G/sub 1/ cells, DE delayed recruitment of apparently quiescent (G/sub 0/) cells by 2 hours, and reduced the synchrony of the cohort of cells recruited into active proliferation. The kinetic analysis system consists of four elements: tissue culture methods for propagating and harvesting cell populations; an elutriation centrifugation system for bulk synchronization of cells in various phases of the mitotic cycle; a flow cytometer (FCM), coupled with appropriate staining protocols, to enable rapid analysis of the DNA distribution of any given cell population; and data reduction and analysis methods for extracting information from the DNA histograms produced by the FCM. The elements of the system are discussed. A mathematical analysis of DNA histograms obtained by FCM is presented. The analysis leads to the detailed implementation of a new modeling approach. The new modeling approach is applied to the estimation of cell cycle kinetic parameters from time series of DNA histograms, and methods for the reduction and interpretation of such series are suggested.

  8. Interphase APC/C-Cdc20 inhibition by cyclin A2-Cdk2 ensures efficient mitotic entry

    DEFF Research Database (Denmark)

    Hein, Jamin B; Nilsson, Jakob

    2016-01-01

    window of the cell cycle, if any, is unknown. Here we show that cyclin A2-Cdk2 binds and phosphorylates Cdc20 in interphase and this inhibits APC/C-Cdc20 activity. Preventing Cdc20 phosphorylation results in pre-mature activation of the APC/C-Cdc20 and several substrates, including cyclin B1 and A2, are...... destabilized which lengthens G2 and slows mitotic entry. Expressing non-degradable cyclin A2 but not cyclin B1 restores mitotic entry in these cells. We have thus uncovered a novel positive feedback loop centred on cyclin A2-Cdk2 inhibition of interphase APC/C-Cdc20 to allow further cyclin A2 accumulation and...

  9. A Decrease in Ambient Temperature Induces Post-Mitotic Enlargement of Palisade Cells in North American Lake Cress.

    Directory of Open Access Journals (Sweden)

    Rumi Amano

    Full Text Available In order to maintain organs and structures at their appropriate sizes, multicellular organisms orchestrate cell proliferation and post-mitotic cell expansion during morphogenesis. Recent studies using Arabidopsis leaves have shown that compensation, which is defined as post-mitotic cell expansion induced by a decrease in the number of cells during lateral organ development, is one example of such orchestration. Some of the basic molecular mechanisms underlying compensation have been revealed by genetic and chimeric analyses. However, to date, compensation had been observed only in mutants, transgenics, and γ-ray-treated plants, and it was unclear whether it occurs in plants under natural conditions. Here, we illustrate that a shift in ambient temperature could induce compensation in Rorippa aquatica (Brassicaceae, a semi-aquatic plant found in North America. The results suggest that compensation is a universal phenomenon among angiosperms and that the mechanism underlying compensation is shared, in part, between Arabidopsis and R. aquatica.

  10. 53BP1 nuclear bodies form around DNA lesions generated by mitotic transmission of chromosomes under replication stress

    DEFF Research Database (Denmark)

    Lukas, Claudia; Savic, Velibor; Bekker-Jensen, Simon;

    2011-01-01

    Completion of genome duplication is challenged by structural and topological barriers that impede progression of replication forks. Although this can seriously undermine genome integrity, the fate of DNA with unresolved replication intermediates is not known. Here, we show that mild replication...... bodies shield chromosomal fragile sites sequestered in these compartments against erosion. Together, these data indicate that restoration of DNA or chromatin integrity at loci prone to replication problems requires mitotic transmission to the next cell generations....... increases after genetic ablation of BLM, a DNA helicase associated with dissolution of entangled DNA. Conversely, 53BP1 nuclear bodies are partially suppressed by knocking down SMC2, a condensin subunit required for mechanical stability of mitotic chromosomes. Finally, we provide evidence that 53BP1 nuclear...

  11. Maternal embryonic leucine zipper kinase is stabilized in mitosis by phosphorylation and is partially degraded upon mitotic exit

    International Nuclear Information System (INIS)

    MELK (maternal embryonic leucine zipper kinase) is a cell cycle dependent protein kinase involved in diverse cell processes including cell proliferation, apoptosis, cell cycle and mRNA processing. Noticeably, MELK expression is increased in cancerous tissues, upon cell transformation and in mitotically-blocked cells. The question of how MELK protein level is controlled is therefore important. Here, we show that MELK protein is restricted to proliferating cells derived from either cancer or normal tissues and that MELK protein level is severely decreased concomitantly with other cell cycle proteins in cells which exit the cell cycle. Moreover, we demonstrate in human HeLa cells and Xenopus embryos that approximately half of MELK protein is degraded upon mitotic exit whereas another half remains stable during interphase. We show that the stability of MELK protein in M-phase is dependent on its phosphorylation state.

  12. Effect of lymphocytes culture variations on the mitotic index and on the dicentric yield following gamma radiation exposure

    International Nuclear Information System (INIS)

    Fundamentals of biological dosimetry are described in the International Atomic Energy Agency manual, but all over the world each laboratory is using its own protocol. To test the influence of protocol variations, some blood samples were exposed to 0.5 Gy of gamma radiation and mitotic index and dicentric rates were measured under different experimental conditions. The effect of seven parameters [bromodeoxyuridine (BrdU), phytohaemagglutinin and colcemide concentrations, blood and medium volumes, culture duration and incubation temperature] was tested using a Placket and Burman experimental design. The analysis reveals that the mitotic index was influenced by the concentration of BrdU, medium and blood volumes, the culture duration and the temperature. However, none of the factors has a significant impact on the yield of dicentrics. The dicentric assay is robust against reagent variations within the range tested. These results could be used by relevant laboratories as elements of their procedures robustness in any event requiring such demonstration. (authors)

  13. The effect of X-rays on the mitotic activity of the adrenal gland, jejunum, lymph node and epidermis of the mouse

    Energy Technology Data Exchange (ETDEWEB)

    Knowlton, N.P. Jr.; Hempelmann, L.H.

    1949-04-19

    In the search for a suitable method of determining quantitatively the effects of mammalian tissue of sublethal doses of ionizing radiation, the following study of the mitotic indices of various mouse tissues following whole body irradiation was undertaken.

  14. Mitotic phosphorylation of SOX2 mediated by Aurora kinase A is critical for the stem-cell like cell maintenance in PA-1 cells.

    Science.gov (United States)

    Qi, Dandan; Wang, Qianqian; Yu, Min; Lan, Rongfeng; Li, Shuiming; Lu, Fei

    2016-08-01

    Transcription factor SOX2 is multiple phosphorylated. However, the kinase and the timing regulating SOX2 phosphorylation remains poorly understood. Here we reported mitotic phosphorylation of SOX2 by Aurora kinase A (AURKA). AURKA inhibitors (VX680, Aurora kinase Inhibitor I) but not PLK1 inhibitors (BI2536, CBB2001) eliminate the mitotic phosphorylation of SOX2. Consistently, siRNA inhibition of AURKA can eliminate mitotic SOX2 phosphorylation. Ser220 and Ser251 are two sites that identified for mitotic phosphorylation on SOX2. Moreover, SOX2 mutants (S220A and S251A) can promote SOX2 induced OCT4 re-expression in differentiated cells. These findings reveal a novel regulation mechanism of SOX2 phosphorylation mediated by AURKA in mitosis and its function in stem cell pluripotency maintenance in cancer cells. PMID:27249336

  15. Novel Mad2-targeting miR-493-3p controls mitotic fidelity and cancer cells’ sensitivity to paclitaxel

    OpenAIRE

    Tambe, Mahesh; Pruikkonen, Sofia; Mäki-Jouppila, Jenni; Ping CHEN; Elgaaen, Bente Vilming; Straume, Anne Hege; Huhtinen, Kaisa; Cárpen, Olli; Lønning, Per Eystein; Davidson, Ben; Hautaniemi, Sampsa; Kallio, Marko J.

    2016-01-01

    The molecular pathways that contribute to the proliferation and drug response of cancer cells are highly complex and currently insufficiently characterized. We have identified a previously unknown microRNA-based mechanism that provides cancer cells means to stimulate tumorigenesis via increased genomic instability and, at the same time, evade the action of clinically utilized microtubule drugs. We demonstrate miR-493-3p to be a novel negative regulator of mitotic arrest deficient-2 (MAD2), an...

  16. Mitotic defects lead to pervasive aneuploidy and accompany loss of RB1 activity in mouse LmnaDhe dermal fibroblasts.

    Directory of Open Access Journals (Sweden)

    C Herbert Pratt

    Full Text Available BACKGROUND: Lamin A (LMNA is a component of the nuclear lamina and is mutated in several human diseases, including Emery-Dreifuss muscular dystrophy (EDMD; OMIM ID# 181350 and the premature aging syndrome Hutchinson-Gilford progeria syndrome (HGPS; OMIM ID# 176670. Cells from progeria patients exhibit cell cycle defects in both interphase and mitosis. Mouse models with loss of LMNA function have reduced Retinoblastoma protein (RB1 activity, leading to aberrant cell cycle control in interphase, but how mitosis is affected by LMNA is not well understood. RESULTS: We examined the cell cycle and structural phenotypes of cells from mice with the Lmna allele, Disheveled hair and ears (Lmna(Dhe. We found that dermal fibroblasts from heterozygous Lmna(Dhe (Lmna(Dhe/+ mice exhibit many phenotypes of human laminopathy cells. These include severe perturbations to the nuclear shape and lamina, increased DNA damage, and slow growth rates due to mitotic delay. Interestingly, Lmna(Dhe/+ fibroblasts also had reduced levels of hypophosphorylated RB1 and the non-SMC condensin II-subunit D3 (NCAP-D3, a mitosis specific centromere condensin subunit that depends on RB1 activity. Mitotic check point control by mitotic arrest deficient-like 1 (MAD2L1 also was perturbed in Lmna(Dhe/+ cells. Lmna(Dhe/+ fibroblasts were consistently aneuploid and had higher levels of micronuclei and anaphase bridges than normal fibroblasts, consistent with chromosome segregation defects. CONCLUSIONS: These data indicate that RB1 may be a key regulator of cellular phenotype in laminopathy-related cells, and suggest that the effects of LMNA on RB1 include both interphase and mitotic cell cycle control.

  17. Combining Magnetic Sorting of Mother Cells and Fluctuation Tests to Analyze Genome Instability During Mitotic Cell Aging in Saccharomyces cerevisiae

    OpenAIRE

    Patterson, Melissa N.; Maxwell, Patrick H.

    2014-01-01

    Saccharomyces cerevisiae has been an excellent model system for examining mechanisms and consequences of genome instability. Information gained from this yeast model is relevant to many organisms, including humans, since DNA repair and DNA damage response factors are well conserved across diverse species. However, S. cerevisiae has not yet been used to fully address whether the rate of accumulating mutations changes with increasing replicative (mitotic) age due to technical constraints. For i...

  18. Cooperative Action of Cdk1/cyclin B and SIRT1 Is Required for Mitotic Repression of rRNA Synthesis

    OpenAIRE

    Voit, Renate; Seiler, Jeanette; Grummt, Ingrid

    2015-01-01

    Author Summary In metazoans, transcription is arrested during mitosis. Previous studies have established that mitotic repression of cellular transcription is mediated by Cdk1/cyclin B-dependent phosphorylation of basal transcription factors that nucleate transcription complex formation. Repression of rDNA transcription at the onset of mitosis is brought about by inactivation of the TBP-containing transcription factor SL1 by Cdk1/cyclin B-dependent phosphorylation of the TAFI110 subunit, which...

  19. Bovine Papillomavirus Type 1 Genomes and the E2 Transactivator Protein Are Closely Associated with Mitotic Chromatin

    OpenAIRE

    Skiadopoulos, Mario H.; Alison A McBride

    1998-01-01

    The bovine papillomavirus type 1 E2 transactivator protein is required for viral transcriptional regulation and DNA replication and may be important for long-term episomal maintenance of viral genomes within replicating cells (M. Piirsoo, E. Ustav, T. Mandel, A. Stenlund, and M. Ustav, EMBO J. 15:1–11, 1996). We have evidence that, in contrast to most other transcriptional transactivators, the E2 transactivator protein is associated with mitotic chromosomes in dividing cells. The shorter E2-T...

  20. Dimerization of the Papillomavirus E2 Protein Is Required for Efficient Mitotic Chromosome Association and Brd4 Binding▿

    OpenAIRE

    Cardenas-Mora, Juan; Spindler, Jonathan E.; Jang, Moon Kyoo; Alison A McBride

    2008-01-01

    The E2 proteins of several papillomaviruses link the viral genome to mitotic chromosomes to ensure retention and the efficient partitioning of genomes into daughter cells following cell division. Bovine papillomavirus type 1 E2 binds to chromosomes in a complex with Brd4, a cellular bromodomain protein. Interaction with Brd4 is also important for E2-mediated transcriptional regulation. The transactivation domain of E2 is crucial for interaction with the Brd4 protein; proteins lacking or mutat...

  1. Navitoclax (ABT-263) accelerates apoptosis during drug-induced mitotic arrest by antagonizing Bcl-xL

    OpenAIRE

    Shi, Jue; Zhou, Yuan; Huang, Hsiao-Chun; Mitchison, Timothy J.

    2011-01-01

    Combining microtubule-targeting anti-mitotic drugs with targeted apoptosis potentiators is a promising new chemotherapeutic strategy to treat cancer. In this study we investigate the cellular mechanism by which Navitoclax (previously called ABT-263), a Bcl-2 family inhibitor, potentiates apoptosis triggered by paclitaxel and an inhibitor of Kinesin-5 (KSP), across a panel of epithelial cancer lines. Using time-lapse microscopy, we show that Navitoclax has little effect on cell death during in...

  2. Upregulation of meiosis-specific genes in lymphoma cell lines following genotoxic insult and induction of mitotic catastrophe

    International Nuclear Information System (INIS)

    We have previously reported that p53 mutated radioresistant lymphoma cell lines undergo mitotic catastrophe after irradiation, resulting in metaphase arrest and the generation of endopolyploid cells. A proportion of these endopolyploid cells then undergo a process of de-polyploidisation, stages of which are partially reminiscent of meiotic prophase. Furthermore, expression of meiosis-specific proteins of the cancer/testis antigens group of genes has previously been reported in tumours. We therefore investigated whether expression of meiosis-specific genes was associated with the polyploidy response in our tumour model. Three lymphoma cell lines, Namalwa, WI-L2-NS and TK6, of varying p53 status were exposed to a single 10 Gy dose of gamma radiation and their responses assessed over an extended time course. DNA flow cytometry and mitotic counts were used to assess the kinetics and extent of polyploidisation and mitotic progression. Expression of meiotic genes was analysed using RT-PCR and western blotting. In addition, localisation of the meiotic cohesin REC8 and its relation to centromeres was analysed by immunofluorescence. The principal meiotic regulator MOS was found to be significantly post-transcriptionally up-regulated after irradiation in p53 mutated but not p53 wild-type lymphoma cells. The maximum expression of MOS coincided with the maximal fraction of metaphase arrested cells and was directly proportional to both the extent of the arrest and the number of endopolyploid cells that subsequently emerged. The meiotic cohesin REC8 was also found to be up-regulated after irradiation, linking sister chromatid centromeres in the metaphase-arrested and subsequent giant cells. Finally, RT-PCR revealed expression of the meiosis-prophase genes, DMC1, STAG3, SYCP3 and SYCP1. We conclude that multiple meiotic genes are aberrantly activated during mitotic catastrophe in p53 mutated lymphoma cells after irradiation. Furthermore, we suggest that the coordinated expression

  3. Paclitaxel targets FOXM1 to regulate KIF20A in mitotic catastrophe and breast cancer paclitaxel resistance.

    Science.gov (United States)

    Khongkow, P; Gomes, A R; Gong, C; Man, E P S; Tsang, J W-H; Zhao, F; Monteiro, L J; Coombes, R C; Medema, R H; Khoo, U S; Lam, E W-F

    2016-02-25

    FOXM1 has been implicated in taxane resistance, but the molecular mechanism involved remains elusive. In here, we show that FOXM1 depletion can sensitize breast cancer cells and mouse embryonic fibroblasts into entering paclitaxel-induced senescence, with the loss of clonogenic ability, and the induction of senescence-associated β-galactosidase activity and flat cell morphology. We also demonstrate that FOXM1 regulates the expression of the microtubulin-associated kinesin KIF20A at the transcriptional level directly through a Forkhead response element (FHRE) in its promoter. Similar to FOXM1, KIF20A expression is downregulated by paclitaxel in the sensitive MCF-7 breast cancer cells and deregulated in the paclitaxel-resistant MCF-7Tax(R) cells. KIF20A depletion also renders MCF-7 and MCF-7Tax(R) cells more sensitive to paclitaxel-induced cellular senescence. Crucially, resembling paclitaxel treatment, silencing of FOXM1 and KIF20A similarly promotes abnormal mitotic spindle morphology and chromosome alignment, which have been shown to induce mitotic catastrophe-dependent senescence. The physiological relevance of the regulation of KIF20A by FOXM1 is further highlighted by the strong and significant correlations between FOXM1 and KIF20A expression in breast cancer patient samples. Statistical analysis reveals that both FOXM1 and KIF20A protein and mRNA expression significantly associates with poor survival, consistent with a role of FOXM1 and KIF20A in paclitaxel action and resistance. Collectively, our findings suggest that paclitaxel targets the FOXM1-KIF20A axis to drive abnormal mitotic spindle formation and mitotic catastrophe and that deregulated FOXM1 and KIF20A expression may confer paclitaxel resistance. These findings provide insights into the underlying mechanisms of paclitaxel resistance and have implications for the development of predictive biomarkers and novel chemotherapeutic strategies for paclitaxel resistance. PMID:25961928

  4. High throughput screening of natural products for anti-mitotic effects in MDA-MB-231 human breast carcinoma cells.

    Science.gov (United States)

    Mazzio, E; Badisa, R; Mack, N; Deiab, S; Soliman, K F A

    2014-06-01

    Some of the most effective anti-mitotic microtubule-binding agents, such as paclitaxel (Taxus brevifolia) were originally discovered through robust National Cancer Institute botanical screenings. In this study, a high-through put microarray format was utilized to screen 897 aqueous extracts of commonly used natural products (0.00015-0.5 mg/mL) relative to paclitaxel for anti-mitotic effects (independent of toxicity) on proliferation of MDA-MB-231 cells. The data obtained showed that less than 1.34 % of the extracts tested showed inhibitory growth (IG50 ) properties Phoradendron flavescens), Tou Gu Cao [symbol: see text] Speranskia herb (Speranskia tuberculata), Bentonite clay, Bunge root (Pulsatilla chinensis), Brucea fruit (Brucea javanica), Madder root (Rubia tinctorum), Gallnut of Chinese Sumac (Melaphis chinensis), Elecampane root (Inula Helenium), Yuan Zhi [symbol: see text] root (Polygala tenuifolia), Pagoda Tree fruit (Melia Toosendan), Stone root (Collinsonia Canadensis), and others such as American Witchhazel, Arjun, and Bladderwrack. The strongest tumoricidal herbs identified from amongst the subset evaluated for anti-mitotic properties were wild yam (Dioscorea villosa), beth root (Trillium Pendulum), and alkanet root (Lithospermum canescens). Additional data was obtained on a lesser-recognized herb: (S. tuberculata), which showed growth inhibition on BT-474 (human ductal breast carcinoma) and Ishikawa (human endometrial adenocarcinoma) cells with ability to block replicative DNA synthesis, leading to G2 arrest in MDA-MB-231 cells. In conclusion, these findings present relative potency of anti-mitotic natural plants that are effective against human breast carcinoma MDA-MB-231 cell division. PMID:24105850

  5. Slx5/Slx8 Promotes Replication Stress Tolerance by Facilitating Mitotic Progression.

    Science.gov (United States)

    Thu, Yee Mon; Van Riper, Susan Kaye; Higgins, LeeAnn; Zhang, Tianji; Becker, Jordan Robert; Markowski, Todd William; Nguyen, Hai Dang; Griffin, Timothy Jon; Bielinsky, Anja Katrin

    2016-05-10

    Loss of minichromosome maintenance protein 10 (Mcm10) causes replication stress. We uncovered that S. cerevisiae mcm10-1 mutants rely on the E3 SUMO ligase Mms21 and the SUMO-targeted ubiquitin ligase complex Slx5/8 for survival. Using quantitative mass spectrometry, we identified changes in the SUMO proteome of mcm10-1 mutants and revealed candidates regulated by Slx5/8. Such candidates included subunits of the chromosome passenger complex (CPC), Bir1 and Sli15, known to facilitate spindle assembly checkpoint (SAC) activation. We show here that Slx5 counteracts SAC activation in mcm10-1 mutants under conditions of moderate replication stress. This coincides with the proteasomal degradation of sumoylated Bir1. Importantly, Slx5-dependent mitotic relief was triggered not only by Mcm10 deficiency but also by treatment with low doses of the alkylating drug methyl methanesulfonate. Based on these findings, we propose a model in which Slx5/8 allows for passage through mitosis when replication stress is tolerable. PMID:27134171

  6. B microchromosomes in the family Curimatidae (Characiformes: mitotic and meiotic behavior

    Directory of Open Access Journals (Sweden)

    Tatiane Ramos Sampaio

    2011-11-01

    Full Text Available In the present work, six curimatid species were analyzed: Cyphocharax voga (Hensel, 1870, C. spilotus (Vari, 1987, C. saladensis (Meinken, 1933, C. modestus (Fernández-Yépez, 1948, Steindachnerina biornata (Braga & Azpelicueta, 1987 and S. insculpta (Fernández-Yépez, 1948 collected from two hydrographic basins. All samples presented 2n=54 meta-submetacentric (m-sm chromosomes and FN equal to 108, and 1 or 2 B microchromosomes in the mitotic and meiotic cells of the six sampled populations showing inter-and intraindividual variation. The analysis of the meiotic cells in C. saladensis, C. spilotus, and C. voga showed a modal number of 54 chromosomes in the spermatogonial metaphases and 27 bivalents in the pachytene, diplotene, diakinesis and in metaphase I stages, and 27 chromosomes in metaphase II; in C. modestus, S. biornata, and S. insculpta, spermatogonial metaphases with 54 chromosomes and pachytene and metaphase I with 27 bivalents were observed. The B microchromosome was observed as univalent in the spermatogonial metaphase of C. spilotus, in the pachytene stage in the other species, with the exception of C. saladensis, and S. biornata in metaphase I. New occurrences of the B microchromosome in C. voga, C. saladensis and S. biornata were observed, confirming that the presence of this type of chromosome is a striking characteristic of this group of fish.

  7. Slx5/Slx8 Promotes Replication Stress Tolerance by Facilitating Mitotic Progression

    Directory of Open Access Journals (Sweden)

    Yee Mon Thu

    2016-05-01

    Full Text Available Loss of minichromosome maintenance protein 10 (Mcm10 causes replication stress. We uncovered that S. cerevisiae mcm10-1 mutants rely on the E3 SUMO ligase Mms21 and the SUMO-targeted ubiquitin ligase complex Slx5/8 for survival. Using quantitative mass spectrometry, we identified changes in the SUMO proteome of mcm10-1 mutants and revealed candidates regulated by Slx5/8. Such candidates included subunits of the chromosome passenger complex (CPC, Bir1 and Sli15, known to facilitate spindle assembly checkpoint (SAC activation. We show here that Slx5 counteracts SAC activation in mcm10-1 mutants under conditions of moderate replication stress. This coincides with the proteasomal degradation of sumoylated Bir1. Importantly, Slx5-dependent mitotic relief was triggered not only by Mcm10 deficiency but also by treatment with low doses of the alkylating drug methyl methanesulfonate. Based on these findings, we propose a model in which Slx5/8 allows for passage through mitosis when replication stress is tolerable.

  8. Spatial Rule-Based Modeling: A Method and Its Application to the Human Mitotic Kinetochore

    Directory of Open Access Journals (Sweden)

    Jan Huwald

    2013-07-01

    Full Text Available A common problem in the analysis of biological systems is the combinatorial explosion that emerges from the complexity of multi-protein assemblies. Conventional formalisms, like differential equations, Boolean networks and Bayesian networks, are unsuitable for dealing with the combinatorial explosion, because they are designed for a restricted state space with fixed dimensionality. To overcome this problem, the rule-based modeling language, BioNetGen, and the spatial extension, SRSim, have been developed. Here, we describe how to apply rule-based modeling to integrate experimental data from different sources into a single spatial simulation model and how to analyze the output of that model. The starting point for this approach can be a combination of molecular interaction data, reaction network data, proximities, binding and diffusion kinetics and molecular geometries at different levels of detail. We describe the technique and then use it to construct a model of the human mitotic inner and outer kinetochore, including the spindle assembly checkpoint signaling pathway. This allows us to demonstrate the utility of the procedure, show how a novel perspective for understanding such complex systems becomes accessible and elaborate on challenges that arise in the formulation, simulation and analysis of spatial rule-based models.

  9. Cell shape impacts on the positioning of the mitotic spindle with respect to the substratum.

    Science.gov (United States)

    Lázaro-Diéguez, Francisco; Ispolatov, Iaroslav; Müsch, Anne

    2015-04-01

    All known mechanisms of mitotic spindle orientation rely on astral microtubules. We report that even in the absence of astral microtubules, metaphase spindles in MDCK and HeLa cells are not randomly positioned along their x-z dimension, but preferentially adopt shallow β angles between spindle pole axis and substratum. The nonrandom spindle positioning is due to constraints imposed by the cell cortex in flat cells that drive spindles that are longer and/or wider than the cell's height into a tilted, quasidiagonal x-z position. In rounder cells, which are taller, fewer cortical constraints make the x-z spindle position more random. Reestablishment of astral microtubule-mediated forces align the spindle poles with cortical cues parallel to the substratum in all cells. However, in flat cells, they frequently cause spindle deformations. Similar deformations are apparent when confined spindles rotate from tilted to parallel positions while MDCK cells progress from prometaphase to metaphase. The spindle disruptions cause the engagement of the spindle assembly checkpoint. We propose that cell rounding serves to maintain spindle integrity during its positioning. PMID:25657320

  10. Mitotic spindle asymmetry in rodents and primates:2D versus 3D measurement methodologies

    Directory of Open Access Journals (Sweden)

    Delphine eDelaunay

    2015-02-01

    Full Text Available Recent data have uncovered that spindle size asymmetry (SSA is a key component of asymmetric cell division in the mouse cerebral cortex (Delaunay et al., 2014. In the present study we show that SSA also occurs during cortical progenitor divisions in the ventricular zone of the macaque cerebral cortex, pointing to a conserved mechanism in the mammalian lineage. Because SSA magnitude is smaller in cortical precursors than in invertebrate neuroblasts, the unambiguous demonstration of volume differences between the two half spindles is considered to require 3D reconstruction of the mitotic spindle (Delaunay et al., 2014. Although straightforward, the 3D analysis of SSA is time consuming, which is likely to hinder SSA identification and prevent further explorations of SSA related mechanisms in generating asymmetric cell division. We therefore set out to develop an alternative method for accurately measuring spindle asymmetry. Based on the mathematically demonstrated linear relationship between 2D and 3D analysis, we show that 2D assessment of spindle size in metaphase cells is as accurate and reliable as 3D reconstruction provided a specific procedure is applied. We have examined the experimental accuracy of the two methods by applying them to different sets of in vivo and in vitro biological data, including mouse and primate cortical precursors. Linear regression analysis demonstrates that the results from 2D and 3D reconstructions are equally powerful. We therefore provide a reliable and efficient technique to measure SSA in mammalian cells.

  11. ATF-2 immunoreactivity in post-mitotic and terminally differentiated human odontoblasts.

    Science.gov (United States)

    Keklikoglu, Nurullah; Akinci, Sevtap

    2015-09-01

    Activating transcription factor 2 (ATF-2/CRE-BP1; cAMP-responsive element binding protein 1) is a member of nuclear transcription factor activator protein-1 (AP-1) family. AP-1 regulates cellular processes including growth, proliferation, differentiation and apoptosis. However, biological relationship of cellular process to each member of the AP-1 family is not clear yet. The objective of the present study was to compare the ATF-2 immunoreactivity in the post-mitotic and terminally differentiated odontoblasts and in the pulpal fibroblasts which can be divided by mitosis when required. Fibroblasts at various stages of differentiation co-exist in the human dental pulp. ATF-2 was investigated immunohistochemically in 20 permanent human teeth. According to the findings obtained, the mean percentage of ATF-2 positive cells was 68.5 ± 19.2% in the odontoblasts and 22.8 ± 13.7% in the pulpal fibroblasts. The comparison of ATF-2 positivity revealed a statistically significant difference between odontoblasts and pulpal fibroblasts. These findings have suggested that ATF-2 is more associated with cell survival rather than cell proliferation, and revealed much of effectiveness in maintaining terminal differentiation than the various differentiation stages of the cells. PMID:25417007

  12. Withaferin-A induces mitotic catastrophe and growth arrest in prostate cancer cells

    Science.gov (United States)

    Roy, Ram V; Suman, Suman; Das, Trinath P.; Luevano, Joe; Damodaran, Chendil

    2014-01-01

    Cell cycle deregulation is strongly associated with the pathogenesis of prostate cancer (CaP). Clinical trials of cell cycle regulators that target either the G0/G1 or G2/M phase to inhibit the growth of cancers including CaP are increasing. In this study, we determined the cell-cycle regulatory potential of the herbal molecule Withaferin-A (WA) on CaP cells. WA induced irreversible G2/M arrest in both CaP cell lines (PC3 and DU145) for 48 h. The G2/M arrest was accompanied by upregulation of phosphorylated Wee1, phophorylated histone H3, p21 and Aurora-B. On the other hand, downregulation of cyclins (E2, A, and B1) and phorphorylated Cdc2 (Tyr15) was observed in WA-treated CaP cells. In addition, decreased levels of phosphorylated Chk1 (Ser345) and Chk2 (Thr68) were evident in WA-treated CaP cells. Our results suggest that activation of Cdc2 leads to accumulation in M-phase, with abnormal duplication, and initiation of mitotic catastrophe that results in cell death. In conclusion, these results clearly highlight the potential of WA as a regulator of the G2/M phase of the cell cycle and as a therapeutic agent for CaP. PMID:24079846

  13. Comprehensive proteomic analysis of interphase and mitotic 14-3-3-binding proteins.

    Science.gov (United States)

    Meek, Sarah E M; Lane, William S; Piwnica-Worms, Helen

    2004-07-30

    14-3-3 proteins regulate the cell division cycle and play a pivotal role in blocking cell cycle advancement after activation of the DNA replication and DNA damage checkpoints. Here we describe a global proteomics analysis to identify proteins that bind to 14-3-3s during interphase and mitosis. 14-3-3-binding proteins were purified from extracts of interphase and mitotic HeLa cells using specific peptide elution from 14-3-3 zeta affinity columns. Proteins that specifically bound and eluted from the affinity columns were identified by microcapillary high pressure liquid chromatography tandem mass spectrometry analysis. Several known and novel 14-3-3-interacting proteins were identified in this screen. Identified proteins are involved in cell cycle regulation, signaling, metabolism, protein synthesis, nucleic acid binding, chromatin structure, protein folding, proteolysis, nucleolar function, and nuclear transport as well as several other cellular processes. In some cases 14-3-3 binding was cell cycle-dependent, whereas in other cases the binding was shown to be cell cycle-independent. This study adds to the growing list of human 14-3-3-binding proteins and implicates a role for 14-3-3 proteins in a plethora of essential biological processes. PMID:15161933

  14. Cell cycle dependence of mitotic delay in X-irradiated normal and ataxia-telangiectasia fibroblasts

    International Nuclear Information System (INIS)

    An investigation has been made of the relationship between suppression in the mitotic index and inhibition of DNA synthesis in normal (N) and ataxia-telangiectasia (A-T) skin fibroblasts, using tritiated thymidine as a marker of the cell cycle. The delay in progression of X-irradiated cells through the cell cycle, which is more pronounced in normal than in A-T fibroblasts, was greatest for cells in G2 at the time of irradiation. The greater effect of radiation on the initiation of DNA synthesis in N than in A-T cells was reflected in the shape of the percent labelled mitosis curves after 3H-thymidine treatment. The duration of the S phase in unirradiated A-T cells was greater than in N cells. It is pointed out that any explanation of the underlying defect in A-T must account not only for the reduced radiosensitivity of DNA synthesis but for the lesser delay in G2. The authors claim that their data support the hypothesis that DNA is the principal target for radiation-induced G2 delay. (U.K.)

  15. Human papillomavirus type 16 E7 perturbs DREAM to promote cellular proliferation and mitotic gene expression.

    Science.gov (United States)

    DeCaprio, J A

    2014-07-31

    The study of the small DNA tumor viruses continues to provide valuable new insights into oncogenesis and fundamental biological processes. Although much has already been revealed about how the human papillomaviruses (HPVs) can transform cells and contribute to cervical and oropharyngeal cancer, there clearly is much more to learn. In this issue of Oncogene, Pang et al., doi:10.1038/onc.2013.426, demonstrate that the high-risk HPV16 E7 oncogene can promote cellular proliferation by interacting with the DREAM (DP, RB-like, E2F and MuvB) complex at two distinct phases of the cell cycle. Consistent with earlier work, HPV16 E7 can bind to the retinoblastoma tumor suppressor (RB) family member p130 (RBL2) protein and promote its proteasome-mediated destruction thereby disrupting the DREAM complex and can prevent exit from the cell cycle into quiescence. In addition, they demonstrate that HPV16 E7 can bind to MuvB core complex in association with BMYB and FOXM1 and activate gene expression during the G2 and M phase of the cell cycle. Thus, HPV16 E7 acts to prevent exit from the cell cycle entry and promotes mitotic proliferation and may account for the high levels of FOXM1 often observed in poor-risk cervical cancers. PMID:24166507

  16. Human papillomavirus type 16 E7 perturbs DREAM to promote cellular proliferation and mitotic gene expression

    Science.gov (United States)

    DeCaprio, James A.

    2014-01-01

    Study of the small DNA tumor viruses continues to provide valuable new insights into oncogenesis and fundamental biological processes. While much has already been revealed about how the human papillomaviruses (HPVs) can transform cells and contribute to cervical and oropharyngeal cancer, there clearly is much more to learn. In this issue of Oncogene, Pang et al. demonstrate that the high-risk HPV16 E7 oncogene can promote cellular proliferation by interacting with the DREAM (DP, RB-like, E2F and MuvB) complex at two distinct phases of the cell cycle (1). Consistent with earlier work, HPV16 E7 can bind to the retinoblastoma tumor suppressor (RB) family member p130 (RBL2) protein and promote its proteasome-mediated destruction thereby disrupting the DREAM complex and prevent exit from the cell cycle into quiescence. In addition, they demonstrate that HPV16 E7 can bind to MuvB core complex in association with BMYB and FOXM1 and activate gene expression during the G2 and M phase of the cell cycle. Thus, HPV16 E7 acts to prevent exit from the cell cycle entry and promotes mitotic proliferation and may account for the high levels of FOXM1 often observed in poor risk cervical cancers. PMID:24166507

  17. Non-anti-mitotic concentrations of taxol reduce breast cancer cell invasiveness

    International Nuclear Information System (INIS)

    Taxol is widely used in breast cancer chemotherapy. Its effects are primarily attributed to its anti-mitotic activity. Microtubule perturbators also exert antimetastatic activities which cannot be explained solely by the inhibition of proliferation. Voltage-dependent sodium channels (NaV) are abnormally expressed in the highly metastatic breast cancer cell line MDA-MB-231 and not in MDA-MB-468 cell line. Inhibiting NaV activity with tetrodotoxin is responsible for an approximately 0.4-fold reduction of MDA-MB-231 cell invasiveness. In this study, we focused on the effect of a single, 2-h application of 10 nM taxol on the two cell lines MDA-MB-231 and MDA-MB-468. At this concentration, taxol had no effect on proliferation after 7 days and on migration in any cell line. However it led to a 40% reduction of transwell invasion of MDA-MB-231 cells. There was no additive effect when taxol and tetrodotoxin were simultaneously applied. NaV activity, as assessed by patch-clamp, indicates that it was changed by taxol pre-treatment. We conclude that taxol can exert anti-tumoral activities, in cells expressing NaV, at low doses that have no effect on cell proliferation. This effect might be due to a modulation of signalling pathways involving sodium channels.

  18. UV-induced mitotic co-segregation of genetic markers in Candida albicans: Evidence for linkage

    International Nuclear Information System (INIS)

    Parasexual genetic studies of the medically important yeast Candida albicans were performed using the method of UV-induced mitotic segregation. UV-irradiation of the Hoffmann-La Roche type culture of C. albicans yielded a limited spectrum of mutants at a relatively high fequency. This observation suggested natural heterozygosity. Canavanine-sensitive (CanS) segregants were induced at a frequency of 7.6 . 10-3. Double mutants that were both CanS and methionine (Met-) auxotrophs were induced at a frequency of 7.4 . 10-3. The single Met- segregant class was missing indicating linkage. UV-induced CanS or Met-CanS segregants occurred occasionally in twin-sectored colonies. Analyses of the sectors as well as the observed and missing classes of segregants indicated that genes met and can are linked in the cis configuration. The proposed gene order is: centromere - met - can. Thus, it is concluded that the Hoffmann-La Roche strain of C. albicans is naturally heterozygous at two linked loci. These findings are consistent with diploidy. (orig.)

  19. Establishment and mitotic characterization of new Drosophila acentriolar cell lines from DSas-4 mutant

    Directory of Open Access Journals (Sweden)

    Nicolas Lecland

    2013-01-01

    In animal cells the centrosome is commonly viewed as the main cellular structure driving microtubule (MT assembly into the mitotic spindle apparatus. However, additional pathways, such as those mediated by chromatin and augmin, are involved in the establishment of functional spindles. The molecular mechanisms involved in these pathways remain poorly understood, mostly due to limitations inherent to current experimental systems available. To overcome these limitations we have developed six new Drosophila cell lines derived from Drosophila homozygous mutants for DSas-4, a protein essential for centriole biogenesis. These cells lack detectable centrosomal structures, astral MT, with dispersed pericentriolar proteins D-PLP, Centrosomin and γ-tubulin. They show poorly focused spindle poles that reach the plasma membrane. Despite being compromised for functional centrosome, these cells could successfully undergo mitosis. Live-cell imaging analysis of acentriolar spindle assembly revealed that nascent MTs are nucleated from multiple points in the vicinity of chromosomes. These nascent MTs then grow away from kinetochores allowing the expansion of fibers that will be part of the future acentriolar spindle. MT repolymerization assays illustrate that acentriolar spindle assembly occurs “inside-out” from the chromosomes. Colchicine-mediated depolymerization of MTs further revealed the presence of a functional Spindle Assembly Checkpoint (SAC in the acentriolar cells. Finally, pilot RNAi experiments open the potential use of these cell lines for the molecular dissection of anastral pathways in spindle and centrosome assembly.

  20. Mitotic recombination in callus of Arabidopsis thaliana (L.) Heynh. after fast-neutron treatment

    International Nuclear Information System (INIS)

    A threefold heterozygote plant, with marker genes as, py and er linked on chromosome 2, was used as material for this study. Six-week-old callus from leaves of this plant was irradiated with 15, 20 and 30 Gy of fast neutrons. A normal segregation was observed in the F2 progenies of about 65% of the callus-derived plants. On the other hand, a statistically significant deviation from the expected ratio of 3:1 was found in progenies of 17 other callus-derived plants. Only wild-type plants were observed in progenies of two plants from irradiated callus. Similar wild-type plants were present, at much higher frequencies than expected, in progenies of 15 other plants from irradiated callus. The mitotic recombination between the er gene and centromere in callus cells could be a reason for the appearance of only wild-type forms in progenies of callus-derived plants. These and other results presented in the paper suggest that fast-neutron irradiation may significantly increase the level of somatic crossing-over in callus cells. (author)

  1. Knob-associated tandem repeats on mitotic chromosomes in maize, Zea diploperennis and their hybrids

    Institute of Scientific and Technical Information of China (English)

    XIONG Zhiyong; GAO Yuan; HE Guanyuan; GU Mingguang; GUO Lequn; SONG Yunchun

    2004-01-01

    Knob-associated tandem repeats, 180-bp repeats and TR-1 elements, together with 45S rDNA were located on mitotic chromosomes of Zea diploperennis (DP),maize inbred line F102 and their hybrid. In DP, knobs on the short arm of chromosomes 1 and 4 and on the long arm of the chromosomes 4 and 5 are composed predominantly of the 180-bp repeats. In addition, 180-bp repeats existed together with TR-1 elements were also detected on the short arm of chromosomes 2 and 5 and on the long arm of the chromosomes 2, 6, 7, 8 and 9. In maize inbred line F102, 180-bp repeats were present in chromosomes 7S and one homologue of chromosomes 8L. TR-1 elements appeared on satellite of chromosome 6 and no detectable hybridization site co-located with 180-bp repeats was observed in maize F102. Polymorphism of size, number, and distribution of 180-bp and TR-1 signals were revealed among different chromosomes in these two species and heteromorphism existed between some homologous chromosomes in the same species.Using these excellent landmarks, the interspecific hybrid of maize and DP were identified. The results suggest that comparative analysis of 180-bp repeats and TR-1 elements may help understand the genome organization and the evolution in Zea.

  2. Mitotic segregation in intergeneric hybrids of yeast to give novel genetic segregants

    International Nuclear Information System (INIS)

    Two strains of yeast, Yarrowia lipolytica and Saccharomycopsis fibuligera, have a filamentous growth form in addition to budding cells. Y. lipolytica produces lipases and is used in the production of citric acid while W. fibuligera produces amylases and is used in the production of rice wine. In the present report, we made a study of the following: (i) karyotypes of the two yeast strains to obtain a better understanding of their genetic relatedness, (ii) genetic crosses between the two strains to produce intergeneric hybrids, and (iii) pattern of genetic segregation of the intergeneric hybrids via the mitotic process. The results of our studies showed that the two yeast strains were genetically related and that putative intergeneric hybrids were obtained by a genetic crossing of the strains. The hybrids were relatively stable in mitosis as compared to their parent strains. However, in prolonged vegetative propagation, the hybrids gave rise to genetic segregants, most of which were of either of the two parent phenotypes. A number of the segregants had phenotypes which combined those of the parental types. Of considerable significance was that yet a few others were novel as they exhibited phenotypes not hitherto seen for both parent strains. (author). 9 refs, 5 tabs

  3. Lysine-specific demethylase-1 (LSD1) is compartmentalized at nuclear chromocenters in early post-mitotic cells of the olfactory sensory neuronal lineage.

    Science.gov (United States)

    Kilinc, Seda; Savarino, Alyssa; Coleman, Julie H; Schwob, James E; Lane, Robert P

    2016-07-01

    Mammalian olfaction depends on the development of specialized olfactory sensory neurons (OSNs) that each express one odorant receptor (OR) protein from a large family of OR genes encoded in the genome. The lysine-specific demethylase-1 (LSD1) protein removes activating H3K4 or silencing H3K9 methylation marks at gene promoters and is required for proper OR regulation. We show that LSD1 protein exhibits variable organization within nuclei of developing OSNs, and tends to consolidate into a single dominant compartment at the edges of chromocenters within nuclei of early post-mitotic cells of the mouse olfactory epithelium (MOE). Using an immortalized cell line derived from developing olfactory placode, we show that consolidation of LSD1 appears to be cell-cycle regulated, with a peak occurrence in early G1. LSD1 co-compartmentalizes with CoREST, a protein known to collaborate with LSD1 to carry out a variety of chromatin-modifying functions. We show that LSD1 compartments co-localize with 1-3 OR loci at the exclusion of most OR genes, and commonly associate with Lhx2, a transcription factor involved in OR regulation. Together, our data suggests that LSD1 is sequestered into a distinct nuclear space that might restrict a histone-modifying function to a narrow developmental time window and/or range of OR gene targets. PMID:26947098

  4. Exposure of Human Lung Cancer Cells to 8-Chloro-Adenosine Induces G2/M Arrest and Mitotic Catastrophe

    Directory of Open Access Journals (Sweden)

    Hong-Yu Zhang

    2004-11-01

    Full Text Available 8-Chloro-adenosine (8-CI-Ado is a potent chemotherapeutic agent whose cytotoxicity in a variety of tumor cell lines has been widely investigated. However, the molecular mechanisms are uncertain. In this study, we found that exposure of human lung cancer cell lines A549 (p53-wt and H1299 (p53-depleted to 8-CI-Ado induced cell arrest in the G2/M phase, which was accompanied by accumulation of binucleated and polymorphonucleated cells resulting from aberrant mitosis and failed cytokinesis. Western blotting showed the loss of phosphorylated forms of Cdc2 and Cdc25C that allowed progression into mitosis. Furthermore, the increase in Ser10-phosphorylated histone H3-positive cells revealed by fluorescence-activated cell sorting suggested that the agent-targeted cells were able to exit the G2 phase and enter the M phase. Immunocytochemistry showed that microtubule and microfilament arrays were changed in exposed cells, indicating that the dynamic instability of microtubules and microfilaments was lost, which may correlate with mitotic dividing failure. Aberrant mitosis resulted in mitotic catastrophe followed by varying degrees of apoptosis, depending on the cell lines. Thus, 8-CI-Ado appears to exert its cytotoxicity toward cells in culture by inducing mitotic catastrophe.

  5. C646, a selective small molecule inhibitor of histone acetyltransferase p300, radiosensitizes lung cancer cells by enhancing mitotic catastrophe

    International Nuclear Information System (INIS)

    Background and purpose: Chromatin remodeling through histone modifications, including acetylation, plays an important role in the appropriate response to DNA damage induced by ionizing radiation (IR). Here we investigated the radiosensitizing effect of C646, a selective small molecule inhibitor of p300 histone acetyltransferase, and explored the underlying mechanisms. Materials and methods: A549, H157 and H460 human non-small cell lung carcinoma (NSCLC) cells, and HFL-III human lung fibroblasts were assessed by clonogenic survival assay. Apoptosis and necrosis were assessed by annexin V staining. Senescence was assessed by Senescence-associated β-galactosidase staining. Mitotic catastrophe was assessed by evaluating nuclear morphology with DAPI staining. Cell cycle profiles were analyzed by flow cytometry. Protein expression was analyzed by immunoblotting. Results: C646 sensitized A549, H460 and H157 cells to IR with a dose enhancement ratio at 10% surviving fraction of 1.4, 1.2 and 1.2, respectively. C646 did not radiosensitize HFL-III cells. In A549 cells, but not in HFL-III cells, C646 (i) enhanced mitotic catastrophe but not apoptosis, necrosis, or senescence after IR; (ii) increased the hyperploid cell population after IR; and (iii) suppressed the phosphorylation of CHK1 after IR. Conclusions: C646 radiosensitizes NSCLC cells by enhancing mitotic catastrophe through the abrogation of G2 checkpoint maintenance

  6. Bovine papillomavirus type 1 genomes and the E2 transactivator protein are closely associated with mitotic chromatin.

    Science.gov (United States)

    Skiadopoulos, M H; McBride, A A

    1998-03-01

    The bovine papillomavirus type 1 E2 transactivator protein is required for viral transcriptional regulation and DNA replication and may be important for long-term episomal maintenance of viral genomes within replicating cells (M. Piirsoo, E. Ustav, T. Mandel, A. Stenlund, and M. Ustav, EMBO J. 15:1-11, 1996). We have evidence that, in contrast to most other transcriptional transactivators, the E2 transactivator protein is associated with mitotic chromosomes in dividing cells. The shorter E2-TR and E8/E2 repressor proteins do not bind to mitotic chromatin, and the N-terminal transactivation domain of the E2 protein is necessary for the association. However, the DNA binding function of E2 is not required. We have found that bovine papillomavirus type 1 genomes are also associated with mitotic chromosomes, and we propose a model in which E2-bound viral genomes are transiently associated with cellular chromosomes during mitosis to ensure that viral genomes are segregated to daughter cells in approximately equal numbers. PMID:9499063

  7. Tumor environmental factors glucose deprivation and lactic acidosis induce mitotic chromosomal instability--an implication in aneuploid human tumors.

    Directory of Open Access Journals (Sweden)

    Chunyan Dai

    Full Text Available Mitotic chromosomal instability (CIN plays important roles in tumor progression, but what causes CIN is incompletely understood. In general, tumor CIN arises from abnormal mitosis, which is caused by either intrinsic or extrinsic factors. While intrinsic factors such as mitotic checkpoint genes have been intensively studied, the impact of tumor microenvironmental factors on tumor CIN is largely unknown. We investigate if glucose deprivation and lactic acidosis--two tumor microenvironmental factors--could induce cancer cell CIN. We show that glucose deprivation with lactic acidosis significantly increases CIN in 4T1, MCF-7 and HCT116 scored by micronuclei, or aneuploidy, or abnormal mitosis, potentially via damaging DNA, up-regulating mitotic checkpoint genes, and/or amplifying centrosome. Of note, the feature of CIN induced by glucose deprivation with lactic acidosis is similar to that of aneuploid human tumors. We conclude that tumor environmental factors glucose deprivation and lactic acidosis can induce tumor CIN and propose that they are potentially responsible for human tumor aneuploidy.

  8. Killing the umpire: cooperative defects in mitotic checkpoint and BRCA2 genes on the road to transformation

    International Nuclear Information System (INIS)

    Recent findings from mouse models of BRCA2 genetic lesions have provided intriguing insights and important questions concerning modes of tumor development in familial breast and ovarian cancers. Fibroblasts from mice homozygous for the BRCA2Tr allele grow poorly and display an array of chromosomal abnormalities that are consistent with a role for BRCA2 in DNA repair. This growth defect can be overcome and cellular transformation promoted by the expression of defective, dominant negative alleles of p53 and of the mitotic checkpoint gene Bub1, both of which are known to induce chromosome instability. These findings are mirrored in the genetic lesions sustained in tumors found in the rare BRCA2Tr/Trmice that survive to adulthood, which include defects in p53 as well as the mitotic checkpoint proteins Bub1 and Mad3L. Together, these data hint that tumors in these mice evolve from an unusually intense selective pressure to remove DNA damage checkpoints, which in turn might be facilitated by chromosomal abolition of mitotic checkpoints and the consequent increase in shuffling of genetic information. How these genetic lesions co-operate to yield transformed cells and how these data relate to BRCA1 and BRCA2 defects in the human population are important questions raised by this work

  9. Aurora A Kinase Regulates Mammary Epithelial Cell Fate by Determining Mitotic Spindle Orientation in a Notch-Dependent Manner

    Directory of Open Access Journals (Sweden)

    Joseph L. Regan

    2013-07-01

    Full Text Available Cell fate determination in the progeny of mammary epithelial stem/progenitor cells remains poorly understood. Here, we have examined the role of the mitotic kinase Aurora A (AURKA in regulating the balance between basal and luminal mammary lineages. We find that AURKA is highly expressed in basal stem cells and, to a lesser extent, in luminal progenitors. Wild-type AURKA expression promoted luminal cell fate, but expression of an S155R mutant reduced proliferation, promoted basal fate, and inhibited serial transplantation. The mechanism involved regulation of mitotic spindle orientation by AURKA and the positioning of daughter cells after division. Remarkably, this was NOTCH dependent, as NOTCH inhibitor blocked the effect of wild-type AURKA expression on spindle orientation and instead mimicked the effect of the S155R mutant. These findings directly link AURKA, NOTCH signaling, and mitotic spindle orientation and suggest a mechanism for regulating the balance between luminal and basal lineages in the mammary gland.

  10. Cold-sensitive mutants of p34cdc2 that suppress a mitotic catastrophe phenotype in fission yeast.

    Science.gov (United States)

    Ayscough, K; Hayles, J; MacNeill, S A; Nurse, P

    1992-04-01

    The p34cdc2 protein kinase plays a central role in the regulation of the eukaryotic cell cycle, being required both in late G1 for the commitment to S-phase and in late G2 for the initiation of mitosis. p34cdc2 also determines the precise timing of entry into mitosis in fission yeast, where a number of gene products that regulate p34cdc2 activity have been identified and characterised. To investigate further the mitotic role of p34cdc2 in this organism we have isolated new cold-sensitive p34cdc2 mutants. These are defective only in their G2 function and are extragenic suppressors of the lethal premature entry into mitosis brought about by mutating the mitotic inhibitor p107wee1 and overproducing the mitotic activator p80cdc25. One of the mutant proteins p34cdc2-E8 is only functional in the absence of p107wee1, and all the mutant strains have reduced histone H1 kinase activity in vitro. Each mutant allele has been cloned and sequenced, and the lesions responsible for the cold-sensitive phenotypes identified. All the mutations were found to map to regions that are conserved between the fission yeast p34cdc2 and functional homologues from higher eukaryotes. PMID:1316996

  11. Radiation-induced chromosome aberrations occurring during the first and second mitotic divisions of human lymphocytes after exposure to X-rays

    International Nuclear Information System (INIS)

    The study described elucidates the relationship between the number of chromosome aberrations/cell and the length of the individual culture periods. The BrdUrd staining technique used here permitted separate analyses of the aberration frequencies during the first and second mitotic divisions of lymphocyte chromosomes cultivated for different periods of time. The results thus obtained were subjected to further analysis to quantify errors attributable to non-separate observations of such mitotic processes and to determine the number of surviving cells observed to have dicentric chromosomes during the first mitotic division. An analysis of the results obtained for the first mitotic division alone failed to reveal any connection between the number of chromosome aberrations and the culture periods. The aberration rates established for the second mitotic division were clearly related to the culture periods. A steady state was reached after approx. 54 h, but the values determined here were much lower than those calculated for the first mitotic division. (orig./MG)

  12. X-ray induced increase of mitotic crossovers in Nicotiana tabacum

    International Nuclear Information System (INIS)

    The mutant, sulfur, of Nicotiana tabacum L. is an incompletely dominant chlorophyll deficiency locus. Single dark green (D) or aurea (A), and double (Db) spots developed spontaneously in leaves of Su/su plants. When Su/su seedlings were exposed to 150 kVP X-rays (50, 150 or 300 R), the frequency of each kind of spot increased with exposure dose. If D, A, and Db spots were summed, the frequencies of spots per leaf were 0.458 +- 0.069, 2.317 +- 0.219, 11.95 +- 1.212, and 35.29 +-5.221 for 0 (control), 50, 150, and 300 R, respectively. Based on the number of leaves with increased spot frequency at 300 R, there were seven leaf initials present at the time of treatment, each at a different developmental stage. The mutation response curve obtained from the eventual seven leaves was similar to curves of mutation induction in other plant species. Uniform sensitivity of cells in leaf primordia to X-irradiation was inferred since D and A spot areas as percentages of the leaf blade area respectively remained equal from leaf to leaf. Spot frequency was also examined geographically over the leaf blade. Right versus left halves had equal frequency spotting; distal halves, with more cells by actual count than proximal halves, had a greater spot frequency. The dose responses of D and A spots had significant linear and non-linear components. The Db spot frequency in these experiments was used to calculate the frequency of mitotic crossing over. The values range from 3.34 x 10-6 to 9.64 x 10-4, extending an earlier estimate. (author)

  13. Mitotic evolution of Plasmodium falciparum shows a stable core genome but recombination in antigen families.

    Directory of Open Access Journals (Sweden)

    Selina E R Bopp

    Full Text Available Malaria parasites elude eradication attempts both within the human host and across nations. At the individual level, parasites evade the host immune responses through antigenic variation. At the global level, parasites escape drug pressure through single nucleotide variants and gene copy amplification events conferring drug resistance. Despite their importance to global health, the rates at which these genomic alterations emerge have not been determined. We studied the complete genomes of different Plasmodium falciparum clones that had been propagated asexually over one year in the presence and absence of drug pressure. A combination of whole-genome microarray analysis and next-generation deep resequencing (totaling 14 terabases revealed a stable core genome with only 38 novel single nucleotide variants appearing in seventeen evolved clones (avg. 5.4 per clone. In clones exposed to atovaquone, we found cytochrome b mutations as well as an amplification event encompassing the P. falciparum multidrug resistance associated protein (mrp1 on chromosome 1. We observed 18 large-scale (>1 kb on average deletions of telomere-proximal regions encoding multigene families, involved in immune evasion (9.5×10(-6 structural variants per base pair per generation. Six of these deletions were associated with chromosomal crossovers generated during mitosis. We found only minor differences in rates between genetically distinct strains and between parasites cultured in the presence or absence of drug. Using these derived mutation rates for P. falciparum (1.0-9.7×10(-9 mutations per base pair per generation, we can now model the frequency at which drug or immune resistance alleles will emerge under a well-defined set of assumptions. Further, the detection of mitotic recombination events in var gene families illustrates how multigene families can arise and change over time in P. falciparum. These results will help improve our understanding of how P. falciparum

  14. Inhibition of Survivin and Aurora B Kinase Sensitizes Mesothelioma Cells by Enhancing Mitotic Arrests

    International Nuclear Information System (INIS)

    Purpose: Survivin, a member of the inhibitor of apoptosis gene family, has also been shown to regulate mitosis. It binds Aurora B kinase and the inner centromere protein to form the chromosome passenger complex. Both Aurora B and survivin are overexpressed in many tumors. In this study, we examined whether irradiation affected survivin and Aurora B expression in mesothelioma cells, and how inhibition of these molecules affected radiosensitivity. Methods and Materials: ZM447439 and survivin antisense oligonucleotides were used to inhibit survivin and Aurora B kinase respectively. Western blot was performed to determine the expression of survivin, Aurora B, phosphorylated-histone H3 (Ser 10), and caspase cleavage. Multinucleated cells were counted using flow cytometry, and cell survival after treatment was determined using clonogenic assay. Results: At 3-Gy irradiation an increase was observed in levels of survivin and Aurora B as well as the kinase activity of Aurora B, with an increase in G2/M phase. The radiation-induced upregulation of these molecules was effectively attenuated by antisense oligonucleotides against survivin and a small-molecule inhibitor of Aurora B, ZM447439. Dual inhibition of survivin and Aurora B synergistically radiosensitized mesothelioma cells with a dose enhancement ratio of 2.55. This treatment resulted in increased formation of multinucleated cells after irradiation but did not increase levels of cleaved caspase 3. Conclusion: Inhibition of survivin and Aurora B induces mitotic cell arrest in mesothelioma cells after irradiation. These two proteins may be potential therapeutic targets for the enhancement of radiotherapy in malignant pleural mesothelioma

  15. Phosphoproteomic Profiling Reveals Epstein-Barr Virus Protein Kinase Integration of DNA Damage Response and Mitotic Signaling.

    Science.gov (United States)

    Li, Renfeng; Liao, Gangling; Nirujogi, Raja Sekhar; Pinto, Sneha M; Shaw, Patrick G; Huang, Tai-Chung; Wan, Jun; Qian, Jiang; Gowda, Harsha; Wu, Xinyan; Lv, Dong-Wen; Zhang, Kun; Manda, Srikanth S; Pandey, Akhilesh; Hayward, S Diane

    2015-12-01

    Epstein-Barr virus (EBV) is etiologically linked to infectious mononucleosis and several human cancers. EBV encodes a conserved protein kinase BGLF4 that plays a key role in the viral life cycle. To provide new insight into the host proteins regulated by BGLF4, we utilized stable isotope labeling by amino acids in cell culture (SILAC)-based quantitative proteomics to compare site-specific phosphorylation in BGLF4-expressing Akata B cells. Our analysis revealed BGLF4-mediated hyperphosphorylation of 3,046 unique sites corresponding to 1,328 proteins. Frequency analysis of these phosphosites revealed a proline-rich motif signature downstream of BGLF4, indicating a broader substrate recognition for BGLF4 than its cellular ortholog cyclin-dependent kinase 1 (CDK1). Further, motif analysis of the hyperphosphorylated sites revealed enrichment in ATM, ATR and Aurora kinase substrates while functional analyses revealed significant enrichment of pathways related to the DNA damage response (DDR), mitosis and cell cycle. Phosphorylation of proteins associated with the mitotic spindle assembly checkpoint (SAC) indicated checkpoint activation, an event that inactivates the anaphase promoting complex/cyclosome, APC/C. Furthermore, we demonstrated that BGLF4 binds to and directly phosphorylates the key cellular proteins PP1, MPS1 and CDC20 that lie upstream of SAC activation and APC/C inhibition. Consistent with APC/C inactivation, we found that BGLF4 stabilizes the expression of many known APC/C substrates. We also noted hyperphosphorylation of 22 proteins associated the nuclear pore complex, which may contribute to nuclear pore disassembly and SAC activation. A drug that inhibits mitotic checkpoint activation also suppressed the accumulation of extracellular EBV virus. Taken together, our data reveal that, in addition to the DDR, manipulation of mitotic kinase signaling and SAC activation are mechanisms associated with lytic EBV replication. All MS data have been deposited in

  16. Carbon-ion beam irradiation kills X-ray-resistant p53-null cancer cells by inducing mitotic catastrophe.

    Directory of Open Access Journals (Sweden)

    Napapat Amornwichet

    Full Text Available BACKGROUND AND PURPOSE: To understand the mechanisms involved in the strong killing effect of carbon-ion beam irradiation on cancer cells with TP53 tumor suppressor gene deficiencies. MATERIALS AND METHODS: DNA damage responses after carbon-ion beam or X-ray irradiation in isogenic HCT116 colorectal cancer cell lines with and without TP53 (p53+/+ and p53-/-, respectively were analyzed as follows: cell survival by clonogenic assay, cell death modes by morphologic observation of DAPI-stained nuclei, DNA double-strand breaks (DSBs by immunostaining of phosphorylated H2AX (γH2AX, and cell cycle by flow cytometry and immunostaining of Ser10-phosphorylated histone H3. RESULTS: The p53-/- cells were more resistant than the p53+/+ cells to X-ray irradiation, while the sensitivities of the p53+/+ and p53-/- cells to carbon-ion beam irradiation were comparable. X-ray and carbon-ion beam irradiations predominantly induced apoptosis of the p53+/+ cells but not the p53-/- cells. In the p53-/- cells, carbon-ion beam irradiation, but not X-ray irradiation, markedly induced mitotic catastrophe that was associated with premature mitotic entry with harboring long-retained DSBs at 24 h post-irradiation. CONCLUSIONS: Efficient induction of mitotic catastrophe in apoptosis-resistant p53-deficient cells implies a strong cancer cell-killing effect of carbon-ion beam irradiation that is independent of the p53 status, suggesting its biological advantage over X-ray treatment.

  17. Phosphorylation of HPV-16 E2 at serine 243 enables binding to Brd4 and mitotic chromosomes.

    Directory of Open Access Journals (Sweden)

    Szu-Wei Chang

    Full Text Available The papillomavirus E2 protein is involved in the maintenance of persistent infection and known to bind either to cellular factors or directly to mitotic chromosomes in order to partition the viral genome into the daughter cells. However, how the HPV-16 E2 protein acts to facilitate partitioning of the viral genome remains unclear. In this study, we found that serine 243 of HPV-16 E2, located in the hinge region, is crucial for chromosome binding during mitosis. Bromodomain protein 4 (Brd4 has been identified as a cellular binding target through which the E2 protein of bovine papillomavirus type 1 (BPV-1 tethers the viral genome to mitotic chromosomes. Mutation analysis showed that, when the residue serine 243 was substituted by glutamic acid or aspartic acid, whose negative charges mimic the effect of constitutive phosphorylation, the protein still can interact with Brd4 and colocalize with Brd4 in condensed metaphase and anaphase chromosomes. However, substitution by the polar uncharged residues asparagine or glutamine abrogated Brd4 and mitotic chromosome binding. Moreover, following treatment with the inhibitor JQ1 to release Brd4 from the chromosomes, Brd4 and E2 formed punctate foci separate from the chromosomes, further supporting the hypothesis that the association of the HPV-16 E2 protein with the chromosomes is Brd4-dependent. In addition, the S243A E2 protein has a shorter half-life than the wild type, indicating that phosphorylation of the HPV-16 E2 protein at serine 243 also increases its half-life. Thus, phosphorylation of serine 243 in the hinge region of HPV-16 E2 is essential for interaction with Brd4 and required for host chromosome binding.

  18. Temperature-dependent Index of Mitotic Interval (τ0) for Chromosome Manipulation in Korean Bullhead, Pseudobagrus fulvidraco

    OpenAIRE

    Lim, Sang Gu; Han, Hyoung Kyun; Gil, Hyun Woo; Park, In-Seok

    2012-01-01

    Korean bullhead (Pseudobagrus fulvidraco) was collected from the Kum River areas of Kangkyung-eup, Nonsan city, Chungcheongnam-do, Korea, from April to June, 2012 and was fertilized in order to observe egg development and temperature-related cleavage rates and mitotic intervals (τ0). The fertilized eggs were separative, demersal and light yellowish with 1.5±0.06 mm in diameter, and did not contain oil globules. The first cleavage stages were 90 min, 80 min, 60 min and 50 min at 21°C , 24°C , ...

  19. Ophiopogonin B induces apoptosis, mitotic catastrophe and autophagy in A549 cells.

    Science.gov (United States)

    Chen, Meijuan; Guo, Yuanyuan; Zhao, Ruolin; Wang, Xiaoxia; Jiang, Miao; Fu, Haian; Zhang, Xu

    2016-07-01

    Ophiopogonin B (OP-B), a saponin compound isolated from Radix Ophiopogon japonicus, was verified to inhibit cell proliferation in numerous non-small cell lung cancer (NSCLC) cells in our previous study. However, the precise mechanisms of action have remained unclear. In the present study, we mainly investigated the effects of OP-B on adenocarcinoma A549 cells to further elaborate the underlying mechanisms of OP-B in different NSCLC cell lines. Detection by high content screening (HCS) and TUNEL assay verified that OP-B induced apoptosis in this cell line, while detection of Caspase-3, Bcl-2 and Bax showed that OP-B induced cell death was caspase and mitochondrial independent. Further experiments showed that OP-B induced cell cycle arrest in the S and G2/M phases by inhibiting the expression of Myt1 and phosphorylation of Histone H3 (Ser10), which resulted in mitotic catastrophe in the cells. Transmission electron microscopy (TEM) observation of cell micro-morphology combined with detection of Atgs by western blot analysis showed that OP-B induced autophagy in this cell line. Autophagy inhibition by the lysosome inhibitor CQ or Beclin1-siRNA knockdown both attenuated cell viability, demonstrated that autophagy also being the vital reason resulted in cell death. More importantly, the xenograft model using A549 cells provided further evidence of the inhibition of OP-B on tumor proliferation. Immunohistochemistry detection of LC3 and Tunel assay both verified that high dose of OP-B (75 mg/kg) induced autophagy and apoptosis in vivo, and western blot detection of p-Histone H3 (Ser10), Survivin and XIAP further indicated the molecular mechanism of OP-B in vivo. As our findings revealed, multiple types of cell death overlapped in OP-B treated A549 cells, it displayed multitarget characteristics of the compounds extracted from the Chinese herbal, which may be used as candidate anticancer medicine in clinic. PMID:27175570

  20. Performance of 181 chemicals in a Drosophila assay predominantly monitoring interchromosomal mitotic recombination.

    Science.gov (United States)

    Vogel, E W; Nivard, M J

    1993-01-01

    An evaluation is presented of the effects of 181 chemicals in the (white/white+) (w/w+) eye mosaic assay, an in vivo short-term test measuring genetic damage in somatic cells of Drosophila after treatment of larvae. The genetic principle of this system is loss of heterozygosity for the wild-type reporter gene w+, an event predominantly resulting from homologous interchromosomal mitotic recombination between the two X chromosomes of female genotypes. The w/w+ eye mosaic test detects a broad spectrum of DNA modifications, since all distinct classes of genotoxins are monitored. Non-DNA-reactive chemicals are in principle not detected by this system. Occasional positive responses obtained for chemicals such as amitrole, ethionine and hexachloeroethane are probably not related to the mechanism responsible for their tumorigenicity. The principle outcome of this analysis is the necessity for classification of responses into three categories. (i) Positive, '++'. The 92 chemicals (Tables II and III) falling into this category were clearly recombinagenic in the assay, meaning that dose-response relations were obtained (or could have been established as was evident from the strong responses obtained at one or two exposure doses). Among the 92 chemicals were 49 promutagens including volatile chemicals such as vinyl bromide and vinyl chloride. (ii) Marginally positive, '+w'. The definition of a weakly positive response is the absence of a dose-response relationship due to the fact that a weak but reproducible effect, in most cases no more than a doubling of the spontaneous clone frequency, is inherently related to toxicity. The 40 chemicals (Tables IV and V) belonging to this category mainly represented four distinct types. (a) Procarcinogens, such as 2-acetylaminofluorene, dibenz[a,h]anthracene, p-dimethylaminoazobenzene, 2-naphthylamine and safrole, for which metabolic conversion was the apparent problem in the assay. (b) Electrophilic chemicals of high nucleophilic

  1. Optimizing the Utility of Allium cepa L. var. aggregatum (sibuyas Tagalog for the Allium Test by Elucidating its Mitotic Periodicity and Rhythmicity Under Varying Light Conditions

    Directory of Open Access Journals (Sweden)

    Ambrocio Melvin A. Matias

    2011-06-01

    Full Text Available The occurrence of pattern of mitotic activity has long been studied in different plants; in the onion Allium cepa, determination of its mitotic activity has led to its utilization in the Allium test for cytotoxicity and mutagenicity of test substances. In this study, the pattern of mitotic activity of A. cepa var. aggregatum and the effect of light exposure on mitotic activity were determined to test the utility of A. cepa L. var. aggregatum as an alternative to the common onion, A. cepa, for the Allium test. Bulblets of A. cepa var. aggregatum were allowed to root for three days in tap water under three different set-ups: constant light exposure set-up (Light, constant dark set-up (Dark and 12 hours light-12 hours dark set-up (Light-Dark. The root tips from the bulblets were then excised and subjected to microscopic observation for the mitotic index (MI every hour after the third day. The results showed no significant difference observed across the three set-ups. However, MI for the Dark and Light set-ups were periodic, showing peaks or maxima of MI falling between 11 AM and 12 PM, whereas that of Light-Dark set-up was rhythmic, having an hourly fluctuation, but also showed maximum between 11 AM and 12 PM. It is recommended that A. cepa var. aggregatum root tips be excised between 11 AM and 12 PM for the Allium test.

  2. The proteolysis of mitotic cyclins in mammalian cells persists from the end of mitosis until the onset of S phase.

    Science.gov (United States)

    Brandeis, M; Hunt, T

    1996-10-01

    We have studied how the cell cycle-specific oscillations of mitotic B-type cyclins are generated in mouse fibroblasts. A reporter enzyme comprising the N-terminus of a B-type cyclin fused to bacterial chloramphenicol acetyl transferase (CAT) was degraded at the end of mitosis like endogenous cyclins. Point mutations in the destruction box of this construct completely abolished its mitotic instability. When the destructible reporter was driven by the cyclin B2 promoter, CAT activity mimicked the oscillations in the level of the endogenous cyclin B2. These oscillations were largely conserved when the reporter was transcribed constitutively from the SV40 promoter. Pulse-chase experiments or addition of the proteasome inhibitors lactacystin and ALLN showed that cyclin synthesis continued after the end of mitosis. The destruction box-specific degradation of cyclins normally ceases at the onset of S phase, and is active in fibroblasts arrested in G0 and in differentiated C2 myoblasts. We were able to reproduce this proteolysis in vitro in extracts of synchronized cells. Extracts of G1 cells degraded cyclin B1 whereas p27Kip1 was stable, in contrast, cyclin B1 remained stable and p27Kip1 was degraded in extracts of S phase cells. PMID:8895573

  3. Novel Mad2-targeting miR-493-3p controls mitotic fidelity and cancer cells' sensitivity to paclitaxel.

    Science.gov (United States)

    Tambe, Mahesh; Pruikkonen, Sofia; Mäki-Jouppila, Jenni; Chen, Ping; Elgaaen, Bente Vilming; Straume, Anne Hege; Huhtinen, Kaisa; Cárpen, Olli; Lønning, Per Eystein; Davidson, Ben; Hautaniemi, Sampsa; Kallio, Marko J

    2016-03-15

    The molecular pathways that contribute to the proliferation and drug response of cancer cells are highly complex and currently insufficiently characterized. We have identified a previously unknown microRNA-based mechanism that provides cancer cells means to stimulate tumorigenesis via increased genomic instability and, at the same time, evade the action of clinically utilized microtubule drugs. We demonstrate miR-493-3p to be a novel negative regulator of mitotic arrest deficient-2 (MAD2), an essential component of the spindle assembly checkpoint that monitors the fidelity of chromosome segregation. The microRNA targets the 3' UTR of Mad2 mRNA thereby preventing translation of the Mad2 protein. In cancer cells, overexpression of miR-493-3p induced a premature mitotic exit that led to increased frequency of aneuploidy and cellular senescence in the progeny cells. Importantly, excess of the miR-493-3p conferred resistance of cancer cells to microtubule drugs. In human neoplasms, miR-493-3p and Mad2 expression alterations correlated with advanced ovarian cancer forms and high miR-493-3p levels were associated with reduced survival of ovarian and breast cancer patients with aggressive tumors, especially in the paclitaxel therapy arm. Our results suggest that intratumoral profiling of miR-493-3p and Mad2 levels can have diagnostic value in predicting the efficacy of taxane chemotherapy. PMID:26943585

  4. Genome-wide high-resolution mapping of UV-induced mitotic recombination events in Saccharomyces cerevisiae.

    Directory of Open Access Journals (Sweden)

    Yi Yin

    2013-10-01

    Full Text Available In the yeast Saccharomyces cerevisiae and most other eukaryotes, mitotic recombination is important for the repair of double-stranded DNA breaks (DSBs. Mitotic recombination between homologous chromosomes can result in loss of heterozygosity (LOH. In this study, LOH events induced by ultraviolet (UV light are mapped throughout the genome to a resolution of about 1 kb using single-nucleotide polymorphism (SNP microarrays. UV doses that have little effect on the viability of diploid cells stimulate crossovers more than 1000-fold in wild-type cells. In addition, UV stimulates recombination in G1-synchronized cells about 10-fold more efficiently than in G2-synchronized cells. Importantly, at high doses of UV, most conversion events reflect the repair of two sister chromatids that are broken at approximately the same position whereas at low doses, most conversion events reflect the repair of a single broken chromatid. Genome-wide mapping of about 380 unselected crossovers, break-induced replication (BIR events, and gene conversions shows that UV-induced recombination events occur throughout the genome without pronounced hotspots, although the ribosomal RNA gene cluster has a significantly lower frequency of crossovers.

  5. Calbindin and parvalbumin are early markers of non-mitotically regenerating hair cells in the bullfrog vestibular otolith organs

    Science.gov (United States)

    Steyger, P. S.; Burton, M.; Hawkins, J. R.; Schuff, N. R.; Baird, R. A.

    1997-01-01

    Earlier studies have demonstrated hair cell regeneration in the absence of cell proliferation, and suggested that supporting cells could phenotypically convert into hair cells following hair cell loss. Because calcium-binding proteins are involved in gene up-regulation, cell growth, and cell differentiation, we wished to determine if these proteins were up-regulated in scar formations and regenerating hair cells following gentamicin treatment. Calbindin and parvalbumin immunolabeling was examined in control or gentamicin-treated (GT) bullfrog saccular and utricular explants cultured for 3 days in amphibian culture medium or amphibian culture medium supplemented with aphidicolin, a blocker of nuclear DNA replication in eukaryotic cells. In control cultures, calbindin and parvalbumin immunolabeled the hair bundles and, less intensely, the cell bodies of mature hair cells. In GT or mitotically-blocked GT (MBGT) cultures, calbindin and parvalbumin immunolabeling was also seen in the hair bundles, cuticular plates, and cell bodies of hair cells with immature hair bundles. Thus, these antigens were useful markers for both normal and regenerating hair cells. Supporting cell immunolabeling was not seen in control cultures nor in the majority of supporting cells in GT cultures. In MBGT cultures, calbindin and parvalbumin immunolabeling was up-regulated in the cytosol of single supporting cells participating in scar formations and in supporting cells with hair cell-like characteristics. These data provide further evidence that non-mitotic hair cell regeneration in cultures can be accomplished by the conversion of supporting cells into hair cells.

  6. A retinoblastoma orthologue is a major regulator of S-phase, mitotic, and developmental gene expression in Dictyostelium.

    Directory of Open Access Journals (Sweden)

    Kimchi Strasser

    Full Text Available BACKGROUND: The retinoblastoma tumour suppressor, Rb, has two major functions. First, it represses genes whose products are required for S-phase entry and progression thus stabilizing cells in G1. Second, Rb interacts with factors that induce cell-cycle exit and terminal differentiation. Dictyostelium lacks a G1 phase in its cell cycle but it has a retinoblastoma orthologue, rblA. METHODOLOGY/PRINCIPAL FINDINGS: Using microarray analysis and mRNA-Seq transcriptional profiling, we show that RblA strongly represses genes whose products are involved in S phase and mitosis. Both S-phase and mitotic genes are upregulated at a single point in late G2 and again in mid-development, near the time when cell cycling is reactivated. RblA also activates a set of genes unique to slime moulds that function in terminal differentiation. CONCLUSIONS: Like its mammalian counterpart Dictyostelium, RblA plays a dual role, regulating cell-cycle progression and transcriptional events leading to terminal differentiation. In the absence of a G1 phase, however, RblA functions in late G2 controlling the expression of both S-phase and mitotic genes.

  7. Change of mitotic behavior and ultra structure of 'Fuju' (Citrus reticulata Blanco) stem-apex clones after space flight

    International Nuclear Information System (INIS)

    By using conventional squash stain technique and ultrathin sectioning technique, the effects of space flight on mitotic behavior and ultrastructure were studied in the shoot apical meristem of 'Fuju' (Citrus reticulata Blanco), which had been carried by China's seed-breeding satellite, Shijian-8. The results showed that space flight had effect on the mutagenesis of stem-apical meristem. Abnormal mitosis with various degrees had been detected in 13 mutant clones, of which mitotic aberrations in clone '08004' were significantly higher than the others. The aberration rate of numerical abnormalities of chromosomes at metaphase, lagging chromosome, micronucleus, C-spindle, S-spindle and polyarch spindle in the clone '08004' was 0.34%, 0.669%, 0.86%, 0.17%, 1.20% and 1.03%, respectively. The ultrastructure of mesophyll cell in most clones was unchanged, but nucleus chromatin agglutination, chloroplast thylakoid disintegrated, autophagosome appeared, cell vacuolated, plasmolysis and the formation of apoptotic body were found in the clone '08004', suggesting that programmed cell death (PCD) Nevertheless, no change in the mitochondrial structure was observed until terminal phase of PCD. (authors)

  8. Influence of 1-p-D-arabinofuranosylcytosine on mitotic activity of apical meristem of onion (Allium cepa L. roots

    Directory of Open Access Journals (Sweden)

    M. Kuraś

    2015-05-01

    Full Text Available The influence of increasing cytosine arabinoside (ara-C concentration (50, 100, 300 and 500 μg/ml on the mitotic activity of the apical meristem of onion adventitious roots was investigated during 24-h incubation in ara-C and postincubation in water. Incubation in ara-C inhibits reversibly mitosis, the degree of inhibition being dependent on the concentration used. 50 μg/ml ara-C causes only a slight and transitional mitotic depression, while 100—500 μg/ml reduces the per cent of mitoses in various degrees after 12-h incubation whereas after 24 h all concentrations (100—500 reduce mitosis to the same level of about 2 per cent (80% inhibition. During postincubation in water there occurs after treatment with 100—500 μg/ml ara-C, a wave of highly synchronized mitoses with a peak after 12 h. The highest frequency of synchronized mitoses (60°/o appeared in roots treated with 300 μg/ml ara-C. The main wave is preceded by a small one with a peak after 2-4 h of postincubation. This seems to be the consequence of phases S and G2 inhibition by ara-C. It was found that under the influence of ara-C the per cent of mitoses with chromosome aberration and of cells with micronuclei increased only slightly.

  9. Live-Cell Imaging Visualizes Frequent Mitotic Skipping During Senescence-Like Growth Arrest in Mammary Carcinoma Cells Exposed to Ionizing Radiation

    Energy Technology Data Exchange (ETDEWEB)

    Suzuki, Masatoshi, E-mail: msuzuki@nagasaki-u.ac.jp [Department of Radiation Medical Sciences, Atomic Bomb Disease Institute, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki (Japan); Yamauchi, Motohiro; Oka, Yasuyoshi; Suzuki, Keiji; Yamashita, Shunichi [Department of Radiation Medical Sciences, Atomic Bomb Disease Institute, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki (Japan)

    2012-06-01

    Purpose: Senescence-like growth arrest in human solid carcinomas is now recognized as the major outcome of radiotherapy. This study was designed to analyze cell cycle during the process of senescence-like growth arrest in mammary carcinoma cells exposed to X-rays. Methods and Materials: Fluorescent ubiquitination-based cell cycle indicators were introduced into the human mammary carcinoma cell line MCF-7. Cell cycle was sequentially monitored by live-cell imaging for up to 5 days after exposure to 10 Gy of X-rays. Results: Live-cell imaging revealed that cell cycle transition from G2 to G1 phase without mitosis, so-called mitotic skipping, was observed in 17.1% and 69.8% of G1- and G2-irradiated cells, respectively. Entry to G1 phase was confirmed by the nuclear accumulation of mKO{sub 2}-hCdt1 as well as cyclin E, which was inversely correlated to the accumulation of G2-specific markers such as mAG-hGeminin and CENP-F. More than 90% of cells skipping mitosis were persistently arrested in G1 phase and showed positive staining for the senescent biochemical marker, which is senescence-associated ss-galactosidase, indicating induction of senescence-like growth arrest accompanied by mitotic skipping. While G2 irradiation with higher doses of X-rays induced mitotic skipping in approximately 80% of cells, transduction of short hairpin RNA (shRNA) for p53 significantly suppressed mitotic skipping, suggesting that ionizing radiation-induced mitotic skipping is associated with p53 function. Conclusions: The present study found the pathway of senescence-like growth arrest in G1 phase without mitotic entry following G2-irradiation.

  10. Live-Cell Imaging Visualizes Frequent Mitotic Skipping During Senescence-Like Growth Arrest in Mammary Carcinoma Cells Exposed to Ionizing Radiation

    International Nuclear Information System (INIS)

    Purpose: Senescence-like growth arrest in human solid carcinomas is now recognized as the major outcome of radiotherapy. This study was designed to analyze cell cycle during the process of senescence-like growth arrest in mammary carcinoma cells exposed to X-rays. Methods and Materials: Fluorescent ubiquitination-based cell cycle indicators were introduced into the human mammary carcinoma cell line MCF-7. Cell cycle was sequentially monitored by live-cell imaging for up to 5 days after exposure to 10 Gy of X-rays. Results: Live-cell imaging revealed that cell cycle transition from G2 to G1 phase without mitosis, so-called mitotic skipping, was observed in 17.1% and 69.8% of G1- and G2-irradiated cells, respectively. Entry to G1 phase was confirmed by the nuclear accumulation of mKO2-hCdt1 as well as cyclin E, which was inversely correlated to the accumulation of G2-specific markers such as mAG-hGeminin and CENP-F. More than 90% of cells skipping mitosis were persistently arrested in G1 phase and showed positive staining for the senescent biochemical marker, which is senescence-associated ß-galactosidase, indicating induction of senescence-like growth arrest accompanied by mitotic skipping. While G2 irradiation with higher doses of X-rays induced mitotic skipping in approximately 80% of cells, transduction of short hairpin RNA (shRNA) for p53 significantly suppressed mitotic skipping, suggesting that ionizing radiation-induced mitotic skipping is associated with p53 function. Conclusions: The present study found the pathway of senescence-like growth arrest in G1 phase without mitotic entry following G2-irradiation.

  11. Inmunoterapia local Local immunotherapy

    Directory of Open Access Journals (Sweden)

    E. Lasa

    2003-01-01

    Full Text Available La inmunoterapia específica, junto con la evitación del alergeno y el tratamiento sintomático, forma parte del tratamiento de la patología alérgica. La modalidad más antigua, más conocida y mejor estudiada es la inmunoterapia subcutánea (ITSC, cuya eficacia tanto a corto como a largo plazo, ha sido ampliamente demostrada en numerosos estudios. Sin embargo, a pesar de haberse demostrado segura, no está exenta de efectos adversos y precisa ser administrada bajo supervisión de personal médico. Esto ha animado a buscar nuevas vías de administración de eficacia similar, con un buen perfil de seguridad, y de buena cumplimentación por parte del paciente. De las distintas alternativas estudiadas la más relevante es la inmunoterapia sublingual (ITSL. En ésta, se administra el antígeno en forma de gotas debajo de la lengua. Existen diferentes pautas de administración en función del alergeno implicado. La dosis óptima de tratamiento está aún sin determinar, hallándose en este momento en un rango amplio de dosis respecto a la inmunoterapia subcutánea. Su mecanismo de acción es poco conocido aunque en diversos estudios se han observado cambios inmunológicos. La ITSL ha mostrado un buen perfil de seguridad con escasos efectos secundarios, habitualmente de carácter local. Asimismo se han realizado distintos ensayos clínicos en los que se ha demostrado su eficacia en el tratamiento de la alergia respiratoria tanto en niños como en adultos. Por ello, aunque aún existen datos sin resolver respecto a esta vía de administración de inmunoterapia, ha sido propuesta por la OMS como una alternativa válida a la ITSC.Specific immunotherapy, together with avoidance of the allergen and symptomatic treatment, forms part of the treatment of allergic pathology. The oldest, best known and most studied form is subcutaneous immunotherapy (SCIT, whose efficacy, both in the short and the long term, has been widely demonstrated in numerous studies

  12. Documenting localities

    CERN Document Server

    Cox, Richard J

    1996-01-01

    Now in paperback! Documenting Localities is the first effort to summarize the past decade of renewed discussion about archival appraisal theory and methodology and to provide a practical guide for the documentation of localities.This book discusses the continuing importance of the locality in American historical research and archival practice, traditional methods archivists have used to document localities, and case studies in documenting localities. These chapters draw on a wide range of writings from archivists, historians, material culture specialists, historic preservationists

  13. Structure of a Blinkin-BUBR1 complex reveals an interaction crucial for kinetochore-mitotic checkpoint regulation via an unanticipated binding Site

    DEFF Research Database (Denmark)

    Bolanos-Garcia, Victor M; Lischetti, Tiziana; Matak-Vinković, Dijana;

    2011-01-01

    The maintenance of genomic stability relies on the spindle assembly checkpoint (SAC), which ensures accurate chromosome segregation by delaying the onset of anaphase until all chromosomes are properly bioriented and attached to the mitotic spindle. BUB1 and BUBR1 kinases are central for this proc...

  14. Incorporation of thymidine into onion root meristematic cell nuclei in presence of hydroxyurea and its role in recovery of mitotic activity

    International Nuclear Information System (INIS)

    Hydroxyurea treatment of onion roots induced mitotic block which was released by transfer of bulbs to water, and also to some extent by addition of cold or 3H-thymidine to hydroxyurea solutions. In presence of hydroxyurea there was noted very intense incorporation of 3H-thymidine into cell nuclei, giving labelling index of 40-70%. However, all the mitotic figures appearing in presence of hydroxyurea and 3H-thymidine were unlabelled. On the other hand, labelled mitotic figures were obtained when roots incubated with 3H-thymidine in presence of hydroxyurea had been transferred to water. Incorporation of 3H-uridine was unaffected by hydroxyurea. The results show that hydroxyurea arrests onion root meristematic cells, either in the S phase and the G2 phase. Enhanced incorporation of 3H-thymidine in the presence of hydroxyurea, and release by added thymidine of the mitotic block indicate that hydroxyurea induces in onion root meristematic cells a particular shortage of thymidylate. (author)

  15. CORRELATION BETWEEN THE NUMBER OF MITOTIC FIGURES AND THE PERCENTAGE OF Ki67-POSITIVE CELLS IN NON-HODGKIN’S LYMPHOMAS

    Institute of Scientific and Technical Information of China (English)

    黄高升; FellerAC; LennertK

    1994-01-01

    Proliferative activity in 106 cases of non-Hodgkin's lymphoma was estimated using the monoclonal antibody Ki67 and by countin the number of mitotic figures.The percentage of Ki67-positive cells was compared with the median mumber of mitotic figures per square millimeter.Both Ki67 positivity and the number of mitotic figures were found to be greater in high grade than in low grade lymphomas,although there was an overlap between the two grades of malignancy.A close correlation was found between the number of mitotic figured and the percentage of Ki67-positive cells not only in all lymphoma types taken together (rs=0.834,P<0.001),but also in B-cell lymphoma(rs=0.818,P<0.001)and T-cell lymphoma(rs=0.764,P<0.001)taken seprately.Thus,both methods are useful for the estimation of proliferative activity,but each method has is advantages and disadvan-tages.

  16. Significant decrease of ADP release rate underlies the potent activity of dimethylenastron to inhibit mitotic kinesin Eg5 and cancer cell proliferation

    Energy Technology Data Exchange (ETDEWEB)

    Sun, Linlin [Lung Cancer Institute, Tianjin Medical University General Hospital, 154 Anshan Road, Heping District, Tianjin 300052 (China); Sun, Xiaodong; Xie, Songbo; Yu, Haiyang [Department of Genetics and Cell Biology, College of Life Sciences, Nankai University, 94 Weijin Road, Tianjin 300071 (China); Zhong, Diansheng, E-mail: Zhongdsh@hotmail.com [Lung Cancer Institute, Tianjin Medical University General Hospital, 154 Anshan Road, Heping District, Tianjin 300052 (China); Department of Oncology, Tianjin Medical University General Hospital, 154 Anshan Road, Heping District, Tianjin 300052 (China)

    2014-05-09

    Highlights: • DIMEN displays higher anti-proliferative activity than enastron. • DIMEN induced mitotic arrest and apoptosis more significantly than enastron. • DIMEN blocked the conformational change of ADP-binding pocket more effectively. • DIMEN hindered ADP release more potently than enastron. - Abstract: Eg5 is a mitotic kinesin that plays a crucial role in the formation of bipolar mitotic spindles, by hydrolyzing ATP to push apart anti-parallel microtubules. Dimethylenastron is potent specific small molecule inhibitor of Eg5. The mechanism by which dimethylenastron inhibits Eg5 function remains unclear. By comparing with enastron, here we report that dimethylenastron prevents the growth of pancreatic and lung cancer cells more effectively, by halting mitotic progression and triggering apoptosis. We analyze their interactions with ADP-bound Eg5 crystal structure, and find that dimethylenastron binds Eg5 motor domain with higher affinity. In addition, dimethylenastron allosterically blocks the conformational change of the “sandwich”-like ADP-binding pocket more effectively. We subsequently use biochemical approach to reveal that dimethylenastron slows ADP release more significantly than enastron. These data thus provide biological, structural and mechanistic insights into the potent inhibitory activity of dimethylenastron.

  17. RNA interference-mediated silencing of mitotic kinesin KIF14 disrupts cell cycle progression and induces cytokinesis failure.

    Science.gov (United States)

    Carleton, Michael; Mao, Mao; Biery, Matthew; Warrener, Paul; Kim, Sammy; Buser, Carolyn; Marshall, C Gary; Fernandes, Christine; Annis, James; Linsley, Peter S

    2006-05-01

    KIF14 is a microtubule motor protein whose elevated expression is associated with poor-prognosis breast cancer. Here we demonstrate KIF14 accumulation in mitotic cells, where it associated with developing spindle poles and spindle microtubules. Cells at later stages of mitosis were characterized by the concentration of KIF14 at the midbody. Time-lapse microscopy revealed that strong RNA interference (RNAi)-mediated silencing of KIF14 induced cytokinesis failure, causing several rounds of endoreduplication and resulting in multinucleated cells. Additionally, less efficacious KIF14-specific short interfering RNAs (siRNAs) induced multiple phenotypes, all of which resulted in acute apoptosis. Our data demonstrate the ability of siRNA-mediated silencing to generate epiallelic hypomorphs associated with KIF14 depletion. Furthermore, the link we observed between siRNA efficacy and phenotypic outcome indicates that distinct stages during cell cycle progression are disrupted by the differential modulation of KIF14 expression. PMID:16648480

  18. Engraftment potential of human fetal hematopoietic cells in NOD/SCID mice is not restricted to mitotically quiescent cells.

    Science.gov (United States)

    Wilpshaar, Jannine; Bhatia, Mickie; Kanhai, Humphrey H H; Breese, Robert; Heilman, Doug K; Johnson, Cynthia S; Falkenburg, J H Frederik; Srour, Edward F

    2002-07-01

    During fetal development, there is a continued demand for large numbers of primitive and mature hematopoietic cells. This demand may require that all potential hematopoietic stem cells (HSCs) migrate effectively to emerging hematopoietic sites and subsequently contribute to blood cell production, regardless of their cell cycle status. We recently established that umbilical cord blood cells in the G(1) phase of the cell cycle have a repopulating potential similar to cells in G(0), suggesting that cycling prenatal and neonatal HSCs may have the same functional capabilities described for quiescent, but not cycling, cells from adult sources. To establish the relationship between cell cycle status and hematopoietic potential at early stages of human ontogeny, the in vivo engraftment potential of mitotically defined fetal liver (FL) and fetal bone marrow (FBM) cells were examined in NOD/SCID recipients. Following transplantation of the same numbers of G(0), G(1), or S/G(2)+M CD34(+) cells from FL, equivalent percentages of recipient mice were chimeric (55%, 60%, and 60%, respectively). FBM-derived CD34(+) cells in all phases of the cell cycle engrafted in conditioned recipients and sustained human hematopoiesis, albeit at lower levels than their FL-derived counterparts. Multilineage differentiation was evident in all transplanted mice independent of the source or cell cycle status of graft cells. In addition, levels of chimerism in mice transplanted with fetal blood-derived G(0) or G(1) CD34(+) lineage-depleted cells were similar. These results support the assertion that mitotically quiescent and cycling fetal hematopoietic cells contain marrow-repopulating stem cells capable of multilineage engraftment in NOD/SCID mouse recipients. PMID:12070016

  19. Sensitivity to sodium arsenite in human melanoma cells depends upon susceptibility to arsenite-induced mitotic arrest

    International Nuclear Information System (INIS)

    Arsenic induces clinical remission in patients with acute promyelocytic leukemia and has potential for treatment of other cancers. The current study examines factors influencing sensitivity to arsenic using human malignant melanoma cell lines. A375 and SK-Mel-2 cells were sensitive to clinically achievable concentrations of arsenite, whereas SK-Mel-3 and SK-Mel-28 cells required supratherapeutic levels for toxicity. Inhibition of glutathione synthesis, glutathione S-transferase (GST) activity, and multidrug resistance protein (MRP) transporter function attenuated arsenite resistance, consistent with studies suggesting that arsenite is extruded from the cell as a glutathione conjugate by MRP-1. However, MRP-1 was not overexpressed in resistant lines and GST-π was only slightly elevated. ICP-MS analysis indicated that arsenite-resistant SK-Mel-28 cells did not accumulate less arsenic than arsenite-sensitive A375 cells, suggesting that resistance was not attributable to reduced arsenic accumulation but rather to intrinsic properties of resistant cell lines. The mode of arsenite-induced cell death was apoptosis. Arsenite-induced apoptosis is associated with cell cycle alterations. Cell cycle analysis revealed arsenite-sensitive cells arrested in mitosis whereas arsenite-resistant cells did not, suggesting that induction of mitotic arrest occurs at lower intracellular arsenic concentrations. Higher intracellular arsenic levels induced cell cycle arrest in the S-phase and G2-phase in SK-Mel-3 and SK-Mel-28 cells, respectively. The lack of arsenite-induced mitotic arrest in resistant cell lines was associated with a weakened spindle checkpoint resulting from reduced expression of spindle checkpoint protein BUBR1. These data suggest that arsenite has potential for treatment of solid tumors but a functional spindle checkpoint is a prerequisite for a positive response to its clinical application

  20. Chemogenetic evaluation of the mitotic kinesin CENP-E reveals a critical role in triple-negative breast cancer.

    Science.gov (United States)

    Kung, Pei-Pei; Martinez, Ricardo; Zhu, Zhou; Zager, Michael; Blasina, Alessandra; Rymer, Isha; Hallin, Jill; Xu, Meirong; Carroll, Christopher; Chionis, John; Wells, Peter; Kozminski, Kirk; Fan, Jeffery; Guicherit, Oivin; Huang, Buwen; Cui, Mei; Liu, Chaoting; Huang, Zhongdong; Sistla, Anand; Yang, Jennifer; Murray, Brion W

    2014-08-01

    Breast cancer patients with tumors lacking the three diagnostic markers (ER, PR, and HER2) are classified as triple-negative (primarily basal-like) and have poor prognosis because there is no disease-specific therapy available. To address this unmet medical need, gene expression analyses using more than a thousand breast cancer samples were conducted, which identified elevated centromere protein E (CENP-E) expression in the basal-a molecular subtype relative to other subtypes. CENP-E, a mitotic kinesin component of the spindle assembly checkpoint, is shown to be induced in basal-a tumor cell lines by the mitotic spindle inhibitor drug docetaxel. CENP-E knockdown by inducible shRNA reduces basal-a breast cancer cell viability. A potent, selective CENP-E inhibitor (PF-2771) was used to define the contribution of CENP-E motor function to basal-like breast cancer. Mechanistic evaluation of PF-2771 in basal-a tumor cells links CENP-E-dependent molecular events (e.g., phosphorylation of histone H3 Ser-10; phospho-HH3-Ser10) to functional outcomes (e.g., chromosomal congression defects). Across a diverse panel of breast cell lines, CENP-E inhibition by PF-2771 selectively inhibits proliferation of basal breast cancer cell lines relative to premalignant ones and its response correlates with the degree of chromosomal instability. Pharmacokinetic-pharmacodynamic efficacy analysis in a basal-a xenograft tumor model shows that PF-2771 exposure is well correlated with increased phospho-HH3-Ser10 levels and tumor growth regression. Complete tumor regression is observed in a patient-derived, basal-a breast cancer xenograft tumor model treated with PF-2771. Tumor regression is also observed with PF-2771 in a taxane-resistant basal-a model. Taken together, CENP-E may be an effective therapeutic target for patients with triple-negative/basal-a breast cancer. PMID:24928852

  1. PUL21a-Cyclin A2 interaction is required to protect human cytomegalovirus-infected cells from the deleterious consequences of mitotic entry.

    Directory of Open Access Journals (Sweden)

    Martin Eifler

    2014-10-01

    Full Text Available Entry into mitosis is accompanied by dramatic changes in cellular architecture, metabolism and gene expression. Many viruses have evolved cell cycle arrest strategies to prevent mitotic entry, presumably to ensure sustained, uninterrupted viral replication. Here we show for human cytomegalovirus (HCMV what happens if the viral cell cycle arrest mechanism is disabled and cells engaged in viral replication enter into unscheduled mitosis. We made use of an HCMV mutant that, due to a defective Cyclin A2 binding motif in its UL21a gene product (pUL21a, has lost its ability to down-regulate Cyclin A2 and, therefore, to arrest cells at the G1/S transition. Cyclin A2 up-regulation in infected cells not only triggered the onset of cellular DNA synthesis, but also promoted the accumulation and nuclear translocation of Cyclin B1-CDK1, premature chromatin condensation and mitotic entry. The infected cells were able to enter metaphase as shown by nuclear lamina disassembly and, often irregular, metaphase spindle formation. However, anaphase onset was blocked by the still intact anaphase promoting complex/cyclosome (APC/C inhibitory function of pUL21a. Remarkably, the essential viral IE2, but not the related chromosome-associated IE1 protein, disappeared upon mitotic entry, suggesting an inherent instability of IE2 under mitotic conditions. Viral DNA synthesis was impaired in mitosis, as demonstrated by the abnormal morphology and strongly reduced BrdU incorporation rates of viral replication compartments. The prolonged metaphase arrest in infected cells coincided with precocious sister chromatid separation and progressive fragmentation of the chromosomal material. We conclude that the Cyclin A2-binding function of pUL21a contributes to the maintenance of a cell cycle state conducive for the completion of the HCMV replication cycle. Unscheduled mitotic entry during the course of the HCMV replication has fatal consequences, leading to abortive infection and

  2. Increased radiation tolerance of mouse tongue epithelium after local conditioning

    International Nuclear Information System (INIS)

    The effect of local stimulation on mitotic activity and radiation tolerance was studied in mouse tongue mucosa. Silver nitrate solution (0.5-20%) was used for local conditioning. Silver nitrate conditioning did not alter the radiosensitivity of the epithelium to single local doses, but shortened the latency to denudation from 11 to 8 days. In contrast, a clear increase in tolerance to fractionated irradiation, delivering 5x2.5, 5x3.5, 5x4.5 Gy or 3x5.2 Gy in 7 days, was observed, equivalent to about four, two, one and two extra dose fractions. This approach may be a suitable way to increase radiation tolerance of oral mucosa in clinical radiotherapy. (author)

  3. Local Budgeting

    OpenAIRE

    Shah, Anwar

    2007-01-01

    This publication, Local Budgeting, provides a comprehensive guide for local administrators who are involved in designing and implementing budgetary institutions and who wish to improve efficiency and equity in service delivery and to strengthen internal and external accountability. It details principles and practices to improve fiscal management. It reviews techniques available in developi...

  4. Cellular and molecular effects of 1GeV/n iron ion exposure on post-mitotic human neurons

    Science.gov (United States)

    Guida, Peter; Vazquez, Marcelo E.; Guida, Peter; Kim, Angela

    During space travel, astronauts will be exposed to high energy, high atomic number (HZE) radiation. The potential for damage to cells of the central nervous system following exposure to HZE particle radiation has been characterized as a potential critical risk. Unfortunately, there are very few working model systems of human neurons and as a result, data describing the effects of HZE radiation on them is scarce. To begin risk assessment studies, we utilized an in vitro model consisting of terminally differentiated, post-mitotic human neurons (hNT cells). Previous studies have shown that transplantation of these cells into numerous rodent models of neurological diseases has resulted in successful mitigation of the related disorders, thereby demonstrating their functional relevance. Following exposure of these cells to 1GeV/n Fe ions at the NASA Space Radiation Laboratory, we measured the induction and repair of DNA damage (as revealed by g-H2AX foci), cytotoxicity, gene expression changes, and the induction of apoptosis and its pharmacological reduction. Fluorescence microscopy techniques revealed that there was a dose-dependent induction of g- H2AX foci in hNT cells, with a peak effect 4 hours after exposure (which is significantly longer than for reports using mitotic cells). DNA repair was evident in that the levels of g-H2AX foci were reduced to those in unirradiated cells by 24 hours post-irradiation. Cytotoxicity was also induced in a dose-dependent manner as detected by the fluorescent-based Live/Dead assay. Analysis of the status of the apoptosis-inducing gene p53 showed that the levels of this protein increased significantly 4-8 hours after exposure to Fe ions. By 3 days post-irradiation, annexin V staining demonstrated a dose-dependent induction of apoptosis in the hNT cells. Pre-treatment with two different concentrations of the growth factor TGF-b were effective in reducing the levels of Fe ion-induced apoptosis to statistically significant degrees.

  5. Override of the radiation-induced mitotic block in human tumour cells by methylxanthines and its relationship to the potentiation of cytotoxicity

    Energy Technology Data Exchange (ETDEWEB)

    Musk, S.R.R.; Steel, G.G. (Institute of Cancer Research, Sutton (UK). Surrey Branch)

    1990-06-01

    Caffeine, theophylline, theobromine and paraxanthine, were tested for ability to override mitotic block induced by ionizing radiation in the human bladder carcinoma cell line RT112. All were found to partially override the block, at a concentration of 1mM in the order caffeine > theophylline > theobromine = paraxanthine. At a concentration of 1 mM only caffeine was found to potentiate cell killing as well as causing block override; at higher concentrations all had a significant effect on survival but little or no further influence on the degree of block override. It is concluded that override of a mitotic block is not in itself sufficient to cause increased killing when irradiated cells are incubated in the presence of caffeine, and that caffeine exerts its potentiating effect by directly inhibiting repair of damage in DNA or by causing override of radiation-induced inhibition of DNA synthesis. (author).

  6. Override of the radiation-induced mitotic block in human tumour cells by methylxanthines and its relationship to the potentiation of cytotoxicity

    International Nuclear Information System (INIS)

    Caffeine, theophylline, theobromine and paraxanthine, were tested for ability to override mitotic block induced by ionizing radiation in the human bladder carcinoma cell line RT112. All were found to partially override the block, at a concentration of 1mM in the order caffeine > theophylline > theobromine = paraxanthine. At a concentration of 1 mM only caffeine was found to potentiate cell killing as well as causing block override; at higher concentrations all had a significant effect on survival but little or no further influence on the degree of block override. It is concluded that override of a mitotic block is not in itself sufficient to cause increased killing when irradiated cells are incubated in the presence of caffeine, and that caffeine exerts its potentiating effect by directly inhibiting repair of damage in DNA or by causing override of radiation-induced inhibition of DNA synthesis. (author)

  7. Influence of the radiation (Co60) in pre-implants rabbit embryos: effect on atypic mitotic index and embryo pole development

    International Nuclear Information System (INIS)

    We studied the effect of ionizing irradiation on 12 New Zealand rabbits (65 embryos), at three different times: at match time (zero hour), two days after and four days after, with two different irradiation doses: five c Gy and ten c Gy. Six rabbits (36 blastocysts) were used as controls. the matching instant was the zero hour. Exactly six days after (± 60 minutes) the embryos of each rabbit was picked up by flushing the uterus with culture media. the embryos were fixed in methanol for 48 hours, and colored with acid Mayer hematoxylin. The following embryo parameters were studied: embryo pole development; percentage of abnormal mitotic figures. irradiation time was associated with lower scores of embryo pole development, but not with irradiation dose. There were no gross abnormalities of embryo pole. The abnormal mitotic cells was affected both by the time and dose of irradiation. (author)

  8. Mitotic silencing of human rRNA synthesis: inactivation of the promoter selectivity factor SL1 by cdc2/cyclin B-mediated phosphorylation.

    OpenAIRE

    Heix, J; Vente, A.; Voit, R; Budde, A; Michaelidis, T M; Grummt, I

    1998-01-01

    We have used a reconstituted cell-free transcription system to investigate the molecular basis of mitotic repression of RNA polymerase I (pol I) transcription. We demonstrate that SL1, the TBP-containing promoter-binding factor, is inactivated by cdc2/cyclin B-directed phosphorylation, and reactivated by dephosphorylation. Transcriptional inactivation in vitro is accompanied by phosphorylation of two subunits, e.g. TBP and hTAFI110. To distinguish whether transcriptional repression is due to ...

  9. Paclitaxel-induced microtubule stabilization causes mitotic block and apoptotic-like cell death in a paclitaxel-sensitive strain of Saccharomyces cerevisiae

    OpenAIRE

    Foland, Travis B.; Dentler, William L.; SUPRENANT, KATHY A.; Gupta, Mohan L.; Himes, Richard H.

    2005-01-01

    Wild-type Saccharomyces cerevisiae tubulin does not bind the anti-mitotic microtubule stabilizing agent paclitaxel. Previously, we introduced mutations into the S. cerevisiae gene for β-tubulin that imparted paclitaxel binding to the protein, but the mutant strain was not sensitive to paclitaxel and other microtubule-stabilizing agents, due to the multiple ABC transporters in the membranes of budding yeast. Here, we introduced the mutated β-tubulin gene into a S. cerevisiae strain with dimini...

  10. Identification of a BET Family Bromodomain/Casein Kinase II/TAF-Containing Complex as a Regulator of Mitotic Condensin Function

    OpenAIRE

    Hyun-Soo Kim; Rituparna Mukhopadhyay; Scott B. Rothbart; Andrea C. Silva; Vincent Vanoosthuyse; Ernest Radovani; Thomas Kislinger; Assen Roguev; Colm J. Ryan; Jiewei Xu; Harlizawati Jahari; Kevin G. Hardwick; Jack F. Greenblatt; Nevan J. Krogan; Jeffrey S. Fillingham

    2014-01-01

    Condensin is a central regulator of mitotic genome structure with mutants showing poorly condensed chromosomes and profound segregation defects. Here, we identify NCT, a complex comprising the Nrc1 BET-family tandem bromodomain protein (SPAC631.02), casein kinase II (CKII), and several TAFs, as a regulator of condensin function. We show that NCT and condensin bind similar genomic regions but only briefly colocalize during the periods of chromosome condensation and decondensation. This pattern...

  11. 3D-QSAR Studies of Dihydropyrazole and Dihydropyrrole Derivatives as Inhibitors of Human Mitotic Kinesin Eg5 Based on Molecular Docking

    OpenAIRE

    Wenjuan Yang; Zhihua Lin; Yuanqiang Wang; Jin Liu; Mao Shu; Xingyan Luo

    2012-01-01

    Human mitotic kinesin Eg5 plays an essential role in mitoses and is an interesting drug target against cancer. To find the correlation between Eg5 and its inhibitors, structure-based 3D-quantitative structure–activity relationship (QSAR) studies were performed on a series of dihydropyrazole and dihydropyrrole derivatives using comparative molecular field analysis (CoMFA) and comparative molecular similarity indices analysis (CoMSIA) methods. Based on the LigandFit docking results, predictive ...

  12. Carotene and cytochrome c dependent mitotic recombination and killing in response to visible light and UV-C in the smut fungus Ustilago violacea

    International Nuclear Information System (INIS)

    The carotene (carotenoid hydrocarbons) and cytochrome c compositions for the ph/ph+ pink diploid strain of Ustilago violacea and three newly developed color variants, white (w), orange (o), and yellow (y) were quantitatively determined using high performance liquid chromatography of lipid extracts and difference spectroscopy of alkali extracts. In addition, the effect of high intensity incandescent and far UV (UV-C) radiation on survival and mitotic recombination in all four ph/ph+ strains were studied. (author)

  13. Localized Scleroderma

    Science.gov (United States)

    ... educational offerings from your local chapter. • Discounted registration fees to the annual National Patient Education Conference. Please consider joining the Scleroderma Foundation today. A membership form is attached on the reverse side of ...

  14. Net Locality

    DEFF Research Database (Denmark)

    de Souza e Silva, Adriana Araujo; Gordon, Eric

    Provides an introduction to the new theory of Net Locality and the profound effect on individuals and societies when everything is located or locatable. Describes net locality as an emerging form of location awareness central to all aspects of digital media, from mobile phones, to Google Maps, to...... of emerging technologies, from GeoCities to GPS, Wi-Fi, Wiki Me, and Google Android....

  15. Ontology Localization

    OpenAIRE

    Espinoza Mejía, Mauricio; Montiel-Ponsoda, Elena; Aguado de Cea, G.; A. GÓMEZ-PÉREZ

    2012-01-01

    In the context of the Semantic Web, resources on the net can be enriched by well-defined, machine-understandable metadata describing their associated conceptual meaning. These metadata consisting of natural language descriptions of concepts are the focus of the activity we describe in this chapter, namely, ontology localization. In the framework of the NeOn Methodology, ontology localization is defined as the activity of adapting an ontology to a particular language and culture. This adaptati...

  16. hSNF5/INI1 inactivation is mainly associated with homozygous deletions and mitotic recombinations in rhabdoid tumors.

    Science.gov (United States)

    Rousseau-Merck, M F; Versteege, I; Legrand, I; Couturier, J; Mairal, A; Delattre, O; Aurias, A

    1999-07-01

    The chromatin-remodeling hSNF5/INI1 gene has recently been shown to act as a tumor suppressor gene in rhabdoid tumors (RTs). In an attempt to further characterize the main chromosomal mechanisms involved in hSNF5/INI1 inactivation in RTs, we report here the molecular cytogenetic data obtained in 12 cell lines harboring hSNF5/INI1 mutations and/or deletions in relation to the molecular genetic analysis using polymorphic markers extended to both extremities of chromosome 22q. On the whole, mitotic recombination occurring in the proximal part of chromosome 22q, as demonstrated in five cases, and nondisjunction/duplication, highly suspected in two cases (processes leading respectively to partial or complete isodisomy), appear to be major mechanisms associated with hSNF5/INI1 inactivation. Such isodisomy accompanies each of the RTs exhibiting two cytogenetically normal chromosomes 22. This results in homozygosity for the mutation at the hSNF5/INI1 locus. An alternate mechanism accounting for hSNF5/INI1 inactivation observed in these tumors is homozygous deletion in the rhabdoid consensus region. This was observed in each of the four tumors carrying a chromosome 22q abnormality and, in particular, in the three tumors with chromosomal translocations. Only one case of our series illustrates the mutation/deletion classical model proposed for the double-hit inactivation of a tumor suppressor gene. PMID:10397258

  17. Abrupt onset of mutations in a developmentally regulated gene during terminal differentiation of post-mitotic photoreceptor neurons in mice.

    Directory of Open Access Journals (Sweden)

    Ivette M Sandoval

    Full Text Available For sensitive detection of rare gene repair events in terminally differentiated photoreceptors, we generated a knockin mouse model by replacing one mouse rhodopsin allele with a form of the human rhodopsin gene that causes a severe, early-onset form of retinitis pigmentosa. The human gene contains a premature stop codon at position 344 (Q344X, cDNA encoding the enhanced green fluorescent protein (EGFP at its 3' end, and a modified 5' untranslated region to reduce translation rate so that the mutant protein does not induce retinal degeneration. Mutations that eliminate the stop codon express a human rhodopsin-EGFP fusion protein (hRho-GFP, which can be readily detected by fluorescence microscopy. Spontaneous mutations were observed at a frequency of about one per retina; in every case, they gave rise to single fluorescent rod cells, indicating that each mutation occurred during or after the last mitotic division. Additionally, the number of fluorescent rods did not increase with age, suggesting that the rhodopsin gene in mature rod cells is less sensitive to mutation than it is in developing rods. Thus, there is a brief developmental window, coinciding with the transcriptional activation of the rhodopsin locus, in which somatic mutations of the rhodopsin gene abruptly begin to appear.

  18. Dose-response of x-ray-induced anaphase aberrations in the mitotic root tip chromosomes of allium

    International Nuclear Information System (INIS)

    A simplified Allium root mitotic chromosome aberration assay by using only the aberrant anaphases (fragments, laggards and bridges) as the end-points were developed by Rank and Nielsen (1993) for screening water soluble chemicals and complex mixtures. A dose-response curve was established by Meir et al., (1994) using a known clastogen, 4-nitroquinolene-N-oxide between the dose range of 0.1-0.5 ug/ml. In order to further validate this assay for clastogen detection, a series of X-ray dose response experiments was carried out. Allium roots were germinated in tapwater for 48 h and treated with a series of 10, 20, 30, 40, 50, 60 R (80 Kvp, 5 ma, dose rate 60 R/min) dosages. After an 18 hr recovery time, the root tips were hydrolyzed in 45% acetic and 1 N HC1 acid (9:1 ratio) solution under 50 degrees C for 5 min and stained with aceto-carmine. Each of the data points were derived from scoring 7-10 slides (15-50 anaphases/slide). The corrrelation coefficient, slope and intercept values of the dose-response curve are: 0.954, 0.515 and 1.155 respectively

  19. Stimulatory effects of gamma irradiation on phytochemical properties, mitotic behaviour, and nutritional composition of sainfoin (Onobrychis viciifolia Scop.).

    Science.gov (United States)

    Mohajer, Sadegh; Taha, Rosna Mat; Lay, Ma Ma; Esmaeili, Arash Khorasani; Khalili, Mahsa

    2014-01-01

    Sainfoin (Onobrychis viciifolia Scop. Syn. Onobrychis sativa L.) is a bloat-safe forage crop with high levels of tannins, which is renowned for its medicinal qualities in grazing animals. Mutagenesis technique was applied to investigate the influence of gamma irradiation at 30, 60, 90, and 120 Gy on mitotic behavior, in vitro growth factors, phytochemical and nutritional constituents of sainfoin. Although a percentage of plant necrosis and non-growing seed were enhanced by irradiation increment, the germination speed was significantly decreased. It was observed that gamma irradiated seeds had higher value of crude protein and dry matter digestibility compared to control seeds. Toxicity of copper was reduced in sainfoin irradiated seeds at different doses of gamma rays. Anthocyanin content also decreased in inverse proportion to irradiation intensity. Accumulation of phenolic and flavonoid compounds was enhanced by gamma irradiation exposure in leaf cells. HPLC profiles differed in peak areas of the two important alkaloids, Berberine and Sanguinarine, in 120 Gy irradiated seeds compared to control seeds. There were positive correlations between irradiation dose and some abnormality divisions such as laggard chromosome, micronucleus, binucleated cells, chromosome bridge, and cytomixis. In reality, radiocytological evaluation was proven to be essential in deducing the effectiveness of gamma irradiation to induce somaclonal variation in sainfoin. PMID:25147870

  20. Integration of biological data by kernels on graph nodes allows prediction of new genes involved in mitotic chromosome condensation.

    Science.gov (United States)

    Hériché, Jean-Karim; Lees, Jon G; Morilla, Ian; Walter, Thomas; Petrova, Boryana; Roberti, M Julia; Hossain, M Julius; Adler, Priit; Fernández, José M; Krallinger, Martin; Haering, Christian H; Vilo, Jaak; Valencia, Alfonso; Ranea, Juan A; Orengo, Christine; Ellenberg, Jan

    2014-08-15

    The advent of genome-wide RNA interference (RNAi)-based screens puts us in the position to identify genes for all functions human cells carry out. However, for many functions, assay complexity and cost make genome-scale knockdown experiments impossible. Methods to predict genes required for cell functions are therefore needed to focus RNAi screens from the whole genome on the most likely candidates. Although different bioinformatics tools for gene function prediction exist, they lack experimental validation and are therefore rarely used by experimentalists. To address this, we developed an effective computational gene selection strategy that represents public data about genes as graphs and then analyzes these graphs using kernels on graph nodes to predict functional relationships. To demonstrate its performance, we predicted human genes required for a poorly understood cellular function-mitotic chromosome condensation-and experimentally validated the top 100 candidates with a focused RNAi screen by automated microscopy. Quantitative analysis of the images demonstrated that the candidates were indeed strongly enriched in condensation genes, including the discovery of several new factors. By combining bioinformatics prediction with experimental validation, our study shows that kernels on graph nodes are powerful tools to integrate public biological data and predict genes involved in cellular functions of interest. PMID:24943848

  1. Visualization and Characterization of High-Order Chromatin Fibers under Light Microscope during Interphase and Mitotic Stages in Plants

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    Using genomic in situ hybridization with genomic DNA.high-order chromatin fibers were successfully exhibited under a light microscope through the cell cycle in barley,rice,maize and field bean.From the interphase to prophase and metaphase of mitosis,the fibers were basically similar.Each was estimated to be around 200 nm in diameter,but the strength of signals was not the same along the fiber length.Through the cell cycle a series of dynamic distribution changes occurred in the fibers.In the interphase,they were unraveled.At the early prophase they were arranged with parallel and mirror symmetry.During late-prophase and metaphase,the fibers were bundled and became different visible chromosomes.The parallel coiling and mirror symmetry structures were visible clearly until the metaphase.In anaphase they disappeared.During telophase,in peripheral regions of congregated chromosome group,borderlines of the chromosomes disappeared and the fibers were unraveled.This demonstrated that mitotic chromosomes are assembled and organized by parallel and adjacent coiling of the fibers and the fibers should be the highest order structure for DNA coiling.

  2. Generation of large homozygous chromosomal segments by mitotic recombination during lymphomagenesis in F{sub 1} hybrid mice

    Energy Technology Data Exchange (ETDEWEB)

    Hong, Doo-Pyo; Mori, Nobuko; Umesako, Seiichi; Okumoto, Masaaki [Osaka Prefectural Univ., Sakai (Japan). Research Inst. for Advanced Science and Technology; Kubo, Kihei; Tsugawa, Naomi [Osaka Prefectural Univ., Sakai (Japan). Graduate School of Agriculture and Biological Sciences; Song, Chang-Woo [Korea Research Inst. of Chemical Technology, Taejon (Korea, Republic of)

    2002-06-01

    The loss of heterozygosity (LOH) has been reported in numerous neoplasms in both human and animals, and has often been observed in chromosomal regions, which contain tumor-suppressor genes. We previously found frequent LOH on chromosomes 4, 12 and 19 in radiation-induced lymphomas from (BALB/cHeA x STS/A)F{sub 1} hybrid mice by allelotype analysis at polymorphic microsatellite loci. In this study, to elucidate the nature of allelic losses, we refined the loss regions on chromosomes 4, 12 and 19 of the tumors from the F{sub 1} mice and then analyzed them cytogenetically. The results represent evidence of a wide range of allelic losses owing to mitotic recombination on chromosomes 4 and 19 in the tumors, possibly reflecting functional losses of putative tumor-suppressor genes. It is suggested that the generation of these large homozygous chromosomal segments probably containing the affected genes is one of the genetic alterations responsible for tumorigenesis. (author)

  3. In silico analysis of deleterious single nucleotide polymorphisms in human BUB1 mitotic checkpoint serine/threonine kinase B gene.

    Science.gov (United States)

    Akhoundi, Fatemeh; Parvaneh, Nikpour; Modjtaba, Emadi-Baygi

    2016-09-01

    One of the major challenges in the analysis of human genetic variation is to distinguish mutations that are functionally neutral from those that contribute to disease. BubR1 is a key protein mediating spindle-checkpoint activation that plays a role in the inhibition of the anaphase-promoting complex/cyclosome (APC/C), delaying the onset of anaphase and ensuring proper chromosome segregation. Owing to the importance of BUB1B gene in mitotic checkpoint a functional analysis using different in silico approaches was undertaken to explore the possible associations between genetic mutations and phenotypic variation. In this work we found that 3 nsSNPs I82N, P334L and R814H have a functional effect on protein function and stability. A literature search revealed that R814H was already implicated in human diseases. Additionally, 2 SNPs in the 5' UTR region was predicted to exhibit a pattern change in the internal ribosome entry site (IRES), and eight MicroRNA binding sites were found to be highly affected due to 3' UTR SNPs. These in silico predictions will provide useful information in selecting the target SNPs that are likely to have functional impact on the BUB1B gene. PMID:27331020

  4. Cryo-EM of Mitotic Checkpoint Complex-Bound APC/C Reveals Reciprocal and Conformational Regulation of Ubiquitin Ligation.

    Science.gov (United States)

    Yamaguchi, Masaya; VanderLinden, Ryan; Weissmann, Florian; Qiao, Renping; Dube, Prakash; Brown, Nicholas G; Haselbach, David; Zhang, Wei; Sidhu, Sachdev S; Peters, Jan-Michael; Stark, Holger; Schulman, Brenda A

    2016-08-18

    The mitotic checkpoint complex (MCC) coordinates proper chromosome biorientation on the spindle with ubiquitination activities of CDC20-activated anaphase-promoting complex/cyclosome (APC/C(CDC20)). APC/C(CDC20) and two E2s, UBE2C and UBE2S, catalyze ubiquitination through distinct architectures for linking ubiquitin (UB) to substrates and elongating polyUB chains, respectively. MCC, which contains a second molecule of CDC20, blocks APC/C(CDC20)-UBE2C-dependent ubiquitination of Securin and Cyclins, while differentially determining or inhibiting CDC20 ubiquitination to regulate spindle surveillance, checkpoint activation, and checkpoint termination. Here electron microscopy reveals conformational variation of APC/C(CDC20)-MCC underlying this multifaceted regulation. MCC binds APC/C-bound CDC20 to inhibit substrate access. However, rotation about the CDC20-MCC assembly and conformational variability of APC/C modulate UBE2C-catalyzed ubiquitination of MCC's CDC20 molecule. Access of UBE2C is limiting for subsequent polyubiquitination by UBE2S. We propose that conformational dynamics of APC/C(CDC20)-MCC modulate E2 activation and determine distinctive ubiquitination activities as part of a response mechanism ensuring accurate sister chromatid segregation. PMID:27522463

  5. Involvement of kinesin family member 2C/mitotic centromere-associated kinesin overexpression in mammary carcinogenesis.

    Science.gov (United States)

    Shimo, Arata; Tanikawa, Chizu; Nishidate, Toshihiko; Lin, Meng-Lay; Matsuda, Koichi; Park, Jae-Hyun; Ueki, Tomomi; Ohta, Tomohiko; Hirata, Koichi; Fukuda, Mamoru; Nakamura, Yusuke; Katagiri, Toyomasa

    2008-01-01

    To elucidate the molecular mechanisms of mammary carcinogenesis and discover novel therapeutic targets for breast cancer, we previously carried out genome-wide expression profile analysis of 81 breast cancer cases by means of cDNA microarray coupled with laser microbeam microdissection of cancer cells. Among the dozens of transactivated genes, in the present study we focused on the functional significance of kinesin family member 2C (KIF2C)/mitotic centromere-associated kinesin (MCAK) in the growth of breast cancer cells. Northern blot and immunohistochemical analyses confirmed KIF2C/MCAK overexpression in breast cancer cells, and showed that it is expressed at undetectable levels in normal human tissues except the testis, suggesting KIF2C/MCAK to be a cancer-testis antigen. Western blot analysis using breast cancer cell lines revealed a significant increase in the endogenous KIF2C/MCAK protein level and its phosphorylation in G(2)/M phase. Treatment of breast cancer cells with small interfering RNA against KIF2C/MCAK effectively suppressed KIF2C/MCAK expression and inhibited the growth of the breast cancer cell lines T47D and HBC5. In addition, we found that KIF2C/MCAK expression was significantly suppressed by ectopic introduction of p53. These findings suggest that overexpression of KIF2C/MCAK might be involved in breast carcinogenesis and is a promising therapeutic target for breast cancers. PMID:17944972

  6. Mitotically active cellular fibroma of ovary: a clinicopathologic analysis%卵巢核分裂象活跃的富于细胞性纤维瘤的临床病理学观察

    Institute of Scientific and Technical Information of China (English)

    毕蕊; 赵燕; 杨文涛

    2013-01-01

    Objective To study the clinicopathologic characteristics of mitotically active cellular fibroma (MACF) of the ovary.Methods The clinicopathologic features of 11 cases of MACF from 2008 to 2012 years were retrospectively reviewed and immunohistochemic EnVision method was performed.Results The age of patients ranged from 21 to 65 years (mean =46 years).The maximum tumor diameter varied from 6 to 16 cm (mean =9.8 cm).The tumors were densely cellular and composed of intersecting bundles of spindle cells,sometimes associated with storiform pattern.The mitotic count ranged from 4 to 20 per 10 high-power field (mean =8.4).Atypical mitotic figures were not identified and coagulative tumor necrosis was not found.There was at most mild to moderate degree of nuclear atypia.Immunohistochemically,8 cases showed positive stain for vimentin and 5 cases expressed diffusely or focally WT-1.In 6 cases,Ki-67 positive index ranged from 5%-30%,including 1 case (10%),and 1 case (30%) in which mitotic figures were 20/10 HPF.CK,AE1/AE3,CD117,CD34,CD99 and desmin were negative in the detected 5 cases.Reticular fiber staining in 2 cases showed dense reticular fibers enveloping individual tumor cells.Ten patients who followed-up range from 4 to 38 months were alive without evidence of recurrence or disease progression.One patient was alive with local recurrence 94 months after surgery.Conclusions Cellular fibromatous neoplasm with mitotic count of≥4/10 HPF but relatively bland nuclear features should be considered as MACF rather than ovarian fibrosarcoma.MACF is a group of ovarian tumor which carries a low malignant potential and occasional chance of recurrence.%目的 探讨卵巢核分裂象活跃的富于细胞性纤维瘤的临床病理学特征及鉴别诊断.方法 收集2008至2012年间11例卵巢核分裂象活跃的富于细胞性纤维瘤患者的临床病理资料,光镜观察并进行免疫组织化学EnVision法染色.结果 11例患者发病年龄21~ 65岁,平均46

  7. Redistribution of the nuclear mitotic apparatus protein (NuMA) during mitosis and nuclear assembly. Properties of purified NuMA protein.

    Science.gov (United States)

    Price, C M; Pettijohn, D E

    1986-10-01

    Monoclonal antibodies and human autoimmune sera specific for the nuclear mitotic apparatus protein (NuMA protein) were applied to study the structure of this protein and its intracellular distribution. The NuMA protein was purified using immuno-affinity columns. Studies on this large (250 kD) nuclear protein indicated that it is a highly asymmetric phosphoprotein. It is present in all mammalian cells examined and in those of some non-mammals. Immunofluorescence studies on fixed cells demonstrated that its intracellular distribution is essentially the same in all species at all stages of the cell cycle. Immunoblot (western blot) analysis showed that the size of the NuMA protein varies slightly in different species. At the onset of mitosis the NuMA protein redistributes from the nucleus to two centrosomal structures that later will become part of the mitotic spindle pole. This occurs at the time of nuclear breakdown and eventually leads to an accumulation of the NuMA protein at the polar region of the mitotic spindle. After anaphase the protein redistributes from the spindle polar region into the reforming nucleus and concentrates initially at the site where nuclear lamins and perichomatin have been reported to assemble. Living cells microinjected with fluorescent anti-NuMA antibodies were studied to examine parameters that effect the redistribution of the NuMA protein in vivo. These experiments indicate that microtubule assembly is essential for the NuMA protein to accumulate in the polar region. PMID:3527729

  8. Three-dimensional tracking of plus-tips by lattice light-sheet microscopy permits the quantification of microtubule growth trajectories within the mitotic apparatus

    Science.gov (United States)

    Yamashita, Norio; Morita, Masahiko; Legant, Wesley R.; Chen, Bi-Chang; Betzig, Eric; Yokota, Hideo; Mimori-Kiyosue, Yuko

    2015-10-01

    Mitotic apparatus, which comprises hundreds of microtubules, plays an essential role in cell division, ensuring the correct segregation of chromosomes into each daughter cell. To gain insight into its regulatory mechanisms, it is essential to detect and analyze the behavior of individual microtubule filaments. However, the discrimination of discrete microtubule filaments within the mitotic apparatus is beyond the capabilities of conventional light microscopic technologies. Recently, we detected three-dimensional (3-D) microtubule growth dynamics within the cellular cytoplasmic space using lattice light-sheet microscopy in conjunction with microtubule growth marker protein end-binding 1, a microtubule plus-end-tracking protein, which was fused to green fluorescent protein (EB1-GFP). This technique enables high-resolution 3-D imaging at subsecond intervals. We adapted mathematical computing and geometric representation techniques to analyze spatial variations in microtubule growth dynamics within the mitotic spindle apparatus. Our analytical approach enabled the different dynamic properties of individual microtubules to be determined, including the direction and speed of their growth, and their growth duration within a 3-D spatial map. Our analysis framework provides an important step toward a more comprehensive understanding of the mechanisms driving cellular machinery at the whole-cell level.

  9. Three-dimensional tracking of plus-tips by lattice light-sheet microscopy permits the quantification of microtubule growth trajectories within the mitotic apparatus.

    Science.gov (United States)

    Yamashita, Norio; Morita, Masahiko; Legant, Wesley R; Chen, Bi-Chang; Betzig, Eric; Yokota, Hideo; Mimori-Kiyosue, Yuko

    2015-10-01

    Mitotic apparatus, which comprises hundreds of microtubules, plays an essential role in cell division, ensuring the correct segregation of chromosomes into each daughter cell. To gain insight into its regulatory mechanisms, it is essential to detect and analyze the behavior of individual microtubule filaments. However, the discrimination of discrete microtubule filaments within the mitotic apparatus is beyond the capabilities of conventional light microscopic technologies. Recently, we detected three-dimensional (3-D) microtubule growth dynamics within the cellular cytoplasmic space using lattice light-sheet microscopy in conjunction with microtubule growth marker protein end-binding 1, a microtubule plus-end-tracking protein, which was fused to green fluorescent protein (EB1-GFP). This technique enables high-resolution 3-D imaging at subsecond intervals. We adapted mathematical computing and geometric representation techniques to analyze spatial variations in microtubule growth dynamics within the mitotic spindle apparatus. Our analytical approach enabled the different dynamic properties of individual microtubules to be determined, including the direction and speed of their growth, and their growth duration within a 3-D spatial map. Our analysis framework provides an important step toward a more comprehensive understanding of the mechanisms driving cellular machinery at the whole-cell level. PMID:26527322

  10. Mutation in the bimD gene of Aspergillus nidulans confers a conditional mitotic block and sensitivity to DNA damaging agents

    International Nuclear Information System (INIS)

    Mutation in the bimD gene of Aspergillus nidulans results in a mitotic block in anaphase characterized by a defective mitosis. Mutation in bimD also confers, at temperatures permissive for the mitotic arrest phenotype, an increased sensitivity to DNA damaging agents, including methyl methanesulfonate and ultraviolet light. In order to better understand the relationship between DNA damage and mitotic progression, the authors cloned the bimD gene from Aspergillus. A cosmid containing the bimD gene was identified among pools of cosmids by cotransformation with the nutritional selective pyrG gene of a strain carrying the recessive, temperature-sensitive lethal bimD6 mutation. The bimD gene encodes a predicted polypeptide of 166,000 daltons in mass and contains amino acid sequence motifs similar to those found in some DNA-binding transcription factors. These sequences include a basic domain followed by a leucine zipper, which together are called a bZIP motif, and a carboxyl-terminal domain enriched in acidic amino acids. Overexpression of the wild-type bimD protein resulted in an arrest of the nuclear division cycle that was reversible and determined to be in either the G1 or S phase of the cell cycle. The data suggest that bimD may play an essential regulatory role relating to DNA metabolism which is required for a successful mitosis. 7l refs., 7 figs., 1 tab

  11. Genotoxic profile of inhibitors of topoisomerases I (camptothecin) and II (etoposide) in a mitotic recombination and sex-chromosome loss somatic eye assay of Drosophila melanogaster.

    Science.gov (United States)

    Sortibrán, América Nitxin Castañeda; Téllez, María Guadalupe Ordaz; Rodríguez-Arnaiz, Rosario

    2006-04-30

    Genotoxic carcinogens which interact with DNA may produce double-strand breaks as normal intermediates of homologous mitotic recombination, and may give rise to structural chromosome aberrations and inter-chromosomal deletion-recombination. The genotoxic profile of two inhibitors of DNA topoisomerases were evaluated using an in vivo somatic w/w+ eye assay of Drosophila melanogaster for the detection of loss of heterozygosity (LOH) by homologous mitotic recombination, intra-chromosomal recombination and structural chromosomal aberrations. We studied camptothecin (CPT) as a topoisomerase-I-interactive agent and etoposide (ETOP) as a topoisomerase II inhibitor. These drugs act by stabilizing a ternary complex consisting of topoisomerases covalently linked to DNA at single-strand or at double-strand breaks, thereby preventing the relegation step of the breakage/rejoining reaction mediated by the enzyme. The genotoxic profiles were determined from the appearance of eye tissue in adult flies, in which LOH and expression of the reporter gene white produced light clones. The results demonstrated that both compounds were significantly genotoxic, with CPT being more effective than ETOP. Inter-chromosomal mitotic recombination was the major mechanism responsible for the induction of light spots by both compounds in XX females. Loss of the ring X chromosome (rX), was significantly enhanced by CPT, and this topoisomerase blocker also produced intra-chromosomal recombination (XY males). PMID:16529987

  12. Elevated level of spindle checkprotein MAD2 correlates with cellular mitotic arrest, but not with aneuploidy and clinicopathological characteristics in gastric cancer

    Institute of Scientific and Technical Information of China (English)

    Chew-Wun Wu; Chin-Wen Chi; Tze-Sing Huang

    2004-01-01

    AIM: To study the relevance of spindle assembly checkprotein MAD2 to cellular mitotic status, aneuploidy and other clinicopathological characteristics in gastric cancer.METHODS: Western blot analyses were performed to analyze the protein levels of MAD2 and cyclin B1 in the tumorous and adjacent nontumorous tissues of 34 gastric cancer patients. Cell cycle distribution and DNA ploidy of cancer tissues were also determined by flow cytometry.Conventional statistical methods were adopted to determine the relevance of abnormal MAD2 level to mitotic status,aneuploidy and clinicopathological parameters.RESULTS: Out of 34 gastric cancer patients 25 (74%)exhibited elevated MAD2 levels in their tumorous tissues compared with the corresponding nontumorous tissues.Elevation of MAD2 levels significantly correlated with the increased levels of cydin B1 expression and G2/M-phase distribution (P = 0.038 and P = 0.033, respectively), but was not relevant to aneuploidy. The gastric cancer patients with elevated MAD2 levels showed a tendency toward better disease-free and overall survival (P>0.05). However, no association was found between elevated MAD2 levels and patients' clinicopathological characteristics.CONCLUSION: Elevation of MAD2 level is present in 74%of gastric cancer patients, and correlates with increased mitotic checkpoint activity. However, elevation of MAD2level is not associated with patients' aneuploidy and any of the clinicopathological characteristics.

  13. 2-(3-Methoxyphenyl)-5-methyl-1,8-naphthyridin-4(1H)-one (HKL-1) induces G2/M arrest and mitotic catastrophe in human leukemia HL-60 cells

    International Nuclear Information System (INIS)

    2-(3-Methoxyphenyl)-5-methyl-1,8-naphthyridin-4(1H)-one (HKL-1), a 2-phenyl-1,8-naphthyridin-4-one (2-PN) derivative, was synthesized and evaluated as an effective antimitotic agent in our laboratory. However, the molecular mechanisms are uncertain. In this study, HKL-1 was demonstrated to induce multipolar spindles, sustain mitotic arrest and generate multinucleated cells, all of which indicate mitotic catastrophe, in human leukemia HL-60 cells. Western blotting showed that HKL-1 induces mitotic catastrophe in HL-60 cells through regulating mitotic phase-specific kinases (down-regulating CDK1, cyclin B1, CENP-E, and aurora B) and regulating the expression of Bcl-2 family proteins (down-regulating Bcl-2 and up-regulating Bax and Bak), followed by caspase-9/-3 cleavage. These findings suggest that HKL-1 appears to exert its cytotoxicity toward HL-60 cells in culture by inducing mitotic catastrophe. Highlights: ► HKL-1 is a potential antimitotic agent against HL-60 cells. ► HKL-1 induces spindle disruption and sustained resulted in mitotic catastrophe. ► CENP-E and aurora B protein expressions significantly reduced. ► Bcl-2 family protein expressions altered and caspase-9/-3 activation. ► HKL-1 is an attractive candidate for possible use as a novel antimitotic agent.

  14. Locals Collection

    Directory of Open Access Journals (Sweden)

    Stephen Hastings-King

    2010-03-01

    Full Text Available A locals collection is a set of parameters that are used to delimit data-mining operations. This piece uses a collection of locals from around Essex Massachusetts to shape and delimit an interrogation of post-reality in contemporary America. It explores the notion of crisis, the possibility of a crisis of empire that may or may not emerge in a media-space that does not allow crisis of empire to be mentioned and relations this maybe-crisis to the various levels of economic dysfunction that have become evident since late 2008. But mostly this piece explores ways in which particular stories about particular people do and do not link/link to these larger-scale narratives. This is the first of a potential series of locals collections that will mine the American post-real.

  15. Localized scleroderma.

    Science.gov (United States)

    Kreuter, Alexander

    2012-01-01

    Localized scleroderma (also called morphea) is a term encompassing a spectrum of sclerotic autoimmune diseases that primarily affect the skin, but also might involve underlying structures such as the fat, fascia, muscle, and bones. Its exact pathogenesis is still unknown, but several trigger factors in genetically predisposed individuals might initially lead to an immunologically triggered release of pro-inflammatory cytokines, resulting in a profound dysregulation of the connective tissue metabolism and ultimately to induction of fibrosis. To date, there are no specific serological markers available for localized scleroderma. Within the last years, several validated clinical scores have been introduced as potential outcome measures for the disease. Given the rarity of localized scleroderma, only few evidence-based therapeutical treatment options exist. So far, the most robust data is available for ultraviolet A1 phototherapy in disease that is restricted to the skin, and methotrexate alone or in combination with systemic corticosteroids in more severe disease that additionally affects extracutaneous structures. This practical review summarizes relevant information on the epidemiology, pathogenesis, clinical subtypes and classifications, differential diagnoses, clinical scores and outcome measures, and current treatment strategies of localized scleroderma. PMID:22741933

  16. Local food:

    DEFF Research Database (Denmark)

    Sundbo, Donna Isabella Caroline

    2013-01-01

    consumers, who emphasise social properties of production more than providers. Experience marketing and events may create diffusion of a local food concept, but only when communication is massive and long-lasting. So far this has not been the case, hence there is no agreement in either theory or among...

  17. Local language

    NARCIS (Netherlands)

    Monique Turkenburg

    2002-01-01

    Original title: Taal lokaal. Children of immigrants living in the Netherlands have for years had the opportunity to receive lessons in their mother tongue at primary school. Since 1998 this has been referred to as minority language teaching (OALT in Dutch), and has been the responsibility of local

  18. SGO1 maintains bovine meiotic and mitotic centromeric cohesions of sister chromatids and directly affects embryo development.

    Directory of Open Access Journals (Sweden)

    Feng-Xia Yin

    Full Text Available Shugoshin (SGO is a critical factor that enforces cohesion from segregation of paired sister chromatids during mitosis and meiosis. It has been studied mainly in invertebrates. Knowledge of SGO(s in a mammalian system has only been reported in the mouse and Hela cells. In this study, the functions of SGO1 in bovine oocytes during meiotic maturation, early embryonic development and somatic cell mitosis were investigated. The results showed that SGO1 was expressed from germinal vesicle (GV to the metaphase II stage. SGO1 accumulated on condensed and scattered chromosomes from pre-metaphase I to metaphase II. The over-expression of SGO1 did not interfere with the process of homologous chromosome separation, although once separated they were unable to move to the opposing spindle poles. This often resulted in the formation of oocytes with 60 replicated chromosomes. Depletion of SGO1 in GV oocytes affected chromosomal separation resulting in abnormal chromosome alignment at a significantly higher proportion than in control oocytes. Knockdown of SGO1 expression significantly decreased the embryonic developmental rate and quality. To further confirm the function(s of SGO1 during mitosis, bovine embryonic fibroblast cells were transfected with SGO1 siRNAs. SGO1 depletion induced the premature dissociation of chromosomal cohesion at the centromere and along the chromosome arm giving rise to abnormal appearing mitotic patterns. The results of this study infer that SGO1 is involved in the centromeric cohesion of sister chromatids and chromosomal movement towards the spindle poles. Depletion of SGO1 causes arrestment of cell division in meiosis and mitosis.

  19. Mitotic chromosomal abnormalities in diploid and colchicine-induced tetraploid jute (Corchorus capsularis L.) following gamma-rays treatment

    International Nuclear Information System (INIS)

    The purpose of the present study was to detect and explain the various types of chromosomal abnormalities resulting from seed irradiation and to evaluate the frequency of abnormal cells in different doses and thereby establish the relative radiosensitivity between the 2x (2n = 14) and c-4x (2n = 28) (colchicine-induced) Corchorus capsularis. Air-dried seeds of the two ploidy levels were exposed to 60, 70 and 80 KR gamma-rays (60Co) and different stages of the mitotic division of the treated and control root tips were studied. There was no chromosomal abnormality in the untreated root-tip cells of both the types. The aberrations in the irradiated samples included: a) only bridges, b) only laggards and c) both bridges and laggards in the anaphase of root-tip cells of both the ploidy levels, d) much more variation in length of metaphase chromosomes in c-4x capsularis. Plausible causes of the origin and occurrence of the abnormalities have been discussed in the light of the findings of other workers in the field. There was increase in the frequency of abnormal cells with the increase in radiation dose in both the ploidy levels but at a given dose c-4x capsularis produced more aberrations than the 2x diploid. It was concluded that the artificially induced autotetoraploid was more radio-sensitive than the 2x type. This perhaps, was due to the artificial nature of origin of the autotetraploid and its lack of sufficient adaptation to protect itself from radiation effect. (author)

  20. Cytokinesis-block micronucleus assay evolves into a 'cytome' assay of chromosomal instability, mitotic dysfunction and cell death

    International Nuclear Information System (INIS)

    The cytokinesis-block micronucleus (CBMN) assay was originally developed as an ideal system for measuring micronuclei (MNi) however it can also be used to measure nucleoplasmic bridges (NPBs), nuclear buds (NBUDs), cell death (necrosis or apoptosis) and nuclear division rate. Current evidence suggests that (a) NPBs originate from dicentric chromosomes in which the centromeres have been pulled to the opposite poles of the cell at anaphase and are therefore indicative of DNA mis-repair, chromosome rearrangement or telomere end-fusions, (b) NPBs may break to form MNi, (c) the nuclear budding process is the mechanism by which cells remove amplified and/or excess DNA and is therefore a marker of gene amplification and/or altered gene dosage, (d) cell cycle checkpoint defects result in micronucleus formation and (e) hypomethylation of DNA, induced nutritionally or by inhibition of DNA methyl transferase can lead to micronucleus formation either via chromosome loss or chromosome breakage. The strong correlation between micronucleus formation, nuclear budding and NPBs (r = 0.75-0.77, P < 0.001) induced by either folic acid deficiency or exposure to ionising radiation is supportive of the hypothesis that folic acid deficiency and/or ionising radiation cause genomic instability and gene amplification by the initiation of breakage-fusion-bridge cycles. In its comprehensive mode, the CBMN assay measures all cells including necrotic and apoptotic cells as well as number of nuclei per cell to provide a measure of cytotoxicity and mitotic activity. The CBMN assay has in fact evolved into a 'cytome' method for measuring comprehensively chromosomal instability phenotype and altered cellular viability caused by genetic defects and/or nutrional deficiencies and/or exogenous genotoxins thus opening up an exciting future for the use of this methodology in the emerging fields of nutrigenomics and toxicogenomics and their combinations

  1. Evaluation of radioprotection properties of propolis by chromosomal alterations, cell proliferation kinetics, mitotic index and sister chromatid exchange

    International Nuclear Information System (INIS)

    A consequence of ionizing radiation is the induction of chromosomal alterations. This causality relation involves that chromosomal alterations can be considered a good indicator of the radiological damage. Some chemical agents can modulate the tissue response to radiation. These compounds are useful because they show certain selectivity, protecting the healthy tissues (radioprotectors) or increasing the sensibility of tissues to radiations (radiosensibilizators). Propolis substance has showed radioprotection properties which are performed in the following study. Propolis is a product of extraordinary interest for both medicine and pharmaceutical industry, since it is assumed to show diverse beneficial health effects. Among many other attributes of EEP (propolis ethanolic extract), it exhibits antioxidant and radical free scavenger properties. In a previous study, human peripheral blood lymphocytes were exposed to 2 Gy of γ rays in presence and absence of EEP, and the analysis showed a reduction in the frequency of dicentrics and rings, with a maximum protection of 50%. The proposed concentration for radioprotection would be between 120-500 μg.ml-1. The cytotoxic effect has been evaluated analyzing the EEP effect in the cellular division cycle. Propolis ethanolic extract (EEP) has been obtained and samples of peripheral blood have been cultured in the presence of increasing concentrations of EEP. In order to quantify it, two indexes have been used, the mitotic index and cell proliferation index. For both indexes the cytotoxic effect takes place from 750 μg.ml-1 concentrations onwards. Similar results were obtained for the analysis of chromosomal aberrations. Finally, propolis effect in lymphocytes by sister chromatid exchange test has been presented for higher concentrations of EEP. (author)

  2. pH-induced changes in mitotic and developmental patterns in sea urchin embryogenesis. I. Exposure of embryos.

    Science.gov (United States)

    Pagano, G; Cipollaro, M; Corsale, G; Esposito, A; Ragucci, E; Giordano, G G

    1985-01-01

    The effects of different pH conditions have been investigated on sea urchin larval development following exposure of embryos to controlled, though changing, decreases or increases of seawater pH. The pH of filtered natural seawater was initially adjusted with 1 N HCl of 1 N KOH and then was altered back to its normal values (8.0-8.2) by the exchange with atmospheric CO2 and subsequent carbonic acid equilibrium. During cultures, pH was regularly monitored. When developing embryos were reared in different pH conditions, larval differentiation was sharply affected by an apparently moderate pH decrease, such as 0.5 pH units. However, even pH decreases as small as 0.2 pH units from the normal value showed reproducible damage to embryogenesis. This damage appeared to be early and irreversible, since the exposure of cleaving embryos resulted in more severe developmental defects than exposure of posthatching blastulae. Moreover, mitotic abnormalities were observed following early exposure of embryos to decreased pH. Increased pH, up to 8.6 (approximately 0.5 pH units above normal value), failed to exert any adverse effect on subsequent development. Moreover, an initial pH increase (8.5-8.7) resulted in the final adjustment of culture pH to 8.1-8.2, thus providing optimal conditions for rearing embryos. Two attempts to stabilize culture pH were performed by decreasing gaseous exchanges or by using Tris as a buffering agent. Both approaches appeared to be impractical, thus ruling out any further attempts. The results point out the hazards of acid contamination in restricted bodies of seawater, leading to apparently "moderate" decreases in pH, which can result in severe damage to some marine organisms, both adult and larval forms. PMID:2859664

  3. Auxin/AID versus conventional knockouts: distinguishing the roles of CENP-T/W in mitotic kinetochore assembly and stability

    Science.gov (United States)

    Wood, Laura; Booth, Daniel G.; Vargiu, Giulia; Ohta, Shinya; deLima Alves, Flavia; Samejima, Kumiko; Fukagawa, Tatsuo; Rappsilber, Juri; Earnshaw, William C.

    2016-01-01

    Most studies using knockout technologies to examine protein function have relied either on shutting off transcription (conventional conditional knockouts with tetracycline-regulated gene expression or gene disruption) or destroying the mature mRNA (RNAi technology). In both cases, the target protein is lost at a rate determined by its intrinsic half-life. Thus, protein levels typically fall over at least 1–3 days, and cells continue to cycle while exposed to a decreasing concentration of the protein. Here we characterise the kinetochore proteome of mitotic chromosomes isolated from a cell line in which the essential kinetochore protein CENP-T is present as an auxin-inducible degron (AID) fusion protein that is fully functional and able to support the viability of the cells. Stripping of the protein from chromosomes in early mitosis via targeted proteasomal degradation reveals the dependency of other proteins on CENP-T for their maintenance in kinetochores. We compare these results with the kinetochore proteome of conventional CENP-T/W knockouts. As the cell cycle is mostly formed from G1, S and G2 phases a gradual loss of CENP-T/W levels is more likely to reflect dependencies associated with kinetochore assembly pre-mitosis and upon entry into mitosis. Interestingly, a putative super-complex involving Rod-Zw10-zwilch (RZZ complex), Spindly, Mad1/Mad2 and CENP-E requires the function of CENP-T/W during kinetochore assembly for its stable association with the outer kinetochore, but once assembled remains associated with chromosomes after stripping of CENP-T during mitosis. This study highlights the different roles core kinetochore components may play in the assembly of kinetochores (upon entry into mitosis) versus the maintenance of specific components (during mitosis). PMID:26791246

  4. Evaluation of techniques for C and ASG banding of the mitotic chromosomes of Anastrepha species (Diptera, Tephritidae

    Directory of Open Access Journals (Sweden)

    Denise Selivon

    1997-12-01

    Full Text Available Methods previously described by Canovai et al. (Caryologia 47: 241-247, 1994 which produced C and ASG bands in mitotic chromosomes of Ceratitis capitata were applied to the chromosomes of several Anastrepha species. Metaphase plate yield was substantially increased by use of imaginal disks together with cerebral ganglia. The C-bands were quite prominent allowing the resolution of tiny blocks of heterochromatin. The ASG method produced G-like banded chromosomes, which permitted recognition of each individual chromosome. These simple techniques do not require special equipment and may be valuable for karyotype variability studies in fruit flies and other DipteraTécnicas descritas anteriormente por Canovai et al. (Caryologia 47: 241-247, 1994, que produzem bandas C e ASG nos cromossomos mitóticos de Ceratitis capitata, foram testadas com os cromossomos de várias espécies de Anastrepha. O número de metáfases analisáveis aumentou significativamente quando, além dos gânglios cerebrais, discos imaginais foram utilizados nas preparações. O bandamento tipo C produzido foi bastante conspícuo, permitindo inclusive a resolução de pequenos blocos de heterocromatina. O método ASG produziu uma fração significativa de metáfases cujos cromossomos apresentavam um nítido bandamento do tipo G, que permitiu diferenciar cada cromossomo do cariótipo. Essas técnicas simples, aplicadas sem a necessidade de equipamento especial, podem ser úteis para estudos de variabilidade cariotípica intra ou inter-específica

  5. STUDIES ON THE PHYTOCHEMISTRY, SPECTROSCOPIC CHARACTERIZATION AND SCREENING FOR ANTI-MITOTIC EFFICACY OF SALICORNIA BRACHIATA ROXB

    Directory of Open Access Journals (Sweden)

    G.V.Pavan Kumar

    2014-10-01

    Full Text Available Salicorniabrachiata is a euhalopytic plant belonging to the family Chenopodiaceae. The present study investigates the phytochemistry, characterization and antimitotic activity of ethanolic extract of S.brachiata.Plants popularly known as Sea asparagus are cooked and eaten or pickled. It is also a good fodder for cattle, sheep and goat. Plant material is also used as raw material in paper and board factories. Its seeds yield high quality edible oil which is highly polyunsaturated and similar to safflower oil in fatty acid.S.brachiata was collected from the back waters of Bapatla,Guntur district. The collected plant material was shade dried and pulverized. The plant material Was studied for phytochemistry,spectroscopic analysis i.e.,UV- Visible, FT-IR and anti mitotic activity.S. brachiata has been prescribed in traditional medicines for the treatment of intestinal ailments, nephropathy, and hepatitis in Oriental countries. In addition, S.brachiata has recently reported to be effective on the atherosclerosis, hyperlipidemia, and diabetes. A variety of pharmacological experiments have revealed that solvent-extracted fractions of S.brachiata exhibited anti-oxidative, anti-microbial, anti-proliferative, and anti-inflammatory activities,supporting rationale behind its several traditional uses.The phytochemical analysis indicates the presence of Tannins and Flavonoids in the plant. UV-Vis Spectrum, used for the quantitative analysis of the plant extract showed peaks at 280 and 290 nm. Identification of the functional groups was performed by FT-IR spectroscopy which confirmed the presence of phenolic, alcoholic and aromatic compounds.

  6. Local supertwistors

    International Nuclear Information System (INIS)

    Geometry of local supertwistors is investigated. It is proved that the Yang-Mills equations for the introduced ansatz for supertwistor connection are equivalent to free bach equations, describing the dynamics of N=1 conformal supergravity. Analogous interpretation of the dynamics of N=1 conformal supergravity coupled to a vector superfield is proposed. It is proved that any complex conformally right or left flat superspace automatically satisfies the Bach equations

  7. Actin and Arp2/3 localize at the centrosome of interphase cells

    Energy Technology Data Exchange (ETDEWEB)

    Hubert, Thomas; Vandekerckhove, Joel; Gettemans, Jan, E-mail: jan.gettemans@vib-ugent.be

    2011-01-07

    Research highlights: {yields} Actin was detected at the centrosome with the anti-actin antibody 1C7 that recognizes antiparallel ('lower dimer') actin dimers. {yields} Centrosomal actin was found in interphase but not mitotic MDA-MB-231 cells. {yields} Neither the anti-actin antibody C4 that binds to globular, monomer actin, nor the anti-actin antibody 2G2 that recognizes the nuclear conformation of actin detect actin at the centrosome. {yields} The Arp2/3 complex transiently localizes at the pericentriolar matrix but not at the centrioles of interphase HEK 293T cells. -- Abstract: Although many actin binding proteins such as cortactin and the Arp2/3 activator WASH localize at the centrosome, the presence and conformation of actin at the centrosome has remained elusive. Here, we report the localization of actin at the centrosome in interphase but not in mitotic MDA-MB-231 cells. Centrosomal actin was detected with the anti-actin antibody 1C7 that recognizes antiparallel ('lower dimer') actin dimers. In addition, we report the transient presence of the Arp2/3 complex at the pericentriolar matrix but not at the centrioles of interphase HEK 293T cells. Overexpression of an Arp2/3 component resulted in expansion of the pericentriolar matrix and selective accumulation of the Arp2/3 component in the pericentriolar matrix. Altogether, we hypothesize that the centrosome transiently recruits Arp2/3 to perform processes such as centrosome separation prior to mitotic entry, whereas the observed constitutive centrosomal actin staining in interphase cells reinforces the current model of actin-based centrosome reorientation toward the leading edge in migrating cells.

  8. Mitotic Events in Cerebellar Granule Progenitor Cells that Expand Cerebellar Surface Area Are Critical for Normal Cerebellar Cortical Lamination in Mice

    Science.gov (United States)

    Chang, Joshua C.; Leung, Mark; Gokozan, Hamza Numan; Gygli, Patrick Edwin; Catacutan, Fay Patsy; Czeisler, Catherine; Otero, José Javier

    2015-01-01

    Late embryonic and postnatal cerebellar folial surface area expansion promotes cerebellar cortical cytoarchitectural lamination. We developed a streamlined sampling scheme to generate unbiased estimates of murine cerebellar surface area and volume using stereological principles. We demonstrate that during the proliferative phase of the external granule layer (EGL) and folial surface area expansion, EGL thickness does not change and thus is a topological proxy for progenitor self-renewal. The topological constraints indicate that during proliferative phases, migration out of the EGL is balanced by self-renewal. Progenitor self-renewal must, therefore, include mitotic events yielding either 2 cells in the same layer to increase surface area (β-events) and mitotic events yielding 2 cells, with 1 cell in a superficial layer and 1 cell in a deeper layer (α-events). As the cerebellum grows, therefore, β-events lie upstream of α-events. Using a mathematical model constrained by the measurements of volume and surface area, we could quantify inter-mitotic times for β-events on a per-cell basis in post-natal mouse cerebellum. Furthermore, we found that loss of CCNA2, which decreases EGL proliferation and secondarily induces cerebellar cortical dyslamination, shows preserved α-type events. Thus, CCNA2-null cerebellar granule progenitor cells are capable of self-renewal of the EGL stem cell niche; this is concordant with prior findings of extensive apoptosis in CCNA2-null mice. Similar methodologies may provide another layer of depth to the interpretation of results from stereological studies. PMID:25668568

  9. Synergistic role of fission yeast Alp16GCP6 and Mzt1MOZART1 in γ-tubulin complex recruitment to mitotic spindle pole bodies and spindle assembly.

    Science.gov (United States)

    Masuda, Hirohisa; Toda, Takashi

    2016-06-01

    In fission yeast, γ-tubulin ring complex (γTuRC)-specific components Gfh1(GCP4), Mod21(GCP5), and Alp16(GCP6) are nonessential for cell growth. Of these deletion mutants, only alp16Δ shows synthetic lethality with temperature-sensitive mutants of Mzt1(MOZART1), a component of the γTuRC required for recruitment of the complex to microtubule-organizing centers. γ-Tubulin small complex levels at mitotic spindle pole bodies (SPBs, the centrosome equivalent in fungi) and microtubule levels for preanaphase spindles are significantly reduced in alp16Δ cells but not in gfh1Δ or mod21Δ cells. Furthermore, alp16Δ cells often form monopolar spindles and frequently lose a minichromosome when the spindle assembly checkpoint is inactivated. Alp16(GCP6) promotes Mzt1-dependent γTuRC recruitment to mitotic SPBs and enhances spindle microtubule assembly in a manner dependent on its expression levels. Gfh1(GCP4) and Mod21(GCP5) are not required for Alp16(GCP6)-dependent γTuRC recruitment. Mzt1 has an additional role in the activation of the γTuRC for spindle microtubule assembly. The ratio of Mzt1 to γTuRC levels for preanaphase spindles is higher than at other stages of the cell cycle. Mzt1 overproduction enhances spindle microtubule assembly without affecting γTuRC levels at mitotic SPBs. We propose that Alp16(GCP6) and Mzt1 act synergistically for efficient bipolar spindle assembly to ensure faithful chromosome segregation. PMID:27053664

  10. Local supertwistors

    International Nuclear Information System (INIS)

    The geometry of local supertwistors is investigated. An ansatz on the form of the supertwistor superconnection is introduced. Because of this restriction on the form of such a superconnection the Yang-Mills equations for the superconnection turn out to be equivalent to the free Bach equations describing the dynamics of simple conformal supergravity. It is shown that the equations of motion of conformal supergravity interacting with a vector superfield admit an analogous interpretation. It is proved that an arbitrary conformally right-flat or left-flat superspace is automatically a solution of the Bach equations

  11. Influence of duration of fixation on the yield of chromosome aberrations in human lumphocyte culture exposed to γ-radiation at different mitotic cycle stages

    International Nuclear Information System (INIS)

    A comparative study was made of the yield of chromosome aberrations in human lymphocyte culture exposed to 60Co- γ-rays (2 Gy) at different mitotic cycle stages the cells being fixed after 52 and 60 hr. It was shown that with the latter fixation time (60 hr) the frequency of chromosome aberrations after irradiation in G1 stage was substantially lower than that with the former one (52 hr) and, vice versa, it was higher after irradiation in S and G2 stages. The authors discuss the probable causes of the distinctions observed

  12. The fission yeast protein p73res2 is an essential component of the mitotic MBF complex and a master regulator of meiosis.

    OpenAIRE

    Ayté, J; Leis, J F; DeCaprio, J A

    1997-01-01

    Depending on environmental conditions, Schizosaccharomyces pombe can remain in the stationary phase or enter into either premitotic or premeiotic DNA synthesis. This decision point is known as Start. In the mitotic cell cycle, regulation of G1/S-specific gene expression is dependent upon the MBF (Mlu1 binding factor) complex, known to contain p85cdc10 and p72res1. Here we demonstrate that p73res2 controls cell cycle progression via its participation in the MBF complex, interacting directly wi...

  13. Influence of duration of fixation on the yield of chromosome aberrations in human lumphocyte culture exposed to. gamma. -radiation at different mitotic cycle stages

    Energy Technology Data Exchange (ETDEWEB)

    Sevan' kaev, A.V.; Bogatykh, B.A.; Lychev, V.A. (Akademiya Meditsinskikh Nauk SSSR, Obninsk. Nauchno-Issledovatel' skij Inst. Meditsinskoj Radiologii)

    A comparative study was made of the yield of chromosome aberrations in human lymphocyte culture exposed to /sup 60/Co- ..gamma..-rays (2 Gy) at different mitotic cycle stages the cells being fixed after 52 and 60 hr. It was shown that with the latter fixation time (60 hr) the frequency of chromosome aberrations after irradiation in G/sub 1/ stage was substantially lower than that with the former one (52 hr) and, vice versa, it was higher after irradiation in S and G/sub 2/ stages. The authors discuss the probable causes of the distinctions observed.

  14. Emi1 is needed to couple DNA replication with mitosis but does not regulate activation of the mitotic APC/C

    OpenAIRE

    Di Fiore, Barbara; Pines, Jonathon

    2007-01-01

    Ubiquitin-mediated proteolysis is critical for the alternation between DNA replication and mitosis and for the key regulatory events in mitosis. The anaphase-promoting complex/cyclosome (APC/C) is a conserved ubiquitin ligase that has a fundamental role in regulating mitosis and the cell cycle in all eukaryotes. In vertebrate cells, early mitotic inhibitor 1 (Emi1) has been proposed as an important APC/C inhibitor whose destruction may trigger activation of the APC/C at mitosis. However, in t...

  15. Acceleration of mitotic cycle in meristem cells of seedlings roots by gamma irradiation of pea and maize seeds at stimulating doses

    International Nuclear Information System (INIS)

    The seeds were irradiated with 500 or 1,000 rads of 60Co γ-rays and laid on filter paper for germination. After 2-3 d the seedlings were placed for 30 min. into a nutrient solution containing 3H-thymidine. Afterwards the seedlings were placed in a non-radioactive nutrient solution and every 2 h the root tip was fixed and prepared for autoradiography for determination of cell division. Irradiation of both the pea and maize seeds resulted in a reduction of the duration of the mitotic cycle, the individual phases being shortened to a different extent. Stimulating radiation doses increased the activity of cell division. (MG)

  16. Mitotic chromosome structure

    International Nuclear Information System (INIS)

    Mounting evidence is compiling linking the physical organizational structure of chromosomes and the nuclear structure to biological function. At the base of the physical organizational structure of both is the concept of loop formation. This implies that physical proximity within chromosomes is provided for otherwise distal genomic regions and thus hierarchically organizing the chromosomes. Together with entropy many experimental observations can be explained with these two concepts. Among the observations that can be explained are the measured physical extent of the chromosomes, their shape, mechanical behavior, the segregation into territories (chromosomal and territories within chromosomes), the results from chromosome conformation capture experiments, as well as linking gene expression to structural organization.

  17. Local Professionals for Local Market

    Institute of Scientific and Technical Information of China (English)

    Wen Xiaojie

    2010-01-01

    @@ In the past three decades,the Chinese hotel industry has developed at a rapid pace,with the number of hotels-especially high star hotelsgrowing fast.In Beijing alone,there arc nearly 60 five-star hotels.With the development of the hotel industry.China has also begun to see the number of local hotel professionals,including senior managers,increase.Wen Xiaojie,Deputy General Manager and Owner's Representative of Sofitel Wanda Beijing,is among the most outstanding senior hotel managers.

  18. Computational fragment-based drug design to explore the hydrophobic sub-pocket of the mitotic kinesin Eg5 allosteric binding site.

    Science.gov (United States)

    Oguievetskaia, Ksenia; Martin-Chanas, Laetitia; Vorotyntsev, Artem; Doppelt-Azeroual, Olivia; Brotel, Xavier; Adcock, Stewart A; de Brevern, Alexandre G; Delfaud, Francois; Moriaud, Fabrice

    2009-08-01

    Eg5, a mitotic kinesin exclusively involved in the formation and function of the mitotic spindle has attracted interest as an anticancer drug target. Eg5 is co-crystallized with several inhibitors bound to its allosteric binding pocket. Each of these occupies a pocket formed by loop 5/helix alpha2 (L5/alpha2). Recently designed inhibitors additionally occupy a hydrophobic pocket of this site. The goal of the present study was to explore this hydrophobic pocket with our MED-SuMo fragment-based protocol, and thus discover novel chemical structures that might bind as inhibitors. The MED-SuMo software is able to compare and superimpose similar interaction surfaces upon the whole protein data bank (PDB). In a fragment-based protocol, MED-SuMo retrieves MED-Portions that encode protein-fragment binding sites and are derived from cross-mining protein-ligand structures with libraries of small molecules. Furthermore we have excluded intra-family MED-Portions derived from Eg5 ligands that occupy the hydrophobic pocket and predicted new potential ligands by hybridization that would fill simultaneously both pockets. Some of the latter having original scaffolds and substituents in the hydrophobic pocket are identified in libraries of synthetically accessible molecules by the MED-Search software. PMID:19533373

  19. Mitotic protein kinase CDK1 phosphorylation of mRNA translation regulator 4E-BP1 Ser83 may contribute to cell transformation

    Energy Technology Data Exchange (ETDEWEB)

    Velasquez, Celestino; Cheng, Erdong; Shuda, Masahiro; Lee-Oesterreich, Paula J.; Pogge von Strandmann, Lisa; Gritsenko, Marina A.; Jacobs, Jon M.; Moore, Patrick S.; Chang, Yuan

    2016-07-26

    mTOR-directed 4E-BP1 phosphorylation promotes cap-dependent translation and tumorigen-esis. During mitosis, CDK1 substitutes for mTOR and fully phosphorylates 4E-BP1 at canoni-cal as well a non-canonical S83 site resulting in a mitosis-specific hyperphosphorylated δ isoform. Colocalization studies with a phospho-S83 specific antibody indicate that 4E-BP1 S83 phosphorylation accumulates at centrosomes during prophase, peaks at metaphase, and decreases through telophase. While S83 phosphorylation of 4E-BP1 does not affect in vitro cap-dependent translation, nor eIF4G/4E-BP1 cap-binding, expression of an alanine substitution mutant 4E-BP1.S83A partially reverses rodent cell transformation induced by Merkel cell polyomavirus (MCV) small T (sT) antigen viral oncoprotein. In contrast to inhibitory mTOR 4E-BP1 phosphorylation, these findings suggest that mitotic CDK1-directed phosphorylation of δ-4E-BP1 may yield a gain-of-function, distinct from translation regulation, that may be important in tumorigenesis and mitotic centrosome function.

  20. Mitotic regulator Nlp interacts with XPA/ERCC1 complexes and regulates nucleotide excision repair (NER) in response to UV radiation.

    Science.gov (United States)

    Ma, Xiao-Juan; Shang, Li; Zhang, Wei-Min; Wang, Ming-Rong; Zhan, Qi-Min

    2016-04-10

    Cellular response to DNA damage, including ionizing radiation (IR) and UV radiation, is critical for the maintenance of genomic fidelity. Defects of DNA repair often result in genomic instability and malignant cell transformation. Centrosomal protein Nlp (ninein-like protein) has been characterized as an important cell cycle regulator that is required for proper mitotic progression. In this study, we demonstrate that Nlp is able to improve nucleotide excision repair (NER) activity and protects cells against UV radiation. Upon exposure of cells to UVC, Nlp is translocated into the nucleus. The C-terminus (1030-1382) of Nlp is necessary and sufficient for its nuclear import. Upon UVC radiation, Nlp interacts with XPA and ERCC1, and enhances their association. Interestingly, down-regulated expression of Nlp is found to be associated with human skin cancers, indicating that dysregulated Nlp might be related to the development of human skin cancers. Taken together, this study identifies mitotic protein Nlp as a new and important member of NER pathway and thus provides novel insights into understanding of regulatory machinery involved in NER. PMID:26805762

  1. Mitotic protein kinase CDK1 phosphorylation of mRNA translation regulator 4E-BP1 Ser83 may contribute to cell transformation.

    Science.gov (United States)

    Velásquez, Celestino; Cheng, Erdong; Shuda, Masahiro; Lee-Oesterreich, Paula J; Pogge von Strandmann, Lisa; Gritsenko, Marina A; Jacobs, Jon M; Moore, Patrick S; Chang, Yuan

    2016-07-26

    Mammalian target of rapamycin (mTOR)-directed eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1) phosphorylation promotes cap-dependent translation and tumorigenesis. During mitosis, cyclin-dependent kinase 1 (CDK1) substitutes for mTOR and fully phosphorylates 4E-BP1 at canonical sites (T37, T46, S65, and T70) and the noncanonical S83 site, resulting in a mitosis-specific hyperphosphorylated δ isoform. Colocalization studies with a phospho-S83 specific antibody indicate that 4E-BP1 S83 phosphorylation accumulates at centrosomes during prophase, peaks at metaphase, and decreases through telophase. Although S83 phosphorylation of 4E-BP1 does not affect general cap-dependent translation, expression of an alanine substitution mutant 4E-BP1.S83A partially reverses rodent cell transformation induced by Merkel cell polyomavirus small T antigen viral oncoprotein. In contrast to inhibitory mTOR 4E-BP1 phosphorylation, these findings suggest that mitotic CDK1-directed phosphorylation of δ-4E-BP1 may yield a gain of function, distinct from translation regulation, that may be important in tumorigenesis and mitotic centrosome function. PMID:27402756

  2. Localized Excitations from Localized Unitary Operators

    CERN Document Server

    Sivaramakrishnan, Allic

    2016-01-01

    Localized unitary operators are basic probes of locality and causality in quantum systems: localized unitary operators create localized excitations in entangled states. Working with an explicit form, we explore the properties of these operators in quantum mechanics and quantum field theory. We show that, unlike unitary operators, local non-unitary operators generically create non-local excitations. We present a local picture for quantum systems in which localized experimentalists can only act through localized Hamiltonian deformations, and therefore localized unitary operators. We demonstrate that localized unitary operators model certain quantum quenches exactly. We show how the Reeh-Schlieder theorem follows intuitively from basic properties of entanglement, non-unitary operators, and the local picture. We show that a recent quasi-particle picture for excited-state entanglement entropy in conformal field theories is not universal for all local operators. We prove a causality relation for entanglement entrop...

  3. Local Sustainability

    International Nuclear Information System (INIS)

    The current polemic about the possibilities of sustainable development has led to a renovated interest for the topic of the sustainability of the communities and the local sustainability. In front of the global sustainability whose conditions have been exposed by systemic ecologists and for macro economists, the sustainability of specific places arises in the planet whose conditions are object of study of the ecology of landscapes, of the ecological economy, of the cultural anthropology, of the environmental sociology and naturally, of the integral environmentalism. In this discussion the Colombian case charges unusual interest to be one of the few countries of Latin America, where a very dense net of municipalities exists, each one with its urban helmet and with a position and some functions defined by the political constitution of the nation. This net of municipalities and of urban helmets it also constitutes net of alternative to the current macro-cephalic situation. As well as Bogota grew, in a hundred years, of less than a hundred thousand inhabitants to six million inhabitants, each one of these municipalities contains a potential of growth that depends on the characteristics of its ecological, social, economic and politic sustainability

  4. Quantum Locality?

    Energy Technology Data Exchange (ETDEWEB)

    Stapp, Henry

    2011-11-10

    vagaries that he cites do not upset the proof in question. It is show here in detail why the precise statement of this theorem justifies the specified application of CQT. It is also shown, in response to his challenge, why a putative proof of locality that he has proposed is not valid.

  5. Inhibitor-3 ensures bipolar mitotic spindle attachment by limiting association of SDS22 with kinetochore-bound protein phosphatase-1

    OpenAIRE

    Eiteneuer, Annika; Seiler, Jonas; Weith, Matthias; Beullens, Monique; Lesage, Bart; Krenn, Veronica; Musacchio, Andrea; Bollen, Mathieu; Meyer, Hemmo

    2014-01-01

    Faithful chromosome segregation during mitosis is tightly regulated by opposing activities of Aurora B kinase and protein phosphatase-1 (PP1). PP1 function at kinetochores has been linked to SDS22, but the exact localization of SDS22 and how it affects PP1 are controversial. Here, we confirm that SDS22 is required for PP1 activity, but show that SDS22 does not normally localize to kinetochores. Instead, SDS22 is kept in solution by formation of a ternary complex with PP1 and inhibitor-3 (I3)....

  6. Anti-mitotic potential of 7-diethylamino-3(2 Prime -benzoxazolyl)-coumarin in 5-fluorouracil-resistant human gastric cancer cell line SNU620/5-FU

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Nam Hyun [Department of Pharmacology, Kwandong University College of Medicine, Gangneung 210-701 (Korea, Republic of); Kim, Su-Nam [KIST Gangneung Institute, Gangneung 210-340 (Korea, Republic of); Oh, Joa Sub [College of Pharmacy, Dankook University, Cheonan 330-714 (Korea, Republic of); Lee, Seokjoon [Department of Basic Science, Kwandong University College of Medicine, Gangneung 210-701 (Korea, Republic of); Kim, Yong Kee, E-mail: yksnbk@sookmyung.ac.kr [College of Pharmacy, Sookmyung Women' s University, Seoul 140-742 (Korea, Republic of)

    2012-02-24

    Highlights: Black-Right-Pointing-Pointer DBC exerts antiproliferative potential against 5FU-resistant human gastric cancer cells. Black-Right-Pointing-Pointer This effect is mediated by destabilization of microtubules and subsequent mitotic arrest. Black-Right-Pointing-Pointer DBC enhances apoptosis via caspase activation and downregulation of antiapoptotic genes. -- Abstract: In this study, we investigate an anti-mitotic potential of the novel synthetic coumarin-based compound, 7-diethylamino-3(2 Prime -benzoxazolyl)-coumarin, in 5-fluorouracil-resistant human gastric cancer cell line SNU-620-5FU and its parental cell SNU-620. It exerts the anti-proliferative effects with similar potencies against both cancer cells, which is mediated by destabilization of microtubules and subsequent mitotic arrest. Furthermore, this compound enhances caspase-dependent apoptotic cell death via decreased expression of anti-apoptotic genes. Taken together, our data strongly support anti-mitotic potential of 7-diethylamino-3(2 Prime -benzoxazolyl)-coumarin against drug-resistant cancer cells which will prompt us to further develop as a novel microtubule inhibitor for drug-resistant cancer chemotherapy.

  7. Protein synthesis rate measured with l-[1-11C]tyrosine positron emission tomography correlates with mitotic activity and MIB-1 antibody-detected proliferation in human soft tissue sarcomas

    International Nuclear Information System (INIS)

    Protein synthesis rate (PSR) can be assessed in vivo using positron emission tomography with l-[1-11C]tyrosine (TYR-PET). Biological activity of soft tissue sarcomas (STS) can be measured in vitro by the mitotic rate and number of proliferating cells. In STS the grade of malignancy, in which the mitotic index plays a major role, is considered to be the major standard in predicting biological tumour behaviour. This study was designed to test the validity of TYR-PET in relation to different histopathological features. In 21 patients with untreated STS, the PSR was measured using TYR-PET. The number of mitoses was counted and tumours were graded according to the grading system of Coindre et al. (Cancer 1986; 58:306-309). Proliferative activity was assessed by immunohistological detection of the Ki-67 nuclear antigen using MIB-1 monoclonal antibody. To test the association between the PSR and these tumour parameters, a correlation analysis was performed. A significant (P<0.05) correlation was found between PSR and the Ki-67 proliferation index (R = 0.54), and between PSR and mitotic rate (R = 0.64). There was no correlation between PSR and tumour grade. The present study in malignant soft tissue tumours relates in vivo tumour metabolism as established with TYR-PET to tumour activity measured in vitro and indicates that the non-invasive method of TYR-PET can estimate the mitotic and proliferative activity in STS. (orig.)

  8. Intelligent CAD System for Automatic Detection of Mitotic Cells from Breast Cancer Histology Slide Images Based on Teaching-Learning-Based Optimization

    Directory of Open Access Journals (Sweden)

    Ramin Nateghi

    2014-01-01

    Full Text Available This paper introduces a computer-assisted diagnosis (CAD system for automatic mitosis detection from breast cancer histopathology slide images. In this system, a new approach for reducing the number of false positives is proposed based on Teaching-Learning-Based optimization (TLBO. The proposed CAD system is implemented on the histopathology slide images acquired by Aperio XT scanner (scanner A. In TLBO algorithm, the number of false positives (falsely detected nonmitosis candidates as mitosis ones is defined as a cost function and, by minimizing it, many of nonmitosis candidates will be removed. Then some color and texture (textural features such as those derived from cooccurrence and run-length matrices are extracted from the remaining candidates and finally mitotic cells are classified using a specific support vector machine (SVM classifier. The simulation results have proven the claims about the high performance and efficiency of the proposed CAD system.

  9. LOCAL BUDGET PROCESS AND LOCAL ECONOMIC DEVELOPMENT

    OpenAIRE

    Cristinel Ichim

    2009-01-01

    The local budget process is in close connection with the economic development of the territorial-administrative units, firstly because the former implies the collection of budget revenues based on which local authorities provide public services to citizens and ensure the provision of such services by third parties. Also, through the implementation of expenditure, local government creates and maintains a favorable climate for local citizens and local traders for them to thrive and stimulates t...

  10. Local energy: a basis for local electronegativity and local hardness

    OpenAIRE

    Gal, Tamas

    2011-01-01

    The traditional approach to establishing a local measure of chemical hardness, by defining a local hardness concept through the derivative of the chemical potential with respect to the electron density, has been found to have limited chemical applicability, and has proved to be an unfeasible approach in principle. Here, we propose a new approach via a unique local energy concept. This local energy is shown to emerge from the Hamilton-Jacobi kind of construction of Schrodinger's quantum mechan...

  11. The HSP90 inhibitor NVP-AUY922 radiosensitizes by abrogation of homologous recombination resulting in mitotic entry with unresolved DNA damage.

    Directory of Open Access Journals (Sweden)

    Shane Zaidi

    Full Text Available Heat shock protein 90 (HSP90 is a molecular chaperone responsible for the conformational maintenance of a number of client proteins that play key roles in cell cycle arrest, DNA damage repair and apoptosis following radiation. HSP90 inhibitors exhibit antitumor activity by modulating the stabilisation and activation of HSP90 client proteins. We sought to evaluate NVP-AUY922, the most potent HSP90 inhibitor yet reported, in preclinical radiosensitization studies.NVP-AUY922 potently radiosensitized cells in vitro at low nanomolar concentrations with a concurrent depletion of radioresistance-linked client proteins. Radiosensitization by NVP-AUY922 was verified for the first time in vivo in a human head and neck squamous cell carcinoma xenograft model in athymic mice, as measured by delayed tumor growth and increased surrogate end-point survival (p = <0.0001. NVP-AUY922 was shown to ubiquitously inhibit resolution of dsDNA damage repair correlating to delayed Rad51 foci formation in all cell lines tested. Additionally, NVP-AUY922 induced a stalled mitotic phenotype, in a cell line-dependent manner, in HeLa and HN5 cell lines irrespective of radiation exposure. Cell cycle analysis indicated that NVP-AUY922 induced aberrant mitotic entry in all cell lines tested in the presence of radiation-induced DNA damage due to ubiquitous CHK1 depletion, but resultant downstream cell cycle effects were cell line dependent.These results identify NVP-AUY922 as the most potent HSP90-mediated radiosensitizer yet reported in vitro, and for the first time validate it in a clinically relevant in vivo model. Mechanistic analysis at clinically achievable concentrations demonstrated that radiosensitization is mediated by the combinatorial inhibition of cell growth and survival pathways, ubiquitous delay in Rad51-mediated homologous recombination and CHK1-mediated G(2/M arrest, but that the contribution of cell cycle perturbation to radiosensitization may be cell line

  12. Mitotic spindle defects and chromosome mis-segregation induced by LDL/cholesterol-implications for Niemann-Pick C1, Alzheimer's disease, and atherosclerosis.

    Directory of Open Access Journals (Sweden)

    Antoneta Granic

    Full Text Available Elevated low-density lipoprotein (LDL-cholesterol is a risk factor for both Alzheimer's disease (AD and Atherosclerosis (CVD, suggesting a common lipid-sensitive step in their pathogenesis. Previous results show that AD and CVD also share a cell cycle defect: chromosome instability and up to 30% aneuploidy-in neurons and other cells in AD and in smooth muscle cells in atherosclerotic plaques in CVD. Indeed, specific degeneration of aneuploid neurons accounts for 90% of neuronal loss in AD brain, indicating that aneuploidy underlies AD neurodegeneration. Cell/mouse models of AD develop similar aneuploidy through amyloid-beta (Aß inhibition of specific microtubule motors and consequent disruption of mitotic spindles. Here we tested the hypothesis that, like upregulated Aß, elevated LDL/cholesterol and altered intracellular cholesterol homeostasis also causes chromosomal instability. Specifically we found that: 1 high dietary cholesterol induces aneuploidy in mice, satisfying the hypothesis' first prediction, 2 Niemann-Pick C1 patients accumulate aneuploid fibroblasts, neurons, and glia, demonstrating a similar aneugenic effect of intracellular cholesterol accumulation in humans 3 oxidized LDL, LDL, and cholesterol, but not high-density lipoprotein (HDL, induce chromosome mis-segregation and aneuploidy in cultured cells, including neuronal precursors, indicating that LDL/cholesterol directly affects the cell cycle, 4 LDL-induced aneuploidy requires the LDL receptor, but not Aß, showing that LDL works differently than Aß, with the same end result, 5 cholesterol treatment disrupts the structure of the mitotic spindle, providing a cell biological mechanism for its aneugenic activity, and 6 ethanol or calcium chelation attenuates lipoprotein-induced chromosome mis-segregation, providing molecular insights into cholesterol's aneugenic mechanism, specifically through its rigidifying effect on the cell membrane, and potentially explaining why ethanol

  13. Microcystin-LR induces mitotic spindle assembly disorders in Vicia faba by protein phosphatase inhibition and not reactive oxygen species induction.

    Science.gov (United States)

    Garda, Tamás; Kónya, Zoltán; Tándor, Ildikó; Beyer, Dániel; Vasas, Gábor; Erdődi, Ferenc; Vereb, György; Papp, Georgina; Riba, Milán; M-Hamvas, Márta; Máthé, Csaba

    2016-07-20

    We aimed to reveal the mechanisms of mitotic spindle anomalies induced by microcystin-LR (MCY-LR), a cyanobacterial toxin in Vicia faba, a well-known model in plant cell and molecular biology. MCY-LR inhibits type 1 and 2A phosphoserine/threonine specific protein phosphatases (PP1 and PP2A) and induces reactive oxygen species (ROS) formation. The cytoskeleton is one of the main targets of the cyanotoxin during cytopathogenesis. Histochemical-immunohistochemical and biochemical methods were used. A significant number of MCY-LR induced spindle alterations are described for the first time. Disrupted, multipolar spindles and missing kinetochore fibers were detected both in metaphase and anaphase cells. Additional polar microtubule (MT) bundles, hyperbundling of spindle MTs, monopolar spindles, C-S- shaped, additional and asymmetric spindles were detected in metaphase, while midplane kinetochore fibers were detected in anaphase cells only. Several spindle anomalies induced mitotic disorders, i.e. they occurred concomitantly with altered sister chromatid separation. Alterations were dependent on the MCY-LR dose and exposure time. Under long-term (2 and mainly 6 days') exposure they were detected in the concentration range of 0.1-20μgmL(-1) MCY-LR that inhibited PP1 and PP2A significantly without significant ROS induction. Elevated peroxidase/catalase activities indicated that MCY-LR treated V. faba plants showed efficient defense against oxidative stress. Thus, although the elevation of ROS is known to induce cytoskeletal aberrations in general, this study shows that long-term protein phosphatase inhibition is the primary cause of MCY-LR induced spindle disorders. PMID:27186862

  14. A mitotic function for the high-mobility group protein HMG20b regulated by its interaction with the BRC repeats of the BRCA2 tumor suppressor.

    Science.gov (United States)

    Lee, M; Daniels, M J; Garnett, M J; Venkitaraman, A R

    2011-07-28

    The inactivation of BRCA2, a suppressor of breast, ovarian and other epithelial cancers, triggers instability in chromosome structure and number, which are thought to arise from defects in DNA recombination and mitotic cell division, respectively. Human BRCA2 controls DNA recombination via eight BRC repeats, evolutionarily conserved motifs of ∼35 residues, that interact directly with the recombinase RAD51. How BRCA2 controls mitotic cell division is debated. Several studies by different groups report that BRCA2 deficiency affects cytokinesis. Moreover, its interaction with HMG20b, a protein of uncertain function containing a promiscuous DNA-binding domain and kinesin-like coiled coils, has been implicated in the G2-M transition. We show here that HMG20b depletion by RNA interference disturbs the completion of cell division, suggesting a novel function for HMG20b. In vitro, HMG20b binds directly to the BRC repeats of BRCA2, and exhibits the highest affinity for BRC5, a motif that binds poorly to RAD51. Conversely, the BRC4 repeat binds strongly to RAD51, but not to HMG20b. In vivo, BRC5 overexpression inhibits the BRCA2-HMG20b interaction, recapitulating defects in the completion of cell division provoked by HMG20b depletion. In contrast, BRC4 inhibits the BRCA2-RAD51 interaction and the assembly of RAD51 at sites of DNA damage, but not the completion of cell division. Our findings suggest that a novel function for HMG20b in cytokinesis is regulated by its interaction with the BRC repeats of BRCA2, and separate this unexpected function for the BRC repeats from their known activity in DNA recombination. We propose that divergent tumor-suppressive pathways regulating chromosome segregation as well as chromosome structure may be governed by the conserved BRC motifs in BRCA2. PMID:21399666

  15. Non-locality

    CERN Document Server

    van Hateren, J H

    2015-01-01

    The fundamental theories of physics are local theories, depending on local interactions of local variables. It is not clear if and how strictly local theories can produce non-local variables that have causal efficacy. Yet, non-local efficacy appears to exist, such as in the form of memory (non-locality through time) and causally effective spatial structures (non-locality through space). Here it is shown, by construction, how such non-locality can be produced from elementary components: non-isolated systems, multiplicative noise, self-replication, and elimination. A theory is derived that explains how non-locality can arise from strictly local interactions.

  16. Golgi localization and dynamics of hyaluronan binding protein 1 (HABP1/p32/C1QBP) during the cell cycle

    Institute of Scientific and Technical Information of China (English)

    Aniruddha SENGUPTA; Bhaswati BANERJEE; Rakesh K. TYAGI; Kasturi DATTA

    2005-01-01

    Hyaluronan binding protein 1 (HABP1) is a negatively charged multifunctional mammalian protein with a unique structural fold. Despite the fact that HABP1 possesses mitochondrial localization signal, it has also been localized to other cellular compartments. Using indirect immunofluorescence, we examined the sub-cellular localization of HABP1 and its dynamics during mitosis. We wanted to determine whether it distributes in any distinctive manner after mitotic nuclear envelope disassembly or is dispersed randomly throughout the cell. Our results reveal the golgi localization of HABP1 and demonstrate its complete dispersion throughout the cell during mitosis. This distinctive distribution pattern of HABP1 during mitosis resembles its ligand hyaluronan, suggesting that in concert with each other the two molecules play critical roles in this dynamic process.

  17. Local Government, local development and citizen participation

    OpenAIRE

    Meldon, Jeanne; Kenny, Michael; Walsh, Jim

    2002-01-01

    Traditionally the local government system had engaged citizens only in limited passive participation. It was no coincidence that the local development structures put in place to counteract the inadequacies in the local government system introduced different and innovative, opportunities for citizen participation. The scope for engagement in the processes of local government and governance has been significantly extended through a number of pilot initiatives as detailed in the preceding sectio...

  18. Local Duality for 2-Dimensional Local Ring

    Indian Academy of Sciences (India)

    Belgacem Draouil

    2008-11-01

    We prove a local duality for some schemes associated to a 2-dimensional complete local ring whose residue field is an -dimensional local field in the sense of Kato–Parshin. Our results generalize the Saito works in the case =0 and are applied to study the Bloch–Ogus complex for such rings in various cases.

  19. Locally Finite Root Supersystems

    OpenAIRE

    YOUSOFZADEH, Malihe

    2013-01-01

    We introduce the notion of locally finite root supersystems as a generalization of both locally finite root systems and generalized root systems. We classify irreducible locally finite root supersystems.

  20. Induction of chromosome instability and stomach cancer by altering the expression pattern of mitotic checkpoint genes in mice exposed to areca-nut

    International Nuclear Information System (INIS)

    There are strong indications for a causal association between areca-nut consumption and cancers. In Meghalaya, India, the variety of areca-nut is used as raw and unprocessed form whose chemical composition and pharmacological actions have been reported. Yet we know little on the initial pathway involved in areca-nut associated carcinogenesis since it is difficult to assess its effects on genetic alterations without interference of other compounding factors. Therefore, present study was undertaken in mice to verify the ability of raw areca-nut (RAN) to induce cancer and to monitor the expression of certain genes involved in carcinogenesis. This study was not intended to isolate any active ingredients from the RAN and to look its action. Three groups of mice (n = 25 in each) were taken and used at different time-points for different experimental analysis. The other three groups of mice (n = 15 in each) were considered for tumor induction studies. In each set, two groups were administered RAN-extract ad libitum in drinking water with or without lime. The expression of certain genes was assessed by conventional RT-PCR and immunoblotting. The mice were given the whole RAN-extract with and without lime in order to mimic the human consumption style of RAN. Histological preparation of stomach tissue revealed that RAN induced stomach cancer. A gradual increase in the frequency of precocious anaphase and aneuploid cells was observed in the bone marrow cells with a greater increment following RAN + lime administeration. Levels of p53, Bax, Securin and p65 in esophageal and stomach cells were elevated during early days of RAN exposure while those of different mitotic checkpoint proteins were downregulated. Apoptotic cell death was detected in non-cancerous stomach cells but not in tumor cells which showed overexpression of Bax and absence of PARP. Present study suggested (a) RAN induces stomach cancer, however, presence of lime promoted higher cell transformation and thereby

  1. Meiotic and Mitotic Phenotypes Conferred by the blm1-1 Mutation in Saccharomyces cerevisiae and MSH4 Suppression of the Bleomycin Hypersusceptibility

    Directory of Open Access Journals (Sweden)

    Carol Wood Moore

    2003-01-01

    Full Text Available Abstract: Oxidative damage can lead to a number of diseases, and can be fatal. The blm1-1 mutation of Saccharomyces cerevisiae confers hypersusceptibility to lethal effects of the oxidative, anticancer and antifungal agent, bleomycin. For the current report, additional defects conferred by the mutation in meiosis and mitosis were investigated. The viability of spores produced during meiosis by homozygous normal BLM1/BLM1, heterozygous BLM1/blm1-1, and homozygous mutant blm1-1/blm1-1 diploid strains was studied and compared. Approximately 88% of the tetrads derived from homozygous blm1-1/blm1-1 mutant diploid cells only produced one or two viable spores. In contrast, just one tetrad among all BLM1/BLM1 and BLM1/blm1-1 tetrads only produced one or two viable spores. Rather, 94% of BLM1/BLM1 tetrads and 100% of BLM1/blm1-1 tetrads produced asci with four or three viable spores. Thus, at least one copy of the BLM1 gene is essential for the production of four viable spores after meiosis. During mitotic growth, mutant blm1-1 strains grew at reduced rates and produced cells with high frequencies of unusual morphologies compared to wild-type strains. These results indicated BLM1 is also essential for normal mitotic growth. We also investigated the suppression by the MSH4 gene, a meiosis-specific MutS homolog, of the bleomycin hypersusceptibility of blm1-1 mutant cells, and the relationship of MSH4 to BLM1. We screened a genomic library, and isolated the MSH4 gene on the basis of its ability to suppress lethal effects of bleomycin in blm1-1 cells. However, genetic mapping studies indicated that BLM1 and MSH4 are not the same gene. The possibility that chromosomal nondisjunction could be the basis for the inability of blm1-1/blm1-1 mutant cells to produce four viable spores after meiosis is discussed.

  2. Mitotic Illegitimate Recombination Is a Mechanism for Novel Changes in High-Molecular-Weight Glutenin Subunits in Wheat-Rye Hybrids

    Science.gov (United States)

    Yuan, Zhongwei; Liu, Dengcai; Zhang, Lianquan; Zhang, Li; Chen, Wenjie; Yan, Zehong; Zheng, Youliang; Zhang, Huaigang; Yen, Yang

    2011-01-01

    Wide hybrids can have novel traits or changed expression of a quantitative trait that their parents do not have. These phenomena have long been noticed, yet the mechanisms are poorly understood. High-molecular-weight glutenin subunits (HMW-GS) are seed storage proteins encoded by Glu-1 genes that only express in endosperm in wheat and its related species. Novel HMW-GS compositions have been observed in their hybrids. This research elucidated the molecular mechanisms by investigating the causative factors of novel HMW-GS changes in wheat-rye hybrids. HMW-GS compositions in the endosperm and their coding sequences in the leaves of F1 and F2 hybrids between wheat landrace Shinchunaga and rye landrace Qinling were investigated. Missing and/or additional novel HMW-GSs were observed in the endosperm of 0.5% of the 2078 F1 and 22% of 36 F2 hybrid seeds. The wildtype Glu-1Ax null allele was found to have 42 types of short repeat sequences of 3-60 bp long that appeared 2 to 100 times. It also has an in-frame stop codon in the central repetitive region. Analyzing cloned allele sequences of HMW-GS coding gene Glu-1 revealed that deletions involving the in-frame stop codon had happened, resulting in novel ∼1.8-kb Glu-1Ax alleles in some F1 and F2 plants. The cloned mutant Glu-1Ax alleles were expressed in Escherichia coli, and the HMW-GSs produced matched the novel HMW-GSs found in the hybrids. The differential changes between the endosperm and the plant of the same hybrids and the data of E. coli expression of the cloned deletion alleles both suggested that mitotic illegitimate recombination between two copies of a short repeat sequence had resulted in the deletions and thus the changed HMW-GS compositions. Our experiments have provided the first direct evidence to show that mitotic illegitimate recombination is a mechanism that produces novel phenotypes in wide hybrids. PMID:21887262

  3. Mitotic illegitimate recombination is a mechanism for novel changes in high-molecular-weight glutenin subunits in wheat-rye hybrids.

    Directory of Open Access Journals (Sweden)

    Zhongwei Yuan

    Full Text Available Wide hybrids can have novel traits or changed expression of a quantitative trait that their parents do not have. These phenomena have long been noticed, yet the mechanisms are poorly understood. High-molecular-weight glutenin subunits (HMW-GS are seed storage proteins encoded by Glu-1 genes that only express in endosperm in wheat and its related species. Novel HMW-GS compositions have been observed in their hybrids. This research elucidated the molecular mechanisms by investigating the causative factors of novel HMW-GS changes in wheat-rye hybrids. HMW-GS compositions in the endosperm and their coding sequences in the leaves of F(1 and F(2 hybrids between wheat landrace Shinchunaga and rye landrace Qinling were investigated. Missing and/or additional novel HMW-GSs were observed in the endosperm of 0.5% of the 2078 F(1 and 22% of 36 F(2 hybrid seeds. The wildtype Glu-1Ax null allele was found to have 42 types of short repeat sequences of 3-60 bp long that appeared 2 to 100 times. It also has an in-frame stop codon in the central repetitive region. Analyzing cloned allele sequences of HMW-GS coding gene Glu-1 revealed that deletions involving the in-frame stop codon had happened, resulting in novel ∼1.8-kb Glu-1Ax alleles in some F(1 and F(2 plants. The cloned mutant Glu-1Ax alleles were expressed in Escherichia coli, and the HMW-GSs produced matched the novel HMW-GSs found in the hybrids. The differential changes between the endosperm and the plant of the same hybrids and the data of E. coli expression of the cloned deletion alleles both suggested that mitotic illegitimate recombination between two copies of a short repeat sequence had resulted in the deletions and thus the changed HMW-GS compositions. Our experiments have provided the first direct evidence to show that mitotic illegitimate recombination is a mechanism that produces novel phenotypes in wide hybrids.

  4. Dose rate, mitotic cycle duration, and sensitivity of cell transitions from G1 → S and G2 → M to protracted gamma radiation in root meristems

    International Nuclear Information System (INIS)

    Experiments were designed to determine the relative radiosensitivity of the cell transition points of G1 → S and G2 → M in root meristems of several plant species. Label and mitotic indices and microspectrophotometry were used to measure the proportions of cells in each mitotic cycle stage in root meristems after protracted gamma radiation. The criterion of radiosensitivity was the dose rate needed to produce a tissue with less than 1 percent cells in S and none in M after 3 days of continuous exposure. The results show that DNA is the primary radiation target in proliferative root meristems and that the cycle duration stipulates the time interval of vulnerability. In each species, nonrandom reproducible cell proportions were established with 2C:4C:8C amounts of nuclear DNA after 3 days of exposure. Roots of Helianthus annuus, Crepis capillaris, and Tradescantia clone 02 had 80 percent cells with a 2C amount of DNA and 20 percent had a 4C amount of DNA. In these species the transition point of G1 → S was more radiosensitive than G2 → M. Roots of Pisum sativum and Triticum aestivum had cell proportions at 2C:4C:8C amounts of DNA in frequencies of 0.10 to 0.20:0.40 to 0.60:0.30 to 0.40. In these two species 0.30 to 0.40 cells underwent radiation-induced endoreduplication that resulted from a rapid inhibition of cell transit from G2 → M and a slower impairment of G1 → S. Cells increased from 2C to 4C and from 4C to 8C amounts of DNA during irradiation. The proportions of nuclei with 2C:4C:8C amounts of DNA were dependent in part upon the relative radiosensitivity of the G1 → S and G2 → M control points. The data show the relative radiosensitivity of the transition points from G1 → S and from G2 → M was species specific and unrelated to the cycle duration and mean nuclear DNA content of the plant species

  5. Resveratrol abrogates the Temozolomide-induced G2 arrest leading to mitotic catastrophe and reinforces the Temozolomide-induced senescence in glioma cells

    International Nuclear Information System (INIS)

    Temozolomide (TMZ) is the most widely used drug to treat glioblastoma (GBM), which is the most common and aggressive primary tumor of the Central Nervous System and one of the hardest challenges in oncotherapy. TMZ is an alkylating agent that induces autophagy, apoptosis and senescence in GBM cells. However, therapy with TMZ increases survival after diagnosis only from 12 to 14.4 months, making the development of combined therapies to treat GBM fundamental. One candidate for GBM therapy is Resveratrol (Rsv), which has additive toxicity with TMZ in several glioma cells in vitro and in vivo. However, the mechanism of Rsv and TMZ additive toxicity, which is the aim of the present work, is not clear, especially concerning cell cycle dynamics and long term effects. Glioma cell lines were treated with Rsv and TMZ, alone or in combinations, and the induction and the role of autophagy, apoptosis, cell cycle dynamics, protein expression and phosphorylation status were measured. We further evaluated the long term senescence induction and clonogenic capacity. As expected, temozolomide caused a G2 cell cycle arrest and extensive DNA damage response. Rsv did not reduced this response, even increasing pATM, pChk2 and gammaH2Ax levels, but abrogated the temozolomide-induced G2 arrest, increasing levels of cyclin B and pRb(S807/811) and reducing levels of pWee1(S642) and pCdk1(Y15). This suggests a cellular state of forced passage through G2 checkpoint despite large DNA damage, a scenario that may produce mitotic catastrophe. Indeed, the proportion of cells with high nuclear irregularity increased from 6 to 26% in 48 h after cotreatment. At a long term, a reduction in clonogenic capacity was observed, accompanied by a large induction of senescence. The presence of Rsv forces cells treated with TMZ through mitosis leading to mitotic catastrophe and senescence, reducing the clonogenic capacity of glioma cells and increasing the chronic effects of temozolomide

  6. Early localization of NPA58, a rat nuclear pore-associated protein, to the reforming nuclear envelope during mitosis

    Indian Academy of Sciences (India)

    Radhika Ganeshan; Nandini Rangaraj; Veena K Parnaik

    2001-03-01

    We have studied the mitotic reassembly of the nuclear envelope, using antibodies to nuclear marker proteins and NPA58 in F-111 rat fibroblast cells. In earlier studies we have proposed that NPA58, a 58 kDa rat nuclear protein, is involved in nuclear protein import. In this report, NPA58 is shown to be localized on the cytoplasmic face of the envelope in interphase cells, in close association with nuclear pores. In mitotic cells NPA58 is dispersed in the cytoplasm till anaphase. The targeting of NPA58 to the reforming nuclear envelope in early telophase coincides with the recruitment of a well-characterized class of nuclear pore proteins recognized by the antibody mAb 414, and occurs prior to the incorporation of lamin B1 into the envelope. Significant protein import activity is detectable only after localization of NPA58 in the newly-formed envelope. The early targeting of NPA58 is consistent with its proposed role in nuclear transport.

  7. AtPPR2, an Arabidopsis pentatricopeptide repeat protein, binds to plastid 23S rRNA and plays an important role in the first mitotic division during gametogenesis and in cell proliferation during embryogenesis

    OpenAIRE

    Lu, Yuqing; Li, Cong; Wang, Hai; Chen, Hao; Berg, Howard; Xia, Yiji

    2011-01-01

    Pentatricopeptide repeat (PPR) proteins are mainly involved in regulating post-transcriptional processes in mitochondria and plastids, including chloroplasts. Mutations in the Arabidopsis PPR2 gene have previously been found to cause defects in seed development and reduced transmission through male and female gametophytes. However, the exact function of AtPPR2 has not been defined. We found that a loss-of-function mutation of AtPPR2 leads to arrest of the first mitotic division during both ma...

  8. Loss of CCDC6, the first identified RET partner gene, affects pH2AX S139 levels and accelerates mitotic entry upon DNA damage.

    Directory of Open Access Journals (Sweden)

    Francesco Merolla

    Full Text Available CCDC6 was originally identified in chimeric genes caused by chromosomal translocation involving the RET proto-oncogene in some thryoid tumors mostly upon ionizing radiation exposure. Recognised as a pro-apoptotic phosphoprotein that negatively regulates CREB1-dependent transcription, CCDC6 is an ATM substrate that is responsive to genotoxic stress. Here we report that following genotoxic stress, loss or inactivation of CCDC6 in cancers that carry the CCDC6 fusion, accelerates the dephosphorylation of pH2AX S139, resulting in defective G2 arrest and premature mitotic entry. Moreover, we show that CCDC6 depleted cells appear to repair DNA damaged in a shorter time compared to controls, based on reporter assays in cells. High-troughput proteomic screening predicted the interaction between the CCDC6 gene product and the catalytic subunit of Serin-Threonin Protein Phosphatase 4 (PP4c recently identified as the evolutionarily conserved pH2AX S139 phosphatase that is activated upon DNA Damage. We describe the interaction between CCDC6 and PP4c and we report the modulation of PP4c enzymatic activity in CCDC6 depleted cells. We discuss the functional significance of CCDC6-PP4c interactions and hypothesize that CCDC6 may act in the DNA Damage Response by negatively modulating PP4c activity. Overall, our data suggest that in primary tumours the loss of CCDC6 function could influence genome stability and thereby contribute to carcinogenesis.

  9. 3D-QSAR Studies of Dihydropyrazole and Dihydropyrrole Derivatives as Inhibitors of Human Mitotic Kinesin Eg5 Based on Molecular Docking

    Directory of Open Access Journals (Sweden)

    Wenjuan Yang

    2012-02-01

    Full Text Available Human mitotic kinesin Eg5 plays an essential role in mitoses and is an interesting drug target against cancer. To find the correlation between Eg5 and its inhibitors, structure-based 3D-quantitative structure–activity relationship (QSAR studies were performed on a series of dihydropyrazole and dihydropyrrole derivatives using comparative molecular field analysis (CoMFA and comparative molecular similarity indices analysis (CoMSIA methods. Based on the LigandFit docking results, predictive 3D-QSAR models were established, with cross-validated coefficient values (q2 up to 0.798 for CoMFA and 0.848 for CoMSIA, respectively. Furthermore, the CoMFA and CoMSIA models were mapped back to the binding sites of Eg5, which could provide a better understanding of vital interactions between the inhibitors and the kinase. Ligands binding in hydrophobic part of the inhibitor-binding pocket were found to be crucial for potent ligand binding and kinases selectivity. The analyses may be used to design more potent EG5 inhibitors and predict their activities prior to synthesis.

  10. Structure–Biological Function Relationship Extended to Mitotic Arrest-Deficient 2-Like Protein Mad2 Native and Mutants-New Opportunity for Genetic Disorder Control

    Directory of Open Access Journals (Sweden)

    Speranta Avram

    2014-11-01

    Full Text Available Overexpression of mitotic arrest-deficient proteins Mad1 and Mad2, two components of spindle assembly checkpoint, is a risk factor for chromosomal instability (CIN and a trigger of many genetic disorders. Mad2 transition from inactive open (O-Mad2 to active closed (C-Mad2 conformations or Mad2 binding to specific partners (cell-division cycle protein 20 (Cdc20 or Mad1 were targets of previous pharmacogenomics studies. Here, Mad2 binding to Cdc20 and the interconversion rate from open to closed Mad2 were predicted and the molecular features with a critical contribution to these processes were determined by extending the quantitative structure-activity relationship (QSAR method to large-size proteins such as Mad2. QSAR models were built based on available published data on 23 Mad2 mutants inducing CIN-related functional changes. The most relevant descriptors identified for predicting Mad2 native and mutants action mechanism and their involvement in genetic disorders are the steric (van der Waals area and solvent accessible area and their subdivided and energetic van der Waals energy descriptors. The reliability of our QSAR models is indicated by significant values of statistical coefficients: Cross-validated correlation q2 (0.53–0.65 and fitted correlation r2 (0.82–0.90. Moreover, based on established QSAR equations, we rationally design and analyze nine de novo Mad2 mutants as possible promoters of CIN.

  11. Mitotic recombination and compound-heterozygous mutations are predominant NF1-inactivating mechanisms in children with juvenile myelomonocytic leukemia and neurofibromatosis type 1.

    Science.gov (United States)

    Steinemann, Doris; Arning, Larissa; Praulich, Inka; Stuhrmann, Manfred; Hasle, Henrik; Stary, Jan; Schlegelberger, Brigitte; Niemeyer, Charlotte M; Flotho, Christian

    2010-02-01

    Children with neurofibromatosis type 1 (NF-1), being constitutionally deficient for one allele of the NF1 gene, are at greatly increased risk of juvenile myelomonocytic leukemia (JMML). NF1 is a negative regulator of RAS pathway activity, which has a central role in JMML. To further clarify the role of biallelic NF1 gene inactivation in the pathogenesis of JMML, we investigated the somatic NF1 lesion in 10 samples from children with JMML/NF-1. We report that two-thirds of somatic events involved loss of heterozygosity (LOH) at the NF1 locus, predominantly caused by segmental uniparental disomy of large parts of chromosome arm 17q. One-third of leukemias showed compound-heterozygous NF1-inactivating mutations. A minority of cases exhibited somatic interstitial deletions. The findings reinforce the emerging role of somatic mitotic recombination as a leukemogenic mechanism. In addition, they support the concept that biallelic NF1 inactivation in hematopoietic progenitor cells is required for transformation to JMML in children with NF-1. PMID:20015894

  12. I element distribution in mitotic heterochromatin of Drosophila melanogaster reactive strains: identification of a specific site which is correlated with the reactivity levels.

    Science.gov (United States)

    Dimitri, P; Bucheton, A

    2005-01-01

    The I factor is a Drosophila melanogaster LINE-like element that efficiently transposes in the genetic system of I-R hybrid dysgenesis. It has been suggested that some of the I-related sequences located in the heterochromatin of D. melanogaster are involved in the regulation of I factor activity. In this work we have performed fluorescent in situ hybridization (FISH) mapping of I element sequences in mitotic heterochromatin of nine differentially reactive D. melanogaster strains. The results of our analysis showed that a single hybridization site mapping to region h28 of the distal heterochromatin of the X chromosome is present in three strains with low or intermediate levels of reactivity, while it is undetectable in six highly reactive strains. Together, these observations suggest a negative correlation between I sequences located at h28 and the level of reactivity. To this regard, it is intriguing that flamenco and COM, two loci that regulate the activity of D. melanogaster endogenous retroviruses also map to the distal heterochromatin of the X chromosome. Our data represent the first experimental evidence in favour of a silencing effect exerted by naturally occurring I element sequences located in pericentromeric heterochromatin. PMID:16093668

  13. The SFP1 gene product of Saccharomyces cerevisiae regulates G2/M transitions during the mitotic cell cycle and DNA-damage response

    International Nuclear Information System (INIS)

    In eukaryotic cells, checkpoint pathways arrest cell-cycle progression if a particular event has failed to complete appropriately or if an important intracellular structure is defective or damaged. Saccharomyces cerevisiae strains that lack the SFP1 gene fail to arrest at the G2 DNA-damage checkpoint in response to genomic injury, but maintain their ability to arrest at the replication and spindle-assembly checkpoints. sfp1D mutants are characterized by a premature entrance into mitosis during a normal (undamaged) cell cycle, while strains that overexpress Sfp1p exhibit delays in G2. Sfp1p therefore acts as a repressor of the G2/M transition, both in the normal cell cycle and in the G2 checkpoint pathway. Sfp1 is a nuclear protein with two Cys2His2 zinc-finger domains commonly found in transcription factors. We propose that Sfp1p regulates the expression of gene products involved in the G2/M transition during the mitotic cell cycle and the DNA-damage response. In support of this model, overexpression of Sfp1p induces the expression of the PDS1 gene, which is known to encode a protein that regulates the G2 checkpoint. (author)

  14. Identification of a BET family bromodomain/casein kinase II/TAF-containing complex as a regulator of mitotic condensin function.

    Science.gov (United States)

    Kim, Hyun-Soo; Mukhopadhyay, Rituparna; Rothbart, Scott B; Silva, Andrea C; Vanoosthuyse, Vincent; Radovani, Ernest; Kislinger, Thomas; Roguev, Assen; Ryan, Colm J; Xu, Jiewei; Jahari, Harlizawati; Hardwick, Kevin G; Greenblatt, Jack F; Krogan, Nevan J; Fillingham, Jeffrey S; Strahl, Brian D; Bouhassira, Eric E; Edelmann, Winfried; Keogh, Michael-Christopher

    2014-03-13

    Condensin is a central regulator of mitotic genome structure with mutants showing poorly condensed chromosomes and profound segregation defects. Here, we identify NCT, a complex comprising the Nrc1 BET-family tandem bromodomain protein (SPAC631.02), casein kinase II (CKII), and several TAFs, as a regulator of condensin function. We show that NCT and condensin bind similar genomic regions but only briefly colocalize during the periods of chromosome condensation and decondensation. This pattern of NCT binding at the core centromere, the region of maximal condensin enrichment, tracks the abundance of acetylated histone H4, as regulated by the Hat1-Mis16 acetyltransferase complex and recognized by the first Nrc1 bromodomain. Strikingly, mutants in NCT or Hat1-Mis16 restore the formation of segregation-competent chromosomes in cells containing defective condensin. These results are consistent with a model where NCT targets CKII to chromatin in a cell-cycle-directed manner in order to modulate the activity of condensin during chromosome condensation and decondensation. PMID:24565511

  15. Identification of a BET Family Bromodomain/Casein Kinase II/TAF-Containing Complex as a Regulator of Mitotic Condensin Function

    Directory of Open Access Journals (Sweden)

    Hyun-Soo Kim

    2014-03-01

    Full Text Available Condensin is a central regulator of mitotic genome structure with mutants showing poorly condensed chromosomes and profound segregation defects. Here, we identify NCT, a complex comprising the Nrc1 BET-family tandem bromodomain protein (SPAC631.02, casein kinase II (CKII, and several TAFs, as a regulator of condensin function. We show that NCT and condensin bind similar genomic regions but only briefly colocalize during the periods of chromosome condensation and decondensation. This pattern of NCT binding at the core centromere, the region of maximal condensin enrichment, tracks the abundance of acetylated histone H4, as regulated by the Hat1-Mis16 acetyltransferase complex and recognized by the first Nrc1 bromodomain. Strikingly, mutants in NCT or Hat1-Mis16 restore the formation of segregation-competent chromosomes in cells containing defective condensin. These results are consistent with a model where NCT targets CKII to chromatin in a cell-cycle-directed manner in order to modulate the activity of condensin during chromosome condensation and decondensation.

  16. Locally orderless registration

    DEFF Research Database (Denmark)

    Darkner, Sune; Sporring, Jon

    2013-01-01

    approach is based on local intensity histograms and built upon the technique of Locally Orderless Images. Histograms by Locally Orderless Images are well posed and offer explicit control over the 3 inherent and unavoidable scales: the spatial resolution, intensity levels, and spatial extent of local...

  17. Mitotic phosphorylation of Bloom helicase at Thr182 is required for its proteasomal degradation and maintenance of chromosomal stability.

    Science.gov (United States)

    Kharat, S S; Tripathi, V; Damodaran, A P; Priyadarshini, R; Chandra, S; Tikoo, S; Nandhakumar, R; Srivastava, V; Priya, S; Hussain, M; Kaur, S; Fishman, J B; Sengupta, S

    2016-02-25

    Mutations in Bloom helicase (BLM) lead to Bloom Syndrome (BS). BS is characterized by multiple clinical manifestations including predisposition to a wide spectrum of cancers. Studies have revealed the mechanism of BLM recruitment after stalled replication and its role during the repair of DNA damage. We now provide evidence that BLM undergoes K48-linked ubiquitylation and subsequent degradation during mitosis due to the E3 ligase, Fbw7α. Fbw7α carries out its function after GSK3β- and CDK2/cyclin A2-dependent phosphorylation events on Thr171 and Ser175 of BLM which lies within a well-defined phosphodegron, a sequence which is conserved in all primates. Phosphorylation on BLM Thr171 and Ser175 depends on prior phosphorylation at Thr182 by Chk1/Chk2. Thr182 phosphorylation not only controls BLM ubiquitylation and degradation during mitosis but is also a determinant for its localization on the ultrafine bridges. Consequently lack of Thr182 phosphorylation leads to multiple manifestations of chromosomal instability including increased levels of DNA damage, lagging chromatin, micronuclei formation, breaks and quadriradials. Hence Thr182 phosphorylation on BLM has two functions-it regulates BLM turnover during mitosis and also helps to maintain the chromosomal stability. PMID:26028025

  18. Defining local food

    DEFF Research Database (Denmark)

    Eriksen, Safania Normann

    2013-01-01

    Despite evolving local food research, there is no consistent definition of “local food.” Various understandings are utilized, which have resulted in a diverse landscape of meaning. The main purpose of this paper is to examine how researchers within the local food systems literature define local...... food, and how these definitions can be used as a starting point to identify a new taxonomy of local food based on three domains of proximity....

  19. Local perturbations perturb—exponentially–locally

    International Nuclear Information System (INIS)

    We elaborate on the principle that for gapped quantum spin systems with local interaction, “local perturbations [in the Hamiltonian] perturb locally [the groundstate].” This principle was established by Bachmann et al. [Commun. Math. Phys. 309, 835–871 (2012)], relying on the “spectral flow technique” or “quasi-adiabatic continuation” [M. B. Hastings, Phys. Rev. B 69, 104431 (2004)] to obtain locality estimates with sub-exponential decay in the distance to the spatial support of the perturbation. We use ideas of Hamza et al. [J. Math. Phys. 50, 095213 (2009)] to obtain similarly a transformation between gapped eigenvectors and their perturbations that is local with exponential decay. This allows to improve locality bounds on the effect of perturbations on the low lying states in certain gapped models with a unique “bulk ground state” or “topological quantum order.” We also give some estimate on the exponential decay of correlations in models with impurities where some relevant correlations decay faster than one would naively infer from the global gap of the system, as one also expects in disordered systems with a localized groundstate

  20. New model. Local financing for local energy

    International Nuclear Information System (INIS)

    While evoking the case of the VMH Energies company in the Poitou-Charentes region, and indicating the difference between France and Germany in terms of wind and photovoltaic energy production potential, of number of existing local companies, and of citizen-based funding, this article shows that renewable energies could put the energy production financing in France into question again, with a more important participation of local communities and of their inhabitants. The author describes how the law on energy transition makes this possible, notably with the strengthening of citizen participation. The author evokes some French local experiments and the case of Germany where this participation is already very much developed

  1. Identification of Local Melon (Cucumis melo L. var. Bartek Based on Chromosomal Characters

    Directory of Open Access Journals (Sweden)

    BUDI SETIADI DARYONO

    2011-12-01

    Full Text Available Bartek is one of local melon varieties mainly cultivated in Pemalang, Central Java. Bartek has three variations of fruits; Long-Green, Ellips-Green, and Yellow. Chromosome characterization of the Bartek was investigated to determine the genetic variation. The main purpose of this research was to determine the genetic characters of Bartek including chromosome number, mitosis, cell cycle, and karyotype. Squash method was used for chromosome preparation. The results showed that all of Bartek observed in this study have similar diploid (2n chromosome number = 24. According to the total number of chromosome, Bartek is closer to melon than cucumber. The mitotic analysis exhibited that the Bartek has similar karyotype formula, 2n = 2x = 24m. Based on the R value of the three kinds of Bartek (R < 0.27, it indicated that three kinds of Bartek were considered to be originated from similar species and one of melon varieties (Cucumis melo L. var. Bartek.

  2. Localized Prostate Cancer

    Science.gov (United States)

    ... a decision aid for men with clinically localized prostate cancer (available at http://effectivehealthcare.ahrq.gov/prostate_da) ... A Decision Aid for Men With Clinically Localized Prostate Cancer Page 1 of 24 Introduction Men with clinically ...

  3. Local Debt Scare

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    China steps up a clampdown on the financing vehicles of local governments control over local governments’ financing vehicles will be tightened to fend off what some economists warn could turn into a mountain

  4. Time to Go Local!

    Science.gov (United States)

    ... Bar Home Current Issue Past Issues Time to Go Local! Past Issues / Winter 2007 Table of Contents ... MedlinePlus.gov health topic pages, you will find "Go Local" links that take you to information about ...

  5. Increased therapeutic potential of an experimental anti-mitotic inhibitor SB715992 by genistein in PC-3 human prostate cancer cell line

    International Nuclear Information System (INIS)

    Kinesin spindle proteins (KSP) are motor proteins that play an essential role in mitotic spindle formation. HsEg5, a KSP, is responsible for the formation of the bipolar spindle, which is critical for proper cell division during mitosis. The function of HsEg5 provides a novel target for the manipulation of the cell cycle and the induction of apoptosis. SB715992, an experimental KSP inhibitor, has been shown to perturb bipolar spindle formation, thus making it an excellent candidate for anti-cancer agent. Our major objective was a) to investigate the cell growth inhibitory effects of SB715992 on PC-3 human prostate cancer cell line, b) to investigate whether the growth inhibitory effects of SB715992 could be enhanced when combined with genistein, a naturally occurring isoflavone and, c) to determine gene expression profile to establish molecular mechanism of action of SB715992. PC-3 cells were treated with varying concentration of SB715992, 30 μM of genistein, and SB715992 plus 30 μM of genistein. After treatments, PC-3 cells were assayed for cell proliferation, induction of apoptosis, and alteration in gene and protein expression using cell inhibition assay, apoptosis assay, microarray analysis, real-time RT-PCR, and Western Blot analysis. SB715992 inhibited cell proliferation and induced apoptosis in PC-3 cells. SB715992 was found to regulate the expression of genes related to the control of cell proliferation, cell cycle, cell signaling pathways, and apoptosis. In addition, our results showed that combination treatment with SB715992 and genistein caused significantly greater cell growth inhibition and induction of apoptosis compared to the effects of either agent alone. Our results clearly show that SB715992 is a potent anti-tumor agent whose therapeutic effects could be enhanced by genistein. Hence, we believe that SB715992 could be a novel agent for the treatment of prostate cancer with greater success when combined with a non-toxic natural agent like

  6. A novel microtubule inhibitor, MT3-037, causes cancer cell apoptosis by inducing mitotic arrest and interfering with microtubule dynamics

    Science.gov (United States)

    Chang, Ling-Chu; Yu, Yung-Luen; Hsieh, Min-Tsang; Wang, Sheng-Hung; Chou, Ruey-Hwang; Huang, Wei-Chien; Lin, Hui-Yi; Hung, Hsin-Yi; Huang, Li-Jiau; Kuo, Sheng-Chu

    2016-01-01

    We investigated the anticancer potential of a new synthetic compound, 7-(3-fluorophenyl)-4-methylpyrido-[2,3-d]pyrimidin-5(8H)-one (MT3-037). We found that MT3-037 effectively decreased the cancer cell viability by inducing apoptosis. MT3-037 treatments led to cell cycle arrest at M phase, with a marked increase in both expression of cyclin B1 and cyclin-dependent kinase 1 (CDK1) as well as in CDK1 kinase activity. Key proteins that regulate mitotic spindle dynamics, including survivin, Aurora A/B kinases, and polo-like kinase 1 (PLK1), were activated in MT3-037-treated cells. MT3-037-induced apoptosis was accompanied by activation of a pro-apoptotic factor, FADD, and the inactivation of apoptosis inhibitors, Bcl-2 and Bcl-xL, resulting in the cleavage/activation of caspases. The activation of c-Jun N-terminal kinase (JNK) was associated with MT3-037-induced CDK1 and Aurora A/B activation and apoptosis. Immunofluorescence staining of tubulin indicated that MT3-037 altered tubulin networks in cancer cells. Moreover, an in vitro tubulin polymerization assay revealed that MT3-037 inhibited the tubulin polymerization by direct binding to tubulin. Molecular docking studies and binding site completion assays revealed that MT3-037 binds to the colchicine-binding site. Furthermore, MT3-037 significantly inhibited the tumor growth in both MDAMB-468 and Erlotinib-resistant MDA-MB-468 xenograft mouse models. In addition, MT3-037 inhibited the angiogenesis and disrupted the tube formation by human endothelial cells. Our study demonstrates that MT3-037 is a potential tubulin-disrupting agent for antitumor therapy.

  7. Anti-mitotic activity towards sea urchin embryos in extracts from the marine haptophycean Phaeocystis pouchetii (Hariot) Lagerheim collected along the coast of northern Norway.

    Science.gov (United States)

    Hansen, Espen; Eilertsen, Hans Chr; Ernstsen, Arild; Genevière, Anne-Marie

    2003-06-01

    The marine bloom-forming alga Phaeocystis pouchetii is suspected to produce some toxic compound responsible for reduced growth, fecundity and survival of other marine organisms. Sea urchin early development was used as a model to investigate the degree and nature of toxicity. Colonial cells of P. pouchetii were collected during its spring-bloom along the coast of northern Norway and maintained in culture for a short period of time in order to evaluate the concentration of toxic compounds present inside the cells or excreted to the surrounding seawater medium. Cells were harvested by filtration and toxins were extracted separately from the collected cells and the filtrate using organic solvents. We found that extracts from the filtered seawater at a concentration corresponding to 9.0 x 10(5) cells ml(-1) completely blocked cell divisions in embryos of the sea urchin Sphaerechinus granularis, whereas extracts from intact algal cells were only mildly cytotoxic. When the extracts from seawater culture medium were purified by RP-HPLC, cytotoxic activity towards S. granularis embryos was recovered in three consecutive fractions. Moreover, unfertilised eggs incubated in the active HPLC fractions became unproductive, whereas incubation of sperm gave a reduced fertilisation rate. This anti-proliferative effect was further characterized by immunofluorescence staining of sea urchin embryos. DNA labelling revealed that incubating sea urchin embryos in the purified algal extracts inhibited both pronuclei migration and fusion. Incorporation and detection of the DNA-base analogue 5-bromo-2-deoxyuridine showed that DNA-replication was blocked. Furthermore, staining of alpha-tubulin subunits demonstrated that embryonic tubulin organisation was altered. We conclude that P. pouchetii produce some anti-mitotic compound, and that senescent colonial cells to a great extent excrete this compound to their surroundings. PMID:12782080

  8. Local network assessment

    Science.gov (United States)

    Glen, D. V.

    1985-04-01

    Local networks, related standards activities of the Institute of Electrical and Electronics Engineers the American National Standards Institute and other elements are presented. These elements include: (1) technology choices such as topology, transmission media, and access protocols; (2) descriptions of standards for the 802 local area networks (LAN's); high speed local networks (HSLN's) and military specification local networks; and (3) intra- and internetworking using bridges and gateways with protocols Interconnection (OSI) reference model. The convergence of LAN/PBX technology is also described.

  9. Application of a new dual localization-affinity purification tag reveals novel aspects of protein kinase biology in Aspergillus nidulans.

    Directory of Open Access Journals (Sweden)

    Colin P De Souza

    Full Text Available Filamentous fungi occupy critical environmental niches and have numerous beneficial industrial applications but devastating effects as pathogens and agents of food spoilage. As regulators of essentially all biological processes protein kinases have been intensively studied but how they regulate the often unique biology of filamentous fungi is not completely understood. Significant understanding of filamentous fungal biology has come from the study of the model organism Aspergillus nidulans using a combination of molecular genetics, biochemistry, cell biology and genomic approaches. Here we describe dual localization-affinity purification (DLAP tags enabling endogenous N or C-terminal protein tagging for localization and biochemical studies in A. nidulans. To establish DLAP tag utility we endogenously tagged 17 protein kinases for analysis by live cell imaging and affinity purification. Proteomic analysis of purifications by mass spectrometry confirmed association of the CotA and NimXCdk1 kinases with known binding partners and verified a predicted interaction of the SldABub1/R1 spindle assembly checkpoint kinase with SldBBub3. We demonstrate that the single TOR kinase of A. nidulans locates to vacuoles and vesicles, suggesting that the function of endomembranes as major TOR cellular hubs is conserved in filamentous fungi. Comparative analysis revealed 7 kinases with mitotic specific locations including An-Cdc7 which unexpectedly located to mitotic spindle pole bodies (SPBs, the first such localization described for this family of DNA replication kinases. We show that the SepH septation kinase locates to SPBs specifically in the basal region of apical cells in a biphasic manner during mitosis and again during septation. This results in gradients of SepH between G1 SPBs which shift along hyphae as each septum forms. We propose that SepH regulates the septation initiation network (SIN specifically at SPBs in the basal region of G1 cells and that

  10. Sound Localization by Cetaceans

    OpenAIRE

    Branstetter, Brian K.; Mercado, Eduardo III

    2006-01-01

    Cetaceans (whales and dolphins) use acoustic cues to determine the locations and identities of environmental stimuli within their underwater habitats. Dolphins evolved unique auditory systems for spatially differentiating ultrasonic signals, whereas the larger baleen whales appear to have evolved different mechanisms for localizing lower frequency sound sources. Many of the cues that terrestrial mammals use to localize sounds in air are less well suited for localizing sounds underwater. Never...

  11. Locally finite profinite rings

    OpenAIRE

    Dobrowolski, Jan; Krupiński, Krzysztof

    2013-01-01

    We investigate the structure of locally finite profinite rings. We classify (Jacobson-) semisimple locally finite profinite rings as products of complete matrix rings of bounded cardinality over finite fields, and we prove that the Jacobson radical of any locally finite profinite ring is nil of finite nilexponent. Our results apply to the context of small compact $G$-rings, where we also obtain a description of possible actions of $G$ on the underlying ring.

  12. Local Social Networks

    OpenAIRE

    Sapuppo, Antonio; Sørensen, Lene Tolstrup

    2011-01-01

    Online social networks have become essential for many users in their daily communication. Through a combination of the online social networks with opportunistic networks, a new concept arises: Local Social Networks. The target of local social networks is to promote social networking benefits in physical environment in order to leverage personal affinities in the users' surroundings. The purpose of this paper is to present and discuss the concept of local social networks as a new social commun...

  13. Change of mitotic cycle and DNA repair in embryonic cells of rat, immortalized by E1 A oncogene and transformated by E1 A and c-Ha-Ras oncogenes under ionizing radiation action

    International Nuclear Information System (INIS)

    Comparison investigation into the repair of mitotic cycle and the reunion of DN single- and double-strand breaks in gamma-ray irradiated initial E1 A oncogene immortalized and E1 A and c-Ha-Ras oncogene transformed (mutant form) lines of rat embryonic fibroblasts was carried out. Possible involvement of Ras gene product in DNA repair speed governing and absence of tumor suppression function of p 53 protein in the embryonic and E1 A oncogene immortalized cells of rat fibroblast, as well as, presence of the mentioned function of p 53 protein in E1 A and c-Ha-Ras oncogene transformed cells were studied

  14. Local Social Networks

    DEFF Research Database (Denmark)

    Sapuppo, Antonio; Sørensen, Lene Tolstrup

    2011-01-01

    Online social networks have become essential for many users in their daily communication. Through a combination of the online social networks with opportunistic networks, a new concept arises: Local Social Networks. The target of local social networks is to promote social networking benefits...... in physical environment in order to leverage personal affinities in the users' surroundings. The purpose of this paper is to present and discuss the concept of local social networks as a new social communication system. Particularly, the preliminary architecture and the prototype of local social networks...

  15. Perturbation of cytokinin and ethylene-signalling pathways explain the strong rooting phenotype exhibited by Arabidopsis expressing the Schizosaccharomyces pombe mitotic inducer, cdc25

    Directory of Open Access Journals (Sweden)

    Spadafora Natasha D

    2012-03-01

    Full Text Available Abstract Background Entry into mitosis is regulated by cyclin dependent kinases that in turn are phosphoregulated. In most eukaryotes, phosphoregulation is through WEE1 kinase and CDC25 phosphatase. In higher plants a homologous CDC25 gene is unconfirmed and hence the mitotic inducer Schizosaccharomyces pombe (Sp cdc25 has been used as a tool in transgenic plants to probe cell cycle function. Expression of Spcdc25 in tobacco BY-2 cells accelerates entry into mitosis and depletes cytokinins; in whole plants it stimulates lateral root production. Here we show, for the first time, that alterations to cytokinin and ethylene signaling explain the rooting phenotype elicited by Spcdc25 expression in Arabidopsis. Results Expressing Spcdc25 in Arabidopsis results in increased formation of lateral and adventitious roots, a reduction of primary root width and more isodiametric cells in the root apical meristem (RAM compared with wild type. Furthermore it stimulates root morphogenesis from hypocotyls when cultured on two way grids of increasing auxin and cytokinin concentrations. Microarray analysis of seedling roots expressing Spcdc25 reveals that expression of 167 genes is changed by > 2-fold. As well as genes related to stress responses and defence, these include 19 genes related to transcriptional regulation and signaling. Amongst these was the up-regulation of genes associated with ethylene synthesis and signaling. Seedlings expressing Spcdc25 produced 2-fold more ethylene than WT and exhibited a significant reduction in hypocotyl length both in darkness or when exposed to 10 ppm ethylene. Furthermore in Spcdc25 expressing plants, the cytokinin receptor AHK3 was down-regulated, and endogenous levels of iPA were reduced whereas endogeous IAA concentrations in the roots increased. Conclusions We suggest that the reduction in root width and change to a more isodiametric cell phenotype in the RAM in Spcdc25 expressing plants is a response to ethylene over

  16. Variation of spontaneous and induced mitotic recombination in different Drosophila populations: a pilot study on the effects of polyaromatic hydrocarbons in six newly constructed tester strains.

    Science.gov (United States)

    Vogel, E W; Nivard, M J; Zijlstra, J A

    1991-01-01

    system is predominantly the result of mitotic recombination. PMID:1944343

  17. Cell biology of mitotic recombination

    DEFF Research Database (Denmark)

    Lisby, Michael; Rothstein, Rodney

    2015-01-01

    Homologous recombination provides high-fidelity DNA repair throughout all domains of life. Live cell fluorescence microscopy offers the opportunity to image individual recombination events in real time providing insight into the in vivo biochemistry of the involved proteins and DNA molecules as w...

  18. Local resilience of graphs

    OpenAIRE

    Sudakov, Benny; Vu, Van

    2007-01-01

    In this paper, we initiate a systematic study of graph resilience. The (local) resilience of a graph G with respect to a property P measures how much one has to change G (locally) in order to destroy P. Estimating the resilience leads to many new and challenging problems. Here we focus on random and pseudo-random graphs and prove several sharp results.

  19. Efficiency of local search

    NARCIS (Netherlands)

    Brueggemann, Tobias

    2006-01-01

    Local search heuristics are an important class of algorithms for obtaining good solutions for hard combinatorial optimization problems. Important issues for these heuristics are solution quality and the time needed to obtain a good solution. Roughly speaking, a local search heuristic starts with an

  20. Locally Orderless Registration Code

    DEFF Research Database (Denmark)

    2012-01-01

    This is code for the TPAMI paper "Locally Orderless Registration". The code requires intel threadding building blocks installed and is provided for 64 bit on mac, linux and windows.......This is code for the TPAMI paper "Locally Orderless Registration". The code requires intel threadding building blocks installed and is provided for 64 bit on mac, linux and windows....

  1. Local Exhaust Ventilation

    DEFF Research Database (Denmark)

    Madsen, Ulla; Breum, N. O.; Nielsen, Peter V.

    Capture efficiency of a local exhaust system, e.g. a kitchen hood, should include only contaminants being direct captured. In this study basic concepts of local exhaust capture efficiency are given, based on the idea of a control box. A validated numerical model is used for estimation of the...

  2. Understanding localized corrosion

    Directory of Open Access Journals (Sweden)

    G.S. Frankel

    2008-10-01

    Full Text Available The breakdown of a protective passive film leading to accelerated dissolution at localized sites is an important practical issue and a vexing scientific problem. The small dimensions, short timescale, and dynamic interplay between a heterogeneous surface and changing potential and solution concentration gradients complicate the development of a complete understanding of the phenomena. This review touches on some of the recent developments in the field, including scanning tunneling microscopy imaging of the earliest stages of pitting which supports a new model explaining the localization of attack, pitting in thin aqueous layers relevant to atmospheric corrosion, the factors controlling crevice corrosion, and predictive modeling of localized corrosion.

  3. Alegerile locale 2000

    Directory of Open Access Journals (Sweden)

    Sorin Dan ȘANDOR

    2000-10-01

    Full Text Available The article presents the Romanian local elections held in June 2000. While local elections are often used by Romanian parties as a preliminary test of future general election, the electoral system used made them more then that. We have in here an incumbency effect that made mayors’ re-election possible unregarding to their political affiliation, we have a different electoral campaign, with different vehicles of propaganda, and possible strange local coalitions among rival parties due to the fact that the presidents of county councils and deputy mayors are elected by the councilors.

  4. Local variances in biomonitoring

    International Nuclear Information System (INIS)

    The present study was undertaken to explore possibilities to judge survey quality on basis of a limited and restricted number of a-priori observations. Here, quality is defined as the ratio between survey and local variance (signal-to-noise ratio). The results indicate that the presented surveys do not permit such judgement; the discussion also suggests that the 5-fold local sampling strategies do not merit any sound judgement. As it stands, uncertainties in local determinations may largely obscure possibilities to judge survey quality. The results further imply that surveys will benefit from procedures, controls and approaches in sampling and sample handling, to assess both average, variance and the nature of the distribution of elemental concentrations in local sites. This reasoning is compatible with the idea of the site as a basic homogeneous survey unit, which is implicitly and conceptually underlying any survey performed. (author)

  5. Stereotype locally convex spaces

    Energy Technology Data Exchange (ETDEWEB)

    Akbarov, S S

    2000-08-31

    We give complete proofs of some previously announced results in the theory of stereotype (that is, reflexive in the sense of Pontryagin duality) locally convex spaces. These spaces have important applications in topological algebra and functional analysis.

  6. Stereotype locally convex spaces

    Science.gov (United States)

    Akbarov, S. S.

    2000-08-01

    We give complete proofs of some previously announced results in the theory of stereotype (that is, reflexive in the sense of Pontryagin duality) locally convex spaces. These spaces have important applications in topological algebra and functional analysis.

  7. Stereotype locally convex spaces

    International Nuclear Information System (INIS)

    We give complete proofs of some previously announced results in the theory of stereotype (that is, reflexive in the sense of Pontryagin duality) locally convex spaces. These spaces have important applications in topological algebra and functional analysis

  8. Interface localization near criticality

    CERN Document Server

    Delfino, Gesualdo

    2016-01-01

    The theory of interface localization in near-critical planar systems at phase coexistence is formulated from first principles. We show that mutual delocalization of two interfaces, amounting to interfacial wetting, occurs when the bulk correlation length critical exponent $\

  9. Time to Go Local!

    Science.gov (United States)

    ... Consumer Health Information, NLM On MedlinePlus.gov health topic pages, you will find "Go Local" links that take ... you visit one of the over 700 health topic pages on MedlinePlus.gov , you will see a box ...

  10. Local Debt Scare

    Institute of Scientific and Technical Information of China (English)

    HU YUE

    2010-01-01

    @@ Control over local governments' financing vehicles will be tightened to fend off what some economistswarn could turn into a mountain of hidden debts,the State Council vowed at an executive meeting on May 26.

  11. Local Relativistic Exact Decoupling

    CERN Document Server

    Peng, Daoling

    2012-01-01

    We present a systematic hierarchy of approximations for {\\it local} exact-decoupling of four-component quantum chemical Hamiltonians based on the Dirac equation. Our ansatz reaches beyond the trivial local approximation that is based on a unitary transformation of only the atomic block-diagonal part of the Hamiltonian. Systematically, off-diagonal Hamiltonian matrix blocks can be subjected to a unitary transformation to yield relativistically corrected matrix elements. The full hierarchy is investigated with respect to the accuracy reached for the electronic energy and molecular properties on a balanced test molecule set that comprises molecules with heavy elements in different bonding situations. Our atomic (local) assembly of the unitary transformation needed for exact decoupling provides an excellent local approximation for any relativistic exact-decoupling approach. Its order-$N^2$ scaling can be further reduced to linear scaling by employing the neighboring-atomic-blocks approximation. Therefore, it is a...

  12. Focus on Local Reaction.

    Science.gov (United States)

    Howe, Rod

    1999-01-01

    Discusses how the student newspaper "The Lance" (at Westside High School in Omaha, Nebraska) covered the shootings at Columbine High School. Notes that the staff localized the event and brought the student body into the story. (RS)

  13. Desempleo y justicia local

    Directory of Open Access Journals (Sweden)

    Elster, Jon

    2003-04-01

    Full Text Available In this paper Jon Elster presents a set of criteria of local justice that regulate dismissals. In situations where work is limited and companies adopt means of employment redundancy, the decision of dismissing certain workers —when this one tries not to be arbitrary— can be based on local criteria of fairness like merit, efficiency, age or other analysed in this work. So, after defining the concept of local justice and presenting some examples, the paper focuses on the importance of the work to organize and structure the life of people. Unemployment has serious consequences for the persons who suffer it and the author advises that it is necessary to subordinate dismissals to criteria of local justice.

    En este artículo se presentan una serie de criterios de justicia local para regular los despidos. En situaciones en las que el trabajo es escaso y las empresas adoptan medidas de regulación de empleo, la decisión de despedir a unos trabajadores o a otros se puede basar, cuando se intenta que no sea arbitraria, en criterios locales de equidad como el mérito, la eficacia, la edad u otros que se analizan en este trabajo. Así, tras definir el concepto de justicia local y presentar algunos ejemplos, el artículo se centra en la importancia del trabajo para organizar y estructurar la vida de la gente. Las graves consecuencias que tiene, por ello, el desempleo para quien lo sufre, aconseja que los despidos se sometan a criterios de justicia local.

  14. Local Governance and Corruption

    OpenAIRE

    Profiroiu, Marius; Andrei, Tudorel; Gheorghe POPESCU; Alina PROFIROIU

    2006-01-01

    This paper attempts to examine, from the Romanian perspective, the degree to which decentralization process and improvement of local governance contributes to the reduction of corruption in the short and medium term. Through the methodology that is used the paper is consistent with the international trend that endeavors to analyze the impact of corruption on economic and social processes at the local level. In addition, recent research on corruption issues has focused upon the measurement of ...

  15. Parathyroid adenoma Localization

    OpenAIRE

    Nasiri, Shirzad; Soroush, Ahmadreza; Hashemi, Amir Pejman; Hedayat, Anushiravan; Donboli, Kianoush; Mehrkhani, Farhad

    2012-01-01

    Background Bilateral neck exploration is the gold standard for parathyroid adenoma localization in primary hyperparathyroidism. But surgeons do not have adequate experience for accurate surgical exploration and new methods are developed for surgery like unilateral exploration and minimally invasive surgery, thus, preoperative localization could reduces time and stress in surgical performance. Method 80 patients with documented primary hyperparathyroidism and with raised serum calcium and para...

  16. The Local Interstellar Medium

    OpenAIRE

    Ferlet, R.

    2006-01-01

    The Local Interstellar Medium (LISM) is a unique environment that presents an opportunity to study general interstellar phenomena in great detail and in three dimensions. In particular, high resolution optical and ultraviolet spectroscopy have proven to be powerful tools for addressing fundamental questions concerning the physical conditions and three-dimensional (3D) morphology of this local material. After reviewing our current understanding of the structure of gas in the solar neighborhood...

  17. Locality and reality revisited

    OpenAIRE

    Kent, Adrian

    2002-01-01

    Bell gave the now standard definition of a local hidden variable theory and showed that such theories cannot reproduce the predictions of quantum mechanics without violating his ``free will'' criterion: experimenters' measurement choices can be assumed to be uncorrelated with properties of the measured system prior to measurement. An alternative is considered here: a probabilistic theory of hidden variables underlying quantum mechanics could be statistically local, in the sense that it suppli...

  18. Local approximate inference algorithms

    OpenAIRE

    Jung, Kyomin; Shah, Devavrat

    2006-01-01

    We present a new local approximation algorithm for computing Maximum a Posteriori (MAP) and log-partition function for arbitrary exponential family distribution represented by a finite-valued pair-wise Markov random field (MRF), say $G$. Our algorithm is based on decomposition of $G$ into {\\em appropriately} chosen small components; then computing estimates locally in each of these components and then producing a {\\em good} global solution. We show that if the underlying graph $G$ either excl...

  19. Local Relativistic Exact Decoupling

    OpenAIRE

    Peng, Daoling; Reiher, Markus

    2012-01-01

    We present a systematic hierarchy of approximations for {\\it local} exact-decoupling of four-component quantum chemical Hamiltonians based on the Dirac equation. Our ansatz reaches beyond the trivial local approximation that is based on a unitary transformation of only the atomic block-diagonal part of the Hamiltonian. Systematically, off-diagonal Hamiltonian matrix blocks can be subjected to a unitary transformation to yield relativistically corrected matrix elements. The full hierarchy is i...

  20. On locally coherent hearts

    OpenAIRE

    Saorín, Manuel

    2016-01-01

    We show that, under particular conditions, if a t-structure in the unbounded derived category of a locally coherent Grothendieck category restricts to the bounded derived category of its category of finitely presented objects, then its heart is itself a locally coherent Grothendieck category. Those particular conditions are always satisfied when the Grothendieck category is arbitrary and one considers the t-structure associated to a torsion pair in the category of finitely presented objects. ...

  1. Localized Tracheal Amyloidosis

    OpenAIRE

    Juričevski, Ivan; Vrčić, Mihovil; Vrčić, Alka; Budimir, Ivan; Križanac, Šimun; Tuđman, Zdenko; Varga, Damir

    2005-01-01

    Amyloidosis is a disorder characterized by localized or diffuse deposition of fibrillary proteins in the extracellular space, causing progressive damage to tissue structure and organ function. Any organ system of the body may be involved by amyloidosis. A case is presented of localized tracheal amyloidosis in a 62-year-old man treated for active lung tuberculosis. Among other procedures, diagnostic workup included bronchoscopy, which revealed tumor-like lesions of tracheal mucosa. Histologic ...

  2. Subcellular localization and regulation of type-1C and type-5 phosphodiesterases

    International Nuclear Information System (INIS)

    We investigated the subcellular localization of PDE5 in in vitro human myometrial cells. We demonstrated for First time that PDE5 is localized in discrete cytoplasmic foci and vesicular compartments corresponding to centrosomes. We also found that PDE5 intracellular localization is not cell- or species-specific, as it is conserved in different animal and human cells. PDE5 protein levels are strongly regulated by the mitotic activity of the smooth muscle cells (SMCs), as they were increased in quiescent, contractile myometrial cultures, and conditions in which proliferation was inhibited. In contrast, PDE1C levels decreased in all conditions that inhibited proliferation. This mirrored the enzymatic activity of both PDE5 and PDE1C. Increasing cGMP intracellular levels by dbcGMP or sildenafil treatments did not block proliferation, while dbcAMP inhibited myometrial cell proliferation. Together, these results suggest that PDE5 regulation of cGMP intracellular levels is not involved in the control of SMC cycle progression, but may represent one of the markers of the contractile phenotype

  3. Local sustainable development – implications for local public finances

    OpenAIRE

    FLORIN OPREA; ELENA CIGU

    2012-01-01

    Local sustainable development implications on local public finances are complex because this phenomenon explained by development strategies mobilizes local resources towards achieving the three dimensions that define the phenomenon. This paper aims to develop these implications in Romania, showing that the local development strategy determines the reorganization of local government finances in identifying resources to cover expenses, which are directed towards achieving the goals.

  4. Monaural Sound Localization Revisited

    Science.gov (United States)

    Wightman, Frederic L.; Kistler, Doris J.

    1997-01-01

    Research reported during the past few decades has revealed the importance for human sound localization of the so-called 'monaural spectral cues.' These cues are the result of the direction-dependent filtering of incoming sound waves accomplished by the pinnae. One point of view about how these cues are extracted places great emphasis on the spectrum of the received sound at each ear individually. This leads to the suggestion that an effective way of studying the influence of these cues is to measure the ability of listeners to localize sounds when one of their ears is plugged. Numerous studies have appeared using this monaural localization paradigm. Three experiments are described here which are intended to clarify the results of the previous monaural localization studies and provide new data on how monaural spectral cues might be processed. Virtual sound sources are used in the experiments in order to manipulate and control the stimuli independently at the two ears. Two of the experiments deal with the consequences of the incomplete monauralization that may have contaminated previous work. The results suggest that even very low sound levels in the occluded ear provide access to interaural localization cues. The presence of these cues complicates the interpretation of the results of nominally monaural localization studies. The third experiment concerns the role of prior knowledge of the source spectrum, which is required if monaural cues are to be useful. The results of this last experiment demonstrate that extraction of monaural spectral cues can be severely disrupted by trial-to-trial fluctuations in the source spectrum. The general conclusion of the experiments is that, while monaural spectral cues are important, the monaural localization paradigm may not be the most appropriate way to study their role.

  5. LOCAL TADPOLE GALAXIES

    International Nuclear Information System (INIS)

    Tadpole galaxies have a giant star-forming region at the end of an elongated intensity distribution. Here we use Sloan Digital Sky Survey data to determine the ages, masses, and surface densities of the heads and tails in 14 local tadpoles selected from the Kiso and Michigan surveys of UV-bright galaxies, and we compare them to tadpoles previously studied in the Hubble Ultra Deep Field. The young stellar mass in the head scales linearly with rest-frame galaxy luminosity, ranging from ∼105 M☉ at galaxy absolute magnitude U = –13 mag to 109 M☉ at U = –20 mag. The corresponding head surface density increases from several M ☉ pc–2 locally to 10-100 M☉ pc–2 at high redshift, and the star formation rate (SFR) per unit area in the head increases from ∼0.01 M☉ yr–1 kpc–2 locally to ∼1 M☉ yr–1 kpc–2 at high z. These local values are normal for star-forming regions, and the increases with redshift are consistent with other cosmological SFRs, most likely reflecting an increase in gas abundance. The tails in the local sample look like bulge-free galaxy disks. Their photometric ages decrease from several Gyr to several hundred Myr with increasing z, and their surface densities are more constant than the surface densities of the heads. The far-outer intensity profiles in the local sample are symmetric and exponential. We suggest that most local tadpoles are bulge-free galaxy disks with lopsided star formation, perhaps from environmental effects such as ram pressure or disk impacts, or from a Jeans length comparable to half the disk size.

  6. 大鼠海马tau蛋白过度磷酸化时细胞周期蛋白cyclin D1的表达%Expression of mitotic protein cyclin D1 during tau hyperphosphorylation in rat hippocampus

    Institute of Scientific and Technical Information of China (English)

    李妍; 董为人; 王海红; 刘忠英; 晏芳

    2011-01-01

    目的:探讨大鼠海马tau蛋白过度磷酸化时细胞周期蛋白cyclin D1的表达.方法:用cAMP-依赖性蛋白激酶 A (PKA) 的激动剂Forskolin注射大鼠侧脑室,免疫印迹和免疫组织化学技术检测大鼠海马tau蛋白的磷酸化水平,同时采用免疫印迹、免疫组织化学和RT-PCR方法检测海马cyclin D1和cyclin D1 mRNA.结果:侧脑室注射Forskolin 48h,大鼠海马tau蛋白Ser214、Ser396和Ser202/Thr205位点的磷酸化水平升高,同时检测到细胞周期蛋白cyclin D1和cyclin D1 mRNA.结论:侧脑室注射Forskolin诱导大鼠海马 tau蛋白过度磷酸化的同时,大鼠海马出现细胞周期蛋白cyclin D1的表达,提示了tau蛋白过度磷酸化与细胞周期相关蛋白cyclin D1之间可能的内在联系.%Objective: To investigate the expression of mitotic protein cyclin D1 during tau hyperphosphorylation. Methods: Forskolin (80 μmol/L, 40μl) was injected into the lateral ventricle by stereotaxic injection. Tau phosphorylation in the hippocampus of rats was measured by Western blot and immunohistochemistry. The expression of cyclin D1 protein and the level of cyclin D1 mRNA in the hippocampus of rats were detected by Western blot, irnmunohistochemistry and RT-PCR technique. Results: The level of tau phosphorylation at Ser214, Ser396 and Ser202/Thr205 sites was significantly elevated at 48 h after administration of Forskolin. Meanwhile cyclin D1 protein and cyclin D1 mRNA were detected in Forskolin injection group. Conclusion: Mitotic protein cyclin D1 expressed in the hippocampus of rats during tau hyperphosphorylation induced by injection of Forskolin into the lateral ventricle of the rats. Present results suggest a positive correlation between tau hyperphosphorylation and the mitotic protein cyclin D1.

  7. The endocrine dyscrasia that accompanies menopause and andropause induces aberrant cell cycle signaling that triggers re-entry of post-mitotic neurons into the cell cycle, neurodysfunction, neurodegeneration and cognitive disease.

    Science.gov (United States)

    Atwood, Craig S; Bowen, Richard L

    2015-11-01

    This article is part of a Special Issue "SBN 2014". Sex hormones are physiological factors that promote neurogenesis during embryonic and fetal development. During childhood and adulthood these hormones support the maintenance of brain structure and function via neurogenesis and the formation of dendritic spines, axons and synapses required for the capture, processing and retrieval of information (memories). Not surprisingly, changes in these reproductive hormones that occur with menopause and during andropause are strongly correlated with neurodegeneration and cognitive decline. In this connection, much evidence now indicates that Alzheimer's disease (AD) involves aberrant re-entry of post-mitotic neurons into the cell cycle. Cell cycle abnormalities appear very early in the disease, prior to the appearance of plaques and tangles, and explain the biochemical, neuropathological and cognitive changes observed with disease progression. Intriguingly, a recent animal study has demonstrated that induction of adult neurogenesis results in the loss of previously encoded memories while decreasing neurogenesis after memory formation during infancy mitigated forgetting. Here we review the biochemical, epidemiological and clinical evidence that alterations in sex hormone signaling associated with menopause and andropause drive the aberrant re-entry of post-mitotic neurons into an abortive cell cycle that leads to neurite retraction, neuron dysfunction and neuron death. When the reproductive axis is in balance, gonadotropins such as luteinizing hormone (LH), and its fetal homolog, human chorionic gonadotropin (hCG), promote pluripotent human and totipotent murine embryonic stem cell and neuron proliferation. However, strong evidence supports menopausal/andropausal elevations in the LH:sex steroid ratio as driving aberrant mitotic events. These include the upregulation of tumor necrosis factor; amyloid-β precursor protein processing towards the production of mitogenic Aβ; and

  8. Local homotopy theory

    CERN Document Server

    Jardine, John F

    2015-01-01

    This monograph on the homotopy theory of topologized diagrams of spaces and spectra gives an expert account of a subject at the foundation of motivic homotopy theory and the theory of topological modular forms in stable homotopy theory. Beginning with an introduction to the homotopy theory of simplicial sets and topos theory, the book covers core topics such as the unstable homotopy theory of simplicial presheaves and sheaves, localized theories, cocycles, descent theory, non-abelian cohomology, stacks, and local stable homotopy theory. A detailed treatment of the formalism of the subject is interwoven with explanations of the motivation, development, and nuances of ideas and results. The coherence of the abstract theory is elucidated through the use of widely applicable tools, such as Barr's theorem on Boolean localization, model structures on the category of simplicial presheaves on a site, and cocycle categories. A wealth of concrete examples convey the vitality and importance of the subject in topology, n...

  9. Locality and causality revisited

    CERN Document Server

    Kent, A

    2002-01-01

    Bell gave the now standard definition of a local hidden variable theory and showed that such theories cannot reproduce the predictions of quantum mechanics without violating his ``free will'' criterion: experimenters' measurement choices can be assumed to be uncorrelated with properties of the measured system prior to measurement. An alternative is considered here: a probabilistic theory of hidden variables underlying quantum mechanics could be statistically local, in the sense that it supplies global configuration probabilities which are defined by expressions involving only local terms. This allows Bell correlations without relying on {\\it either} a conspiracy theory in which prior common causes correlate the system state with experimenters' choices {\\it or} a reverse causation principle in which experimenters' choices affect the earlier system states. In particular, there is no violation of the free will criterion. It gives a different perspective on Bell correlations, in which the puzzle is not that appar...

  10. Tackling Health Inequalities Locally

    DEFF Research Database (Denmark)

    Diderichsen, Finn; Scheele, Christian Elling; Little, Ingvild Gundersen

    equality and growth (64). Health equity has been anexplicit political goal in Scandinavia for decades. Nevertheless, in the health domain, average improvement has not been followed by reduced inequality – at least not between socioeconomic groups. It has in other words turned out to be a challenge...... of translating small inequalities in wealth into small inequalities in health. Denmark, Norway and Sweden all have legislation that indifferent ways offers local governments key roles in public health. This is partly due to local governments’ responsibility for many policy areas of great relevance to health...... and health equity. National governments have thus largely made the WHO and EU recommendation of ‘Health in All Policies’a local responsibility. In his analysis for the Nordic Council of Ministers, former Swedish Minister of Health Bo Könberg identifies tackling health inequalities as one of 14 prioritised...

  11. Theoretical Considerations On Local Budgets

    OpenAIRE

    Ionel Eduard Ionescu; Cristian Constantin Oprea

    2012-01-01

    This paper provides an overview of local budgets, or otherwise highlights the importance of these tools - local budgets, in the financial autonomy mechanism. Local communities know better than central authorities, their possibilities of obtaining financial resources and needs on expenses for providing local partner services. All these resources and needs are identified in local budgets.

  12. Fast Local Computation Algorithms

    OpenAIRE

    Rubinfeld, Ronitt; Tamir, Gil; Vardi, Shai; Xie, Ning

    2011-01-01

    For input $x$, let $F(x)$ denote the set of outputs that are the "legal" answers for a computational problem $F$. Suppose $x$ and members of $F(x)$ are so large that there is not time to read them in their entirety. We propose a model of {\\em local computation algorithms} which for a given input $x$, support queries by a user to values of specified locations $y_i$ in a legal output $y \\in F(x)$. When more than one legal output $y$ exists for a given $x$, the local computation algorithm should...

  13. Fermion localization and causality

    International Nuclear Information System (INIS)

    The velocity of a fermion (neutrino) propagation from its source to its detector is investigated. It is shown that a simple approach to the problem in terms of the quantum packet spreading is not applicable because the fermion cannot be localized in a bounded space region. Another approach is elaborated which uses relativistic quantum field description of the fermion source and detector. It is demonstrated that velocity of the fermion propagation does not exceed the knight velocity C within the precision of the source and detector localization. 41 refs.; 1 fig

  14. Localizing the relativistic electron

    International Nuclear Information System (INIS)

    A causally well-behaved solution of the localization problem for the free electron is given, with natural space-time transformation properties, in terms of Dirac's position operator x. It is shown that, although x is not an observable in the usual sense, and has no positive-energy (generalized) eigenstates, the four-vector density ((x,t),j(x,t)/c) is observable, and can be localized arbitrarily precisely about any point in space, at any instant of time, using only positive-energy states. A suitable spin operator can be diagonalized at the same time. (author)

  15. Localizing the relativistic electron

    Energy Technology Data Exchange (ETDEWEB)

    Bracken, A.J. [School of Theoretical Physics, Dublin Institute for Advanced Studies, Dublin (Ireland); Melloy, G.F. [Department of Mathematics, University of Queensland, Brisbane (Australia)

    1999-08-27

    A causally well-behaved solution of the localization problem for the free electron is given, with natural space-time transformation properties, in terms of Dirac's position operator x. It is shown that, although x is not an observable in the usual sense, and has no positive-energy (generalized) eigenstates, the four-vector density ((x,t),j(x,t)/c) is observable, and can be localized arbitrarily precisely about any point in space, at any instant of time, using only positive-energy states. A suitable spin operator can be diagonalized at the same time. (author)

  16. Ergodicity breaking and localization

    Science.gov (United States)

    Geneston, Elvis; Tuladhar, Rohisha; Beig, M. T.; Bologna, Mauro; Grigolini, Paolo

    2016-07-01

    We study the joint action of the non-Poisson renewal events (NPR) yielding Continuous-time random walk (CTRW) with index α produce subdiffusion. The joint action of NPR and FBM, on the contrary, may lead to localization when FBM is a source of superdiffusion. The joint action of NPR and FBM is equivalent to extending the CTRW to the case where the jumps of the runner are correlated and we argue that the the memory-induced localization requires a refinement of the theoretical perspective about determinism and randomness.

  17. Calming Local Debt Fears

    Institute of Scientific and Technical Information of China (English)

    2011-01-01

    Local governments in China, barred from directly selling bonds and taking out bank loans, have set up thousands of financing vehicles to raise money to fund infrastructure projects. A first-ever audit found that these local government financing vehicles have accumulated a record breaking debt of 10.7 trillion yuan ($1.7 trillion).But there’s no need to panic over this debt, said Xu Lin, Director of the Department of Fiscal and Financial Affairs of the National Development and Reform Commission (NDRC), in a question-andanswer statement posted on the NDRC’s website on August 29.Edited excerpts follow:

  18. Calming Local Debt Fears

    Institute of Scientific and Technical Information of China (English)

    2011-01-01

    Local governments in China,barred from directly selling bonds and taking out bank loans,have set up thousands of financing vehicles to raise money to fund infrastructure projects.A first-ever audit found that these local government financing vehicles have accumulated a record breaking debt of 10.7 trillion yuan ($1.7 trillion).But there's no need to panic over this debt,said Xu Lin,Director of the Department of Fiscal and Financial Affairs of the National Development and Reform Commission (NDRC),in a question-and-answer statement posted on the NDRC's website on August 29.

  19. Localizing the Relativistic Electron

    CERN Document Server

    Bracken, A J

    1999-01-01

    A causally well-behaved solution of the localization problem for the free electron is given, with natural space-time transformation properties, in terms of Dirac's position operator. It is shown that, although this operator does not represent an observable in the usual sense, and has no positive-energy (generalized) eigenstates, the associated 4-vector density is observable, and can be localized arbitrarily precisely about any point in space, at any instant of time, using only positive-energy states. A suitable spin operator can be diagonalized at the same time.

  20. Who Buys Local Food?

    OpenAIRE

    Zepeda, Lydia; Li, Jinghan

    2006-01-01

    Using data from a national survey of food shoppers, a Lancaster-Weinstein model is estimated using probit analysis to investigate the characteristics of local food buyers. Because there is no standard for what “local food†is, consumer research is used to define the term fairly narrowly as buying from farmers’ markets, buying directly from farmers, and Community Supported Agriculture (CSA) membership. The results reveal that income and demographic characteristics are not dominant factors...

  1. Local current measurements

    Czech Academy of Sciences Publication Activity Database

    Klapetek, P.; Fejfar, Antonín; Rezek, Bohuslav

    Oxford: Elsevier, 2013 - (Klapetek, P.), s. 221-245. (Micro and Nano Technologies). ISBN 978-1-4557-3058-2 R&D Projects: GA MŠk(CZ) LM2011026 Institutional support: RVO:68378271 Keywords : STM * AFM * local electronic properties * nano structures Subject RIV: BM - Solid Matter Physics ; Magnetism http://www.sciencedirect.com/science/book/9781455730582

  2. Statistics of Local Extremes

    DEFF Research Database (Denmark)

    Larsen, Gunner Chr.; Bierbooms, W.; Hansen, Kurt Schaldemose

    2003-01-01

    theoretical expression for the probability density function associated with local extremes of a stochasticprocess is presented. The expression is basically based on the lower four statistical moments and a bandwidth parameter. The theoretical expression is subsequently verified by comparison with simulated...

  3. Beyond local climate

    DEFF Research Database (Denmark)

    D'haen, Sarah Ann Lise; Nielsen, Jonas Østergaard; Lambin, Eric F.

    2014-01-01

    At the household level, nonfarm activities are thought to help rural poor households buffer against agricultural risks related to local climate variability by providing them with cash to buy food in the case of harvest shortfalls. Over the recent decades, households in rural Sub-Sahara have been...

  4. Stroke? Localized, otogenic meningitis!

    DEFF Research Database (Denmark)

    Ingolfsdottir, Harpa Maria; Thomasen, Per Caye

    2011-01-01

    We report the case of a patient admitted with aphasia, treated for a stroke. Subsequently, it was revealed that the symptoms were caused by complicated otitis media with localized meningitis. This case draws attention to the possible intracranial spread of infection when neurological symptoms occur...

  5. The Local Fractional Bootstrap

    DEFF Research Database (Denmark)

    Bennedsen, Mikkel; Hounyo, Ulrich; Lunde, Asger;

    new resampling method, the local fractional bootstrap, relies on simulating an auxiliary fractional Brownian motion that mimics the fine properties of high frequency differences of the Brownian semistationary process under the null hypothesis. We prove the first order validity of the bootstrap method...

  6. Test of Einstein locality

    International Nuclear Information System (INIS)

    Einstein locality imples that spacelike separated coherent quantum systems do not interfere, and that all interferences must propagate with at most the speed of light. It then follows that the reaction e+e-→K0antiK0 must yield some Ksub(s)Ksuu(s) decays in apparent violation of P and C conservation. (author)

  7. Local control room

    CERN Multimedia

    1972-01-01

    Local control room in the ejection building : all electronics pertaining to proton distribution and concomitants such as beam gymnastics and diagnostics at high energies will eventually be gathered here. Shown is the first of two rows of fast ejection electronic racks. It includes only what is necessary for operation.

  8. Local microwave background radiation

    OpenAIRE

    Soares, Domingos

    2006-01-01

    An inquiry on a possible local origin for the Microwave Background Radiation is made. Thermal MBR photons are contained in a system called {\\it magnetic bottle} which is due to Earth magnetic field and solar wind particles, mostly electrons. Observational tests are anticipated.

  9. REC46 gene of Saccharomyces cerevisiae controls mitotic chromosomal stability, recombination and sporulation: cell-type and life cycle stage specific expression of the rec46-1 mutation

    International Nuclear Information System (INIS)

    Studies of chromosomal recombination during mitosis and meiosis of Saccharomyces cerevisiae have demonstrated that recombination at these two distinct stages of the yeast life cycle proceeds by mechanisms that appear similar but involve discrete mitosis-specific and meiosis-specific properties. UV radiation induced REC mutants are being employed as a genetic tool to identify the partial reactions comprising recombination and the involvement of individual REC gene products in mitotic and meiotic recombination. The sequence of molecular events that results in genetic recombination in eukaryotes is presently ill-defined. Genetic characterization of REC gene mutants and biochemical analyses of them for discrete defects in DNA metabolic proteins and enzymes (in collaboration with the laboratory of Junko Hosoda) are beginning to remedy this gap in the authors knowledge. This report summarizes the genetic properties of the rec46-1 mutation

  10. Symbolic local information transfer

    Science.gov (United States)

    Nakajima, K.; Haruna, T.

    2013-06-01

    Recently, the permutation-information theoretic approach has been used in a broad range of research fields. In particular, in the study of high-dimensional dynamical systems, it has been shown that this approach can be effective in characterizing global properties, including the complexity of their spatiotemporal dynamics. Here, we show that this approach can also be applied to reveal local spatiotemporal profiles of distributed computations existing at each spatiotemporal point in the system. J. T. Lizier et al. have recently introduced the concept of local information dynamics, which consists of information storage, transfer, and modification. This concept has been intensively studied with regard to cellular automata, and has provided quantitative evidence of several characteristic behaviors observed in the system. In this paper, by focusing on the local information transfer, we demonstrate that the application of the permutation-information theoretic approach, which introduces natural symbolization methods, makes the concept easily extendible to systems that have continuous states. We propose measures called symbolic local transfer entropies, and apply these measures to two test models, the coupled map lattice (CML) system and the Bak-Sneppen model (BS-model), to show their relevance to spatiotemporal systems that have continuous states. In the CML, we demonstrate that it can be successfully used as a spatiotemporal filter to stress a coherent structure buried in the system. In particular, we show that the approach can clearly stress out defect turbulences or Brownian motion of defects from the background, which gives quantitative evidence suggesting that these moving patterns are the information transfer substrate in the spatiotemporal system. We then show that these measures reveal qualitatively different properties from the conventional approach using the sliding window method, and are also robust against external noise. In the BS-model, we demonstrate that

  11. Bilateral lesions of suprachiasmatic nuclei affect circadian rhythms in [3H]-thymidine incorporation into deoxyribonucleic acid in mouse intestinal tract, mitotic index of corneal epithelium, and serum corticosterone

    International Nuclear Information System (INIS)

    Investigations into the role of the suprachiasmatic nuclei (SCN) in the coordination of circadian rhythms have presented differing results. Several reports have shown that ablation of the suprachiasmatic nuclei (SCNA) alters the phase and amplitude of rhythms but does not abolish them. The present study investigates the effect of SCNA on the rhythms in cell proliferation in various regions of the intestinal tract as measured by the incorporation of [3H]-thymidine into deoxyribonucleic acid, in the mitotic activity of the corneal epithelium, and in serum corticosterone levels. The study involved mice with verified lesions of the SCN (six to 13 mice per time point) and control groups of both sham-operated and unoperated mice (seven of each per time point). The mice were killed in groups that represented seven time points over a single 24 hr span (3 hr intervals with the 0800 hr sampled both at start and end of the series). The tissues examined were the tongue, esophagus, gastric stomach, and colon for DNA synthesis, the corneal epithelium for mitotic index, and blood serum for corticosterone level. The most consistent result of SCNA was a phase advance in the rhythms in cell proliferation in the tongue, esophagus, gastric stomach, colon, and corneal epithelium. A reduction in rhythm amplitude occurred in the tongue, esophagus, and corneal epithelium; however, there was an amplitude increase for the stomach, colon, and serum corticosterone. The mesor (rhythm-adjusted mean) was increased by SCNA in all tissues except the corneal epithelium. These findings further support the role of the suprachiasmatic nuclear area in the control of rhythms in cell proliferation and corticosterone production, by acting as a ''phase-resetter'' and as a modulator of rhythm amplitude

  12. Localization and vector spherical harmonics

    Science.gov (United States)

    von Brecht, James H.

    2016-01-01

    This paper establishes the following localization property for vector spherical harmonics: a wide class of non-local, vector-valued operators reduce to local, multiplication-type operations when applied to a vector spherical harmonic. As localization occurs in a very precise, quantifiable and explicitly computable fashion, the localization property provides a set of useful formulae for analyzing vector-valued fractional diffusion and non-local differential equations defined on S d - 1. As such analyses require a detailed understanding of operators for which localization occurs, we provide several applications of the result in the context of non-local differential equations.

  13. Interacting factors and cellular localization of SR protein-specific kinase Dsk1

    Energy Technology Data Exchange (ETDEWEB)

    Tang, Zhaohua, E-mail: ztang@jsd.claremont.edu [W.M. Keck Science Center, The Claremont Colleges, Claremont, CA 91711 (United States); Luca, Maria; Taggart-Murphy, Laura; Portillio, Jessica; Chang, Cathey; Guven, Ayse [W.M. Keck Science Center, The Claremont Colleges, Claremont, CA 91711 (United States); Lin, Ren-Jang [Department of Molecular and Cellular Biology, Beckman Research Institute of the City of Hope, Duarte, CA 91010 (United States); Murray, Johanne; Carr, Antony [Genome Damage and Stability Center, University of Sussex, Falmer, BN1 9RQ (United Kingdom)

    2012-10-01

    Schizosaccharomyces pombe Dsk1 is an SR protein-specific kinase (SRPK), whose homologs have been identified in every eukaryotic organism examined. Although discovered as a mitotic regulator with protein kinase activity toward SR splicing factors, it remains largely unknown about what and how Dsk1 contributes to cell cycle and pre-mRNA splicing. In this study, we investigated the Dsk1 function by determining interacting factors and cellular localization of the kinase. Consistent with its reported functions, we found that pre-mRNA processing and cell cycle factors are prominent among the proteins co-purified with Dsk1. The identification of these factors led us to find Rsd1 as a novel Dsk1 substrate, as well as the involvement of Dsk1 in cellular distribution of poly(A){sup +} RNA. In agreement with its role in nuclear events, we also found that Dsk1 is mainly localized in the nucleus during G{sub 2} phase and at mitosis. Furthermore, we revealed the oscillation of Dsk1 protein in a cell cycle-dependent manner. This paper marks the first comprehensive analysis of in vivo Dsk1-associated proteins in fission yeast. Our results reflect the conserved role of SRPK family in eukaryotic organisms, and provide information about how Dsk1 functions in pre-mRNA processing and cell-division cycle.

  14. AtKP1, a kinesin-like protein, mainly localizes to mitochondria in Arabidopsis thaliana

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    Kinesins and kinesin-like proteins (KLPs) constitute a large family of microtubule-based motors that play important roles in many fundamental cellular and developmental processes. To date, a number of kinesins or KLPs have been identified in plants including Arabidopsis thaliana. Here, a polyclonal antibody against AtKP1 (kinesin-like protein 1 in A.thaliana) was raised by injection the expressed AtKP1 specific C-terminal polypeptides in rabbits, and immunoblot analysis was conducted with the affinity-purified anti-AtKP1 antibody. The results indicated that this antibody recognized the AtKP1 fusion proteins expressed in E. coli and proteins of ~125 kDa in the soluble fractions of Arabidopsis extracts. The molecular weight was consistent with the calculated molecular weight based on deduced amino acids sequence of AtKP1. To acquire the subcellular localization of the protein, AtKP1 in Arabidopsis root cells was observed by indirect immunofluorescence microscopy. AtKP1 was localized to particle-like organelles in interphase or dividing cells, but not to mitotic microtubule arrays. Relatively more AtKP1 was found in isolated mitochondria fraction on immunoblot of the subcellular fractions. The AtKP1 protein could not be released following a 0.6 M KI washing,indicating that AtKP1 is tightly bind to mitochondria and might function associated with this kind of organelles.

  15. Interacting factors and cellular localization of SR protein-specific kinase Dsk1

    International Nuclear Information System (INIS)

    Schizosaccharomyces pombe Dsk1 is an SR protein-specific kinase (SRPK), whose homologs have been identified in every eukaryotic organism examined. Although discovered as a mitotic regulator with protein kinase activity toward SR splicing factors, it remains largely unknown about what and how Dsk1 contributes to cell cycle and pre-mRNA splicing. In this study, we investigated the Dsk1 function by determining interacting factors and cellular localization of the kinase. Consistent with its reported functions, we found that pre-mRNA processing and cell cycle factors are prominent among the proteins co-purified with Dsk1. The identification of these factors led us to find Rsd1 as a novel Dsk1 substrate, as well as the involvement of Dsk1 in cellular distribution of poly(A)+ RNA. In agreement with its role in nuclear events, we also found that Dsk1 is mainly localized in the nucleus during G2 phase and at mitosis. Furthermore, we revealed the oscillation of Dsk1 protein in a cell cycle-dependent manner. This paper marks the first comprehensive analysis of in vivo Dsk1-associated proteins in fission yeast. Our results reflect the conserved role of SRPK family in eukaryotic organisms, and provide information about how Dsk1 functions in pre-mRNA processing and cell-division cycle.

  16. The Local Interstellar Medium

    CERN Document Server

    Redfield, S

    2006-01-01

    The Local Interstellar Medium (LISM) is a unique environment that presents an opportunity to study general interstellar phenomena in great detail and in three dimensions. In particular, high resolution optical and ultraviolet spectroscopy have proven to be powerful tools for addressing fundamental questions concerning the physical conditions and three-dimensional (3D) morphology of this local material. After reviewing our current understanding of the structure of gas in the solar neighborhood, I will discuss the influence that the LISM can have on stellar and planetary systems, including LISM dust deposition onto planetary atmospheres and the modulation of galactic cosmic rays through the astrosphere - the balancing interface between the outward pressure of the magnetized stellar wind and the inward pressure of the surrounding interstellar medium. On Earth, galactic cosmic rays may play a role as contributors to ozone layer chemistry, planetary electrical discharge frequency, biological mutation rates, and cl...

  17. Local versus national

    DEFF Research Database (Denmark)

    Milan, Christian; Nielsen, Mads Pagh; Bojesen, Carsten

    2014-01-01

    individual energy supply systems based on on-site weather and building conditions, as well as considering the expected energy consumption profile. However, local planning processes are problematic if they do not take regional or national impacts into account. Given the grid connection, the local building...... solution also has an impact on a national scale by exchanging electricity. Therefore it is important to implement respective grid loads into the planning process in order to avoid technology choices, which might counteract grid stability or cost inefficiencies at other sites. The aim of this paper is to...... adapt the earlier proposed methodology by integrating flexible national electricity prices and thus taking account for the aforementioned effects. The methodology is applied in a case study for a single family house under Danish conditions. The results show that the system configuration might not...

  18. Local Governance and Corruption

    Directory of Open Access Journals (Sweden)

    Marius PROFIROIU

    2006-06-01

    Full Text Available This paper attempts to examine, from the Romanian perspective, the degree to which decentralization process and improvement of local governance contributes to the reduction of corruption in the short and medium term. Through the methodology that is used the paper is consistent with the international trend that endeavors to analyze the impact of corruption on economic and social processes at the local level. In addition, recent research on corruption issues has focused upon the measurement of the level of corruption level and on its impact on the growing rate of the GDP (Mauro [1995]1, Abed and Davoodi2, Krueger [1974]3, on the impact upon some of the national economical sectors (Tanzi [1998]4, Shang-Jin Wei [2001]5, or on the decentralization processes (Shah [2006]6.

  19. Global-local interactions

    DEFF Research Database (Denmark)

    Larsen, Jytte Agergaard; Fold, Niels; Gough, Katherine

    2009-01-01

    although migration patterns and economic growth are conditioned by world market dynamics, local socioeconomic outcomes within frontier regions are diverse. Frontier formation is far from a homogenous process that slowly incorporates all localities and communities in the same way. Dak Lak Province, in the......Due to their dependence on a single crop, agricultural frontiers are often considered to be formed through phases of 'boom and bust'. These phases are closely related to fluctuations in world market prices of the commodity that constitutes the frontier's economic basis. This paper demonstrates how...... Central Highlands of Vietnam, produces more than 50% of Vietnam's coffee. As Vietnam is the world's largest exporter of Robusta coffee, Dak Lak is highly embedded in the dynamics of the world coffee market. Planned settlement in Dak Lak started in the 1950s and has continued in phases orchestrated by the...

  20. Local Quantum Gravity

    CERN Document Server

    Christiansen, Nicolai; Meibohm, Jan; Pawlowski, Jan M; Reichert, Manuel

    2015-01-01

    We investigate the ultraviolet behaviour of quantum gravity within a functional renormalisation group approach. The present setup includes the full ghost and graviton propagators and, for the first time, the dynamical graviton three-point function. The latter gives access to the coupling of dynamical gravitons and makes the system minimally self-consistent. The resulting phase diagram confirms the asymptotic safety scenario in quantum gravity with a non-trivial UV fixed point. A well-defined Wilsonian block spinning requires locality of the flow in momentum space. This property is discussed in the context of functional renormalisation group flows. We show that momentum locality of graviton correlation functions is non-trivially linked to diffeomorphism invariance, and is realised in the present setup.