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Sample records for activity cell cycle

  1. The Cell Cycle: An Activity Using Paper Plates to Represent Time Spent in Phases of the Cell Cycle

    ERIC Educational Resources Information Center

    Scherer, Yvette D.

    2014-01-01

    In this activity, students are given the opportunity to combine skills in math and geometry for a biology lesson in the cell cycle. Students utilize the data they collect and analyze from an online onion-root-tip activity to create a paper-plate time clock representing a 24-hour cell cycle. By dividing the paper plate into appropriate phases of…

  2. Staphylococcal Enterotoxin O Exhibits Cell Cycle Modulating Activity

    PubMed Central

    Hodille, Elisabeth; Alekseeva, Ludmila; Berkova, Nadia; Serrier, Asma; Badiou, Cedric; Gilquin, Benoit; Brun, Virginie; Vandenesch, François; Terman, David S.; Lina, Gerard

    2016-01-01

    Maintenance of an intact epithelial barrier constitutes a pivotal defense mechanism against infections. Staphylococcus aureus is a versatile pathogen that produces multiple factors including exotoxins that promote tissue alterations. The aim of the present study is to investigate the cytopathic effect of staphylococcal exotoxins SEA, SEG, SEI, SElM, SElN and SElO on the cell cycle of various human cell lines. Among all tested exotoxins only SEIO inhibited the proliferation of a broad panel of human tumor cell lines in vitro. Evaluation of a LDH release and a DNA fragmentation of host cells exposed to SEIO revealed that the toxin does not induce necrosis or apoptosis. Analysis of the DNA content of tumor cells synchronized by serum starvation after exposure to SEIO showed G0/G1 cell cycle delay. The cell cycle modulating feature of SEIO was confirmed by the flow cytometry analysis of synchronized cells exposed to supernatants of isogenic S. aureus strains wherein only supernatant of the SElO producing strain induced G0/G1 phase delay. The results of yeast-two-hybrid analysis indicated that SEIO’s potential partner is cullin-3, involved in the transition from G1 to S phase. In conclusion, we provide evidence that SEIO inhibits cell proliferation without inducing cell death, by delaying host cell entry into the G0/G1 phase of the cell cycle. We speculate that this unique cell cycle modulating feature allows SEIO producing bacteria to gain advantage by arresting the cell cycle of target cells as part of a broader invasive strategy. PMID:27148168

  3. Staphylococcal Enterotoxin O Exhibits Cell Cycle Modulating Activity.

    PubMed

    Hodille, Elisabeth; Alekseeva, Ludmila; Berkova, Nadia; Serrier, Asma; Badiou, Cedric; Gilquin, Benoit; Brun, Virginie; Vandenesch, François; Terman, David S; Lina, Gerard

    2016-01-01

    Maintenance of an intact epithelial barrier constitutes a pivotal defense mechanism against infections. Staphylococcus aureus is a versatile pathogen that produces multiple factors including exotoxins that promote tissue alterations. The aim of the present study is to investigate the cytopathic effect of staphylococcal exotoxins SEA, SEG, SEI, SElM, SElN and SElO on the cell cycle of various human cell lines. Among all tested exotoxins only SEIO inhibited the proliferation of a broad panel of human tumor cell lines in vitro. Evaluation of a LDH release and a DNA fragmentation of host cells exposed to SEIO revealed that the toxin does not induce necrosis or apoptosis. Analysis of the DNA content of tumor cells synchronized by serum starvation after exposure to SEIO showed G0/G1 cell cycle delay. The cell cycle modulating feature of SEIO was confirmed by the flow cytometry analysis of synchronized cells exposed to supernatants of isogenic S. aureus strains wherein only supernatant of the SElO producing strain induced G0/G1 phase delay. The results of yeast-two-hybrid analysis indicated that SEIO's potential partner is cullin-3, involved in the transition from G1 to S phase. In conclusion, we provide evidence that SEIO inhibits cell proliferation without inducing cell death, by delaying host cell entry into the G0/G1 phase of the cell cycle. We speculate that this unique cell cycle modulating feature allows SEIO producing bacteria to gain advantage by arresting the cell cycle of target cells as part of a broader invasive strategy. PMID:27148168

  4. RACK1 inhibits colonic cell growth by regulating Src activity at cell cycle checkpoints.

    PubMed

    Mamidipudi, V; Dhillon, N K; Parman, T; Miller, L D; Lee, K C; Cartwright, C A

    2007-05-01

    Previously, we showed that Src tyrosine kinases are activated early in the development of human colon cancer and are suppressed as intestinal cells differentiate. We identified RACK1 as an endogenous substrate, binding partner and inhibitor of Src. Here we show (by overexpressing RACK1, depleting Src or RACK1 and utilizing cell-permeable peptides that perturb RACK1's interaction with Src) that RACK1 regulates growth of colon cells by suppressing Src activity at G(1) and mitotic checkpoints, and consequently delaying cell cycle progression. Activated Src rescues RACK1-inhibited growth of HT-29 cells. Conversely, inhibiting Src abolishes growth promoted by RACK1 depletion in normal cells. Two potential mechanisms whereby RACK1 regulates mitotic exit are identified: suppression of Src-mediated Sam68 phosphorylation and maintenance of the cyclin-dependent kinase (CDK) 1-cyclin B complex in an active state. Our results reveal novel mechanisms of cell cycle control in G(1) and mitosis of colon cells. The significance of this work lies in the discovery of a mechanism by which the growth of colon cancer cells can be slowed, by RACK1 suppression of an oncogenic kinase at critical cell cycle checkpoints. Small molecules that mimic RACK1 function may provide a powerful new approach to the treatment of colon cancer. PMID:17072338

  5. 5-ASA Affects Cell Cycle Progression in Colorectal Cells by Reversibly Activating a Replication Checkpoint

    PubMed Central

    LUCIANI, M. GLORIA; CAMPREGHER, CHRISTOPH; FORTUNE, JOHN M.; KUNKEL, THOMAS A.; GASCHE, CHRISTOPH

    2007-01-01

    Background & Aims Individuals with inflammatory bowel disease are at risk of developing colorectal cancer (CRC). Epidemiologic, animal, and laboratory studies suggest that 5-amino-salicylic acid (5-ASA) protects from the development of CRC by altering cell cycle progression and by inducing apoptosis. Our previous results indicate that 5-ASA improves replication fidelity in colorectal cells, an effect that is active in reducing mutations. In this study, we hypothesized that 5-ASA restrains cell cycle progression by activating checkpoint pathways in colorectal cell lines, which would prevent tumor development and improve genomic stability. Methods CRC cells with different genetic backgrounds such as HT29, HCT116, HCT116p53−/−, HCT116+chr3, and LoVo were treated with 5-ASA for 2–96 hours. Cell cycle progression, phosphorylation, and DNA binding of cell cycle checkpoint proteins were analyzed. Results We found that 5-ASA at concentrations between 10 and 40 mmol/L affects cell cycle progression by inducing cells to accumulate in the S phase. This effect was independent of the hMLH1, hMSH2, and p53 status because it was observed to a similar extent in all cell lines under investigation. Moreover, wash-out experiments demonstrated reversibility within 48 hours. Although p53 did not have a causative role, p53 Ser15 was strongly phosphorylated. Proteins involved in the ATM-and-Rad3-related kinase (ATR)-dependent S-phase checkpoint response (Chk1 and Rad17) were also phosphorylated but not ataxia telengectasia mutated kinase. Conclusions Our data demonstrate that 5-ASA causes cells to reversibly accumulate in S phase and activate an ATR-dependent checkpoint. The activation of replication checkpoint may slow down DNA replication and improve DNA replication fidelity, which increases the maintenance of genomic stability and counteracts carcinogenesis. PMID:17241873

  6. Cdk1 activity acts as a quantitative platform for coordinating cell cycle progression with periodic transcription

    PubMed Central

    Banyai, Gabor; Baïdi, Feriel; Coudreuse, Damien; Szilagyi, Zsolt

    2016-01-01

    Cell proliferation is regulated by cyclin-dependent kinases (Cdks) and requires the periodic expression of particular gene clusters in different cell cycle phases. However, the interplay between the networks that generate these transcriptional oscillations and the core cell cycle machinery remains largely unexplored. In this work, we use a synthetic regulable Cdk1 module to demonstrate that periodic expression is governed by quantitative changes in Cdk1 activity, with different clusters directly responding to specific activity levels. We further establish that cell cycle events neither participate in nor interfere with the Cdk1-driven transcriptional program, provided that cells are exposed to the appropriate Cdk1 activities. These findings contrast with current models that propose self-sustained and Cdk1-independent transcriptional oscillations. Our work therefore supports a model in which Cdk1 activity serves as a quantitative platform for coordinating cell cycle transitions with the expression of critical genes to bring about proper cell cycle progression. PMID:27045731

  7. TGEV nucleocapsid protein induces cell cycle arrest and apoptosis through activation of p53 signaling

    SciTech Connect

    Ding, Li; Huang, Yong; Du, Qian; Dong, Feng; Zhao, Xiaomin; Zhang, Wenlong; Xu, Xingang; Tong, Dewen

    2014-03-07

    Highlights: • TGEV N protein reduces cell viability by inducing cell cycle arrest and apoptosis. • TGEV N protein induces cell cycle arrest and apoptosis by regulating p53 signaling. • TGEV N protein plays important roles in TGEV-induced cell cycle arrest and apoptosis. - Abstract: Our previous studies showed that TGEV infection could induce cell cycle arrest and apoptosis via activation of p53 signaling in cultured host cells. However, it is unclear which viral gene causes these effects. In this study, we investigated the effects of TGEV nucleocapsid (N) protein on PK-15 cells. We found that TGEV N protein suppressed cell proliferation by causing cell cycle arrest at the S and G2/M phases and apoptosis. Characterization of various cellular proteins that are involved in regulating cell cycle progression demonstrated that the expression of N gene resulted in an accumulation of p53 and p21, which suppressed cyclin B1, cdc2 and cdk2 expression. Moreover, the expression of TGEV N gene promoted translocation of Bax to mitochondria, which in turn caused the release of cytochrome c, followed by activation of caspase-3, resulting in cell apoptosis in the transfected PK-15 cells following cell cycle arrest. Further studies showed that p53 inhibitor attenuated TGEV N protein induced cell cycle arrest at S and G2/M phases and apoptosis through reversing the expression changes of cdc2, cdk2 and cyclin B1 and the translocation changes of Bax and cytochrome c induced by TGEV N protein. Taken together, these results demonstrated that TGEV N protein might play an important role in TGEV infection-induced p53 activation and cell cycle arrest at the S and G2/M phases and apoptosis occurrence.

  8. Cell cycle-dependent regulation of RNA polymerase II basal transcription activity.

    PubMed Central

    Yonaha, M; Chibazakura, T; Kitajima, S; Yasukochi, Y

    1995-01-01

    Regulation of transcription by RNA polymerase II (pol II) in eukaryotic cells requires both basal and regulatory transcription factors. In this report we have investigated in vitro pol II basal transcription activity during the cell cycle by using nuclear extracts from synchronized HeLa cells. It is shown that pol II basal transcription activity is low in the S and G2 phases and high in early G1 phase and TFIID is the rate limiting component of pol II basal transcription activity during the cell cycle. Further analyses reveal that TFIID exists as a less active form in the S and G2 phases and nuclear extracts from S and G2 phase cells contain a heat-sensitive repressor(s) of TATA box binding protein (TBP). These results suggest that pol II basal transcription activity is regulated by a qualitative change in the TFIID complex, which could involve repression of TBP, during the cell cycle. Images PMID:7479063

  9. Activation of the methylation cycle in cells reprogrammed into a stem cell-like state

    PubMed Central

    Bosch-Barrera, Joaquim; Alarcón, Tomás; Joven, Jorge; Menendez, Javier A.

    2015-01-01

    Generation of induced pluripotent stem (iPS) cells and cancer biogenesis share similar metabolic switches. Most studies have focused on how the establishment of a cancer-like glycolytic phenotype is necessary for the optimal routing of somatic cells for achieving stemness. However, relatively little effort has been dedicated towards elucidating how one-carbon (1C) metabolism is retuned during acquisition of stem cell identity. Here we used ultra-high pressure liquid chromatography coupled to an electrospray ionization source and a triple-quadrupole mass spectrometer [UHPLC-ESI-QqQ-MS/MS] to quantitatively examine the methionine/folate bi-cyclic 1C metabolome during nuclear reprogramming of somatic cells into iPS cells. iPS cells optimize the synthesis of the universal methyl donor S-adenosylmethionine (SAM), apparently augment the ability of the redox balance regulator NADPH in SAM biosynthesis, and greatly increase their methylation potential by triggering a high SAM:S-adenosylhomocysteine (SAH) ratio. Activation of the methylation cycle in iPS cells efficiently prevents the elevation of homocysteine (Hcy), which could alter global DNA methylation and induce mitochondrial toxicity, oxidative stress and inflammation. In this regard, the methyl donor choline is also strikingly accumulated in iPS cells, suggesting perhaps an overactive intersection of the de novo synthesis of choline with the methionine-Hcy cycle. Activation of methylogenesis and maintenance of an optimal SAM:Hcy ratio might represent an essential function of 1C metabolism to provide a labile pool of methyl groups and NADPH-dependent redox products required for successfully establishing and maintaining an embryonic-like DNA methylation imprint in stem cell states. PMID:26909364

  10. A cell cycle-controlled redox switch regulates the topoisomerase IV activity

    PubMed Central

    Narayanan, Sharath; Janakiraman, Balaganesh; Kumar, Lokesh

    2015-01-01

    Topoisomerase IV (topo IV), an essential factor during chromosome segregation, resolves the catenated chromosomes at the end of each replication cycle. How the decatenating activity of the topo IV is regulated during the early stages of the chromosome cycle despite being in continuous association with the chromosome remains poorly understood. Here we report a novel cell cycle-regulated protein in Caulobacter crescentus, NstA (negative switch for topo IV decatenation activity), that inhibits the decatenation activity of the topo IV during early stages of the cell cycle. We demonstrate that in C. crescentus, NstA acts by binding to the ParC DNA-binding subunit of topo IV. Most importantly, we uncover a dynamic oscillation of the intracellular redox state during the cell cycle, which correlates with and controls NstA activity. Thus, we propose that predetermined dynamic intracellular redox fluctuations may act as a global regulatory switch to control cellular development and cell cycle progression and may help retain pathogens in a suitable cell cycle state when encountering redox stress from the host immune response. PMID:26063575

  11. SB225002 Induces Cell Death and Cell Cycle Arrest in Acute Lymphoblastic Leukemia Cells through the Activation of GLIPR1

    PubMed Central

    Leal, Paulo C.; Bhasin, Manoj K.; Zenatti, Priscila Pini; Nunes, Ricardo J.; Yunes, Rosendo A.; Nowill, Alexandre E.; Libermann, Towia A.; Zerbini, Luiz Fernando; Yunes, José Andrés

    2015-01-01

    Acute Lymphoblastic Leukemia (ALL) is the most frequent childhood malignancy. In the effort to find new anti-leukemic agents, we evaluated the small drug SB225002 (N-(2-hydroxy-4-nitrophenyl)-N’-(2-bromophenyl)urea). Although initially described as a selective antagonist of CXCR2, later studies have identified other cellular targets for SB225002, with potential medicinal use in cancer. We found that SB225002 has a significant pro-apoptotic effect against both B- and T-ALL cell lines. Cell cycle analysis demonstrated that treatment with SB225002 induces G2-M cell cycle arrest. Transcriptional profiling revealed that SB225002-mediated apoptosis triggered a transcriptional program typical of tubulin binding agents. Network analysis revealed the activation of genes linked to the JUN and p53 pathways and inhibition of genes linked to the TNF pathway. Early cellular effects activated by SB225002 included the up-regulation of GLIPR1, a p53-target gene shown to have pro-apoptotic activities in prostate and bladder cancer. Silencing of GLIPR1 in B- and T-ALL cell lines resulted in increased resistance to SB225002. Although SB225002 promoted ROS increase in ALL cells, antioxidant N-Acetyl Cysteine pre-treatment only modestly attenuated cell death, implying that the pro-apoptotic effects of SB225002 are not exclusively mediated by ROS. Moreover, GLIPR1 silencing resulted in increased ROS levels both in untreated and SB225002-treated cells. In conclusion, SB225002 induces cell cycle arrest and apoptosis in different B- and T-ALL cell lines. Inhibition of tubulin function with concurrent activation of the p53 pathway, in particular, its downstream target GLIPR1, seems to underlie the anti-leukemic effect of SB225002. PMID:26302043

  12. SB225002 Induces Cell Death and Cell Cycle Arrest in Acute Lymphoblastic Leukemia Cells through the Activation of GLIPR1.

    PubMed

    de Vasconcellos, Jaíra Ferreira; Laranjeira, Angelo Brunelli Albertoni; Leal, Paulo C; Bhasin, Manoj K; Zenatti, Priscila Pini; Nunes, Ricardo J; Yunes, Rosendo A; Nowill, Alexandre E; Libermann, Towia A; Zerbini, Luiz Fernando; Yunes, José Andrés

    2015-01-01

    Acute Lymphoblastic Leukemia (ALL) is the most frequent childhood malignancy. In the effort to find new anti-leukemic agents, we evaluated the small drug SB225002 (N-(2-hydroxy-4-nitrophenyl)-N'-(2-bromophenyl)urea). Although initially described as a selective antagonist of CXCR2, later studies have identified other cellular targets for SB225002, with potential medicinal use in cancer. We found that SB225002 has a significant pro-apoptotic effect against both B- and T-ALL cell lines. Cell cycle analysis demonstrated that treatment with SB225002 induces G2-M cell cycle arrest. Transcriptional profiling revealed that SB225002-mediated apoptosis triggered a transcriptional program typical of tubulin binding agents. Network analysis revealed the activation of genes linked to the JUN and p53 pathways and inhibition of genes linked to the TNF pathway. Early cellular effects activated by SB225002 included the up-regulation of GLIPR1, a p53-target gene shown to have pro-apoptotic activities in prostate and bladder cancer. Silencing of GLIPR1 in B- and T-ALL cell lines resulted in increased resistance to SB225002. Although SB225002 promoted ROS increase in ALL cells, antioxidant N-Acetyl Cysteine pre-treatment only modestly attenuated cell death, implying that the pro-apoptotic effects of SB225002 are not exclusively mediated by ROS. Moreover, GLIPR1 silencing resulted in increased ROS levels both in untreated and SB225002-treated cells. In conclusion, SB225002 induces cell cycle arrest and apoptosis in different B- and T-ALL cell lines. Inhibition of tubulin function with concurrent activation of the p53 pathway, in particular, its downstream target GLIPR1, seems to underlie the anti-leukemic effect of SB225002. PMID:26302043

  13. RCC1-dependent activation of Ran accelerates cell cycle and DNA repair, inhibiting DNA damage-induced cell senescence.

    PubMed

    Cekan, Pavol; Hasegawa, Keisuke; Pan, Yu; Tubman, Emily; Odde, David; Chen, Jin-Qiu; Herrmann, Michelle A; Kumar, Sheetal; Kalab, Petr

    2016-04-15

    The coordination of cell cycle progression with the repair of DNA damage supports the genomic integrity of dividing cells. The function of many factors involved in DNA damage response (DDR) and the cell cycle depends on their Ran GTPase-regulated nuclear-cytoplasmic transport (NCT). The loading of Ran with GTP, which is mediated by RCC1, the guanine nucleotide exchange factor for Ran, is critical for NCT activity. However, the role of RCC1 or Ran⋅GTP in promoting cell proliferation or DDR is not clear. We show that RCC1 overexpression in normal cells increased cellular Ran⋅GTP levels and accelerated the cell cycle and DNA damage repair. As a result, normal cells overexpressing RCC1 evaded DNA damage-induced cell cycle arrest and senescence, mimicking colorectal carcinoma cells with high endogenous RCC1 levels. The RCC1-induced inhibition of senescence required Ran and exportin 1 and involved the activation of importin β-dependent nuclear import of 53BP1, a large NCT cargo. Our results indicate that changes in the activity of the Ran⋅GTP-regulated NCT modulate the rate of the cell cycle and the efficiency of DNA repair. Through the essential role of RCC1 in regulation of cellular Ran⋅GTP levels and NCT, RCC1 expression enables the proliferation of cells that sustain DNA damage. PMID:26864624

  14. RCC1-dependent activation of Ran accelerates cell cycle and DNA repair, inhibiting DNA damage–induced cell senescence

    PubMed Central

    Cekan, Pavol; Hasegawa, Keisuke; Pan, Yu; Tubman, Emily; Odde, David; Chen, Jin-Qiu; Herrmann, Michelle A.; Kumar, Sheetal; Kalab, Petr

    2016-01-01

    The coordination of cell cycle progression with the repair of DNA damage supports the genomic integrity of dividing cells. The function of many factors involved in DNA damage response (DDR) and the cell cycle depends on their Ran GTPase–regulated nuclear–cytoplasmic transport (NCT). The loading of Ran with GTP, which is mediated by RCC1, the guanine nucleotide exchange factor for Ran, is critical for NCT activity. However, the role of RCC1 or Ran⋅GTP in promoting cell proliferation or DDR is not clear. We show that RCC1 overexpression in normal cells increased cellular Ran⋅GTP levels and accelerated the cell cycle and DNA damage repair. As a result, normal cells overexpressing RCC1 evaded DNA damage–induced cell cycle arrest and senescence, mimicking colorectal carcinoma cells with high endogenous RCC1 levels. The RCC1-induced inhibition of senescence required Ran and exportin 1 and involved the activation of importin β–dependent nuclear import of 53BP1, a large NCT cargo. Our results indicate that changes in the activity of the Ran⋅GTP–regulated NCT modulate the rate of the cell cycle and the efficiency of DNA repair. Through the essential role of RCC1 in regulation of cellular Ran⋅GTP levels and NCT, RCC1 expression enables the proliferation of cells that sustain DNA damage. PMID:26864624

  15. Induction of Apoptosis and Antiproliferative Activity of Naringenin in Human Epidermoid Carcinoma Cell through ROS Generation and Cell Cycle Arrest

    PubMed Central

    Jafri, Asif; Ahmad, Sheeba; Afzal, Mohammad; Arshad, Md

    2014-01-01

    A natural predominant flavanone naringenin, especially abundant in citrus fruits, has a wide range of pharmacological activities. The search for antiproliferative agents that reduce skin carcinoma is a task of great importance. The objective of this study was to analyze the anti-proliferative and apoptotic mechanism of naringenin using MTT assay, DNA fragmentation, nuclear condensation, change in mitochondrial membrane potential, cell cycle kinetics and caspase-3 as biomarkers and to investigate the ability to induce reactive oxygen species (ROS) initiating apoptotic cascade in human epidermoid carcinoma A431 cells. Results showed that naringenin exposure significantly reduced the cell viability of A431 cells (p<0.01) with a concomitant increase in nuclear condensation and DNA fragmentation in a dose dependent manner. The intracellular ROS generation assay showed statistically significant (p<0.001) dose-related increment in ROS production for naringenin. It also caused naringenin-mediated epidermoid carcinoma apoptosis by inducing mitochondrial depolarization. Cell cycle study showed that naringenin induced cell cycle arrest in G0/G1 phase of cell cycle and caspase-3 analysis revealed a dose dependent increment in caspase-3 activity which led to cell apoptosis. This study confirms the efficacy of naringenin that lead to cell death in epidermoid carcinoma cells via inducing ROS generation, mitochondrial depolarization, nuclear condensation, DNA fragmentation, cell cycle arrest in G0/G1 phase and caspase-3 activation. PMID:25330158

  16. c-Myc activates multiple metabolic networks to generate substrates for cell-cycle entry.

    SciTech Connect

    Morrish, Fionnuala M.; Isern, Nancy; Sadilek, Martin; Jeffrey, Mark; Hockenbery, David M.

    2009-05-18

    Cell proliferation requires the coordinated activity of cytosolic and mitochondrial metabolic pathways to provide ATP and building blocks for DNA, RNA, and protein synthesis. Many metabolic pathway genes are targets of the c-myc oncogene and cell cycle regulator. However, the contribution of c-Myc to the activation of cytosolic and mitochondrial metabolic networks during cell cycle entry is unknown. Here, we report the metabolic fates of [U-13C] glucose in serum-stimulated myc-/- and myc+/+ fibroblasts by 13C isotopomer NMR analysis. We demonstrate that endogenous c-myc increased 13C-labeling of ribose sugars, purines, and amino acids, indicating partitioning of glucose carbons into C1/folate and pentose phosphate pathways, and increased tricarboxylic acid cycle turnover at the expense of anaplerotic flux. Myc expression also increased global O-linked GlcNAc protein modification, and inhibition of hexosamine biosynthesis selectively reduced growth of Myc-expressing cells, suggesting its importance in Myc-induced proliferation. These data reveal a central organizing role for the Myc oncogene in the metabolism of cycling cells. The pervasive deregulation of this oncogene in human cancers may be explained by its role in directing metabolic networks required for cell proliferation.

  17. Postnatal telomere dysfunction induces cardiomyocyte cell-cycle arrest through p21 activation.

    PubMed

    Aix, Esther; Gutiérrez-Gutiérrez, Óscar; Sánchez-Ferrer, Carlota; Aguado, Tania; Flores, Ignacio

    2016-06-01

    The molecular mechanisms that drive mammalian cardiomyocytes out of the cell cycle soon after birth remain largely unknown. Here, we identify telomere dysfunction as a critical physiological signal for cardiomyocyte cell-cycle arrest. We show that telomerase activity and cardiomyocyte telomere length decrease sharply in wild-type mouse hearts after birth, resulting in cardiomyocytes with dysfunctional telomeres and anaphase bridges and positive for the cell-cycle arrest protein p21. We further show that premature telomere dysfunction pushes cardiomyocytes out of the cell cycle. Cardiomyocytes from telomerase-deficient mice with dysfunctional telomeres (G3 Terc(-/-)) show precocious development of anaphase-bridge formation, p21 up-regulation, and binucleation. In line with these findings, the cardiomyocyte proliferative response after cardiac injury was lost in G3 Terc(-/-) newborns but rescued in G3 Terc(-/-)/p21(-/-) mice. These results reveal telomere dysfunction as a crucial signal for cardiomyocyte cell-cycle arrest after birth and suggest interventions to augment the regeneration capacity of mammalian hearts. PMID:27241915

  18. Glutathione cycle activity and pyridine nucleotide levels in oxidant-induced injury of cells.

    PubMed Central

    Schraufstätter, I U; Hinshaw, D B; Hyslop, P A; Spragg, R G; Cochrane, C G

    1985-01-01

    Exposure of target cells to a bolus of H2O2 induced cell lysis after a latent period of several hours, which was prevented only when the H2O2 was removed within the first 30 min of injury by addition of catalase. This indicated that early metabolic events take place that are important in the fate of the cell exposed to oxidants. In this study, we described two early and independent events of H2O2-induced injury in P388D1 macrophagelike tumor cells: activation of the glutathione cycle and depletion of cellular NAD. Glutathione cycle and hexose monophosphate shunt (HMPS) were activated within seconds after the addition of H2O2. High HMPS activity maintained glutathione that was largely reduced. However, when HMPS activity was inhibited--by glucose depletion or by incubation at 4 degrees C--glutathione remained in the oxidized state. Total pyridine nucleotide levels were diminished when cells were exposed to H2O2, and the breakdown product, nicotinamide, was recovered in the extracellular medium. Intracellular NAD levels fell by 80% within 20 min of exposure of cells to H2O2. The loss of NADP(H) and stimulation of the HMPS could be prevented when the glutathione cycle was inhibited by either blocking glutathione synthesis with buthionine sulfoximine (BSO) or by inhibiting glutathione reductase with (1,3-bis) 2 chlorethyl-1-nitrosourea. The loss of NAD developed independently of glutathione cycle and HMPS activity, as it also occurred in BSO-treated cells. PMID:3840176

  19. Conditional inactivation of PDCD2 induces p53 activation and cell cycle arrest

    PubMed Central

    Granier, Celine J.; Wang, Wei; Tsang, Tiffany; Steward, Ruth; Sabaawy, Hatem E.; Bhaumik, Mantu; Rabson, Arnold B.

    2014-01-01

    ABSTRACT PDCD2 (programmed cell death domain 2) is a highly conserved, zinc finger MYND domain-containing protein essential for normal development in the fly, zebrafish and mouse. The molecular functions and cellular activities of PDCD2 remain unclear. In order to better understand the functions of PDCD2 in mammalian development, we have examined PDCD2 activity in mouse blastocyst embryos, as well as in mouse embryonic stem cells (ESCs) and embryonic fibroblasts (MEFs). We have studied mice bearing a targeted PDCD2 locus functioning as a null allele through a splicing gene trap, or as a conditional knockout, by deletion of exon2 containing the MYND domain. Tamoxifen-induced knockout of PDCD2 in MEFs, as well as in ESCs, leads to defects in progression from the G1 to the S phase of cell cycle, associated with increased levels of p53 protein and p53 target genes. G1 prolongation in ESCs was not associated with induction of differentiation. Loss of entry into S phase of the cell cycle and marked induction of nuclear p53 were also observed in PDCD2 knockout blastocysts. These results demonstrate a unique role for PDCD2 in regulating the cell cycle and p53 activation during early embryonic development of the mouse. PMID:25150276

  20. Conditional inactivation of PDCD2 induces p53 activation and cell cycle arrest.

    PubMed

    Granier, Celine J; Wang, Wei; Tsang, Tiffany; Steward, Ruth; Sabaawy, Hatem E; Bhaumik, Mantu; Rabson, Arnold B

    2014-01-01

    PDCD2 (programmed cell death domain 2) is a highly conserved, zinc finger MYND domain-containing protein essential for normal development in the fly, zebrafish and mouse. The molecular functions and cellular activities of PDCD2 remain unclear. In order to better understand the functions of PDCD2 in mammalian development, we have examined PDCD2 activity in mouse blastocyst embryos, as well as in mouse embryonic stem cells (ESCs) and embryonic fibroblasts (MEFs). We have studied mice bearing a targeted PDCD2 locus functioning as a null allele through a splicing gene trap, or as a conditional knockout, by deletion of exon2 containing the MYND domain. Tamoxifen-induced knockout of PDCD2 in MEFs, as well as in ESCs, leads to defects in progression from the G1 to the S phase of cell cycle, associated with increased levels of p53 protein and p53 target genes. G1 prolongation in ESCs was not associated with induction of differentiation. Loss of entry into S phase of the cell cycle and marked induction of nuclear p53 were also observed in PDCD2 knockout blastocysts. These results demonstrate a unique role for PDCD2 in regulating the cell cycle and p53 activation during early embryonic development of the mouse. PMID:25150276

  1. Inhibition of protein kinase B activity induces cell cycle arrest and apoptosis during early G₁ phase in CHO cells.

    PubMed

    van Opstal, Angélique; Bijvelt, José; van Donselaar, Elly; Humbel, Bruno M; Boonstra, Johannes

    2012-04-01

    Inhibition of PKB (protein kinase B) activity using a highly selective PKB inhibitor resulted in inhibition of cell cycle progression only if cells were in early G1 phase at the time of addition of the inhibitor, as demonstrated by time-lapse cinematography. Addition of the inhibitor during mitosis up to 2 h after mitosis resulted in arrest of the cells in early G1 phase, as deduced from the expression of cyclins D and A and incorporation of thymidine. After 24 h of cell cycle arrest, cells expressed the cleaved caspase-3, a central mediator of apoptosis. These results demonstrate that PKB activity in early G1 phase is required to prevent the induction of apoptosis. Using antibodies, it was demonstrated that active PKB translocates to the nucleus during early G1 phase, while an even distribution of PKB was observed through cytoplasm and nucleus during the end of G1 phase. PMID:22251027

  2. Cell cycle effect on the activity of deoxynucleoside analogue metabolising enzymes

    SciTech Connect

    Fyrberg, Anna; Albertioni, Freidoun; Lotfi, Kourosh . E-mail: koulo@imv.liu.se

    2007-06-15

    Deoxynucleoside analogues (dNAs) are cytotoxic towards both replicating and indolent malignancies. The impact of fluctuations in the metabolism of dNAs in relation to cell cycle could have strong implications regarding the activity of dNAs. Deoxycytidine kinase (dCK) and deoxyguanosine kinase (dGK) are important enzymes for phosphorylation/activation of dNAs. These drugs can be dephosphorylated/deactivated by 5'-nucleotidases (5'-NTs) and elevated activities of 5'-NTs and decreased dCK and/or dGK activities represent resistance mechanisms towards dNAs. The activities of dCK, dGK, and three 5'-NTs were investigated in four human leukemic cell lines in relationship to cell cycle progression and cytotoxicity of dNAs. Synchronization of cell cultures to arrest in G0/G1 by serum-deprivation was performed followed by serum-supplementation for cell cycle progression. The activities of dCK and dGK increased up to 3-fold in CEM, HL60, and MOLT-4 cells as they started to proliferate, while the activity of cytosolic nucleotidase I was reduced in proliferating cells. CEM, HL60, and MOLT-4 cells were also more sensitive to cladribine, cytarabine, 9-{beta}-D-arabinofuranosylguanine and clofarabine than K562 cells which demonstrated lower levels and less alteration of these enzymes and were least susceptible to the cytotoxic effects of most dNAs. The results suggest that, in the cell lines studied, the proliferation process is associated with a general shift in the direction of activation of dNAs by inducing activities of dCK/dGK and reducing the activity of cN-I which is favourable for the cytotoxic effects of cladribine, cytarabine and, 9-{beta}-D-arabinofuranosylguanine. These results emphasize the importance of cellular proliferation and dNA metabolism by both phosphorylation and dephosphorylation for susceptibility to dNAs. It underscores the need to understand the mechanisms of action and resistance to dNAs in order to increase efficacy of dNAs treatment by new rational.

  3. Natural Compounds' Activity against Cancer Stem-Like or Fast-Cycling Melanoma Cells

    PubMed Central

    Majchrzak, Kinga; Hartman, Mariusz; Czyz, Malgorzata

    2014-01-01

    Background Accumulating evidence supports the concept that melanoma is highly heterogeneous and sustained by a small subpopulation of melanoma stem-like cells. Those cells are considered as responsible for tumor resistance to therapies. Moreover, melanoma cells are characterized by their high phenotypic plasticity. Consequently, both melanoma stem-like cells and their more differentiated progeny must be eradicated to achieve durable cure. By reevaluating compounds in heterogeneous melanoma populations, it might be possible to select compounds with activity not only against fast-cycling cells but also against cancer stem-like cells. Natural compounds were the focus of the present study. Methods We analyzed 120 compounds from The Natural Products Set II to identify compounds active against melanoma populations grown in an anchorage-independent manner and enriched with cells exerting self-renewing capacity. Cell viability, cell cycle arrest, apoptosis, gene expression, clonogenic survival and label-retention were analyzed. Findings Several compounds efficiently eradicated cells with clonogenic capacity and nanaomycin A, streptonigrin and toyocamycin were effective at 0.1 µM. Other anti-clonogenic but not highly cytotoxic compounds such as bryostatin 1, siomycin A, illudin M, michellamine B and pentoxifylline markedly reduced the frequency of ABCB5 (ATP-binding cassette, sub-family B, member 5)-positive cells. On the contrary, treatment with maytansine and colchicine selected for cells expressing this transporter. Maytansine, streptonigrin, toyocamycin and colchicine, even if highly cytotoxic, left a small subpopulation of slow-dividing cells unaffected. Compounds selected in the present study differentially altered the expression of melanocyte/melanoma specific microphthalmia-associated transcription factor (MITF) and proto-oncogene c-MYC. Conclusion Selected anti-clonogenic compounds might be further investigated as potential adjuvants targeting melanoma stem

  4. Cell cycle-specific cleavage of Scc2 regulates its cohesin deposition activity

    PubMed Central

    Woodman, Julie; Fara, Tyler; Dzieciatkowska, Monika; Trejo, Michael; Luong, Nancy; Hansen, Kirk C.; Megee, Paul C.

    2014-01-01

    Sister chromatid cohesion (SCC), efficient DNA repair, and the regulation of some metazoan genes require the association of cohesins with chromosomes. Cohesins are deposited by a conserved heterodimeric loading complex composed of the Scc2 and Scc4 proteins in Saccharomyces cerevisiae, but how the Scc2/Scc4 deposition complex regulates the spatiotemporal association of cohesin with chromosomes is not understood. We examined Scc2 chromatin association during the cell division cycle and found that the affinity of Scc2 for chromatin increases biphasically during the cell cycle, increasing first transiently in late G1 phase and then again later in G2/M. Inactivation of Scc2 following DNA replication reduces cellular viability, suggesting that this post S-phase increase in Scc2 chromatin binding affinity is biologically relevant. Interestingly, high and low Scc2 chromatin binding levels correlate strongly with the presence of full-length or amino-terminally cleaved forms of Scc2, respectively, and the appearance of the cleaved Scc2 species is promoted in vitro either by treatment with specific cell cycle-staged cellular extracts or by dephosphorylation. Importantly, Scc2 cleavage eliminates Scc2–Scc4 physical interactions, and an scc2 truncation mutant that mimics in vivo Scc2 cleavage is defective for cohesin deposition. These observations suggest a previously unidentified mechanism for the spatiotemporal regulation of cohesin association with chromosomes through cell cycle regulation of Scc2 cohesin deposition activity by Scc2 dephosphorylation and cleavage. PMID:24778232

  5. Cell-cycle-regulated activation of Akt kinase by phosphorylation at its carboxyl terminus

    PubMed Central

    Liu, Pengda; Begley, Michael; Michowski, Wojciech; Inuzuka, Hiroyuki; Ginzberg, Miriam; Gao, Daming; Tsou, Peiling; Gan, Wenjian; Papa, Antonella; Kim, Byeong Mo; Wan, Lixin; Singh, Amrik; Zhai, Bo; Yuan, Min; Wang, Zhiwei; Gygi, Steven P.; Lee, Tae Ho; Lu, Kun-Ping; Toker, Alex; Pandolfi, Pier Paolo; Asara, John M.; Kirschner, Marc W.; Sicinski, Piotr; Cantley, Lewis; Wei, Wenyi

    2014-01-01

    Akt, also known as protein kinase B, plays key roles in cell proliferation, survival and metabolism. Akt hyperactivation contributes to many pathophysiological conditions, including human cancers1–3, and is closely associated with poor prognosis and chemo- or radio-therapeutic resistance4. Phosphorylation of Akt at S473 (ref. 5) and T308 (ref. 6) activates Akt. However, it remains unclear whether further mechanisms account for full Akt activation, and whether Akt hyperactivation is linked to misregulated cell cycle progression, another cancer hallmark7. Here we report that Akt activity fluctuates across the cell cycle, mirroring cyclin A expression. Mechanistically, phosphorylation of S477 and T479 at the Akt extreme carboxy terminus by cyclin-dependent kinase 2 (Cdk2)/cyclin A or mTORC2, under distinct physiological conditions, promotes Akt activation through facilitating, or functionally compensating for, S473 phosphorylation. Furthermore, deletion of the cyclin A2 allele in the mouse olfactory bulb leads to reduced S477/T479 phosphorylation and elevated cellular apoptosis. Notably, cyclin A2-deletion-induced cellular apoptosis in mouse embryonic stem cells is partly rescued by S477D/T479E-Akt1, supporting a physiological role for cyclin A2 in governing Akt activation. Together, the results of our study show Akt S477/T479 phosphorylation to be an essential layer of the Akt activation mechanism to regulate its physiological functions, thereby providing a new mechanistic link between aberrant cell cycle progression and Akt hyperactivation in cancer. PMID:24670654

  6. Oscillation of APC/C activity during cell cycle arrest promotes centrosome amplification

    PubMed Central

    Prosser, Suzanna L.; Samant, Mugdha D.; Baxter, Joanne E.; Morrison, Ciaran G.; Fry, Andrew M.

    2014-01-01

    Centrosome duplication is licensed by the disengagement, or ‘uncoupling’, of centrioles during late mitosis. However, arrest of cells in G2 can trigger premature centriole disengagement. Here, we show that premature disengagement results from untimely activation of the APC/C leading to securin degradation and release of active separase. APC/C activation during G2 arrest is dependent on Plk1-mediated degradation of the APC/C inhibitor, Emi1, but Plk1 also has a second APC/C-independent role in promoting disengagement. Importantly, APC/C and Plk1 activity also stimulate centriole disengagement in response to hydroxyurea or DNA damage-induced cell cycle arrest and this leads to centrosome amplification. However, the re-duplication of disengaged centrioles is dependent on Cdk2 activity and Cdk2 activation coincides with a subsequent inactivation of the APC/C and re-accumulation of cyclin A. Release from these arrests leads to mitotic entry but, due to the presence of disengaged and/or amplified centrosomes, formation of abnormal mitotic spindles that lead to chromosome missegregation. Thus, oscillation of APC/C activity during cell cycle arrest promotes both centrosome amplification and genome instability. PMID:22956538

  7. RSS1 regulates the cell cycle and maintains meristematic activity under stress conditions in rice

    PubMed Central

    Ogawa, Daisuke; Abe, Kiyomi; Miyao, Akio; Kojima, Mikiko; Sakakibara, Hitoshi; Mizutani, Megumi; Morita, Haruka; Toda, Yosuke; Hobo, Tokunori; Sato, Yutaka; Hattori, Tsukaho; Hirochika, Hirohiko; Takeda, Shin

    2011-01-01

    Plant growth and development are sustained by continuous cell division in the meristems, which is perturbed by various environmental stresses. For the maintenance of meristematic functions, it is essential that cell division be coordinated with cell differentiation. However, it is unknown how the proliferative activities of the meristems and the coordination between cell division and differentiation are maintained under stressful conditions. Here we show that a rice protein, RSS1, whose stability is controlled by cell cycle phases, contributes to the vigour of meristematic cells and viability under salinity conditions. These effects of RSS1 are exerted by regulating the G1–S transition, possibly through an interaction of RSS1 with protein phosphatase 1, and are mediated by the phytohormone, cytokinin. RSS1 is conserved widely in plant lineages, except eudicots, suggesting that RSS1-dependent mechanisms might have been adopted in specific lineages during the evolutionary radiation of angiosperms. PMID:21505434

  8. E. adenophorum Induces Cell Cycle and Apoptosis of Renal Cells through Mitochondrial Pathway and Caspase Activation in Saanen Goat

    PubMed Central

    Hu, Yanchun; Luo, Biao; Wu, Lei; Qiao, Yan; Mo, Quan; Xu, Ruiguang; Zhou, Yancheng; Ren, Zhihua; Zuo, Zhicai; Deng, Junliang; Peng, Guangneng; He, Wei; Wei, Yahui

    2015-01-01

    The cytotoxicity effects of E. adenophorum on cell cycle and apoptosis of renal cells in Saanen goat was evaluated by TUNEL, DAPI, AO/EB staining, DNA fragmentation assay, Caspase activity, Western-blot, qRT-PCR and flow cytometry analysis. 16 saanen goats randomly divided into four groups were fed on 0%, 40%, 60% and 80% E. adenophorum diets. The Results showed that E. adenophorum induced typical apoptotic features of renal cells. E. adenophorum significantly suppressed renal cells viability, caused cell cycle activity arrest and induced typical apoptotic features in a dose-dependent manner. However, the protein levels of Fas/FasL, Bid and caspase-8 did not appear significant changes in the process of E. adenophorum-induced apoptosis. Moreover, E. adenophorum administration slightly decreased Bcl-2 expression, promoted Bax translocation to mitochondria, triggered the release of Cyt c from mitochondria into cytosol and activated caspase-9, -3, and cleaved PARP. The mitochondrial p53 translocation was significantly activated, accompanied by a significant increase in the loss of ΔΨm, Cyt c release and caspase-9 activation. Above all, these data suggest that E. adenophorum induces renal cells apoptosis via the activation of mitochondria-mediated apoptosis pathway in renal cells. These findings may provide new insights to understand the mechanisms involved in E. adenophorum-caused cytotoxicity of renal cells. PMID:26382060

  9. Impact of Mitochondria-Mediated Apoptosis in U251 Cell Cycle Arrest in G1 Stage and Caspase Activation

    PubMed Central

    Zhang, Lei; Liang, Peng; Zhang, Rui

    2015-01-01

    Background Most mitochondria-mediated apoptosis has some relevance to the cell cycle, but there is still a lack of investigations about U251 cell cycle in human brain glioma cells. In this study, we aimed to clarify the correlation of mitochondria-mediated apoptosis with the U251 cell cycle and its influence on apoptosis, through observing the impact of mitochondria-mediated apoptosis in U251cell specificity cycle arrest and Caspase activation. Material/Methods AnnexinV/PI and API were used to label the brain glioma cells for flow cytometry analysis of U251 cell apoptosis and cell cycle. RT-PCR and Western blot were performed to detect Caspase-3 and Caspase-9 activation. Results Peripheral blood in stationary phase is not sensitive to apoptosis induction, but U251 cells have obvious apoptosis. Mitochondria-mediated apoptosis mainly occurs in the G1 phase of the cell cycle. Caspase-3 and Caspase-9 mRNAs and proteins expression increased significantly after the cells were treated by mitochondrial apoptosis-related gene Bax induction. Conclusions Mitochondria-mediated apoptosis is related to the U251 cell cycle with specific G1 stage arrest. Caspase activation occurs in the process of cell apoptosis. PMID:26594875

  10. Omacetaxine mepesuccinate induces apoptosis and cell cycle arrest, promotes cell differentiation, and reduces telomerase activity in diffuse large B-cell lymphoma cells

    PubMed Central

    ZHANG, LINA; CHEN, ZHENZHU; ZUO, WENLI; ZHU, XINGHU; LI, YUFU; LIU, XINJIAN; WEI, XUDONG

    2016-01-01

    Clinical studies have demonstrated that omacetaxine mepesuccinate exerts beneficial effects on acute myelogenous leukemia. It has been suggested that omacetaxine mepesuccinate, used alone or with interferon-α or cytarabine, induces remission in patients with chronic myelogenous leukemia. These effects are possibly mediated by its ability to induce apoptosis of leukemia cells and inhibit the activity of telomerase. To determine whether omacetaxine mepesuccinate is beneficial in diffuse large B-cell lymphoma (DLBCL), two DLBCL cell lines [a germinal center B cell-like subtype (GCB) and an activated B cell-like subtype (ABC)] were treated with omacetaxine mepesuccinate at various concentrations for different durations. The present study indicated that omacetaxine mepesuccinate exerts proapoptotic effects in the two cell types in a dose- and time-dependent manner. The ABC subtype demonstrated increased sensitivity compared with the GCB subtype. At 40 ng/ml, omacetaxine mepesuccinate exhibited a marked proapoptotic effect on DLBCL cells compared with the other tumor cells investigated. Furthermore, omacetaxine mepesuccinate induced cell cycle arrest at G0/G1 phase, and promoted cell terminal differentiation of pro-B cells. The present study also demonstrated that omacetaxine mepesuccinate exerted its antitumor effect by reducing telomerase activity. In conclusion, the present study demonstrated that omacetaxine mepesuccinate may induce apoptosis and cell cycle arrest, promote cell differentiation, and reduce telomerase activity in DLBCL cells, thus aiding the development of omacetaxine mepesuccinate-based DLBCL therapeutic strategies. PMID:26935769

  11. Suppression of cell cycle progression by a fungal lectin: activation of cyclin-dependent kinase inhibitors.

    PubMed

    Liua, W; Ho, J C; Ng, T

    2001-01-01

    The antiproliferative activity of a fungal lectin (VVL) isolated from the mushroom, Volvariella volvacea, was studied using a battery of cultured tumor cell lines. It was revealed that [(3)H]thymidine incorporation into the cell lines was markedly reduced at 0.32 microM VVL. When S180 mouse sarcoma cells were incubated for 48 hr with doses of VVL ranging from 0.32 to 0.8 microM, prominent blebs on the cell surface and large vacuoles in the cytoplasm, but not apoptotic bodies, were observed under a fluorescence microscopy. VVL did not exert ribosome-inactivating activity or induce any changes in the expression of cyclins A, D1, and E. However, it did activate the expression of cyclin kinase inhibitors, namely p21, p27, p53, and Rb, in a dose-dependent manner. Flow cytometric analysis demonstrated an accumulation of cells in the G2/M phase in a time- and dose-dependent manner, indicating that VVL arrested cell proliferation by blocking cell cycle progression in the G2/M phase. PMID:11137706

  12. Effectiveness and Student Perceptions of an Active Learning Activity Using a Headline News Story to Enhance In-Class Learning of Cell Cycle Regulation

    ERIC Educational Resources Information Center

    Dirks-Naylor, Amie J.

    2016-01-01

    An active learning activity was used to engage students and enhance in-class learning of cell cycle regulation in a PharmD level integrated biological sciences course. The aim of the present study was to determine the effectiveness and perception of the in-class activity. After completion of a lecture on the topic of cell cycle regulation,…

  13. Omacetaxine mepesuccinate induces apoptosis and cell cycle arrest, promotes cell differentiation, and reduces telomerase activity in diffuse large B‑cell lymphoma cells.

    PubMed

    Zhang, Lina; Chen, Zhenzhu; Zuo, Wenli; Zhu, Xinghu; Li, Yufu; Liu, Xinjian; Wei, Xudong

    2016-04-01

    Clinical studies have demonstrated that omacetaxine mepesuccinate exerts beneficial effects on acute myelogenous leukemia. It has been suggested that omacetaxine mepesuccinate, used alone or with interferon‑α or cytarabine, induces remission in patients with chronic myelogenous leukemia. These effects are possibly mediated by its ability to induce apoptosis of leukemia cells and inhibit the activity of telomerase. To determine whether omacetaxine mepesuccinate is beneficial in diffuse large B‑cell lymphoma (DLBCL), two DLBCL cell lines [a germinal center B cell‑like subtype (GCB) and an activated B cell‑like subtype (ABC)] were treated with omacetaxine mepesuccinate at various concentrations for different durations. The present study indicated that omacetaxine mepesuccinate exerts proapoptotic effects in the two cell types in a dose‑ and time‑dependent manner. The ABC subtype demonstrated increased sensitivity compared with the GCB subtype. At 40 ng/ml, omacetaxine mepesuccinate exhibited a marked proapoptotic effect on DLBCL cells compared with the other tumor cells investigated. Furthermore, omacetaxine mepesuccinate induced cell cycle arrest at G0/G1 phase, and promoted cell terminal differentiation of pro‑B cells. The present study also demonstrated that omacetaxine mepesuccinate exerted its antitumor effect by reducing telomerase activity. In conclusion, the present study demonstrated that omacetaxine mepesuccinate may induce apoptosis and cell cycle arrest, promote cell differentiation, and reduce telomerase activity in DLBCL cells, thus aiding the development of omacetaxine mepesuccinate‑based DLBCL therapeutic strategies. PMID:26935769

  14. Piperine Causes G1 Phase Cell Cycle Arrest and Apoptosis in Melanoma Cells through Checkpoint Kinase-1 Activation

    PubMed Central

    Fofaria, Neel M.; Kim, Sung-Hoon; Srivastava, Sanjay K.

    2014-01-01

    In this study, we determined the cytotoxic effects of piperine, a major constituent of black and long pepper in melanoma cells. Piperine treatment inhibited the growth of SK MEL 28 and B16 F0 cells in a dose and time-dependent manner. The growth inhibitory effects of piperine were mediated by cell cycle arrest of both the cell lines in G1 phase. The G1 arrest by piperine correlated with the down-regulation of cyclin D1 and induction of p21. Furthermore, this growth arrest by piperine treatment was associated with DNA damage as indicated by phosphorylation of H2AX at Ser139, activation of ataxia telangiectasia and rad3-related protein (ATR) and checkpoint kinase 1 (Chk1). Pretreatment with AZD 7762, a Chk1 inhibitor not only abrogated the activation of Chk1 but also piperine mediated G1 arrest. Similarly, transfection of cells with Chk1 siRNA completely protected the cells from G1 arrest induced by piperine. Piperine treatment caused down-regulation of E2F1 and phosphorylation of retinoblastoma protein (Rb). Apoptosis induced by piperine was associated with down-regulation of XIAP, Bid (full length) and cleavage of Caspase-3 and PARP. Furthermore, our results showed that piperine treatment generated ROS in melanoma cells. Blocking ROS by tiron protected the cells from piperine mediated cell cycle arrest and apoptosis. These results suggest that piperine mediated ROS played a critical role in inducing DNA damage and activation of Chk1 leading to G1 cell cycle arrest and apoptosis. PMID:24804719

  15. Platelet-activating factor induces cell cycle arrest and disrupts the DNA damage response in mast cells

    PubMed Central

    Puebla-Osorio, N; Damiani, E; Bover, L; Ullrich, S E

    2015-01-01

    Platelet-activating factor (PAF) is a potent phospholipid modulator of inflammation that has diverse physiological and pathological functions. Previously, we demonstrated that PAF has an essential role in ultraviolet (UV)-induced immunosuppression and reduces the repair of damaged DNA, suggesting that UV-induced PAF is contributing to skin cancer initiation by inducing immune suppression and also affecting a proper DNA damage response. The exact role of PAF in modulating cell proliferation, differentiation or transformation is unclear. Here, we investigated the mechanism(s) by which PAF affects the cell cycle and impairs early DNA damage response. PAF arrests proliferation in transformed and nontransformed human mast cells by reducing the expression of cyclin-B1 and promoting the expression of p21. PAF-treated cells show a dose-dependent cell cycle arrest mainly at G2–M, and a decrease in the DNA damage response elements MCPH1/BRIT-1 and ataxia telangiectasia and rad related (ATR). In addition, PAF disrupts the localization of p-ataxia telangiectasia mutated (p-ATM), and phosphorylated-ataxia telangiectasia and rad related (p-ATR) at the site of DNA damage. Whereas the potent effect on cell cycle arrest may imply a tumor suppressor activity for PAF, the impairment of proper DNA damage response might implicate PAF as a tumor promoter. The outcome of these diverse effects may be dependent on specific cues in the microenvironment. PMID:25950475

  16. Downregulation of cell division cycle 25 homolog C reduces the radiosensitivity and proliferation activity of esophageal squamous cell carcinoma.

    PubMed

    Yin, Yachao; Dou, Xiaoyan; Duan, Shimiao; Zhang, Lei; Xu, Quanjing; Li, Hongwei; Li, Duojie

    2016-09-30

    Radiation therapy is one of the most important methods of contemporary cancer treatment. Cells in the G2 and M phases are more sensitive to radiation therapy, and cell division cycle 25 homolog C (CDC25C) is essential in shifting the cell cycle between these two phases. In this study, the knockdown of CDC25C in human esophageal squamous carcinoma EC9706 cells was mediated by transfecting shRNA against human CDC25C-subcloning into pGV248. The levels of CDC25C mRNA and protein expression were assessed by reverse transcription-polymerase chain reaction (RT-PCR) and western blotting, respectively. Moreover, cell proliferation and radiosensitivity were measured. Stable CDC25C-knockdown EC9706 cell lines were successfully established. Furthermore, the proliferation of both control and CDC25C-shRNA-EC9706 cells was inhibited after the cells were treated with increasing X-ray doses, and the proliferation of the control cells was affected more significantly (p<0.05). Moreover, cell colony formation assays allowed us to reach the same conclusion. Taken together, our experiments demonstrated that the knockdown of CDC25C can reduce both the radiotherapy sensitivity and the proliferation activity of EC9706 cells. Thus, CDC25C might be a potential biomarker for radiotherapy treatment. PMID:27188256

  17. Effects of activated fibroblasts on phenotype modulation, EGFR signalling and cell cycle regulation in OSCC cells.

    PubMed

    Berndt, Alexander; Büttner, Robert; Gühne, Stefanie; Gleinig, Anna; Richter, Petra; Chen, Yuan; Franz, Marcus; Liebmann, Claus

    2014-04-01

    Crosstalk between carcinoma associated fibroblasts (CAFs) and oral squamous cell carcinoma (OSCC) cells is suggested to mediate phenotype transition of cancer cells as a prerequisite for tumour progression, to predict patients' outcome, and to influence the efficacy of EGFR inhibitor therapies. Here we investigate the influence of activated fibroblasts as a model for CAFs on phenotype and EGFR signalling in OSCC cells in vitro. For this, immortalised hTERT-BJ1 fibroblasts were activated with TGFβ1 and PDGFAB to generate a myofibroblast or proliferative phenotype, respectively. Conditioned media (FCMTGF, FCMPDGF) were used to stimulate PE/CA-PJ15 OSCC cells. Results were compared to the effect of conditioned media of non-stimulated fibroblasts (FCMB). FCMTGF stimulation leads to an up-regulation of vimentin in the OSCC cells and an enhancement of invasive behaviour, indicating EMT-like effects. Similarly, FCMTGF≫FCMPDGF induced up-regulation of EGFR, but not of ErbB2/ErbB3. In addition, we detected an increase in basal activities of ERK, PI3K/Akt and Stat3 (FCMTGF>FCMPDGF) accompanied by protein interaction of vimentin with pERK. These effects are correlated with an increased proliferation. In summary, our results suggest that the activated myofibroblast phenotype provides soluble factors which are able to induce EMT-like phenomena and to increase EGFR signalling as well as cell proliferation in OSCC cells. Our results indicate a possible influence of activated myofibroblasts on EGFR-inhibitor therapy. Therefore, CAFs may serve as promising novel targets for combined therapy strategies. PMID:24394543

  18. Cell cycle arrest and apoptosis, two alternative mechanisms for PMKT2 killer activity.

    PubMed

    Santos, Antonio; Alonso, Alejandro; Belda, Ignacio; Marquina, Domingo

    2013-01-01

    Pichia membranifaciens CYC 1086 secretes a unique 30kDa killer toxin (PMKT2) that inhibits a variety of spoilage yeasts and fungi of agronomical interest. The cytocidal effect of PMKT2 on Saccharomyces cerevisiae cells was studied. Metabolic events associated with the loss of S. cerevisiae viability caused by PMKT2 were qualitatively identical to those reported for K28 killer toxin activity, but different to those reported for PMKT. At higher doses, none of the cellular events accounting for the action of PMKT, the killer toxin secreted by P. membranifaciens CYC 1106, was observed for PMKT2. Potassium leakage, sodium influx and the decrease of intracellular pH were not among the primary effects of PMKT2. We report here that this protein is unable to form ion-permeable channels in liposome membranes, suggesting that channel formation is not the mechanism of cytotoxic action of PMKT2. Nevertheless, flow cytometry studies have revealed a cell cycle arrest at an early S-phase with an immature bud and pre-replicated 1n DNA content. By testing the sensitivity of cells arrested at different stages in the cell cycle, we hoped to identify the execution point for lethality more precisely. Cells arrested at the G1-phase by α-factor or arrested at G2-phase by the spindle poison methyl benzimidazol-2-yl-carbamate (MBC) were protected against the toxin. Cells released from the arrest in both cases were killed by PMKT2 at a similar rate. Nevertheless, cells released from MBC-arrest were able to grow for a short time, and then viability dropped rapidly. These findings suggest that cells released from G2-phase are initially able to divide, but die in the presence of PMKT2 after initiating the S-phase in a new cycle, adopting a terminal phenotype within that cycle. By contrast, low doses of PMKT and PMKT2 were able to generate the same cellular response. The evidence presented here shows that treating yeast with low doses of PMKT2 leads to the typical membranous, cytoplasmic

  19. Antiproliferative activity of bicyclic benzimidazole nucleosides: synthesis, DNA-binding and cell cycle analysis.

    PubMed

    Sontakke, Vyankat A; Lawande, Pravin P; Kate, Anup N; Khan, Ayesha; Joshi, Rakesh; Kumbhar, Anupa A; Shinde, Vaishali S

    2016-04-26

    An efficient route was developed for synthesis of bicyclic benzimidazole nucleosides from readily available d-glucose. The key reactions were Vörbruggen glycosylation and ring closing metathesis (RCM). Primarily, to understand the mode of DNA binding, we performed a molecular docking study and the binding was found to be in the minor groove region. Based on the proposed binding model, UV-visible and fluorescence spectroscopic techniques using calf thymus DNA (CT-DNA) demonstrated a non-intercalative mode of binding. Antiproliferative activity of nucleosides was tested against MCF-7 and MDA-MB-231 breast cancer cell lines and found to be active at low micromolar concentrations. Compounds and displayed significant antiproliferative activity as compared to and with the reference anticancer drug, doxorubicin. Cell cycle analysis showed that nucleoside induced cell cycle arrest at the S-phase. Confocal microscopy has been performed to validate the induction of cellular apoptosis. Based on these findings, such modified bicyclic benzimidazole nucleosides will make a significant contribution to the development of anticancer drugs. PMID:27074628

  20. Effects of activated fibroblasts on phenotype modulation, EGFR signalling and cell cycle regulation in OSCC cells

    SciTech Connect

    Berndt, Alexander; Büttner, Robert; Gühne, Stefanie; Gleinig, Anna; Richter, Petra; Chen, Yuan; Franz, Marcus; Liebmann, Claus

    2014-04-01

    Crosstalk between carcinoma associated fibroblasts (CAFs) and oral squamous cell carcinoma (OSCC) cells is suggested to mediate phenotype transition of cancer cells as a prerequisite for tumour progression, to predict patients’ outcome, and to influence the efficacy of EGFR inhibitor therapies. Here we investigate the influence of activated fibroblasts as a model for CAFs on phenotype and EGFR signalling in OSCC cells in vitro. For this, immortalised hTERT-BJ1 fibroblasts were activated with TGFβ1 and PDGFAB to generate a myofibroblast or proliferative phenotype, respectively. Conditioned media (FCM{sub TGF}, FCM{sub PDGF}) were used to stimulate PE/CA-PJ15 OSCC cells. Results were compared to the effect of conditioned media of non-stimulated fibroblasts (FCM{sub B}). FCM{sub TGF} stimulation leads to an up-regulation of vimentin in the OSCC cells and an enhancement of invasive behaviour, indicating EMT-like effects. Similarly, FCM{sub TGF}≫FCM{sub PDGF} induced up-regulation of EGFR, but not of ErbB2/ErbB3. In addition, we detected an increase in basal activities of ERK, PI3K/Akt and Stat3 (FCM{sub TGF}>FCM{sub PDGF}) accompanied by protein interaction of vimentin with pERK. These effects are correlated with an increased proliferation. In summary, our results suggest that the activated myofibroblast phenotype provides soluble factors which are able to induce EMT-like phenomena and to increase EGFR signalling as well as cell proliferation in OSCC cells. Our results indicate a possible influence of activated myofibroblasts on EGFR-inhibitor therapy. Therefore, CAFs may serve as promising novel targets for combined therapy strategies. - Highlights: • A cell culture model for cancer associated fibroblasts is described. • The mutual interaction with OSCC cells leads to up-regulation of EGFR in tumour cells. • mCAF induces EGFR downstream signalling with increased proliferation in OSCC. • Erk activation is associated with protein interaction with vimentin

  1. BRCA2 Coordinates the Activities of Cell-Cycle Kinases to Promote Genome Stability

    PubMed Central

    Yata, Keiko; Bleuyard, Jean-Yves; Nakato, Ryuichiro; Ralf, Christine; Katou, Yuki; Schwab, Rebekka A.; Niedzwiedz, Wojciech; Shirahige, Katsuhiko; Esashi, Fumiko

    2014-01-01

    Summary Numerous human genome instability syndromes, including cancer, are closely associated with events arising from malfunction of the essential recombinase Rad51. However, little is known about how Rad51 is dynamically regulated in human cells. Here, we show that the breast cancer susceptibility protein BRCA2, a key Rad51 binding partner, coordinates the activity of the central cell-cycle drivers CDKs and Plk1 to promote Rad51-mediated genome stability control. The soluble nuclear fraction of BRCA2 binds Plk1 directly in a cell-cycle- and CDK-dependent manner and acts as a molecular platform to facilitate Plk1-mediated Rad51 phosphorylation. This phosphorylation is important for enhancing the association of Rad51 with stressed replication forks, which in turn protects the genomic integrity of proliferating human cells. This study reveals an elaborate but highly organized molecular interplay between Rad51 regulators and has significant implications for understanding tumorigenesis and therapeutic resistance in patients with BRCA2 deficiency. PMID:24835992

  2. Benfluron Induces Cell Cycle Arrest, Apoptosis and Activation of p53 Pathway in MOLT-4 Leukemic Cells.

    PubMed

    Seifrtová, M; Cochlarová, T; Havelek, R; Řezáčová, M

    2015-01-01

    The aim of our study was to determine the effect of potential anti-tumour agent benfluron on human leukemic cells MOLT-4 and elucidate the molecular mechanisms of response of tumour cells to this chemotherapeutic agent. It has been shown that the mechanisms of action of benfluron are complex, but the molecular pathways of the cytostatic effect have remained unknown and the present study contributes to their elucidation. In this work, benfluron reduced viability of the treated cells and induced caspase-mediated apoptosis. The programmed cell death was associated with activation of caspases 8, 9 and 3/7. Moreover, exposure of cells to benfluron resulted in accumulation of the cells primarily in late S and G2/M phases. The changes in the levels of key proteins show that benfluron provoked activation of p53 and induced phosphorylation of p53 on serine 15 and serine 392. The application of benfluron led to phosphorylation of Chk1 on serine 345 and phosphorylation of Chk2 on threonine 68 in the treated cells. Higher doses of benfluron caused phosphorylation of ERK1/2 on threonine 202 and tyrosine 204, whereas JNK and p38 kinases were not activated. In conclusion, benfluron induces apoptosis, cell cycle arrest in late S and G2/M phases, and activates various signalling pathways of the DNA damage response. PMID:26441204

  3. Enteric pathogens deploy cell cycle inhibiting factors to block the bactericidal activity of Perforin-2

    PubMed Central

    McCormack, Ryan M; Lyapichev, Kirill; Olsson, Melissa L; Podack, Eckhard R; Munson, George P

    2015-01-01

    Perforin-2 (MPEG1) is an effector of the innate immune system that limits the proliferation and spread of medically relevant Gram-negative, -positive, and acid fast bacteria. We show here that a cullin-RING E3 ubiquitin ligase (CRL) complex containing cullin-1 and βTrCP monoubiquitylates Perforin-2 in response to pathogen associated molecular patterns such as LPS. Ubiquitylation triggers a rapid redistribution of Perforin-2 and is essential for its bactericidal activity. Enteric pathogens such as Yersinia pseudotuberculosis and enteropathogenic Escherichia coli disarm host cells by injecting cell cycle inhibiting factors (Cifs) into mammalian cells to deamidate the ubiquitin-like protein NEDD8. Because CRL activity is dependent upon NEDD8, Cif blocks ubiquitin dependent trafficking of Perforin-2 and thus, its bactericidal activity. Collectively, these studies further underscore the biological significance of Perforin-2 and elucidate critical molecular events that culminate in Perforin-2-dependent killing of both intracellular and extracellular, cell-adherent bacteria. DOI: http://dx.doi.org/10.7554/eLife.06505.001 PMID:26418746

  4. Neferine, an alkaloid from lotus seed embryo, inhibits human lung cancer cell growth by MAPK activation and cell cycle arrest.

    PubMed

    Poornima, Paramasivan; Weng, Ching Feng; Padma, Viswanadha Vijaya

    2014-01-01

    Neferine is the major bisbenzylisoquinoline alkaloid isolated from the seed embryo of a traditional medicinal plant Nelumbo nucifera (Lotus). Epidemiological studies have revealed the therapeutic potential of lotus seed embryo. Although several mechanisms have been proposed, a clear anticancer action mechanism of neferine on lung cancer cells is still not known. Lung cancer is the most common cause of cancer death in the world, and the patients with advanced stage of nonsmall lung cancer require adjunct chemotherapy after surgical resection for the eradication of cancer cells. In this study, the effects of neferine were evaluated and characterized in A549 cells. Neferine induced apoptosis in a dose-dependent manner with the hypergeneration of reactive oxygen species, activation of MAPKs, lipid peroxidation, depletion of cellular antioxidant pool, loss of mitochondrial membrane potential, and intracellular calcium accumulation. Furthermore, neferine treatment leads to the inhibition of nuclear factor kappaB and Bcl2, upregulation of Bax and Bad, release of cytochrome C, activation of caspase cascade, and DNA fragmentation. In addition, neferine could induce p53 and its effector protein p21 and downregulation of cell cycle regulatory protein cyclin D1 thereby inducing G1 cell cycle arrest. These results suggest a novel function of neferine as an apoptosis inducer in lung cancer cells. PMID:23983146

  5. Systematic characterization of cell cycle phase-dependent protein dynamics and pathway activities by high-content microscopy-assisted cell cycle phenotyping.

    PubMed

    Bruhn, Christopher; Kroll, Torsten; Wang, Zhao-Qi

    2014-12-01

    Cell cycle progression is coordinated with metabolism, signaling and other complex cellular functions. The investigation of cellular processes in a cell cycle stage-dependent manner is often the subject of modern molecular and cell biological research. Cell cycle synchronization and immunostaining of cell cycle markers facilitate such analysis, but are limited in use due to unphysiological experimental stress, cell type dependence and often low flexibility. Here, we describe high-content microscopy-assisted cell cycle phenotyping (hiMAC), which integrates high-resolution cell cycle profiling of asynchronous cell populations with immunofluorescence microscopy. hiMAC is compatible with cell types from any species and allows for statistically powerful, unbiased, simultaneous analysis of protein interactions, modifications and subcellular localization at all cell cycle stages within a single sample. For illustration, we provide a hiMAC analysis pipeline tailored to study DNA damage response and genomic instability using a 3-4-day protocol, which can be adjusted to any other cell cycle stage-dependent analysis. PMID:25458086

  6. Prion-induced neurotoxicity: Possible role for cell cycle activity and DNA damage response

    PubMed Central

    Bujdoso, Raymond; Landgraf, Matthias; Jackson, Walker S; Thackray, Alana M

    2015-01-01

    Protein misfolding neurodegenerative diseases arise through neurotoxicity induced by aggregation of host proteins. These conditions include Alzheimer’s disease, Huntington’s disease, Parkinson’s disease, motor neuron disease, tauopathies and prion diseases. Collectively, these conditions are a challenge to society because of the increasing aged population and through the real threat to human food security by animal prion diseases. It is therefore important to understand the cellular and molecular mechanisms that underlie protein misfolding-induced neurotoxicity as this will form the basis for designing strategies to alleviate their burden. Prion diseases are an important paradigm for neurodegenerative conditions in general since several of these maladies have now been shown to display prion-like phenomena. Increasingly, cell cycle activity and the DNA damage response are recognised as cellular events that participate in the neurotoxic process of various neurodegenerative diseases, and their associated animal models, which suggests they are truly involved in the pathogenic process and are not merely epiphenomena. Here we review the role of cell cycle activity and the DNA damage response in neurodegeneration associated with protein misfolding diseases, and suggest that these events contribute towards prion-induced neurotoxicity. In doing so, we highlight PrP transgenic Drosophila as a tractable model for the genetic analysis of transmissible mammalian prion disease. PMID:26279981

  7. TOUSLED Kinase Activity Oscillates during the Cell Cycle and Interacts with Chromatin Regulators1

    PubMed Central

    Ehsan, Hashimul; Reichheld, Jean-Philippe; Durfee, Tim; Roe, Judith L.

    2004-01-01

    The TOUSLED (TSL)-like nuclear protein kinase family is highly conserved in plants and animals. tsl loss of function mutations cause pleiotropic defects in both leaf and flower development, and growth and initiation of floral organ primordia is abnormal, suggesting that basic cellular processes are affected. TSL is more highly expressed in exponentially growing Arabidopsis culture cells than in stationary, nondividing cells. While its expression remains constant throughout the cell cycle in dividing cells, TSL kinase activity is higher in enriched late G2/M-phase and G1-phase populations of Arabidopsis suspension culture cells compared to those in S-phase. tsl mutants also display an aberrant pattern and increased expression levels of the mitotic cyclin gene CycB1;1, suggesting that TSL represses CycB1;1 expression at certain times during development or that cells are delayed in mitosis. TSL interacts with and phosphorylates one of two Arabidopsis homologs of the nucleosome assembly/silencing protein Asf1 and histone H3, as in humans, and a novel plant SANT/myb-domain protein, TKI1, suggesting that TSL plays a role in chromatin metabolism. PMID:15047893

  8. The Chlamydomonas Cell Cycle

    PubMed Central

    Cross, Frederick R.; Umen, James G.

    2015-01-01

    The position of Chlamydomonas within the eukaryotic phylogeny makes it a unique model in at least two important ways: as a representative of the critically important, early-diverging lineage leading to plants, and as a microbe retaining important features of the last eukaryotic common ancestor (LECA) that have been lost in the highly studied yeast lineages. Its cell biology has been studied for many decades, and it has well-developed experimental genetic tools, both classical (Mendelian) and molecular. Unlike land plants, it is a haploid with very few gene duplicates, making it ideal for loss-of-function genetic studies. The Chlamydomonas cell cycle has a striking temporal and functional separation between cell growth and rapid cell divisions, probably connected to the interplay between diurnal cycles that drive photosynthetic cell growth with the cell division cycle; it also exhibits a highly choreographed interaction between the cell cycle and its centriole/basal body/flagellar cycle. Here we review the current status of studies of the Chlamydomonas cell cycle. We begin with an overview of cell cycle control in the well-studied yeast and animal systems, which has yielded a canonical, well-supported model. We discuss briefly what is known about similarities and differences in plant cell cycle control compared to this model. We next review the cytology and cell biology of the multiple fission cell cycle of Chlamydomonas. Lastly we review recent genetic approaches and insights into Chlamydomonas cell cycle regulation that have been enabled by a new generation of genomics-based tools. PMID:25690512

  9. Inflammasome activation by cell volume regulation and inflammation-associated hyponatremia: A vicious cycle.

    PubMed

    Kim, J H; Park, J H; Eisenhut, M; Yu, J W; Shin, J I

    2016-08-01

    Inflammasomes are caspase-1-activating molecular platforms that produce active interleukin (IL)-1β and are implicated in various central nervous system (CNS) diseases. These multi-protein complexes can be activated by exposure of cells to low osmolality. The inflammasome nucleotide-binding and oligomerization domain-like receptor pyrin domain-containing protein 3 (NLRP3) is hereby the main sensor of cellular osmolality. IL-1β was found to stimulate the secretion of antidiuretic hormone (ADH) from the posterior pituitary gland either by action of prostaglandins or indirectly by causing the release of IL-6. Based on these findings, we hypothesize that the hyponatremia caused by a wide range of CNS diseases is able to induce significant cell swelling with induction of a hyposmotic intracellular environment, which activates the NLRP3 inflammasome, causing the release of IL-1β and induced by IL-1β, IL-6, which increases the production of ADH that leads to more profound hyponatremia. Supportive evidence for this hypothesis is the finding that IL-1 injection can induce ADH release and hyposmotic effect of ADH induced hyponatremia can, via the mechanical effect of cell swelling, activate transient receptor potential channels, which via transforming growth factor β-activated kinase 1 activate NLRP3. Implications of this hypothesis, if confirmed, would include that hyponatremia can be exacerbated through this vicious cycle but also that the inflammasomes are key mediators of this process. Confirmation of this hypothesis would have implications for prevention and clinical management of changes in patients sodium levels related to syndrome of inappropriate antidiuretic hormone secretion (SIADH) with interventions targeting inflammatory mediator production and function of inflammasomes with the potential of prevention of permanent brain damage in a wide range of CNS diseases. PMID:27372869

  10. Fluctuation of the CaS -sequestering activity of permeabilized sea urchin embryos during the cell cycle

    SciTech Connect

    Suprynowicz, F.A.; Mazia, D.

    1985-04-01

    The authors have followed the sequestration of CaS by intracellular compartments in sea urchin embryos through the first cell cycles. To gain biochemical access to these compartments, the embryos were permeabilized by brief exposure to an intense electric field. Sequestration was determined as the retention of tracer, UVCa, after filtration of aliquots on Millipore filters. The permeabilized cells sequester CaS at a constant rate for at least 20 min. The CaS -sequestering activities of embryos that are permeabilized at successive stages of the first cell cycle (one-cell stage) progressively increase to 5 times the initial level. The rate of sequestration is maximal during telophase and, in some populations of zygotes, is nearly as great throughout prophase. Over the course of the second cell cycle (two-cell stage), the activity undergoes a 2-fold oscillation that bears the same temporal relationship to mitosis as the previous fluctuation.

  11. Chloroquine enhances the chemotherapeutic activity of 5-fluorouracil in a colon cancer cell line via cell cycle alteration.

    PubMed

    Choi, Jung-Hye; Yoon, Jin Sun; Won, Young-Woong; Park, Byeong-Bae; Lee, Young Yiul

    2012-07-01

    Autophagy is a conserved catabolic process that degrades cytoplasmic proteins and organelles for recycling. The role of autophagy in tumorigenesis is controversial because autophagy can be either protective or damaging to tumor cells, and its effects may change during tumor progression. A number of cancer cell lines have been exposed to chloroquine, an anti-malarial drug, with the aim of inhibiting cell growth and inducing cell death. In addition, chloroquine inhibits a late phase of autophagy. This study was conducted to investigate the anti-cancer effect of autophagy inhibition, using chloroquine together with 5-fluorouracil (5-FU) in a colon cancer cell line. Human colon cancer DLD-1 cells were treated with 5-FU (10 μΜ) or chloroquine (100 μΜ), or a combination of both. Autophagy was evaluated by western blot analysis of microtubule-associated protein light chain3 (LC3). Proliferative activity, alterations of the cell cycle, and apoptosis were measured by MTT assays, flow cytometry, and western blotting. LC3-II protein increased after treatment with 5-FU, and chloroquine potentiated the cytotoxicity of 5-FU. MTT assays showed that 5-FU inhibited proliferation of the DLD-1 cells and that chloroquine enhanced this inhibitory effect of 5-FU. The combination of 5-FU and chloroquine induced G1 arrest, up-regulation of p27 and p53, and down-regulation of CDK2 and cyclin D1. These results suggest that chloroquine may potentiate the anti-cancer effect of 5-FU via cell cycle inhibition. Chloroquine potentiates the anti-cancer effect of 5-FU in colon cancer cells. Supplementation of conventional chemotherapy with chloroquine may provide a new cancer therapy modality. PMID:22716215

  12. Ziyuglycoside II induces cell cycle arrest and apoptosis through activation of ROS/JNK pathway in human breast cancer cells.

    PubMed

    Zhu, Xue; Wang, Ke; Zhang, Kai; Zhu, Ling; Zhou, Fanfan

    2014-05-16

    Ziyuglycoside II, a triterpenoid saponin compound extracted from Sanguisorba officinalis L., has been reported to have a wide range of clinical applications including anti-cancer effect. In this study, the anti-proliferative effect of ziyuglycoside II in two classic human breast cancer cell lines, MCF-7 and MDA-MB-231, was extensively investigated. Our study indicated that ziyuglycoside II could effectively induce G2/M phase arrest and apoptosis in both cell lines. Cell cycle blocking was associated with the down-regulation of Cdc25C, Cdc2, cyclin A and cyclin B1 as well as the up-regulation of p21/WAF1, phospho-Cdc25C and phospho-Cdc2. Ziyuglycoside II treatment also induced reactive oxygen species (ROS) production and apoptosis by activating the extrinsic/Fas/FasL pathway as well as the intrinsic/mitochondrial pathway. More importantly, the c-Jun NH2-terminal kinase (JNK), a downstream target of ROS, was found to be a critical mediator of ziyuglycoside II-induced cell apoptosis. Further knockdown of JNK by siRNA could inhibit ziyuglycoside II-mediated apoptosis with attenuating the up-regulation of Bax and Fas/FasL as well as the down-regulation of Bcl-2. Taken together, the cell death of breast cancer cells in response to ziyuglycoside II was dependent upon cell cycle arrest and cell apoptosis via a ROS-dependent JNK activation pathway. Our findings may significantly contribute to the understanding of the anti-proliferative effect of ziyuglycoside II, in particular to breast carcinoma and provide novel insights into the potential application of such compound in breast cancer therapy. PMID:24680927

  13. JNK is constitutively active in mantle cell lymphoma: cell cycle deregulation and polyploidy by JNK inhibitor SP600125.

    PubMed

    Wang, Miao; Atayar, Cigdem; Rosati, Stefano; Bosga-Bouwer, Anneke; Kluin, Philip; Visser, Lydia

    2009-05-01

    Mantle cell lymphoma (MCL) is characterized by genetic instability and a poor prognosis. Many blastoid variants are (hypo)tetraploid and have an even worse prognosis. We investigated the role of signalling by mitogen-activated protein kinases (MAPKs) in MCL. As compared to normal tonsil B cells, MCL cells showed higher activation of the JNK MAPK in both an MAPK array and a sandwich ELISA assay. Immunohistochemistry showed overexpression of phospho (p)-JNK (Thr183/Tyr185) in 30 of 37 MCL cases. Inhibition of p-JNK with SP600125 resulted in growth arrest in all four MCL cell lines (Jeko-1, HBL-2, UPN-1, Granta-519), which could be partly reversed by the addition of CD40L and IL-4. Furthermore, SP600125 led to G2/M phase arrest on day 1 and a striking increase in endoreduplication on day 2 and day 3, which was confirmed by karyotype analysis. G2/M arrest was associated with down-regulation of EGR1 and p21 protein expression. SP600125-induced polyploidy could be blocked by the BCL-2 inhibitor YC137. These data suggest that constitutive JNK activity is necessary to promote proliferation and maintain diploidy in MCL. JNK inhibition leads to cell cycle deregulation and endoreduplication, mimicking the tetraploid state seen in a subset of MCL cases. Thus, our data also provide an experimental model to study polyploid MCL cells. PMID:19206150

  14. Changes in the activities of key enzymes of glycolysis during the cell cycle in yeast: a rectification.

    PubMed

    de Koning, W; Groeneveld, K; Oehlen, L J; Berden, J A; van Dam, K

    1991-04-01

    Activities of glycolytic enzymes were determined in elutriation fractionated cultures of Saccharomyces cerevisiae grown on different carbon sources. Almost pure fractions of single cells at the G1 state of cell division were obtained for some of the growth conditions tested, whereas other stages were enriched in particular fractions. Specific activities of glucose-6-phosphate dehydrogenase and alcohol dehydrogenase were found to be constant during the cell cycle, as reported by van Doorn et al. (1988a), Journal of Bacteriology 170, 4808-4815, and (1988b), Journal of General Microbiology 134, 785-790. In contrast to the earlier reports, the activities of hexokinase, phosphofructokinase, pyruvate kinase and trehalase were also constant in different states of the cell cycle. For hexokinase and phosphofructokinase it was shown that the apparent specific activity in a cell-free extract strongly diminished when extracts contained less that 0.5-1 mg protein ml-1. In the experiments of van Doorn et al. (1988a) the protein content of the outer fractions was up to 20 times lower than that of the central fractions, suggesting an alternative explanation for the observed changes in enzyme activities during the cell cycle. Therefore, we want to rectify the observations presented by van Doorn et al. (1988a), and conclude that the activities of the glycolytic enzymes do not vary greatly during the cell cycle of S. cervisiae. PMID:1856683

  15. Cell Cycle Regulators Guide Mitochondrial Activity in Radiation-Induced Adaptive Response

    PubMed Central

    Alexandrou, Aris T.

    2014-01-01

    Abstract Significance: There are accruing concerns on potential genotoxic agents present in the environment including low-dose ionizing radiation (LDIR) that naturally exists on earth's surface and atmosphere and is frequently used in medical diagnosis and nuclear industry. Although its long-term health risk is being evaluated and remains controversial, LDIR is shown to induce temporary but significant adaptive responses in mammalian cells and animals. The mechanisms guiding the mitochondrial function in LDIR-induced adaptive response represent a unique communication between DNA damage and cellular metabolism. Elucidation of the LDIR-regulated mitochondrial activity may reveal new mechanisms adjusting cellular function to cope with hazardous environmental stress. Recent Advances: Key cell cycle regulators, including Cyclin D1/CDK4 and Cyclin B1/cyclin-dependent kinase 1 (CDK1) complexes, are actively involved in the regulation of mitochondrial functions via phosphorylation of their mitochondrial targets. Accumulating new evidence supports a concept that the Cyclin B1/CDK1 complex acts as a mediator in the cross talk between radiation-induced DNA damage and mitochondrial functions to coordinate cellular responses to low-level genotoxic stresses. Critical Issues: The LDIR-mediated mitochondrial activity via Cyclin B1/CDK1 regulation is an irreplaceable network that is able to harmonize vital cellular functions with adjusted mitochondrial metabolism to enhance cellular homeostasis. Future Directions: Further investigation of the coordinative mechanism that regulates mitochondrial activities in sublethal stress conditions, including LDIR, will reveal new insights of how cells cope with genotoxic injury and will be vital for future targeted therapeutic interventions that reduce environmental injury and cancer risk. Antioxid. Redox Signal. 20, 1463–1480. PMID:24180340

  16. Aristolochic acid-induced apoptosis and G2 cell cycle arrest depends on ROS generation and MAP kinases activation.

    PubMed

    Romanov, Victor; Whyard, Terry C; Waltzer, Wayne C; Grollman, Arthur P; Rosenquist, Thomas

    2015-01-01

    Ingestion of aristolochic acids (AAs) contained in herbal remedies results in a renal disease and, frequently, urothelial malignancy. The genotoxicity of AA in renal cells, including mutagenic DNA adducts formation, is well documented. However, the mechanisms of AA-induced tubular atrophy and renal fibrosis are largely unknown. To better elucidate some aspects of this process, we studied cell cycle distribution and cell survival of renal epithelial cells treated with AAI at low and high doses. A low dose of AA induces cell cycle arrest in G2/M phase via activation of DNA damage checkpoint pathway ATM-Chk2-p53-p21. DNA damage signaling pathway is activated more likely via increased production of reactive oxygen species (ROS) caused by AA treatment then via DNA damage induced directly by AA. Higher AA concentration induced cell death partly via apoptosis. Since mitogen-activated protein kinases play an important role in cell survival, death and cell cycle progression, we assayed their function in AA-treated renal tubular epithelial cells. ERK1/2 and p38 but not JNK were activated in cells treated with AA. In addition, pharmacological inhibition of ERK1/2 and p38 as well as suppression of ROS generation with N-acetyl-L-cysteine resulted in the partial relief of cells from G2/M checkpoint and a decline of apoptosis level. Cell cycle arrest may be a mechanism for DNA repair, cell survival and reprogramming of epithelial cells to the fibroblast type. An apoptosis of renal epithelial cells at higher AA dose might be necessary to provide space for newly reprogrammed fibrotic cells. PMID:24792323

  17. UVA-activated 8-methoxypsoralen (PUVA) causes G2/M cell cycle arrest in Karpas 299 T-lymphoma cells.

    PubMed

    Bartosová, Jitka; Kuzelová, Katerina; Pluskalová, Michaela; Marinov, Iuri; Halada, Petr; Gasová, Zdenka

    2006-10-01

    We investigated the effect of UVA-activated 8-methoxypsoralen (PUVA) on the cell line Karpas 299 derived from anaplastic large-cell lymphoma (ALCL) expressing chimeric fusion protein nucleophosmin-anaplastic lymphoma kinase (NPM/ALK). NPM/ALK activates phosphatidylinositol 3 kinase (PI3K)/Akt pathway responsible for the cell protection from apoptosis. We found that PUVA treatment first induced G2/M cell cycle arrest resulting in a decrease in the cell proliferation rate. The mitochondrial apoptosis was triggered immediately following PUVA treatment, as we judged from the unmasking of mitochondrial membrane antigen 7A6. However, the mitochondrial membrane depolarization was not observed and caspase-3 was only slightly activated. The late apoptotic events were lacking: neither translocation of phosphatidylserine to the outer side of plasma membrane nor DNA fragmentation occurred. We revealed that PUVA enhanced the expression of peroxiredoxin, stress protein endoplasmin and galectin-3. Galectin-3 has been shown to protect mitochondrial membrane integrity and prevent cytochrome c release thereby blocking the effector stage of apoptosis. We suggest that the elevated level of this protein following PUVA treatment acts in synergy with the constitutively expressed chimeric kinase NPM/ALK to block the apoptosis. PMID:16735125

  18. Members Only: Hypoxia-Induced Cell-Cycle Activation in Cardiomyocytes.

    PubMed

    Sharma, Arun; Wu, Sean M

    2015-09-01

    A low level of cardiomyocyte turnover occurs in the adult mammalian heart, but the source of these new cells remains unknown. Kimura et al., 2015 utilized a novel hypoxia-induced fate mapping system to identify a rare population of adult cardiomyocytes undergoing cell-cycle entry and expansion in healthy adult hearts and following ischemic injury. PMID:26331604

  19. The flavonoid quercetin transiently inhibits the activity of taxol and nocodazole through interference with the cell cycle

    PubMed Central

    Samuel, Temesgen; Fadlalla, Khalda; Turner, Timothy; Yehualaeshet, Teshome E.

    2010-01-01

    Quercetin is a flavonoid with anticancer properties. In this study, we examined the effects of quercetin on cell cycle, viability and proliferation of cancer cells, either singly or in combination with the microtubule-targeting drugs taxol and nocodazole. Although quercetin induced cell death in a dose dependent manner, 12.5-50μM quercetin inhibited the activity of both taxol and nocodazole to induce G2/M arrest in various cell lines. Quercetin also partially restored drug-induced loss in viability of treated cells for up to 72 hours. This antagonism of microtubule-targeting drugs was accompanied by a delay in cell cycle progression and inhibition of the buildup of cyclin-B1 at the microtubule organizing center of treated cells. However, quercetin did not inhibit the microtubule targeting of taxol or nocodazole. Despite the short-term protection of cells by quercetin, colony formation and clonogenicity of HCT116 cells were still suppressed by quercetin or quercetin-taxol combination. The status of cell adherence to growth matrix was critical in determining the sensitivity of HCT116 cells to quercetin. We conclude that while long-term exposure of cancer cells to quercetin may prevent cell proliferation and survival, the interference of quercetin with cell cycle progression diminishes the efficacy of microtubule-targeting drugs to arrest cells at G2/M. PMID:21058190

  20. Waves of Cdk1 Activity in S Phase Synchronize the Cell Cycle in Drosophila Embryos.

    PubMed

    Deneke, Victoria E; Melbinger, Anna; Vergassola, Massimo; Di Talia, Stefano

    2016-08-22

    Embryos of most metazoans undergo rapid and synchronous cell cycles following fertilization. While diffusion is too slow for synchronization of mitosis across large spatial scales, waves of Cdk1 activity represent a possible process of synchronization. However, the mechanisms regulating Cdk1 waves during embryonic development remain poorly understood. Using biosensors of Cdk1 and Chk1 activities, we dissect the regulation of Cdk1 waves in the Drosophila syncytial blastoderm. We show that Cdk1 waves are not controlled by the mitotic switch but by a double-negative feedback between Cdk1 and Chk1. Using mathematical modeling and surgical ligations, we demonstrate a fundamental distinction between S phase Cdk1 waves, which propagate as active trigger waves in an excitable medium, and mitotic Cdk1 waves, which propagate as passive phase waves. Our findings show that in Drosophila embryos, Cdk1 positive feedback serves primarily to ensure the rapid onset of mitosis, while wave propagation is regulated by S phase events. PMID:27554859

  1. Solar activity secular cycles

    NASA Astrophysics Data System (ADS)

    Kramynin, A. P.; Mordvinov, A. V.

    2013-12-01

    Long-term variations in solar activity secular cycles have been studied using a method for the expansion of reconstructed sunspot number series Sn( t) for 11400 years in terms of natural orthogonal functions. It has been established that three expansion components describe more than 98% of all Sn( t) variations. In this case, the contribution of the first expansion component is about 92%. The averaged form of the 88year secular cycle has been determined based on the form of the first expansion coordinate function. The quasi-periodicities modulating the secular cycle have been revealed based on the time function conjugate to the first function. The quasi-periodicities modulating the secular cycle coincide with those observed in the Sn( t) series spectrum. A change in the secular cycle form and the time variations in this form are described by the second and third expansion components, the contributions of which are about 4 and 2%, respectively. The variations in the steepness of the secular cycle branches are more pronounced in the 200-year cycle, and the secular cycle amplitude varies more evidently in the 2300-year cycle.

  2. Changes of Constituents and Activity to Apoptosis and Cell Cycle During Fermentation of Tea

    PubMed Central

    Zhao, Hang; Zhang, Min; Zhao, Lu; Ge, Ya-kun; Sheng, Jun; Shi, Wei

    2011-01-01

    Tea is believed to be beneficial for health, and the effects of the fermentation process on its contributions to apoptosis and cell cycle arrest of gastric cancer cells have not been completely investigated. In this study, the chemical components in green tea, black tea and pu-erh tea aqueous extracts were analyzed and compared. The polysaccharide and caffeine levels were substantially higher in the fermented black tea and pu-erh tea, while the polyphenol level was higher in the unfermented green tea. Hence, a treatment of tea aqueous extract and the components, which are emerging as promising anticancer agents, were pursued to determine whether this treatment could lead to enhance apoptosis and cell cycle arrest. In the human gastric cancer cell line SGC-7901, the cell viability and flow cytometry analysis for apoptotic cells indicated effects in a dose-dependent inhibition manner for the three tea treatment groups. The apoptosis rates were found to be elevated after 48 h of treatment with 31.2, 125, and 500 μg/mL of green tea extract, the higher catechins content may be involved in the mechanism. Cell cycle was arrested in S phase in the fermented black tea and pu-erh tea, and the populations were significantly decreased in G2/M phases, possibly due to the oxidation of tea polyphenols, which causes an increase of theabrownins. CCC-HEL-1 normal cells were not sensitive to tea extract. These findings suggest that the fermentation process causes changes of the compounds which might be involved in the changes of cell proliferation inhibition, apoptosis induction and cell cycle arrest. PMID:21673927

  3. Myc and cell cycle control.

    PubMed

    Bretones, Gabriel; Delgado, M Dolores; León, Javier

    2015-05-01

    Soon after the discovery of the Myc gene (c-Myc), it became clear that Myc expression levels tightly correlate to cell proliferation. The entry in cell cycle of quiescent cells upon Myc enforced expression has been described in many models. Also, the downregulation or inactivation of Myc results in the impairment of cell cycle progression. Given the frequent deregulation of Myc oncogene in human cancer it is important to dissect out the mechanisms underlying the role of Myc on cell cycle control. Several parallel mechanisms account for Myc-mediated stimulation of the cell cycle. First, most of the critical positive cell cycle regulators are encoded by genes induced by Myc. These Myc target genes include Cdks, cyclins and E2F transcription factors. Apart from its direct effects on the transcription, Myc is able to hyperactivate cyclin/Cdk complexes through the induction of Cdk activating kinase (CAK) and Cdc25 phosphatases. Moreover, Myc antagonizes the activity of cell cycle inhibitors as p21 and p27 through different mechanisms. Thus, Myc is able to block p21 transcription or to induce Skp2, a protein involved in p27 degradation. Finally, Myc induces DNA replication by binding to replication origins and by upregulating genes encoding proteins required for replication initiation. Myc also regulates genes involved in the mitotic control. A promising approach to treat tumors with deregulated Myc is the synthetic lethality based on the inhibition of Cdks. Thus, the knowledge of the Myc-dependent cell cycle regulatory mechanisms will help to discover new therapeutic approaches directed against malignancies with deregulated Myc. This article is part of a Special Issue entitled: Myc proteins in cell biology and pathology. PMID:24704206

  4. New activators and inhibitors in the hair cycle clock: targeting stem cells' state of competence.

    PubMed

    Plikus, Maksim V

    2012-05-01

    The timing mechanism of the hair cycle remains poorly understood. However, it has become increasingly clear that the telogen-to-anagen transition is controlled jointly by at least the bone morphogenic protein (BMP), WNT, fibroblast growth factor (FGF), and transforming growth factor (TGF)-β signaling pathways. New research shows that Fgf18 signaling in hair follicle stem cells synergizes BMP-mediated refractivity, whereas Tgf-β2 signaling counterbalances it. Loss of Fgf18 signaling markedly accelerates anagen initiation, whereas loss of Tgf-β2 signaling significantly delays it, supporting key roles for these pathways in hair cycle timekeeping. PMID:22499035

  5. Specific cell cycle synchronization with butyrate and cell cycle analysis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Synchronized cells have been invaluable for many kinds of cell cycle and cell proliferation studies. Butyrate induces cell cycle arrest and apoptosis in MDBK cells. To explore the possibility of using butyrate-blocked cells to obtain synchronized cells, we investigated the property of the cell cyc...

  6. Ecdysteroid promotes cell cycle progression in the Bombyx wing disc through activation of c-Myc.

    PubMed

    Moriyama, Minoru; Osanai, Kohji; Ohyoshi, Tomokazu; Wang, Hua-Bing; Iwanaga, Masashi; Kawasaki, Hideki

    2016-03-01

    Developmental switching from growth to metamorphosis in imaginal primordia is an essential process of adult body planning in holometabolous insects. Although it is disciplined by a sequential action of the ecdysteroid, molecular mechanisms linking to cell proliferation are poorly understood. In the present study, we investigated the expression control of cell cycle-related genes by the ecdysteroid using the wing disc of the final-instar larvae of the silkworm, Bombyx mori. We found that the expression level of c-myc was remarkably elevated in the post-feeding cell proliferation phase, which coincided with a small increase in ecdysteroid titer. An in vitro wing disc culture showed that supplementation of the moderate level of the ecdysteroid upregulated c-myc expression within an hour and subsequently increased the expression of cell cycle core regulators, including A-, B-, D-, and E-type cyclin genes, Cdc25 and E2F1. We demonstrated that c-myc upregulation by the ecdysteroid was not inhibited in the presence of a protein synthesis inhibitor, suggesting a possibility that the ecdysteroid directly stimulates c-myc expression. Finally, results from the administration of a c-Myc inhibitor demonstrated that c-Myc plays an essential role in 20E-inducible cell proliferation. These findings suggested a novel pathway for ecdysteroid-inducible cell proliferation in insects, and it is likely to be conserved between insects and mammals in terms of steroid hormone regulation. PMID:26696544

  7. DNA Damage Response Checkpoint Activation Drives KP1019 Dependent Pre-Anaphase Cell Cycle Delay in S. cerevisiae

    PubMed Central

    Bierle, Lindsey A.; Reich, Kira L.; Taylor, Braden E.; Blatt, Eliot B.; Middleton, Sydney M.; Burke, Shawnecca D.; Stultz, Laura K.; Hanson, Pamela K.; Partridge, Janet F.; Miller, Mary E.

    2015-01-01

    Careful regulation of the cell cycle is required for proper replication, cell division, and DNA repair. DNA damage–including that induced by many anticancer drugs–results in cell cycle delay or arrest, which can allow time for repair of DNA lesions. Although its molecular mechanism of action remains a matter of debate, the anticancer ruthenium complex KP1019 has been shown to bind DNA in biophysical assays and to damage DNA of colorectal and ovarian cancer cells in vitro. KP1019 has also been shown to induce mutations and induce cell cycle arrest in Saccharomyces cerevisiae, suggesting that budding yeast can serve as an appropriate model for characterizing the cellular response to the drug. Here we use a transcriptomic approach to verify that KP1019 induces the DNA damage response (DDR) and find that KP1019 dependent expression of HUG1 requires the Dun1 checkpoint; both consistent with KP1019 DDR in budding yeast. We observe a robust KP1019 dependent delay in cell cycle progression as measured by increase in large budded cells, 2C DNA content, and accumulation of Pds1 which functions to inhibit anaphase. Importantly, we also find that deletion of RAD9, a gene required for the DDR, blocks drug-dependent changes in cell cycle progression, thereby establishing a causal link between the DDR and phenotypes induced by KP1019. Interestingly, yeast treated with KP1019 not only delay in G2/M, but also exhibit abnormal nuclear position, wherein the nucleus spans the bud neck. This morphology correlates with short, misaligned spindles and is dependent on the dynein heavy chain gene DYN1. We find that KP1019 creates an environment where cells respond to DNA damage through nuclear (transcriptional changes) and cytoplasmic (motor protein activity) events. PMID:26375390

  8. "Constructing" the Cell Cycle in 3D

    ERIC Educational Resources Information Center

    Koc, Isil; Turan, Merve

    2012-01-01

    The cycle of duplication and division, known as the "cell cycle," is the essential mechanism by which all living organisms reproduce. This activity allows students to develop an understanding of the main events that occur during the typical eukaryotic cell cycle mostly in the process of mitotic phase that divides the duplicated genetic material…

  9. Cell Cycle Regulation and Melanoma.

    PubMed

    Xu, Wen; McArthur, Grant

    2016-06-01

    Dysregulation of cell cycle control is a hallmark of melanomagenesis. Agents targeting the G1-S and G2-M checkpoints, as well as direct anti-mitotic agents, have all shown promising preclinical activity in melanoma. However, in vivo, standalone single agents targeting cell cycle regulation have only demonstrated modest efficacy in unselected patients. The advent of specific CDK 4/6 inhibitors targeting the G1-S transition, with an improved therapeutic index, is a significant step forward. Potential synergy exists with the combination of CDK4/6 inhibitors with existing therapies targeting the MAPK pathway, particularly in subsets of metastatic melanomas such as NRAS and BRAF mutants. This reviews summaries of the latest developments in both preclinical and clinical data with cell cycle-targeted therapies in melanoma. PMID:27106898

  10. Local RhoA activation induces cytokinetic furrows independent of spindle position and cell cycle stage.

    PubMed

    Wagner, Elizabeth; Glotzer, Michael

    2016-06-20

    The GTPase RhoA promotes contractile ring assembly and furrow ingression during cytokinesis. Although many factors that regulate RhoA during cytokinesis have been characterized, the spatiotemporal regulatory logic remains undefined. We have developed an optogenetic probe to gain tight spatial and temporal control of RhoA activity in mammalian cells and demonstrate that cytokinetic furrowing is primarily regulated at the level of RhoA activation. Light-mediated recruitment of a RhoGEF domain to the plasma membrane leads to rapid induction of RhoA activity, leading to assembly of cytokinetic furrows that partially ingress. Furthermore, furrow formation in response to RhoA activation is not temporally or spatially restricted. RhoA activation is sufficient to generate furrows at both the cell equator and cell poles, in both metaphase and anaphase. Remarkably, furrow formation can be initiated in rounded interphase cells, but not adherent cells. These results indicate that RhoA activation is sufficient to induce assembly of functional contractile rings and that cell rounding facilitates furrow formation. PMID:27298323

  11. Force-specific activation of Smad1/5 regulates vascular endothelial cell cycle progression in response to disturbed flow.

    PubMed

    Zhou, Jing; Lee, Pei-Ling; Tsai, Chien-Sung; Lee, Chih-I; Yang, Tung-Lin; Chuang, Han-Sheng; Lin, Wei-Wen; Lin, Ting-Er; Lim, Seh Hong; Wei, Shu-Yi; Chen, Yuh-Lien; Chien, Shu; Chiu, Jeng-Jiann

    2012-05-15

    Vascular endothelial cells (ECs) are constantly exposed to blood flow-induced shear stress, but the mechanism of force-specific activation of their signaling to modulate cellular function remains unclear. We have demonstrated that bone morphogenetic protein receptor (BMPR)-specific Smad1/5 can be force-specifically activated by oscillatory shear stress (OSS) in ECs to cause cell cycle progression. Smad1/5 is highly activated in ECs of atherosclerotic lesions in diseased human coronary arteries from patients with end-stage heart failure undergoing heart transplantation and from apolipoprotein E-deficient mice. Application of OSS (0.5 ± 4 dyn/cm(2)) causes the sustained activation of Smad1/5 in ECs through activations of mammalian target of rapamycin and p70S6 kinase, leading to up-regulation of cyclin A and down-regulations of p21(CIP1) and p27(KIP1) and, hence, EC cycle progression. En face examination of rat aortas reveals high levels of phospho-Smad1/5 in ECs of the inner, but not the outer, curvature of aortic arch, nor the straight segment of thoracic aorta [corrected]. Immunohistochemical and en face examinations of the experimentally stenosed abdominal aorta in rats show high levels of phospho-Smad1/5 in ECs at poststenotic sites, where OSS occurs. These OSS activations of EC Smad1/5 in vitro and in vivo are not inhibited by the BMP-specific antagonist Noggin and, hence, are independent of BMP ligand. Transfecting ECs with Smad1/5-specific small interfering RNAs inhibits the OSS-induced EC cycle progression. Our findings demonstrate the force-specificity of the activation of Smad1/5 and its contribution to cell cycle progression in ECs induced by disturbed flow. PMID:22550179

  12. Dichlorodiphenyltrichloroethane technical mixture regulates cell cycle and apoptosis genes through the activation of CAR and ERα in mouse livers

    SciTech Connect

    Kazantseva, Yuliya A.; Yarushkin, Andrei A.; Pustylnyak, Vladimir O.

    2013-09-01

    Dichlorodiphenyltrichloroethane (DDT) is a widely used organochlorine pesticide and a xenoestrogen that promotes rodent hepatomegaly and tumours. A recent study has shown significant correlation between DDT serum concentration and liver cancer incidence in humans, but the underlying mechanisms remain elusive. We hypothesised that a mixture of DDT isomers could exert effects on the liver through pathways instead of classical ERs. The acute effects of a DDT mixture containing the two major isomers p,p′-DDT (85%) and o,p′-DDT (15%) on CAR and ERα receptors and their cell cycle and apoptosis target genes were studied in mouse livers. ChIP results demonstrated increased CAR and ERα recruitment to their specific target gene binding sites in response to the DDT mixture. The results of real-time RT-PCR were consistent with the ChIP data and demonstrated that the DDT was able to activate both CAR and ERα in mouse livers, leading to target gene transcriptional increases including Cyp2b10, Gadd45β, cMyc, Mdm2, Ccnd1, cFos and E2f1. Western blot analysis demonstrated increases in cell cycle progression proteins cMyc, Cyclin D1, CDK4 and E2f1 and anti-apoptosis proteins Mdm2 and Gadd45β. In addition, DDT exposure led to Rb phosphorylation. Increases in cell cycle progression and anti-apoptosis proteins were accompanied by a decrease in p53 content and its transcriptional activity. However, the DDT was unable to stimulate the β-catenin signalling pathway, which can play an important role in hepatocyte proliferation. Thus, our results indicate that DDT treatment may result in cell cycle progression and apoptosis inhibition through CAR- and ERα-mediated gene activation in mouse livers. These findings suggest that the proliferative and anti-apoptotic conditions induced by CAR and ERα activation may be important contributors to the early stages of hepatocarcinogenesis as produced by DDT in rodent livers. - Highlights: • DDT activated both CAR and ERα and their cell

  13. Triptolide abrogates growth of colon cancer and induces cell cycle arrest by inhibiting transcriptional activation of E2F.

    PubMed

    Oliveira, Amanda R; Beyer, Georg; Chugh, Rohit; Skube, Steven J; Majumder, Kaustav; Banerjee, Sulagna; Sangwan, Veena; Li, Lihua; Dawra, Rajinder K; Subramanian, Subbaya; Saluja, Ashok K; Dudeja, Vikas

    2015-06-01

    Despite significant progress in diagnostics and therapeutics, over 50 thousand patients die from colorectal cancer annually. Hence, there is urgent need for new lines of treatment. Triptolide, a natural compound isolated from the Chinese herb Tripterygium wilfordii, is effective against multiple cancers. We have synthesized a water soluble analog of triptolide, named Minnelide, which is currently in phase I trial against pancreatic cancer. The aims of the current study were to evaluate whether triptolide/Minnelide is effective against colorectal cancer and to elucidate the mechanism by which triptolide induces cell death in colorectal cancer. Efficacy of Minnelide was evaluated in subcutaneous xenograft and liver metastasis model of colorectal cancer. For mechanistic studies, colon cancer cell lines HCT116 and HT29 were treated with triptolide and the effect on viability, caspase activation, annexin positivity, lactate dehydrogenase release, and cell cycle progression was evaluated. Effect of triptolide on E2F transcriptional activity, mRNA levels of E2F-dependent genes, E2F1- retinoblastoma protein (Rb) binding, and proteins levels of regulator of G1-S transition was also measured. DNA binding of E2F1 was evaluated by chromatin immunoprecipitation assay. Triptolide decreased colon cancer cell viability in a dose- and time-dependent fashion. Minnelide markedly inhibited the growth of colon cancer in the xenograft and liver metastasis model of colon cancer and more than doubles the median survival of animals with liver metastases from colon cancer. Mechanistically, we demonstrate that at low concentrations triptolide induces apoptotic cell death but at higher concentrations it induces cell cycle arrest. Our data suggest that triptolide is able to induce G1 cell cycle arrest by inhibiting transcriptional activation of E2F1. Our data also show that triptolide downregulates E2F activity by potentially modulating events downstream of DNA binding. Therefore, we conclude

  14. Triptolide Abrogates Growth of Colon Cancer and Induces Cell Cycle Arrest by Inhibiting Transcriptional Activation of E2F

    PubMed Central

    Chugh, Rohit; Skube, Steven J; Majumder, Kaustav; Banerjee, Sulagna; Sangwan, Veena; Li, Lihua; Dawra, Rajinder; Subramanian, Subbaya; Saluja, Ashok; Dudeja, Vikas

    2016-01-01

    Background Despite significant progress in diagnostics and therapeutics, over fifty thousand patients die from colorectal cancer annually. Hence there is urgent need for new lines of treatment. Triptolide, a natural compound isolated from the Chinese herb Tripterygium wilfordii, is effective against multiple cancers. We have synthesized a water soluble analog of triptolide, named Minnelide, which is currently in phase I trial against pancreatic cancer. The aims of the current study were to evaluate whether triptolide/Minnelide is effective against colorectal cancer and to elucidate the mechanism by which triptolide induces cell death in colorectal cancer. Methods Efficacy of Minnelide was evaluated in subcutaneous xenograft and liver metastasis model of colorectal cancer. For mechanistic studies colon cancer cell lines HCT116 and HT29 were treated with triptolide and the effect on viability, caspase activation, annexin positivity, lactate dehydrogenase(LDH) release and cell cycle progression was evaluated. Effect of triptolide on E2F transcriptional activity, mRNA levels of E2F dependent genes, E2F1-Rb binding and proteins levels of regulator of G1-S transition was also measured. DNA binding of E2F1 was evaluated by chromatin immunoprecipitation assay. Results Triptolide decreased colon cancer cell viability in a dose- and time-dependent fashion. Minnelide markedly inhibited the growth of colon cancer in the xenograft and liver metastasis model of colon cancer and more than doubles the median survival of animals with liver metastases from colon cancer. Mechanistically we demonstrate that at low concentrations, triptolide induces apoptotic cell death but at higher concentrations it induces cell cycle arrest. Our data suggest that triptolide is able to induce G1 cell cycle arrest by inhibiting transcriptional activation of E2F1. Our data also show that triptolide downregulates E2F activity by potentially modulating events downstream of DNA binding. Conclusion

  15. Fisetin inhibits the activities of cyclin-dependent kinases leading to cell cycle arrest in HT-29 human colon cancer cells.

    PubMed

    Lu, Xianghua; Jung, Jae in; Cho, Han Jin; Lim, Do Young; Lee, Hyun Sook; Chun, Hyang Sook; Kwon, Dae Young; Park, Jung Han Yoon

    2005-12-01

    Fisetin, a natural flavonol present in edible vegetables, fruits, and wine, was reported to exert anticarcinogenic effects. The objective of the current study was to examine the effect of fisetin on the cell cycle progression of the human colon cancer cell line HT-29. HT-29 cells were cultured in serum-free medium with 0, 20, 40, or 60 micromol/L fisetin. Fisetin dose dependently inhibited both cell growth and DNA synthesis (P < 0.05), with a 79 +/- 1% decrease in cell number observed 72 h after the addition of 60 micromol/L fisetin. Perturbed cell cycle progression from the G(1) to S phase was observed at 8 h with 60 micromol/L fisetin treatment, whereas a G(2)/M phase arrest was observed after 24 h (P < 0.05). The phosphorylation state of the retinoblastoma proteins shifted from hyperphosphorylated to hypophosphorylated in cells treated with 40 micromol/L fisetin. (P < 0.05). Fisetin decreased the activities of cyclin-dependent kinases (CDK)2 and CDK4; these effects were likely attributable to decreases in the levels of cyclin E and D1 and an increase in p21(CIP1/WAF1) levels (P < 0.05). However, fisetin also inhibited CDk4 activity in a cell-free system (P < 0.05), indicating that it may directly inhibit CDk4 activity. The protein levels of cell division cycles (CDC)2 and CDC25C and the activity of CDC2 were also decreased in fisetin-treated cells (P < 0.05). These results indicate that inhibition of cell cycle progression in HT-29 cells after treatment with fisetin can be explained, at least in part, by modification of CDK activities. PMID:16317137

  16. Umbelliferone exhibits anticancer activity via the induction of apoptosis and cell cycle arrest in HepG2 hepatocellular carcinoma cells.

    PubMed

    Yu, Shi-Min; Hu, Dong-Hui; Zhang, Jian-Jun

    2015-09-01

    Hepatocellular carcinoma (HCC) is a highly malignant tumor, associated with poor patient prognoses, and high rates of morbidity and mortality. To date, the therapeutic strategies available for the treatment of HCC remain limited. The present study aimed to elucidate the anticancer activity of umbelliferone, a naturally occurring coumarin derivative isolated from Ferula communis, against the HepG2 HCC cell line. A 3‑(4,5‑dimthylthaizol‑2‑yl)‑2,5, diphenyltetrazolium bromide assay was used to evaluate cell viability following umbelliferone treatment, and the effects of umbelliferone on cell cycle progression and apoptosis were evaluated using flow cytometry. The presence of morphological features characteristic of apoptosis, including cell shrinkage, membrane blebbing, nuclear condensation and apoptotic body formation, were evaluated in HepG2 cells following umbelliferone treatment. Cell cycle analysis conducted via propidium iodide (PI) staining indicated that umbelliferone treatment induced cell cycle arrest at S phase in HepG2 cells. Analysis with Annexin V and PI staining revealed that umbelliferone induced apoptotic events in HepG2 cells in a concentration‑dependant manner (0‑50 µM). Umbelliferone also induced dose‑dependant DNA fragmentation. In conclusion, umbelliferone was found to exhibit significant anticancer effects via the induction of apoptosis, cell cycle arrest and DNA fragmentation in HepG2 cancer cells. PMID:25997538

  17. Characterization of Photosystem II Activity and Heterogeneity during the Cell Cycle of the Green Alga Scenedesmus quadricauda1

    PubMed Central

    Kaftan, David; Meszaros, Tibor; Whitmarsh, John; Nedbal, Ladislav

    1999-01-01

    The photosynthetic activity of the green alga Scenedesmus quadricauda was investigated during synchronous growth in light/dark cycles. The rate of O2 evolution increased 2-fold during the first 3 to 4 h of the light period, remained high for the next 3 to 4 h, and then declined during the last half of the light period. During cell division, which occurred at the beginning of the dark period, the ability of the cells to evolve O2 was at a minimum. To determine if photosystem II (PSII) controls the photosynthetic capacity of the cells during the cell cycle we measured PSII activity and heterogeneity. Measurements of electron-transport activity revealed two populations of PSII, active centers that contribute to carbon reduction and inactive centers that do not. Measurements of PSII antenna sizes also revealed two populations, PSIIα and PSIIβ, which differ from one another by their antenna size. During the early light period the photosynthetic capacity of the cells doubled, the O2-evolving capacity of PSII was nearly constant, the proportion of PSIIβ centers decreased to nearly zero, and the proportion of inactive PSII centers remained constant. During the period of minimum photosynthetic activity 30% of the PSII centers were insensitive to the inhibitor 3-(3,4-dichlorophenyl)-1,1-dimethylurea, which may be related to reorganization of the thylakoid membrane. We conclude from these results that PSII does not limit the photosynthetic activity of the cells during the first half of the light period. However, the decline in photosynthetic activity observed during the last half of the light period can be accounted for by limited PSII activity. PMID:10364394

  18. Activity Cycles in Stars

    NASA Technical Reports Server (NTRS)

    Hathaway, David H.

    2009-01-01

    Starspots and stellar activity can be detected in other stars using high precision photometric and spectrometric measurements. These observations have provided some surprises (starspots at the poles - sunspots are rarely seen poleward of 40 degrees) but more importantly they reveal behaviors that constrain our models of solar-stellar magnetic dynamos. The observations reveal variations in cycle characteristics that depend upon the stellar structure, convection zone dynamics, and rotation rate. In general, the more rapidly rotating stars are more active. However, for stars like the Sun, some are found to be inactive while nearly identical stars are found to be very active indicating that periods like the Sun's Maunder Minimum (an inactive period from 1645 to 1715) are characteristic of Sun-like stars.

  19. New chlorogenin hexasaccharide isolated from Agave fourcroydes with cytotoxic and cell cycle inhibitory activities.

    PubMed

    Ohtsuki, Takashi; Koyano, Takashi; Kowithayakorn, Thaworn; Sakai, Shinobu; Kawahara, Nobuo; Goda, Yukihiro; Yamaguchi, Naoto; Ishibashi, Masami

    2004-07-15

    A new chlorogenin hexasaccharide (1) was isolated from leaves of Agave fourcroydes (Agavaceae). The structure of the new saponin was elucidated as chlorogenin 3-O-[alpha-L-rhamnopyranosyl-(1-->4)-beta-D-glucopyranosyl-(1-->3)-[beta-D-glucopyranosyl-(1-->3)-beta-D-glucopyranosyl-(1-->2)]-beta-D-glucopyranosyl-(1-->4)-beta-D-galactopyranoside] (1) by spectroscopic analysis and the result of acidic hydrolysis. The new saponin (1) as well as known hexasaccharides (3 and 5) isolated here showed cytotoxicity against HeLa cells, and 1 exhibited a cell cycle inhibitory effect at the G2/M stage at the concentration of 7.5 and 10 microg/mL. PMID:15210151

  20. Prazosin displays anticancer activity against human prostate cancers: targeting DNA and cell cycle.

    PubMed

    Lin, Ssu-Chia; Chueh, Shih-Chieh; Hsiao, Che-Jen; Li, Tsia-Kun; Chen, Tzu-Hsuan; Liao, Cho-Hwa; Lyu, Ping-Chiang; Guh, Jih-Hwa

    2007-10-01

    Quinazoline-based alpha1-adrenoceptor antagonists, in particular doxazosin and terazosin, are suggested to display antineoplastic activity against prostate cancers. However, there are few studies elucidating the effect of prazosin. In this study, prazosin displayed antiproliferative activity superior to that of other alpha1-blockers, including doxazosin, terazosin, tamsulosin, and phentolamine. Prazosin induced G2 checkpoint arrest and subsequent apoptosis in prostate cancer PC-3, DU-145, and LNCaP cells. In p53-null PC-3 cells, prazosin induced an increase in DNA strand breaks and ATM/ATR checkpoint pathways, leading to the activation of downstream signaling cascades, including Cdc25c phosphorylation at Ser216, nuclear export of Cdc25c, and cyclin-dependent kinase (Cdk) 1 phosphorylation at Tyr15. The data, together with sustained elevated cyclin A levels (other than cyclin B1 levels), suggested that Cdk1 activity was inactivated by prazosin. Moreover, prazosin triggered mitochondria-mediated and caspase-executed apoptotic pathways in PC-3 cells. The oral administration of prazosin significantly reduced tumor mass in PC-3-derived cancer xenografts in nude mice. In summary, we suggest that prazosin is a potential antitumor agent that induces cell apoptosis through the induction of DNA damage stress, leading to Cdk1 inactivation and G2 checkpoint arrest. Subsequently, mitochondria-mediated caspase cascades are triggered to induce apoptosis in PC-3 cells. PMID:17971903

  1. Mitochondrial adenosine triphosphatase of the fission yeast, Schizosaccharomyces pombe 972h-. Changes in activity and oligomycin-sensitivity during the cell cycle of catabolite-repressed and -de-repressed cells.

    PubMed Central

    Edwards, S W; Lloyd, D

    1977-01-01

    1. Changes in activity of ATPase (adenosine triphosphatase) during the cell cycle of Schizosaccharomyces pombe were analysed in cell-free extracts of cells harvested from different stages of growth of synchronous cultures and also after cell-cycle fractionation. 2. Oligomycin-sensitive ATPase oscillates in both glucose-repressed synchronous cultures and shows four maxima of activity approximately equally spaced through the cell cycle. The amplitude of the oscillations accounts for between 13 and 80% of the total activity at different times in the cell cycle. 3. Oligomycin sensitivity varies over a fourfold range at different stages of the cell cycle. 4. The periodicity of maximum oligomycin sensitivity is one-quarter of a cell cycle. 5. These results were confirmed for the first three-quarters of the cell cycle by cell-cycle fractionation. 6. In cells growing synchronously with glycerol, ATPase activity increases in a stepwise pattern, with two steps per cell cycle; the first of these occurs at 0.54 of the cell cycle and the second at 0.95. 7. These results are discussed in relation to previously obtained data on the development of mitochondrial activities during the cell cycle. PMID:139890

  2. Effectiveness and student perceptions of an active learning activity using a headline news story to enhance in-class learning of cell cycle regulation.

    PubMed

    Dirks-Naylor, Amie J

    2016-06-01

    An active learning activity was used to engage students and enhance in-class learning of cell cycle regulation in a PharmD level integrated biological sciences course. The aim of the present study was to determine the effectiveness and perception of the in-class activity. After completion of a lecture on the topic of cell cycle regulation, students completed a 10-question multiple-choice quiz before and after engaging in the activity. The activity involved reading of a headline news article published by ScienceDaily.com entitled "One Gene Lost Equals One limb Regained." The name of the gene was deleted from the article and, thus, the end goal of the activity was to determine the gene of interest by the description in the story. The activity included compiling a list of all potential gene candidates before sufficient information was given to identify the gene of interest (p21). A survey was completed to determine student perceptions of the activity. Quiz scores improved by an average of 20% after the activity (40.1 ± 1.95 vs. 59.9 ± 2.14,P< 0.0001,n= 96). Students enjoyed the activity, found the news article interesting, and believed that the activity improved their understanding of cell cycle regulation. The majority of students agreed that the in-class activity piqued their interest for learning the subject matter and also agreed that if they understand a concept during class, they are more likely to want to study that concept outside of class. In conclusion, the activity improved in-class understanding and enhanced interest in cell cycle regulation. PMID:27068993

  3. Characterization of functionally active interleukin-18/eGFP fusion protein expression during cell cycle phases in recombinant chicken DF1 Cells.

    PubMed

    Wu, Hsing Chieh; Chen, Yu San; Shien, Jui Hung; Shen, Pin Chun; Lee, Long Huw

    2016-05-01

    The dependence of foreign gene expression on cell cycle phases in mammalian cells has been described. In this study, a DF1/chIL-18a cell line that stably expresses the fusion protein chIL-18 was constructed and the enhanced green fluorescence protein connected through a (G4 S)3 linker sequence investigated the relationship between cell cycle phases and fusion protein production. DF1/chIL-18a cells (1 × 10(5) ) were inoculated in 60-mm culture dishes containing 5 mL of media to achieve 50%-60% confluence and were cultured in the presence of the cycle-specific inhibitors 10058-F4, aphidicolin, and colchicine for 24 and 48 h. The percentage of cell density and mean fluorescence intensity in each cell cycle phase were assessed using flow cytometry. The inhibitors effectively arrested cell growth. The fusion protein production rate was higher in the S phase than in the G0/G1 and G2/M phases. When cell cycle progression was blocked in the G0/G1, S, and G2/M phases by the addition of 10058-F4, aphidicolin, and colchicine, respectively, the aphidicolin-induced single cells showed higher fusion protein levels than did the 10058-F4- or colchicine-induced phase cells and the uninduced control cells. Although the cells did not proliferate after the drug additions, the amount of total fusion protein accumulated in aphidicolin-treated cells was similar to that in the untreated cultures. Fusion protein is biologically active because it induces IFN-γ production in splenocyte cultures of chicken. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:581-591, 2016. PMID:26850993

  4. Inhibition of p70 S6 Kinase (S6K1) Activity by A77 1726 and Its Effect on Cell Proliferation and Cell Cycle Progress12

    PubMed Central

    Doscas, Michelle E.; Williamson, Ashley J.; Usha, Lydia; Bogachkov, Yedida; Rao, Geetha S.; Xiao, Fei; Wang, Yimin; Ruby, Carl; Kaufman, Howard; Zhou, Jingsong; Williams, James W.; Li, Yi; Xu, Xiulong

    2014-01-01

    Leflunomide is a novel immunomodulatory drug prescribed for treating rheumatoid arthritis. It inhibits the activity of protein tyrosine kinases and dihydroorotate dehydrogenase, a rate-limiting enzyme in the pyrimidine nucleotide synthesis pathway. Here, we report that A77 1726, the active metabolite of leflunomide, inhibited the phosphorylation of ribosomal protein S6 and two other substrates of S6K1, insulin receptor substrate-1 and carbamoyl phosphate synthetase 2, in an A375 melanoma cell line. A77 1726 increased the phosphorylation of AKT, p70 S6 (S6K1), ERK1/2, and MEK through the feedback activation of the IGF-1 receptor–mediated signaling pathway. Invitro kinase assay revealed that leflunomide and A77 1726 inhibited S6K1 activity with IC50 values of approximately 55 and 80 μM, respectively. Exogenous uridine partially blocked A77 1726–induced inhibition of A375 cell proliferation. S6K1 knockdown led to the inhibition of A375 cell proliferation but did not potentiate the antiproliferative effect of A77 1726. A77 1726 stimulated bromodeoxyuridine incorporation in A375 cells but arrested the cell cycle in the S phase, which was reversed by addition of exogenous uridine or by MAP kinase pathway inhibitors but not by rapamycin and LY294002 (a phosphoinositide 3-kinase inhibitor). These observations suggest that A77 1726 accelerates cell cycle entry into the S phase through MAP kinase activation and that pyrimidine nucleotide depletion halts the completion of the cell cycle. Our study identified a novel molecular target of A77 1726 and showed that the inhibition of S6K1 activity was in part responsible for its antiproliferative activity. Our study also provides a novel mechanistic insight into A77 1726–induced cell cycle arrest in the S phase. PMID:25379019

  5. Regulation of store-operated Ca{sup 2+} entry activity by cell cycle dependent up-regulation of Orai2 in brain capillary endothelial cells

    SciTech Connect

    Kito, Hiroaki; Yamamura, Hisao; Suzuki, Yoshiaki; Yamamura, Hideto; Ohya, Susumu; Asai, Kiyofumi; Imaizumi, Yuji

    2015-04-10

    Store-operated Ca{sup 2+} entry (SOCE) via Orai1 and STIM1 complex is supposed to have obligatory roles in the regulation of cellular functions of vascular endothelial cells, while little is known about the contribution of Orai2. Quantitative PCR and Western blot analyses indicated the expression of Orai2 and STIM2, in addition to Orai1 and STIM1 in bovine brain capillary endothelial cell line, t-BBEC117. During the exponential growth of t-BBEC117, the knockdown of Orai1 and STIM1 significantly reduced the SOCE activity, whereas Orai2 and STIM2 siRNAs had no effect. To examine whether endogenous SOCE activity contributes to the regulation of cell cycle progression, t-BBEC117 were synchronized using double thymidine blockage. At the G2/M phase, Ca{sup 2+} influx via SOCE was decreased and Orai2 expression was increased compared to the G0/G1 phase. When Orai2 was knocked down at the G2/M phase, the decrease in SOCE was removed, and cell proliferation was partly attenuated. Taken together, Orai1 significantly contributes to cell proliferation via the functional expression, which is presumably independent of the cell cycle phases. In construct, Orai2 is specifically up-regulated during the G2/M phase, negatively modulates the SOCE activity, and may contribute to the regulation of cell cycle progression in brain capillary endothelial cells. - Highlights: • Orai1 is essential for SOCE activity in brain capillary endothelial cells (BCECs). • Cell cycle independent expression of Orai1 regulated SOCE and cell proliferation. • Orai2 was up-regulated only at G2/M phase and this consequently reduced SOCE. • Orai2 as well as Orai1 is a key player controlling SOCE and proliferation in BCECs.

  6. Atmospheric particulate matter (PM10) exposure-induced cell cycle arrest and apoptosis evasion through STAT3 activation via PKCζ and Src kinases in lung cells.

    PubMed

    Reyes-Zárate, Elizabeth; Sánchez-Pérez, Yesennia; Gutiérrez-Ruiz, María Concepción; Chirino, Yolanda I; Osornio-Vargas, Álvaro Román; Morales-Bárcenas, Rocío; Souza-Arroyo, Verónica; García-Cuellar, Claudia María

    2016-07-01

    Atmospheric particulate matter with aerodynamic diameter ≤10 μm (PM10) is a risk factor for the development of lung cancer, but cellular pathways are not completely understood. STAT3 is a p21(Waf1/Cip1) transcription factor and is associated with proliferation and cell survival and is upregulated in lung cancer. PM10 exposure induces p21(Waf1/Cip1) expression, which could be related to STAT3 activation. The aims of this work were to investigate whether STAT3 was activated on lung epithelial cells after PM10 exposure and to determine whether or not STAT3 could have an impact on cell cycle distribution and cell survival. Our results showed that PM10 induced STAT3 activation through Src and PKCζ kinases, and it is partially responsible for the p21(Waf1/Cip1) induction that was also observed. Moreover, PM10 induced G1-G0 cell cycle arrest. The inhibition of STAT3 phosphorylation prevented cell cycle arrest and triggered apoptosis. These results suggest that PM10 exposure might activate a survival pathway related to STAT3 activation, similar to what has been described as part of the immune system and apoptosis evasion during tumor promotion and development. PMID:27131825

  7. Temperature and the cell cycle.

    PubMed

    Francis, D; Barlow, P W

    1988-01-01

    that at 5 degrees C, nucleoli are large compared with those at 10-25 degrees C. These observations are consistent with high levels of RNA polymerase and cellular RNA found at low compared with high temperatures. These responses may be important in sustaining growth at 5 degrees C. Finally, the effects of temperature on developmental transitions are discussed. The picture that emerges is that more is known about low, as opposed to high, temperature as a morphogenetic switch but virtually nothing is known about cell cycle activity during such transitions.(ABSTRACT TRUNCATED AT 400 WORDS) PMID:3077857

  8. Microtubule depolymerization activates the Epstein-Barr virus lytic cycle through protein kinase C pathways in nasopharyngeal carcinoma cells.

    PubMed

    Liu, Yi-Ru; Huang, Sheng-Yen; Chen, Jen-Yang; Wang, Lily Hui-Ching

    2013-12-01

    Elevated levels of antibodies against Epstein-Barr virus (EBV) and the presence of viral DNA in plasma are reliable biomarkers for the diagnosis of nasopharyngeal carcinoma (NPC) in high-prevalence areas, such as South-East Asia. The presence of these viral markers in the circulation suggests that a minimal level of virus reactivation may have occurred in an infected individual, although the underlying mechanism of reactivation remains to be elucidated. Here, we showed that treatment with nocodazole, which provokes the depolymerization of microtubules, induces the expression of two EBV lytic cycle proteins, Zta and EA-D, in EBV-positive NPC cells. This effect was independent of mitotic arrest, as viral reactivation was not abolished in cells synchronized at interphase. Notably, the induction of Zta by nocodazole was mediated by transcriptional upregulation via protein kinase C (PKC). Pre-treatment with inhibitors for PKC or its downstream signalling partners p38 mitogen-activated protein kinase (MAPK) and c-Jun N-terminal kinase (JNK) abolished the nocodazole-mediated induction of Zta and EA-D. Interestingly, the effect of nocodazole, as well as colchicine and vinblastine, on lytic gene expression occurred only in NPC epithelial cells but not in cells derived from lymphocytes. These results establish a novel role of microtubule integrity in controlling the EBV life cycle through PKC and its downstream pathways, which represents a tissue-specific mechanism for controlling the life-cycle switch of EBV. PMID:24062531

  9. Gene copy number and cell cycle arrest

    NASA Astrophysics Data System (ADS)

    Ghosh, Bhaswar; Bose, Indrani

    2006-03-01

    The cell cycle is an orderly sequence of events which ultimately lead to the division of a single cell into two daughter cells. In the case of DNA damage by radiation or chemicals, the damage checkpoints in the G1 and G2 phases of the cell cycle are activated. This results in an arrest of the cell cycle so that the DNA damage can be repaired. Once this is done, the cell continues with its usual cycle of activity. We study a mathematical model of the DNA damage checkpoint in the G2 phase which arrests the transition from the G2 to the M (mitotic) phase of the cell cycle. The tumor suppressor protein p53 plays a key role in activating the pathways leading to cell cycle arrest in mammalian systems. If the DNA damage is severe, the p53 proteins activate other pathways which bring about apoptosis, i.e., programmed cell death. Loss of the p53 gene results in the proliferation of cells containing damaged DNA, i.e., in the growth of tumors which may ultimately become cancerous. There is some recent experimental evidence which suggests that the mutation of a single copy of the p53 gene (in the normal cell each gene has two identical copies) is sufficient to trigger the formation of tumors. We study the effect of reducing the gene copy number of the p53 and two other genes on cell cycle arrest and obtain results consistent with experimental observations.

  10. The Src homology 2 protein Shb promotes cell cycle progression in murine hematopoietic stem cells by regulation of focal adhesion kinase activity

    SciTech Connect

    Gustafsson, Karin; Heffner, Garrett; Wenzel, Pamela L.; Curran, Matthew; Grawé, Jan; McKinney-Freeman, Shannon L.; Daley, George Q.; Welsh, Michael

    2013-07-15

    The widely expressed adaptor protein Shb has previously been reported to contribute to T cell function due to its association with the T cell receptor and furthermore, several of Shb's known interaction partners are established regulators of blood cell development and function. In addition, Shb deficient embryonic stem cells displayed reduced blood cell colony formation upon differentiation in vitro. The aim of the current study was therefore to explore hematopoietic stem and progenitor cell function in the Shb knockout mouse. Shb deficient bone marrow contained reduced relative numbers of long-term hematopoietic stem cells (LT-HSCs) that exhibited lower proliferation rates. Despite this, Shb knockout LT-HSCs responded promptly by entering the cell cycle in response to genotoxic stress by 5-fluorouracil treatment. In competitive LT-HSC transplantations, Shb null cells initially engrafted as well as the wild-type cells but provided less myeloid expansion over time. Moreover, Shb knockout bone marrow cells exhibited elevated basal activities of focal adhesion kinase/Rac1/p21-activated kinase signaling and reduced responsiveness to Stem Cell Factor stimulation. Consequently, treatment with a focal adhesion kinase inhibitor increased Shb knockout LT-HSC proliferation. The altered signaling characteristics thus provide a plausible mechanistic explanation for the changes in LT-HSC proliferation since these signaling intermediates have all been shown to participate in LT-HSC cell cycle control. In summary, the loss of Shb dependent signaling in bone marrow cells, resulting in elevated focal adhesion kinase activity and reduced proliferative responses in LT-HSCs under steady state hematopoiesis, confers a disadvantage to the maintenance of LT-HSCs over time. -- Highlights: • Shb is an adaptor protein operating downstream of tyrosine kinase receptors. • Shb deficiency reduces hematopoietic stem cell proliferation. • The proliferative effect of Shb occurs via increased

  11. Aloe emodin induces G2/M cell cycle arrest and apoptosis via activation of caspase-6 in human colon cancer cells.

    PubMed

    Suboj, Priya; Babykutty, Suboj; Srinivas, Priya; Gopala, Srinivas

    2012-01-01

    Aloe emodin (AE), a natural anthraquinone, is reported to have antiproliferative activity in various cancer cell lines. In this study, we analyzed the molecular mechanisms involved in the growth-inhibitory activity of this hydroxyanthraquinone in colon cancer cell, WiDr. In our observation AE inhibited cell proliferation by arresting the cell cycle at the G2/M phase and inhibiting cyclin B1. AE appreciably induced cell death specifically through the induction of apoptosis and by activating caspases 9/6. Apoptotic execution was found to be solely dependent on caspase-6 rather than caspase-3 or caspase-7. This is the first study indicating that the AE induces apoptosis specifically through the activation of caspase-6. PMID:22343391

  12. The cytotoxic activities of 7-isopentenyloxycoumarin on 5637 cells via induction of apoptosis and cell cycle arrest in G2/M stage

    PubMed Central

    2014-01-01

    Background Bladder cancer is the second common malignancy of genitourinary tract, and transitional cell carcinomas (TCCs) account for 90% of all bladder cancers. Due to acquired resistance of TCC cells to a wide range of chemotherapeutic agents, there is always a need for search on new compounds for treatment of these cancers. Coumarins represent a group of natural compounds, which some of them have exerted valuable anti-tumor activities. The current study was designed to evaluate anti-tumor properties and mechanism of action of 7-isopentenyloxycoumarin, a prenyloxycoumarin, on 5637 cells (a TCC cell line). Results MTT results revealed that the cytotoxic effects of 7-isopentenyloxycoumarin on 5637 cancerous cells were more prominent in comparison to HDF-1 normal cells. This coumarin increased the amount of chromatin condensation and DNA damage in 5637 cells by 58 and 33%, respectively. The results also indicated that it can induce apoptosis most probably via activation of caspase-3 in these cells. Moreover, propidium iodide staining revealed that 7-isopentenyloxycoumarin induced cell cycle arrest at G2/M stage, after 24 h of treatment. Conclusion Our results indicated that 7-isopentenyloxycoumarin had selective toxic effects on this bladder cancer cell line and promoted its effects by apoptosis induction and cell cycle arrest. This coumarin can be considered for further studies to reveal its exact mechanism of action and also its anti-cancer effects in vivo. PMID:24393601

  13. Activation of the cell cycle machinery and the isoflavonoid biosynthesis pathway by active Rhizobium meliloti Nod signal molecules in Medicago microcallus suspensions.

    PubMed Central

    Savouré, A; Magyar, Z; Pierre, M; Brown, S; Schultze, M; Dudits, D; Kondorosi, A; Kondorosi, E

    1994-01-01

    We have shown that treatment of Medicago microcallus suspensions with the cognate Rhizobium meliloti Nod signal molecule NodRm-IV(C16:2,S) can modify gene expression both qualitatively and quantitatively. At concentrations of 10(-6) - 10(-9) M, this host specific plant morphogen but not the inactive non-sulfated molecule stimulated cell cycle progression as indicated by the significantly enhanced thymidine incorporation, elevated number of S phase cells, increase in kinase activity of the p34cdc2-related complexes and enhancement of the level of expression of several cell cycle marker genes, the histone H3-1, the cdc2Ms and the cyclin cycMs2. The presented data suggest that at least part of the physiological role of the Nod factor may be linked to molecular events involved in the control of the plant cell division cycle. In situ hybridization experiments with antisense H3-1 RNA probe indicated that only certain cells of the calli were able to respond to the Nod factor. High (10(-6) M) but not low (10(-9) M) concentrations of the active Nod factors induced the expression of the isoflavone reductase gene (IFR), a marker gene of the isoflavonoid biosynthesis pathway in most callus cells. Our results indicate that Medicago cell responses to the Nod signal molecules can be investigated in suspension cultures. Images PMID:8131743

  14. Quercetin, a Natural Flavonoid Interacts with DNA, Arrests Cell Cycle and Causes Tumor Regression by Activating Mitochondrial Pathway of Apoptosis

    PubMed Central

    Srivastava, Shikha; Somasagara, Ranganatha R.; Hegde, Mahesh; Nishana, Mayilaadumveettil; Tadi, Satish Kumar; Srivastava, Mrinal; Choudhary, Bibha; Raghavan, Sathees C.

    2016-01-01

    Naturally occurring compounds are considered as attractive candidates for cancer treatment and prevention. Quercetin and ellagic acid are naturally occurring flavonoids abundantly seen in several fruits and vegetables. In the present study, we evaluate and compare antitumor efficacies of quercetin and ellagic acid in animal models and cancer cell lines in a comprehensive manner. We found that quercetin induced cytotoxicity in leukemic cells in a dose-dependent manner, while ellagic acid showed only limited toxicity. Besides leukemic cells, quercetin also induced cytotoxicity in breast cancer cells, however, its effect on normal cells was limited or none. Further, quercetin caused S phase arrest during cell cycle progression in tested cancer cells. Quercetin induced tumor regression in mice at a concentration 3-fold lower than ellagic acid. Importantly, administration of quercetin lead to ~5 fold increase in the life span in tumor bearing mice compared to that of untreated controls. Further, we found that quercetin interacts with DNA directly, and could be one of the mechanisms for inducing apoptosis in both, cancer cell lines and tumor tissues by activating the intrinsic pathway. Thus, our data suggests that quercetin can be further explored for its potential to be used in cancer therapeutics and combination therapy. PMID:27068577

  15. Plasma membrane/cell wall perturbation activates a novel cell cycle checkpoint during G1 in Saccharomyces cerevisiae.

    PubMed

    Kono, Keiko; Al-Zain, Amr; Schroeder, Lea; Nakanishi, Makoto; Ikui, Amy E

    2016-06-21

    Cellular wound healing or the repair of plasma membrane/cell wall damage (plasma membrane damage) occurs frequently in nature. Although various cellular perturbations, such as DNA damage, spindle misalignment, and impaired daughter cell formation, are monitored by cell cycle checkpoint mechanisms in budding yeast, whether plasma membrane damage is monitored by any of these checkpoints remains to be addressed. Here, we define the mechanism by which cells sense membrane damage and inhibit DNA replication. We found that the inhibition of DNA replication upon plasma membrane damage requires GSK3/Mck1-dependent degradation of Cdc6, a component of the prereplicative complex. Furthermore, the CDK inhibitor Sic1 is stabilized in response to plasma membrane damage, leading to cell integrity maintenance in parallel with the Mck1-Cdc6 pathway. Cells defective in both Cdc6 degradation and Sic1 stabilization failed to grow in the presence of plasma membrane damage. Taking these data together, we propose that plasma membrane damage triggers G1 arrest via Cdc6 degradation and Sic1 stabilization to promote the cellular wound healing process. PMID:27274080

  16. p38α Activates Purine Metabolism to Initiate Hematopoietic Stem/Progenitor Cell Cycling in Response to Stress.

    PubMed

    Karigane, Daiki; Kobayashi, Hiroshi; Morikawa, Takayuki; Ootomo, Yukako; Sakai, Mashito; Nagamatsu, Go; Kubota, Yoshiaki; Goda, Nobuhito; Matsumoto, Michihiro; Nishimura, Emi K; Soga, Tomoyoshi; Otsu, Kinya; Suematsu, Makoto; Okamoto, Shinichiro; Suda, Toshio; Takubo, Keiyo

    2016-08-01

    Hematopoietic stem cells (HSCs) maintain quiescence by activating specific metabolic pathways, including glycolysis. We do not yet have a clear understanding of how this metabolic activity changes during stress hematopoiesis, such as bone marrow transplantation. Here, we report a critical role for the p38MAPK family isoform p38α in initiating hematopoietic stem and progenitor cell (HSPC) proliferation during stress hematopoiesis in mice. We found that p38MAPK is immediately phosphorylated in HSPCs after a hematological stress, preceding increased HSPC cycling. Conditional deletion of p38α led to defective recovery from hematological stress and a delay in initiation of HSPC proliferation. Mechanistically, p38α signaling increases expression of inosine-5'-monophosphate dehydrogenase 2 in HSPCs, leading to altered levels of amino acids and purine-related metabolites and changes in cell-cycle progression in vitro and in vivo. Our studies have therefore uncovered a p38α-mediated pathway that alters HSPC metabolism to respond to stress and promote recovery. PMID:27345838

  17. Anticancer and apoptotic activities of oleanolic acid are mediated through cell cycle arrest and disruption of mitochondrial membrane potential in HepG2 human hepatocellular carcinoma cells.

    PubMed

    Zhu, Yue-Yong; Huang, Hong-Yan; Wu, Yin-Lian

    2015-10-01

    Hepatocellular carcinoma (HCC) is an aggressive form of cancer, with high rates of morbidity and mortality, a poor prognosis and limited therapeutic options. The objective of the present study was to demonstrate the anticancer activity of oleanolic acid in HepG2 human HCC cells. Cell viability was evaluated using an MTT assay, following administration of various doses of oleanolic acid. The effect of oleanolic acid on cell cycle phase distribution and mitochondrial membrane potential was evaluated using flow cytometry with propidium iodide and rhodamine‑123 DNA‑binding cationic fluorescent dyes. Fluorescence microscopy was employed to detect morphological changes in HepG2 cells following oleanolic acid treatment. The results revealed that oleanolic acid induced a dose‑dependent, as well as time‑dependent inhibition in the growth of HepG2 cancer cells. Following acridine orange and ethidium bromide staining, treatment with various doses (0, 5, 25 and 50 µM) of oleanolic acid induced typical morphological changes associated with apoptosis, including cell shrinkage, membrane blebbing, nuclear condensation and apoptotic body formation. Cell cycle analysis revealed that oleanolic acid induced cell cycle arrest in HepG2 cells at the sub‑G1 (apoptotic) phase of the cell cycle, in a dose‑dependent manner. Staining with Annexin V‑fluorescein isothiocyanate and propidium iodide revealed that apoptosis occurred early in these cells. Oleanolic acid treatment also resulted in fragmentation of nuclear DNA in a dose‑dependent manner, producing the typical features of DNA laddering on an agarose gel. The results also demonstrated that oleanolic acid treatment resulted in a potent loss of mitochondrial membrane potential, which also occurred in a dose‑dependent manner. Therefore, oleanolic acid may be used as a therapeutic agent in the treatment of human HCC. PMID:26151733

  18. E. adenophorum induces Cell Cycle Arrest and Apoptosis of Splenocytes through the Mitochondrial Pathway and Caspase Activation in Saanen Goats

    PubMed Central

    He, Yajun; Mo, Quan; Hu, Yanchun; Chen, Weihong; Luo, Biao; Wu, Lei; Qiao, Yan; Xu, Ruiguang; Zhou, Yancheng; Zuo, Zhicai; Deng, Junliang; He, Wei; Wei, Yahui

    2015-01-01

    The precise cytotoxicity of E. Adenophorum in relation to the cell cycle and apoptosis of splenocytes in Saanen goats remains unclear. In the present study, 16 Saanen goats were randomly divided into four groups, which were fed on 0%, 40%, 60% and 80% E. adenophorum diets. The results of TUNEL, DAPI and AO/EB staining, flow cytometry analysis and DNA fragmentation assays showed that E. adenophorum induced typical apoptotic features in splenocytes, suppressed splenocyte viability, and caused cell cycle arrest in a dose-dependent manner. However, westernblot, ELISA, qRT-PCR and caspase activity analyses showed that E. adenophoruminhibited Bcl-2 expression, promoted Bax translocation to the mitochondria, triggered the release of Cytc from the mitochondria into the cytosol, and activated caspase-9 and -3 and the subsequent cleavage of PARP. Moreover, in E. adenophorum-induced apoptosis, the protein levels of Fas, Bid, FasL and caspase-8 showed no significant changes. E. adenophorum treatment induced the collapse of ΔΨm. Moreover, these data suggested that E. adenophorum induces splenocyte apoptosis via the activation of the mitochondrial apoptosis pathway in splenocytes. These findings provide new insights into the mechanisms underlying the effects of E. adenophorum cytotoxicity on splenocytes. PMID:26527166

  19. Nek2 activation of Kif24 ensures cilium disassembly during the cell cycle.

    PubMed

    Kim, Sehyun; Lee, Kwanwoo; Choi, Jung-Hwan; Ringstad, Niels; Dynlacht, Brian David

    2015-01-01

    Many proteins are known to promote ciliogenesis, but mechanisms that promote primary cilia disassembly before mitosis are largely unknown. Here we identify a mechanism that favours cilium disassembly and maintains the disassembled state. We show that co-localization of the S/G2 phase kinase, Nek2 and Kif24 triggers Kif24 phosphorylation, inhibiting cilia formation. We show that Kif24, a microtubule depolymerizing kinesin, is phosphorylated by Nek2, which stimulates its activity and prevents the outgrowth of cilia in proliferating cells, independent of Aurora A and HDAC6. Our data also suggest that cilium assembly and disassembly are in dynamic equilibrium, but Nek2 and Kif24 can shift the balance toward disassembly. Further, Nek2 and Kif24 are overexpressed in breast cancer cells, and ablation of these proteins restores ciliation in these cells, thereby reducing proliferation. Thus, Kif24 is a physiological substrate of Nek2, which regulates cilia disassembly through a concerted mechanism involving Kif24-mediated microtubule depolymerization. PMID:26290419

  20. Nek2 activation of Kif24 ensures cilium disassembly during the cell cycle

    PubMed Central

    Kim, Sehyun; Lee, Kwanwoo; Choi, Jung-Hwan; Ringstad, Niels; Dynlacht, Brian David

    2015-01-01

    Many proteins are known to promote ciliogenesis, but mechanisms that promote primary cilia disassembly before mitosis are largely unknown. Here we identify a mechanism that favours cilium disassembly and maintains the disassembled state. We show that co-localization of the S/G2 phase kinase, Nek2 and Kif24 triggers Kif24 phosphorylation, inhibiting cilia formation. We show that Kif24, a microtubule depolymerizing kinesin, is phosphorylated by Nek2, which stimulates its activity and prevents the outgrowth of cilia in proliferating cells, independent of Aurora A and HDAC6. Our data also suggest that cilium assembly and disassembly are in dynamic equilibrium, but Nek2 and Kif24 can shift the balance toward disassembly. Further, Nek2 and Kif24 are overexpressed in breast cancer cells, and ablation of these proteins restores ciliation in these cells, thereby reducing proliferation. Thus, Kif24 is a physiological substrate of Nek2, which regulates cilia disassembly through a concerted mechanism involving Kif24-mediated microtubule depolymerization. PMID:26290419

  1. Cell cycle gene expression under clinorotation

    NASA Astrophysics Data System (ADS)

    Artemenko, Olga

    2016-07-01

    Cyclins and cyclin-dependent kinase (CDK) are main regulators of the cell cycle of eukaryotes. It's assumes a significant change of their level in cells under microgravity conditions and by other physical factors actions. The clinorotation use enables to determine the influence of gravity on simulated events in the cell during the cell cycle - exit from the state of quiet stage and promotion presynthetic phase (G1) and DNA synthesis phase (S) of the cell cycle. For the clinorotation effect study on cell proliferation activity is the necessary studies of molecular mechanisms of cell cycle regulation and development of plants under altered gravity condition. The activity of cyclin D, which is responsible for the events of the cell cycle in presynthetic phase can be controlled by the action of endogenous as well as exogenous factors, but clinorotation is one of the factors that influence on genes expression that regulate the cell cycle.These data can be used as a model for further research of cyclin - CDK complex for study of molecular mechanisms regulation of growth and proliferation. In this investigation we tried to summarize and analyze known literature and own data we obtained relatively the main regulators of the cell cycle in altered gravity condition.

  2. Intrinsic caspase-8 activation mediates sensitization of erlotinib-resistant tumor cells to erlotinib/cell-cycle inhibitors combination treatment

    PubMed Central

    Orzáez, M; Guevara, T; Sancho, M; Pérez-Payá, E

    2012-01-01

    Inhibitors of the tyrosine kinase activity of epidermal growth factor receptor, as erlotinib, have an established role in treating several cancer types. However, resistance to erlotinib, particularly in breast cancer cell lines, and erlotinib treatment-associated disorders have also been described. Also, methods and combination therapies that could reverse resistance and ameliorate non-desirable effects represent a clinical challenge. Here, we show that the ATP non-competitive CDK2/cyclin A inhibitor NBI1 sensitizes erlotinib-resistant tumor cells to the combination treatment (co-treatment) for apoptosis-mediated cell death. Furthermore, in erlotinib-sensitive cells, the effective dose of erlotinib was lower in the presence of NBI1. The analysis in the breast cancer MDA-MB-468 erlotinib-resistant and in lung cancer A549 cell lines of the molecular mechanism underlying the apoptosis induced by co-treatment highlighted that the accumulation of DNA defects and depletion of cIAP and XIAP activates the ripoptosome that ultimately activates caspases-8 and -10 and apoptosis. This finding could have significant implications for future treatment strategies in clinical settings. PMID:23096116

  3. The Arabidopsis Cell Division Cycle

    PubMed Central

    Gutierrez, Crisanto

    2009-01-01

    Plant cells have evolved a complex circuitry to regulate cell division. In many aspects, the plant cell cycle follows a basic strategy similar to other eukaryotes. However, several key issues are unique to plant cells. In this chapter, both the conserved and unique cellular and molecular properties of the plant cell cycle are reviewed. In addition to division of individual cells, the specific characteristic of plant organogenesis and development make that cell proliferation control is of primary importance during development. Therefore, special attention should be given to consider plant cell division control in a developmental context. Proper organogenesis depends on the formation of different cell types. In plants, many of the processes leading to cell differentiation rely on the occurrence of a different cycle, termed the endoreplication cycle, whereby cells undergo repeated full genome duplication events in the absence of mitosis and increase their ploidy. Recent findings are focusing on the relevance of changes in chromatin organization for a correct cell cycle progression and, conversely, in the relevance of a correct functioning of chromatin remodelling complexes to prevent alterations in both the cell cycle and the endocycle. PMID:22303246

  4. Cucurbitacin-I (JSI-124) activates the JNK/c-Jun signaling pathway independent of apoptosis and cell cycle arrest in B Leukemic Cells

    PubMed Central

    2011-01-01

    Background Cucurbitacin-I (JSI-124) is potent inhibitor of JAK/STAT3 signaling pathway and has anti-tumor activity in a variety of cancer including B cell leukemia. However, other molecular targets of JSI-124 beyond the JAK/STAT3 pathway are not fully understood. Methods BJAB, I-83, NALM-6 and primary CLL cells were treated with JSI-124 as indicated. Apoptosis was measured using flow cytometry for accumulation of sub-G1 phase cells (indicator of apoptosis) and Annexin V/PI staining. Cell cycle was analyzed by FACS for DNA content of G1 and G2 phases. Changes in phosphorylation and protein expression of p38, Erk1/2, JNK, c-Jun, and XIAP were detected by Western blot analysis. STAT3 and c-Jun genes were knocked out using siRNA transfection. VEGF expression was determined by mRNA and protein levels by RT-PCR and western blotting. Streptavidin Pull-Down Assay was used to determine c-Jun binding to the AP-1 DNA binding site. Results Herein, we show that JSI-124 activates c-Jun N-terminal kinase (JNK) and increases both the expression and serine phosphorylation of c-Jun protein in the B leukemic cell lines BJAB, I-83 and NALM-6. JSI-124 also activated MAPK p38 and MAPK Erk1/2 albeit at lower levels than JNK activation. Inhibition of the JNK signaling pathway failed to effect cell cycle arrest or apoptosis induced by JSI-124 but repressed JSI-124 induced c-Jun expression in these leukemia cells. The JNK pathway activation c-Jun leads to transcriptional activation of many genes. Treatment of BJAB, I-83, and NALM-6 cells with JSI-124 lead to an increase of Vascular Endothelial Growth Factor (VEGF) at both the mRNA and protein level. Knockdown of c-Jun expression and inhibition of JNK activation significantly blocked JSI-124 induced VEGF expression. Pretreatment with recombinant VEGF reduced JSI-124 induced apoptosis. Conclusions Taken together, our data demonstrates that JSI-124 activates the JNK signaling pathway independent of apoptosis and cell cycle arrest, leading to

  5. MST-312 induces G2/M cell cycle arrest and apoptosis in APL cells through inhibition of telomerase activity and suppression of NF-κB pathway.

    PubMed

    Fatemi, Ahmad; Safa, Majid; Kazemi, Ahmad

    2015-11-01

    Telomerase-targeted therapy for cancer has received great attention because telomerase is expressed in almost all cancer cells but is inactive in most normal somatic cells. This study was aimed to investigate the effects of telomerase inhibitor MST-312, a chemically modified derivative of epigallocatechin gallate (EGCG), on acute promyelocytic leukemia (APL) cells. Our results showed that MST-312 exerted a dose-dependent short-term cytotoxic effect on APL cells, with G2/M cell cycle arrest. Moreover, MST-312 induced apoptosis of APL cells in caspase-mediated manner. Telomeric repeat amplification protocol (TRAP) assay revealed significant reduction in telomerase activity of APL cells following short-term exposure to MST-312. Interestingly, MST-312-induced telomerase inhibition was coupled with suppression of NF-κB activity as evidenced by inhibition of IκBα phosphorylation and its degradation and decreased NF-κB DNA binding activity. In addition, gene expression analysis showed downregulation of genes regulated by NF-κB, such as antiapoptotic (survivin, Bcl-2, Mcl-1), proliferative (c-Myc), and telomerase-related (hTERT) genes. Importantly, MST-312 did not show any apoptotic effect in normal human peripheral blood mononuclear cells (PBMCs). In conclusion, our data suggest that dual inhibition of telomerase activity and NF-κB pathway by MST-312 represents a novel treatment strategy for APL. PMID:26022158

  6. Tetrahydrocurcumin induces G2/M cell cycle arrest and apoptosis involving p38 MAPK activation in human breast cancer cells.

    PubMed

    Kang, Ning; Wang, Miao-Miao; Wang, Ying-Hui; Zhang, Zhe-Nan; Cao, Hong-Rui; Lv, Yuan-Hao; Yang, Yang; Fan, Peng-Hui; Qiu, Feng; Gao, Xiu-Mei

    2014-05-01

    Curcumin (CUR) is a major naturally-occurring polyphenol of Curcuma species, which is commonly used as a yellow coloring and flavoring agent in foods. In recent years, it has been reported that CUR exhibits significant anti-tumor activity in vivo. However, the pharmacokinetic features of CUR have indicated poor oral bioavailability, which may be related to its extensive metabolism. The CUR metabolites might be responsible for the antitumor pharmacological effects in vivo. Tetrahydrocurcumin (THC) is one of the major metabolites of CUR. In the present study, we examined the efficacy and associated mechanism of action of THC in human breast cancer MCF-7 cells for the first time. Here, THC exhibited significant cell growth inhibition by inducing MCF-7 cells to undergo mitochondrial apoptosis and G2/M arrest. Moreover, co-treatment of MCF-7 cells with THC and p38 MAPK inhibitor, SB203580, effectively reversed the dissipation in mitochondrial membrane potential (Δψm), and blocked THC-mediated Bax up-regulation, Bcl-2 down-regulation, caspase-3 activation as well as p21 up-regulation, suggesting p38 MAPK might mediate THC-induced apoptosis and G2/M arrest. Taken together, these results indicate THC might be an active antitumor form of CUR in vivo, and it might be selected as a potentially effective agent for treatment of human breast cancer. PMID:24593988

  7. A novel quantitative model of cell cycle progression based on cyclin-dependent kinases activity and population balances.

    PubMed

    Pisu, Massimo; Concas, Alessandro; Cao, Giacomo

    2015-04-01

    Cell cycle regulates proliferative cell capacity under normal or pathologic conditions, and in general it governs all in vivo/in vitro cell growth and proliferation processes. Mathematical simulation by means of reliable and predictive models represents an important tool to interpret experiment results, to facilitate the definition of the optimal operating conditions for in vitro cultivation, or to predict the effect of a specific drug in normal/pathologic mammalian cells. Along these lines, a novel model of cell cycle progression is proposed in this work. Specifically, it is based on a population balance (PB) approach that allows one to quantitatively describe cell cycle progression through the different phases experienced by each cell of the entire population during its own life. The transition between two consecutive cell cycle phases is simulated by taking advantage of the biochemical kinetic model developed by Gérard and Goldbeter (2009) which involves cyclin-dependent kinases (CDKs) whose regulation is achieved through a variety of mechanisms that include association with cyclins and protein inhibitors, phosphorylation-dephosphorylation, and cyclin synthesis or degradation. This biochemical model properly describes the entire cell cycle of mammalian cells by maintaining a sufficient level of detail useful to identify check point for transition and to estimate phase duration required by PB. Specific examples are discussed to illustrate the ability of the proposed model to simulate the effect of drugs for in vitro trials of interest in oncology, regenerative medicine and tissue engineering. PMID:25601491

  8. A role for Hsp90 in cell cycle control: Wee1 tyrosine kinase activity requires interaction with Hsp90.

    PubMed Central

    Aligue, R; Akhavan-Niak, H; Russell, P

    1994-01-01

    Wee1 protein kinase regulates the length of G2 phase by carrying out the inhibitory tyrosyl phosphorylation of Cdc2-cyclin B kinase. Mutations were isolated that suppressed the G2 cell cycle arrest caused by overproduction of Wee1. One class of swo (suppressor of wee1 overproduction) mutation, exemplified by swo1-26, also caused a temperature sensitive lethal phenotype in a wee1+ background. The swo1+ gene encodes a member of the Hsp90 family of stress proteins. Swo1 is essential for viability at all temperatures. Swo1 coimmunoprecipitates with Wee1, showing that the two proteins interact. The swo1-26 mutant undergoes premature mitosis when grown at a semi-permissive temperature. These data strongly indicate that formation of active Wee1 tyrosine kinase requires interaction with Swo1, perhaps in a manner analogous to the previously demonstrated interaction between Hsp90 and v-src tyrosine kinase. These observations demonstrate a unexpected role for Hsp90 in cell cycle control. Images PMID:7813446

  9. Closing the cell cycle circle in yeast: G2 cyclin proteolysis initiated at mitosis persists until the activation of G1 cyclins in the next cycle.

    PubMed

    Amon, A; Irniger, S; Nasmyth, K

    1994-07-01

    It is thought that DNA replication and mitosis in yeast are triggered by oscillations in the level of G1-specific (CLN1 and CLN2) and G2-specific (CLB1-CLB4) cyclins, which determine the substrate specificity of the CDC28 protein kinase. It is not understood how the time and order of appearance of different cyclin types are determined. We show here that CLB2 proteolysis, which is important for transition from mitosis to G1, is not confined to a narrow window at the end of mitosis as previously thought but continues until reactivation of CDC28 by CLN cyclins toward the end of the subsequent G1 period. Thus, cell cycle-regulated proteolysis prevents accumulation of G2-specific CLB cyclins during G1 and thereby ensures that the CLN-associated forms of the CDC28 kinase are activated without interference from CLB cyclins. Accumulation of CLN cyclins leads to inactivation of CLB cyclin proteolysis, which is a precondition for subsequent activation of G2-specific B-type cyclins. PMID:8020094

  10. Combined Treatment of MCF-7 Cells with AICAR and Methotrexate, Arrests Cell Cycle and Reverses Warburg Metabolism through AMP-Activated Protein Kinase (AMPK) and FOXO1

    PubMed Central

    Fodor, Tamás; Szántó, Magdolna; Abdul-Rahman, Omar; Nagy, Lilla; Dér, Ádám; Kiss, Borbála; Bai, Peter

    2016-01-01

    Cancer cells are characterized by metabolic alterations, namely, depressed mitochondrial oxidation, enhanced glycolysis and pentose phosphate shunt flux to support rapid cell growth, which is called the Warburg effect. In our study we assessed the metabolic consequences of a joint treatment of MCF-7 breast cancer cells with AICAR, an inducer of AMP-activated kinase (AMPK) jointly with methotrexate (MTX), a folate-analog antimetabolite that blunts de novo nucleotide synthesis. MCF7 cells, a model of breast cancer cells, were resistant to the individual application of AICAR or MTX, however combined treatment of AICAR and MTX reduced cell proliferation. Prolonged joint application of AICAR and MTX induced AMPK and consequently enhanced mitochondrial oxidation and reduced the rate of glycolysis. These metabolic changes suggest an anti-Warburg rearrangement of metabolism that led to the block of the G1/S and the G2/M transition slowing down cell cycle. The slowdown of cell proliferation was abolished when mitotropic transcription factors, PGC-1α, PGC-1β or FOXO1 were silenced. In human breast cancers higher expression of AMPKα and FOXO1 extended survival. AICAR and MTX exerts similar additive antiproliferative effect on other breast cancer cell lines, such as SKBR and 4T1 cells, too. Our data not only underline the importance of Warburg metabolism in breast cancer cells but nominate the AICAR+MTX combination as a potential cytostatic regime blunting Warburg metabolism. Furthermore, we suggest the targeting of AMPK and FOXO1 to combat breast cancer. PMID:26919657

  11. Combined Treatment of MCF-7 Cells with AICAR and Methotrexate, Arrests Cell Cycle and Reverses Warburg Metabolism through AMP-Activated Protein Kinase (AMPK) and FOXO1.

    PubMed

    Fodor, Tamás; Szántó, Magdolna; Abdul-Rahman, Omar; Nagy, Lilla; Dér, Ádám; Kiss, Borbála; Bai, Peter

    2016-01-01

    Cancer cells are characterized by metabolic alterations, namely, depressed mitochondrial oxidation, enhanced glycolysis and pentose phosphate shunt flux to support rapid cell growth, which is called the Warburg effect. In our study we assessed the metabolic consequences of a joint treatment of MCF-7 breast cancer cells with AICAR, an inducer of AMP-activated kinase (AMPK) jointly with methotrexate (MTX), a folate-analog antimetabolite that blunts de novo nucleotide synthesis. MCF7 cells, a model of breast cancer cells, were resistant to the individual application of AICAR or MTX, however combined treatment of AICAR and MTX reduced cell proliferation. Prolonged joint application of AICAR and MTX induced AMPK and consequently enhanced mitochondrial oxidation and reduced the rate of glycolysis. These metabolic changes suggest an anti-Warburg rearrangement of metabolism that led to the block of the G1/S and the G2/M transition slowing down cell cycle. The slowdown of cell proliferation was abolished when mitotropic transcription factors, PGC-1α, PGC-1β or FOXO1 were silenced. In human breast cancers higher expression of AMPKα and FOXO1 extended survival. AICAR and MTX exerts similar additive antiproliferative effect on other breast cancer cell lines, such as SKBR and 4T1 cells, too. Our data not only underline the importance of Warburg metabolism in breast cancer cells but nominate the AICAR+MTX combination as a potential cytostatic regime blunting Warburg metabolism. Furthermore, we suggest the targeting of AMPK and FOXO1 to combat breast cancer. PMID:26919657

  12. Anticancer Activity of Novel Daphnane Diterpenoids from Daphne genkwa through Cell-Cycle Arrest and Suppression of Akt/STAT/Src Signalings in Human Lung Cancer Cells.

    PubMed

    Jo, Si-Kyoung; Hong, Ji-Young; Park, Hyen Joo; Lee, Sang Kook

    2012-11-01

    Although the immense efforts have been made for cancer prevention, early diagnosis, and treatment, cancer morbidity and mortality has not been decreased during last forty years. Especially, lung cancer is top-ranked in cancer-associated human death. Therefore, effective strategy is strongly required for the management of lung cancer. In the present study, we found that novel daphnane diterpenoids, yuanhualine (YL), yuanhuahine (YH) and yuanhuagine (YG) isolated from the flower of Daphne genkwa (Thymelaeaceae), exhibited potent anti-proliferative activities against human lung A549 cells with the IC50 values of 7.0, 15.2 and 24.7 nM, respectively. Flow cytometric analysis revealed that the daphnane diterpenoids induced cell-cycle arrest in the G0/G1 as well as G2/M phase in A549 cells. The cell-cycle arrests were well correlated with the expression of checkpoint proteins including the up-regulation of cyclin-dependent kinase inhibitor p21 and p53 and down-regulation of cyclin A, cyclin B1, cyclin E, cyclin dependent kinase 4, cdc2, phosphorylation of Rb and cMyc expression. In the analysis of signal transduction molecules, the daphnane diterpenoids suppressed the activation of Akt, STAT3 and Src in human lung cancer cells. The daphnane diterpenoids also exerted the potent anti-proliferative activity against anticancer-drug resistant cancer cells including gemcitabine-resistant A549, gefitinib-, erlotinib-resistant H292 cells. Synergistic effects in the growth inhibition were also observed when yuanhualine was combined with gemcitabine, gefitinib or erlotinib in A549 cells. Taken together, these findings suggest that the novel daphnane diterpenoids might provide lead candidates for the development of therapeutic agents for human lung cancers. PMID:24009843

  13. Anti-proliferative activity and cell cycle arrest induced by evodiamine on paclitaxel-sensitive and -resistant human ovarian cancer cells

    PubMed Central

    Zhong, Zhang-Feng; Tan, Wen; Wang, Sheng-Peng; Qiang, Wen-An; Wang, Yi-Tao

    2015-01-01

    Chemo-resistance is the main factor for poor prognosis in human ovarian epithelial cancer. Active constituents derived from Chinese medicine with anti-cancer potential might circumvent this obstacle. In our present study, evodiamine (EVO) derived from Evodia rutaecarpa (Juss.) Benth suppressed the proliferation of human epithelial ovarian cancer, A2780 and the related paclitaxel-resistant cell lines and did not cause cytotoxicity, as confirmed by the significant decline of clone formation and the representative alterations of CFDA-SE fluorescence. Meanwhile, EVO induced cell cycle arrest in a dose- and time-dependent manner. This disturbance might be mediated by the cooperation of Cyclin B1 and Cdc2, including the up-regulation of Cyclin B1, p27, and p21, and activation failure of Cdc2 and pRb. MAPK signaling pathway regulation also assisted in this process. Furthermore, chemo-sensitivity potential was enhanced as indicated in A2780/PTXR cells by the down-regulation of MDR-1 expression, accompanied by MDR-1 function suppression. Taken together, we confirmed initially that EVO exerted an anti-proliferative effect on human epithelial ovarian cancer cells, A2780/WT and A2780/PTXR, induced G2/M phase cell cycle arrest, and improved chemo-resistance. Overall, we found that EVO significantly suppressed malignant proliferation in human epithelial ovarian cancer, thus proving to be a potential anti-cancer agent in the future. PMID:26553648

  14. Cell cycle control in Alphaproteobacteria.

    PubMed

    Collier, Justine

    2016-04-01

    Alphaproteobacteria include many medically and environmentally important organisms. Despite the diversity of their niches and lifestyles, from free-living to host-associated, they usually rely on very similar mechanisms to control their cell cycles. Studies on Caulobacter crescentus still lay the foundation for understanding the molecular details of pathways regulating DNA replication and cell division and coordinating these two processes with other events of the cell cycle. This review highlights recent discoveries on the regulation and the mode of action of conserved global regulators and small molecules like c-di-GMP and (p)ppGpp, which play key roles in cell cycle control. It also describes several newly identified mechanisms that modulate cell cycle progression in response to stresses or environmental conditions. PMID:26871482

  15. The cell cycle and pluripotency.

    PubMed

    Hindley, Christopher; Philpott, Anna

    2013-04-15

    PSCs (pluripotent stem cells) possess two key properties that have made them the focus of global research efforts in regenerative medicine: they have unlimited expansion potential under conditions which favour their preservation as PSCs and they have the ability to generate all somatic cell types upon differentiation (pluripotency). Conditions have been defined in vitro in which pluripotency is maintained, or else differentiation is favoured and is directed towards specific somatic cell types. However, an unanswered question is whether or not the core cell cycle machinery directly regulates the pluripotency and differentiation properties of PSCs. If so, then manipulation of the cell cycle may represent an additional tool by which in vitro maintenance or differentiation of PSCs may be controlled in regenerative medicine. The present review aims to summarize our current understanding of links between the core cell cycle machinery and the maintenance of pluripotency in ESCs (embryonic stem cells) and iPSCs (induced PSCs). PMID:23535166

  16. Metabolic cycle, cell cycle, and the finishing kick to Start

    PubMed Central

    Futcher, Bruce

    2006-01-01

    Slowly growing budding yeast store carbohydrate, then liquidate it in late G1 phase of the cell cycle, superimposing a metabolic cycle on the cell cycle. This metabolic cycle may separate biochemically incompatible processes. Alternatively it may provide a burst of energy and material for commitment to the cell cycle. Stored carbohydrate could explain the size requirement for cells passing the Start point. PMID:16677426

  17. Cell Cycle Regulation by Checkpoints

    PubMed Central

    Barnum, Kevin J.; O’Connell, Matthew J.

    2016-01-01

    Cell cycle checkpoints are surveillance mechanisms that monitor the order, integrity, and fidelity of the major events of the cell cycle. These include growth to the appropriate cell size, the replication and integrity of the chromosomes, and their accurate segregation at mitosis. Many of these mechanisms are ancient in origin and highly conserved, and hence have been heavily informed by studies in simple organisms such as the yeasts. Others have evolved in higher organisms, and control alternative cell fates with significant impact on tumor suppression. Here, we consider these different checkpoint pathways and the consequences of their dysfunction on cell fate. PMID:24906307

  18. Cell cycle inhibitory activity of Piper longum against A549 cell line and its protective effect against metal-induced toxicity in rats.

    PubMed

    Sharma, Amit Kumar; Kumar, Shashank; Chashoo, Gousia; Saxena, Ajit K; Pandey, Abhay K

    2014-10-01

    Anticancer potential of Piper longum fruit against human cancer cell lines (DU-145 prostate, A549 lung, THP-1 leukemia, IGR-OVI-1 ovary and MCF-7 breast) as well as its in vitro and in vivo biochemical efficacy in A1Cl3-induced hepatotoxicity were evaluated in the rats. Dried samples were extracted with several solvents using soxhlet apparatus. Flavonoid content in chloroform, benzene, ethyl alcohol and aqueous extracts of fruit was 19, 14, 12 and 11 μg quercetin equivalent/mg of sample, respectively. Hexane extracts exhibited 90-92% cytotoxicity against most of the test cell lines (A549, THP-1, IGR-OVI-1 and MCF-7), while benzene extract displayed 84-87% cytotoxicity against MCF-7, IGR-OV-1 and THP-1 cell lines. Among extracts, hexane, benzene and acetone extracts demonstrated considerable cytotoxicity (91-95%) against A549 (lung cancer) cell line in Sulforhodamine B dye (SRB) assay. Cell cycle analysis revealed that hexane, benzene and acetone extracts produced 41, 63 and 43% sub-G1 DNA fraction, demonstrating cell cycle inhibitory potential of these extracts against A549 cell line. Chloroform, ethyl alcohol and aqueous extracts displayed 71, 64 and 65% membrane protective activity, respectively in lipid peroxidation inhibition assay. P. longum fruit extracts also ameliorated A1Cl3-induced hepatotoxicity, as indicated by alterations observed in serum enzymes ALP, SGOT and SGPT activity, as well as creatinine and bilirubin contents. In conclusion, study established the cytotoxic and hepatoprotective activity in P. longum extracts. PMID:25630105

  19. Regulation of TBK1 activity by Optineurin contributes to cell cycle-dependent expression of the interferon pathway.

    PubMed

    Weil, Robert; Laplantine, Emmanuel; Génin, Pierre

    2016-06-01

    The innate immune system has evolved to detect and neutralize viral invasions. Triggering of this defense mechanism relies on the production and secretion of soluble factors that stimulate intracellular antiviral defense mechanisms. The Tank Binding Kinase 1 (TBK1) is a serine/threonine kinase in the innate immune signaling pathways including the antiviral response and the host defense against cytosolic infection by bacteries. Given the critical roles of TBK1, important regulatory mechanisms are required to regulate its activity. Among these, Optineurin (Optn) was shown to negatively regulate the interferon response, in addition to its important role in membrane trafficking, protein secretion, autophagy and cell division. As Optn does not carry any enzymatic activity, its functions depend on its precise subcellular localization and its interaction with other proteins, especially with components of the innate immune pathway. This review highlights advances in our understanding of Optn mechanisms of action with focus on the relationships between Optn and TBK1 and their implication in host defense against pathogens. Specifically, how the antiviral immune system is controlled during the cell cycle by the Optn/TBK1 axis and the physiological consequences of this regulatory mechanism are described. This review may serve to a better understanding of the relationships between the different functions of Optn, including those related to immune responses and its associated pathologies such as primary open-angle glaucoma, amyotrophic lateral sclerosis and Paget's disease of bone. PMID:26976762

  20. Nucleosome architecture throughout the cell cycle

    PubMed Central

    Deniz, Özgen; Flores, Oscar; Aldea, Martí; Soler-López, Montserrat; Orozco, Modesto

    2016-01-01

    Nucleosomes provide additional regulatory mechanisms to transcription and DNA replication by mediating the access of proteins to DNA. During the cell cycle chromatin undergoes several conformational changes, however the functional significance of these changes to cellular processes are largely unexplored. Here, we present the first comprehensive genome-wide study of nucleosome plasticity at single base-pair resolution along the cell cycle in Saccharomyces cerevisiae. We determined nucleosome organization with a specific focus on two regulatory regions: transcription start sites (TSSs) and replication origins (ORIs). During the cell cycle, nucleosomes around TSSs display rearrangements in a cyclic manner. In contrast to gap (G1 and G2) phases, nucleosomes have a fuzzier organization during S and M phases, Moreover, the choreography of nucleosome rearrangements correlate with changes in gene expression during the cell cycle, indicating a strong association between nucleosomes and cell cycle-dependent gene functionality. On the other hand, nucleosomes are more dynamic around ORIs along the cell cycle, albeit with tighter regulation in early firing origins, implying the functional role of nucleosomes on replication origins. Our study provides a dynamic picture of nucleosome organization throughout the cell cycle and highlights the subsequent impact on transcription and replication activity. PMID:26818620

  1. Mechanism of antifungal activity of antimicrobial peptide APP, a cell-penetrating peptide derivative, against Candida albicans: intracellular DNA binding and cell cycle arrest.

    PubMed

    Li, Lirong; Sun, Jin; Xia, Shufang; Tian, Xu; Cheserek, Maureen Jepkorir; Le, Guowei

    2016-04-01

    We investigated the antifungal properties and anti-candidal mechanism of antimicrobial peptide APP. The minimum inhibitory concentration of APP was 8 μM against Candida albicans and Aspeogillus flavus, the concentration against Saccharomyces cerevisiae and Cryptococcus neoformans was 16 μM, while 32 μM inhibited Aspergilla niger and Trichopyton rubrum. APP caused slight depolarization (12.32 ± 0.87%) of the membrane potential of intact C. albicans cells when it exerted its anti-candidal activity and only caused 21.52 ± 0.48% C. albicans cell membrane damage. APP interacted with cell wall membrane, caused potassium efflux and nucleotide leakage. However, confocal fluorescence microscopy experiment and flow cytometry confirmed that FITC-labeled APP penetrated C. albicans cell membrane with 52.31 ± 1.88% cell-penetrating efficiency and accumulated in the cytoplasm. Then, APP interact with C. albicans genomic DNA and completely suppressed DNA migration above weight ratio (peptide/DNA) of 2, and significantly arrested cell cycles during the S-phase (S-phase cell population was 27.09 ± 0.73%, p < 0.05) after penetrating the cell membrane. Results indicated that APP kills C. albicans for efficient cell-penetrating efficiency, strong DNA-binding affinity and significant physiological changes inducing S-phase arrest in intracellular environment. PMID:26743655

  2. Benzo[a]pyrene-induced cell cycle progression occurs via ERK-induced Chk1 pathway activation in human lung cancer cells.

    PubMed

    Wang, Bing-Yen; Wu, Sung-Yu; Tang, Sheau-Chung; Lai, Chien-Hung; Ou, Chu-Chyn; Wu, Ming-Fang; Hsiao, Yi-Min; Ko, Jiunn-Liang

    2015-03-01

    Benzo[a]pyrene (B[a]P) is a potent lung carcinogen derived from tobacco smoking and environmental contamination. This study aimed to investigate the signal transduction pathway responsible for B[a]P-induced non-small cell lung cancer (NSCLC) development. We exposed the human NSCLC cell lines Calu-1, CL3, H1299, CH27, H23, and H1355 to B[a]P and assessed cell cycle progression using flow cytometry. Expression of cell cycle mediators was measured using Western blot analyses and electrophoretic mobility shift assays (EMSAs). B[a]P exposure dramatically induced S-phase accumulation in H1355 cells. Phospho-p53 (Ser15 and Ser20), phospho-ERK, phospho-p38, and Bax were significantly increased in H1355 cells whereas phospho-Rb was decreased in these cells. In addition, B[a]P induced phosphorylation of checkpoint kinase-1 (Chk1) but not Chk2. EMSA experiments revealed a slower migrating band after c-Myc bound the E-box in response to B[a]P treatment, which was abolished upon the addition of the ERK inhibitor PD98059 in H1355 cells. Phospho-ERK inhibition and dominant negative mutant Chk1 expression reversed B[a]P-induced S phase accumulation and downregulated phospho-Chk1 and phospho-ERK expression. Taken together, these results suggest that activation of ERK and its downstream mediator Chk1 may contribute to B[a]P-induced S phase accumulation in H1355 cells. The results could help in the development of lung cancer treatments that target the Chk1 pathway through ERK. PMID:25769181

  3. Age-related neurogenesis decline in the subventricular zone is associated with specific cell cycle regulation changes in activated neural stem cells

    PubMed Central

    Daynac, Mathieu; Morizur, Lise; Chicheportiche, Alexandra; Mouthon, Marc-André; Boussin, François D.

    2016-01-01

    Although neural stem cells (NSCs) sustain continuous neurogenesis throughout the adult lifespan of mammals, they progressively exhibit proliferation defects that contribute to a sharp reduction in subventricular neurogenesis during aging. However, little is known regarding the early age-related events in neurogenic niches. Using a fluorescence-activated cell sorting technique that allows for the prospective purification of the main neurogenic populations from the subventricular zone (SVZ), we demonstrated an early decline in adult neurogenesis with a dramatic loss of progenitor cells in 4 month-old young adult mice. Whereas the activated and quiescent NSC pools remained stable up to 12 months, the proliferative status of activated NSCs was already altered by 6 months, with an overall extension of the cell cycle resulting from a specific lengthening of G1. Whole genome analysis of activated NSCs from 2- and 6-month-old mice further revealed distinct transcriptomic and molecular signatures, as well as a modulation of the TGFβ signalling pathway. Our microarray study constitutes a cogent identification of new molecular players and signalling pathways regulating adult neurogenesis and its early modifications. PMID:26893147

  4. Age-related neurogenesis decline in the subventricular zone is associated with specific cell cycle regulation changes in activated neural stem cells.

    PubMed

    Daynac, Mathieu; Morizur, Lise; Chicheportiche, Alexandra; Mouthon, Marc-André; Boussin, François D

    2016-01-01

    Although neural stem cells (NSCs) sustain continuous neurogenesis throughout the adult lifespan of mammals, they progressively exhibit proliferation defects that contribute to a sharp reduction in subventricular neurogenesis during aging. However, little is known regarding the early age-related events in neurogenic niches. Using a fluorescence-activated cell sorting technique that allows for the prospective purification of the main neurogenic populations from the subventricular zone (SVZ), we demonstrated an early decline in adult neurogenesis with a dramatic loss of progenitor cells in 4 month-old young adult mice. Whereas the activated and quiescent NSC pools remained stable up to 12 months, the proliferative status of activated NSCs was already altered by 6 months, with an overall extension of the cell cycle resulting from a specific lengthening of G1. Whole genome analysis of activated NSCs from 2- and 6-month-old mice further revealed distinct transcriptomic and molecular signatures, as well as a modulation of the TGFβ signalling pathway. Our microarray study constitutes a cogent identification of new molecular players and signalling pathways regulating adult neurogenesis and its early modifications. PMID:26893147

  5. Fangchinoline inhibits rat aortic vascular smooth muscle cell proliferation and cell cycle progression through inhibition of ERK1/2 activation and c-fos expression.

    PubMed

    Zhang, Yong-He; Fang, Lian-Hua; Ku, Bao-Shan

    2003-11-01

    Fangchinoline (FAN; a plant alkaloid isolated from Stephania tetrandrae) is a nonspecific Ca(2+) channel blocker. The objective of the present study was to investigate the effect of FAN on the growth factor-induced proliferation of primary cultured rat aortic smooth muscle cells (RASMCs). FAN significantly inhibited both 5% fetal bovine serum (FBS)- and 50ng/mL platelet-derived growth factor (PDGF)-BB-induced proliferation, [3H]thymidine incorporation into DNA and phosphorylation of extracellular signal-regulated kinase 1/2. In accordance with these findings, FAN revealed blocking of the FBS-inducible progression through G(0)/G(1) to S phase of the cell cycle in synchronized cells and caused a 62% decrease in the early elevation of c-fos expression induced after 5% FBS addition. Furthermore, significant antiproliferative activity of FAN is observed at concentrations below those required to achieve significant inhibition of Ca(2+) channels by FAN. These results suggest that FAN reduced both FBS- and PDGF-BB-induced RASMCs proliferation by perturbing cell cycle progression. This antiproliferative effect of FAN is dependent on the MAP kinase pathway, but cannot be limited to its Ca(2+) modulation. PMID:14563495

  6. Noscapine Increases the Sensitivity of Drug-Resistant Ovarian Cancer Cell Line SKOV3/DDP to Cisplatin by Regulating Cell Cycle and Activating Apoptotic Pathways.

    PubMed

    Shen, Wei; Liang, Bingfeng; Yin, Jie; Li, Xiurong; Cheng, Jianxin

    2015-05-01

    Cisplatin is a first-line chemotherapy drug against ovarian cancer. However, its strong toxic side effects and the development of cisplatin resistance in human cancer cells seriously influence the effects of chemotherapy and quality of life in patients. Noscapine (Nos), a non-toxic benzylisoquinoline alkaloid extracted from opium, has been recently reported to have anti-cancer activity, but the mechanism of that effect has not been clearly established. In the present study, we investigated cytotoxicity of Nos in combination with cisplatin (DDP) in drug-resistant human ovarian cancer cell line SKOV3/DDP in vitro and in vivo null mice xenograft model. Cell proliferation was measured by MTT assay, flow cytometry was used to analyze cell cycle and apoptosis, protein expression of several apoptotic factors was investigated by flow cytometry and immunohistochemical method, and their mRNA expression levels were determined by real-time PCR. In vitro experiments showed that Nos significantly inhibited proliferation of SKOV3/DDP cells. DDP/Nos-combined treatment notably enhanced DDP-induced inhibition of cell proliferation and increased the pro-apoptotic effect of DDP in SKOV3/DDP cells. DDP/Nos administration increased the proportion of G2/M cells, reduced both protein and mRNA expression of anti-apoptotic factors XIAP, surviving and NF-kB, and augmented protein and mRNA levels of pro-apoptotic caspase-3. In vivo experiments revealed that Nos/DDP treatment increased the apoptotic rate of xenograft tumors in null mice. Tumor volume decreased from 1.733 ± 0.155 g in mice treated with DDP alone to 1.191 ± 0.106 g in animals treated with Nos/DDP. These observations suggest that Nos increases the anti-cancer activity of DDP against the drug-resistant ovarian cancer cell line SKOV3/DDP by modulating the cell cycle and activating apoptotic pathways. The study provides a new chemotherapy strategy for the treatment of DDP-resistant human ovarian cancer. PMID:25510462

  7. Autoradiography and the Cell Cycle.

    ERIC Educational Resources Information Center

    Jones, C. Weldon

    1992-01-01

    Outlines the stages of a cell biology "pulse-chase" experiment in which the students apply autoradiography techniques to learn about the concept of the cell cycle. Includes (1) seed germination and plant growth; (2) radioactive labeling and fixation of root tips; (3) feulgen staining of root tips; (4) preparation of autoradiograms; and (5)…

  8. Myricetin and methyl eugenol combination enhances the anticancer activity, cell cycle arrest and apoptosis induction of cis-platin against HeLa cervical cancer cell lines

    PubMed Central

    Yi, Jin-Ling; Shi, Song; Shen, Yan-Li; Wang, Ling; Chen, Hai-Yan; Zhu, Jun; Ding, Yan

    2015-01-01

    Drug combination therapies are common practice in the treatment of cancer. In this study, we evaluated the anticancer effects of myricetin (MYR), methyl eugenol (MEG) and cisplatin (CP) both separately as well as in combination against cervical cancer (HeLa) cells. To demonstrate whether MYR and MEG enhance the anticancer activity of CP against cervical cancer cells, we treated HeLa cells with MYR and MEG alone or in combination with cisplatin and evaluated cell growth and apoptosis using MTT (3 (4, 5 dimethyl thiazol 2yl) 2, 5 diphenyltetrazolium bromide) assay, LDH release assay, flow cytometry and fluorescence microscopy. The results revealed that, as compared to single drug treatment, the combination of MYR or MEG with CP resulted in greater effect in inhibiting cancer cell growth and inducing apoptosis. Cell apoptosis induction, Caspase-3 activity, cell cycle arrest and mitochondrial membrane potential loss were systematically studied to reveal the mechanisms of synergy between MYR, MEG and CP. Combination of MYR or MEG with CP resulted in more potent apoptosis induction as revealed by fluorescence microscopy using Hoechst 33258 and AO-ETBR staining. The combination treatment also increased the number of cells in G0/G1 phase dramatically as compared to single drug treatment. Mitochondrial membrane potential loss (ΛΨm) as well as Caspase-3 activity was much higher in combination treatment as compared to single drug treatment. Findings of this investigation suggest that MYR and MEG combined with cisplatin is a potential clinical chemotherapeutic approach in human cervical cancer. PMID:25972998

  9. Microcystin-LR promotes proliferation by activating Akt/S6K1 pathway and disordering apoptosis and cell cycle associated proteins phosphorylation in HL7702 cells.

    PubMed

    Liu, Jinghui; Wang, Hao; Wang, Beilei; Chen, Tao; Wang, Xiaofeng; Huang, Pu; Xu, Lihong; Guo, Zonglou

    2016-01-01

    Our previous studies had shown that MC-LR inhibited PP2A activity and hyperphosphorylated PP2A substrates at 24 h exposure in HL7702 cells. Although the cytoskeleton was rearranged, the cellular effects were not observed. The purpose of the present study with HL7702 cell exposed to MC-LR for 1-72 h was to further uncover the adverse effects of MC-LR comprehensively. The results showed that there were no obvious difference in apoptosis rate and cell-cycle distribution but the cell proliferation was changed since 36 h exposure while the uptake of MC-LR and its binding to PP2A/C kept unchanged since 1h exposure. PP2A activity had not manifested continued decline compare to 24h exposure and PP2A regulator α4 was found to release its associated PP2A/C since 1h exposure. The increasing of p-Akt-T308, p-Akt-S473, p-S6K1, p-S6, and p-4E-BP1 since 1h MC-LR exposure indicated that Akt/S6K1 cascade had been activated as early as 1h MC-LR treatment. And, PI3K/Akt inhibitor (LY294002) blocked MC-LR-induced Akt/S6K1 activation and proliferation. Besides, MC-LR also led to hyperphosphorylation of c-Myc, c-Jun, Bcl-2 and Bad and activation of Cdk1. Our study indicated that MC-LR exposure promoted HL7702 cell proliferation and the main mechanism was the activation of Akt/S6K1 cascade. Meanwhile, hyperphosphorylation of Bcl-2, Bad, c-Myc and c-Jun might also be involved. And, the inhibition of PP2A was the major reason for these molecular changes. PMID:26506538

  10. Small ubiquitin-related modifier ligase activity of Mms21 is required for maintenance of chromosome integrity during the unperturbed mitotic cell division cycle in Saccharomyces cerevisiae.

    PubMed

    Rai, Ragini; Varma, Satya P M V; Shinde, Nikhil; Ghosh, Shilpa; Kumaran, Srikala P; Skariah, Geena; Laloraya, Shikha

    2011-04-22

    The SUMO ligase activity of Mms21/Nse2, a conserved member of the Smc5/6 complex, is required for resisting extrinsically induced genotoxic stress. We report that the Mms21 SUMO ligase activity is also required during the unchallenged mitotic cell cycle in Saccharomyces cerevisiae. SUMO ligase-defective cells were slow growing and spontaneously incurred DNA damage. These cells required caffeine-sensitive Mec1 kinase-dependent checkpoint signaling for survival even in the absence of extrinsically induced genotoxic stress. SUMO ligase-defective cells were sensitive to replication stress and displayed synthetic growth defects with DNA damage checkpoint-defective mutants such as mec1, rad9, and rad24. MMS21 SUMO ligase and mediator of replication checkpoint 1 gene (MRC1) were epistatic with respect to hydroxyurea-induced replication stress or methyl methanesulfonate-induced DNA damage sensitivity. Subjecting Mms21 SUMO ligase-deficient cells to transient replication stress resulted in enhancement of cell cycle progression defects such as mitotic delay and accumulation of hyperploid cells. Consistent with the spontaneous activation of the DNA damage checkpoint pathway observed in the Mms21-mediated sumoylation-deficient cells, enhanced frequency of chromosome breakage and loss was detected in these mutant cells. A mutation in the conserved cysteine 221 that is engaged in coordination of the zinc ion in Loop 2 of the Mms21 SPL-RING E3 ligase catalytic domain resulted in strong replication stress sensitivity and also conferred slow growth and Mec1 dependence to unchallenged mitotically dividing cells. Our findings establish Mms21-mediated sumoylation as a determinant of cell cycle progression and maintenance of chromosome integrity during the unperturbed mitotic cell division cycle in budding yeast. PMID:21324902

  11. Circulating leptin and NF-κB activation in peripheral blood mononuclear cells across the menstrual cycle.

    PubMed

    Faustmann, Gernot; Tiran, Beate; Maimari, Theopisti; Kieslinger, Petra; Obermayer-Pietsch, Barbara; Gruber, Hans-Jürgen; Roob, Johannes M; Winklhofer-Roob, Brigitte M

    2016-07-01

    Using the menstrual cycle as a model, this study focused on longitudinal changes and associations within a physiological network known to play a role in female fertility, including, as biologically active nodes, NF-κB, leptin and adiponectin, β-carotene, adipose tissue, and progesterone. In 28 women, leptin, adiponectin, β-carotene, and progesterone concentrations, NF-κB p65 and p50 activation in peripheral blood mononuclear cells (known to possess estrogen, progesterone and leptin receptors), total body fat (TBF) and subcutaneous adipose tissue (SAT) mass were determined at early (T1) and late follicular (T2) and mid (T3) and late (T4) luteal phase. Leptin and adiponectin concentrations were higher, while NF-κB p65 activation was lower at T3 compared with T1. NF-κB p65 activation was inversely related to leptin concentrations at T1, T3, and T4. β-Carotene was inversely related to leptin (T1,T2,T4) and SAT (T1,T3,T4). NF-κB p50 activation was inversely related to TBF (T4) and SAT (T3,T4), and leptin was positively related to TBF and SAT (T1-T4). Progesterone was inversely related to leptin (T2,T3), adiponectin (T3), TBF (T3,T4), and SAT (T2,T3,T4). By providing evidence of luteal phase-specific reduced NF-κB p65 activation in women under physiological conditions, this study bridges the gap between existing evidence of a Th1-Th2 immune response shift induced by reduced NF-κB p65 activation and a Th1-Th2 shift previously observed at luteal phase. For the first time, inverse regressions suggest inhibitory effects of leptin on NF-κB p65 activation at luteal phase, along with inhibitory effects of leptin as well as adiponectin on progesterone production in corpus luteum. © 2016 The Authors BioFactors published by Wiley Periodicals, Inc. on behalf of International Union of Biochemistry and Molecular Biology. 24(4):376-387, 2016. PMID:27093900

  12. MLN2238, a proteasome inhibitor, induces caspase-dependent cell death, cell cycle arrest, and potentiates the cytotoxic activity of chemotherapy agents in rituximab-chemotherapy-sensitive or rituximab-chemotherapy-resistant B-cell lymphoma preclinical models.

    PubMed

    Gu, Juan J; Hernandez-Ilizaliturri, Francisco J; Mavis, Cory; Czuczman, Natalie M; Deeb, George; Gibbs, John; Skitzki, Joseph J; Patil, Ritesh; Czuczman, Myron S

    2013-11-01

    To further develop therapeutic strategies targeting the proteasome system, we studied the antitumor activity and mechanisms of action of MLN2238, a reversible proteasome inhibitor, in preclinical lymphoma models. Experiments were conducted in rituximab-chemotherapy-sensitive cell lines, rituximab-chemotherapy-resistant cell lines (RRCL), and primary B-cell lymphoma cells. Cells were exposed to MLN2238 or caspase-dependent inhibitors, and differences in cell viability, alterations in apoptotic protein levels, effects on cell cycle, and the possibility of synergy when combined with chemotherapeutic agents were evaluated. MLN2238 showed more potent dose-dependent and time-dependent cytotoxicity and inhibition of cell proliferation in lymphoma cells than bortezomib. Our data suggest that MLN2238 can induce caspase-independent cell death in RRCL. MLN2238 (and to a much lesser degree bortezomib) reduced RRCL S phase and induced cell cycle arrest in the G2/M phase. Exposure of rituximab-chemotherapy-sensitive cell lines and RRCL to MLN2238 potentiated the cytotoxic effects of gemcitabine, doxorubicin, and paclitaxel and overcame resistance to chemotherapy in RRCL. MLN2238 is a potent proteasome inhibitor active in rituximab-chemotherapy-sensitive and rituximab-chemotherapy-resistant cell models and potentiates the antitumor activity of chemotherapy agents and has the potential of becoming an effective therapeutic agent in the treatment of therapy-resistant B-cell lymphoma. PMID:23995855

  13. Improved Gene Targeting through Cell Cycle Synchronization

    PubMed Central

    Tsakraklides, Vasiliki; Brevnova, Elena; Stephanopoulos, Gregory; Shaw, A. Joe

    2015-01-01

    Gene targeting is a challenge in organisms where non-homologous end-joining is the predominant form of recombination. We show that cell division cycle synchronization can be applied to significantly increase the rate of homologous recombination during transformation. Using hydroxyurea-mediated cell cycle arrest, we obtained improved gene targeting rates in Yarrowia lipolytica, Arxula adeninivorans, Saccharomyces cerevisiae, Kluyveromyces lactis and Pichia pastoris demonstrating the broad applicability of the method. Hydroxyurea treatment enriches for S-phase cells that are active in homologous recombination and enables previously unattainable genomic modifications. PMID:26192309

  14. Properties of stellar activity cycles

    NASA Astrophysics Data System (ADS)

    Korhonen, Heidi

    2015-08-01

    The current photometric datasets, that span decades, allow for studying long-term magentic cycles on active stars. Complementary Ca H&K observations give information also on the cycles of normal solar-like stars, which have significantly smaller, and less easily detectable, spots. In recent years, high precision space-based observations, for example from the Kepler satellite, have allowed also to study the sunspot-like spot sizes in other stars. In this talk I will review what is known about the properties of the cyclic stellar activity in other stars than our Sun, and also discuss the future prospects in this field.

  15. The ethanol extract of Scutellaria baicalensis and the active compounds induce cell cycle arrest and apoptosis including upregulation of p53 and Bax in human lung cancer cells

    SciTech Connect

    Gao Jiayu; Morgan, Winston A.; Sanchez-Medina, Alberto; Corcoran, Olivia

    2011-08-01

    Despite a lack of scientific authentication, Scutellaria baicalensis is clinically used in Chinese medicine as a traditional adjuvant to chemotherapy of lung cancer. In this study, cytotoxicity assays demonstrated that crude ethanolic extracts of S. baicalensis were selectively toxic to human lung cancer cell lines A549, SK-LU-1 and SK-MES-1 compared with normal human lung fibroblasts. The active compounds baicalin, baicalein and wogonin did not exhibit such selectivity. Following exposure to the crude extracts, cellular protein expression in the cancer cell lines was assessed using 2D gel electrophoresis coupled with MALDI-TOF-MS/Protein Fingerprinting. The altered protein expression indicated that cell growth arrest and apoptosis were potential mechanisms of cytotoxicity. These observations were supported by PI staining cell cycle analysis using flow cytometry and Annexin-V apoptotic analysis by fluorescence microscopy of cancer cells treated with the crude extract and pure active compounds. Moreover, specific immunoblotting identification showed the decreased expression of cyclin A results in the S phase arrest of A549 whereas the G{sub 0}/G{sub 1} phase arrest in SK-MES-1 cells results from the decreased expression of cyclin D1. Following treatment, increased expression in the cancer cells of key proteins related to the enhancement of apoptosis was observed for p53 and Bax. These results provide further insight into the molecular mechanisms underlying the clinical use of this herb as an adjuvant to lung cancer therapy. - Research Highlights: > Scutellaria baicalensis is a clinical adjuvant to lung cancer chemotherapy in China. > Scutellaria ethanol extracts selectively toxic to A549, SK-LU-1 and SK-MES-1. > Baicalin, baicalein and wogonin were toxic to all lung cancer cell lines. > Proteomics identified increased p53 and BAX in response to Scutellaria extracts.

  16. Activity of dorsal raphe cells across the sleep–waking cycle and during cataplexy in narcoleptic dogs

    PubMed Central

    Wu, M-F; John, J; Boehmer, L N; Yau, D; Nguyen, G B; Siegel, J M

    2004-01-01

    Cataplexy, a symptom associated with narcolepsy, represents a unique dissociation of behavioural states. During cataplectic attacks, awareness of the environment is maintained, as in waking, but muscle tone is lost, as in REM sleep. We have previously reported that, in the narcoleptic dog, noradrenergic cells of the locus coeruleus cease discharge during cataplexy. In the current study, we report on the activity of serotonergic cells of the dorsal raphe nucleus. The discharge patterns of serotonergic dorsal raphe cells across sleep–waking states did not differ from those of dorsal raphe and locus coeruleus cells recorded in normal rats, cats and monkeys, with tonic discharge in waking, reduced activity in non-REM sleep and cessation of activity in REM sleep. However, in contrast with locus coeruleus cells, dorsal raphe REM sleep-off neurones did not cease discharge during cataplexy. Instead, discharge continued at a level significantly higher than that seen in REM sleep and comparable to that seen in non-REM sleep. We also identified several cells in the dorsal raphe whose pattern of activity was the opposite of that of the presumed serotonergic cells. These cells were maximally active in REM sleep and minimally active in waking and increased activity during cataplexy. The difference between noradrenergic and serotonergic cell discharge profiles in cataplexy suggests different roles for these cell groups in the normal regulation of environmental awareness and muscle tone and in the pathophysiology of narcolepsy. PMID:14678502

  17. Cell Cycle Regulation of DNA Replication

    PubMed Central

    Sclafani, R. A.; Holzen, T. M.

    2008-01-01

    Eukaryotic DNA replication is regulated to ensure all chromosomes replicate once and only once per cell cycle. Replication begins at many origins scattered along each chromosome. Except for budding yeast, origins are not defined DNA sequences and probably are inherited by epigenetic mechanisms. Initiation at origins occurs throughout the S phase according to a temporal program that is important in regulating gene expression during development. Most replication proteins are conserved in evolution in eukaryotes and archaea, but not in bacteria. However, the mechanism of initiation is conserved and consists of origin recognition, assembly of pre-replication (pre-RC) initiative complexes, helicase activation, and replisome loading. Cell cycle regulation by protein phosphorylation ensures that pre-RC assembly can only occur in G1 phase, whereas helicase activation and loading can only occur in S phase. Checkpoint regulation maintains high fidelity by stabilizing replication forks and preventing cell cycle progression during replication stress or damage. PMID:17630848

  18. Natural flavonoids targeting deregulated cell cycle progression in cancer cells.

    PubMed

    Singh, Rana Pratap; Agarwal, Rajesh

    2006-03-01

    The prolonged duration requiring alteration of multi-genetic and epigenetic molecular events for cancer development provides a strong rationale for cancer prevention, which is developing as a potential strategy to arrest or reverse carcinogenic changes before the appearance of the malignant disease. Cell cycle progression is an important biological event having controlled regulation in normal cells, which almost universally becomes aberrant or deregulated in transformed and neoplastic cells. In this regard, targeting deregulated cell cycle progression and its modulation by various natural and synthetic agents are gaining widespread attention in recent years to control the unchecked growth and proliferation in cancer cells. In fact, a vast number of experimental studies convincingly show that many phytochemicals halt uncontrolled cell cycle progression in cancer cells. Among these phytochemicals, natural flavonoids have been identified as a one of the major classes of natural anticancer agents exerting antineoplastic activity via cell cycle arrest as a major mechanism in various types of cancer cells. This review is focused at the modulatory effects of natural flavonoids on cell cycle regulators including cyclin-dependent kinases and their inhibitors, cyclins, p53, retinoblastoma family of proteins, E2Fs, check-point kinases, ATM/ATR and survivin controlling G1/S and G2/M check-point transitions in cell cycle progression, and discusses how these molecular changes could contribute to the antineoplastic effects of natural flavonoids. PMID:16515531

  19. Ferulago angulata activates intrinsic pathway of apoptosis in MCF-7 cells associated with G1 cell cycle arrest via involvement of p21/p27

    PubMed Central

    Karimian, Hamed; Moghadamtousi, Soheil Zorofchian; Fadaeinasab, Mehran; Golbabapour, Shahram; Razavi, Mahboubeh; Hajrezaie, Maryam; Arya, Aditya; Abdulla, Mahmood Ameen; Mohan, Syam; Ali, Hapipah Mohd; Noordin, Mohamad Ibrahim

    2014-01-01

    Ferulago angulata is a medicinal plant that is traditionally known for its anti-inflammatory and antiulcer properties. The present study was aimed to evaluate its anticancer activity and the possible mechanism of action using MCF-7 as an in vitro model. F. angulata leaf extracts were prepared using solvents in the order of increasing polarity. As determined by MTT assay, F. angulata leaves hexane extract (FALHE) revealed the strongest cytotoxicity against MCF-7 cells with the half maximal inhibitory concentration (IC50) value of 5.3±0.82 μg/mL. The acute toxicity study of FALHE provided evidence of the safety of the plant extract. Microscopic and flow cytometric analysis using annexin-V probe showed an induction of apoptosis in MCF-7 by FALHE. Treatment of MCF-7 cells with FALHE encouraged the intrinsic pathway of apoptosis, with cell death transducing signals that reduced the mitochondrial membrane potential with cytochrome c release from mitochondria to cytosol. The released cytochrome c triggered the activation of caspase-9. Meanwhile, the overexpression of caspase-8 suggested the involvement of an extrinsic pathway in the induced apoptosis at the late stage of treatment. Moreover, flow cytometric analysis showed that FALHE treatment significantly arrested MCF-7 cells in the G1 phase, which was associated with upregulation of p21 and p27 assessed by quantitative polymerase chain reaction. Immunofluorescence and the quantitative polymerase chain reaction analysis of MCF-7 cells after treatment with FALHE revealed an upregulation of Bax and a downregulation of Bcl-2 proteins. These findings proposed that FALHE suppressed the proliferation of MCF-7 cells via cell cycle arrest and the induction of apoptosis through intrinsic pathway. PMID:25278746

  20. Activity cycles of M dwarfs

    NASA Astrophysics Data System (ADS)

    Savanov, I. S.

    2012-09-01

    We have determined activity cycles for coolest M dwarfs using photometry from the ASAS survey. The time scales of brightness variations were determined for the program stars using calculated amplitude power spectra and wavelet spectra. Most of ther program stars display periodicities in their light-curve variations, with periods from hundreds of days to years. Analysis of diagrams plotting P cyc/ P rot versus 1/ P rot in logarithmic coordinates shows that the data for all our program objects fit the general relation quite well. No differences in the activity cycles are found for our sample stars, which have different masses and thus internal structures, some having convective envelopes and others being totally convective. Our analysis indicates that the slope i of this relation is close to unity, regardless of whether it is determined from all data, from data for the shortest cycles, or from data for the longest cycles. This value of i differs from values in the literature for stars of other spectral types. Our analysis of the P cyc- P rot relation indicates that the activity cycles for the studied sample of M dwarfs do not depend on the rotation periods of these objects. The data for the studied objects do not agree with any of the relations for relatively young (active) stars or older (less active) stars. The studied M dwarfs probably form another branch of low-mass stars that display more random, irregular magnetic activity on their surfaces, which is generated and supported by the distributed dynamo mechanism or a small-scale dynamo mechanism.

  1. Docetaxel enhances apoptosis and G2/M cell cycle arrest by suppressing mitogen-activated protein kinase signaling in human renal clear cell carcinoma.

    PubMed

    Han, T D; Shang, D H; Tian, Y

    2016-01-01

    Tremendous efforts have been made in renal cell carcinoma (RCC) patients' research; however, clinical findings in patients have been disappointing. The aims of our study were to identify better or alternative therapeutic methods that can reverse chemotherapy resistance and to enhance sensitivity to docetaxel (DOX)-based chemotherapy drugs. We evaluated the anti-proliferative effect of DOX against RCC cells. DOX was found to suppress proliferation of RCC cells under in vitro and in vivo settings. Flow cytometric analysis revealed that DOX suppressed cell growth by induction of both apoptosis and G2/M cell cycle arrest in a dose-dependent manner. Various patterns of gene expression were observed by cluster analysis. In addition, based on network analysis using the ingenuity pathway analysis software, DOX was found to suppress phosphorylation of extracellular signal-regulated kinase 1/2 and p38, suggesting that the mitogen-activated protein kinase signaling pathway plays a vital role in the anti-proliferative effect of DOX against RCC. PMID:26909952

  2. Wogonoside induces growth inhibition and cell cycle arrest via promoting the expression and binding activity of GATA-1 in chronic myelogenous leukemia cells.

    PubMed

    Li, Hui; Hui, Hui; Xu, Jingyan; Yang, Hao; Zhang, Xiaoxiao; Liu, Xiao; Zhou, Yuxin; Li, Zhiyu; Guo, Qinglong; Lu, Na

    2016-06-01

    GATA-1, a zinc finger transcription factor, has been demonstrated to play a key role in the progression of leukemia. In this study, we investigate the effects of wogonoside, a naturally bioactive flavonoid derived from Scutellaria baicalensis Georgi, on cell growth and cell cycle in chronic myeloid leukemia (CML) cells, and uncover its underlying mechanisms. The experimental design comprised CML cell lines K562, imatinib-resistant K562 (K562r) cells, and primary CML cells, treated in vitro or in vivo, respectively, with wogonoside; growth and cell cycle were then evaluated. We found that wogonoside could induce growth inhibition and G0/G1 cell cycle arrest in both normal and K562r cells. Wogonoside promotes the expression of GATA-1 and facilitates the binding to methyl ethyl ketone (MEK) and p21 promoter, thus inhibiting MEK/extracellular signal-regulated kinase signaling and cell cycle checkpoint proteins, including CDK2, CDK4, cyclin A, and cyclin D1, and increasing p21 expression. Furthermore, in vivo studies showed that administration of wogonoside decreased CML cells and prolonged survival in NOD/SCID mice with CML cell xenografts. In conclusion, these results clearly revealed the inhibitory effect of wogonoside on the growth in CML cells and suggested that wogonoside may act as a promising drug for the treatment of imatinib-resistant CML. PMID:26104856

  3. Effect of active fraction of Eriocaulon sieboldianum on human leukemia K562 cells via proliferation inhibition, cell cycle arrest and apoptosis induction.

    PubMed

    Fan, Yanhua; Lu, Hongyuan; An, Li; Wang, Changli; Zhou, Zhipeng; Feng, Fan; Ma, Hongda; Xu, Yongnan; Zhao, Qingchun

    2016-04-01

    Eriocaulon sieboldianum (Sieb. & Zucc. ex Steud.), a genus of Eriocaulon in the Eriocaulaceae family, is an edible and medicinal plant used in traditional Chinese medicine. It was processed into healthcare beverages for expelling wind-heat, protecting eyes, and reducing blood fat. Also, it has been used with other herbs as Traditional Chinese herbal compound to treat cancer as adjuvants in tumor therapy in China. However, the active fractions and precise cellular mechanisms of E. sieboldianum extract remain to be illustrated. The goal of this study was to investigate the effects of the active fraction of E. sieboldianum on the growth of K562 cells and understand the possible mechanisms of its action. Our findings suggested that the fraction E3 of E. sieboldianum could effectively inhibit the activity of Aurora kinase and induce apoptosis via blocking cell cycle, up-regulating the expression of proapoptotic proteins including p53 and Bax and reducing the expression of Bcl-2. The levels of Cytochrome C, cleaved caspase-9, cleaved caspase-3 and cleaved PARP were also found to be increased after treatment with fraction E3 of E. sieboldianum. This study could improve the development of E. sieboldianum and raise its application value in cancer adjuvant therapy. Considering it is both a dietary supplement and a traditional Chinese herbal medicine which exhibits anticancer activities, it can be developed into functional food. PMID:26923230

  4. Loss of DDB1 Leads to Transcriptional p53 Pathway Activation in Proliferating Cells, Cell Cycle Deregulation, and Apoptosis in Zebrafish Embryos

    PubMed Central

    Hu, Zhilian; Holzschuh, Jochen; Driever, Wolfgang

    2015-01-01

    DNA damage-binding protein 1 (DDB1) is a large subunit of the heterodimeric DDB complex that recognizes DNA lesions and initiates the nucleotide excision repair process. DDB1 is also a component of the CUL4 E3 ligase complex involved in a broad spectrum of cellular processes by targeted ubiquitination of key regulators. Functions of DDB1 in development have been addressed in several model organisms, however, are not fully understood so far. Here we report an ENU induced mutant ddb1 allele (ddb1m863) identified in zebrafish (Danio rerio), and analyze its effects on development. Zebrafish ddb1 is expressed broadly, both maternally and zygotically, with enhanced expression in proliferation zones. The (ddb1m863 mutant allele affects the splice acceptor site of exon 20, causing a splicing defect that results in truncation of the 1140 amino acid protein after residue 800, lacking part of the β-propeller domain BPC and the C-terminal helical domain CTD. ddb1m863 zygotic mutant embryos have a pleiotropic phenotype, including smaller and abnormally shaped brain, head skeleton, eyes, jaw, and branchial arches, as well as reduced dopaminergic neuron groups. However, early forming tissues develop normally in zygotic ddb1m863 mutant embryos, which may be due to maternal rescue. In ddb1m863 mutant embryos, pcna-expressing proliferating cell populations were reduced, concurrent with increased apoptosis. We also observed a concomitant strong up-regulation of transcripts of the tumor suppressor p53 (tp53) and the cell cycle inhibitor cdkn1a (p21a/bCIP1/WAF1) in proliferating tissues. In addition, transcription of cyclin genes ccna2 and ccnd1 was deregulated in ddb1m863 mutants. Reduction of p53 activity by anti-sense morpholinos alleviated the apoptotic phenotype in ddb1m863 mutants. These results imply that Ddb1 may be involved in maintaining proper cell cycle progression and viability of dividing cells during development through transcriptional mechanisms regulating genes

  5. Cell cycle arrest or survival signaling through αv integrins, activation of PKC and ERK1/2 lead to anoikis resistance of ovarian cancer spheroids.

    PubMed

    Carduner, Ludovic; Picot, Cédric R; Leroy-Dudal, Johanne; Blay, Lyvia; Kellouche, Sabrina; Carreiras, Franck

    2014-01-15

    Ovarian cancer is the most lethal gynecologic cancer mainly due to spheroids organization of cancer cells that disseminate within the peritoneal cavity. We have investigated the molecular mechanisms by which ovarian cancer spheroids resist anoikis, choosing as models the 2 well-characterized human ovarian cancer cell lines IGROV1 and SKOV3. These cell lines have the propensity to float as clusters, and were isolated from tumor tissue and ascites, respectively. To form spheroids, IGROV1 and SKOV3 ovarian adenocarcinoma cells were maintained under anchorage-independent culture conditions, in which both lines survive at least a week. A short apoptotic period prior to a survival signaling commitment was observed for IGROV1 cells whereas SKOV3 cells entered G0/G1 phase of the cell cycle. This difference in behavior was due to different signals. With regard to SKOV3 cells, activation of p38 and an increase in p130/Rb occurred once anchorage-independent culture was established. Analyses of the survival signaling pathway switched on by IGROV1 cells showed that activation of ERK1/2 was required to evade apoptosis, an effect partly dependent on PKC activation and αv integrins. αv-integrin expression is essential for survival through activation of ERK1/2 phosphorylation. The above data indicate that ovarian cancer cells can resist anoikis in the spheroid state by arrest in the cell cycle or through activation of αv-integrin-ERK-mediated survival signals. Such signaling might result in the selection of resistant cells within disseminating spheroids, favoring further relapse in ovarian cancers. PMID:24291221

  6. c-Myc Regulates Cyclin D-Cdk4 and -Cdk6 Activity but Affects Cell Cycle Progression at Multiple Independent Points

    PubMed Central

    Mateyak, Maria K.; Obaya, Alvaro J.; Sedivy, John M.

    1999-01-01

    c-myc is a cellular proto-oncogene associated with a variety of human cancers and is strongly implicated in the control of cellular proliferation, programmed cell death, and differentiation. We have previously reported the first isolation of a c-myc-null cell line. Loss of c-Myc causes a profound growth defect manifested by the lengthening of both the G1 and G2 phases of the cell cycle. To gain a clearer understanding of the role of c-Myc in cellular proliferation, we have performed a comprehensive analysis of the components that regulate cell cycle progression. The largest defect observed in c-myc−/− cells is a 12-fold reduction in the activity of cyclin D1-Cdk4 and -Cdk6 complexes during the G0-to-S transition. Downstream events, such as activation of cyclin E-Cdk2 and cyclin A-Cdk2 complexes, are delayed and reduced in magnitude. However, it is clear that c-Myc affects the cell cycle at multiple independent points, because restoration of the Cdk4 and -6 defect does not significantly increase growth rate. In exponentially cycling cells the absence of c-Myc reduces coordinately the activities of all cyclin–cyclin-dependent kinase complexes. An analysis of cyclin-dependent kinase complex regulators revealed increased expression of p27KIP1 and decreased expression of Cdk7 in c-myc−/− cells. We propose that c-Myc functions as a crucial link in the coordinate adjustment of growth rate to environmental conditions. PMID:10373516

  7. Isolation of austroinulin possessing cell cycle inhibition activity from Blumea glomerata and revision of its absolute configuration.

    PubMed

    Ohtsuki, Takashi; Koyano, Takashi; Kowithayakorn, Thaworn; Yamaguchi, Naoto; Ishibashi, Masami

    2004-12-01

    A labdane-diterpene, austroinulin (1), together with several known flavonoids and sesquiterpenes were isolated from leaves of Blumea glomerata (Compositae). Austroinulin (1) and most of the flavonoids showed cytotoxicity against HeLa cells, while austroinulin (1) exhibited a cell cycle inhibition effect at the G1 stage at the concentration of 15.2 microg/mL (47.2 microM). The absolute configuration of 1 was revised as 5S,6R,7S,8S,9R,10R on the basis of the modified Mosher's method. PMID:15643553

  8. Synthesis, Anticancer Activity, Effect on Cell Cycle Profile, and Apoptosis-Inducing Ability of Novel Hexahydrocyclooctathieno[2,3-d]pyrimidine Derivatives.

    PubMed

    Kassab, Asmaa Elsayed; Gedawy, Ehab Mohamed; El-Malah, Afaf Ali; Abdelghany, Tamer Mohamed; Abdel-Bakky, Mohamed Sadek

    2016-01-01

    A novel series of hexahydrocyclooctathieno[2,3-d]pyrimidines was synthesized. Investigation of the anticancer activity of these derivatives revealed that compounds 2a and b showed broad-spectrum anticancer activity in nanomolar to micromolar concentrations. In particular, compound 2b showed a concentration required for 50% inhibition of cell growth (GI50) value of less than 1 µM against 20 cancer cell lines. Compounds 2a and b induced G2/M- and S-phase cell cycle arrest in human colon adenocarcinoma (HCT116) and human breast adenocarcinoma (MCF7) cell lines with a concomitant increase in the pre-G cell population in a time-dependent manner. Furthermore, compound 2b increased the nuclear expression of the proapoptotic protein cleaved caspase-3, indicating that apoptosis has an important role, at least in part, in the cancer cell death induced by the new compounds. PMID:27150481

  9. Cell heterogeneity during the cell cycle

    SciTech Connect

    Darzynkiewicz, Z.; Crissman, H.; Traganos, F.; Steinkamp, J.

    1982-12-01

    Using flow cytometry, populations of Chinese hamster ovary cells, asynchronous and synchronized in the cycle, were measured with respect to cellular RNA- and protein-content, as well as cell light scatter properties. Heterogeneities of cell populations were expressed as coefficients of variation (c.v.) in percent of the respective mean values. Populations of cells immediately after mitosis have about 15% higher c.v. than mitotic cell populations, regardless of whether RNA, proteins, or light scatter are measured. These data indicate that cytoplasmic constituents are unequally distributed into the daughter cells during cytokinesis and that unequal cytokinesis generates intercellular metabolic variability during the cycle. An additional increase in heterogeneity, although of smaller degree, occurs during G/sub 2/ phase. Populations of S-phase cells are the most uniform, having 20-30% lower c.v. than the postmitotic cells. Cell progression through S does not involve any significant increase in intercellular variability with respect to RNA or protein content. In unperturbed exponentially growing cultures a critical RNA content is required for G/sub 1/ cells prior to their entrance into S. The cell residence times in the equalization compartments are exponentially distributed, which may reflect the randomness generated by the uneven division of metabolic constituents to daughter cells during cytokinesis. The cell heterogeneities were presently estimated at two metabolic levels, transcription (RNA content) and translation (proteins). The most uniform were populations stained for RNA and the highest variability was observed after staining of proteins. This suggests that the regulatory mechanisms equalizing cells in the cell cycle may operate primarily at the level of DNA transcription.

  10. Cell cycle-specific effects of lovastatin.

    PubMed Central

    Jakóbisiak, M; Bruno, S; Skierski, J S; Darzynkiewicz, Z

    1991-01-01

    Lovastatin (LOV), the drug recently introduced to treat hypercholesteremia, inhibits the synthesis of mevalonic acid. The effects of LOV on the cell cycle progression of the human bladder carcinoma T24 cell line expressing activated p21ras were investigated. At a concentration of 2-10 microM, LOV arrested cells in G1 and also prolonged--or arrested a minor fraction of cells in--the G2 phase of the cell cycle; at a concentration of 50 microM, LOV was cytotoxic. The cytostatic effects were reversed by addition of exogenous mevalonate. Cells arrested in the cycle by LOV were viable for up to 72 hr and did not show any changes in RNA or protein content or chromatin condensation, which would be typical of either unbalanced growth or deep quiescence. The expression of the proliferation-associated nuclear proteins Ki-67 and p105 in these cells was reduced by up to 72% and 74%, respectively, compared with exponentially growing control cells. After removal of LOV, the cells resumed progression through the cycle; they entered S phase asynchronously after a lag of approximately 6 hr. Because mevalonate is essential for the posttranslational modification (isoprenylation) of p21ras, which in turn allows this protein to become attached to the cell membrane, the data suggest that the LOV-induced G1 arrest may be a consequence of the loss of the signal transduction capacity of p21ras. Indeed, while exposure of cells to LOV had no effect on the cellular content of p21ras (detected immunocytochemically), it altered the intracellular location of this protein, causing its dissociation from the cell membrane and translocation toward the cytoplasm and nucleus. However, it is also possible that inhibition of isoprenylation of proteins other than p21ras (e.g., nuclear lamins) by LOV may be responsible for the observed suppression of growth of T24 cells. Images PMID:1673788

  11. Cell cycle checkpoint regulators reach a zillion

    PubMed Central

    Yasutis, Kimberly M.; Kozminski, Keith G.

    2013-01-01

    Entry into mitosis is regulated by a checkpoint at the boundary between the G2 and M phases of the cell cycle (G2/M). In many organisms, this checkpoint surveys DNA damage and cell size and is controlled by both the activation of mitotic cyclin-dependent kinases (Cdks) and the inhibition of an opposing phosphatase, protein phosphatase 2A (PP2A). Misregulation of mitotic entry can often lead to oncogenesis or cell death. Recent research has focused on discovering the signaling pathways that feed into the core checkpoint control mechanisms dependent on Cdk and PP2A. Herein, we review the conserved mechanisms of the G2/M transition, including recently discovered upstream signaling pathways that link cell growth and DNA replication to cell cycle progression. Critical consideration of the human, frog and yeast models of mitotic entry frame unresolved and emerging questions in this field, providing a prediction of signaling molecules and pathways yet to be discovered. PMID:23598718

  12. Acacetin inhibits cell growth and cell cycle progression, and induces apoptosis in human prostate cancer cells: structure-activity relationship with linarin and linarin acetate.

    PubMed

    Singh, Rana P; Agrawal, Puja; Yim, Dongsool; Agarwal, Chapla; Agarwal, Rajesh

    2005-04-01

    This study was carried out to assess the anticancer efficacy of linarin (LN), linarin acetate (LA) and acacetin (AC), the flavonoid compounds with the same flavone ring structure but different substitution, against human prostate cancer (PCA), LNCaP and DU145 cells. LN was isolated and purified from Chrysanthemum zawadskii; LA was chemically synthesized from LN, and AC obtained commercially. In each case, the cells were treated with these agents at 25-100 microM doses for 24-72 h. LN and LA showed moderate cell growth inhibition with different time kinetics as compared to AC. LN caused up to a 5-fold increase in cell death and LA enhanced cell death by up to 4-fold with the increase in treatment time in both cell lines. AC showed a time- as well as dose-dependent stronger cell growth inhibition (20-70%) accompanied by cell death as compared to LN and LA in both the cell lines. LN or LA did not show any profound effect on cell cycle arrest except for a moderate G1 arrest, whereas, AC showed a stronger G1 and/or G2-M arrest depending on the doses and treatment times. G1 arrest was associated with an increase in Cip1/p21 and a decrease in CDK2, CDK4 and CDK6 protein levels. G2-M arrest was associated with a decrease in Cdc25C, Cdc2/p34 and cyclin B1, which were more prominent in LNCaP compared to DU145 cells. LN, LA and AC induced cell death was associated with significant increase in apoptosis induction (up to 5-6-fold) accompanied by poly-(ADP-ribose) polymerase cleavage. Overall, AC showed more potent anticancer efficacy among these three flavonoids, which was diminished when its flavone ring was modified by disaccharide rhamnose substitution at C7 (LN) or acetylation of this substituted group (LA). These findings, for the first time, revealed the structural determinants in anticancer efficacy and mechanisms of these three flavonoids against human PCA cells. PMID:15637089

  13. Chk1 keeps cell cycle transcription active in response to replication stress by interfering with E2F6-dependent repression

    PubMed Central

    Bertoli, Cosetta; Klier, Steffi; McGowan, Clare; Wittenberg, Curt; de Bruin, Robertus A. M.

    2014-01-01

    Background In eukaryotic cells, detection of replication stress results in the activation of the DNA replication checkpoint, a signaling cascade whose central players are the kinases ATR and Chk1. The checkpoint response prevents the accumulation of DNA damage and ensures cell viability by delaying progression into mitosis. However, the role and mechanism of the replication checkpoint transcriptional response in human cells, which is p53-independent, is largely unknown. Results We show that, in response to DNA replication stress, the regular E2F-dependent cell cycle transcriptional program is maintained at high levels and we establish the mechanisms governing such transcriptional upregulation. E2F6, a repressor of E2F-dependent G1/S transcription, replaces the activating E2Fs at promoters to repress transcription in cells progressing into S-phase in unperturbed conditions. Following replication stress, the checkpoint kinase Chk1 phosphorylates E2F6 leading to its dissociation from promoters. This promotes E2F-dependent transcription, which mediates cell survival by preventing DNA damage and cell death. Conclusions This work reveals, for the first time, that the regular cell cycle transcriptional program is part of the DNA replication checkpoint response in human cells and establishes the molecular mechanism involved. We show that maintaining high levels of G1/S cell cycle transcription in response to replication stress contributes to two key functions of the DNA replication checkpoint response, namely preventing genomic instability and cell death. Given the critical role of replication stress in oncogene transformation, a detailed understanding of the molecular mechanisms involved in the checkpoint response will contribute to a better insight into cancer development. PMID:23954429

  14. Stellar activity cycles and asteroseismology

    NASA Astrophysics Data System (ADS)

    Salabert, D.

    2011-12-01

    The success of helioseismology is due to its capability to accurately measure the p-mode parameters of the solar eigenmode spectrum, which allow us to infer unique information about the internal structure and dynamics of the Sun from its surface all the way down to the core. It has contributed greatly to a clearer understanding of the Sun and provided insights into the complex solar magnetism, by means for instance of the variability of the characteristics of the p-mode spectrum. Indeed, variations in the mean strength of the solar magnetic field lead to significant shifts in the frequencies of even the lowest-degree p modes with high levels of correlation with solar surface activity proxies. These frequency shifts are explained to arise from structural changes in the outer layers of the Sun during the 11-year activity cycle, which is understood to be driven by a dynamo process. However, clear differences between p-mode frequencies and solar surface activity during the unusually extended minimum of cycle 23 were observed. The origin of the p-mode variability is thus far from being properly understood and a better comprehension of its relationship with solar and stellar activity cycles will help us in our understanding of the dynamo processes. Spectroscopic measurements of Ca H and K emission lines revealed magnetic activity variations in a large sample of solar-type stars with timescales ranging from 2.5 and 25 years. This broad range of cycle periods is thought to reflect differences in the rotational properties and the depths of the surface convection zones with various masses and ages. However, spectroscopic measurements are only good proxies of surface magnetic fields. The recent discovery of variations with magnetic activity in the p-mode oscillation frequencies of the solar-like star HD 49933 observed by CoRoT, with a frequency dependence comparable in shape to the one observed in the Sun, opens a new era in the study of the physical phenomena involved in the

  15. The cell cycle and acute kidney injury

    PubMed Central

    Price, Peter M.; Safirstein, Robert L.; Megyesi, Judit

    2009-01-01

    Acute kidney injury (AKI) activates pathways of cell death and cell proliferation. Although seemingly discrete and unrelated mechanisms, these pathways can now be shown to be connected and even to be controlled by similar pathways. The dependence of the severity of renal-cell injury on cell cycle pathways can be used to control and perhaps to prevent acute kidney injury. This review is written to address the correlation between cellular life and death in kidney tubules, especially in acute kidney injury. PMID:19536080

  16. Synchronized Cell Cycle Arrest Promotes Osteoclast Differentiation

    PubMed Central

    Kwon, Minsuk; Kim, Jin-Man; Lee, Kyunghee; Park, So-Young; Lim, Hyun-Sook; Kim, Taesoo; Jeong, Daewon

    2016-01-01

    Osteoclast progenitors undergo cell cycle arrest before differentiation into osteoclasts, induced by exposure to macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor-κB ligand (RANKL). The role of such cell cycle arrest in osteoclast differentiation has remained unclear, however. We here examined the effect of synchronized cell cycle arrest on osteoclast formation. Osteoclast progenitors deprived of M-CSF in culture adopted a uniform morphology and exhibited cell cycle arrest at the G0–G1 phase in association with both down-regulation of cyclins A and D1 as well as up-regulation of the cyclin-dependent kinase inhibitor p27Kip1. Such M-CSF deprivation also promoted the differentiation of osteoclast progenitors into multinucleated osteoclasts expressing high levels of osteoclast marker proteins such as NFATc1, c-Fos, Atp6v0d2, cathepsin K, and integrin β3 on subsequent exposure to M-CSF and RANKL. Our results suggest that synchronized arrest and reprogramming of osteoclast progenitors renders them poised to respond to inducers of osteoclast formation. Further characterization of such effects may facilitate induction of the differentiation of heterogeneous and multipotent cells into desired cell lineages. PMID:27517906

  17. Synchronized Cell Cycle Arrest Promotes Osteoclast Differentiation.

    PubMed

    Kwon, Minsuk; Kim, Jin-Man; Lee, Kyunghee; Park, So-Young; Lim, Hyun-Sook; Kim, Taesoo; Jeong, Daewon

    2016-01-01

    Osteoclast progenitors undergo cell cycle arrest before differentiation into osteoclasts, induced by exposure to macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor-κB ligand (RANKL). The role of such cell cycle arrest in osteoclast differentiation has remained unclear, however. We here examined the effect of synchronized cell cycle arrest on osteoclast formation. Osteoclast progenitors deprived of M-CSF in culture adopted a uniform morphology and exhibited cell cycle arrest at the G₀-G₁ phase in association with both down-regulation of cyclins A and D1 as well as up-regulation of the cyclin-dependent kinase inhibitor p27(Kip1). Such M-CSF deprivation also promoted the differentiation of osteoclast progenitors into multinucleated osteoclasts expressing high levels of osteoclast marker proteins such as NFATc1, c-Fos, Atp6v0d2, cathepsin K, and integrin β3 on subsequent exposure to M-CSF and RANKL. Our results suggest that synchronized arrest and reprogramming of osteoclast progenitors renders them poised to respond to inducers of osteoclast formation. Further characterization of such effects may facilitate induction of the differentiation of heterogeneous and multipotent cells into desired cell lineages. PMID:27517906

  18. Single-cell vs. bulk activity properties of coastal bacterioplankton over an annual cycle in a temperate ecosystem.

    PubMed

    Morán, Xosé Anxelu G; Calvo-Díaz, Alejandra

    2009-01-01

    The connections between single-cell activity properties of heterotrophic planktonic bacteria and whole community metabolism are still poorly understood. Here, we show flow cytometry single-cell analysis of membrane-intact (live), high nucleic acid (HNA) content and actively respiring (CTC+) bacteria with samples collected monthly during 2006 in northern Spain coastal waters. Bulk activity was assessed by measuring 3H-Leucine incorporation and specific growth rates. Consistently, different single-cell relative abundances were found, with 60-100% for live, 30-84% for HNA and 0.2-12% for CTC+ cells. Leucine incorporation rates (2-153 pmol L(-1) h(-1)), specific growth rates (0.01-0.29 day(-1)) and the total and relative abundances of the three single-cell groups showed marked seasonal patterns. Distinct depth distributions during summer stratification and different relations with temperature, chlorophyll and bacterial biovolume suggest the existence of different controlling factors on each single-cell property. Pooled leucine incorporation rates were similarly correlated with the abundance of all physiological groups, while specific growth rates were only substantially explained by the percentage of CTC+ cells. However, the ability to reduce CTC proved notably better than the other two single-cell properties at predicting bacterial bulk rates within seasons, suggesting a tight linkage between bacterial individual respiration and biomass production at the community level. PMID:19120458

  19. PLK-1: Angel or devil for cell cycle progression.

    PubMed

    Kumar, Shiv; Sharma, Ashish Ranjan; Sharma, Garima; Chakraborty, Chiranjib; Kim, Jaebong

    2016-04-01

    PLK-1 is a key player in the eukaryotic cell cycle. Cell cycle progression is precisely controlled by cell cycle regulatory kinases. PLK-1 is a mitotic kinase that actively regulates the G2/M transition, mitosis, mitotic exit, and cytokinesis. During cell cycle progression, PLK-1 controls various events related to the cell cycle maturation, directly and/or indirectly. On the contrary, aberrant expression of PLK-1 is strongly associated with tumorigenesis and its poor prognosis. The misexpression of PLK-1 causes the abnormalities including aneuploidy, mitotic defects, leading to tumorigenesis through inhibiting the p53 and pRB genes. Therefore, we reviewed the role of PLK-1 in the cell cycle progression and in the tumorigenesis either as a cell cycle regulator or on an attractive anti-cancer drug target. PMID:26899266

  20. Antitumor Activity of Tenacissoside H on Esophageal Cancer through Arresting Cell Cycle and Regulating PI3K/Akt-NF-κB Transduction Cascade

    PubMed Central

    Jia, Yong-sen; Hu, Xue-qin; Gabriella, Hegyi; Qin, Li-juan; Meggyeshazi, Nora

    2015-01-01

    Objective. The purpose of the study was to elucidate the molecular mechanism of tenacissoside H (TDH) inhibiting esophageal carcinoma infiltration and proliferation. Methods. In vitro, EC9706 cells were treated with TDH. Cells proliferation and cell cycle were assayed. PI3K and NF-κB mRNAs expression were determined by real time PCR. In vivo, model of nude mice with tumor was established. Mice were treated with TDH. Inhibition ratio of tumor volume was calculated. PCNA expression was examined. Protein expression in PI3K/Akt-NF-κB signaling pathway was determined. Results. In vitro, TDH significantly inhibited cells proliferation in a time-and-dose-dependent manner. TDH arrested the cell cycle in S phase and significantly inhibited PI3K and NF-κB mRNA expression, compared with blank controlled group (P < 0.05). In vivo, TDH strongly inhibits tumor growth and volume. PCNA expression was significantly decreased after treatment of TDH. TDH downregulated proteins expression in PI3K/Akt-NF-κB transduction cascade (P < 0.05). Conclusion. TDH inhibited esophageal carcinoma infiltration and proliferation both in vitro and in vivo. The anticancer activity has relation to arresting the cell cycle at the S phase, inhibited the PCNA expression of transplanted tumors in nude mice, and regulated the protein expression in the PI3K/Akt-NF-κB transduction cascade. PMID:26495015

  1. Functional interplay between the cell cycle and cell phenotypes.

    PubMed

    Chen, Wei-Chiang; Wu, Pei-Hsun; Phillip, Jude M; Khatau, Shyam B; Choi, Jae Min; Dallas, Matthew R; Konstantopoulos, Konstantinos; Sun, Sean X; Lee, Jerry S H; Hodzic, Didier; Wirtz, Denis

    2013-03-01

    Cell cycle distribution of adherent cells is typically assessed using flow cytometry, which precludes the measurements of many cell properties and their cycle phase in the same environment. Here we develop and validate a microscopy system to quantitatively analyze the cell-cycle phase of thousands of adherent cells and their associated cell properties simultaneously. This assay demonstrates that population-averaged cell phenotypes can be written as a linear combination of cell-cycle fractions and phase-dependent phenotypes. By perturbing the cell cycle through inhibition of cell-cycle regulators or changing nuclear morphology by depletion of structural proteins, our results reveal that cell cycle regulators and structural proteins can significantly interfere with each other's prima facie functions. This study introduces a high-throughput method to simultaneously measure the cell cycle and phenotypes at single-cell resolution, which reveals a complex functional interplay between the cell cycle and cell phenotypes. PMID:23319145

  2. Antitumoral activity of the mithralog EC-8042 in triple negative breast cancer linked to cell cycle arrest in G2

    PubMed Central

    Pandiella, Atanasio; Morís, Francisco; Ocaña, Alberto; Núñez, Luz-Elena; Montero, Juan C.

    2015-01-01

    Triple negative breast cancer (TNBC) is an aggressive form of breast cancer. Despite response to chemotherapy, relapses are frequent and resistance to available treatments is often observed in the metastatic setting. Therefore, identification of new therapeutic strategies is required. Here we have investigated the effect of the mithramycin analog EC-8042 (demycarosil-3D-β-D-digitoxosyl mithramycin SK) on TNBC. The drug caused a dose-dependent inhibition of proliferation of a set of TNBC cell lines in vitro, and decreased tumor growth in mice xenografted with TNBC cells. Mechanistically, EC-8042 caused an arrest in the G2 phase of the cell cycle, coincident with an increase in pCDK1 and Wee1 levels in cells treated with the drug. In addition, prolonged treatment with the drug also causes apoptosis, mainly through caspase-independent routes. Importantly, EC-8042 synergized with drugs commonly used in the therapy of TNBC in vitro, and potentiated the antitumoral effect of docetaxel in vivo. Together, these data suggest that the mithralog EC-8042 exerts an antitumoral action on TNBC cells and reinforces the action of standard of care drugs used in the therapy of this disease. These characteristics, together with a better toxicology profile of EC-8042 with respect to mithramycin, open the possibility of its clinical evaluation. PMID:26439989

  3. Therapeutic strategy for hair regeneration: Hair cycle activation, niche environment modulation, wound-induced follicle neogenesis and stem cell engineering

    PubMed Central

    Chueh, Shan-Chang; Lin, Sung-Jan; Chen, Chih-Chiang; Lei, Mingxing; Wang, Ling Mei; Widelitz, Randall B.; Hughes, Michael W.; Jiang, Ting-Xing; Chuong, Cheng Ming

    2013-01-01

    Introduction There are major new advancements in the fields of stem cell biology, developmental biology, regenerative hair cycling, and tissue engineering. The time is ripe to integrate, translate and apply these findings to tissue engineering and regenerative medicine. Readers will learn about new progress in cellular and molecular aspects of hair follicle development, regeneration and potential therapeutic opportunities these advances may offer. Areas covered Here we use hair follicle formation to illustrate this progress and to identify targets for potential strategies in therapeutics. Hair regeneration is discussed in four different categories. (1) Intra-follicle regeneration (or renewal) is the basic production of hair fibers from hair stem cells and dermal papillae in existing follicles. (2) Chimeric follicles via epithelial-mesenchymal recombination to identify stem cells and signaling centers. (3) Extra-follicular factors including local dermal and systemic factors can modulate the regenerative behavior of hair follicles, and may be relatively easy therapeutic targets. (4) Follicular neogenesis means the de novo formation of new follicles. In addition, scientists are working to engineer hair follicles, which require hair forming competent epidermal cells and hair inducing dermal cells. Expert opinion Ideally self-organizing processes similar to those occurring during embryonic development should be elicited with some help from biomaterials. PMID:23289545

  4. Antitumoral activity of the mithralog EC-8042 in triple negative breast cancer linked to cell cycle arrest in G2.

    PubMed

    Pandiella, Atanasio; Morís, Francisco; Ocaña, Alberto; Núñez, Luz-Elena; Montero, Juan C

    2015-10-20

    Triple negative breast cancer (TNBC) is an aggressive form of breast cancer. Despite response to chemotherapy, relapses are frequent and resistance to available treatments is often observed in the metastatic setting. Therefore, identification of new therapeutic strategies is required. Here we have investigated the effect of the mithramycin analog EC-8042 (demycarosil-3D-β-D-digitoxosyl mithramycin SK) on TNBC. The drug caused a dose-dependent inhibition of proliferation of a set of TNBC cell lines in vitro, and decreased tumor growth in mice xenografted with TNBC cells. Mechanistically, EC-8042 caused an arrest in the G2 phase of the cell cycle, coincident with an increase in pCDK1 and Wee1 levels in cells treated with the drug. In addition, prolonged treatment with the drug also causes apoptosis, mainly through caspase-independent routes. Importantly, EC-8042 synergized with drugs commonly used in the therapy of TNBC in vitro, and potentiated the antitumoral effect of docetaxel in vivo. Together, these data suggest that the mithralog EC-8042 exerts an antitumoral action on TNBC cells and reinforces the action of standard of care drugs used in the therapy of this disease. These characteristics, together with a better toxicology profile of EC-8042 with respect to mithramycin, open the possibility of its clinical evaluation. PMID:26439989

  5. Cell cycle arrest in Metformin treated breast cancer cells involves activation of AMPK, downregulation of cyclin D1, and requires p27Kip1 or p21Cip1

    PubMed Central

    Zhuang, Yongxian; Miskimins, W Keith

    2008-01-01

    Background The antihyperglycemic drug metformin may have beneficial effects on the prevention and treatment of cancer. Metformin is known to activate AMP-activated protein kinase (AMPK). It has also been shown to inhibit cyclin D1 expression and proliferation of some cultured cancer cells. However, the mechanisms of action by which metformin mediates cell cycle arrest are not completely understood. Results In this study, metformin was found to inhibit proliferation of most cultured breast cancer cell lines. This was independent of estrogen receptor, HER2, or p53 status. Inhibition of cell proliferation was associated with arrest within G0/G1 phase of the cell cycle. As in previous studies, metformin treatment led to activation of (AMPK) and downregulation of cyclin D1. However, these events were not sufficient for cell cycle arrest because they were also observed in the MDA-MB-231 cell line, which is not sensitive to growth arrest by metformin. In sensitive breast cancer lines, the reduction in cyclin D1 led to release of sequestered CDK inhibitors, p27Kip1 and p21Cip1, and association of these inhibitors with cyclin E/CDK2 complexes. The metformin-resistant cell line MDA-MB-231 expresses significantly lower levels of p27Kip1 and p21Cip1 than the metformin-sensitive cell line, MCF7. When p27Kip1 or p21Cip1 were overexpressed in MDA-MB-231, the cells became sensitive to cell cycle arrest in response to metformin. Conclusion Cell cycle arrest in response to metformin requires CDK inhibitors in addition to AMPK activation and cyclin D1 downregulation. This is of interest because many cancers are associated with loss or downregulation of CDK inhibitors and the results may be relevant to the development of anti-tumor reagents that target the AMPK pathway. PMID:19046439

  6. p28-Mediated Activation of p53 in G2-M Phase of the Cell Cycle Enhances the Efficacy of DNA Damaging and Antimitotic Chemotherapy.

    PubMed

    Yamada, Tohru; Das Gupta, Tapas K; Beattie, Craig W

    2016-04-15

    p28 is an anionic cell-penetrating peptide of 28 amino acids that activates wild-type and mutated p53, leading subsequently to selective inhibition of CDK2 and cyclin A expression and G2-M cell-cycle arrest. In this study, we investigated the cytotoxic effects of p28 treatment alone and in combination with DNA-damaging and antimitotic agents on human cancer cells. p28 enhanced the cytotoxic activity of lower concentrations (IC20-50) of DNA-damaging drugs (doxorubicin, dacarbazine, temozolamide) or antimitotic drugs (paclitaxel and docetaxel) in a variety of cancer cells expressing wild-type or mutated p53. Mechanistic investigations revealed that p28 induced a post-translational increase in the expression of wild-type or mutant p53 and p21, resulting in cell-cycle inhibition at the G2-M phase. The enhanced activity of these anticancer agents in combination with p28 was facilitated through the p53/p21/CDK2 pathway. Taken together, these results highlight a new approach to maximize the efficacy of chemotherapeutic agents while reducing dose-related toxicity. Cancer Res; 76(8); 2354-65. ©2016 AACR. PMID:26921335

  7. Fluorescence detection of telomerase activity in cancer cell extracts based on autonomous exonuclease III-assisted isothermal cycling signal amplification.

    PubMed

    Ding, Caifeng; Li, Xiaoqian; Wang, Wei; Chen, Yaoyao

    2016-09-15

    Based on the extension reaction of a telomerase substrate (TS) primer in the presence of the telomerase, strand-displacement process to perform more stable longer duplex chain, and stepwise hydrolysis of mononucleotides from the blunt or the recessed 3'-hydroxyl termini of duplex DNA in the presence of Exonuclease III (Exo III), an amplified fluorescence detection of telomerase activity in the cancer cells was described in this manuscript. A fluorescence probe DNA, a quencher DNA, and a TS primer were mixed to construct a three-chain DNA structure and a two-chain DNA structure because the amount of the TS primer was less than the other two DNA. In the presence of the telomerase, the quencher DNA was replaced from the probe DNA and the telomerase activity could be determined with the fluorescence enhancement. The telomerase activity in HeLa extracts equivalent to 6-2000 cells was detected by this method. Moreover, the strategy was further proved by using telomerase extracted from Romas cells. With the multiple rounds of isothermal strand displacement and the hydrolysis process, constituted consecutive of signal amplification for the novel detection paradigm that allowed measuring of telomerase activity in crude cancer cell extracts confirmed the reliability and practicality of the protocol, which reveal this platform holds great promise in the biochemical assay for the telomerase activity in early diagnosis for cancers. PMID:27108253

  8. Calcium efflux activity of plasma membrane Ca2+ ATPase-4 (PMCA4) mediates cell cycle progression in vascular smooth muscle cells.

    PubMed

    Afroze, Talat; Yang, Ge; Khoshbin, Amir; Tanwir, Mansoor; Tabish, Taha; Momen, Abdul; Husain, Mansoor

    2014-03-01

    We explored the role played by plasma membrane calcium ATPase-4 (PMCA4) and its alternative splice variants in the cell cycle of vascular smooth muscle cells (VSMC). A novel variant (PMCA4e) was discovered. Quantitative real-time-PCR-quantified PMCA4 splice variant proportions differed in specific organs. The PMCA4a:4b ratio in uninjured carotid arteries (∼1:1) was significantly reduced by wire denudation injury (to ∼1:3) by modulation of alternative splicing, as confirmed by novel antibodies against PMCA4a/e and PMCA4b. Laser capture microdissection localized this shift to the media and adventitia. Primary carotid VSMC from PMCA4 knock-out (P4KO) mice showed impaired [(3)H]thymidine incorporation and G1 phase arrest as compared with wild type (P4WT). Electroporation of expression constructs encoding PMCA4a, PMCA4b, and a PMCA4b mutant lacking PDZ binding rescued this phenotype of P4KO cells, whereas a mutant with only 10% of normal Ca(2+) efflux activity could not. Microarray of early G1-synchronized VSMC showed 39-fold higher Rgs16 (NFAT (nuclear factor of activated T-cells) target; MAPK inhibitor) and 69-fold higher Decorin (G1 arrest marker) expression in P4KO versus P4WT. Validation by Western blot also revealed decreased levels of Cyclin D1 and NFATc3 in P4KO. Microarrays of P4KO VSMC rescued by PMCA4a or PMCA4b expression showed reversal of perturbed Rgs16, Decorin, and NFATc3 expression levels. However, PMCA4a rescue caused a 44-fold reduction in AP-2β, a known anti-proliferative transcription factor, whereas PMCA4b rescue resulted in a 50-fold reduction in p15 (Cyclin D1/Cdk4 inhibitor). We conclude that Ca(2+) efflux activity of PMCA4 underlies G1 progression in VSMC and that PMCA4a and PMCA4b differentially regulate specific downstream mediators. PMID:24448801

  9. Glutathione transferases P1/P2 regulate the timing of signaling pathway activations and cell cycle progression during mouse liver regeneration.

    PubMed

    Pajaud, J; Ribault, C; Ben Mosbah, I; Rauch, C; Henderson, C; Bellaud, P; Aninat, C; Loyer, P; Morel, F; Corlu, A

    2015-01-01

    Glutathione transferases (GST) are phase II enzymes catalyzing the detoxification of endogenous noxious compounds and xenobiotics. They also regulate phosphorylation activities of MAPKinases in a catalytic-independent manner. Previous studies have demonstrated the regulation of JNK-dependent pathway by GSTP1/2. Considering the crucial role of JNK in the early steps of the hepatocyte cell cycle, we sought to determine whether GSTP1/2 were essential for hepatocyte proliferation following partial hepatectomy (PH). Using a conventional double knockout mouse model for the Gstp1 and Gstp2 genes, we found that the lack of GSTP1/P2 reduced the rate of DNA replication and mitotic index during the first wave of hepatocyte proliferation. The lowered proliferation was associated with the decrease in TNFalpha and IL-6 plasma concentrations, reduced hepatic HGF expression and delayed and/or altered activation of STAT3, JNK and ERK1/2 signaling pathways. In addition, the expression and/or activation of cell cycle regulators such as Cyclin D1, CDK4, E2F1 and MCM7 was postponed demonstrating that the absence of GSTP1/2 delayed the entry into and progression through the G1 phase of the cell cycle and impaired the synchrony of proliferation in hepatocytes following PH. Furthermore, while JNK and its downstream targets c-Jun and ATF2 were activated during the early steps of the liver regeneration in wild-type animals, the constitutively active JNK found in the quiescent liver of Gstp1/2 knockout mice underwent a decrease in its activity after PH. Transient induction of antioxidant enzymes and nitric oxide synthase were also delayed or repressed during the regenerative response. Altogether our results demonstrate that GSTP1/2 are a critical regulators of hepatocyte proliferation in the initial phases of liver regeneration. PMID:25590808

  10. Glutathione transferases P1/P2 regulate the timing of signaling pathway activations and cell cycle progression during mouse liver regeneration

    PubMed Central

    Pajaud, J; Ribault, C; Ben Mosbah, I; Rauch, C; Henderson, C; Bellaud, P; Aninat, C; Loyer, P; Morel, F; Corlu, A

    2015-01-01

    Glutathione transferases (GST) are phase II enzymes catalyzing the detoxification of endogenous noxious compounds and xenobiotics. They also regulate phosphorylation activities of MAPKinases in a catalytic-independent manner. Previous studies have demonstrated the regulation of JNK-dependent pathway by GSTP1/2. Considering the crucial role of JNK in the early steps of the hepatocyte cell cycle, we sought to determine whether GSTP1/2 were essential for hepatocyte proliferation following partial hepatectomy (PH). Using a conventional double knockout mouse model for the Gstp1 and Gstp2 genes, we found that the lack of GSTP1/P2 reduced the rate of DNA replication and mitotic index during the first wave of hepatocyte proliferation. The lowered proliferation was associated with the decrease in TNFalpha and IL-6 plasma concentrations, reduced hepatic HGF expression and delayed and/or altered activation of STAT3, JNK and ERK1/2 signaling pathways. In addition, the expression and/or activation of cell cycle regulators such as Cyclin D1, CDK4, E2F1 and MCM7 was postponed demonstrating that the absence of GSTP1/2 delayed the entry into and progression through the G1 phase of the cell cycle and impaired the synchrony of proliferation in hepatocytes following PH. Furthermore, while JNK and its downstream targets c-Jun and ATF2 were activated during the early steps of the liver regeneration in wild-type animals, the constitutively active JNK found in the quiescent liver of Gstp1/2 knockout mice underwent a decrease in its activity after PH. Transient induction of antioxidant enzymes and nitric oxide synthase were also delayed or repressed during the regenerative response. Altogether our results demonstrate that GSTP1/2 are a critical regulators of hepatocyte proliferation in the initial phases of liver regeneration. PMID:25590808

  11. Pokemon enhances proliferation, cell cycle progression and anti-apoptosis activity of colorectal cancer independently of p14ARF-MDM2-p53 pathway.

    PubMed

    Zhao, Yi; Yao, Yun-hong; Li, Li; An, Wei-fang; Chen, Hong-zen; Sun, Li-ping; Kang, Hai-xian; Wang, Sen; Hu, Xin-rong

    2014-12-01

    Pokemon has been showed to directly suppress p14(ARF) expression and also to overexpress in multiple cancers. However, p14(ARF)-MDM2-p53 pathway is usually aberrant in colorectal cancer (CRC). The aim is to confirm whether Pokemon plays a role in CRC and explore whether Pokemon works through p14(ARF)-MDM2-p53 pathway in CRC. Immunohistochemistry for Pokemon, p14(ARF) and Mtp53 protein was applied to 45 colorectal epitheliums (CREs), 42 colorectal adenomas (CRAs) and 66 CRCs. Pokemon was knocked down with RNAi technique in CRC cell line Lovo to detect mRNA expression of p14(ARF) with qRT-PCR, cell proliferation with CCK8 assay, and cell cycle and apoptosis with flowcytometry analysis. The protein expression rates were significantly higher in CRC (75.8%) than in CRE (22.2 %) or CRA (38.1%) for Pokemon and higher in CRC (53.0%) than in CRE (0) or CRA (4.8%) for Mtp53, but not significantly different in CRC (86.4 %) versus CRE (93.3%) or CRA (90.5 %) for p14(ARF). Higher expression rate of Pokemon was associated with lymph node metastasis and higher Duke's stage. After knockdown of Pokemon in Lovo cells, the mRNA level of p14(ARF) was not significantly changed, the cell proliferation ability was decreased by 20.6%, cell cycle was arrested by 55.7% in G0/G1 phase, and apoptosis rate was increased by 19.0%. Pokemon enhanced the oncogenesis of CRC by promoting proliferation, cell cycle progression and anti-apoptosis activity of CRC cells independently of p14(ARF)-MDM2-p53 pathway. This finding provided a novel idea for understanding and further studying the molecular mechanism of Pokemon on carcinogenesis of CRC. PMID:25367850

  12. Analysis of cell cycle position in mammalian cells.

    PubMed

    Cecchini, Matthew J; Amiri, Mehdi; Dick, Frederick A

    2012-01-01

    The regulation of cell proliferation is central to tissue morphogenesis during the development of multicellular organisms. Furthermore, loss of control of cell proliferation underlies the pathology of diseases like cancer. As such there is great need to be able to investigate cell proliferation and quantitate the proportion of cells in each phase of the cell cycle. It is also of vital importance to indistinguishably identify cells that are replicating their DNA within a larger population. Since a cell's decision to proliferate is made in the G1 phase immediately before initiating DNA synthesis and progressing through the rest of the cell cycle, detection of DNA synthesis at this stage allows for an unambiguous determination of the status of growth regulation in cell culture experiments. DNA content in cells can be readily quantitated by flow cytometry of cells stained with propidium iodide, a fluorescent DNA intercalating dye. Similarly, active DNA synthesis can be quantitated by culturing cells in the presence of radioactive thymidine, harvesting the cells, and measuring the incorporation of radioactivity into an acid insoluble fraction. We have considerable expertise with cell cycle analysis and recommend a different approach. We Investigate cell proliferation using bromodeoxyuridine/fluorodeoxyuridine (abbreviated simply as BrdU) staining that detects the incorporation of these thymine analogs into recently synthesized DNA. Labeling and staining cells with BrdU, combined with total DNA staining by propidium iodide and analysis by flow cytometry offers the most accurate measure of cells in the various stages of the cell cycle. It is our preferred method because it combines the detection of active DNA synthesis, through antibody based staining of BrdU, with total DNA content from propidium iodide. This allows for the clear separation of cells in G1 from early S phase, or late S phase from G2/M. Furthermore, this approach can be utilized to investigate the effects

  13. ZO-2 silencing induces renal hypertrophy through a cell cycle mechanism and the activation of YAP and the mTOR pathway.

    PubMed

    Domínguez-Calderón, Alaide; Ávila-Flores, Antonia; Ponce, Arturo; López-Bayghen, Esther; Calderón-Salinas, José-Víctor; Luis Reyes, José; Chávez-Munguía, Bibiana; Segovia, José; Angulo, Carla; Ramírez, Leticia; Gallego-Gutiérrez, Helios; Alarcón, Lourdes; Martín-Tapia, Dolores; Bautista-García, Pablo; González-Mariscal, Lorenza

    2016-05-15

    Renal compensatory hypertrophy (RCH) restores normal kidney function after disease or loss of kidney tissue and is characterized by an increase in organ size due to cell enlargement and not to cell proliferation. In MDCK renal epithelial cells, silencing of the tight junction protein zona occludens 2 (ZO-2 KD) induces cell hypertrophy by two mechanisms: prolonging the time that cells spend at the G1 phase of the cell cycle due to an increase in cyclin D1 level, and augmenting the rate of protein synthesis. The latter is triggered by the nuclear accumulation and increased transcriptional activity of Yes-associated protein (YAP), the main target of the Hippo pathway, which results in decreased expression of phosphatase and tensin homologue. This in turn increased the level of phosphatidylinositol (3,4,5)-triphosphate, which transactivates the Akt/mammalian target of rapamycin pathway, leading to activation of the kinase S6K1 and increased synthesis of proteins and cell size. In agreement, in a rat model of uninephrectomy, RCH is accompanied by decreased expression of ZO-2 and nuclear expression of YAP. Our results reveal a novel role of ZO-2 as a modulator of cell size. PMID:27009203

  14. ZO-2 silencing induces renal hypertrophy through a cell cycle mechanism and the activation of YAP and the mTOR pathway

    PubMed Central

    Domínguez-Calderón, Alaide; Ávila-Flores, Antonia; Ponce, Arturo; López-Bayghen, Esther; Calderón-Salinas, José-Víctor; Luis Reyes, José; Chávez-Munguía, Bibiana; Segovia, José; Angulo, Carla; Ramírez, Leticia; Gallego-Gutiérrez, Helios; Alarcón, Lourdes; Martín-Tapia, Dolores; Bautista-García, Pablo; González-Mariscal, Lorenza

    2016-01-01

    Renal compensatory hypertrophy (RCH) restores normal kidney function after disease or loss of kidney tissue and is characterized by an increase in organ size due to cell enlargement and not to cell proliferation. In MDCK renal epithelial cells, silencing of the tight junction protein zona occludens 2 (ZO-2 KD) induces cell hypertrophy by two mechanisms: prolonging the time that cells spend at the G1 phase of the cell cycle due to an increase in cyclin D1 level, and augmenting the rate of protein synthesis. The latter is triggered by the nuclear accumulation and increased transcriptional activity of Yes-associated protein (YAP), the main target of the Hippo pathway, which results in decreased expression of phosphatase and tensin homologue. This in turn increased the level of phosphatidylinositol (3,4,5)-triphosphate, which transactivates the Akt/mammalian target of rapamycin pathway, leading to activation of the kinase S6K1 and increased synthesis of proteins and cell size. In agreement, in a rat model of uninephrectomy, RCH is accompanied by decreased expression of ZO-2 and nuclear expression of YAP. Our results reveal a novel role of ZO-2 as a modulator of cell size. PMID:27009203

  15. Cell cycle regulation of the activity and subcellular localization of Plk1, a human protein kinase implicated in mitotic spindle function.

    PubMed

    Golsteyn, R M; Mundt, K E; Fry, A M; Nigg, E A

    1995-06-01

    Correct assembly and function of the mitotic spindle during cell division is essential for the accurate partitioning of the duplicated genome to daughter cells. Protein phosphorylation has long been implicated in controlling spindle function and chromosome segregation, and genetic studies have identified several protein kinases and phosphatases that are likely to regulate these processes. In particular, mutations in the serine/threonine-specific Drosophila kinase polo, and the structurally related kinase Cdc5p of Saccharomyces cerevisae, result in abnormal mitotic and meiotic divisions. Here, we describe a detailed analysis of the cell cycle-dependent activity and subcellular localization of Plk1, a recently identified human protein kinase with extensive sequence similarity to both Drosophila polo and S. cerevisiae Cdc5p. With the aid of recombinant baculoviruses, we have established a reliable in vitro assay for Plk1 kinase activity. We show that the activity of human Plk1 is cell cycle regulated, Plk1 activity being low during interphase but high during mitosis. We further show, by immunofluorescent confocal laser scanning microscopy, that human Plk1 binds to components of the mitotic spindle at all stages of mitosis, but undergoes a striking redistribution as cells progress from metaphase to anaphase. Specifically, Plk1 associates with spindle poles up to metaphase, but relocalizes to the equatorial plane, where spindle microtubules overlap (the midzone), as cells go through anaphase. These results indicate that the association of Plk1 with the spindle is highly dynamic and that Plk1 may function at multiple stages of mitotic progression. Taken together, our data strengthen the notion that human Plk1 may represent a functional homolog of polo and Cdc5p, and they suggest that this kinase plays an important role in the dynamic function of the mitotic spindle during chromosome segregation. PMID:7790358

  16. Cell cycle of globose basal cells in rat olfactory epithelium.

    PubMed

    Huard, J M; Schwob, J E

    1995-05-01

    The olfactory epithelium of adult mammals has the unique property of generating olfactory sensory neurons throughout life. Cells of the basal compartment, which include horizontal and globose basal cells, are responsible for the ongoing process of neurogenesis in this system. We report here that the globose basal cells in olfactory epithelium of rats, as in mice, are the predominant type of proliferating cell, and account for 97.6% of the actively dividing cells in the basal compartment of the normal epithelium. Globose basal cells have not been fully characterized in terms of their proliferative properties, and the dynamic aspects of neurogenesis are not well understood. As a consequence, it is uncertain whether cell kinetic properties are under any regulation that could affect the rate of neurogenesis. To address this gap in our knowledge, we have determined the duration of both the synthesis phase (S-phase) and the full cell cycle of globose basal cells in adult rats. The duration of the S-phase was found to be 9 hr in experiments utilizing sequential injections of either IdU followed by BrdU or 3H-thy followed by BrdU. The duration of the cell cycle was determined by varying the time interval between the injections of 3H-thy and BrdU and tracking the set of cells that exit S shortly after the first injection. With this paradigm, the interval required for these cells to traverse G2, M, G1, and a second S-phase, is equivalent to the duration of one mitotic cycle and equals 17 hr. These observations serve as the foundation to assess whether the cell cycle duration is subject to regulation in response to experimental injury, and whether such regulation is partly responsible for changes in the rate of neurogenesis in such settings. PMID:7647371

  17. Escherichia coli cytolethal distending toxin blocks the HeLa cell cycle at the G2/M transition by preventing cdc2 protein kinase dephosphorylation and activation.

    PubMed Central

    Comayras, C; Tasca, C; Pérès, S Y; Ducommun, B; Oswald, E; De Rycke, J

    1997-01-01

    Cytolethal distending toxins (CDT) constitute an emerging heterogeneous family of bacterial toxins whose common biological property is to inhibit the proliferation of cells in culture by blocking their cycle at G2/M phase. In this study, we investigated the molecular mechanisms underlying the block caused by CDT from Escherichia coli on synchronized HeLa cell cultures. To this end, we studied specifically the behavior of the two subunits of the complex that determines entry into mitosis, i.e., cyclin B1, the regulatory unit, and cdc2 protein kinase, the catalytic unit. We thus demonstrate that CDT causes cell accumulation in G2 and not in M, that it does not slow the progression of cells through S phase, and that it does not affect the normal increase of cyclin B1 from late S to G2. On the other hand, we show that CDT inhibits the kinase activity of cdc2 by preventing its dephosphorylation, an event which, in normal cells, triggers mitosis. This inhibitory activity was demonstrated for the three partially related CDTs so far described for E. coli. Moreover, we provide evidence that cells exposed to CDT during G2 and M phases are blocked only at the subsequent G2 phase. This observation means that the toxin triggers a mechanism of cell arrest that is initiated in S phase and therefore possibly related to the DNA damage checkpoint system. PMID:9393800

  18. 3,39-Diindolylmethane Ameliorates Staphylococcal Enterotoxin B–Induced Acute Lung Injury through Alterations in the Expression of MicroRNA that Target Apoptosis and Cell-Cycle Arrest in Activated T Cells.

    PubMed

    Elliott, David M; Nagarkatti, Mitzi; Nagarkatti, Prakash S

    2016-04-01

    3,39-Diindolylmethane (DIM), a natural indole found in cruciferous vegetables, has significant anti-cancer and anti-inflammatory properties. In this current study, we investigated the effects of DIM on acute lung injury (ALI) induced by exposure to staphylococcal enterotoxin B (SEB). We found that pretreatment of mice with DIM led to attenuation of SEB-induced inflammation in the lungs, vascular leak, and IFN-g secretion. Additionally, DIM could induce cell-cycle arrest and cell death in SEB-activated T cells in a concentration-dependent manner. Interestingly, microRNA (miRNA) microarray analysis uncovered an altered miRNA profile in lung-infiltrating mononuclear cells after DIM treatment of SEB-exposed mice. Moreover, computational analysis of miRNA gene targets and regulation networks indicated that DIM alters miRNA in the cell death and cell-cycle progression pathways. Specifically, DIM treatment significantly downregulated several miRNA and a correlative increase associated gene targets. Furthermore, overexpression and inhibition studies demonstrated that DIM-induced cell death, at least in part, used miR-222. Collectively, these studies demonstrate for the first time that DIM treatment attenuates SEB-induced ALI and may do so through the induction of microRNAs that promote apoptosis and cell-cycle arrest in SEB-activated T cells. PMID:26818958

  19. [Cell cycle, mitosis and therapeutic applications].

    PubMed

    Levy, Antonin; Albiges-Sauvin, Laurence; Massard, Christophe; Soria, Jean-Charles; Deutsch, Eric

    2011-10-01

    Genomic DNA is constantly under stress of endogenous and exogenous DNA damaging agents. Without proper care, the DNA damage causes an alteration of the genomic structure and can lead to cell death or the occurrence of mutations involved in tumorigenesis. During the process of evolution, organisms have acquired a series of response mechanisms and repair of DNA damage, thereby ensuring the maintenance of genome stability and faithful transmission of genetic information. The checkpoints are the major mechanisms by which a cell can respond to DNA damage, either by actively stopping the cell cycle or by induction of apoptosis. Two parallel signalling pathways, ATM and ATR respond to genotoxic stress by activating their downstream target proteins including the two effectors kinases CHK1 and CHK2. Promising preliminary data render these proteins potential targets for therapeutic development against cancer. PMID:21669563

  20. The ceramide-activated protein phosphatase Sit4p controls lifespan, mitochondrial function and cell cycle progression by regulating hexokinase 2 phosphorylation.

    PubMed

    Barbosa, António Daniel; Pereira, Clara; Osório, Hugo; Moradas-Ferreira, Pedro; Costa, Vítor

    2016-06-17

    Sit4p is the catalytic subunit of a ceramide-activated PP2A-like phosphatase that regulates cell cycle, mitochondrial function, oxidative stress resistance and chronological lifespan in yeast. In this study, we show that hexokinase 2 (Hxk2p) is hyperphosphorylated in sit4Δ mutants grown in glucose medium by a Snf1p-independent mechanism and Hxk2p-S15A mutation suppresses phenotypes associated with SIT4 deletion, namely growth arrest at G1 phase, derepression of mitochondrial respiration, H2O2 resistance and lifespan extension. Consistently, the activation of Sit4p in isc1Δ mutants, which has been associated with premature aging, leads to Hxk2p hypophosphorylation, and the expression of Hxk2p-S15E increases the lifespan of isc1Δ cells. The overall results suggest that Hxk2p functions downstream of Sit4p in the control of cell cycle, mitochondrial function, oxidative stress resistance and chronological lifespan. PMID:27163342

  1. Analysis of the Schizosaccharomyces pombe Cell Cycle.

    PubMed

    Hagan, Iain M; Grallert, Agnes; Simanis, Viesturs

    2016-01-01

    Schizosaccharomyces pombe cells are rod shaped, and they grow by tip elongation. Growth ceases during mitosis and cell division; therefore, the length of a septated cell is a direct measure of the timing of mitotic commitment, and the length of a wild-type cell is an indicator of its position in the cell cycle. A large number of documented stage-specific changes can be used as landmarks to characterize cell cycle progression under specific experimental conditions. Conditional mutations can permanently or transiently block the cell cycle at almost any stage. Large, synchronously dividing cell populations, essential for the biochemical analysis of cell cycle events, can be generated by induction synchrony (arrest-release of a cell cycle mutant) or selection synchrony (centrifugal elutriation or lactose-gradient centrifugation). Schizosaccharomyces pombe cell cycle studies routinely combine particular markers, mutants, and synchronization procedures to manipulate the cycle. We describe these techniques and list key landmarks in the fission yeast mitotic cell division cycle. PMID:27587785

  2. Bergamottin isolated from Citrus bergamia exerts in vitro and in vivo antitumor activity in lung adenocarcinoma through the induction of apoptosis, cell cycle arrest, mitochondrial membrane potential loss and inhibition of cell migration and invasion.

    PubMed

    Wu, Hui-Juan; Wu, Hong-Bo; Zhao, Yan-Qiu; Chen, Li-Juan; Zou, Hong-Zhi

    2016-07-01

    The objective of the present study was to investigate the in vitro and in vivo anticancer properties of bergamottin, a natural furanocoumarin, against human non-small cell lung carcinoma (NSCLC) A549 cells. We also studied its effect on cell proliferation, cell cycle arrest, cell invasion, cell migration as well as cell apoptosis. Antiproliferative activity of bergamottin was estimated by the MTT assay. Phase contrast and fluorescence microscopy as well as flow cytometry using Annexin V-FITC assay were used to study induction of apoptosis by bergamottin in these cells. The effects of bergamottin on cell cycle phase distribution as well as on mitochondrial membrane potential were also demonstrated using flow cytometry. In vitro wound healing assay was used to study the effect of bergamottin on cell migration. The effects of bergamottin on tumor progression were also observed using a nude mouse model. The mice were divided into 4 groups and treated with bergamottin injected intraperitoneally. Bergamottin induced dose-dependent as well as time-dependent cytotoxic effects as well as inhibition of colony formation in the A549 cancer cells. Bergamottin also suppressed cancer cell invasion as well as cancer cell migration. Phase contrast microscopy and fluorescence microscopy revealed that bergamottin induced cell shrinkage, chromatin condensation and the cells became rounded and detached from each other. Bergamottin also induced a potent cell cycle arrest at the G2/M phase of the cell cycle. Experiments in mice showed that 25, 50 and 100 mg/kg bergamottin injection reduced the tumor weight from 1.61 g in the phosphate-buffered saline (PBS)-treated group (control) to 1.21, 0.42 and 0.15 g in the bergamottin-treated groups, respectively. The results of the present study revealed that bergamottin was able to inhibit lung cancer cell growth both in a cell model and a xenograft mouse model by inducing apoptosis, mitochondrial membrane potential loss, G2/M cell cycle

  3. Assaying Cell Cycle Status Using Flow Cytometry.

    PubMed

    Kim, Kang Ho; Sederstrom, Joel M

    2015-01-01

    In this unit, two protocols are described for analyzing cell cycle status using flow cytometry. The first is based on the simultaneous analysis of proliferation-specific marker (Ki-67) and cellular DNA content, which discriminate resting/quiescent cell populations (G0 cell) and quantify cell cycle distribution (G1, S, or G2/M), respectively. The second is based on differential staining of DNA and RNA through co-staining of Hoechst 33342 and Pyronin Y, which is also useful to identify G0 cells from G1 cells. Along with these methods for analyzing cell cycle status, two additional methods for cell proliferation assays with recent updates of newly developed fluorophores, which allow multiplex analysis of cell cycle status, cell proliferation, and a gene of interest using flow cytometry, are outlined. PMID:26131851

  4. Lidamycin induces marked G2 cell cycle arrest in human colon carcinoma HT-29 cells through activation of p38 MAPK pathway.

    PubMed

    Liu, Xia; Bian, Chunjing; Ren, Kaihuan; Jin, Haixia; Li, Baowei; Shao, Rong-Guang

    2007-03-01

    Lidamycin (LDM), a member of the enediyne antibiotic family, is presently undergoing phase I clinical trials in P.R. China. In this study, we investigated the mechanisms of LDM-induced cell cycle arrest in order to support its use in clinical cancer therapy. Using human colon carcinoma HT-29 cells, we observed that LDM induced G2 cell cycle arrest in a time- and dose-dependent manner. LDM-induced G2 arrest was associated with increasing phosphorylation of Chk1, Chk2, Cdc25C, Cdc2 and expression of Cdc2 and cyclin B1. In addition, cytoplasmic localization of cyclin B1 was also involved in LDM-induced G2 arrest. Moreover, we found that p38 MAPK pathway contributed to LDM-induced G2 arrest. Inhibition of p38 MAPK by its inhibitor SB203580 not only attenuated LDM-induced G2 arrest but also potentiated LDM-induced apoptosis, which was accompanied by decreasing phosphorylation of Cdc2 and increasing expression of FasL and phosphorylation of JNK. Finally, we demonstrated that cells at G1 phase were more sensitive to LDM. Together, our findings suggest that p38 MAPK signaling pathway is involved in LDM-induced G2 arrest, at least partly, and a combination of LDM with p38 MAPK inhibitor may represent a new strategy for human colon cancer therapy. PMID:17273739

  5. Fission Yeast Cell Cycle Synchronization Methods.

    PubMed

    Tormos-Pérez, Marta; Pérez-Hidalgo, Livia; Moreno, Sergio

    2016-01-01

    Fission yeast cells can be synchronized by cell cycle arrest and release or by size selection. Cell cycle arrest synchronization is based on the block and release of temperature-sensitive cell cycle mutants or treatment with drugs. The most widely used approaches are cdc10-129 for G1; hydroxyurea (HU) for early S-phase; cdc25-22 for G2, and nda3-KM311 for mitosis. Cells can also be synchronized by size selection using centrifugal elutriation or a lactose gradient. Here we describe the methods most commonly used to synchronize fission yeast cells. PMID:26519320

  6. Targeting cell cycle regulators in hematologic malignancies

    PubMed Central

    Aleem, Eiman; Arceci, Robert J.

    2015-01-01

    Hematologic malignancies represent the fourth most frequently diagnosed cancer in economically developed countries. In hematologic malignancies normal hematopoiesis is interrupted by uncontrolled growth of a genetically altered stem or progenitor cell (HSPC) that maintains its ability of self-renewal. Cyclin-dependent kinases (CDKs) not only regulate the mammalian cell cycle, but also influence other vital cellular processes, such as stem cell renewal, differentiation, transcription, epigenetic regulation, apoptosis, and DNA repair. Chromosomal translocations, amplification, overexpression and altered CDK activities have been described in different types of human cancer, which have made them attractive targets for pharmacological inhibition. Mouse models deficient for one or more CDKs have significantly contributed to our current understanding of the physiological functions of CDKs, as well as their roles in human cancer. The present review focuses on selected cell cycle kinases with recent emerging key functions in hematopoiesis and in hematopoietic malignancies, such as CDK6 and its role in MLL-rearranged leukemia and acute lymphocytic leukemia, CDK1 and its regulator WEE-1 in acute myeloid leukemia (AML), and cyclin C/CDK8/CDK19 complexes in T-cell acute lymphocytic leukemia. The knowledge gained from gene knockout experiments in mice of these kinases is also summarized. An overview of compounds targeting these kinases, which are currently in clinical development in various solid tumors and hematopoietic malignances, is presented. These include the CDK4/CDK6 inhibitors (palbociclib, LEE011, LY2835219), pan-CDK inhibitors that target CDK1 (dinaciclib, flavopiridol, AT7519, TG02, P276-00, terampeprocol and RGB 286638) as well as the WEE-1 kinase inhibitor, MK-1775. The advantage of combination therapy of cell cycle inhibitors with conventional chemotherapeutic agents used in the treatment of AML, such as cytarabine, is discussed. PMID:25914884

  7. Association of Transcription Factor IIA with TATA Binding Protein Is Required for Transcriptional Activation of a Subset of Promoters and Cell Cycle Progression in Saccharomyces cerevisiae

    PubMed Central

    Ozer, Josef; Lezina, Larissa E.; Ewing, Joshua; Audi, Salma; Lieberman, Paul M.

    1998-01-01

    The general transcription factor IIA (TFIIA) interacts with the TATA binding protein (TBP) and promoter DNA to mediate transcription activation in vitro. To determine if this interaction is generally required for activation of all class II genes in vivo, we have constructed substitution mutations in yeast TFIIA which compromise its ability to bind TBP. Substitution mutations in the small subunit of TFIIA (Toa2) at residue Y69 or W76 significantly impaired the ability of TFIIA to stimulate TBP-promoter binding in vitro. Gene replacement of wild-type TOA2 with a W76E or Y69A/W76A mutant was lethal in Saccharomyces cerevisiae, while the Y69F/W76F mutant exhibited extremely slow growth at 30°C. Both the Y69A and W76A mutants were conditionally lethal at higher temperatures. Light microscopy indicated that viable toa2 mutant strains accumulate as equal-size dumbbells and multibudded clumps. Transcription of the cell cycle-regulatory genes CLB1, CLB2, CLN1, and CTS1 was significantly reduced in the toa2 mutant strains, while the noncycling genes PMA1 and ENO2 were only modestly affected, suggesting that these toa2 mutant alleles disrupt cell cycle progression. The differential effect of these toa2 mutants on gene transcription was examined for a number of other genes. toa2 mutant strains supported high levels of CUP1, PHO5, TRP3, and GAL1 gene activation, but the constitutive expression of DED1 was significantly reduced. Activator-induced start site expression for HIS3, GAL80, URA1, and URA3 promoters was defective in toa2 mutant strains, suggesting that the TFIIA-TBP complex is important for promoters which require an activator-dependent start site selection from constitutive to regulated expression. We present evidence to indicate that transcription defects in toa2 mutants can be both activator and promoter dependent. These results suggest that the association of TFIIA with TBP regulates activator-induced start site selection and cell cycle progression in S

  8. Activation of nuclear PTEN by inhibition of Notch signaling induces G2/M cell cycle arrest in gastric cancer.

    PubMed

    Kim, S-J; Lee, H-W; Baek, J-H; Cho, Y-H; Kang, H G; Jeong, J S; Song, J; Park, H-S; Chun, K-H

    2016-01-14

    Mutation in PTEN has not yet been detected, but its function as a tumor suppressor is inactivated in many cancers. In this study we determined that, activated Notch signaling disables PTEN by phosphorylation and thereby contributes to gastric tumorigenesis. Notch inhibition by small interfering RNA or γ-secretase inhibitor (GSI) induced mitotic arrest and apoptosis in gastric cancer cells. Notch inhibition induced dephosphorylation in the C-terminal domain of PTEN, which led to PTEN nuclear localization. Overexpression of activated Notch1-induced phosphorylation of PTEN and reversed GSI-induced mitotic arrest. Dephosphorylated nuclear PTEN caused prometaphase arrest by interaction with the cyclin B1-CDK1 complex, resulting in their accumulation in the nucleus and subsequent apoptosis. We found a correlation between high expression levels of Notch1 and low survival rates and, similarly, between reduced nuclear PTEN expression and increasing the TNM classification of malignant tumours stages in malignant tissues from gastric cancer patients. The growth of Notch1-depleted gastric tumors was significantly retarded in xenografted mice, and in addition, PTEN deletion restored growth similar to control tumors. We also demonstrated that combination treatment with GSI and chemotherapeutic agents significantly reduced the orthotopically transplanted gastric tumors in mice without noticeable toxicity. Overall, our findings suggest that inhibition of Notch signaling can be employed as a PTEN activator, making it a potential target for gastric cancer therapy. PMID:25823029

  9. Basal p21 controls population heterogeneity in cycling and quiescent cell cycle states

    PubMed Central

    Overton, K. Wesley; Spencer, Sabrina L.; Noderer, William L.; Meyer, Tobias; Wang, Clifford L.

    2014-01-01

    Phenotypic heterogeneity within a population of genetically identical cells is emerging as a common theme in multiple biological systems, including human cell biology and cancer. Using live-cell imaging, flow cytometry, and kinetic modeling, we showed that two states—quiescence and cell cycling—can coexist within an isogenic population of human cells and resulted from low basal expression levels of p21, a Cyclin-dependent kinase (CDK) inhibitor (CKI). We attribute the p21-dependent heterogeneity in cell cycle activity to double-negative feedback regulation involving CDK2, p21, and E3 ubiquitin ligases. In support of this mechanism, analysis of cells at a point before cell cycle entry (i.e., before the G1/S transition) revealed a p21–CDK2 axis that determines quiescent and cycling cell states. Our findings suggest a mechanistic role for p21 in generating heterogeneity in both normal tissues and tumors. PMID:25267623

  10. Stroma-Derived Connective Tissue Growth Factor Maintains Cell Cycle Progression and Repopulation Activity of Hematopoietic Stem Cells In Vitro

    PubMed Central

    Istvánffy, Rouzanna; Vilne, Baiba; Schreck, Christina; Ruf, Franziska; Pagel, Charlotta; Grziwok, Sandra; Henkel, Lynette; Prazeres da Costa, Olivia; Berndt, Johannes; Stümpflen, Volker; Götze, Katharina S.; Schiemann, Matthias; Peschel, Christian; Mewes, Hans-Werner; Oostendorp, Robert A.J.

    2015-01-01

    Summary Hematopoietic stem cells (HSCs) are preserved in co-cultures with UG26-1B6 stromal cells or their conditioned medium. We performed a genome-wide study of gene expression changes of UG26-1B6 stromal cells in contact with Lineage− SCA-1+ KIT+ (LSK) cells. This analysis identified connective tissue growth factor (CTGF) to be upregulated in response to LSK cells. We found that co-culture of HSCs on CTGF knockdown stroma (shCtgf) shows impaired engraftment and long-term quality. Further experiments demonstrated that CD34− CD48− CD150+ LSK (CD34− SLAM) cell numbers from shCtgf co-cultures increase in G0 and senescence and show delayed time to first cell division. To understand this observation, a CTGF signaling network model was assembled, which was experimentally validated. In co-culture experiments of CD34− SLAM cells with shCtgf stromal cells, we found that SMAD2/3-dependent signaling was activated, with increasing p27Kip1 expression and downregulating cyclin D1. Our data support the view that LSK cells modulate gene expression in the niche to maintain repopulating HSC activity. PMID:26527384

  11. Cell cycle: proteomics gives it a spin.

    PubMed

    Archambault, Vincent

    2005-08-01

    The eukaryotic cell division cycle has been studied at the molecular level for over 30 years, most fruitfully in model organisms. In the past 5 years, developments in mass spectrometry-based proteomics have been applied to the study of protein interactions and post-translational modifications involving key cell cycle regulators such as cyclin-dependent kinases and the anaphase-promoting complex, as well as effectors such as centrosomes, the kinetochore and DNA replication forks. In addition, innovations in chemical biology, functional proteomics and bioinformatics have been employed to study the cell cycle at the proteome level. This review surveys the contributions of proteomics to cell cycle research. The near future should see the application of more quantitative proteomic approaches to probe the dynamic aspects of the molecular system that underlie the cell cycle in model organisms and in human cells. PMID:16097893

  12. Cell cycle control and seed development

    PubMed Central

    Dante, Ricardo A.; Larkins, Brian A.; Sabelli, Paolo A.

    2014-01-01

    Seed development is a complex process that requires coordinated integration of many genetic, metabolic, and physiological pathways and environmental cues. Different cell cycle types, such as asymmetric cell division, acytokinetic mitosis, mitotic cell division, and endoreduplication, frequently occur in sequential yet overlapping manner during the development of the embryo and the endosperm, seed structures that are both products of double fertilization. Asymmetric cell divisions in the embryo generate polarized daughter cells with different cell fates. While nuclear and cell division cycles play a key role in determining final seed cell numbers, endoreduplication is often associated with processes such as cell enlargement and accumulation of storage metabolites that underlie cell differentiation and growth of the different seed compartments. This review focuses on recent advances in our understanding of different cell cycle mechanisms operating during seed development and their impact on the growth, development, and function of seed tissues. Particularly, the roles of core cell cycle regulators, such as cyclin-dependent-kinases and their inhibitors, the Retinoblastoma-Related/E2F pathway and the proteasome-ubiquitin system, are discussed in the contexts of different cell cycle types that characterize seed development. The contributions of nuclear and cellular proliferative cycles and endoreduplication to cereal endosperm development are also discussed. PMID:25295050

  13. Anticancer activity of taraxerol acetate in human glioblastoma cells and a mouse xenograft model via induction of autophagy and apoptotic cell death, cell cycle arrest and inhibition of cell migration

    PubMed Central

    HONG, JING-FANG; SONG, YING-FANG; LIU, ZHENG; ZHENG, ZHAO-CONG; CHEN, HONG-JIE; WANG, SHOU-SEN

    2016-01-01

    The aim of the present study was to investigate the in vitro and in vivo anticancer and apoptotic effects of taraxerol acetate in U87 human glioblastoma cells. The effects on cell cycle phase distribution, cell cycle-associated proteins, autophagy, DNA fragmentation and cell migration were assessed. Cell viability was determined using the MTT assay, and phase contrast and fluorescence microscopy was utilized to determine the viability and apoptotic morphological features of the U87 cells. Flow cytometry using propidium iodide and Annexin V-fluorescein isothiocyanate demonstrated the effect of taraxerol acetate on the cell cycle phase distribution and apoptosis induction. Western blot analysis was performed to investigate the effect of the taraxerol acetate on cell cycle-associated proteins and autophagy-linked LC3B-II proteins. The results demonstrated that taraxerol acetate induced dose- and time-dependent cytotoxic effects in the U87 cells. Apoptotic induction following taraxerol acetate treatment was observed and the percentage of apoptotic cells increased from 7.3% in the control cells, to 16.1, 44.1 and 76.7% in the 10, 50 and 150 µM taraxerol acetate-treated cells, respectively. Furthermore, taraxerol acetate treatment led to sub-G1 cell cycle arrest with a corresponding decrease in the number of S-phase cells. DNA fragments were observed as a result of the gel electrophoresis experiment following taraxerol acetate treatment. To investigate the inhibitory effects of taraxerol acetate on the migration of U87 cell, a wound healing assay was conducted. The number of cells that migrated to the scratched area decreased significantly following treatment with taraxerol acetate. In addition, taraxerol acetate inhibited tumor growth in a mouse xenograft model. Administration of 0.25 and 0.75 µg/g taraxerol acetate reduced the tumor weight from 1.2 g in the phosphate-buffered saline (PBS)-treated group (control) to 0.81 and 0.42 g, respectively. Similarly, 0.25 and 0

  14. Cytotoxicity of diacetoxyscirpenol is associated with apoptosis by activation of caspase-8 and interruption of cell cycle progression by down-regulation of cdk4 and cyclin B1 in human Jurkat T cells

    SciTech Connect

    Jun, Do Youn; Kim, Jun Seok; Park, Hae Sun; Song, Woo Sun; Bae, Young Seuk; Kim, Young Ho . E-mail: ykim@knu.ac.kr

    2007-07-15

    To understand the mechanism underlying T-cell toxicity of diacetoxyscirpenol (DAS) from Fusarium sambucinum, its apoptogenic as well as growth retardation activity was investigated in human Jurkat T cells. Exposure to DAS (0.01-0.15 {mu}M) caused apoptotic DNA fragmentation along with caspase-8 activation, Bid cleavage, mitochondrial cytochrome c release, activation of caspase-9 and caspase-3, and PARP degradation, without any alteration in the levels of Fas or FasL. Under these conditions, necrosis was not accompanied. The cytotoxicity of DAS was not blocked by the anti-Fas neutralizing antibody ZB-4. Although the DAS-induced apoptotic events were completely prevented by overexpression of Bcl-xL, the cells overexpressing Bcl-xL were unable to divide in the presence of DAS, resulting from the failure of cell cycle progression possibly due to down-regulation in the protein levels of cdk4 and cyclin B1. The DAS-mediated apoptosis and activation of caspase-8, -9, and -3 were abrogated by either pan-caspase inhibitor (z-VAD-fmk) or caspase-8 inhibitor (z-IETD-fmk). While the DAS-mediated apoptosis and activation of caspase-9 and caspase-3 were slightly suppressed by the mitochondrial permeability transition pore inhibitor (CsA), both caspase-8 activation and Bid cleavage were not affected by CsA. The activated normal peripheral T cells possessed a similar susceptibility to the cytotoxicity of DAS. These results demonstrate that the T-cell toxicity of DAS is attributable to not only apoptosis initiated by caspase-8 activation and subsequent mitochondrion-dependent or -independent activation of caspase cascades, which can be regulated by Bcl-xL, but also interruption of cell cycle progression caused by down-regulation of cdk4 and cyclin B1 proteins.

  15. Interleukin-24 mediates apoptosis in human B-cells through early activation of cell cycle arrest followed by late induction of the mitochondrial apoptosis pathway.

    PubMed

    Hadife, Nader; Nemos, Christophe; Frippiat, Jean-Pol; Hamadé, Tala; Perrot, Aurore; Dalloul, Ali

    2013-03-01

    Interleukin (IL)-24 has death-promoting effects on various proliferating cells including B-cells from chronic lymphocytic leukemia (CLL) and germinal center B-cells, but its molecular mechanisms are poorly understood. Using a B-cell differentiation model and mRNA profiling, we found that recombinant (r)IL-24 stimulated genes of the mitochondrial apoptotic pathway (Bax, Bid, Casp8, COX6C, COX7B) after 36 h, whereas the transcription of genes involved in DNA replication and metabolism was inhibited within 6 h. Unexpectedly, insulin-like growth factor 1 (IGF1), a hormone known to promote cell growth, was stimulated by IL-24. Activated B-cells express receptor for IGF1, to which they become sensitized and undergo apoptosis, a mechanism similar in this respect to IL-24-induced cell death. Furthermore, inhibition of the IGF1 pathway reversed the effects of IL-24. IL-24-mediated apoptosis was also antagonized by pifithrin-alpha, an inhibitor of p53 transactivation. Altogether, these results disclose sequential molecular signals generated by IL-24 in activated B-cells. PMID:22860893

  16. Microgravity induces pelvic bone loss through osteoclastic activity, osteocytic osteolysis, and osteoblastic cell cycle inhibition by CDKN1a/p21.

    PubMed

    Blaber, Elizabeth A; Dvorochkin, Natalya; Lee, Chialing; Alwood, Joshua S; Yousuf, Rukhsana; Pianetta, Piero; Globus, Ruth K; Burns, Brendan P; Almeida, Eduardo A C

    2013-01-01

    Bone is a dynamically remodeled tissue that requires gravity-mediated mechanical stimulation for maintenance of mineral content and structure. Homeostasis in bone occurs through a balance in the activities and signaling of osteoclasts, osteoblasts, and osteocytes, as well as proliferation and differentiation of their stem cell progenitors. Microgravity and unloading are known to cause osteoclast-mediated bone resorption; however, we hypothesize that osteocytic osteolysis, and cell cycle arrest during osteogenesis may also contribute to bone loss in space. To test this possibility, we exposed 16-week-old female C57BL/6J mice (n = 8) to microgravity for 15-days on the STS-131 space shuttle mission. Analysis of the pelvis by µCT shows decreases in bone volume fraction (BV/TV) of 6.29%, and bone thickness of 11.91%. TRAP-positive osteoclast-covered trabecular bone surfaces also increased in microgravity by 170% (p = 0.004), indicating osteoclastic bone degeneration. High-resolution X-ray nanoCT studies revealed signs of lacunar osteolysis, including increases in cross-sectional area (+17%, p = 0.022), perimeter (+14%, p = 0.008), and canalicular diameter (+6%, p = 0.037). Expression of matrix metalloproteinases (MMP) 1, 3, and 10 in bone, as measured by RT-qPCR, was also up-regulated in microgravity (+12.94, +2.98 and +16.85 fold respectively, p<0.01), with MMP10 localized to osteocytes, and consistent with induction of osteocytic osteolysis. Furthermore, expression of CDKN1a/p21 in bone increased 3.31 fold (p<0.01), and was localized to osteoblasts, possibly inhibiting the cell cycle during tissue regeneration as well as conferring apoptosis resistance to these cells. Finally the apoptosis inducer Trp53 was down-regulated by -1.54 fold (p<0.01), possibly associated with the quiescent survival-promoting function of CDKN1a/p21. In conclusion, our findings identify the pelvic and femoral region of the mouse skeleton as an active site of rapid bone

  17. Microgravity Induces Pelvic Bone Loss through Osteoclastic Activity, Osteocytic Osteolysis, and Osteoblastic Cell Cycle Inhibition by CDKN1a/p21

    PubMed Central

    Blaber, Elizabeth A.; Dvorochkin, Natalya; Lee, Chialing; Alwood, Joshua S.; Yousuf, Rukhsana; Pianetta, Piero; Globus, Ruth K.; Burns, Brendan P.; Almeida, Eduardo A. C.

    2013-01-01

    Bone is a dynamically remodeled tissue that requires gravity-mediated mechanical stimulation for maintenance of mineral content and structure. Homeostasis in bone occurs through a balance in the activities and signaling of osteoclasts, osteoblasts, and osteocytes, as well as proliferation and differentiation of their stem cell progenitors. Microgravity and unloading are known to cause osteoclast-mediated bone resorption; however, we hypothesize that osteocytic osteolysis, and cell cycle arrest during osteogenesis may also contribute to bone loss in space. To test this possibility, we exposed 16-week-old female C57BL/6J mice (n = 8) to microgravity for 15-days on the STS-131 space shuttle mission. Analysis of the pelvis by µCT shows decreases in bone volume fraction (BV/TV) of 6.29%, and bone thickness of 11.91%. TRAP-positive osteoclast-covered trabecular bone surfaces also increased in microgravity by 170% (p = 0.004), indicating osteoclastic bone degeneration. High-resolution X-ray nanoCT studies revealed signs of lacunar osteolysis, including increases in cross-sectional area (+17%, p = 0.022), perimeter (+14%, p = 0.008), and canalicular diameter (+6%, p = 0.037). Expression of matrix metalloproteinases (MMP) 1, 3, and 10 in bone, as measured by RT-qPCR, was also up-regulated in microgravity (+12.94, +2.98 and +16.85 fold respectively, p<0.01), with MMP10 localized to osteocytes, and consistent with induction of osteocytic osteolysis. Furthermore, expression of CDKN1a/p21 in bone increased 3.31 fold (p<0.01), and was localized to osteoblasts, possibly inhibiting the cell cycle during tissue regeneration as well as conferring apoptosis resistance to these cells. Finally the apoptosis inducer Trp53 was down-regulated by −1.54 fold (p<0.01), possibly associated with the quiescent survival-promoting function of CDKN1a/p21. In conclusion, our findings identify the pelvic and femoral region of the mouse skeleton as an active site of rapid bone

  18. Insulin-like growth factor-I extends in vitro replicative life span of skeletal muscle satellite cells by enhancing G1/S cell cycle progression via the activation of phosphatidylinositol 3'-kinase/Akt signaling pathway

    NASA Technical Reports Server (NTRS)

    Chakravarthy, M. V.; Abraha, T. W.; Schwartz, R. J.; Fiorotto, M. L.; Booth, F. W.

    2000-01-01

    Interest is growing in methods to extend replicative life span of non-immortalized stem cells. Using the insulin-like growth factor I (IGF-I) transgenic mouse in which the IGF-I transgene is expressed during skeletal muscle development and maturation prior to isolation and during culture of satellite cells (the myogenic stem cells of mature skeletal muscle fibers) as a model system, we elucidated the underlying molecular mechanisms of IGF-I-mediated enhancement of proliferative potential of these cells. Satellite cells from IGF-I transgenic muscles achieved at least five additional population doublings above the maximum that was attained by wild type satellite cells. This IGF-I-induced increase in proliferative potential was mediated via activation of the phosphatidylinositol 3'-kinase/Akt pathway, independent of mitogen-activated protein kinase activity, facilitating G(1)/S cell cycle progression via a down-regulation of p27(Kip1). Adenovirally mediated ectopic overexpression of p27(Kip1) in exponentially growing IGF-I transgenic satellite cells reversed the increase in cyclin E-cdk2 kinase activity, pRb phosphorylation, and cyclin A protein abundance, thereby implicating an important role for p27(Kip1) in promoting satellite cell senescence. These observations provide a more complete dissection of molecular events by which increased local expression of a growth factor in mature skeletal muscle fibers extends replicative life span of primary stem cells than previously known.

  19. Protein tyrosine nitration in the cell cycle

    SciTech Connect

    Jia, Min; Mateoiu, Claudia; Souchelnytskyi, Serhiy

    2011-09-23

    Highlights: {yields} Enrichment of 3-nitrotyrosine containing proteins from cells synchronized in different phases of the cell cycle. {yields} Identification of 76 tyrosine nitrated proteins that change expression during the cell cycle. {yields} Nineteen identified proteins were previously described as regulators of cell proliferation. -- Abstract: Nitration of tyrosine residues in proteins is associated with cell response to oxidative/nitrosative stress. Tyrosine nitration is relatively low abundant post-translational modification that may affect protein functions. Little is known about the extent of protein tyrosine nitration in cells during progression through the cell cycle. Here we report identification of proteins enriched for tyrosine nitration in cells synchronized in G0/G1, S or G2/M phases of the cell cycle. We identified 27 proteins in cells synchronized in G0/G1 phase, 37 proteins in S phase synchronized cells, and 12 proteins related to G2/M phase. Nineteen of the identified proteins were previously described as regulators of cell proliferation. Thus, our data indicate which tyrosine nitrated proteins may affect regulation of the cell cycle.

  20. High-Cycle-Life Lithium Cell

    NASA Technical Reports Server (NTRS)

    Yen, S. P. S.; Carter, B.; Shen, D.; Somoano, R.

    1985-01-01

    Lithium-anode electrochemical cell offers increased number of charge/ discharge cycles. Cell uses components selected for compatibility with electrolyte solvent: These materials are wettable and chemically stable. Low vapor pressure and high electrochemical stability of solvent improve cell packaging, handling, and safety. Cell operates at modest temperatures - less than 100 degrees C - and is well suited to automotive, communications, and other applications.

  1. Anticancer activity of taraxerol acetate in human glioblastoma cells and a mouse xenograft model via induction of autophagy and apoptotic cell death, cell cycle arrest and inhibition of cell migration.

    PubMed

    Hong, Jing-Fang; Song, Ying-Fang; Liu, Zheng; Zheng, Zhao-Cong; Chen, Hong-Jie; Wang, Shou-Sen

    2016-06-01

    The aim of the present study was to investigate the in vitro and in vivo anticancer and apoptotic effects of taraxerol acetate in U87 human glioblastoma cells. The effects on cell cycle phase distribution, cell cycle-associated proteins, autophagy, DNA fragmentation and cell migration were assessed. Cell viability was determined using the MTT assay, and phase contrast and fluorescence microscopy was utilized to determine the viability and apoptotic morphological features of the U87 cells. Flow cytometry using propidium iodide and Annexin V-fluorescein isothiocyanate demonstrated the effect of taraxerol acetate on the cell cycle phase distribution and apoptosis induction. Western blot analysis was performed to investigate the effect of the taraxerol acetate on cell cycle‑associated proteins and autophagy‑linked LC3B‑II proteins. The results demonstrated that taraxerol acetate induced dose‑ and time‑dependent cytotoxic effects in the U87 cells. Apoptotic induction following taraxerol acetate treatment was observed and the percentage of apoptotic cells increased from 7.3% in the control cells, to 16.1, 44.1 and 76.7% in the 10, 50 and 150 µM taraxerol acetate‑treated cells, respectively. Furthermore, taraxerol acetate treatment led to sub‑G1 cell cycle arrest with a corresponding decrease in the number of S‑phase cells. DNA fragments were observed as a result of the gel electrophoresis experiment following taraxerol acetate treatment. To investigate the inhibitory effects of taraxerol acetate on the migration of U87 cell, a wound healing assay was conducted. The number of cells that migrated to the scratched area decreased significantly following treatment with taraxerol acetate. In addition, taraxerol acetate inhibited tumor growth in a mouse xenograft model. Administration of 0.25 and 0.75 µg/g taraxerol acetate reduced the tumor weight from 1.2 g in the phosphate‑buffered saline (PBS)‑treated group (control) to 0.81 and 0.42

  2. Cell cycle control, checkpoint mechanisms, and genotoxic stress.

    PubMed Central

    Shackelford, R E; Kaufmann, W K; Paules, R S

    1999-01-01

    The ability of cells to maintain genomic integrity is vital for cell survival and proliferation. Lack of fidelity in DNA replication and maintenance can result in deleterious mutations leading to cell death or, in multicellular organisms, cancer. The purpose of this review is to discuss the known signal transduction pathways that regulate cell cycle progression and the mechanisms cells employ to insure DNA stability in the face of genotoxic stress. In particular, we focus on mammalian cell cycle checkpoint functions, their role in maintaining DNA stability during the cell cycle following exposure to genotoxic agents, and the gene products that act in checkpoint function signal transduction cascades. Key transitions in the cell cycle are regulated by the activities of various protein kinase complexes composed of cyclin and cyclin-dependent kinase (Cdk) molecules. Surveillance control mechanisms that check to ensure proper completion of early events and cellular integrity before initiation of subsequent events in cell cycle progression are referred to as cell cycle checkpoints and can generate a transient delay that provides the cell more time to repair damage before progressing to the next phase of the cycle. A variety of cellular responses are elicited that function in checkpoint signaling to inhibit cyclin/Cdk activities. These responses include the p53-dependent and p53-independent induction of Cdk inhibitors and the p53-independent inhibitory phosphorylation of Cdk molecules themselves. Eliciting proper G1, S, and G2 checkpoint responses to double-strand DNA breaks requires the function of the Ataxia telangiectasia mutated gene product. Several human heritable cancer-prone syndromes known to alter DNA stability have been found to have defects in checkpoint surveillance pathways. Exposures to several common sources of genotoxic stress, including oxidative stress, ionizing radiation, UV radiation, and the genotoxic compound benzo[a]pyrene, elicit cell cycle

  3. Menstrual cycle and reproductive aging alters immune reactivity, NGF expression, antioxidant enzyme activities, and intracellular signaling pathways in the peripheral blood mononuclear cells of healthy women.

    PubMed

    Priyanka, Hannah P; Sharma, Utsav; Gopinath, Srinivasan; Sharma, Varun; Hima, Lalgi; ThyagaRajan, Srinivasan

    2013-08-01

    Reproductive senescence in women is a process that begins with regular menstrual cycles and culminates in menopause followed by gradual development of diseases such as autoimmune diseases, osteoporosis, neurodegenerative diseases, and hormone-dependent cancers. The age-associated impairment in the functions of neuroendocrine system and immune system results in menopause which contributes to subsequent development of diseases and cancer. The aim of this study is to characterize the alterations in immune responses, compensatory factors such as nerve growth factor (NGF) and antioxidant enzyme activities, and the molecular mechanisms of actions in the peripheral blood mononuclear cells (PBMCs) of young (follicular and luteal phases), middle-aged, and old healthy women. Peripheral blood mononuclear cells were isolated from young women in follicular and luteal phases of the menstrual cycle (n=20; 22.6±2.9 yrs), middle-aged women (n=19; 47.1±3.8 yrs; perimenopausal) and old (n=16; 63.2±4.7 yrs; post-menopausal) women and analyzed for Concanavalin (Con A)-induced proliferation of lymphocytes and cytokine (IL-2 and IFN-γ) production, expression of NGF, p-NF-κB, p-ERK, p-CREB, and p-Akt, antioxidant enzymes [superoxide dismutase (SOD), catalase, and glutathione peroxidase (GPx), glutathione-S-transferase (GST)], extent of lipid peroxidation, and nitric oxide (NO) production. Serum gonadal hormones (17β-estradiol and progesterone) were also measured. A characteristic age- and menstrual cycle-related change was observed in the serum gonadal hormone secretion (estrogen and progesterone), T lymphocyte proliferation and IFN-γ production. Salient features include the age-related decline observed in target-derived growth factors (lymphocyte NGF expression), signaling molecules (p-ERK/ERK and p-CREB/CREB ratios) and compensatory factors such as the activities of plasma and PBMC antioxidant enzymes (SOD and catalase) and NO production. Further, an age-associated increase in p

  4. Different cell cycle modulation by celecoxib at different concentrations.

    PubMed

    Kim, Young-Mee; Pyo, Hongryull

    2013-03-01

    Abstract Different cyclooxygenase (COX)-2 inhibitors were known to cause different cell cycle changes. We investigated whether this different effect on cell cycle change was due to concentration-dependent effect. We investigated the effects of celecoxib, a COX-2 selective inhibitor, on cell cycle regulation in irradiated cancer cells that express high or low levels of COX-2. Four stably COX-2 knocked-down or overexpressed cell lines were treated with various concentrations of celecoxib with or without radiation. Celecoxib differentially modulated the cell cycle according to the concentrations applied. G1 arrest was induced at lower concentrations, whereas G2/M arrest was induced at higher concentrations in each cell line tested. Radiation-induced G2/M arrest was enhanced at lower concentrations but reduced at higher concentrations. The cutoff values to divide lower and higher concentrations were cell-type specific. Celecoxib treatment activated Cdc25C and inhibited p21 expression in both unirradiated and irradiated cells, regardless of COX-2 expression. Apoptosis was induced in irradiated cells 48 hours after treatment with celecoxib dependent of COX-2. These results imply that celecoxib deactivates the G2 checkpoint via both Cdc25C- and p21-dependent pathways in irradiated cells, which subsequently die by secondary apoptosis. Cell cycle modulating effects in irradiated cells resulting from treatment with celecoxib may have clinical importance with regard to the potential application of celecoxib in cancer patients undergoing radiotherapy. PMID:23268707

  5. Arginine starvation in colorectal carcinoma cells: Sensing, impact on translation control and cell cycle distribution.

    PubMed

    Vynnytska-Myronovska, Bozhena O; Kurlishchuk, Yuliya; Chen, Oleh; Bobak, Yaroslav; Dittfeld, Claudia; Hüther, Melanie; Kunz-Schughart, Leoni A; Stasyk, Oleh V

    2016-02-01

    Tumor cells rely on a continued exogenous nutrient supply in order to maintain a high proliferative activity. Although a strong dependence of some tumor types on exogenous arginine sources has been reported, the mechanisms of arginine sensing by tumor cells and the impact of changes in arginine availability on translation and cell cycle regulation are not fully understood. The results presented herein state that human colorectal carcinoma cells rapidly exhaust the internal arginine sources in the absence of exogenous arginine and repress global translation by activation of the GCN2-mediated pathway and inhibition of mTOR signaling. Tumor suppressor protein p53 activation and G1/G0 cell cycle arrest support cell survival upon prolonged arginine starvation. Cells with the mutant or deleted TP53 fail to stop cell cycle progression at defined cell cycle checkpoints which appears to be associated with reduced recovery after durable metabolic stress triggered by arginine withdrawal. PMID:26751966

  6. Comparison of the activity of three different HSP70 inhibitors on apoptosis, cell cycle arrest, autophagy inhibition, and HSP90 inhibition

    PubMed Central

    Budina-Kolomets, Anna; Balaburski, Gregor M; Bondar, Anastasia; Beeharry, Neil; Yen, Tim; Murphy, Maureen E

    2014-01-01

    The chaperone HSP70 promotes the survival of cells exposed to many different types of stresses, and is also potently anti-apoptotic. The major stress-induced form of this protein, HSP70–1, is overexpressed in a number of human cancers, yet is negligibly expressed in normal cells. Silencing of the gene encoding HSP70–1 (HSPA1A) is cytotoxic to transformed but not normal cells. Therefore, HSP70 is considered to be a promising cancer drug target, and there has been active interest in the identification and characterization of HSP70 inhibitors for cancer therapy. Because HSP70 behaves in a relatively non-specific manner in the control of protein folding, to date there are no reliably-identified “clients” of this protein, nor is there consensus as to what the phenotypic effects of HSP70 inhibitors are on a cancer cell. Here for the first time we compare three recently-identified HSP70 inhibitors, PES-Cl, MKT-077, and Ver-155008, for their ability to impact some of the known and reported functions of this chaperone; specifically, the ability to inhibit autophagy, to influence the level of HSP90 client proteins, to induce cell cycle arrest, and to inhibit the enzymatic activity of the anaphase-promoting complex/cyclosome (APC/C). We report that all three of these compounds can inhibit autophagy and cause reduced levels of HSP90 client proteins; however, only PES-Cl can inhibit the APC/C and induce G2/M arrest. Possible reasons for these differences, and the implications for the further development of these prototype compounds as anti-cancer agents, are discussed. PMID:24100579

  7. Comparison of the activity of three different HSP70 inhibitors on apoptosis, cell cycle arrest, autophagy inhibition, and HSP90 inhibition.

    PubMed

    Budina-Kolomets, Anna; Balaburski, Gregor M; Bondar, Anastasia; Beeharry, Neil; Yen, Tim; Murphy, Maureen E

    2014-02-01

    The chaperone HSP70 promotes the survival of cells exposed to many different types of stresses, and is also potently anti-apoptotic. The major stress-induced form of this protein, HSP70-1, is overexpressed in a number of human cancers, yet is negligibly expressed in normal cells. Silencing of the gene encoding HSP70-1 (HSPA1A) is cytotoxic to transformed but not normal cells. Therefore, HSP70 is considered to be a promising cancer drug target, and there has been active interest in the identification and characterization of HSP70 inhibitors for cancer therapy. Because HSP70 behaves in a relatively non-specific manner in the control of protein folding, to date there are no reliably-identified "clients" of this protein, nor is there consensus as to what the phenotypic effects of HSP70 inhibitors are on a cancer cell. Here for the first time we compare three recently-identified HSP70 inhibitors, PES-Cl, MKT-077, and Ver-155008, for their ability to impact some of the known and reported functions of this chaperone; specifically, the ability to inhibit autophagy, to influence the level of HSP90 client proteins, to induce cell cycle arrest, and to inhibit the enzymatic activity of the anaphase-promoting complex/cyclosome (APC/C). We report that all three of these compounds can inhibit autophagy and cause reduced levels of HSP90 client proteins; however, only PES-Cl can inhibit the APC/C and induce G 2/M arrest. Possible reasons for these differences, and the implications for the further development of these prototype compounds as anti-cancer agents, are discussed. PMID:24100579

  8. Feedback and Modularity in Cell Cycle Control

    NASA Astrophysics Data System (ADS)

    Skotheim, Jan

    2009-03-01

    Underlying the wonderful diversity of natural forms is the ability of an organism to grow into its appropriate shape. Regulation ensures that cells grow, divide and differentiate so that the organism and its constitutive parts are properly proportioned and of suitable size. Although the size-control mechanism active in an individual cell is of fundamental importance to this process, it is difficult to isolate and study in complex multi-cellular systems and remains poorly understood. This motivates our use of the budding yeast model organism, whose Start checkpoint integrates multiple internal (e.g. cell size) and external signals into an irreversible decision to enter the cell cycle. We have endeavored to address the following two questions: What makes the Start transition irreversible? How does a cell compute its own size? I will report on the progress we have made. Our work is part of an emerging framework for understanding biological control circuits, which will allow us to discern the function of natural systems and aid us in engineering synthetic systems.

  9. Classic “broken cell” techniques and newer live cell methods for cell cycle assessment

    PubMed Central

    Henderson, Lindsay; Bortone, Dante S.; Lim, Curtis

    2013-01-01

    Many common, important diseases are either caused or exacerbated by hyperactivation (e.g., cancer) or inactivation (e.g., heart failure) of the cell division cycle. A better understanding of the cell cycle is critical for interpreting numerous types of physiological changes in cells. Moreover, new insights into how to control it will facilitate new therapeutics for a variety of diseases and new avenues in regenerative medicine. The progression of cells through the four main phases of their division cycle [G0/G1, S (DNA synthesis), G2, and M (mitosis)] is a highly conserved process orchestrated by several pathways (e.g., transcription, phosphorylation, nuclear import/export, and protein ubiquitination) that coordinate a core cell cycle pathway. This core pathway can also receive inputs that are cell type and cell niche dependent. “Broken cell” methods (e.g., use of labeled nucleotide analogs) to assess for cell cycle activity have revealed important insights regarding the cell cycle but lack the ability to assess living cells in real time (longitudinal studies) and with single-cell resolution. Moreover, such methods often require cell synchronization, which can perturb the pathway under study. Live cell cycle sensors can be used at single-cell resolution in living cells, intact tissue, and whole animals. Use of these more recently available sensors has the potential to reveal physiologically relevant insights regarding the normal and perturbed cell division cycle. PMID:23392113

  10. The Hydrologic Cycle Distributed Active Archive Center

    NASA Technical Reports Server (NTRS)

    Hardin, Danny M.; Goodman, H. Michael

    1995-01-01

    The Marshall Space Flight Center Distributed Active Archive Center in Huntsville, Alabama supports the acquisition, production, archival and dissemination of data relevant to the study of the global hydrologic cycle. This paper describes the Hydrologic Cycle DAAC, surveys its principle data holdings, addresses future growth, and gives information for accessing the data sets.

  11. Subversion of cell cycle regulatory mechanisms by HIV

    PubMed Central

    Rice, Andrew P.; Kimata, Jason T.

    2015-01-01

    To establish a productive infection, HIV-1 must counteract cellular innate immune mechanisms and redirect cellular process towards viral replication. Recent studies have discovered that HIV-1 and other primate immunodeficiency viruses subvert cell cycle regulatory mechanisms to achieve these ends. The viral Vpr and Vpx proteins target cell cycle controls to counter innate immunity. The cell cycle-related protein Cyclin L2 is also utilized to counter innate immunity. The viral Tat protein utilizes Cyclin T1 to activated proviral transcription, and regulation of Cyclin T1 levels in CD4+ T cells has important consequences for viral replication and latency. This review will summarize this emerging evidence that primate immunodeficiency viruses subvert cell cycle regulatory mechanisms to enhance replication. PMID:26067601

  12. Antiproliferative activity of the isoindigo 5'-Br in HL-60 cells is mediated by apoptosis, dysregulation of mitochondrial functions and arresting cell cycle at G0/G1 phase.

    PubMed

    Saleh, Ayman M; El-Abadelah, Mustafa M; Aziz, Mohammad Azhar; Taha, Mutasem O; Nasr, Amre; Rizvi, Syed A A

    2015-06-01

    Our new compound, 5'-Br [(E)-1-(5'-bromo-2'-oxoindolin-3'-ylidene)-6-ethyl-2,3,6,9-tetrahydro-2,9-dioxo-1H-pyrrolo[3,2-f]quinoline-8-carboxylic acid], had shown strong, selective antiproliferative activity against different cancer cell lines. Here, we aim to comprehensively characterize the mechanisms associated with its cytotoxicity in the human promyelocytic leukemia HL-60 cells. We focused at studying the involvement of apoptotic pathway and cell cycle effects. 5'-Br significantly inhibited proliferation by inducing caspase-dependent apoptosis. Involvement of caspase independent mechanism is also possible due to observed inability of z-VAD-FMK to rescue apoptotic cells. 5'-Br was found to trigger intrinsic apoptotic pathway as indicated by depolarization of the mitochondrial inner membrane, decreased level of cellular ATP, modulated expression and phosphorylation of Bcl-2 leading to loss of its association with Bax, and increased release of cytochrome c. 5'-Br treated cells were found arrested at G0/G1 phase with modulation in protein levels of cyclins, dependent kinases and their inhibitors. Expression and enzymatic activity of CDK2 and CDK4 was found inhibited. Retinoblastoma protein (Rb) phosphorylation was also inhibited whereas p21 protein levels were increased. These results suggest that the antiproliferative mechanisms of action of 5'-Br could involve apoptotic pathways, dysregulation of mitochondrial functions and disruption of cell cycle checkpoint. PMID:25790909

  13. How the cell cycle impacts chromatin architecture and influences cell fate

    PubMed Central

    Ma, Yiqin; Kanakousaki, Kiriaki; Buttitta, Laura

    2015-01-01

    Since the earliest observations of cells undergoing mitosis, it has been clear that there is an intimate relationship between the cell cycle and nuclear chromatin architecture. The nuclear envelope and chromatin undergo robust assembly and disassembly during the cell cycle, and transcriptional and post-transcriptional regulation of histone biogenesis and chromatin modification is controlled in a cell cycle-dependent manner. Chromatin binding proteins and chromatin modifications in turn influence the expression of critical cell cycle regulators, the accessibility of origins for DNA replication, DNA repair, and cell fate. In this review we aim to provide an integrated discussion of how the cell cycle machinery impacts nuclear architecture and vice-versa. We highlight recent advances in understanding cell cycle-dependent histone biogenesis and histone modification deposition, how cell cycle regulators control histone modifier activities, the contribution of chromatin modifications to origin firing for DNA replication, and newly identified roles for nucleoporins in regulating cell cycle gene expression, gene expression memory and differentiation. We close with a discussion of how cell cycle status may impact chromatin to influence cell fate decisions, under normal contexts of differentiation as well as in instances of cell fate reprogramming. PMID:25691891

  14. Metformin inhibits cell cycle progression of B-cell chronic lymphocytic leukemia cells

    PubMed Central

    Bruno, Silvia; Ledda, Bernardetta; Tenca, Claudya; Ravera, Silvia; Orengo, Anna Maria; Mazzarello, Andrea Nicola; Pesenti, Elisa; Casciaro, Salvatore; Racchi, Omar; Ghiotto, Fabio; Marini, Cecilia; Sambuceti, Gianmario; DeCensi, Andrea; Fais, Franco

    2015-01-01

    B-cell chronic lymphocytic leukemia (CLL) was believed to result from clonal accumulation of resting apoptosis-resistant malignant B lymphocytes. However, it became increasingly clear that CLL cells undergo, during their life, iterative cycles of re-activation and subsequent clonal expansion. Drugs interfering with CLL cell cycle entry would be greatly beneficial in the treatment of this disease. 1, 1-Dimethylbiguanide hydrochloride (metformin), the most widely prescribed oral hypoglycemic agent, inexpensive and well tolerated, has recently received increased attention for its potential antitumor activity. We wondered whether metformin has apoptotic and anti-proliferative activity on leukemic cells derived from CLL patients. Metformin was administered in vitro either to quiescent cells or during CLL cell activation stimuli, provided by classical co-culturing with CD40L-expressing fibroblasts. At doses that were totally ineffective on normal lymphocytes, metformin induced apoptosis of quiescent CLL cells and inhibition of cell cycle entry when CLL were stimulated by CD40-CD40L ligation. This cytostatic effect was accompanied by decreased expression of survival- and proliferation-associated proteins, inhibition of signaling pathways involved in CLL disease progression and decreased intracellular glucose available for glycolysis. In drug combination experiments, metformin lowered the apoptotic threshold and potentiated the cytotoxic effects of classical and novel antitumor molecules. Our results indicate that, while CLL cells after stimulation are in the process of building their full survival and cycling armamentarium, the presence of metformin affects this process. PMID:26265439

  15. Metformin inhibits cell cycle progression of B-cell chronic lymphocytic leukemia cells.

    PubMed

    Bruno, Silvia; Ledda, Bernardetta; Tenca, Claudya; Ravera, Silvia; Orengo, Anna Maria; Mazzarello, Andrea Nicola; Pesenti, Elisa; Casciaro, Salvatore; Racchi, Omar; Ghiotto, Fabio; Marini, Cecilia; Sambuceti, Gianmario; DeCensi, Andrea; Fais, Franco

    2015-09-01

    B-cell chronic lymphocytic leukemia (CLL) was believed to result from clonal accumulation of resting apoptosis-resistant malignant B lymphocytes. However, it became increasingly clear that CLL cells undergo, during their life, iterative cycles of re-activation and subsequent clonal expansion. Drugs interfering with CLL cell cycle entry would be greatly beneficial in the treatment of this disease. 1, 1-Dimethylbiguanide hydrochloride (metformin), the most widely prescribed oral hypoglycemic agent, inexpensive and well tolerated, has recently received increased attention for its potential antitumor activity. We wondered whether metformin has apoptotic and anti-proliferative activity on leukemic cells derived from CLL patients. Metformin was administered in vitro either to quiescent cells or during CLL cell activation stimuli, provided by classical co-culturing with CD40L-expressing fibroblasts. At doses that were totally ineffective on normal lymphocytes, metformin induced apoptosis of quiescent CLL cells and inhibition of cell cycle entry when CLL were stimulated by CD40-CD40L ligation. This cytostatic effect was accompanied by decreased expression of survival- and proliferation-associated proteins, inhibition of signaling pathways involved in CLL disease progression and decreased intracellular glucose available for glycolysis. In drug combination experiments, metformin lowered the apoptotic threshold and potentiated the cytotoxic effects of classical and novel antitumor molecules. Our results indicate that, while CLL cells after stimulation are in the process of building their full survival and cycling armamentarium, the presence of metformin affects this process. PMID:26265439

  16. Redox regulation of cardiomyocyte cell cycling via an ERK1/2 and c-Myc-dependent activation of cyclin D2 transcription

    PubMed Central

    Murray, Thomas V.A.; Smyrnias, Ioannis; Schnelle, Moritz; Mistry, Rajesh K.; Zhang, Min; Beretta, Matteo; Martin, Daniel; Anilkumar, Narayana; de Silva, Shana M.; Shah, Ajay M.; Brewer, Alison C.

    2015-01-01

    Adult mammalian cardiomyocytes have a very limited capacity to proliferate, and consequently the loss of cells after cardiac stress promotes heart failure. Recent evidence suggests that administration of hydrogen peroxide (H2O2), can regulate redox-dependent signalling pathway(s) to promote cardiomyocyte proliferation in vitro, but the potential relevance of such a pathway in vivo has not been tested. We have generated a transgenic (Tg) mouse model in which the H2O2-generating enzyme, NADPH oxidase 4 (Nox4), is overexpressed within the postnatal cardiomyocytes, and observed that the hearts of 1–3 week old Tg mice pups are larger in comparison to wild type (Wt) littermate controls. We demonstrate that the cardiomyocytes of Tg mouse pups have increased cell cycling capacity in vivo as determined by incorporation of 5-bromo-2′-deoxyuridine. Further, microarray analyses of the transcriptome of these Tg mouse hearts suggested that the expression of cyclin D2 is significantly increased. We investigated the molecular mechanisms which underlie this more proliferative phenotype in isolated neonatal rat cardiomyocytes (NRCs) in vitro, and demonstrate that Nox4 overexpression mediates an H2O2-dependent activation of the ERK1/2 signalling pathway, which in turn phosphorylates and activates the transcription factor c-myc. This results in a significant increase in cyclin D2 expression, which we show to be mediated, at least in part, by cis-acting c-myc binding sites within the proximal cyclin D2 promoter. Overexpression of Nox4 in NRCs results in an increase in their proliferative capacity that is ablated by the silencing of cyclin D2. We further demonstrate activation of the ERK1/2 signalling pathway, increased phosphorylation of c-myc and significantly increased expression of cyclin D2 protein in the Nox4 Tg hearts. We suggest that this pathway acts to maintain the proliferative capacity of cardiomyocytes in Nox4 Tg pups in vivo and so delays their exit from the cell

  17. Fuel cell and advanced turbine power cycle

    SciTech Connect

    White, D.J.

    1995-10-19

    Solar Turbines, Incorporated (Solar) has a vested interest in the integration of gas turbines and high temperature fuel cells and in particular, solid oxide fuel cells (SOFCs). Solar has identified a parallel path approach to the technology developments needed for future products. The primary approach is to move away from the simple cycle industrial machines of the past and develop as a first step more efficient recuperated engines. This move was prompted by the recognition that the simple cycle machines were rapidly approaching their efficiency limits. Improving the efficiency of simple cycle machines is and will become increasingly more costly. Each efficiency increment will be progressively more costly than the previous step.

  18. Retinal progenitor cells, differentiation, and barriers to cell cycle reentry.

    PubMed

    Davis, Denise M; Dyer, Michael A

    2010-01-01

    Neurogenesis in the retina occurs via the coordination of proliferation, cell cycle exit and differentiation of retinal progenitor cells. Until recently, it was widely assumed that once a retinal progenitor cell produced a postmitotic neuron, there was no possibility for cell-cycle re-entry. However, recent studies have shown that mature differentiated horizontal neurons with reduced Rb pathway function can re-enter the cell cycle and proliferate while maintaining their differentiated features. This chapter will explore the molecular and cellular mechanisms that help to keep differentiated retinal neurons and glia postmitotic. We propose that there are cell-type specific barriers to cell-cycle re-entry by differentiated neurons and these may include apoptosis, chromatin/epigenetics mechanisms, cellular morphology and/or metabolic demands that are distinct across cell populations. Our data suggest that differentiated neurons span a continuum of cellular properties related to their ability to re-enter the cell cycle and undergo cytokinesis while maintaining their differentiated features. A deeper understanding of these processes may allow us to begin to explain the cell type specificity of neuronal cell death and tumor susceptibility. For example, neurons that have more barriers to cell-cycle re-entry may be less likely to form tumors but more likely to undergo degeneration. Conversely, neurons that have fewer barriers to cell-cycle re-entry may be more likely to form tumors but less likely to undergo degeneration. PMID:20959166

  19. Artemisinin triggers a G1 cell cycle arrest of human Ishikawa endometrial cancer cells and inhibits Cyclin Dependent Kinase-4 promoter activity and expression by disrupting NF-kB transcriptional signaling

    PubMed Central

    Tran, Kalvin Q.; Tin, Antony S.; Firestone, Gary L.

    2014-01-01

    Relatively little is known about the anti-proliferative effects of Artemisinin, a naturally occurring anti-malarial compound from Artemisia annua, or sweet wormwood, in human endometrial cancer cells. Artemisinin induced a G1 cell cycle arrest in cultured human Ishikawa endometrial cancer cells and down regulated CDK2 and CDK4 transcript and protein levels. Analysis of CDK4 promoter-luciferase reporter constructs showed that the artemisinin ablation of CDK4 gene expression was accounted for by the loss of CDK4 promoter activity. Chromatin immunoprecipitation demonstrated that artemisinin inhibited nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) subunit p65 and p50 interactions with the endogenous Ishikawa cell CDK4 promoter. Coimmunoprecipitation revealed that artemisinin disrupts endogenous p65 and p50 nuclear translocation via increased protein-protein interactions with IκB-α, an NF-κB inhibitor, and disrupts its interaction with the CDK4 promoter, leading to a loss of CDK4 gene expression. Artemisinin treatment stimulated the cellular levels of IκB-α protein without altering the level of IκB-α transcripts. Finally, expression of exogenous p65 resulted in the accumulation of this NF-κB subunit in the nucleus of artemisinin treated and untreated cells, reversed the artemisinin down-regulation of CDK4 protein expression and promoter activity and prevented the artemisinin induced G1 cell cycle arrest. Taken together, our results demonstrate that a key event in the artemisinin anti-proliferative effects in endometrial cancer cells is the transcriptional down-regulation of CDK4 expression by disruption of NF-κB interactions with the CDK4 promoter. PMID:24296733

  20. Artemisinin triggers a G1 cell cycle arrest of human Ishikawa endometrial cancer cells and inhibits cyclin-dependent kinase-4 promoter activity and expression by disrupting nuclear factor-κB transcriptional signaling.

    PubMed

    Tran, Kalvin Q; Tin, Antony S; Firestone, Gary L

    2014-03-01

    Relatively little is known about the antiproliferative effects of artemisinin, a naturally occurring antimalarial compound from Artemisia annua, or sweet wormwood, in human endometrial cancer cells. Artemisinin induced a G1 cell cycle arrest in cultured human Ishikawa endometrial cancer cells and downregulated cyclin-dependent kinase-2 (CDK2) and CDK4 transcript and protein levels. Analysis of CDK4 promoter-luciferase reporter constructs showed that the artemisinin ablation of CDK4 gene expression was accounted for by the loss of CDK4 promoter activity. Chromatin immunoprecipitation demonstrated that artemisinin inhibited nuclear factor κ-light-chain-enhancer of activated B cells (NF-κB) subunit p65 and p50 interactions with the endogenous Ishikawa cell CDK4 promoter. Coimmunoprecipitation revealed that artemisinin disrupts endogenous p65 and p50 nuclear translocation through increased protein-protein interactions with IκB-α, an NF-κB inhibitor, and disrupts its interaction with the CDK4 promoter, leading to a loss of CDK4 gene expression. Artemisinin treatment stimulated the cellular levels of IκB-α protein without altering the level of IκB-α transcripts. Finally, expression of exogenous p65 resulted in the accumulation of this NF-κB subunit in the nucleus of artemisinin-treated and artemisinin-untreated cells, reversed the artemisinin downregulation of CDK4 protein expression and promoter activity, and prevented the artemisinin-induced G1 cell cycle arrest. Taken together, our results demonstrate that a key event in the artemisinin antiproliferative effects in endometrial cancer cells is the transcriptional downregulation of CDK4 expression by disruption of NF-κB interactions with the CDK4 promoter. PMID:24296733

  1. Arabidopsis thalianaMRE11 is essential for activation of cell cycle arrest, transcriptional regulation and DNA repair upon the induction of double-stranded DNA breaks.

    PubMed

    Šamanić, I; Cvitanić, R; Simunić, J; Puizina, J

    2016-07-01

    Given the fundamental role of MRE11 in many aspects of DNA metabolism and signalling in eukaryotes, we analysed the impact of several MRE11 mutations on DNA damage response (DDR) and DNA repair in Arabidopsis thaliana. Three different atmre11 and an atatm-2 mutant lines, together with the wild type (WT), were compared using a new Arabidopsis genotoxic assay for in situ evaluation of genome integrity and DNA damage repair efficiency after double strand break (DSB) induction. The results showed that, despite the phenotypic differences and different lengths of the putative truncated AtMRE11 proteins, all three atmre11 and the atatm-2 mutant lines exhibited common hypersensitivity to bleomycin treatment, where they only slightly reduced mitotic activity, indicating a G2/M checkpoint abrogation. In contrast to the WT, which reduced the frequency of chromosomal aberrations throughout the recovery period after treatment, none of the three atmre11 and atatm-2 mutants recovered. Moreover, atmre11-3 mutants, similarly to atatm-2 mutants, failed to transcriptionally induce several DDR genes and had altered expression of the CYCB1;1::GUS protein. Nevertheless, numerous chromosomal fusions in the atmre11 mutants, observed after DNA damage induction, suggest intensive DNA repair activity. These results indicate that functional and full-length AtMRE11 is essential for activation of the cell cycle arrest, transcriptional regulation and DNA repair upon induction of DSB. PMID:27007017

  2. The Cell Cycle Switch Computes Approximate Majority

    NASA Astrophysics Data System (ADS)

    Cardelli, Luca; Csikász-Nagy, Attila

    2012-09-01

    Both computational and biological systems have to make decisions about switching from one state to another. The `Approximate Majority' computational algorithm provides the asymptotically fastest way to reach a common decision by all members of a population between two possible outcomes, where the decision approximately matches the initial relative majority. The network that regulates the mitotic entry of the cell-cycle in eukaryotes also makes a decision before it induces early mitotic processes. Here we show that the switch from inactive to active forms of the mitosis promoting Cyclin Dependent Kinases is driven by a system that is related to both the structure and the dynamics of the Approximate Majority computation. We investigate the behavior of these two switches by deterministic, stochastic and probabilistic methods and show that the steady states and temporal dynamics of the two systems are similar and they are exchangeable as components of oscillatory networks.

  3. Thrombin Induces Tumor Cell Cycle Activation and Spontaneous Growth by Down-regulation of p27Kip1, in Association with the Up-regulation of Skp2 and MiR-222

    PubMed Central

    Hu, Liang; Ibrahim, Sherif; Liu, Cynthia; Skaar, Jeffrey; Pagano, Michele; Karpatkin, Simon

    2009-01-01

    The effect of thrombin on tumor cell cycle activation and spontaneous growth was examined in synchronized serum-starved tumor cell lines and a model of spontaneous prostate cancer development in TRAMP mice. BrdUrd incorporation and propidium iodide staining of prostate LNCaP cells arrested in G0 and treated with thrombin or serum revealed a 48- and 29-fold increase in S phase cells, respectively, at 8 hours. Similar results were obtained with TRAMP cells and a glioblastoma cell line, T98G. Cell cycle kinases and inhibitors in synchronized tumor cells revealed high levels of p27Kip1 and low levels of Skp2 and cyclins D1 and A. Addition of thrombin, TFLLRN, or serum down-regulated p27Kip1 with concomitant induction of Skp2, Cyclin D1, and Cyclin A with similar kinetics. LNCaP p27Kip1-transfected cells or Skp2 knockdown cells were refractory to thrombin-induced cell cycle activation. MicroRNA 222, an inhibitor of p27Kip1, was robustly up-regulated by thrombin. The in vitro observations were tested in vivo with transgenic TRAMP mice. Repetitive thrombin injection enhanced prostate tumor volume 6- to 8-fold (P < 0.04). Repetitive hirudin, a specific potent antithrombin, decreased tumor volume 13- to 24-fold (P < 0.04). Thus, thrombin stimulates tumor cell growth in vivo by down-regulation of p27Kip1. PMID:19351827

  4. Cell Cycle Synchronization in Xenopus Egg Extracts.

    PubMed

    Gillespie, Peter J; Neusiedler, Julia; Creavin, Kevin; Chadha, Gaganmeet Singh; Blow, J Julian

    2016-01-01

    Many important discoveries in cell cycle research have been made using cell-free extracts prepared from the eggs of the South African clawed frog Xenopus laevis. These extracts efficiently support the key nuclear functions of the eukaryotic cell cycle in vitro under apparently the same controls that exist in vivo. The Xenopus cell-free system is therefore uniquely suited to the study of the mechanisms, dynamics and integration of cell cycle regulated processes at a biochemical level. Here, we describe methods currently in use in our laboratory for the preparation of Xenopus egg extracts and demembranated sperm nuclei. We detail how these extracts can be used to study the key transitions of the eukaryotic cell cycle and describe conditions under which these transitions can be manipulated by addition of drugs that either retard or advance passage. In addition, we describe in detail essential techniques that provide a practical starting point for investigating the function of proteins involved in the operation of the eukaryotic cell cycle. PMID:26254920

  5. Crataegus azarolus Leaves Induce Antiproliferative Activity, Cell Cycle Arrest, and Apoptosis in Human HT-29 and HCT-116 Colorectal Cancer Cells.

    PubMed

    Mustapha, Nadia; Pinon, Aline; Limami, Youness; Simon, Alain; Ghedira, Kamel; Hennebelle, Thierry; Chekir-Ghedira, Leila

    2016-05-01

    Limited success has been achieved in extending the survival of patients with metastatic colorectal cancer (CRC). There is a strong need for novel agents in the treatment and prevention of CRC. Therefore, in the present study we evaluated the antiproliferative and pro-apoptotic potential of Crataegus azarolus ethyl acetate extract in HCT-116 and HT-29 human colorectal cancer cell lines. Moreover, we attempted to investigate the signaling pathways that should be involved in its cytotoxic effect. The Crataegus azarolus ethyl acetate extract-induced growth inhibitory effect was associated with DNA fragmentation, sub-G1 peak, loss of mitochondrial potential, and poly (ADP-ribose) polymerase (PARP) cleavage. In addition, ethyl acetate extract of Crataegus azarolus induced the cleavage of caspase-8. It has no effect on steady-state levels of total Bcl-2 protein. Whereas Bax levels decreased significantly in a dose-dependent manner in both tested cell lines. Taken together, these findings confirm the involvement of the extrinsic pathway of apoptosis. The apoptotic cell death induced by ethyl acetate extract of Crataegus azarolus was accompanied by an enhancement of the p21 expression but not through p53 activation in human colorectal cancer cells. The above-mentioned data provide insight into the molecular mechanisms of Crataegus azarolus ethyl acetate extract-induced apoptosis in CRC. Therefore, this compound should be a potential anticancer agent for the treatment of CRC. PMID:26495895

  6. Cell Cycle Phase-Specific Drug Resistance as an Escape Mechanism of Melanoma Cells.

    PubMed

    Beaumont, Kimberley A; Hill, David S; Daignault, Sheena M; Lui, Goldie Y L; Sharp, Danae M; Gabrielli, Brian; Weninger, Wolfgang; Haass, Nikolas K

    2016-07-01

    The tumor microenvironment is characterized by cancer cell subpopulations with heterogeneous cell cycle profiles. For example, hypoxic tumor zones contain clusters of cancer cells that arrest in G1 phase. It is conceivable that neoplastic cells exhibit differential drug sensitivity based on their residence in specific cell cycle phases. In this study, we used two-dimensional and organotypic melanoma culture models in combination with fluorescent cell cycle indicators to investigate the effects of cell cycle phases on clinically used drugs. We demonstrate that G1-arrested melanoma cells, irrespective of the underlying cause mediating G1 arrest, are resistant to apoptosis induced by the proteasome inhibitor bortezomib or the alkylating agent temozolomide. In contrast, G1-arrested cells were more sensitive to mitogen-activated protein kinase pathway inhibitor-induced cell death. Of clinical relevance, pretreatment of melanoma cells with a mitogen-activated protein kinase pathway inhibitor, which induced G1 arrest, resulted in resistance to temozolomide or bortezomib. On the other hand, pretreatment with temozolomide, which induced G2 arrest, did not result in resistance to mitogen-activated protein kinase pathway inhibitors. In summary, we established a model to study the effects of the cell cycle on drug sensitivity. Cell cycle phase-specific drug resistance is an escape mechanism of melanoma cells that has implications on the choice and timing of drug combination therapies. PMID:26970356

  7. Burn to cycle: energetics of cell-cycle control and stem cell maintenance.

    PubMed

    Mans, Laurie D; Haramis, Anna-Pavlina G

    2014-01-01

    Stem cells have the unique ability to both maintain the stem cell population via self-renewal and give rise to differentiated cells. The balance between these options is very delicate and important for the short- and long-term maintenance of tissue homeostasis in an organism. Pathways involved in integrating environmental cues and in directing energy metabolism play an important role in the fate decisions of stem cells. In this review, we give an overview of the effects of cellular and systemic metabolic states on stem-cell fate in both embryonic and in adult stem cell populations, with a particular emphasis on cell-cycle regulation. We discuss the major pathways implicated in sensing energetic status and regulating metabolism, including: the mTOR pathway, Forkhead-box-O transcription factors (FoxOs), Sirtuins, reactive oxygen species (ROS), AMP-activated kinase (AMPK) and LKB1, the mTOR pathway and hypoxia inducible factors (HIFs). Given the importance of a correct balance between self-renewal and differentiation, understanding the mechanisms that drive stem-cell fate in different metabolic conditions will provide more insight in stem cell biology in both health and disease. PMID:24896332

  8. Cycle life test of secondary spacecraft cells

    NASA Astrophysics Data System (ADS)

    Harkness, J. D.

    1980-04-01

    The results of the life cycling program on rechargeable calls are reported. Information on required data, the use of which the data will be put, application details, including orbital description, charge control methods, load rquirements, etc., are given. Cycle tests were performed on 660 sealed, nickel cadmium cells. The cells consisted of seven sample classifications ranging form 3.0 to 20 amp. hours. Nickel cadmium, silver cadmium, and silver zinc sealed cells, excluding synchronous orbit and accelerated test packs were added. The capacities of the nickel cadmium cells, the silver cadmium and the silver zinc cells differed in range of amp hrs. The cells were cylced under different load, charge control, and temperature conditions. All cell packs are recharged by use of a pack voltage limit. All charging is constant current until the voltage limit is reached.

  9. Cycle life test of secondary spacecraft cells

    NASA Technical Reports Server (NTRS)

    Harkness, J. D.

    1980-01-01

    The results of the life cycling program on rechargeable calls are reported. Information on required data, the use of which the data will be put, application details, including orbital description, charge control methods, load rquirements, etc., are given. Cycle tests were performed on 660 sealed, nickel cadmium cells. The cells consisted of seven sample classifications ranging form 3.0 to 20 amp. hours. Nickel cadmium, silver cadmium, and silver zinc sealed cells, excluding synchronous orbit and accelerated test packs were added. The capacities of the nickel cadmium cells, the silver cadmium and the silver zinc cells differed in range of amp hrs. The cells were cylced under different load, charge control, and temperature conditions. All cell packs are recharged by use of a pack voltage limit. All charging is constant current until the voltage limit is reached.

  10. Juvenile Hormone Activates the Transcription of Cell-division-cycle 6 (Cdc6) for Polyploidy-dependent Insect Vitellogenesis and Oogenesis.

    PubMed

    Wu, Zhongxia; Guo, Wei; Xie, Yingtian; Zhou, Shutang

    2016-03-01

    Although juvenile hormone (JH) is known to prevent insect larval metamorphosis and stimulate adult reproduction, the molecular mechanisms of JH action in insect reproduction remain largely unknown. Earlier, we reported that the JH-receptor complex, composed of methoprene-tolerant and steroid receptor co-activator, acts on mini-chromosome maintenance (Mcm) genes Mcm4 and Mcm7 to promote DNA replication and polyploidy for the massive vitellogenin (Vg) synthesis required for egg production in the migratory locust (Guo, W., Wu, Z., Song, J., Jiang, F., Wang, Z., Deng, S., Walker, V. K., and Zhou, S. (2014) PLoS Genet. 10, e1004702). In this study we have investigated the involvement of cell-division-cycle 6 (Cdc6) in JH-dependent vitellogenesis and oogenesis, as Cdc6 is essential for the formation of prereplication complex. We demonstrate here that Cdc6 is expressed in response to JH and methoprene-tolerant, and Cdc6 transcription is directly regulated by the JH-receptor complex. Knockdown of Cdc6 inhibits polyploidization of fat body and follicle cells, resulting in the substantial reduction of Vg expression in the fat body as well as severely impaired oocyte maturation and ovarian growth. Our data indicate the involvement of Cdc6 in JH pathway and a pivotal role of Cdc6 in JH-mediated polyploidization, vitellogenesis, and oogenesis. PMID:26728459

  11. Looking at plant cell cycle from the chromatin window

    PubMed Central

    Desvoyes, Bénédicte; Fernández-Marcos, María; Sequeira-Mendes, Joana; Otero, Sofía; Vergara, Zaida; Gutierrez, Crisanto

    2014-01-01

    The cell cycle is defined by a series of complex events, finely coordinated through hormonal, developmental and environmental signals, which occur in a unidirectional manner and end up in producing two daughter cells. Accumulating evidence reveals that chromatin is not a static entity throughout the cell cycle. In fact, there are many changes that include nucleosome remodeling, histone modifications, deposition and exchange, among others. Interestingly, it is possible to correlate the occurrence of several of these chromatin-related events with specific processes necessary for cell cycle progression, e.g., licensing of DNA replication origins, the E2F-dependent transcriptional wave in G1, the activation of replication origins in S-phase, the G2-specific transcription of genes required for mitosis or the chromatin packaging occurring in mitosis. Therefore, an emerging view is that chromatin dynamics must be considered as an intrinsic part of cell cycle regulation. In this article, we review the main features of several key chromatin events that occur at defined times throughout the cell cycle and discuss whether they are actually controlling the transit through specific cell cycle stages. PMID:25120553

  12. The inhibition of activated hepatic stellate cells proliferation by arctigenin through G0/G1 phase cell cycle arrest: persistent p27(Kip1) induction by interfering with PI3K/Akt/FOXO3a signaling pathway.

    PubMed

    Li, Ao; Wang, Jun; Wu, Mingjun; Zhang, Xiaoxun; Zhang, Hongzhi

    2015-01-15

    Proliferation of hepatic stellate cells (HSCs) is vital for the development of fibrosis during liver injury. In this study, we describe that arctigenin (ATG), a major bioactive component of Fructus Arctii, exhibited selective cytotoxic activity via inhibiting platelet-derived growth factor-BB (PDGF-BB)-activated HSCs proliferation and arrested cell cycle at G0/G1 phase, which could not be observed in normal human hepatocytes in vitro. The cyclin-dependent kinase (CDK) 4/6 activities could be strongly inhibited by ATG through down-regulation of cyclin D1 and CDK4/6 expression in early G1 phase arrest. In the ATG-treated HSCs, the expression level of p27(Kip1) and the formation of CDK2-p27(Kip1) complex were also increased. p27(Kip1) silencing significantly attenuated the effect of ATG, including cell cycle arrest and suppression of proliferation in activated HSCs. We also found that ATG suppressed PDGF-BB-induced phosphorylation of Akt and its downstream transcription factor Forkhead box O 3a (FOXO3a), decreased binding of FOXO3a to 14-3-3 protein, and stimulated nuclear translocation of FOXO3a in activated HSCs. Furthermore, knockdown of FOXO3a expression by FOXO3a siRNA attenuated ATG-induced up-regulation of p27(Kip1) in activated HSCs. All the above findings suggested that ATG could increase the levels of p27(Kip1) protein through inhibition of Akt and improvement of FOXO3a activity, in turn inhibited the CDK2 kinase activity, and eventually caused an overall inhibition of HSCs proliferation. PMID:25498792

  13. Cell Cycle Regulation in the Developing Lens

    PubMed Central

    Griep, Anne E.

    2007-01-01

    Regulation of cell proliferation is a critical aspect of the development of multicellular organisms. The ocular lens is an excellent model system in which to unravel the mechanisms controlling cell proliferation during development. In recent years, several cell cycle regulators have been shown to be essential for maintaining normal patterns of lens cell proliferation. Additionally, many growth factor signaling pathways and cell adhesion factors have been shown to have the capacity to regulate lens cell proliferation. Given this complexity, understanding the cross talk between these many signaling pathways and how they are coordinated are important directions for the future. PMID:17218126

  14. Astaxanthin Inhibits Proliferation and Induces Apoptosis and Cell Cycle Arrest of Mice H22 Hepatoma Cells

    PubMed Central

    Shao, Yiye; Ni, Yanbo; Yang, Jing; Lin, Xutao; Li, Jun; Zhang, Lixia

    2016-01-01

    Background It is widely recognized that astaxanthin (ASX), a member of the carotenoid family, has strong biological activities including antioxidant, anti-inflammation, and immune-modulation activities. Previous studies have confirmed that ASX can effectively inhibit hepatoma cells in vitro. Material/Methods MTT was used to assay proliferation of mice H22 cells, and flow cytometry was used to determine apoptosis and cell cycle arrest of H22 cells in vitro and in vivo. Moreover, anti-tumor activity of ASX was observed in mice. Results ASX inhibited the proliferation of H22 cells, promoted cell necrosis, and induced cell cycle arrest in G2 phase in vitro and in vivo. Conclusions This study indicated that ASX can inhibit proliferation and induce apoptosis and cell cycle arrest in mice H22 hepatoma cells in vitro and in vivo. PMID:27333866

  15. Astaxanthin Inhibits Proliferation and Induces Apoptosis and Cell Cycle Arrest of Mice H22 Hepatoma Cells.

    PubMed

    Shao, Yiye; Ni, Yanbo; Yang, Jing; Lin, Xutao; Li, Jun; Zhang, Lixia

    2016-01-01

    BACKGROUND It is widely recognized that astaxanthin (ASX), a member of the carotenoid family, has strong biological activities including antioxidant, anti-inflammation, and immune-modulation activities. Previous studies have confirmed that ASX can effectively inhibit hepatoma cells in vitro. MATERIAL AND METHODS MTT was used to assay proliferation of mice H22 cells, and flow cytometry was used to determine apoptosis and cell cycle arrest of H22 cells in vitro and in vivo. Moreover, anti-tumor activity of ASX was observed in mice. RESULTS ASX inhibited the proliferation of H22 cells, promoted cell necrosis, and induced cell cycle arrest in G2 phase in vitro and in vivo. CONCLUSIONS This study indicated that ASX can inhibit proliferation and induce apoptosis and cell cycle arrest in mice H22 hepatoma cells in vitro and in vivo. PMID:27333866

  16. Flavonoids: from cell cycle regulation to biotechnology.

    PubMed

    Woo, Ho-Hyung; Jeong, Byeong Ryong; Hawes, Martha C

    2005-03-01

    Flavonoids have been proposed to play diverse roles in plant growth and development, including defense, symbiosis, pollen development and male fertility, polar auxin transport, and protection against ultraviolet radiation. Recently, a new role in cell cycle regulation has emerged. Genetic alteration of glucuronide metabolism by altered expression of a Pisum sativum UDP-glucuronosyltransferase (PsUGT1) results in an altered cell cycle in pea, alfalfa, and Arabidopsis. In alfalfa, altered expression of PsUGT1 results in accumulation of a flavonoid-like compound that suppresses growth of cultured cells. The results are consistent with the hypothesis that PsUGT1 functions by controlling cellular levels of a factor controlling cell cycle (FCC). PMID:15834800

  17. Cell cycling with the SEB: a personal view.

    PubMed

    Bryant, John

    2014-06-01

    This review, written from a personal perspective, traces firstly the development of plant cell cycle research from the 1970s onwards, with some focus on the work of the author and of Dr Dennis Francis. Secondly there is a discussion of the support for and discussion of plant cell cycle research in the SEB, especially through the activities of the Cell Cycle Group within the Society's Cell Biology Section. In the main part of the review, selected aspects of DNA replication that have of been of special interest to the author are discussed. These are DNA polymerases and associated proteins, pre-replication events, regulation of enzymes and other proteins, nature and activation of DNA replication origins, and DNA endoreduplication. For all these topics, there is mention of the author's own work, followed by a brief synthesis of current understanding and a look to possible future developments. PMID:24493805

  18. NONO couples the circadian clock to the cell cycle

    PubMed Central

    Kowalska, Elzbieta; Ripperger, Juergen A.; Hoegger, Dominik C.; Bruegger, Pascal; Buch, Thorsten; Birchler, Thomas; Mueller, Anke; Albrecht, Urs; Contaldo, Claudio; Brown, Steven A.

    2013-01-01

    Mammalian circadian clocks restrict cell proliferation to defined time windows, but the mechanism and consequences of this interrelationship are not fully understood. Previously we identified the multifunctional nuclear protein NONO as a partner of circadian PERIOD (PER) proteins. Here we show that it also conveys circadian gating to the cell cycle, a connection surprisingly important for wound healing in mice. Specifically, although fibroblasts from NONO-deficient mice showed approximately normal circadian cycles, they displayed elevated cell doubling and lower cellular senescence. At a molecular level, NONO bound to the p16-Ink4A cell cycle checkpoint gene and potentiated its circadian activation in a PER protein-dependent fashion. Loss of either NONO or PER abolished this activation and circadian expression of p16-Ink4A and eliminated circadian cell cycle gating. In vivo, lack of NONO resulted in defective wound repair. Because wound healing defects were also seen in multiple circadian clock-deficient mouse lines, our results therefore suggest that coupling of the cell cycle to the circadian clock via NONO may be useful to segregate in temporal fashion cell proliferation from tissue organization. PMID:23267082

  19. Integrin α(V)β(3)-targeted magnetic nanohybrids with enhanced antitumor efficacy, cell cycle arrest ability, and encouraging anti-cell-migration activity.

    PubMed

    Ding, Guo-Bin; Wang, Yan; Guo, Yi; Xu, Li

    2014-10-01

    Organic/inorganic nanohybrids, which integrate advantages of the biocompatibility of organic polymers and diversified functionalities of inorganic nanoparticles, have been extensively investigated in recent years. Herein, we report the construction of arginine-glycine-aspartic acid-cysteine (RGDC) tetrapeptide functionalized and 10-hydroxycamptothecin (HCPT)-encapsulated magnetic nanohybrids (RFHEMNs) for integrin αVβ3-targeted drug delivery. The obtained RFHEMNs were near-spherical in shape with a homogeneous size about 50 nm, and exhibited a superparamagnetic behavior. In vitro drug release study showed a sustained and pH-dependent release profile. Cell viability tests revealed that RFHEMNs displayed a significant enhancement of cytotoxicity against αVβ3-overexpressing A549 cells, as compared to free HCPT and nontargeting micelles. Flow cytometry analysis indicated that this cytotoxic effect was associated with dose-dependent S phase arrest. Finally, RFHEMNs exerted encouraging anti-cell-migration activity as determined by an in vitro wound-healing assay and a transwell assay. Overall, we envision that this tumor-targeting nanoscale drug delivery system may be of great application potential in chemotherapy of primary tumor and their metastases. PMID:25207865

  20. Activity of the human immunodeficiency virus type 1 cell cycle-dependent internal ribosomal entry site is modulated by IRES trans-acting factors.

    PubMed

    Vallejos, Maricarmen; Deforges, Jules; Plank, Terra-Dawn M; Letelier, Alejandro; Ramdohr, Pablo; Abraham, Christopher G; Valiente-Echeverría, Fernando; Kieft, Jeffrey S; Sargueil, Bruno; López-Lastra, Marcelo

    2011-08-01

    The 5' leader of the human immunodeficiency virus type 1 (HIV-1) genomic RNA harbors an internal ribosome entry site (IRES) that is functional during the G2/M phase of the cell cycle. Here we show that translation initiation mediated by the HIV-1 IRES requires the participation of trans-acting cellular factors other than the canonical translational machinery. We used 'standard' chemical and enzymatic probes and an 'RNA SHAPE' analysis to model the structure of the HIV-1 5' leader and we show, by means of a footprinting assay, that G2/M extracts provide protections to regions previously identified as crucial for HIV-1 IRES activity. We also assessed the impact of mutations on IRES function. Strikingly, mutations did not significantly affect IRES activity suggesting that the requirement for pre-formed stable secondary or tertiary structure within the HIV-1 IRES may not be as strict as has been described for other viral IRESes. Finally, we used a proteomic approach to identify cellular proteins within the G2/M extracts that interact with the HIV-1 5' leader. Together, data show that HIV-1 IRES-mediated translation initiation is modulated by cellular proteins. PMID:21482538

  1. Activity of the human immunodeficiency virus type 1 cell cycle-dependent internal ribosomal entry site is modulated by IRES trans-acting factors

    PubMed Central

    Vallejos, Maricarmen; Deforges, Jules; Plank, Terra-Dawn M.; Letelier, Alejandro; Ramdohr, Pablo; Abraham, Christopher G.; Valiente-Echeverría, Fernando; Kieft, Jeffrey S.; Sargueil, Bruno; López-Lastra, Marcelo

    2011-01-01

    The 5′ leader of the human immunodeficiency virus type 1 (HIV-1) genomic RNA harbors an internal ribosome entry site (IRES) that is functional during the G2/M phase of the cell cycle. Here we show that translation initiation mediated by the HIV-1 IRES requires the participation of trans-acting cellular factors other than the canonical translational machinery. We used ‘standard’ chemical and enzymatic probes and an ‘RNA SHAPE’ analysis to model the structure of the HIV-1 5′ leader and we show, by means of a footprinting assay, that G2/M extracts provide protections to regions previously identified as crucial for HIV-1 IRES activity. We also assessed the impact of mutations on IRES function. Strikingly, mutations did not significantly affect IRES activity suggesting that the requirement for pre-formed stable secondary or tertiary structure within the HIV-1 IRES may not be as strict as has been described for other viral IRESes. Finally, we used a proteomic approach to identify cellular proteins within the G2/M extracts that interact with the HIV-1 5′ leader. Together, data show that HIV-1 IRES-mediated translation initiation is modulated by cellular proteins. PMID:21482538

  2. A possible activity cycle in Proxima Centauri

    NASA Astrophysics Data System (ADS)

    Cincunegui, C.; Díaz, R. F.; Mauas, P. J. D.

    2007-01-01

    Context: Several late-type stars present activity cycles resembling the Solar one. This fact has been observed mostly in stars ranging from F to K, i.e., in stars with a radiative core and an outer convective layer. Aims: This work aims at studying whether an activity cycle can be detected in the dM5.5e star Proxima Centauri, which is supposed to be completely convective. Methods: We present periodical medium-resolution echelle observations covering the complete visual range, which were taken at the CASLEO Argentinean Observatory. These observations are distributed over 7 years. We discarded the spectra that present flare activity, and analyze the remaining activity levels using four different statistical techniques to look for a period of activity. Results: We find strong evidence of a cyclic activity, with a period of ~442 days. We also estimate that the Ca ~II S index varies around 130% due to activity variations outside of flares.

  3. Parvovirus infection-induced cell death and cell cycle arrest

    PubMed Central

    Chen, Aaron Yun; Qiu, Jianming

    2011-01-01

    The cytopathic effects induced during parvovirus infection have been widely documented. Parvovirus infection-induced cell death is often directly associated with disease outcomes (e.g., anemia resulting from loss of erythroid progenitors during parvovirus B19 infection). Apoptosis is the major form of cell death induced by parvovirus infection. However, nonapoptotic cell death, namely necrosis, has also been reported during infection of the minute virus of mice, parvovirus H-1 and bovine parvovirus. Recent studies have revealed multiple mechanisms underlying the cell death during parvovirus infection. These mechanisms vary in different parvoviruses, although the large nonstructural protein (NS)1 and the small NS proteins (e.g., the 11 kDa of parvovirus B19), as well as replication of the viral genome, are responsible for causing infection-induced cell death. Cell cycle arrest is also common, and contributes to the cytopathic effects induced during parvovirus infection. While viral NS proteins have been indicated to induce cell cycle arrest, increasing evidence suggests that a cellular DNA damage response triggered by an invading single-stranded parvoviral genome is the major inducer of cell cycle arrest in parvovirus-infected cells. Apparently, in response to infection, cell death and cell cycle arrest of parvovirus-infected cells are beneficial to the viral cell lifecycle (e.g., viral DNA replication and virus egress). In this article, we will discuss recent advances in the understanding of the mechanisms underlying parvovirus infection-induced cell death and cell cycle arrest. PMID:21331319

  4. Genome-wide examination of myoblast cell cycle withdrawal duringdifferentiation

    SciTech Connect

    Shen, Xun; Collier, John Michael; Hlaing, Myint; Zhang, Leanne; Delshad, Elizabeth H.; Bristow, James; Bernstein, Harold S.

    2002-12-02

    Skeletal and cardiac myocytes cease division within weeks of birth. Although skeletal muscle retains limited capacity for regeneration through recruitment of satellite cells, resident populations of adult myocardial stem cells have not been identified. Because cell cycle withdrawal accompanies myocyte differentiation, we hypothesized that C2C12 cells, a mouse myoblast cell line previously used to characterize myocyte differentiation, also would provide a model for studying cell cycle withdrawal during differentiation. C2C12 cells were differentiated in culture medium containing horse serum and harvested at various time points to characterize the expression profiles of known cell cycle and myogenic regulatory factors by immunoblot analysis. BrdU incorporation decreased dramatically in confluent cultures 48 hr after addition of horse serum, as cells started to form myotubes. This finding was preceded by up-regulation of MyoD, followed by myogenin, and activation of Bcl-2. Cyclin D1 was expressed in proliferating cultures and became undetectable in cultures containing 40 percent fused myotubes, as levels of p21(WAF1/Cip1) increased and alpha-actin became detectable. Because C2C12 myoblasts withdraw from the cell cycle during myocyte differentiation following a course that recapitulates this process in vivo, we performed a genome-wide screen to identify other gene products involved in this process. Using microarrays containing approximately 10,000 minimally redundant mouse sequences that map to the UniGene database of the National Center for Biotechnology Information, we compared gene expression profiles between proliferating, differentiating, and differentiated C2C12 cells and verified candidate genes demonstrating differential expression by RT-PCR. Cluster analysis of differentially expressed genes revealed groups of gene products involved in cell cycle withdrawal, muscle differentiation, and apoptosis. In addition, we identified several genes, including DDAH2 and Ly

  5. Piperlongumine Suppresses Proliferation of Human Oral Squamous Cell Carcinoma through Cell Cycle Arrest, Apoptosis and Senescence.

    PubMed

    Chen, San-Yuan; Liu, Geng-Hung; Chao, Wen-Ying; Shi, Chung-Sheng; Lin, Ching-Yen; Lim, Yun-Ping; Lu, Chieh-Hsiang; Lai, Peng-Yeh; Chen, Hau-Ren; Lee, Ying-Ray

    2016-01-01

    Oral squamous cell carcinoma (OSCC), an aggressive cancer originating in the oral cavity, is one of the leading causes of cancer deaths in males worldwide. This study investigated the antitumor activity and mechanisms of piperlongumine (PL), a natural compound isolated from Piper longum L., in human OSCC cells. The effects of PL on cell proliferation, the cell cycle, apoptosis, senescence and reactive oxygen species (ROS) levels in human OSCC cells were investigated. PL effectively inhibited cell growth, caused cell cycle arrest and induced apoptosis and senescence in OSCC cells. Moreover, PL-mediated anti-human OSCC behavior was inhibited by an ROS scavenger N-acetyl-l-cysteine (NAC) treatment, suggesting that regulation of ROS was involved in the mechanism of the anticancer activity of PL. These findings suggest that PL suppresses tumor growth by regulating the cell cycle and inducing apoptosis and senescence and is a potential chemotherapy agent for human OSCC cells. PMID:27120594

  6. Piperlongumine Suppresses Proliferation of Human Oral Squamous Cell Carcinoma through Cell Cycle Arrest, Apoptosis and Senescence

    PubMed Central

    Chen, San-Yuan; Liu, Geng-Hung; Chao, Wen-Ying; Shi, Chung-Sheng; Lin, Ching-Yen; Lim, Yun-Ping; Lu, Chieh-Hsiang; Lai, Peng-Yeh; Chen, Hau-Ren; Lee, Ying-Ray

    2016-01-01

    Oral squamous cell carcinoma (OSCC), an aggressive cancer originating in the oral cavity, is one of the leading causes of cancer deaths in males worldwide. This study investigated the antitumor activity and mechanisms of piperlongumine (PL), a natural compound isolated from Piper longum L., in human OSCC cells. The effects of PL on cell proliferation, the cell cycle, apoptosis, senescence and reactive oxygen species (ROS) levels in human OSCC cells were investigated. PL effectively inhibited cell growth, caused cell cycle arrest and induced apoptosis and senescence in OSCC cells. Moreover, PL-mediated anti-human OSCC behavior was inhibited by an ROS scavenger N-acetyl-l-cysteine (NAC) treatment, suggesting that regulation of ROS was involved in the mechanism of the anticancer activity of PL. These findings suggest that PL suppresses tumor growth by regulating the cell cycle and inducing apoptosis and senescence and is a potential chemotherapy agent for human OSCC cells. PMID:27120594

  7. Synthesis, Biological Evaluation, and Structure–Activity Relationships of Novel Substituted N-Phenyl Ureidobenzenesulfonate Derivatives Blocking Cell Cycle Progression in S-Phase and Inducing DNA Double-Strand Breaks

    PubMed Central

    2012-01-01

    Twenty-eight new substituted N-phenyl ureidobenzenesulfonate (PUB-SO) and 18 N-phenylureidobenzenesulfonamide (PUB-SA) derivatives were prepared. Several PUB-SOs exhibited antiproliferative activity at the micromolar level against the HT-29, M21, and MCF-7 cell lines and blocked cell cycle progression in S-phase similarly to cisplatin. In addition, PUB-SOs induced histone H2AX (γH2AX) phosphorylation, indicating that these molecules induce DNA double-strand breaks. In contrast, PUB-SAs were less active than PUB-SOs and did not block cell cycle progression in S-phase. Finally, PUB-SOs 4 and 46 exhibited potent antitumor activity in HT-1080 fibrosarcoma cells grafted onto chick chorioallantoic membranes, which was similar to cisplatin and combretastatin A-4 and without significant toxicity toward chick embryos. These new compounds are members of a promising new class of anticancer agents. PMID:22694057

  8. Polychlorinated Biphenyl Quinone Metabolite Promotes p53-Dependent DNA Damage Checkpoint Activation, S-Phase Cycle Arrest and Extrinsic Apoptosis in Human Liver Hepatocellular Carcinoma HepG2 Cells.

    PubMed

    Song, Xiufang; Li, Lingrui; Shi, Qiong; Lehmler, Hans-Joachim; Fu, Juanli; Su, Chuanyang; Xia, Xiaomin; Song, Erqun; Song, Yang

    2015-11-16

    Polychlorinated biphenyls (PCBs) are a group of persistent organic pollutants. The toxic behavior and mechanism of PCBs individuals and congeners have been extensively investigated. However, there is only limited information on their metabolites. Our previous studies have shown that a synthetic PCB metabolite, PCB29-pQ, causes oxidative damage with the evidence of cytotoxicity, genotoxicity, and mitochondrial-derived intrinsic apoptosis. Here, we investigate the effects of PCB29-pQ on DNA damage checkpoint activation, cell cycle arrest, and death receptor-related extrinsic apoptosis in human liver hepatocellular carcinoma HepG2 cells. Our results illustrate that PCB29-pQ increases the S-phase cell population by down-regulating cyclins A/D1/E, cyclin-dependent kinases (CDK 2/4/6), and cell division cycle 25A (CDC25A) and up-regulating p21/p27 protein expressions. PCB29-pQ also induces apoptosis via the up-regulation of Fas/FasL and the activation of caspase 8/3. Moreover, p53 plays a pivotal role in PCB29-pQ-induced cell cycle arrest and apoptosis via the activation of ATM/Chk2 and ATR/Chk1 checkpoints. Cell cycle arrest and apoptotic cell death were attenuated by the pretreatment with antioxidant N-acetyl-cysteine (NAC). Taken together, these results demonstrate that PCB29-pQ induces oxidative stress and promotes p53-dependent DNA damage checkpoint activation, S-phase cycle arrest, and extrinsic apoptosis in HepG2 cells. PMID:26451628

  9. Modeling of Sonos Memory Cell Erase Cycle

    NASA Technical Reports Server (NTRS)

    Phillips, Thomas A.; MacLeond, Todd C.; Ho, Fat D.

    2010-01-01

    Silicon-oxide-nitride-oxide-silicon (SONOS) nonvolatile semiconductor memories (NVSMS) have many advantages. These memories are electrically erasable programmable read-only memories (EEPROMs). They utilize low programming voltages, endure extended erase/write cycles, are inherently resistant to radiation, and are compatible with high-density scaled CMOS for low power, portable electronics. The SONOS memory cell erase cycle was investigated using a nonquasi-static (NQS) MOSFET model. The SONOS floating gate charge and voltage, tunneling current, threshold voltage, and drain current were characterized during an erase cycle. Comparisons were made between the model predictions and experimental device data.

  10. Human Fucci Pancreatic Beta Cell Lines: New Tools to Study Beta Cell Cycle and Terminal Differentiation

    PubMed Central

    Carlier, Géraldine; Maugein, Alicia; Cordier, Corinne; Pechberty, Séverine; Garfa-Traoré, Meriem; Martin, Patrick; Scharfmann, Raphaël; Albagli, Olivier

    2014-01-01

    Regulation of cell cycle in beta cells is poorly understood, especially in humans. We exploited here the recently described human pancreatic beta cell line EndoC-βH2 to set up experimental systems for cell cycle studies. We derived 2 populations from EndoC-βH2 cells that stably harbor the 2 genes encoding the Fucci fluorescent indicators of cell cycle, either from two vectors, or from a unique bicistronic vector. In proliferating non-synchronized cells, the 2 Fucci indicators revealed cells in the expected phases of cell cycle, with orange and green cells being in G1 and S/G2/M cells, respectively, and allowed the sorting of cells in different substeps of G1. The Fucci indicators also faithfully red out alterations in human beta cell proliferative activity since a mitogen-rich medium decreased the proportion of orange cells and inflated the green population, while reciprocal changes were observed when cells were induced to cease proliferation and increased expression of some beta cell genes. In the last situation, acquisition of a more differentiated beta cell phenotype correlates with an increased intensity in orange fluorescence. Hence Fucci beta cell lines provide new tools to address important questions regarding human beta cell cycle and differentiation. PMID:25259951

  11. K+ channels and cell cycle progression in tumor cells

    PubMed Central

    Ouadid-Ahidouch, Halima; Ahidouch, Ahmed

    2013-01-01

    K+ ions play a major role in many cellular processes. The deregulation of K+ signaling is associated with a variety of diseases such as hypertension, atherosclerosis, or diabetes. K+ ions are important for setting the membrane potential, the driving force for Ca2+ influx, and regulate volume of growing cells. Moreover, it is increasingly recognized that K+ channels control cell proliferation through a novel signaling mechanisms triggered and modulated independently of ion fluxes. In cancer, aberrant expression, regulation and/or sublocalization of K+ channels can alter the downstream signals that converge on the cell cycle machinery. Various K+ channels are involved in cell cycle progression and are needed only at particular stages of the cell cycle. Consistent with this idea, the expression of Eag1 and HERG channels fluctuate along the cell cycle. Despite of acquired knowledge, our understanding of K+ channels functioning in cancer cells requires further studies. These include identifying the molecular mechanisms controlling the cell cycle machinery. By understanding how K+ channels regulate cell cycle progression in cancer cells, we will gain insights into how cancer cells subvert the need for K+ signal and its downstream targets to proliferate. PMID:23970866

  12. A possible activity cycle in Proxima Centauri

    NASA Astrophysics Data System (ADS)

    Cincunegui, C.; Díaz, R. F.; Mauas, P. J. D.

    Several late-type stars (stars with a radiative core and an outer convective layer) present activity cycles resembling the Solar one. This work aims at studying whether an activity cycle can be detected in the dM5.5e star Proxima Centauri, which is supposed to be completely convective. We present periodical medium-resolution echelle observations covering the complete visual range, which were taken at the CASLEO Argentinean Observatory. These observations are distributed along 7 years. We analize the activity levels to look for a period of activity. We find strong evidence of a cyclic activity, with a period of ˜442 days. We also estimated that the Ca II S index varies around 130% due to activity variations outside of flares.

  13. Global water cycle and solar activity variations

    NASA Astrophysics Data System (ADS)

    Al-Tameemi, Muthanna A.; Chukin, Vladimir V.

    2016-05-01

    The water cycle is the most active and most important component in the circulation of global mass and energy in the Earth system. Furthermore, water cycle parameters such as evaporation, precipitation, and precipitable water vapour play a major role in global climate change. In this work, we attempt to determine the impact of solar activity on the global water cycle by analyzing the global monthly values of precipitable water vapour, precipitation, and the Solar Modulation Potential in 1983-2008. The first object of this study was to calculate global evaporation for the period 1983-2008. For this purpose, we determined the water cycle rate from satellite data, and precipitation/evaporation relationship from 10 years of Planet Simulator model data. The second object of our study was to investigate the relationship between the Solar Modulation Potential (solar activity index) and the evaporation for the period 1983-2008. The results showed that there is a relationship between the solar modulation potential and the evaporation values for the period of study. Therefore, we can assume that the solar activity has an impact on the global water cycle.

  14. Does Arabidopsis thaliana DREAM of cell cycle control?

    PubMed

    Fischer, Martin; DeCaprio, James A

    2015-08-01

    Strict temporal control of cell cycle gene expression is essential for all eukaryotes including animals and plants. DREAM complexes have been identified in worm, fly, and mammals, linking several distinct transcription factors to coordinate gene expression throughout the cell cycle. In this issue of The EMBO Journal, Kobayashi et al (2015) identify distinct activator and repressor complexes for genes expressed during the G2 and M phases in Arabidopsis that can be temporarily separated during proliferating and post‐mitotic stages of development. The complexes incorporate specific activator and repressor MYB and E2F transcription factors and indicate the possibility of the existence of multiple DREAM complexes in plants. PMID:26089020

  15. Cell cycle regulators and their abnormalities in breast cancer.

    PubMed Central

    Fernández, P L; Jares, P; Rey, M J; Campo, E; Cardesa, A

    1998-01-01

    One of the main properties of cancer cells is their increased and deregulated proliferative activity. It is now well known that abnormalities in many positive and negative modulators of the cell cycle are frequent in many cancer types, including breast carcinomas. Abnormalities such as defective function of the retinoblastoma gene and cyclin-dependent kinase inhibitors (for example, p16, p21, and p27), as well as upregulation of cyclins, are often seen in breast tumours. These abnormalities are sometimes coincidental, and newly described interplays between them suggest the existence of a complex regulatory web in the cell cycle. PMID:10193510

  16. Nanosecond pulsed electric fields and the cell cycle

    NASA Astrophysics Data System (ADS)

    Mahlke, Megan A.

    Exposure to nanosecond pulsed electrical fields (nsPEFs) can cause poration of external and internal cell membranes, DNA damage, and disassociation of cytoskeletal components, all of which are capable of disrupting a cell's ability to replicate. The phase of the cell cycle at the time of exposure is linked to differential sensitivities to nsPEFs across cell lines, as DNA structure, membrane elasticity, and cytoskeletal structure change dramatically during the cell cycle. Additionally, nsPEFs are capable of activating cell cycle checkpoints, which could lead to apoptosis or slow population growth. NsPEFs are emerging as a method for treating tumors via apoptotic induction; therefore, investigating the relevance of nsPEFs and the cell cycle could translate into improved efficacy in tumor treatment. Populations of Jurkat and Chinese Hamster Ovary (CHO) cells were examined post-exposure (10 ns pulse trains at 150kV/cm) by analysis of DNA content via propidium iodide staining and flow cytometric analysis at various time points (1, 6, and 12h post-exposure) to determine population distribution in cell cycle phases. Additionally, CHO and Jurkat cells were synchronized in G1/S and G2/M phases, pulsed, and analyzed to evaluate the role of cell cycle phase in survival of nsPEFs. CHO populations appeared similar to sham populations post-nsPEFs but exhibited arrest in the G1 phase at 6h after exposure. Jurkat cells exhibited increased cell death after nsPEFs compared to CHO cells but did not exhibit checkpoint arrest at any observed time point. The G1/S phase checkpoint is partially controlled by the action of p53; the lack of an active p53 response in Jurkat cells could contribute to their ability to pass this checkpoint and resist cell cycle arrest. Both cell lines exhibited increased sensitivity to nsPEFs in G2/M phase. Live imaging of CHO cells after nsPEF exposure supports the theory of G1/S phase arrest, as a reduced number of cells undergo mitosis within 24 h when

  17. Delayed Cell Cycle Progression in Selenoprotein W-depleted Cells Is Regulated by a Mitogen-activated Protein Kinase Kinase 4-p38/c-Jun NH2-terminal Kinase-p53 Pathway*

    PubMed Central

    Hawkes, Wayne Chris; Alkan, Zeynep

    2012-01-01

    Selenoprotein W (SEPW1) is a ubiquitous, highly conserved thioredoxin-like protein whose depletion causes a transient p53- and p21Cip1-dependent G1-phase cell cycle arrest in breast and prostate epithelial cells. SEPW1 depletion increases phosphorylation of Ser-33 in p53, which is associated with decreased p53 ubiquitination and stabilization of p53. We report here that delayed cell cycle progression, Ser-33 phosphorylation, and p53 nuclear accumulation from SEPW1 depletion require mitogen-activated protein kinase kinase 4 (MKK4). Silencing MKK4 rescued G1 arrest, Ser-33 phosphorylation, and nuclear accumulation of p53 induced by SEPW1 depletion, but silencing MKK3, MKK6, or MKK7 did not. SEPW1 silencing did not change the phosphorylation state of MKK4 but increased total MKK4 protein. Silencing p38γ, p38δ, or JNK2 partially rescued G1 arrest from SEPW1 silencing, suggesting they signal downstream from MKK4. These results imply that SEPW1 silencing increases MKK4, which activates p38γ, p38δ, and JNK2 to phosphorylate p53 on Ser-33 and cause a transient G1 arrest. PMID:22730327

  18. Potassium channels in cell cycle and cell proliferation

    PubMed Central

    Urrego, Diana; Tomczak, Adam P.; Zahed, Farrah; Stühmer, Walter; Pardo, Luis A.

    2014-01-01

    Normal cell-cycle progression is a crucial task for every multicellular organism, as it determines body size and shape, tissue renewal and senescence, and is also crucial for reproduction. On the other hand, dysregulation of the cell-cycle progression leading to uncontrolled cell proliferation is the hallmark of cancer. Therefore, it is not surprising that it is a tightly regulated process, with multifaceted and very complex control mechanisms. It is now well established that one of those mechanisms relies on ion channels, and in many cases specifically on potassium channels. Here, we summarize the possible mechanisms underlying the importance of potassium channels in cell-cycle control and briefly review some of the identified channels that illustrate the multiple ways in which this group of proteins can influence cell proliferation and modulate cell-cycle progression. PMID:24493742

  19. Chemical dissection of the cell cycle: probes for cell biology and anti-cancer drug development

    PubMed Central

    Senese, S; Lo, Y C; Huang, D; Zangle, T A; Gholkar, A A; Robert, L; Homet, B; Ribas, A; Summers, M K; Teitell, M A; Damoiseaux, R; Torres, J Z

    2014-01-01

    Cancer cell proliferation relies on the ability of cancer cells to grow, transition through the cell cycle, and divide. To identify novel chemical probes for dissecting the mechanisms governing cell cycle progression and cell division, and for developing new anti-cancer therapeutics, we developed and performed a novel cancer cell-based high-throughput chemical screen for cell cycle modulators. This approach identified novel G1, S, G2, and M-phase specific inhibitors with drug-like properties and diverse chemotypes likely targeting a broad array of processes. We further characterized the M-phase inhibitors and highlight the most potent M-phase inhibitor MI-181, which targets tubulin, inhibits tubulin polymerization, activates the spindle assembly checkpoint, arrests cells in mitosis, and triggers a fast apoptotic cell death. Importantly, MI-181 has broad anti-cancer activity, especially against BRAFV600E melanomas. PMID:25321469

  20. SAFT nickel hydrogen cell cycling status

    NASA Technical Reports Server (NTRS)

    Borthomieu, Yannick; Duquesne, Didier

    1994-01-01

    An overview of the NiH2 cell development is given. The NiH2 SAFT system is an electrochemical (single or dual) stack (IPV). The stack is mounted in an hydroformed Inconel 718 vessel operating at high pressure, equipped with 'rabbit ears' ceramic brazed electrical feedthroughs. The cell design is described: positive electrode, negative electrode, and stack configuration. Overviews of low earth orbit and geostationary earth orbit cyclings are provided. DPA results are also provided. The cycling and DPA results demonstrate that SAFT NiH2 is characterized by high reliability and very stable performances.

  1. How active was solar cycle 22?

    NASA Technical Reports Server (NTRS)

    Hoegy, W. R.; Pesnell, W. D.; Woods, T. N.; Rottman, G. J.

    1993-01-01

    Solar EUV observations from the Langmuir probe on Pioneer Venus Orbiter suggest that at EUV wavelengths solar cycle 22 was more active than solar cycle 21. The Langmuir probe, acting as a photodiode, measured the integrated solar EUV flux over a 13 1/2 year period from January 1979 to June 1992, the longest continuous solar EUV measurement. The Ipe EUV flux correlated very well with the SME measurement of L-alpha during the lifetime of SME and with the UARS SOLSTICE L-alpha from October 1991 to June 1992 when the Ipe measurement ceased. Starting with the peak of solar cycle 21, there was good general agreement of Ipe EUV with the 10.7 cm, Ca K, and He 10830 solar indices, until the onset of solar cycle 22. From 1989 to the start of 1992, the 10.7 cm flux exhibited a broad maximum consisting of two peaks of nearly equal magnitude, whereas Ipe EUV exhibited a strong increase during this time period making the second peak significantly higher than the first. The only solar index that exhibits the same increase in solar activity as Ipe EUV and L-alpha during the cycle 22 peak is the total magnetic flux. The case for high activity during this peak is also supported by the presence of very high solar flare intensity.

  2. Creatine kinase in cell cycle regulation and cancer.

    PubMed

    Yan, Yong-Bin

    2016-08-01

    The phosphocreatine-creatine kinase (CK) shuttle system is increasingly recognized as a fundamental mechanism for ATP homeostasis in both excitable and non-excitable cells. Many intracellular processes are ATP dependent. Cell division is a process requiring a rapid rate of energy turnover. Cell cycle regulation is also a key point to understanding the mechanisms underlying cancer progression. It has been known for about 40 years that aberrant CK levels are associated with various cancers and for over 30 years that CK is involved in mitosis regulation. However, the underlying molecular mechanisms have not been investigated sufficiently until recently. By maintaining ATP at sites of high-energy demand, CK can regulate cell cycle progression by affecting the intracellular energy status as well as by influencing signaling pathways that are essential to activate cell division and cytoskeleton reorganization. Aberrant CK levels may impair cell viability under normal or stressed conditions and induce cell death. The involvement of CK in cell cycle regulation and cellular energy metabolism makes it a potential diagnostic biomarker and therapeutic target in cancer. To understand the multiple physiological/pathological functions of CK, it is necessary to identify CK-binding partners and regulators including proteins, non-coding RNAs and participating endogenous small molecular weight chemical compounds. This review will focus on molecular mechanisms of CK in cell cycle regulation and cancer progression. It will also discuss the implications of recent mechanistic studies, the emerging problems and future challenges of the multifunctional enzyme CK. PMID:27020776

  3. Inhibition of DNA Methylation Alters Chromatin Organization, Nuclear Positioning and Activity of 45S rDNA Loci in Cycling Cells of Q. robur

    PubMed Central

    Horvat, Tomislav; Maglica, Željka; Vojta, Aleksandar; Zoldoš, Vlatka

    2014-01-01

    Around 2200 copies of genes encoding ribosomal RNA (rRNA) in pedunculate oak, Quercus robur, are organized into two rDNA loci, the major (NOR-1) and the minor (NOR-2) locus. We present the first cytogenetic evidence indicating that the NOR-1 represents the active nucleolar organizer responsible for rRNA synthesis, while the NOR-2 probably stays transcriptionally silent and does not participate in the formation of the nucleolus in Q. robur, which is a situation resembling the well-known phenomenon of nucleolar dominance. rDNA chromatin topology analyses in cycling root tip cells by light and electron microscopy revealed the minor locus to be highly condensed and located away from the nucleolus, while the major locus was consistently associated with the nucleolus and often exhibited different levels of condensation. In addition, silver precipitation was confined exclusively to the NOR-1 locus. Also, NOR-2 was highly methylated at cytosines and rDNA chromatin was marked with histone modifications characteristic for repressive state. After treatment of the root cells with the methylation inhibitor 5-aza-2′-deoxycytidine, we observed an increase in the total level of rRNA transcripts and a decrease in DNA methylation level at the NOR-2 locus. Also, NOR-2 sites relocalized with respect to the nuclear periphery/nucleolus, however, the relocation did not affect the contribution of this locus to nucleolar formation, nor did it affect rDNA chromatin decondensation, strongly suggesting that NOR-2 has lost the function of rRNA synthesis and nucleolar organization. PMID:25093501

  4. Inhibition of DNA methylation alters chromatin organization, nuclear positioning and activity of 45S rDNA loci in cycling cells of Q. robur.

    PubMed

    Bočkor, Vedrana Vičić; Barišić, Darko; Horvat, Tomislav; Maglica, Željka; Vojta, Aleksandar; Zoldoš, Vlatka

    2014-01-01

    Around 2200 copies of genes encoding ribosomal RNA (rRNA) in pedunculate oak, Quercus robur, are organized into two rDNA loci, the major (NOR-1) and the minor (NOR-2) locus. We present the first cytogenetic evidence indicating that the NOR-1 represents the active nucleolar organizer responsible for rRNA synthesis, while the NOR-2 probably stays transcriptionally silent and does not participate in the formation of the nucleolus in Q. robur, which is a situation resembling the well-known phenomenon of nucleolar dominance. rDNA chromatin topology analyses in cycling root tip cells by light and electron microscopy revealed the minor locus to be highly condensed and located away from the nucleolus, while the major locus was consistently associated with the nucleolus and often exhibited different levels of condensation. In addition, silver precipitation was confined exclusively to the NOR-1 locus. Also, NOR-2 was highly methylated at cytosines and rDNA chromatin was marked with histone modifications characteristic for repressive state. After treatment of the root cells with the methylation inhibitor 5-aza-2'-deoxycytidine, we observed an increase in the total level of rRNA transcripts and a decrease in DNA methylation level at the NOR-2 locus. Also, NOR-2 sites relocalized with respect to the nuclear periphery/nucleolus, however, the relocation did not affect the contribution of this locus to nucleolar formation, nor did it affect rDNA chromatin decondensation, strongly suggesting that NOR-2 has lost the function of rRNA synthesis and nucleolar organization. PMID:25093501

  5. Classification of a frameshift/extended and a stop mutation in WT1 as gain-of-function mutations that activate cell cycle genes and promote Wilms tumour cell proliferation

    PubMed Central

    Busch, Maike; Schwindt, Heinrich; Brandt, Artur; Beier, Manfred; Görldt, Nicole; Romaniuk, Paul; Toska, Eneda; Roberts, Stefan; Royer, Hans-Dieter; Royer-Pokora, Brigitte

    2014-01-01

    The WT1 gene encodes a zinc finger transcription factor important for normal kidney development. WT1 is a suppressor for Wilms tumour development and an oncogene for diverse malignant tumours. We recently established cell lines from primary Wilms tumours with different WT1 mutations. To investigate the function of mutant WT1 proteins, we performed WT1 knockdown experiments in cell lines with a frameshift/extension (p.V432fsX87 = Wilms3) and a stop mutation (p.P362X = Wilms2) of WT1, followed by genome-wide gene expression analysis. We also expressed wild-type and mutant WT1 proteins in human mesenchymal stem cells and established gene expression profiles. A detailed analysis of gene expression data enabled us to classify the WT1 mutations as gain-of-function mutations. The mutant WT1Wilms2 and WT1Wilms3 proteins acquired an ability to modulate the expression of a highly significant number of genes from the G2/M phase of the cell cycle, and WT1 knockdown experiments showed that they are required for Wilms tumour cell proliferation. p53 negatively regulates the activity of a large number of these genes that are also part of a core proliferation cluster in diverse human cancers. Our data strongly suggest that mutant WT1 proteins facilitate expression of these cell cycle genes by antagonizing transcriptional repression mediated by p53. We show that mutant WT1 can physically interact with p53. Together the findings show for the first time that mutant WT1 proteins have a gain-of-function and act as oncogenes for Wilms tumour development by regulating Wilms tumour cell proliferation. PMID:24619359

  6. Control points within the cell cycle

    SciTech Connect

    Van't Hof, J.

    1984-01-01

    Evidence of the temporal order of chromosomal DNA replication argues favorably for the view that the cell cycle is controlled by genes acting in sequence whose time of expression is determined by mitosis and the amount of nuclear DNA (2C vs 4C) in the cell. Gl and G2 appear to be carbohydrate dependent in that cells starved of either carbohydrate of phosphate fail to make these transitions. Cells deprived of nitrate, however, fail only at Gl to S transition indicating that the controls that operate in G1 differ from those that operate in G2. 46 references, 5 figures.

  7. Anti-hepatocellular carcinoma activity using human HepG2 cells and hepatotoxicity of 6-substituted methyl 3-aminothieno[3,2-b]pyridine-2-carboxylate derivatives: in vitro evaluation, cell cycle analysis and QSAR studies.

    PubMed

    Abreu, Rui M V; Ferreira, Isabel C F R; Calhelha, Ricardo C; Lima, Raquel T; Vasconcelos, M Helena; Adega, Filomena; Chaves, Raquel; Queiroz, Maria-João R P

    2011-12-01

    Hepatocellular carcinoma (HCC) is a highly complex cancer, resistant to commonly used treatments and new therapeutic agents are urgently needed. A total of thirty-two thieno[3,2-b]pyridine derivatives of two series: methyl 3-amino-6-(hetero)arylthieno[3,2-b]pyridine-2-carboxylates (1a-1t) and methyl 3-amino-6-[(hetero)arylethynyl]thieno[3,2-b]pyridine-2-carboxylates (2a-2n), previously prepared by some of us, were evaluated as new potential anti-HCC agents by studying their in vitro cell growth inhibition on human HepG2 cells and hepatotoxicity using a porcine liver primary cell culture (PLP1). The presence of amino groups linked to a benzene moiety emerges as the key element for the anti-HCC activity. The methyl 3-amino-6-[(3-aminophenyl)ethynyl]thieno[3,2-b]pyridine-2-carboxylate (2f) is the most potent compound presenting GI(50) values on HepG2 cells of 1.2 μM compared to 2.9 μM of the positive control ellipticine, with no observed hepatotoxicity (PLP1 GI(50) > 125 μM against 3.3 μM of ellipticine). Moreover this compound changes the cell cycle profile of the HepG2 cells, causing a decrease in the % of cells in the S phase and a cell cycle arrest in the G2/M phase. QSAR studies were also performed and the correlations obtained using molecular and 1D descriptors revealed the importance of the presence of amino groups and hydrogen bond donors for anti-HCC activity, and hydrogen bond acceptors for hepatotoxicity. The best correlations were obtained with 3D descriptors belonging to different subcategories for anti-HCC activity and hepatotoxicity, respectively. These results point to different molecular mechanisms of action of the compounds in anti-HCC activity and hepatotoxicity. This work presents some promising thieno[3,2-b]pyridine derivatives for potential use in the therapy of HCC. These compounds can also be used as scaffolds for further synthesis of more potent analogs. PMID:22014996

  8. Esculetin, a natural coumarin compound, evokes Ca(2+) movement and activation of Ca(2+)-associated mitochondrial apoptotic pathways that involved cell cycle arrest in ZR-75-1 human breast cancer cells.

    PubMed

    Chang, Hong-Tai; Chou, Chiang-Ting; Lin, You-Sheng; Shieh, Pochuen; Kuo, Daih-Huang; Jan, Chung-Ren; Liang, Wei-Zhe

    2016-04-01

    Esculetin (6,7-dihydroxycoumarin), a derivative of coumarin compound, is found in traditional medicinal herbs. It has been shown that esculetin triggers diverse cellular signal transduction pathways leading to regulation of physiology in different models. However, whether esculetin affects Ca(2+) homeostasis in breast cancer cells has not been explored. This study examined the underlying mechanism of cytotoxicity induced by esculetin and established the relationship between Ca(2+) signaling and cytotoxicity in human breast cancer cells. The results showed that esculetin induced concentration-dependent rises in the intracellular Ca(2+) concentration ([Ca(2+)]i) in ZR-75-1 (but not in MCF-7 and MDA-MB-231) human breast cancer cells. In ZR-75-1 cells, this Ca(2+) signal response was reduced by removing extracellular Ca(2+) and was inhibited by the store-operated Ca(2+) channel blocker 2-aminoethoxydiphenyl borate (2-APB). In Ca(2+)-free medium, pre-treatment with the endoplasmic reticulum Ca(2+) pump inhibitor thapsigargin (TG) abolished esculetin-induced [Ca(2+)]i rises. Conversely, incubation with esculetin abolished TG-induced [Ca(2+)]i rises. Esculetin induced cytotoxicity that involved apoptosis, as supported by the reduction of mitochondrial membrane potential and the release of cytochrome c and the proteolytic activation of caspase-9/caspase-3, which were partially reversed by pre-chelating cytosolic Ca(2+) with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid-acetoxymethyl ester (BAPTA-AM). Moreover, esculetin increased the percentage of cells in G2/M phase and regulated the expressions of p53, p21, CDK1, and cyclin B1. Together, in ZR-75-1 cells, esculetin induced [Ca(2+)]i rises by releasing Ca(2+) from the ER and causing Ca(2+) influx through 2-APB-sensitive store-operated Ca(2+) entry. Furthermore, esculetin activated Ca(2+)-associated mitochondrial apoptotic pathways that involved G2/M cell cycle arrest. Graphical abstract The summary of esculetin

  9. Cytological cycles and fates in Psidium myrtoides are altered towards new cell metabolism and functionalities by the galling activity of Nothotrioza myrtoidis.

    PubMed

    Carneiro, R G S; Isaias, R M S

    2015-03-01

    The morphogenesis of galls occurs by the redifferentiation of cells that assume new functions in the modified host plant organs. The redifferentiated cells in the galls of Nothotrioza myrtoidis on Psidium myrtoides have low complexity metabolism and are photosynthesis-deficient. These galls were studied in search for evidences of the establishment of new cell cycles and fates and cytological gradients that corroborate their metabolic profile. Young and mature leaves of P. myrtoides and leaf galls induced by N. myrtoidis at different developmental stages were collected along 24 months and analyzed under light and transmission electron microscopy. The leaves of P. myrtoides are long-lasting and did not senesce within the analyzed period, while the galls have a shorter cycle, and senesce within 1 year. A homogenous parenchyma is established by a "standby-redifferentiation" of the chlorophyllous tissues, and sclerenchyma cells redifferentiate from parenchyma cells in the outer cortex of the mature galls. The lack of organelles, the underdeveloped lamellation of chloroplasts, and the occurrence of few plastoglobules are related to the photosynthetic deficiency of the galls. No cytological gradients were observed, but the organelle-rich cells of the vascular and perivascular parenchymas are similar to those of the nutritive tissues of galls induced by other insect taxa. These cells nearest to the feeding sites of N. myrtoidis present higher metabolism and well-developed apparatus for the prevention of oxidative stress. The features herein described corroborate the low metabolic profile of the galls as the cell cycles and fates of P. myrtoides are manipulated for completely new functionalities. PMID:25272990

  10. Mitochondrial dynamics and the cell cycle

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Nuclear-mitochondrial (NM) communication impacts many aspects of plant development including vigor, sterility and viability. Dynamic changes in mitochondrial number, shape, size, and cellular location takes place during the cell cycle possibly impacting the process itself and leading to distribution...

  11. Cell Cycle Progression of Human Cells Cultured in Rotating Bioreactor

    NASA Technical Reports Server (NTRS)

    Parks, Kelsey

    2009-01-01

    Space flight has been shown to alter the astronauts immune systems. Because immune performance is complex and reflects the influence of multiple organ systems within the host, scientists sought to understand the potential impact of microgravity alone on the cellular mechanisms critical to immunity. Lymphocytes and their differentiated immature form, lymphoblasts, play an important and integral role in the body's defense system. T cells, one of the three major types of lymphocytes, play a central role in cell-mediated immunity. They can be distinguished from other lymphocyte types, such as B cells and natural killer cells by the presence of a special receptor on their cell surface called T cell receptors. Reported studies have shown that spaceflight can affect the expression of cell surface markers. Cell surface markers play an important role in the ability of cells to interact and to pass signals between different cells of the same phenotype and cells of different phenotypes. Recent evidence suggests that cell-cycle regulators are essential for T-cell function. To trigger an effective immune response, lymphocytes must proliferate. The objective of this project is to investigate the changes in growth of human cells cultured in rotating bioreactors and to measure the growth rate and the cell cycle distribution for different human cell types. Human lymphocytes and lymphoblasts will be cultured in a bioreactor to simulate aspects of microgravity. The bioreactor is a cylindrical culture vessel that incorporates the aspects of clinostatic rotation of a solid fluid body around a horizontal axis at a constant speed, and compensates gravity by rotation and places cells within the fluid body into a sustained free-fall. Cell cycle progression and cell proliferation of the lymphocytes will be measured for a number of days. In addition, RNA from the cells will be isolated for expression of genes related in cell cycle regulations.

  12. Identification of a novel EGF-sensitive cell cycle checkpoint

    SciTech Connect

    Walker, Francesca . E-mail: francesca.walker@ludwig.edu.au; Zhang Huihua; Burgess, Antony W.

    2007-02-01

    The site of action of growth factors on mammalian cell cycle has been assigned to the boundary between the G1 and S phases. We show here that Epidermal Growth Factor (EGF) is also required for mitosis. BaF/3 cells expressing the EGFR (BaF/wtEGFR) synthesize DNA in response to EGF, but arrest in S-phase. We have generated a cell line (BaF/ERX) with defective downregulation of the EGFR and sustained activation of EGFR signalling pathways: these cells undergo mitosis in an EGF-dependent manner. The transit of BaF/ERX cells through G2/M strictly requires activation of EGFR and is abolished by AG1478. This phenotype is mimicked by co-expression of ErbB2 in BaF/wtEGFR cells, and abolished by inhibition of the EGFR kinase, suggesting that sustained signalling of the EGFR, through impaired downregulation of the EGFR or heterodimerization, is required for completion of the cycle. We have confirmed the role of EGFR signalling in the G2/M phase of the cell cycle using a human tumor cell line which overexpresses the EGFR and is dependent on EGFR signalling for growth. These findings unmask an EGF-sensitive checkpoint, helping to understand the link between sustained EGFR signalling, proliferation and the acquisition of a radioresistant phenotype in cancer cells.

  13. Polyamines and the Cell Cycle of Catharanthus roseus Cells in Culture 1

    PubMed Central

    Maki, Hisae; Ando, Satoshi; Kodama, Hiroaki; Komamine, Atsushi

    1991-01-01

    Investigation was made on the effect of partial depletion of polyamines (PAs), induced by treatment with inhibitors of the biosynthesis of PAs, on the distribution of cells at each phase of the cell cycle in Catharanthus roseus (L.) G. Don. cells in suspension cultures, using flow cytometry. More cells treated with inhibitors of arginine decarboxylase (ADC) and ornithine decarboxylase (ODC) were accumulated in the G1 phase than those in the control, while the treatment with an inhibitor of spermidine (SPD) synthase showed no effect on the distribution of cells. The endogenous levels of the PAs, putrescine (PUT), SPD, and spermine (SPM), were determined during the cell cycle in synchronous cultures of C. roseus. Two peaks of endogenous level of PAs, in particular, of PUT and SPD, were observed during the cell cycle. Levels of PAs increased markedly prior to synthesis of DNA in the S phase and prior to cytokinesis. Activities of ADC and ODC were also assayed during the cell cycle. Activities of ADC was much higher than that of ODC throughout the cell cycle, but both activities of ODC and ADC changed in concert with changes in levels of PAs. Therefore, it is suggested that these enzymes may regulate PA levels during the cell cycle. These results indicate that inhibitors of PUT biosynthesis caused the suppression of cell proliferation by prevention of the progression of the cell cycle, probably from the G1 to the S phase, and PUT may play more important roles in the progression of the cell cycle than other PAs. PMID:16668290

  14. The Effect of Spaceflight on Cartilage Cell Cycle and Differentiation

    NASA Technical Reports Server (NTRS)

    Doty, Stephen B.; Stiner, Dalina; Telford, William G.

    2000-01-01

    In vivo studies have shown that spaceflight results in loss of bone and muscle. In an effort to understand the mechanisms of these changes, cell cultures of cartilage, bone and muscle have been subjected to spaceflight to study the microgravity effects on differentiated cells. However it now seems possible that the cell differentiation process itself may be the event(s) most affected by spaceflight. For example, osteoblast-like cells have been shown to have reduced cellular activity in microgravity due to an underdifferentiated state (Carmeliet, et al, 1997). And reduced human lymphocyte growth in spaceflight was related to increased apoptosis (Lewis, et al, 1998). Which brings us to the question of whether reduced cellular activity in space is due to an effect on the differentiated cell, an effect on the cell cycle and cell proliferation, or an effect on cell death. This question has not been specifically addressed on previous flights and was the question behind die present study.

  15. SAMHD1 controls cell cycle status, apoptosis and HIV-1 infection in monocytic THP-1 cells.

    PubMed

    Bonifati, Serena; Daly, Michele B; St Gelais, Corine; Kim, Sun Hee; Hollenbaugh, Joseph A; Shepard, Caitlin; Kennedy, Edward M; Kim, Dong-Hyun; Schinazi, Raymond F; Kim, Baek; Wu, Li

    2016-08-01

    SAMHD1 limits HIV-1 infection in non-dividing myeloid cells by decreasing intracellular dNTP pools. HIV-1 restriction by SAMHD1 in these cells likely prevents activation of antiviral immune responses and modulates viral pathogenesis, thus highlighting a critical role of SAMHD1 in HIV-1 physiopathology. Here, we explored the function of SAMHD1 in regulating cell proliferation, cell cycle progression and apoptosis in monocytic THP-1 cells. Using the CRISPR/Cas9 technology, we generated THP-1 cells with stable SAMHD1 knockout. We found that silencing of SAMHD1 in cycling cells stimulates cell proliferation, redistributes cell cycle population in the G1/G0 phase and reduces apoptosis. These alterations correlated with increased dNTP levels and more efficient HIV-1 infection in dividing SAMHD1 knockout cells relative to control. Our results suggest that SAMHD1, through its dNTPase activity, affects cell proliferation, cell cycle distribution and apoptosis, and emphasize a key role of SAMHD1 in the interplay between cell cycle regulation and HIV-1 infection. PMID:27183329

  16. Induction of selective cytotoxicity and apoptosis in human T4-lymphoblastoid cell line (CEMss) by boesenbergin a isolated from boesenbergia rotunda rhizomes involves mitochondrial pathway, activation of caspase 3 and G2/M phase cell cycle arrest

    PubMed Central

    2013-01-01

    Background Boesenbergia rotunda (Roxb.) Schlecht (family zingiberaceae) is a rhizomatous herb that is distributed from north-eastern India to south-east Asia, especially in Indonesia, Thailand and Malaysia. Previous research has shown that the crude extract of this plant has cytotoxic properties. The current study examines the cytotoxic properties of boesenbergin A isolated from Boesenbergia rotunda. Methods MTT assay was used to check the cytotoxicity of boesenbergin A. The morphological assessment of apoptosis was monitored using normal and fluorescence microscopy. The early and late phase of apoptosis was investigated using annexin V and DNA laddering assays, respectively. The mitochondrial membrane potential (MMP) was assessed by fluorescence microscopy. Human apoptosis proteome profiler assays were performed to investigate the mechanism of cell death. In addition, the protein levels of Bax, Bcl2 and HSP 70 were also analyzed using western blot. Assays of caspase =-3/7, -8 and =-9 were carried out in order to test for induction during treatment. Lastly, cell cycle progression was analyzed using flow cytometry. Results Boesenbergin A was found to have the highest toxicity towards CEMss cancer cells (IC50 = 8 μg/ml). The morphology of CEMss cells after treatment showed evidence of apoptosis that included blebbing and chromatin condensation. The annexin V assay revealed that early apoptosis is induced after treatment. The DNA laddering assay confirmed that DNA fragmentation had occurred during late apoptosis. The cell cycle analysis indicated that boesenbergin A was able to induce G2/M phase arrest in CEMss cells. The activity of caspases -3/7, -8 and -9 was increased after treatment which indicates both intrinsic and extrinsic pathways are induced during apoptosis. The involvement of mitochondria was established by increased mitochondrial membrane potential and up and down regulation of Bcl2 and Bax proteins as well as HSP70. Conclusion In conclusion, the

  17. Carnosol, a dietary diterpene, displays growth inhibitory effects in human prostate cancer PC3 cells leading to G2-phase cell cycle arrest and targets the 5'-AMP-activated protein kinase (AMPK) pathway

    PubMed Central

    Johnson, Jeremy J.; Syed, Deeba N.; Heren, Chenelle R.; Suh, Yewseok; Adhami, Vaqar M.; Mukhtar, Hasan

    2010-01-01

    Purpose The anti-cancer effect of carnosol was investigated in human prostate cancer PC3 cells. Methods Biochemical analysis and protein array data of carnosol treated PC3 cells were analyzed. Results We evaluated carnosol for its potential anti-cancer properties in the PC3 cells. Using an MTT assay we found that carnosol (10 – 70 µM) decreases cell viability in a time and dose dependent manner. Next, we evaluated the effect of carnosol (20–60 uM) effect using flow cytometry as well as biochemical analysis and found induction of G2-phase cell cycle arrest. To establish a more precise mechanism, we performed a protein array that evaluated 638 proteins involved in cell signaling pathways. The protein array identified 5'-AMP-activated protein kinase (AMPK), a serine/threonine protein kinase involved in the regulation of cellular energy balance as a potential target. Further downstream effects consistent with cancer inhibition included the modulation of the mTOR/HSP70S6k/4E-BP1 pathway. Additionally, we found that carnosol targeted the PI3K/Akt pathway in a dose dependent manner. Conclusions These results suggest that carnosol targets multiple signaling pathways that include the AMPK pathway. The ability of carnosol to inhibit prostate cancer in vitro suggests carnosol may be a novel agent for the management of PCa. PMID:18286356

  18. POSSIBLE CHROMOSPHERIC ACTIVITY CYCLES IN AD LEO

    SciTech Connect

    Buccino, Andrea P.; Petrucci, Romina; Mauas, Pablo J. D.; Jofré, Emiliano

    2014-01-20

    AD Leo (GJ 388) is an active dM3 flare star that has been extensively observed both in the quiescent and flaring states. Since this active star is near the fully convective boundary, studying its long-term chromospheric activity in detail could be an appreciable contribution to dynamo theory. Here, using the Lomb-Scargle periodogram, we analyze the Ca II K line-core fluxes derived from CASLEO spectra obtained between 2001 and 2013 and the V magnitude from the ASAS database between 2004 and 2010. From both of these totally independent time series, we obtain a possible activity cycle with a period of approximately seven years and a less significant shorter cycle of approximately two years. A tentative interpretation is that a dynamo operating near the surface could be generating the longer cycle, while a second dynamo operating in the deep convection zone could be responsible for the shorter one. Based on the long duration of our observing program at CASLEO and the fact that we observe different spectral features simultaneously, we also analyze the relation between simultaneous measurements of the Na I index (R{sub D}{sup ′}), Hα, and Ca II K fluxes at different activity levels of AD Leo, including flares.

  19. Cell cycle nucleic acids, polypeptides and uses thereof

    DOEpatents

    Gordon-Kamm, William J.; Lowe, Keith S.; Larkins, Brian A.; Dilkes, Brian R.; Sun, Yuejin

    2007-08-14

    The invention provides isolated nucleic acids and their encoded proteins that are involved in cell cycle regulation. The invention further provides recombinant expression cassettes, host cells, transgenic plants, and antibody compositions. The present invention provides methods and compositions relating to altering cell cycle protein content, cell cycle progression, cell number and/or composition of plants.

  20. Indirect-fired gas turbine dual fuel cell power cycle

    SciTech Connect

    Micheli, P.L.; Williams, M.C.; Sudhoff, F.A.

    1998-04-01

    The present invention relates generally to an integrated fuel cell power plant, and more specifically to a combination of cycles wherein a first fuel cell cycle tops an indirect-fired gas turbine cycle and a second fuel cell cycle bottoms the gas turbine cycle so that the cycles are thermally integrated in a tandem operating arrangement. The United States Government has rights in this invention pursuant to the employer-employee relationship between the United States Department of Energy and the inventors.

  1. A role for homologous recombination proteins in cell cycle regulation

    PubMed Central

    Kostyrko, Kaja; Bosshard, Sandra; Urban, Zuzanna; Mermod, Nicolas

    2015-01-01

    Eukaryotic cells respond to DNA breaks, especially double-stranded breaks (DSBs), by activating the DNA damage response (DDR), which encompasses DNA repair and cell cycle checkpoint signaling. The DNA damage signal is transmitted to the checkpoint machinery by a network of specialized DNA damage-recognizing and signal-transducing molecules. However, recent evidence suggests that DNA repair proteins themselves may also directly contribute to the checkpoint control. Here, we investigated the role of homologous recombination (HR) proteins in normal cell cycle regulation in the absence of exogenous DNA damage. For this purpose, we used Chinese Hamster Ovary (CHO) cells expressing the Fluorescent ubiquitination-based cell cycle indicators (Fucci). Systematic siRNA-mediated knockdown of HR genes in these cells demonstrated that the lack of several of these factors alters cell cycle distribution, albeit differentially. The knock-down of MDC1, Rad51 and Brca1 caused the cells to arrest in the G2 phase, suggesting that they may be required for the G2/M transition. In contrast, inhibition of the other HR factors, including several Rad51 paralogs and Rad50, led to the arrest in the G1/G0 phase. Moreover, reduced expression of Rad51B, Rad51C, CtIP and Rad50 induced entry into a quiescent G0-like phase. In conclusion, the lack of many HR factors may lead to cell cycle checkpoint activation, even in the absence of exogenous DNA damage, indicating that these proteins may play an essential role both in DNA repair and checkpoint signaling. PMID:26125600

  2. FUEL CELL/MICRO-TURBINE COMBINED CYCLE

    SciTech Connect

    Larry J. Chaney; Mike R. Tharp; Tom W. Wolf; Tim A. Fuller; Joe J. Hartvigson

    1999-12-01

    A wide variety of conceptual design studies have been conducted that describe ultra-high efficiency fossil power plant cycles. The most promising of these ultra-high efficiency cycles incorporate high temperature fuel cells with a gas turbine. Combining fuel cells with a gas turbine increases overall cycle efficiency while reducing per kilowatt emissions. This study has demonstrated that the unique approach taken to combining a fuel cell and gas turbine has both technical and economic merit. The approach used in this study eliminates most of the gas turbine integration problems associated with hybrid fuel cell turbine systems. By using a micro-turbine, and a non-pressurized fuel cell the total system size (kW) and complexity has been reduced substantially from those presented in other studies, while maintaining over 70% efficiency. The reduced system size can be particularly attractive in the deregulated electrical generation/distribution environment where the market may not demand multi-megawatt central stations systems. The small size also opens up the niche markets to this high efficiency, low emission electrical generation option.

  3. Cell shape dynamics during the staphylococcal cell cycle

    PubMed Central

    Monteiro, João M.; Fernandes, Pedro B.; Vaz, Filipa; Pereira, Ana R.; Tavares, Andreia C.; Ferreira, Maria T.; Pereira, Pedro M.; Veiga, Helena; Kuru, Erkin; VanNieuwenhze, Michael S.; Brun, Yves V.; Filipe, Sérgio R.; Pinho, Mariana G.

    2015-01-01

    Staphylococcus aureus is an aggressive pathogen and a model organism to study cell division in sequential orthogonal planes in spherical bacteria. However, the small size of staphylococcal cells has impaired analysis of changes in morphology during the cell cycle. Here we use super-resolution microscopy and determine that S. aureus cells are not spherical throughout the cell cycle, but elongate during specific time windows, through peptidoglycan synthesis and remodelling. Both peptidoglycan hydrolysis and turgor pressure are required during division for reshaping the flat division septum into a curved surface. In this process, the septum generates less than one hemisphere of each daughter cell, a trait we show is common to other cocci. Therefore, cell surface scars of previous divisions do not divide the cells in quadrants, generating asymmetry in the daughter cells. Our results introduce a need to reassess the models for division plane selection in cocci. PMID:26278781

  4. Solid oxide fuel cell combined cycles

    SciTech Connect

    Bevc, F.P.; Lundberg, W.L.; Bachovchin, D.M.

    1996-12-31

    The integration of the solid oxide fuel cell and combustion turbine technologies can result in combined-cycle power plants, fueled with natural gas, that have high efficiencies and clean gaseous emissions. Results of a study are presented in which conceptual designs were developed for 3 power plants based upon such an integration, and ranging in rating from 3 to 10 MW net ac. The plant cycles are described and characteristics of key components summarized. Also, plant design-point efficiency estimates are presented as well as values of other plant performance parameters.

  5. Modeling of SONOS Memory Cell Erase Cycle

    NASA Technical Reports Server (NTRS)

    Phillips, Thomas A.; MacLeod, Todd C.; Ho, Fat H.

    2011-01-01

    Utilization of Silicon-Oxide-Nitride-Oxide-Silicon (SONOS) nonvolatile semiconductor memories as a flash memory has many advantages. These electrically erasable programmable read-only memories (EEPROMs) utilize low programming voltages, have a high erase/write cycle lifetime, are radiation hardened, and are compatible with high-density scaled CMOS for low power, portable electronics. In this paper, the SONOS memory cell erase cycle was investigated using a nonquasi-static (NQS) MOSFET model. Comparisons were made between the model predictions and experimental data.

  6. Polyphenol-rich extract of Salvia chinensis exhibits anticancer activity in different cancer cell lines, and induces cell cycle arrest at the G0/G1-phase, apoptosis and loss of mitochondrial membrane potential in pancreatic cancer cells

    PubMed Central

    ZHAO, QUAN; HUO, XUE-CHEN; SUN, FU-DONG; DONG, RUI-QIAN

    2015-01-01

    Pancreatic cancer (PC) is one of the most aggressive types of human malignancy, which has an overall 5-year survival rate of <2%. PC is the fourth most common cause of cancer-associated mortality in the western world. At present, there is almost no effective treatment available for the treatment of PC. The aim of the present study was to evaluate the anticancer potential of a polyphenol enriched extract obtained from Salvia chinensis, a Chinese medicinal plant. An MTT assay was used to evaluate the cell viability of five cancer cell lines and one normal cell line. In addition, the effects of the extract on apoptotic induction, cell cycle phase distribution, DNA damage and loss of mitochondrial membrane potential (ΛΨm) were evaluated in MiapaCa-2 human PC cells. The effects of the extract on cell cycle phase distribution and ΛΨm were assessed by flow cytometry, using propidium iodide and rhodamine-123 DNA-binding fluorescent dyes, respectively. Fluorescence microscopy, using 4′,6-diamidino-2-phenylindole as a staining agent, was performed in order to detect the morphological changes of the MiapaCa-2 cancer cells and the presence of apoptotic bodies following treatment with the extract. The results of the present study demonstrated that the polyphenol-rich extract from S. chinensis induced potent cytotoxicity in the MCF-7 human breast cancer cells, A549 human lung cancer cells, HCT-116 and COLO 205 human colon cancer cells, and MiapaCa-2 human PC cells. The COLO 205 and MCF-7 cancer cell lines were the most susceptible to treatment with the extract, which exhibited increased rate of growth inhibition. Fluorescence microscopy revealed characteristic morphological features of apoptosis and detected the appearance of apoptotic bodies following treatment with the extract in the PC cells. Flow cytometric analysis demonstrated that the extract induced G0/G1 cell cycle arrest in a dose-dependent manner. In addition, treatment with the extract induced a significant and

  7. The Heliosphere Through the Solar Activity Cycle

    NASA Technical Reports Server (NTRS)

    Balogh, A.; Lanzerotti, L. J.; Suess, S. T.

    2006-01-01

    Understanding how the Sun changes though its 11-year sunspot cycle and how these changes affect the vast space around the Sun the heliosphere has been one of the principal objectives of space research since the advent of the space age. This book presents the evolution of the heliosphere through an entire solar activity cycle. The last solar cycle (cycle 23) has been the best observed from both the Earth and from a fleet of spacecraft. Of these, the joint ESA-NASA Ulysses probe has provided continuous observations of the state of the heliosphere since 1990 from a unique vantage point, that of a nearly polar orbit around the Sun. Ulysses results affect our understanding of the heliosphere from the interior of the Sun to the interstellar medium - beyond the outer boundary of the heliosphere. Written by scientists closely associated with the Ulysses mission, the book describes and explains the many different aspects of changes in the heliosphere in response to solar activity. In particular, the authors describe the rise in solar ESA and NASA have now unamiously agreed a third extension to operate the highly successful Ulysses spacecraft until March 2008 and, in 2007 and 2008, the European-built space probe will fly over the poles of the Sun for a third time. This will enable Ulysses to add an important chapter to its survey of the high-latitude heliosphere and this additional material would be included in a 2nd edition of this book.

  8. Regulation of the Cell Division Cycle in Trypanosoma brucei

    PubMed Central

    2012-01-01

    The cell division cycle is tightly regulated by the activation and inactivation of a series of proteins that control the replication and segregation of organelles to the daughter cells. During the past decade, we have witnessed significant advances in our understanding of the cell cycle in Trypanosoma brucei and how the cycle is regulated by various regulatory proteins. However, many other regulators, especially those unique to trypanosomes, remain to be identified, and we are just beginning to delineate the signaling pathways that drive the transitions through different cell cycle stages, such as the G1/S transition, G2/M transition, and mitosis-cytokinesis transition. Trypanosomes appear to employ both evolutionarily conserved and trypanosome-specific molecules to regulate the various stages of its cell cycle, including DNA replication initiation, spindle assembly, chromosome segregation, and cytokinesis initiation and completion. Strikingly, trypanosomes lack some crucial regulators that are well conserved across evolution, such as Cdc6 and Cdt1, which are involved in DNA replication licensing, the spindle motor kinesin-5, which is required for spindle assembly, the central spindlin complex, which has been implicated in cytokinesis initiation, and the actomyosin contractile ring, which is located at the cleavage furrow. Conversely, trypanosomes possess certain regulators, such as cyclins, cyclin-dependent kinases, and mitotic centromere-associated kinesins, that are greatly expanded and likely play diverse cellular functions. Overall, trypanosomes apparently have integrated unique regulators into the evolutionarily conserved pathways to compensate for the absence of those conserved molecules and, additionally, have evolved certain cell cycle regulatory pathways that are either different from its human host or distinct between its own life cycle forms. PMID:22865501

  9. Polydatin inhibits growth of lung cancer cells by inducing apoptosis and causing cell cycle arrest.

    PubMed

    Zhang, Yusong; Zhuang, Zhixiang; Meng, Qinghui; Jiao, Yang; Xu, Jiaying; Fan, Saijun

    2014-01-01

    Polydatin (PD), a small natural compound from Polygonum cuspidatum, has a number of biological functions. However, the anticancer activity of PD has been poorly investigated. In the present study, thiazolyl blue tetrazolium bromide assay was used to evaluate the inhibitory effect of PD on cell growth. Cell cycle distribution and apoptosis were investigated by flow cytometry. In addition, the expression of several proteins associated with apoptosis and cell cycle were analyzed by western blot analysis. The results demonstrated that PD significantly inhibits the proliferation of A549 and NCI-H1975 lung cancer cell lines and causes dose-dependent apoptosis. Cell cycle analysis revealed that PD induces S phase cell cycle arrest. Western blot analysis showed that the expression of Bcl-2 decreased as that of Bax increased, and the expression of cyclin D1 was also suppressed. The results suggest that PD has potential therapeutic applications in the treatment of lung cancer. PMID:24348867

  10. Westinghouse fuel cell combined cycle systems

    SciTech Connect

    Veyo, S.

    1996-12-31

    Efficiency (voltage) of the solid oxide fuel cell (SOFC) should increase with operating pressure, and a pressurized SOFC could function as the heat addition process in a Brayton cycle gas turbine (GT) engine. An overall cycle efficiency of 70% should be possible. In cogeneration, half of the waste heat from a PSOFC/GT should be able to be captured in process steam and hot water, leading to a fuel effectiveness of about 85%. In order to make the PSOFC/GT a commercial reality, satisfactory operation of the SOFC at elevated pressure must be verified, a pressurized SOFC generator module must be designed, built, and tested, and the combined cycle and parameters must be optimized. A prototype must also be demonstrated. This paper describes progress toward making the PSOFC/GT a reality.

  11. 4D chromatin dynamics in cycling cells

    PubMed Central

    Strickfaden, Hilmar; Zunhammer, Andreas; van Koningsbruggen, Silvana; Köhler, Daniela

    2010-01-01

    This live cell study of chromatin dynamics in four dimensions (space and time) in cycling human cells provides direct evidence for three hypotheses first proposed by Theodor Boveri in seminal studies of fixed blastomeres from Parascaris equorum embryos: (I) Chromosome territory (CT) arrangements are stably maintained during interphase. (II) Chromosome proximity patterns change profoundly during prometaphase. (III) Similar CT proximity patterns in pairs of daughter nuclei reflect symmetrical chromosomal movements during anaphase and telophase, but differ substantially from the arrangement in mother cell nucleus. Hypothesis I could be confirmed for the majority of interphase cells. A minority, however, showed complex, rotational movements of CT assemblies with large-scale changes of CT proximity patterns, while radial nuclear arrangements were maintained. A new model of chromatin dynamics is proposed. It suggests that long-range DNA-DNA interactions in cell nuclei may depend on a combination of rotational CT movements and locally constrained chromatin movements. PMID:21327076

  12. The Yeast Cyclin-Dependent Kinase Routes Carbon Fluxes to Fuel Cell Cycle Progression.

    PubMed

    Ewald, Jennifer C; Kuehne, Andreas; Zamboni, Nicola; Skotheim, Jan M

    2016-05-19

    Cell division entails a sequence of processes whose specific demands for biosynthetic precursors and energy place dynamic requirements on metabolism. However, little is known about how metabolic fluxes are coordinated with the cell division cycle. Here, we examine budding yeast to show that more than half of all measured metabolites change significantly through the cell division cycle. Cell cycle-dependent changes in central carbon metabolism are controlled by the cyclin-dependent kinase (Cdk1), a major cell cycle regulator, and the metabolic regulator protein kinase A. At the G1/S transition, Cdk1 phosphorylates and activates the enzyme Nth1, which funnels the storage carbohydrate trehalose into central carbon metabolism. Trehalose utilization fuels anabolic processes required to reliably complete cell division. Thus, the cell cycle entrains carbon metabolism to fuel biosynthesis. Because the oscillation of Cdk activity is a conserved feature of the eukaryotic cell cycle, we anticipate its frequent use in dynamically regulating metabolism for efficient proliferation. PMID:27203178

  13. Predicting stem cell fate changes by differential cell cycle progression patterns.

    PubMed

    Roccio, Marta; Schmitter, Daniel; Knobloch, Marlen; Okawa, Yuya; Sage, Daniel; Lutolf, Matthias P

    2013-01-15

    Stem cell self-renewal, commitment and reprogramming rely on a poorly understood coordination of cell cycle progression and execution of cell fate choices. Using existing experimental paradigms, it has not been possible to probe this relationship systematically in live stem cells in vitro or in vivo. Alterations in stem cell cycle kinetics probably occur long before changes in phenotypic markers are apparent and could be used as predictive parameters to reveal changes in stem cell fate. To explore this intriguing concept, we developed a single-cell tracking approach that enables automatic detection of cell cycle phases in live (stem) cells expressing fluorescent ubiquitylation-based cell-cycle indicator (FUCCI) probes. Using this tool, we have identified distinctive changes in lengths and fluorescence intensities of G1 (red fluorescence) and S/G2-M (green) that are associated with self-renewal and differentiation of single murine neural stem/progenitor cells (NSCs) and embryonic stem cells (ESCs). We further exploited these distinctive features using fluorescence-activated cell sorting to select for desired stem cell fates in two challenging cell culture settings. First, as G1 length was found to nearly double during NSC differentiation, resulting in progressively increasing red fluorescence intensity, we successfully purified stem cells from heterogeneous cell populations by their lower fluorescence. Second, as ESCs are almost exclusively marked by the green (S/G2-M) FUCCI probe due to their very short G1, we substantially augmented the proportion of reprogramming cells by sorting green cells early on during reprogramming from a NSC to an induced pluripotent stem cell state. Taken together, our studies begin to shed light on the crucial relationship between cell cycle progression and fate choice, and we are convinced that the presented approach can be exploited to predict and manipulate cell fate in a wealth of other mammalian cell systems. PMID:23193167

  14. Novel functions of core cell cycle regulators in neuronal migration.

    PubMed

    Godin, Juliette D; Nguyen, Laurent

    2014-01-01

    The cerebral cortex is one of the most intricate regions of the brain, which required elaborated cell migration patterns for its development. Experimental observations show that projection neurons migrate radially within the cortical wall, whereas interneurons migrate along multiple tangential paths to reach the developing cortex. Tight regulation of the cell migration processes ensures proper positioning and functional integration of neurons to specific cerebral cortical circuits. Disruption of neuronal migration often lead to cortical dysfunction and/or malformation associated with neurological disorders. Unveiling the molecular control of neuronal migration is thus fundamental to understand the physiological or pathological development of the cerebral cortex. Generation of functional cortical neurons is a complex and stratified process that relies on decision of neural progenitors to leave the cell cycle and generate neurons that migrate and differentiate to reach their final position in the cortical wall. Although accumulating work shed some light on the molecular control of neuronal migration, we currently do not have a comprehensive understanding of how cell cycle exit and migration/differentiation are coordinated at the molecular level. The current chapter tends to lift the veil on this issue by discussing how core cell cycle regulators, and in particular p27(Kip1) acts as a multifunctional protein to control critical steps of neuronal migration through activities that go far beyond cell cycle regulation. PMID:24243100

  15. Recent Advances in GEO Water Cycle Activities

    NASA Astrophysics Data System (ADS)

    Lawford, R. G.

    2009-12-01

    Over the past few years GEO (Group on Earth Observations) efforts within the Water Societal Benefit Area (SBA) have been coordinated by the Science Committee of the former Integrated Global Observing Strategy Partnership (IGOS-P) IGWCO (Integrated Global Water Cycle Observations) theme. Within this framework a number of projects related to data system design, product development, and capacity building are being carried out. GEO has recently consolidated the Water SBA activities into three tasks, namely Droughts, Floods and Water Resource Management; Capacity Building for Water Resource Management (in Asia, Africa and the Americas); and Integrated Products for Water Resource Management and Research. In order to strengthen interactions with the GEO and its User Interface Committee, a Water Cycle Community of Practice (COP) was initiated. In addition, within the past year, the IGWCO Science Committee has decided to also function as a Community of Practice in collaboration with the existing Water Cycle COP. This overview will provide background and an update on the GEO Water SBA activities with an emphasis of the way in which these activities are being integrated within the three tasks. It will also describe activities that are planned for 2010 to facilitate this integration. Recent advances related to drought monitoring, capacity and network building, and observational and data systems will be highlighted. New water-related activities arising from collaborations between US GEO and Canada GEO, and through activities within the GEO Architecture and Data Committee, will also be described. This presentation will conclude with a longer-term outlook for water within the GEO framework and provide some guidance for interested experts on how they can become involved in helping to implement these plans.

  16. Apoptosis in male germ cells in response to cyclin A1-deficiency and cell cycle arrest.

    PubMed

    Salazar, Glicella; Liu, Dong; Liao, Ching; Batkiewicz, Leah; Arbing, Rachel; Chung, Sanny S W; Lele, Karen; Wolgemuth, Debra J

    2003-10-15

    Male mice homozygous for a mutated allele of the cyclin A1 gene (Ccna1) are sterile due to a block in cell cycle progression before the first meiotic division. Meiosis arrest in Ccna1(-/-) spermatocytes is associated with desynapsis abnormalities, lowered MPF activity, and apoptosis as evidenced by TUNEL-positive staining. With time, adult testicular tubules exhibit severe degeneration: some tubules in the older animals are almost devoid of germ cells at various stages of spermatogenesis. The mechanisms by which the cells sense the cell cycle arrest and the regulation of the decision to undergo cell death are under investigation. PMID:14555236

  17. α-Mangostin Induces Apoptosis and Cell Cycle Arrest in Oral Squamous Cell Carcinoma Cell

    PubMed Central

    Kwak, Hyun-Ho; Park, Bong-Soo

    2016-01-01

    Mangosteen has long been used as a traditional medicine and is known to have antibacterial, antioxidant, and anticancer effects. Although the effects of α-mangostin, a natural compound extracted from the pericarp of mangosteen, have been investigated in many studies, there is limited data on the effects of the compound in human oral squamous cell carcinoma (OSCC). In this study, α-mangostin was assessed as a potential anticancer agent against human OSCC cells. α-Mangostin inhibited cell proliferation and induced cell death in OSCC cells in a dose- and time-dependent manner with little to no effect on normal human PDLF cells. α-Mangostin treatment clearly showed apoptotic evidences such as nuclear fragmentation and accumulation of annexin V and PI-positive cells on OSCC cells. α-Mangostin treatment also caused the collapse of mitochondrial membrane potential and the translocation of cytochrome c from the mitochondria into the cytosol. The expressions of the mitochondria-related proteins were activated by α-mangostin. Treatment with α-mangostin also induced G1 phase arrest and downregulated cell cycle-related proteins (CDK/cyclin). Hence, α-mangostin specifically induces cell death and inhibits proliferation in OSCC cells via the intrinsic apoptosis pathway and cell cycle arrest at the G1 phase, suggesting that α-mangostin may be an effective agent for the treatment of OSCC. PMID:27478478

  18. Temporal Organization of the Cell Cycle

    PubMed Central

    Tyson, John J.; Novak, Bela

    2009-01-01

    Summary The coordination of growth, DNA replication and division in proliferating cells can be adequately explained by a ‘clock + checkpoint’ model. The clock is an underlying circular sequence of states; the checkpoints ensure that the cycle proceeds without mistakes. From the molecular complexities of the control system in modern eukaryotes, we isolate a simple network of positive and negative feedbacks that embodies a clock + checkpoints. The model accounts for the fundamental physiological properties of mitotic cell divisions, evokes a new view of the meiotic program, and suggests how the control system may have evolved in the first place. PMID:18786381

  19. Cell cycle population effects in perturbation studies

    PubMed Central

    O'Duibhir, Eoghan; Lijnzaad, Philip; Benschop, Joris J; Lenstra, Tineke L; van Leenen, Dik; Groot Koerkamp, Marian JA; Margaritis, Thanasis; Brok, Mariel O; Kemmeren, Patrick; Holstege, Frank CP

    2014-01-01

    Growth condition perturbation or gene function disruption are commonly used strategies to study cellular systems. Although it is widely appreciated that such experiments may involve indirect effects, these frequently remain uncharacterized. Here, analysis of functionally unrelated Saccharyomyces cerevisiae deletion strains reveals a common gene expression signature. One property shared by these strains is slower growth, with increased presence of the signature in more slowly growing strains. The slow growth signature is highly similar to the environmental stress response (ESR), an expression response common to diverse environmental perturbations. Both environmental and genetic perturbations result in growth rate changes. These are accompanied by a change in the distribution of cells over different cell cycle phases. Rather than representing a direct expression response in single cells, both the slow growth signature and ESR mainly reflect a redistribution of cells over different cell cycle phases, primarily characterized by an increase in the G1 population. The findings have implications for any study of perturbation that is accompanied by growth rate changes. Strategies to counter these effects are presented and discussed. PMID:24952590

  20. Elutriation for Cell Cycle Synchronization in Fission Yeast.

    PubMed

    Kume, Kazunori

    2016-01-01

    Cell synchronization is a powerful technique for studying the eukaryotic cell cycle events precisely. The fission yeast is a rod-shaped cell whose growth is coordinated with the cell cycle. Monitoring the cellular growth of fission yeast is a relatively simple way to measure the cell cycle stage of a cell. Here, we describe a detailed method of unperturbed cell synchronization, named centrifugal elutriation, for fission yeast. PMID:26254921

  1. Cell cycle regulation of Golgi membrane dynamics.

    PubMed

    Tang, Danming; Wang, Yanzhuang

    2013-06-01

    The Golgi apparatus is a membranous organelle in the cell that plays essential roles in protein and lipid trafficking, sorting, processing, and modification. Its basic structure is a stack of closely aligned flattened cisternae. In mammalian cells, dozens of Golgi stacks are often laterally linked into a ribbon-like structure. Biogenesis of the Golgi during cell division occurs through a sophisticated disassembly and reassembly process that can be divided into three distinct but cooperative steps, including the deformation and reformation of the Golgi cisternae, stacks, and ribbon. Here, we review our current understanding of the protein machineries that control these three steps in the cycle of mammalian cell division: GRASP65 and GRASP55 in Golgi stack and ribbon formation; ubiquitin and AAA ATPases in postmitotic Golgi membrane fusion; and golgins and cytoskeleton in Golgi ribbon formation. PMID:23453991

  2. Titer of trastuzumab produced by a Chinese hamster ovary cell line is associated with tricarboxylic acid cycle activity rather than lactate metabolism.

    PubMed

    Ishii, Yoichi; Imamoto, Yasufumi; Yamamoto, Rie; Tsukahara, Masayoshi; Wakamatsu, Kaori

    2015-04-01

    Achieving high productivity and quality is the final goal of therapeutic antibody development, but the productivity and quality of antibodies are known to be substantially dependent on the nature of the cell lines expressing the antibodies. We characterized two contrasting cell lines that produce trastuzumab, namely cell line A with a high titer and a low aggregate content and cell line B with a low titer and a high aggregate content to identify the causes of the differences. We observed the following differences: cell growth (A > B), proportion of defucosylated oligosaccharides on antibodies (A < B), and proportion of covalent antibody aggregates (A > B). Our results suggest that the high monoclonal antibody (mAb) titers in cell line A is associated with the high proliferation and is not caused by the lactate metabolism shift (switching from lactate production to net lactate consumption). Rather, these differences can be accounted for by the following: levels of tricarboxylic acid cycle intermediates (A > B), ammonium ion levels (A ≤ B), and oxidative stress (A > B). PMID:25449760

  3. Stochastic cycle selection in active flow networks.

    PubMed

    Woodhouse, Francis G; Forrow, Aden; Fawcett, Joanna B; Dunkel, Jörn

    2016-07-19

    Active biological flow networks pervade nature and span a wide range of scales, from arterial blood vessels and bronchial mucus transport in humans to bacterial flow through porous media or plasmodial shuttle streaming in slime molds. Despite their ubiquity, little is known about the self-organization principles that govern flow statistics in such nonequilibrium networks. Here we connect concepts from lattice field theory, graph theory, and transition rate theory to understand how topology controls dynamics in a generic model for actively driven flow on a network. Our combined theoretical and numerical analysis identifies symmetry-based rules that make it possible to classify and predict the selection statistics of complex flow cycles from the network topology. The conceptual framework developed here is applicable to a broad class of biological and nonbiological far-from-equilibrium networks, including actively controlled information flows, and establishes a correspondence between active flow networks and generalized ice-type models. PMID:27382186

  4. Cell cycle-dependence of HL-60 cell deformability.

    PubMed Central

    Tsai, M A; Waugh, R E; Keng, P C

    1996-01-01

    In this study, the role of cytoskeleton in HL-60 deformability during the cell cycle was investigated. G1, S, and G2/M cell fractions were separated by centrifugal elutriation. Cell deformability was evaluated by pipette aspiration. Tested at the same aspiration pressures, S cells were found to be less deformable than G1 cells. Moreover, HL-60 cells exhibited power-law fluid behavior: mu = mu c(gamma m/ gamma c)-b, where mu is cytoplasmic viscosity, gamma m is mean shear rate, mu c is the characteristic viscosity at the characteristic shear rate gamma c, and b is a material constant. At a given shear rate, S cells (mu c = 276 +/- 14 Pa.s, b = 0.51 +/- 0.03) were more viscous than G1 cells (mu c = 197 +/- 25, b = 0.53 +/- 0.02). To evaluate the relative importance of different cytoskeletal components in these cell cycle-dependent properties, HL-60 cells were treated with 30 microM dihydrocytochalasin B (DHB) to disrupt F-actin or 100 microM colchicine to collapse microtubules. DHB dramatically softened both G1 and S cells, which reduced the material constants mu c by approximately 65% and b by 20-30%. Colchicine had a limited effect on G1 cells but significantly reduced mu c of S cells (approximately 25%). Thus, F-actin plays the predominate role in determining cell mechanical properties, but disruption of microtubules may also influence the behavior of proliferating cells in a cell cycle-dependent fashion. Images FIGURE 1 PMID:8785361

  5. GEO Water Cycle Activities and Plans

    NASA Astrophysics Data System (ADS)

    Lawford, R.; Koike, T.; Ishida, C.; Grabs, W.

    2008-12-01

    The Group on Earth Observations (GEO) consists of more than 70 countries and 40 international organizations which are working together to develop the Global Earth Observation System of Systems (GEOSS). Since its launch in 2004, GEO has stimulated a wide range of activities related to data systems and their architecture, the development of science and technology to support observational programs, user interactions and interfaces, and capacity building. GEO tasks directed at Water Resources Management, one of the nine GEO Societal Benefit areas, are an integral part of these developments. They draw heavily upon the activities of the Integrated Global Water Cycle Observations (IGWCO) theme and on the activities and infrastructure provided through GEO and its committees. Within the GEO framework the water related activities have been focused on four specific tasks namely integrated data set development; information for floods, droughts and water management; water quality, and capacity building. Currently these efforts are being facilitated by the IGWCO theme that was formed under the former Integrated Global Observing Strategy Partnership (IGOS-P). With the dissolution of this partnership, other mechanisms, including the GEO Water Cycle Community of Practice, are being considered as new opportunitites for coordinating the work of the theme and the water-related GEO tasks. This talk provides a description of the GEO water tasks and reviews the progress that has been made in addressing them. It also provides a perspective on new opportunities and briefly describes some of the mechanisms, such as the Water Cycle Community of Practice, that could be expanded to coordinate a more comprehensive set of water tasks and greater community involvement.

  6. MicroRNAs and cell cycle of malignant glioma.

    PubMed

    Ouyang, Qing; Xu, Lunshan; Cui, Hongjuan; Xu, Minhui; Yi, Liang

    2016-01-01

    The control of malignant glioma cell cycle by microRNAs (miRNAs) is well established. The deregulation of miRNAs in glioma may contribute to tumor proliferation by directly targeting the critical cell-cycle regulators. Tumor suppressive miRNAs inhibit cell cycle through repressing the expression of positive cell-cycle regulators. However, oncogenic miRNAs promote the cell-cycle progression by targeting cell-cycle negative regulators. Recent studies have identified that transcription factors had involved in the expression of miRNAs. Transcription factors and miRNAs are implicated in regulatory network of glioma cell cycle, the deregulation of these transcription factors might be a cause of the deregulation of miRNAs. Abnormal versions of miRNAs have been implicated in the cell cycle of glioma. Based on those, miRNAs are excellent biomarker candidates and potential targets for therapeutic intervention in glioma. PMID:26000816

  7. Cell Division and Targeted Cell Cycle Arrest Opens and Stabilizes Basement Membrane Gaps

    PubMed Central

    Matus, David Q.; Chang, Emily; Makohon-Moore, Sasha C.; Hagedorn, Mary A.; Chi, Qiuyi; Sherwood, David R.

    2014-01-01

    Large gaps in basement membrane (BM) occur during organ remodeling and cancer cell invasion. Whether dividing cells, which temporarily reduce their attachment to BM, influence these breaches is unknown. Here we analyse uterine-vulval attachment during development across 21 species of rhabditid nematodes and find that the BM gap that forms between these organs is always bounded by a non-dividing vulval cell. Through cell cycle manipulation and live cell imaging in Caenorhabditis elegans, we show that actively dividing vulval cells facilitate enlargement of this breach by promoting BM movement. In contrast, targeted cell-cycle arrest halts BM movement and limits gap opening. Further, we demonstrate that the BM component laminin accumulates at the BM gap edge and promotes increased integrin levels in non-dividing vulval cells, stabilizing gap position. Together, these studies reveal that cell division can be used as a mechanism to regulate BM breaches, thus controlling the exchange of cells between tissues. PMID:24924309

  8. In vitro and in vivo anti-tumor activity of CoQ0 against melanoma cells: inhibition of metastasis and induction of cell-cycle arrest and apoptosis through modulation of Wnt/β-catenin signaling pathways

    PubMed Central

    Hseu, You-Cheng; Thiyagarajan, Varadharajan; Tsou, Hsiao-Tung; Lin, Kai-Yuan; Chen, Hui-Jye; Lin, Chung-Ming; Liao, Jiuun-Wang; Yang, Hsin-Ling

    2016-01-01

    Coenzyme Q0 (CoQ0, 2,3-dimethoxy-5-methyl-1,4-benzoquinone), a novel quinone derivative, has been shown to modulate cellular redox balance. However, effect of this compound on melanoma remains unclear. This study examined the in vitro or in vivo anti-tumor, apoptosis, and anti-metastasis activities of CoQ0 (0-20 μM) through inhibition of Wnt/β-catenin signaling pathway. CoQ0 exhibits a significant cytotoxic effect on melanoma cell lines (B16F10, B16F1, and A2058), while causing little toxicity toward normal (HaCaT) cells. The suppression of β-catenin was seen with CoQ0 administration accompanied by a decrease in the expression of Wnt/β-catenin transcriptional target c-myc, cyclin D1, and survivin through GSK3β-independent pathway. We found that CoQ0 treatment caused G1 cell-cycle arrest by reducing the levels of cyclin E and CDK4. Furthermore, CoQ0 treatment induced apoptosis through caspase-9/-3 activation, PARP degradation, Bcl-2/Bax dysregulation, and p53 expression. Notably, non- or sub-cytotoxic concentrations of CoQ0 markedly inhibited migration and invasion, accompanied by the down-regulation of MMP-2 and -9, and up-regulation of TIMP-1 and -2 expressions in highly metastatic B16F10 cells. Furthermore, the in vivo study results revealed that CoQ0 treatment inhibited the tumor growth in B16F10 xenografted nude mice. Histological analysis and western blotting confirmed that CoQ0 significantly decreased the xenografted tumor progression as demonstrated by induction of apoptosis, suppression of β-catenin, and inhibition of cell cycle-, apoptotic-, and metastatic-regulatory proteins. The data suggest that CoQ0 unveils a novel mechanism by down-regulating Wnt/β-catenin pathways and could be used as a potential lead compound for melanoma chemotherapy. PMID:26968952

  9. Mitochondrial Regulation of Cell Cycle and Proliferation

    PubMed Central

    Antico Arciuch, Valeria Gabriela; Elguero, María Eugenia; Poderoso, Juan José

    2012-01-01

    Abstract Eukaryotic mitochondria resulted from symbiotic incorporation of α-proteobacteria into ancient archaea species. During evolution, mitochondria lost most of the prokaryotic bacterial genes and only conserved a small fraction including those encoding 13 proteins of the respiratory chain. In this process, many functions were transferred to the host cells, but mitochondria gained a central role in the regulation of cell proliferation and apoptosis, and in the modulation of metabolism; accordingly, defective organelles contribute to cell transformation and cancer, diabetes, and neurodegenerative diseases. Most cell and transcriptional effects of mitochondria depend on the modulation of respiratory rate and on the production of hydrogen peroxide released into the cytosol. The mitochondrial oxidative rate has to remain depressed for cell proliferation; even in the presence of O2, energy is preferentially obtained from increased glycolysis (Warburg effect). In response to stress signals, traffic of pro- and antiapoptotic mitochondrial proteins in the intermembrane space (B-cell lymphoma-extra large, Bcl-2-associated death promoter, Bcl-2 associated X-protein and cytochrome c) is modulated by the redox condition determined by mitochondrial O2 utilization and mitochondrial nitric oxide metabolism. In this article, we highlight the traffic of the different canonical signaling pathways to mitochondria and the contributions of organelles to redox regulation of kinases. Finally, we analyze the dynamics of the mitochondrial population in cell cycle and apoptosis. Antioxid. Redox Signal. 16, 1150–1180. PMID:21967640

  10. Cell cycle-dependent regulation of extra-adrenal glucocorticoid synthesis in murine intestinal epithelial cells.

    PubMed

    Atanasov, Atanas G; Leiser, Dominic; Roesselet, Corinne; Noti, Mario; Corazza, Nadia; Schoonjans, Kristina; Brunner, Thomas

    2008-12-01

    Glucocorticoids are anti-inflammatory steroids with important applications in the treatment of inflammatory diseases. Endogenous glucocorticoids are mainly produced by the adrenal glands, although there is increasing evidence for extra-adrenal sources. Recent findings show that intestinal crypt cells produce glucocorticoids, which contribute to the maintenance of intestinal immune homeostasis. Intestinal glucocorticoid synthesis is critically regulated by the transcription factor liver receptor homologue-1 (LRH-1). As expression of steroidogenic enzymes and LRH-1 is restricted to the proliferating cells of the crypts, we aimed to investigate the role of the cell cycle in the regulation of LRH-1 activity and intestinal glucocorticoid synthesis. We here show that either pharmacological or molecular modulation of cell cycle progression significantly inhibited expression of steroidogenic enzymes and synthesis of glucocorticoids in intestinal epithelial cells. Synchronization of intestinal epithelial cells in the cell cycle revealed that expression of steroidogenic enzymes is preferentially induced at the G(1)/S stage. Differentiation of immature intestinal epithelial cells to mature nonproliferating cells also resulted in reduced expression of steroidogenic enzymes. This cell cycle-related effect on intestinal steroidogenesis was found to be mediated through the regulation of LRH-1 transcriptional activity. This mechanism may restrict intestinal glucocorticoid synthesis to the proliferating cells of the crypts. PMID:18711026

  11. ADP-ribosyltransferase in isolated nuclei during the cell cycle of Physarum polycephalum.

    PubMed Central

    Gröbner, P; Loidl, P

    1985-01-01

    ADP-ribosyltransferase was measured in isolated nuclei of Physarum polycephalum. Activity was determined with and without exogenous DNA and histones. During the synchronous cell cycle the activity measured with exogenous substrates exhibited a typical peak enzyme pattern with a maximum of activity in S-phase, whereas activity measured without exogenous substrates displayed a step enzyme pattern. Both activities doubled in each cell cycle. PMID:3002325

  12. Coronal Activity and Extended Solar Cycles

    NASA Astrophysics Data System (ADS)

    Altrock, R. C.

    2012-12-01

    Wilson et al. (1988, Nature 333, 748) discussed a number of solar parameters, which appear at high latitudes and gradually migrate towards the equator, merging with the sunspot "butterfly diagram". They found that this concept had been identified by earlier investigators extending back to 1957. They named this process the "Extended Solar Cycle" (ESC). Altrock (1997, Solar Phys. 170, 411) found that this process continued in Fe XIV 530.3 nm emission features. In cycles 21 - 23 solar maximum occurred when the number of Fe XIV emission regions per day > 0.19 (averaged over 365 days and both hemispheres) first reached latitudes 18°, 21° and 21°, for an average of 20° ± 1.7°. Other recent studies have shown that Torsional Oscillation (TO) negative-shear zones are co-located with the ESC from at least 50° down to the equator and also in the zones where the Rush to the Poles occur. These phenomena indicate that coronal activity occurring up to 50° and higher latitudes is related to TO shear zones, another indicator that the ESC is an important solar process. Another high-latitude process, which appears to be connected with the ESC, is the "Rush to the Poles" ("Rush") of polar crown prominences and their associated coronal emission, including Fe XIV. The Rush is is a harbinger of solar maximum (cf. Altrock, 2003, Solar Phys. 216, 343). Solar maximum in cycles 21 - 23 occurred when the center line of the Rush reached a critical latitude. These latitudes were 76°, 74° and 78°, respectively, for an average of 76° ± 2°. Applying the above conclusions to Cycle 24 is difficult due to the unusual nature of this cycle. Cycle 24 displays an intermittent "Rush" that is only well-defined in the northern hemisphere. In 2009 an initial slope of 4.6°/yr was found in the north, compared to an average of 9.4 ± 1.7 °/yr in the previous three cycles. This early fit to the Rush would have reached 76° at 2014.6. However, in 2010 the slope increased to 7.5°/yr (an increase

  13. Cell cycle control in isoproterenol-induced murine salivary acinar cell proliferation.

    PubMed

    Zeng, T; Yamamoto, H; Bowen, E; Broverman, R L; Nguyen, K H; Humphreys-Beher, M G

    1996-11-01

    The eukaryotic cell cycle is a summary of a complex network of signal transduction pathways resulting in both DNA replication and cell division. Cyclin-dependent kinases (CDKs) control the cell cycle in all eukaryotes, whereas other proteins, known as cyclins, act as their regulatory subunits. Chronic injection with isoproterenol (ISO) can induce acinar cell proliferation in rodent salivary glands. Cyclins and CDK proteins from control and ISO-treated murine parotid acinar cells were detected by using Western blotting techniques. By comparing the expression of these cell cycle regulatory kinases in the parotid acinar cell transition from a quiescent state to a hypertrophic state, we found rapid increases in the protein levels of all CDKs, cyclin D and proliferating cell nuclear antigen (PCNA). The highest protein levels for CDKs and cyclins appeared at about 72 hr of ISO stimulation and were coincident with the highest rate of increase in gland wet weight. After 72 hr, the increase of both cell cycle protein and gland wet weight began to subside. By using a co-immunoprecipitation method, the following cell cycle regulators (CDK-cyclin complexes) were detected, CDK4-cyclin D, CDK2-cyclin E, CDK2-cyclin A, and cdc2-cyclin B, along with an increase in kinase activity over control untreated animals. Additionally, we detected significant decreases in the newly isolated CDK inhibitor (CKI) p27kip but not Wee 1 kinase. The increased levels of CKI correlated with a decrease in kinase activity of CDK/cyclin complexes by 144 hr of chronic isoproterenol treatment. Our data suggest that the holoenzymes for cell cycle control (cyclin-CDK complexes) function as a final regulatory mechanism leading to salivary gland acinar cell proliferation. The gradual decline in protein levels of the CDKs and cyclins after 3 days of chronic treatment further indicates that ISO-induced proliferation of parotid acinar cells is self-limiting and non-tumorigenic. PMID:9375366

  14. Delayed cell cycle progression in selenoprotein W depleted cells is regulated by a mitogen-activated protein kinase kinase 4–p38–p53 pathway

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Selenoprotein W (SEPW1) is a ubiquitous, highly conserved thioredoxin-like protein whose depletion causes a p53- and p21Cip1-dependent G1-phase cell cycle arrest in breast and prostate epithelial cells. SEPW1 depletion increases phosphorylation of Ser33 in p53, which is associated with decreased p53...

  15. Initiation of stem cell differentiation involves cell cycle-dependent regulation of developmental genes by Cyclin D.

    PubMed

    Pauklin, Siim; Madrigal, Pedro; Bertero, Alessandro; Vallier, Ludovic

    2016-02-15

    Coordination of differentiation and cell cycle progression represents an essential process for embryonic development and adult tissue homeostasis. These mechanisms ultimately determine the quantities of specific cell types that are generated. Despite their importance, the precise molecular interplays between cell cycle machinery and master regulators of cell fate choice remain to be fully uncovered. Here, we demonstrate that cell cycle regulators Cyclin D1-3 control cell fate decisions in human pluripotent stem cells by recruiting transcriptional corepressors and coactivator complexes onto neuroectoderm, mesoderm, and endoderm genes. This activity results in blocking the core transcriptional network necessary for endoderm specification while promoting neuroectoderm factors. The genomic location of Cyclin Ds is determined by their interactions with the transcription factors SP1 and E2Fs, which result in the assembly of cell cycle-controlled transcriptional complexes. These results reveal how the cell cycle orchestrates transcriptional networks and epigenetic modifiers to instruct cell fate decisions. PMID:26883361

  16. Initiation of stem cell differentiation involves cell cycle-dependent regulation of developmental genes by Cyclin D

    PubMed Central

    Pauklin, Siim; Madrigal, Pedro; Bertero, Alessandro; Vallier, Ludovic

    2016-01-01

    Coordination of differentiation and cell cycle progression represents an essential process for embryonic development and adult tissue homeostasis. These mechanisms ultimately determine the quantities of specific cell types that are generated. Despite their importance, the precise molecular interplays between cell cycle machinery and master regulators of cell fate choice remain to be fully uncovered. Here, we demonstrate that cell cycle regulators Cyclin D1–3 control cell fate decisions in human pluripotent stem cells by recruiting transcriptional corepressors and coactivator complexes onto neuroectoderm, mesoderm, and endoderm genes. This activity results in blocking the core transcriptional network necessary for endoderm specification while promoting neuroectoderm factors. The genomic location of Cyclin Ds is determined by their interactions with the transcription factors SP1 and E2Fs, which result in the assembly of cell cycle-controlled transcriptional complexes. These results reveal how the cell cycle orchestrates transcriptional networks and epigenetic modifiers to instruct cell fate decisions. PMID:26883361

  17. High levels of endogenous nitric oxide produced after burn injury in rats arrest activated T lymphocytes in the first G1 phase of the cell cycle and then induce their apoptosis

    SciTech Connect

    Valenti, Lionel M.; Chancerelle, Yves; Sousa, Martine de; Anh Tuan Dinh-Xuan

    2005-05-15

    Major physical traumas provoke a systemic inflammatory response and immune dysfunction. In a model of thermal injury in rats, we previously showed that an overproduction of nitric oxide (NO) was responsible for the collapse of lymphoproliferative responses. In the present work, we performed a time-course analysis of cell proliferation and cell death parameters in order to establish the sequence of events triggered by the high NO output in Wistar/Han rat splenocytes activated with Con A, 10 days after burn injury. We demonstrate that activated T cells from burned rats never divided whereas normal T cells underwent four division cycles. However, T cells from both burned and normal rat entered the G1 phase as shown by increase of cell size, mitochondria hyperpolarization, and expression of cyclin D1. Burned rat T cells progressed to the late G1 phase as shown by expression of the nuclear Ki-67 antigen, but they never entered the S phase. They underwent apoptosis as shown by morphological parameters, disruption of transmembrane mitochondrial potential, and DNA fragmentation. Persistent accumulation of the p53 protein accompanied these phenomena. NO synthase inhibitors antagonize alterations of cell proliferation and cell death parameters in burned rat T cells and accelerated p53 turnover.

  18. Does Arabidopsis thaliana DREAM of cell cycle control?

    PubMed Central

    Fischer, Martin; DeCaprio, James A

    2015-01-01

    Strict temporal control of cell cycle gene expression is essential for all eukaryotes including animals and plants. DREAM complexes have been identified in worm, fly, and mammals, linking several distinct transcription factors to coordinate gene expression throughout the cell cycle. In this issue of The EMBO Journal, Kobayashi et al (2015) identify distinct activator and repressor complexes for genes expressed during the G2 and M phases in Arabidopsis that can be temporarily separated during proliferating and post-mitotic stages of development. The complexes incorporate specific activator and repressor MYB and E2F transcription factors and indicate the possibility of the existence of multiple DREAM complexes in plants. PMID:26089020

  19. Cell Cycle Control by a Minimal Cdk Network

    PubMed Central

    Gérard, Claude; Tyson, John J.; Coudreuse, Damien; Novák, Béla

    2015-01-01

    In present-day eukaryotes, the cell division cycle is controlled by a complex network of interacting proteins, including members of the cyclin and cyclin-dependent protein kinase (Cdk) families, and the Anaphase Promoting Complex (APC). Successful progression through the cell cycle depends on precise, temporally ordered regulation of the functions of these proteins. In light of this complexity, it is surprising that in fission yeast, a minimal Cdk network consisting of a single cyclin-Cdk fusion protein can control DNA synthesis and mitosis in a manner that is indistinguishable from wild type. To improve our understanding of the cell cycle regulatory network, we built and analysed a mathematical model of the molecular interactions controlling the G1/S and G2/M transitions in these minimal cells. The model accounts for all observed properties of yeast strains operating with the fusion protein. Importantly, coupling the model’s predictions with experimental analysis of alternative minimal cells, we uncover an explanation for the unexpected fact that elimination of inhibitory phosphorylation of Cdk is benign in these strains while it strongly affects normal cells. Furthermore, in the strain without inhibitory phosphorylation of the fusion protein, the distribution of cell size at division is unusually broad, an observation that is accounted for by stochastic simulations of the model. Our approach provides novel insights into the organization and quantitative regulation of wild type cell cycle progression. In particular, it leads us to propose a new mechanistic model for the phenomenon of mitotic catastrophe, relying on a combination of unregulated, multi-cyclin-dependent Cdk activities. PMID:25658582

  20. Mechanisms of Cell Cycle Control Revealed by a Systematic and Quantitative Overexpression Screen in S. cerevisiae

    PubMed Central

    Niu, Wei; Li, Zhihua; Zhan, Wenjing; Iyer, Vishwanath R.; Marcotte, Edward M.

    2008-01-01

    Regulation of cell cycle progression is fundamental to cell health and reproduction, and failures in this process are associated with many human diseases. Much of our knowledge of cell cycle regulators derives from loss-of-function studies. To reveal new cell cycle regulatory genes that are difficult to identify in loss-of-function studies, we performed a near-genome-wide flow cytometry assay of yeast gene overexpression-induced cell cycle delay phenotypes. We identified 108 genes whose overexpression significantly delayed the progression of the yeast cell cycle at a specific stage. Many of the genes are newly implicated in cell cycle progression, for example SKO1, RFA1, and YPR015C. The overexpression of RFA1 or YPR015C delayed the cell cycle at G2/M phases by disrupting spindle attachment to chromosomes and activating the DNA damage checkpoint, respectively. In contrast, overexpression of the transcription factor SKO1 arrests cells at G1 phase by activating the pheromone response pathway, revealing new cross-talk between osmotic sensing and mating. More generally, 92%–94% of the genes exhibit distinct phenotypes when overexpressed as compared to their corresponding deletion mutants, supporting the notion that many genes may gain functions upon overexpression. This work thus implicates new genes in cell cycle progression, complements previous screens, and lays the foundation for future experiments to define more precisely roles for these genes in cell cycle progression. PMID:18617996

  1. Citral inhibits cell proliferation and induces apoptosis and cell cycle arrest in MCF-7 cells.

    PubMed

    Chaouki, Wahid; Leger, David Y; Liagre, Bertrand; Beneytout, Jean-Louis; Hmamouchi, Mohamed

    2009-10-01

    Many natural components of plants extract are studied for their beneficial effects on health and particularly on carcinogenesis chemoprevention. In this study, we investigated the effect of citral (3,7-dimethyl-2,6-octadienal), a key component of essential oils extracted from several herbal plants, on the proliferation rate, cell cycle distribution, and apoptosis of the human breast cancer cell line MCF-7. The effects of this compound were also tested on cyclo-oxygenase activity. Citral treatment caused inhibition of MCF-7 cell growth (IC(50)-48 h: 18 x 10(-5)m), with a cycle arrest in G(2)/M phase and apoptosis induction. Moreover, we observed a decrease in prostaglandin E(2) synthesis 48 h after citral treatment. These findings suggest that citral has a potential chemopreventive effect. PMID:19656204

  2. Cell cycle modulation by a multitargeted antifolate, LY231514, increases the cytotoxicity and antitumor activity of gemcitabine in HT29 colon carcinoma.

    PubMed

    Tonkinson, J L; Worzalla, J F; Teng, C H; Mendelsohn, L G

    1999-08-01

    The proliferation rate of HT29 colon carcinoma cells was decreased by the multitargeted antifolate (MTA), LY231514. This effect correlated with a buildup of cells near the G1-S interface after 24 h of incubation, and a synchronized progression of the population through S phase during the next 24 h. MTA treatment (0.03-3 microM) was minimally cytotoxic (20-30%) to HT29 cells after a 24-h exposure, and no dose response was observed. In contrast, the nucleoside analogue gemcitabine (GEM) was cytotoxic (IC50, 0.071 +/- 0.011 microM; IC90, 0.648 +/- 0.229 microM) after a 24-h exposure. We hypothesized that pretreatment of these cells with MTA would increase the potency of GEM by synchronizing the population for DNA synthesis. The cytotoxicity of GEM increased 2-7-fold when MTA was administered 24 h before GEM (IC50, 0.032 +/- 0.009 microM; IC90, 0.094 +/- 0.019 microM). In addition, an increase in cell kill for the combination compared with GEM alone (IC99, 12 microM for GEM alone; IC99, 0.331 microM for combination) was observed. No increase in potency or cell kill was observed when the two compounds were added simultaneously. MTA pretreatment also potentiated the cytotoxicity of a 1-h exposure to GEM. These cell-based observations were extended to evaluate the schedule-dependent interaction of these two agents in vivo using a nude mouse HT29 xenograft tumor model. At the doses tested, MTA alone (100 mg/kg) had a marginal effect on tumor growth delay, whereas GEM (80 mg/kg) produced a statistically significant tumor growth delay. In combination, the increase in tumor growth delay was greatest when MTA was administered before GEM, compared with simultaneous drug administration or the reverse sequence, e.g., GEM followed by MTA. The effect of sequential administration of MTA followed by GEM was greater than additive, indicating synergistic interaction of these agents. Thus, in vitro, MTA induced cell cycle effects on HT29 cells that resulted in potentiation of the

  3. Ethanol Mediates Cell Cycle Arrest and Apoptosis in SK-N-SH Neuroblastoma Cells

    PubMed Central

    Lee, Maria; Song, Byoung-Joon; Kwon, Yongil

    2014-01-01

    Background: The mechanisms of cell or organ damage by chronic alcohol consumption are still poorly understood. The present study aimed to investigate the role of the mitogen-activated protein kinases during ethanol-induced damage to SK-N-SH neuroblastoma cells. Methods: Cells were treated with ethanol and subsequently analyzed for cell morphology, viability, and DNA fragmentation. Immunoblot analysis was performed to assess various proteins levels associated with cell cycle arrest and apoptosis after ethanol exposure. Results: Ethanol induced time- and dose-dependent cell death in SK-N-SH cells and increased c-Jun N-terminal protein kinase (JNK) activity in a time- and concentration dependent manner. In contrast, p38 kinase activity increased transiently. After treatment with JNK or p38 kinase inhibitors, ethanol-induced cell death significantly reduced. Ethanol-induced cell death was accompanied by increased cytochrome c release and caspase 3 activity observed at 12 h. In contrast, the level of anti-apoptotic Bcl-2 protein did not change. Ethanol also increased the phosphorylation of p53 and p53 activation was followed by an increase in the p21 tumor suppressor protein accompanied by a gradual decrease in phospho-Rb protein. Conclusion: Our results suggest that ethanol mediates apoptosis of neuroblastoma cells by stimulating p53-related cell cycle arrest mediated through activation of the JNK-related pathway. PMID:25337571

  4. Regulation of Sp1 by cell cycle related proteins

    PubMed Central

    Tapias, Alicia; Ciudad, Carlos J.; Roninson, Igor B.; Noé, Véronique

    2009-01-01

    Sp1 transcription factor regulates the expression of multiple genes, including the Sp1 gene itself. We analyzed the ability of different cell cycle regulatory proteins to interact with Sp1 and to affect Sp1 promoter activity. Using an antibody array, we observed that CDK4, SKP2, Rad51, BRCA2 and p21 could interact with Sp1 and we confirmed these interactions by co-immunoprecipitation. CDK4, SKP2, Rad51, BRCA2 and p21 also activated the Sp1 promoter. Among the known Sp1-interacting proteins, E2F-DP1, Cyclin D1, Stat3 and Rb activated the Sp1 promoter, whereas p53 and NFκB inhibited it. The proteins that regulated Sp1 gene expression were shown by positive chromatin immunoprecipitation to be bound to the Sp1 promoter. Moreover, SKP2, BRCA2, p21, E2F-DP1, Stat3, Rb, p53 and NFκB had similar effects on an artificial promoter containing only Sp1 binding sites. Transient transfections of CDK4, Rad51, E2F-DP1, p21 and Stat3 increased mRNA expression from the endogenous Sp1 gene in HeLa cells whereas overexpression of NFκB, and p53 decreased Sp1 mRNA levels. p21 expression from a stably integrated inducible promoter in HT1080 cells activated Sp1 expression at the promoter and mRNA levels, but at the same time it decreased Sp1 protein levels due to the activation of Sp1 degradation. The observed multiple effects of cell cycle regulators on Sp1 suggest that Sp1 may be a key mediator of cell cycle associated changes in gene expression. PMID:18769160

  5. Alteration of cell cycle progression by Sindbis virus infection

    SciTech Connect

    Yi, Ruirong; Saito, Kengo; Isegawa, Naohisa; Shirasawa, Hiroshi

    2015-07-10

    We examined the impact of Sindbis virus (SINV) infection on cell cycle progression in a cancer cell line, HeLa, and a non-cancerous cell line, Vero. Cell cycle analyses showed that SINV infection is able to alter the cell cycle progression in both HeLa and Vero cells, but differently, especially during the early stage of infection. SINV infection affected the expression of several cell cycle regulators (CDK4, CDK6, cyclin E, p21, cyclin A and cyclin B) in HeLa cells and caused HeLa cells to accumulate in S phase during the early stage of infection. Monitoring SINV replication in HeLa and Vero cells expressing cell cycle indicators revealed that SINV which infected HeLa cells during G{sub 1} phase preferred to proliferate during S/G{sub 2} phase, and the average time interval for viral replication was significantly shorter in both HeLa and Vero cells infected during G{sub 1} phase than in cells infected during S/G{sub 2} phase. - Highlights: • SINV infection was able to alter the cell cycle progression of infected cancer cells. • SINV infection can affect the expression of cell cycle regulators. • SINV infection exhibited a preference for the timing of viral replication among the cell cycle phases.

  6. Mitochondrial dynamics and the cell cycle

    PubMed Central

    Kianian, Penny M. A.; Kianian, Shahryar F.

    2014-01-01

    Nuclear-mitochondrial (NM) communication impacts many aspects of plant development including vigor, sterility, and viability. Dynamic changes in mitochondrial number, shape, size, and cellular location takes place during the cell cycle possibly impacting the process itself and leading to distribution of this organelle into daughter cells. The genes that underlie these changes are beginning to be identified in model plants such as Arabidopsis. In animals disruption of the drp1 gene, a homolog to the plant drp3A and drp3B, delays mitochondrial division. This mutation results in increased aneuploidy due to chromosome mis-segregation. It remains to be discovered if a similar outcome is observed in plants. Alloplasmic lines provide an opportunity to understand the communication between the cytoplasmic organelles and the nucleus. Examples of studies in these lines, especially from the extensive collection in wheat, point to the role of mitochondria in chromosome movement, pollen fertility and other aspects of development. PMID:24904617

  7. Flip-flop cycles in solar and stellar activity

    NASA Astrophysics Data System (ADS)

    Berdyugina, S. V.

    2006-08-01

    Doppler images and long time series of photometric observations of cool active stars reveal permanent active longitudes on their surfaces. They are found to alternate their dominant activity quasi-periodically which indicates a new type of the activity cycles, flip-flop cycles. In this talk I will review properties of active longitudes and flip-flop cycles on different types of active stars including the Sun.

  8. Tcf3 and cell cycle factors contribute to butyrate resistance in colorectal cancer cells

    SciTech Connect

    Chiaro, Christopher; Lazarova, Darina L.; Bordonaro, Michael

    2012-11-09

    Highlights: Black-Right-Pointing-Pointer We investigate mechanisms responsible for butyrate resistance in colon cancer cells. Black-Right-Pointing-Pointer Tcf3 modulates butyrate's effects on Wnt activity and cell growth in resistant cells. Black-Right-Pointing-Pointer Tcf3 modulation of butyrate's effects differ by cell context. Black-Right-Pointing-Pointer Cell cycle factors are overexpressed in the resistant cells. Black-Right-Pointing-Pointer Reversal of altered gene expression can enhance the anti-cancer effects of butyrate. -- Abstract: Butyrate, a fermentation product of dietary fiber, inhibits clonal growth in colorectal cancer (CRC) cells dependent upon the fold induction of Wnt activity. We have developed a CRC cell line (HCT-R) that, unlike its parental cell line, HCT-116, does not respond to butyrate exposure with hyperactivation of Wnt signaling and suppressed clonal growth. PCR array analyses revealed Wnt pathway-related genes, the expression of which differs between butyrate-sensitive HCT-116 CRC cells and their butyrate-resistant HCT-R cell counterparts. We identified overexpression of Tcf3 as being partially responsible for the butyrate-resistant phenotype, as this DNA-binding protein suppresses the hyperinduction of Wnt activity by butyrate. Consequently, Tcf3 knockdown in HCT-R cells restores their sensitivity to the effects of butyrate on Wnt activity and clonal cell growth. Interestingly, the effects of overexpressed Tcf3 differ between HCT-116 and HCT-R cells; thus, in HCT-116 cells Tcf3 suppresses proliferation without rendering the cells resistant to butyrate. In HCT-R cells, however, the overexpression of Tcf3 inhibits Wnt activity, and the cells are still able to proliferate due to the higher expression levels of cell cycle factors, particularly those driving the G{sub 1} to S transition. Knowledge of the molecular mechanisms determining the variable sensitivity of CRC cells to butyrate may assist in developing approaches that prevent or

  9. Coupling of the cell cycle and apoptotic machineries in developing T cells.

    PubMed

    Xue, Ling; Sun, Yuefang; Chiang, Leslie; He, Bo; Kang, Chulho; Nolla, Hector; Winoto, Astar

    2010-03-01

    Proliferation and apoptosis are diametrically opposite processes. Expression of certain genes like c-Myc, however, can induce both, pointing to a possible linkage between them. Developing CD4(+)CD8(+) thymocytes are intrinsically sensitive to apoptosis, but the molecular basis is not known. We have found that these noncycling cells surprisingly express many cell cycle proteins. We generated transgenic mice expressing a CDK2 kinase-dead (CDK2-DN) protein in the T cell compartment. Analysis of these mice showed that the CDK2-DN protein acts as a dominant negative mutant in mature T cells as expected, but surprisingly, it acts as a dominant active protein in CD4(+)CD8(+) thymocytes. The levels of CDK2 kinase activity, cyclin E, cyclin A, and other cell cycle proteins in transgenic CD4(+)CD8(+) thymocytes are increased. Concurrently, caspase levels are elevated, and apoptosis is significantly enhanced in vitro and in vivo. E2F-1, the unique E2F member capable of inducing apoptosis when overexpressed, is specifically up-regulated in transgenic CD4(+)CD8(+) thymocytes but not in other T cell populations. These results demonstrate that the cell cycle and apoptotic machineries are normally linked, and expression of cell cycle proteins in developing T cells contributes to their inherent 1sensitivity to apoptosis. PMID:20068041

  10. Cell cycle constraints on capsulation and bacteriophage susceptibility

    PubMed Central

    Ardissone, Silvia; Fumeaux, Coralie; Bergé, Matthieu; Beaussart, Audrey; Théraulaz, Laurence; Radhakrishnan, Sunish Kumar; Dufrêne, Yves F; Viollier, Patrick H

    2014-01-01

    Despite the crucial role of bacterial capsules in pathogenesis, it is still unknown if systemic cues such as the cell cycle can control capsule biogenesis. In this study, we show that the capsule of the synchronizable model bacterium Caulobacter crescentus is cell cycle regulated and we unearth a bacterial transglutaminase homolog, HvyA, as restriction factor that prevents capsulation in G1-phase cells. This capsule protects cells from infection by a generalized transducing Caulobacter phage (φCr30), and the loss of HvyA confers insensitivity towards φCr30. Control of capsulation during the cell cycle could serve as a simple means to prevent steric hindrance of flagellar motility or to ensure that phage-mediated genetic exchange happens before the onset of DNA replication. Moreover, the multi-layered regulatory circuitry directing HvyA expression to G1-phase is conserved during evolution, and HvyA orthologues from related Sinorhizobia can prevent capsulation in Caulobacter, indicating that alpha-proteobacteria have retained HvyA activity. DOI: http://dx.doi.org/10.7554/eLife.03587.001 PMID:25421297

  11. Possible roles of the endocytic cycle in cell motility.

    PubMed

    Traynor, David; Kay, Robert R

    2007-07-15

    Starving, highly motile Dictyostelium cells maintain an active endocytic cycle, taking up their surface about every 11 minutes. Cell motility depends on a functional NSF (N-ethylmaleimide sensitive factor) protein--also essential for endocytosis and membrane trafficking generally--and we, therefore, investigated possible ways in which the endocytic cycle might be required for cell movement. First, NSF, and presumably membrane trafficking, are not required for the initial polarization of the leading edge in a cyclic-AMP gradient. Second, we can detect no evidence for membrane flow from the leading edge, as photobleached or photoactivated marks in the plasma membrane move forward roughly in step with the leading edge, rather than backwards from it. Third, we find that the surface area of a cell--measured from confocal reconstructions--constantly fluctuates during movement as it projects pseudopodia and otherwise changes shape; increases of 20-30% can often occur over a few minutes. These fluctuations cannot be explained by reciprocal changes in filopodial surface area and they substantially exceed the 2-3% by which membranes can stretch. We propose that the endocytic cycle has a key function in motility by allowing adjustment of cell surface area to match changes in shape and that, without this function, movement is severely impaired. PMID:17606987

  12. Preclinical characterization of the CDK4/6 inhibitor LY2835219: in-vivo cell cycle-dependent/independent anti-tumor activities alone/in combination with gemcitabine.

    PubMed

    Gelbert, Lawrence M; Cai, Shufen; Lin, Xi; Sanchez-Martinez, Concepcion; Del Prado, Miriam; Lallena, Maria Jose; Torres, Raquel; Ajamie, Rose T; Wishart, Graham N; Flack, Robert Steven; Neubauer, Blake Lee; Young, Jamie; Chan, Edward M; Iversen, Philip; Cronier, Damien; Kreklau, Emiko; de Dios, Alfonso

    2014-10-01

    The G1 restriction point is critical for regulating the cell cycle and is controlled by the Rb pathway (CDK4/6-cyclin D1-Rb-p16/ink4a). This pathway is important because of its inactivation in a majority of human tumors. Transition through the restriction point requires phosphorylation of retinoblastoma protein (Rb) by CDK4/6, which are highly validated cancer drug targets. We present the identification and characterization of a potent CDK4/6 inhibitor, LY2835219. LY2835219 inhibits CDK4 and CDK6 with low nanomolar potency, inhibits Rb phosphorylation resulting in a G1 arrest and inhibition of proliferation, and its activity is specific for Rb-proficient cells. In vivo target inhibition studies show LY2835219 is a potent inhibitor of Rb phosphorylation, induces a complete cell cycle arrest and suppresses expression of several Rb-E2F-regulated proteins 24 hours after a single dose. Oral administration of LY2835219 inhibits tumor growth in human tumor xenografts representing different histologies in tumor-bearing mice. LY2835219 is effective and well tolerated when administered up to 56 days in immunodeficient mice without significant loss of body weight or tumor outgrowth. In calu-6 xenografts, LY2835219 in combination with gemcitabine enhanced in vivo antitumor activity without a G1 cell cycle arrest, but was associated with a reduction of ribonucleotide reductase expression. These results suggest LY2835219 may be used alone or in combination with standard-of-care cytotoxic therapy. In summary, we have identified a potent, orally active small-molecule inhibitor of CDK4/6 that is active in xenograft tumors. LY2835219 is currently in clinical development. PMID:24919854

  13. DNA methylation is stable during replication and cell cycle arrest

    PubMed Central

    Vandiver, Amy R.; Idrizi, Adrian; Rizzardi, Lindsay; Feinberg, Andrew P.; Hansen, Kasper D.

    2015-01-01

    DNA methylation is an epigenetic modification with important functions in development. Large-scale loss of DNA methylation is a hallmark of cancer. Recent work has identified large genomic blocks of hypomethylation associated with cancer, EBV transformation and replicative senescence, all of which change the proportion of actively proliferating cells within the population measured. We asked if replication or cell-cycle arrest affects the global levels of methylation or leads to hypomethylated blocks as observed in other settings. We used fluorescence activated cell sorting to isolate primary dermal fibroblasts in G0, G1 and G2 based on DNA content and Ki67 staining. We additionally examined G0 cells arrested by contact inhibition for one week to determine the effects of extended arrest. We analyzed genome wide DNA methylation from sorted cells using whole genome bisulfite sequencing. This analysis demonstrated no global changes or large-scale hypomethylated blocks in any of the examined cell cycle phases, indicating that global levels of methylation are stable with replication and arrest. PMID:26648411

  14. Synthesis, anticancer activity and effects on cell cycle profile and apoptosis of novel thieno[2,3-d]pyrimidine and thieno[3,2-e] triazolo[4,3-c]pyrimidine derivatives.

    PubMed

    Kandeel, Manal M; Refaat, Hanan M; Kassab, Asmaa E; Shahin, Inas G; Abdelghany, Tamer M

    2015-01-27

    Motivated by the widely reported anticancer activity of thieno[2,3-d]pyrimidines a series of 24 new 2-substitutedhexahydrocycloocta[4,5] thieno[2,3-d]pyrimidines with different substituents at C-4 position and hexahydrocycloocta[4,5]thieno[3,2-e]-1,2,4-triazolo[4,3-c]pyrimidines were synthesized. The anticancer activity of 17 compounds were evaluated by National Cancer Institute (USA) using a two stage process utilizing 59 different human tumor cell lines representing leukemia, melanoma, cancers of lung, colon, central nervous system (CNS), ovary, kidney, prostate as well as breast. Compound 9c showed broad spectrum potent anticancer activity in nano molar to micro molar range against 56 human tumor cell lines with GI50 less than 10 μM ranging from 0.495 to 5.57 μM, also it is worth mentioning that compound 9c had the marked highest selectivity against the two cell lines T-47D and MDA-MB-468 belonging to breast cancer with GI50 = 0.495 and 0.568 μM respectively, and its effect was further studied on cell cycle progression and induction of apoptosis in the MDA-MB-468 cell line. Results showed that compound 9c induced cell cycle arrest at G2/M phase and also, showed accumulation of cells in pre-G1 phase which may result from, degradation or fragmentation of the genetic materials indicating a possible role of apoptosis in compound 9c-induced cancer cell death and cytotoxicity and verifying this compound as promising selective anticancer lead. Compound 6c was selective against K-562, SR and MOLT-4 cell lines belonging to leukemia showing growth inhibition percentages 86.38, 65.76 and 60.40 at a single dose test, at the same time it showed lethal activity against HOP-92 representing non-small cell lung cancer. Interestingly, leukemia SR, CNS cancer SNB-75 and renal cancer UO-31 cell lines proved to be sensitive to compound 6d with growth inhibition percentages 52.86, 50.94 and 53.99 respectively. Additionally, compound 6d demonstrated lethal activity to HOP-92

  15. Selection of mammalian cells based on their cell-cycle phase using dielectrophoresis

    PubMed Central

    Kim, Unyoung; Shu, Chih-Wen; Dane, Karen Y.; Daugherty, Patrick S.; Wang, Jean Y. J.; Soh, H. T.

    2007-01-01

    An effective, noninvasive means of selecting cells based on their phase within the cell cycle is an important capability for biological research. Current methods of producing synchronous cell populations, however, tend to disrupt the natural physiology of the cell or suffer from low synchronization yields. In this work, we report a microfluidic device that utilizes the dielectrophoresis phenomenon to synchronize cells by exploiting the relationship between the cell's volume and its phase in the cell cycle. The dielectrophoresis activated cell synchronizer (DACSync) device accepts an asynchronous mixture of cells at the inlet, fractionates the cell populations according to the cell-cycle phase (G1/S and G2/M), and elutes them through different outlets. The device is gentle and efficient; it utilizes electric fields that are 1–2 orders of magnitude below those used in electroporation and enriches asynchronous tumor cells in the G1 phase to 96% in one round of sorting, in a continuous flow manner at a throughput of 2 × 105 cells per hour per microchannel. This work illustrates the feasibility of using laminar flow and electrokinetic forces for the efficient, noninvasive separation of living cells. PMID:18093921

  16. Effect of Docosahexaenoic Acid on Cell Cycle Pathways in Breast Cell Lines With Different Transformation Degree.

    PubMed

    Rescigno, Tania; Capasso, Anna; Tecce, Mario Felice

    2016-06-01

    n-3 polyunsaturated fatty acids (PUFAs), such as eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), abundant in fish, have been shown to affect development and progression of some types of cancer, including breast cancer. The aim of our study was to further analyze and clarify the effects of these nutrients on the molecular mechanisms underlying breast cancer. Following treatments with DHA we examined cell viability, death, cell cycle, and some molecular effects in breast cell lines with different transformation, phenotypic, and biochemical characteristics (MCF-10A, MCF-7, SK-BR-3, ZR-75-1). These investigations showed that DHA is able to affect cell viability, proliferation, and cell cycle progression in a different way in each assayed breast cell line. The activation of ERK1/2 and STAT3 pathways and the expression and/or activation of molecules involved in cell cycle regulation such as p21(Waf1/Cip1) and p53, are very differently regulated by DHA treatments in each cell model. DHA selectively: (i) arrests non tumoral MCF-10A breast cells in G0 /G1 cycle phase, activating p21(Waf1/Cip1) , and p53, (ii) induces to death highly transformed breast cells SK-BR-3, reducing ERK1/2 and STAT3 phosphorylation and (iii) only slightly affects each analyzed process in MCF-7 breast cell line with transformation degree lower than SK-BR-3 cells. These findings suggest a more relevant inhibitory role of DHA within early development and late progression of breast cancer cell transformation and a variable effect in the other phases, depending on individual molecular properties and degree of malignancy of each clinical case. J. Cell. Physiol. 231: 1226-1236, 2016. © 2015 Wiley Periodicals, Inc. PMID:26480024

  17. Anticancer effect of arsenite on cell migration, cell cycle and apoptosis in human pancreatic cancer cells

    PubMed Central

    HORIBE, YOHEI; ADACHI, SEIJI; YASUDA, ICHIRO; YAMAUCHI, TAKAHIRO; KAWAGUCHI, JUNJI; KOZAWA, OSAMU; SHIMIZU, MASAHITO; MORIWAKI, HISATAKA

    2016-01-01

    The standard treatment for advanced pancreatic cancer is chemotherapy, but its clinical outcome remains unsatisfactory. Therefore, the development of novel treatments for this malignancy is urgently required. In the present study, the anticancer effect of arsenite on platelet-derived growth factor (PDGF)-BB-induced migration, cell cycle and apoptosis was investigated in pancreatic cancer cells (AsPC-1 and BxPC-3), and compared with the effect on normal pancreatic epithelial (PE) cells. In the cell migration assay, arsenite clearly inhibited PDGF-BB-induced cell migration in AsPC-1 cells, but not in BxPC-3 or PE cells. Arsenite also caused cell apoptosis in AsPC-1 cells, but not in BxPC-3 or PE cells. In AsPC-1 cells, the levels of cyclin D1 and phosphorylated retinoblastoma protein decreased following treatment with arsenite, but this was not observed in BxPC-3 cells. To further examine the differences between these two cell lines, the effect of arsenite on upstream p44/p42 mitogen-activated protein kinase (MAPK) and Akt was investigated. PDGF-BB caused phosphorylation of p44/p42 MAPK and Akt in both cell lines. Pretreatment with arsenite significantly suppressed PDGF-BB-induced phosphorylation of Akt, but not of p44/p42 MAPK in AsPC-1 cells. By contrast, arsenite did not affect these molecules in BxPC-3 cells. Since the inhibition of the Akt signaling pathway markedly reduced PDGF-BB-induced migration in AsPC-1 cells, the present results strongly suggest that arsenite inhibits PDGF-BB-induced migration by suppressing the Akt signaling pathway in AsPC-1 cells. Therefore, arsenite may be a useful tool for the treatment of patients with certain types of pancreatic cancer, without causing adverse effects on normal pancreatic cells. PMID:27347121

  18. Capacity fade of Sony 18650 cells cycled at elevated temperatures. Part I. Cycling performance

    NASA Astrophysics Data System (ADS)

    Ramadass, P.; Haran, Bala; White, Ralph; Popov, Branko N.

    The capacity fade of Sony 18650 Li-ion cells increases with increase in temperature. After 800 cycles, the cells cycled at RT and 45 °C showed a capacity fade of 30 and 36%, respectively. The cell cycled at 55 °C showed a capacity loss of about 70% after 490 cycles. The rate capability of the cells continues to decrease with cycling. Impedance measurements showed an overall increase in the cell resistance with cycling and temperature. Impedance studies of the electrode materials showed an increased positive electrode resistance when compared to that of the negative electrode for cells cycled at RT and 45 °C. However, cells cycled at 50 and 55 °C exhibit higher negative electrode resistance. The increased capacity fade for the cells cycled at high temperatures can be explained by taking into account the repeated film formation over the surface of anode, which results in increased rate of lithium loss and also in a drastic increase in the negative electrode resistance with cycling.

  19. Activity Cycles in the Hyades and Praesepe

    NASA Astrophysics Data System (ADS)

    Baliunas, Sallie L.

    The giant stars in the Hyades present a well-studied group of stars of spectral type KO III. Their optical properties are quite similar, if not identical. All rotate with the same, slow period. Yet their chromospheric and coronal emission is different one from another, by as much as a factor of ten. We conjecture that this disparity results from sampling during different phases of long-term activity cycles which are present among dwarf stars. Some variation on a three-year timescale has been observed, as well as during phases of rotation modulation, however, at levels too small to explain the discrepancy of the emission strengths between the stars. We propose to investigate the range of chromospheric activity from these giants which are similar in the visible three ways: (a) reobserve the Hyades to search for variability on at least a seven-year timescale; (b) reobserve another young cluster, Praesepe, with four KO III stars similar to those in the Hyades to search for variability on a five-year timescale; (c) extend the sampling to four Hyades moving group stars with similar photospheric properties. The ultraviolet spectra provided by IUE represent the longest time frame, seven years, over which to search for long-term activity variations.

  20. Sphingosine-1-phosphate receptor 3 influences cell cycle progression in muscle satellite cells

    PubMed Central

    Fortier, Mathieu; Figeac, Nicolas; White, Robert B.; Knopp, Paul; Zammit, Peter S.

    2013-01-01

    Skeletal muscle retains a resident stem cell population called satellite cells, which are mitotically quiescent in mature muscle, but can be activated to produce myoblast progeny for muscle homeostasis, hypertrophy and repair. We have previously shown that satellite cell activation is partially controlled by the bioactive phospholipid, sphingosine-1-phosphate, and that S1P biosynthesis is required for muscle regeneration. Here we investigate the role of sphingosine-1-phosphate receptor 3 (S1PR3) in regulating murine satellite cell function. S1PR3 levels were high in quiescent myogenic cells before falling during entry into cell cycle. Retrovirally-mediated constitutive expression of S1PR3 led to suppressed cell cycle progression in satellite cells, but did not overtly affect the myogenic program. Conversely, satellite cells isolated from S1PR3-null mice exhibited enhanced proliferation ex-vivo. In vivo, acute cardiotoxin-induced muscle regeneration was enhanced in S1PR3-null mice, with bigger muscle fibres compared to control mice. Importantly, genetically deleting S1PR3 in the mdx mouse model of Duchenne muscular dystrophy produced a less severe muscle dystrophic phenotype, than when signalling though S1PR3 was operational. In conclusion, signalling though S1PR3 suppresses cell cycle progression to regulate function in muscle satellite cells. PMID:23911934

  1. Induction of cell cycle arrest in prostate cancer cells by the dietary compound isoliquiritigenin.

    PubMed

    Lee, Yeo Myeong; Lim, Do Young; Choi, Hyun Ju; Jung, Jae In; Chung, Won-Yoon; Park, Jung Han Yoon

    2009-02-01

    Isoliquiritigenin (ISL), a flavonoid chalcone that is present in licorice, shallot, and bean sprouts, is known to have antitumorigenic activities. The present study examined whether ISL alters prostate cancer cell cycle progression. DU145 human and MatLyLu (MLL) rat prostate cancer cells were cultured with various concentrations of ISL. In both DU145 and MLL cells treated with ISL, the percentage of cells in the G1 phase increased, and the incorporation of [(3)H]thymidine decreased. ISL decreased the protein levels of cyclin D1, cyclin E, and cyclin-dependent kinase (CDK) 4, whereas cyclin A and CDK2 expressions were unaltered in cells treated with ISL. The expression of the CDK inhibitor p27(KIP1) was increased in cells treated with 20 micromol/L ISL. In addition, treatment of cells with 20 micromol/L ISL for 24 hours led to G2/M cell cycle arrest. Cell division control (CDC) 2 protein levels remained unchanged. The protein levels of phospho-CDC2 (Tyr15) and cyclin B1 were increased, and the CDC25C level was decreased by ISL dose-dependently. We demonstrate that ISL promotes cell cycle arrest in DU145 and MLL cells, thereby providing insights into the mechanisms underlying its antitumorigenic activities. PMID:19298190

  2. Complex regulatory pathways coordinate cell cycle progression and development in Caulobacter crescentus

    PubMed Central

    Brown, Pamela J.B.; Hardy, Gail G.; Trimble, Michael J.; Brun, Yves V.

    2008-01-01

    Caulobacter crescentus has become the predominant bacterial model system to study the regulation of cell cycle progression. Stage specific processes such as chromosome replication and segregation, and cell division are coordinated with the development of four polar structures: the flagellum, pili, stalk, and holdfast. The production, activation, localization, and proteolysis of specific regulatory proteins at precise times during the cell cycle culminate in the ability of the cell to produce two physiologically distinct daughter cells. We examine the recent advances that have enhanced our understanding of the mechanisms of temporal and spatial regulation that occur during cell cycle progression. PMID:18929067

  3. p53 Promotes Cell Survival Due to the Reversibility of its Cell Cycle Checkpoints

    PubMed Central

    Lukin, Dana J.; Carvajal, Luis A.; Liu, Wen-jun; Resnick-Silverman, Lois; Manfredi, James J.

    2014-01-01

    The tumor suppressor p53 (TP53) has a well-studied role in triggering cell cycle checkpoint in response to DNA damage. Previous studies have suggested that functional p53 enhances chemosensitivity. In contrast, data are presented to show that p53 can be required for cell survival following DNA damage due to activation of reversible cell cycle checkpoints. The cellular outcome to DNA damage is determined by the duration and extent of the stimulus in a p53-dependent manner. In response to transient or low levels of DNA damage, p53 triggers a reversible G2 arrest whereas a sustained p53-dependent cell cycle arrest and senescence follows prolonged or high levels of DNA damage. Regardless of the length of treatment, p53-null cells arrest in G2, but ultimately adapt and proceed into mitosis. Interestingly, they fail to undergo cytokinesis, become multinucleated, and then die from apoptosis. Upon transient treatment with DNA damaging agents, wild-type p53 cells reversibly arrest and repair the damage, whereas p53-null cells fail to do so and die. These data indicate that p53 can promote cell survival by inducing reversible cell cycle arrest, thereby allowing for DNA repair. Thus, transient treatments may exploit differences between wild-type p53 and p53-null cells. PMID:25158956

  4. Programmed cell cycle arrest is required for infection of corn plants by the fungus Ustilago maydis.

    PubMed

    Castanheira, Sónia; Mielnichuk, Natalia; Pérez-Martín, José

    2014-12-01

    Ustilago maydis is a plant pathogen that requires a specific structure called infective filament to penetrate the plant tissue. Although able to grow, this filament is cell cycle arrested on the plant surface. This cell cycle arrest is released once the filament penetrates the plant tissue. The reasons and mechanisms for this cell cycle arrest are unknown. Here, we have tried to address these questions. We reached three conclusions from our studies. First, the observed cell cycle arrest is the result of the cooperation of at least two distinct mechanisms: one involving the activation of the DNA damage response (DDR) cascade; and the other relying on the transcriptional downregulation of Hsl1, a kinase that modulates the G2/M transition. Second, a sustained cell cycle arrest during the infective filament step is necessary for the virulence in U. maydis, as a strain unable to arrest the cell cycle was severely impaired in its ability to infect corn plants. Third, production of the appressorium, a structure required for plant penetration, is incompatible with an active cell cycle. The inability to infect plants by strains defective in cell cycle arrest seems to be caused by their failure to induce the appressorium formation process. In summary, our findings uncover genetic circuits to arrest the cell cycle during the growth of this fungus on the plant surface, thus allowing the penetration into plant tissue. PMID:25411209

  5. Effects of mistletoe (Viscum album L.) extracts Iscador on cell cycle and survival of tumor cells.

    PubMed

    Harmsma, Marjan; Ummelen, Monique; Dignef, Wendy; Tusenius, Karel Jan; Ramaekers, Frans C S

    2006-06-01

    The molecular and cellular mechanisms by which mistletoe (Viscum album L.) extracts exert cytotoxic and immunomodulatory anti-tumoral effects are largely unknown. In this study the hypothesis that Iscador preparations induce tumor regression by cell cycle inhibition and/or interference with apoptotic signaling pathways in cancer cells was investigated. Also a possible effect on angiogenesis, which is a prerequisite for tumor growth in vivo, is studied in endothelial cell cultures. Furthermore, it was examined which apoptotic signaling route(s) is (are) activated by Iscador by studying specific pro-apoptotic proteins in cultured cells. To characterize these properties, 9 human cancer cell lines of different origin, one epidermis derived cell line and 2 endothelial cell cultures were incubated with different concentrations of Iscador Quercus Spezial and Iscador Malus Spezial. Cell cycle kinetic parameters were measured by bromodeoxyuridine (BrdU) pulse labeling and tubulin staining. Apoptotic responses were detected by M30 Cyto-Death or Annexin V/propidium iodide assays. Characterization of the apoptotic pathway(s) was performed by staining cells for amongst others active caspase 3 and cytochrome C (mitochondrial pathway), as well as active caspase 8 (death receptor pathway). The sensitivity to Iscador treatment varies strongly between different cell lines and also ing those derived from small cell lung cancer, and adenocarcinoma of the lung and breast, as well as endothelial cell cultures, Iscador caused early cell cycle inhibition followed by apoptosis in a dose dependent manner. Amongst the low responders are cell lines derived from colorectal carcinoma. In general Iscador Malus exerted a stronger response than Iscador Quercus. Apoptosis was induced by activating the mitochondrial but not the death receptor dependent pathway, at least in case of Iscador Quercus. Iscador Malus also seemed to induce apoptosis via the death receptor route, which may explain the

  6. Linalool Induces Cell Cycle Arrest and Apoptosis in Leukemia Cells and Cervical Cancer Cells through CDKIs

    PubMed Central

    Chang, Mei-Yin; Shieh, Den-En; Chen, Chung-Chi; Yeh, Ching-Sheng; Dong, Huei-Ping

    2015-01-01

    Plantaginaceae, a popular traditional Chinese medicine, has long been used for treating various diseases from common cold to cancer. Linalool is one of the biologically active compounds that can be isolated from Plantaginaceae. Most of the commonly used cytotoxic anticancer drugs have been shown to induce apoptosis in susceptible tumor cells. However, the signaling pathway for apoptosis remains undefined. In this study, the cytotoxic effect of linalool on human cancer cell lines was investigated. Water-soluble tetrazolium salts (WST-1) based colorimetric cellular cytotoxicity assay, was used to test the cytotoxic ability of linalool against U937 and HeLa cells, and flow cytometry (FCM) and genechip analysis were used to investigate the possible mechanism of apoptosis. These results demonstrated that linalool exhibited a good cytotoxic effect on U937 and HeLa cells, with the IC50 value of 2.59 and 11.02 μM, respectively, compared with 5-FU with values of 4.86 and 12.31 μM, respectively. After treating U937 cells with linalool for 6 h, we found an increased sub-G1 peak and a dose-dependent phenomenon, whereby these cells were arrested at the G0/G1 phase. Furthermore, by using genechip analysis, we observed that linalool can promote p53, p21, p27, p16, and p18 gene expression. Therefore, this study verified that linalool can arrest the cell cycle of U937 cells at the G0/G1 phase and can arrest the cell cycle of HeLa cells at the G2/M phase. Its mechanism facilitates the expression of the cyclin-dependent kinases inhibitors (CDKIs) p53, p21, p27, p16, and p18, as well as the non-expression of cyclin-dependent kinases (CDKs) activity. PMID:26703569

  7. KOH concentration effect on cycle life of nickel-hydrogen cells. III - Cycle life test

    NASA Technical Reports Server (NTRS)

    Lim, H. S.; Verzwyvelt, S. A.

    1988-01-01

    A cycle life test of Ni/H2 cells containing electrolytes of various KOH concentrations and a sintered type nickel electrode was carried out at 23 C using a 45 min accelerated low earth orbit (LEO) cycle regime at 80 percent depth of discharge. One of three cells containing 26 percent KOH has achieved over 28,000 cycles, and the other two 19,000 cycles, without a sign of failure. Two other cells containing 31 percent KOH electrolyte, which is the concentration presently used in aerospace cells, failed after 2,979 and 3,620 cycles. This result indicates that the cycle life of the present type of Ni/H2 cells may be extended by a factor of 5 to 10 simply by lowering the KOH concentration. Long cycle life of a Ni/H2 battery at high depth-of-discharge operation is desired, particularly for an LEO spacecraft application. Typically, battery life of about 30,000 cycles is required for a five year mission in an LEO. Such a cycle life with presently available cells can be assured only at a very low depth-of-discharge operation. Results of testing already show that the cycle life of an Ni/H2 cell is tremendously improved by simply using an electrolyte of low KOH concentration.

  8. Centrosome/Cell Cycle Uncoupling and Elimination in the Endoreduplicating Intestinal Cells of C. elegans

    PubMed Central

    Lu, Yu; Roy, Richard

    2014-01-01

    The centrosome cycle is most often coordinated with mitotic cell division through the activity of various essential cell cycle regulators, consequently ensuring that the centriole is duplicated once, and only once, per cell cycle. However, this coupling can be altered in specific developmental contexts; for example, multi-ciliated cells generate hundreds of centrioles without any S-phase requirement for their biogenesis, while Drosophila follicle cells eliminate their centrosomes as they begin to endoreduplicate. In order to better understand how the centrosome cycle and the cell cycle are coordinated in a developmental context we use the endoreduplicating intestinal cell lineage of C. elegans to address how novel variations of the cell cycle impact this important process. In C. elegans, the larval intestinal cells undergo one nuclear division without subsequent cytokinesis, followed by four endocycles that are characterized by successive rounds of S-phase. We monitored the levels of centriolar/centrosomal markers and found that centrosomes lose their pericentriolar material following the nuclear division that occurs during the L1 stage and is thereafter never re-gained. The centrioles then become refractory to S phase regulators that would normally promote duplication during the first endocycle, after which they are eliminated during the L2 stage. Furthermore, we show that SPD-2 plays a central role in the numeral regulation of centrioles as a potential target of CDK activity. On the other hand, the phosphorylation on SPD-2 by Polo-like kinase, the transcriptional regulation of genes that affect centriole biogenesis, and the ubiquitin/proteasome degradation pathway, contribute collectively to the final elimination of the centrioles during the L2 stage. PMID:25360893

  9. Cell cycle regulation of a Xenopus Wee1-like kinase.

    PubMed Central

    Mueller, P R; Coleman, T R; Dunphy, W G

    1995-01-01

    Using a polymerase chain reaction-based strategy, we have isolated a gene encoding a Wee1-like kinase from Xenopus eggs. The recombinant Xenopus Wee1 protein efficiently phosphorylates Cdc2 exclusively on Tyr-15 in a cyclin-dependent manner. The addition of exogenous Wee1 protein to Xenopus cell cycle extracts results in a dose-dependent delay of mitotic initiation that is accompanied by enhanced tyrosine phosphorylation of Cdc2. The activity of the Wee1 protein is highly regulated during the cell cycle: the interphase, underphosphorylated form of Wee1 (68 kDa) phosphorylates Cdc2 very efficiently, whereas the mitotic, hyperphosphorylated version (75 kDa) is weakly active as a Cdc2-specific tyrosine kinase. The down-modulation of Wee1 at mitosis is directly attributable to phosphorylation, since dephosphorylation with protein phosphatase 2A restores its kinase activity. During interphase, the activity of this Wee1 homolog does not vary in response to the presence of unreplicated DNA. The mitosis-specific phosphorylation of Wee1 is due to at least two distinct kinases: the Cdc2 protein and another activity (kinase X) that may correspond to an MPM-2 epitope kinase. These studies indicate that the down-regulation of Wee1-like kinase activity at mitosis is a multistep process that occurs after other biochemical reactions have signaled the successful completion of S phase. Images PMID:7749193

  10. Ghrelin regulates cell cycle-related gene expression in cultured hippocampal neural stem cells.

    PubMed

    Chung, Hyunju; Park, Seungjoon

    2016-08-01

    We have previously demonstrated that ghrelin stimulates the cellular proliferation of cultured adult rat hippocampal neural stem cells (NSCs). However, little is known about the molecular mechanisms by which ghrelin regulates cell cycle progression. The purpose of this study was to investigate the potential effects of ghrelin on cell cycle regulatory molecules in cultured hippocampal NSCs. Ghrelin treatment increased proliferation assessed by CCK-8 proliferation assay. The expression levels of proliferating cell nuclear antigen and cell division control 2, well-known cell-proliferating markers, were also increased by ghrelin. Fluorescence-activated cell sorting analysis revealed that ghrelin promoted progression of cell cycle from G0/G1 to S phase, whereas this progression was attenuated by the pretreatment with specific inhibitors of MEK/extracellular signal-regulated kinase 1/2, phosphoinositide 3-kinase/Akt, mammalian target of rapamycin, and janus kinase 2/signal transducer and activator of transcription 3. Ghrelin-induced proliferative effect was associated with increased expression of E2F1 transcription factor in the nucleus, as determined by Western blotting and immunofluorescence. We also found that ghrelin caused an increase in protein levels of positive regulators of cell cycle, such as cyclin A and cyclin-dependent kinase (CDK) 2. Moreover, p27(KIP1) and p57(KIP2) protein levels were reduced when cell were exposed to ghrelin, suggesting downregulation of CDK inhibitors may contribute to proliferative effect of ghrelin. Our data suggest that ghrelin targets both cell cycle positive and negative regulators to stimulate proliferation of cultured hippocampal NSCs. PMID:27325242

  11. SUMOylation-mediated regulation of cell cycle progression and cancer

    PubMed Central

    Eifler, Karolin; Vertegaal, Alfred C.O.

    2016-01-01

    SUMOylation plays critical roles during cell cycle progression. Many important cell cycle regulators, including many oncogenes and tumor suppressors, are functionally regulated via SUMOylation. The dynamic SUMOylation pattern observed throughout the cell cycle is ensured via distinct spatial and temporal regulation of the SUMO machinery. Additionally, SUMOylation cooperates with other post-translational modifications to mediate cell cycle progression. Deregulation of these SUMOylation and deSUMOylation enzymes causes severe defects in cell proliferation and genome stability. Different types of cancers were recently shown to be dependent on a functioning SUMOylation system, a finding that could potentially be exploited in anti-cancer therapies. PMID:26601932

  12. A primer for studying cell cycle dynamics of the human hair follicle.

    PubMed

    Purba, Talveen S; Brunken, Lars; Hawkshaw, Nathan J; Peake, Michael; Hardman, Jonathan; Paus, Ralf

    2016-09-01

    The cell cycle is of major importance to human hair follicle (HF) biology. Not only is continuously active cell cycling required to facilitate healthy hair growth in anagen VI HFs, but perturbations in the cell cycle are likely to be of significance in HF pathology (i.e. in scarring, non-scarring, chemotherapy-induced and androgenic alopecias). However, cell cycle dynamics of the human hair follicle (HF) are poorly understood in contrast to what is known in mouse. The current Methods Review aims at helping to close this gap by presenting a primer that introduces immunohistological/immunofluorescent techniques to study the cell cycle in the human HF. Moreover, this primer encourages the exploitation of the human HF as a powerful and clinically relevant tool to investigate mammalian cell cycle biology in situ. To achieve this, we describe methods to study markers of general 'proliferation' (nuclei count, Ki-67 expression), apoptosis (terminal deoxynucleotidyl transferase dUTP nick-end labelling, cleaved caspase 3), mitosis (phospho-histone H3, 'pS780'), DNA synthesis (5-ethynyl-2'-deoxyuridine) and cell cycle regulation (cyclins) in the human HF. In addition, we provide specific examples of dual immunolabelling for instructive cell cycle analyses and for investigating the cell cycle behaviour of specific HF keratinocyte subpopulations, such as keratin 15+ stem/progenitor cells. PMID:27094702

  13. Gold nanoparticle sensitize radiotherapy of prostate cancer cells by regulation of the cell cycle

    NASA Astrophysics Data System (ADS)

    Roa, Wilson; Zhang, Xiaojing; Guo, Linghong; Shaw, Andrew; Hu, Xiuying; Xiong, Yeping; Gulavita, Sunil; Patel, Samir; Sun, Xuejun; Chen, Jie; Moore, Ronald; Xing, James Z.

    2009-09-01

    Glucose-capped gold nanoparticles (Glu-GNPs) have been used to improve cellular targeting and radio-sensitization. In this study, we explored the mechanism of Glu-GNP enhanced radiation sensitivity in radiation-resistant human prostate cancer cells. Cell survival and proliferation were measured using MTT and clonogenic assay. Flow cytometry with staining by propidium iodide (PI) was performed to study the cell cycle changes induced by Glu-GNPs, and western blotting was used to determine the expression of p53 and cyclin proteins that correlated to cell cycle regulation. With 2 Gy of ortho-voltage irradiation, Glu-GNP showed a 1.5-2.0 fold enhancement in growth inhibition when compared to x-rays alone. Comparing the cell cycle change, Glu-GNPs induced acceleration in the G0/G1 phase and accumulation of cells in the G2/M phase at 29.8% versus 18.4% for controls at 24 h. G2/M arrest was accompanied by decreased expression of p53 and cyclin A, and increased expression of cyclin B1 and cyclin E. In conclusion, Glu-GNPs trigger activation of the CDK kinases leading to cell cycle acceleration in the G0/G1 phase and accumulation in the G2/M phase. This activation is accompanied by a striking sensitization to ionizing radiation, which may have clinical implications.

  14. A decrease in solar and geomagnetic activity from cycle 19 to cycle 24

    NASA Astrophysics Data System (ADS)

    Gvishiani, A. D.; Starostenko, V. I.; Sumaruk, Yu. P.; Soloviev, A. A.; Legostaeva, O. V.

    2015-05-01

    Variations in the solar and geomagnetic activity from cycle 19 to cycle 24 were considered based on data from the magnetic observatories of the Russian-Ukrainian INTERMAGNET segment and international centers of data on solar-terrestrial physics. It has been indicated that activity decreases over the course of time. This is especially evident during the cycle 24 growth phase. The possible causes and consequences of a decrease in geomagnetic activity were analyzed.

  15. A stochastic spatiotemporal model of a response-regulator network in the Caulobacter crescentus cell cycle

    NASA Astrophysics Data System (ADS)

    Li, Fei; Subramanian, Kartik; Chen, Minghan; Tyson, John J.; Cao, Yang

    2016-06-01

    The asymmetric cell division cycle in Caulobacter crescentus is controlled by an elaborate molecular mechanism governing the production, activation and spatial localization of a host of interacting proteins. In previous work, we proposed a deterministic mathematical model for the spatiotemporal dynamics of six major regulatory proteins. In this paper, we study a stochastic version of the model, which takes into account molecular fluctuations of these regulatory proteins in space and time during early stages of the cell cycle of wild-type Caulobacter cells. We test the stochastic model with regard to experimental observations of increased variability of cycle time in cells depleted of the divJ gene product. The deterministic model predicts that overexpression of the divK gene blocks cell cycle progression in the stalked stage; however, stochastic simulations suggest that a small fraction of the mutants cells do complete the cell cycle normally.

  16. Monoterpenes inhibit cell growth, cell cycle progression, and cyclin D1 gene expression in human breast cancer cell lines.

    PubMed

    Bardon, S; Picard, K; Martel, P

    1998-01-01

    Monoterpenes are found in the essential oils of many commonly consumed fruits and vegetables. These compounds have been shown to exert chemopreventive and chemotherapeutic activities in mammary tumor models and represent a new class of breast cancer therapeutic agents. In this study, we investigated the effects of limonene and limonene-related monoterpenes, perillyl alcohol and perillic acid, on cell growth, cell cycle progression, and expression of cyclin D1 cell cycle-regulatory gene in T-47D, MCF-7, and MDA-MB-231 breast cancer cell lines. Our results revealed that limonene-related monoterpenes caused a dose-dependent inhibition of cell proliferation. Of the three monoterpenes tested, perillyl alcohol was the most potent and limonene was the least potent inhibitor of cell growth. The enantiomeric composition of limonene and perillyl alcohol did not interfere with their effect on cell growth. Sensitivity of breast cancer cell lines to monoterpenes was in the following order: T-47D > MCF-7 > MDA-MB-231. Growth inhibition induced by perillyl alcohol and perillic acid was associated with a fall in the proportion of cells in the S phase and an accumulation of cells in the G1 phase of the cell cycle. Finally, we showed that the effects of limonene-related monoterpenes on cell proliferation and cell cycle progression were preceded by a decrease in cyclin D1 mRNA levels. PMID:9824849

  17. Two ZNF509 (ZBTB49) isoforms induce cell-cycle arrest by activating transcription of p21/CDKN1A and RB upon exposure to genotoxic stress

    PubMed Central

    Jeon, Bu-Nam; Kim, Min-Kyeong; Yoon, Jae-Hyeon; Kim, Min-Young; An, Haemin; Noh, Hee-Jin; Choi, Won-Il; Koh, Dong-In; Hur, Man-Wook

    2014-01-01

    ZNF509 is unique among POK family proteins in that four isoforms are generated by alternative splicing. Short ZNF509 (ZNF509S1, -S2 and -S3) isoforms contain one or two out of the seven zinc-fingers contained in long ZNF509 (ZNF509L). Here, we investigated the functions of ZNF509 isoforms in response to DNA damage, showing isoforms to be induced by p53. Intriguingly, to inhibit proliferation of HCT116 and HEK293 cells, we found that ZNF509L activates p21/CDKN1A transcription, while ZNF509S1 induces RB. ZNF509L binds to the p21/CDKN1A promoter either alone or by interacting with MIZ-1 to recruit the co-activator p300 to activate p21/CDKN1A transcription. In contrast, ZNF509S1 binds to the distal RB promoter to interact and interfere with the MIZF repressor, resulting in derepression and transcription of RB. Immunohistochemical analysis revealed that ZNF509 is highly expressed in normal epithelial cells, but was completely repressed in tumor tissues of the colon, lung and skin, indicating a possible role as a tumor suppressor. PMID:25245946

  18. Indirect-fired gas turbine dual fuel cell power cycle

    DOEpatents

    Micheli, Paul L.; Williams, Mark C.; Sudhoff, Frederick A.

    1996-01-01

    A fuel cell and gas turbine combined cycle system which includes dual fuel cell cycles combined with a gas turbine cycle wherein a solid oxide fuel cell cycle operated at a pressure of between 6 to 15 atms tops the turbine cycle and is used to produce CO.sub.2 for a molten carbonate fuel cell cycle which bottoms the turbine and is operated at essentially atmospheric pressure. A high pressure combustor is used to combust the excess fuel from the topping fuel cell cycle to further heat the pressurized gas driving the turbine. A low pressure combustor is used to combust the excess fuel from the bottoming fuel cell to reheat the gas stream passing out of the turbine which is used to preheat the pressurized air stream entering the topping fuel cell before passing into the bottoming fuel cell cathode. The CO.sub.2 generated in the solid oxide fuel cell cycle cascades through the system to the molten carbonate fuel cell cycle cathode.

  19. Loratadine dysregulates cell cycle progression and enhances the effect of radiation in human tumor cell lines

    PubMed Central

    2010-01-01

    Background The histamine receptor-1 (H1)-antagonist, loratadine has been shown to inhibit growth of human colon cancer xenografts in part due to cell cycle arrest in G2/M. Since this is a radiation sensitive phase of the cell cycle, we sought to determine if loratadine modifies radiosensitivity in several human tumor cell lines with emphasis on human colon carcinoma (HT29). Methods Cells were treated with several doses of loratadine at several time points before and after exposure to radiation. Radiation dose modifying factors (DMF) were determined using full radiation dose response survival curves. Cell cycle phase was determined by flow cytometry and the expression of the cell cycle-associated proteins Chk1, pChk1ser345, and Cyclin B was analyzed by western blot. Results Loratadine pre-treatment of exponentially growing cells (75 μM, 24 hours) increased radiation-induced cytotoxicity yielding a radiation DMF of 1.95. However, treatment of plateau phase cells also yielded a DMF of 1.3 suggesting that mechanisms other than cell cycle arrest also contribute to loratadine-mediated radiation modification. Like irradiation, loratadine initially induced G2/M arrest and activation of the cell-cycle associated protein Chk1 to pChk1ser345, however a subsequent decrease in expression of total Chk1 and Cyclin B correlated with abrogation of the G2/M checkpoint. Analysis of DNA repair enzyme expression and DNA fragmentation revealed a distinct pattern of DNA damage in loratadine-treated cells in addition to enhanced radiation-induced damage. Taken together, these data suggest that the observed effects of loratadine are multifactorial in that loratadine 1) directly damages DNA, 2) activates Chk1 thereby promoting G2/M arrest making cells more susceptible to radiation-induced DNA damage and, 3) downregulates total Chk1 and Cyclin B abrogating the radiation-induced G2/M checkpoint and allowing cells to re-enter the cell cycle despite the persistence of damaged DNA. Conclusions

  20. A prediction of geomagnetic activity for solar cycle 23

    NASA Astrophysics Data System (ADS)

    Cliver, E. W.; Ling, A. G.; Wise, J. E.; Lanzerotti, L. J.

    1999-04-01

    Using a database of 13 solar cycles of geomagnetic aa data, we obtained correlations between cycle averages of geomagnetic activity (and sunspot number) and the numbers of days with disturbance levels above certain aa thresholds. We then used a precursor-type relation to predict an average aa index of 23.1 nT for cycle 23 and inserted this average aa value into the above correlations to forecast the integral size distribution of geomagnetic activity for the new cycle. The predicted size distribution is similar to that observed for cycles 21 and 22 but most closely resembles that of solar cycle 18 (1944-1954), which was slightly smaller than cycles 21 and 22. Our prediction agrees reasonably well with the ``climatology-based'' forecast made by the intergovernmental panel tasked to predict geomagnetic activity for the coming solar cycle and is significantly different from their ``precursor-based'' prediction.

  1. Tudor Staphylococcal Nuclease (Tudor-SN), a Novel Regulator Facilitating G1/S Phase Transition, Acting as a Co-activator of E2F-1 in Cell Cycle Regulation*

    PubMed Central

    Su, Chao; Zhang, Chunyan; Tecle, Adiam; Fu, Xue; He, Jinyan; Song, Juan; Zhang, Wei; Sun, Xiaoming; Ren, Yuanyuan; Silvennoinen, Olli; Yao, Zhi; Yang, Xi; Wei, Minxin; Yang, Jie

    2015-01-01

    Tudor staphylococcal nuclease (Tudor-SN) is a multifunctional protein implicated in a variety of cellular processes. In the present study, we identified Tudor-SN as a novel regulator in cell cycle. Tudor-SN was abundant in proliferating cells whereas barely expressed in terminally differentiated cells. Functional analysis indicated that ectopic overexpression of Tudor-SN promoted the G1/S transition, whereas knockdown of Tudor-SN caused G1 arrest. Moreover, the live-cell time-lapse experiment demonstrated that the cell cycle of MEF−/− (knock-out of Tudor-SN in mouse embryonic fibroblasts) was prolonged compared with wild-type MEF+/+. We noticed that Tudor-SN was constantly expressed in every cell cycle phase, but was highly phosphorylated in the G1/S border. Further study revealed that Tudor-SN was a potential substrate of Cdk2/4/6, supportively, we found the physical interaction of endogenous Tudor-SN with Cdk4/6 in G1 and the G1/S border, and with Cdk2 in the G1/S border and S phase. In addition, roscovitine (Cdk1/2/5 inhibitor) or CINK4 (Cdk4/6 inhibitor) could inhibit the phosphorylation of Tudor-SN, whereas ectopic overexpression of Cdk2/4/6 increased the Tudor-SN phosphorylation. The underlying molecular mechanisms indicated that Tudor-SN could physically interact with E2F-1 in vivo, and could enhance the physical association of E2F-1 with GCN5 (a cofactor of E2F-1, which possesses histone acetyltransferase activity), and promote the binding ability of E2F-1 to the promoter region of its target genes CYCLIN A and E2F-1, and as a result, facilitate the gene transcriptional activation. Taken together, Tudor-SN is identified as a novel co-activator of E2F-1, which could facilitate E2F-1-mediated gene transcriptional activation of target genes, which play essential roles in G1/S transition. PMID:25627688

  2. Morphogenesis checkpoint kinase Swe1 is the executor of lipolysis-dependent cell-cycle progression.

    PubMed

    Chauhan, Neha; Visram, Myriam; Cristobal-Sarramian, Alvaro; Sarkleti, Florian; Kohlwein, Sepp D

    2015-03-10

    Cell growth and division requires the precise duplication of cellular DNA content but also of membranes and organelles. Knowledge about the cell-cycle-dependent regulation of membrane and storage lipid homeostasis is only rudimentary. Previous work from our laboratory has shown that the breakdown of triacylglycerols (TGs) is regulated in a cell-cycle-dependent manner, by activation of the Tgl4 lipase by the major cyclin-dependent kinase Cdc28. The lipases Tgl3 and Tgl4 are required for efficient cell-cycle progression during the G1/S (Gap1/replication phase) transition, at the onset of bud formation, and their absence leads to a cell-cycle delay. We now show that defective lipolysis activates the Swe1 morphogenesis checkpoint kinase that halts cell-cycle progression by phosphorylation of Cdc28 at tyrosine residue 19. Saturated long-chain fatty acids and phytosphingosine supplementation rescue the cell-cycle delay in the Tgl3/Tgl4 lipase-deficient strain, suggesting that Swe1 activity responds to imbalanced sphingolipid metabolism, in the absence of TG degradation. We propose a model by which TG-derived sphingolipids are required to activate the protein phosphatase 2A (PP2A(Cdc55)) to attenuate Swe1 phosphorylation and its inhibitory effect on Cdc28 at the G1/S transition of the cell cycle. PMID:25713391

  3. Graded requirement for the spliceosome in cell cycle progression

    PubMed Central

    Karamysheva, Zemfira; Díaz-Martínez, Laura A; Warrington, Ross; Yu, Hongtao

    2015-01-01

    Genome stability is ensured by multiple surveillance mechanisms that monitor the duplication, segregation, and integrity of the genome throughout the cell cycle. Depletion of components of the spliceosome, a macromolecular machine essential for mRNA maturation and gene expression, has been associated with increased DNA damage and cell cycle defects. However, the specific role for the spliceosome in these processes has remained elusive, as different cell cycle defects have been reported depending on the specific spliceosome subunit depleted. Through a detailed cell cycle analysis after spliceosome depletion, we demonstrate that the spliceosome is required for progression through multiple phases of the cell cycle. Strikingly, the specific cell cycle phenotype observed after spliceosome depletion correlates with the extent of depletion. Partial depletion of a core spliceosome component results in defects at later stages of the cell cycle (G2 and mitosis), whereas a more complete depletion of the same component elicits an early cell cycle arrest in G1. We propose a quantitative model in which different functional dosages of the spliceosome are required for different cell cycle transitions. PMID:25892155

  4. Intermittent Stem Cell Cycling Balances Self-Renewal and Senescence of the C. elegans Germ Line.

    PubMed

    Cinquin, Amanda; Chiang, Michael; Paz, Adrian; Hallman, Sam; Yuan, Oliver; Vysniauskaite, Indre; Fowlkes, Charless C; Cinquin, Olivier

    2016-04-01

    Self-renewing organs often experience a decline in function in the course of aging. It is unclear whether chronological age or external factors control this decline, or whether it is driven by stem cell self-renewal-for example, because cycling cells exhaust their replicative capacity and become senescent. Here we assay the relationship between stem cell cycling and senescence in the Caenorhabditis elegans reproductive system, defining this senescence as the progressive decline in "reproductive capacity," i.e. in the number of progeny that can be produced until cessation of reproduction. We show that stem cell cycling diminishes remaining reproductive capacity, at least in part through the DNA damage response. Paradoxically, gonads kept under conditions that preclude reproduction keep cycling and producing cells that undergo apoptosis or are laid as unfertilized gametes, thus squandering reproductive capacity. We show that continued activity is in fact beneficial inasmuch as gonads that are active when reproduction is initiated have more sustained early progeny production. Intriguingly, continued cycling is intermittent-gonads switch between active and dormant states-and in all likelihood stochastic. Other organs face tradeoffs whereby stem cell cycling has the beneficial effect of providing freshly-differentiated cells and the detrimental effect of increasing the likelihood of cancer or senescence; stochastic stem cell cycling may allow for a subset of cells to preserve proliferative potential in old age, which may implement a strategy to deal with uncertainty as to the total amount of proliferation to be undergone over an organism's lifespan. PMID:27077385

  5. The Cell Cycle Timing of Human Papillomavirus DNA Replication

    PubMed Central

    Reinson, Tormi; Henno, Liisi; Toots, Mart; Ustav, Mart; Ustav, Mart

    2015-01-01

    Viruses manipulate the cell cycle of the host cell to optimize conditions for more efficient viral genome replication. One strategy utilized by DNA viruses is to replicate their genomes non-concurrently with the host genome; in this case, the viral genome is amplified outside S phase. This phenomenon has also been described for human papillomavirus (HPV) vegetative genome replication, which occurs in G2-arrested cells; however, the precise timing of viral DNA replication during initial and stable replication phases has not been studied. We developed a new method to quantitate newly synthesized DNA levels and used this method in combination with cell cycle synchronization to show that viral DNA replication is initiated during S phase and is extended to G2 during initial amplification but follows the replication pattern of cellular DNA during S phase in the stable maintenance phase. E1 and E2 protein overexpression changes the replication time from S only to both the S and G2 phases in cells that stably maintain viral episomes. These data demonstrate that the active synthesis and replication of the HPV genome are extended into the G2 phase to amplify its copy number and the duration of HPV genome replication is controlled by the level of the viral replication proteins E1 and E2. Using the G2 phase for genome amplification may be an important adaptation that allows exploitation of changing cellular conditions during cell cycle progression. We also describe a new method to quantify newly synthesized viral DNA levels and discuss its benefits for HPV research. PMID:26132923

  6. Cell cycle and centromere FISH studies in premature centromere division

    PubMed Central

    Corona-Rivera, Alfredo; Salamanca-Gomez, Fabio; Bobadilla-Morales, Lucina; Corona-Rivera, Jorge R; Palomino-Cueva, Cesar; Garcia-Cobian, Teresa A; Corona-Rivera, Enrique

    2005-01-01

    Background Mitotic configurations consistent in split centromeres and splayed chromatids in all or most of the chromosomes or premature centromere division (PCD) have been described in three categories. (1) Low frequency of PCD observed in colchicines-treated lymphocyte cultures from normal individuals. (2) High frequency of PCD with mosaic variegated aneuploidy. (3) High frequency of PCD as a sole chromosome abnormality observed in individuals with no recognizable clinical pattern. We report four members of a family with the third category of PCD. Methods Cell cycle duration assessed by average generation time using differential sister chromatid stain analysis and FISH studies of DNA centromere sequences in PCD individuals, are included and compared with previously reported PCD individuals from 9 families. Results We observed PCD in colchicine-treated cultures from the propositus, his father, and two paternal aunts but not in his mother and four other paternal and maternal family members, as well as in untreated cultures from the propositus and his father. We observed cytological evidence of active centromeres by Cd stain. Significative cell cycle time reduction in anaphases of PCD individuals (average generation time of 21.8 h;SD 0.4) with respect to individuals without PCD (average generation time of 31.8 h;SD 3.9) was observed (P < 0.005, Student t-test for independent samples). Increased cell proliferation kinetics was observed in anaphasic cells of individuals with PCD, by differential sister chromatid stain analysis. FISH studies revealed the presence of alpha satellite DNA from chromosomes 1, 13, 21/18, X, all centromeres, and CENP-B box sequences in metaphasic and anaphasic cells from PCD individuals. Conclusion This report examines evidences of a functional relationship between PCD and cell cycle impairment. It seems that essential centromere integrity is present in these cases. PMID:16174301

  7. The Cell Cycle Timing of Human Papillomavirus DNA Replication.

    PubMed

    Reinson, Tormi; Henno, Liisi; Toots, Mart; Ustav, Mart; Ustav, Mart

    2015-01-01

    Viruses manipulate the cell cycle of the host cell to optimize conditions for more efficient viral genome replication. One strategy utilized by DNA viruses is to replicate their genomes non-concurrently with the host genome; in this case, the viral genome is amplified outside S phase. This phenomenon has also been described for human papillomavirus (HPV) vegetative genome replication, which occurs in G2-arrested cells; however, the precise timing of viral DNA replication during initial and stable replication phases has not been studied. We developed a new method to quantitate newly synthesized DNA levels and used this method in combination with cell cycle synchronization to show that viral DNA replication is initiated during S phase and is extended to G2 during initial amplification but follows the replication pattern of cellular DNA during S phase in the stable maintenance phase. E1 and E2 protein overexpression changes the replication time from S only to both the S and G2 phases in cells that stably maintain viral episomes. These data demonstrate that the active synthesis and replication of the HPV genome are extended into the G2 phase to amplify its copy number and the duration of HPV genome replication is controlled by the level of the viral replication proteins E1 and E2. Using the G2 phase for genome amplification may be an important adaptation that allows exploitation of changing cellular conditions during cell cycle progression. We also describe a new method to quantify newly synthesized viral DNA levels and discuss its benefits for HPV research. PMID:26132923

  8. The Analysis of Cell Cycle, Proliferation, and Asymmetric Cell Division by Imaging Flow Cytometry.

    PubMed

    Filby, Andrew; Day, William; Purewal, Sukhveer; Martinez-Martin, Nuria

    2016-01-01

    Measuring cellular DNA content by conventional flow cytometry (CFC) and fluorescent DNA-binding dyes is a highly robust method for analysing cell cycle distributions within heterogeneous populations. However, any conclusions drawn from single-parameter DNA analysis alone can often be confounded by the asynchronous nature of cell proliferation. We have shown that by combining fluorescent DNA stains with proliferation tracking dyes and antigenic staining for mitotic cells one can elucidate the division history and cell cycle position of any cell within an asynchronously dividing population. Furthermore if one applies this panel to an imaging flow cytometry (IFC) system then the spatial information allows resolution of the four main mitotic phases and the ability to study molecular distributions within these populations. We have employed such an approach to study the prevalence of asymmetric cell division (ACD) within activated immune cells by measuring the distribution of key fate determining molecules across the plane of cytokinesis in a high-throughput, objective, and internally controlled manner. Moreover the ability to perform high-resolution, temporal dissection of the cell division process lends itself perfectly to investigating the influence chemotherapeutic agents exert on the proliferative capacity of transformed cell lines. Here we describe the method in detail and its application to both ACD and general cell cycle analysis. PMID:27460238

  9. HIV protease inhibitor nelfinavir inhibits growth of human melanoma cells by induction of cell cycle arrest.

    PubMed

    Jiang, Wei; Mikochik, Peter J; Ra, Jin H; Lei, Hanqin; Flaherty, Keith T; Winkler, Jeffrey D; Spitz, Francis R

    2007-02-01

    HIV protease inhibitors (HIV PI) are a class of antiretroviral drugs that are designed to target the viral protease. Unexpectedly, this class of drugs is also reported to have antitumor activity. In this study, we have evaluated the in vitro activity of nelfinavir, a HIV PI, against human melanoma cells. Nelfinavir inhibits the growth of melanoma cell lines at low micromolar concentrations that are clinically attainable. Nelfinavir promotes apoptosis and arrests cell cycle at G(1) phase. Cell cycle arrest is attributed to inhibition of cyclin-dependent kinase 2 (CDK2) and concomitant dephosphorylation of retinoblastoma tumor suppressor. We further show that nelfinavir inhibits CDK2 through proteasome-dependent degradation of Cdc25A phosphatase. Our results suggest that nelfinavir is a promising candidate chemotherapeutic agent for advanced melanoma, for which novel and effective therapies are urgently needed. PMID:17283158

  10. Single-cell analysis of transcription kinetics across the cell cycle.

    PubMed

    Skinner, Samuel O; Xu, Heng; Nagarkar-Jaiswal, Sonal; Freire, Pablo R; Zwaka, Thomas P; Golding, Ido

    2016-01-01

    Transcription is a highly stochastic process. To infer transcription kinetics for a gene-of-interest, researchers commonly compare the distribution of mRNA copy-number to the prediction of a theoretical model. However, the reliability of this procedure is limited because the measured mRNA numbers represent integration over the mRNA lifetime, contribution from multiple gene copies, and mixing of cells from different cell-cycle phases. We address these limitations by simultaneously quantifying nascent and mature mRNA in individual cells, and incorporating cell-cycle effects in the analysis of mRNA statistics. We demonstrate our approach on Oct4 and Nanog in mouse embryonic stem cells. Both genes follow similar two-state kinetics. However, Nanog exhibits slower ON/OFF switching, resulting in increased cell-to-cell variability in mRNA levels. Early in the cell cycle, the two copies of each gene exhibit independent activity. After gene replication, the probability of each gene copy to be active diminishes, resulting in dosage compensation. PMID:26824388

  11. Cell cycle control of polyomavirus-induced transformation.

    PubMed Central

    Chen, H H; Fluck, M M

    1993-01-01

    The cell cycle dependence of polyomavirus transformation was analyzed in infections of nonpermissive Fischer rat (FR3T3) cells released from G0. A 5- to 100-fold (average, ca. 20-fold) difference in relative frequency of transformation was found for cells infected in the early G1 phase of the cell cycle compared with cells infected in G2. Differences in the relative level of early viral gene expression in those two cell populations were equivalent to those obtained for transformation frequencies. The difference in transformation potential was accounted for only in part by a cell cycle control of viral adsorption (2- to 15-fold effect). Furthermore, in cells infected in the early G1 phase, viral gene expression was induced as a big synchronous burst of large transcripts of variable sizes, delayed till the G1 phase of the cell cycle after that in which infection took place. Thus, the results demonstrate that the abortive infection cycle of G0-released FR3T3 cells is cell cycle regulated at least at two steps: adsorption and another early step, nuclear transport, decapsidation, up to or including the transcription of the viral early genes. The cell cycle regulation of these steps results in a similar regulation of the abortive and stable transformation processes, although it is more pronounced for the latter. A model implicating c-fos and c-jun is proposed. Images PMID:8383223

  12. Cell shape, cytoskeletal mechanics, and cell cycle control in angiogenesis

    NASA Technical Reports Server (NTRS)

    Ingber, D. E.; Prusty, D.; Sun, Z.; Betensky, H.; Wang, N.

    1995-01-01

    Capillary endothelial cells can be switched between growth and differentiation by altering cell-extracellular matrix interactions and thereby, modulating cell shape. Studies were carried out to determine when cell shape exerts its growth-regulatory influence during cell cycle progression and to explore the role of cytoskeletal structure and mechanics in this control mechanism. When G0-synchronized cells were cultured in basic fibroblast growth factor (FGF)-containing defined medium on dishes coated with increasing densities of fibronectin or a synthetic integrin ligand (RGD-containing peptide), cell spreading, nuclear extension, and DNA synthesis all increased in parallel. To determine the minimum time cells must be adherent and spread on extracellular matrix (ECM) to gain entry into S phase, cells were removed with trypsin or induced to retract using cytochalasin D at different times after plating. Both approaches revealed that cells must remain extended for approximately 12-15 h and hence, most of G1, in order to enter S phase. After this restriction point was passed, normally 'anchorage-dependent' endothelial cells turned on DNA synthesis even when round and in suspension. The importance of actin-containing microfilaments in shape-dependent growth control was confirmed by culturing cells in the presence of cytochalasin D (25-1000 ng ml-1): dose-dependent inhibition of cell spreading, nuclear extension, and DNA synthesis resulted. In contrast, induction of microtubule disassembly using nocodazole had little effect on cell or nuclear spreading and only partially inhibited DNA synthesis. Interestingly, combination of nocodazole with a suboptimal dose of cytochalasin D (100 ng ml-1) resulted in potent inhibition of both spreading and growth, suggesting that microtubules are redundant structural elements which can provide critical load-bearing functions when microfilaments are partially compromised. Similar synergism between nocodazole and cytochalasin D was observed

  13. Differential expression of cell cycle regulators in CDK5-dependent medullary thyroid carcinoma tumorigenesis.

    PubMed

    Pozo, Karine; Hillmann, Antje; Augustyn, Alexander; Plattner, Florian; Hai, Tao; Singh, Tanvir; Ramezani, Saleh; Sun, Xiankai; Pfragner, Roswitha; Minna, John D; Cote, Gilbert J; Chen, Herbert; Bibb, James A; Nwariaku, Fiemu E

    2015-05-20

    Medullary thyroid carcinoma (MTC) is a neuroendocrine cancer of thyroid C-cells, for which few treatment options are available. We have recently reported a role for cyclin-dependent kinase 5 (CDK5) in MTC pathogenesis. We have generated a mouse model, in which MTC proliferation is induced upon conditional overexpression of the CDK5 activator, p25, in C-cells, and arrested by interrupting p25 overexpression. Here, we identify genes and proteins that are differentially expressed in proliferating versus arrested benign mouse MTC. We find that downstream target genes of the tumor suppressor, retinoblastoma protein, including genes encoding cell cycle regulators such as CDKs, cyclins and CDK inhibitors, are significantly upregulated in malignant mouse tumors in a CDK5-dependent manner. Reducing CDK5 activity in human MTC cells down-regulated these cell cycle regulators suggesting that CDK5 activity is critical for cell cycle progression and MTC proliferation. Finally, the same set of cell cycle proteins was consistently overexpressed in human sporadic MTC but not in hereditary MTC. Together these findings suggest that aberrant CDK5 activity precedes cell cycle initiation and thus may function as a tumor-promoting factor facilitating cell cycle protein expression in MTC. Targeting aberrant CDK5 or its downstream effectors may be a strategy to halt MTC tumorigenesis. PMID:25900242

  14. Solar cell activation system

    SciTech Connect

    Apelian, L.

    1983-07-05

    A system for activating solar cells involves the use of phosphorescent paint, the light from which is amplified by a thin magnifying lens and used to activate solar cells. In a typical system, a member painted with phosphorescent paint is mounted adjacent a thin magnifying lens which focuses the light on a predetermined array of sensitive cells such as selenium, cadmium or silicon, mounted on a plastic board. A one-sided mirror is mounted adjacent the cells to reflect the light back onto said cells for purposes of further intensification. The cells may be coupled to rechargeable batteries or used to directly power a small radio or watch.

  15. Influence of lycopene on cell viability, cell cycle, and apoptosis of human prostate cancer and benign hyperplastic cells.

    PubMed

    Soares, Nathalia da Costa Pereira; Teodoro, Anderson Junger; Oliveira, Felipe Leite; Santos, Carlos Antonio do Nascimento; Takiya, Christina Maeda; Junior, Oswaldo Saback; Bianco, Mario; Junior, Antonio Palumbo; Nasciutti, Luiz Eurico; Ferreira, Luciana Bueno; Gimba, Etel Rodrigues Pereira; Borojevic, Radovan

    2013-01-01

    Prostate cancer is the most common malignancy in men and the second leading cause of cancer-related mortality in men of the Western world. Lycopene has received attention because of its expcted potential to prevent cancer. In the present study, we evaluated the influence of lycopene on cell viability, cell cycle, and apoptosis of human prostate cancer cells and benign prostate hyperplastic cells. Using MTT assay, we observed a decrease of cell viability in all cancer cell lines after treatment with lycopene, which decreased the percentage of cells in G0/G1 phase and increased in S and G2/M phases after 96 h of treatment in metastatic prostate cancer cell lineages. Flow citometry analysis of cell cycle revealed lycopene promoted cell cycle arrest in G0/G1 phase after 48 and 96 h of treatment in a primary cancer cell line. Using real time PCR assay, lycopene also induced apoptosis in prostate cancer cells with altered gene expression of Bax and Bcl-2. No effect was observed in benign prostate hyperplasia cells. These results suggest an effect of lycopene on activity of human prostate cancer cells. PMID:24053141

  16. PGC-1α regulates the cell cycle through ATP and ROS in CH1 cells* #

    PubMed Central

    Fu, Xu-feng; Yao, Kun; Du, Xing; Li, Yan; Yang, Xiu-yu; Yu, Min; Li, Mei-zhang; Cui, Qing-hua

    2016-01-01

    Peroxisome proliferator-activated receptor-γ coactivator 1α (PGC-1α) is a transcriptional co-activator involved in mitochondrial biogenesis, respiratory capacity, and oxidative phosphorylation (OXPHOS). PGC-1α plays an important role in cellular metabolism and is associated with tumorigenesis, suggesting an involvement in cell cycle progression. However, the underlying mechanisms mediating its involvement in these processes remain unclear. To elucidate the signaling pathways involved in PGC-1α function, we established a cell line, CH1 PGC-1α, which stably overexpresses PGC-1α. Using this cell line, we found that over-expression of PGC-1α stimulated extra adenosine triphosphate (ATP) and reduced reactive oxygen species (ROS) production. These effects were accompanied by up-regulation of the cell cycle checkpoint regulators CyclinD1 and CyclinB1. We hypothesized that ATP and ROS function as cellular signals to regulate cyclins and control cell cycle progression. Indeed, we found that reduction of ATP levels down-regulated CyclinD1 but not CyclinB1, whereas elevation of ROS levels down-regulated CyclinB1 but not CyclinD1. Furthermore, both low ATP levels and elevated ROS levels inhibited cell growth, but PGC-1α was maintained at a constant level. Together, these results demonstrate that PGC-1α regulates cell cycle progression through modulation of CyclinD1 and CyclinB1 by ATP and ROS. These findings suggest that PGC-1α potentially coordinates energy metabolism together with the cell cycle. PMID:26834014

  17. Effects of flavonoids on the growth and cell cycle of cancer cells.

    PubMed

    Choi, S U; Ryu, S Y; Yoon, S K; Jung, N P; Park, S H; Kim, K H; Choi, E J; Lee, C O

    1999-01-01

    In this study, we investigated the cytotoxicities of flavone (F01), 3-hydroxyflavone (F02), 6- hydroxyflavone (F03), 7-hydroxyflavone (F04), 3,6-dihydroxyflavone (F05), 5,7-dihydroxyflavone (F06) and 5,6,7-trihydroxyflavone (F07) to human cancer cells including P- glycoprotein (Pgp)-expressing HCT15 cells and its multidrug resistant subline, HCT15/CL02 cells. We also examined the effects of those flavonoids on the cell cycle of these cancer cells. HCT15/CL02 cells did not reveal resistance to all the flavonoids tested in comparison with HCT15 cells. In cell cycle analysis, all the flavonoids tested, except F01 and F04, reduced the G0/G1 population of SF295 cells at growth inhibitory concentrations, and increased G2/M (F02, F03 and F06) or S (F05 and F07) populations. In addition, F02 and F03 decreased the G2/M and G0/G1 population, and increased the S and G2/M population in HCT15 cells, respectively. Meanwhile, in HCT15/CL02 cells, F02 and F03 decreased the G0/G1 populations and increased the S population. In conclusion, we deemed that the flavonoids tested had diverse cytotoxic mechanisms, and exerted their cell growth inhibitory or killing activity by distinctive ways in different cells. PMID:10697540

  18. Animal Models for Studying the In Vivo Functions of Cell Cycle CDKs.

    PubMed

    Risal, Sanjiv; Adhikari, Deepak; Liu, Kui

    2016-01-01

    Multiple Cdks (Cdk4, Cdk6, and Cdk2) and a mitotic Cdk (Cdk1) are involved in cell cycle progression in mammals. Cyclins, Cdk inhibitors, and phosphorylations (both activating and inhibitory) at different cellular levels tightly modulate the activities of these kinases. Based on the results of biochemical studies, it was long believed that different Cdks functioned at specific stages during cell cycle progression. However, deletion of all three interphase Cdks in mice affected cell cycle entry and progression only in certain specialized cells such as hematopoietic cells, beta cells of the pancreas, pituitary lactotrophs, and cardiomyocytes. These genetic experiments challenged the prevailing biochemical model and established that Cdks function in a cell-specific, but not a stage-specific, manner during cell cycle entry and the progression of mitosis. Recent in vivo studies have further established that Cdk1 is the only Cdk that is both essential and sufficient for driving the resumption of meiosis during mouse oocyte maturation. These genetic studies suggest a minimal-essential cell cycle model in which Cdk1 is the central regulator of cell cycle progression. Cdk1 can compensate for the loss of the interphase Cdks by forming active complexes with A-, B-, E-, and D-type Cyclins in a stepwise manner. Thus, Cdk1 plays an essential role in both mitosis and meiosis in mammals, whereas interphase Cdks are dispensable. PMID:26231715

  19. From the cell cycle to population cycles in phytoplankton-nutrient interactions

    SciTech Connect

    Pascual, M.; Caswell, H.

    1997-04-01

    The internal demographic structure of a population influences its dynamics and its response to the environment. Most models for phytoplankton ignore internal structure and group all cells in a single variable such as total biomass or density. However, a cell does have a life history, the cell division cycle. We investigate the significance of the cell cycle to phytoplankton population dynamics in a variable nutrient environment, using chemostate models. Following the transition point hypothesis, nutrient uptake affects cell development only within a limited segment of the cell cycle. Simulation results demonstrate oscillations in cell numbers and population structure generated by this interaction. When nutrient input is varied periodically, the population displays an aperiodic response with frequencies different from that of the forcing. These results also hold for a model that includes nutrient storage by the cells. These dynamics differ from those of traditional chemostate models and from cell cycle models driven by light cycles. Resource control of cell cycle progression may explain the time delays previously postulated to explain oscillatory transients in chemostate experiments. 78 refs., 22 figs.

  20. Metal–Arene Complexes with Indolo[3,2-c]-quinolines: Effects of Ruthenium vs Osmium and Modifications of the Lactam Unit on Intermolecular Interactions, Anticancer Activity, Cell Cycle, and Cellular Accumulation

    PubMed Central

    2013-01-01

    Six novel ruthenium(II)– and osmium(II)–arene complexes with three modified indolo[3,2-c]quinolines have been synthesized in situ starting from 2-aminoindoloquinolines and 2-pyridinecarboxaldehyde in the presence of [M(p-cymene)Cl2]2 (M = Ru, Os) in ethanol. All complexes have been characterized by elemental analysis, spectroscopic techniques (1H, 13C NMR, IR, UV–vis), and ESI mass spectrometry, while four complexes were investigated by X-ray diffraction. The complexes have been tested for antiproliferative activity in vitro in A549 (non-small cell lung), SW480 (colon), and CH1 (ovarian) human cancer cell lines and showed IC50 values between 1.3 and >80 μM. The effects of Ru vs Os and modifications of the lactam unit on intermolecular interactions, antiproliferative activity, and cell cycle are reported. One ruthenium complex and its osmium analogue have been studied for anticancer activity in vivo applied both intraperitoneally and orally against the murine colon carcinoma model CT-26. Interestingly, the osmium(II) complex displayed significant growth-inhibitory activity in contrast to its ruthenium counterpart, providing stimuli for further investigation of this class of compounds as potential antitumor drugs. PMID:23431223

  1. Coordinating cell proliferation and differentiation: Antagonism between cell cycle regulators and cell type-specific gene expression

    PubMed Central

    Ruijtenberg, Suzan; van den Heuvel, Sander

    2016-01-01

    ABSTRACT Cell proliferation and differentiation show a remarkable inverse relationship. Precursor cells continue division before acquiring a fully differentiated state, while terminal differentiation usually coincides with proliferation arrest and permanent exit from the division cycle. Mechanistic insight in the temporal coordination between cell cycle exit and differentiation has come from studies of cells in culture and genetic animal models. As initially described for skeletal muscle differentiation, temporal coordination involves mutual antagonism between cyclin-dependent kinases that promote cell cycle entry and transcription factors that induce tissue-specific gene expression. Recent insights highlight the contribution of chromatin-regulating complexes that act in conjunction with the transcription factors and determine their activity. In particular SWI/SNF chromatin remodelers contribute to dual regulation of cell cycle and tissue-specific gene expression during terminal differentiation. We review the concerted regulation of the cell cycle and cell type-specific transcription, and discuss common mutations in human cancer that emphasize the clinical importance of proliferation versus differentiation control. PMID:26825227

  2. Coordinating cell proliferation and differentiation: Antagonism between cell cycle regulators and cell type-specific gene expression.

    PubMed

    Ruijtenberg, Suzan; van den Heuvel, Sander

    2016-01-01

    Cell proliferation and differentiation show a remarkable inverse relationship. Precursor cells continue division before acquiring a fully differentiated state, while terminal differentiation usually coincides with proliferation arrest and permanent exit from the division cycle. Mechanistic insight in the temporal coordination between cell cycle exit and differentiation has come from studies of cells in culture and genetic animal models. As initially described for skeletal muscle differentiation, temporal coordination involves mutual antagonism between cyclin-dependent kinases that promote cell cycle entry and transcription factors that induce tissue-specific gene expression. Recent insights highlight the contribution of chromatin-regulating complexes that act in conjunction with the transcription factors and determine their activity. In particular SWI/SNF chromatin remodelers contribute to dual regulation of cell cycle and tissue-specific gene expression during terminal differentiation. We review the concerted regulation of the cell cycle and cell type-specific transcription, and discuss common mutations in human cancer that emphasize the clinical importance of proliferation versus differentiation control. PMID:26825227

  3. Glucocorticoid receptor-mediated cell cycle arrest is achieved through distinct cell-specific transcriptional regulatory mechanisms.

    PubMed Central

    Rogatsky, I; Trowbridge, J M; Garabedian, M J

    1997-01-01

    Glucocorticoids inhibit proliferation of many cell types, but the events leading from the activated glucocorticoid receptor (GR) to growth arrest are not understood. Ectopic expression and activation of GR in human osteosarcoma cell lines U2OS and SAOS2, which lack endogenous receptors, result in a G1 cell cycle arrest. GR activation in U2OS cells represses expression of the cyclin-dependent kinases (CDKs) CDK4 and CDK6 as well as their regulatory partner, cyclin D3, leading to hypophosphorylation of the retinoblastoma protein (Rb). We also demonstrate a ligand-dependent reduction in the expression of E2F-1 and c-Myc, transcription factors involved in the G1-to-S-phase transition. Mitogen-activated protein kinase, CDK2, cyclin E, and the CDK inhibitors (CDIs) p27 and p21 are unaffected by receptor activation in U2OS cells. The receptor's N-terminal transcriptional activation domain is not required for growth arrest in U2OS cells. In Rb-deficient SAOS2 cells, however, the expression of p27 and p21 is induced upon receptor activation. Remarkably, in SAOS2 cells that express a GR deletion derivative lacking the N-terminal transcriptional activation domain, induction of CDI expression is abolished and the cells fail to undergo ligand-dependent cell cycle arrest. Similarly, murine S49 lymphoma cells, which, like SAOS2 cells, lack Rb, require the N-terminal activation domain for growth arrest and induce CDI expression upon GR activation. These cell-type-specific differences in receptor domains and cellular targets linking GR activation to cell cycle machinery suggest two distinct regulatory mechanisms of GR-mediated cell cycle arrest: one involving transcriptional repression of G1 cyclins and CDKs and the other involving enhanced transcription of CDIs by the activated receptor. PMID:9154817

  4. Destructive physical analysis results of Ni/H2 cells cycled in LEO regime

    NASA Technical Reports Server (NTRS)

    Lim, Hong S.; Zelter, Gabriela R.; Smithrick, John J.; Hall, Stephen W.

    1991-01-01

    Six 48-Ah individual pressure vessel (IPV) Ni/H2 cells containing 26 and 31 percent KOH electrolyte were life cycle tested in low Earth orbit. All three cells containing 31 percent KOH failed (3729, 4165, and 11,355 cycles), while those with 26 percent KOH were cycled over 14,000 times in the continuing test. Destructive physical analysis (DPA) of the failed cells included visual inspections, measurements of electrode thickness, scanning electron microscopy, chemical analysis, and measurements of nickel electrode capacity in an electrolyte flooded cell. The cycling failure was due to a decrease of nickel electrode capacity. As possible causes of the capacity decrease, researchers observed electrode expansion, rupture, and corrosion of the nickel electrode substrate, active material redistribution, and accumulation of electrochemically undischargeable active material with cycling.

  5. Long life nickel electrodes for a nickel-hydrogen cell: Cycle life tests

    NASA Technical Reports Server (NTRS)

    Lim, H. S.; Verzwyvelt, S. A.

    1985-01-01

    In order to develop a long life nickel electrode for a Ni/H2 cell, the cycle life of nickel electrodes was tested in Ni/H2 boiler plate cells. A 19 test cell matrix was made of various nickel electrode designs including three levels each of plaque mechanical strength, median pore size of the plaque, and active material loading. Test cells were cycled to the end of their life (0.5v) in a 45 minute low Earth orbit cycle regime at 80% depth-of-discharge. It is shown that the active material loading level affects the cycle life the most with the optimum loading at 1.6 g/cc void. Mechanical strength does not affect the cycle life noticeably in the bend strength range of 400 to 700 psi. It is found that the best plaque is made of INCO nickel powder type 287 and has median pore size of 13 micron.

  6. Cytoplasmic sequestration of cyclin D1 associated with cell cycle withdrawal of neuroblastoma cells

    SciTech Connect

    Sumrejkanchanakij, Piyamas; Eto, Kazuhiro; Ikeda, Masa-Aki . E-mail: mikeda.emb@tmd.ac.jp

    2006-02-03

    The regulation of D-type cyclin-dependent kinase activity is critical for neuronal differentiation and apoptosis. We recently showed that cyclin D1 is sequestered in the cytoplasm and that its nuclear localization induces apoptosis in postmitotic primary neurons. Here, we further investigated the role of the subcellular localization of cyclin D1 in cell cycle withdrawal during the differentiation of N1E-115 neuroblastoma cells. We show that cyclin D1 became predominantly cytoplasmic after differentiation. Targeting cyclin D1 expression to the nucleus induced phosphorylation of Rb and cdk2 kinase activity. Furthermore, cyclin D1 nuclear localization promoted differentiated N1E-115 cells to reenter the cell cycle, a process that was inhibited by p16{sup INK4a}, a specific inhibitor of D-type cyclin activity. These results indicate that cytoplasmic sequestration of cyclin D1 plays a role in neuronal cell cycle withdrawal, and suggests that the abrogation of machinery involved in monitoring aberrant nuclear cyclin D1 activity contributes to neuronal tumorigenesis.

  7. Diosgenin induces G2/M cell cycle arrest and apoptosis in human hepatocellular carcinoma cells.

    PubMed

    Li, Yongjian; Wang, Xiaorong; Cheng, Silu; Du, Juan; Deng, Zhengting; Zhang, Yani; Liu, Qun; Gao, Jingdong; Cheng, Binbin; Ling, Changquan

    2015-02-01

    Diosgenin is a major compound of Dioscoreaceae plants such as yam, which is used as a drug in Traditional Chinese Medicine, and a common vegetable worldwide. The anticancer effect of diosgenin has been reported in various tumor cells, including leukemia, gastric, colorectal, and breast cancer. However, the activity of diosgenin on hepatocellular carcinoma (HCC) and the underlying mechanism have not been completely investigated. Therefore, we investigated the efficacy and associated mechanisms of diosgenin in HCC cells. Flow cytometric analysis was performed to determine the presence of cell cycle arrest and apopotic cells. Diosgenin significantly inhibited the growth of Bel-7402, SMMC-7721 and HepG2 HCC cells in a concentration-dependent manner. Diosgenin treatment for 24 h induced G2/M cell cycle arrest and apoptosis of hepatoma cells. Diosgenin inhibited Akt phosphorylation and upregulated p21 and p27 expression, but did not alter the expression of p53, suggesting diosgenin-induced upregulation of p21 and p57 is p53-independent in HCC cells. Diosgenin induced HCC cell apoptosis by activating caspase cascades -3, -8 and -9. However, diosgenin did not affect Bcl-2 and Bax levels. In conclusion, results of the present study suggest that diosgenin may be an active anti-HCC agent obtained from natural plants and provide new insights in understanding the mechanisms of diosgenin. PMID:25434486

  8. Meridional Flow Variations in Cycles 23 and 24: Active Latitude Control of Sunspot Cycle Amplitudes

    NASA Technical Reports Server (NTRS)

    Hathaway, David H.; Upton, Lisa

    2013-01-01

    We have measured the meridional motions of magnetic elements observed in the photosphere over sunspot cycles 23 and 24 using magnetograms from SOHO/MDI and SDO/HMI. Our measurements confirm the finding of Komm, Howard, and Harvey (1993) that the poleward meridional flow weakens at cycle maxima. Our high spatial and temporal resolution analyses show that this variation is in the form of a superimposed inflow toward the active latitudes. This inflow is weaker in cycle 24 when compared to the inflow in 23, the stronger cycle. This systematic modulation of the meridional flow can modulate the amplitude of the following sunspot cycle through its influence on the Sun's polar fields.

  9. Cycle life of nickel-hydrogen cells. II - Accelerated cycle life test

    NASA Technical Reports Server (NTRS)

    Lim, H. S.; Verzwyvelt, S. A.

    1986-01-01

    A cycle life test of nickel-hydrogen (Ni/H2) cells containing electrolytes of various KOH concentrations and a sintered-type nickel electrode were carried out at 23 C using a 45-min accelerated low earth orbit (LEO) cycle regime at 80 percent depth of discharge. Ten cells containing 21 to 36 percent KOH were tested. Since this accelerated test regime accelerated the cycle life roughly twice as fast as a typical LEO regime, the present results indicate that the cells with 26 percent KOH may last over 5 years in an 80 percent depth-of-discharge cycling in an LEO regime. Cells with lower KOH concentrations (21 to 23.5 percent) also showed longer cycle life than those with KOH concentrations of 31 percent or higher, although the life was shorter than those with 26 percent KOH.

  10. Thermal stress cycling of GaAs solar cells

    NASA Technical Reports Server (NTRS)

    Francis, Robert W.

    1987-01-01

    Thermal stress cycling was performed on gallium arsenide solar cells to investigate their electrical, mechanical, and structural integrity. Cells were cycled under low Earth orbit (LEO) simulated temperature conditions in vacuum. Cell evaluations consisted of power output values, spectral response, optical microscopy and ion microprobe mass analysis, and depth profiles on both front surface inter-grid areas and metallization contact grid lines. Cells were examined for degradation after 500, 5,000, 10,000 and 15,245 thermal cycles. No indication of performance degradation was found for any vendor's cell lot.

  11. Modeling cell-cycle synchronization during embryogenesis in Xenopus laevis

    NASA Astrophysics Data System (ADS)

    McIsaac, R. Scott; Huang, K. C.; Sengupta, Anirvan; Wingreen, Ned

    2010-03-01

    A widely conserved aspect of embryogenesis is the ability to synchronize nuclear divisions post-fertilization. How is synchronization achieved? Given a typical protein diffusion constant of 10 μm^2sec, and an embryo length of 1mm, it would take diffusion many hours to propagate a signal across the embryo. Therefore, synchrony cannot be attained by diffusion alone. We hypothesize that known autocatalytic reactions of cell-cycle components make the embryo an ``active medium'' in which waves propagate much faster than diffusion, enforcing synchrony. We report on robust spatial synchronization of components of the core cell cycle circuit based on a mathematical model previously determined by in vitro experiments. In vivo, synchronized divisions are preceded by a rapid calcium wave that sweeps across the embryo. Experimental evidence supports the hypothesis that increases in transient calcium levels lead to derepression of a negative feedback loop, allowing cell divisions to start. Preliminary results indicate a novel relationship between the speed of the initial calcium wave and the ability to achieve synchronous cell divisions.

  12. (p)ppGpp and the bacterial cell cycle.

    PubMed

    Nazir, Aanisa; Harinarayanan, Rajendran

    2016-06-01

    Genes of the Rel/Spo homolog (RSH) superfamily synthesize and/or hydrolyse the modified nucleotides pppGpp/ ppGpp (collectively referred to as (p)ppGpp) and are prevalent across diverse bacteria and in plant chloroplasts. Bacteria accumulate (p)ppGpp in response to nutrient deprivation (generically called the stringent response) and elicit appropriate adaptive responses mainly through the regulation of transcription. Although at different concentrations (p)ppGpp affect the expression of distinct set of genes, the two well-characterized responses are reduction in expression of the protein synthesis machinery and increase in the expression of genes coding for amino acid biosynthesis. In Escherichia coli, the cellular (p)ppGpp level inversely correlates with the growth rate and increasing its concentration decreases the steady state growth rate in a defined growth medium. Since change in growth rate must be accompanied by changes in cell cycle parameters set through the activities of the DNA replication and cell division apparatus, (p)ppGpp could coordinate protein synthesis (cell mass increase) with these processes. Here we review the role of (p)ppGpp in bacterial cell cycle regulation. PMID:27240988

  13. RGC-32 is a novel regulator of the T-lymphocyte cell cycle.

    PubMed

    Tegla, Cosmin A; Cudrici, Cornelia D; Nguyen, Vinh; Danoff, Jacob; Kruszewski, Adam M; Boodhoo, Dallas; Mekala, Armugam P; Vlaicu, Sonia I; Chen, Ching; Rus, Violeta; Badea, Tudor C; Rus, Horea

    2015-06-01

    We have previously shown that RGC-32 is involved in cell cycle regulation in vitro. To define the in vivo role of RGC-32, we generated RGC-32 knockout mice. These mice developed normally and did not spontaneously develop overt tumors. To assess the effect of RGC-32 deficiency on cell cycle activation in T cells, we determined the proliferative rates of CD4(+) and CD8(+) T cells from the spleens of RGC-32(-/-) mice, as compared to wild-type (WT, RGC-32(+/+)) control mice. After stimulation with anti-CD3/anti-CD28, CD4(+) T cells from RGC-32(-/-) mice displayed a significant increase in [(3)H]-thymidine incorporation when compared to WT mice. In addition, both CD4(+) and CD8(+) T cells from RGC-32(-/-) mice displayed a significant increase in the proportion of proliferating Ki67(+) cells, indicating that in T cells, RGC-32 has an inhibitory effect on cell cycle activation induced by T-cell receptor/CD28 engagement. Furthermore, Akt and FOXO1 phosphorylation induced in stimulated CD4(+) T-cells from RGC-32(-/-) mice were significantly higher, indicating that RGC-32 inhibits cell cycle activation by suppressing FOXO1 activation. We also found that IL-2 mRNA and protein expression were significantly increased in RGC-32(-/-) CD4(+) T cells when compared to RGC-32(+/+) CD4(+) T cells. In addition, the effect of RGC-32 on the cell cycle and IL-2 expression was inhibited by pretreatment of the samples with LY294002, indicating a role for phosphatidylinositol 3-kinase (PI3K). Thus, RGC-32 is involved in controlling the cell cycle of T cells in vivo, and this effect is mediated by IL-2 in a PI3K-dependent fashion. PMID:25770350

  14. Timing the Drosophila Mid-Blastula Transition: A Cell Cycle-Centered View.

    PubMed

    Yuan, Kai; Seller, Charles A; Shermoen, Antony W; O'Farrell, Patrick H

    2016-08-01

    At the mid-blastula transition (MBT), externally developing embryos refocus from increasing cell number to elaboration of the body plan. Studies in Drosophila reveal a sequence of changes in regulators of Cyclin:Cdk1 that increasingly restricts the activity of this cell cycle kinase to slow cell cycles during early embryogenesis. By reviewing these events, we provide an outline of the mechanisms slowing the cell cycle at and around the time of MBT. The perspectives developed should provide a guiding paradigm for the study of other MBT changes as the embryo transits from maternal control to a regulatory program centered on the expression of zygotic genes. PMID:27339317

  15. The marine-derived fungal metabolite, terrein, inhibits cell proliferation and induces cell cycle arrest in human ovarian cancer cells.

    PubMed

    Chen, Yi-Fei; Wang, Shu-Ying; Shen, Hong; Yao, Xiao-Fen; Zhang, Feng-Li; Lai, Dongmei

    2014-12-01

    The difficulties faced in the effective treatment of ovarian cancer are multifactorial, but are mainly associated with relapse and drug resistance. Cancer stem-like cells have been reported to be an important contributor to these hindering factors. In this study, we aimed to investigate the anticancer activities of a bioactive fungal metabolite, namely terrein, against the human epithelial ovarian cancer cell line, SKOV3, primary human ovarian cancer cells and ovarian cancer stem-like cells. Terrein was separated and purified from the fermentation metabolites of the marine sponge-derived fungus, Aspergillus terreus strain PF26. Its anticancer activities against ovarian cancer cells were investigated by cell proliferation assay, cell migration assay, cell apoptosis and cell cycle assays. The ovarian cancer stem-like cells were enriched and cultured in a serum-free in vitro suspension system. Terrein inhibited the proliferation of the ovarian cancer cells by inducing G2/M phase cell cycle arrest. The underlying mechanisms involved the suppression of the expression of LIN28, an important marker gene of stemness in ovarian cancer stem cells. Of note, our study also demonstrated the ability of terrein to inhibit the proliferation of ovarian cancer stem-like cells, in which the expression of LIN28 was also downregulated. Our findings reveal that terrein (produced by fermention) may prove to be a promising drug candidate for the treatment of ovarian cancer by inhibiting the proliferation of cancer stem-like cells. PMID:25318762

  16. Protein Kinase C Signaling Mediates a Program of Cell Cycle Withdrawal in the Intestinal Epithelium

    PubMed Central

    Frey, Mark R.; Clark, Jennifer A.; Leontieva, Olga; Uronis, Joshua M.; Black, Adrian R.; Black, Jennifer D.

    2000-01-01

    Members of the protein kinase C (PKC) family of signal transduction molecules have been widely implicated in regulation of cell growth and differentiation, although the underlying molecular mechanisms involved remain poorly defined. Using combined in vitro and in vivo intestinal epithelial model systems, we demonstrate that PKC signaling can trigger a coordinated program of molecular events leading to cell cycle withdrawal into G0. PKC activation in the IEC-18 intestinal crypt cell line resulted in rapid downregulation of D-type cyclins and differential induction of p21waf1/cip1 and p27kip1, thus targeting all of the major G1/S cyclin-dependent kinase complexes. These events were associated with coordinated alterations in expression and phosphorylation of the pocket proteins p107, pRb, and p130 that drive cells to exit the cell cycle into G0 as indicated by concomitant downregulation of the DNA licensing factor cdc6. Manipulation of PKC isozyme levels in IEC-18 cells demonstrated that PKCα alone can trigger hallmark events of cell cycle withdrawal in intestinal epithelial cells. Notably, analysis of the developmental control of cell cycle regulatory molecules along the crypt–villus axis revealed that PKCα activation is appropriately positioned within intestinal crypts to trigger this program of cell cycle exit–specific events in situ. Together, these data point to PKCα as a key regulator of cell cycle withdrawal in the intestinal epithelium. PMID:11076962

  17. Rhizoma Paridis Saponins Induces Cell Cycle Arrest and Apoptosis in Non-Small Cell Lung Carcinoma A549 Cells

    PubMed Central

    Zhang, Jue; Yang, Yixi; Lei, Lei; Tian, Mengliang

    2015-01-01

    Background As a traditional Chinese medicine herb, Chonglou (Paris polyphylla var. chinensis) has been used as anticancer medicine in China in recent decades, as it can induce cell cycle arrest and apoptosis in numerous cancer cells. The saponins extract from the rhizoma of Chonglou [Rhizoma Paridis saponins (RPS)] is known as the main active component for anticancer treatment. However, the molecular mechanism of the anticancer effect of RPS is unknown. Material/Methods The present study evaluated the effect of RPS in non-small-cell lung cancer (NSCLC) A549 cells using the 3-(4,5-dimethylthiazol-2-yl) -2,5-diphenyl tetrazolium bromide (MTT) assay and flow cytometry. Subsequently, the expression of several genes associated with cell cycle and apoptosis were detected by reverse transcription-quantitative polymerase chain reaction (qRT-PCR) and Western blotting. Results RPS was revealed to inhibit cell growth, causing a number of cells to accumulate in the G 1 phase of the cell cycle, leading to apoptosis. In addition, the effect was dose-dependent. Moreover, the results of qRT-PCR and Western blotting showed that p53 and cyclin-dependent kinase 2 (CDK2) were significantly downregulated, and that BCL2, BAX, and p21 were upregulated, by RPS treatment. Conclusions We speculated that the RPS could act on a pathway, including p53, p21, BCL2, BAX, and CDK2, and results in G1 cell cycle arrest and apoptosis in NSCLC cells. PMID:26311066

  18. Induction of apoptosis and cell-cycle arrest in human colon cancer cells by meclizine.

    PubMed

    Lin, Jiunn-Chang; Ho, Yuan-Soon; Lee, Jie-Jen; Liu, Chien-Liang; Yang, Tsen-Long; Wu, Chih-Hsiung

    2007-06-01

    Meclizine (MEC), a histamine H1 antagonist, is used for the treatment of motion sickness and vertigo. In this study, we demonstrate that MEC dose-dependently induced apoptosis in human colon cancer cell lines (COLO 205 and HT 29 cells). Results of a DNA ladder assay revealed that DNA ladders appeared with MEC treatment in COLO 205 cells at dosage of >50 microM. In addition, the total cell number decreased dose-dependently after treatment with MEC in COLO 205 and HT 29 cells. Using flow cytometry, the percentage of COLO 205 cells arrested at G0/G1 phase increased dose-dependently. Analysis of changes in cell-cycle arrest-associated proteins with Western blotting showed that p53 and p21 were upregulated after treatment with MEC. The kinase activities of cyclin-dependent kinase 2 (CDK2) and CDK4 were suppressed in MEC-treated cells. As for apoptosis, MEC may induce upregulation of p53 and downregulation of Bcl-2, thus causing the release of cytochrome C from mitochondria and the translocation of apoptosis-inducing factor (AIF) to the nucleus. This resulted in the activation of caspase 3, 8, and 9. Our results provide the molecular basis of MEC-induced apoptosis and cell-cycle arrest in human colon cancer cells. PMID:17222494

  19. Regulation of the Embryonic Cell Cycle During Mammalian Preimplantation Development.

    PubMed

    Palmer, N; Kaldis, P

    2016-01-01

    The preimplantation development stage of mammalian embryogenesis consists of a series of highly conserved, regulated, and predictable cell divisions. This process is essential to allow the rapid expansion and differentiation of a single-cell zygote into a multicellular blastocyst containing cells of multiple developmental lineages. This period of development, also known as the germinal stage, encompasses several important developmental transitions, which are accompanied by dramatic changes in cell cycle profiles and dynamics. These changes are driven primarily by differences in the establishment and enforcement of cell cycle checkpoints, which must be bypassed to facilitate the completion of essential cell cycle events. Much of the current knowledge in this area has been amassed through the study of knockout models in mice. These mouse models are powerful experimental tools, which have allowed us to dissect the relative dependence of the early embryonic cell cycles on various aspects of the cell cycle machinery and highlight the extent of functional redundancy between members of the same gene family. This chapter will explore the ways in which the cell cycle machinery, their accessory proteins, and their stimuli operate during mammalian preimplantation using mouse models as a reference and how this allows for the usually well-defined stages of the cell cycle to be shaped and transformed during this unique and critical stage of development. PMID:27475848

  20. Capacity-cycle life behavior in secondary lithium cells

    NASA Technical Reports Server (NTRS)

    Somoano, R. B.; Carter, B. J.; Shen, D.; Yen, S. P. S.

    1985-01-01

    The practical utilization of high energy density rechargeable lithium cells is dependent upon maintaining high capacity for the duration of the required cycle life. However, a critical, yet generic problem with room temperature lithium systems is that the capacity often declines considerably during the early stages of cycling. The results of our studies are reported on electrolyte degradation which is observed after cells have undergone 300 and 700 deep cycles with 3-methylsulfolane- and 2-methyltetrahydrofuran-LiAsF6 electrolytes, respectively.

  1. SPARC expression induces cell cycle arrest via STAT3 signaling pathway in medulloblastoma cells

    SciTech Connect

    Chetty, Chandramu; Dontula, Ranadheer; Gujrati, Meena; Lakka, Sajani S.

    2012-01-13

    Highlights: Black-Right-Pointing-Pointer Ectopic expression of SPARC impaired cell proliferation in medulloblastoma cells. Black-Right-Pointing-Pointer SPARC expression induces STAT3 mediated cell cycle arrest in medulloblastoma cells. Black-Right-Pointing-Pointer SPARC expression significantly inhibited pre-established tumor growth in nude-mice. -- Abstract: Dynamic cell interaction with ECM components has profound influence in cancer progression. SPARC is a component of the ECM, impairs the proliferation of different cell types and modulates tumor cell aggressive features. We previously reported that SPARC expression significantly impairs medulloblastoma tumor growth in vivo. In this study, we demonstrate that expression of SPARC inhibits medulloblastoma cell proliferation. MTT assay indicated a dose-dependent reduction in tumor cell proliferation in adenoviral mediated expression of SPARC full length cDNA (Ad-DsRed-SP) in D425 and UW228 cells. Flow cytometric analysis showed that Ad-DsRed-SP-infected cells accumulate in the G2/M phase of cell cycle. Further, immunoblot and immunoprecipitation analyses revealed that SPARC induced G2/M cell cycle arrest was mediated through inhibition of the Cyclin-B-regulated signaling pathway involving p21 and Cdc2 expression. Additionally, expression of SPARC decreased STAT3 phosphorylation at Tyr-705; constitutively active STAT3 expression reversed SPARC induced G2/M arrest. Ad-DsRed-SP significantly inhibited the pre-established orthotopic tumor growth and tumor volume in nude-mice. Immunohistochemical analysis of tumor sections from mice treated with Ad-DsRed-SP showed decreased immunoreactivity for pSTAT3 and increased immunoreactivity for p21 compared to tumor section from mice treated with mock and Ad-DsRed. Taken together our studies further reveal that STAT3 plays a key role in SPARC induced G2/M arrest in medulloblastoma cells. These new findings provide a molecular basis for the mechanistic understanding of the

  2. Punctuated evolution and transitional hybrid network in an ancestral cell cycle of fungi

    PubMed Central

    Medina, Edgar M; Turner, Jonathan J; Gordân, Raluca; Skotheim, Jan M; Buchler, Nicolas E

    2016-01-01

    Although cell cycle control is an ancient, conserved, and essential process, some core animal and fungal cell cycle regulators share no more sequence identity than non-homologous proteins. Here, we show that evolution along the fungal lineage was punctuated by the early acquisition and entrainment of the SBF transcription factor through horizontal gene transfer. Cell cycle evolution in the fungal ancestor then proceeded through a hybrid network containing both SBF and its ancestral animal counterpart E2F, which is still maintained in many basal fungi. We hypothesize that a virally-derived SBF may have initially hijacked cell cycle control by activating transcription via the cis-regulatory elements targeted by the ancestral cell cycle regulator E2F, much like extant viral oncogenes. Consistent with this hypothesis, we show that SBF can regulate promoters with E2F binding sites in budding yeast. DOI: http://dx.doi.org/10.7554/eLife.09492.001 PMID:27162172

  3. Punctuated evolution and transitional hybrid network in an ancestral cell cycle of fungi.

    PubMed

    Medina, Edgar M; Turner, Jonathan J; Gordân, Raluca; Skotheim, Jan M; Buchler, Nicolas E

    2016-01-01

    Although cell cycle control is an ancient, conserved, and essential process, some core animal and fungal cell cycle regulators share no more sequence identity than non-homologous proteins. Here, we show that evolution along the fungal lineage was punctuated by the early acquisition and entrainment of the SBF transcription factor through horizontal gene transfer. Cell cycle evolution in the fungal ancestor then proceeded through a hybrid network containing both SBF and its ancestral animal counterpart E2F, which is still maintained in many basal fungi. We hypothesize that a virally-derived SBF may have initially hijacked cell cycle control by activating transcription via the cis-regulatory elements targeted by the ancestral cell cycle regulator E2F, much like extant viral oncogenes. Consistent with this hypothesis, we show that SBF can regulate promoters with E2F binding sites in budding yeast. PMID:27162172

  4. A Krebs Cycle Component Limits Caspase Activation Rate through Mitochondrial Surface Restriction of CRL Activation.

    PubMed

    Aram, Lior; Braun, Tslil; Braverman, Carmel; Kaplan, Yosef; Ravid, Liat; Levin-Zaidman, Smadar; Arama, Eli

    2016-04-01

    How cells avoid excessive caspase activity and unwanted cell death during apoptotic caspase-mediated removal of large cellular structures is poorly understood. We investigate caspase-mediated extrusion of spermatid cytoplasmic contents in Drosophila during spermatid individualization. We show that a Krebs cycle component, the ATP-specific form of the succinyl-CoA synthetase β subunit (A-Sβ), binds to and activates the Cullin-3-based ubiquitin ligase (CRL3) complex required for caspase activation in spermatids. In vitro and in vivo evidence suggests that this interaction occurs on the mitochondrial surface, thereby limiting the source of CRL3 complex activation to the vicinity of this organelle and reducing the potential rate of caspase activation by at least 60%. Domain swapping between A-Sβ and the GTP-specific SCSβ (G-Sβ), which functions redundantly in the Krebs cycle, show that the metabolic and structural roles of A-Sβ in spermatids can be uncoupled, highlighting a moonlighting function of this Krebs cycle component in CRL activation. PMID:27052834

  5. Caveolin-2 regulation of the cell cycle in response to insulin in Hirc-B fibroblast cells

    SciTech Connect

    Kim, Sangmin; Pak, Yunbae . E-mail: ybpak@nongae.gsnu.ac.kr

    2005-04-29

    The regulatory function of caveolin-2 in cell cycle regulation by insulin was investigated in human insulin receptor-overexpressed rat 1 fibroblast (Hirc-B) cells. Insulin increased induction of the caveolin-2 gene in a time-dependent manner. Direct interaction between ERK and caveolin-2 was confirmed by immunoprecipitation and phosphorylated ERK increased the specific interaction in response to insulin. That insulin induced their nuclear co-localization over time was demonstrated by immunofluorescence microscopy. Insulin increased the S phase in the cell cycle by 6-fold. When recombinant caveolin-1 was transiently expressed, a decrease in the S phase was detected by flow-cytometry. The results indicate that the up-regulation of caveolin-2 in response to insulin activates the downstream signal cascades in the cell cycle, chiefly the increased phosphorylation of ERK, the nuclear translocation of phosphorylated ERK, and the subsequent activation of G0/G1 to S phase transition of the cell cycle. The results also suggest that DNA synthesis and the activation of the cell cycle by insulin are achieved concomitantly with an increase in the interaction between caveolin-2 and phosphorylated ERK, and the nuclear translocation of that complex. Taken together, we conclude that caveolin-2 positively regulates the insulin-induced cell cycle through activation of and direct interaction with ERK in Hirc-B cells.

  6. The ORC1 cycle in human cells: I. cell cycle-regulated oscillation of human ORC1.

    PubMed

    Tatsumi, Yasutoshi; Ohta, Satoshi; Kimura, Hiroshi; Tsurimoto, Toshiki; Obuse, Chikashi

    2003-10-17

    Components of ORC (the origin recognition complex) are highly conserved among eukaryotes and are thought to play an essential role in the initiation of DNA replication. The level of the largest subunit of human ORC (ORC1) during the cell cycle was studied in several human cell lines with a specific antibody. In all cell lines, ORC1 levels oscillate: ORC1 starts to accumulate in mid-G1 phase, reaches a peak at the G1/S boundary, and decreases to a basal level in S phase. In contrast, the levels of other ORC subunits (ORCs 2-5) remain constant throughout the cell cycle. The oscillation of ORC1, or the ORC1 cycle, also occurs in cells expressing ORC1 ectopically from a constitutive promoter. Furthermore, the 26 S proteasome inhibitor MG132 blocks the decrease in ORC1, suggesting that the ORC1 cycle is mainly due to 26 S proteasome-dependent degradation. Arrest of the cell cycle in early S phase by hydroxyurea, aphidicolin, or thymidine treatment is associated with basal levels of ORC1, indicating that ORC1 proteolysis starts in early S phase and is independent of S phase progression. These observations indicate that the ORC1 cycle in human cells is highly linked with cell cycle progression, allowing the initiation of replication to be coordinated with the cell cycle and preventing origins from refiring. PMID:12909627

  7. Formula G1: Cell cycle in the driver's seat of stem cell fate determination.

    PubMed

    Julian, Lisa M; Carpenedo, Richard L; Rothberg, Janet L Manias; Stanford, William L

    2016-04-01

    Cell cycle dynamics has emerged as a key regulator of stem cell fate decisions. In particular, differentiation decisions are associated with the G1 phase, and recent evidence suggests that self-renewal is actively regulated outside of G1. The mechanisms underlying these phenomena are largely unknown, but direct control of gene regulatory programs by the cell cycle machinery is heavily implicated. A recent study sheds important mechanistic insight by demonstrating that in human embryonic stem cells (hESCs) the Cyclin-dependent kinase CDK2 controls a wide-spread epigenetic program that drives transcription at differentiation-related gene promoters specifically in G1. Here, we discuss this finding and explore whether similar mechanisms are likely to function in multipotent stem cells. The implications of this discovery toward our understanding of stem cell-related disease are discussed, and we postulate novel mechanisms that position the cell cycle as a regulator of cell fate gene networks at epigenetic, transcriptional and post-transcriptional levels. PMID:26857166

  8. Xanthohumol induces apoptosis and S phase cell cycle arrest in A549 non-small cell lung cancer cells

    PubMed Central

    Yong, Wai Kuan; Ho, Yen Fong; Malek, Sri Nurestri Abd

    2015-01-01

    Background: Xanthohumol, a major prenylated chalcone found in female hop plant, Humulus lupulus, was reported to have various chemopreventive and anti-cancer properties. However, its apoptotic effect on human alveolar adenocarcinoma cell line (A549) of non-small cell lung cancer (NSCLC) was unknown. Objective: This study aimed to investigate the effects of xanthohumol on apoptosis in A549 human NSCLC cells. Materials and Methods: A549 cell proliferation was determined by sulforhodamine B assay. Morphological changes of the cells were studied via phase contrast and fluorescent microscopy. Induction of apoptosis was assessed by Annexin-V fluorescein isothiocyanate/propidium iodide (Annexin V-FITC/PI) staining, DNA fragmentation (TUNEL) assay mitochondrial membrane potential assay, cell cycle analysis, and caspase activity studies. Results: Xanthohumol was found to decrease cell proliferation in A549 cells but had relatively low cytotoxicity on normal human lung fibroblast cell line (MRC-5). Typical cellular and nuclear apoptotic features were also observed in A549 cells treated with xanthohumol. Onset of apoptosis in A549 cells was further confirmed by externalization of phosphatidylserine, changes in mitochondrial membrane potential, and DNA fragmentation in the cells after treatment. Xanthohumol induced accumulation of cells in sub G1 and S phase based on cell cycle analysis and also increased the activities of caspase-3, -8, and -9. Conclusion: This work suggests that xanthohumol as an apoptosis inducer, may be a potent therapeutic compound for NSCLC. PMID:26664015

  9. Microarray-Based Analysis of Cell-Cycle Gene Expression During Spermatogenesis in the Mouse1

    PubMed Central

    Roy Choudhury, Dipanwita; Small, Chris; Wang, Yufeng; Mueller, Paul R.; Rebel, Vivienne I.; Griswold, Michael D.; McCarrey, John R.

    2010-01-01

    Mammalian spermatogenesis is a continuum of cellular differentiation in a lineage that features three principal stages: 1) a mitotically active stage in spermatogonia, 2) a meiotic stage in spermatocytes, and 3) a postreplicative stage in spermatids. We used a microarray-based approach to identify changes in expression of cell-cycle genes that distinguish 1) mitotic type A spermatogonia from meiotic pachytene spermatocytes and 2) pachytene spermatocytes from postreplicative round spermatids. We detected expression of 550 genes related to cell-cycle function in one or more of these cell types. Although a majority of these genes were expressed during all three stages of spermatogenesis, we observed dramatic changes in levels of individual transcripts between mitotic spermatogonia and meiotic spermatocytes and between meiotic spermatocytes and postreplicative spermatids. Our results suggest that distinct cell-cycle gene regulatory networks or subnetworks are associated with each phase of the cell cycle in each spermatogenic cell type. In addition, we observed expression of different members of certain cell-cycle gene families in each of the three spermatogenic cell types investigated. Finally, we report expression of 221 cell-cycle genes that have not previously been annotated as part of the cell cycle network expressed during spermatogenesis, including eight novel genes that appear to be testis-specific. PMID:20631398

  10. Apoptosis and cell-cycle arrest in human and murine tumor cells are initiated by isoprenoids.

    PubMed

    Mo, H; Elson, C E

    1999-04-01

    Diverse classes of phytochemicals initiate biological responses that effectively lower cancer risk. One class of phytochemicals, broadly defined as pure and mixed isoprenoids, encompasses an estimated 22,000 individual components. A representative mixed isoprenoid, gamma-tocotrienol, suppresses the growth of murine B16(F10) melanoma cells, and with greater potency, the growth of human breast adenocarcinoma (MCF-7) and human leukemic (HL-60) cells. beta-Ionone, a pure isoprenoid, suppresses the growth of B16 cells and with greater potency, the growth of MCF-7, HL-60 and human colon adenocarcinoma (Caco-2) cells. Results obtained with diverse cell lines differing in ras and p53 status showed that the isoprenoid-mediated suppression of growth is independent of mutated ras and p53 functions. beta-Ionone suppressed the growth of human colon fibroblasts (CCD-18Co) but only when present at three-fold the concentration required to suppress the growth of Caco-2 cells. The isoprenoids initiated apoptosis and, concomitantly arrested cells in the G1 phase of the cell cycle. Both suppress 3-hydroxy-3-methylglutaryl CoA reductase activity. beta-Ionone and lovastatin interfered with the posttranslational processing of lamin B, an activity essential to assembly of daughter nuclei. This interference, we postulate, renders neosynthesized DNA available to the endonuclease activities leading to apoptotic cell death. Lovastatin-imposed mevalonate starvation suppressed the glycosylation and translocation of growth factor receptors to the cell surface. As a consequence, cells were arrested in the G1 phase of the cell cycle. This rationale may apply to the isoprenoid-mediated G1-phase arrest of tumor cells. The additive and potentially synergistic actions of these isoprenoids in the suppression of tumor cell proliferation and initiation of apoptosis coupled with the mass action of the diverse isoprenoid constituents of plant products may explain, in part, the impact of fruit, vegetable

  11. Revealing the cellular localization of STAT1 during the cell cycle by super-resolution imaging

    NASA Astrophysics Data System (ADS)

    Gao, Jing; Wang, Feng; Liu, Yanhou; Cai, Mingjun; Xu, Haijiao; Jiang, Junguang; Wang, Hongda

    2015-03-01

    Signal transducers and activators of transcription (STATs) can transduce cytokine signals and regulate gene expression. The cellular localization and nuclear trafficking of STAT1, a representative of the STAT family with multiple transcriptional functions, is tightly related with transcription process, which usually happens in the interphase of the cell cycle. However, these priority questions regarding STAT1 distribution and localization at the different cell-cycle stages remain unclear. By using direct stochastic optical reconstruction microscopy (dSTORM), we found that the nuclear expression level of STAT1 increased gradually as the cell cycle carried out, especially after EGF stimulation. Furthermore, STAT1 formed clusters in the whole cell during the cell cycle, with the size and the number of clusters also increasing significantly from G1 to G2 phase, suggesting that transcription and other cell-cycle related activities can promote STAT1 to form more and larger clusters for fast response to signals. Our work reveals that the cellular localization and clustering distribution of STAT1 are associated with the cell cycle, and further provides an insight into the mechanism of cell-cycle regulated STAT1 signal transduction.

  12. Revealing the cellular localization of STAT1 during the cell cycle by super-resolution imaging.

    PubMed

    Gao, Jing; Wang, Feng; Liu, Yanhou; Cai, Mingjun; Xu, Haijiao; Jiang, Junguang; Wang, Hongda

    2015-01-01

    Signal transducers and activators of transcription (STATs) can transduce cytokine signals and regulate gene expression. The cellular localization and nuclear trafficking of STAT1, a representative of the STAT family with multiple transcriptional functions, is tightly related with transcription process, which usually happens in the interphase of the cell cycle. However, these priority questions regarding STAT1 distribution and localization at the different cell-cycle stages remain unclear. By using direct stochastic optical reconstruction microscopy (dSTORM), we found that the nuclear expression level of STAT1 increased gradually as the cell cycle carried out, especially after EGF stimulation. Furthermore, STAT1 formed clusters in the whole cell during the cell cycle, with the size and the number of clusters also increasing significantly from G1 to G2 phase, suggesting that transcription and other cell-cycle related activities can promote STAT1 to form more and larger clusters for fast response to signals. Our work reveals that the cellular localization and clustering distribution of STAT1 are associated with the cell cycle, and further provides an insight into the mechanism of cell-cycle regulated STAT1 signal transduction. PMID:25762114

  13. Revealing the cellular localization of STAT1 during the cell cycle by super-resolution imaging

    PubMed Central

    Gao, Jing; Wang, Feng; Liu, Yanhou; Cai, Mingjun; Xu, Haijiao; Jiang, Junguang; Wang, Hongda

    2015-01-01

    Signal transducers and activators of transcription (STATs) can transduce cytokine signals and regulate gene expression. The cellular localization and nuclear trafficking of STAT1, a representative of the STAT family with multiple transcriptional functions, is tightly related with transcription process, which usually happens in the interphase of the cell cycle. However, these priority questions regarding STAT1 distribution and localization at the different cell-cycle stages remain unclear. By using direct stochastic optical reconstruction microscopy (dSTORM), we found that the nuclear expression level of STAT1 increased gradually as the cell cycle carried out, especially after EGF stimulation. Furthermore, STAT1 formed clusters in the whole cell during the cell cycle, with the size and the number of clusters also increasing significantly from G1 to G2 phase, suggesting that transcription and other cell-cycle related activities can promote STAT1 to form more and larger clusters for fast response to signals. Our work reveals that the cellular localization and clustering distribution of STAT1 are associated with the cell cycle, and further provides an insight into the mechanism of cell-cycle regulated STAT1 signal transduction. PMID:25762114

  14. Regulation of Neuronal Cell Cycle and Apoptosis by MicroRNA 34a.

    PubMed

    Modi, Prashant Kumar; Jaiswal, Surbhi; Sharma, Pushkar

    2016-01-01

    The cell cycle of neurons remains suppressed to maintain the state of differentiation and aberrant cell cycle reentry results in loss of neurons, which is a feature in neurodegenerative disorders like Alzheimer's disease (AD). Present studies revealed that the expression of microRNA 34a (miR-34a) needs to be optimal in neurons, as an aberrant increase or decrease in its expression causes apoptosis. miR-34a keeps the neuronal cell cycle under check by preventing the expression of cyclin D1 and promotes cell cycle arrest. Neurotoxic amyloid β1-42 peptide (Aβ42) treatment of cortical neurons suppressed miR-34a, resulting in unscheduled cell cycle reentry, which resulted in apoptosis. The repression of miR-34a was a result of degradation of TAp73, which was mediated by aberrant activation of the MEK extracellular signal-regulated kinase (ERK) pathway by Aβ42. A significant decrease in miR-34a and TAp73 was observed in the cortex of a transgenic (Tg) mouse model of AD, which correlated well with cell cycle reentry observed in the neurons of these animals. Importantly, the overexpression of TAp73α and miR-34a reversed cell cycle-related neuronal apoptosis (CRNA). These studies provide novel insights into how modulation of neuronal cell cycle machinery may lead to neurodegeneration and may contribute to the understanding of disorders like AD. PMID:26459758

  15. Rest-Activity Cycles in Childhood and Adolescent Depression.

    ERIC Educational Resources Information Center

    Armitage, Roseanne; Hoffmann, Robert; Emslie, Graham; Rintelman, Jeanne; Moore, Jarrette; Lewis, Kelly

    2004-01-01

    Objective: To quantify circadian rhythms in rest-activity cycles in depressed children and adolescents. Method: Restactivity cycles were evaluated by actigraphy over five consecutive 24-hour periods in 100 children and adolescents, including 59 outpatients with major depressive disorder (MDD) and 41 healthy normal controls. Total activity, total…

  16. Geomagnetic Activity Indicates Large Amplitude for Sunspot Cycle 24

    NASA Technical Reports Server (NTRS)

    Hathaway, D. H.; Wilson, R. M.

    2006-01-01

    The level of geomagnetic activity near the time of solar activity minimum has been shown to be a reliable indicator for the amplitude of the following solar activity maximum. The geomagnetic activity index aa can be split into two components: one associated with solar flares, prominence eruptions, and coronal mass ejections which follows the solar activity cycle and a second component associated with recurrent high speed solar wind streams which is out of phase with the solar activity cycle. This second component often peaks before solar activity minimum and has been one of the most reliable indicators for the amplitude of the following maximum. The size of the recent maximum in this second component indicates that solar activity cycle 24 will be much higher than average - similar in size to cycles 21 and 22.

  17. Hydrogen peroxide inhibits transforming growth factor-β1-induced cell cycle arrest by promoting Smad3 linker phosphorylation through activation of Akt-ERK1/2-linked signaling pathway

    SciTech Connect

    Choi, Jiyeon; Park, Seong Ji; Jo, Eun Ji; Lee, Hui-Young; Hong, Suntaek; Kim, Seong-Jin; Kim, Byung-Chul

    2013-06-14

    Highlights: •H{sub 2}O{sub 2} inhibits TGF-β1-induced cell cycle arrest. •H{sub 2}O{sub 2} induces Smad3 linker phosphorylation through Akt-ERK1/2 pathway. •H{sub 2}O{sub 2}-mediated suppression of TGF-β signal requires Smad3 linker phosphorylation. •This is a first report about interplay between H{sub 2}O{sub 2} and growth inhibition pathway. -- Abstract: Hydrogen peroxide (H{sub 2}O{sub 2}) functions as a second messenger in growth factor receptor-mediated intracellular signaling cascade and is tumorigenic by virtue of its ability to promote cell proliferation; however, the mechanisms underlying the growth stimulatory action of H{sub 2}O{sub 2} are less understood. Here we report an important mechanism for antagonistic effects of H{sub 2}O{sub 2} on growth inhibitory response to transforming growth factor-β1 (TGF-β1). In Mv1Lu and HepG2 cells, pretreatment of H{sub 2}O{sub 2} (0.05–0.2 mM) completely blocked TGF-β1-mediated induction of p15{sup INK4B} expression and increase of its promoter activity. Interestingly, H{sub 2}O{sub 2} selectively suppressed the transcriptional activation potential of Smad3, not Smad2, in the absence of effects on TGF-β1-induced phosphorylation of the COOH-tail SSXS motif of Smad3 and its nuclear translocation. Mechanism studies showed that H{sub 2}O{sub 2} increases the phosphorylation of Smad3 at the middle linker region in a concentration- and time-dependent manner and this effect is mediated by activation of extracellular signal-activated kinase 1/2 through Akt. Furthermore, expression of a mutant Smad3 in which linker phosphorylation sites were ablated significantly abrogated the inhibitory effects of H{sub 2}O{sub 2} on TGF-β1-induced increase of p15{sup INK4B}-Luc reporter activity and blockade of cell cycle progression from G1 to S phase. These findings for the first time define H{sub 2}O{sub 2} as a signaling molecule that modulate Smad3 linker phosphorylation and its transcriptional activity, thus providing

  18. Backup pathways of NHEJ in cells of higher eukaryotes: cell cycle dependence.

    PubMed

    Iliakis, George

    2009-09-01

    DNA double-strand breaks (DSBs) induced by ionizing radiation (IR) in cells of higher eukaryotes are predominantly repaired by a pathway of non-homologous end joining (NHEJ) utilizing Ku, DNA-PKcs, DNA ligase IV, XRCC4 and XLF/Cernunnos (D-NHEJ) as central components. Work carried out in our laboratory and elsewhere shows that when this pathway is chemically or genetically compromised, cells do not shunt DSBs to homologous recombination repair (HRR) but instead use another form of NHEJ operating as a backup (B-NHEJ). Here I review our efforts to characterize this repair pathway and discuss its dependence on the cell cycle as well as on the growth conditions. I present evidence that B-NHEJ utilizes ligase III, PARP-1 and histone H1. When B-NHEJ is examined throughout the cell cycle, significantly higher activity is observed in G2 phase that cannot be attributed to HRR. Furthermore, the activity of B-NHEJ is compromised when cells enter the plateau phase of growth. Together, these observations uncover a repair pathway with unexpected biochemical constitution and interesting cell cycle and growth factor regulation. They generate a framework for investigating the mechanistic basis of HRR contribution to DSB repair. PMID:19604590

  19. A Bioengineered Hydrogel System Enables Targeted and Sustained Intramyocardial Delivery of Neuregulin, Activating the Cardiomyocyte Cell Cycle and Enhancing Ventricular Function in a Murine Model of Ischemic Cardiomyopathy

    PubMed Central

    Cohen, Jeffrey E.; Purcell, Brendan P.; MacArthur, John W.; Mu, Anbin; Shudo, Yasuhiro; Patel, Jay B.; Brusalis, Christopher M.; Trubelja, Alen; Fairman, Alexander S.; Edwards, Bryan B.; Davis, Mollie S.; Hung, George; Hiesinger, William; Atluri, Pavan; Margulies, Kenneth B.; Burdick, Jason A.; Woo, Y. Joseph

    2014-01-01

    Background Neuregulin (NRG) is a member of the epidermal growth factor family possessing a critical role in cardiomyocyte development and proliferation. Systemic administration of NRG demonstrated efficacy in cardiomyopathy animal models, leading to clinical trials employing daily NRG infusions. This approach is hindered by requiring daily infusions and off-target exposure. Therefore, this study aimed to encapsulate NRG in a hydrogel (HG) to be directly delivered to the myocardium, accomplishing sustained localized NRG delivery. Methods and Results NRG was encapsulated in HG and release over 14 days confirmed by ELISA in vitro. Sprague-Dawly rats were utilized for cardiomyocyte isolation. Cells were stimulated by PBS, NRG, HG, or NRG-HG and evaluated for proliferation. Cardiomyocytes demonstrated EdU and phosphorylated histone-H3 (PH3) positivity in the NRG-HG group only. For in vivo studies, 2 month old mice (n=60) underwent LAD ligation and were randomized to the 4 treatment groups mentioned. Only NRG-HG treated mice demonstrated PH3 and Ki67 positivity along with decreased caspase-3 activity compared to all controls. NRG was detected in myocardium 6 days following injection without evidence of off-target exposure in NRG-HG animals. At 2 weeks, the NRG-HG group exhibited enhanced LVEF, decreased LV area, and augmented borderzone thickness. Conclusions Targeted and sustained delivery of NRG directly to the myocardial borderzone augments cardiomyocyte mitotic activity, decreases apoptosis, and greatly enhances LV function in a model of ICM. This novel approach to NRG administration avoids off-target exposure and represents a clinically translatable strategy in myocardial regenerative therapeutics. PMID:24902740

  20. Genipin as a novel chemical activator of EBV lytic cycle.

    PubMed

    Son, Myoungki; Lee, Minjung; Ryu, Eunhyun; Moon, Aree; Jeong, Choon-Sik; Jung, Yong Woo; Park, Gyu Hwan; Sung, Gi-Ho; Cho, Hyosun; Kang, Hyojeung

    2015-02-01

    Epstein-Barr virus (EBV) is a ubiquitous gammaherpesvirus that causes acute infection and establishes life-long latency. EBV causes several human cancers, including Burkitt's lymphoma, nasopharyngeal and gastric carcinoma. Antiviral agents can be categorized as virucides, antiviral chemotherapeutic agents, and immunomodulators. Most antiviral agents affect actively replicating viruses, but not their latent forms. Novel antiviral agents must be active on both the replicating and the latent forms of the virus. Gardenia jasminoides is an evergreen flowering plant belonging to the Rubiaceae family and is most commonly found growing wild in Vietnam, Southern China, Taiwan, Japan, Myanmar, and India. Genipin is an aglycone derived from an iridoid glycoside called geniposide, which is present in large quantities in the fruit of G. jasminoides. In this study, genipin was evaluated for its role as an antitumor and antiviral agent that produces inhibitory effects against EBV and EBV associated gastric carcinoma (EBVaGC). In SNU719 cells, one of EBVaGCs, genipin caused significant cytotoxicity (70 μM), induced methylation on EBV C promoter and tumor suppressor gene BCL7A, arrested cell-cycle progress (S phases), upregulated EBV latent/lytic genes in a dose-dependent manner, stimulated EBV progeny production, activated EBV F promoter for EBV lytic activation, and suppressed EBV infection. These results indicated that genipin could be a promising candidate for antiviral and antitumor agents against EBV and EBVaGC. PMID:25626372

  1. Analysis of Cell Cycle Phase Response Captures the Synchronization Phenomena and Reveals a Novel Cell Cycle Network Topology

    NASA Astrophysics Data System (ADS)

    Li, Ying; Lin, Yihan; Scherer, Norbert; Dinner, Aaron

    2011-03-01

    Cell cycle progression requires a succession of temporally-regulated sub-processes, including chromosome replication and cell division, which are each controlled by their own regulatory modules. The modular design of cell cycle regulatory network allows robust environmental responses and evolutionary adaptations. It is emerging that some of the cell cycle modules involve their own autonomous periodic dynamics. As a consequence, the realization of robust coordination among these modules becomes challenging since each module could potentially run out of sync. We believe that an insight into this puzzle resides in the coupling between the contributing regulatory modules. Here, we measured the phase response curve (PRC) of the cell cycle oscillator by driving the expression of a master regulator of the cell cycle in a pulsatile manner and measuring the single cell phase response. We constructed a return map that quantitatively explains the synchronization phenomena that were caused by periodic chemical perturbation. To capture the measured phase response, we derived a minimalist coupled oscillator model that generalizes the basic topology of the cell cycle network. This diode-like coupling suggests that the cell is engineered to ensure complete coordination of constituent events with the cell cycle.

  2. Targeting of cytosolic phospholipase A2α impedes cell cycle re-entry of quiescent prostate cancer cells

    PubMed Central

    Yao, Mu; Xie, Chanlu; Kiang, Mei-Yee; Teng, Ying; Harman, David; Tiffen, Jessamy; Wang, Qian; Sved, Paul; Bao, Shisan; Witting, Paul; Holst, Jeff; Dong, Qihan

    2015-01-01

    Cell cycle re-entry of quiescent cancer cells has been proposed to be involved in cancer progression and recurrence. Cytosolic phospholipase A2α (cPLA2α) is an enzyme that hydrolyzes membrane glycerophospholipids to release arachidonic acid and lysophospholipids that are implicated in cancer cell proliferation. The aim of this study was to determine the role of cPLA2α in cell cycle re-entry of quiescent prostate cancer cells. When PC-3 and LNCaP cells were rendered to a quiescent state, the active form of cPLA2α with a phosphorylation at Ser505 was lower compared to their proliferating state. Conversely, the phospho-cPLA2α levels were resurgent during the induction of cell cycle re-entry. Pharmacological inhibition of cPLA2α with Efipladib upon induction of cell cycle re-entry inhibited the re-entry process, as manifested by refrained DNA synthesis, persistent high proportion of cells in G0/G1 and low percentage of cells in S and G2/M phases, together with a stagnant recovery of Ki-67 expression. Simultaneously, Efipladib prohibited the emergence of Skp2 while maintained p27 at a high level in the nuclear compartment during cell cycle re-entry. Inhibition of cPLA2α also prevented an accumulation of cyclin D1/CDK4, cyclin E/CDK2, phospho-pRb, pre-replicative complex proteins CDC6, MCM7, ORC6 and DNA synthesis-related protein PCNA during induction of cell cycle re-entry. Moreover, a pre-treatment of the prostate cancer cells with Efipladib during induction of cell cycle re-entry subsequently compromised their tumorigenic capacity in vivo. Hence, cPLA2α plays an important role in cell cycle re-entry by quiescent prostate cancer cells. PMID:26416244

  3. Hydrogen peroxide inhibits transforming growth factor-β1-induced cell cycle arrest by promoting Smad3 linker phosphorylation through activation of Akt-ERK1/2-linked signaling pathway.

    PubMed

    Choi, Jiyeon; Park, Seong Ji; Jo, Eun Ji; Lee, Hui-Young; Hong, Suntaek; Kim, Seong-Jin; Kim, Byung-Chul

    2013-06-14

    Hydrogen peroxide (H2O2) functions as a second messenger in growth factor receptor-mediated intracellular signaling cascade and is tumorigenic by virtue of its ability to promote cell proliferation; however, the mechanisms underlying the growth stimulatory action of H2O2 are less understood. Here we report an important mechanism for antagonistic effects of H2O2 on growth inhibitory response to transforming growth factor-β1 (TGF-β1). In Mv1Lu and HepG2 cells, pretreatment of H2O2 (0.05-0.2 mM) completely blocked TGF-β1-mediated induction of p15(INK4B) expression and increase of its promoter activity. Interestingly, H2O2 selectively suppressed the transcriptional activation potential of Smad3, not Smad2, in the absence of effects on TGF-β1-induced phosphorylation of the COOH-tail SSXS motif of Smad3 and its nuclear translocation. Mechanism studies showed that H2O2 increases the phosphorylation of Smad3 at the middle linker region in a concentration- and time-dependent manner and this effect is mediated by activation of extracellular signal-activated kinase 1/2 through Akt. Furthermore, expression of a mutant Smad3 in which linker phosphorylation sites were ablated significantly abrogated the inhibitory effects of H2O2 on TGF-β1-induced increase of p15(INK4B)-Luc reporter activity and blockade of cell cycle progression from G1 to S phase. These findings for the first time define H2O2 as a signaling molecule that modulate Smad3 linker phosphorylation and its transcriptional activity, thus providing a potential mechanism whereby H2O2 antagonizes the cytostatic function of TGF-β1. PMID:23685151

  4. Cell Cycle Related Differentiation of Bone Marrow Cells into Lung Cells

    SciTech Connect

    Dooner, Mark; Aliotta, Jason M.; Pimental, Jeffrey; Dooner, Gerri J.; Abedi, Mehrdad; Colvin, Gerald; Liu, Qin; Weier, Heinz-Ulli; Dooner, Mark S.; Quesenberry, Peter J.

    2007-12-31

    Green-fluorescent protein (GFP) labeled marrow cells transplanted into lethally irradiated mice can be detected in the lungs of transplanted mice and have been shown to express lung specific proteins while lacking the expression of hematopoietic markers. We have studied marrow cells induced to transit cell cycle by exposure to IL-3, IL-6, IL-11 and steel factor at different times of culture corresponding to different phases of cell cycle. We have found that marrow cells at the G1/S interface have a 3-fold increase in cells which assume a lung phenotype and that this increase is no longer seen in late S/G2. These cells have been characterized as GFP{sup +} CD45{sup -} and GFP{sup +} cytokeratin{sup +}. Thus marrow cells with the capacity to convert into cells with a lung phenotype after transplantation show a reversible increase with cytokine induced cell cycle transit. Previous studies have shown the phenotype of bone marrow stem cells fluctuates reversibly as these cells traverse cell cycle, leading to a continuum model of stem cell regulation. The present studies indicate that marrow stem cell production of nonhematopoietic cells also fluctuates on a continuum.

  5. Eye Selector Logic for a Coordinated Cell Cycle Exit

    PubMed Central

    Lopes, Carla S.; Casares, Fernando

    2015-01-01

    Organ-selector transcription factors control simultaneously cell differentiation and proliferation, ensuring the development of functional organs and their homeostasis. How this is achieved at the molecular level is still unclear. Here we have investigated how the transcriptional pulse of string/cdc25 (stg), the universal mitotic trigger, is regulated during Drosophila retina development as an example of coordinated deployment of differentiation and proliferation programs. We identify the eye specific stg enhancer, stg-FMW, and show that Pax6 selector genes, in cooperation with Eya and So, two members of the retinal determination network, activate stg-FMW, establishing a positive feed-forward loop. This loop is negatively modulated by the Meis1 protein, Hth. This regulatory logic is reminiscent of that controlling the expression of differentiation transcription factors. Our work shows that subjecting transcription factors and key cell cycle regulators to the same regulatory logic ensures the coupling between differentiation and proliferation programs during organ development. PMID:25695251

  6. Cycle life test. [of secondary spacecraft cells

    NASA Technical Reports Server (NTRS)

    Harkness, J. D.

    1977-01-01

    Statistical information concerning cell performance characteristics and limitations of secondary spacecraft cells is presented. Weaknesses in cell design as well as battery weaknesses encountered in various satellite programs are reported. Emphasis is placed on improving the reliability of space batteries.

  7. Protein phosphatase 2A is expressed in response to colony-stimulating factor 1 in macrophages and is required for cell cycle progression independently of extracellular signal-regulated protein kinase activity.

    PubMed Central

    Wilson, N J; Moss, S T; Csar, X F; Ward, A C; Hamilton, J A

    1999-01-01

    Colony-stimulating factor 1 (CSF-1) is required for the development of monocytes/macrophages from progenitor cells and for the survival and activation of mature macrophages. The receptor for CSF-1 is the product of the c-fms proto-oncogene, which, on binding ligand, can stimulate a mitogenic response in the appropriate cells. To investigate which genes are regulated in response to CSF-1-stimulation in murine bone-marrow-derived macrophages (BMM), we employed mRNA differential display reverse transcriptase-mediated PCR to identify cDNA species induced by CSF-1. Both Northern and Western blot analyses confirmed the increased expression of one of the cDNA species identified as coding for the catalytic subunit of protein phosphatase 2A (PP2A), an observation not previously reported during the response to a growth factor. To determine the significance of the increased expression of PP2A in response to CSF-1, the PP2A inhibitor okadaic acid (OA) was added to CSF-1-treated BMM and found to inhibit DNA synthesis in a dose-dependent manner. Further analysis with flow cytometry in the presence of OA led to the novel conclusion that PP2A activity is critical for CSF-1-driven BMM cell cycle progression in both early G1 and S phases. Surprisingly, in the light of previous studies with other cells, the PP2A-dependent proliferation could be dissociated from activation by extracellular signal-regulated protein kinase (ERK) in macrophages because OA did not affect either the basal or CSF-1-induced ERK activity in BMM. Two-dimensional SDS/PAGE analysis of lysates of 32P-labelled BMM, which had been treated with CSF-1 in the presence or absence of OA, identified candidate substrates for PP2A. PMID:10215588

  8. Infection with Toxoplasma gondii results in dysregulation of the host cell cycle

    PubMed Central

    Molestina, Robert E.; El-Guendy, Nadia; Sinai, Anthony P.

    2009-01-01

    SUMMARY Mammalian cells infected with Toxoplasma gondii are characterized by a profound reprogramming of gene expression. We examined whether such transcriptional responses were linked to changes in the cell cycle of the host. Human foreskin fibroblasts (HFF) in the G0/G1 phase of the cell cycle were infected with T. gondii and FACS analysis of DNA content was performed. Cell cycle profiles revealed a promotion into the S phase followed by an arrest towards the G2/M boundary with infection. This response was markedly different from that of growth factor stimulation which caused cell cycle entry and completion. Transcriptional profiles of T. gondii-infected HFF showed sustained increases in transcripts associated with a G1/S transition and DNA synthesis coupled to an abrogation of cell cycle regulators critical in G2/M transition relative to growth factor stimulation. These divergent responses correlated with a distinct temporal modulation of the critical cell cycle regulator kinase ERK by infection. While the kinetics of ERK phosphorylation by EGF showed rapid and sustained activation, infected cells displayed an oscillatory pattern of activation. Our results suggest that T. gondii infection induces and maintains a “proliferation response” in the infected cell which may fulfill critical growth requirements of the parasite during intracellular residence. PMID:18182087

  9. Inferences on Stellar Activity and Stellar Cycles from Asteroseismology

    NASA Astrophysics Data System (ADS)

    Chaplin, William J.; Basu, Sarbani

    2014-12-01

    The solar activity cycle can be studied using many different types of observations, such as counting sunspots, measuring emission in the Ca II H&K lines, magnetograms, radio emissions, etc. One of the more recent ways of studying solar activity is to use the changing properties of solar oscillations. Stellar activity cycles are generally studied using the Ca II lines, or sometimes using photometry. Asteroseismology is potentially an exciting means of studying these cycles. In this article we examine whether or not asteroseismic data can be used for this purpose, and what the asteroseismic signatures of stellar activity are. We also examine how asteroseismology may help in more indirect ways.

  10. 6-Shogaol induces cell cycle arrest and apoptosis in human hepatoma cells through pleiotropic mechanisms.

    PubMed

    Wu, Jung-Ju; Omar, Hany A; Lee, Ying-Ray; Teng, Yen-Ni; Chen, Pin-Shern; Chen, Yu-Chung; Huang, Hsiao-Shan; Lee, Kuan-Han; Hung, Jui-Hsiang

    2015-09-01

    Shogaols are a group of the active constituents of ginger that have been identified to have various biological activities. The aim of the current study was to investigate the antitumor activity of 6-shogaol in hepatocellular carcinoma (HCC) and the possible involvement of reactive oxygen species as a putative mechanism of action. HCC cell lines, HepG2 and Huh-7, were used to study the in vitro anti-cancer activity of 6-shogaol via the application of various molecular biology techniques. Results showed that 6-shogaol effectively inhibited the cell viability, caused cell cycle arrest at G2/M phase and induced apoptosis in HCC cells as indicated by MTT assay, DAPI nuclear staining, annexin V assay, cell cycle analysis, and activation of caspase-3. Western blot analysis revealed the ability of 6-shogaol to target cancer survival signaling pathways mediated by mitogen-activated protein kinase (MAPK), 5' AMP-activated protein kinase (AMPK) and Akt. In addition, 6-Shogaol induced alteration of cyclin proteins expression and caused cleavage of protein kinase C delta. Furthermore, 6-Shogaol was able to induce the production of reactive oxygen species and endoplasmic reticulum (ER) stress-associated proteins and the consequent activation of autophagy in HepG2 cells. Taken together, the current study highlights evidences that 6-shogaol induces apoptosis, modulates cyclins expression and targets cancer survival signaling pathways in HCC cell lines, at least in part, via the production of reactive oxygen species. These findings support 6-shogaol's clinical promise as a potential candidate for HCC therapy. PMID:26101062

  11. A Src inhibitor regulates the cell cycle of human pluripotent stem cells and improves directed differentiation

    PubMed Central

    Engquist, Elise N.; Mehanna, Elie; Lui, Kathy O.; Tsankov, Alexander M.

    2015-01-01

    Driving human pluripotent stem cells (hPSCs) into specific lineages is an inefficient and challenging process. We show that a potent Src inhibitor, PP1, regulates expression of genes involved in the G1 to S phase transition of the cell cycle, activates proteins in the retinoblastoma family, and subsequently increases the differentiation propensities of hPSCs into all three germ layers. We further demonstrate that genetic suppression of Src regulates the activity of the retinoblastoma protein and enhances the differentiation potential of hPSCs across all germ layers. These positive effects extend beyond the initial germ layer specification and enable efficient differentiation at subsequent stages of differentiation. PMID:26416968

  12. A Src inhibitor regulates the cell cycle