WorldWideScience

Sample records for cell cycle-related cyclin

  1. Cyclin K and cyclin D1b are oncogenic in myeloma cells

    Directory of Open Access Journals (Sweden)

    Renoir Jack-Michel

    2010-05-01

    Full Text Available Abstract Background Aberrant expression of cyclin D1 is a common feature in multiple myeloma (MM and always associated with mantle cell lymphoma (MCL. CCND1 gene is alternatively spliced to produce two cyclin D1 mRNA isoforms which are translated in two proteins: cyclin D1a and cyclin D1b. Both isoforms are present in MM cell lines and primary cells but their relative role in the tumorigenic process is still elusive. Results To test the tumorigenic potential of cyclin D1b in vivo, we generated cell clones derived from the non-CCND1 expressing MM LP-1 cell line, synthesizing either cyclin D1b or cyclin K, a structural homolog and viral oncogenic form of cyclin D1a. Immunocompromised mice injected s.c. with LP-1K or LP-1D1b cells develop tumors at the site of injection. Genome-wide analysis of LP-1-derived cells indicated that several cellular processes were altered by cyclin D1b and/or cyclin K expression such as cell metabolism, signal transduction, regulation of transcription and translation. Importantly, cyclin K and cyclin D1b have no major action on cell cycle or apoptosis regulatory genes. Moreover, they impact differently cell functions. Cyclin K-expressing cells have lost their migration properties and display enhanced clonogenic capacities. Cyclin D1b promotes tumorigenesis through the stimulation of angiogenesis. Conclusions Our study indicates that cyclin D1b participates into MM pathogenesis via previously unrevealed actions.

  2. Translational control of cyclins

    Directory of Open Access Journals (Sweden)

    Lai Ming-Chih

    2011-02-01

    Full Text Available Abstract Regulation of cyclin levels is important for many cell cycle-related processes and can occur at several different steps of gene expression. Translational regulation of cyclins, which occurs by a variety of regulatory mechanisms, permits a prompt response to signal transduction pathways induced by environmental stimuli. This review will summarize translational control of cyclins and its influence on cell cycle progression.

  3. Cell cyclins: triggering elements of cancer or not?

    Directory of Open Access Journals (Sweden)

    Pateras Ioannis

    2010-12-01

    Full Text Available Abstract Cyclins are indispensable elements of the cell cycle and derangement of their function can lead to cancer formation. Recent studies have also revealed more mechanisms through which cyclins can express their oncogenic potential. This review focuses on the aberrant expression of G1/S cyclins and especially cyclin D and cyclin E; the pathways through which they lead to tumour formation and their involvement in different types of cancer. These elements indicate the mechanisms that could act as targets for cancer therapy.

  4. Cyclin A2: a genuine cell cycle regulator?

    Science.gov (United States)

    Bendris, Nawal; Loukil, Abdelhalim; Cheung, Caroline; Arsic, Nikola; Rebouissou, Cosette; Hipskind, Robert; Peter, Marion; Lemmers, Bénédicte; Blanchard, Jean Marie

    2012-12-01

    Abstract Cyclin A2 belongs to the core cell cycle regulators and participates in the control of both S phase and mitosis. However, several observations suggest that it is also endowed with other functions, and our recent data shed light on its involvement in cytoskeleton dynamic and cell motility. From the transcription of its gene to its posttranslational modifications, cyclin A2 regulation reveals the complexity of the regulatory network shaping cell cycle progression. We summarize our current knowledge on this cell cycle regulator and discuss recent findings raising the possibility that cyclin A2 might play a much broader role in epithelial tissues homeostasis.

  5. Rising cyclin-CDK levels order cell cycle events.

    Directory of Open Access Journals (Sweden)

    Catherine Oikonomou

    Full Text Available Diverse mitotic events can be triggered in the correct order and time by a single cyclin-CDK. A single regulator could confer order and timing on multiple events if later events require higher cyclin-CDK than earlier events, so that gradually rising cyclin-CDK levels can sequentially trigger responsive events: the "quantitative model" of ordering.This 'quantitative model' makes predictions for the effect of locking cyclin at fixed levels for a protracted period: at low cyclin levels, early events should occur rapidly, while late events should be slow, defective, or highly variable (depending on threshold mechanism. We titrated the budding yeast mitotic cyclin Clb2 within its endogenous expression range to a stable, fixed level and measured time to occurrence of three mitotic events: growth depolarization, spindle formation, and spindle elongation, as a function of fixed Clb2 level. These events require increasingly more Clb2 according to their normal order of occurrence. Events occur efficiently and with low variability at fixed Clb2 levels similar to those observed when the events normally occur. A second prediction of the model is that increasing the rate of cyclin accumulation should globally advance timing of all events. Moderate (<2-fold overexpression of Clb2 accelerates all events of mitosis, resulting in consistently rapid sequential cell cycles. However, this moderate overexpression also causes a significant frequency of premature mitoses leading to inviability, suggesting that Clb2 expression level is optimized to balance the fitness costs of variability and catastrophe.We conclude that mitotic events are regulated by discrete cyclin-CDK thresholds. These thresholds are sequentially triggered as cyclin increases, yielding reliable order and timing. In many biological processes a graded input must be translated into discrete outputs. In such systems, expression of the central regulator is likely to be tuned to an optimum level, as we

  6. Reversible regulation of cell cycle-related genes by epigallocatechin gallate for hibernation of neonatal human tarsal fibroblasts.

    Science.gov (United States)

    Bae, Jung Yoon; Kanamune, Jun; Han, Dong-Wook; Matsumura, Kazuaki; Hyon, Suong-Hyu

    2009-01-01

    We investigated the hibernation effect of epigallocatechin-3-O-gallate (EGCG) on neonatal human tarsal fibroblasts (nHTFs) by analyzing the expression of cell cycle-related genes. EGCG application to culture media moderately inhibited the growth of nHTFs, and the removal of EGCG from culture media led to complete recovery of cell growth. EGCG resulted in a slight decrease in the cell population of the S and G(2)/M phases of cell cycle with concomitant increase in that of the G(0)/G(1) phase, but this cell cycle profile was restored to the initial level after EGCG removal. The expression of cyclin D1 (CCND1), CCNE2, CCN-dependent kinase 6 (CDK6), and CDK2 was restored, whereas that of CCNA, CCNB1, and CDK1 was irreversibly attenuated. The expression of a substantial number of genes analyzed by cDNA microarray was affected by EGCG application, and these affected expression levels were restored to the normal levels after EGCG removal. We also found the incorporation of FITC-EGCG into the cytosol of nHTFs and its further nuclear translocation, which might lead to the regulation of the exogenous signals directed to genes for cellular responses including proliferation and cell cycle progression. These results suggest that EGCG temporarily affects not only genes related to the cell cycle but also various other cellular functions.

  7. Cyclin D

    NARCIS (Netherlands)

    Bernards, R.A.

    2001-01-01

    D type cyclins belong to a family of related proteins that bind to and activate several protein kinases named cyclin-dependent kinases (CDKs), which are involved in regulation of the cell division cycle.

  8. Potential gene regulatory role for cyclin D3 in muscle cells

    Indian Academy of Sciences (India)

    2015-06-27

    Jun 27, 2015 ... Cyclin D3 is important for muscle development and regeneration, and is involved in post-mitotic arrest of muscle cells. Cyclin D3 also has cell-cycle-independent functions such as regulation of specific genes in other tissues. Ectopic expression of cyclin D3 in myoblasts, where it is normally undetectable, ...

  9. Potential gene regulatory role for cyclin D3 in muscle cells

    Indian Academy of Sciences (India)

    Cyclin D3 is important for muscle development and regeneration, and is involved in post-mitotic arrest of muscle cells. Cyclin D3 also has cell-cycle-independent functions such as regulation of specific genes in other tissues. Ectopic expression of cyclin D3 in myoblasts, where it is normally undetectable, promotes muscle ...

  10. Slug regulates Cyclin D1 expression by ubiquitin-proteasome pathway in prostate cancer cells.

    Science.gov (United States)

    Ding, G X; Liu, J; Feng, C C; Jiang, H W; Xu, J F; Ding, Q

    2012-09-01

    Cyclin D1 is an important cell cycle regulatory proteins, which is a functional target of Slug in the regulation of cell growth of prostate cancer cells. But the pathway of these two factors interacting with each other is unclear. The infectde PCa Cells were treated with proteasome inhibitor MG-132. Expression level of Slug, HA-cyclin D1 and other protein was examined by Western blot. Increasing doses of adenovirus expressing human Slug were added to DU-145 cells separately, but there were no significantly difference on expressions of Slug and cyclin D1. We found that the protein expressions of HA-Cyclin D1 (wide-type) were all reduced through high expression of Slug, which is dose-dependent. However, there is no change for HA-Cyclin D1 (mutant) expression in PC-3 with pMIGW-Cyclin D1-HA T286A. The protein expression of HA-Cyclin D1 were all reduced three days after infection by adding adenovirus expressing human Slug to PC-3 carrying pMIGW-Cyclin D1-HA vector compared to negative control, which is dose-dependent. However, there is no change for HA-Cyclin D1 expression in PC-3 with pMIGW-Cyclin D1-HA treated by MG-132. We found that forced expression of Slug inhibited proliferation of prostate cancer cells through downregulation of cyclin D1 expression. And Slug regulates cyclin D1 expression by ubiquitin-proteasome pathway in PCa cells.

  11. Overexpression of Zwint predicts poor prognosis and promotes the proliferation of hepatocellular carcinoma by regulating cell-cycle-related proteins

    Directory of Open Access Journals (Sweden)

    Ying H

    2018-02-01

    Full Text Available Hanning Ying,1,2 Zhiyao Xu,3 Mingming Chen,1,2 Senjun Zhou,1,2 Xiao Liang,1,2 Xiujun Cai1,2 1Department of General Surgery, 2Key Laboratory of Endoscopic Technique Research of Zhejiang Province, 3Central Lab of Biomedical Research Center, School of Medicine, Sir Run Run Shaw Hospital, Zhejiang University, Hangzhou, China Introduction: Zwint, a centromere-complex component required for the mitotic spindle checkpoint, has been reported to be overexpressed in different human cancers, but it has not been studied in human hepatocellular carcinoma (HCC.Materials and methods: The role of Zwint in hepatocellular carcinoma cell proliferation capacities was evaluated by using cell counting kit-8 (CCK8, flow cytometry, clone formation and tumor formation assay in nude mice. Western blot analysis and qPCR assay were performed to assess Zwint interacting with cell-cycle-related proteins.Results: We report that ZWINT mRNA and protein expression were upregulated in HCC samples and cell lines. An independent set of 106 HCC-tissue pairs and corresponding noncancerous tissues was evaluated for Zwint expression using immunohistochemistry, and elevated Zwint expression in HCC tissues was significantly correlated with clinicopathological features, such as tumor size and number. Kaplan–Meier survival and Cox regression analysis revealed that high expression of Zwint was correlated with poor overall survival and a greater tendency for tumor recurrence. Ectopic expression of Zwint promoted HCC-cell proliferation, and Zwint expression affected the expression of several cell-cycle proteins, including PCNA, cyclin B1, Cdc25C and CDK1.Conclusion: Our findings suggest that upregulation of Zwint may contribute to the progression of HCC and may be a prognostic biomarker and potential therapeutic target for treating HCC. Keywords: Zwint, hepatocellular carcinoma, HCC, prognosis, cell proliferation, cell cycle

  12. The 3' untranslated region of the cyclin B mRNA is not sufficient to enhance the synthesis of cyclin B during a mitotic block in human cells.

    Directory of Open Access Journals (Sweden)

    Dominik Schnerch

    Full Text Available Antimitotic agents are frequently used to treat solid tumors and hematologic malignancies. However, one major limitation of antimitotic approaches is mitotic slippage, which is driven by slow degradation of cyclin B during a mitotic block. The extent to which cyclin B levels decline is proposed to be governed by an equilibrium between cyclin B synthesis and degradation. It was recently shown that the 3' untranslated region (UTR of the murine cyclin B mRNA contributes to the synthesis of cyclin B during mitosis in murine cells. Using a novel live-cell imaging-based technique allowing us to study synthesis and degradation of cyclin B simultaneously at the single cell level, we tested here the role of the human cyclin B 3'UTR in regulating cyclin B synthesis during mitosis in human cells. We observed that the cyclin B 3'UTR was not sufficient to enhance cyclin B synthesis in human U2Os, HeLa or hTERT RPE-1 cells. A better understanding of how the equilibrium of cyclin B is regulated in mitosis may contribute to the development of improved therapeutic approaches to prevent mitotic slippage in cancer cells treated with antimitotic agents.

  13. Cyclin F suppresses B-Myb activity to promote cell cycle checkpoint control

    DEFF Research Database (Denmark)

    Klein, Ditte Kjærsgaard; Hoffmann, Saskia; Ahlskog, Johanna K

    2015-01-01

    an important role in checkpoint control following ionizing radiation. Cyclin F-depleted cells initiate checkpoint signalling after ionizing radiation, but fail to maintain G2 phase arrest and progress into mitosis prematurely. Importantly, cyclin F suppresses the B-Myb-driven transcriptional programme...... that promotes accumulation of crucial mitosis-promoting proteins. Cyclin F interacts with B-Myb via the cyclin box domain. This interaction is important to suppress cyclin A-mediated phosphorylation of B-Myb, a key step in B-Myb activation. In summary, we uncover a regulatory mechanism linking the F-box protein...

  14. Flowcytometric evaluation of cell cycle regulators (cyclins and cyclin dependent kinase inhibitors expressed on bone marrow cells of patients with chronic myelogenous leukemia and multiple myeloma

    Directory of Open Access Journals (Sweden)

    Selami Koçak Toprak

    2012-03-01

    Full Text Available OBJECTIVE: Etiopathology of malignancy can be demonstrated by the comparison of the quantified changes in the different phases of the cycle about cyclins and cyclin dependent kinase inhibitors (CDKI in healthy and malignant proliferated cells. The aim of this study is to analyze flow cytometric expression of cell cycle regulating elements in the malignant diseases with low and high proliferative signature. METHODS: The levels of cyclin D, E, A, B and CDKI's p16, p21 were studied by flowcytometry in patients with chronic myeloid leukemia (CML (n=16, multiple myeloma (MM (n=13 and control subjects (n=15. RESULTS: The distributions of the cell cycle S phase were 10, 63%, 6, 72% and 3, 59%; for CML, MM and control subjects, respectively. Among all the cyclins expressed during the S phase, cyclin D expression was the lowest, in CML patients. While the distribution of cyclins and CDKI’s was similar between MM and control groups in G2/M phase; cyclins expressions were parallel in all three phases in MM and chronic myeloid leukemia groups. CONCLUSION: CML and MM are diseases presenting with variable degrees of proliferation. The increase of cyclins in cell cycle phases in patient group was not associated with the augmentation of the expression of CDKI’s. This finding may contribute the mechanisms effective in the etiopathogenesis of hematological malignancy.

  15. Cyclin-dependent kinases regulate apoptosis of intestinal epithelial cells

    Science.gov (United States)

    Bhattacharya, Sujoy; Ray, Ramesh M.; Johnson, Leonard R.

    2014-01-01

    Homeostasis of the gastrointestinal epithelium is dependent upon a balance between cell proliferation and apoptosis. Cyclin-dependent kinases (Cdks) are well known for their role in cell proliferation. Previous studies from our group have shown that polyamine-depletion of intestinal epithelial cells (IEC-6) decreases cyclin-dependent kinase 2 (Cdk2) activity, increases p53 and p21Cip1 protein levels, induces G1 arrest, and protects cells from camptothecin (CPT)-induced apoptosis. Although emerging evidence suggests that members of the Cdk family are involved in the regulation of apoptosis, their roles directing apoptosis of IEC-6 cells are not known. In this study, we report that inhibition of Cdk1, 2, and 9 (with the broad range Cdk inhibitor, AZD5438) in proliferating IEC-6 cells triggered DNA damage, activated p53 signaling, inhibited proliferation, and induced apoptosis. By contrast, inhibition of Cdk2 (with NU6140) increased p53 protein and activity, inhibited proliferation, but had no effect on apoptosis. Notably, AZD5438 sensitized, whereas, NU6140 rescued proliferating IEC-6 cells from CPT-induced apoptosis. However, in colon carcinoma (Caco2) cells with mutant p53, treatment with either AZD5438 or NU6140 blocked proliferation, albeit more robustly with AZD5438. Both Cdk inhibitors induced apoptosis in Caco2 cells in a p53-independent manner. In serum starved quiescent IEC-6 cells, both AZD5438 and NU6140 decreased TNF- /CPT-induced activation of p53 and, consequently, rescued cells from apoptosis, indicating that sustained Cdk activity is required for apoptosis of quiescent cells. Furthermore, AZD5438 partially reversed the protective effect of polyamine depletion whereas NU6140 had no effect. Together, these results demonstrate that Cdks possess opposing roles in the control of apoptosis in quiescent and proliferating cells. In addition, Cdk inhibitors uncouple proliferation from apoptosis in a p53-dependent manner. PMID:24242917

  16. Targeting cyclin B1 inhibits proliferation and sensitizes breast cancer cells to taxol

    International Nuclear Information System (INIS)

    Androic, Ilija; Krämer, Andrea; Yan, Ruilan; Rödel, Franz; Gätje, Regine; Kaufmann, Manfred; Strebhardt, Klaus; Yuan, Juping

    2008-01-01

    Cyclin B1, the regulatory subunit of cyclin-dependent kinase 1 (Cdk1), is essential for the transition from G2 phase to mitosis. Cyclin B1 is very often found to be overexpressed in primary breast and cervical cancer cells as well as in cancer cell lines. Its expression is correlated with the malignancy of gynecological cancers. In order to explore cyclin B1 as a potential target for gynecological cancer therapy, we studied the effect of small interfering RNA (siRNA) on different gynecological cancer cell lines by monitoring their proliferation rate, cell cycle profile, protein expression and activity, apoptosis induction and colony formation. Tumor formation in vivo was examined using mouse xenograft models. Downregulation of cyclin B1 inhibited proliferation of several breast and cervical cancer cell lines including MCF-7, BT-474, SK-BR-3, MDA-MB-231 and HeLa. After combining cyclin B1 siRNA with taxol, we observed an increased apoptotic rate accompanied by an enhanced antiproliferative effect in breast cancer cells. Furthermore, control HeLa cells were progressively growing, whereas the tumor growth of HeLa cells pre-treated with cyclin B1 siRNA was strongly inhibited in nude mice, indicating that cyclin B1 is indispensable for tumor growth in vivo. Our data support the notion of cyclin B1 being essential for survival and proliferation of gynecological cancer cells. Concordantly, knockdown of cyclin B1 inhibits proliferation in vitro as well as in vivo. Moreover, targeting cyclin B1 sensitizes breast cancer cells to taxol, suggesting that specific cyclin B1 targeting is an attractive strategy for the combination with conventionally used agents in gynecological cancer therapy

  17. Targeting cyclin B1 inhibits proliferation and sensitizes breast cancer cells to taxol

    Directory of Open Access Journals (Sweden)

    Strebhardt Klaus

    2008-12-01

    Full Text Available Abstract Background Cyclin B1, the regulatory subunit of cyclin-dependent kinase 1 (Cdk1, is essential for the transition from G2 phase to mitosis. Cyclin B1 is very often found to be overexpressed in primary breast and cervical cancer cells as well as in cancer cell lines. Its expression is correlated with the malignancy of gynecological cancers. Methods In order to explore cyclin B1 as a potential target for gynecological cancer therapy, we studied the effect of small interfering RNA (siRNA on different gynecological cancer cell lines by monitoring their proliferation rate, cell cycle profile, protein expression and activity, apoptosis induction and colony formation. Tumor formation in vivo was examined using mouse xenograft models. Results Downregulation of cyclin B1 inhibited proliferation of several breast and cervical cancer cell lines including MCF-7, BT-474, SK-BR-3, MDA-MB-231 and HeLa. After combining cyclin B1 siRNA with taxol, we observed an increased apoptotic rate accompanied by an enhanced antiproliferative effect in breast cancer cells. Furthermore, control HeLa cells were progressively growing, whereas the tumor growth of HeLa cells pre-treated with cyclin B1 siRNA was strongly inhibited in nude mice, indicating that cyclin B1 is indispensable for tumor growth in vivo. Conclusion Our data support the notion of cyclin B1 being essential for survival and proliferation of gynecological cancer cells. Concordantly, knockdown of cyclin B1 inhibits proliferation in vitro as well as in vivo. Moreover, targeting cyclin B1 sensitizes breast cancer cells to taxol, suggesting that specific cyclin B1 targeting is an attractive strategy for the combination with conventionally used agents in gynecological cancer therapy.

  18. Effects of cyclin D1 gene silencing on cell proliferation, cell cycle, and apoptosis of hepatocellular carcinoma cells.

    Science.gov (United States)

    Chen, Jin; Li, Xue; Cheng, Qi; Ning, Deng; Ma, Jie; Zhang, Zhi-Ping; Chen, Xiao-Ping; Jiang, Li

    2018-02-01

    This study aims to investigate the effects of Cyclin D1 silencing on cell cycle, cell proliferation, and apoptosis of hepatocellular carcinoma cells (HCC). Cells were divided into the blank group, negative control group (HCC cells transfected with control shRNA), Cyclin D1 shRNA group (HCC cells transfected with Cyclin D1 shRNA), and the normal group (human normal liver L-02 cells). Expressions of Cyclin D1, Caspase-3, Bcl-2, and C-myc were detected by RT-qPCR and Western blotting. Cell proliferation was detected by Cell Counting Kit-8. Cell cycle and apoptosis were detected by flow cytometry. Tumor xenograft in nude mice was performed to detect in vivo tumorigenesis. HCC tissues and HCC cells exhibited elevated expression levels of Cyclin D1. Cyclin D1 expression levels was found to be correlated with tumor size and tumor staging. Compared with the normal group, the blank group showed enhanced cell proliferation, a reduction in the amount of cells in G0/G1 phase, increased number cells in S and G2/M phase, reduced apoptosis, elevated expressions of Cyclin D1, Bcl-2, and C-myc, decreased Caspase-3 activity and significant tumorigenicity. In comparison with the blank group, the Cyclin D1 shRNA group revealed weakened cell proliferation, reduced cells in S and G2/M phase, increased cells in G0/G1 phase, increased Annexin V positive cell ratio, decreased expression of Cyclin D1, Bcl-2, and C-myc, elevated Caspase-3 activity and inhibited tumorigenicity. In conclusion, Cyclin D1 gene silencing suppresses cell proliferation and inhibits cell apoptosis, which may be a new target approach in the treatment and management for HCC. © 2017 Wiley Periodicals, Inc.

  19. Stimulation of hERG1 channel activity promotes a calcium-dependent degradation of cyclin E2, but not cyclin E1, in breast cancer cells.

    Science.gov (United States)

    Perez-Neut, Mathew; Shum, Andrew; Cuevas, Bruce D; Miller, Richard; Gentile, Saverio

    2015-01-30

    Cyclin E2 gene amplification, but not cyclin E1, has been recently defined as marker for poor prognosis in breast cancer, and appears to play a major role in proliferation and therapeutic resistance in several breast cancer cells. Our laboratory has previously reported that stimulation of the hERG1 potassium channel with selective activators led to down-regulation of cyclin E2 in breast cancer cells. In this work, we demonstrate that stimulation of hERG1 promotes an ubiquitin-proteasome-dependent degradation of cyclin E2 in multiple breast cancer cell lines representing Luminal A, HER2+ and Trastuzumab-resistant breast cancer cells. In addition we have also reveal that hERG1 stimulation induces an increase in intracellular calcium that is required for cyclin E2 degradation. This novel function for hERG1 activity was specific for cyclin E2, as cyclins A, B, D E1 were unaltered by the treatment. Our results reveal a novel mechanism by which hERG1 activation impacts the tumor marker cyclin E2 that is independent of cyclin E1, and suggest a potential therapeutic use for hERG1 channel activators.

  20. miR-340 inhibits glioblastoma cell proliferation by suppressing CDK6, cyclin-D1 and cyclin-D2

    Energy Technology Data Exchange (ETDEWEB)

    Li, Xuesong; Gong, Xuhai [Department of Neurology, Daqing Oilfield General Hospital, Daqing, Heilongjiang 163001 (China); Chen, Jing [Department of Neurology, Daqing Longnan Hospital, Daqing, Heilongjiang, 163001 China (China); Zhang, Jinghui [Department of Cardiology, The Fourth Hospital of Harbin City, Harbin, Heilongjiang 150026 (China); Sun, Jiahang [Department of Neurosurgery, The Second Affiliated Hospital of Harbin Medical University, Harbin, Heilongjiang 150086 (China); Guo, Mian, E-mail: guomian_hyd@163.com [Department of Neurosurgery, The Second Affiliated Hospital of Harbin Medical University, Harbin, Heilongjiang 150086 (China)

    2015-05-08

    Glioblastoma development is often associated with alteration in the activity and expression of cell cycle regulators, such as cyclin-dependent kinases (CKDs) and cyclins, resulting in aberrant cell proliferation. Recent studies have highlighted the pivotal roles of miRNAs in controlling the development and growth of glioblastoma. Here, we provide evidence for a function of miR-340 in the inhibition of glioblastoma cell proliferation. We found that miR-340 is downregulated in human glioblastoma tissue samples and several established glioblastoma cell lines. Proliferation and neurosphere formation assays revealed that miR-340 plays an oncosuppressive role in glioblastoma, and that its ectopic expression causes significant defect in glioblastoma cell growth. Further, using bioinformatics, luciferase assay and western blot, we found that miR-340 specifically targets the 3′UTRs of CDK6, cyclin-D1 and cyclin-D2, leading to the arrest of glioblastoma cells in the G0/G1 cell cycle phase. Confirming these results, we found that re-introducing CDK6, cyclin-D1 or cyclin-D2 expression partially, but significantly, rescues cells from the suppression of cell proliferation and cell cycle arrest mediated by miR-340. Collectively, our results demonstrate that miR-340 plays a tumor-suppressive role in glioblastoma and may be useful as a diagnostic biomarker and/or a therapeutic avenue for glioblastoma. - Highlights: • miR-340 is downregulated in glioblastoma samples and cell lines. • miR-340 inhibits glioblastoma cell proliferation. • miR-340 directly targets CDK6, cyclin-D1, and cyclin-D2. • miR-340 regulates glioblastoma cell proliferation via CDK6, cyclin-D1 and cyclin-D2.

  1. Cyclin D1 fine-tunes the neurogenic output of embryonic retinal progenitor cells

    Directory of Open Access Journals (Sweden)

    Choi Yoon

    2009-05-01

    Full Text Available Abstract Background Maintaining the correct balance of proliferation versus differentiation in retinal progenitor cells (RPCs is essential for proper development of the retina. The cell cycle regulator cyclin D1 is expressed in RPCs, and mice with a targeted null allele at the cyclin D1 locus (Ccnd1-/- have microphthalmia and hypocellular retinas, the latter phenotype attributed to reduced RPC proliferation and increased photoreceptor cell death during the postnatal period. How cyclin D1 influences RPC behavior, especially during the embryonic period, is unclear. Results In this study, we show that embryonic RPCs lacking cyclin D1 progress through the cell cycle at a slower rate and exit the cell cycle at a faster rate. Consistent with enhanced cell cycle exit, the relative proportions of cell types born in the embryonic period, such as retinal ganglion cells and photoreceptor cells, are increased. Unexpectedly, cyclin D1 deficiency decreases the proportions of other early born retinal neurons, namely horizontal cells and specific amacrine cell types. We also found that the laminar positioning of horizontal cells and other cell types is altered in the absence of cyclin D1. Genetically replacing cyclin D1 with cyclin D2 is not efficient at correcting the phenotypes due to the cyclin D1 deficiency, which suggests the D-cyclins are not fully redundant. Replacement with cyclin E or inactivation of cyclin-dependent kinase inhibitor p27Kip1 restores the balance of RPCs and retinal cell types to more normal distributions, which suggests that regulation of the retinoblastoma pathway is an important function for cyclin D1 during embryonic retinal development. Conclusion Our findings show that cyclin D1 has important roles in RPC cell cycle regulation and retinal histogenesis. The reduction in the RPC population due to a longer cell cycle time and to an enhanced rate of cell cycle exit are likely to be the primary factors driving retinal hypocellularity

  2. Promoter de-methylation of cyclin D2 by sulforaphane in prostate cancer cells.

    Science.gov (United States)

    Hsu, Anna; Wong, Carmen P; Yu, Zhen; Williams, David E; Dashwood, Roderick H; Ho, Emily

    2011-01-01

    Sulforaphane (SFN), an isothiocyanate derived from cruciferous vegetables, induces potent anti-proliferative effects in prostate cancer cells. One mechanism that may contribute to the anti-proliferative effects of SFN is the modulation of epigenetic marks, such as inhibition of histone deacetylase (HDAC) enzymes. However, the effects of SFN on other common epigenetic marks such as DNA methylation are understudied. Promoter hyper-methylation of cyclin D2, a major regulator of cell cycle, is correlated with prostate cancer progression, and restoration of cyclin D2 expression exerts anti-proliferative effects on LnCap prostate cancer cells. Our study aimed to investigate the effects of SFN on DNA methylation status of cyclin D2 promoter, and how alteration in promoter methylation impacts cyclin D2 gene expression in LnCap cells. We found that SFN significantly decreased the expression of DNA methyltransferases (DNMTs), especially DNMT1 and DNMT3b. Furthermore, SFN significantly decreased methylation in cyclin D2 promoter regions containing c-Myc and multiple Sp1 binding sites. Reduced methlyation of cyclin D2 promoter corresponded to an increase in cyclin D2 transcript levels, suggesting that SFN may de-repress methylation-silenced cyclin D2 by impacting epigenetic pathways. Our results demonstrated the ability of SFN to epigenetically modulate cyclin D2 expression, and provide novel insights into the mechanisms by which SFN may regulate gene expression as a prostate cancer chemopreventive agent.

  3. Promoter de-methylation of cyclin D2 by sulforaphane in prostate cancer cells

    Directory of Open Access Journals (Sweden)

    Hsu Anna

    2011-10-01

    Full Text Available Abstract Sulforaphane (SFN, an isothiocyanate derived from cruciferous vegetables, induces potent anti-proliferative effects in prostate cancer cells. One mechanism that may contribute to the anti-proliferative effects of SFN is the modulation of epigenetic marks, such as inhibition of histone deacetylase (HDAC enzymes. However, the effects of SFN on other common epigenetic marks such as DNA methylation are understudied. Promoter hyper-methylation of cyclin D2, a major regulator of cell cycle, is correlated with prostate cancer progression, and restoration of cyclin D2 expression exerts anti-proliferative effects on LnCap prostate cancer cells. Our study aimed to investigate the effects of SFN on DNA methylation status of cyclin D2 promoter, and how alteration in promoter methylation impacts cyclin D2 gene expression in LnCap cells. We found that SFN significantly decreased the expression of DNA methyltransferases (DNMTs, especially DNMT1 and DNMT3b. Furthermore, SFN significantly decreased methylation in cyclin D2 promoter regions containing c-Myc and multiple Sp1 binding sites. Reduced methlyation of cyclin D2 promoter corresponded to an increase in cyclin D2 transcript levels, suggesting that SFN may de-repress methylation-silenced cyclin D2 by impacting epigenetic pathways. Our results demonstrated the ability of SFN to epigenetically modulate cyclin D2 expression, and provide novel insights into the mechanisms by which SFN may regulate gene expression as a prostate cancer chemopreventive agent.

  4. Analysis of cell cycle-related proteins in gastric intramucosal differentiated-type cancers based on mucin phenotypes: a novel hypothesis of early gastric carcinogenesis based on mucin phenotype

    Directory of Open Access Journals (Sweden)

    Matsushita Hiroo

    2010-06-01

    Full Text Available Abstract Background Abnormalities of cell cycle regulators are common features in human cancers, and several of these factors are associated with the early development of gastric cancers. However, recent studies have shown that gastric cancer tumorigenesis was characterized by mucin expression. Thus, expression patterns of cell cycle-related proteins were investigated in the early phase of differentiated-type gastric cancers to ascertain any mechanistic relationships with mucin phenotypes. Methods Immunostaining for Cyclins D1, A, E, and p21, p27, p53 and β-catenin was used to examine impairments of the cell cycle in 190 gastric intramucosal differentiated-type cancers. Mucin phenotypes were determined by the expressions of MUC5AC, MUC6, MUC2 and CD10. A Ki-67 positive rate (PR was also examined. Results Overexpressions of p53, cyclin D1 and cyclin A were significantly more frequent in a gastric phenotype than an intestinal phenotype. Cyclin A was overexpressed in a mixed phenotype compared with an intestinal phenotype, while p27 overexpression was more frequent in an intestinal phenotype than in a mixed phenotype. Reduction of p21 was a common feature of the gastric intramucosal differentiated-type cancers examined. Conclusions Our results suggest that the levels of some cell cycle regulators appear to be associated with mucin phenotypes of early gastric differentiated-type cancers.

  5. Cell Cycle Related Differentiation of Bone Marrow Cells into Lung Cells

    Energy Technology Data Exchange (ETDEWEB)

    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.

  6. Alfalfa cyclins: differential expression during the cell cycle and in plant organs.

    Science.gov (United States)

    Hirt, H; Mink, M; Pfosser, M; Bögre, L; Györgyey, J; Jonak, C; Gartner, A; Dudits, D; Heberle-Bors, E

    1992-12-01

    Cell division in eukaryotes is mediated by the action of the mitosis promoting factor, which is composed of the CDC2 protein kinase and one of the various mitotic cyclins. We have recently isolated a cdc2 gene from alfalfa. Here, we report the isolation of two cyclin genes, cycMs1 and cycMs2, from alfalfa. The cycMs2 gene shows highest similarity to type B cyclins. In contrast, the predicted amino acid sequence of the cycMs1 gene shows similar homology scores to cyclins of all types (25 to 35%). Both genes are expressed in dividing suspension cultured cells but cease to be expressed when the cells enter stationary phase. In synchronized alfalfa suspension cultured cells, the mRNAs of cycMs1 and cycMs2 show maximal expression in the G2 and M phases. Transcripts of cycMs2 are found only in late G2 and M phase cells, an expression pattern typical for cyclin B genes, whereas cycMs1 appears with the onset of G2. This pattern indicates that alfalfa cycMs1 and cycMs2 belong to different classes of cyclins. In young leaves, expression of both genes is high, whereas in mature leaves no transcripts can be detected, indicating that the two cyclin genes are true cell division markers at the mRNA level. In other organs, a more complex expression pattern of the two cyclin genes was found.

  7. Alfalfa cyclins: differential expression during the cell cycle and in plant organs

    OpenAIRE

    Hirt, H.; Mink, M.; Pfosser, M.; Bogre, L.; Gyorgyey, J.; Jonak, C.; Gartner, A.; Dudits, D.; Heberle-Bors, E.

    1992-01-01

    Cell division in eukaryotes is mediated by the action of the mitosis promoting factor, which is composed of the CDC2 protein kinase and one of the various mitotic cyclins. We have recently isolated a cdc2 gene from alfalfa. Here, we report the isolation of two cyclin genes, cycMs1 and cycMs2, from alfalfa. The cycMs2 gene shows highest similarity to type B cyclins. In contrast, the predicted amino acid sequence of the cycMs1 gene shows similar homology scores to cyclins of all types (25 to 35...

  8. Cyclin D1 overexpression, cell cycle progression and radiosensitivity in MBP cells

    International Nuclear Information System (INIS)

    Wu Lijun; Yu Zengliang

    2000-11-01

    Clones that exhibited a minimum of 7-8 fold cyclin D1 level above the parent cell lines or the vector control were obtained after transfected with the entire coding sequence of human 1.1 kb cyclin D1 cDNA. Studies showed that there was no significant difference in Radiosensitivity between over-expressing cyclin D1 and control cultures from either mouse or human origin. Using flow cytometry to access cell cycle distribution in the exponentially growth cultures of MCF10F-D1-21 and MCF10F-V-3, it was found that there was a 50 percent increase in the proportion of G2/M phase cells and 5.3 percent decrease in the proportion of G0/G1 phase cells in MCF10F-D1-21 comparing with MCF10F-V-3, though they were with the same proportion of cells in S phase

  9. Epidermal growth factor induces cyclin D1 in human pancreatic carcinoma: evidence for a cyclin D1-dependent cell cycle progression.

    Science.gov (United States)

    Poch, B; Gansauge, F; Schwarz, A; Seufferlein, T; Schnelldorfer, T; Ramadani, M; Beger, H G; Gansauge, S

    2001-10-01

    We recently showed that cyclin D1 is overexpressed in human pancreatic carcinoma cells, and that this overexpression correlates significantly with a poor prognosis. To assess the interrelations of epidermal growth factor (EGF), EGF receptor (EGFR), and cyclin D1 in human pancreatic carcinoma. In pancreatic carcinoma cell lines (BxPC-3, AsPC-1), cell cycle analysis revealed an increase in cells in the S/G1 phase between 18 and 30 hours after stimulation with 50 ng/mL EGF. Cyclin D1 mRNA increased after 2 hours, corresponding to an increase in cyclin D1 protein, with the maximum level between 7.5 and 10 hours after stimulation, as demonstrated by Western blot analysis. We performed immunohistochemical analysis on 61 adenocarcinoma tissues for the expression of EGF, EGFR, and cyclin D1 and demonstrated an overexpression in the tumor cells in 51%, 54%, and 62.3%, respectively, whereas normal human pancreas stained negative for all of the three factors. Interestingly, EGF and EGFR expression correlated significantly with the cyclin D1 expression in human pancreatic tumor cells (p < 0.001 and p < 0.01, respectively). These results demonstrate that cyclin D1 overexpression in the tumor cells of pancreatic carcinoma tissue is at least partly dependent on the mitogenic effects of EGF signaling through the EGFR.

  10. Fluorescent peptide biosensor for probing the relative abundance of cyclin-dependent kinases in living cells.

    Directory of Open Access Journals (Sweden)

    Laetitia Kurzawa

    Full Text Available Cyclin-dependant kinases play a central role in coordinating cell growth and division, and in sustaining proliferation of cancer cells, thereby constituting attractive pharmacological targets. However, there are no direct means of assessing their relative abundance in living cells, current approaches being limited to antigenic and proteomic analysis of fixed cells. In order to probe the relative abundance of these kinases directly in living cells, we have developed a fluorescent peptide biosensor with biligand affinity for CDKs and cyclins in vitro, that retains endogenous CDK/cyclin complexes from cell extracts, and that bears an environmentally-sensitive probe, whose fluorescence increases in a sensitive fashion upon recognition of its targets. CDKSENS was introduced into living cells, through complexation with the cell-penetrating carrier CADY2 and applied to assess the relative abundance of CDK/Cyclins through fluorescence imaging and ratiometric quantification. This peptide biosensor technology affords direct and sensitive readout of CDK/cyclin complex levels, and reports on differences in complex formation when tampering with a single CDK or cyclin. CDKSENS further allows for detection of differences between different healthy and cancer cell lines, thereby enabling to distinguish cells that express high levels of these heterodimeric kinases, from cells that present decreased or defective assemblies. This fluorescent biosensor technology provides information on the overall status of CDK/Cyclin complexes which cannot be obtained through antigenic detection of individual subunits, in a non-invasive fashion which does not require cell fixation or extraction procedures. As such it provides promising perspectives for monitoring the response to therapeutics that affect CDK/Cyclin abundance, for cell-based drug discovery strategies and fluorescence-based cancer diagnostics.

  11. Rictor regulates FBXW7-dependent c-Myc and cyclin E degradation in colorectal cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Guo, Zheng [Markey Cancer Center, The University of Kentucky, 800 Rose Street, Lexington, KY 40536 (United States); Guangdong Provincial Key Laboratory of Gastroenterology, Department of Gastroenterology, Nanfang Hospital, Southern Medical University, 1838 Guangzhou Dadao Bei, Guangzhou 510515 (China); Zhou, Yuning [Markey Cancer Center, The University of Kentucky, 800 Rose Street, Lexington, KY 40536 (United States); Evers, B. Mark [Markey Cancer Center, The University of Kentucky, 800 Rose Street, Lexington, KY 40536 (United States); Department of Surgery, The University of Kentucky, 800 Rose Street, Lexington, KY 40536 (United States); Wang, Qingding, E-mail: qingding.wang@uky.edu [Markey Cancer Center, The University of Kentucky, 800 Rose Street, Lexington, KY 40536 (United States); Department of Surgery, The University of Kentucky, 800 Rose Street, Lexington, KY 40536 (United States)

    2012-02-10

    Highlights: Black-Right-Pointing-Pointer Rictor associates with FBXW7 to form an E3 complex. Black-Right-Pointing-Pointer Knockdown of rictor decreases ubiquitination of c-Myc and cylin E. Black-Right-Pointing-Pointer Knockdown of rictor increases protein levels of c-Myc and cylin E. Black-Right-Pointing-Pointer Overexpression of rictor induces the degradation of c-Myc and cyclin E proteins. Black-Right-Pointing-Pointer Rictor regulation of c-Myc and cyclin E requires FBXW7. -- Abstract: Rictor (Rapamycin-insensitive companion of mTOR) forms a complex with mTOR and phosphorylates and activates Akt. Activation of Akt induces expression of c-Myc and cyclin E, which are overexpressed in colorectal cancer and play an important role in colorectal cancer cell proliferation. Here, we show that rictor associates with FBXW7 to form an E3 complex participating in the regulation of c-Myc and cyclin E degradation. The Rictor-FBXW7 complex is biochemically distinct from the previously reported mTORC2 and can be immunoprecipitated independently of mTORC2. Moreover, knocking down of rictor in serum-deprived colorectal cancer cells results in the decreased ubiquitination and increased protein levels of c-Myc and cyclin E while overexpression of rictor induces the degradation of c-Myc and cyclin E proteins. Genetic knockout of FBXW7 blunts the effects of rictor, suggesting that rictor regulation of c-Myc and cyclin E requires FBXW7. Our findings identify rictor as an important component of FBXW7 E3 ligase complex participating in the regulation of c-Myc and cyclin E protein ubiquitination and degradation. Importantly, our results suggest that elevated growth factor signaling may contribute to decrease rictor/FBXW7-mediated ubiquitination of c-Myc and cyclin E, thus leading to accumulation of cyclin E and c-Myc in colorectal cancer cells.

  12. Rictor regulates FBXW7-dependent c-Myc and cyclin E degradation in colorectal cancer cells

    International Nuclear Information System (INIS)

    Guo, Zheng; Zhou, Yuning; Evers, B. Mark; Wang, Qingding

    2012-01-01

    Highlights: ► Rictor associates with FBXW7 to form an E3 complex. ► Knockdown of rictor decreases ubiquitination of c-Myc and cylin E. ► Knockdown of rictor increases protein levels of c-Myc and cylin E. ► Overexpression of rictor induces the degradation of c-Myc and cyclin E proteins. ► Rictor regulation of c-Myc and cyclin E requires FBXW7. -- Abstract: Rictor (Rapamycin-insensitive companion of mTOR) forms a complex with mTOR and phosphorylates and activates Akt. Activation of Akt induces expression of c-Myc and cyclin E, which are overexpressed in colorectal cancer and play an important role in colorectal cancer cell proliferation. Here, we show that rictor associates with FBXW7 to form an E3 complex participating in the regulation of c-Myc and cyclin E degradation. The Rictor–FBXW7 complex is biochemically distinct from the previously reported mTORC2 and can be immunoprecipitated independently of mTORC2. Moreover, knocking down of rictor in serum-deprived colorectal cancer cells results in the decreased ubiquitination and increased protein levels of c-Myc and cyclin E while overexpression of rictor induces the degradation of c-Myc and cyclin E proteins. Genetic knockout of FBXW7 blunts the effects of rictor, suggesting that rictor regulation of c-Myc and cyclin E requires FBXW7. Our findings identify rictor as an important component of FBXW7 E3 ligase complex participating in the regulation of c-Myc and cyclin E protein ubiquitination and degradation. Importantly, our results suggest that elevated growth factor signaling may contribute to decrease rictor/FBXW7-mediated ubiquitination of c-Myc and cyclin E, thus leading to accumulation of cyclin E and c-Myc in colorectal cancer cells.

  13. Cyclin D3 expression in non-Hodgkin lymphoma. Correlation with other cell cycle regulators and clinical features

    DEFF Research Database (Denmark)

    Møller, Michael Boe; Nielsen, O; Pedersen, Niels Tinggaard

    2001-01-01

    analyzed immunohistochemically for cyclin D3 expression. In 43 lymphomas (21.7%), cyclin D3 was overexpressed. T-cell lymphomas more frequently overexpressed cyclin D3 than B-cell lymphomas. Furthermore, cyclin D3-overexpressing indolent lymphomas were associated with higher proliferation rate, higher p21......Waf1 expression, lower p27Kip1 expression, and altered p53. Cyclin D3 overexpression identified a subgroup of patients with indolent B-cell lymphoma with adverse clinical features: patients were older, more frequently had "B" symptoms and extranodal involvement, and were more frequently in the high...

  14. A conserved cyclin-binding domain determines functional interplay between anaphase-promoting complex-Cdh1 and cyclin A-Cdk2 during cell cycle progression

    DEFF Research Database (Denmark)

    Lukas, C; Kramer, E R; Peters, J M

    2001-01-01

    Periodic activity of the anaphase-promoting complex (APC) ubiquitin ligase determines progression through multiple cell cycle transitions by targeting cell cycle regulators for destruction. At the G(1)/S transition, phosphorylation-dependent dissociation of the Cdh1-activating subunit inhibits...... the APC, allowing stabilization of proteins required for subsequent cell cycle progression. Cyclin-dependent kinases (CDKs) that initiate and maintain Cdh1 phosphorylation have been identified. However, the issue of which cyclin-CDK complexes are involved has been a matter of debate, and the mechanism...... of how cyclin-CDKs interact with APC subunits remains unresolved. Here we substantiate the evidence that mammalian cyclin A-Cdk2 prevents unscheduled APC reactivation during S phase by demonstrating its periodic interaction with Cdh1 at the level of endogenous proteins. Moreover, we identified...

  15. Rescue of cyclin D1 deficiency by knockin cyclin E

    NARCIS (Netherlands)

    Geng, Y.; Whoriskey, W.; Park, M.Y.; Bronson, R.T.; Medema, R.H.; Li, T.; Weinberg, R.A.; Sicinski, P.

    1999-01-01

    D-type cyclins and cyclin E represent two very distinct classes of mammalian G1 cyclins. We have generated a mouse strain in which the coding sequences of the cyclin D1 gene (Ccnd1) have been deleted and replaced by those of human cyclin E (CCNE). In the tissues and cells of these mice, the

  16. Implications of caspase-dependent proteolytic cleavage of cyclin A1 in DNA damage-induced cell death

    Energy Technology Data Exchange (ETDEWEB)

    Woo, Sang Hyeok; Seo, Sung-Keum [Division of Radiation Cancer Research, Korea Institute of Radiological and Medical Sciences, 215-4 Gongneung-dong, Nowon-gu, Seoul (Korea, Republic of); An, Sungkwan; Choe, Tae-Boo [Department of Microbiological Engineering, Kon-Kuk University, Gwangjin-gu, Seoul (Korea, Republic of); Hong, Seok-Il [Department of Laboratory Medicine, Korea Cancer Center Hospital, Korea Institute of Radiological and Medical Sciences, 215-4 Gongneung-dong, Nowon-gu, Seoul (Korea, Republic of); Lee, Yun-Han, E-mail: yhlee87@yuhs.ac [Department of Radiation Oncology, College of Medicine, Yonsei University, 250 Seongsan-no, Seodaemun-gu, Seoul (Korea, Republic of); Park, In-Chul, E-mail: parkic@kcch.re.kr [Division of Radiation Cancer Research, Korea Institute of Radiological and Medical Sciences, 215-4 Gongneung-dong, Nowon-gu, Seoul (Korea, Republic of)

    2014-10-24

    Highlights: • Caspase-1 mediates doxorubicin-induced downregulation of cyclin A1. • Active caspase-1 effectively cleaved cyclin A1 at D165. • Cyclin A1 expression is involved in DNA damage-induced cell death. - Abstract: Cyclin A1 is an A-type cyclin that directly binds to CDK2 to regulate cell-cycle progression. In the present study, we found that doxorubicin decreased the expression of cyclin A1 at the protein level in A549 lung cancer cells, while markedly downregulating its mRNA levels. Interestingly, doxorubicin upregulated caspase-1 in a concentration-dependent manner, and z-YAVD-fmk, a specific inhibitor of caspase-1, reversed the doxorubicin-induced decrease in cyclin A1 in A549 lung cancer and MCF7 breast cancer cells. Active caspase-1 effectively cleaved cyclin A1 at D165 into two fragments, which in vitro cleavage assays showed were further cleaved by caspase-3. Finally, we found that overexpression of cyclin A1 significantly reduced the cytotoxicity of doxorubicin, and knockdown of cyclin A1 by RNA interference enhanced the sensitivity of cells to ionizing radiation. Our data suggest a new mechanism for the downregulation of cyclin A1 by DNA-damaging stimuli that could be intimately involved in the cell death induced by DNA damage-inducing stimuli, including doxorubicin and ionizing radiation.

  17. Cyclin D1 Expression and Its Correlation with Histopathological Differentiation in Oral Squamous Cell Carcinoma

    Directory of Open Access Journals (Sweden)

    Swati Saawarn

    2012-01-01

    Full Text Available Background. Cyclin D1 regulates the G1 to S transition of cell cycle. Its deregulation or overexpression may lead to disturbance in the normal cell cycle control and tumour formation. Overexpression of cyclin D1 has been reported in various tumors of diverse histogenesis. This case control retrospective study was carried out to study the immunohistochemical reactivity and expression of cyclin D1 and its association with site, clinical staging, and histopathological differentiation of oral squamous cell carcinoma (OSCC. Methods. Forty formalin-fixed paraffin-embedded tissue blocks of biopsy specimens of oral squamous cell carcinoma were immunohistochemically evaluated for expression of cyclin D1. Results. Cyclin D1 expression was seen in 45% cases of OSCC. It did not correlate with site and clinical staging. Highest expression was seen in well-differentiated, followed by moderately differentiated, and poorly differentiated squamous cell carcinomas, with a statistically significant correlation. Conclusion. Cyclin D1 expression significantly increases with increase in differentiation.

  18. MicroRNA-195 inhibits the proliferation of human glioma cells by directly targeting cyclin D1 and cyclin E1.

    Directory of Open Access Journals (Sweden)

    Wang Hui

    Full Text Available Glioma proliferation is a multistep process during which a sequence of genetic and epigenetic alterations randomly occur to affect the genes controlling cell proliferation, cell death and genetic stability. microRNAs are emerging as important epigenetic modulators of multiple target genes, leading to abnormal cellular signaling involving cellular proliferation in cancers.In the present study, we found that expression of miR-195 was markedly downregulated in glioma cell lines and human primary glioma tissues, compared to normal human astrocytes and matched non-tumor associated tissues. Upregulation of miR-195 dramatically reduced the proliferation of glioma cells. Flow cytometry analysis showed that ectopic expression of miR-195 significantly decreased the percentage of S phase cells and increased the percentage of G1/G0 phase cells. Overexpression of miR-195 dramatically reduced the anchorage-independent growth ability of glioma cells. Furthermore, overexpression of miR-195 downregulated the levels of phosphorylated retinoblastoma (pRb and proliferating cell nuclear antigen (PCNA in glioma cells. Conversely, inhibition of miR-195 promoted cell proliferation, increased the percentage of S phase cells, reduced the percentage of G1/G0 phase cells, enhanced anchorage-independent growth ability, upregulated the phosphorylation of pRb and PCNA in glioma cells. Moreover, we show that miR-195 inhibited glioma cell proliferation by downregulating expression of cyclin D1 and cyclin E1, via directly targeting the 3'-untranslated regions (3'-UTR of cyclin D1 and cyclin E1 mRNA. Taken together, our results suggest that miR-195 plays an important role to inhibit the proliferation of glioma cells, and present a novel mechanism for direct miRNA-mediated suppression of cyclin D1 and cyclin E1 in glioma.

  19. A novel role for the cell cycle regulatory complex cyclin D1?CDK4 in gluconeogenesis

    OpenAIRE

    Hosooka, Tetsuya; Ogawa, Wataru

    2015-01-01

    Dysregulation of gluconeogenesis is a key pathological feature of type 2 diabetes. However, the molecular mechanisms underlying the regulation of gluconeogenesis remain unclear. Bhalla et?al. recently reported that cyclin D1 suppresses hepatic gluconeogenesis through CDK4?dependent phosphorylation of PGC1alpha and consequent inhibition of its activity. The cyclin D1?CDK4 might thus serve as an important link between the cell cycle and control of energy metabolism through modulation of PGC1alp...

  20. A novel role for the cell cycle regulatory complex cyclin D1-CDK4 in gluconeogenesis.

    Science.gov (United States)

    Hosooka, Tetsuya; Ogawa, Wataru

    2016-01-01

    Dysregulation of gluconeogenesis is a key pathological feature of type 2 diabetes. However, the molecular mechanisms underlying the regulation of gluconeogenesis remain unclear. Bhalla et al. recently reported that cyclin D1 suppresses hepatic gluconeogenesis through CDK4-dependent phosphorylation of PGC1alpha and consequent inhibition of its activity. The cyclin D1-CDK4 might thus serve as an important link between the cell cycle and control of energy metabolism through modulation of PGC1alpha activity.

  1. PKCeta enhances cell cycle progression, the expression of G1 cyclins and p21 in MCF-7 cells.

    Science.gov (United States)

    Fima, E; Shtutman, M; Libros, P; Missel, A; Shahaf, G; Kahana, G; Livneh, E

    2001-10-11

    Protein kinase C encodes a family of enzymes implicated in cellular differentiation, growth control and tumor promotion. However, not much is known with respect to the molecular mechanisms that link protein kinase C to cell cycle control. Here we report that the expression of PKCeta in MCF-7 cells, under the control of a tetracycline-responsive inducible promoter, enhanced cell growth and affected the cell cycle at several points. The induced expression of another PKC isoform, PKCdelta, in MCF-7 cells had opposite effects and inhibited their growth. PKCeta expression activated cellular pathways in these cells that resulted in the increased expression of the G1 phase cyclins, cyclin D and cyclin E. Expression of the cyclin-dependent kinase inhibitor p21(WAF1) was also specifically elevated in PKCeta expressing cells, but its overall effects were not inhibitory. Although, the protein levels of the cyclin-dependent kinase inhibitor p27(KIP1) were not altered by the induced expression of PKCeta, the cyclin E associated Cdk2 kinase activity was in correlation with the p27(KIP1) bound to the cyclin E complex and not by p21(WAF1) binding. PKCeta expression enhanced the removal of p27(KIP1) from this complex, and its re-association with the cyclin D/Cdk4 complex. Reduced binding of p27(KIP1) to the cyclin D/Cdk4 complex at early time points of the cell cycle also enhanced the activity of this complex, while at later time points the decrease in bound p21(WAF1) correlated with its increased activity in PKCeta-expressing cells. Thus, PKCeta induces altered expression of several cell cycle functions, which may contribute to its ability to affect cell growth.

  2. Progesterone Receptor–Cyclin D1 Complexes Induce Cell Cycle–Dependent Transcriptional Programs in Breast Cancer Cells

    Science.gov (United States)

    Dressing, Gwen E.; Knutson, Todd P.; Schiewer, Matthew J.; Daniel, Andrea R.; Hagan, Christy R.; Diep, Caroline H.; Knudsen, Karen E.

    2014-01-01

    The progesterone receptor (PR) and its coactivators are direct targets of activated cyclin-dependent kinases (CDKs) in response to peptide growth factors, progesterone, and deregulation of cell cycle inhibitors. Herein, using the T47D breast cancer model, we probed mechanisms of cell cycle–dependent PR action. In the absence of exogenous progestin, the PR is specifically phosphorylated during the G2/M phase. Accordingly, numerous PR target genes are cell cycle regulated, including HSPB8, a heat-shock protein whose high expression is associated with tamoxifen resistance. Progestin-induced HSPB8 expression required cyclin D1 and was insensitive to antiestrogens but blocked by antiprogestins or inhibition of specificity factor 1 (SP1). HSPB8 expression increased with or without ligand when cells were G2/M synchronized or contained high levels of cyclin D1. Knockdown of PRs abrogated ligand-independent HSPB8 expression in synchronized cells. Notably, PRs and cyclin D1 copurified in whole-cell lysates of transiently transfected COS-1 cells and in PR-positive T47D breast cancer cells expressing endogenous cyclin D1. PRs, cyclin D1, and SP1 were recruited to the HSPB8 promoter in progestin-treated T47D breast cancer cells. Mutation of PR Ser345 to Ala (S345A) or inhibition of CDK2 activity using roscovitine disrupted PR/cyclin D1 interactions with DNA and blocked HSPB8 mRNA expression. Interaction of phosphorylated PRs with SP1 and cyclin D1 provides a mechanism for targeting transcriptionally active PRs to selected gene promoters relevant to breast cancer progression. Understanding the functional linkage between PRs and cell cycle regulatory proteins will provide keys to targeting novel PR/cyclin D1 cross talk in both hormone-responsive disease and HSPB8-high refractory disease with high HSPB8 expression. PMID:24606123

  3. Differentiation-inducing factor-1 suppresses gene expression of cyclin D1 in tumor cells

    International Nuclear Information System (INIS)

    Yasmin, Tania; Takahashi-Yanaga, Fumi; Mori, Jun; Miwa, Yoshikazu; Hirata, Masato; Watanabe, Yutaka; Morimoto, Sachio; Sasaguri, Toshiyuki

    2005-01-01

    To determine the mechanism by which differentiation-inducing factor-1 (DIF-1), a morphogen of Dictyostelium discoideum, inhibits tumor cell proliferation, we examined the effect of DIF-1 on the gene expression of cyclin D1. DIF-1 strongly reduced the expression of cyclin D1 mRNA and correspondingly decreased the amount of β-catenin in HeLa cells and squamous cell carcinoma cells. DIF-1 activated glycogen synthase kinase-3β (GSK-3β) and inhibition of GSK-3β attenuated the DIF-1-induced β-catenin degradation, indicating the involvement of GSK-3β in this effect. Moreover, DIF-1 reduced the activities of T-cell factor (TCF)/lymphoid enhancer factor (LEF) reporter plasmid and a reporter gene driven by the human cyclin D1 promoter. Eliminating the TCF/LEF consensus site from the cyclin D1 promoter diminished the effect of DIF-1. These results suggest that DIF-1 inhibits Wnt/β-catenin signaling, resulting in the suppression of cyclin D1 promoter activity

  4. Mantle cell lymphoma in cyclin D1 transgenic mice with Bim-deficient B cells.

    Science.gov (United States)

    Katz, Samuel G; Labelle, James L; Meng, Hailong; Valeriano, Regina P; Fisher, Jill K; Sun, Heather; Rodig, Scott J; Kleinstein, Steven H; Walensky, Loren D

    2014-02-06

    Mantle cell lymphoma (MCL) is a highly aggressive B-cell lymphoma resistant to conventional chemotherapy. Although defined by the characteristic t(11;14) translocation, MCL has not been recapitulated in transgenic mouse models of cyclin D1 overexpression alone. Indeed, several genetic aberrations have been identified in MCL that may contribute to its pathogenesis and chemoresistance. Of particular interest is the frequent biallelic deletion of the proapoptotic BCL-2 family protein BIM. BIM exerts its pro-death function via its α-helical BH3 death domain that has the dual capacity to inhibit antiapoptotic proteins such as BCL-2 and MCL-1 and directly trigger proapoptotic proteins such as the mitochondrial executioner protein BAX. To evaluate a functional role for Bim deletion in the pathogenesis of MCL, we generated cyclin D1-transgenic mice harboring Bim-deficient B cells. In response to immunization, Eμ(CycD1)CD19(CRE)Bim(fl/fl) mice manifested selective expansion of their splenic mantle zone compartment. Three distinct immune stimulation regimens induced lymphomas with histopathologic and molecular features of human MCL in a subset of mice. Thus, deletion of Bim in B cells, in the context of cyclin D1 overexpression, disrupts a critical control point in lymphoid maturation and predisposes to the development of MCL. This genetic proof of concept for MCL pathogenesis suggests an opportunity to reactivate the death pathway by pharmacologic mimicry of proapoptotic BIM.

  5. Cyclin D1 in ASM Cells from Asthmatics Is Insensitive to Corticosteroid Inhibition.

    Science.gov (United States)

    Allen, Jodi C; Seidel, Petra; Schlosser, Tobias; Ramsay, Emma E; Ge, Qi; Ammit, Alaina J

    2012-01-01

    Hyperplasia of airway smooth muscle (ASM) is a feature of the remodelled airway in asthmatics. We examined the antiproliferative effectiveness of the corticosteroid dexamethasone on expression of the key regulator of G(1) cell cycle progression-cyclin D1-in ASM cells from nonasthmatics and asthmatics stimulated with the mitogen platelet-derived growth factor BB. While cyclin D1 mRNA and protein expression were repressed in cells from nonasthmatics in contrast, cyclin D1 expression in asthmatics was resistant to inhibition by dexamethasone. This was independent of a repressive effect on glucocorticoid receptor translocation. Our results corroborate evidence demonstrating that corticosteroids inhibit mitogen-induced proliferation only in ASM cells from subjects without asthma and suggest that there are corticosteroid-insensitive proliferative pathways in asthmatics.

  6. A critical role for FBXW8 and MAPK in cyclin D1 degradation and cancer cell proliferation.

    Directory of Open Access Journals (Sweden)

    Hiroshi Okabe

    2006-12-01

    Full Text Available Cyclin D1 regulates G1 progression. Its transcriptional regulation is well understood. However, the mechanism underlying cyclin D1 ubiquitination and its subsequent degradation is not yet clear. We report that cyclin D1 undergoes increased degradation in the cytoplasm during S phase in a variety of cancer cells. This is mediated by phosphorylation at Thr286 through the activity of the Ras/Raf/MEK/ERK cascade and the F-box protein FBXW8, which is an E3 ligase. The majority of FBXW8 is expressed in the cytoplasm during G1 and S phase. In contrast, cyclin D1 accumulates in the nucleus during G1 phase and exits into the cytoplasm in S phase. Increased cyclin D1 degradation is linked to association with FBXW8 in the cytoplasm, and enhanced phosphorylation of cyclin D1 through sustained ERK1/2 signaling. Depletion of FBXW8 caused a significant accumulation of cyclin D1, as well as sequestration of CDK1 in the cytoplasm. This resulted in a severe reduction of cell proliferation. These effects could be rescued by constitutive nuclear expression of cyclin D1-T286A. Thus, FBXW8 plays an essential role in cancer cell proliferation through proteolysis of cyclin D1. It may present new opportunities to develop therapies targeting destruction of cyclin D1 or its regulator E3 ligase selectively.

  7. Rb and FZR1/Cdh1 determine CDK4/6-cyclin D requirement in C. elegans and human cancer cells

    NARCIS (Netherlands)

    The, Inge; Ruijtenberg, Suzan; Bouchet, Benjamin P; Cristobal, Alba; Prinsen, Martine B W; van Mourik, Tim; Koreth, John; Xu, Huihong; Heck, Albert J R; Akhmanova, Anna; Cuppen, Edwin; Boxem, Mike; Muñoz, Javier; van den Heuvel, Sander

    2015-01-01

    Cyclin-dependent kinases 4 and 6 (CDK4/6) in complex with D-type cyclins promote cell cycle entry. Most human cancers contain overactive CDK4/6-cyclin D, and CDK4/6-specific inhibitors are promising anti-cancer therapeutics. Here, we investigate the critical functions of CDK4/6-cyclin D kinases,

  8. Cell cycle regulation of the cyclin A gene promoter is mediated by a variant E2F site

    DEFF Research Database (Denmark)

    Schulze, A; Zerfass, K; Spitkovsky, D

    1995-01-01

    Cyclin A is involved in the control of S phase and mitosis in mammalian cells. Expression of the cyclin A gene in nontransformed cells is characterized by repression of its promoter during the G1 phase of the cell cycle and its induction at S-phase entry. We show that this mode of regulation...

  9. Cell cycle regulation of the cyclin A gene promoter is mediated by a variant E2F site

    DEFF Research Database (Denmark)

    Schulze, A; Zerfass, K; Spitkovsky, D

    1995-01-01

    Cyclin A is involved in the control of S phase and mitosis in mammalian cells. Expression of the cyclin A gene in nontransformed cells is characterized by repression of its promoter during the G1 phase of the cell cycle and its induction at S-phase entry. We show that this mode of regulation...... is mediated by the transcription factor E2F, which binds to a specific site in the cyclin A promoter. It differs from the prototype E2F site in nucleotide sequence and protein binding; it is bound by E2F complexes containing cyclin E and p107 but not pRB. Ectopic expression of cyclin D1 triggers premature...... activation of the cyclin A promoter by E2F, and this effect is blocked by the tumor suppressor protein p16INK4....

  10. Amplification of cyclin D1 in squamous cell carcinoma of the head and neck and the prognostic value of chromosomal abnormalities and cyclin D1 overexpression

    NARCIS (Netherlands)

    Akervall, J. A.; Michalides, R. J.; Mineta, H.; Balm, A.; Borg, A.; Dictor, M. R.; Jin, Y.; Loftus, B.; Mertens, F.; Wennerberg, J. P.

    1997-01-01

    Abnormalities of chromosome band 11q13 are frequent in squamous cell carcinoma of the head and neck (SCCHN). The oncogene CCND1 is located at 11q13 and encodes cyclin D1, a cell cycle-regulating protein. The authors investigated the clinical relevance and associations between amplification and

  11. VHL-mediated hypoxia regulation of cyclin D1 in renal carcinoma cells.

    Science.gov (United States)

    Bindra, Ranjit S; Vasselli, James R; Stearman, Robert; Linehan, W Marston; Klausner, Richard D

    2002-06-01

    Renal cell carcinoma is associated with mutation of the von Hippel-Lindau (VHL) tumor suppressor gene. Cell lines derived from these tumors cannot exit the cell cycle when deprived of growth factors, and the ability to exit the cell cycle can be restored by the reintroduction of wild-type protein VHL (pVHL). Here, we report that cyclin D1 is overexpressed and remains inappropriately high in during contact inhibition in pVHL-deficient cell lines. In addition, hypoxia increased the expression of cyclin D1 specifically in pVHL-negative cell lines into which pVHL expression was restored. Hypoxic-induction of cyclin D1 was not observed in other pVHL-positive cell lines. This suggests a model whereby in some kidney cell types, pVHL may regulate a proliferative response to hypoxia, whereas the loss of pVHL leads to constitutively elevated cyclin D1 and abnormal proliferation under normal growth conditions.

  12. Three-dimensional collagen represses cyclin E1 via β1 integrin in invasive breast cancer cells.

    Science.gov (United States)

    Wu, Yuehan; Guo, Xun; Brandt, Yekaterina; Hathaway, Helen J; Hartley, Rebecca S

    2011-06-01

    The behavior of breast epithelial cells is influenced by their microenvironment which includes stromal cells and extracellular matrix (ECM). During cancer progression, the tissue microenvironment fails to control proliferation and differentiation, resulting in uncontrolled growth and invasion. Upon invasion, the ECM encountered by breast cancer cells changes from primarily laminin and collagen IV to primarily collagen I. We show here that culturing invasive breast cancer cells in 3-dimensional (3D) collagen I inhibits proliferation through direct regulation of cyclin E1, a G(1)/S regulator that is overexpressed in breast cancer. When the breast cancer cell line MDA-MB-231 was cultured within 3D collagen I gels, the G(1)/S transition was inhibited as compared to cells cultured on conventional 2D collagen or plastic dishes. Cells in 3D collagen downregulated cyclin E1 protein and mRNA, with no change in cyclin D1 level. Cyclin D1 was primarily cytoplasmic in 3D cultures, and this was accompanied by decreased phosphorylation of Rb, a nuclear target for both cyclin E1- and cyclin D1-associated kinases. Positive regulators of cyclin E1 expression, the transcription factor c-Myc and cold-inducible RNA binding protein (CIRP), were decreased in 3D collagen cultures, while the collagen I receptor β1 integrin was greatly increased. Inhibition of β1 integrin function rescued proliferation and cyclin E1 expression as well as c-Myc expression and Rb phosphorylation, but cyclin D1 remained cytoplasmic. We conclude that cyclin E1 is repressed independent of effects on cyclin D1 in a 3D collagen environment and dependent on β1 integrin interaction with collagen I, reducing proliferation of invasive breast cancer cells.

  13. Cell cycle phase specificity of putative cyclin-dependent kinase variants in synchronized alfalfa cells.

    Science.gov (United States)

    Magyar, Z; Mészáros, T; Miskolczi, P; Deák, M; Fehér, A; Brown, S; Kondorosi, E; Athanasiadis, A; Pongor, S; Bilgin, M; Bakó, L; Koncz, C; Dudits, D

    1997-02-01

    The eukaryotic cell division cycle is coordinated by cyclin-dependent kinases (CDKs), represented by a single major serine/threonine kinase in yeasts (Cdc2/CDC28) and a family of kinases (CDK1 to CDK8) in human cells. Previously, two cdc2 homologs, cdc2MsA and cdc2MsB, have been identified in alfalfa (Medicago sativa). By isolating cDNAs using a cdc2MsA probe, we demonstrate here that at least four additional cdc2 homologous genes are expressed in the tetraploid alfalfa. Proteins encoded by the new cdc2MsC to cdc2MsF cDNAs share the characteristic functional domains of CDKs with the conserved and plant-specific sequence elements. Transcripts from cdc2MsA, cdc2MsB, cdc2MsC, and cdc2MsE genes are synthesized throughout the cell cycle, whereas the amounts of cdc2MsD and cdc2MsF mRNAs peak during G2-to-M phases. The translation of Cdc2MsA/B, Cdc2MsD, and Cdc2MsF proteins follows the pattern of transcript accumulation. The multiplicity of kinase complexes with cell cycle phase-dependent activities was revealed by in vitro phosphorylation experiments. Proteins bound to p13suc1-Sepharose or immunoprecipitated with Cdc2MsA/B antibodies from cells at G1-to-S and G2-to-M phase boundaries showed elevated kinase activities. the Cdc2MsF antibodies separated a G2-to-M phase-related kinase complex. Detection of histone H1 phosphorylation activities in fractions immunoprecipitated with antimitotic cyclin (CyclinMs2) antibodies from G2-to-M phase cells indicates the complex formation between this cyclin and a kinase partner in alfalfa. The observed fluctuation of transcript levels, amounts, and activities of kinases in different cell cycle phases reflects a multilevel regulatory system during cell cycle progression in plants.

  14. PARP-2 regulates cell cycle-related genes through histone deacetylation and methylation independently of poly(ADP-ribosyl)ation

    International Nuclear Information System (INIS)

    Liang, Ya-Chen; Hsu, Chiao-Yu; Yao, Ya-Li; Yang, Wen-Ming

    2013-01-01

    Highlights: ► PARP-2 acts as a transcription co-repressor independently of PARylation activity. ► PARP-2 recruits HDAC5, 7, and G9a and generates repressive chromatin. ► PARP-2 is recruited to the c-MYC promoter by DNA-binding factor YY1. ► PARP-2 represses cell cycle-related genes and alters cell cycle progression. -- Abstract: Poly(ADP-ribose) polymerase-2 (PARP-2) catalyzes poly(ADP-ribosyl)ation (PARylation) and regulates numerous nuclear processes, including transcription. Depletion of PARP-2 alters the activity of transcription factors and global gene expression. However, the molecular action of how PARP-2 controls the transcription of target promoters remains unclear. Here we report that PARP-2 possesses transcriptional repression activity independently of its enzymatic activity. PARP-2 interacts and recruits histone deacetylases HDAC5 and HDAC7, and histone methyltransferase G9a to the promoters of cell cycle-related genes, generating repressive chromatin signatures. Our findings propose a novel mechanism of PARP-2 in transcriptional regulation involving specific protein–protein interactions and highlight the importance of PARP-2 in the regulation of cell cycle progression

  15. PARP-2 regulates cell cycle-related genes through histone deacetylation and methylation independently of poly(ADP-ribosyl)ation

    Energy Technology Data Exchange (ETDEWEB)

    Liang, Ya-Chen; Hsu, Chiao-Yu [Institute of Molecular Biology, National Chung Hsing University, Taichung 40227, Taiwan (China); Yao, Ya-Li [Department of Biotechnology, Asia University, Taichung 41354, Taiwan (China); Yang, Wen-Ming, E-mail: yangwm@nchu.edu.tw [Institute of Molecular Biology, National Chung Hsing University, Taichung 40227, Taiwan (China)

    2013-02-01

    Highlights: ► PARP-2 acts as a transcription co-repressor independently of PARylation activity. ► PARP-2 recruits HDAC5, 7, and G9a and generates repressive chromatin. ► PARP-2 is recruited to the c-MYC promoter by DNA-binding factor YY1. ► PARP-2 represses cell cycle-related genes and alters cell cycle progression. -- Abstract: Poly(ADP-ribose) polymerase-2 (PARP-2) catalyzes poly(ADP-ribosyl)ation (PARylation) and regulates numerous nuclear processes, including transcription. Depletion of PARP-2 alters the activity of transcription factors and global gene expression. However, the molecular action of how PARP-2 controls the transcription of target promoters remains unclear. Here we report that PARP-2 possesses transcriptional repression activity independently of its enzymatic activity. PARP-2 interacts and recruits histone deacetylases HDAC5 and HDAC7, and histone methyltransferase G9a to the promoters of cell cycle-related genes, generating repressive chromatin signatures. Our findings propose a novel mechanism of PARP-2 in transcriptional regulation involving specific protein–protein interactions and highlight the importance of PARP-2 in the regulation of cell cycle progression.

  16. The regulation of SIRT2 function by cyclin-dependent kinases affects cell motility.

    NARCIS (Netherlands)

    Pandithage, R.; Lilischkis, R.; Harting, K.; Wolf, A.; Jedamzik, B.; Luscher-Firzlaff, J.; Vervoorts, J.; Lasonder, E.; Kremmer, E.; Knoll, B.; Luscher, B.

    2008-01-01

    Cyclin-dependent kinases (Cdks) fulfill key functions in many cellular processes, including cell cycle progression and cytoskeletal dynamics. A limited number of Cdk substrates have been identified with few demonstrated to be regulated by Cdk-dependent phosphorylation. We identify on protein

  17. p21/Cyclin E pathway modulates anticlastogenic function of Bmi-1 in cancer cells

    Science.gov (United States)

    Deng, Wen; Zhou, Yuan; Tiwari, Agnes FY; Su, Hang; Yang, Jie; Zhu, Dandan; Lau, Victoria Ming Yi; Hau, Pok Man; Yip, Yim Ling; Cheung, Annie LM; Guan, Xin-Yuan; Tsao, Sai Wah

    2015-01-01

    Apart from regulating stem cell self-renewal, embryonic development and proliferation, Bmi-1 has been recently reported to be critical in the maintenance of genome integrity. In searching for novel mechanisms underlying the anticlastogenic function of Bmi-1, we observed, for the first time, that Bmi-1 positively regulates p21 expression. We extended the finding that Bmi-1 deficiency induced chromosome breaks in multiple cancer cell models. Interestingly, we further demonstrated that knockdown of cyclin E or ectopic overexpression of p21 rescued Bmi-1 deficiency-induced chromosome breaks. We therefore conclude that p21/cyclin E pathway is crucial in modulating the anticlastogenic function of Bmi-1. As it is well established that the overexpression of cyclin E potently induces genome instability and p21 suppresses the function of cyclin E, the novel and important implication from our findings is that Bmi-1 plays an important role in limiting genomic instability in cylin E-overexpressing cancer cells by positive regulation of p21. PMID:25131797

  18. Colorectal cancer cell-derived microvesicles are enriched in cell cycle-related mRNAs that promote proliferation of endothelial cells

    Directory of Open Access Journals (Sweden)

    Kim Yoon-Keun

    2009-11-01

    Full Text Available Abstract Background Various cancer cells, including those of colorectal cancer (CRC, release microvesicles (exosomes into surrounding tissues and peripheral circulation. These microvesicles can mediate communication between cells and affect various tumor-related processes in their target cells. Results We present potential roles of CRC cell-derived microvesicles in tumor progression via a global comparative microvesicular and cellular transcriptomic analysis of human SW480 CRC cells. We first identified 11,327 microvesicular mRNAs involved in tumorigenesis-related processes that reflect the physiology of donor CRC cells. We then found 241 mRNAs enriched in the microvesicles above donor cell levels, of which 27 were involved in cell cycle-related processes. Network analysis revealed that most of the cell cycle-related microvesicle-enriched mRNAs were associated with M-phase activities. The integration of two mRNA datasets showed that these M-phase-related mRNAs were differentially regulated across CRC patients, suggesting their potential roles in tumor progression. Finally, we experimentally verified the network-driven hypothesis by showing a significant increase in proliferation of endothelial cells treated with the microvesicles. Conclusion Our study demonstrates that CRC cell-derived microvesicles are enriched in cell cycle-related mRNAs that promote proliferation of endothelial cells, suggesting that microvesicles of cancer cells can be involved in tumor growth and metastasis by facilitating angiogenesis-related processes. This information will help elucidate the pathophysiological functions of tumor-derived microvesicles, and aid in the development of cancer diagnostics, including colorectal cancer.

  19. Copper Uptake in Mammary Epithelial Cells Activates Cyclins and Triggers Antioxidant Response

    Directory of Open Access Journals (Sweden)

    Nathália Villa dos Santos

    2015-01-01

    Full Text Available The toxicologic effects of copper (Cu on tumor cells have been studied during the past decades, and it is suggested that Cu ion may trigger antiproliferative effects in vitro. However, in normal cells the toxicologic effects of high exposures of free Cu are not well understood. In this work, Cu uptake, the expression of genes associated with cell cycle regulation, and the levels of ROS production and related oxidative processes were evaluated in Cu-treated mammary epithelial MCF10A nontumoral cells. We have shown that the Cu additive is associated with the activation of cyclin D1 and cyclin B1, as well as cyclin-dependent kinase 2 (CDK2. These nontumor cells respond to Cu-induced changes in the oxidative balance by increase of the levels of reduced intracellular glutathione (GSH, decrease of reactive oxygen species (ROS generation, and accumulation during progression of the cell cycle, thus preventing the cell abnormal proliferation or death. Taken together, our findings revealed an effect that contributes to prevent a possible damage of normal cells exposed to chemotherapeutic effects of drugs containing the Cu ion.

  20. Immunohistochemical Expression of Cyclin B1 in Epithelial Hyperplasia, Dysplasia and Oral Squamous Cell Carcinomas - A Comparative Study.

    Science.gov (United States)

    Niranjan, Kochli Channappa; Tayaar, Amsavardani; Kumar, G S; Krishnapillai, Rekha; Hallikeri, Kaveri; Hunasgi, Santosh

    2016-09-01

    Cyclin B1 is important in the cell cycle progression from G2 to M phase. Cyclin B1 binds to CDC2, which then becomes dephosphorylated and gets relocated to the nucleus, ensuring the transition toward mitosis. Over expression of Cyclin B1, has been reported more recently in breast, colon, prostate, oral and esophageal carcinomas. Thus, the aim of the present study was to examine the expression of Cyclin B1 in hyperplasia, dysplasia and Oral Squamous Cell Carcinomas (OSCC). A total of 64 histopathologically diagnosed cases of epithelial hyperplasias, dysplastic oral epithelium and OSCC were included in the study. Immunohistochemical procedure was carried out using the monoclonal mouse Cyclin B1 antibody (Clone V-152). The Cyclin B1 positive tumor cells counted were expressed as percentage of positive tumor cells. Nuclear and cytoplasmic labeling index (n&cLI) were calculated. The results were tabulated and statistically analyzed by Kruskal Wallis test- One Way ANOVA and Mann Whitney U- test. Combined n&cLI was considered only in 28.57% of epithelial hyperplasias, 40.7% of oral epithelial dysplasias and 72% of OSCC showed over expression of Cyclin B1 with p value being 0.029. Cyclin B1 expression was not significantly different between the grades of dysplasia, between the grades of OSCC and between the marginal groups. The present study demonstrates more than 50% of the study group showing less than 20% of nuclear staining. The importance of such variations within a type of lesion requires further investigation, since Cyclin B1 has proved useful in many studies from esophageal and laryngeal squamous cell carcinoma as a prognostic indicator, an indicator of recurrence and as an indicator for tumor sensitivity to radiotherapy. Further studies are to be extended towards evaluating the role of Cyclin B1 as a prognostic indicator.

  1. Peripheral T-cell Lymphoma with Cyclin D1 overexpression: a case report

    Directory of Open Access Journals (Sweden)

    Aquino Gabriella

    2012-07-01

    Full Text Available Abstract Peripheral T-cell lymphomas not otherwise specified are generally considered aggressive non-Hodgkin lymphomas, because of poor natural outcome and response to therapy. They show a complex karyotype without any specific genetic hallmark. We report a case of peripheral T-cell lymphoma not otherwise specified with heterogeneous nuclear Cyclin D1 immunohistochemical overexpression, due to gene copy gain, a phenomenon similar to that observed in Mantle Cell Lymphoma characterized by t(11;14(q13;q32. In this case report we underline the diagnostic pitfall rapresented by Cyclin D1 immunoistochemical overexpression in a T-cell lymphoma. Several pitfalls could lead to misinterpretation of diagnosis, therefore, we underlined the need to integrate the classical histology and immunohistochemistry with molecular tests as clonality or Fluorescence in situ hybridization. Virtual slide The virtual slides for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/1117747619703769

  2. Peripheral T-cell Lymphoma with Cyclin D1 overexpression: a case report.

    Science.gov (United States)

    Aquino, Gabriella; Franco, Renato; Ronconi, Fioravante; Anniciello, Annamaria; Russo, Luigi; De Chiara, Annarosaria; Panico, Luigi

    2012-07-06

    Peripheral T-cell lymphomas not otherwise specified are generally considered aggressive non-Hodgkin lymphomas, because of poor natural outcome and response to therapy. They show a complex karyotype without any specific genetic hallmark. We report a case of peripheral T-cell lymphoma not otherwise specified with heterogeneous nuclear cyclin D1 immunohistochemical overexpression, due to gene copy gain, a phenomenon similar to that observed in mantle cell lymphoma characterized by t(11;14)(q13;q32). In this case report we underline the diagnostic pitfall represented by cyclin D1 immunohistochemical overexpression in a T-cell lymphoma. Several pitfalls could lead to misinterpretation of diagnosis, therefore, we underlined the need to integrate the classical histology and immunohistochemistry with molecular tests as clonality or fluorescence in situ hybridization. The virtual slides for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/1117747619703769.

  3. Protocatechualdehyde possesses anti-cancer activity through downregulating cyclin D1 and HDAC2 in human colorectal cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Jeong, Jin Boo [Department of Nutrition and Food Science, University of Maryland, College Park, MD 20742 (United States); Lee, Seong-Ho, E-mail: slee2000@umd.edu [Department of Nutrition and Food Science, University of Maryland, College Park, MD 20742 (United States)

    2013-01-04

    Highlights: Black-Right-Pointing-Pointer Protocatechualdehyde (PCA) suppressed cell proliferation and induced apoptosis in human colorectal cancer cells. Black-Right-Pointing-Pointer PCA enhanced transcriptional downregulation of cyclin D1 gene. Black-Right-Pointing-Pointer PCA suppressed HDAC2 expression and activity. Black-Right-Pointing-Pointer These findings suggest that anti-cancer activity of PCA may be mediated by reducing HDAC2-derived cyclin D1 expression. -- Abstract: Protocatechualdehyde (PCA) is a naturally occurring polyphenol found in barley, green cavendish bananas, and grapevine leaves. Although a few studies reported growth-inhibitory activity of PCA in breast and leukemia cancer cells, the underlying mechanisms are still poorly understood. Thus, we performed in vitro study to investigate if treatment of PCA affects cell proliferation and apoptosis in human colorectal cancer cells and define potential mechanisms by which PCA mediates growth arrest and apoptosis of cancer cells. Exposure of PCA to human colorectal cancer cells (HCT116 and SW480 cells) suppressed cell growth and induced apoptosis in dose-dependent manner. PCA decreased cyclin D1 expression in protein and mRNA level and suppressed luciferase activity of cyclin D1 promoter, indicating transcriptional downregulation of cyclin D1 gene by PCA. We also observed that PCA treatment attenuated enzyme activity of histone deacetylase (HDAC) and reduced expression of HDAC2, but not HDAC1. These findings suggest that cell growth inhibition and apoptosis by PCA may be a result of HDAC2-mediated cyclin D1 suppression.

  4. Protocatechualdehyde possesses anti-cancer activity through downregulating cyclin D1 and HDAC2 in human colorectal cancer cells

    International Nuclear Information System (INIS)

    Jeong, Jin Boo; Lee, Seong-Ho

    2013-01-01

    Highlights: ► Protocatechualdehyde (PCA) suppressed cell proliferation and induced apoptosis in human colorectal cancer cells. ► PCA enhanced transcriptional downregulation of cyclin D1 gene. ► PCA suppressed HDAC2 expression and activity. ► These findings suggest that anti-cancer activity of PCA may be mediated by reducing HDAC2-derived cyclin D1 expression. -- Abstract: Protocatechualdehyde (PCA) is a naturally occurring polyphenol found in barley, green cavendish bananas, and grapevine leaves. Although a few studies reported growth-inhibitory activity of PCA in breast and leukemia cancer cells, the underlying mechanisms are still poorly understood. Thus, we performed in vitro study to investigate if treatment of PCA affects cell proliferation and apoptosis in human colorectal cancer cells and define potential mechanisms by which PCA mediates growth arrest and apoptosis of cancer cells. Exposure of PCA to human colorectal cancer cells (HCT116 and SW480 cells) suppressed cell growth and induced apoptosis in dose-dependent manner. PCA decreased cyclin D1 expression in protein and mRNA level and suppressed luciferase activity of cyclin D1 promoter, indicating transcriptional downregulation of cyclin D1 gene by PCA. We also observed that PCA treatment attenuated enzyme activity of histone deacetylase (HDAC) and reduced expression of HDAC2, but not HDAC1. These findings suggest that cell growth inhibition and apoptosis by PCA may be a result of HDAC2-mediated cyclin D1 suppression.

  5. Slug is temporally regulated by cyclin E in cell cycle and controls genome stability.

    Science.gov (United States)

    Wang, W-L; Huang, H-C; Kao, S-H; Hsu, Y-C; Wang, Y-T; Li, K-C; Chen, Y-J; Yu, S-L; Wang, S-P; Hsiao, T-H; Yang, P-C; Hong, T-M

    2015-02-26

    The transcriptional repressor Slug is best known to control epithelial-mesenchymal transition (EMT) and promote cancer invasion/metastasis. In this study, we demonstrate that Slug is temporally regulated during cell cycle progression. At G1/S transition, cyclin E-cyclin-dependent kinase 2 mediates the phosphorylation of Slug at Ser-54 and Ser-104, resulting in its ubiquitylation and degradation. Non-phosphorylatable Slug is markedly stabilized at G1/S transition compared with wild-type Slug and greatly leads to downregulation of DNA synthesis and checkpoint-related proteins, including TOP1, DNA Ligase IV and Rad17, reduces cell proliferation, delays S-phase progression and contributes to genome instability. Our results indicate that Slug has multifaceted roles in cancer progression by controlling both EMT and genome stability.

  6. Resibufogenin Induces G1-Phase Arrest through the Proteasomal Degradation of Cyclin D1 in Human Malignant Tumor Cells.

    Directory of Open Access Journals (Sweden)

    Masami Ichikawa

    Full Text Available Huachansu, a traditional Chinese medicine prepared from the dried toad skin, has been used in clinical studies for various cancers in China. Resibufogenin is a component of huachansu and classified as bufadienolides. Resibufogenin has been shown to exhibit the anti-proliferative effect against cancer cells. However, the molecular mechanism of resibufogenin remains unknown. Here we report that resibufogenin induces G1-phase arrest with hypophosphorylation of retinoblastoma (RB protein and down-regulation of cyclin D1 expression in human colon cancer HT-29 cells. Since the down-regulation of cyclin D1 was completely blocked by a proteasome inhibitor MG132, the suppression of cyclin D1 expression by resibufogenin was considered to be in a proteasome-dependent manner. It is known that glycogen synthase kinase-3β (GSK-3β induces the proteasomal degradation of cyclin D1. The addition of GSK-3β inhibitor SB216763 inhibited the reduction of cyclin D1 caused by resibufogenin. These effects on cyclin D1 by resibufogenin were also observed in human lung cancer A549 cells. These findings suggest that the anti-proliferative effect of resibufogenin may be attributed to the degradation of cyclin D1 caused by the activation of GSK-3β.

  7. MicroRNA-16 Modulates HuR Regulation of Cyclin E1 in Breast Cancer Cells

    Directory of Open Access Journals (Sweden)

    Xun Guo

    2015-03-01

    Full Text Available RNA binding protein (RBPs and microRNAs (miRNAs or miRs are post-transcriptional regulators of gene expression that are implicated in development of cancers. Although their individual roles have been studied, the crosstalk between RBPs and miRNAs is under intense investigation. Here, we show that in breast cancer cells, cyclin E1 upregulation by the RBP HuR is through specific binding to regions in the cyclin E1 mRNA 3' untranslated region (3'UTR containing U-rich elements. Similarly, miR-16 represses cyclin E1, dependent on its cognate binding sites in the cyclin E1 3'UTR. Evidence in the literature indicates that HuR can regulate miRNA expression and recruit or dissociate RNA-induced silencing complexes (RISC. Despite this, miR-16 and HuR do not affect the other’s expression level or binding to the cyclin E1 3'UTR. While HuR overexpression partially blocks miR-16 repression of a reporter mRNA containing the cyclin E1 3'UTR, it does not block miR-16 repression of endogenous cyclin E1 mRNA. In contrast, miR-16 blocks HuR-mediated upregulation of cyclin E1. Overall our results suggest that miR-16 can override HuR upregulation of cyclin E1 without affecting HuR expression or association with the cyclin E1 mRNA.

  8. Sp1 phosphorylation by cyclin-dependent kinase 1/cyclin B1 represses its DNA-binding activity during mitosis in cancer cells.

    Science.gov (United States)

    Chuang, J-Y; Wang, S-A; Yang, W-B; Yang, H-C; Hung, C-Y; Su, T-P; Chang, W-C; Hung, J-J

    2012-11-22

    Sp1 is important for the transcription of many genes. Our previous studies have shown that Sp1 is degraded in normal cell, but it is preserved in cancer cells during mitosis and exists a priori in the daughter cells, ready to engage in gene transcription and thereby contributes to the proliferation and survival of cancer cells. The mechanism by which Sp1 is preserved in cancer cells during mitosis remains unknown. In this study, we observed that Sp1 strongly colocalized with cyclin-dependent kinase 1 (CDK1)/cyclin B1 during mitosis. Moreover, we showed that Sp1 is a novel mitotic substrate of CDK1/cyclin B1 and is phosphorylated by it at Thr 739 before the onset of mitosis. Phospho-Sp1 reduced its DNA-binding ability and facilitated the chromatin condensation process during mitosis. Mutation of Thr739 to alanine resulted in Sp1 remaining in the chromosomes, delayed cell-cycle progression, and eventually led to apoptosis. Screening of Sp1-associated proteins during mitosis by using liquid chromatography/mass spectrometry indicated the tethering of Sp1 to myosin/F-actin. Furthermore, phospho-Sp1 and myosin/F-actin appeared to exist as a congregated ring at the periphery of the chromosome. However, at the end of mitosis and the beginning of interphase, Sp1 was dephosphorylated by PP2A and returned to the chromatin. These results indicate that cancer cells use CDK1 and PP2A to regulate the movement of Sp1 in and out of the chromosomes during cell-cycle progression, which may benefit cancer-cell proliferation.

  9. Sp1 phosphorylation by cyclin-dependent kinase 1/cyclin B1 represses its DNA-binding activity during mitosis in cancer cells

    Science.gov (United States)

    Chuang, J-Y; Wang, S-A; Yang, W-B; Yang, H-C; Hung, C-Y; Su, T-P; Chang, W-C; Hung, J-J

    2013-01-01

    Sp1 is important for the transcription of many genes. Our previous studies have shown that Sp1 is degraded in normal cell, but it is preserved in cancer cells during mitosis and exists a priori in the daughter cells, ready to engage in gene transcription and thereby contributes to the proliferation and survival of cancer cells. The mechanism by which Sp1 is preserved in cancer cells during mitosis remains unknown. In this study, we observed that Sp1 strongly colocalized with cyclin-dependent kinase 1 (CDK1)/cyclin B1 during mitosis. Moreover, we showed that Sp1 is a novel mitotic substrate of CDK1/cyclin B1 and is phosphorylated by it at Thr 739 before the onset of mitosis. Phospho-Sp1 reduced its DNA-binding ability and facilitated the chromatin condensation process during mitosis. Mutation of Thr739 to alanine resulted in Sp1 remaining in the chromosomes, delayed cell-cycle progression, and eventually led to apoptosis. Screening of Sp1-associated proteins during mitosis by using liquid chromatography/mass spectrometry indicated the tethering of Sp1 to myosin/F-actin. Furthermore, phospho-Sp1 and myosin/F-actin appeared to exist as a congregated ring at the periphery of the chromosome. However, at the end of mitosis and the beginning of interphase, Sp1 was dephosphorylated by PP2A and returned to the chromatin. These results indicate that cancer cells use CDK1 and PP2A to regulate the movement of Sp1 in and out of the chromosomes during cell-cycle progression, which may benefit cancer-cell proliferation. PMID:22266860

  10. Multiple cyclin-dependent kinase complexes and phosphatases control G2/M progression in alfalfa cells.

    Science.gov (United States)

    Mészáros, T; Miskolczi, P; Ayaydin, F; Pettkó-Szandtner, A; Peres, A; Magyar, Z; Horváth, G V; Bakó, L; Fehér, A; Dudits, D

    2000-08-01

    Reversible phosphorylation of proteins by kinases and phosphatases plays a key regulatory role in several eukaryotic cellular functions including the control of the division cycle. Increasing numbers of sequence and biochemical data show the involvement of cyclin-dependent kinases (CDKs) and cyclins in regulation of the cell cycle progression in higher plants. The complexity represented by different types of CDKs and cyclins in a single species such as alfalfa, indicates that multicomponent regulatory pathways control G2/M transition. A set of cdc2-related genes (cdc2Ms A, B, D and F) was expressed in G2 and M cells. Phosphorylation assays also revealed that at least three kinase complexes (Cdc2Ms A/B, D and F) were successively active in G2/M cells after synchronization. Interaction between alfalfa mitotic cyclin (Medsa;CycB2;1) and a kinase partner has been reported previously. The present yeast two-hybrid analyses showed differential interaction between defined D-type cyclins and Cdc2Ms kinases functioning in G2/M phases. Localization of Cdc2Ms F kinase to the preprophase band (PPB), the perinuclear ring in early prophase, the mitotic spindle and the phragmoplast indicated a pivotal role for this kinase in mitotic plant cells. So far limited research efforts have been devoted to the functions of phosphatases in the control of plant cell division. A homologue of dual phosphatase, cdc25, has not been cloned yet from alfalfa; however tyrosine phosphorylation was indicated in the case of Cdc2Ms A kinase and the p(13suc1)-bound kinase activity was increased by treatment of this complex with recombinant Drosophila Cdc25. The potential role of serine/threonine phosphatases can be concluded from inhibitor studies based on okadaic acid or endothall. Endothall elevated the kinase activity of p(13suc1)-bound fractions in G2-phase alfalfa cells. These biochemical data are in accordance with observed cytological abnormalities. The present overview with selected original data

  11. CD8 T-cell responses against cyclin B1 in breast cancer patients with tumors overexpressing p53

    DEFF Research Database (Denmark)

    Sørensen, Rikke Baek; Andersen, Rikke Sick; Svane, Inge Marie

    2009-01-01

    CD8 T-cell response against at least one of the peptides; strongest reactivity was detected against the CB9L2 peptide. Because the level of cyclin B1 has been shown to be influenced by the level of p53, which in turn is elevated in cancer cells because of point mutation, we analyzed the level of p53...... protein in biopsies from the patients by immune histochemistry. Combined data showed that anti-cyclin B1 reactivity was predominantly detected in patients with tumors characterized by elevated expression of p53. Interestingly, no reactivity was detected against six peptides derived from the p53 protein....... CONCLUSIONS: Our data support the notion of cyclin B1 as a prominent target for immunologic recognition in cancer patients harboring p53-mutated cancer cells. Because mutation of p53 is one of the most frequent genetic alterations in human cancers, this suggests that immunotherapy based on targeting of cyclin...

  12. Phosphorylation of XIAP by CDK1–cyclin-B1 controls mitotic cell death

    Science.gov (United States)

    Hou, Ying; Allan, Lindsey A.

    2017-01-01

    ABSTRACT Regulation of cell death is crucial for the response of cancer cells to drug treatments that cause arrest in mitosis, and is likely to be important for protection against chromosome instability in normal cells. Prolonged mitotic arrest can result in cell death by activation of caspases and the induction of apoptosis. Here, we show that X-linked inhibitor of apoptosis (XIAP) plays a key role in the control of mitotic cell death. Ablation of XIAP expression sensitises cells to prolonged mitotic arrest caused by a microtubule poison. XIAP is stable during mitotic arrest, but its function is controlled through phosphorylation by the mitotic kinase CDK1–cyclin-B1 at S40. Mutation of S40 to a phosphomimetic residue (S40D) inhibits binding to activated effector caspases and abolishes the anti-apoptotic function of XIAP, whereas a non-phosphorylatable mutant (S40A) blocks apoptosis. By performing live-cell imaging, we show that phosphorylation of XIAP reduces the threshold for the onset of cell death in mitosis. This work illustrates that mitotic cell death is a form of apoptosis linked to the progression of mitosis through control by CDK1–cyclin-B1. PMID:27927753

  13. [Quercetin regulates cell cycle-related gene expression in a model of glucose-oxygen deprivation in astrocytes].

    Science.gov (United States)

    Yao, Fang; Zhang, Lanlan; Yuan, Zhaohu; Zeng, Yong; Wu, Bingyi

    2013-09-01

    To study the effect of quercetin on gene expression in astrocytes after glucose-oxygen deprivation and the underlying mechanism. The primary cultured astrocytes were randomly divided into glucose-oxygen deprivation group (only treated with glucose-oxygen deprivation for 4 hours) and glucose-oxygen deprivation combined with quercetin-treated group (glucose-oxygen deprivation for 4 hours combined with quercetin treatment for 24 hours). Their mRNA expressions were analyzed by the large-scale oligo microarray. The differential genes obtained were further confirmed by real-time quantitative PCR (qRT-PCR). Compared with the glucose-oxygen deprivation group, the glucose-oxygen deprivation combined with quercetin-treated group presented the changes in the expressions of 31 genes that were related to cell cycle, of which 5 genes were up-regulated and 26 were down-regulated. Six of those differential genes were confirmed by qRT-PCR and the result of their differential expressions was consistent with that by large-scale oligo microarray. Quercetin can regulate some of cell cycle-related genes in astrocytes after glucose-oxygen deprivation.

  14. γ-Carboxymuconolactone decarboxylase: a novel cell cycle-related basal body protein in the early branching eukaryote Trichomonas vaginalis.

    Science.gov (United States)

    Cheng, Wei-Hung; Huang, Kuo-Yang; Huang, Po-Jung; Lee, Chi-Ching; Yeh, Yuan-Ming; Ku, Fu-Man; Lin, Rose; Cheng, Mei-Ling; Chiu, Cheng-Hsun; Tang, Petrus

    2017-09-26

    γ-Carboxymuconolactone decarboxylase (CMD) participates in the β-ketoadipate pathway, which catalyzes aromatic compounds to produce acetyl- or succinyl-CoA, in prokaryotes and yeast. Our previous study demonstrated that expression of a CMD homologue that contains two signatures (dualCMD) is negatively regulated by iron in Trichomonas vaginalis. However, we were not able to identify the components of the β-ketoadipate pathway in the parasite's genome. These observations prompted us to investigate the biological functions of this novel CMD homologue in T. vaginalis. The specific anti-TvCMD1 antibody was generated, and the expression of TvCMD1 in T. vaginalis cultured under iron-rich and iron-deficient were evaluated. Phylogenetic, metabolomic and substrate induction (protocatechuate and benzoate) analysis were conducted to clarify the function of dualCMD in trichomonad cells. Subcellular localization of TvCMD1 was observed by confocal microscopy. The cell cycle-related role of TvCMD1 was assessed by treating cells with G2/M inhibitor nocodazole. We confirmed that T. vaginalis is not able to catabolize the aromatic compounds benzoate and protocatechuate, which are known substrates of the β-ketoadipate pathway. Using immunofluorescence microscopy, we found that TvCMD1 is spatially associated with the basal body, a part of the cytoskeletal organizing center in T. vaginalis. TvCMD1 accumulated upon treatment with the G2/M inhibitor nocodazole. Additionally, TvCMD1 was expressed and transported to/from the basal body during cytokinesis, suggesting that TvCMD1 plays a role in cell division. We demonstrated that TvCMD1 is unlikely to participate in the β-ketoadipate pathway and demonstrated that it is a novel basal body-localizing (associated) protein. This model sheds light on the importance of genes that are acquired laterally in the coevolution of ancient protists, which surprisingly functions in cell cycle regulation of T. vaginalis.

  15. cycMs3, a novel B-type alfalfa cyclin gene, is induced in the G0-to-G1 transition of the cell cycle

    OpenAIRE

    Meskiene, I.; Bogre, L.; Dahl, M.; Pirck, M.; Ha, D. T.; Swoboda, I.; Heberle-Bors, E.; Ammerer, G.; Hirt, H.

    1995-01-01

    Cyclins are key regulators of the cell cycle in all eukaryotes. We have previously isolated two B-type cyclin genes, cycMs1 and cycMs2, from alfalfa that are primarily expressed during the G2-to-M phase transition and are most likely mitotic cyclin genes. Here, we report the isolation of a novel alfalfa cyclin gene, termed cycMs3 (for cyclin Medicago sativa), by selecting for mating type alpha-pheromone-induced cell cycle arrest suppression in yeast. The central region of the predicted amino ...

  16. The transcription factor NFAT5 is required for cyclin expression and cell cycle progression in cells exposed to hypertonic stress.

    Directory of Open Access Journals (Sweden)

    Katherine Drews-Elger

    Full Text Available BACKGROUND: Hypertonicity can perturb cellular functions, induce DNA damage-like responses and inhibit proliferation. The transcription factor NFAT5 induces osmoprotective gene products that allow cells to adapt to sustained hypertonic conditions. Although it is known that NFAT5-deficient lymphocytes and renal medullary cells have reduced proliferative capacity and viability under hypertonic stress, less is understood about the contribution of this factor to DNA damage responses and cell cycle regulation. METHODOLOGY/PRINCIPAL FINDINGS: We have generated conditional knockout mice to obtain NFAT5(-/- T lymphocytes, which we used as a model of proliferating cells to study NFAT5-dependent responses. We show that hypertonicity triggered an early, NFAT5-independent, genotoxic stress-like response with induction of p53, p21 and GADD45, downregulation of cyclins, and cell cycle arrest. This was followed by an NFAT5-dependent adaptive phase in wild-type cells, which induced an osmoprotective gene expression program, downregulated stress markers, resumed cyclin expression and proliferation, and displayed enhanced NFAT5 transcriptional activity in S and G2/M. In contrast, NFAT5(-/- cells failed to induce osmoprotective genes and exhibited poorer viability. Although surviving NFAT5(-/- cells downregulated genotoxic stress markers, they underwent cell cycle arrest in G1/S and G2/M, which was associated with reduced expression of cyclins E1, A2 and B1. We also show that pathologic hypertonicity levels, as occurring in plasma of patients and animal models of osmoregulatory disorders, inhibited the induction of cyclins and aurora B kinase in response to T cell receptor stimulation in fresh NFAT5(-/- lymphocytes. CONCLUSIONS/SIGNIFICANCE: We conclude that NFAT5 facilitates cell proliferation under hypertonic conditions by inducing an osmoadaptive response that enables cells to express fundamental regulators needed for cell cycle progression.

  17. Nuclear accumulation of cyclin D1 following long-term fractionated exposures to low-dose ionizing radiation in normal human diploid cells.

    Science.gov (United States)

    Shimura, Tsutomu; Hamada, Nobuyuki; Sasatani, Megumi; Kamiya, Kenji; Kunugita, Naoki

    2014-01-01

    Cyclin D1 is a mitogenic sensor that responds to growth signals from the extracellular environment and regulates the G 1-to-S cell cycle transition. When cells are acutely irradiated with a single dose of 10 Gy, cyclin D1 is degraded, causing cell cycle arrest at the G 1/S checkpoint. In contrast, cyclin D1 accumulates in human tumor cells that are exposed to long-term fractionated radiation (0.5 Gy/fraction of X-rays). In this study we investigated the effect of fractionated low-dose radiation exposure on cyclin D1 localization in 3 strains of normal human fibroblasts. To specifically examine the nuclear accumulation of cyclin D1, cells were treated with a hypotonic buffer containing detergent to remove cytoplasmic cyclin D1. Proliferating cell nuclear antigen (PCNA) immunofluorescence was used to identify cells in S phase. With this approach, we observed S-phase nuclear retention of cyclin D1 following low-dose fractionated exposures, and found that cyclin D1 nuclear retention increased with exposure time. Cells that retained nuclear cyclin D1 were more likely to have micronuclei than non-retaining cells, indicating that the accumulation of nuclear cyclin D1 was associated with genomic instability. Moreover, inhibition of the v-akt murine thymoma viral oncogene homolog (AKT) pathway facilitated cyclin D1 degradation and eliminated cyclin D1 nuclear retention in cells exposed to fractionated radiation. Thus, cyclin D1 may represent a useful marker for monitoring long-term effects associated with exposure to low levels of radiation.

  18. The D-type alfalfa cyclin gene cycMs4 complements G1 cyclin-deficient yeast and is induced in the G1 phase of the cell cycle.

    Science.gov (United States)

    Dahl, M; Meskiene, I; Bögre, L; Ha, D T; Swoboda, I; Hubmann, R; Hirt, H; Heberle-Bors, E

    1995-11-01

    Cyclins are key regulators of the cell cycle in all eukaryotes. In alfalfa, we have previously isolated three B-type cyclins. The closely related cycMs1 and cycMs2 genes are expressed primarily during the G2 and M phases and are most likely mitotic cyclins; expression of the cycMs3 gene is induced in the G0-to-G1 transition, when cells reenter the cell cycle. By complementation of G1 cyclin-deficient yeast cells, a novel alfalfa cyclin, designated cycMs4, was isolated. The predicted amino acid sequence of the cycMs4 gene is most similar to that of the Arabidopsis cyclin delta 3 gene. CycMs4 and cyclin delta 3 belong to the class of D-type cyclins and contain PEST-rich regions and a retinoblastoma binding motif. When comparing expression levels in different organs, cycMs4 transcripts were present predominantly in roots. Whereas expression of the cycMs4 gene was cell cycle-regulated in suspension-cultured cells, transcription in roots was observed to depend also on the positional context of the cell. When differentiated G0-arrested leaf cells were induced to resume cell division by treatment with plant hormones, cycMs4 transcription was induced before the onset of DNA synthesis. Whereas this induction was preceded by that of the cycMs3 gene, cycMs2 expression occurred later and at the same time as mitotic activity. These data suggest that cycMs4 plays a role in the G1-to-S transition and provide a model to investigate the plant cell cycle at the molecular level.

  19. Canonical and Alternative Pathways in Cyclin-Dependent Kinase 1/Cyclin B Inactivation upon M-Phase Exit in Xenopus laevis Cell-Free Extracts

    Directory of Open Access Journals (Sweden)

    Jacek Z. Kubiak

    2011-01-01

    Full Text Available Cyclin-Dependent Kinase 1 (CDK1 is the major M-phase kinase known also as the M-phase Promoting Factor or MPF. Studies performed during the last decade have shown many details of how CDK1 is regulated and also how it regulates the cell cycle progression. Xenopus laevis cell-free extracts were widely used to elucidate the details and to obtain a global view of the role of CDK1 in M-phase control. CDK1 inactivation upon M-phase exit is a primordial process leading to the M-phase/interphase transition during the cell cycle. Here we discuss two closely related aspects of CDK1 regulation in Xenopus laevis cell-free extracts: firstly, how CDK1 becomes inactivated and secondly, how other actors, like kinases and phosphatases network and/or specific inhibitors, cooperate with CDK1 inactivation to assure timely exit from the M-phase.

  20. Involvement of the D-type cyclins in germ cell proliferation and differentiation in the mouse

    NARCIS (Netherlands)

    Beumer, T. L.; Roepers-Gajadien, H. L.; Gademan, I. S.; Kal, H. B.; de rooij, D. G.

    2000-01-01

    Using immunohistochemistry, the expression of the D-type cyclin proteins was studied in the developing and adult mouse testis. Both during testicular development and in adult testis, cyclin D(1) is expressed only in proliferating gonocytes and spermatogonia, indicating a role for cyclin D(1) in

  1. Cyclin D1 is a direct transcriptional target of GATA3 in neuroblastoma tumor cells

    NARCIS (Netherlands)

    Molenaar, J. J.; Ebus, M. E.; Koster, J.; Santo, E.; Geerts, D.; Versteeg, R.; Caron, H. N.

    2010-01-01

    Almost all neuroblastoma tumors express excess levels of Cyclin D1 (CCND1) compared to normal tissues and other tumor types. Only a small percentage of these neuroblastoma tumors have high-level amplification of the Cyclin D1 gene. The other neuroblastoma tumors have equally high Cyclin D1

  2. SCTR regulates cell cycle-related genes toward anti-proliferation in normal breast cells while having pro-proliferation activity in breast cancer cells.

    Science.gov (United States)

    Kang, Seongeun; Kim, Byungtak; Kang, Han-Sung; Jeong, Gookjoo; Bae, Hansol; Lee, Hyunkyung; Lee, Seungyeon; Kim, Sun Jung

    2015-11-01

    Secretin receptor (SCTR), the G-protein coupled receptor (GPCR) for secretin, has been observed to be upregulated in a few tumor types while downregulated in others, promoting or suppressing the proliferation of tumor cells, respectively. However, little is known about the molecular regulatory mechanism of dysregulation in cancer. In the present study, an analysis of the biological pathways affected by methylation in breast cancer using the methylome databases revealed that GPCRs played a major part in the affected pathway. SCTR, one of the dysregulated GPCRs, showed hypermethylation (pcells identified the G2/M stage checkpoint as the top-scored pathway. Cell cycle-related genes were all upregulated or downregulated suppressing cell proliferation. However, the overexpression of SCTR in MCF-7 cells led to a 35% increase of the cell proliferation index and 2.1-fold increase of cellular migration. Our findings indicate that SCTR suppresses the proliferation of normal breast cells, while the gene stimulates the proliferation and migration of cancer cells being downregulated by promoter methylation.

  3. Elevated cyclin A associated kinase activity promotes sensitivity of metastatic human cancer cells to DNA antimetabolite drug.

    Science.gov (United States)

    Wang, Jin; Yin, Hailin; Panandikar, Ashwini; Gandhi, Varsha; Sen, Subrata

    2015-08-01

    Drug resistance is a major obstacle in successful systemic therapy of metastatic cancer. We analyzed the involvement of cell cycle regulatory proteins in eliciting response to N (phosphonoacetyl)-L-aspartate (PALA), an inhibitor of de novo pyrimidine synthesis, in two metastatic variants of human cancer cell line MDA-MB-435 isolated from lung (L-2) and brain (Br-1) in nude mouse, respectively. L-2 and Br-l cells markedly differed in their sensitivity to PALA. While both cell types displayed an initial S phase delay/arrest, Br-l cells proliferated but most L-2 cells underwent apoptosis. There was distinct elevation in cyclin A, and phosphorylated Rb proteins concomitant with decreased expression of bcl-2 protein in the PALA treated L-2 cells undergoing apoptosis. Markedly elevated cyclin A associated and cdk2 kinase activities together with increased E2F1-DNA binding were detected in these L-2 cells. Induced ectopic cyclin A expression sensitized Br-l cells to PALA by activating an apoptotic pathway. Our findings demonstrate that elevated expression of cyclin A and associated kinase can activate an apoptotic pathway in cells exposed to DNA antimetabolites. Abrogation of this pathway can lead to resistance against these drugs in metastatic variants of human carcinoma cells.

  4. Differentiation-inducing factor-1 induces cyclin D1 degradation through the phosphorylation of Thr286 in squamous cell carcinoma

    International Nuclear Information System (INIS)

    Mori, Jun; Takahashi-Yanaga, Fumi; Miwa, Yoshikazu; Watanabe, Yutaka; Hirata, Masato; Morimoto, Sachio; Shirasuna, Kanemitsu; Sasaguri, Toshiyuki

    2005-01-01

    Differentiation-inducing factors (DIFs) are morphogens which induce cell differentiation in Dictyostelium. We reported that DIF-1 and DIF-3 inhibit proliferation and induce differentiation in mammalian cells. In this study, we investigated the effect of DIF-1 on oral squamous cell carcinoma cell lines NA and SAS, well differentiated and poorly differentiated cell lines, respectively. Although DIF-1 did not induce the expression of cell differentiation makers in these cell lines, it inhibited the proliferation of NA and SAS in a dose-dependent manner by restricting the cell cycle in the G 0 /G 1 phase. DIF-1 induced cyclin D1 degradation, but this effect was prevented by treatment with lithium chloride and SB216763, the inhibitors of glycogen synthase kinase-3β (GSK-3β). Depletion of endogenous GSK-3β by RNA interference also attenuated the effect of DIF-1 on cyclin D1 degradation. Therefore, we investigated the effect of DIF-1 on GSK-3β and found that DIF-1 dephosphorylated GSK-3β on Ser 9 and induced the nuclear translocation of GSK-3β, suggesting that DIF-1 activated GSK-3β. Then, we examined the effect of DIF-1 on cyclin D1 mutants (Thr286Ala, Thr288Ala, and Thr286/288Ala). We revealed that Thr286Ala and Thr286/288Ala mutants were highly resistant to DIF-1-induced degradation compared with wild-type cyclin D1, indicating that the phosphorylation of Thr 286 was critical for cyclin D1 degradation induced by DIF-1. These results suggest that DIF-1 induces degradation of cyclin D1 through the GSK-3β-mediated phosphorylation of Thr 286

  5. CREG Promotes the Proliferation of Human Umbilical Vein Endothelial Cells through the ERK/Cyclin E Signaling Pathway

    Directory of Open Access Journals (Sweden)

    Xinliang Ma

    2013-09-01

    Full Text Available Cellular repressor of E1A-stimulated genes (CREG is a recently discovered secreted glycoprotein involved in homeostatic modulation. We previously reported that CREG is abundantly expressed in the adult vascular endothelium and dramatically downregulated in atherosclerotic lesions. In addition, CREG participates in the regulation of apoptosis, inflammation and wound healing of vascular endothelial cells. In the present study, we attempted to investigate the effect of CREG on the proliferation of vascular endothelial cells and to decipher the underlying molecular mechanisms. Overexpression of CREG in human umbilical vein endothelial cells (HUVEC was obtained by infection with adenovirus carrying CREG. HUVEC proliferation was investigated by flow cytometry and 5-bromo-2'-deoxy-uridine (BrdU incorporation assays. The expressions of cyclins, cyclin-dependent kinases and signaling molecules were also examined. In CREG-overexpressing cells, we observed a marked increase in the proportion of the S and G2 population and a decrease in the G0/G1 phase population. The number of BrdU positively-stained cells also increased, obviously. Furthermore, silencing of CREG expression by specific short hairpin RNA effectively inhibited the proliferation of human umbilical vein endothelial cells (HUVEC. CREG overexpression induced the expression of cyclin E in both protein and mRNA levels to regulate cell cycle progression. Further investigation using inhibitor blocking analysis identified that ERK activation mediated the CREG modulation of the proliferation and cyclin E expression in HUVEC. In addition, blocking vascular endothelial growth factor (VEGF in CREG-overexpressed HUVEC and supplementation of VEGF in CREG knocked-down HUVEC identified that the pro-proliferative effect of CREG was partially mediated by VEGF-induced ERK/cyclin E activation. These results suggest a novel role of CREG to promote HUVEC proliferation through the ERK/cyclin E signaling pathway.

  6. Zinc fingers and homeoboxes 2 inhibits hepatocellular carcinoma cell proliferation and represses expression of Cyclins A and E.

    Science.gov (United States)

    Yue, Xuetian; Zhang, Zhenyu; Liang, Xiaohong; Gao, Lifen; Zhang, Xiaoning; Zhao, Di; Liu, Xiao; Ma, Hongxin; Guo, Min; Spear, Brett T; Gong, Yaoqin; Ma, Chunhong

    2012-06-01

    Zinc-fingers and homeoboxes 2 (ZHX2) represses transcription of several genes associated with liver cancer. However, little is known about the role of ZHX2 in the development of hepatocellular carcinoma (HCC). We investigated the mechanisms by which ZHX2 might affect proliferation of HCC cells. We overexpressed and knocked down ZHX2 in HCC cells and analyzed the effects on proliferation, colony formation, and the cell cycle. We also analyzed the effects of ZHX2 overexpression in growth of HepG2.2.15 tumor xenografts in nude mice. Chromatin immunoprecipitation and luciferase reporter assays were used to measure binding of ZHX2 target promoters. Levels of ZHX2 in HCC samples were evaluated by immunohistochemistry. ZHX2 overexpression significantly reduced proliferation of HCC cells and growth of tumor xenografts in mice; it led to G1 arrest and reduced levels of Cyclins A and E in HCC cell lines. ZHX2 bound to promoter regions of CCNA2 (which encodes Cyclin A) and CCNE1 (which encodes Cyclin E) and inhibited their transcription. Knockdown of Cyclin A or Cyclin E reduced the increased proliferation mediated by ZHX2 knockdown. Nuclear localization of ZHX2 was required for it to inhibit proliferation of HCC cells in culture and in mice. Nuclear localization of ZHX2 was reduced in human HCC samples, even in small tumors (diameter, <5 cm), compared with adjacent nontumor tissues. Moreover, reduced nuclear levels of ZHX2 correlated with reduced survival times of patients, high levels of tumor microvascularization, and hepatocyte proliferation. ZHX2 inhibits HCC cell proliferation by preventing expression of Cyclins A and E, and reduces growth of xenograft tumors in mice. Loss of nuclear ZHX2 might be an early step in the development of HCC. Copyright © 2012 AGA Institute. Published by Elsevier Inc. All rights reserved.

  7. Speeding through cell cycle roadblocks: Nuclear cyclin D1-dependent kinase and neoplastic transformation

    Directory of Open Access Journals (Sweden)

    Diehl J Alan

    2008-09-01

    Full Text Available Abstract Mitogenic induction of cyclin D1, the allosteric regulator of CDK4/6, is a key regulatory event contributing to G1 phase progression. Following the G1/S transition, cyclin D1 activation is antagonized by GSK3β-dependent threonine-286 (Thr-286 phosphorylation, triggering nuclear export and subsequent cytoplasmic degradation mediated by the SCFFbx4-αBcrystallin E3 ubiquitin ligase. Although cyclin D1 overexpression occurs in numerous malignancies, overexpression of cyclin D1 alone is insufficient to drive transformation. In contrast, cyclin D1 mutants refractory to phosphorylation-dependent nuclear export and degradation are acutely transforming. This raises the question of whether overexpression of cyclin D1 is a significant contributor to tumorigenesis or an effect of neoplastic transformation. Significantly, recent work strongly supports a model wherein nuclear accumulation of cyclin D1-dependent kinase during S-phase is a critical event with regard to transformation. The identification of mutations within SCFFbx4-αBcrystallin ligase in primary tumors provides mechanistic insight into cyclin D1 accumulation in human cancer. Furthermore, analysis of mouse models expressing cyclin D1 mutants refractory to degradation indicate that nuclear cyclin D1/CDK4 kinase triggers DNA re-replication and genomic instability. Collectively, these new findings provide a mechanism whereby aberrations in post-translational regulation of cyclin D1 establish a cellular environment conducive to mutations that favor neoplastic growth.

  8. Overexpression of cyclin D1 induces the reprogramming of differentiated epidermal cells into stem cell-like cells.

    Science.gov (United States)

    Zhao, Along; Yang, Leilei; Ma, Kui; Sun, Mengli; Li, Lei; Huang, Jin; Li, Yang; Zhang, Cuiping; Li, Haihong; Fu, Xiaobing

    2016-01-01

    It has been reported that Wnt/β-catenin is critical for dedifferentiation of differentiated epidermal cells. Cyclin D1 (CCND1) is a β-catenin target gene. In this study, we provide evidence that overexpression of CCND1 induces reprogramming of epidermal cells into stem cell-like cells. After introducing CCND1 gene into differentiated epidermal cells, we found that the large flat-shaped cells with a small nuclear-cytoplasmic ratio changed into small round-shaped cells with a large nuclear-cytoplasmic ratio. The expressions of CK10, β1-integrin, Oct4 and Nanog in CCND1 induced cells were remarkably higher than those in the control group (P cells exhibited a high colony-forming ability and a long-term proliferative potential. When the induced cells were implanted into a wound of laboratory animal model, the wound healing was accelerated. These results suggested that overexpression of CCND1 induced the reprogramming of differentiated epidermal cells into stem cell-like cells. This study may also offer a new approach to yield epidermal stem cells for wound repair and regeneration.

  9. Regulation of Microtubule, Apoptosis, and Cell Cycle-Related Genes by Taxotere in Prostate Cancer Cells Analyzed by Microarray

    Directory of Open Access Journals (Sweden)

    Yiwei Li

    2004-03-01

    Full Text Available Taxotere showed antitumor activity against solid tumors including prostate cancer. However, the molecular mechanism(s of action of Taxotere has not been fully elucidated. In order to establish such molecular mechanism(s in both hormone-insensitive (PC3 and hormone-sensitive (LNCaP prostate cancer cells, comprehensive gene expression profiles were obtained by Affymetrix Human Genome U133A Array. The RNA from the cells treated with 2 nM Taxotere was subjected to microarray analysis. We found that a total of 166, 365, and 1785 genes showed greater than twofold change in PC3 cells after 6, 36, and 72 hours of treatment, respectively compared to 57, 823, and 964 genes in LNCaP cells. The expression of tubulin was decreased, whereas the expression of microtubuleassociated proteins was increased in Taxotere-treated prostate cancer cells, confirming the microtubuletargeting effect of Taxotere. Clustering analysis showed downregulation of some genes for cell proliferation and cell cycle. In contrast, Taxotere upregulated some genes that are related to induction of apoptosis and cell cycle arrest. From these results, we conclude that Taxotere caused alterations of a large number of genes, many of which may contribute to the molecular mechanism(s by which Taxotere affects prostate cancer cells. Further molecular studies are needed in order to determine the cause and effect relationships between these genes altered by Taxotere. Nevertheless, our results could be further exploited for devising strategies to optimize therapeutic effects of Taxotere for the treatment of prostate cancer.

  10. The influence of arsenic trioxide on the cell cycle, apoptosis and expression of cyclin D1 in the Jurkat cell line.

    Science.gov (United States)

    Zuryń, Agnieszka; Litwiniec, Anna; Gagat, Maciej; Drzewucka, Joanna; Gackowska, Lidia; Grzanka, Alina

    2014-10-01

    Cyclin D1 drives cell cycle progression at the G1/S transition and is believed to play a significant role in tumorigenesis, contributing to efficient proliferation of many cancer cells. Consequently, it is also recognized as an end-point biomarker of therapeutic outcome for different treatment modalities in cancer. In this study we aimed to evaluate the expression and localization of cyclin D1 in arsenic trioxide (ATO) treated Jurkat cells (lymphoblastic leukemia cell line) and to correlate these results with the extent of cell death and/or cell cycle alterations. Jurkat cells were incubated with increasing concentrations of ATO (0.2, 0.6 and 1.0μM) for 24h in standard cell culture conditions. To reach our goal we performed annexin V/PI labeling for detection of cell death and RNase/PI labeling for evaluation of cell cycle distribution, which were followed by the respective flow cytometric analyses of ATO-treated Jurkat cells. Transmission electron microscopy was applied for visualization of the cell ultrastructure. For cyclin D1 estimation a biparametric cyclinD1/cell cycle assay was done and localization of the protein was shown after immuno-labeling using light microscopy (ABC procedure) and confocal fluorescence microscopy. We found that there were no significant changes in the percentages of cyclin D1-positive cells after the treatment with ATO, but at the same time mean fluorescence intensity reflecting cyclin D1 content was gradually increasing along with the cell cycle progression, irrespective of the applied dose of the drug. On the other hand, we found a nuclear-cytoplasmic shift of this protein as a major treatment-related response, which was in good accord with an increased rate of cell death and suggested that cyclin D1 cytoplasmic degradation is an important determinant of the therapeutic efficiency of ATO in the Jurkat cell line. Copyright © 2014 Elsevier GmbH. All rights reserved.

  11. The Rts1 regulatory subunit of protein phosphatase 2A is required for control of G1 cyclin transcription and nutrient modulation of cell size.

    Directory of Open Access Journals (Sweden)

    Karen Artiles

    2009-11-01

    Full Text Available The key molecular event that marks entry into the cell cycle is transcription of G1 cyclins, which bind and activate cyclin-dependent kinases. In yeast cells, initiation of G1 cyclin transcription is linked to achievement of a critical cell size, which contributes to cell-size homeostasis. The critical cell size is modulated by nutrients, such that cells growing in poor nutrients are smaller than cells growing in rich nutrients. Nutrient modulation of cell size does not work through known critical regulators of G1 cyclin transcription and is therefore thought to work through a distinct pathway. Here, we report that Rts1, a highly conserved regulatory subunit of protein phosphatase 2A (PP2A, is required for normal control of G1 cyclin transcription. Loss of Rts1 caused delayed initiation of bud growth and delayed and reduced accumulation of G1 cyclins. Expression of the G1 cyclin CLN2 from an inducible promoter rescued the delayed bud growth in rts1Delta cells, indicating that Rts1 acts at the level of transcription. Moreover, loss of Rts1 caused altered regulation of Swi6, a key component of the SBF transcription factor that controls G1 cyclin transcription. Epistasis analysis revealed that Rts1 does not work solely through several known critical upstream regulators of G1 cyclin transcription. Cells lacking Rts1 failed to undergo nutrient modulation of cell size. Together, these observations demonstrate that Rts1 is a key player in pathways that link nutrient availability, cell size, and G1 cyclin transcription. Since Rts1 is highly conserved, it may function in similar pathways in vertebrates.

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

    Science.gov (United States)

    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. Copyright © 2016 Elsevier Inc. All rights reserved.

  13. Glucose Induces Mouse β-Cell Proliferation via IRS2, MTOR, and Cyclin D2 but Not the Insulin Receptor

    Science.gov (United States)

    Stamateris, Rachel E.; Sharma, Rohit B.; Kong, Yahui; Ebrahimpour, Pantea; Panday, Deepika; Ranganath, Pavana; Zou, Baobo; Levitt, Helena; Parambil, Nisha Abraham; O’Donnell, Christopher P.; García-Ocaña, Adolfo

    2016-01-01

    An important goal in diabetes research is to understand the processes that trigger endogenous β-cell proliferation. Hyperglycemia induces β-cell replication, but the mechanism remains debated. A prime candidate is insulin, which acts locally through the insulin receptor. Having previously developed an in vivo mouse hyperglycemia model, we tested whether glucose induces β-cell proliferation through insulin signaling. By using mice lacking insulin signaling intermediate insulin receptor substrate 2 (IRS2), we confirmed that hyperglycemia-induced β-cell proliferation requires IRS2 both in vivo and ex vivo. Of note, insulin receptor activation was not required for glucose-induced proliferation, and insulin itself was not sufficient to drive replication. Glucose and insulin caused similar acute signaling in mouse islets, but chronic signaling differed markedly, with mammalian target of rapamycin (MTOR) and extracellular signal–related kinase (ERK) activation by glucose and AKT activation by insulin. MTOR but not ERK activation was required for glucose-induced proliferation. Cyclin D2 was necessary for glucose-induced β-cell proliferation. Cyclin D2 expression was reduced when either IRS2 or MTOR signaling was lost, and restoring cyclin D2 expression rescued the proliferation defect. Human islets shared many of these regulatory pathways. Taken together, these results support a model in which IRS2, MTOR, and cyclin D2, but not the insulin receptor, mediate glucose-induced proliferation. PMID:26740601

  14. Fluorescence energy transfer monitoring of protein-protein interaction in human cells: the Cyclin T1-HIV1 Tat case.

    Science.gov (United States)

    Ferrari, Aldo; Cinelli, Riccardo A. G.; Pellegrini, Vittorio; Beltram, Fabio; Marcello, Alessandro; Tyagi, Mudit; Giacca, Mauro

    2001-03-01

    The human immunodeficiency virus type 1 (HIV-1) Tat protein promotes transcriptional elongation of viral RNAs. Here we show that human Cyclin T1 directly binds Tat in cultured cells. By mapping fluorescence resonance energy transfer (FRET) in different cellular compartments we shall present a quantitative analysis of this interaction. The matched tagging pair consists of two optically matched variants of the green fluorescent protein: the enhanced GFP and the blue fluorescent protein. Strong energy transfer was observed between Cyclin T1 and Tat both in the cytoplasm and in specific subnuclear regions. We shall argue that such high-resolution optical studies can provide significant new insight in molecular processes and demonstrate that, for the specific case-study presented, they lead to a model by which Tat recruits Cyclin T1 out of the nuclear compartments where the protein resides to promote transcriptional activation.

  15. Selective Cyclin-Dependent Kinase Inhibitors Discriminating between Cell Cycle and Transcriptional Kinases Future Reality or Utopia?

    Czech Academy of Sciences Publication Activity Database

    Wesierska-Gadek, J.; Kryštof, Vladimír

    2009-01-01

    Roč. 1171, - (2009), s. 228-241 ISSN 0077-8923 R&D Projects: GA ČR GA204/08/0511 Institutional research plan: CEZ:AV0Z50380511 Keywords : cell cycle * CYC202 * cyclin-dependent kinase Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.670, year: 2009

  16. Overexpression of cyclin D1 indicates a poor prognosis in squamous cell carcinoma of the head and neck

    NARCIS (Netherlands)

    Michalides, R. J.; van Veelen, N. M.; Kristel, P. M.; Hart, A. A.; Loftus, B. M.; Hilgers, F. J.; Balm, A. J.

    1997-01-01

    To evaluate the overexpression of cyclin D1 and p53 as a prognostic marker of squamous cell carcinoma of the head and neck and to investigate whether deregulation of these genes is associated with an unfavorable course of disease. Retrospective study. Formalin-fixed, paraffin-embedded tumor

  17. Cyclin D-cdk6 complex is targeted by p21WAF in growth-arrested lymphoma cells

    Czech Academy of Sciences Publication Activity Database

    Tvrdík, Daniel; Djaborkhel, Rashed; Nagy, Alexandr; Eckschlager, T.; Raška, Ivan; Müller, Julius

    2002-01-01

    Roč. 140, - (2002), s. 49-56 ISSN 1047-8477 R&D Projects: GA ČR GA304/01/0564 Institutional research plan: CEZ:AV0Z5039906; CEZ:MSM 111100003 Keywords : cyclin-dependent kinase * follicular lymphoma * retinoblastoma protein Subject RIV: EA - Cell Biology Impact factor: 4.194, year: 2002

  18. The human cyclin B1 protein modulates sensitivity of DNA mismatch repair deficient prostate cancer cell lines to alkylating agents.

    Science.gov (United States)

    Rasmussen, L J; Rasmussen, M; Lützen, A; Bisgaard, H C; Singh, K K

    2000-05-25

    DNA damage caused by alkylating agents results in a G2 checkpoint arrest. DNA mismatch repair (MMR) deficient cells are resistant to killing by alkylating agents and are unable to arrest the cell cycle in G2 phase after alkylation damage. We investigated the response of two MMR-deficient prostate cancer cell lines DU145 and LNCaP to the alkylating agent MNNG. Our studies reveal that DU145 cancer cells are more sensitive to killing by MNNG than LNCaP. Investigation of the underlying reasons for lower resistance revealed that the DU145 cells contain low endogenous levels of cyclin B1. We provide direct evidence that the endogenous level of cyclin B1 modulates the sensitivity of MMR-deficient prostate cancer cells to alkylating agents.

  19. Inverse relationship between TCTP/RhoA and p53/ /cyclin A/actin expression in ovarian cancer cells Inverse relationship between TCTP/RhoA and p53/ /cyclin A/actin expression in ovarian cancer cells

    Directory of Open Access Journals (Sweden)

    Malgorzata Kloc

    2012-10-01

    Full Text Available The translationally controlled tumor protein (TCTP plays a role in cell growth, cell cycle and cancer
    progression. TCTP controls negatively the stability of the p53 tumor suppressor protein and interacts with the
    cellular cytoskeleton. The deregulation of the actin and cytokeratin cytoskeleton is responsible for the increased
    migratory activity of tumor cells and is linked with poor patient outcome. Recent studies indicate that cyclin A,
    a key regulator of cell cycle, controls actin organization and negatively regulates cell motility via regulation of RhoA
    expression. We studied the organization of actin and cytokeratin cytoskeleton and the expression of TCTP, p53,
    cyclin A, RhoA and actin in HIO180 non-transformed ovarian epithelial cells, and OVCAR3 and SKOV3 (expressing
    low level of inducible p53 ovarian epithelial cancer cells with different metastatic potential. Immunostaining
    and ultrastructural analyses illustrated a dramatic difference in the organization of the cytokeratin and actin
    filaments in non-transformed versus cancer cell lines. We also determined that there is an inverse relationship between
    the level of TCTP/RhoA and actin/p53/cyclin A expression in ovarian cancer cell lines. This previously unidentified
    negative relationship between TCTP/RhoA and actin/p53/cyclin A may suggest that this interaction is linked
    with the high aggressiveness of ovarian cancers.The translationally controlled tumor protein (TCTP plays a role in cell growth, cell cycle and cancer
    progression. TCTP controls negatively the stability of the p53 tumor suppressor protein and interacts with the
    cellular cytoskeleton. The deregulation of the actin and cytokeratin cytoskeleton is responsible for the increased
    migratory activity of tumor cells and is linked with poor patient outcome. Recent studies indicate that cyclin A,
    a key regulator of cell cycle, controls actin organization

  20. Impaired liver regeneration is associated with reduced cyclin B1 in natural killer T cell-deficient mice.

    Science.gov (United States)

    Ben Ya'acov, Ami; Meir, Hadar; Zolotaryova, Lydia; Ilan, Yaron; Shteyer, Eyal

    2017-03-23

    It has been shown that the proportion of natural killer T cells is markedly elevated during liver regeneration and their activation under different conditions can modulate this process. As natural killer T cells and liver injury are central in liver regeneration, elucidating their role is important. The aim of the current study is to explore the role of natural killer T cells in impaired liver regeneration. Concanvalin A was injected 4 days before partial hepatectomy to natural killer T cells- deficient mice or to anti CD1d1-treated mice. Ki-67 and proliferating cell nuclear antigen were used to measure hepatocytes proliferation. Expression of hepatic cyclin B1 and proliferating cell nuclear antigen were evaluated by Western Blot and liver injury was assessed by ALT and histology. Natural killer T cells- deficient or mice injected with anti CD1d antibodies exhibited reduced liver regeneration. These mice were considerably resistant to ConA-induced liver injury. In the absence of NKT cells hepatic proliferating cell nuclear antigen and cyclin B1 decreased in mice injected with Concanvalin A before partial hepatectomy. This was accompanied with reduced serum interleukin-6 levels. Natural killer T cells play an important role in liver regeneration, which is associated with cyclin B1 and interleukin-6.

  1. Overexpression of cyclin D1 correlates with recurrence in a group of forty-seven operable squamous cell carcinomas of the head and neck

    NARCIS (Netherlands)

    Michalides, R.; van Veelen, N.; Hart, A.; Loftus, B.; Wientjens, E.; Balm, A.

    1995-01-01

    We evaluated the prognostic significance of overexpression of cyclin D1 in 47 patients with surgically resected squamous cell carcinomas of the head and neck. Overexpression of cyclin D1 was detected immunohistochemically using an affinity-purified polyclonal antibody directed against the

  2. Hypoxia Upregulates Mitotic Cyclins Which Contribute to the Multipotency of Human Mesenchymal Stem Cells by Expanding Proliferation Lifespan.

    Science.gov (United States)

    Lee, Janet; Kim, Hyun-Soo; Kim, Su-Min; Kim, Dong-Ik; Lee, Chang-Woo

    2018-02-21

    Hypoxic culture is widely recognized as a method to efficiently expand human mesenchymal stem cells (MSCs) without loss of stem cell properties. However, the molecular basis of how hypoxia priming benefits MSC expansion remains unclear. In this report, our systemic quantitative proteomic and RT-PCR analyses revealed the involvement of hypoxic conditioning activated genes in the signaling process of the mitotic cell cycle. Introduction of screened two mitotic cyclins, CCNA2 and CCNB1, significantly extended the proliferation lifespan of MSCs in normoxic condition. Our results provide important molecular evidence that multipotency of human MSCs by hypoxic conditioning is determined by the mitotic cell cycle duration. Thus, the activation of mitotic cyclins could be a potential strategy to the application of stem cell therapy.

  3. Protective effect of epigenetic silencing of CyclinD1 against spinal cord injury using bone marrow-derived mesenchymal stem cells in rats.

    Science.gov (United States)

    Wang, Yuan; Kong, Qing-Jie; Sun, Jin-Chuan; Xu, Xi-Ming; Yang, Yong; Liu, Ning; Shi, Jian-Gang

    2018-07-01

    This study focuses on the protective effect of epigenetic silencing of CyclinD1 against spinal cord injury (SCI) using bone marrow-derived mesenchymal stem cells (BMSCs) in rats. Eighty-eight adult female Wistar rats were randomly assigned into the sham group, the control group, the si-CyclinD1 + BMSCs group and the BMSCs group. CyclinD1 protein and mRNA expressions after siRNA transfection were detected by Western blotting and qRT-PCR. The siRNA-CyclinD1 BMSCs were transplanted into rats in the si-CyclinD1 + BMSCs group using stereotaxic method 6 hr after SCI. Hindlimb locomotor performance was determined using inclined plane test and Basso-Beattie-Bresnahan (BBB) locomotor rating scale. Expressions of glial fibrillary acidic protein (GFAP) and nerve growth factor (NGF) were detected by immunohistochemistry. Inclined plane and BBB scores in the control, si-CyclinD1 + BMSCs, and BMSCs groups were significantly lower than the sham group, but these scores were evidently decreased in the control group and increased in the si-CyclinD1 + BMSCs group compared with the BMSCs group. The repair degree of spinal cord tissues of rats in the si-CyclinD1 + BMSCs group was obvious than the BMSCs group. GFAP and NGF protein expressions were markedly decreased in the control, si-CyclinD1 + BMSCs and BMSCs groups when compared with the sham group. GFAP- and NGF-positive cells were significantly increased in the si-CyclinD1 + BMSCs group while decreased in the control group. Our study provides evidence that epigenetic silencing of CyclinD1 using BMSCs might accelerate the repair of SCI in rats. © 2017 Wiley Periodicals, Inc.

  4. Cyclin-dependent kinase inhibitor 3 is overexpressed in hepatocellular carcinoma and promotes tumor cell proliferation

    International Nuclear Information System (INIS)

    Xing, Chunyang; Xie, Haiyang; Zhou, Lin; Zhou, Wuhua; Zhang, Wu; Ding, Songming; Wei, Bajin; Yu, Xiaobo; Su, Rong; Zheng, Shusen

    2012-01-01

    Highlights: ► CDKN3 is commonly overexpressed in HCC and is associated with poor clinical outcome. ► Overexpression of CDKN3 could stimulate the proliferation of HCC cells by promoting G1/S transition. ► CDKN3 could inhibit the expression of p21 in HCC cells. ► Overexpression of CDKN3 has no effect on apoptosis and invasion of HCC cells. ► We identified 61 genes co-expressed with CDKN3, and BIRC5 was located at the center of the co-expression network. -- Abstract: Cyclin-dependent kinase inhibitor 3 (CDKN3) belongs to the protein phosphatases family and has a dual function in cell cycling. The function of this gene has been studied in several kinds of cancers, but its role in human hepatocellular carcinoma (HCC) remains to be elucidated. In this study, we found that CDKN3 was frequently overexpressed in both HCC cell lines and clinical samples, and this overexpression was correlated with poor tumor differentiation and advanced tumor stage. Functional studies showed that overexpression of CDKN3 could promote cell proliferation by stimulating G1-S transition but has no impact on cell apoptosis and invasion. Microarray-based co-expression analysis identified a total of 61 genes co-expressed with CDKN3, with most of them involved in cell proliferation, and BIRC5 was located at the center of CDKN3 co-expression network. These results suggest that CDKN3 acts as an oncogene in human hepatocellular carcinoma and antagonism of CDKN3 may be of interest for the treatment of HCC.

  5. Cellular distribution of cell cycle-related molecules in the renal tubules of rats treated with renal carcinogens for 28 days: relationship between cell cycle aberration and carcinogenesis.

    Science.gov (United States)

    Taniai, Eriko; Hayashi, Hitomi; Yafune, Atsunori; Watanabe, Maiko; Akane, Hirotoshi; Suzuki, Kazuhiko; Mitsumori, Kunitoshi; Shibutani, Makoto

    2012-09-01

    Some renal carcinogens can induce karyomegaly, which reflects aberrant cell division in the renal tubules, from the early stages of exposure. To clarify the cell cycle-related changes during the early stages of renal carcinogenesis, we performed immunohistochemical analysis of tubular cells in male F344 rats treated with carcinogenic doses of representative renal carcinogens for 28 days. For this purpose, the karyomegaly-inducing carcinogens ochratoxin A (OTA), ferric nitrilotriacetic acid, and monuron, and the non-karyomegaly-inducing carcinogens tris(2-chloroethyl) phosphate and potassium bromate were examined. For comparison, a karyomegaly-inducing non-carcinogen, p-nitrobenzoic acid, and a non-carcinogenic non-karyomegaly-inducing renal toxicant, acetaminophen, were also examined. The outer stripe of the outer medulla (OSOM) and the cortex + OSOM were subjected to morphometric analysis of immunoreactive proximal tubular cells. Renal carcinogens, irrespective of their karyomegaly-inducing potential, increased proximal tubular cell proliferation accompanied by an increase in topoisomerase IIα-immunoreactive cells, suggesting a reflection of cell proliferation. Karyomegaly-inducing carcinogens increased nuclear Cdc2-, γH2AX-, and phosphorylated Chk2-immunoreactive cells in both areas, the former two acting in response to DNA damage and the latter one suggestive of sustained G₂. OTA, an OSOM-targeting carcinogen, could easily be distinguished from untreated controls and non-carcinogens by evaluation of molecules responding to DNA damage and G₂/M transition in the OSOM. Thus, all renal carcinogens examined facilitated proximal tubular proliferation by repeated short-term treatment. Among these, karyomegaly-inducing carcinogens may cause DNA damage and G₂ arrest in the target tubular cells.

  6. Change in expression of cyclin G2 in kidney cancer cell and its significance.

    Science.gov (United States)

    Cui, D W; Sun, G G; Cheng, Y J

    2014-04-01

    This study aims to analyze the expression and clinical significance of cyclin G2 (CCNG2) in kidney carcinoma, and the biological effect in its cell line by CCNG2 overexpression. Immunohistochemistry and western blot were used to analyze CCNG2 protein expression in 63 cases of kidney cancer and normal tissues to study the relationship between CCNG2 expression and clinical factors. CCNG2 lentiviral vector and empty vector were respectively transfected into kidney ACHN cell line. During immunohistochemistry, the level of CCNG2 protein expression was found to be significantly lower in kidney cancer tissue than normal tissues (P kidney cancer tissue was respectively found to be significantly lower than in normal tissues (P 0.05), but it was correlated with lymph node metastasis, clinic stage, and histological grade (P kidney cancer and correlated significantly with lymph node metastasis, clinical stage, histological grade, and poor overall survival, suggesting that CCNG2 may play important roles as a negative regulator to kidney cancer ACHN cell by promoting degradation of CDK2.

  7. Differential regulation of cyclin-dependent kinase inhibitors in neuroblastoma cells

    International Nuclear Information System (INIS)

    Qiao, Lan; Paul, Pritha; Lee, Sora; Qiao, Jingbo; Wang, Yongsheng; Chung, Dai H.

    2013-01-01

    Highlights: •GRP-R signaling differentially regulated the expression of p21 and p27. •Silencing GRP/GRP-R downregulated p21, while p27 expression was upregulated. •Inhibition of GRP/GRP-R signaling enhanced PTEN expression, correlative to the increased expression of p27. •PTEN and p27 co-localized in cytoplasm and silencing PTEN decreased p27 expression. -- Abstract: Gastrin-releasing peptide (GRP) and its receptor (GRP-R) are highly expressed in undifferentiated neuroblastoma, and they play critical roles in oncogenesis. We previously reported that GRP activates the PI3K/AKT signaling pathway to promote DNA synthesis and cell cycle progression in neuroblastoma cells. Conversely, GRP-R silencing induces cell cycle arrest. Here, we speculated that GRP/GRP-R signaling induces neuroblastoma cell proliferation via regulation of cyclin-dependent kinase (CDK) inhibitors. Surprisingly, we found that GRP/GRP-R differentially induced expressions of p21 and p27. Silencing GRP/GRP-R decreased p21, but it increased p27 expressions in neuroblastoma cells. Furthermore, we found that the intracellular localization of p21 and p27 in the nuclear and cytoplasmic compartments, respectively. In addition, we found that GRP/GRP-R silencing increased the expression and accumulation of PTEN in the cytoplasm of neuroblastoma cells where it co-localized with p27, thus suggesting that p27 promotes the function of PTEN as a tumor suppressor by stabilizing PTEN in the cytoplasm. GRP/GRP-R regulation of CDK inhibitors and tumor suppressor PTEN may be critical for tumoriogenesis of neuroblastoma

  8. Differential regulation of cyclin-dependent kinase inhibitors in neuroblastoma cells

    Energy Technology Data Exchange (ETDEWEB)

    Qiao, Lan [Department of Pediatric Surgery, Vanderbilt University Medical Center, Nashville, TN 37232 (United States); Department of Pharmaceutical Sciences, Jilin University, Changchun 130021 (China); Paul, Pritha; Lee, Sora [Department of Pediatric Surgery, Vanderbilt University Medical Center, Nashville, TN 37232 (United States); Department of Cancer Biology, Vanderbilt University Medical Center, Nashville, TN 37232 (United States); Qiao, Jingbo [Department of Pediatric Surgery, Vanderbilt University Medical Center, Nashville, TN 37232 (United States); Wang, Yongsheng [Department of Pharmaceutical Sciences, Jilin University, Changchun 130021 (China); Chung, Dai H., E-mail: dai.chung@vanderbilt.edu [Department of Pediatric Surgery, Vanderbilt University Medical Center, Nashville, TN 37232 (United States); Department of Cancer Biology, Vanderbilt University Medical Center, Nashville, TN 37232 (United States)

    2013-05-31

    Highlights: •GRP-R signaling differentially regulated the expression of p21 and p27. •Silencing GRP/GRP-R downregulated p21, while p27 expression was upregulated. •Inhibition of GRP/GRP-R signaling enhanced PTEN expression, correlative to the increased expression of p27. •PTEN and p27 co-localized in cytoplasm and silencing PTEN decreased p27 expression. -- Abstract: Gastrin-releasing peptide (GRP) and its receptor (GRP-R) are highly expressed in undifferentiated neuroblastoma, and they play critical roles in oncogenesis. We previously reported that GRP activates the PI3K/AKT signaling pathway to promote DNA synthesis and cell cycle progression in neuroblastoma cells. Conversely, GRP-R silencing induces cell cycle arrest. Here, we speculated that GRP/GRP-R signaling induces neuroblastoma cell proliferation via regulation of cyclin-dependent kinase (CDK) inhibitors. Surprisingly, we found that GRP/GRP-R differentially induced expressions of p21 and p27. Silencing GRP/GRP-R decreased p21, but it increased p27 expressions in neuroblastoma cells. Furthermore, we found that the intracellular localization of p21 and p27 in the nuclear and cytoplasmic compartments, respectively. In addition, we found that GRP/GRP-R silencing increased the expression and accumulation of PTEN in the cytoplasm of neuroblastoma cells where it co-localized with p27, thus suggesting that p27 promotes the function of PTEN as a tumor suppressor by stabilizing PTEN in the cytoplasm. GRP/GRP-R regulation of CDK inhibitors and tumor suppressor PTEN may be critical for tumoriogenesis of neuroblastoma.

  9. MicroRNA-193b represses cell proliferation and regulates cyclin D1 in melanoma.

    Science.gov (United States)

    Chen, Jiamin; Feilotter, Harriet E; Paré, Geneviève C; Zhang, Xiao; Pemberton, Joshua G W; Garady, Cherif; Lai, Dulcie; Yang, Xiaolong; Tron, Victor A

    2010-05-01

    Cutaneous melanoma is an aggressive form of human skin cancer characterized by high metastatic potential and poor prognosis. To better understand the role of microRNAs (miRNAs) in melanoma, the expression of 470 miRNAs was profiled in tissue samples from benign nevi and metastatic melanomas. We identified 31 miRNAs that were differentially expressed (13 up-regulated and 18 down-regulated) in metastatic melanomas relative to benign nevi. Notably, miR-193b was significantly down-regulated in the melanoma tissues examined. To understand the role of miR-193b in melanoma, functional studies were undertaken. Overexpression of miR-193b in melanoma cell lines repressed cell proliferation. Gene expression profiling identified 314 genes down-regulated by overexpression of miR-193b in Malme-3M cells. Eighteen of these down-regulated genes, including cyclin D1 (CCND1), were also identified as putative miR-193b targets by TargetScan. Overexpression of miR-193b in Malme-3M cells down-regulated CCND1 mRNA and protein by > or = 50%. A luciferase reporter assay confirmed that miR-193b directly regulates CCND1 by binding to the 3'untranslated region of CCND1 mRNA. These studies indicate that miR-193b represses cell proliferation and regulates CCND1 expression and suggest that dysregulation of miR-193b may play an important role in melanoma development.

  10. cycMs3, a novel B-type alfalfa cyclin gene, is induced in the G0-to-G1 transition of the cell cycle.

    Science.gov (United States)

    Meskiene, I; Bögre, L; Dahl, M; Pirck, M; Ha, D T; Swoboda, I; Heberle-Bors, E; Ammerer, G; Hirt, H

    1995-06-01

    Cyclins are key regulators of the cell cycle in all eukaryotes. We have previously isolated two B-type cyclin genes, cycMs1 and cycMs2, from alfalfa that are primarily expressed during the G2-to-M phase transition and are most likely mitotic cyclin genes. Here, we report the isolation of a novel alfalfa cyclin gene, termed cycMs3 (for cyclin Medicago sativa), by selecting for mating type alpha-pheromone-induced cell cycle arrest suppression in yeast. The central region of the predicted amino acid sequence of the cycMs3 gene is most similar to the cyclin box of yeast B-type and mammalian A- and B-type cyclins. In situ hybridization showed that cycMs3 mRNA can be detected only in proliferating cells and not in differentiated alfalfa cells. When differentiated G0-arrested cells were induced to reenter the cell cycle in the G1 phase and resume cell division by treatment with plant hormones, cycMs3 transcript levels increased long before the onset of DNA synthesis. In contrast, histone H3-1 mRNA and cycMs2 transcripts were not observed before DNA replication and mitosis, respectively. In addition, cycMs3 mRNA was found in all stages of the cell cycle in synchronously dividing cells, whereas the cycMs2 and histone H3-1 genes showed a G2-to-M phase- or S phase-specific transcription pattern, respectively. These data suggest that the role of cyclin CycMs3 differs from that of CycMs1 and CycMs2. We propose that CycMs3 helps control reentry of quiescent G0-arrested cells into the G1 phase of the cell cycle.

  11. Stabilization of cyclin E and cdk2 mRNAs at G1/S transition in Rat-1A cells emerging from the G0 state.

    Science.gov (United States)

    Oda, S; Nishida, J; Nakabeppu, Y; Sekiguchi, M

    1995-04-06

    mRNAs for cyclin E and Cdk2 have a role in the commitment to DNA replication in the cell cycle, and are induced in Rat-1A cells by serum stimulation. Cyclin E and cdk2 genes are transcribed in quiescent cells, but their transcripts rapidly turn over and levels are kept low. The rate of transcription of the cdk2 gene is slightly increased after serum stimulation, while that of cyclin E is fairly constant. At the G1/S transition of serum-stimulated cells, transient stabilization of the two types of mRNAs occurs, an event which may lead to induction of each mRNA. Artificial expression of an immediate-early protein delta FosB results in proliferation of quiescent Rat-1A cells, and this is accompanied by an efficient induction of cyclin E and cdk2 mRNAs. In delta FosB-expressing cells, two types of mRNAs are stabilized to the same extent seen in serum-stimulated cells. The expression of cyclin E and cdk2 genes is upregulated by stabilization of their transcripts, at least in part. We propose that delta FosB may have a role in regulation of progression of the cell cycle in serum-stimulated Rat-1A cells by triggering stabilization of mRNAs for cyclin E and Cdk2.

  12. Rottlerin inhibits the nuclear factor kappaB/cyclin-D1 cascade in MCF-7 breast cancer cells

    Czech Academy of Sciences Publication Activity Database

    Torricelli, C.; Fortino, V.; Capurro, E.; Valacchi, G.; Pacini, A.; Muscettola, M.; Souček, Karel; Maioli, E.

    2008-01-01

    Roč. 82, 11-12 (2008), s. 638-643 ISSN 0024-3205 R&D Projects: GA ČR(CZ) GA310/07/0961; GA AV ČR(CZ) 1QS500040507 Institutional research plan: CEZ:AV0Z50040507; CEZ:AV0Z50040702 Keywords : Rottlerin * MCF-7 cells * cyclin -D1 Subject RIV: BO - Biophysics Impact factor: 2.583, year: 2008

  13. Isolation of Proteins Interacting with the Cyclin D1-CDK6 Complex from Normal and Tumorigenic Human Breast Cells Using a Novel Yeast Three-Hybrid System

    National Research Council Canada - National Science Library

    Nichols, Michael

    1998-01-01

    ...% of human breast cancer cases. Cyclin D1, through its association with the catalytic subunit CDK6 and CDK4, controls G1 cell cycle progression, presumably by phosphorylating a substrate protein...

  14. Comparative proteome profiling of MCF10A and 184A1 human breast epithelial cells emphasized involvement of CDK4 and cyclin D3 in cell proliferation.

    Science.gov (United States)

    Bhaskaran, Nimesh; Lin, Kah Wai; Gautier, Aude; Woksepp, Hanna; Hellman, Ulf; Souchelnytskyi, Serhiy

    2009-01-01

    Acquiring high proliferation rate is crucial for carcinogenic transformation of cells. We report here proteome profiling of human breast epithelial cells with low (184A1) and high (MCF10A) proliferation rates. We identified 183 proteins in 184A1 and 318 proteins in MCF10A cells. These datasets provide the most comprehensive proteome annotations of 184A1 and MCF10A cells. Proteins were taken for identification from 2-D gels in a systematic and unbiased way. Functional clustering of the identified proteins showed similarities in distribution of proteins to the same functional domains, indicating similarities in proteomes of 184A1 and MCF10A cells. Among observed differences in protein expression, we validated correlation of expression of endogenous cyclin-dependent kinase 4 (CDK4), cyclin D3, cdc25B, and p38γ with cell proliferation. Furthermore, down-regulation of CDK4 and cyclin D3 with specific siRNA inhibited cell proliferation, which emphasized the role of CDK4 and cyclin D3 in enhancement of cell proliferation rate of human breast epithelial cells. Copyright © 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. Synergistic effect of Bcl-2 and cyclin A2 on adverse recurrence-free survival in stage I non-small cell lung cancer.

    Science.gov (United States)

    Ko, Eunkyung; Kim, Yujin; Cho, Eun Yoon; Han, Jungho; Shim, Young Mog; Park, Joobae; Kim, Duk-Hwan

    2013-03-01

    The prognostic significance of cyclin A2 overexpression in non-small cell lung cancer (NSCLC) is controversial. To understand the effect of cyclin A2 on recurrence in NSCLC, we retrospectively analyzed the expression of Bcl-2, cyclin A2, E-cadherin, Ki-67, and p53 using immunohistochemistry in 635 NSCLCs. Overexpression of cyclin A2 was found in 466 (73%) of 635 NSCLCs, and recurrence occurred in 291 (46%) of 635 NSCLCs with a median follow-up of 5.4 years. The relationship between recurrence and cyclin A2 overexpression was not homogenous by pathologic stage (Breslow-Day test for homogeneity, P = 0.007). Overexpression of cyclin A2 was associated with poor recurrence-free survival (RFS) in 374 stage I NSCLCs (P = 0.02), and RFS was worse in patient with negative expression of Bcl-2 than those with positive expression of Bcl-2. Cox proportional hazard analysis showed that stage I NSCLC patients with overexpression of cyclin A2 and negative expression of Bcl-2 had poorer RFS (hazard ratio = 3.86, 95% confidence interval = 1.07-15.77; P = 0.03) than those with normal expression of cyclin A2 and Bcl-2, after adjusting for age, adjuvant radiotherapy, and histology. Neural network and generalized linear model including cyclin A2 and Bcl-2 showed best performance in the prediction of recurrence; error rates for neural network and generalized linear model were 15% and 12%, respectively. Negative effect of cyclin A2 on RFS in stage I NSCLC was aggravated by negative expression of Bcl-2.

  16. Altered expression of the cell cycle regulatory protein cyclin D1 in the rat dentate gyrus after adrenalectomy-induced granular cell lass

    NARCIS (Netherlands)

    Postigo, JA; Van der Werf, YD; Korf, J; Krugers, HJ

    1998-01-01

    The loss of dentate gyrus (DG) granular cells after removal of the rat adrenal glands (ADX) is mediated by a process that is apoptotic in nature. The present study was initiated to compare changes in the immunocytochemical distribution of the cell-cycle regulatory protein cyclin D1, which has been

  17. A new class of cyclin dependent kinase in Chlamydomonas is required for coupling cell size to cell division

    Science.gov (United States)

    Li, Yubing; Liu, Dianyi; López-Paz, Cristina; Olson, Bradley JSC; Umen, James G

    2016-01-01

    Proliferating cells actively control their size by mechanisms that are poorly understood. The unicellular green alga Chlamydomonas reinhardtii divides by multiple fission, wherein a ‘counting’ mechanism couples mother cell-size to cell division number allowing production of uniform-sized daughters. We identified a sizer protein, CDKG1, that acts through the retinoblastoma (RB) tumor suppressor pathway as a D-cyclin-dependent RB kinase to regulate mitotic counting. Loss of CDKG1 leads to fewer mitotic divisions and large daughters, while mis-expression of CDKG1 causes supernumerous mitotic divisions and small daughters. The concentration of nuclear-localized CDKG1 in pre-mitotic cells is set by mother cell size, and its progressive dilution and degradation with each round of cell division may provide a link between mother cell-size and mitotic division number. Cell-size-dependent accumulation of limiting cell cycle regulators such as CDKG1 is a potentially general mechanism for size control. DOI: http://dx.doi.org/10.7554/eLife.10767.001 PMID:27015111

  18. Accelerated cell cycle progression in osteoblasts overexpressing the c-fos proto-oncogene: induction of cyclin A and enhanced CDK2 activity.

    Science.gov (United States)

    Sunters, Andrew; Thomas, David P; Yeudall, W Andrew; Grigoriadis, Agamemnon E

    2004-03-12

    Transgenic mice overexpressing the c-Fos oncoprotein develop osteosarcomas that are associated with deregulated expression of cell cycle genes. Here we have generated osteoblast cell lines expressing c-fos under the control of a tetracycline-regulatable promoter to investigate the role of c-Fos in osteoblast cell cycle control in vitro. Three stable subclones, AT9.2, AT9.3, and AT9.7, derived from MC3T3-E1 mouse osteoblasts, expressed high levels of exogenous c-fos mRNA and protein in the absence of tetracycline. Functional contribution of ectopic c-Fos to AP-1 complexes was confirmed by electromobility shift assays and transactivation of AP-1 reporter constructs. Induction of exogenous c-Fos in quiescent AT9.2 cells caused accelerated S-phase entry following serum stimulation, resulting in enhanced growth rate. Ectopic c-Fos resulted in increased expression of cyclins A and E protein levels, and premature activation of cyclin A-, cyclin E-, and cyclin-dependent kinase (CDK) 2-associated kinase activities, although cyclin D levels and CDK4 activity were not affected significantly in these cell lines. The enhanced CDK2 kinase activity was associated with a rapid, concomitant dissociation of p27 from CDK2-containing complexes. Deregulated cyclin A expression and CDK2 activity was also observed in primary mouse osteoblasts overexpressing c-Fos, but not in fibroblasts, and c-Fos transgenic tumor-derived osteosarcoma cells constitutively expressed high levels of cyclin A protein. These data suggest that overexpression of c-Fos in osteoblasts results in accelerated S phase entry as a result of deregulated cyclin A/E-CDK2 activity. This represents a novel role for c-Fos in osteoblast growth control and may provide c-Fos-overexpressing osteoblasts with a growth advantage during tumorigenesis.

  19. Curcumol induces cell cycle arrest in colon cancer cells via reactive oxygen species and Akt/ GSK3β/cyclin D1 pathway.

    Science.gov (United States)

    Wang, Juan; Li, Xu-Mei; Bai, Zhun; Chi, Bi-Xia; Wei, Yan; Chen, Xu

    2018-01-10

    Curcuma kwangsiensis S. G. Lee & C. F. Liang (Guangxi ezhu, in Chinese) belongs to the Zingiberaceae family, has been used as a traditionally Chinese medicine nearly 2000 year. Curcumol is one of the guaiane-type sesquiterpenoid hemiketal isolated from medicine plant Curcuma kwangsiensis S. G. Lee & C. F. Liang, which has been reported possesses anti-cancer effects. Our previous study found that the most contribution to inhibit nasopharyngeal carcinoma cell growth was curcumol. To assess the effect of curcumol on cell cycle arrest against human colon cancer cells (CRC) cells (LoVo and SW480) and explore its mechanism in vitro and in vivo. Curcumol was dissolved in absolute ethyl alcohol. The concentration of absolute ethyl alcohol in the control group or in experimental samples was always 1/500 (v/v) of the final medium volume. LoVo and SW480 cells were treated with different concentrations of curcumol (0, 53, 106, 212 and 424μM). And then the cell cycle of each group was examined by flow cytometry. The protein levels of PI3K, p-Akt, cyclin D1, cyclin E, CDK2, CDK4 and GSK3β were determined by Western blot. The mRNA expression of PI3K, Akt, cyclin D1, CDK4, P27, p21, and P16 in the treated cells were analyzed by real-time RT-PCR. In addition, the antitumor activity of curcumol was evaluated in nude mice bearing orthotopic tumor implants. Curcumol induced cell cycle arrest in G1/S phase. RT-qPCR and Western blot data showed that curcumol enhanced the expression of GSK3β, P27, p21 and P16, and decreased the levels of PI3K, phosphorylated Akt (p-Akt), cyclin D1, CDK4, cyclin E and CDK2. Furthermore, curcumol induced reactive oxygen species (ROS) generation in LoVo cells, and ROS scavenger N-acetylcysteine (NAC) significantly reversed curcumol-induced cell growth inhibition. Besides, curcumol also prevented the growth of human colon cancer cells xenografts in nude mouse, accompanied by the reduction of PI3K, Akt, cyclin D1, CDK4, cycln E and significant increase of

  20. The p-ERK–p-c-Jun–cyclinD1 pathway is involved in proliferation of smooth muscle cells after exposure to cigarette smoke extract

    Energy Technology Data Exchange (ETDEWEB)

    Li, Tianjia [Department of Vascular surgery, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, 5 Dong Dan San Tiao, Beijing 100005 (China); Song, Ting [Nursing Department of Orthopedics 3rd Ward, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, 5 Dong Dan San Tiao, Beijing 100005 (China); Ni, Leng; Yang, Genhuan; Song, Xitao; Wu, Lifei [Department of Vascular surgery, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, 5 Dong Dan San Tiao, Beijing 100005 (China); Liu, Bao, E-mail: liubao72@yahoo.com.cn [Department of Vascular surgery, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, 5 Dong Dan San Tiao, Beijing 100005 (China); Liu, Changwei, E-mail: liucw@vip.sina.com [Department of Vascular surgery, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, 5 Dong Dan San Tiao, Beijing 100005 (China)

    2014-10-24

    Highlights: • Smooth muscle cells proliferated after exposure to cigarette smoke extract. • The p-ERK, p-c-Jun, and cyclinD1 expressions increased in the process. • The p-ERK inhibitor, U0126, can reverse these effects. • The p-ERK → p-c-Jun → cyclinD1 pathway is involved in the process. - Abstract: An epidemiological survey has shown that smoking is closely related to atherosclerosis, in which excessive proliferation of vascular smooth muscle cells (SMCs) plays a key role. To investigate the mechanism underlying this unusual smoking-induced proliferation, cigarette smoke extract (CSE), prepared as smoke-bubbled phosphate-buffered saline (PBS), was used to induce effects mimicking those exerted by smoking on SMCs. As assessed by Cell Counting Kit-8 detection (an improved MTT assay), SMC viability increased significantly after exposure to CSE. Western blot analysis demonstrated that p-ERK, p-c-Jun, and cyclinD1 expression increased. When p-ERK was inhibited using U0126 (inhibitor of p-ERK), cell viability decreased and the expression of p-c-Jun and cyclinD1 was reduced accordingly, suggesting that p-ERK functions upstream of p-c-Jun and cyclinD1. When a c-Jun over-expression plasmid was transfected into SMCs, the level of cyclinD1 in these cells increased. Moreover, when c-Jun was knocked down by siRNA, cyclinD1 levels decreased. In conclusion, our findings indicate that the p-ERK–p-c-Jun–cyclinD1 pathway is involved in the excessive proliferation of SMCs exposed to CSE.

  1. The p-ERK–p-c-Jun–cyclinD1 pathway is involved in proliferation of smooth muscle cells after exposure to cigarette smoke extract

    International Nuclear Information System (INIS)

    Li, Tianjia; Song, Ting; Ni, Leng; Yang, Genhuan; Song, Xitao; Wu, Lifei; Liu, Bao; Liu, Changwei

    2014-01-01

    Highlights: • Smooth muscle cells proliferated after exposure to cigarette smoke extract. • The p-ERK, p-c-Jun, and cyclinD1 expressions increased in the process. • The p-ERK inhibitor, U0126, can reverse these effects. • The p-ERK → p-c-Jun → cyclinD1 pathway is involved in the process. - Abstract: An epidemiological survey has shown that smoking is closely related to atherosclerosis, in which excessive proliferation of vascular smooth muscle cells (SMCs) plays a key role. To investigate the mechanism underlying this unusual smoking-induced proliferation, cigarette smoke extract (CSE), prepared as smoke-bubbled phosphate-buffered saline (PBS), was used to induce effects mimicking those exerted by smoking on SMCs. As assessed by Cell Counting Kit-8 detection (an improved MTT assay), SMC viability increased significantly after exposure to CSE. Western blot analysis demonstrated that p-ERK, p-c-Jun, and cyclinD1 expression increased. When p-ERK was inhibited using U0126 (inhibitor of p-ERK), cell viability decreased and the expression of p-c-Jun and cyclinD1 was reduced accordingly, suggesting that p-ERK functions upstream of p-c-Jun and cyclinD1. When a c-Jun over-expression plasmid was transfected into SMCs, the level of cyclinD1 in these cells increased. Moreover, when c-Jun was knocked down by siRNA, cyclinD1 levels decreased. In conclusion, our findings indicate that the p-ERK–p-c-Jun–cyclinD1 pathway is involved in the excessive proliferation of SMCs exposed to CSE

  2. Transcriptional profiling of arbuscular mycorrhizal roots exposed to high levels of phosphate reveals the repression of cell cycle-related genes and secreted protein genes in Rhizophagus irregularis.

    Science.gov (United States)

    Sugimura, Yusaku; Saito, Katsuharu

    2017-02-01

    The development of arbuscular mycorrhiza (AM) is strongly suppressed under high-phosphate (Pi) conditions. To investigate AM fungal responses during the suppression of AM by high Pi, we performed an RNA-seq analysis of Rhizophagus irregularis colonizing Lotus japonicus roots at different levels of Pi (20, 100, 300, and 500 μM). AM fungal colonization decreased markedly under high-Pi conditions. In total, 163 fungal genes were differentially expressed among the four Pi treatments. Among these genes, a cell cycle-regulatory gene, cyclin-dependent kinase CDK1, and several DNA replication- and mitosis-related genes were repressed under high-Pi conditions. More than 20 genes encoding secreted proteins were also downregulated by high-Pi conditions, including the strigolactone-induced putative secreted protein 1 gene that enhances AM fungal colonization. In contrast, the expression of genes related to aerobic respiration and transport in R. irregularis were largely unaffected. Our data suggest that high Pi suppresses the expression of genes associated with fungal cell cycle progression or that encode secreted proteins that may be required for intercellular hyphal growth and arbuscule formation. However, high Pi has little effect on the transcriptional regulation of the primary metabolism or transport in preformed fungal structures.

  3. Endoglin inhibits ERK-induced c-Myc and cyclin D1 expression to impede endothelial cell proliferation

    Energy Technology Data Exchange (ETDEWEB)

    Pan, Christopher C.; Bloodworth, Jeffrey C. [Division of Pharmacology, Columbus, OH 43210 (United States); Mythreye, Karthikeyan [Duke University, Department of Medicine, Durham, NC 27708 (United States); Lee, Nam Y., E-mail: lee.5064@osu.edu [Division of Pharmacology, Columbus, OH 43210 (United States); Davis Heart and Lung Research Institute, Columbus, OH 43210 (United States)

    2012-08-03

    Highlights: Black-Right-Pointing-Pointer Endoglin inhibits ERK activation in endothelial cells. Black-Right-Pointing-Pointer Endoglin is a regulator of c-Myc and cyclin D1 expression. Black-Right-Pointing-Pointer {beta}-arrestin2 interaction with endoglin is required for ERK/c-Myc repression. Black-Right-Pointing-Pointer Endoglin impedes cellular proliferation by targeting ERK-induced mitogenic signaling. -- Abstract: Endoglin is an endothelial-specific transforming growth factor beta (TGF-{beta}) co-receptor essential for angiogenesis and vascular remodeling. Endoglin regulates a wide range of cellular processes, including cell adhesion, migration, and proliferation, through TGF-{beta} signaling to canonical Smad and Smad-independent pathways. Despite its overall pro-angiogenic role in the vasculature, the underlying mechanism of endoglin action is poorly characterized. We previously identified {beta}-arrestin2 as a binding partner that causes endoglin internalization from the plasma membrane and inhibits ERK signaling towards endothelial migration. In the present study, we examined the mechanistic role of endoglin and {beta}-arrestin2 in endothelial cell proliferation. We show that endoglin impedes cell growth through sustained inhibition of ERK-induced c-Myc and cyclin D1 expression in a TGF-{beta}-independent manner. The down-regulation of c-Myc and cyclin D1, along with growth-inhibition, are reversed when the endoglin/{beta}-arrestin2 interaction is disrupted. Given that TGF-{beta}-induced Smad signaling potently represses c-Myc in most cell types, our findings here show a novel mechanism by which endoglin augments growth-inhibition by targeting ERK and key downstream mitogenic substrates.

  4. Endoglin inhibits ERK-induced c-Myc and cyclin D1 expression to impede endothelial cell proliferation

    International Nuclear Information System (INIS)

    Pan, Christopher C.; Bloodworth, Jeffrey C.; Mythreye, Karthikeyan; Lee, Nam Y.

    2012-01-01

    Highlights: ► Endoglin inhibits ERK activation in endothelial cells. ► Endoglin is a regulator of c-Myc and cyclin D1 expression. ► β-arrestin2 interaction with endoglin is required for ERK/c-Myc repression. ► Endoglin impedes cellular proliferation by targeting ERK-induced mitogenic signaling. -- Abstract: Endoglin is an endothelial-specific transforming growth factor beta (TGF-β) co-receptor essential for angiogenesis and vascular remodeling. Endoglin regulates a wide range of cellular processes, including cell adhesion, migration, and proliferation, through TGF-β signaling to canonical Smad and Smad-independent pathways. Despite its overall pro-angiogenic role in the vasculature, the underlying mechanism of endoglin action is poorly characterized. We previously identified β-arrestin2 as a binding partner that causes endoglin internalization from the plasma membrane and inhibits ERK signaling towards endothelial migration. In the present study, we examined the mechanistic role of endoglin and β-arrestin2 in endothelial cell proliferation. We show that endoglin impedes cell growth through sustained inhibition of ERK-induced c-Myc and cyclin D1 expression in a TGF-β-independent manner. The down-regulation of c-Myc and cyclin D1, along with growth-inhibition, are reversed when the endoglin/β-arrestin2 interaction is disrupted. Given that TGF-β-induced Smad signaling potently represses c-Myc in most cell types, our findings here show a novel mechanism by which endoglin augments growth-inhibition by targeting ERK and key downstream mitogenic substrates.

  5. Rb and FZR1/Cdh1 determine CDK4/6-cyclin D requirement in C. elegans and human cancer cells.

    Science.gov (United States)

    The, Inge; Ruijtenberg, Suzan; Bouchet, Benjamin P; Cristobal, Alba; Prinsen, Martine B W; van Mourik, Tim; Koreth, John; Xu, Huihong; Heck, Albert J R; Akhmanova, Anna; Cuppen, Edwin; Boxem, Mike; Muñoz, Javier; van den Heuvel, Sander

    2015-01-06

    Cyclin-dependent kinases 4 and 6 (CDK4/6) in complex with D-type cyclins promote cell cycle entry. Most human cancers contain overactive CDK4/6-cyclin D, and CDK4/6-specific inhibitors are promising anti-cancer therapeutics. Here, we investigate the critical functions of CDK4/6-cyclin D kinases, starting from an unbiased screen in the nematode Caenorhabditis elegans. We found that simultaneous mutation of lin-35, a retinoblastoma (Rb)-related gene, and fzr-1, an orthologue to the APC/C co-activator Cdh1, completely eliminates the essential requirement of CDK4/6-cyclin D (CDK-4/CYD-1) in C. elegans. CDK-4/CYD-1 phosphorylates specific residues in the LIN-35 Rb spacer domain and FZR-1 amino terminus, resembling inactivating phosphorylations of the human proteins. In human breast cancer cells, simultaneous knockdown of Rb and FZR1 synergistically bypasses cell division arrest induced by the CDK4/6-specific inhibitor PD-0332991. Our data identify FZR1 as a candidate CDK4/6-cyclin D substrate and point to an APC/C(FZR1) activity as an important determinant in response to CDK4/6-inhibitors.

  6. Coffee Polyphenols Change the Expression of STAT5B and ATF-2 Modifying Cyclin D1 Levels in Cancer Cells

    Directory of Open Access Journals (Sweden)

    Carlota Oleaga

    2012-01-01

    Full Text Available Background. Epidemiological studies suggest that coffee consumption reduces the risk of cancer, but the molecular mechanisms of its chemopreventive effects remain unknown. Objective. To identify differentially expressed genes upon incubation of HT29 colon cancer cells with instant caffeinated coffee (ICC or caffeic acid (CA using whole-genome microarrays. Results. ICC incubation of HT29 cells caused the overexpression of 57 genes and the underexpression of 161, while CA incubation induced the overexpression of 12 genes and the underexpression of 32. Using Venn-Diagrams, we built a list of five overexpressed genes and twelve underexpressed genes in common between the two experimental conditions. This list was used to generate a biological association network in which STAT5B and ATF-2 appeared as highly interconnected nodes. STAT5B overexpression was confirmed at the mRNA and protein levels. For ATF-2, the changes in mRNA levels were confirmed for both ICC and CA, whereas the decrease in protein levels was only observed in CA-treated cells. The levels of cyclin D1, a target gene for both STAT5B and ATF-2, were downregulated by CA in colon cancer cells and by ICC and CA in breast cancer cells. Conclusions. Coffee polyphenols are able to affect cyclin D1 expression in cancer cells through the modulation of STAT5B and ATF-2.

  7. DACH1 regulates cell cycle progression of myeloid cells through the control of cyclin D, Cdk 4/6 and p21{sup Cip1}

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Jae-Woong; Kim, Hyeng-Soo; Kim, Seonggon; Hwang, Junmo; Kim, Young Hun; Lim, Ga Young [School of Life Science and Biotechnology, Kyungpook National University, Daegu 702-701 (Korea, Republic of); Sohn, Wern-Joo [Department of Biochemistry, School of Dentistry, IHBR, Kyungpook National University, Daegu 700-412 (Korea, Republic of); Yoon, Suk-Ran [Cell Therapy Research Center, Korea Research Institute of Bioscience and Biotechnology, Yuseong-gu, Daejeon 305-806 (Korea, Republic of); Kim, Jae-Young [Department of Biochemistry, School of Dentistry, IHBR, Kyungpook National University, Daegu 700-412 (Korea, Republic of); Park, Tae Sung [Department of Laboratory Medicine, Kyung Hee University School of Medicine, 1 Hoegi-dong, Dongdaemun-gu, Seoul 130-702 (Korea, Republic of); Park, Kwon Moo [Department of Anatomy, Kyungpook National University School of Medicine, Daegu 700-422 (Korea, Republic of); Ryoo, Zae Young [School of Life Science and Biotechnology, Kyungpook National University, Daegu 702-701 (Korea, Republic of); Lee, Sanggyu, E-mail: slee@knu.ac.kr [School of Life Science and Biotechnology, Kyungpook National University, Daegu 702-701 (Korea, Republic of)

    2012-03-30

    Highlights: Black-Right-Pointing-Pointer DACH1 increases cyclin D, F and Cdk 1, 4, 6 in mouse myeloid progenitor cells. Black-Right-Pointing-Pointer The knockdown of DACH1 blocked the cell cycle progression of HL-60 cells. Black-Right-Pointing-Pointer The novel effect of DACH1 related with cell cycle regulation and leukemogenesis. -- Abstract: The cell-fate determination factor Dachshund, a component of the Retinal Determination Gene Network (RDGN), has a role in breast tumor proliferation through the repression of cyclin D1 and several key regulators of embryonic stem cell function, such as Nanog and Sox2. However, little is known about the role of DACH1 in a myeloid lineage as a cell cycle regulator. Here, we identified the differential expression levels of extensive cell cycle regulators controlled by DACH1 in myeloid progenitor cells. The forced expression of DACH1 induced p27{sup Kip1} and repressed p21{sup Cip1}, which is a pivotal characteristic of the myeloid progenitor. Furthermore, DACH1 significantly increased the expression of cyclin D1, D3, F, and Cdk 1, 4, and 6 in myeloid progenitor cells. The knockdown of DACH1 blocked the cell cycle progression of HL-60 promyeloblastic cells through the decrease of cyclin D1, D3, F, and Cdk 1, 4, and 6 and increase in p21{sup Cip1}, which in turn decreased the phosphorylation of the Rb protein. The expression of Sox2, Oct4, and Klf4 was significantly up-regulated by the forced expression of DACH1 in mouse myeloid progenitor cells.

  8. CDK2 Phosphorylation on Threonine39 by AKT and Its Implication on Cyclin Binding, Cellular Localization, and Cell Cycle Progression

    National Research Council Canada - National Science Library

    Da Silva, Thiago

    2007-01-01

    .... Our hypothesis is that Cdk2 exists in two freely exchangeable conformations: that seen in the active, cyclin bound crystal and that of the inactive monomeric Cdk2, with the latter predominating in the absence of cyclin...

  9. Cell Cycle Regulatory Proteins p27(kip), Cyclins Dl and E and Proliferative Activity in Oncocytic (Hurthle Cell) Lesions of the Thyroid.

    Science.gov (United States)

    Maynes, Lincoln J.; Hutzler, Michael J.; Patwardhan, Nilima A.; Wang, Songtao; Khan, Ashraf

    2000-01-01

    Cyclins are prime cell-cycle regulators central to the control of cell proliferation in eukaryotic cells. The formation of cyclin/cyclin-dependent kinases (CDK) complexes activates the kinases and initiates a cascade of events, which directs cells through the cell cycle. CDK inhibitors (CDKIs) such as p27(kip1) inhibit cyclln-CDK complexes and function as negative regulators of the cell cycle. Previous studies have shown that p27(kip1) is decreased In malignant relative to benign thyroid tumors, but its role and Interaction with other cell cycle regulatory proteins have not been well established In oncocytic lesions of the thyroid. We studied the expression of p27(kip1), cyclins D1 and E, and Ki67 In 20 cases of oncocytic adenoma (AD). 6 cases of oncocytic carcinoma (CA). 8 cases of Hashimoto's thyroiditis (HT). and 9 cases of nodular goiter with oncocytic change (NG) by Immunohistochemlstry. In the latter two lesions only oncocytic cells were evaluated. The positive staining was stratified Into four groups. Statistical analysis was done using the Kruslcal-Wallis one-way analysis of variance test, and, when significant the Dunn multiple-comparisons procedure was used to determine pairwise differences. AllI 20 AD were p27(kip1) posItive, 10 were 4+, 2 were 3+, and the remaining 8 were 1+. In contrast all 6 CA showed 4+ p27(kip1) staining, of the 8 HT 2 were 4+, two 3+, three1+, and I was negative.All 9 NG were p27 positive, 7 showed 4+, one 3+, and one 1+ staining. On pairwise comparison differences in p27(kip1) staining between AD and CA and between HT and CA were statistically significant (p=0.0243 and p=0.0142, respectively). In all but one case Ki67 expression was either very low (<3%) or negative. No significant differences were seen in the expression of cyclin D1 or cyclin E among the groups observed. In conclusion, the increased p27(kip1) expression in malignant oncocytlc tumors relative to benign oncocytic lesions is unlike any other malignant progression

  10. Decreased MiR-17 in glioma cells increased cell viability and migration by increasing the expression of Cyclin D1, p-Akt and Akt.

    Directory of Open Access Journals (Sweden)

    Guangwei Sun

    Full Text Available The activating mutations of micro RNA (miR-17 have been revealed in tumors such as human non-Hodgkin's lymphoma and T cell leukemia. However, it is unclear about the role of miR-17 in glioma cells. The current study aimed to investigate effects of miR-17 mimics or inhibitor on the viability and migration of rat glioma C6 cells, and explore possible mechanisms.The expression of miR-17 in rat glioma C6 cells and normal brain tissue was detected by quantitative PCR. Protein expression of Cyclin D1 in rat glioma C6 cells and normal brain tissue was measured by Western Blot. Glioma C6 cells were transfected with MiR-17 mimics or inhibitor. Cells that were not transfected (Lipofectamine only and cells that were transfected with nonsense RNA negative control served as control. MTT assay was utilized to detect cell viability, and cell wound scratch assay was utilized to examine the migration index. In addition, protein expression of Cyclin D1, p-Akt and Akt in MiR-17 mimics or inhibitor-transfected glioma C6 cells was detected by Western Blot. This study had been approved by the Medical Ethics Committee of the First Affiliated Hospital of Soochow University. All applicable international, national, and/or institutional guidelines for the care and use of animals were followed.The expression of miR-17 was significantly lower, whereas the expression of Cyclin D1 was significantly higher in glioma C6 cells compared to normal brain tissue. MiR-17 mimics decreased the viability and migration of glioma C6 cells markedly at 48 h. In addition, MiR-17 inhibitor increased the viability and migration of glioma C6 cells at 24 and 48 h. The protein expression of Cyclin D1, p-Akt and Akt in glioma C6 cells decreased after transfection with miR-17 mimics for 72 h, and increased after transfection with miR-17 inhibitor for 72 h.The reduced miR-17 levels in glioma cells increased cell viability and migration, which correlates with increased expression of Cyclin D1, p

  11. Immortalization of Fetal Bovine Colon Epithelial Cells by Expression of Human Cyclin D1, Mutant Cyclin Dependent Kinase 4, and Telomerase Reverse Transcriptase: An In Vitro Model for Bacterial Infection.

    Directory of Open Access Journals (Sweden)

    Kengo Kuroda

    Full Text Available Cattle are the economically important animals in human society. They are essential for the production of livestock products such as milk and meats. The production efficiency of livestock products is negatively impacted by infection with zoonotic pathogens. To prevent and control infectious diseases, it is important to understand the interaction between cattle tissue and pathogenic bacteria. In this study, we established an in vitro infection model of an immortalized bovine colon-derived epithelial cell line by transducing the cells with lentiviral vectors containing genes encoding cell cycle regulators cyclin D1, mutant cyclin dependent kinase 4 (CDK4, and human telomerase reverse transcriptase (TERT. The established cell line showed continuous cell proliferation, expression of epithelial markers, and an intact karyotype, indicating that the cells maintained their original nature as colon-derived epithelium. Furthermore, we exposed the established cell line to two strains of Salmonella enterica and EHEC. Interestingly, S. Typhimurium showed higher affinity for the established cell line and invaded the cytoplasm than S. Enteritidis. Quantitative RT-PCR revealed that gene expression of Toll-like receptor 1 (TLR1, TLR 2 and TLR 3, whereas TLR 4, 5 and 6 were not detectable in established cells. Our established immortalized colon-derived epithelial cell should be a useful tool for studies evaluating the molecular mechanisms underlying bacterial infection.

  12. Immortalization of Fetal Bovine Colon Epithelial Cells by Expression of Human Cyclin D1, Mutant Cyclin Dependent Kinase 4, and Telomerase Reverse Transcriptase: An In Vitro Model for Bacterial Infection.

    Science.gov (United States)

    Kuroda, Kengo; Kiyono, Tohru; Isogai, Emiko; Masuda, Mizuki; Narita, Moe; Okuno, Katsuya; Koyanagi, Yukako; Fukuda, Tomokazu

    2015-01-01

    Cattle are the economically important animals in human society. They are essential for the production of livestock products such as milk and meats. The production efficiency of livestock products is negatively impacted by infection with zoonotic pathogens. To prevent and control infectious diseases, it is important to understand the interaction between cattle tissue and pathogenic bacteria. In this study, we established an in vitro infection model of an immortalized bovine colon-derived epithelial cell line by transducing the cells with lentiviral vectors containing genes encoding cell cycle regulators cyclin D1, mutant cyclin dependent kinase 4 (CDK4), and human telomerase reverse transcriptase (TERT). The established cell line showed continuous cell proliferation, expression of epithelial markers, and an intact karyotype, indicating that the cells maintained their original nature as colon-derived epithelium. Furthermore, we exposed the established cell line to two strains of Salmonella enterica and EHEC. Interestingly, S. Typhimurium showed higher affinity for the established cell line and invaded the cytoplasm than S. Enteritidis. Quantitative RT-PCR revealed that gene expression of Toll-like receptor 1 (TLR1), TLR 2 and TLR 3, whereas TLR 4, 5 and 6 were not detectable in established cells. Our established immortalized colon-derived epithelial cell should be a useful tool for studies evaluating the molecular mechanisms underlying bacterial infection.

  13. Purification, characterization, and kinetic mechanism of cyclin D1. CDK4, a major target for cell cycle regulation.

    Science.gov (United States)

    Konstantinidis, A K; Radhakrishnan, R; Gu, F; Rao, R N; Yeh, W K

    1998-10-09

    The cyclin D1.CDK4-pRb (retinoblastoma protein) pathway plays a central role in the cell cycle, and its deregulation is correlated with many types of cancers. As a major drug target, we purified dimeric cyclin D1.CDK4 complex to near-homogeneity by a four-step procedure from a recombinant baculovirus-infected insect culture. We optimized the kinase activity and stability and developed a reproducible assay. We examined several catalytic and kinetic properties of the complex and, via steady-state kinetics, derived a kinetic mechanism with a peptide (RbING) and subsequently investigated the mechanistic implications with a physiologically relevant protein (Rb21) as the phosphoacceptor. The complex bound ATP 130-fold tighter when Rb21 instead of RbING was used as the phosphoacceptor. By using staurosporine and ADP as inhibitors, the kinetic mechanism of the complex appeared to be a "single displacement or Bi-Bi" with Mg2+.ATP as the leading substrate and phosphorylated RbING as the last product released. In addition, we purified a cyclin D1-CDK4 fusion protein to homogeneity by a three-step protocol from another recombinant baculovirus culture and observed similar kinetic properties and mechanisms as those from the complex. We attempted to model staurosporine in the ATP-binding site of CDK4 according to our kinetic data. Our biochemical and modeling data provide validation of both the complex and fusion protein as highly active kinases and their usefulness in antiproliferative inhibitor discovery.

  14. Expression of cyclin D1 correlates with p27KIP1and regulates the degree of oral dysplasia and squamous cell carcinoma differentiation.

    Science.gov (United States)

    Guan, Guangzhao; Bakr, Mahmoud M; Firth, Norman; Love, Robert M

    2018-02-01

    The aim of this study was to identify an association or link between cyclin D1 and p27 KIP1 protein expression and dysplastic changes or progression. Oral mucosal biopsies with a diagnosis of non-neoplastic tissue (gingivitis) (n = 10), mild to moderate oral epithelial dysplasia (n = 12), and oral squamous cell carcinoma (n = 11) were evaluated by using immunohistochemistry. Scanning software was used to determine cyclin D1 and p27 KIP1 intensity of expression, location, and pattern. A significant increase in expression of cyclin D1 and a decrease in expression of p27 KIP1 proteins were identified in oral epithelial dysplasia and less differentiated oral squamous cell carcinoma (OSCC). There was a more diffuse distribution of cyclin D1 protein expression extending from the basal cell layer into the prickle cell layers in epithelial dysplasia and extending within all epithelial layers in OSCC. Cases of oral epithelial dysplasia had moderate infrequent expression of p27 KIP1 . There were no p27 KIP1 -positive cells in OSCC. The percentage of cells with both nuclear and cytoplasmic cyclin D1 staining was higher in OSCC specimens than control groups and oral epithelial dysplasia. The expression of both cyclin D1 and p27 KIP1 correlated with the grade of oral epithelial dysplasia and degree of OSCC differentiation. The results obtained will be verified through a basic follow-up of the cases to determine the prognosis/progression of oral dysplasia. Copyright © 2018 Elsevier Inc. All rights reserved.

  15. GM-CSF Induces Cyclin D1 Expression and Proliferation of Endothelial Progenitor Cells via PI3K and MAPK Signaling

    Directory of Open Access Journals (Sweden)

    Chaolin Qiu

    2014-03-01

    Full Text Available Background/Aims: Endothelial progenitor cells (EPCs, which can be isolated from the bone marrow or the peripheral blood, have generated interest because of their capacity to migrate to sites of vascularization and endothelialization and differentiate into endothelial cells in a process termed neovasculogenesis. EPCs are therefore possible regenerative tools for the treatment of vascular diseases and potential targets for the inhibition of angiogenesis during tumor development. Here, we investigated the mechanisms underlying the effect of granulocyte-macrophage colony-stimulating factor (GM-CSF on the acceleration of EPC proliferation and colony formation. Methods: EPCs were isolated, identified and cultured in the presence of GM-CSF. The effect of GM-CSF on endothelial cell colony formation and proliferation was examine by colony formation assay and MTT assay, separately. Cell cycle was analyzed by flow cytometry. The expression of cyclin D1 and cyclin E were detected by western bloting. JAK/Stat, PI3K/Akt and MAPK signaling were analyzed. Results: GM-CSF accelerated the G1/S phase transition in EPCs by upregulating the expression of cyclins D1 and E. The GM-CSF induced increase in the levels of cyclin D1 and the subsequent phosphorylation of the retinoblastoma (Rb protein activated E2F-1, resulting in the upregulation of the transcription of cyclin E. Furthermore, the induction of cyclin D1 expression and cell cycle progression by GM-CSF was mediated by the PI3K/Akt, JNK and ERK signaling pathways through the phosphorylation of GSK3β or the activation of AP-1 transcription factors. Conclusion: Our findings shed light on the mechanisms underlying the effect of GM-CSF on the modulation of cell cycle progression in EPCs, which is important considering their role in vascular repair and their therapeutic potential in several diseases.

  16. Expression of a TGF-{beta} regulated cyclin-dependent kinase inhibitor in normal and immortalized airway epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Tierney, L.A.; Bloomfield, C.; Johnson, N.F. [and others

    1995-12-01

    Tumors arising from epithelial cells, including lung cancers are frequently resistant to factors that regulate growth and differentiation in normal in normal cells. Once such factor is transforming growth factor-{Beta} (TGF-{Beta}). Escape from the growth-inhibitory effects of TGF-{Beta} is thought to be a key step in the transformation of airway epithelial cells. most lung cancer cell lines require serum for growth. In contrast, normal human bronchial epithelial (NHBE) cells are exquisitely sensitive to growth-inhibitory and differentiating effects of TGF-{Beta}. The recent identification of a novel cyclin-dependent kinase inhibitor, p15{sup INK4B}, which is regulated by TGF-{Beta}, suggests a mechanism by which TGF-{Beta} mediates growth arrest in NHBE cells. The purpose of this study was two-fold: (1) to determine if p15{sup INK4B} is induced by TGF-{Beta} in NHBE cells or immortalized bronchial epithelial (R.1) cells and if that induction corresponds to a G1/S cell-cycle arrest; (2) to determine the temporal relationship between p15{sup INK4B} induction, cell-cycle arrest, and the phosphorylation state of the pRB because it is thought that p15{sup INK4B} acts indirectly by preventing phosphorylation of the RB gene product. In this study, expression of p15{sup INK4B} was examined in NHBE cells and R.1 cells at different time intervals following TGF-{Beta} treatment. The expression of this kinase inhibitor and its relationship to the cell and the pRb phosphorylation state were examined in cells that were both sensitive (NHBE) and resistant (R.1) to the effects of TGF-{Beta}. These results suggest that the cyclin-dependent kinase inhibitor, p15{sup INK4B}, is involved in airway epithelial cell differentiation and that loss or reduction of expression plays a role in the resistance of transformed or neoplastic cells to the growth-inhibitory effects of TGF-{Beta}.

  17. p53 and cyclin G cooperate in mediating genome stability in somatic cells of Drosophila.

    Science.gov (United States)

    Bayer, Fabienne E; Zimmermann, Mirjam; Fischer, Patrick; Gromoll, Christian; Preiss, Anette; Nagel, Anja C

    2017-12-20

    One of the key players in genome surveillance is the tumour suppressor p53 mediating the adaptive response to a multitude of stress signals. Here we identify Cyclin G (CycG) as co-factor of p53-mediated genome stability. CycG has been shown before to be involved in double-strand break repair during meiosis. Moreover, it is also important for mediating DNA damage response in somatic tissue. Here we find it in protein complexes together with p53, and show that the two proteins interact physically in vitro and in vivo in response to ionizing irradiation. In contrast to mammals, Drosophila Cyclin G is no transcriptional target of p53. Genetic interaction data reveal that p53 activity during DNA damage response requires the presence of CycG. Morphological defects caused by overexpression of p53 are ameliorated in cycG null mutants. Moreover, using a p53 biosensor we show that p53 activity is impeded in cycG mutants. As both p53 and CycG are likewise required for DNA damage repair and longevity we propose that CycG plays a positive role in mediating p53 function in genome surveillance of Drosophila.

  18. Signal transducer and activator of transcription 5 activation is sufficient to drive transcriptional induction of cyclin D2 gene and proliferation of rat pancreatic beta-cells

    DEFF Research Database (Denmark)

    Friedrichsen, Birgitte N; Richter, Henrijette E; Hansen, Johnny A

    2003-01-01

    in a time-dependent manner by hGH in INS-1 cells. Inhibition of protein synthesis by coincubation with cycloheximide did not affect the hGH-induced increase of cyclin D2 mRNA levels at 4 h. Expression of a dominant negative STAT5 mutant, STAT5aDelta749, partially inhibited cyclin D2 protein levels. INS-1...... cells transiently transfected with a cyclin D2 promoter-reporter construct revealed a 3- to 5-fold increase of transcriptional activity in response to hGH stimulation. Furthermore, coexpression of a constitutive active STAT5 mutant (either CA-STAT5a or CA-STAT5b) was sufficient to drive transactivation...

  19. Cyclin T1-dependent genes in activated CD4 T and macrophage cell lines appear enriched in HIV-1 co-factors.

    Directory of Open Access Journals (Sweden)

    Wendong Yu

    2008-09-01

    Full Text Available HIV-1 is dependent upon cellular co-factors to mediate its replication cycle in CD4(+ T cells and macrophages, the two major cell types infected by the virus in vivo. One critical co-factor is Cyclin T1, a subunit of a general RNA polymerase II elongation factor known as P-TEFb. Cyclin T1 is targeted directly by the viral Tat protein to activate proviral transcription. Cyclin T1 is up-regulated when resting CD4(+ T cells are activated and during macrophage differentiation or activation, conditions that are also necessary for high levels of HIV-1 replication. Because Cyclin T1 is a subunit of a transcription factor, the up-regulation of Cyclin T1 in these cells results in the induction of cellular genes, some of which might be HIV-1 co-factors. Using shRNA depletions of Cyclin T1 and transcriptional profiling, we identified 54 cellular mRNAs that appear to be Cyclin T1-dependent for their induction in activated CD4(+ T Jurkat T cells and during differentiation and activation of MM6 cells, a human monocytic cell line. The promoters for these Cyclin T1-dependent genes (CTDGs are over-represented in two transcription factor binding sites, SREBP1 and ARP1. Notably, 10 of these CTDGs have been reported to be involved in HIV-1 replication, a significant over-representation of such genes when compared to randomly generated lists of 54 genes (p value<0.00021. The results of siRNA depletion and dominant-negative protein experiments with two CTDGs identified here, CDK11 and Casein kinase 1 gamma 1, suggest that these genes are involved either directly or indirectly in HIV-1 replication. It is likely that the 54 CTDGs identified here include novel HIV-1 co-factors. The presence of CTDGs in the protein space that was available for HIV-1 to sample during its evolution and acquisition of Tat function may provide an explanation for why CTDGs are enriched in viral co-factors.

  20. Protein expression of B-cell lymphoma gene 6 (BCL-6) in invasive breast cancer is associated with cyclin D1 and hypoxia-inducible factor-1α (HIF-1α)

    NARCIS (Netherlands)

    Bos, R.; Diest, P.J. van; Groep, P. van der; Greijer, A.E.; Hermsen, M.A.J.A.; Heijnen, I.; Meijer, G.A.; Baak, J.P.A.; Pinedo, H.M.; Wall, E. van der; Shvarts, A.

    2003-01-01

    B-cell lymphoma gene (BCL-6) upregulation contributes to immortalization of mouse embryo fibroblast and primary B cells via upregulation of cyclin D1. As cyclin D1 overexpression is a common phenomenon in different cancers, BCL-6 protein overexpression may not be restricted to lymphomas. In this

  1. Dienogest, a synthetic progestin, inhibits the proliferation of immortalized human endometrial epithelial cells with suppression of cyclin D1 gene expression.

    Science.gov (United States)

    Shimizu, Yutaka; Takeuchi, Takashi; Mita, Shizuka; Mizuguchi, Kiyoshi; Kiyono, Tohru; Inoue, Masaki; Kyo, Satoru

    2009-10-01

    Dienogest is a specific progesterone receptor agonist with potent oral endometrial activity and is used in the treatment of endometriosis. In this study, we examined the direct effects of dienogest on the proliferation of human endometrial epithelial cells using an immortalized cell line. 5-Bromo-2'-deoxyuridine incorporation into the cells was inhibited by dienogest and by progesterone (P(4)) in dose-dependent fashion at concentrations of 10(-8) mol/l or higher. To identify the target genes of dienogest and P(4), we screened the expression of 84 genes related to cell cycle regulation by real-time polymerase chain reaction after 6 h of treatment at a concentration of 10(-7) mol/l. Results showed that only cyclin D1 expression was significantly down-regulated, although expression of the other genes did not significantly change after dienogest or P(4) treatment compared with the control. In a time-course study during the first 24 h after drug treatment, dienogest and P(4) each produced a lasting decrease in the expression of cyclin D1 mRNA, followed by a decrease in cyclin E1 mRNA but not an increase in the expression of cell cycle inhibitor genes (p21, p27 and p53). These findings suggest that dienogest directly inhibits the proliferation of human endometrial epithelial cells with suppression of cyclin D1 gene expression.

  2. The Cyclin-Dependent Kinase Ortholog pUL97 of Human Cytomegalovirus Interacts with Cyclins

    Directory of Open Access Journals (Sweden)

    Laura Graf

    2013-12-01

    Full Text Available The human cytomegalovirus (HCMV-encoded protein kinase, pUL97, is considered a cyclin-dependent kinase (CDK ortholog, due to shared structural and functional characteristics. The primary mechanism of CDK activation is binding to corresponding cyclins, including cyclin T1, which is the usual regulatory cofactor of CDK9. This study provides evidence of direct interaction between pUL97 and cyclin T1 using yeast two-hybrid and co-immunoprecipitation analyses. Confocal immunofluorescence revealed partial colocalization of pUL97 with cyclin T1 in subnuclear compartments, most pronounced in viral replication centres. The distribution patterns of pUL97 and cyclin T1 were independent of HCMV strain and host cell type. The sequence domain of pUL97 responsible for the interaction with cyclin T1 was between amino acids 231–280. Additional co-immunoprecipitation analyses showed cyclin B1 and cyclin A as further pUL97 interaction partners. Investigation of the pUL97-cyclin T1 interaction in an ATP consumption assay strongly suggested phosphorylation of pUL97 by the CDK9/cyclin T1 complex in a substrate concentration-dependent manner. This is the first demonstration of interaction between a herpesviral CDK ortholog and cellular cyclins.

  3. Prevalence and clinical implications of cyclin D1 expression in diffuse large B-cell lymphoma (DLBCL) treated with immunochemotherapy

    DEFF Research Database (Denmark)

    Ok, Chi Young; Xu-Monette, Zijun Y; Tzankov, Alexandar

    2014-01-01

    oncogene E3 ubiquitin protein ligase (MDM2), MDM4, and tumor protein 53 (TP53) were rare or absent. Gene expression profiling did not reveal any striking differences with respect to cyclin D1 in DLBCL. CONCLUSIONS: Compared with patients who had cyclin D1-negative DLBCL, men were more commonly affected......1-positive according to immunohistochemistry were also assessed for rearrangements of the cyclin D1 gene (CCND1) using fluorescence in situ hybridization. Gene expression profiling was performed to compare patients who had DLBCL with and without cyclin D1 expression. RESULTS: In total, 30 patients...... (2.1%) who had DLBCL that expressed cyclin D1 and lacked CCND1 gene rearrangements were identified. Patients with cyclin D1-positive DLBCL had a median age of 57 years (range, 16.0-82.6 years). There were 23 males and 7 females. Twelve patients (40%) had bulky disease. None of them expressed CD5. Two...

  4. p27KIP1 blocks cyclin E-dependent transactivation of cyclin A gene expression

    DEFF Research Database (Denmark)

    Zerfass-Thome, K; Schulze, A; Zwerschke, W

    1997-01-01

    Cyclin E is necessary and rate limiting for the passage of mammalian cells through the G1 phase of the cell cycle. Control of cell cycle progression by cyclin E involves cdk2 kinase, which requires cyclin E for catalytic activity. Expression of cyclin E/cdk2 leads to an activation of cyclin A gene...... expression, as monitored by reporter gene constructs derived from the human cyclin A promoter. Promoter activation by cyclin E/cdk2 requires an E2F binding site in the cyclin A promoter. We show here that cyclin E/cdk2 kinase can directly bind to E2F/p107 complexes formed on the cyclin A promoter-derived E2F...... binding site, and this association is controlled by p27KIP1, most likely through direct protein-protein interaction. These observation suggest that cyclin E/cdk2 associates with E2F/p107 complexes in late G1 phase, once p27KIP1 has decreased below a critical threshold level. Since a kinase-negative mutant...

  5. Peroxisome proliferator-activated receptors (PPAR) agonists affect cell viability, apoptosis and expression of cell cycle related proteins in cell lines of glial brain tumors

    Czech Academy of Sciences Publication Activity Database

    Straková, N.; Ehrmann, J.; Bartoš, Jan; Malíková, J.; Doležel, Jaroslav; Kolář, Z.

    2005-01-01

    Roč. 52, - (2005), s. 126-136 ISSN 0028-2685 Institutional research plan: CEZ:AV0Z50380511; CEZ:AV0Z5038910 Keywords : PPAR * glioplasma * cell cycle Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 0.731, year: 2005

  6. Aryl hydrocarbon receptor regulates cell cycle progression in human breast cancer cells via a functional interaction with cyclin-dependent kinase 4.

    Science.gov (United States)

    Barhoover, Melissa A; Hall, Julie M; Greenlee, William F; Thomas, Russell S

    2010-02-01

    The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor with constitutive activities and those induced by xenobiotic ligands, such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). One unexplained cellular role for the AHR is its ability to promote cell cycle progression in the absence of exogenous ligands, whereas treatment with exogenous ligands induces cell cycle arrest. Within the cell cycle, progression from G(1) to S phase is controlled by sequential phosphorylation of the retinoblastoma protein (RB1) by cyclin D-cyclin-dependent kinase (CDK) 4/6 complexes. In this study, the functional interactions between the AHR, CDK4, and cyclin D1 (CCND1) were investigated as a potential mechanism for the cell cycle regulation by the AHR. Time course cell cycle and molecular experiments were performed in human breast cancer cells. The results demonstrated that the AHR and CDK4 interact within the cell cycle, and the interaction was disrupted upon TCDD treatment. The disruption was temporally correlated with G(1) cell cycle arrest and decreased phosphorylation of RB1. Biochemical reconstitution assays using in vitro-translated protein recapitulated the AHR and CDK4 interaction and showed that CCND1 was also part of the complex. In vitro assays for CDK4 kinase activity demonstrated that RB1 phosphorylation by the AHR/CDK4/CCND1 complex was reduced in the presence of TCDD. The results suggest that the AHR interacts in a complex with CDK4 and CCND1 in the absence of exogenous ligands to facilitate cell cycle progression. This interaction is disrupted by exogenous ligands, such as TCDD, to induce G(1) cell cycle arrest.

  7. Casearin D inhibits ERK phosphorylation and induces downregulation of cyclin D1 in HepG2 cells.

    Science.gov (United States)

    Ferreira-Silva, Guilherme Álvaro; Lages, Carla Carolina Lopes; Sartorelli, Patricia; Hasegawa, Flávia Rie; Soares, Marisi Gomes; Ionta, Marisa

    2017-02-01

    Cancer is a public health problem which represents the second cause of death in the world. In this framework, it is necessary to identify novel compounds with antineoplastic potential. Plants are an important source for discovering novel compounds with pharmacological potential. In this study, we aimed to investigate the antiproliferative potential of isolated compounds from Casearia sylvestris on tumor cell lines. Crude extract effectively reduced cell viability of 4 tumor cell lines (HepG2, A549, U251-MG, and HT-144) after 48h treatment. HepG2 and HT-144 were the most responsive cells. Three fractions (aqueous ethanol, n-hexane and ethyl acetate) were tested against HepG2 and HT-144 cells and we observed that compounds with antiproliferative activity were concentrated in n-hexane and ethyl acetate fractions. The casearins A, G and J were isolated from n-hexane fraction, while casearin D was obtained from ethyl acetate fraction. We demonstrated that casearin D significantly inhibited the clonogenic capacity of HepG2 cells after 24h exposure indicating its antiproliferative activity. In addition, G1/S transition cell cycle arrest in HepG2 cells was also observed. These effects are related, at least in part, to ability of the casearin D in reducing ERK phosphorylation and cyclin D1 expression levels. Copyright © 2016 Elsevier B.V. All rights reserved.

  8. Lineage specific composition of cyclin D-CDK4/CDK6-p27 complexes reveals distinct functions of CDK4, CDK6 and individual D-type cyclins in differentiating cells of embryonic origin

    Czech Academy of Sciences Publication Activity Database

    Bryja, Vítězslav; Pacherník, J.; Vondráček, Jan; Souček, Karel; Čajánek, L.; Horváth, Viktor; Holubcová, Z.; Dvořák, Petr; Hampl, Aleš

    2008-01-01

    Roč. 41, č. 6 (2008), s. 875-893 ISSN 0960-7722 R&D Projects: GA AV ČR(CZ) 1QS500040507; GA ČR(CZ) GA301/05/0463; GA ČR(CZ) GA204/07/0834; GA MŠk(CZ) 1M0538 Institutional research plan: CEZ:AV0Z50040507; CEZ:AV0Z50040702; CEZ:AV0Z50390512; CEZ:AV0Z50390703 Keywords : G1/S transition * cyclin D * embryonic cells differentiation Subject RIV: AQ - Safety, Health Protection, Human - Machine Impact factor: 2.423, year: 2008

  9. Baicalein induces G1 arrest in oral cancer cells by enhancing the degradation of cyclin D1 and activating AhR to decrease Rb phosphorylation

    Energy Technology Data Exchange (ETDEWEB)

    Cheng, Ya-Hsin, E-mail: yhcheng@mail.cmu.edu.tw [Department of Physiology, School of Medicine, China Medical University, Taichung 40402, Taiwan, ROC (China); Li, Lih-Ann; Lin, Pinpin; Cheng, Li-Chuan [Division of Environmental Health and Occupational Medicine, National Health Research Institutes, Zhunan, Miaoli 35053, Taiwan, ROC (China); Hung, Chein-Hui [Graduate Institute of Clinical Medicine Sciences, Chang Gung University, Puizi City, Chiayi 613, Taiwan, ROC (China); Chang, Nai Wen [Department of Biochemistry, School of Medicine, China Medical University, Taichung, Taiwan, ROC (China); Lin, Chingju [Department of Physiology, School of Medicine, China Medical University, Taichung 40402, Taiwan, ROC (China)

    2012-09-15

    Baicalein is a flavonoid, known to have anti-inflammatory and anti-cancer effects. As an aryl hydrocarbon receptor (AhR) ligand, baicalein at high concentrations blocks AhR-mediated dioxin toxicity. Because AhR had been reported to play a role in regulating the cell cycle, we suspected that the anti-cancer effect of baicalein is associated with AhR. This study investigated the molecular mechanism involved in the anti-cancer effect of baicalein in oral cancer cells HSC-3, including whether such effect would be AhR-mediated. Results revealed that baicalein inhibited cell proliferation and increased AhR activity in a dose-dependent manner. Cell cycle was arrested at the G1 phase and the expression of CDK4, cyclin D1, and phosphorylated retinoblastoma (pRb) was decreased. When the AhR was suppressed by siRNA, the reduction of pRb was partially reversed, accompanied by a decrease of cell population at G1 phase and an increase at S phase, while the reduction of cyclin D1 and CDK4 did not change. This finding suggests that the baicalein activation of AhR is indeed associated with the reduction of pRb, but is independent of the reduction of cyclin D1 and CDK4. When cells were pre-treated with LiCl, the inhibitor of GSK-3β, the decrease of cyclin D1 was blocked and the reduction of pRb was recovered. The data indicates that in HSC-3 the reduction of pRb is both mediated by baicalein through activation of AhR and facilitation of cyclin D1 degradation, which causes cell cycle arrest at the G1 phase, and results in the inhibition of cell proliferation. -- Highlights: ► Baicalein causes the G1 phase arrest by decreasing Rb phosphorylation. ► Baicalein modulates AhR-mediated cell proliferation. ► Both AhR activation and cyclin D1 degradation results in hypophosphorylation of Rb. ► Baicalein facilitates cyclin D1 degradation by signalling the GSK-3β pathway.

  10. Identification of an hexapeptide that binds to a surface pocket in cyclin A and inhibits the catalytic activity of the complex cyclin-dependent kinase 2-cyclin A.

    Science.gov (United States)

    Canela, Núria; Orzáez, Mar; Fucho, Raquel; Mateo, Francesca; Gutierrez, Ricardo; Pineda-Lucena, Antonio; Bachs, Oriol; Pérez-Payá, Enrique

    2006-11-24

    The protein-protein complexes formed between different cyclins and cyclin-dependent kinases (CDKs) are central to cell cycle regulation. These complexes represent interesting points of chemical intervention for the development of antineoplastic molecules. Here we describe the identification of an all d-amino acid hexapeptide, termed NBI1, that inhibits the kinase activity of the cyclin-dependent kinase 2 (cdk2)-cyclin A complex through selective binding to cyclin A. The mechanism of inhibition is non-competitive for ATP and non-competitive for protein substrates. In contrast to the existing CDKs peptide inhibitors, the hexapeptide NBI1 interferes with the formation of the cdk2-cyclin A complex. Furthermore, a cell-permeable derivative of NBI1 induces apoptosis and inhibits proliferation of tumor cell lines. Thus, the NBI1-binding site on cyclin A may represent a new target site for the selective inhibition of activity cdk2-cyclin A complex.

  11. Transcriptional and post-transcriptional down-regulation of cyclin D1 contributes to C6 glioma cell differentiation induced by forskolin.

    Science.gov (United States)

    He, Songmin; Zhu, Wenbo; Zhou, Yuxi; Huang, Yijun; Ou, Yanqiu; Li, Yan; Yan, Guangmei

    2011-09-01

    Malignant gliomas are the most common and lethal intracranial tumors, and differentiation therapy shows great potential to be a promising candidate for their treatment. Here, we have elaborated that a PKA activator, forskolin, represses cell growth via cell cycle arrest in the G0/G1 phase and induces cell differentiation characteristic with elongated processes and restoration of GFAP expression. In mechanisms, we verified that forskolin significantly diminishes the mRNA and protein level of a key cell cycle regulator cyclin D1, and maintenance of low cyclin D1 expression level was required for forskolin-induced proliferation inhibition and differentiation by gain and loss of function approaches. In addition, that forskolin down-regulated the cyclin D1 by proteolytic (post-transcriptional) mechanisms was dependent on GSK-3β activation at Ser9. The pro-differentiation activity of forskolin and related molecular mechanisms imply that forskolin can be developed into a candidate for the future in differentiation therapy of glioma, and cyclin D1 is a promising target for pro-differentiation strategy. Copyright © 2011 Wiley-Liss, Inc.

  12. Addition of rituximab to chemotherapy overcomes the negative prognostic impact of cyclin E expression in diffuse large B-cell lymphoma

    DEFF Research Database (Denmark)

    Frei, E; Visco, C; Xu-Monette, Z Y

    2013-01-01

    High levels of cyclin E (CCNE) are accompanied by shorter survival in cyclophosphamide, hydroxydaunorubicin, oncovin and prednisone (CHOP)-treated diffuse large B-cell lymphomas (DLBCL), independent of the international prognostic index (IPI). Data on the prognostic role of CCNE in the 'rituximab...

  13. P276-00, a cyclin-dependent kinase inhibitor, modulates cell cycle and induces apoptosis in vitro and in vivo in mantle cell lymphoma cell lines

    Directory of Open Access Journals (Sweden)

    Shirsath Nitesh P

    2012-10-01

    Full Text Available Abstract Background Mantle cell lymphoma (MCL is a well-defined aggressive lymphoid neoplasm characterized by proliferation of mature B-lymphocytes that have a remarkable tendency to disseminate. This tumor is considered as one of the most aggressive lymphoid neoplasms with poor responses to conventional chemotherapy and relatively short survival. Since cyclin D1 and cell cycle control appears as a natural target, small-molecule inhibitors of cyclin-dependent kinases (Cdks and cyclins may play important role in the therapy of this disorder. We explored P276-00, a novel selective potent Cdk4-D1, Cdk1-B and Cdk9-T1 inhibitor discovered by us against MCL and elucidated its potential mechanism of action. Methods The cytotoxic effect of P276-00 in three human MCL cell lines was evaluated in vitro. The effect of P276-00 on the regulation of cell cycle, apoptosis and transcription was assessed, which are implied in the pathogenesis of MCL. Flow cytometry, western blot, immunoflourescence and siRNA studies were performed. The in vivo efficacy and effect on survival of P276-00 was evaluated in a Jeko-1 xenograft model developed in SCID mice. PK/PD analysis of tumors were performed using LC-MS and western blot analysis. Results P276-00 showed a potent cytotoxic effect against MCL cell lines. Mechanistic studies confirmed down regulation of cell cycle regulatory proteins with apoptosis. P276-00 causes time and dose dependent increase in the sub G1 population as early as from 24 h. Reverse transcription PCR studies provide evidence that P276-00 treatment down regulated transcription of antiapoptotic protein Mcl-1 which is a potential pathogenic protein for MCL. Most importantly, in vivo studies have revealed significant efficacy as a single agent with increased survival period compared to vehicle treated. Further, preliminary combination studies of P276-00 with doxorubicin and bortezomib showed in vitro synergism. Conclusion Our studies thus provide

  14. Insulin Promotes the Proliferation of Human Umbilical Cord Matrix-Derived Mesenchymal Stem Cells by Activating the Akt-Cyclin D1 Axis

    Directory of Open Access Journals (Sweden)

    Peng Li

    2017-01-01

    Full Text Available Background. The functions of insulin in mesenchymal stem cells (MSC remain poorly understood. Methods. MSC from human umbilical cord matrix (UCM cultured in serum-free media (SFM with or without insulin were subjected to various molecular biological analyses to determine their proliferation and growth states, expression levels of Akt-cyclin D1 signaling molecules, and in vitro differentiation capacities. Results. Insulin accelerated the G1-S cell cycle progression of UCM-MSC and significantly stimulated their proliferation and growth in SFM. The pro-proliferative action of insulin was associated with augmented cyclin D1 and phosphorylated Akt expression levels. Akt inactivation remarkably abrogated insulin-induced increases in cyclin D1 expression and cell proliferation, indicating that insulin enhances the proliferation of UCM-MSC via acceleration of the G1-S transition mediated by the Akt-cyclin D1 pathway. Additionally, the UCM-MSC propagated in SFM supplemented with insulin exhibited similar specific surface antigen profiles and differentiation capacities as those generated in conventional media containing fetal bovine serum. Conclusions. These findings suggest that insulin acts solely to promote UCM-MSC proliferation without affecting their immunophenotype and differentiation potentials and thus have important implications for utilizing insulin to expand clinical-grade MSC in vitro.

  15. Localization of two mammalian cyclin dependent kinases during mammalian meiosis

    NARCIS (Netherlands)

    Ashley, T.; Walpita, D.; de rooij, D. G.

    2001-01-01

    Mammalian meiotic progression, like mitotic cell cycle progression, is regulated by cyclins and cyclin dependent kinases (CDKs). However, the unique requirements of meiosis (homologous synapsis, reciprocal recombination and the dual divisions that segregate first homologues, then sister chromatids)

  16. Interferon regulatory factor-1 together with reactive oxygen species promotes the acceleration of cell cycle progression by up-regulating the cyclin E and CDK2 genes during high glucose-induced proliferation of vascular smooth muscle cells.

    Science.gov (United States)

    Zhang, Xi; Liu, Long; Chen, Chao; Chi, Ya-Li; Yang, Xiang-Qun; Xu, Yan; Li, Xiao-Tong; Guo, Shi-Lei; Xiong, Shao-Hu; Shen, Man-Ru; Sun, Yu; Zhang, Chuan-Sen; Hu, Kai-Meng

    2013-10-14

    The high glucose-induced proliferation of vascular smooth muscle cells (VSMCs) plays an important role in the development of diabetic vascular diseases. In a previous study, we confirmed that Interferon regulatory factor-1 (Irf-1) is a positive regulator of the high glucose-induced proliferation of VSMCs. However, the mechanisms remain to be determined. The levels of cyclin/CDK expression in two cell models involving Irf-1 knockdown and overexpression were quantified to explore the relationship between Irf-1 and its downstream effectors under normal or high glucose conditions. Subsequently, cells were treated with high glucose/NAC, normal glucose/H₂O₂, high glucose/U0126 or normal glucose/H₂O₂/U0126 during an incubation period. Then proliferation, cyclin/CDK expression and cell cycle distribution assays were performed to determine whether ROS/Erk1/2 signaling pathway was involved in the Irf-1-induced regulation of VSMC growth under high glucose conditions. We found that Irf-1 overexpression led to down-regulation of cyclin D1/CDK4 and inhibited cell cycle progression in VSMCs under normal glucose conditions. In high glucose conditions, Irf-1 overexpression led to an up-regulation of cyclin E/CDK2 and an acceleration of cell cycle progression, whereas silencing of Irf-1 suppressed the expression of both proteins and inhibited the cell cycle during the high glucose-induced proliferation of VSMCs. Treatment of VSMCs with antioxidants prevented the Irf-1 overexpression-induced proliferation of VSMCs, the up-regulation of cyclin E/CDK2 and the acceleration of cell cycle progression in high glucose conditions. In contrast, under normal glucose conditions, H₂O₂ stimulation and Irf-1 overexpression induced cell proliferation, up-regulated cyclin E/CDK2 expression and promoted cell cycle acceleration. In addition, overexpression of Irf-1 promoted the activation of Erk1/2 and when VSMCs overexpressing Irf-1 were treated with U0126, the specific Erk1/2 inhibitor

  17. Multi-tasking Sulf1/Sulf2 enzymes do not only facilitate extracellular cell signalling but also participate in cell cycle related nuclear events.

    Science.gov (United States)

    Krishnakumar, Kavithanjali; Chakravorty, Ishani; Foy, Wendy; Allen, Steve; Justo, Tiago; Mukherjee, Abir; Dhoot, Gurtej K

    2018-03-01

    This study demonstrates highly dynamic spatial and temporal pattern of SULF1/SULF2 expression in a number of neuronal cell types growing in normal culture medium that included their transient nuclear mobilisation. Their nuclear translocation became particularly apparent during cell proliferation as both SULF1/SULF2 demonstrated not only cell membrane associated expression, their known site of function but also transient nuclear mobilisation during nuclear cell division. Nuclear localisation was apparent not only by immunocytochemical staining but also confirmed by immunoblotting staining of isolated nuclear fractions of C6, U87 and N2A cells. Immunocytochemical analysis demonstrated rapid nuclear exit of both SULF1/SULF2 following cell division that was slightly delayed but not blocked in a fraction of the polyploid cells observed in C6 cells. The overexpression of both Sulf1 and Sulf2 genes in C6 and U87 cells markedly promoted in vitro growth of these cells accompanied by nuclear mobilisation while inhibition of both these genes inhibited cell proliferation with little or no nuclear SULF1/SULF2 mobilisation. SULF1/SULF2 activity in these cells thus demonstrated a clear co-ordination of extracellular cell signalling with nuclear events related to cell proliferation. Crown Copyright © 2018. Published by Elsevier Inc. All rights reserved.

  18. CG0009, a novel glycogen synthase kinase 3 inhibitor, induces cell death through cyclin D1 depletion in breast cancer cells.

    Directory of Open Access Journals (Sweden)

    Hyun Mi Kim

    Full Text Available Glycogen synthase kinase 3α/β (GSK3α/β is a constitutively active serine/threonine kinase involved in multiple physiological processes, such as protein synthesis, stem cell maintenance and apoptosis, and acts as a key suppressor of the Wnt-β-catenin pathway. In the present study, we examined the therapeutic potential of a novel GSK3 inhibitor, CG0009, in the breast cancer cell lines, BT549, HS578T, MDA-MB-231, NCI/ADR-RES, T47D, MCF7 and MDA-MB-435, from the NCI-60 cancer cell line panel. Assessment of cytotoxicity, apoptosis and changes in estrogen-signaling proteins was performed using cell viability assays, Western blotting and quantitative real-time PCR. CG0009 enhanced the inactivating phosphorylation of GSK3α at Ser21 and GSK3β at Ser9 and simultaneously decreased activating phosphorylation of GSK3β at Tyr216, and induced caspase-dependent apoptosis independently of estrogen receptor α (ERα expression status, which was not observed with the other GSK3 inhibitors examined, including SB216763, kenpaullone and LiCl. CG0009 treatment (1 µmol/L completely ablated cyclin D1 expression in a time-dependent manner in all the cell lines examined, except T47D. CG0009 alone significantly activated p53, leading to relocation of p53 and Bax to the mitochondria. GSK3 inhibition by CG0009 led to slight upregulation of the β-catenin target genes, c-Jun and c-Myc, but not cyclin D1, indicating that CG0009-mediated cyclin D1 depletion overwhelms the pro-survival signal of β-catenin, resulting in cell death. Our findings suggest that the novel GSK3 inhibitor, CG0009, inhibits breast cancer cell growth through cyclin D1 depletion and p53 activation, and may thus offer an innovative therapeutic approach for breast cancers resistant to hormone-based therapy.

  19. Sulforaphane, a Dietary Isothiocyanate, Induces G2/M Arrest in Cervical Cancer Cells through CyclinB1 Downregulation and GADD45β/CDC2 Association

    Directory of Open Access Journals (Sweden)

    Ya-Min Cheng

    2016-09-01

    Full Text Available Globally, cervical cancer is the most common malignancy affecting women. The main treatment methods for this type of cancer include conization or hysterectomy procedures. Sulforaphane (SFN is a natural, compound-based drug derived from dietary isothiocyanates which has previously been shown to possess potent anti-tumor and chemopreventive effects against several types of cancer. The present study investigated the effects of SFN on anti-proliferation and G2/M phase cell cycle arrest in cervical cancer cell lines (Cx, CxWJ, and HeLa. We found that cytotoxicity is associated with an accumulation of cells in the G2/M phases of the cell-cycle. Treatment with SFN led to cell cycle arrest as well as the down-regulation of Cyclin B1 expression, but not of CDC2 expression. In addition, the effects of GADD45β gene activation in cell cycle arrest increase proportionally with the dose of SFN; however, mitotic delay and the inhibition of proliferation both depend on the dosage of SFN used to treat cancer cells. These results indicate that SFN may delay the development of cancer by arresting cell growth in the G2/M phase via down-regulation of Cyclin B1 gene expression, dissociation of the cyclin B1/CDC2 complex, and up-regulation of GADD45β proteins.

  20. Rsf-1 is overexpressed in non-small cell lung cancers and regulates cyclinD1 expression and ERK activity

    International Nuclear Information System (INIS)

    Li, Qingchang; Dong, Qianze; Wang, Enhua

    2012-01-01

    Highlights: ► Rsf-1 expression is elevated in non-small cell lung cancers. ► Rsf-1 depletion inhibits proliferation and increased apoptosis in lung cancer cells. ► Rsf-1 depletion decreases the level of cyclinD1 and phosphor-ERK expression. -- Abstract: Rsf-1 (HBXAP) was recently reported to be overexpressed in various cancers and associated with the malignant behavior of cancer cells. However, the expression of Rsf-1 in primary lung cancer and its biological roles in non-small cell lung cancer (NSCLC) have not been reported. The molecular mechanism of Rsf-1 in cancer aggressiveness remains ambiguous. In the present study, we analyzed the expression pattern of Rsf-1 in NSCLC tissues and found that Rsf-1 was overexpressed at both the mRNA and protein levels. There was a significant association between Rsf-1 overexpression and TNM stage (p = 0.0220) and poor differentiation (p = 0.0013). Furthermore, knockdown of Rsf-1 expression in H1299 and H460 cells with high endogenous Rsf-1 expression resulted in a decrease of colony formation ability and inhibition of cell cycle progression. Rsf-1 knockdown also induced apoptosis in these cell lines. Further analysis showed that Rsf-1 knockdown decreased cyclin D1 expression and phospho-ERK levels. In conclusion, Rsf-1 is overexpressed in NSCLC and contributes to malignant cell growth by cyclin D1 and ERK modulation, which makes Rsf-1 a candidate therapeutic target in lung cancer.

  1. BAFF induces spleen CD4{sup +} T cell proliferation by down-regulating phosphorylation of FOXO3A and activates cyclin D2 and D3 expression

    Energy Technology Data Exchange (ETDEWEB)

    Ji, Fang; Chen, Rongjing [Department of Orthodontics, Ninth People' s Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai Key Laboratory of Stomatology, Shanghai (China); Liu, Baojun [Laboratory of Lung, Inflammation and Cancers, Huashan Hospital, Fudan University, Shanghai (China); Zhang, Xiaoping [Department of Nuclear Medicine, Shanghai 10th People' s Hospital, Tongji University School of Medicine, Shanghai 200072 (China); Han, Junli; Wang, Haining [Department of General Dentistry, Ninth People' s Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai Key Laboratory of Stomatology, Shanghai (China); Shen, Gang [Department of Orthodontics, Ninth People' s Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai Key Laboratory of Stomatology, Shanghai (China); Tao, Jiang, E-mail: taojiang2012@yahoo.cn [Department of General Dentistry, Ninth People' s Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai Key Laboratory of Stomatology, Shanghai (China)

    2012-09-07

    Highlights: Black-Right-Pointing-Pointer Firstly analyze the mechanism of BAFF and anti-CD3 co-stimulation on purified mouse splenic CD4{sup +} T cells. Black-Right-Pointing-Pointer Carrying out siRNA technology to study FOXO3A protein function. Black-Right-Pointing-Pointer Helpful to understand the T cell especially CD4{sup +} T cell's role in immunological reaction. -- Abstract: The TNF ligand family member 'B cell-activating factor belonging to the TNF family' (BAFF, also called BLyS, TALL-1, zTNF-4, and THANK) is an important survival factor for B and T cells. In this study, we show that BAFF is able to induce CD4{sup +} spleen T cell proliferation when co-stimulated with anti-CD3. Expression of phosphorylated FOXO3A was notably down-regulated and cyclins D2 and D3 were up-regulated and higher in the CD4{sup +} T cells when treated with BAFF and anti-CD3, as assessed by Western blotting. Furthermore, after FOXO3A was knocked down, expression of cyclin D1 was unchanged, compared with control group levels, but the expression of cyclins D2 and D3 increased, compared with the control group. In conclusion, our results suggest that BAFF induced CD4{sup +} spleen T cell proliferation by down-regulating the phosphorylation of FOXO3A and then activating cyclin D2 and D3 expression, leading to CD4{sup +} T cell proliferation.

  2. Accelerated turnover of taste bud cells in mice deficient for the cyclin-dependent kinase inhibitor p27Kip1

    Directory of Open Access Journals (Sweden)

    Perna Marla K

    2011-04-01

    Full Text Available Abstract Background Mammalian taste buds contain several specialized cell types that coordinately respond to tastants and communicate with sensory nerves. While it has long been appreciated that these cells undergo continual turnover, little is known concerning how adequate numbers of cells are generated and maintained. The cyclin-dependent kinase inhibitor p27Kip1 has been shown to influence cell number in several developing tissues, by coordinating cell cycle exit during cell differentiation. Here, we investigated its involvement in the control of taste cell replacement by examining adult mice with targeted ablation of the p27Kip1 gene. Results Histological and morphometric analyses of fungiform and circumvallate taste buds reveal no structural differences between wild-type and p27Kip1-null mice. However, when examined in functional assays, mutants show substantial proliferative changes. In BrdU incorporation experiments, more S-phase-labeled precursors appear within circumvallate taste buds at 1 day post-injection, the earliest time point examined. After 1 week, twice as many labeled intragemmal cells are present, but numbers return to wild-type levels by 2 weeks. Mutant taste buds also contain more TUNEL-labeled cells and 50% more apoptotic bodies than wild-type controls. In normal mice, p27 Kip1 is evident in a subset of receptor and presynaptic taste cells beginning about 3 days post-injection, correlating with the onset of taste cell maturation. Loss of gene function, however, does not alter the proportions of distinct immunohistochemically-identified cell types. Conclusions p27Kip1 participates in taste cell replacement by regulating the number of precursor cells available for entry into taste buds. This is consistent with a role for the protein in timing cell cycle withdrawal in progenitor cells. The equivalence of mutant and wild-type taste buds with regard to cell number, cell types and general structure contrasts with the hyperplasia

  3. Effects of prostratin on Cyclin T1/P-TEFb function and the gene expression profile in primary resting CD4+ T cells

    Directory of Open Access Journals (Sweden)

    Rice Andrew P

    2006-10-01

    Full Text Available Abstract Background The latent reservoir of human immunodeficiency virus type 1 (HIV-1 in resting CD4+ T cells is a major obstacle to the clearance of infection by highly active antiretroviral therapy (HAART. Recent studies have focused on searches for adjuvant therapies to activate this reservoir under conditions of HAART. Prostratin, a non tumor-promoting phorbol ester, is a candidate for such a strategy. Prostratin has been shown to reactivate latent HIV-1 and Tat-mediated transactivation may play an important role in this process. We examined resting CD4+ T cells from healthy donors to determine if prostratin induces Cyclin T1/P-TEFb, a cellular kinase composed of Cyclin T1 and Cyclin-dependent kinase-9 (CDK9 that mediates Tat function. We also examined effects of prostratin on Cyclin T2a, an alternative regulatory subunit for CDK9, and 7SK snRNA and the HEXIM1 protein, two factors that associate with P-TEFb and repress its kinase activity. Results Prostratin up-regulated Cyclin T1 protein expression, modestly induced CDK9 protein expression, and did not affect Cyclin T2a protein expression. Although the kinase activity of CDK9 in vitro was up-regulated by prostratin, we observed a large increase in the association of 7SK snRNA and the HEXIM1 protein with CDK9. Using HIV-1 reporter viruses with and without a functional Tat protein, we found that prostratin stimulation of HIV-1 gene expression appears to require a functional Tat protein. Microarray analyses were performed and several genes related to HIV biology, including APOBEC3B, DEFA1, and S100 calcium-binding protein genes, were found to be regulated by prostratin. Conclusion Prostratin induces Cyclin T1 expression and P-TEFb function and this is likely to be involved in prostratin reactivation of latent HIV-1 proviruses. The large increase in association of 7SK and HEXIM1 with P-TEFb following prostratin treatment may reflect a requirement in CD4+ T cells for a precise balance between

  4. Cinacalcet HCl suppresses Cyclin D1 oncogene-derived parathyroid cell proliferation in a murine model for primary hyperparathyroidism.

    Science.gov (United States)

    Imanishi, Yasuo; Kawata, Takehisa; Kenko, Takao; Wada, Michihito; Nagano, Nobuo; Miki, Takami; Arnold, Andrew; Inaba, Masaaki

    2011-07-01

    Cinacalcet HCl (cinacalcet) is a calcimimetic compound, which suppresses parathyroid (PTH) hormone secretion from parathyroid glands in both primary hyperparathyroidism (PHPT) and secondary hyperparathyroidism (SHPT). We previously reported the suppressive effect of cinacalcet on PTH secretion in vivo in a PHPT model mouse, in which parathyroid-targeted overexpression of the cyclin D1 oncogene caused chronic biochemical hyperparathyroidism and parathyroid cell hyperplasia. Although cinacalcet suppressed parathyroid cell proliferation in SHPT in 5/6-nephrectomized uremic rats, its effect on PHPT has not yet been determined. In this study, the effect of cinacalcet on parathyroid cell proliferation was analyzed in PHPT mice. Cinacalcet (1 mg/g) was mixed into the rodent diet and orally administrated to 80-week-old PHPT mice for 10 days before death. 5-Bromo-2'-deoxyuridine (BrdU, 6 mg/day) was infused by an osmotic pump for 5 days before death, followed by immunostaining of the thyroid-parathyroid complex using an anti-BrdU antibody to estimate parathyroid cell proliferation. Compared to untreated PHPT mice, cinacalcet significantly suppressed both serum calcium and PTH. The proportion of BrdU-positive cells to the total cell number in the parathyroid glands increased considerably in untreated PHPT mice (9.5 ± 3.1%) compared to wild-type mice (0.7 ± 0.1%) and was significantly suppressed by cinacalcet (1.2 ± 0.2%). Cinacalcet did not affect apoptosis in the parathyroid cells of PHPT mice. These data suggest that cinacalcet suppressed both serum PTH levels and parathyroid cell proliferation in vivo in PHPT.

  5. [miR-497 suppresses proliferation of human cervical carcinoma HeLa cells by targeting cyclin E1].

    Science.gov (United States)

    Han, Jiming; Huo, Manpeng; Mu, Mingtao; Liu, Junjun; Zhang, Jing

    2014-06-01

    To evaluate the effect of miR-497 on proliferation of human cervical carcinoma HeLa cells and target relationship between miR-497 and cyclin E1 (CCNE1). Pre-miR-497 sequences were synthesized and cloned into pcDNATM6.2-GW to construct recombinant plasmid pcDNATM6.2-GW-pre-miR-497 and identified by real-time quantitative PCR (qRT-PCR). In addition, sequences of the wild-type CCNE1 (WT-CCNE1) and mutant CCNE1 (MT-CCNE1) were respectively cloned into pmirGLO vectors. MTT assay was used to explore the impact of miR-497 on the proliferation of HeLa cells. Furthermore, the target effect of miR-497 on the CCNE1 was identified by dual-luciferase reporter assay system, qRT-PCR and Western blotting. The recombinant plasmids pcDNATM6.2-GW-pre-miR-497 and pmirGLO-WT-CCNE1, pmirGLO-MT-CCNE1 were successfully constructed, and the miR-497 expression level in HeLa cells transfected with pre-miR-497 was significantly higher than that in the neg-miR group (PHeLa cells (PHeLa cells with pre-miR-497 transfection (PHeLa cells transfected with pre-miR-497 (PHeLa cells could suppress cell proliferation by targeting CCNE1.

  6. Cyclin D1 expression in prostate carcinoma

    International Nuclear Information System (INIS)

    Pereira, R.A.; Ravinal, R.C.; Costa, R.S.; Lima, M.S.; Tucci, S.; Muglia, V.F.; Reis, R.B. Dos; Silva, G.E.B.

    2014-01-01

    The purpose of this study was to investigate the relationship between cyclin D1 expression and clinicopathological parameters in patients with prostate carcinoma. We assessed cyclin D1 expression by conventional immunohistochemistry in 85 patients who underwent radical prostatectomy for prostate carcinoma and 10 normal prostate tissue samples retrieved from autopsies. We measured nuclear immunostaining in the entire tumor area and based the results on the percentage of positive tumor cells. The preoperative prostate-specific antigen (PSA) level was 8.68±5.16 ng/mL (mean±SD). Cyclin D1 staining was positive (cyclin D1 expression in >5% of tumor cells) in 64 cases (75.4%) and negative (cyclin D1 expression in ≤5% of tumor cells) in 21 cases (including 15 cases with no immunostaining). Normal prostate tissues were negative for cyclin D1. Among patients with a high-grade Gleason score (≥7), 86% of patients demonstrated cyclin D1 immunostaining of >5% (P<0.05). In the crude analysis of cyclin D1 expression, the high-grade Gleason score group showed a mean expression of 39.6%, compared to 26.9% in the low-grade Gleason score group (P<0.05). Perineural invasion tended to be associated with cyclin D1 expression (P=0.07), whereas cyclin D1 expression was not associated with PSA levels or other parameters. Our results suggest that high cyclin D1 expression could be a potential marker for tumor aggressiveness

  7. Cyclin D1 expression in prostate carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Pereira, R.A.; Ravinal, R.C.; Costa, R.S.; Lima, M.S. [Universidade de São Paulo, Faculdade de Medicina de Ribeirão Preto, Departamento de Patologia, Ribeirão Preto, SP, Brasil, Departamento de Patologia, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP (Brazil); Tucci, S. [Universidade de São Paulo, Faculdade de Medicina de Ribeirão Preto, Departamento de Cirurgia e Anatomia, Divisão de Urologia, Ribeirão Preto, SP, Brasil, Divisão de Urologia, Departamento de Cirurgia e Anatomia, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP (Brazil); Muglia, V.F. [Universidade de São Paulo, Faculdade de Medicina de Ribeirão Preto, Departamento de Medicina Interna (Centro de Ciência da Imagem), Ribeirão Preto, SP, Brasil, Departamento de Medicina Interna (Centro de Ciência da Imagem), Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP (Brazil); Reis, R.B. Dos [Universidade de São Paulo, Faculdade de Medicina de Ribeirão Preto, Departamento de Cirurgia e Anatomia, Divisão de Urologia, Ribeirão Preto, SP, Brasil, Divisão de Urologia, Departamento de Cirurgia e Anatomia, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP (Brazil); Silva, G.E.B. [Universidade de São Paulo, Faculdade de Medicina de Ribeirão Preto, Departamento de Patologia, Ribeirão Preto, SP, Brasil, Departamento de Patologia, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP (Brazil)

    2014-05-09

    The purpose of this study was to investigate the relationship between cyclin D1 expression and clinicopathological parameters in patients with prostate carcinoma. We assessed cyclin D1 expression by conventional immunohistochemistry in 85 patients who underwent radical prostatectomy for prostate carcinoma and 10 normal prostate tissue samples retrieved from autopsies. We measured nuclear immunostaining in the entire tumor area and based the results on the percentage of positive tumor cells. The preoperative prostate-specific antigen (PSA) level was 8.68±5.16 ng/mL (mean±SD). Cyclin D1 staining was positive (cyclin D1 expression in >5% of tumor cells) in 64 cases (75.4%) and negative (cyclin D1 expression in ≤5% of tumor cells) in 21 cases (including 15 cases with no immunostaining). Normal prostate tissues were negative for cyclin D1. Among patients with a high-grade Gleason score (≥7), 86% of patients demonstrated cyclin D1 immunostaining of >5% (P<0.05). In the crude analysis of cyclin D1 expression, the high-grade Gleason score group showed a mean expression of 39.6%, compared to 26.9% in the low-grade Gleason score group (P<0.05). Perineural invasion tended to be associated with cyclin D1 expression (P=0.07), whereas cyclin D1 expression was not associated with PSA levels or other parameters. Our results suggest that high cyclin D1 expression could be a potential marker for tumor aggressiveness.

  8. Aspirin regulation of c-myc and cyclinD1 proteins to overcome tamoxifen resistance in estrogen receptor-positive breast cancer cells.

    Science.gov (United States)

    Cheng, Ran; Liu, Ya-Jing; Cui, Jun-Wei; Yang, Man; Liu, Xiao-Ling; Li, Peng; Wang, Zhan; Zhu, Li-Zhang; Lu, Si-Yi; Zou, Li; Wu, Xiao-Qin; Li, Yu-Xia; Zhou, You; Fang, Zheng-Yu; Wei, Wei

    2017-05-02

    Tamoxifen is still the most commonly used endocrine therapy drug for estrogen receptor (ER)-positive breast cancer patients and has an excellent outcome, but tamoxifen resistance remains a great impediment to successful treatment. Recent studies have prompted an anti-tumor effect of aspirin. Here, we demonstrated that aspirin not only inhibits the growth of ER-positive breast cancer cell line MCF-7, especially when combined with tamoxifen, but also has a potential function to overcome tamoxifen resistance in MCF-7/TAM. Aspirin combined with tamoxifen can down regulate cyclinD1 and block cell cycle in G0/G1 phase. Besides, tamoxifen alone represses c-myc, progesterone receptor (PR) and cyclinD1 in MCF-7 cell line but not in MCF-7/TAM, while aspirin combined with tamoxifen can inhibit the expression of these proteins in the resistant cell line. When knocking down c-myc in MCF-7/TAM, cells become more sensitive to tamoxifen, cell cycle is blocked as well, indicating that aspirin can regulate c-myc and cyclinD1 proteins to overcome tamoxifen resistance. Our study discovered a novel role of aspirin based on its anti-tumor effect, and put forward some kinds of possible mechanisms of tamoxifen resistance in ER-positive breast cancer cells, providing a new strategy for the treatment of ER-positive breast carcinoma.

  9. Mutation analysis of the negative regulator cyclin G2 in gastric cancer

    African Journals Online (AJOL)

    Cyclin G2 is an unconventional cyclin which might have a potential negative role in carcinogenesis. In this study, the effect of cyclin G2 overexpression on gastric cell proliferation and expression levels of cyclin G2 in normal gastric cells and gastric cancer cells were investigated. Moreover, mutation analysis was performed ...

  10. Cyclin D3 regulates proliferation and apoptosis of leukemic T cell lines

    NARCIS (Netherlands)

    Boonen, G.J.J.C.; Oirschot, B.A. van; Diepen, A. van; Mackus, W.J.M.; Verdonck, L.F.; Rijksen, G.; Medema, R.H.

    1999-01-01

    Activation of the T cell receptor in leukemic T cell lines or T cell hybridomas causes growth inhibition. A similar growth inhibition is seen when protein kinase C is activated through addition of phorbol myristate acetate. This inhibition is due to an arrest of cell cycle progression in G1 combined

  11. Mig-6 is down-regulated in HCC and inhibits the proliferation of HCC cells via the P-ERK/Cyclin D1 pathway.

    Science.gov (United States)

    Li, Zixuan; Qu, Lianyue; Luo, Wenting; Tian, Yulong; Zhai, Huan; Xu, Ke; Zhong, Hongshan

    2017-06-01

    The ablation of Mig-6 has been shown to induce tumor formation in various tissues. However, the relationships between Mig-6 expression, clinical pathological factors, and prognosis have not been clarified in hepatocellular carcinoma (HCC), and the mechanism by which Mig-6 regulates the proliferation of HCC cells has not been reported. In this study, we investigated the clinical significance of the loss of Mig-6 expression in HCC and the mechanism underlying the inhibition of cell proliferation by Mig-6. The down-regulation of Mig-6 correlated significantly with large tumors, a more advanced BCLC stage, and a more advanced TNM stage, and low Mig-6 expression predicted significantly reduced survival. Low Mig-6 expression and high Cyclin D1 expression were independent predictors for survival. The overexpression of Mig-6 led to significant G 1 arrest and growth inhibition in HCC cells, possibly through the inhibition P-ERK and Cyclin D1. These results indicate that Mig-6 expression is low in HCC, which predicts a poor prognosis. Mig-6 may regulate cell proliferation and the cell cycle through the P-ERK/Cyclin D1 pathway. Copyright © 2017 Elsevier Inc. All rights reserved.

  12. Crystal Structure of Human Cyclin K, A Positive Regulator of Cyclin-Dependent Kinase 9

    Energy Technology Data Exchange (ETDEWEB)

    Baek,K.; Brown, R.; Birrane, G.; Ladias, J.

    2007-01-01

    K and the closely related cyclins T1, T2a, and T2b interact with cyclin-dependent kinase 9 (CDK9) forming multiple nuclear complexes, referred to collectively as positive transcription elongation factor b (P-TEFb). Through phosphorylation of the C-terminal domain of the RNA polymerase II largest subunit, distinct P-TEFb species regulate the transcriptional elongation of specific genes that play central roles in human physiology and disease development, including cardiac hypertrophy and human immunodeficiency virus-1 pathogenesis. We have determined the crystal structure of human cyclin K (residues 11-267) at 1.5 {angstrom} resolution, which represents the first atomic structure of a P-TEFb subunit. The cyclin K fold comprises two typical cyclin boxes with two short helices preceding the N-terminal box. A prominent feature of cyclin K is an additional helix (H4a) in the first cyclin box that obstructs the binding pocket for the cell-cycle inhibitor p27{sup Kip1}. Modeling of CDK9 bound to cyclin K provides insights into the structural determinants underlying the formation and regulation of this complex. A homology model of human cyclin T1 generated using the cyclin K structure as a template reveals that the two proteins have similar structures, as expected from their high level of sequence identity. Nevertheless, their CDK9-interacting surfaces display significant structural differences, which could potentially be exploited for the design of cyclin-targeted inhibitors of the CDK9-cyclin K and CDK9-cyclin T1 complexes.

  13. Mitochondrial reactive oxygen species perturb AKT/cyclin D1 cell cycle signaling via oxidative inactivation of PP2A in lowdose irradiated human fibroblasts.

    Science.gov (United States)

    Shimura, Tsutomu; Sasatani, Megumi; Kamiya, Kenji; Kawai, Hidehiko; Inaba, Yohei; Kunugita, Naoki

    2016-01-19

    Here we investigated the cellular response of normal human fibroblasts to repeated exposure to low-dose radiation. In contrast to acute single radiation, low-dose fractionated radiation (FR) with 0.01 Gy/fraction or 0.05 Gy/fraction for 31 days increased in mitochondrial mass, decreased cellular levels of the antioxidant glutathione and caused persistent accumulation of mitochondrial reactive oxygen species (ROS). Excess ROS promoted oxidative inactivation of protein phosphatase PP2A which in turn led to disruption of normal negative feed-back control of AKT/cyclin D1 signaling in cells treated with long-term FR. The resulting abnormal nuclear accumulation of cyclin D1 causes growth retardation, cellular senescence and genome instability in low-dose irradiated cells. Thus, loss of redox control and subsequently elevated levels of ROS perturb signal transduction as a result of oxidative stress. Our study highlights a specific role of mitochondrial ROS in perturbation of AKT/cyclin D1 cell cycle signaling after low-dose long-term FR. The antioxidants N-acetyl-L-cysteine, TEMPO and mitochondrial-targeted antioxidant Mito-TEMPO provided protection against the harmful cell cycle perturbations induced by low-dose long-term FR.

  14. Cytotoxic activity of simvastatin in T47D breast cancer cell lines and its effect on cyclin D1 expression and apoptosis

    OpenAIRE

    Putra, Bayu; Wahyuningsih, Mae Sri Hartati; Sholikhah, Eti Nurwening

    2017-01-01

    Background: Statins (HMG-CoA Inhibitors) is a drug used for decreasing plasma cholesterol levels and used in therapy to prevent coronary artery disease. Research in animals and epidemiological studies showed that statin therapy can decrease risk against cancer associated with cholesterol. Based on that result then research of cytotoxic activity against simvastatin knowing cultur T47D breast cancer cells and his influence in decreasing expression of cyclin D1 and induction of apoptosis has bee...

  15. DYRK1A-mediated Cyclin D1 Degradation in Neural Stem Cells Contributes to the Neurogenic Cortical Defects in Down Syndrome

    Directory of Open Access Journals (Sweden)

    Sònia Najas

    2015-02-01

    Full Text Available Alterations in cerebral cortex connectivity lead to intellectual disability and in Down syndrome, this is associated with a deficit in cortical neurons that arises during prenatal development. However, the pathogenic mechanisms that cause this deficit have not yet been defined. Here we show that the human DYRK1A kinase on chromosome 21 tightly regulates the nuclear levels of Cyclin D1 in embryonic cortical stem (radial glia cells, and that a modest increase in DYRK1A protein in transgenic embryos lengthens the G1 phase in these progenitors. These alterations promote asymmetric proliferative divisions at the expense of neurogenic divisions, producing a deficit in cortical projection neurons that persists in postnatal stages. Moreover, radial glial progenitors in the Ts65Dn mouse model of Down syndrome have less Cyclin D1, and Dyrk1a is the triplicated gene that causes both early cortical neurogenic defects and decreased nuclear Cyclin D1 levels in this model. These data provide insights into the mechanisms that couple cell cycle regulation and neuron production in cortical neural stem cells, emphasizing that the deleterious effect of DYRK1A triplication in the formation of the cerebral cortex begins at the onset of neurogenesis, which is relevant to the search for early therapeutic interventions in Down syndrome.

  16. The human HECA interacts with cyclins and CDKs to antagonize Wnt-mediated proliferation and chemoresistance of head and neck cancer cells

    International Nuclear Information System (INIS)

    Dowejko, Albert; Bauer, Richard; Bauer, Karin; Müller-Richter, Urs D.A.; Reichert, Torsten E.

    2012-01-01

    There is a growing evidence that the human homologue of the Drosophila headcase (HECA) plays an important role in human carcinogenesis. So far specific protein interaction partners and affected signaling pathways of HECA are still elusive. In a recent study we showed that HECA overexpression in oral squamous-cell carcinoma (OSCC) keratinocytes has tumor suppressive effects resulting in a recuperation of cell cycle control concerning the entry and progression of S-phase, G2- and M-phase. Currently, quantitative RT-PCR and immunohistochemical analysis of primary tumor tissue from OSCC patients demonstrate that HECA expression is markedly decreased compared to normal control patients with abundant HECA expression. Additionally, there is nearly no HECA expression in OSCC metastases. Here, we show that HECA expression is negatively controlled by the Wnt-pathway and TCF4, a Wnt related transcription factor, binds to the HECA promoter. Furthermore, immunocytochemistry reveals colocalization of HECA with the cyclin dependent kinase CDK9. Immunoprecipitation experiments and proximity ligation assays further reveal an interaction of HECA with CDK2, CDK9, Cyclin A and Cyclin K, a direct transcriptional target of the p53 tumor suppressor. Silencing HECA in OSCC cell lines leads to a significant increase of cell division and a markedly increased resistance against the chemotherapeutic cisplatin. On the contrary, HECA overexpressing OSCC cell lines show decreased resistance of OSCC cells against cisplatin. Therefore, HECA could be considered as future therapeutic agent against Wnt-dependent tumor progression. -- Highlights: ► HECA is a new cell cycle regulator with anti-tumor features in head and neck cancer. ► During tumor progression HECA mRNA and protein expression decrease. ► The HECA promotor is a direct target of the Wnt/beta-catenin/TCF-pathway. ► The HECA protein antagonizes Wnt-mediated cell proliferation through interaction with major cell cycle factors.

  17. Modulation of Cyclins, p53 and Mitogen-Activated Protein Kinases Signaling in Breast Cancer Cell Lines by 4-(3,4,5-Trimethoxyphenoxybenzoic Acid

    Directory of Open Access Journals (Sweden)

    Kuan-Han Lee

    2014-01-01

    Full Text Available Despite the advances in cancer therapy and early detection, breast cancer remains a leading cause of cancer-related deaths among females worldwide. The aim of the current study was to investigate the antitumor activity of a novel compound, 4-(3,4,5-trimethoxyphenoxybenzoic acid (TMPBA and its mechanism of action, in breast cancer. Results indicated the relatively high sensitivity of human breast cancer cell-7 and MDA-468 cells towards TMPBA with IC50 values of 5.9 and 7.9 µM, respectively compared to hepatocarcinoma cell line Huh-7, hepatocarcinoma cell line HepG2, and cervical cancer cell line Hela cells. Mechanistically, TMPBA induced apoptotic cell death in MCF-7 cells as indicated by 4',6-diamidino-2-phenylindole (DAPI nuclear staining, cell cycle analysis and the activation of caspase-3. Western blot analysis revealed the ability of TMPBA to target pathways mediated by mitogen-activated protein (MAP kinases, 5' adenosine monophosphate-activated protein kinase (AMPK, and p53, of which the concerted action underlined its antitumor efficacy. In addition, TMPBA induced alteration of cyclin proteins’ expression and consequently modulated the cell cycle. Taken together, the current study underscores evidence that TMPBA induces apoptosis in breast cancer cells via the modulation of cyclins and p53 expression as well as the modulation of AMPK and mitogen-activated protein kinases (MAPK signaling. These findings support TMPBA’s clinical promise as a potential candidate for breast cancer therapy.

  18. The Role of Cyclin D1 in the Chemoresistance of Mantle Cell Lymphoma

    Science.gov (United States)

    2016-09-01

    the tumor bank of the Pathology Department of City of Hope as de-identified samples after approval by the Institutional Review Board. Cells were...previous studies for this cell line.[21,31] Aberrant CCND1 signaling is considered central to MCL pathogenesis and major efforts to target this...kinase-coordinated phosphorylation of endogenous pocket proteins differentially regulates their interactions with E2F4 and E2F1 and gene expression. J

  19. Induction of G1 cell cycle arrest and cyclin D1 down-regulation in response to pericarp extract of Baneh in human breast cancer T47D cells.

    Science.gov (United States)

    Rezaei, Parisa Fathi; Fouladdel, Shamileh; Ghaffari, Seyed Mahmood; Amin, Gholamreza; Azizi, Ebrahim

    2012-12-28

    Natural products from plants have an important role in the development and production of new drugs mainly for cancer therapy. More recently, we have shown that the pericarp methanolic extract of Pistacia atlantica sub kurdica (with local name of Baneh) as a rich source of active biological components with high antioxidant and radical scavenging activities, has ability to cease proliferation and induce apoptosis in T47D human breast cancer cells. The present study aimed to clarify whether Baneh extract able to alter cell cycle progression of T47D cells or not. In order to study the possible effect of Baneh extract on cell cycle of T47D cells, we evaluated cell cycle distribution and its regulatory proteins by flow cytometry and western blot analysis respectively. Baneh extract induced G0/G1 cell cycle arrest in conjunction with a marked decrease in expression of cyclin D1 and cdk4 that was strongly dependent on time of exposure. In parallel, Dox-treated T47D cells in early time points were accumulated on S phase, but after 48 h cell cycle progression was inhibited on G2/M. Dox promoted striking accumulation of cyclin B1 rapidly and enhanced cyclin A abundance. Taken together, our results establish that the antitumor activity of the pericarp extract of Baneh partly is mediated via cell cycle arrest and downregulation of cyclin D1 and cdk4 expression. These findings warrant further evaluation regarding the mechanism(s) of action of this promising anticancer agent.

  20. Cyclin D1 represses p300 transactivation through a cyclin-dependent kinase-independent mechanism.

    Science.gov (United States)

    Fu, Maofu; Wang, Chenguang; Rao, Mahadev; Wu, Xiaofang; Bouras, Toula; Zhang, Xueping; Li, Zhiping; Jiao, Xuanmao; Yang, Jianguo; Li, Anping; Perkins, Neil D; Thimmapaya, Bayar; Kung, Andrew L; Munoz, Alberto; Giordano, Antonio; Lisanti, Michael P; Pestell, Richard G

    2005-08-19

    Cyclin D1 encodes a regulatory subunit, which with its cyclin-dependent kinase (Cdk)-binding partner forms a holoenzyme that phosphorylates and inactivates the retinoblastoma protein. In addition to its Cdk binding-dependent functions, cyclin D1 regulates cellular differentiation in part by modifying several transcription factors and nuclear receptors. The molecular mechanism through which cyclin D1 regulates the function of transcription factors involved in cellular differentiation remains to be clarified. The histone acetyltransferase protein p300 is a co-integrator required for regulation of multiple transcription factors. Here we show that cyclin D1 physically interacts with p300 and represses p300 transactivation. We demonstrated further that the interaction of the two proteins occurs at the peroxisome proliferator-activated receptor gamma-responsive element of the lipoprotein lipase promoter in the context of the local chromatin structure. We have mapped the domains in p300 and cyclin D1 involved in this interaction. The bromo domain and cysteine- and histidine-rich domains of p300 were required for repression by cyclin D1. Cyclin D1 repression of p300 was independent of the Cdk- and retinoblastoma protein-binding domains of cyclin D1. Cyclin D1 inhibits histone acetyltransferase activity of p300 in vitro. Microarray analysis identified a signature of genes repressed by cyclin D1 and induced by p300 that promotes cellular differentiation and induces cell cycle arrest. Together, our results suggest that cyclin D1 plays an important role in cellular proliferation and differentiation through regulation of p300.

  1. Effects of BP-14, a novel cyclin-dependent kinase inhibitor, on anaplastic thyroid cancer cells

    Czech Academy of Sciences Publication Activity Database

    Allegri, L.; Baldan, F.; Mio, F.; Puppin, C.; Russo, D.; Kryštof, Vladimír; Damante, G.

    2016-01-01

    Roč. 35, č. 4 (2016), s. 2413-2418 ISSN 1021-335X R&D Projects: GA ČR(CZ) GA15-15264S Institutional support: RVO:61389030 Keywords : mTOR * thyroid cancer * cell proliferation Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.662, year: 2016

  2. Genome-wide analysis of cyclin family in rice (Oryza Sativa L.).

    Science.gov (United States)

    La, Honggui; Li, Jun; Ji, Zhendong; Cheng, Yanjun; Li, Xiuli; Jiang, Shuye; Venkatesh, Prasanna Nori; Ramachandran, Srinivasan

    2006-04-01

    The cyclins together with highly conserved cyclin-dependent kinases regulate cell cycle progression in plants. Although extensive and systematic study on cell cycle mechanisms and cyclin functions in yeasts and animals has been carried out, only a small number of plant cyclins have been characterized and classified functionally and phylogenetically. We identified several types of cyclin genes in the rice genome and characterized them by phylogenetic, tandem and segmental duplications analyses. Our results indicated that there were at least 49 predicted rice cyclin genes in the rice genome, and they were distributed on 12 chromosomes. Of these cyclins, one possessed only cyclin_C domain and no cyclin_N domain, and the remaining 48 cyclins with cyclin_N domains were classified as nine types based on evolutionary relationships. Eight of these nine types were common between rice and Arabidopsis, whereas only one, known as F-type cyclins, was unique to rice. No homologues of the F-type cyclins in plants could be retrieved from the public databases, and reverse transcription-PCR analysis supported an existence of the F-type cyclin genes. Sequence alignment suggested that the cyclin genes in the rice genome experienced a mass of gene tandem and segmental duplications occurred on seven chromosomes related to the origins of new cyclin genes. Our study provided an opportunity to facilitate assessment and classification of new members, serving as a guide for further functional elucidation of rice cyclins.

  3. Promoter- and cell-specific epigenetic regulation of CD44, Cyclin D2, GLIPR1 and PTEN by Methyl-CpG binding proteins and histone modifications

    International Nuclear Information System (INIS)

    Müller, Imke; Wischnewski, Frank; Pantel, Klaus; Schwarzenbach, Heidi

    2010-01-01

    The aim of the current study was to analyze the involvement of methyl-CpG binding proteins (MBDs) and histone modifications on the regulation of CD44, Cyclin D2, GLIPR1 and PTEN in different cellular contexts such as the prostate cancer cells DU145 and LNCaP, and the breast cancer cells MCF-7. Since global chromatin changes have been shown to occur in tumours and regions of tumour-associated genes are affected by epigenetic modifications, these may constitute important regulatory mechanisms for the pathogenesis of malignant transformation. In DU145, LNCaP and MCF-7 cells mRNA expression levels of CD44, Cyclin D2, GLIPR1 and PTEN were determined by quantitative RT-PCR at the basal status as well as after treatment with demethylating agent 5-aza-2'-deoxycytidine and/or histone deacetylase inhibitor Trichostatin A. Furthermore, genomic DNA was bisulfite-converted and sequenced. Chromatin immunoprecipitation was performed with the stimulated and unstimulated cells using antibodies for MBD1, MBD2 and MeCP2 as well as 17 different histone antibodies. Comparison of the different promoters showed that MeCP2 and MBD2a repressed promoter-specifically Cyclin D2 in all cell lines, whereas in MCF-7 cells MeCP2 repressed cell-specifically all methylated promoters. Chromatin immunoprecipitation showed that all methylated promoters associated with at least one MBD. Treatment of the cells by the demethylating agent 5-aza-2'-deoxycytidine (5-aza-CdR) caused dissociation of the MBDs from the promoters. Only MBD1v1 bound and repressed methylation-independently all promoters. Real-time amplification of DNA immunoprecipitated by 17 different antibodies showed a preferential enrichment for methylated lysine of histone H3 (H3K4me1, H3K4me2 and H3K4me3) at the particular promoters. Notably, the silent promoters were associated with unmodified histones which were acetylated following treatment by 5-aza-CdR. This study is one of the first to reveal the histone code and MBD profile

  4. Promoter- and cell-specific epigenetic regulation of CD44, Cyclin D2, GLIPR1 and PTEN by Methyl-CpG binding proteins and histone modifications

    Directory of Open Access Journals (Sweden)

    Schwarzenbach Heidi

    2010-06-01

    Full Text Available Abstract Background The aim of the current study was to analyze the involvement of methyl-CpG binding proteins (MBDs and histone modifications on the regulation of CD44, Cyclin D2, GLIPR1 and PTEN in different cellular contexts such as the prostate cancer cells DU145 and LNCaP, and the breast cancer cells MCF-7. Since global chromatin changes have been shown to occur in tumours and regions of tumour-associated genes are affected by epigenetic modifications, these may constitute important regulatory mechanisms for the pathogenesis of malignant transformation. Methods In DU145, LNCaP and MCF-7 cells mRNA expression levels of CD44, Cyclin D2, GLIPR1 and PTEN were determined by quantitative RT-PCR at the basal status as well as after treatment with demethylating agent 5-aza-2'-deoxycytidine and/or histone deacetylase inhibitor Trichostatin A. Furthermore, genomic DNA was bisulfite-converted and sequenced. Chromatin immunoprecipitation was performed with the stimulated and unstimulated cells using antibodies for MBD1, MBD2 and MeCP2 as well as 17 different histone antibodies. Results Comparison of the different promoters showed that MeCP2 and MBD2a repressed promoter-specifically Cyclin D2 in all cell lines, whereas in MCF-7 cells MeCP2 repressed cell-specifically all methylated promoters. Chromatin immunoprecipitation showed that all methylated promoters associated with at least one MBD. Treatment of the cells by the demethylating agent 5-aza-2'-deoxycytidine (5-aza-CdR caused dissociation of the MBDs from the promoters. Only MBD1v1 bound and repressed methylation-independently all promoters. Real-time amplification of DNA immunoprecipitated by 17 different antibodies showed a preferential enrichment for methylated lysine of histone H3 (H3K4me1, H3K4me2 and H3K4me3 at the particular promoters. Notably, the silent promoters were associated with unmodified histones which were acetylated following treatment by 5-aza-CdR. Conclusions This study is one

  5. Functional p53 in cells contributes to the anticancer effect of the cyclin-dependent kinase inhibitor roscovitine

    Czech Academy of Sciences Publication Activity Database

    Paprskářová, Martina; Kryštof, Vladimír; Jorda, Radek; Džubák, P.; Hajdúch, M.; Wesierska-Gadek, J.; Strnad, Miroslav

    2009-01-01

    Roč. 107, č. 3 (2009), s. 428-437 ISSN 0730-2312 R&D Projects: GA ČR GA204/08/0511 Institutional research plan: CEZ:AV0Z50380511 Keywords : APOPTOSIS * CYCLIN-DEPENDENT KINASE * OLOMOUCINE II * p53 Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.935, year: 2009

  6. Nerve growth factor-induced accumulation of PC12 cells expressing cyclin D1: evidence for a G1 phase block.

    Science.gov (United States)

    van Grunsven, L A; Thomas, A; Urdiales, J L; Machenaud, S; Choler, P; Durand, I; Rudkin, B B

    1996-02-15

    The anti-proliferative effect of nerve growth factor (NGF) on the rat pheochromocytoma cell line PC12 has been previously shown to be accompanied by the accumulation of cells in either the G1 phase with a 2c DNA content, or with a 4c DNA content characteristic for G2/M, as evidenced by flow cytometric analysis of DNA distribution using propidium iodide. Herein, these apparently conflicting results are clarified. The present studies indicate that a simple DNA distribution profile obtained by this technique can confound interpretation of the biological effects of NGF on cell-cycle distribution due to the presence of tetraploid cells. Using cyclin D1 and incorporation of bromodeoxyuridine as markers of respectively, G1 and S phase, we show that PC12 cultures can have a considerable amount of tetraploid cells which, when in the G1 phase, have a 4c DNA content and express cyclin D1. During exposure to NGF, this population increases, reflecting the accumulation of cells in the G1 phase of the cell cycle. The data presented, support the possibility that events affecting the expression or action of G1 regulatory proteins may be involved in the molecular mechanism of the anti-mitogenic effect of NGF.

  7. Inhibition of protein kinase C in PHA-activated PBMC treated with anti-HLA class I monoclonal antibody is associated to decreased proliferation and expression of cell cycle related genes.

    Science.gov (United States)

    Rita, S; Michele, R; Patrizia, O; Alessansdra, B; Sara, B; Enrico, G

    1994-01-01

    Treatment of PHA-activated PBMC with anti-HLA class I monoclonal antibody (mAb 01.65) shows: 1) depletion of particulate protein Kinase C (PKC) and partial reduction of cytosolic PKC after only 10 min.; 2) inhibition of tritiated thymidine (3H-Td) incorporation; 3) slowing down of cell cycle; 4) reduced expression of four cell cycle related genes. These findings suggest that the depletion of PKC is reflected on the cell cycle progression and expression of cell cycle related genes. We studied, in PHA-activated PBMC cultures, the effect of N-N-Staurosporine (StSp) acting as PKC inhibitor at nanomolar concentrations, alone and combined with mAb 01.65. StSp, inhibits the proliferative response of PHA stimulated PBMC in a competitive fashion with mAb 01.65. We report here that StSp alone and combined with mAb 01.65 affects also the expression of c-myc and cdc2 and the membrane expression of two proliferation markers: IL-2R (CD25) and TfR (CD71).

  8. MicroRNA-26a/cyclin-dependent kinase 5 axis controls proliferation, apoptosis and in vivo tumor growth of diffuse large B-cell lymphoma cell lines.

    Science.gov (United States)

    Farina, Floriana Maria; Inguscio, Alessandra; Kunderfranco, Paolo; Cortesi, Alice; Elia, Leonardo; Quintavalle, Manuela

    2017-06-22

    Diffuse large B-cell lymphoma (DLBCL) is the most frequent type of non-Hodgkin lymphoma. Despite a favorable therapeutic response to first-line chemo-immunotherapy, still 30-40% of patients is refractory, or relapse after this treatment. Thus, alternative strategies must be sought. Previous studies have indicated that cyclin-dependent kinase 5 (CDK5), a serine/threonine protein kinase, is involved in tumor development and progression, and it may represent a potential therapeutic target. However, its role in modulating DLBCL growth and progression remains largely unexplored. In this study, we show that CDK5 and its activator, cyclin-dependent kinase 5 activator 1 (CDK5R1 or p35), are overexpressed in DLBCL cell lines and that signal transducer and activator of transcription 3 (STAT3) phosphorylation and activity is dependent on CDK5 expression in DLBCL. Using public data sets, we also demonstrate that patients with DLBCL show a higher expression of CDK5 compared with healthy individuals. By using loss-of-function approaches, we demonstrate that CDK5's activity regulates proliferation and survival of DLBCL cells. MicroRNAs (miRNAs or miRs) are small noncoding RNAs that negatively regulating gene expression and are involved in cancer initiation and progression. We identify miR-26a as direct regulator of p35 expression and CDK5 activity. We show that miR-26a expression is lower in DLBCL cell lines compared to B lymphocytes and that its ectopic expression leads to a drastic reduction of DLBCL tumor growth in vivo and decreased proliferation, cell-cycle progression, and survival in vitro. Remarkably, concomitant overexpression of a 3'-UTR-truncated form of p35 promoted tumor growth in vivo and cell proliferation, cell-cycle progression, and cell survival in vitro. In conclusion, these results demonstrate an important role for miR-26a and CDK5 together in the survival and growth of DLBCL cells, suggesting the existence of potential novel therapeutic targets for the

  9. Molecular evolution of cyclin proteins in animals and fungi

    Directory of Open Access Journals (Sweden)

    Afonnikov Dmitry A

    2011-07-01

    Full Text Available Abstract Background The passage through the cell cycle is controlled by complexes of cyclins, the regulatory units, with cyclin-dependent kinases, the catalytic units. It is also known that cyclins form several families, which differ considerably in primary structure from one eukaryotic organism to another. Despite these lines of evidence, the relationship between the evolution of cyclins and their function is an open issue. Here we present the results of our study on the molecular evolution of A-, B-, D-, E-type cyclin proteins in animals and fungi. Results We constructed phylogenetic trees for these proteins, their ancestral sequences and analyzed patterns of amino acid replacements. The analysis of infrequently fixed atypical amino acid replacements in cyclins evidenced that accelerated evolution proceeded predominantly during paralog duplication or after it in animals and fungi and that it was related to aromorphic changes in animals. It was shown also that evolutionary flexibility of cyclin function may be provided by consequential reorganization of regions on protein surface remote from CDK binding sites in animal and fungal cyclins and by functional differentiation of paralogous cyclins formed in animal evolution. Conclusions The results suggested that changes in the number and/or nature of cyclin-binding proteins may underlie the evolutionary role of the alterations in the molecular structure of cyclins and their involvement in diverse molecular-genetic events.

  10. Genome-Wide Analysis of the Cyclin Gene Family in Tomato

    Directory of Open Access Journals (Sweden)

    Tingyan Zhang

    2013-12-01

    Full Text Available Cyclins play important roles in cell division and cell expansion. They also interact with cyclin-dependent kinases to control cell cycle progression in plants. Our genome-wide analysis identified 52 expressed cyclin genes in tomato. Phylogenetic analysis of the deduced amino sequences of tomato and Arabidopsis cyclin genes divided them into 10 types, A-, B-, C-, D-, H-, L-, T-, U-, SDS- and J18. Pfam analysis indicated that most tomato cyclins contain a cyclin-N domain. C-, H- and J18 types only contain a cyclin-C domain, and U-type cyclins contain another potential cyclin domain. All of the cyclin genes are distributed throughout the tomato genome except for chromosome 8, and 30 of them were found to be segmentally duplicated; they are found on the duplicate segments of chromosome 1, 2, 3, 4, 5, 6, 10, 11 and 12, suggesting that tomato cyclin genes experienced a mass of segmental duplication. Quantitative real-time polymerase chain reaction analysis indicates that the expression patterns of tomato cyclin genes were significantly different in vegetative and reproductive stages. Transcription of most cyclin genes can be enhanced or repressed by exogenous application of gibberellin, which implies that gibberellin maybe a direct regulator of cyclin genes. The study presented here may be useful as a guide for further functional research on tomato cyclins.

  11. The human HECA interacts with cyclins and CDKs to antagonize Wnt-mediated proliferation and chemoresistance of head and neck cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Dowejko, Albert, E-mail: Albert.Dowejko@klinik.uni-regensburg.de [Department of Oral and Maxillofacial Surgery, University Hospital Regensburg, Franz-Josef-Strauss-Allee 11, 93053 Regensburg (Germany); Bauer, Richard; Bauer, Karin [Department of Oral and Maxillofacial Surgery, University Hospital Regensburg, Franz-Josef-Strauss-Allee 11, 93053 Regensburg (Germany); Mueller-Richter, Urs D.A. [Department of Oral and Maxillofacial Plastic Surgery, University of Wuerzburg, Pleicherwall 2, 97070 Wuerzburg (Germany); Reichert, Torsten E. [Department of Oral and Maxillofacial Surgery, University Hospital Regensburg, Franz-Josef-Strauss-Allee 11, 93053 Regensburg (Germany)

    2012-03-10

    There is a growing evidence that the human homologue of the Drosophila headcase (HECA) plays an important role in human carcinogenesis. So far specific protein interaction partners and affected signaling pathways of HECA are still elusive. In a recent study we showed that HECA overexpression in oral squamous-cell carcinoma (OSCC) keratinocytes has tumor suppressive effects resulting in a recuperation of cell cycle control concerning the entry and progression of S-phase, G2- and M-phase. Currently, quantitative RT-PCR and immunohistochemical analysis of primary tumor tissue from OSCC patients demonstrate that HECA expression is markedly decreased compared to normal control patients with abundant HECA expression. Additionally, there is nearly no HECA expression in OSCC metastases. Here, we show that HECA expression is negatively controlled by the Wnt-pathway and TCF4, a Wnt related transcription factor, binds to the HECA promoter. Furthermore, immunocytochemistry reveals colocalization of HECA with the cyclin dependent kinase CDK9. Immunoprecipitation experiments and proximity ligation assays further reveal an interaction of HECA with CDK2, CDK9, Cyclin A and Cyclin K, a direct transcriptional target of the p53 tumor suppressor. Silencing HECA in OSCC cell lines leads to a significant increase of cell division and a markedly increased resistance against the chemotherapeutic cisplatin. On the contrary, HECA overexpressing OSCC cell lines show decreased resistance of OSCC cells against cisplatin. Therefore, HECA could be considered as future therapeutic agent against Wnt-dependent tumor progression. -- Highlights: Black-Right-Pointing-Pointer HECA is a new cell cycle regulator with anti-tumor features in head and neck cancer. Black-Right-Pointing-Pointer During tumor progression HECA mRNA and protein expression decrease. Black-Right-Pointing-Pointer The HECA promotor is a direct target of the Wnt/beta-catenin/TCF-pathway. Black-Right-Pointing-Pointer The HECA protein

  12. Lidocaine inhibits NIH-3T3 cell multiplication by increasing the expression of cyclin-dependent kinase inhibitor 1A (p21).

    Science.gov (United States)

    Desai, Sukumar P; Kojima, Koji; Vacanti, Charles A; Kodama, Shohta

    2008-11-01

    We explored molecular mechanisms by which lidocaine inhibits growth in the murine embryonic fibroblast cell line NIH-3T3. Local anesthetics can adversely affect cell growth in vitro. Their effects on wound healing are controversial. We examined the effects and novel mechanisms by which lidocaine affects in vitro multiplication of the murine fibroblast cell line NIH-3T3. NIH-3T3 cells were grown in culture with lidocaine [0, 0.05, 0.5, 1, 2, and 5 mM]. Cell multiplication was assessed by determining cell counts on subsequent days, while mechanisms by which inhibition occurred were evaluated by bromodeoxyuridine uptake, gene expression using polymerase chain reaction array, and Western blot analysis to verify increased levels of affected proteins. Lidocaine caused dose-dependent inhibition of multiplication of NIH-3T3 cells. Effects ranged from no inhibition [0.05 and 0.5 mM] and mild inhibition [1 mM], to severe inhibition [2 and 5 mM] [P = 0.006]. Lidocaine 2 mM inhibited bromodeoxyuridine uptake at day 3.5 [P = 0.02 versus control, and P = 0.0495 vs 1 mM lidocaine]. On day 1.5, lidocaine upregulated expression of cyclin-D1 and cyclin-dependent kinase inhibitor 1A [p21]. On day 2.5, lidocaine increased the levels of p21 protein. Low concentrations of lidocaine, as would be seen in plasma after spinal, epidural, or plexus anesthesia, do not significantly affect multiplication of fibroblasts. Higher doses of lidocaine arrest cell multiplication at the S-phase of the growth cycle by upregulation of p21, an extremely potent inhibitor of cell multiplication. Higher concentrations, as would be seen after tissue infiltration, severely inhibit fibroblast multiplication and thus may impair wound healing.

  13. The role of cell cycle in retinal development: cyclin-dependent kinase inhibitors co-ordinate cell-cycle inhibition, cell-fate determination and differentiation in the developing retina.

    Science.gov (United States)

    Bilitou, Aikaterini; Ohnuma, Shin-ichi

    2010-03-01

    The mature retina is formed through multi-step developmental processes, including eye field specification, optic vesicle evagination, and cell-fate determination. Co-ordination of these developmental events with cell-proliferative activity is essential to achieve formation of proper retinal structure and function. In particular, the molecular and cellular dynamics of the final cell cycle significantly influence the identity that a cell acquires, since cell fate is largely determined at the final cell cycle for the production of postmitotic cells. This review summarizes our current understanding of the cellular mechanisms that underlie the co-ordination of cell-cycle and cell-fate determination, and also describes a molecular role of cyclin-dependent kinase inhibitors (CDKIs) as co-ordinators of cell-cycle arrest, cell-fate determination and differentiation. Copyright (c) 2010 Wiley-Liss, Inc.

  14. The effect of Cyclin-dependent kinase 5 on voltage-dependent calcium channels in PC12 cells varies according to channel type and cell differentiation state.

    Science.gov (United States)

    Furusawa, Kotaro; Asada, Akiko; Saito, Taro; Hisanaga, Shin-ichi

    2014-08-01

    Cyclin-dependent kinase 5 (Cdk5) is a Ser/Thr kinase that plays an important role in the release of neurotransmitter from pre-synaptic terminals triggered by Ca(2+) influx into the pre-synaptic cytoplasm through voltage-dependent Ca(2+) channels (VDCCs). It is reported that Cdk5 regulates L-, P/Q-, or N-type VDCC, but there is conflicting data as to the effect of Cdk5 on VDCC activity. To clarify the mechanisms involved, we examined the role of Cdk5 in regulating the Ca(2+) -channel property of VDCCs, using PC12 cells expressing endogenous, functional L-, P/Q-, and N-type VDCCs. The Ca(2+) influx, induced by membrane depolarization with high K(+) , was monitored with a fluorescent Ca(2+) indicator protein in both undifferentiated and nerve growth factor (NGF)-differentiated PC12 cells. Overall, Ca(2+) influx was increased by expression of Cdk5-p35 in undifferentiated PC12 cells but suppressed in differentiated PC12 cells. Moreover, we found that different VDCCs are distinctly regulated by Cdk5-p35 depending on the differentiation states of PC12 cells. These results indicate that Cdk5-p35 regulates L-, P/Q-, or N-type VDCCs in a cellular context-dependent manner. Calcium (Ca(2+) ) influx through voltage-dependent Ca(2+) channels (VDCCs) triggers neurotransmitter release from pre-synaptic terminal of neurons. The channel activity of VDCCs is regulated by Cdk5-p35, a neuronal Ser/Thr kinase. However, there have been debates about the regulation of VDCCs by Cdk5. Using PC12 cells, we show that Cdk5-p35 regulates VDCCs in a type (L, P/Q, and N) and differentiation-dependent manner. NGF = nerve growth factor. © 2014 International Society for Neurochemistry.

  15. Increased expression of cyclin B1 mRNA coincides with diminished G{sub 2}-phase arrest in irradiated HeLa cells treated with staurosporine or caffeine

    Energy Technology Data Exchange (ETDEWEB)

    Bernhard, E.J.; Maity, A.; McKenna, W.G.; Muschel, R.J. [Univ. of Pennsylvania School of Medicine, Philadelphia, PA (United States)

    1994-12-01

    The irradiation of cells results in delayed progression through the G{sub 2} phase of the cell cycle. Treatment of irradiated HeLa cells with caffeine greatly reduces the G{sub 2}-phase delay, while caffeine does not alter progression of cells through the cell cycle in unirradiated cells. In this report we demonstrate that treatment of HeLa cells with the kinase inhibitor staurosporine, but not with the inhibitor H7, also results in a reduction of the G{sub 2}-phase arrest after irradiation. Cell cycle progression in unirradiated cells is unaffected by 4.4 nM (2ng/ml) staurosporine, which releases the radiation-induced G{sub 2}-phase arrest. In HeLa cells, the G{sub 2}-phase delay after irradiation in S phase is accompanied by decreased expression of cyclin B1 mRNA. Coincident with the reduction in G{sub 2}-phase delay, we observed an increase in cyclin B1 mRNA accumulation in irradiated, staurosporine-treated cells compared to cells treated with irradiation alone. Caffeine treatment of irradiated HeLa cells also resulted in an elevation in the levels of cyclin B1 message. These results support the hypothesis that diminished cyclin B1 mRNA levels influence G{sub 2}-phase arrest to some degree. The findings that both staurosporine and caffeine treatments reverse the depression in cyclin B1 expression suggest that these two compounds may act on a common pathway of cell cycle control in response to radiation injury. 33 refs., 6 figs.

  16. Apoptosis, proliferation and p53, cyclin D1, and retinoblastoma gene expression in relation to radiation response in transitional cell carcinoma of the bladder

    International Nuclear Information System (INIS)

    Moonen, Luc; Ong, Francisca; Gallee, Maarten; Verheij, Marcel; Horenblas, Simon; Hart, Augustinus A.M.; Bartelink, Harry

    2001-01-01

    Purpose: To determine whether the apoptotic index, the Ki67 index, and the expression of the p53, cyclin D1, and retinoblastoma genes correlate with local control, overall survival, and time to distant metastases in invasive bladder cancer treated with external beam radiation. Methods and Materials: Paraffin-embedded pretreatment biopsies from 83 patients with invasive transitional cell carcinoma of the bladder were scored morphologically for apoptosis and immunohistochemically for Ki67, p53, cyclin D1, and retinoblastoma gene expression. Survival analysis methods were used to assess overall survival, local control, and freedom from distant metastases. A multiple proportional hazard (PH) regression analysis was performed to study the prognostic value of the above mentioned biologic parameters (all divided into two categories, except Ki67) in addition to classical prognostic factors such as T stage, histologic grade, multifocality of the tumor, and completeness of transurethral resection. All patients were treated with external beam radiation as sole treatment. Median follow-up for the 19 patients still living was 7.5 years. Results: Apoptotic index varied from 0% to 3.4% with a mean of 0.8% and a median of 0.6%. Ki67 index varied from 0% to 60% with a mean of 14% and a median of 12%. P53 protein was detectable in 61% of the tumors. Overexpression of cyclin D1 was observed in 39% of the tumors and loss of retinoblastoma protein in 23% of the tumors. High Ki67 index was found to be significantly associated with p53 expression (p=0.04) and cyclin D1 overexpression (p=0.023). Cyclin D1 overexpression was found more often in Rb-positive tumors than in Rb-negative tumors (p=0.006). Other associations between the markers are less clear. Biologic markers were not correlated with T stage or grade. In the PH analysis local control was found to be significantly better for tumors with wild-type p53 (p=0.028). Also, tumors with an apoptotic index above the median value (0

  17. PPARγ ligands suppress the feedback loop between E2F2 and cyclin-E1

    International Nuclear Information System (INIS)

    Komatsu, Yoko; Ito, Ichiaki; Wayama, Mitsutoshi; Fujimura, Akiko; Akaogi, Kensuke; Machida, Hikaru; Nakajima, Yuka; Kuroda, Takao; Ohmori, Kazuji; Murayama, Akiko; Kimura, Keiji; Yanagisawa, Junn

    2008-01-01

    PPARγ is a nuclear hormone receptor that plays a key role in the induction of peroxisome proliferation. A number of studies showed that PPARγ ligands suppress cell cycle progression; however, the mechanism remains to be determined. Here, we showed that PPARγ ligand troglitazone inhibited G1/S transition in colon cancer cells, LS174T. Troglitazone did not affect on either expression of CDK inhibitor (p18) or Wnt signaling pathway, indicating that these pathways were not involved in the troglitazone-dependent cell cycle arrest. GeneChip and RT-PCR analyses revealed that troglitazone decreased mRNA levels of cell cycle regulatory factors E2F2 and cyclin-E1 whose expression is activated by E2F2. Down-regulation of E2F2 by troglitazone results in decrease of cyclin-E1 transcription, which could inhibit phosphorylation of Rb protein, and consequently evoke the suppression of E2F2 transcriptional activity. Thus, we propose that troglitazone suppresses the feedback loop containing E2F2, cyclin-E1, and Rb protein

  18. 17-AAG mediated targeting of Hsp90 limits tert activity in peritoneal sarcoma related malignant ascites by downregulating cyclin D1 during cell cycle entry.

    Science.gov (United States)

    Chaklader, M; Das, P; Pereira, J A; Law, A; Chattopadhyay, S; Chatterjee, R; Mondal, A; Law, S

    2012-07-01

    Peritoneal or retro-peritoneal sarcomatosis related malignant ascites formation is a rare but serious consequence of the locoregional metastatic event. The present work aimed to study the effect of the Hsp90 inhibitor (17-AAG), an ansamycin analog, on cell cycle and DNA replication specific chaperone-clients interaction in the event of peritoneal sarcoma related malignant ascites formation in mouse model at the late stage of malignant growth. We administered 17-AAG, an Hsp90 inhibitor, divided doses (330 μg/kg b.w./day for first five days then next ten days with166 μg/kg b.w./day) through intra-peritoneal route of inbred Swiss albino mice bearing full grown peritoneal malignant ascites of sarcoma-180. Our study was evaluated by peripheral blood hemogram analysis, malignant ascitic cytology, cell viability test, survival time and mitotic indexing. Furthermore, flowcytometric HSP90, TERT, CyclinD1, PCNA and GM-CSF expression analysis has been considered for special objective of the study. Our experimental efforts reduced the aggressive proliferation of malignant ascites by drastic downregulation of TERT and cyclin D1 on the verge of cell cycle entry along with DNA replication processivity factor PCNA by directly modulating their folding machinery - heat shock protein 90. Consequently, we observed that malignant ascitic cells became error prone during the event of karyokinesis and produced micronucleus containing malignant cells with low viability. Peripheral neutrophilia due to over-expression of GM-CSF by the peritoneal malignant ascites were also controlled by the treatment with 17-AAG and overall, the treatment modality improved the median survival time. Finally we can conclude that 17AAG administration might serve as a prospective pharmacological agent for the management of peritoneal sarcoma related malignant ascites and throws light towards prolonged survival of the patients concerned.

  19. Role of Cyclin D1 and cdk Inhibitors in Breast Cancer Pathogenesis

    National Research Council Canada - National Science Library

    Roovers, Kristin

    2002-01-01

    Cyclin Dl is frequently overexpressed in breast cancer tumors. Thus, an understanding of cyclin Dl regulation in normal and malignant cells may prove beneficial in the search for therapeutic targets...

  20. Cyclin-dependent kinase inhibition by the KLF6 tumor suppressor protein through interaction with cyclin D1.

    Science.gov (United States)

    Benzeno, Sharon; Narla, Goutham; Allina, Jorge; Cheng, George Z; Reeves, Helen L; Banck, Michaela S; Odin, Joseph A; Diehl, J Alan; Germain, Doris; Friedman, Scott L

    2004-06-01

    Kruppel-like factor 6 (KLF6) is a tumor suppressor gene inactivated in prostate and colon cancers, as well as in astrocytic gliomas. Here, we establish that KLF6 mediates growth inhibition through an interaction with cyclin D1, leading to reduced phosphorylation of the retinoblastoma protein (Rb) at Ser(795). Furthermore, introduction of KLF6 disrupts cyclin D1-cyclin-dependent kinase (cdk) 4 complexes and forces the redistribution of p21(Cip/Kip) onto cdk2, which promotes G(1) cell cycle arrest. Our data suggest that KLF6 converges with the Rb pathway to inhibit cyclin D1/cdk4 activity, resulting in growth suppression.

  1. Nicotine induces cell proliferation in association with cyclin D1 up-regulation and inhibits cell differentiation in association with p53 regulation in a murine pre-osteoblastic cell line

    International Nuclear Information System (INIS)

    Sato, Tsuyoshi; Abe, Takahiro; Nakamoto, Norimichi; Tomaru, Yasuhisa; Koshikiya, Noboru; Nojima, Junya; Kokabu, Shoichiro; Sakata, Yasuaki; Kobayashi, Akio; Yoda, Tetsuya

    2008-01-01

    Recent studies have suggested that nicotine critically affects bone metabolism. Many studies have examined the effects of nicotine on proliferation and differentiation, but the underlying molecular mechanisms remain unclear. We examined cell cycle regulators involved in the proliferation and differentiation of MC3T3-E1 cells. Nicotine induced cell proliferation in association with p53 down-regulation and cyclin D1 up-regulation. In differentiated cells, nicotine reduced alkaline phosphatase activity and mineralized nodule formation in dose-dependent manners. Furthermore, p53 expression was sustained in nicotine-treated cells during differentiation. These findings indicate that nicotine promotes the cell cycle and inhibits differentiation in association with p53 regulation in pre-osteoblastic cells

  2. The Role of Cyclin D1 in Altering Stromal-Epithelial Interactions in Prostate Carcinogenesis

    Science.gov (United States)

    2008-03-01

    Alarid ET, Turner T, Donjacour AA, Boutin EL, Foster BA. Normal and abnormal development of the male urogenital tract : role of androgens, mesenchymal...expression in both epithelium and stroma by retroviral infection and examined the role of cyclin D1 using a tissue recombination model. The data showed...overexpressed Cyclin D1 in BPH-1 cells by retroviral infection to generate BPH-1C7-cyclin D1 cell lines. The consequences of cyclin D1 expression was examined

  3. Autocrine parathyroid hormone-like hormone promotes intrahepatic cholangiocarcinoma cell proliferation via increased ERK/JNK-ATF2-cyclinD1 signaling

    Directory of Open Access Journals (Sweden)

    Jing Tang

    2017-11-01

    Full Text Available Abstract Background and aims Intrahepatic cholangiocarcinoma (ICC is an aggressive tumor with a high fatality rate. It was recently found that parathyroid hormone-like hormone (PTHLH was frequently overexpressed in ICC compared with non-tumor tissue. This study aimed to elucidate the underlying mechanisms of PTHLH in ICC development. Methods The CCK-8 assay, colony formation assays, flow cytometry and a xenograft model were used to examine the role of PTHLH in ICC cells proliferation. Immunohistochemistry (IHC and western blot assays were used to detect target proteins. Luciferase reporter, chromatin immunoprecipitation (ChIP and DNA pull-down assays were used to verify the transcription regulation of activating transcription factor-2 (ATF2. Results PTHLH was significantly upregulated in ICC compared with adjacent and normal tissues. Upregulation of PTHLH indicated a poor pathological differentiation and intrahepatic metastasis. Functional study demonstrated that PTHLH silencing markedly suppressed ICC cells growth, while specific overexpression of PTHLH has the opposite effect. Mechanistically, secreted PTHLH could promote ICC cell growth by activating extracellular signal-related kinase (ERK and c-Jun N-terminal kinase (JNK signaling pathways, and subsequently upregulated ATF2 and cyclinD1 expression. Further study found that the promoter activity of PTHLH were negatively regulated by ATF2, indicating that a negative feedback loop exists. Conclusions Our findings demonstrated that the ICC-secreted PTHLH plays a characteristic growth-promoting role through activating the canonical ERK/JNK-ATF2-cyclinD1 signaling pathways in ICC development. We identified a negative feedback loop formed by ATF2 and PTHLH. In this study, we explored the therapeutic implication for ICC patients.

  4. Urinary bladder lesions after the chernobyl accident. Immunohistochemical assessment of p53, proliferating cell nuclear antigen, cyclin D1 and p21[sup WAF1/Cip1

    Energy Technology Data Exchange (ETDEWEB)

    Romanenko, A.; Zaparin, W.; Vinnichenko, W.; Vozianov, A. (Academy of Medical Sciences of Ukraine, Kiev (Ukraine)); Lee, C.C.R.; Yamamoto, Shinji; Hori, Taka-aki; Wanibuchi, Hideki; Fukushima, Shoji

    1999-02-01

    During the 11-year period subsequent to the Chernobyl accident, the incidence of urinary bladder cancer in Ukraine has increased from 26.2 to 36.1 per 100,000 population. Cesium-137 ([sup 137]Cs) accounts for 80-90% of the incorporated radioactivity in this population, which has been exposed to long-term, low-dose ionizing radiation, and 80% of the more labile pool of cesium is excreted via the urine. The present study was performed to evaluate the histopathological features and the immunohistochemical status of p53, p21[sup WAF1/Cip1], cyclin D1 and PCNA (proliferating cell nuclear antigen) in urinary bladder mucosa of 55 males (49-92 years old) with benign prostatic hyperplasia who underwent surgery in Kiev, Ukraine, in 1995 and 1996. Group I (28 patients) inhabiting radiocontaminated areas of the country, group II (17 patients) from Kiev city with less radiocontamination and a control group III (10 patients) living in so-called ''clean'' areas of Ukraine were compared. In groups I and II, an increase in multiple areas of moderate or severe dysplasia or carcinoma in situ was seen in 42 (93%) of 45 cases. In addition, two small transitional cell carcinomas were found in one patient in each of groups I and II. Nuclear accumulation of p53, PCNA, cyclin D1, and to a lesser extent p21[sup WAF1/Cip1], was significantly increased in both groups I and II as compared with the control group III, indicating possible transformation events or enhancement of repair activities, that may precede the defect in the regulatory pathway itself, at least in the G1 phase of the cell cycle. Our results suggest that early malignant transformation is taking place in the bladder urothelium of people in the radiocontaminated areas of Ukraine and that this could possibly lead sometime in the future to an increased incidence of urinary bladder cancer. (author)

  5. X-linked inhibitor of apoptosis protein (XIAP) regulation of cyclin D1 protein expression and cancer cell anchorage-independent growth via its E3 ligase-mediated protein phosphatase 2A/c-Jun axis.

    Science.gov (United States)

    Cao, Zipeng; Zhang, Ruowen; Li, Jingxia; Huang, Haishan; Zhang, Dongyun; Zhang, Jingjie; Gao, Jimin; Chen, Jingyuan; Huang, Chuanshu

    2013-07-12

    The X-linked inhibitor of apoptosis protein (XIAP) is a well known potent inhibitor of apoptosis; however, it is also involved in other cancer cell biological behavior. In the current study, we discovered that XIAP and its E3 ligase played a crucial role in regulation of cyclin D1 expression in cancer cells. We found that deficiency of XIAP expression resulted in a marked reduction in cyclin D1 expression. Consistently, cell cycle transition and anchorage-independent cell growth were also attenuated in XIAP-deficient cancer cells compared with those of the parental wild-type cells. Subsequent studies demonstrated that E3 ligase activity within the RING domain of XIAP is crucial for its ability to regulate cyclin D1 transcription, cell cycle transition, and anchorage-independent cell growth by up-regulating transactivation of c-Jun/AP-1. Moreover, we found that E3 ligase within RING domain was required for XIAP inhibition of phosphatase PP2A activity by up-regulation of PP2A phosphorylation at Tyr-307 in its catalytic subunit. Such PP2A phosphorylation and inactivation resulted in phosphorylation and activation of its downstream target c-Jun in turn leading to cyclin D1 expression. Collectively, our studies uncovered a novel function of E3 ligase activity of XIAP in the up-regulation of cyclin D1 expression, providing significant insight into the understanding of the biomedical significance of overexpressed XIAP in cancer development, further offering a new molecular basis for utilizing XIAP E3 ligase as a cancer therapeutic target.

  6. Distinction between Asymptomatic Monoclonal B-cell Lymphocytosis with Cyclin D1 Overexpression and Mantle Cell Lymphoma: From Molecular Profiling to Flow Cytometry

    Science.gov (United States)

    Espinet, Blanca; Ferrer, Ana; Bellosillo, Beatriz; Nonell, Lara; Salar, Antonio; Fernández-Rodríguez, Concepción; Puigdecanet, Eulàlia; Gimeno, Javier; Garcia-Garcia, Mar; Carmen Vela, Maria; Luño, Elisa; Collado, Rosa; Navarro, José Tomás; de la Banda, Esmeralda; Abrisqueta, Pau; Arenillas, Leonor; Serrano, Cristina; Lloreta, Josep; Miñana, Belén; Cerutti, Andrea; Florensa, Lourdes; Orfao, Alberto; Sanz, Ferran; Solé, Francesc; Dominguez-Sola, David; Serrano, Sergio

    2015-01-01

    Purpose According to current diagnostic criteria, mantle cell lymphoma (MCL) encompasses the usual, aggressive variants and rare, nonnodal cases with monoclonal asymptomatic lymphocytosis, cyclin D1–positive (MALD1). We aimed to understand the biology behind this clinical heterogeneity and to identify markers for adequate identification of MALD1 cases. Experimental Design We compared 17 typical MCL cases with a homogeneous group of 13 untreated MALD1 cases (median follow-up, 71 months). We conducted gene expression profiling with functional analysis in five MCL and five MALD1. Results were validated in 12 MCL and 8 MALD1 additional cases by quantitative reverse transcription polymerase chain reaction (qRT-PCR) and in 24 MCL and 13 MALD1 cases by flow cytometry. Classification and regression trees strategy was used to generate an algorithm based on CD38 and CD200 expression by flow cytometry. Results We found 171 differentially expressed genes with enrichment of neoplastic behavior and cell proliferation signatures in MCL. Conversely, MALD1 was enriched in gene sets related to immune activation and inflammatory responses. CD38 and CD200 were differentially expressed between MCL and MALD1 and confirmed by flow cytometry (median CD38, 89% vs. 14%; median CD200, 0% vs. 24%, respectively). Assessment of both proteins allowed classifying 85% (11 of 13) of MALD1 cases whereas 15% remained unclassified. SOX11 expression by qRT-PCR was significantly different between MCL and MALD1 groups but did not improve the classification. Conclusion We show for the first time that MALD1, in contrast to MCL, is characterized by immune activation and driven by inflammatory cues. Assessment of CD38/CD200 by flow cytometry is useful to distinguish most cases of MALD1 from MCL in the clinical setting. MALD1 should be identified and segregated from the current MCL category to avoid overdiagnosis and unnecessary treatment. PMID:24352646

  7. Gomisin G Inhibits the Growth of Triple-Negative Breast Cancer Cells by Suppressing AKT Phosphorylation and Decreasing Cyclin D1.

    Science.gov (United States)

    Maharjan, Sony; Park, Byoung Kwon; Lee, Su In; Lim, Yoonho; Lee, Keunwook; Kwon, Hyung-Joo

    2018-03-28

    A type of breast cancer with a defect in three molecular markers such as the estrogen receptor, progesterone receptor, and human epidermal growth factor receptor is called triple-negative breast cancer (TNBC). Many patients with TNBC have a lower survival rate than patients with other types due to a poor prognosis. In this study, we confirmed the anti-cancer effect of a natural compound, Gomisin G, in TNBC cancer cells. Treatment with Gomisin G suppressed the viability of two TNBC cell lines, MDA-MB-231 and MDA-MB-468 but not non-TNBC cell lines such as MCF-7, T47D, and ZR75-1. To investigate the molecular mechanism of this activity, we examined the signal transduction pathways after treatment with Gomisin G in MDA-MB-231 cells. Gomisin G did not induce apoptosis but drastically inhibited AKT phosphorylation and reduced the amount of retinoblastoma tumor suppressor protein (Rb) and phosphorylated Rb. Gomisin G induced in a proteasome-dependent manner a decrease in Cyclin D1. Consequently, Gomisin G causes cell cycle arrest in the G1 phase. In contrast, there was no significant change in T47D cells except for a mild decrease in AKT phosphorylation. These results show that Gomisin G has an anti-cancer activity by suppressing proliferation rather than inducing apoptosis in TNBC cells. Our study suggests that Gomisin G could be used as a therapeutic agent in the treatment of TNBC patients.

  8. Expression of δ-cyclins of Brassica rapa L. embryos by clinorotation

    Science.gov (United States)

    Artemenko, O. A.

    Cyclins is one of the important regulators of cell cycle. There are several types of cyclins exists. They are responding for different phases of cycle and have high homology in plant's and mammalian's cells. δ -cyclins are specific for plants and controlling the presynthetic phase events. These cyclins likes to mammalian D-cyclins and have similar functions. This class consist three types of cyclins -- δ 1, δ 2 and δ 3. Cyclin δ 1 is responding for events in cell, which take place before exiting from stage of quiet (G0). Cyclin δ 1 is responding for entering and outputting from G0, and cyclin δ 3 -- for events, which happen in cell after stage of quiet, by entering to S-phase (phase of DNA's synthesis). In present research was used δ 1- and δ 3-cyclins. For determination of δ -cyclins gene's expression level was excreted RNA from embryos: 3-days (spherical stage), 6-days (heart-shaped stage) and 9-days (generated stage) seedlings of Brassica rapa L. in control and under clinorotation. For definition the cyclins gene's expression level applied Northern Blot Analysis. Obtained data testify about difference in level of gene's expression of cyclin δ 1 between control and clinorotation variants. After three days by pollination the expression of this gene in embryos was observed in control only. By clinorotation the gene's expression was detected on 6 days later, but it level was lower than in control variant. On 9 days it was gently expressed by clinorotation, where as by control it was not detected absolutely. Cyclin δ 3 gene's expression was observed during all time of the experiment. These data also confirm known one about expression δ 1- cyclin, which expressed on beginning of cell cycle only. And δ 3 --cyclin that express during whole presinthetic phase of cell cycle (Sony et al., 1995, Murray, 1994, Inze et al, 1999, Umeda, 2000).

  9. Lack of sik1 in mouse embryonic stem cells impairs cardiomyogenesis by down-regulating the cyclin-dependent kinase inhibitor p57kip2.

    Directory of Open Access Journals (Sweden)

    Antonio Romito

    Full Text Available Sik1 (salt inducible kinase 1 is a serine/threonine kinase that belongs to the stress- and energy-sensing AMP-activated protein kinase family. During murine embryogenesis, sik1 marks the monolayer of future myocardial cells that will populate first the primitive ventricle, and later the primitive atrium suggesting its involvement in cardiac cell differentiation and/or heart development. Despite that observation, the involvement of sik1 in cardiac differentiation is still unknown. We examined the sik1 function during cardiomyocyte differentiation using the ES-derived embryoid bodies. We produced a null embryonic stem cell using a gene-trap cell line carrying an insertion in the sik1 locus. In absence of the sik1 protein, the temporal appearance of cardiomyocytes is delayed. Expression profile analysis revealed sik1 as part of a genetic network that controls the cell cycle, where the cyclin-dependent kinase inhibitor p57(Kip2 is directly involved. Collectively, we provided evidence that sik1-mediated effects are specific for cardiomyogenesis regulating cardiomyoblast cell cycle exit toward terminal differentiation.

  10. Targeting cyclin-dependent kinase 1 (CDK1) but not CDK4/6 or CDK2 is selectively lethal to MYC-dependent human breast cancer cells

    International Nuclear Information System (INIS)

    Kang, Jian; Sergio, C Marcelo; Sutherland, Robert L; Musgrove, Elizabeth A

    2014-01-01

    Although MYC is an attractive therapeutic target for breast cancer treatment, it has proven challenging to inhibit MYC directly, and clinically effective pharmaceutical agents targeting MYC are not yet available. An alternative approach is to identify genes that are synthetically lethal in MYC-dependent cancer. Recent studies have identified several cell cycle kinases as MYC synthetic-lethal genes. We therefore investigated the therapeutic potential of specific cyclin-dependent kinase (CDK) inhibition in MYC-driven breast cancer. Using small interfering RNA (siRNA), MYC expression was depleted in 26 human breast cancer cell lines and cell proliferation evaluated by BrdU incorporation. MYC-dependent and MYC-independent cell lines were classified based on their sensitivity to siRNA-mediated MYC knockdown. We then inhibited CDKs including CDK4/6, CDK2 and CDK1 individually using either RNAi or small molecule inhibitors, and compared sensitivity to CDK inhibition with MYC dependence in breast cancer cells. Breast cancer cells displayed a wide range of sensitivity to siRNA-mediated MYC knockdown. The sensitivity was correlated with MYC protein expression and MYC phosphorylation level. Sensitivity to siRNA-mediated MYC knockdown did not parallel sensitivity to the CDK4/6 inhibitor PD0332991; instead MYC-independent cell lines were generally sensitive to PD0332991. Cell cycle arrest induced by MYC knockdown was accompanied by a decrease in CDK2 activity, but inactivation of CDK2 did not selectively affect the viability of MYC-dependent breast cancer cells. In contrast, CDK1 inactivation significantly induced apoptosis and reduced viability of MYC-dependent cells but not MYC- independent cells. This selective induction of apoptosis by CDK1 inhibitors was associated with up-regulation of the pro-apoptotic molecule BIM and was p53-independent. Overall, these results suggest that further investigation of CDK1 inhibition as a potential therapy for MYC-dependent breast cancer

  11. The pterocarpanquinone LQB-118 inhibits tumor cell proliferation by downregulation of c-Myc and cyclins D1 and B1 mRNA and upregulation of p21 cell cycle inhibitor expression.

    Science.gov (United States)

    Martino, Thiago; Magalhães, Fernanda C J; Justo, Graça A; Coelho, Marsen G P; Netto, Chaquip D; Costa, Paulo R R; Sabino, Kátia C C

    2014-06-15

    The incidence of cancer grows annually worldwide and in Brazil it is the second cause of death. The search for anti-cancer drugs has then become urgent. It depends on the studies of natural and chemical synthesis products. The antitumor action of LQB-118, a pterocarpanquinone structurally related to lapachol, has been demonstrated to induce mechanisms linked to leukemia cell apoptosis. This work investigated some mechanisms of the in vitro antitumor action of LQB-118 on prostate cancer cells. LQB-118 reduced the expression of the c-Myc transcription factor, downregulated the cyclin D1 and cyclin B1 mRNA levels and upregulated the p21 cell cycle inhibitor. These effects resulted in cell cycle arrest in the S and G2/M phases and inhibition of tumor cell proliferation. LQB-118 also induced programmed cell death of the prostate cancer cells, as evidenced by internucleosomal DNA fragmentation and annexin-V positive cells. Except the cell cycle arrest in the S phase and enhanced c-Myc expression, all the mechanisms observed here for the in vitro antitumor action of LQB-118 were also found for Paclitaxel, a traditional antineoplastic drug. These findings suggest new molecular mechanisms for the LQB-118 in vitro antitumor action. Copyright © 2014 Elsevier Ltd. All rights reserved.

  12. Cyclin A2 regulates erythrocyte morphology and numbers.

    Science.gov (United States)

    Jayapal, Senthil Raja; Ang, Heather Yin-Kuan; Wang, Chelsia Qiuxia; Bisteau, Xavier; Caldez, Matias J; Xuan, Gan Xiao; Yu, Weimiao; Tergaonkar, Vinay; Osato, Motomi; Lim, Bing; Kaldis, Philipp

    2016-11-16

    Cyclin A2 is an essential gene for development and in haematopoietic stem cells and therefore its functions in definitive erythropoiesis have not been investigated. We have ablated cyclin A2 in committed erythroid progenitors in vivo using erythropoietin receptor promoter-driven Cre, which revealed its critical role in regulating erythrocyte morphology and numbers. Erythroid-specific cyclin A2 knockout mice are viable but displayed increased mean erythrocyte volume and reduced erythrocyte counts, as well as increased frequency of erythrocytes containing Howell-Jolly bodies. Erythroblasts lacking cyclin A2 displayed defective enucleation, resulting in reduced production of enucleated erythrocytes and increased frequencies of erythrocytes containing nuclear remnants. Deletion of the Cdk inhibitor p27 Kip1 but not Cdk2, ameliorated the erythroid defects resulting from deficiency of cyclin A2, confirming the critical role of cyclin A2/Cdk activity in erythroid development. Loss of cyclin A2 in bone marrow cells in semisolid culture prevented the formation of BFU-E but not CFU-E colonies, uncovering its essential role in BFU-E function. Our data unveils the critical functions of cyclin A2 in regulating mammalian erythropoiesis.

  13. The effect of miR-338-3p on HBx deletion-mutant (HBx-d382 mediated liver-cell proliferation through CyclinD1 regulation.

    Directory of Open Access Journals (Sweden)

    Xiaoyu Fu

    Full Text Available Hepatitis B Virus (HBV DNA integration and HBV X (HBx deletion mutation occurs in HBV-positive liver cancer patients, and C-terminal deletion in HBx gene mutants are highly associated with hepatocarcinogenesis. Our previous study found that the HBx-d382 deletion mutant (deleted at nt 382-400 can down-regulate miR-338-3p expression in HBx-expressing cells. The aim of the present study is to examine the role of miR-338-3p in the HBx-d382-mediated liver-cell proliferation.We established HBx-expressing LO2 cells by Lipofectamine 2000 transfection. A miR-338-3p mimics or inhibitor was transfected into LO2/HBx-d382 and LO2/HBx cells using miR-NC as a control miRNA. In silico analysis of potential miR-338-3p targets revealed that miR-338-3p could target the cell cycle regulatory protein CyclinD1. To confirm that CyclinD1 is negatively regulated by miR-338-3p, we constructed luciferase reporters with wild-type and mutated CyclinD1-3'UTR target sites for miR-338-3p binding. We examined the CyclinD1 expression by real-time PCR and western blot, and proliferation activity by flow cytometric cell cycle analysis, Edu incorporation, and soft agar colony.HBx-d382 exhibited enhanced proliferation and CyclinD1 expression in LO2 cells. miR-338-3p expression inhibited cell proliferation in LO2/HBx-d382 cells (and LO2/HBx cells, and also negatively regulated CyclinD1 protein expression. Of the two putative miR-338-3p binding sites in the CyclinD1-3'UTR region, the effect of miR-338-3p on the second binding site (nt 2397-2403 was required for the inhibition.miR-338-3p can directly regulate CyclinD1 expression through binding to the CyclinD1-3'UTR region, mainly at nt 2397-2403. Down-regulation of miR-338-3p expression is required for liver cell proliferation in both LO2/HBx and LO2/HBx-d382 mutant cells, although the effect is more pronounced in LO2/HBx-d382 cells. Our study elucidated a novel mechanism, from a new miRNA-regulation perspective, underlying the

  14. Cyclin A2 promotes DNA repair in the brain during both development and aging

    OpenAIRE

    Gygli, Patrick E.; Chang, Joshua C.; Gokozan, Hamza N.; Catacutan, Fay P.; Schmidt, Theresa A.; Kaya, Behiye; Goksel, Mustafa; Baig, Faisal S.; Chen, Shannon; Griveau, Amelie; Michowski, Wojciech; Wong, Michael; Palanichamy, Kamalakannan; Sicinski, Piotr; Nelson, Randy J.

    2016-01-01

    Various stem cell niches of the brain have differential requirements for Cyclin A2. Cyclin A2 loss results in marked cerebellar dysmorphia, whereas forebrain growth is retarded during early embryonic development yet achieves normal size at birth. To understand the differential requirements of distinct brain regions for Cyclin A2, we utilized neuroanatomical, transgenic mouse, and mathematical modeling techniques to generate testable hypotheses that provide insight into how Cyclin A2 loss resu...

  15. Transforming growth factor-β1 induces cell cycle arrest by activating atypical cyclin-dependent kinase 5 through up-regulation of Smad3-dependent p35 expression in human MCF10A mammary epithelial cells.

    Science.gov (United States)

    Park, Seong Ji; Yang, Sun Woo; Kim, Byung-Chul

    2016-04-08

    Cyclin-dependent kinases (Cdks) play important roles in control of cell division. Cdk5 is an atypical member of Cdk family with non-cyclin-like regulatory subunit, p35, but its role in cell cycle progression is still unclear. In the present study, we investigated the role of Cdk5/p35 on transforming growth factor-β1 (TGF-β1)-induced cell cycle arrest. In human MCF10A mammary epithelial cells, TGF-β1 induced cell cycle arrest at G1 phase and increased p27KIP1 expression. Interestingly, pretreatment with roscovitine, an inhibitor of Cdk5, or transfection with small interfering (si) RNAs specific to Cdk5 and p35 significantly attenuated the TGF-β1-induced p27KIP1 expression and cell cycle arrest. TGF-β1 increased Cdk5 activity via up-regulation of p35 gene at transcriptional level, and these effects were abolished by transfection with Smad3 siRNA or infection of adenovirus carrying Smad3 mutant at the C-tail (3SA). Chromatin immunoprecipitation assay further revealed that wild type Smad3, but not mutant Smad3 (3SA), binds to the region of the p35 promoter region (-1000--755) in a TGF-β1-dependent manner. These results for the first time demonstrate a role of Cdk5/p35 in the regulation of cell cycle progression modulated by TGF-β1. Copyright © 2016 Elsevier Inc. All rights reserved.

  16. Involvement of cdks and cyclins in muscle differentiation

    Directory of Open Access Journals (Sweden)

    M De Falco

    2009-06-01

    Full Text Available Myocyte differentiation is due to transcription of genes that characterize the phenotypic and biochemical identity of differentiated muscle cells. These are the myogenic regulatory factors (MRFs MyoD, Myf5, myogenin and MRF4. Overexpression of cdk/cyclins has been reported to inhibit the activity of MyoD and prevent myogenic differentiation by different modalities. Unlike other cdk/cyclin complexes, overexpression of cdk9/cyclin T2a, enhances MyoD function and promotes myogenic differentiation. In addition, cyclin T2a interacting with a novel partner, PKNa, is able to strongly enhance the expression of myogenic differentiation markers, such as myogenin and Myosin Heavy Chain. So, cyclin T2a could stimulate myogenic differentiation interacting with different kinase partners Cdk9 or PKNa in a synergistic or antagonistic way.

  17. dp53 Restrains ectopic neural stem cell formation in the Drosophila brain in a non-apoptotic mechanism involving Archipelago and cyclin E.

    Directory of Open Access Journals (Sweden)

    Yingshi Ouyang

    Full Text Available Accumulating evidence suggests that tumor-initiating stem cells or cancer stem cells (CSCs possibly originating from normal stem cells may be the root cause of certain malignancies. How stem cell homeostasis is impaired in tumor tissues is not well understood, although certain tumor suppressors have been implicated. In this study, we use the Drosophila neural stem cells (NSCs called neuroblasts as a model to study this process. Loss-of-function of Numb, a key cell fate determinant with well-conserved mammalian counterparts, leads to the formation of ectopic neuroblasts and a tumor phenotype in the larval brain. Overexpression of the Drosophila tumor suppressor p53 (dp53 was able to suppress ectopic neuroblast formation caused by numb loss-of-function. This occurred in a non-apoptotic manner and was independent of Dacapo, the fly counterpart of the well-characterized mammalian p53 target p21 involved in cellular senescence. The observation that dp53 affected Edu incorporation into neuroblasts led us to test the hypothesis that dp53 acts through regulation of factors involved in cell cycle progression. Our results show that the inhibitory effect of dp53 on ectopic neuroblast formation was mediated largely through its regulation of Cyclin E (Cyc E. Overexpression of Cyc E was able to abrogate dp53's ability to rescue numb loss-of-function phenotypes. Increasing Cyc E levels by attenuating Archipelago (Ago, a recently identified transcriptional target of dp53 and a negative regulator of Cyc E, had similar effects. Conversely, reducing Cyc E activity by overexpressing Ago blocked ectopic neuroblast formation in numb mutant. Our results reveal an intimate connection between cell cycle progression and NSC self-renewal vs. differentiation control, and indicate that p53-mediated regulation of ectopic NSC self-renewal through the Ago/Cyc E axis becomes particularly important when NSC homeostasis is perturbed as in numb loss-of-function condition. This has

  18. Radioiodination of cyclin dependent kinase inhibitor Olomoucine loaded Fe rate at Au nanoparticle and evaluation of the therapeutic efficacy on cancerous cells

    Energy Technology Data Exchange (ETDEWEB)

    Takan, Gokhan; Guldu, Ozge Kozgus; Medine, Emin Ilker [Ege Univ., Izmir (Turkey). Dept. of Nuclear Applications

    2017-06-01

    Magnetic nanoparticles have promising biomedical applications such as drug delivery, novel therapeutics and diagnostic imaging. Magnetic drug delivery combination works on the delivery of magnetic nanoparticles loaded with drug to the target tissue by means of an external magnetic field. Gold coated iron oxide (Fe rate at Au) nanoparticles can provide useful surface chemistry and biological reactivity. Covalent conjugation to the Fe rate at Au nanoparticles through cleavable linkages can be used to deliver drugs to tumor cells, then the drug can be released by an external. In this paper, purine based cyclin dependent kinases (CDKs) inhibitor Olomoucine (Olo) [2-(Hydroxyethylamino)-6-benzylamino-9-methylpurine] was loaded on gold coated iron oxide (Fe rate at Au) nanoparticles and radiolabeled with {sup 131}I to combine magnetic targeted drug delivery and radiotherapy. Fe rate at Au nanoparticles were synthesized by microemulsion method. The characterization of nanoparticles was examined by TEM, VSM and XRD. Amine activation was utilized by cysteamine hydrochloride and then CDI was used for conjugation of Olomoucine. Antiproliferative effect and cytotoxicity of Olomoucine loaded Fe rate at Au nanoparticles (Fe rate at Au-Olo) were investigated on MCF7 and A549 cell lines. Proliferation rate was decreased while uptake of Fe rate at Au-Olo on both cell lines was high in comparison with Olomoucine. Also, enhanced incorporation ratio was observed under external magnetic field.

  19. Molecular analysis of Bcl-2 and cyclin D1 expression in differentially expressing estrogen receptor breast cancer MCF7, T47D and MDA-MB-468 cell lines treated with adriamycin

    Directory of Open Access Journals (Sweden)

    2008-08-01

    Full Text Available Background and purpose of the study: Bcl-2 and Cyclin D1 (CCND1 are key elements in cancer development and progression. Bcl-2 acts as a cell death suppressor and is involved in apoptosis regulation. Cyclin D1 is an important regulator of G1/S phase of the cell cycle progression. In addition, estrogen receptor (ER is an important prognostic factor in breast cancer cells. Therefore it is important to determine the Bcl-2 and CCND1 expression in MCF7, T47D and MDA-MB-468 breast cancer cell lines with different ER status following Adriamycin (ADR treatment. Methods: Cytotoxicity of ADR (250 and 500nM after 1-5 days exposure of the cell lines was evaluated by MTT assay. The mRNA and protein levels of Bcl-2 and cyclin D1 in tested cell lines were also analyzed by RT-PCR and immunocytochemistry (ICC methods. Results: ADR cytotoxicity was highest in MDA-MB-468 and lowest in MCF7 cells in a time-dependent manner. Bcl-2 mRNA increased in MCF7 and decreased in MDA-MB-468 after exposure to ADR but it was less detectable in T47D cells. The expression of CCND1 in MCF7 with high level of ER expression was higher than the other two cell lines in untreated conditions. However, CCND1 mRNA did not show significant changes after ADR treatment. Immunocytochemical analysis did not show significant differences between Bcl-2 protein expression in the presence or absence of ADR in MDA-MB-468 cell line while in T47D and MCF7 cells its expression decreased after exposure to ADR. In addition to nuclear expression of cyclin D1 in all cell lines, strong cytoplasmic expression of cyclin D1 protein was observed only in MCF7 and T47D cells. Conclusion: The tested cell lines with different levels of ER expression showed differential molecular responses to ADR that is important in tumor-targeted cancer therapy.

  20. Adenovirus-mediated gene transfer of MMAC1/PTEN to glioblastoma cells inhibits S phase entry by the recruitment of p27Kip1 into cyclin E/CDK2 complexes.

    Science.gov (United States)

    Cheney, I W; Neuteboom, S T; Vaillancourt, M T; Ramachandra, M; Bookstein, R

    1999-05-15

    Genetic alterations in the MMAC1 tumor suppressor gene (also referred to as PTEN or TEP1) occur in several types of human cancers including glioblastoma. Growth suppression induced by overexpression of MMAC1 in cells with mutant MMAC1 alleles is thought to be mediated by the inhibition of signaling through the phosphatidylinositol 3-kinase pathway. However, the exact biochemical mechanisms by which MMAC1 exerts its growth-inhibitory effects are still unknown. Here we report that recombinant adenovirus-mediated overexpression of MMAC1 in three different MMAC1-mutant glioblastoma cell lines blocked progression from G0/G1 to S phase of the cell cycle. Cell cycle arrest correlated with the recruitment of the cyclin-dependent kinase (CDK) inhibitor, p27Kip1, to cyclin E immunocomplexes, which resulted in a reduction in CDK2 kinase activities and a decrease in levels of endogenous phosphorylated retinoblastoma protein. CDK4 kinase activities were unaffected, as were the levels of the CDK inhibitor p21Cip1 present in cyclin E immunocomplexes. Therefore, overexpression of MMAC1 via adenovirus-mediated gene transfer suppresses tumor cell growth through cell cycle inhibitory mechanisms, and as such, represents a potential therapeutic approach to treating glioblastomas.

  1. The Role of Cyclins and Cyclins Inhibitors in the Multistep Process of HPV-Associated Cervical Carcinoma

    International Nuclear Information System (INIS)

    Bahnassy, A.A.; Mokhtar, N.M.; Zekri, A.; Alam El-Din, H.M.; Aboubaker, A.A.; Kamel, K.; El-Sabah, M.T.

    2006-01-01

    Background: Human papillomavirus (HPV) types 16 and 18 are associated with cervical carcinogenesis. This is possibly achieved through an interaction between HPV oncogenic proteins and some cell cycle regulatory genes. However, the exact pathogenetic mechanisms are not well defined yet. Methods: We investigated 110 subjects (43 invasive squamous cell carcinoma [ISCC], 38 CIN Ill, II CIN II, 18 CIN I) confirmed to be positive for HPV 16 and/or 18 as well as 20 normal cervical tissue (NCT) samples for abnormal expression of cyclin DJ, cyclin E, CDK4, cyclin inhibitors (p2Jwa/; p27, pI6/NK4A) and Ki-67 using immunohistochemistry and differential PCR techniques. Results: There was a significant increase in the expression of Ki-67, cyclin E, CDK4, pJ6/NK4A (p=0003, 0.001,0.001) and a significant decrease in p27K1P/ from NCT to ISCC (p=0.003). There was a significant correlation between altered expression of p27K1P I and p 161NK4A (p KIpl (ρ=0.011) in all studied groups In ISCC, there was significant relationship between standard clinico-pathological prognostic factors and high Ki-67 index, increased cyclin D J and cyclin E, reduced p2 7Kip / and p21 waf Conclusion: I) Aberrations involving p27K/P 1, cyclin E, CDK4 and pJ6/NK4A are considered early events in HPV 16 and IS-associated cervical carcinogenesis (CINI and lI), whereas cyclin DI aberrations are late events (CINIII and ISCC). 2) immunohistochemical tests for pJ61NK4A and cyclin E could help in early diagnosis of cervical carcinoma. 3) Only FIGO stage, cyclin DI, p27K1P1 and Ki-67 are independent prognostic factors that might help in predicting outcome of cervical cancer palients

  2. WOX5 Suppresses CYCLIN D Activity to Establish Quiescence at the Center of the Root Stem Cell Niche

    NARCIS (Netherlands)

    Forzani, C.; Aichinger, E.; Willemsen, V.; Murray, J.A.

    2014-01-01

    In Arabidopsis, stem cells maintain the provision of new cells for root growth. They surround a group of slowly dividing cells named the quiescent center (QC), and, together, they form the stem cell niche (SCN). The QC acts as the signaling center of the SCN, repressing differentiation of the

  3. miR-198 Represses the Proliferation of HaCaT Cells by Targeting Cyclin D2

    Directory of Open Access Journals (Sweden)

    Jian Wang

    2015-07-01

    Full Text Available Background: MiR-198 has been considered as an inhibitor of cell proliferation, invasion, migration and a promoter of apoptosis in most cancer cells, while its effect on non-cancer cells is poorly understood. Methods: The effect of miR-198 transfection on HaCaT cell proliferation was firstly detected using Cell Count Kit-8 and the cell cycle progression was analyzed by flow cytometry. Using bioinformatics analyses and luciferase assay, a new target of miR-198 was searched and identified. Then, the effect of the new target gene of miR-198 on cell proliferation and cell cycle was also detected. Results: Here we showed that miR-198 directly bound to the 3′-UTR of CCND2 mRNA, which was a key regulator in cell cycle progression. Overexpressed miR-198 repressed CCND2 expression at mRNA and protein levels and subsequently led to cell proliferation inhibition and cell cycle arrest in the G1 phase. Transfection ofSiCCND2 in HaCaT cells showed similar inhibitory effects on cell proliferation and cell cycle progression. Conclusion: In conclusion, we have identified that miR-198 inhibited HaCaT cell proliferation by directly targeting CCND2.

  4. Expression of cyclin D{sub 1} during endotoxin-induced aleveolar type II cell hyperplasia in rat lung and the detection of apoptotic cells during the remodeling process

    Energy Technology Data Exchange (ETDEWEB)

    Tesfaigzi, J.; Wood, M.B.; Johnson, N.F.

    1995-12-01

    Our studies have shown that endotoxin intratracheally instilled into the rat lung induces proliferation of alveolar type II cells. In that study, the alveolar type II cells. In that study, the alveolar type II cell hyperplasia occurred 2 d after instillation of endotoxin and persisted for a further 2 d. After hyperplasia, the lung remodeled and returned to a normal state within 24-48 h. Understanding the mechanisms involved in the remodeling process of this transient hyperplasia may be useful to identify molecular changes that are altered in neoplasia. The purpose of the present study was to corroborate induction of epithelial cell hyperplasia by endotoxin and to delineate mechanisms involved in tissue remodeling after endotoxin-induced alveolar type II cell hyperplasia. In conclusion, immonostaining with cyclin D1 and cytokeratin shows that endotoxin induced epithelial cell proliferation and resulted in hyperplasia in the lung which persisted through 4 d post-instillation.

  5. D-type cyclins in adult human testis and testicular cancer

    DEFF Research Database (Denmark)

    Bartkova, J; Rajpert-de Meyts, E; Skakkebaek, N E

    1999-01-01

    D-type cyclins are proto-oncogenic components of the 'RB pathway', a G1/S regulatory mechanism centred around the retinoblastoma tumour suppressor (pRB) implicated in key cellular decisions that control cell proliferation, cell-cycle arrest, quiescence, and differentiation. This study focused...... on immunohistochemical and immunochemical analysis of human adult testis and 32 testicular tumours to examine the differential expression and abundance of cyclins D1, D2, and D3 in relation to cell type, proliferation, differentiation, and malignancy. In normal testis, the cell type-restricted expression patterns were...... dominated by high levels of cyclin D3 in quiescent Leydig cells and the lack of any D-type cyclin in the germ cells, the latter possibly representing the only example of normal mammalian cells proliferating in the absence of these cyclins. Most carcinoma-in-situ lesions appeared to gain expression of cyclin...

  6. Cyclin-Dependent Kinase Inhibitor P1446A Induces Apoptosis in a JNK/p38 MAPK-Dependent Manner in Chronic Lymphocytic Leukemia B-Cells.

    Directory of Open Access Journals (Sweden)

    Cody Paiva

    Full Text Available CDK (cyclin-dependent kinase inhibitors have shown remarkable activity in CLL, where its efficacy has been linked to inhibition of the transcriptional CDKs (7 and 9 and deregulation of RNA polymerase and short-lived pro-survival proteins such as MCL1. Furthermore, ER (endoplasmic reticulum stress has been implicated in CDK inhibition in CLL. Here we conducted a pre-clinical study of a novel orally active kinase inhibitor P1446A in CLL B-cells. P1446A inhibited CDKs at nanomolar concentrations and induced rapid apoptosis of CLL cells in vitro, irrespective of chromosomal abnormalities or IGHV mutational status. Apoptosis preceded inactivation of RNA polymerase, and was accompanied by phosphorylation of stress kinases JNK (c-Jun N-terminal kinase and p38 MAPK (mitogen-activated protein kinase. Pharmacologic inhibitors of JNK/p38 MAPK conferred protection from P1446A-mediated apoptosis. Treatment with P1446A led to a dramatic induction of NOXA in a JNK-dependent manner, and sensitized CLL cells to ABT-737, a BH3-mimetic. We observed concurrent activation of apoptosis stress-inducing kinase 1 (ASK1 and its interaction with inositol-requiring enzyme 1 (IRE1 and tumor necrosis factor receptor-associated factor 2 (TRAF2 in CLL cells treated with P1446A, providing insights into upstream regulation of JNK in this setting. Consistent with previous reports on limited functionality of ER stress mechanism in CLL cells, treatment with P1446A failed to induce an extensive unfolded protein response. This study provides rationale for additional investigations of P1446A in CLL.

  7. Cdk2 Phosphorylation on Threonine39 by AKT and Its Implication on Cyclin Binding, Cellular Localization, and Cell Cycle Progression

    Science.gov (United States)

    2008-10-01

    for mammalian cell lines Cell culture MCF-7 and MEF cells were grown in Dulbeco’s improved Eagle’s medium (DMEM) supplemented with L-glutamine and...factor block was done by arresting cells (typically 400 ml grown in YPD to 8 x 106 cells/ml) for 3h at room temperature with 5 µM α-factor, 31...washing twice with 100 ml pre-warmed YPD and resuspending in the original volume of YPD . At 5 min intervals aliquots were removed, rapidly pelleted

  8. Molecular characterization and expression analysis of Cyclin B and Cell division cycle 2 in gonads of diploid and triploid bighead catfish, Clarias macrocephalus Günther, 1864

    Directory of Open Access Journals (Sweden)

    Anyalak Wachirachaikarn

    2017-04-01

    Full Text Available This study investigated the differential expression of genes associated with reproduction in sterile triploid and normal diploid bighead catfish (Clarias macrocephalus Günther, 1864. The triploid fish were produced using cold shock and were reared in the same conditions as the diploid counterpart. The histomicrographs showed completely retarded triploid gonads across the samples aged 2–12 mth, whereas the gonads of the diploids were in developing stages during 2–4 mth, reached the early maturing stage at 6 mth, matured at 8 mth and showed signs of atresia at 10–12 mth. In parallel, the full-length cDNAs of cyclin B1 (CmCcnb1; 1539 bp in length with an open reading frame (ORF of 1194 bp corresponding to 397 amino acids and cell division cycle 2 (CmCdc2; 1355 bp, an ORF of 909 bp, 302 amino acids of bighead catfish (C. macrocephalus Günther, 1864 were isolated. Phylogenetic analysis revealed that the newly characterized CmCcnb1 should be regarded as a member of cyclin B1 rather than cyclin B2. The expression level of CmCcnb1 mRNA was limited in different stages of the ovaries and testes of triploids. In diploid ovaries, its expression was significantly higher than that in triploid ovaries in fish aged 2 mth (513.43 ± 82.22 fold and in fish aged 8 mth (2430.87 ± 900.06 fold. The CmCcnb1 level in the testes of diploids was significantly greater than that in triploids in fish aged 2 mth (928.85 ± 208.72 fold. Similarly, expression of CmCdc2 mRNA was also reduced in triploids. Its expression was significantly lower than that in diploid females aged 2 mth (7.66 ± 3.42 fold, 4 mth (59.42 ± 10.50 fold and 8 mth (42.74 ± 8.36 fold. In males, significantly greater expression of CmCdc2 was observed at age 6 mth (58.61 ± 34.64 fold and 8 mth (72.70 ± 4.36 fold diploids compared to triploids. The results illustrated that CmCcnb1 and CmCdc2 are functionally involved in oogenesis and spermatogenesis and reduced

  9. Cyclin D1 overexpression modulates radiation-induced G1 arrest and apoptosis

    International Nuclear Information System (INIS)

    Su, M.; Borek, C.; Schmidt, E.V.; Pardo, F.S.

    1996-01-01

    Objective: Cyclin D1 is localized to the nucleus G1 and forms complexes with proliferating cell nuclear antigen(PCNA), p21/WAF1 protein, and cyclin dependent kinases. Since it is known that cyclin D1 is a key regulator of the G1 phase of cell cycle and G1/S regulation is important in cellular response to DNA damage, we sought to further investigate the role of cyclin D1 in the modulation of post-irradiation cell kinetics by activities of cyclin D/CDK4 complexes in cell lines which express different levels of cyclin D1. Method: Rat embryo cells (REC) at early passage underwent transfection by a standard calcium phosphate technique with pMMTVPrad-1. After transfection the cells were selected in medium containing 200μg/ml G418, and a number of individual drug-resistant clones were randomly picked and studied following clonal expansion. Following exposure to 250 kvp x-ray proteins were extracted from exponentially growing cells and were subjected to immunoprecipitation analysis using antihuman cyclin D1 antibody to detected expression of cyclin D1 and western blot analysis using antip21/WAF1 antibody, affinity-purified mAbs against PCNA and p53. The asynchronized and synchronized cells were collected and analysis for cell cycle distribution by flow cytometry. Apoptosis was assessed using an in-situ end labeling technique for fragmented DNA (Apoptag, Oncor). Results: A human cyclin D1 within a pMMTVPrad-1 expression vector was introduced by transfection into the REC. As shown high levels of theMr 35,000 cyclin D1 protein were detected in the transfected clones, i.e., CYD1-5, CYD1-6, and CYD1-pop, whereas low levels (8-10 fold) of endogenous cyclin D1 protein were seen in the mock-transfected clone. Cell cycle analyses following irradiation in vitro is illustrated cyclin D1 shown a relatively more pronounced G1 arrest following administration of 5 Gy ionizing radiation, as compared to mock-transfected cells. Cyclin D1-transfected cells showed significant levels of

  10. Stimulation of pancreatic beta-cell replication by incretins involves transcriptional induction of cyclin D1 via multiple signalling pathways

    DEFF Research Database (Denmark)

    Friedrichsen, Birgitte N; Neubauer, Nicole; Lee, Ying C

    2006-01-01

    The incretin hormones, glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic peptide (GIP), have been suggested to act as beta-cell growth factors and may therefore be of critical importance for the maintenance of a proper beta-cell mass. We have investigated the molecular mechanis...

  11. miR-338-3p Is Down-Regulated by Hepatitis B Virus X and Inhibits Cell Proliferation by Targeting the 3′-UTR Region of CyclinD1

    Directory of Open Access Journals (Sweden)

    Xiaoyu Fu

    2012-07-01

    Full Text Available Hepatitis B virus X protein (HBx is recognized as an oncogene in hepatocellular carcinoma (HCC. HBx regulates microRNA expression, including down-regulating miR-338-3p in LO2 cells. Here, we investigated miR-338-3p function in HBx-mediated hepatocarcinogenesis. In 23 HBV-infected HCC clinical patient tumor and adjacent non-tumor control tissues, 17 and 19 tumors expressed HBx mRNA and protein, respectively. When considered as a group, HBV-infected HCC tumors had lower miR-338-3p expression than controls; however, miR-338-3p was only significantly down-regulated in HBx-positive tumors, indicating that HBx inversely correlated with miR-338-3p. Functional characterization of miR-338-3p indicated that miR-338-3p mimics inhibited cell proliferation by inducing cell cycle arrest at the G1/S phase as assessed by EdU and cell cycle assays in HBx-expressing LO2 cells. CyclinD1, containing two putative miR-338-3p targets, was confirmed as a direct target using 3′-UTR luciferase reporter assays from cells transfected with mutated binding sites. Mutating the 2397–2403 nt binding site conferred the greatest resistance to miR-338-3p suppression of CyclinD1, indicating that miR-338-3p suppresses CyclinD1 at this site. Overall, this study demonstrates that miR-338-3p inhibits proliferation by regulating CyclinD1, and HBx down-regulates miR-338-3p in HCC. This newly identified miR-338-3p/CyclinD1 interaction provides novel insights into HBx-mediated hepatocarcinogenesis and may facilitate therapeutic development against HCC.

  12. Cinnamon and its Components Suppress Vascular Smooth Muscle Cell Proliferation by Up-Regulating Cyclin-Dependent Kinase Inhibitors.

    Science.gov (United States)

    Kwon, Hyeeun; Lee, Jung-Jin; Lee, Ji-Hye; Cho, Won-Kyung; Gu, Min Jung; Lee, Kwang Jin; Ma, Jin Yeul

    2015-01-01

    Cinnamomum cassia bark has been used in traditional herbal medicine to treat a variety of cardiovascular diseases. However, the antiproliferative effect of cinnamon extract on vascular smooth muscle cells (VSMCs) and the corresponding restenosis has not been explored. Hence, after examining the effect of cinnamon extract on VSMC proliferation, we investigated the possible involvement of signal transduction pathways associated with early signal and cell cycle analysis, including regulatory proteins. Besides, to identify the active components, we investigated the components of cinnamon extract on VSMC proliferation. Cinnamon extract inhibited platelet-derived growth factor (PDGF)-BB-induced VSMC proliferation and suppressed the PDGF-stimulated early signal transduction. In addition, cinnamon extract arrested the cell cycle and inhibited positive regulatory proteins. Correspondingly, the protein levels of p21 and p27 not only were increased in the presence of cinnamon extract, also the expression of proliferating cell nuclear antigen (PCNA) was inhibited by cinnamon extract. Besides, among the components of cinnamon extract, cinnamic acid (CA), eugenol (EG) and cinnamyl alcohol significantly inhibited the VSMC proliferation. Overall, the present study demonstrates that cinnamon extract inhibited the PDGF-BB-induced proliferation of VSMCs through a G0/G1 arrest, which down-regulated the expression of cell cycle positive regulatory proteins by up-regulating p21 and p27 expression.

  13. Rational design of a cyclin A fluorescent peptide sensor.

    Science.gov (United States)

    Pazos, Elena; Pérez, Miguel; Gutiérrez-de-Terán, Hugo; Orzáez, Mar; Guevara, Tatiana; Mascareñas, José L; Vázquez, M Eugenio

    2011-10-26

    We report the design and development of a fluorescent sensor specifically designed to target cyclin A, a protein that plays a key role in the regulation of the cell cycle. Computational studies provide a molecular picture that explains the observed emission increase, suggesting that the 4-DMAP fluorophore in the peptide is protected from the bulk solvent when inserted into the hydrophobic binding groove of cyclin A.

  14. Convergence of mitogenic signalling cascades from diverse classes of receptors at the cyclin D-cyclin-dependent kinase-pRb-controlled G1 checkpoint.

    OpenAIRE

    Lukas, J; Bartkova, J; Bartek, J

    1996-01-01

    The commitment of mammalian cells in late G1 to replicate the genome and divide in response to mitogenic growth factors operating via tyrosine kinase receptors depends on phosphorylation of the retinoblastoma protein (pRb), a process controlled by cyclin D-associated cyclin-dependent kinases (cdks) and their inhibitors. This study addressed the issue of whether also other mitogenic signalling cascades require activation of cyclin D-associated kinases or whether any mitogenic pathway can bypas...

  15. G1-phase regulators, cyclin D1, cyclin D2, and cyclin D3: up-regulation at gastrulation and dynamic expression during neurulation.

    Science.gov (United States)

    Wianny, F; Real, F X; Mummery, C L; Van Rooijen, M; Lahti, J; Samarut, J; Savatier, P

    1998-05-01

    Gastrulation in rodents is associated with an increase in the rate of growth and with the start of differentiation within the embryo proper. In an effort to understand the role played by the cell cycle control in these processes, expression of cyclin D1, D2, and D3--three major positive regulators of the G1/S transition--has been investigated by in situ hybrization and RT-PCR. Cyclin D1 and D2 transcripts are first detected in the epiblast at gastrulation, when a proliferative burst occurs, and subsequently in its differentiated derivatives within the embryo proper, indicating that activation of their expression takes place prior to the differentiation of epiblast progenitors. In contrast, cyclin D3 transcript is undetectable in the epiblast itself and its expression is activated exclusively in extraembryonic tissues of both epiblast and trophoblast origin. During neurulation, expression of each cyclin D RNA is dynamically regulated along the anterior-posterior axis. In the hindbrain, cyclin D1 and D2 show distinct segment-specific restricted expression and this pattern is conserved between mouse and chick. These results strongly suggest that D-type cyclins act as developmental regulators.

  16. Restriction point control of the mammalian cell cycle via the cyclin E/Cdk2:p27 complex.

    NARCIS (Netherlands)

    Conradie, R.; Bruggeman, F.J.; Ciliberto, A.; Csikasz-Nagby, A.; Novak, B.; Westerhoff, H.V.; Snoep, J.L.

    2010-01-01

    Numerous top-down kinetic models have been constructed to describe the cell cycle. These models have typically been constructed, validated and analyzed using model species (molecular intermediates and proteins) and phenotypic observations, and therefore do not focus on the individual model processes

  17. Identification of mutations that disrupt phosphorylation-dependent nuclear export of cyclin D1.

    Science.gov (United States)

    Benzeno, S; Lu, F; Guo, M; Barbash, O; Zhang, F; Herman, J G; Klein, P S; Rustgi, A; Diehl, J A

    2006-10-12

    Although cyclin D1 is overexpressed in a significant number of human cancers, overexpression alone is insufficient to promote tumorigenesis. In vitro studies have revealed that inhibition of cyclin D1 nuclear export unmasks its neoplastic potential. Cyclin D1 nuclear export depends upon phosphorylation of a C-terminal residue, threonine 286, (Thr-286) which in turn promotes association with the nuclear exportin, CRM1. Mutation of Thr-286 to a non-phosphorylatable residue results in a constitutively nuclear cyclin D1 protein with significantly increased oncogenic potential. To determine whether cyclin D1 is subject to mutations that inhibit its nuclear export in human cancer, we have sequenced exon 5 of cyclin D1 in primary esophageal carcinoma samples and in cell lines derived from esophageal cancer. Our work reveals that cyclin D1 is subject to mutations in primary human cancer. The mutations identified specifically disrupt phosphorylation of cyclin D1 at Thr-286, thereby enforcing nuclear accumulation of cyclin D1. Through characterization of these mutants, we also define an acidic residue within the C-terminus of cyclin D1 that is necessary for recognition and phosphorylation of cyclin D1 by glycogen synthase kinase-3 beta. Finally, through construction of compound mutants, we demonstrate that cell transformation by the cancer-derived cyclin D1 alleles correlates with their ability to associate with and activate CDK4. Our data reveal that cyclin D1 is subject to mutations in primary human cancer that specifically disrupt phosphorylation-dependent nuclear export of cyclin D1 and suggest that such mutations contribute to the genesis and progression of neoplastic growth.

  18. Cyclin D3 interacts with human activating transcription factor 5 and potentiates its transcription activity

    International Nuclear Information System (INIS)

    Liu Wenjin; Sun Maoyun; Jiang Jianhai; Shen Xiaoyun; Sun Qing; Liu Weicheng; Shen Hailian; Gu Jianxin

    2004-01-01

    The Cyclin D3 protein is a member of the D-type cyclins. Besides serving as cell cycle regulators, D-type cyclins have been reported to be able to interact with several transcription factors and modulate their transcriptional activations. Here we report that human activating transcription factor 5 (hATF5) is a new interacting partner of Cyclin D3. The interaction was confirmed by in vivo coimmunoprecipitation and in vitro binding analysis. Neither interaction between Cyclin D1 and hATF5 nor interaction between Cyclin D2 and hATF5 was observed. Confocal microscopy analysis showed that Cyclin D3 could colocalize with hATF5 in the nuclear region. Cyclin D3 could potentiate hATF5 transcriptional activity independently of its Cdk4 partner. But Cyclin D1 and Cyclin D2 had no effect on hATF5 transcriptional activity. These data provide a new clue to understand the new role of Cyclin D3 as a transcriptional regulator

  19. CCAAT/enhancer-binding protein beta inhibits proliferation in monocytic cells by affecting the retinoblastoma protein/E2F/cyclin E pathway but is not directly required for macrophage morphology.

    Science.gov (United States)

    Gutsch, Romina; Kandemir, Judith D; Pietsch, Daniel; Cappello, Christian; Meyer, Johann; Simanowski, Kathrin; Huber, René; Brand, Korbinian

    2011-07-01

    Monocytic differentiation is orchestrated by complex networks that are not fully understood. This study further elucidates the involvement of transcription factor CCAAT/enhancer-binding protein β (C/EBPβ). Initially, we demonstrated a marked increase in nuclear C/EBPβ-liver-enriched activating protein* (LAP*)/liver-enriched activating protein (LAP) levels and LAP/liver-enriched inhibiting protein (LIP) ratios in phorbol 12-myristate 13-acetate (PMA)-treated differentiating THP-1 premonocytic cells accompanied by reduced proliferation. To directly study C/EBPβ effects on monocytic cells, we generated novel THP-1-derived (low endogenous C/EBPβ) cell lines stably overexpressing C/EBPβ isoforms. Most importantly, cells predominantly overexpressing LAP* (C/EBPβ-long), but not those overexpressing LIP (C/EBPβ-short), exhibited a reduced proliferation, with no effect on morphology. PMA-induced inhibition of proliferation was attenuated in C/EBPβ-short cells. In C/EBPβ(WT) macrophage-like cells (high endogenous C/EBPβ), we measured a reduced proliferation/cycling index compared with C/EBPβ(KO). The typical macrophage morphology was only observed in C/EBPβ(WT), whereas C/EBPβ(KO) stayed round. C/EBPα did not compensate for C/EBPβ effects on proliferation/morphology. Serum reduction, an independent approach known to inhibit proliferation, induced macrophage morphology in C/EBPβ(KO) macrophage-like cells but not THP-1. In PMA-treated THP-1 and C/EBPβ-long cells, a reduced phosphorylation of cell cycle repressor retinoblastoma was found. In addition, C/EBPβ-long cells showed reduced c-Myc expression accompanied by increased CDK inhibitor p27 and reduced cyclin D1 levels. Finally, C/EBPβ-long and C/EBPβ(WT) cells exhibited low E2F1 and cyclin E levels, and C/EBPβ overexpression was found to inhibit cyclin E1 promoter-dependent transcription. Our results suggest that C/EBPβ reduces monocytic proliferation by affecting the retinoblastoma/E2F/cyclin E

  20. Hath1 inhibits proliferation of colon cancer cells probably through up-regulating expression of Muc2 and p27 and down-regulating expression of cyclin D1.

    Science.gov (United States)

    Zhu, Dai-Hua; Niu, Bai-Lin; Du, Hui-Min; Ren, Ke; Sun, Jian-Ming; Gong, Jian-Ping

    2012-01-01

    Previous studies showed that Math1 homologous to human Hath1 can cause mouse goblet cells to differentiate. In this context it is important that the majority of colon cancers have few goblet cells. In the present study, the potential role of Hath1 in colon carcinogenesis was investigated. Sections of paraffin-embedded tissues were used to investigate the goblet cell population of normal colon mucosa, mucosa adjacent colon cancer and colon cancer samples from 48 patients. Hath1 and Muc2 expression in these samples were tested by immunohistochemistry, quantitative real-time reverse transcription -PCR and Western blotting. After the recombinant plasmid, pcDNA3.1(+)-Hath1 had been transfected into HT29 colon cancer cells, three clones were selected randomly to test the levels of Hath1 mRNA, Muc2 mRNA, Hath1, Muc2, cyclin D1 and p27 by quantitative real-time reverse transcription-PCR and Western blotting. Moreover, the proliferative ability of HT29 cells introduced with Hath1 was assessed by means of colony formation assay and xenografting. Expression of Hath1, Muc2, cyclin D1 and p27 in the xenograft tumors was also detected by Western blotting. No goblet cells were to be found in colon cancer and levels of Hath1 mRNA and Hath1, Muc2 mRNA and Muc2 were significantly down-regulated. Hath1 could decrease cyclin D1, increase p27 and Muc2 in HT29 cells and inhibit their proliferation. Hath1 may be an anti-oncogene in colon carcinogenesis.

  1. Subversion of Cell Cycle Regulatory Mechanisms by HIV

    OpenAIRE

    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 util...

  2. K-Ras-Independent Effects of the Farnesyl Transferase Inhibitor L-744,832 on Cyclin B1/Cdc2 Kinase Activity, G2/M Cell Cycle Progression and Apoptosis in Human Pancreatic Ductal Adenocarcinoma Cell

    Directory of Open Access Journals (Sweden)

    Si Young Song

    2000-05-01

    Full Text Available Pancreatic ductal adenocarcinoma is a highly lethal malignancy that is resistant to traditional cytotoxic therapy. High rates of activating codon 12 K-Ras mutations in this disease have generated considerable interest in the therapeutic application of novel farnesyl transferase inhibitors (FTIs. However, a comprehensive analysis of the effects of FTI treatment on pancreatic cancer cells has not been performed. Treatment of five different human pancreatic cancer cell lines with FTI L744,832 resulted in inhibition of anchorage-dependent growth, with wide variation in sensitivity among different lines. Effective growth inhibition by L-744,832 correlated with accumulation of cells with a tetraploid (4N DNA content and high levels of cyclin B1/cdc2 kinase activity, implying cell cycle arrest downstream from the DNA damage -inducible G2/M cell cycle checkpoint. In addition, sensitive cell lines underwent apoptosis as evidenced by changes in nuclear morphology and internucleosomal DNA fragmentation. L-744,832 at a concentration of 1 µM additively enhanced the cytotoxic effect of ionizing radiation, apparently by overriding G2/M checkpoint activation. The effects of FTI treatment on cell growth and cell cycle regulation were associated with changes in posttranslational processing of H-Ras and N-Ras, but not K-Ras. The results confirm the potential therapeutic efficacy of FTI treatment in pancreatic cancer, and suggest that farnesylated proteins other than K-Ras may act as important regulators of G2/M cell cycle kinetics.

  3. Subversion of Cell Cycle Regulatory Mechanisms by HIV.

    Science.gov (United States)

    Rice, Andrew P; Kimata, Jason T

    2015-06-10

    To establish a productive infection, HIV-1 must counteract cellular innate immune mechanisms and redirect cellular processes toward 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 activate 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. Copyright © 2015 Elsevier Inc. All rights reserved.

  4. Cyclin A1 is a transcriptional target of PITX2 and overexpressed in papillary thyroid carcinoma.

    Science.gov (United States)

    Liu, Yan; Huang, Yue; Zhu, Guo-Zhang

    2013-12-01

    Physiological expression of cyclin A1, a unique cell cycle regulator essential for spermatogenesis, is predominantly restricted in male germ cells. Outstandingly, previous studies have also demonstrated the abnormal expression of cyclin A1 in various human tumors. How male germ cell-specific cyclin A1 is transcriptionally activated in tumor cells, however, is elusive. To begin to understand the molecular mechanisms governing the ectopic expression of cyclin A1, we searched for transcription factors and cis-regulatory DNA elements. We found that overexpression of PITX2, a paired-like homeodomain transcription factor and a downstream effector of Wnt/β-catenin signaling, resulted in upregulation of cyclin A1 in HEK293 cells and TPC-1 thyroid cancer cells. On the other hand, PITX2 knockdown in TPC-1 cells caused reduced cyclin A1. Promoter reporter assays with a series of deletion constructs determined that the DNA element from -102 to -96 bp of the cyclin A1 promoter is responsible for PITX2-induced gene expression. The result of chromatin immunoprecipitation revealed the occupancy of PITX2 on the cyclin A1 promoter. Taken together, these findings demonstrate that cyclin A1 is a transcriptional target of PITX2. Consistently, our immunohistochemistry result showed up-regulation of cyclin A1 in human papillary thyroid carcinoma, where overexpressed PITX2 has been endorsed in our recent report. Thus, our study provides new evidence on the regulation of cyclin A1 gene expression and offers a PITX2-cycin A1 pathway for cell cycle regulation.

  5. Identification of extracellular signal-regulated kinase 3 as a new interaction partner of cyclin D3

    International Nuclear Information System (INIS)

    Sun Maoyun; Wei Yuanyan; Yao Luyang; Xie Jianhui; Chen Xiaoning; Wang Hanzhou; Jiang Jianhai; Gu Jianxin

    2006-01-01

    Cyclin D3, like cyclin D1 and D2 isoforms, is a crucial component of the core cell cycle machinery in mammalian cells. It also exhibits its unique properties in many other physiological processes. In the present study, using yeast two-hybrid screening, we identified ERK3, an atypical mitogen-activated protein kinase (MAPK), as a cyclin D3 binding partner. GST pull-down assays showed that cyclin D3 interacts directly and specifically with ERK3 in vitro. The binding of cyclin D3 and ERK3 was further confirmed in vivo by co-immunoprecipitation assay and confocal microscopic analysis. Moreover, carboxy-terminal extension of ERK3 was responsible for its association with intact cyclin D3. These findings further expand distinct roles of cyclin D3 and suggest the potential activity of ERK3 in cell proliferation

  6. Cyclin dependent kinase (CDK) inhibitors as anticancer drugs.

    Science.gov (United States)

    Sánchez-Martínez, Concepción; Gelbert, Lawrence M; Lallena, María José; de Dios, Alfonso

    2015-09-01

    Sustained proliferative capacity is a hallmark of cancer. In mammalian cells proliferation is controlled by the cell cycle, where cyclin-dependent kinases (CDKs) regulate critical checkpoints. CDK4 and CDK6 are considered highly validated anticancer drug targets due to their essential role regulating cell cycle progression at the G1 restriction point. This review provides an overview of recent advances on cyclin dependent kinase inhibitors in general with special emphasis on CDK4 and CDK6 inhibitors and compounds under clinical evaluation. Chemical structures, structure activity relationships, and relevant preclinical properties will be described. Copyright © 2015 Elsevier Ltd. All rights reserved.

  7. Cordyceps militaris Grown on Germinated Soybean Induces G2/M Cell Cycle Arrest through Downregulation of Cyclin B1 and Cdc25c in Human Colon Cancer HT-29 Cells

    Directory of Open Access Journals (Sweden)

    Mohammad Lalmoddin Mollah

    2012-01-01

    Full Text Available Cordyceps militaris (CM is an insect-borne fungus that has been used in traditional Chinese medicine because of its wide range of pharmacological activities. In this paper, we studied CM grown on germinated soybean (GSC and investigated the possible mechanisms underlying antiproliferative effect of GSC on HT-29 human colon cancer cells. In comparison with CM extracts and germinated soybean (GS BuOH extracts, BuOH extracts of GSC showed remarkable inhibitory and antiproliferative effects on HT-29 colon cancer cells. After GSC treatment, HT-29 cells became smaller and irregular in shape. High G2/M phase cell populations were observed in the GSC-treated group. The levels of cyclin B1 and Cdc25 in the GSC-treated group were lower than those in the control group. These findings suggest that GSC BuOH extracts might act as an effective anti-proliferative agent by inducing G2/M cell cycle arrest in colon cancer cells.

  8. Viral cyclins mediate separate phases of infection by integrating functions of distinct mammalian cyclins.

    Directory of Open Access Journals (Sweden)

    Katherine S Lee

    2012-02-01

    Full Text Available Gammaherpesvirus cyclins have expanded biochemical features relative to mammalian cyclins, and promote infection and pathogenesis including acute lung infection, viral persistence, and reactivation from latency. To define the essential features of the viral cyclin, we generated a panel of knock-in viruses expressing various viral or mammalian cyclins from the murine gammaherpesvirus 68 cyclin locus. Viral cyclins of both gammaherpesvirus 68 and Kaposi's sarcoma-associated herpesvirus supported all cyclin-dependent stages of infection, indicating functional conservation. Although mammalian cyclins could not restore lung replication, they did promote viral persistence and reactivation. Strikingly, distinct and non-overlapping mammalian cyclins complemented persistence (cyclin A, E or reactivation from latency (cyclin D3. Based on these data, unique biochemical features of viral cyclins (e.g. enhanced kinase activation are not essential to mediate specific processes during infection. What is essential for, and unique to, the viral cyclins is the integration of the activities of several different mammalian cyclins, which allows viral cyclins to mediate multiple, discrete stages of infection. These studies also demonstrated that closely related stages of infection, that are cyclin-dependent, are in fact genetically distinct, and thus predict that cyclin requirements may be used to tailor potential therapies for virus-associated diseases.

  9. Cyclin K dependent regulation of Aurora B affects apoptosis and proliferation by induction of mitotic catastrophe in prostate cancer.

    Science.gov (United States)

    Schecher, Sabrina; Walter, Britta; Falkenstein, Michael; Macher-Goeppinger, Stephan; Stenzel, Philipp; Krümpelmann, Kristina; Hadaschik, Boris; Perner, Sven; Kristiansen, Glen; Duensing, Stefan; Roth, Wilfried; Tagscherer, Katrin E

    2017-10-15

    Cyclin K plays a critical role in transcriptional regulation as well as cell development. However, the role of Cyclin K in prostate cancer is unknown. Here, we describe the impact of Cyclin K on prostate cancer cells and examine the clinical relevance of Cyclin K as a biomarker for patients with prostate cancer. We show that Cyclin K depletion in prostate cancer cells induces apoptosis and inhibits proliferation accompanied by an accumulation of cells in the G2/M phase. Moreover, knockdown of Cyclin K causes mitotic catastrophe displayed by multinucleation and spindle multipolarity. Furthermore, we demonstrate a Cyclin K dependent regulation of the mitotic kinase Aurora B and provide evidence for an Aurora B dependent induction of mitotic catastrophe. In addition, we show that Cyclin K expression is associated with poor biochemical recurrence-free survival in patients with prostate cancer treated with an adjuvant therapy. In conclusion, targeting Cyclin K represents a novel, promising anti-cancer strategy to induce cell cycle arrest and apoptotic cell death through induction of mitotic catastrophe in prostate cancer cells. Moreover, our results indicate that Cyclin K is a putative predictive biomarker for clinical outcome and therapy response for patients with prostate cancer. © 2017 UICC.

  10. Aminopurvalanol A, a Potent, Selective, and Cell Permeable Inhibitor of Cyclins/Cdk Complexes, Causes the Reduction of in Vitro Fertilizing Ability of Boar Spermatozoa, by Negatively Affecting the Capacitation-Dependent Actin Polymerization

    Directory of Open Access Journals (Sweden)

    Nicola Bernabò

    2017-12-01

    Full Text Available The adoption of high-througput technologies demonstrated that in mature spermatozoa are present proteins that are thought to be not present or active in sperm cells, such as those involved in control of cell cycle. Here, by using an in silico approach based on the application of networks theory, we found that Cyclins/Cdk complexes could play a central role in signal transduction active during capacitation. Then, we tested this hypothesis in the vitro model. With this approach, spermatozoa were incubated under capacitating conditions in control conditions (CTRL or in the presence of Aminopurvalanol A a potent, selective and cell permeable inhibitor of Cyclins/Cdk complexes at different concentrations (2, 10, and 20 μM. We found that this treatment caused dose-dependent inhibition of sperm fertilizing ability. We attribute this event to the loss of acrosome integrity due to the inhibition of physiological capacitation-dependent actin polymerization, rather than to a detrimental effect on membrane lipid remodeling or on other signaling pathways such as tubulin reorganization or MAPKs activation. In our opinion, these data could revamp the knowledge on biochemistry of sperm capacitation and could suggest new perspectives in studying male infertility.

  11. Cyclin-E is a strong predictor of endocrine therapy failure in human breast cancer.

    NARCIS (Netherlands)

    Span, P.N.; Tjan-Heijnen, V.C.; Manders, P.; Beex, L.V.A.M.; Sweep, C.G.J.

    2003-01-01

    Recently, cyclin-E was reported to be the most prominent prognostic factor for breast cancer outcome described so far, even surpassing axillary nodal involvement. Earlier studies on the prognostic value of cyclin-E in breast cancer, however, yielded heterogeneous results. Therefore, we set out to

  12. Analysis of cyclins A, B1, D1 and E in breast cancer in relation to tumour grade and other prognostic factors

    Directory of Open Access Journals (Sweden)

    Collan Yrjö

    2009-07-01

    Full Text Available Abstract Background The cell cycle is promoted by activation of cyclin dependent kinases (Cdks, which are regulated positively by cyclins and negatively by Cdk inhibitors. Proliferation of carcinoma is associated with altered regulation of the cell cycle. Little is known on the combined alterations of cyclins A, B1, D1 and E in breast cancer in relation to the tumour grade and other prognostic factors. Findings Immunohistochemical analysis of cyclins A, B1, D1 and E, estrogen receptor, progesterone receptor, Ki-67, Her-2/neu and CK5/6 was performed on 53 breast carcinomas. mRNA levels of the cyclins were analysed of 12 samples by RT-PCR. The expression of cyclins A, B1 and E correlated with each other, while cyclin D1 correlated with none of these cyclins. Cyclins A, B1 and E showed association with tumour grade, Her-2/neu and Ki-67. Cyclin E had a negative correlation with hormone receptors and a positive correlation with triple negative carcinomas. Cyclin D1 had a positive correlation with ER, PR and non-basal breast carcinomas. Conclusion Cyclin A, B1 and E overexpression correlates to grade, Ki-67 and Her2/neu expression. Overexpression of cyclin D1 has a positive correlation with receptor status and non-basal carcinomas suggesting that cyclin D1 expression might be a marker of good prognosis. Combined analysis of cyclins indicates that cyclin A, B and E expression is similarly regulated, while other factors regulate cyclin D1 expression. The results suggest that the combined immunoreactivity of cyclins A, B1, D and E might be a useful prognostic factor in breast cancer.

  13. Regulation of the retinoblastoma protein-related p107 by G1 cyclin complexes

    NARCIS (Netherlands)

    Beijersbergen, R.L.; Carlée, L.; Kerkhoven, R.M.; Bernards, R.A.

    1995-01-01

    The orderly progression through the cell cycle is mediated by the sequential activation of several cyclin/cyclin-dependent kinase (cdk) complexes. These kinases phosphorylate a number of cellular substrates, among which is the product of the retinoblastoma gene, pRb. Phosphorylation of pRb in late

  14. microRNA-365, down-regulated in colon cancer, inhibits cell cycle progression and promotes apoptosis of colon cancer cells by probably targeting Cyclin D1 and Bcl-2.

    Science.gov (United States)

    Nie, Jing; Liu, Lin; Zheng, Wei; Chen, Lin; Wu, Xin; Xu, Yingxin; Du, Xiaohui; Han, Weidong

    2012-01-01

    Deregulated microRNAs participate in carcinogenesis and cancer progression, but their roles in cancer development remain unclear. In this study, miR-365 expression was found to be downregulated in human colon cancer tissues as compared with that in matched non-neoplastic mucosa tissues, and its downregulation was correlated with cancer progression and poor survival in colon cancer patients. Functional studies revealed that restoration of miR-365 expression inhibited cell cycle progression, promoted 5-fluorouracil-induced apoptosis and repressed tumorigenicity in colon cancer cell lines. Furthermore, bioinformatic prediction and experimental validation were used to identify miR-365 target genes and indicated that the antitumor effects of miR-365 were probably mediated by its targeting and repression of Cyclin D1 and Bcl-2 expression, thus inhibiting cell cycle progression and promoting apoptosis. These results suggest that downregulation of miR-365 in colon cancer may have potential applications in prognosis prediction and gene therapy in colon cancer patients.

  15. Selective induction of cyclin B protein abrogates the G2 delay after irradiation

    International Nuclear Information System (INIS)

    Kao, G.; Muschel, R.J.; Maity, A.; Kunig, A.; McKenna, W.G.

    1996-01-01

    Purpose/Objective: Irradiation of tumor cells commonly results in G2 delay, which has been postulated to allow DNA repair and cell survival. The G2 delay after irradiation is also often marked in some cell lines by delayed expression of cyclin B protein, suggesting a role for cyclin B regulation. Investigations of these hypotheses however has been hampered by the inability to selectively perturb the G2 delay in a physiologic manner. Materials and Methods: We have devised a system, with which we are able to selectively induce cyclin B protein expression in vivo at specific points in the cell cycle, by transfecting Hela cells with an expression vector under control of a dexamethasone-inducible promoter. Experiments were subsequently performed by synchronizing, releasing, irradiating, inducing, and harvesting these cells through the cell cycle. Results: Irradiation with 5 Gy led to a pronounced G2 delay, reflected by markedly slowed progression into mitosis, concomitant with reduced expression of cyclin B protein. Induction of cyclin B after radiation in these cells abrogated the G2 delay by approximately doubling the rate at which the cells re-enter mitosis. Treatment of irradiated untransfected control cells with dexamethasone, in which cyclin B is not induced, led to minimal changes. Studies of effects of cyclin B induction on cyclin B localization (using immunofluorescence), cdc2 phosphorylation and activation will also be presented. Conclusion: This system should allow further investigations into fundamental mechanisms of cell cycle regulation after irradiation and DNA damage. This also provides direct evidence for the first time that cyclin B protein regulation may play a role in the G2 delay following irradiation in Hela cells, perhaps complementing phosphorylation events

  16. Potentiation of in vitro and in vivo antitumor efficacy of doxorubicin by cyclin-dependent kinase inhibitor P276-00 in human non-small cell lung cancer cells

    International Nuclear Information System (INIS)

    Rathos, Maggie J; Khanwalkar, Harshal; Joshi, Kavita; Manohar, Sonal M; Joshi, Kalpana S

    2013-01-01

    In the present study, we show that the combination of doxorubicin with the cyclin-dependent kinase inhibitor P276-00 was synergistic at suboptimal doses in the non-small cell lung carcinoma (NSCLC) cell lines and induces extensive apoptosis than either drug alone in H-460 human NSCLC cells. Synergistic effects of P276-00 and doxorubicin on growth inhibition was studied using the Propidium Iodide (PI) assay. The doses showing the best synergistic effect was determined and these doses were used for further mechanistic studies such as western blotting, cell cycle analysis and RT-PCR. The in vivo efficacy of the combination was evaluated using the H-460 xenograft model. The combination of 100 nM doxorubicin followed by 1200 nM P276-00 showed synergistic effect in the p53-positive and p53-mutated cell lines H-460 and H23 respectively as compared to the p53-null cell line H1299. Abrogation of doxorubicin-induced G2/M arrest and induction of apoptosis was observed in the combination treatment. This was associated with induction of tumor suppressor protein p53 and reduction of anti-apoptotic protein Bcl-2. Furthermore, doxorubicin alone greatly induced COX-2, a NF-κB target and Cdk-1, a target of P276-00, which was downregulated by P276-00 in the combination. Doxorubicin when combined with P276-00 in a sequence-specific manner significantly inhibited tumor growth, compared with either doxorubicin or P276-00 alone in H-460 xenograft model. These findings suggest that this combination may increase the therapeutic index over doxorubicin alone and reduce systemic toxicity of doxorubicin most likely via an inhibition of doxorubicin-induced chemoresistance involving NF-κB signaling and inhibition of Cdk-1 which is involved in cell cycle progression

  17. Down-regulation of hTERT and Cyclin D1 transcription via PI3K/Akt and TGF-β pathways in MCF-7 Cancer cells with PX-866 and Raloxifene

    Energy Technology Data Exchange (ETDEWEB)

    Peek, Gregory W. [Department of Biology, University of Alabama at Birmingham, Birmingham, AL (United States); Tollefsbol, Trygve O., E-mail: trygve@uab.edu [Department of Biology, University of Alabama at Birmingham, Birmingham, AL (United States); Comprehensive Cancer Center, University of Alabama at Birmingham, Birmingham, AL (United States); Comprehensive Center for Healthy Aging, University of Alabama at Birmingham, Birmingham, AL (United States); Comprehensive Diabetes Center, University of Alabama at Birmingham, Birmingham, AL (United States); Nutrition Obesity Research Center, University of Alabama at Birmingham, Birmingham, AL (United States)

    2016-05-15

    Human telomerase reverse transcriptase (hTERT) is the catalytic and limiting component of telomerase and also a transcription factor. It is critical to the integrity of the ends of linear chromosomes and to the regulation, extent and rate of cell cycle progression in multicellular eukaryotes. The level of hTERT expression is essential to a wide range of bodily functions and to avoidance of disease conditions, such as cancer, that are mediated in part by aberrant level and regulation of cell cycle proliferation. Value of a gene in regulation depends on its ability to both receive input from multiple sources and transmit signals to multiple effectors. The expression of hTERT and the progression of the cell cycle have been shown to be regulated by an extensive network of gene products and signaling pathways, including the PI3K/Akt and TGF-β pathways. The PI3K inhibitor PX-866 and the competitive estrogen receptor ligand raloxifene have been shown to modify progression of those pathways and, in combination, to decrease proliferation of estrogen receptor positive (ER+) MCF-7 breast cancer cells. We found that combinations of modulators of those pathways decreased not only hTERT transcription but also transcription of additional essential cell cycle regulators such as Cyclin D1. By evaluating known expression profile signatures for TGF-β pathway diversions, we confirmed additional genes such as heparin-binding epidermal growth factor-like growth factor (HB EGF) by which those pathways and their perturbations may also modify cell cycle progression. - Highlights: • PX-866 and raloxifene affect the PI3K/Akt and TGF-β pathways. • PX-866 and raloxifene down-regulate genes up-regulated in cancer. • PX-866 and raloxifene decrease transcription of hTERT and Cyclin D1. • Pathological transcription signatures can identify new defense mechanisms.

  18. CyclinPred: a SVM-based method for predicting cyclin protein sequences.

    Directory of Open Access Journals (Sweden)

    Mridul K Kalita

    Full Text Available Functional annotation of protein sequences with low similarity to well characterized protein sequences is a major challenge of computational biology in the post genomic era. The cyclin protein family is once such important family of proteins which consists of sequences with low sequence similarity making discovery of novel cyclins and establishing orthologous relationships amongst the cyclins, a difficult task. The currently identified cyclin motifs and cyclin associated domains do not represent all of the identified and characterized cyclin sequences. We describe a Support Vector Machine (SVM based classifier, CyclinPred, which can predict cyclin sequences with high efficiency. The SVM classifier was trained with features of selected cyclin and non cyclin protein sequences. The training features of the protein sequences include amino acid composition, dipeptide composition, secondary structure composition and PSI-BLAST generated Position Specific Scoring Matrix (PSSM profiles. Results obtained from Leave-One-Out cross validation or jackknife test, self consistency and holdout tests prove that the SVM classifier trained with features of PSSM profile was more accurate than the classifiers based on either of the other features alone or hybrids of these features. A cyclin prediction server--CyclinPred has been setup based on SVM model trained with PSSM profiles. CyclinPred prediction results prove that the method may be used as a cyclin prediction tool, complementing conventional cyclin prediction methods.

  19. Cyclin A2 promotes DNA repair in the brain during both development and aging.

    Science.gov (United States)

    Gygli, Patrick E; Chang, Joshua C; Gokozan, Hamza N; Catacutan, Fay P; Schmidt, Theresa A; Kaya, Behiye; Goksel, Mustafa; Baig, Faisal S; Chen, Shannon; Griveau, Amelie; Michowski, Wojciech; Wong, Michael; Palanichamy, Kamalakannan; Sicinski, Piotr; Nelson, Randy J; Czeisler, Catherine; Otero, José J

    2016-07-01

    Various stem cell niches of the brain have differential requirements for Cyclin A2. Cyclin A2 loss results in marked cerebellar dysmorphia, whereas forebrain growth is retarded during early embryonic development yet achieves normal size at birth. To understand the differential requirements of distinct brain regions for Cyclin A2, we utilized neuroanatomical, transgenic mouse, and mathematical modeling techniques to generate testable hypotheses that provide insight into how Cyclin A2 loss results in compensatory forebrain growth during late embryonic development. Using unbiased measurements of the forebrain stem cell niche, we parameterized a mathematical model whereby logistic growth instructs progenitor cells as to the cell-types of their progeny. Our data was consistent with prior findings that progenitors proliferate along an auto-inhibitory growth curve. The growth retardation inCCNA2-null brains corresponded to cell cycle lengthening, imposing a developmental delay. We hypothesized that Cyclin A2 regulates DNA repair and that CCNA2-null progenitors thus experienced lengthened cell cycle. We demonstrate that CCNA2-null progenitors suffer abnormal DNA repair, and implicate Cyclin A2 in double-strand break repair. Cyclin A2's DNA repair functions are conserved among cell lines, neural progenitors, and hippocampal neurons. We further demonstrate that neuronal CCNA2 ablation results in learning and memory deficits in aged mice.

  20. Plasmodium P-Type Cyclin CYC3 Modulates Endomitotic Growth during Oocyst Development in Mosquitoes

    KAUST Repository

    Roques, Magali

    2015-11-13

    Cell-cycle progression and cell division in eukaryotes are governed in part by the cyclin family and their regulation of cyclin-dependent kinases (CDKs). Cyclins are very well characterised in model systems such as yeast and human cells, but surprisingly little is known about their number and role in Plasmodium, the unicellular protozoan parasite that causes malaria. Malaria parasite cell division and proliferation differs from that of many eukaryotes. During its life cycle it undergoes two types of mitosis: endomitosis in asexual stages and an extremely rapid mitotic process during male gametogenesis. Both schizogony (producing merozoites) in host liver and red blood cells, and sporogony (producing sporozoites) in the mosquito vector, are endomitotic with repeated nuclear replication, without chromosome condensation, before cell division. The role of specific cyclins during Plasmodium cell proliferation was unknown. We show here that the Plasmodium genome contains only three cyclin genes, representing an unusual repertoire of cyclin classes. Expression and reverse genetic analyses of the single Plant (P)-type cyclin, CYC3, in the rodent malaria parasite, Plasmodium berghei, revealed a cytoplasmic and nuclear location of the GFP-tagged protein throughout the lifecycle. Deletion of cyc3 resulted in defects in size, number and growth of oocysts, with abnormalities in budding and sporozoite formation. Furthermore, global transcript analysis of the cyc3-deleted and wild type parasites at gametocyte and ookinete stages identified differentially expressed genes required for signalling, invasion and oocyst development. Collectively these data suggest that cyc3 modulates oocyst endomitotic development in Plasmodium berghei.

  1. Plasmodium P-Type Cyclin CYC3 Modulates Endomitotic Growth during Oocyst Development in Mosquitoes.

    Science.gov (United States)

    Roques, Magali; Wall, Richard J; Douglass, Alexander P; Ramaprasad, Abhinay; Ferguson, David J P; Kaindama, Mbinda L; Brusini, Lorenzo; Joshi, Nimitray; Rchiad, Zineb; Brady, Declan; Guttery, David S; Wheatley, Sally P; Yamano, Hiroyuki; Holder, Anthony A; Pain, Arnab; Wickstead, Bill; Tewari, Rita

    2015-11-01

    Cell-cycle progression and cell division in eukaryotes are governed in part by the cyclin family and their regulation of cyclin-dependent kinases (CDKs). Cyclins are very well characterised in model systems such as yeast and human cells, but surprisingly little is known about their number and role in Plasmodium, the unicellular protozoan parasite that causes malaria. Malaria parasite cell division and proliferation differs from that of many eukaryotes. During its life cycle it undergoes two types of mitosis: endomitosis in asexual stages and an extremely rapid mitotic process during male gametogenesis. Both schizogony (producing merozoites) in host liver and red blood cells, and sporogony (producing sporozoites) in the mosquito vector, are endomitotic with repeated nuclear replication, without chromosome condensation, before cell division. The role of specific cyclins during Plasmodium cell proliferation was unknown. We show here that the Plasmodium genome contains only three cyclin genes, representing an unusual repertoire of cyclin classes. Expression and reverse genetic analyses of the single Plant (P)-type cyclin, CYC3, in the rodent malaria parasite, Plasmodium berghei, revealed a cytoplasmic and nuclear location of the GFP-tagged protein throughout the lifecycle. Deletion of cyc3 resulted in defects in size, number and growth of oocysts, with abnormalities in budding and sporozoite formation. Furthermore, global transcript analysis of the cyc3-deleted and wild type parasites at gametocyte and ookinete stages identified differentially expressed genes required for signalling, invasion and oocyst development. Collectively these data suggest that cyc3 modulates oocyst endomitotic development in Plasmodium berghei.

  2. Low-molecular-weight cyclin E: the missing link between biology and clinical outcome

    International Nuclear Information System (INIS)

    Akli, Said; Keyomarsi, Khandan

    2004-01-01

    Cyclin E, a key mediator of transition during the G 1 /S cellular division phase, is deregulated in a wide variety of human cancers. Our group recently reported that overexpression and generation of low-molecular-weight (LMW) isoforms of cyclin E were associated with poor clinical outcome among breast cancer patients. However, the link between LMW cyclin E biology in mediating a tumorigenic phenotype and clinical outcome is unknown. To address this gap in knowledge, we assessed the role of LMW isoforms in breast cancer cells; we found that these forms of cyclin E induced genomic instability and resistance to p21, p27, and antiestrogens in breast cancer. These findings suggest that high levels of LMW isoforms of cyclin E not only can predict failure to endocrine therapy but also are true prognostic indicators because of their influence on cell proliferation and genetic instability

  3. Mesenchymal stem cells promote liver regeneration and prolong survival in small-for-size liver grafts: involvement of C-Jun N-terminal kinase, cyclin D1, and NF-κB.

    Directory of Open Access Journals (Sweden)

    Weijie Wang

    Full Text Available BACKGROUND: The therapeutic potential of mesenchymal stem cells (MSCs has been highlighted recently for treatment of acute or chronic liver injury, by possibly differentiating into hepatocyte-like cells, reducing inflammation, and enhancing tissue repair. Despite recent progress, exact mechanisms of action are not clearly elucidated. In this study, we attempted to explore whether and how MSCs protected hepatocytes and stimulated allograft regeneration in small-for-size liver transplantation (SFSLT. METHODS: SFSLT model was established with a 30% partial liver transplantation (30PLT in rats. The differentiation potential and characteristics of bone marrow derived MSCs were explored in vitro. MSCs were infused transvenously immediately after graft implantation in therapy group. Expressions of apoptosis-, inflammatory-, anti-inflammatory-, and growth factor-related genes were measured by RT-PCR, activities of transcription factors AP-1 and NF-κB were analyzed by EMSA, and proliferative responses of the hepatic graft were evaluated by immunohistochemistry and western blot. RESULTS: MSCs were successfully induced into hepatocyte-like cells, osteoblasts and adipocytes in vitro. MSCs therapy could not only alleviate ischemia reperfusion injury and acute inflammation to promote liver regeneration, but also profoundly improve one week survival rate. It markedly up-regulated the mRNA expressions of HGF, Bcl-2, Bcl-XL, IL-6, IL-10, IP-10, and CXCR2, however, down-regulated TNF-α. Increased activities of AP-1 and NF-κB, as well as elevated expressions of p-c-Jun, cyclin D1, and proliferating cell nuclear antigen (PCNA, were also found in MSCs therapy group. CONCLUSION: These data suggest that MSCs therapy promotes hepatocyte proliferation and prolongs survival in SFSLT by reducing ischemia reperfusion injury and acute inflammation, and sustaining early increased expressions of c-Jun N-terminal Kinase, Cyclin D1, and NF-κB.

  4. Altered expression of cyclin A 1 in muscle of patients with facioscapulohumeral muscle dystrophy (FSHD-1.

    Directory of Open Access Journals (Sweden)

    Anna Pakula

    Full Text Available OBJECTIVES: Cyclin A1 regulates cell cycle activity and proliferation in somatic and germ-line cells. Its expression increases in G1/S phase and reaches a maximum in G2 and M phases. Altered cyclin A1 expression might contribute to clinical symptoms in facioscapulohumeral muscular dystrophy (FSHD. METHODS: Muscle biopsies were taken from the Vastus lateralis muscle for cDNA microarray, RT-PCR, immunohistochemistry and Western blot analyses to assess RNA and protein expression of cyclin A1 in human muscle cell lines and muscle tissue. Muscle fibers diameter was calculated on cryosections to test for hypertrophy. RESULTS: cDNA microarray data showed specifically elevated cyclin A1 levels in FSHD vs. other muscular disorders such as caveolinopathy, dysferlinopathy, four and a half LIM domains protein 1 deficiency and healthy controls. Data could be confirmed with RT-PCR and Western blot analysis showing up-regulated cyclin A1 levels also at protein level. We found also clear signs of hypertrophy within the Vastus lateralis muscle in FSHD-1 patients. CONCLUSIONS: In most somatic human cell lines, cyclin A1 levels are low. Overexpression of cyclin A1 in FSHD indicates cell cycle dysregulation in FSHD and might contribute to clinical symptoms of this disease.

  5. Cyclin D1 represses gluconeogenesis via inhibition of the transcriptional coactivator PGC1α.

    Science.gov (United States)

    Bhalla, Kavita; Liu, Wan-Ju; Thompson, Keyata; Anders, Lars; Devarakonda, Srikripa; Dewi, Ruby; Buckley, Stephanie; Hwang, Bor-Jang; Polster, Brian; Dorsey, Susan G; Sun, Yezhou; Sicinski, Piotr; Girnun, Geoffrey D

    2014-10-01

    Hepatic gluconeogenesis is crucial to maintain normal blood glucose during periods of nutrient deprivation. Gluconeogenesis is controlled at multiple levels by a variety of signal transduction and transcriptional pathways. However, dysregulation of these pathways leads to hyperglycemia and type 2 diabetes. While the effects of various signaling pathways on gluconeogenesis are well established, the downstream signaling events repressing gluconeogenic gene expression are not as well understood. The cell-cycle regulator cyclin D1 is expressed in the liver, despite the liver being a quiescent tissue. The most well-studied function of cyclin D1 is activation of cyclin-dependent kinase 4 (CDK4), promoting progression of the cell cycle. We show here a novel role for cyclin D1 as a regulator of gluconeogenic and oxidative phosphorylation (OxPhos) gene expression. In mice, fasting decreases liver cyclin D1 expression, while refeeding induces cyclin D1 expression. Inhibition of CDK4 enhances the gluconeogenic gene expression, whereas cyclin D1-mediated activation of CDK4 represses the gluconeogenic gene-expression program in vitro and in vivo. Importantly, we show that cyclin D1 represses gluconeogenesis and OxPhos in part via inhibition of peroxisome proliferator-activated receptor γ coactivator-1α (PGC1α) activity in a CDK4-dependent manner. Indeed, we demonstrate that PGC1α is novel cyclin D1/CDK4 substrate. These studies reveal a novel role for cyclin D1 on metabolism via PGC1α and reveal a potential link between cell-cycle regulation and metabolic control of glucose homeostasis. © 2014 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered.

  6. Limited prognostic value of tissue protein expression levels of cyclin E in Danish ovarian cancer patients

    DEFF Research Database (Denmark)

    Heeran, Mel C; Høgdall, Claus K; Kjaer, Susanne K

    2012-01-01

    The primary objective of this study was to assess the expression of cyclin E in tumour tissues from 661 patients with epithelial ovarian tumours. The second was to evaluate whether cyclin E tissue expression levels correlate with clinico-pathological parameters and prognosis of the disease. Using...... tissue arrays (TA), we analysed the cyclin E expression levels in tissues from 168 women with borderline ovarian tumours (BOT) (147 stage I, 4 stage II, 17 stage III) and 493 Ovarian cancer (OC) patients (127 stage I, 45 stage II, 276 stage III, 45 stage IV). Using a 10% cut-off level for cyclin E...... overexpression, 20% of the BOTs were positive with a higher proportion of serous than mucinous tumours. Sixty-two per cent of the OCs were positive for cyclin E expression with the highest percentage found in clear cell carcinomas. Results based on univariate and multivariate survival analyses with a 10% cut...

  7. The role of cyclins and cyclin dependent kinases in development and progression of hepatitis C virus-genotype 4-associated hepatitis and hepatocellular carcinoma.

    Science.gov (United States)

    Bahnassy, Abeer A; Zekri, Abdel-Rahman N; Loutfy, Samah A; Mohamed, Waleed S; Moneim, Amrallah Abdel; Salem, Salem E; Sheta, Marwa M; Omar, Ashraf; Al-Zawahry, Heba

    2011-10-01

    Altered cell cycle regulatory genes expression contributes to HCV-associated liver disease. We sought to assess the role of cyclins and cyclin dependent kinases (CDKs) in HCV-associated CH and HCC. Aberrant expression of cyclins A, E, D1, CDK2 and CDK4 was assessed by immunohistochemistry and differential PCR in HCV-associated CH and HCC with pericarcinomatous foci (PCF). S phase fraction (SPF) was determined by flow cytometry. Results were correlated with overall survival (OS) in HCC patients. In HCC, cyclins A, E, D1, CDK2 and CDK4 protein overexpression was detected in 52.8%, 52.8%, 69%, 47% and 58% compared to 36.1%, 33%, 56%, 27.8%, 55.6% for CH and 36.1%, 27%, 30.6%, 27%, 50% for PCF. Gene amplification was detected in 38.9%, 33% 66%, 33%, 44% of HCC compared to 27.8%, 25%, 44%, 27.8%, 36% in CH and 25%, 22.2%, 38.9%, 27%, 33% in PCF. A significant difference was reported between HCC, CH, NHT regarding cyclins A, E, D1, CDK2 (p=0.007, p=0.002, p=0.047, p=0.002) protein expression (ADD) and cyclin D1 amplification (p=0.009). Cyclins A, E, CDK2 expression was associated with fibrosis in CH (p=0.004, p=0.02, p=0.012). Reduced OS was (ADD) associated with cyclin D1 and cyclin A, grade, stage and metastasis (p=0.001, p=0.02, p=0.018, p=0.01, p=0.001). Increased cyclins A, E, D1, CDK2 and CDK4 expression is important for HCV-associated CH and HCC. Cyclin D1 and cyclin A are prognostic biomarkers associated with reduced OS in HCC. Cyclin D1 aberration could identify high risk groups of CH patients prone to develop HCC. Copyright © 2011 Elsevier Inc. All rights reserved.

  8. The cyclin-dependent kinase inhibitor flavopiridol disrupts sodium butyrate-induced p21WAF1/CIP1 expression and maturation while reciprocally potentiating apoptosis in human leukemia cells.

    Science.gov (United States)

    Rosato, Roberto R; Almenara, Jorge A; Cartee, Leanne; Betts, Vicki; Chellappan, Srikumar P; Grant, Steven

    2002-02-01

    Interactions between the cyclin-dependent kinase inhibitor flavopiridol (FP) and the histone deacetylase inhibitor sodium butyrate (SB) have been examined in human leukemia cells (U937) in relation to differentiation and apoptosis. Whereas 1 mM of SB or 100 nM of FP minimally induced apoptosis (4% and 10%, respectively) at 24 h, simultaneous exposure of U937 cells to these agents dramatically increased cell death (e.g., approximately 60%), reflected by both morphological and Annexin/propidium iodide-staining features, procaspase 3 activation, and poly(ADP-ribose) polymerase cleavage. Similar interactions were observed in human promyelocytic (HL-60), B-lymphoblastic (Raji), and T-lymphoblastic (Jurkat) leukemia cells. Coadministration of FP opposed SB-mediated accumulation of cells in G0G1 and differentiation, reflected by reduced CD11b expression, but instead dramatically increased procaspase-3, procaspase-8, Bid, and poly(ADP-ribose) polymerase cleavage, as well as mitochondrial damage (e.g., loss of mitochondrial membrane potential and cytochrome c release). FP also blocked SB-related p21WAF1-CIP1 induction through a caspase-independent mechanism and triggered the caspase-mediated cleavage of p27KIP1 and retinoblastoma protein. The latter event was accompanied by a marked reduction in retinoblastoma protein/E2F1 complex formation. However, FP did not modify the extent of SB-associated acetylation of histones H3 and H4. Treatment of cells with FP/SB also resulted in the caspase-mediated cleavage of Bcl-2 and caspase-independent down-regulation of Mcl-1. Levels of cyclins A, D1, and E, and X-linked inhibitor of apoptosis also declined in SB/FP-treated cells. Finally, FP/SB coexposure potently induced apoptosis in two primary acute myelogenous leukemia samples. Together, these findings demonstrate that FP, when combined with SB, induces multiple perturbations in cell cycle and apoptosis regulatory proteins, which oppose leukemic cell differentiation but instead

  9. Cyclin-dependent kinases in C. elegans

    Directory of Open Access Journals (Sweden)

    Boxem Mike

    2006-05-01

    Full Text Available Abstract Cell division is an inherent part of organismal development, and defects in this process can lead to developmental abnormalities as well as cancerous growth. In past decades, much of the basic cell-cycle machinery has been identified, and a major challenge in coming years will be to understand the complex interplay between cell division and multicellular development. Inevitably, this requires the use of more complex multicellular model systems. The small nematode Caenorhabditis elegans is an excellent model system to study the regulation of cell division in a multicellular organism, and is poised to make important contributions to this field. The past decade has already seen a surge in cell-cycle research in C. elegans, yielding information on the function of many basic cell-cycle regulators, and making inroads into the developmental control of cell division. This review focuses on the in vivo roles of cyclin-dependent kinases in C. elegans, and highlights novel findings implicating CDKs in coupling development to cell-cycle progression.

  10. Mutational analysis of the p16 family cyclin-dependent kinase inhibitors p15INK4b and p18INK4c in tumor-derived cell lines and primary tumors.

    Science.gov (United States)

    Zariwala, M; Liu, E; Xiong, Y

    1996-01-18

    The growth suppressing activity of the retinoblastoma suspectibility gene product, pRb, is down regulated by cyclin-dependent kinases 4 and 6 (CDK4 and CDK6) whose kinase activity is negatively regulated by CDK inhibitors of the p16 family. We have examined the genomic status of two recently isolated p16-related CDK inhibitors, p15 and p18, in 15 normal and 73 tumor-derived cell lines established from 23 different tissues, as well as 26 invasive primary breast cancers and 20 acute myelogenous leukemias. p15 was found to be homozygously deleted in 22% of the tumor derived cell lines, but no point mutations were found in either the cultured cells or the two types of primary tumors. With the exception of one breast cancer cell line, no deletions or mutations were found in the p18 gene in either cultured cell lines or primary tumors. These results indicate that mutation of the p18 gene occurs rarely in human tumors. Thus, while they share a very similar biochemical mechanism of inhibiting the kinase activity of CDK4 and CDK6, members of the p16 gene family play different roles in controlling cell proliferation and suppressing tumor growth.

  11. Control of G1 in the developing Drosophila eye: rca1 regulates Cyclin A.

    Science.gov (United States)

    Dong, X; Zavitz, K H; Thomas, B J; Lin, M; Campbell, S; Zipursky, S L

    1997-01-01

    In the developing eye of Drosophila melanogaster, cells become synchronized in the G1 phase of the cell cycle just prior to the onset of cellular differentiation and morphogenesis. In roughex (rux) mutants, cells enter S phase precociously because of ectopic activation of a Cyclin A/Cdk complex in early G1. This leads to defects in cell fate and pattern formation, and results in abnormalities in the morphology of the adult eye. A screen for dominant suppressors of the rux eye phenotype led to the identification of mutations in cyclin A, string (cdc25), and new cell cycle genes. One of these genes, regulator of cyclin A (rca1), encodes a novel protein required for both mitotic and meiotic cell cycle progression. rca1 mutants arrest in G2 of embryonic cell cycle 16 with a phenotype very similar to cyclin A loss of function mutants. Expression of rca1 transgenes in G1 or in postmitotic neurons promotes Cyclin A protein accumulation and drives cells into S phase in a Cyclin A-dependent fashion.

  12. Kinase Independent Functions of Cyclin D1 Which Contribute to its Oncogenic Potential In Vivo

    National Research Council Canada - National Science Library

    Landis, Mark

    2002-01-01

    .... Upon phosphorylation, pRb is inactivated, and cells pass from G1 into S phase. We and others have demonstrated that cyclin D1 has other functions, many of which are independent of kinase activity in vino...

  13. Human cyclin T1 expression ameliorates a T-cell-specific transcriptional limitation for HIV in transgenic rats, but is not sufficient for a spreading infection of prototypic R5 HIV-1 strains ex vivo

    Directory of Open Access Journals (Sweden)

    Littman Dan R

    2009-01-01

    Full Text Available Abstract Background Cells derived from native rodents have limits at distinct steps of HIV replication. Rat primary CD4 T-cells, but not macrophages, display a profound transcriptional deficit that is ameliorated by transient trans-complementation with the human Tat-interacting protein Cyclin T1 (hCycT1. Results Here, we generated transgenic rats that selectively express hCycT1 in CD4 T-cells and macrophages. hCycT1 expression in rat T-cells boosted early HIV gene expression to levels approaching those in infected primary human T-cells. hCycT1 expression was necessary, but not sufficient, to enhance HIV transcription in T-cells from individual transgenic animals, indicating that endogenous cellular factors are critical co-regulators of HIV gene expression in rats. T-cells from hCD4/hCCR5/hCycT1-transgenic rats did not support productive infection of prototypic wild-type R5 HIV-1 strains ex vivo, suggesting one or more significant limitation in the late phase of the replication cycle in this primary rodent cell type. Remarkably, we identify a replication-competent HIV-1 GFP reporter strain (R7/3 YU-2 Env that displays characteristics of a spreading, primarily cell-to-cell-mediated infection in primary T-cells from hCD4/hCCR5-transgenic rats. Moreover, the replication of this recombinant HIV-1 strain was significantly enhanced by hCycT1 transgenesis. The viral determinants of this so far unique replicative ability are currently unknown. Conclusion Thus, hCycT1 expression is beneficial to de novo HIV infection in a transgenic rat model, but additional genetic manipulations of the host or virus are required to achieve full permissivity.

  14. Patterns of cyclin A and B1 immunostaining in papillary thyroid carcinoma.

    Science.gov (United States)

    Cyniak-Magierska, Anna; Stasiak, Magdalena; Naze, Maciej; Dedecjus, Marek; Brzeziński, Jan; Lewiński, Andrzej

    2015-01-01

    Cyclin A, encoded by CCNA (cyclin A) gene with locus in chromosome 4q27, and cyclin B1, encoded by CCNB1 (cyclin B1) gene with locus in chromosome 5q12, are proteins that play a key role in the passage through the restriction point in G2 phase of the cell cycle. The aim of the study was to analyse immunohistochemically the expression of cyclins A and B1 in different variants of papillary thyroid carcinoma (PTC). The immunostaining patterns of the proteins in question in the tissue of 40 resected PTC (20 cases of classic variant of PTC, 9 cases of PTC follicular variant and 11 cases of other non-classic variants of PTC) were investigated. On analyzing cyclin A and B1 expression, positive staining in 90% cases of PTC were observed. The study revealed a significant difference in expression of cyclins A and B1 between classic and non-classic variants of PTC. The expression of both examined cyclins was weaker in the classic variant of PTC. In the group of follicular variant of PTC, the expression of cyclins was of medium intensity and in the group of other non-classic variants of PTC, the expression was clearly higher. The results of the presented study suggest that cyclins A and B1 expression may have a characteristic pattern of immunostaining for particular variants of PTC. If the obtained results are confirmed in a larger group of patients, the diagnostic panel constructed of the antibodies against these proteins may increase the diagnostic accuracy in PTC cases.

  15. Cyclin D3 interacts with vitamin D receptor and regulates its transcription activity

    International Nuclear Information System (INIS)

    Jian Yongzhi; Yan Jun; Wang Hanzhou; Chen Chen; Sun Maoyun; Jiang Jianhai; Lu Jieqiong; Yang Yanzhong; Gu Jianxin

    2005-01-01

    D-type cyclins are essential for the progression through the G1 phase of the cell cycle. Besides serving as cell cycle regulators, D-type cyclins were recently reported to have transcription regulation functions. Here, we report that cyclin D3 is a new interacting partner of vitamin D receptor (VDR), a member of the superfamily of nuclear receptors for steroid hormones, thyroid hormone, and the fat-soluble vitamins A and D. The interaction was confirmed with methods of yeast two-hybrid system, in vitro binding analysis and in vivo co-immunoprecipitation. Cyclin D3 interacted with VDR in a ligand-independent manner, but treatment of the ligand, 1,25-dihydroxyvitamin D3, strengthened the interaction. Confocal microscopy analysis showed that ligand-activated VDR led to an accumulation of cyclin D3 in the nuclear region. Cyclin D3 up-regulated transcriptional activity of VDR and this effect was counteracted by overexpression of CDK4 and CDK6. These findings provide us a new clue to understand the transcription regulation functions of D-type cyclins

  16. Structural basis for CDK6 activation by a virus-encoded cyclin

    Energy Technology Data Exchange (ETDEWEB)

    Schulze-Gahmen, Ursula; Kim, Sung-Hou

    2002-01-17

    Cyclin from herpesvirus saimiri (Vcyclin) preferentially forms complexes with cyclin-dependent kinase 6 (CDK6) from primate host cells. These complexes show higher kinase activity than host cell CDK complexes with cellular cyclins and are resistant to cyclin-dependent inhibitory proteins (CDKIs). The crystal structure of human CDK6-Vcyclin in an active state was determined to 3.1 Angstrom resolution to get a better understanding of the structural basis of CDK6 activation by viral cyclins. The unphosphorylated CDK6 complexed to Vcyclin has many features characteristic of cyclinA-activated, phosphorylated CDK2. There are, however, differences in the conformation at the tip of the T-loop and its interactions with Vcyclin. Residues in the N-terminal extension of Vcyclin wrap around the tip of the CDK6 T-loop and form a short b-sheet with the T-loop backbone. These interactions lead to a 20 percent larger buried surface in the CDK6-Vcyclin interface than in the CDK2-cyclinA complex and are probably largely responsible for Vcyclin specificity for CDK6 and resistance of the complex to inhibition by INK-typeCDKIs.

  17. Cyclin D1 overexpression associated with activation of STAT3 in oral carcinoma patients from South India.

    Science.gov (United States)

    Deepak Roshan, V G; Sinto, M S; Thomas, Shaji; Kannan, S

    2018-01-01

    Our previous study showed that overexpression of cyclin D1 protein is associated with poor prognosis in oral squamous cell carcinoma (OSCC) patients. Regarding the alteration in the transactivating pathway regulating cyclin D1 expression is still unclear in OSCC from our population. The major objective of this study is to understand the alteration associated with the transactivation pathway regulating the cyclin D1 overexpression in OSCC patients from our population. Alteration in the transactivation pathway regulating cyclin D1 expression was evaluated in tumor sample from OSCC patients. The findings were further validated using in vitro knockdown model in OSCC cell line. Results from the patients' samples showed that the Phospho-STAT3 has a significant association with cyclin D1 overexpression in OSCC tumor samples. Further knockdown in vitro studies using SCC66 showed a significant correlation between STAT3 and cyclin D1 in OSCC. The results from this study showed that in our population the cyclin D1 overexpression is associated with hyperactivation of STAT3 pathway. Our previous result has shown that the cyclin D1 protein overexpression is associated with poor prognosis in OSCC patients. Hence, STAT3 pathway will be better target for the patients with increased cyclin D1 in OSCC patients from our population.

  18. Differential expression of cyclin Dl in human pituitary tumors: relation to MIB-1 and p27/Kipl labeling indices

    International Nuclear Information System (INIS)

    Hewedi, I.H.; Osman, W.M.; El Mahdy, M.M.

    2011-01-01

    Pituitary tumors are a common form of endocrine neoplasia. However few studies assessed the expression of the principal cyclin regulating checkpoint exit, cyclin Dl. Cyclin Dl expression in pituitary tumors and its possible relation to MIB-1 and p27/K.ipl labeling indices (Us) was explored. Design: We studied a total of 199 pituitaries, including normal pituitaries (n = 7), pituitary adenomas (n = 187), and pituitary carcinoma (n = 5). All tissues were tested as cores of archived tissue microarrays that were immuno stained for cyclin Dl, MIB-1 and p27 using a standard technique. Tissue cores were subjected to automated analysis to evaluate the staining LIs, Results: No cyclin Dl positive cells in the normal anterior pituitary gland was found. Sparse nuclear staining was noted in pituitary tumors. Higher expression of cyclin Dl was noted in pituitary carcinomas compared to adenomas (p < 0.001), in non-functioning adenomas compared to functioning ones (p < 0.001) in macroadenomas versus micro adenomas (p — 0.017) and in recurrent non recurrent adenomas (p < 0.001). Cyclin Dl LI and MIB-1 LI were related among adenomas (p < 0.001) and carcinomas (p = 0.041). p27 LI was neither related to pituitary adenoma recurrence nor invasion. Conclusions: Expression of cyclin Dl in pituitary tumors is related to cell proliferation, recurrence, and metastatic potential. Nuclear cyclin Dl expression is a good marker of aggressive behavior in pituitary tumors

  19. Expression of cyclins A and E in melanocytic skin lesions and its correlation with some clinicopathologic features

    Directory of Open Access Journals (Sweden)

    Ana Alekseenko

    2012-07-01

    Full Text Available Cyclins play a fundamental role in the cell cycle. Recent studies have focused on their role in the development of various malignancies. The objective of this study was to evaluate and compare the expression of cyclins A and E in common nevi, dysplastic nevi and malignant melanomas, and to investigate the relationship between cyclin expression and some pathological parameters such as tumor thickness, ulceration, regression, and mitotic rate, as well as several clinical and phenotypic parameters such as skin phototype, hair and eye color, number of nevi, personal or family melanoma history, and personal history of nonmelanoma skin cancer (NMSC. A total of 102 melanocytic skin lesions, including 30 common nevi, 38 dysplastic nevi and 34 melanomas, were examined. Expression of cyclins was detected by immunohistochemistry and quantified as a percentage of immunostained cell nuclei in each sample. Significant differences in expression of both cyclins were found between all lesion types: the median percentage of cyclin A-positive nuclei was 8.2% in melanomas, 3.4% in dysplastic nevi, and 0.95% in common nevi (p < 0.001. The corresponding percentages for cyclin E were 9.5%, 4.25% and 1.44% (p < 0.001. Expression of both cyclins was significantly higher among patients with a personal history of NMSC. Cyclin A was also significantly overexpressed in patients with a high total nevus count (TNC compared to moderate and low TNC. Expression of cyclins did not significantly correlate with the other clinicopathologic features investigated. These findings indicate the possible involvement of cyclins A and E in the pathogenesis of malignant melanoma. Our results also show a potential diagnostic significance of these cyclins as markers allowing discrimination between dysplastic nevi and melanoma.

  20. Control of cyclin C levels during development of Dictyostelium.

    Directory of Open Access Journals (Sweden)

    David M Greene

    2010-05-01

    Full Text Available Cdk8 and its partner cyclin C form part of the mediator complex which links the basal transcription machinery to regulatory proteins. The pair are required for correct regulation of a subset of genes and have been implicated in control of development in a number of organisms including the social amoeba Dictyostelium discoideum. When feeding, Dictyostelium amoebae are unicellular but upon starvation they aggregate to form a multicellular structure which develops into a fruiting body containing spores. Cells in which the gene encoding Cdk8 has been deleted fail to enter aggregates due to a failure of early gene expression.We have monitored the expression levels of cyclin C protein during development and find levels decrease after the multicellular mound is formed. This decrease is triggered by extracellular cAMP that, in turn, is working in part through an increase in intracellular cAMP. The loss of cyclin C is coincident with a reduction in the association of Cdk8 with a high molecular weight complex in the nucleus. Overexpression of cyclin C and Cdk8 lead to an increased rate of early development, consistent with the levels being rate limiting.Overall these results show that both cyclin C and Cdk8 are regulated during development in response to extracellular signals and the levels of these proteins are important in controlling the timing of developmental processes. These findings have important implications for the role of these proteins in controlling development, suggesting that they are targets for developmental signals to regulate gene expression.

  1. Drosophila Cyclin G and epigenetic maintenance of gene expression during development.

    Science.gov (United States)

    Dupont, Camille A; Dardalhon-Cuménal, Delphine; Kyba, Michael; Brock, Hugh W; Randsholt, Neel B; Peronnet, Frédérique

    2015-01-01

    Cyclins and cyclin-dependent kinases (CDKs) are essential for cell cycle regulation and are functionally associated with proteins involved in epigenetic maintenance of transcriptional patterns in various developmental or cellular contexts. Epigenetic maintenance of transcription patterns, notably of Hox genes, requires the conserved Polycomb-group (PcG), Trithorax-group (TrxG), and Enhancer of Trithorax and Polycomb (ETP) proteins, particularly well studied in Drosophila. These proteins form large multimeric complexes that bind chromatin and appose or recognize histone post-translational modifications. PcG genes act as repressors, counteracted by trxG genes that maintain gene activation, while ETPs interact with both, behaving alternatively as repressors or activators. Drosophila Cyclin G negatively regulates cell growth and cell cycle progression, binds and co-localizes with the ETP Corto on chromatin, and participates with Corto in Abdominal-B Hox gene regulation. Here, we address further implications of Cyclin G in epigenetic maintenance of gene expression. We show that Cyclin G physically interacts and extensively co-localizes on chromatin with the conserved ETP Additional sex combs (ASX), belonging to the repressive PR-DUB complex that participates in H2A deubiquitination and Hox gene silencing. Furthermore, Cyclin G mainly co-localizes with RNA polymerase II phosphorylated on serine 2 that is specific to productive transcription. CycG interacts with Asx, PcG, and trxG genes in Hox gene maintenance, and behaves as a PcG gene. These interactions correlate with modified ectopic Hox protein domains in imaginal discs, consistent with a role for Cyclin G in PcG-mediated Hox gene repression. We show here that Drosophila CycG is a Polycomb-group gene enhancer, acting in epigenetic maintenance of the Hox genes Sex combs reduced (Scr) and Ultrabithorax (Ubx). However, our data suggest that Cyclin G acts alternatively as a transcriptional activator or repressor

  2. The cyclin-dependent kinase inhibitor 5, 6-dichloro-1-beta-D-ribofuranosylbenzimidazole induces nongenotoxic, DNA replication-independent apoptosis of normal and leukemic cells, regardless of their p53 status

    International Nuclear Information System (INIS)

    Turinetto, Valentina; Porcedda, Paola; Orlando, Luca; De Marchi, Mario; Amoroso, Antonio; Giachino, Claudia

    2009-01-01

    Current chemotherapy of human cancers focuses on the DNA damage pathway to induce a p53-mediated cellular response leading to either G1 arrest or apoptosis. However, genotoxic treatments may induce mutations and translocations that result in secondary malignancies or recurrent disease. In addition, about 50% of human cancers are associated with mutations in the p53 gene. Nongenotoxic activation of apoptosis by targeting specific molecular pathways thus provides an attractive therapeutic approach. Normal and leukemic cells were evaluated for their sensitivity to 5, 6-dichloro-1-beta-D-ribofuranosylbenzimidazole (DRB) through cell viability and caspase activation tests. The apoptotic pathway induced by DRB was analysed by immunfluorescence and immunoblot analysis. H2AX phosphorylation and cell cycle analysis were performed to study the dependance of apoptosis on DNA damage and DNA replication, respectively. To investigate the role of p53 in DRB-induced apoptosis, specific p53 inhibitors were used. Statistical analysis on cell survival was performed with the test of independence. Here we report that DRB, an inhibitor of the transcriptional cyclin-dependent kinases (CDKs) 7 and 9, triggers DNA replication-independent apoptosis in normal and leukemic human cells regardless of their p53 status and without inducing DNA damage. Our data indicate that (i) in p53-competent cells, apoptosis induced by DRB relies on a cytosolic accumulation of p53 and subsequent Bax activation, (ii) in the absence of p53, it may rely on p73, and (iii) it is independent of ATM and NBS1 proteins. Notably, even apoptosis-resistant leukemic cells such as Raji were sensitive to DRB. Our results indicate that DRB represents a potentially useful cancer chemotherapeutic strategy that employs both the p53-dependent and -independent apoptotic pathways without inducing genotoxic stress, thereby decreasing the risk of secondary malignancies

  3. Cyclin B Translation Depends on mTOR Activity after Fertilization in Sea Urchin Embryos

    Science.gov (United States)

    Boulben, Sandrine; Glippa, Virginie; Morales, Julia; Cormier, Patrick

    2016-01-01

    The cyclin B/CDK1 complex is a key regulator of mitotic entry. Using PP242, a specific ATP-competitive inhibitor of mTOR kinase, we provide evidence that the mTOR signalling pathway controls cyclin B mRNA translation following fertilization in Sphaerechinus granularis and Paracentrotus lividus. We show that PP242 inhibits the degradation of the cap-dependent translation repressor 4E-BP (eukaryotic initiation factor 4E-Binding Protein). PP242 inhibits global protein synthesis, delays cyclin B accumulation, cyclin B/CDK1 complex activation and consequently entry into the mitotic phase of the cell cycle triggered by fertilization. PP242 inhibits cyclin B mRNA recruitment into active polysomes triggered by fertilization. An amount of cyclin B mRNA present in active polysomes appears to be insensitive to PP242 treatment. Taken together, our results suggest that, following sea urchin egg fertilization, cyclin B mRNA translation is controlled by two independent mechanisms: a PP242-sensitive and an additional PP242-insentitive mechanism. PMID:26962866

  4. The Drosophila PNG kinase complex regulates the translation of cyclin B.

    Science.gov (United States)

    Vardy, Leah; Orr-Weaver, Terry L

    2007-01-01

    The Drosophila PAN GU (PNG) kinase complex regulates the developmental translation of cyclin B. cyclin B mRNA becomes unmasked during oogenesis independent of PNG activity, but PNG is required for translation from egg activation. We find that although polyadenylation of cyclin B augments translation, it is not essential, and a fully elongated poly(A) is not required for translation to proceed. In fact, changes in poly(A) tail length are not sufficient to account for PNG-mediated control of cyclin B translation and of the early embryonic cell cycles. We present evidence that PNG functions instead as an antagonist of PUMILIO-dependent translational repression. Our data argue that changes in poly(A) tail length are not a universal mechanism governing embryonic cell cycles, and that PNG-mediated derepression of translation is an important alternative mechanism in Drosophila.

  5. Establishment of functioning human corneal endothelial cell line with high growth potential.

    Directory of Open Access Journals (Sweden)

    Tadashi Yokoi

    Full Text Available Hexagonal-shaped human corneal endothelial cells (HCEC form a monolayer by adhering tightly through their intercellular adhesion molecules. Located at the posterior corneal surface, they maintain corneal translucency by dehydrating the corneal stroma, mainly through the Na(+- and K(+-dependent ATPase (Na(+/K(+-ATPase. Because HCEC proliferative activity is low in vivo, once HCEC are damaged and their numbers decrease, the cornea begins to show opacity due to overhydration, resulting in loss of vision. HCEC cell cycle arrest occurs at the G1 phase and is partly regulated by cyclin-dependent kinase inhibitors (CKIs in the Rb pathway (p16-CDK4/CyclinD1-pRb. In this study, we tried to activate proliferation of HCEC by inhibiting CKIs. Retroviral transduction was used to generate two new HCEC lines: transduced human corneal endothelial cell by human papillomavirus type E6/E7 (THCEC (E6/E7 and transduced human corneal endothelial cell by Cdk4R24C/CyclinD1 (THCEH (Cyclin. Reverse transcriptase polymerase chain reaction analysis of gene expression revealed little difference between THCEC (E6/E7, THCEH (Cyclin and non-transduced HCEC, but cell cycle-related genes were up-regulated in THCEC (E6/E7 and THCEH (Cyclin. THCEH (Cyclin expressed intercellular molecules including ZO-1 and N-cadherin and showed similar Na(+/K(+-ATPase pump function to HCEC, which was not demonstrated in THCEC (E6/E7. This study shows that HCEC cell cycle activation can be achieved by inhibiting CKIs even while maintaining critical pump function and morphology.

  6. Novel arylazopyrazole inhibitors of cyclin-dependent kinases

    Czech Academy of Sciences Publication Activity Database

    Jorda, Radek; Schütznerová, E.; Cankař, P.; Brychtová, Veronika; Navrátilová, Jana; Kryštof, Vladimír

    2015-01-01

    Roč. 23, č. 9 (2015), s. 1975-1981 ISSN 0968-0896 R&D Projects: GA ČR GAP305/12/0783; GA ČR GA14-19590S; GA MŠk(CZ) LO1204 Institutional support: RVO:61389030 Keywords : Cyclin-dependent kinases * Inhibitor * Cell cycle Subject RIV: CE - Biochemistry Impact factor: 2.923, year: 2015

  7. Repression of c-Myc responsive genes in cycling cells causes G1 arrest through reduction of cyclin E/CDK2 kinase activity

    NARCIS (Netherlands)

    Berns, K.; Hijmans, E.M.; Bernards, R.A.

    1997-01-01

    The c-myc gene encodes a sequence-specific DNA binding protein involved in proliferation and oncogenesis. Activation of c-myc expression in quiescent cells is sufficient to mediate cell cycle entry, whereas inhibition of c-myc expression causes cycling cells to withdraw from the cell cycle. To

  8. Differential expression of peroxisome proliferator activated receptor gamma and cyclin D1 does not affect proliferation of asthma- and non-asthma-derived airway smooth muscle cells

    NARCIS (Netherlands)

    Lau, Justine Y; Oliver, Brian G; Moir, Lyn M; Black, Judith L; Burgess, Janette K

    UNLABELLED: PPARgamma levels in asthma- and non-asthma-derived airway smooth muscle cells and PPARgamma activation-induced cell proliferation were investigated. In the presence of FBS, PPARgamma levels were higher in subconfluent asthma-derived cells but lower in confluent cells compared with

  9. Acteoside inhibits human promyelocytic HL-60 leukemia cell proliferation via inducing cell cycle arrest at G0/G1 phase and differentiation into monocyte.

    Science.gov (United States)

    Lee, Kyung-Won; Kim, Hyoung Ja; Lee, Yong Sup; Park, Hee-Jun; Choi, Jong-Won; Ha, Joohun; Lee, Kyung-Tae

    2007-09-01

    We investigated the in vitro effects of acteoside on the proliferation, cell cycle regulation and differentiation of HL-60 human promyelocytic leukemia cells. Acteoside inhibited the proliferation of HL-60 cells in a concentration- and time-dependent manner with an IC50, approximately 30 microM. DNA flow cytometric analysis indicated that acteoside blocked cell cycle progression at the G1 phase in HL-60 human promyelocytic leukemia cells. Among the G1 phase cell cycle-related proteins, the levels of cyclin-dependent protein kinase (CDK)2, CDK6, cyclin D1, cyclin D2, cyclin D3 and cyclin E were reduced by acteoside, whereas the steady-state level of CDK4 was unaffected. The protein and mRNA levels of CDK inhibitors (cyclin-dependent kinase inhibitors), such as p21(CIP1/WAF1) and p27(KIP1), were gradually increased after acteoside treatment in a time-dependent manner. In addition, acteoside markedly enhanced the binding of p21(CIP1/WAF1) and p27(KIP1) to CDK4 and CDK6, resulting in the reduction of CDK2, CDK4 and CDK6 activities. Moreover, the hypophosphorylated form of retinoblastoma increased, leading to the enhanced binding of protein retinoblastoma (pRb) and E2F1. Our results further suggest that acteoside is a potent inducer of differentiation of HL-60 cells based on biochemical activities and the expression level of CD14 cell surface antigen. In conclusion, the onset of acteoside-induced G1 arrest of HL-60 cells prior to the differentiation appears to be tightly linked to up-regulation of the p21(CIP1/WAF1) and p27(KIP1) levels and decreases in the CDK2, CDK4 and CDK6 activities. These findings, for the first time, reveal the mechanism underlying the anti-proliferative effect of acteoside on human promyelocytic HL-60 cells.

  10. Molecular biological mechanism II. Molecular mechanisms of cell cycle regulation; Molekularbiologische Mechanismen II. Molekulare Mechanismen der Zellzyklusregulation

    Energy Technology Data Exchange (ETDEWEB)

    Jung, T. [Bundesamt fuer Strahlenschutz Neuherberg (Germany). Institut fuer Strahlenhygiene

    2000-07-01

    The cell cycle in eukaryotes is regulated by central cell cycle controlling protein kinase complexes. These protein kinase complexes consist of a catalytic subunit from the cyclin-dependent protein kinase family (CDK), and a regulatory subunit from the cyclin family. Cyclins are characterised by their periodic cell cycle related synthesis and destruction. Each cell cycle phase is characterised by a specific set of CDKs and cyclins. The activity of CDK/cyclin complexes is mainly regulated on four levels. It is controlled by specific phosphorylation steps, the synthesis and destruction of cyclins, the binding of specific inhibitor proteins, and by active control of their intracellular localisation. At several critical points within the cell cycle, named checkpoints, the integrity of the cellular genome is monitored. If damage to the genome or an unfinished prior cell cycle phase is detected, the cell cycle progression is stopped. These cell cycle blocks are of great importance to secure survival of cells. Their primary importance is to prevent the manifestation and heritable passage of a mutated genome to daughter cells. Damage sensing, DNA repair, cell cycle control and apoptosis are closely linked cellular defence mechanisms to secure genome integrity. Disregulation in one of these defence mechanisms are potentially correlated with an increased cancer risk and therefore in at least some cases with an increased radiation sensitivity. (orig.) [German] Der eukaryotische Zellzyklus wird reguliert durch zentrale Zellzyklus-steuernde Proteinkinase Komplexe. Diese Proteinkomplexe betehen jeweils aus einer katalytischen Untereinheit aus der Familie der Cyclin-abhaengigen Proteinkinasen (CDK) und einer regulatorischen Untereinheit, den Cyclinen, deren Name von der im Zellzyklus periodischen Synthese und Proteolyse herstammt. Jede Zellzyklusphase ist charakterisiert durch eine spezifische Kombination bestimmter CDKs und Cycline. Die Aktivitaet der CDK/Cyclin Komplexe

  11. The Cytotoxic Role of Intermittent High Glucose on Apoptosis and Cell Viability in Pancreatic Beta Cells

    Directory of Open Access Journals (Sweden)

    Zhen Zhang

    2014-01-01

    Full Text Available Objectives. Glucose fluctuations are both strong predictor of diabetic complications and crucial factor for beta cell damages. Here we investigated the effect of intermittent high glucose (IHG on both cell apoptosis and proliferation activity in INS-1 cells and the potential mechanisms. Methods. Cells were treated with normal glucose (5.5 mmol/L, constant high glucose (CHG (25 mmol/L, and IHG (rotation per 24 h in 11.1 or 25 mmol/L for 7 days. Reactive oxygen species (ROS, xanthine oxidase (XOD level, apoptosis, cell viability, cell cycle, and expression of cyclinD1, p21, p27, and Skp2 were determined. Results. We found that IHG induced more significant apoptosis than CHG and normal glucose; intracellular ROS and XOD levels were more markedly increased in cells exposed to IHG. Cells treated with IHG showed significant decreased cell viability and increased cell proportion in G0/G1 phase. Cell cycle related proteins such as cyclinD1 and Skp2 were decreased significantly, but expressions of p27 and p21 were increased markedly. Conclusions. This study suggested that IHG plays a more toxic effect including both apoptosis-inducing and antiproliferative effects on INS-1 cells. Excessive activation of cellular stress and regulation of cyclins might be potential mechanism of impairment in INS-1 cells induced by IHG.

  12. Negative effect of cyclin D1 overexpression on recurrence-free survival in stage II-IIIA lung adenocarcinoma and its expression modulation by vorinostat in vitro.

    Science.gov (United States)

    Lee, Eunju; Jin, DongHao; Lee, Bo Bin; Kim, Yujin; Han, Joungho; Shim, Young Mog; Kim, Duk-Hwan

    2015-12-17

    This study was aimed at identifying prognostic biomarkers for stage II-IIIA non-small cell lung cancer (NSCLC) according to histology and at investigating the effect of vorinostat on the expression of these biomarkers. Expression levels of cyclin D1, cyclin A2, cyclin E, and p16 proteins that are involved in the G1-to-S phase progression of cell cycle were analyzed using immunohistochemistry in formalin-fixed paraffin-embedded tissues from 372 samples of stage II-IIIA NSCLC. The effect of vorinostat on the expression of these proteins, impacts on cell cycle, and histone modification was explored in lung cancer cells. Abnormal expression of cyclin A2, cyclin D1, cyclin E, and p16 was found in 66, 47, 34, and 51 % of 372 cases, respectively. Amongst the four proteins, only cyclin D1 overexpression was significantly associated with poor recurrence-free survival (adjusted hazard ratio = 1.87; 95 % confidence interval = 1.12 - 2.69, P = 0.02) in adenocarcinoma but not in squamous cell carcinoma (P = 0.44). Vorinostat inhibited cell cycle progression to the S-phase and induced down-regulation of cyclin D1 in vitro. The down-regulation of cyclin D1 by vorinostat was comparable to a siRNA-mediated knockdown of cyclin D1 in A549 cells, but vorinostat in the presence of benzo[a]pyrene showed a differential effect in different lung cancer cell lines. Cyclin D1 down-regulation by vorinostat was associated with the accumulation of dimethyl-H3K9 at the promoter of the gene. The present study suggests that cyclin D1 may be an independent prognostic factor for recurrence-free survival in stage II-IIIA adenocarcinoma of lung and its expression may be modulated by vorinostat.

  13. Isolation of a full-length mitotic cyclin cDNA clone CycIIIMs from Medicago sativa: chromosomal mapping and expression.

    Science.gov (United States)

    Savouré, A; Fehér, A; Kaló, P; Petrovics, G; Csanádi, G; Szécsi, J; Kiss, G; Brown, S; Kondorosi, A; Kondorosi, E

    1995-03-01

    Cyclins in association with the protein kinase p34cdc2 and related cyclin-dependent protein kinases (cdks) are key regulatory elements in controlling the cell division cycle. Here, we describe the identification and characterization of a full-length cDNA clone of alfalfa mitotic cyclin, termed CycIIIMs. Computer analysis of known plant cyclin gene sequences revealed that this cyclin belongs to the same structural group as the other known partial alfalfa cyclin sequences. Genetic segregation analysis based on DNA-DNA hybridization data showed that the CycIIIMs gene(s) locates in a single chromosomal region on linkage group 5 of the alfalfa genetic map between RFLP markers UO89A and CG13. The assignment of this cyclin to the mitotic cyclin class was based on its cDNA-derived sequence and its differential expression during G2/M cell cycle phase transition of a partially synchronized alfalfa cell culture. Sequence analysis indicated common motifs with both the A- and B-types of mitotic cyclins similarly to the newly described B3-type of animal cyclins.

  14. Accumulation of cyclin B1 requires E2F and cyclin-A-dependent rearrangement of the anaphase-promoting complex

    DEFF Research Database (Denmark)

    Lukas, C; Sørensen, Claus Storgaard; Kramer, E

    1999-01-01

    In mammalian somatic-cell cycles, progression through the G1-phase restriction point and initiation of DNA replication are controlled by the ability of the retinoblastoma tumour-suppressor protein (pRb) family to regulate the E2F/DP transcription factors. Continuing transcription of E2F target...... in the timely accumulation of cyclin B1 and the coordination of cell-cycle progression during the post-restriction point period....... remains unknown. Here, using an in vivo assay to measure protein stability in real time during the cell cycle, we show that repression of E2F activity or inhibition of cyclin-A-dependent kinase in S phase triggers the destruction of cyclin B1 through the re-assembly of APC, the ubiquitin ligase...

  15. Novel functions of plant cyclin-dependent kinase inhibitors, ICK1/KRP1, can act non-cell-autonomously and inhibit entry into mitosis

    DEFF Research Database (Denmark)

    Weinl, Christina; Marquardt, Sebastian; Kuijt, Suzanne J H

    2005-01-01

    numbers of cells consistent with a function of CKIs in blocking the G1-S cell cycle transition. Here, we demonstrate that at least one inhibitor from Arabidopsis, ICK1/KRP1, can also block entry into mitosis but allows S-phase progression causing endoreplication. Our data suggest that plant CKIs act...... independently from ICK1/KRP1-induced endoreplication. Strikingly, we found that endoreplicated cells were able to reenter mitosis, emphasizing the high degree of flexibility of plant cells during development. Moreover, we show that in contrast with animal CDK inhibitors, ICK1/KRP1 can move between cells...

  16. Progesterone arrested cell cycle progression through progesterone receptor isoform A in pancreatic neuroendocrine neoplasm.

    Science.gov (United States)

    Yazdani, Samaneh; Kasajima, Atsuko; Onodera, Yoshiaki; McNamara, Keely May; Ise, Kazue; Nakamura, Yasuhiro; Tachibana, Tomoyoshi; Motoi, Fuyuhiko; Unno, Michiaki; Sasano, Hironobu

    2018-04-01

    In pancreatic neuroendocrine neoplasms (Pan-NEN) progesterone signaling has been shown to have both inhibitory and stimulatory effects on cell proliferation. The ability of progesterone to inhibit tumor proliferation is of particular interest and is suggested to be mediated through the less abundantly expressed progesterone receptor (PR) isoform A (PRA). To date the mechanistic processes underlying this inhibition of proliferation remain unclear. To examine the mechanism of PRA actions, the human Pan-NEN cell line QGP-1, that endogenously expresses PR isoform B (PRB) without PRA, was transfected with PRA. PRA transfection suppressed the majority of cell cycle related genes increased by progesterone including cyclin A2 (CCNA2), cyclin B1 (CCNB1), cyclin-dependent kinase 1 (CDK1) and cyclin-dependent kinase 2 (CDK2). Importantly, following progesterone administration cell cycle distribution was shifted to S and G2/M phases in the naïve cell line but in PRA-transfected cells, this effect was suppressed. To see if these mechanistic insights were confirmed in patient samples PRA, PRB, CCNA2, CCNB, CDK1 and CDK2 immunoreactivities were assessed in Pan-NEN cases. Higher levels of cell cycle markers were associated with higher WHO grade tumors and correlations between the markers suggested formation of cyclin/CDK activated complexes in S and G2/M phases. PRA expression was associated with inverse correlation of all cell cycle markers. Collectively, these results indicate that progesterone signals through PRA negatively regulates cell cycle progression through suppressing S and G2/M phases and downregulation of cell cycle phases specific cyclins/CDKs. Copyright © 2018 Elsevier Ltd. All rights reserved.

  17. Convergence of mitogenic signalling cascades from diverse classes of receptors at the cyclin D-cyclin-dependent kinase-pRb-controlled G1 checkpoint.

    Science.gov (United States)

    Lukas, J; Bartkova, J; Bartek, J

    1996-12-01

    The commitment of mammalian cells in late G1 to replicate the genome and divide in response to mitogenic growth factors operating via tyrosine kinase receptors depends on phosphorylation of the retinoblastoma protein (pRb), a process controlled by cyclin D-associated cyclin-dependent kinases (cdks) and their inhibitors. This study addressed the issue of whether also other mitogenic signalling cascades require activation of cyclin D-associated kinases or whether any mitogenic pathway can bypass the cyclin D-pRb checkpoint. We show that mitogenic signal transduction pathways from three classes of receptors, the membrane tyrosine kinase receptors activated by serum mitogens or epidermal growth factor, estrogen receptors triggered by estradiol, and the cyclic AMP-dependent signalling from G-protein-coupled thyrotropin receptors, all converge and strictly require the cyclin D-cdk activity to induce S phase in human MCF-7 cells and/or primary dog thyrocytes. Combined microinjection and biochemical approaches showed that whereas these three mitogenic cascades are sensitive to the p16 inhibitor of cdk4/6 and/or cyclin D1-neutralizing antibody and able to induce pRb kinase activity, their upstream biochemical routes are distinct as demonstrated by their differential sensitivity to lovastatin and requirements for mitogen-activated protein kinases whose sustained activation is seen only in the growth factor-dependent pathway. Taken together, these results support the candidacy of the cyclin D-cdk-pRb interplay for the convergence step of multiple signalling cascades and a mechanism contributing to the restriction point switch.

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

    Science.gov (United States)

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

    2016-06-01

    Trypanosoma cruzi, the etiologic agent of Chagas disease, is a protozoan parasite with a life cycle that alternates between replicative and non-replicative forms, but the components and mechanisms that regulate its cell cycle are poorly described. In higher eukaryotes, cyclins are proteins that activate cyclin-dependent kinases (CDKs), by associating with them along the different stages of the cell cycle. These cyclin-CDK complexes exert their role as major modulators of the cell cycle by phosphorylating specific substrates. For the correct progression of the cell cycle, the mechanisms that regulate the activity of cyclins and their associated CDKs are diverse and must be controlled precisely. Different types of cyclins are involved in specific phases of the eukaryotic cell cycle, preferentially activating certain CDKs. In this work, we characterized TcCYC6, a putative coding sequence of T. cruzi which encodes a protein with homology to mitotic cyclins. The overexpression of this sequence, fused to a tag of nine amino acids from influenza virus hemagglutinin (TcCYC6-HA), showed to be detrimental for the proliferation of epimastigotes in axenic culture and affected the cell cycle progression. In silico analysis revealed an N-terminal segment similar to the consensus sequence of the destruction box, a hallmark for the degradation of several mitotic cyclins. We experimentally determined that the TcCYC6-HA turnover decreased in the presence of proteasome inhibitors, suggesting that TcCYC6 degradation occurs via ubiquitin-proteasome pathway. The results obtained in this study provide first evidence that TcCYC6 expression and degradation are finely regulated in T. cruzi. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  19. Effect of red and blue light on the timing of cyclin-dependent kinase activity and the timing of cell division in Chlamydomonas reinhardtii

    Czech Academy of Sciences Publication Activity Database

    Oldenhof, H.; Bišová, Kateřina; Ende, H.; Zachleder, Vilém

    2004-01-01

    Roč. 42, - (2004), s. 341-348 ISSN 0981-9428 R&D Projects: GA ČR GA204/02/1438 Institutional research plan: CEZ:AV0Z5020903 Keywords : blue light * chlamydomonas reingardtii * cell cycle Subject RIV: EA - Cell Biology Impact factor: 1.414, year: 2004

  20. H2O2 induces a transient multi-phase cell cycle arrest in mouse fibroblasts through modulating cyclin D and p21Cip1 expression

    NARCIS (Netherlands)

    Barnouin, K.; Dubuisson, M.L.; Child, E.S.; Fernandez de Mattos, S.; Glassford, J.; Medema, R.H.; Mann, D.J.; Lam, E.W.-F.

    2002-01-01

    From the h CRC Laboratories and the Section of Cancer Cell Biology, Imperial College School of Medicine at Hammersmith Hospital, Du Cane Road, London W12 ONN, United Kingdom, the e Ludwig Institute for Cancer Research and Section of Virology and Cell Biology, Imperial College School of Medicine at

  1. Lawsone inhibits cell growth and improves the efficacy of cisplatin in ...

    African Journals Online (AJOL)

    Cell cycle analysis was done by Flow cytometric studies, Immunoblotting studies for protein expression was done, proteins controlling cell cycle such as cyclinD1, cyclin E, cyclin A, cyclin B1 and Cip1/p21 and p53 which also are cyclin dependent inhibitors of protein kinase were estimated. Annexin V staining was done to ...

  2. 2-Methoxy-4-vinylphenol can induce cell cycle arrest by blocking the hyper-phosphorylation of retinoblastoma protein in benzo[a]pyrene-treated NIH3T3 cells

    Energy Technology Data Exchange (ETDEWEB)

    Jeong, Jin Boo [Bioresource Sciences, Andong National University, Andong 760749 (Korea, Republic of); Jeong, Hyung Jin, E-mail: jhj@andong.ac.kr [Bioresource Sciences, Andong National University, Andong 760749 (Korea, Republic of)

    2010-10-01

    Research highlights: {yields} 2M4VP activated the expression of p21 and p15 protein, and down-regulated the expression of cyclin D1 and cyclin E. {yields} 2M4VP inhibited hyper-phosphorylation of Rb protein. {yields} 2M4VP induced cell cycle arrest from G1 to S. {yields} 2M4VP inhibited hyper-proliferation of the cells in BaP-treated cells. {yields} 2M4VP induces growth arrest of BaP-treated cells by blocking hyper-phosphorylation of Rb via regulating the expression of cell cycle-related proteins. -- Abstract: Benzo[a]pyrene (BaP) is an environment carcinogen that can enhance cell proliferation by disturbing the signal transduction pathways in cell cycle regulation. In this study, the effects of 2M4VP on cell proliferation, cell cycle and cell cycle regulatory proteins were studied in BaP-treated NIH 3T3 cells to establish the molecular mechanisms of 2M4VP as anti-proliferative agents. 2M4VP exerted a dose-dependent inhibitory effect on cell growth correlated with a G1 arrest. Analysis of G1 cell cycle regulators expression revealed 2M4VP increased expression of CDK inhibitor, p21Waf1/Cip1 and p15 INK4b, decreased expression of cyclin D1 and cyclin E, and inhibited kinase activities of CDK4 and CDK2. However, 2M4VP did not affect the expression of CDK4 and CDK2. Also, 2M4VP inhibited the hyper-phosphorylation of Rb induced by BaP. Our results suggest that 2M4VP induce growth arrest of BaP-treated NIH 3T3 cells by blocking the hyper-phosphorylation of Rb via regulating the expression of cell cycle-related proteins.

  3. 2-Methoxy-4-vinylphenol can induce cell cycle arrest by blocking the hyper-phosphorylation of retinoblastoma protein in benzo[a]pyrene-treated NIH3T3 cells

    International Nuclear Information System (INIS)

    Jeong, Jin Boo; Jeong, Hyung Jin

    2010-01-01

    Research highlights: → 2M4VP activated the expression of p21 and p15 protein, and down-regulated the expression of cyclin D1 and cyclin E. → 2M4VP inhibited hyper-phosphorylation of Rb protein. → 2M4VP induced cell cycle arrest from G1 to S. → 2M4VP inhibited hyper-proliferation of the cells in BaP-treated cells. → 2M4VP induces growth arrest of BaP-treated cells by blocking hyper-phosphorylation of Rb via regulating the expression of cell cycle-related proteins. -- Abstract: Benzo[a]pyrene (BaP) is an environment carcinogen that can enhance cell proliferation by disturbing the signal transduction pathways in cell cycle regulation. In this study, the effects of 2M4VP on cell proliferation, cell cycle and cell cycle regulatory proteins were studied in BaP-treated NIH 3T3 cells to establish the molecular mechanisms of 2M4VP as anti-proliferative agents. 2M4VP exerted a dose-dependent inhibitory effect on cell growth correlated with a G1 arrest. Analysis of G1 cell cycle regulators expression revealed 2M4VP increased expression of CDK inhibitor, p21Waf1/Cip1 and p15 INK4b, decreased expression of cyclin D1 and cyclin E, and inhibited kinase activities of CDK4 and CDK2. However, 2M4VP did not affect the expression of CDK4 and CDK2. Also, 2M4VP inhibited the hyper-phosphorylation of Rb induced by BaP. Our results suggest that 2M4VP induce growth arrest of BaP-treated NIH 3T3 cells by blocking the hyper-phosphorylation of Rb via regulating the expression of cell cycle-related proteins.

  4. HCdc14A is involved in cell cycle regulation of human brain vascular endothelial cells following injury induced by high glucose, free fatty acids and hypoxia.

    Science.gov (United States)

    Su, Jingjing; Zhou, Houguang; Tao, Yinghong; Guo, Zhuangli; Zhang, Shuo; Zhang, Yu; Huang, Yanyan; Tang, Yuping; Hu, Renming; Dong, Qiang

    2015-01-01

    Cell cycle processes play a vital role in vascular endothelial proliferation and dysfunction. Cell division cycle protein 14 (Cdc14) is an important cell cycle regulatory phosphatase. Previous studies in budding yeast demonstrated that Cdc14 could trigger the inactivation of mitotic cyclin-dependent kinases (Cdks), which are required for mitotic exit and cytokinesis. However, the exact function of human Cdc14 (hCdc14) in cell cycle regulation during vascular diseases is yet to be elucidated. There are two HCdc14 homologs: hCdc14A and hCdc14B. In the current study, we investigated the potential role of hCdc14A in high glucose-, free fatty acids (FFAs)-, and hypoxia-induced injury in cultured human brain vascular endothelial cells (HBVECs). Data revealed that high glucose, FFA, and hypoxia down-regulated hCdc14A expression remarkably, and also affected the expression of other cell cycle-related proteins such as cyclin B, cyclin D, cyclin E, and p53. Furthermore, the combined addition of the three stimuli largely blocked cell cycle progression, decreased cell proliferation, and increased apoptosis. We also determined that hCdc14A was localized mainly to centrosomes during interphase and spindles during mitosis using confocal microscopy, and that it could affect the expression of other cycle-related proteins. More importantly, the overexpression of hCdc14A accelerated cell cycle progression, enhanced cell proliferation, and promoted neoplastic transformation, whereas the knockdown of hCdc14A using small interfering RNA produced the opposite effects. Therefore, these findings provide novel evidence that hCdc14A might be involved in cell cycle regulation in cultured HBVECs during high glucose-, FFA-, and hypoxia-induced injury. Copyright © 2014 Elsevier Inc. All rights reserved.

  5. Cyclin E mRNA overexpression in epithelial ovarian cancers: inverse correlation with p53 protein accumulation.

    Science.gov (United States)

    Sawasaki, T; Shigemasa, K; Shiroyama, Y; Kusuda, T; Fujii, T; Parmley, T H; O'Brien, T J; Ohama, K

    2001-01-01

    We investigated the relationship between cyclin E mRNA overexpression and p53 protein accumulation in epithelial ovarian cancers. mRNA was isolated and cDNA was prepared from 36 epithelial ovarian tumors (three adenomas, three low malignant potential tumors, and 30 carcinomas), and six normal ovaries. The cyclin E mRNA expression levels relative to an internal control, beta-tubulin, were determined by semiquantitative polymerase chain reaction (PCR). Cyclin E and p53 protein expression in ovarian cancer tissues were examined by immunohistochemistry using the same series of samples. Fisher exact test of significance and an unpaired t test were used for statistical analysis. Considerable levels of cyclin E mRNA were detected in all normal ovaries and ovarian tumor samples examined by semiquantitative PCR amplification. mRNA levels of cyclin E were significantly higher in nine of 30 (30%) ovarian cancers compared with those in normal ovaries. The immunohistochemical expression of cyclin E protein was confirmed in the nuclei of tumor cells in 13 of 30 (43%) ovarian cancers. p53 protein accumulation was detected in 12 of 30 (40%) ovarian cancers examined. There was a significant inverse correlation between cyclin E mRNA overexpression and p53 protein accumulation (P ovarian cancers without p53 protein accumulation. Cyclin E might have an important effect on the development of a limited number of ovarian cancers.

  6. Cyclin D1 overexpression increases susceptibility to 4-nitroquinoline 1-oxide induced dysplasia and neoplasia in murine squamous oral epithelium

    Science.gov (United States)

    Wilkey, Jonathan F.; Buchberger, Glenn; Saucier, Kirsten; Patel, Salony M.; Eisenberg, Ellen; Nakagawa, Hiroshi; Michaylira, Carmen; Rustgi, Anil K.; Mallya, Sanjay M.

    2009-01-01

    The cyclin D1 oncogene is frequently amplified/overexpressed in oral squamous cell carcinomas. Mice with overexpression of cyclin D1 targeted to the stratified squamous epithelia of the tongue, esophagus and forestomach develop a phenotype of epithelial dysplasia at these sites. In this study, we examined the effect cyclin D1 overexpression on susceptibility of mice to carcinogen-induced tumorigenesis, using 4-nitroquinoline-1-oxide (4NQO), an established potent oral carcinogen in mice. Cyclin D1 overexpressing mice and non-transgenic littermates were administered 4NQO (20 ppm or 50 ppm in the drinking water) for 8 weeks and monitored for an additional 16 weeks. Histopathological analyses of the tongue revealed significantly higher severity of dysplasia in the cyclin D1 overexpression mice, compared with non-transgenic controls and with untreated controls. Moreover, only the cyclin D1 overexpression mice developed neoplastic lesions in the oro-esophageal epithelia. Examination of the dysplastic and neoplastic lesions reveled abnormal proliferation. Our findings suggest that cyclin D1 overexpression enhances susceptibility to carcinogen-induced oral tumorigenesis. These results underscore the importance of cyclin D1 in the process of oral neoplastic development. Further, they emphasize the value of this transgenic model to study the pathogenesis of oral precancer and cancer and establish it as a model system to test candidate agents for chemoprevention of upper aero-digestive cancer. PMID:19263437

  7. Cyclin D1 and p22ack1 play opposite roles in plant growth and development

    International Nuclear Information System (INIS)

    Cho, Jeong Woo; Park, Sun Chung; Shin, Eun Ah; Kim, Chong Ki; Han, Woong; Sohn, Soo-In; Song, Pill Soon; Wang, Myeong Hyeon

    2004-01-01

    The plant cell division cycle, a highly coordinated process, is continually regulated during the growth and development of plants. In this report, we demonstrate how two cell-cycle regulators act together to control cell proliferation in transgenic Arabidopsis. To identify potential cyclin dependent kinase regulators from Arabidopsis, we employed an two-hybrid screening system to isolate genes encoding G1 specific cyclin-interacting proteins. One of these, p22 ack1 , which encodes a novel 22 kDa protein, binds to cyclin D1. Overexpression of p22 ack1 in transgenic Arabidopsis resulted in growth retardation due to a strong inhibition of cell division in the leaf primordial and meristematic tissue. The leaf shape of p22 ack1 transgenic Arabidopsis was altered from oval in wild-type to dentate. Wild-type phenotype was successfully restored in F1 hybrids by cross-hybridizing the p22 ackl Arabidopsis mutants with cyclin D1. Taken together, these results suggest that p22 ack1 and cyclin D1, which act antagonistically, are major rate-limiting factors for cell division in the leaf meristem

  8. Growth inhibition and cell cycle arrest effects of epigallocatechin gallate in the NBT-II bladder tumour cell line.

    Science.gov (United States)

    Chen, J J; Ye, Z-Q; Koo, M W L

    2004-05-01

    To examine the growth inhibition and cell cycle arrest effects of epigallocatechin gallate (EGCG), a major constituent of green tea polyphenols, on the NBT-II bladder tumour cell line. Growth inhibition and cell cycle arrest effects of EGCG were evaluated by the tetrazolium assay, flow cytometry and apoptotic DNA ladder tests. The cell cycle-related oncogene and protein expressions in NBT-II bladder tumour cells, when incubated with EGCG, were detected with reverse transcription-polymerase chain reaction (RT-PCR) and Western blot analysis. EGCG inhibited growth of the NBT-II bladder tumour cells in a dose- and time-dependent manner. Flow cytometry showed a G0/G1 arrest in cells when cultured with EGCG at doses of 10, 20 or 40 micro mol/L for 48 or 72 h. The apoptotic DNA ladder test showed that EGCG at 10 micro mol/L induced early apoptosis after 48 h of incubation. A down-regulation of cyclin D1 was detected by RT-PCR when the cells were incubated with EGCG (20 micro mol/L for 48 h. EGCG also down-regulated protein expression of cyclin D1, cyclin-dependent kinase 4/6 and phosphorylated retinoblastoma protein, in both a time- and dose-dependent manner, when detected by Western blot. EGCG had growth inhibition and cell-cycle arrest effects in NBT-II bladder tumour cells by down-regulating the cyclin D1, cyclin-dependent kinase 4/6 and retinoblastoma protein machinery for regulating cell-cycle progression.

  9. Involvement of cyclin K posttranscriptional regulation in the formation of Artemia diapause cysts.

    Directory of Open Access Journals (Sweden)

    Yang Zhao

    Full Text Available BACKGROUND: Artemia eggs tend to develop ovoviviparously to yield nauplius larvae in good rearing conditions; while under adverse situations, they tend to develop oviparously and encysted diapause embryos are formed instead. However, the intrinsic mechanisms regulating this process are not well understood. PRINCIPAL FINDING: This study has characterized the function of cyclin K, a regulatory subunit of the positive transcription elongation factor b (P-TEFb in the two different developmental pathways of Artemia. In the diapause-destined embryo, Western blots showed that the cyclin K protein was down-regulated as the embryo entered dormancy and reverted to relatively high levels of expression once development resumed, consistent with the fluctuations in phosphorylation of position 2 serines (Ser2 in the C-terminal domain (CTD of the largest subunit (Rpb1 of RNA polymerase II (RNAP II. Interestingly, the cyclin K transcript levels remained constant during this process. In vitro translation data indicated that the template activity of cyclin K mRNA stored in the postdiapause cyst was repressed. In addition, in vivo knockdown of cyclin K in developing embryos by RNA interference eliminated phosphorylation of the CTD Ser2 of RNAP II and induced apoptosis by inhibiting the extracellular signal-regulated kinase (ERK survival signaling pathway. CONCLUSIONS/SIGNIFICANCE: Taken together, these findings reveal a role for cyclin K in regulating RNAP II activity during diapause embryo development, which involves the post-transcriptional regulation of cyclin K. In addition, a further role was identified for cyclin K in regulating the control of cell survival during embryogenesis through ERK signaling pathways.

  10. Cyclin G2 suppresses estrogen-mediated osteogenesis through inhibition of Wnt/β-catenin signaling.

    Directory of Open Access Journals (Sweden)

    Jinlan Gao

    Full Text Available Estrogen plays an important role in the maintenance of bone formation, and deficiency in the production of estrogen is directly linked to postmenopausal osteoporosis. To date, the underlying mechanisms of estrogen-mediated osteogenic differentiation are not well understood. In this study, a pluripotent mesenchymal precursor cell line C2C12 was used to induce osteogenic differentiation and subjected to detection of gene expressions or to manipulation of cyclin G2 expressions. C57BL/6 mice were used to generate bilateral ovariectomized and sham-operated mice for analysis of bone mineral density and protein expression. We identified cyclin G2, an unconventional member of cyclin, is involved in osteoblast differentiation regulated by estrogen in vivo and in vitro. In addition, the data showed that ectopic expression of cyclin G2 suppressed expression of osteoblast transcription factor Runx2 and osteogenic differentiation marker genes, as well as ALP activity and in vitro extracellular matrix mineralization. Mechanistically, Wnt/β-catenin signaling pathway is essential for cyclin G2 to inhibit osteogenic differentiation. To the best of our knowledge, the current study presents the first evidence that cyclin G2 serves as a negative regulator of both osteogenesis and Wnt/β-catenin signaling. Most importantly, the basal and 17β-estradiol-induced osteogenic differentiation was restored by overexpression of cyclin G2. These results taken together suggest that cyclin G2 may function as an endogenous suppressor of estrogen-induced osteogenic differentiation through inhibition of Wnt/β-catenin signaling.

  11. Phosphorylation of Rad9 at serine 328 by cyclin A-Cdk2 triggers apoptosis via interfering Bcl-xL.

    Directory of Open Access Journals (Sweden)

    Zhuo Zhan

    Full Text Available Cyclin A-Cdk2, a cell cycle regulated Ser/Thr kinase, plays important roles in a variety of apoptoticprocesses. However, the mechanism of cyclin A-Cdk2 regulated apoptosis remains unclear. Here, we demonstrated that Rad9, a member of the BH3-only subfamily of Bcl-2 proteins, could be phosphorylated by cyclin A-Cdk2 in vitro and in vivo. Cyclin A-Cdk2 catalyzed the phosphorylation of Rad9 at serine 328 in HeLa cells during apoptosis induced by etoposide, an inhibitor of topoisomeraseII. The phosphorylation of Rad9 resulted in its translocation from the nucleus to the mitochondria and its interaction with Bcl-xL. The forced activation of cyclin A-Cdk2 in these cells by the overexpression of cyclin A,triggered Rad9 phosphorylation at serine 328 and thereby promoted the interaction of Rad9 with Bcl-xL and the subsequent initiation of the apoptotic program. The pro-apoptotic effects regulated by the cyclin A-Cdk2 complex were significantly lower in cells transfected with Rad9S328A, an expression vector that encodes a Rad9 mutant that is resistant to cyclin A-Cdk2 phosphorylation. These findings suggest that cyclin A-Cdk2 regulates apoptosis through a mechanism that involves Rad9phosphorylation.

  12. The Gcn2 Regulator Yih1 Interacts with the Cyclin Dependent Kinase Cdc28 and Promotes Cell Cycle Progression through G2/M in Budding Yeast.

    Directory of Open Access Journals (Sweden)

    Richard C Silva

    Full Text Available The Saccharomyces cerevisiae protein Yih1, when overexpressed, inhibits the eIF2 alpha kinase Gcn2 by competing for Gcn1 binding. However, deletion of YIH1 has no detectable effect on Gcn2 activity, suggesting that Yih1 is not a general inhibitor of Gcn2, and has no phenotypic defect identified so far. Thus, its physiological role is largely unknown. Here, we show that Yih1 is involved in the cell cycle. Yeast lacking Yih1 displays morphological patterns and DNA content indicative of a delay in the G2/M phases of the cell cycle, and this phenotype is independent of Gcn1 and Gcn2. Accordingly, the levels of phosphorylated eIF2α, which show a cell cycle-dependent fluctuation, are not altered in cells devoid of Yih1. We present several lines of evidence indicating that Yih1 is in a complex with Cdc28. Yih1 pulls down endogenous Cdc28 in vivo and this interaction is enhanced when Cdc28 is active, suggesting that Yih1 modulates the function of Cdc28 in specific stages of the cell cycle. We also demonstrate, by Bimolecular Fluorescence Complementation, that endogenous Yih1 and Cdc28 interact with each other, confirming Yih1 as a bona fide Cdc28 binding partner. Amino acid substitutions within helix H2 of the RWD domain of Yih1 enhance Yih1-Cdc28 association. Overexpression of this mutant, but not of wild type Yih1, leads to a phenotype similar to that of YIH1 deletion, supporting the view that Yih1 is involved through Cdc28 in the regulation of the cell cycle. We further show that IMPACT, the mammalian homologue of Yih1, interacts with CDK1, the mammalian counterpart of Cdc28, indicating that the involvement with the cell cycle is conserved. Together, these data provide insights into the cellular function of Yih1/IMPACT, and provide the basis for future studies on the role of this protein in the cell cycle.

  13. Prevention, Evaluation, and Rehabilitation of Cycling-Related Injury.

    Science.gov (United States)

    Kotler, Dana H; Babu, Ashwin N; Robidoux, Greg

    2016-01-01

    The unique quality of the bicycle is its ability to accommodate a wide variety of injuries and disabilities. Cycling for recreation, transportation, and competition is growing nationwide, and has proven health and societal benefits. The demands of each type of cycling dictate the necessary equipment, as well as potential for injury. Prevention of cycling-related injury in both the athlete and the recreational cyclist involves understanding the common mechanisms for both traumatic and overuse injury, and early correction of strength and flexibility imbalances, technique errors, and bicycle fit.

  14. Signal transducer and activator of transcription 5 activation is sufficient to drive transcriptional induction of cyclin D2 gene and proliferation of rat pancreatic beta-cells

    DEFF Research Database (Denmark)

    Friedrichsen, Birgitte N; Richter, Henrijette E; Hansen, Johnny A

    2003-01-01

    Signal transducer and activator of transcription 5 (STAT5) activation plays a central role in GH- and prolactin-mediated signal transduction in the pancreatic beta-cells. In previous experiments we demonstrated that STAT5 activation is necessary for human (h)GH-stimulated proliferation of INS-1 c...

  15. Potent induction of wild-type p53-dependent transcription in tumour cells by a synthetic inhibitor of cyclin-dependent kinases

    Czech Academy of Sciences Publication Activity Database

    Kotala, V.; Uldrijan, S.; Horký, M.; Trbušek, M.; Strnad, Miroslav; Vojtěšek, B.

    2001-01-01

    Roč. 58, č. 9 (2001), s. 1333-1339 ISSN 1420-682X R&D Projects: GA ČR GA301/02/0475 Institutional research plan: CEZ:AV0Z5038910 Keywords : Roscovitine * transcriptional activity * tumour cells Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 4.539, year: 2001

  16. Cyclin E-Mediated Human Proopiomelanocortin Regulation as a Therapeutic Target for Cushing Disease.

    Science.gov (United States)

    Liu, Ning-Ai; Araki, Takako; Cuevas-Ramos, Daniel; Hong, Jiang; Ben-Shlomo, Anat; Tone, Yukiko; Tone, Masahide; Melmed, Shlomo

    2015-07-01

    Cushing disease, due to pituitary corticotroph tumor ACTH hypersecretion, drives excess adrenal cortisol production with adverse morbidity and mortality. Loss of glucocorticoid negative feedback on the hypothalamic-pituitary-adrenal axis leads to autonomous transcription of the corticotroph precursor hormone proopiomelanocortin (POMC), consequent ACTH overproduction, and adrenal hypercortisolism. We previously reported that R-roscovitine (CYC202, seliciclib), a 2,6,9-trisubstituted purine analog, suppresses cyclin-dependent-kinase 2/cyclin E and inhibits ACTH in mice and zebrafish. We hypothesized that intrapituitary cyclin E signaling regulates corticotroph tumor POMC transcription independently of cell cycle progression. The aim was to investigate whether R-roscovitine inhibits human ACTH in corticotroph tumors by targeting the cyclin-dependent kinase 2/cyclin E signaling pathway. Primary cell cultures of surgically resected human corticotroph tumors were treated with or without R-roscovitine, ACTH measured by RIA and quantitative PCR, and/or Western blot analysis performed to investigate ACTH and lineage-specific transcription factors. Cyclin E and E2F transcription factor 1 (E2F1) small interfering RNA (siRNA) transfection was performed in murine corticotroph tumor AtT20 cells to elucidate mechanisms for drug action. POMC gene promoter activity in response to R-roscovitine treatment was analyzed using luciferase reporter and chromatin immunoprecipitation assays. R-roscovitine inhibits human corticotroph tumor POMC and Tpit/Tbx19 transcription with decreased ACTH expression. Cyclin E and E2F1 exhibit reciprocal positive regulation in corticotroph tumors. R-roscovitine disrupts E2F1 binding to the POMC gene promoter and suppresses Tpit/Tbx19 and other lineage-specific POMC transcription cofactors via E2F1-dependent and -independent pathways. R-roscovitine inhibits human pituitary corticotroph tumor ACTH by targeting the cyclin E/E2F1 pathway. Pituitary cyclin E

  17. Circadian variation in expression of G1 phase cyclins D1 and E and cyclin-dependent kinase inhibitors p16 and p21 in human bowel mucosa

    Science.gov (United States)

    Griniatsos, John; Michail, Othon P; Theocharis, Stamatios; Arvelakis, Antonios; Papaconstantinou, Ioannis; Felekouras, Evangelos; Pikoulis, Emmanouel; Karavokyros, Ioannis; Bakoyiannis, Chris; Marinos, George; Bramis, John; Michail, Panayiotis O

    2006-01-01

    AIM: To evaluate whether the cellular proliferation rate in the large bowel epithelial cells is characterized by circadian rhythm. METHODS: Between January 2003 and December 2004, twenty patients who were diagnosed as suffering from primary, resectable, non-metastatic adenocarcinoma of the lower rectum, infiltrating the sphincter mechanism, underwent abdominoperineal resection, total mesorectal excision and permanent left iliac colostomy. In formalin-fixed and paraffin-embedded biopsy specimens obtained from the colostomy mucosa every six hours (00:00, 06:00, 12:00, 18:00 and 24:00), we studied the expression of G1 phase cyclins (D1 and E) as well as the expression of the G1 phase cyclin-dependent kinase (CDK) inhibitors p16 and p21 as indicators of cell cycle progression in colonic epithelial cells using immunohistochemical methods. RESULTS: The expression of both cyclins showed a similar circadian fashion obtaining their lowest and highest values at 00:00 and 18:00, respectively (P< 0.001). A circadian rhythm in the expression of CDK inhibitor proteins p16 and p21 was also observed, with the lowest levels obtained at 12:00 and 18:00 (P< 0.001), respectively. When the complexes cyclins D1 - p21 and E - p21 were examined, the expression of the cyclins was adversely correlated to the p21 expression throughout the day. When the complexes the cyclins D1 - p16 and E - p16 were examined, high levels of p16 expression were correlated to low levels of cyclin expression at 00:00, 06:00 and 24:00. Meanwhile, the highest expression levels of both cyclins were correlated to high levels of p16 expression at 18:00. CONCLUSION: Colonic epithelial cells seem to enter the G1 phase of the cell cycle during afternoon (between 12:00 and 18:00) with the highest rates obtained at 18:00. From a clinical point of view, the present results suggest that G1-phase specific anticancer therapies in afternoon might maximize their anti-tumor effect while minimizing toxicity

  18. Cyclin D activates the Rb tumor suppressor by mono-phosphorylation.

    Science.gov (United States)

    Narasimha, Anil M; Kaulich, Manuel; Shapiro, Gary S; Choi, Yoon J; Sicinski, Piotr; Dowdy, Steven F

    2014-06-04

    The widely accepted model of G1 cell cycle progression proposes that cyclin D:Cdk4/6 inactivates the Rb tumor suppressor during early G1 phase by progressive multi-phosphorylation, termed hypo-phosphorylation, to release E2F transcription factors. However, this model remains unproven biochemically and the biologically active form(s) of Rb remains unknown. In this study, we find that Rb is exclusively mono-phosphorylated in early G1 phase by cyclin D:Cdk4/6. Mono-phosphorylated Rb is composed of 14 independent isoforms that are all targeted by the E1a oncoprotein, but show preferential E2F binding patterns. At the late G1 Restriction Point, cyclin E:Cdk2 inactivates Rb by quantum hyper-phosphorylation. Cells undergoing a DNA damage response activate cyclin D:Cdk4/6 to generate mono-phosphorylated Rb that regulates global transcription, whereas cells undergoing differentiation utilize un-phosphorylated Rb. These observations fundamentally change our understanding of G1 cell cycle progression and show that mono-phosphorylated Rb, generated by cyclin D:Cdk4/6, is the only Rb isoform in early G1 phase.

  19. Molecular cloning and chromosomal localization of the human cyclin C (CCNC) and cyclin E (CCNE) genes: Deletion of the CCNC gene in human tumors

    Energy Technology Data Exchange (ETDEWEB)

    Li, Haimin; Lahti, J.M.; Kidd, V.J. [St. Jude Children`s Research Hospital, Memphis, TN (United States)] [and others

    1996-03-01

    The human G1-phase cyclins are important regulators of cell cycle progression that interact with various cyclin-dependent kinases and facilitate entry into S-phase. We have confirmed the localization of the human cyclin C (CCNC) gene to chromosome 6q21 and of human cyclin E (CCNE to 19q12). The CCNC gene structure was also determined, and we have shown that it is deleted in a subset of acute lymphoblastic leukemias, including a patient sample containing a t(2;6)(p21;q15), with no apparent cytogenetic deletion. Single-strand conformational polymorphism analysis of the remaining CCNC allele from patients with a deletion of one allele established that there were no further mutations within the exons or the flanking intronic sequences. These results suggest either that haploinsufficiency of the cyclin C protein is sufficient to promote tumorigenesis or that the important tumor suppressor gene is linked to the CCNC locus. 48 refs., 4 figs., 1 tab.

  20. Cyclin E-induced S phase without activation of the pRb/E2F pathway

    DEFF Research Database (Denmark)

    Lukas, J; Herzinger, T; Hansen, Klaus

    1997-01-01

    In cells of higher eukaryotes, cyclin D-dependent kinases Cdk4 and Cdk6 and, possibly, cyclin E-dependent Cdk2 positively regulate the G1- to S-phase transition, by phosphorylating the retinoblastoma protein (pRb), thereby releasing E2F transcription factors that control S-phase genes. Here we...... performed microinjection and transfection experiments using rat R12 fibroblasts, their derivatives conditionally overexpressing cyclins D1 or E, and human U-2-OS cells, to explore the action of G1 cyclins and the relationship of E2F and cyclin E in S-phase induction. We demonstrate that ectopic expression...... that the cyclin E-induced S phase and completion of the cell division cycle can occur in the absence of E2F-mediated transactivation. Together with the ability of cyclin E to overcome a G1 block induced by expression of dominant-negative mutant DP-1, a heterodimeric partner of E2Fs, these results provide evidence...

  1. Cyclin E-induced S phase without activation of the pRb/E2F pathway

    DEFF Research Database (Denmark)

    Lukas, J; Herzinger, T; Hansen, Klaus

    1997-01-01

    In cells of higher eukaryotes, cyclin D-dependent kinases Cdk4 and Cdk6 and, possibly, cyclin E-dependent Cdk2 positively regulate the G1- to S-phase transition, by phosphorylating the retinoblastoma protein (pRb), thereby releasing E2F transcription factors that control S-phase genes. Here we...... performed microinjection and transfection experiments using rat R12 fibroblasts, their derivatives conditionally overexpressing cyclins D1 or E, and human U-2-OS cells, to explore the action of G1 cyclins and the relationship of E2F and cyclin E in S-phase induction. We demonstrate that ectopic expression...... of cyclin E, but not cyclin D1, can override G1 arrest imposed by either the p16INK4a Cdk inhibitor specific for Cdk4 and Cdk6 or a novel phosphorylation-deficient mutant pRb. Several complementary approaches to assess E2F activation, including quantitative reporter assays in live cells, showed...

  2. Stromal and Epithelial Caveolin-1 Both Confer a Protective Effect Against Mammary Hyperplasia and Tumorigenesis : Caveolin-1 Antagonizes Cyclin D1 Function in Mammary Epithelial Cells

    OpenAIRE

    Williams, Terence M.; Sotgia, Federica; Lee, Hyangkyu; Hassan, Ghada; Di Vizio, Dolores; Bonuccelli, Gloria; Capozza, Franco; Mercier, Isabelle; Rui, Hallgeir; Pestell, Richard G.; Lisanti, Michael P.

    2006-01-01

    Here, we investigate the role of caveolin-1 (Cav-1) in breast cancer onset and progression, with a focus on epithelial-stromal interactions, ie, the tumor microenvironment. Cav-1 is highly expressed in adipocytes and is abundant in mammary fat pads (stroma), but it remains unknown whether loss of Cav-1 within mammary stromal cells affects the differentiated state of mammary epithelia via paracrine signaling. To address this issue, we characterized the development of the mammary ductal system ...

  3. Inhibitor of CDK interacting with cyclin A1 (INCA1) regulates proliferation and is repressed by oncogenic signaling

    DEFF Research Database (Denmark)

    Baumer, Nicole; Tickenbrock, Lara; Tschanter, Petra

    2011-01-01

    The cell cycle is driven by the kinase activity of cyclin/CDK complexes which is negatively regulated by CDK inhibitor proteins. Recently, we identified INCA1 as interaction partner and substrate of cyclin A1 in complex with CDK2. On a functional level, we identified a novel cyclin binding site...... in the INCA1 protein. INCA1 inhibited CDK2 activity and cell proliferation. The inihibitory effects depended on the cyclin-interacting domain. Mitogenic and oncogenic signals suppressed INCA1 expression, while it was induced by cell cycle arrest. We established a deletional mouse model that showed increased...... CDK2 activity in spleen with altered spleen architecture in Inca1-/- mice. Inca1-/- embryonic fibroblasts showed an increase in the fraction of S-phase cells. Furthermore, blasts from ALL and AML patients expressed significantly reduced INCA1 levels highlighting its relevance for growth control...

  4. Molecular Dissection of the 8 Phase Transcriptional Program Controlled by Cyclin E-P220 NPAT Signaling Pathway

    National Research Council Canada - National Science Library

    Nalepa, Grzegorz; Harper, Jeffrey

    2005-01-01

    Cyclin E/Cdk2, a central regulator of the G1/S transition, coordinates multiple cell cycle events, including DNA replication, centrosome duplication, and activation of the E2F transcriptional program...

  5. Cyclin A degradation by primate cytomegalovirus protein pUL21a counters its innate restriction of virus replication.

    Directory of Open Access Journals (Sweden)

    Nicolas Caffarelli

    Full Text Available Cyclin A is critical for cellular DNA synthesis and S phase progression of the cell cycle. Human cytomegalovirus (HCMV can reduce cyclin A levels and block cellular DNA synthesis, and cyclin A overexpression can repress HCMV replication. This interaction has only been previously observed in HCMV as murine CMV does not downregulate cyclin A, and the responsible viral factor has not been identified. We previously reported that the HCMV protein pUL21a disrupted the anaphase-promoting complex (APC, but a point mutant abrogating this activity did not phenocopy a UL21a-deficient virus, suggesting that pUL21a has an additional function. Here we identified a conserved arginine-x-leucine (RxL cyclin-binding domain within pUL21a, which allowed pUL21a to interact with cyclin A and target it for proteasome degradation. Homologous pUL21a proteins from both chimpanzee and rhesus CMVs also contained the RxL domain and similarly degraded cyclin A, indicating that this function is conserved in primate CMVs. The RxL point mutation disabled the virus' ability to block cellular DNA synthesis and resulted in a growth defect similar to pUL21a-deficient virus. Importantly, knockdown of cyclin A rescued growth of UL21a-deficient virus. Together, these data show that during evolution, the pUL21a family proteins of primate CMVs have acquired a cyclin-binding domain that targets cyclin A for degradation, thus neutralizing its restriction on virus replication. Finally, the combined proteasome-dependent degradation of pUL21a and its cellular targets suggests that pUL21a may act as a novel suicide protein, targeting its protein cargos for destruction.

  6. Mammalian E-type cyclins control chromosome pairing, telomere stability and CDK2 localization in male meiosis.

    Directory of Open Access Journals (Sweden)

    Laetitia Martinerie

    2014-02-01

    Full Text Available Loss of function of cyclin E1 or E2, important regulators of the mitotic cell cycle, yields viable mice, but E2-deficient males display reduced fertility. To elucidate the role of E-type cyclins during spermatogenesis, we characterized their expression patterns and produced additional deletions of Ccne1 and Ccne2 alleles in the germline, revealing unexpected meiotic functions. While Ccne2 mRNA and protein are abundantly expressed in spermatocytes, Ccne1 mRNA is present but its protein is detected only at low levels. However, abundant levels of cyclin E1 protein are detected in spermatocytes deficient in cyclin E2 protein. Additional depletion of E-type cyclins in the germline resulted in increasingly enhanced spermatogenic abnormalities and corresponding decreased fertility and loss of germ cells by apoptosis. Profound meiotic defects were observed in spermatocytes, including abnormal pairing and synapsis of homologous chromosomes, heterologous chromosome associations, unrepaired double-strand DNA breaks, disruptions in telomeric structure and defects in cyclin-dependent-kinase 2 localization. These results highlight a new role for E-type cyclins as important regulators of male meiosis.

  7. Problem-Solving Test: Analysis of the Role of Cyclin B

    Science.gov (United States)

    Szeberenyi, Jozsef

    2011-01-01

    An experiment is described in this test that was designed to study the role of the cyclin B protein in a cell-free system. The work was performed in the lab of Tim Hunt who, together with Hartwell and Nurse, received the Nobel Prize in Physiology or Medicine in 2001 "for their discoveries of key chemicals that regulate the cell division cycle." It…

  8. Cyclin D1 Expression and the Inhibitory Effect of Celecoxib on Ovarian Tumor Growth in Vivo

    Directory of Open Access Journals (Sweden)

    Ling-Yun Zhai

    2010-10-01

    Full Text Available The report aims to investigate the relationship between the expression of cyclin D1 and Cyclooxgenase-2 (COX-2, thus to explore the molecular mechanisms of the antitumor efficacy of Celecoxib, a COX-2 inhibitor. Human ovarian SKOV-3 carcinoma cell xenograft-bearing mice were treated with Celecoxib by infusing gaster (i.g. twice/day for 21 days. The mRNA levels of COX-2 and cyclin D1 were determined by RT-PCR. The expression of cyclin D1 at the protein level was detected by immunohistochemistry, while COX-2 protein expression was determined by Western blot. A high-dose of Celecoxib (100 mg/kg significantly inhibited tumor growth (P < 0.05, and the expression of cyclin D1 was reduced by 61%. Celecoxib decreased the proliferation cell index by 40% (P < 0.001 and increased apoptotic index by 52% (P < 0.05 in high-dose Celecoxib treated group. Our results suggest that the antitumor efficacy of Celecoxib against ovarian cancer in mice may in part be mediated through suppression of cyclin D1, which may contribute to its ability to suppress proliferation.

  9. Oncolytic adenovirus targeting cyclin E overexpression repressed tumor growth in syngeneic immunocompetent mice

    International Nuclear Information System (INIS)

    Cheng, Pei-Hsin; Rao, Xiao-Mei; Wechman, Stephen L.; Li, Xiao-Feng; McMasters, Kelly M.; Zhou, Heshan Sam

    2015-01-01

    Clinical trials have indicated that preclinical results obtained with human tumor xenografts in mouse models may overstate the potential of adenovirus (Ad)-mediated oncolytic therapies. We have previously demonstrated that the replication of human Ads depends on cyclin E dysregulation or overexpression in cancer cells. ED-1 cell derived from mouse lung adenocarcinomas triggered by transgenic overexpression of human cyclin E may be applied to investigate the antitumor efficacy of oncolytic Ads. Ad-cycE was used to target cyclin E overexpression in ED-1 cells and repress tumor growth in a syngeneic mouse model for investigation of oncolytic virotherapies. Murine ED-1 cells were permissive for human Ad replication and Ad-cycE repressed ED-1 tumor growth in immunocompetent FVB mice. ED-1 cells destroyed by oncolytic Ads in tumors were encircled in capsule-like structures, while cells outside the capsules were not infected and survived the treatment. Ad-cycE can target cyclin E overexpression in cancer cells and repress tumor growth in syngeneic mouse models. The capsule structures formed after Ad intratumoral injection may prevent viral particles from spreading to the entire tumor. The online version of this article (doi:10.1186/s12885-015-1731-x) contains supplementary material, which is available to authorized users

  10. NF-{kappa}B p65 represses {beta}-catenin-activated transcription of cyclin D1

    Energy Technology Data Exchange (ETDEWEB)

    Hwang, Injoo; Choi, Yong Seok; Jeon, Mi-Ya [National Research Lab for RNA Cell Biology, BK21 Graduate Program for RNA Biology, Institute of Nanosensor and Biotechnology and Department of Molecular Biology, Dankook University, Gyeonggi-do 448-701 (Korea, Republic of); Jeong, Sunjoo, E-mail: sjsj@dankook.ac.kr [National Research Lab for RNA Cell Biology, BK21 Graduate Program for RNA Biology, Institute of Nanosensor and Biotechnology and Department of Molecular Biology, Dankook University, Gyeonggi-do 448-701 (Korea, Republic of)

    2010-12-03

    Research highlights: {yields} Cyclin D1 transcription is directly activated by {beta}-catenin; however, {beta}-catenin-induced cyclin D1 transcription is reduced by NF-{kappa}B p65. {yields} Protein-protein interaction between NF-{kappa}B p65 and {beta}-catenin might be responsible for p65-mediated repression of cyclin D1. {yields} One of five putative binding sites, located further upstream of other sites, is the major {beta}-catenin binding site in the cyclin D1 promoter. {yields} NF-{kappa}B binding site in cyclin D1 is occupied not only by p65 but also by {beta}-catenin, which is dynamically regulated by the signal. -- Abstract: Signaling crosstalk between the {beta}-catenin and NF-{kappa}B pathways represents a functional network. To test whether the crosstalk also occurs on their common target genes, the cyclin D1 promoter was used as a model because it contains binding sites for both proteins. {beta}-catenin activated transcription from the cyclin D1 promoter, while co-expression of NF-{kappa}B p65 reduced {beta}-catenin-induced transcription. Chromatin immunoprecipitation revealed lithium chloride-induced binding of {beta}-catenin on one of the T-cell activating factor binding sites. More interestingly, {beta}-catenin binding was greatly reduced by NF-{kappa}B p65, possibly by the protein-protein interaction between the two proteins. Such a dynamic and complex binding of {beta}-catenin and NF-{kappa}B on promoters might contribute to the regulated expression of their target genes.

  11. Basaloid Squamous Cell Carcinoma of the Head and Neck: Subclassification into Basal, Ductal, and Mixed Subtypes Based on Comparison of Clinico-pathologic Features and Expression of p53, Cyclin D1, Epidermal Growth Factor Receptor, p16, and Human Papillomavirus

    Directory of Open Access Journals (Sweden)

    Kyung-Ja Cho

    2017-07-01

    Full Text Available Background Basaloid squamous cell carcinoma (BSCC is a rare variant of squamous cell carcinoma with distinct pathologic characteristics. The histogenesis of BSCC is not fully understood, and the cancer has been suggested to originate from a totipotent primitive cell in the basal cell layer of the surface epithelium or in the proximal duct of secretory glands. Methods Twenty-six cases of head and neck BSCC from Asan Medical Center, Seoul, Korea, reported during a 14-year-period were subclassified into basal, ductal, and mixed subtypes according to the expression of basal (cytokeratin [CK] 5/6, p63 or ductal markers (CK7, CK8/18. The cases were also subject to immunohistochemical study for CK19, p53, cyclin D1, epidermal growth factor receptor (EGFR, and p16 and to in situ hybridization for human papillomavirus (HPV, and the results were clinico-pathologically compared. Results Mixed subtype (12 cases was the most common, and these cases showed hypopharyngeal predilection, older age, and higher expression of CK19, p53, and EGFR than other subtypes. The basal subtype (nine cases showed frequent comedo-necrosis and high expression of cyclin D1. The ductal subtype (five cases showed the lowest expression of p53, cyclin D1, and EGFR. A small number of p16- and/or HPV-positive cases were not restricted to one subtype. BSCC was the cause of death in 19 patients, and the average follow-up period for all patients was 79.5 months. Overall survival among the three subtypes was not significantly different. Conclusions The results of this study suggest a heterogeneous pathogenesis of head and neck BSCC. Each subtype showed variable histology and immunoprofiles, although the clinical implication of heterogeneity was not determined in this study.

  12. Inhibitors of Cyclin-dependent Kinases

    Czech Academy of Sciences Publication Activity Database

    Kryštof, V.; Strnad, Miroslav

    2001-01-01

    Roč. 95, č. 5 (2001), s. 295-300 ISSN 0009-2770 R&D Projects: GA ČR GA301/02/0475; GA MŠk VS96154 Institutional research plan: CEZ:AV0Z5038910 Keywords : Cyclin-dependent Kinase Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 0.317, year: 2001

  13. PARK2 orchestrates cyclins to avoid cancer

    Czech Academy of Sciences Publication Activity Database

    Bartek, Jiří; Hodný, Zdeněk

    2014-01-01

    Roč. 46, č. 6 (2014), s. 527-528 ISSN 1061-4036 Institutional support: RVO:68378050 Keywords : PARK2 * G1/S-phase cyclin * cancer Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 29.352, year: 2014

  14. Identification of Cyclin A Binders with a Fluorescent Peptide Sensor.

    Science.gov (United States)

    Pazos, Elena; Mascareñas, José L; Vázquez, M Eugenio

    2016-01-01

    A peptide sensor that integrates the 4-dimethylaminophthalimide (4-DMAP) fluorophore in a short cyclin A binding sequence displays a large fluorescence emission increase upon interacting with the cyclin A Binding Groove (CBG). Competitive displacement assays of this probe allow the straightforward identification of peptides that interact with the CBG, which could potentially block the recognition of CDK/cyclin A kinase substrates.

  15. Lack of cyclin D1 overexpression in gastric carcinogenesis

    NARCIS (Netherlands)

    Blok, P.; Craanen, M. E.; van Diest, P. J.; Dekker, W.; Tytgat, G. N.

    2000-01-01

    Cyclin D1 overexpression was examined in early gastric carcinomas and precursor lesions with the following aims; (1) to assess the chronology of cyclin D1 overexpression in various stages of gastric carcinogenesis, (2) to correlate cyclin D1 overexpression with the Lauren type, the grade of

  16. Cdc2/cyclin B1 regulates centrosomal Nlp proteolysis and subcellular localization.

    Science.gov (United States)

    Zhao, Xuelian; Jin, Shunqian; Song, Yongmei; Zhan, Qimin

    2010-11-01

    The formation of proper mitotic spindles is required for appropriate chromosome segregation during cell division. Aberrant spindle formation often causes aneuploidy and results in tumorigenesis. However, the underlying mechanism of regulating spindle formation and chromosome separation remains to be further defined. Centrosomal Nlp (ninein-like protein) is a recently characterized BRCA1-regulated centrosomal protein and plays an important role in centrosome maturation and spindle formation. In this study, we show that Nlp can be phosphorylated by cell cycle protein kinase Cdc2/cyclin B1. The phosphorylation sites of Nlp are mapped at Ser185 and Ser589. Interestingly, the Cdc2/cyclin B1 phosphorylation site Ser185 of Nlp is required for its recognition by PLK1, which enable Nlp depart from centrosomes to allow the establishment of a mitotic scaffold at the onset of mitosis . PLK1 fails to dissociate the Nlp mutant lacking Ser185 from centrosome, suggesting that Cdc2/cyclin B1 might serve as a primary kinase of PLK1 in regulating Nlp subcellular localization. However, the phosphorylation at the site Ser589 by Cdc2/cyclin B1 plays an important role in Nlp protein stability probably due to its effect on protein degradation. Furthermore, we show that deregulated expression or subcellular localization of Nlp lead to multinuclei in cells, indicating that scheduled levels of Nlp and proper subcellular localization of Nlp are critical for successful completion of normal cell mitosis, These findings demonstrate that Cdc2/cyclin B1 is a key regulator in maintaining appropriate degradation and subcellular localization of Nlp, providing novel insights into understanding on the role of Cdc2/cyclin B1 in mitotic progression.

  17. Spatial Reorganization of the Endoplasmic Reticulum during Mitosis Relies on Mitotic Kinase Cyclin A in the Early Drosophila Embryo

    Science.gov (United States)

    Bergman, Zane J.; Mclaurin, Justin D.; Eritano, Anthony S.; Johnson, Brittany M.; Sims, Amanda Q.; Riggs, Blake

    2015-01-01

    Mitotic cyclin-dependent kinase with their cyclin partners (cyclin:Cdks) are the master regulators of cell cycle progression responsible for regulating a host of activities during mitosis. Nuclear mitotic events, including chromosome condensation and segregation have been directly linked to Cdk activity. However, the regulation and timing of cytoplasmic mitotic events by cyclin:Cdks is poorly understood. In order to examine these mitotic cytoplasmic events, we looked at the dramatic changes in the endoplasmic reticulum (ER) during mitosis in the early Drosophila embryo. The dynamic changes of the ER can be arrested in an interphase state by inhibition of either DNA or protein synthesis. Here we show that this block can be alleviated by micro-injection of Cyclin A (CycA) in which defined mitotic ER clusters gathered at the spindle poles. Conversely, micro-injection of Cyclin B (CycB) did not affect spatial reorganization of the ER, suggesting CycA possesses the ability to initiate mitotic ER events in the cytoplasm. Additionally, RNAi-mediated simultaneous inhibition of all 3 mitotic cyclins (A, B and B3) blocked spatial reorganization of the ER. Our results suggest that mitotic ER reorganization events rely on CycA and that control and timing of nuclear and cytoplasmic events during mitosis may be defined by release of CycA from the nucleus as a consequence of breakdown of the nuclear envelope. PMID:25689737

  18. Differential expression of cyclin D1, Ki‑67, pRb, and p53 in psoriatic skin lesions and normal skin.

    Science.gov (United States)

    Kim, Sung Ae; Ryu, Young Wook; Kwon, Jun Il; Choe, Mi Sun; Jung, Jin Woong; Cho, Jae We

    2018-01-01

    Psoriasis is a hyperproliferative inflammatory skin disease; therefore, it is highly likely that psoriatic skin lesions may transform into malignancies. However, malignant transformation is not common. We performed immunohistochemical studies using anti‑cyclin D1, anti‑cyclin E, anti‑pRb, anti‑p53, anti‑p16INK4a, and anti‑Ki‑67 antibodies in normal skin, psoriatic epidermal tissue, and squamous cell carcinoma (SCC) tissue. Furthermore, western blot analysis and immunohistochemical staining were performed to ascertain differences in cyclin D1, cyclin E, pRb, and Ki‑67 expression before and after treatment for psoriasis. Cyclin D1 expression was higher in chronic psoriatic lesions than that in normal epidermis. Psoriasis lesions showed a strong intensity of positive nuclear staining for cyclin D1 among several normally stained nuclei in the basal layer. Cyclin E expression in psoriasis was stronger in the granular and spinous layer than in the normal epidermis. Expression levels of pRb and p53 were found to be higher in the psoriasis group compared with the normal epidermis. Total basal layer cell counts for p53WT expression were found to be significantly higher in the psoriasis group compared with the normal group. However, p16 expression was very weak in the normal and psoriasis groups compared with that in the SCC group. Ki‑67 immunoreactivity was significantly higher in psoriasis compared with normal epidermis and was similar with that in the SCC group. According to immunohistochemistry and immunoblot analysis, the expression levels of cyclin D1, cyclin E, pRb, and Ki‑67 in psoriasis lesions decreased after treatment and were similar with those in the normal group. Thus, increased expression of cyclin D1 and cyclin E may be involved in cell cycle progression in psoriatic epidermis, and pRb and p53 may play important roles in the prevention of malignant transformation under the hyperproliferative state in psoriasis.

  19. Alternative splicing variants of human Fbx4 disturb cyclin D1 proteolysis in human cancer

    Energy Technology Data Exchange (ETDEWEB)

    Chu, Xiufeng; Zhang, Ting; Wang, Jie; Li, Meng; Zhang, Xiaolei; Tu, Jing [Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing 100191 (China); Sun, Shiqin [College of Pharmacy, Harbin Medical University-Daqing, Daqing, Heilongjiang 163319 (China); Chen, Xiangmei, E-mail: xm_chen6176@bjmu.edu.cn [Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing 100191 (China); Lu, Fengmin [Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing 100191 (China)

    2014-04-25

    Highlights: • The expression of Fbx4 was significantly lower in HCC tissues. • Novel splicing variants of Fbx4 were identified. • These novel variants are much more abundant in human cancer tissues and cells. • The novel Fbx4 isoforms could promote cell proliferation and migration in vitro. • These isoforms showed less capability for cyclin D1 binding and degradation. - Abstract: Fbx4 is a specific substrate recognition component of SCF ubiquitin ligases that catalyzes the ubiquitination and subsequent degradation of cyclin D1 and Trx1. Two isoforms of human Fbx4 protein, the full length Fbx4α and the C-terminal truncated Fbx4β have been identified, but their functions remain elusive. In this study, we demonstrated that the mRNA level of Fbx4 was significantly lower in hepatocellular carcinoma tissues than that in the corresponding non-tumor tissues. More importantly, we identified three novel splicing variants of Fbx4: Fbx4γ (missing 168–245nt of exon1), Fbx4δ (missing exon6) and a N-terminal reading frame shift variant (missing exon2). Using cloning sequencing and RT-PCR, we demonstrated these novel splice variants are much more abundant in human cancer tissues and cell lines than that in normal tissues. When expressed in Sk-Hep1 and NIH3T3 cell lines, Fbx4β, Fbx4γ and Fbx4δ could promote cell proliferation and migration in vitro. Concordantly, these isoforms could disrupt cyclin D1 degradation and therefore increase cyclin D1 expression. Moreover, unlike the full-length isoform Fbx4α that mainly exists in cytoplasm, Fbx4β, Fbx4γ, and Fbx4δ locate in both cytoplasm and nucleus. Since cyclin D1 degradation takes place in cytoplasm, the nuclear distribution of these Fbx4 isoforms may not be involved in the down-regulation of cytoplasmic cyclin D1. These results define the impact of alternative splicing on Fbx4 function, and suggest that the attenuated cyclin D1 degradation by these novel Fbx4 isoforms provides a new insight for aberrant

  20. Alternative splicing variants of human Fbx4 disturb cyclin D1 proteolysis in human cancer

    International Nuclear Information System (INIS)

    Chu, Xiufeng; Zhang, Ting; Wang, Jie; Li, Meng; Zhang, Xiaolei; Tu, Jing; Sun, Shiqin; Chen, Xiangmei; Lu, Fengmin

    2014-01-01

    Highlights: • The expression of Fbx4 was significantly lower in HCC tissues. • Novel splicing variants of Fbx4 were identified. • These novel variants are much more abundant in human cancer tissues and cells. • The novel Fbx4 isoforms could promote cell proliferation and migration in vitro. • These isoforms showed less capability for cyclin D1 binding and degradation. - Abstract: Fbx4 is a specific substrate recognition component of SCF ubiquitin ligases that catalyzes the ubiquitination and subsequent degradation of cyclin D1 and Trx1. Two isoforms of human Fbx4 protein, the full length Fbx4α and the C-terminal truncated Fbx4β have been identified, but their functions remain elusive. In this study, we demonstrated that the mRNA level of Fbx4 was significantly lower in hepatocellular carcinoma tissues than that in the corresponding non-tumor tissues. More importantly, we identified three novel splicing variants of Fbx4: Fbx4γ (missing 168–245nt of exon1), Fbx4δ (missing exon6) and a N-terminal reading frame shift variant (missing exon2). Using cloning sequencing and RT-PCR, we demonstrated these novel splice variants are much more abundant in human cancer tissues and cell lines than that in normal tissues. When expressed in Sk-Hep1 and NIH3T3 cell lines, Fbx4β, Fbx4γ and Fbx4δ could promote cell proliferation and migration in vitro. Concordantly, these isoforms could disrupt cyclin D1 degradation and therefore increase cyclin D1 expression. Moreover, unlike the full-length isoform Fbx4α that mainly exists in cytoplasm, Fbx4β, Fbx4γ, and Fbx4δ locate in both cytoplasm and nucleus. Since cyclin D1 degradation takes place in cytoplasm, the nuclear distribution of these Fbx4 isoforms may not be involved in the down-regulation of cytoplasmic cyclin D1. These results define the impact of alternative splicing on Fbx4 function, and suggest that the attenuated cyclin D1 degradation by these novel Fbx4 isoforms provides a new insight for aberrant

  1. Requirements of cyclin a for mitosis are independent of its subcellular localization.

    Science.gov (United States)

    Dienemann, Axel; Sprenger, Frank

    2004-06-22

    Cyclin A (CycA), the only essential mitotic cyclin in Drosophila, is cytoplasmic during interphase and accumulates in the nucleus during prophase. We show that interphase localization is mediated by Leptomycin B (LMB)-sensitive nuclear export. This is a feature shared with human CyclinB1, and it is assumed that nuclear accumulation is necessary for mitotic entry. Here, we tested if the unique mitotic function of CycA requires nuclear accumulation. We fused subcellular localization signals to CycA and tested their mitotic capability. Surprisingly, nuclear accumulation was not required, and even a membrane-tethered form of CycA was able to induce mitosis. We noted that Cyclin B (CycB) protein disappears prematurely in CycA mutants, reminiscent of rca1 mutants. Rca1 is an inhibitor of Fizzy-related-APC/C activity, and in rca1 mutants, mitotic cyclins are degraded in G2 of the 16(th) embryonic cell cycle. Overexpression of Rca1 can restore mitosis in CycA mutants, indicating that the mitotic failure of CycA mutants is caused by premature activation of the APC/C. The essential mitotic function of CycA is therefore not the activation of numerous mitotic substrates by Cdk1-dependent phosphorylation. Rather, CycA-dependent kinase activity is required to inhibit one inhibitor of mitosis, the Fzr protein.

  2. Virotherapy targeting cyclin E overexpression in tumors with adenovirus-enhanced cancer-selective promoter.

    Science.gov (United States)

    Cheng, Pei-Hsin; Rao, Xiao-Mei; Duan, Xiaoxian; Li, Xiao-Feng; Egger, Michael E; McMasters, Kelly M; Zhou, H Sam

    2015-02-01

    Oncolytic virotherapy can selectively destroy cancer cells and is a potential approach in cancer treatment. A strategy to increase tumor-specific selectivity is to control the expression of a key regulatory viral gene with a tumor-specific promoter. We have previously found that cyclin E expression is augmented in cancer cells after adenovirus (Ad) infection. Thus, the cyclin E promoter that is further activated by Ad in cancer cells may have unique properties for enhancing oncolytic viral replication. We have shown that high levels of viral E1a gene expression are achieved in cancer cells infected with Ad-cycE, in which the endogenous Ad E1a promoter was replaced with the cyclin E promoter. Ad-cycE shows markedly selective oncolytic efficacy in vitro and destroys various types of cancer cells, including those resistant to ONYX-015/dl1520. Furthermore, Ad-cycE shows a strong capacity to repress A549 xenograft tumor growth in nude mice and significantly prolongs survival. This study suggests the potential of Ad-cycE in cancer therapy and indicates the advantages of using promoters that can be upregulated by virus infection in cancer cells in development of oncolytic viruses. Key messages: Cyclin E promoter activity is high in cancer cells and enhanced by adenovirus infection. Cyclin E promoter is used to control the E1a gene of a tumor-specific oncolytic adenovirus. Ad-cycE efficiently targets cancer cells and induces oncolysis. Ad-cycE significantly repressed xenograft tumor and prolonged survival.

  3. Investigating the Role of Cyclin D1 in the Promotion of Genomic Instability and Breast Cancer

    Science.gov (United States)

    2011-09-01

    with purified cyclin D1T286A/CDK4 and subjected it to mass spectometry ; we also affinity purified MEP50 from cells coexpressing cyclin D1T286A/ CDK4 for...Philadelphia, PA 19104 REPORT DATE : September 2011 TYPE OF REPORT: Annual Summary PREPARED FOR: U.S. Army Medical Research and Materiel Command...OMB control number. PLEASE DO NOT RETURN YOUR FORM TO THE ABOVE ADDRESS. 1. REPORT DATE September 2011 2. REPORT TYPE Annual Summary 3. DATES

  4. Modulation of human melanoma cell proliferation and apoptosis by hydatid cyst fluid of Echinococcus granulosus

    Directory of Open Access Journals (Sweden)

    Gao X

    2018-03-01

    Full Text Available Xiang-Yang Gao,1,* Guang-Hui Zhang,2,* Li Huang3 1Department of Laboratory Medicine, Pu’er People’s Hospital, Pu’er, 2Department of Clinical Laboratory, Xin Hua Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai, 3Department of General Surgery, Shanghai General Hospital, Shanghai, China *These authors contributed equally to this work Objective: The objective of this paper was to assess the effects of hydatid cyst fluid (HCF of Echinococcus granulosus on melanoma A375 cell proliferation and apoptosis.Methods: A375 cells were classified into five groups by in vitro culture: normal group, control group, 10% HCF group, 20% HCF group and 30% HCF group. Trypan blue staining method was employed to detect the toxicity of HCF. Effects of different concentrations of HCF on melanoma A375 cell proliferation at different time points were evaluated using the 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide assay. Flow cytometry and propidium iodide (PI staining were used to detect cell cycle, and Annexin-V/PI double staining method was used to determine A375 cell apoptotic rate. Western blotting was applied to detect the expression of phosphorylated extracellular regulated protein kinases, proliferating cell nuclear antigen (PCNA, cell-cycle-related proteins (cyclin A, cyclin B1, cyclin D1 and cyclin E and apoptosis-related proteins (Bcl-2, Bax and caspase-3.Results: HCF with a high concentration was considered as atoxic to A375 cells. HCF promoted A375 cell proliferation, and the effects got stronger with an increase in concentrations but was retarded after reaching a certain range of concentrations. HCF increased phosphorylation level and expression of extracellular regulated protein kinase, as well as PCNA expression. HCF also promoted the transferring progression of A375 cells from the G0/G1 phase to the S phase to increase the cell number in S phase and increased the expression of cyclin A, cyclin D1 and

  5. Antisense inhibition of cyclin D1 expression is equivalent to flavopiridol for radiosensitization of zebrafish embryos

    International Nuclear Information System (INIS)

    Purpose: Flavopiridol, a small molecule pan-cyclin inhibitor, has been shown to enhance Radiation response of tumor cells both in vitro and in vivo. The clinical utility of flavopiridol, however, is limited by toxicity, previously attributed to pleiotropic inhibitory effects on several targets affecting multiple signal transduction pathways. Here we used zebrafish embryos to investigate radiosensitizing effects of flavopiridol in normal tissues. Methods and Materials: Zebrafish embryos at the 1- to 4-cell stage were treated with 500 nM flavopiridol or injected with 0.5 pmol antisense hydroxylprolyl-phosphono nucleic acid oligomers to reduce cyclin D1 expression, then subjected to ionizing radiation (IR) or no radiation. Results: Flavopiridol-treated embryos demonstrated a twofold increase in mortality after exposure to 40 Gy by 96 hpf and developed distinct radiation-induced defects in midline development (designated as the 'curly up' phenotype) at higher rates when compared with embryos receiving IR only. Cyclin D1-deficient embryos had virtually identical IR sensitivity profiles when compared with embryos treated with flavopiridol. This was particularly evident for the IR-induced curly up phenotype, which was greatly exacerbated by both flavopriridol and cyclin D1 downregulation. Conclusions: Treatment of zebrafish embryos with flavopiridol enhanced radiation sensitivity of zebrafish embryos to a degree that was very similar to that associated with downregulation of cyclin D1 expression. These results are consistent with the hypothesis that inhibition of cyclin D1 is sufficient to account for the radiosensitizing action of flavopiridol in the zebrafish embryo vertebrate model

  6. Structural basis of divergent cyclin-dependent kinase activation by Spy1/RINGO proteins

    Energy Technology Data Exchange (ETDEWEB)

    McGrath, Denise A.; Fifield, Bre-Anne; Marceau, Aimee H.; Tripathi, Sarvind; Porter, Lisa A.; Rubin, Seth M. (UCSC); (Windsor)

    2017-06-30

    Cyclin-dependent kinases (Cdks) are principal drivers of cell division and are an important therapeutic target to inhibit aberrant proliferation. Cdk enzymatic activity is tightly controlled through cyclin interactions, posttranslational modifications, and binding of inhibitors such as the p27 tumor suppressor protein. Spy1/RINGO (Spy1) proteins bind and activate Cdk but are resistant to canonical regulatory mechanisms that establish cell-cycle checkpoints. Cancer cells exploit Spy1 to stimulate proliferation through inappropriate activation of Cdks, yet the mechanism is unknown. We have determined crystal structures of the Cdk2-Spy1 and p27-Cdk2-Spy1 complexes that reveal how Spy1 activates Cdk. We find that Spy1 confers structural changes to Cdk2 that obviate the requirement of Cdk activation loop phosphorylation. Spy1 lacks the cyclin-binding site that mediates p27 and substrate affinity, explaining why Cdk-Spy1 is poorly inhibited by p27 and lacks specificity for substrates with cyclin-docking sites. We identify mutations in Spy1 that ablate its ability to activate Cdk2 and to proliferate cells. Our structural description of Spy1 provides important mechanistic insights that may be utilized for targeting upregulated Spy1 in cancer.

  7. Characterization of the mouse cyclin D3 gene: Exon/intron organization and promoter activity

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Zhengyu; Zhang, Ying; Ravid, K. [Boston Univ. School of Medicine, Boston, MA (United States)] [and others

    1996-07-01

    The three D-type cyclins have been shown to be differentially expressed in a number of cell types, suggesting that they play distinct roles in cell cycle regulation in particular cell lineages. We have determined the complete nucleotide sequence (-1681 to +6582) of the mouse cyclin D3 gene, which encodes a G1 phase cyclin. The gene consists of five exons and four introns, varying in length from 422 to 2472 bp. Primer extension analysis revealed one major transcription initiation site at the position 107 bp 5{prime} upstream of the translation start. The promoter region lacks both canonical {open_quotes}TATA{close_quotes} and {open_quotes}CAAT{close_quotes} boxes. It contains, however, multiple transcription factor recognition by GATA, NF-{kappa}B, ATF, E2F, and TRE/AP1 transcription factors, E box binding myogenic factors, and the IL-6 induced-transcription factor, APRF. Promoter activity of the 1681-bp fragment upstream of the transcription initiation site was confirmed by linking it to a reporter gene and subjecting it to transient expression experiments in various cell types. Promoter activity was high in cell lines that expressed high levels of endogenous D3 mRNA, as indicated by Northern blot analysis, and was significantly reduced when the promoter was truncated to -122 bp. The characterization of the mouse cyclin D3 gene and insight into its promoter region will allow further studies defining the molecular events regulating the expression of this cyclin in proliferating and quiescent cells. 60 refs., 4 figs., 1 tab.

  8. Primary hyperparathyroidism caused by parathyroid-targeted overexpression of cyclin D1 in transgenic mice

    OpenAIRE

    Imanishi, Yasuo; Hosokawa, Yoshitaka; Yoshimoto, Katsuhiko; Schipani, Ernestina; Mallya, Sanjay; Papanikolaou, Alexandros; Kifor, Olga; Tokura, Takehiko; Sablosky, Marilyn; Ledgard, Felicia; Gronowicz, Gloria; Wang, Timothy C.; Schmidt, Emmett V.; Hall, Charles; Brown, Edward M.

    2001-01-01

    The relationship between abnormal cell proliferation and aberrant control of hormonal secretion is a fundamental and poorly understood issue in endocrine cell neoplasia. Transgenic mice with parathyroid-targeted overexpression of the cyclin D1 oncogene, modeling a gene rearrangement found in human tumors, were created to determine whether a primary defect in this cell-cycle regulator can cause an abnormal relationship between serum calcium and parathyroid hormone response, as is typical of hu...

  9. Epigenetically altered miR-193b targets cyclin D1 in prostate cancer

    International Nuclear Information System (INIS)

    Kaukoniemi, Kirsi M; Rauhala, Hanna E; Scaravilli, Mauro; Latonen, Leena; Annala, Matti; Vessella, Robert L; Nykter, Matti; Tammela, Teuvo L J; Visakorpi, Tapio

    2015-01-01

    Micro-RNAs (miRNA) are important regulators of gene expression and often differentially expressed in cancer and other diseases. We have previously shown that miR-193b is hypermethylated in prostate cancer (PC) and suppresses cell growth. It has been suggested that miR-193b targets cyclin D1 in several malignancies. Here, our aim was to determine if miR-193b targets cyclin D1 in prostate cancer. Our data show that miR-193b is commonly methylated in PC samples compared to benign prostate hyperplasia. We found reduced miR-193b expression (P < 0.05) in stage pT3 tumors compared to pT2 tumors in a cohort of prostatectomy specimens. In 22Rv1 PC cells with low endogenous miR-193b expression, the overexpression of miR-193b reduced CCND1mRNA levels and cyclin D1 protein levels. In addition, the exogenous expression of miR-193b decreased the phosphorylation level of RB, a target of the cyclin D1-CDK4/6 pathway. Moreover, according to a reporter assay, miR-193b targeted the 3’UTR of CCND1 in PC cells and the CCND1 activity was rescued by expressing CCND1 lacking its 3’UTR. Immunohistochemical analysis of cyclin D1 showed that castration-resistant prostate cancers have significantly (P = 0.0237) higher expression of cyclin D1 compared to hormone-naïve cases. Furthermore, the PC cell lines 22Rv1 and VCaP, which express low levels of miR-193b and high levels of CCND1, showed significant growth retardation when treated with a CDK4/6 inhibitor. In contrast, the inhibitor had no effect on the growth of PC-3 and DU145 cells with high miR-193b and low CCND1 expression. Taken together, our data demonstrate that miR-193b targets cyclin D1 in prostate cancer

  10. C/EBP{delta} targets cyclin D1 for proteasome-mediated degradation via induction of CDC27/APC3 expression.

    Science.gov (United States)

    Pawar, Snehalata A; Sarkar, Tapasree Roy; Balamurugan, Kuppusamy; Sharan, Shikha; Wang, Jun; Zhang, Youhong; Dowdy, Steven F; Huang, A-Mei; Sterneck, Esta

    2010-05-18

    The transcription factor CCAAT/enhancer binding protein delta (C/EBPdelta, CEBPD, NFIL-6beta) has tumor suppressor function; however, the molecular mechanism(s) by which C/EBPdelta exerts its effect are largely unknown. Here, we report that C/EBPdelta induces expression of the Cdc27 (APC3) subunit of the anaphase promoting complex/cyclosome (APC/C), which results in the polyubiquitination and degradation of the prooncogenic cell cycle regulator cyclin D1, and also down-regulates cyclin B1, Skp2, and Plk-1. In C/EBPdelta knockout mouse embryo fibroblasts (MEF) Cdc27 levels were reduced, whereas cyclin D1 levels were increased even in the presence of activated GSK-3beta. Silencing of C/EBPdelta, Cdc27, or the APC/C coactivator Cdh1 (FZR1) in MCF-10A breast epithelial cells increased cyclin D1 protein expression. Like C/EBPdelta, and in contrast to cyclin D1, Cdc27 was down-regulated in several breast cancer cell lines, suggesting that Cdc27 itself may be a tumor suppressor. Cyclin D1 is a known substrate of polyubiquitination complex SKP1/CUL1/F-box (SCF), and our studies show that Cdc27 directs cyclin D1 to alternative degradation by APC/C. These findings shed light on the role and regulation of APC/C, which is critical for most cellular processes.

  11. Automated image analysis of cyclin D1 protein expression in invasive lobular breast carcinoma provides independent prognostic information.

    Science.gov (United States)

    Tobin, Nicholas P; Lundgren, Katja L; Conway, Catherine; Anagnostaki, Lola; Costello, Sean; Landberg, Göran

    2012-11-01

    The emergence of automated image analysis algorithms has aided the enumeration, quantification, and immunohistochemical analyses of tumor cells in both whole section and tissue microarray samples. To date, the focus of such algorithms in the breast cancer setting has been on traditional markers in the common invasive ductal carcinoma subtype. Here, we aimed to optimize and validate an automated analysis of the cell cycle regulator cyclin D1 in a large collection of invasive lobular carcinoma and relate its expression to clinicopathologic data. The image analysis algorithm was trained to optimally match manual scoring of cyclin D1 protein expression in a subset of invasive lobular carcinoma tissue microarray cores. The algorithm was capable of distinguishing cyclin D1-positive cells and illustrated high correlation with traditional manual scoring (κ=0.63). It was then applied to our entire cohort of 483 patients, with subsequent statistical comparisons to clinical data. We found no correlation between cyclin D1 expression and tumor size, grade, and lymph node status. However, overexpression of the protein was associated with reduced recurrence-free survival (P=.029), as was positive nodal status (Pinvasive lobular carcinoma. Finally, high cyclin D1 expression was associated with increased hazard ratio in multivariate analysis (hazard ratio, 1.75; 95% confidence interval, 1.05-2.89). In conclusion, we describe an image analysis algorithm capable of reliably analyzing cyclin D1 staining in invasive lobular carcinoma and have linked overexpression of the protein to increased recurrence risk. Our findings support the use of cyclin D1 as a clinically informative biomarker for invasive lobular breast cancer. Copyright © 2012 Elsevier Inc. All rights reserved.

  12. Cyclin A1 modulates the expression of vascular endothelial growth factor and promotes hormone-dependent growth and angiogenesis of breast cancer.

    Directory of Open Access Journals (Sweden)

    Azharuddin Sajid Syed Khaja

    Full Text Available Alterations in cellular pathways related to both endocrine and vascular endothelial growth factors (VEGF may contribute to breast cancer progression. Inhibition of the elevated levels of these pathways is associated with clinical benefits. However, molecular mechanisms by which endocrine-related pathways and VEGF signalling cooperatively promote breast cancer progression remain poorly understood. In the present study, we show that the A-type cyclin, cyclin A1, known for its important role in the initiation of leukemia and prostate cancer metastasis, is highly expressed in primary breast cancer specimens and metastatic lesions, in contrasting to its barely detectable expression in normal human breast tissues. There is a statistically significant correlation between cyclin A1 and VEGF expression in breast cancer specimens from two patient cohorts (p<0.01. Induction of cyclin A1 overexpression in breast cancer cell line MCF-7 results in an enhanced invasiveness and a concomitant increase in VEGF expression. In addition, there is a formation of protein-protein complexes between cyclin A1 and estrogen receptor ER-α cyclin A1 overexpression increases ER-α expression in MCF-7 and T47D cells. In mouse tumor xenograft models in which mice were implanted with MCF-7 cells that overexpressed cyclin A1 or control vector, cyclin A1 overexpression results in an increase in tumor growth and angiogenesis, which is coincident with an enhanced expression of VEGF, VEGFR1 and ER-α Our findings unravel a novel role for cyclin A1 in growth and progression of breast cancer, and suggest that multiple cellular pathways, including cell cycle regulators, angiogenesis and estrogen receptor signalling, may cooperatively contribute to breast cancer progression.

  13. Cyclin B1 Destruction Box-Mediated Protein Instability: The Enhanced Sensitivity of Fluorescent-Protein-Based Reporter Gene System

    Directory of Open Access Journals (Sweden)

    Chao-Hsun Yang

    2013-01-01

    Full Text Available The periodic expression and destruction of several cyclins are the most important steps for the exact regulation of cell cycle. Cyclins are degraded by the ubiquitin-proteasome system during cell cycle. Besides, a short sequence near the N-terminal of cyclin B called the destruction box (D-box; CDB is also required. Fluorescent-protein-based reporter gene system is insensitive to analysis because of the overly stable fluorescent proteins. Therefore, in this study, we use human CDB fused with both enhanced green fluorescent protein (EGFP at C-terminus and red fluorescent protein (RFP, DsRed at N-terminus in the transfected human melanoma cells to examine the effects of CDB on different fluorescent proteins. Our results indicated that CDB-fused fluorescent protein can be used to examine the slight gene regulations in the reporter gene system and have the potential to be the system for screening of functional compounds in the future.

  14. NF-κB-dependent transcriptional upregulation of cyclin D1 exerts cytoprotection against hypoxic injury upon EGFR activation

    International Nuclear Information System (INIS)

    Chen, Zhi-Dong; Xu, Liang; Tang, Kan-Kai; Gong, Fang-Xiao; Liu, Jing-Quan; Ni, Yin; Jiang, Ling-Zhi; Hong, Jun; Han, Fang; Li, Qian; Yang, Xiang-Hong; Sun, Ren-Hua; Mo, Shi-Jing

    2016-01-01

    Apoptosis of neural cells is one of the main pathological features in hypoxic/ischemic brain injury. Nuclear factor-κB (NF-κB) might be a potential therapeutic target for hypoxic/ischemic brain injury since NF-κB has been found to be inactivated after hypoxia exposure, yet the underlying molecular mechanisms of NF-κB inactivation are largely unknown. Here we report that epidermal growth factor receptor (EGFR) activation prevents neuron-like PC12 cells apoptosis in response to hypoxia via restoring NF-κB-dependent transcriptional upregulation of cyclin D1. Functionally, EGFR activation by EGF stimulation mitigates hypoxia-induced PC12 cells apoptosis in both dose- and time-dependent manner. Of note, EGFR activation elevates IKKβ phosphorylation, increases IκBα ubiquitination, promotes P65 nuclear translocation and recruitment at cyclin D1 gene promoter as well as upregulates cyclin D1 expression. EGFR activation also abrogates the decrease of IKKβ phosphorylation, reduction of IκBα ubiquitination, blockade of P65 nuclear translocation and recruitment at cyclin D1 gene promoter as well as downregulation of cyclin D1 expression induced by hypoxia. Furthermore, NF-κB-dependent upregulation of cyclin D1 is instrumental for the EGFR-mediated cytoprotection against hypoxic apoptosis. In addition, the dephosphorylation of EGFR induced by either EGF siRNA transfection or anti-HB-EGF neutralization antibody treatment enhances hypoxic cytotoxicity, which are attenuated by EGF administration. Our results highlight the essential role of NF-κB-dependent transcriptional upregulation of cyclin D1 in EGFR-mediated cytoprotective effects under hypoxic preconditioning and support further investigation of EGF in clinical trials of patients with hypoxic/ischemic brain injury. - Highlights: • EGFR activation significantly decreases hypoxia-induced PC12 cells injury. • EGFR activation abrogates the transcriptional repression of cyclin D1 induced by hypoxia in a NF

  15. NF-κB-dependent transcriptional upregulation of cyclin D1 exerts cytoprotection against hypoxic injury upon EGFR activation

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Zhi-Dong [Department of Critical Care Medicine, The First Affiliated Hospital of Huzhou Normal College, Huzhou 313000, Zhejiang (China); Xu, Liang [Department of Critical Care Medicine, Zhejiang Provincial People’s Hospital, Hangzhou 310000, Zhejiang (China); Tang, Kan-Kai [Department of Critical Care Medicine, The First Affiliated Hospital of Huzhou Normal College, Huzhou 313000, Zhejiang (China); Gong, Fang-Xiao; Liu, Jing-Quan; Ni, Yin; Jiang, Ling-Zhi; Hong, Jun; Han, Fang; Li, Qian; Yang, Xiang-Hong [Department of Critical Care Medicine, Zhejiang Provincial People’s Hospital, Hangzhou 310000, Zhejiang (China); Sun, Ren-Hua, E-mail: jqin168@hotmail.com [Department of Critical Care Medicine, Zhejiang Provincial People’s Hospital, Hangzhou 310000, Zhejiang (China); Mo, Shi-Jing, E-mail: msj860307@163.com [Department of Critical Care Medicine, Zhejiang Provincial People’s Hospital, Hangzhou 310000, Zhejiang (China)

    2016-09-10

    Apoptosis of neural cells is one of the main pathological features in hypoxic/ischemic brain injury. Nuclear factor-κB (NF-κB) might be a potential therapeutic target for hypoxic/ischemic brain injury since NF-κB has been found to be inactivated after hypoxia exposure, yet the underlying molecular mechanisms of NF-κB inactivation are largely unknown. Here we report that epidermal growth factor receptor (EGFR) activation prevents neuron-like PC12 cells apoptosis in response to hypoxia via restoring NF-κB-dependent transcriptional upregulation of cyclin D1. Functionally, EGFR activation by EGF stimulation mitigates hypoxia-induced PC12 cells apoptosis in both dose- and time-dependent manner. Of note, EGFR activation elevates IKKβ phosphorylation, increases IκBα ubiquitination, promotes P65 nuclear translocation and recruitment at cyclin D1 gene promoter as well as upregulates cyclin D1 expression. EGFR activation also abrogates the decrease of IKKβ phosphorylation, reduction of IκBα ubiquitination, blockade of P65 nuclear translocation and recruitment at cyclin D1 gene promoter as well as downregulation of cyclin D1 expression induced by hypoxia. Furthermore, NF-κB-dependent upregulation of cyclin D1 is instrumental for the EGFR-mediated cytoprotection against hypoxic apoptosis. In addition, the dephosphorylation of EGFR induced by either EGF siRNA transfection or anti-HB-EGF neutralization antibody treatment enhances hypoxic cytotoxicity, which are attenuated by EGF administration. Our results highlight the essential role of NF-κB-dependent transcriptional upregulation of cyclin D1 in EGFR-mediated cytoprotective effects under hypoxic preconditioning and support further investigation of EGF in clinical trials of patients with hypoxic/ischemic brain injury. - Highlights: • EGFR activation significantly decreases hypoxia-induced PC12 cells injury. • EGFR activation abrogates the transcriptional repression of cyclin D1 induced by hypoxia in a NF

  16. The tumor suppressor, parafibromin, mediates histone H3 K9 methylation for cyclin D1 repression.

    Science.gov (United States)

    Yang, Yong-Jin; Han, Jeung-Whan; Youn, Hong-Duk; Cho, Eun-Jung

    2010-01-01

    Parafibromin, a component of the RNA polymerase II-associated PAF1 complex, is a tumor suppressor linked to hyperparathyroidism-jaw tumor syndrome and sporadic parathyroid carcinoma. Parafibromin induces cell cycle arrest by repressing cyclin D1 via an unknown mechanism. Here, we show that parafibromin interacts with the histone methyltransferase, SUV39H1, and functions as a transcriptional repressor. The central region (128-227 amino acids) of parafibromin is important for both the interaction with SUV39H1 and transcriptional repression. Parafibromin associated with the promoter and coding regions of cyclin D1 and was required for the recruitment of SUV39H1 and the induction of H3 K9 methylation but not H3 K4 methylation. RNA interference analysis showed that SUV39H1 was critical for cyclin D1 repression. These data suggest that parafibromin plays an unexpected role as a repressor in addition to its widely known activity associated with transcriptional activation. Parafibromin as a part of the PAF1 complex might downregulate cyclin D1 expression by integrating repressive H3 K9 methylation during transcription.

  17. Virotherapy Targeting Cyclin E Overexpression in Tumors with Adenovirus-enhanced Cancer Selective Promoter

    Science.gov (United States)

    Cheng, Pei-Hsin; Rao, Xiao-Mei; Duan, Xiaoxian; Li, Xiao-Feng; Egger, Michael E.; McMasters, Kelly M.; Zhou, H. Sam

    2014-01-01

    Oncolytic virotherapy can selectively destroy cancer cells and is a potential approach in cancer treatment. A strategy to increase tumor-specific selectivity is to control the expression of a key regulatory viral gene with a tumor-specific promoter. We have previously found that cyclin E expression is augmented in cancer cells after adenovirus (Ad) infection. Thus, the cyclin E promoter that is further activated by Ad in cancer cells may have unique properties for enhancing oncolytic viral replication. We have shown that high levels of viral E1a gene expression are achieved in cancer cells infected with Ad-cycE, in which the endogenous Ad E1a promoter was replaced with the cyclin E promoter. Ad-cycE shows markedly selective oncolytic efficacy in vitro and destroys various types of cancer cells, including those resistant to ONYX-015/dl1520. Furthermore, Ad-cycE shows a strong capacity to repress A549 xenograft tumor growth in nude mice and significantly prolongs survival. This study suggests the potential of Ad-cycE in cancer therapy and indicates the advantages of using promoters that can be upregulated by virus infection in cancer cells in development of oncolytic viruses. PMID:25376708

  18. MEK inhibition potentiates the activity of Hsp90 inhibitor 17-AAG against pancreatic cancer cells.

    Science.gov (United States)

    Zhang, Tao; Li, Yanyan; Zhu, Zhenkun; Gu, Mancang; Newman, Bryan; Sun, Duxin

    2010-10-04

    The Ras/Raf/MEK/ERK signaling has been implicated in uncontrolled cell proliferation and tumor progression in pancreatic cancer. The purpose of this study is to evaluate the antitumor activity of MEK inhibitor U0126 in combination with Hsp90 inhibitor 17-allylamino-17-demethoxygeldanamycin (17-AAG) in pancreatic cancer cells. Western blotting showed that 17-AAG caused a 2- to 3-fold transient activation of MEK/ERK signaling in pancreatic cancer cells. The activation sustained for 6 h before phospho-ERK (p-ERK) destabilization. The selective MEK inhibitor U0126 completely abolished 17-AAG induced ERK1/2 activation and resulted in more than 80% of phospho-ERK degradation after only 15 min treatment. Moreover, U0126 had complementary effect on 17-AAG regulated oncogenic and cell cycle related proteins. Although 17-AAG downregulated cyclin D1, cyclin E, CDK4 and CDK6, it led to cyclin A and CDK2 accumulation, which was reversed by the addition of U0126. Antiproliferation assay showed that combination of U0126 and 17-AAG resulted in synergistic cytotoxic effect. More importantly, 17-AAG alone only exhibited moderate inhibition of cell migration in vitro, while addition of U0126 dramatically enhanced the inhibitory effect by 2- to 5-fold. Taken together, these data demonstrate that MEK inhibitor U0126 potentiates the activity of Hsp90 inhibitor 17-AAG against pancreatic cancer cells. The combination of Hsp90 and MEK inhibition could provide a promising avenue for the treatment of pancreatic cancer.

  19. A mechanism for the inhibition of neural progenitor cell proliferation by cocaine.

    Directory of Open Access Journals (Sweden)

    Chun-Ting Lee

    2008-06-01

    Full Text Available BACKGROUND: Prenatal exposure of the developing brain to cocaine causes morphological and behavioral abnormalities. Recent studies indicate that cocaine-induced proliferation inhibition and/or apoptosis in neural progenitor cells may play a pivotal role in causing these abnormalities. To understand the molecular mechanism through which cocaine inhibits cell proliferation in neural progenitors, we sought to identify the molecules that are responsible for mediating the effect of cocaine on cell cycle regulation. METHODS AND FINDINGS: Microarray analysis followed by quantitative real-time reverse transcription PCR was used to screen cocaine-responsive and cell cycle-related genes in a neural progenitor cell line where cocaine exposure caused a robust anti-proliferative effect by interfering with the G1-to-S transition. Cyclin A2, among genes related to the G1-to-S cell cycle transition, was most strongly down-regulated by cocaine. Down-regulation of cyclin A was also found in cocaine-treated human primary neural and A2B5+ progenitor cells, as well as in rat fetal brains exposed to cocaine in utero. Reversing cyclin A down-regulation by gene transfer counteracted the proliferation inhibition caused by cocaine. Further, we found that cocaine-induced accumulation of reactive oxygen species, which involves N-oxidation of cocaine via cytochrome P450, promotes cyclin A down-regulation by causing an endoplasmic reticulum (ER stress response, as indicated by increased phosphorylation of eIF2alpha and expression of ATF4. In the developing rat brain, the P450 inhibitor cimetidine counteracted cocaine-induced inhibition of neural progenitor cell proliferation as well as down-regulation of cyclin A. CONCLUSIONS: Our results demonstrate that down-regulation of cyclin A underlies cocaine-induced proliferation inhibition in neural progenitors. The down-regulation of cyclin A is initiated by N-oxidative metabolism of cocaine and consequent ER stress. Inhibition of

  20. Reduced hepatic tumor incidence in cyclin G1-deficient mice

    DEFF Research Database (Denmark)

    Jensen, Michael Rugaard; Factor, Valentina M; Fantozzi, Anna

    2003-01-01

    found that the p53 levels in the cyclin G1-deficient mice are 2-fold higher that in wild-type mice. Moreover, we showed that treatment of mice with the alkylating agent 1,4-bis[N,N'-di(ethylene)-phosphamide]piperazine (Dipin), followed by partial hepatectomy, decreased G1-S transition in cyclin G1-null...

  1. Clinical significance of cyclin-dependent kinase inhibitor p27Kip1 expression and proliferation in non-Hodgkin's lymphoma

    DEFF Research Database (Denmark)

    Møller, Michael Boe; Skjødt, Karsten; Mortensen, Leif Spange

    1999-01-01

    The cyclin-dependent kinase inhibitor p27Kip1 is a negative cell cycle regulator linking extracellular growth-regulatory signals to the cell cycle machinery in G1. We investigated the pattern and prognostic value of p27Kip1 expression in a population-based group of 203 non-Hodgkin's lymphoma (NHL...

  2. Cyclin A2 and CDK2 as Novel Targets of Aspirin and Salicylic Acid: A Potential Role in Cancer Prevention.

    Science.gov (United States)

    Dachineni, Rakesh; Ai, Guoqiang; Kumar, D Ramesh; Sadhu, Satya S; Tummala, Hemachand; Bhat, G Jayarama

    2016-03-01

    Data emerging from the past 10 years have consolidated the rationale for investigating the use of aspirin as a chemopreventive agent; however, the mechanisms leading to its anticancer effects are still being elucidated. We hypothesized that aspirin's chemopreventive actions may involve cell-cycle regulation through modulation of the levels or activity of cyclin A2/cyclin-dependent kinase-2 (CDK2). In this study, HT-29 and other diverse panel of cancer cells were used to demonstrate that both aspirin and its primary metabolite, salicylic acid, decreased cyclin A2 (CCNA2) and CDK2 protein and mRNA levels. The downregulatory effect of either drugs on cyclin A2 levels was prevented by pretreatment with lactacystin, an inhibitor of proteasomes, suggesting the involvement of 26S proteasomes. In-vitro kinase assays showed that lysates from cells treated with salicylic acid had lower levels of CDK2 activity. Importantly, three independent experiments revealed that salicylic acid directly binds to CDK2. First, inclusion of salicylic acid in naïve cell lysates, or in recombinant CDK2 preparations, increased the ability of the anti-CDK2 antibody to immunoprecipitate CDK2, suggesting that salicylic acid may directly bind and alter its conformation. Second, in 8-anilino-1-naphthalene-sulfonate (ANS)-CDK2 fluorescence assays, preincubation of CDK2 with salicylic acid dose-dependently quenched the fluorescence due to ANS. Third, computational analysis using molecular docking studies identified Asp145 and Lys33 as the potential sites of salicylic acid interactions with CDK2. These results demonstrate that aspirin and salicylic acid downregulate cyclin A2/CDK2 proteins in multiple cancer cell lines, suggesting a novel target and mechanism of action in chemoprevention. Biochemical and structural studies indicate that the antiproliferative actions of aspirin are mediated through cyclin A2/CDK2. ©2015 American Association for Cancer Research.

  3. miR-379 regulates cyclin B1 expression and is decreased in breast cancer.

    Directory of Open Access Journals (Sweden)

    Sonja Khan

    Full Text Available MicroRNAs are small non-coding RNA molecules that control gene expression post-transcriptionally, and are known to be altered in many diseases including breast cancer. The aim of this study was to determine the relevance of miR-379 in breast cancer. miR-379 expression was quantified in clinical samples including tissues from breast cancer patients (n=103, healthy controls (n=30 and patients with benign breast disease (n=35. The level of miR-379 and its putative target Cyclin B1 were investigated on all breast tissue specimens by RQ-PCR. Potential relationships with gene expression and patient clinicopathological details were also determined. The effect of miR-379 on Cyclin B1 protein expression and function was investigated using western blot, immunohistochemistry and proliferation assays respectively. Finally, the levels of circulating miR-379 were determined in whole blood from patients with breast cancer (n=40 and healthy controls (n=34. The level of miR-379 expression was significantly decreased in breast cancer (Mean(SEM 1.9 (0.09 Log10 Relative Quantity (RQ compared to normal breast tissues (2.6 (0.16 Log10 RQ, p<0.01. miR-379 was also found to decrease significantly with increasing tumour stage. A significant negative correlation was determined between miR-379 and Cyclin B1 (r=-0.31, p<0.001. Functional assays revealed reduced proliferation (p<0.05 and decreased Cyclin B1 protein levels following transfection of breast cancer cells with miR-379. Circulating miR-379 was not significantly dysregulated in patients with breast cancer compared to healthy controls (p=0.42. This data presents miR-379 as a novel regulator of Cyclin B1 expression, with significant loss of the miRNA observed in breast tumours.

  4. The 19q12 bladder cancer GWAS signal: association with cyclin E function and aggressive disease

    Science.gov (United States)

    Fu, Yi-Ping; Kohaar, Indu; Moore, Lee E.; Lenz, Petra; Figueroa, Jonine D.; Tang, Wei; Porter-Gill, Patricia; Chatterjee, Nilanjan; Scott-Johnson, Alexandra; Garcia-Closas, Montserrat; Muchmore, Brian; Baris, Dalsu; Paquin, Ashley; Ylaya, Kris; Schwenn, Molly; Apolo, Andrea B.; Karagas, Margaret R.; Tarway, McAnthony; Johnson, Alison; Mumy, Adam; Schned, Alan; Guedez, Liliana; Jones, Michael A.; Kida, Masatoshi; Monawar Hosain, GM; Malats, Nuria; Kogevinas, Manolis; Tardon, Adonina; Serra, Consol; Carrato, Alfredo; Garcia-Closas, Reina; Lloreta, Josep; Wu, Xifeng; Purdue, Mark; Andriole, Gerald L.; Grubb, Robert L.; Black, Amanda; Landi, Maria T.; Caporaso, Neil E.; Vineis, Paolo; Siddiq, Afshan; Bueno-de-Mesquita, H. Bas; Trichopoulos, Dimitrios; Ljungberg, Börje; Severi, Gianluca; Weiderpass, Elisabete; Krogh, Vittorio; Dorronsoro, Miren; Travis, Ruth C.; Tjønneland, Anne; Brennan, Paul; Chang-Claude, Jenny; Riboli, Elio; Prescott, Jennifer; Chen, Constance; De Vivo, Immaculata; Govannucci, Edward; Hunter, David; Kraft, Peter; Lindstrom, Sara; Gapstur, Susan M.; Jacobs, Eric J.; Diver, W. Ryan; Albanes, Demetrius; Weinstein, Stephanie J.; Virtamo, Jarmo; Kooperberg, Charles; Hohensee, Chancellor; Rodabough, Rebecca J.; Cortessis, Victoria K.; Conti, David V.; Gago-Dominguez, Manuela; Stern, Mariana C.; Pike, Malcolm C.; Van Den Berg, David; Yuan, Jian-Min; Haiman, Christopher A.; Cussenot, Olivier; Cancel-Tassin, Geraldine; Roupret, Morgan; Comperat, Eva; Porru, Stefano; Carta, Angela; Pavanello, Sofia; Arici, Cecilia; Mastrangelo, Giuseppe; Grossman, H. Barton; Wang, Zhaoming; Deng, Xiang; Chung, Charles C.; Hutchinson, Amy; Burdette, Laurie; Wheeler, William; Fraumeni, Joseph; Chanock, Stephen J.; Hewitt, Stephen M.; Silverman, Debra T.; Rothman, Nathaniel; Prokunina-Olsson, Ludmila

    2014-01-01

    A genome-wide association study (GWAS) of bladder cancer identified a genetic marker rs8102137 within the 19q12 region as a novel susceptibility variant. This marker is located upstream of the CCNE1 gene, which encodes cyclin E, a cell cycle protein. We performed genetic fine mapping analysis of the CCNE1 region using data from two bladder cancer GWAS (5,942 cases and 10,857 controls). We found that the original GWAS marker rs8102137 represents a group of 47 linked SNPs (with r2≥0.7) associated with increased bladder cancer risk. From this group we selected a functional promoter variant rs7257330, which showed strong allele-specific binding of nuclear proteins in several cell lines. In both GWAS, rs7257330 was associated only with aggressive bladder cancer, with a combined per-allele odds ratio (OR) =1.18 (95%CI=1.09-1.27, p=4.67×10−5 vs. OR =1.01 (95%CI=0.93-1.10, p=0.79) for non-aggressive disease, with p=0.0015 for case-only analysis. Cyclin E protein expression analyzed in 265 bladder tumors was increased in aggressive tumors (p=0.013) and, independently, with each rs7257330-A risk allele (ptrend=0.024). Over-expression of recombinant cyclin E in cell lines caused significant acceleration of cell cycle. In conclusion, we defined the 19q12 signal as the first GWAS signal specific for aggressive bladder cancer. Molecular mechanisms of this genetic association may be related to cyclin E over-expression and alteration of cell cycle in carriers of CCNE1 risk variants. In combination with established bladder cancer risk factors and other somatic and germline genetic markers, the CCNE1 variants could be useful for inclusion into bladder cancer risk prediction models. PMID:25320178

  5. Regulatory mechanism of radiation-induced cancer cell death by the change of cell cycle

    Energy Technology Data Exchange (ETDEWEB)

    Jeong, Soo Jin; Jeong, Min Ho; Jang, Ji Yeon [College of Medicine, Donga Univ., Pusan (Korea, Republic of)

    2003-09-01

    In our previous study, we have shown the main cell death pattern induced by irradiation or protein tyrosine kinase (PTK) inhibitors in K562 human myelogenous leukemic cell line. Death of the cells treated with irradiation alone was characterized by mitotic catastrophe and typical radiation-induced apoptosis was accelerated by herbimycin A (HMA). Both types of cell death were inhibited by genistein. In this study, we investigated the effects of HMA and genistein on cell cycle regulation and its correlation with the alterations of radiation-induced cell death. K562 cells in exponential growth phase were used for this study. The cells were irradiated with 10 Gy using 6 MeV Linac (200-300 cGy/min). Immediately after irradiation, cells were treated with 250 nM of HMA or 25{mu}M of genistein. The distributions of cell cycle, the expressions of cell cycle-related protein, the activities of cyclin-dependent kinase, and the yield of senescence and differentiation were analyzed. X-irradiated cells were arrested in the G2 phase of the cell cycle but unlike the p53-positive cells, they were not able to sustain the cell cycle arrest. An accumulation of cells in G2 phase of first cell-cycle post-treatment and an increase of cyclin B1 were correlated with spontaneous, premature, chromosome condensation and mitotic catastrophe. HMA induced rapid G2 checkpoint abrogation and concomitant p53-independent G1 accumulation HMA-induced cell cycle modifications correlated with the increase of cdc2 kinase activity, the decrease of the expressions of cyclins E and A and of CDK2 kinase activity, and the enhancement of radiation-induced apoptosis. Genistein maintained cells that were arrested in the G2-phase, decreased the expressions of cyclin B1 and cdc25C and cdc2 kinase activity, increased the expression of p16, and sustained senescence and megakaryocytic differentiation. The effects of HMA and genistein on the radiation-induced cell death of K562 cells were closely related to the cell

  6. Accelerated Stem Growth Rates and Improved Fiber Properties of Loblolly Pine: Functional Analysis Of CyclinD from Pinus taeda

    Energy Technology Data Exchange (ETDEWEB)

    Dr. John Cairney, School of Biology and Institute of Paper Science and Technology @ Georgia Tech, Georgia Institute of Technology; Dr. Gary Peter, University of Florida; Dr. Ulrika Egertsdotter, Dept. of Forestry, Virgina Tech; Dr. Armin Wagner, New Zealand Forest Research Institute Ltd. (Scion Research.)

    2005-11-30

    A sustained supply of low-cost, high quality raw materials is essential for the future success of the U.S. forest products industry. To maximize stem (trunk) growth, a fundamental understanding of the molecular mechanisms that regulate cell divisions within the cambial meristem is essential. We hypothesize that auxin levels within the cambial meristem regulate cyclin gene expression and this in turn controls cell cycle progression as occurs in all eukaryotic cells. Work with model plant species has shown that ectopic overexpression of cyclins promotes cell division thereby increasing root growth > five times. We intended to test whether ectopic overexpression of cambial cyclins in the cambial zone of loblolly pine also promotes cell division rates that enhance stem growth rates. Results generated in model annual angiosperm systems cannot be reliably extrapolated to perennial gymnosperms, thus while the generation and development of transgenic pine is time consuming, this is the necessary approach for meaningful data. We succeeded in isolating a cyclin D gene and Clustal analysis to the Arabidopsis cyclin D gene family indicates that it is more closely related to cyclin D2 than D1 or D3 Using this gene as a probe we observed a small stimulation of cyclin D expression in somatic embryo culture upon addition of auxin. We hypothesized that trees with more cells in the vascular cambial and expansion zones will have higher cyclin mRNA levels. We demonstrated that in trees under compressive stress where the rates of cambial divisions are increased on the underside of the stem relative to the top or opposite side, there was a 20 fold increase in the level of PtcyclinD1 mRNA on the compressed side of the stem relative to the opposite. This suggests that higher secondary growth rates correlate with PtcyclinD1 expression. We showed that larger diameter trees show more growth during each year and that the increased growth in loblolly pine trees correlates with more cell

  7. Zinc protoporphyrin regulates cyclin D1 expression independent of heme oxygenase inhibition.

    Science.gov (United States)

    La, Ping; Fernando, Amal P; Wang, Zhi; Salahudeen, Ameen; Yang, Guang; Lin, Qing; Wright, Clyde J; Dennery, Phyllis A

    2009-12-25

    Zinc protoporphyrin IX (ZnPP), an endogenous heme analogue that inhibits heme oxygenase (HO) activity, represses tumor growth. It can also translocate into the nucleus and up-regulate heme oxygenase 1 (HMOX1) gene expression. Here, we demonstrate that tumor cell proliferation was inhibited by ZnPP, whereas tin protoporphyrin (SnPP), another equally potent HO-1 inhibitor, had no effect. Microarray analysis on 128 tumorigenesis related genes showed that ZnPP suppressed genes involved in cell proliferation and angiogenesis. Among these genes, CYCLIN D1 (CCND1) was specifically inhibited as were its mRNA and protein levels. Additionally, ZnPP inhibited CCND1 promoter activity through an Sp1 and Egr1 overlapping binding site (S/E). We confirmed that ZnPP modulated the S/E site, at least partially by associating with Sp1 and Egr1 proteins rather than direct binding to DNA targets. Furthermore, administration of ZnPP significantly inhibited cyclin D1 expression and progression of a B-cell leukemia/lymphoma 1 tumor in mice by preferentially targeting tumor cells. These observations show HO independent effects of ZnPP on cyclin D1 expression and tumorigenesis.

  8. Zinc Protoporphyrin Regulates Cyclin D1 Expression Independent of Heme Oxygenase Inhibition*

    Science.gov (United States)

    La, Ping; Fernando, Amal P.; Wang, Zhi; Salahudeen, Ameen; Yang, Guang; Lin, Qing; Wright, Clyde J.; Dennery, Phyllis A.

    2009-01-01

    Zinc protoporphyrin IX (ZnPP), an endogenous heme analogue that inhibits heme oxygenase (HO) activity, represses tumor growth. It can also translocate into the nucleus and up-regulate heme oxygenase 1 (HMOX1) gene expression. Here, we demonstrate that tumor cell proliferation was inhibited by ZnPP, whereas tin protoporphyrin (SnPP), another equally potent HO-1 inhibitor, had no effect. Microarray analysis on 128 tumorigenesis related genes showed that ZnPP suppressed genes involved in cell proliferation and angiogenesis. Among these genes, CYCLIN D1 (CCND1) was specifically inhibited as were its mRNA and protein levels. Additionally, ZnPP inhibited CCND1 promoter activity through an Sp1 and Egr1 overlapping binding site (S/E). We confirmed that ZnPP modulated the S/E site, at least partially by associating with Sp1 and Egr1 proteins rather than direct binding to DNA targets. Furthermore, administration of ZnPP significantly inhibited cyclin D1 expression and progression of a B-cell leukemia/lymphoma 1 tumor in mice by preferentially targeting tumor cells. These observations show HO independent effects of ZnPP on cyclin D1 expression and tumorigenesis. PMID:19850937

  9. The LIM-only protein FHL2 mediates ras-induced transformation through cyclin D1 and p53 pathways.

    Directory of Open Access Journals (Sweden)

    Charlotte Labalette

    Full Text Available BACKGROUND: Four and a half LIM-only protein 2 (FHL2 has been implicated in multiple signaling pathways that regulate cell growth and tissue homeostasis. We reported previously that FHL2 regulates cyclin D1 expression and that immortalized FHL2-null mouse embryo fibroblasts (MEFs display reduced levels of cyclin D1 and low proliferative activity. METHODOLOGY/PRINCIPAL FINDINGS: Here we address the contribution of FHL2 in cell transformation by investigating the effects of oncogenic Ras in FHL2-null context. We show that H-RasV12 provokes cell cycle arrest accompanied by accumulation of p53 and p16(INK4a in immortalized FHL2(-/- MEFs. These features contrast sharply with Ras transforming activity in wild type cell lines. We further show that establishment of FHL2-null cell lines differs from conventional immortalization scheme by retaining functional p19(ARF/p53 checkpoint that is required for cell cycle arrest imposed by Ras. However, after serial passages of Ras-expressing FHL2(-/- cells, dramatic increase in the levels of D-type cyclins and Rb phosphorylation correlates with the onset of cell proliferation and transformation without disrupting the p19(ARF/p53 pathway. Interestingly, primary FHL2-null cells overexpressing cyclin D1 undergo a classical immortalization process leading to loss of the p19(ARF/p53 checkpoint and susceptibility to Ras transformation. CONCLUSIONS/SIGNIFICANCE: Our findings uncover a novel aspect of cellular responses to mitogenic stimulation and illustrate a critical role of FHL2 in the signalling network that implicates Ras, cyclin D1 and p53.

  10. Anti-cancer activity ofBacillus amyloliquefaciensAK-0 through cyclin D1 proteasomal degradationviaGSK3β-dependent phosphorylation of threonine-286.

    Science.gov (United States)

    Park, Gwang Hun; Song, Hun Min; Kim, Young Soo; Jeon, Yongho; Koo, Jin Suk; Jeong, Hyung Jin; Jeong, Jin Boo

    2017-06-01

    Microorganisms have been regarded as important sources of novel bioactive natural products. In this study, we evaluated the anti-cancer activity and the potential mechanism of Bacillus amyloliquefaciens AK-0 newly isolated from the rhizosphere soil of Korean ginseng. The ethyl acetate fraction from the culture medium of B. amyloliquefaciens AK-0 (EA-AK0) inhibited markedly the proliferation of human colorectal cancer cells such as HCT116, SW480, LoVo and HT-29. EA-AK0 effectively decreased cyclin D1 protein level in human colorectal cancer cells, while cyclin D1 mRNA level was not changed by EA-AK0 treatment. Inhibition of proteasomal degradation by MG132 blocked EA-AK0-mediated cyclin D1 downregulation and the half-life of cyclin D1 was decreased in the cells treated with MRB. In addition, EA-AK0 increased threonine-286 (T286) phosphorylation of cyclin D1, and a point mutation of T286 to alanine attenuated cyclin D1 degradation by EA-AK0. Inhibition of GSK3β by LiCl suppressed cyclin D1 phosphorylation and downregulation by EA-AKO. From these results, EA-AK0 may suppress the proliferation of human colorectal cancer cells by inducing cyclin D1 proteasomal degradation through GSK3β-dependent T286 phosphorylation. These results indicate that EA-AK0 could be used for treating colorectal cancer and serve as a potential candidate for anticancer drug development. In addition, these findings will be helpful for expanding the knowledge on the molecular anti-cancer mechanisms of EA-AK0.

  11. Relationship between cyclin G1 and human papilloma virus infection in cervical intraepithelial neoplasia and cervical carcinoma.

    Science.gov (United States)

    Liang, Jing; Bian, Mei-Lu; Chen, Qing-Yun; Liu, Xia; Ou, Hua; Li, Min; Liu, Jun

    2006-06-01

    To evaluate the overexpression of cyclin G1 in cervical intraepithelial neoplasia (CIN) and cervical carcinoma, and the correlation between cyclin G1 and high-risk human papilloma virus (HPV) infection. All of the specimens were obtained from the Department of Pathology of China-Japan Friendship Hospital from January 2000 to August 2004. We detected the expression of cyclin G1 with immunohistochemistry, HPV16/18 infection with in situ hybridization, and high-risk HPV infection with Hybrid capture system II (HC-II) in normal group (25 cases), CIN I (48 cases), CIN II (56 cases), CIN III (54 cases), and invasive cervical squamous-cell carcinoma (SCC, 31 cases). The positive rates of cyclin G1 expression in CIN (77.85%) and SCC cervical tissues (87.10%) were significantly higher than normal (8.00%, P biomarker for detecting CIN and SCC. Cyclin G1 may play an important role in the oncogenesis of CIN and SCC by high-risk HPV infection.

  12. Phosphorylation of pRb by cyclin D kinase is necessary for development of cardiac hypertrophy

    DEFF Research Database (Denmark)

    Hinrichsen, Rebecca; Hansen, A.H.; Haunsø, S.

    2008-01-01

    OBJECTIVES: A number of stimuli induce cardiac hypertrophy and may lead to cardiomyopathy and heart failure. It is believed that cardiomyocytes withdraw from the cell cycle shortly after birth and become terminally differentiated. However, cell cycle regulatory proteins take part in the development...... growth in cardiomyocytes, whereas cyclin E-cdk2 kinase is not necessary for hypertrophy but regulates endoreplication in these cells. The data support the notion that hypertrophic growth of cardiomyocytes involves a partial progression through the G1 phase of the cell cycle Udgivelsesdato: 2008/10...

  13. Calcium-induced chromatin condensation and cyclin phosphorylation during chromatin condensation cycles in ammonia-activated sea urchin eggs.

    Science.gov (United States)

    Patel, R; Twigg, J; Crossley, I; Golsteyn, R; Whitaker, M

    1989-01-01

    Ammonia-activated sea urchin eggs undergo repeated cycles of DNA synthesis, nuclear envelope breakdown (NEB) and chromatin condensation. No mitotic spindle forms, nor do the eggs undergo cytokinesis. Ammonia-activated eggs exhibit a form of the cell cycle in which the nuclear cycle proceeds without segregation of the chromatin into daughter cells. We discuss here experiments that demonstrate that intracellular free calcium concentration controls the S phase-M phase transition in ammonia-activated eggs, as it does in fertilized embryos. Cyclins are proteins that are synthesized throughout the cell cycle and destroyed abruptly during each round of chromatin condensation. We find that cycles of cyclin phosphorylation and destruction occur coincident with chromatin condensation in ammonia-activated eggs. Cyclin phosphorylation also occurs in eggs treated with the tumour promoter, phorbol myristate acetate (PMA). There is no accompanying NEB or chromatin condensation, however, and the nucleus is insensitive to exogenously-generated calcium transients. These latter data indicate that cyclin synthesis and phosphorylation is not a sufficient condition for calcium-induced NEB in sea urchin embryos. PMA must fail to induce one of the necessary cell cycle initiation signals. We suggest that the missing signal is the activation of the cell cycle control protein p34cdc2, which we have shown to be phosphorylated at fertilization and which is phosphorylated in ammonia-activated eggs.

  14. Active Component of Danshen (Salvia miltiorrhiza Bunge, Tanshinone I, Attenuates Lung Tumorigenesis via Inhibitions of VEGF, Cyclin A, and Cyclin B Expressions

    Directory of Open Access Journals (Sweden)

    Yu-Tang Tung

    2013-01-01

    Full Text Available Tanshinone I (T1 and tanshinone II (T2 are the major diterpenes isolated from Danshen (Salvia miltiorrhiza Bunge. Three human lung adenocarcinoma cell lines, A549, CL1-0, and CL1-5, were treated with T1 and T2 for the in vitro antitumor test. Results showed that T1 was more effective than T2 in inhibiting the growth of lung cancer cells via suppressing the expression of VEGF, Cyclin A, and Cyclin B proteins in a dose-dependent manner. Moreover, a transgenic mice model of the human vascular endothelial growth factor-A165 (hVEGF-A165 gene-induced pulmonary tumor was further treated with T1 for the in vivo lung cancer therapy test. T1 significantly attenuated hVEGF-A165 overexpression to normal levels of the transgenic mice (Tg that were pretreated with human monocytic leukemia THP-1 cell-derived conditioned medium (CM. It also suppressed the formation of lung adenocarcinoma tumors (16.7% compared with two placebo groups (50% for Tg/Placebo and 83.3% for Tg/CM/Placebo; P<0.01. This antitumor effect is likely to slow the progression of cells through the S and G2/M phases of the cell cycle. Blocking of the tumor-activated cell cycle pathway may be a critical mechanism for the observed antitumorigenic effects of T1 treatment on vasculogenesis and angiogenesis.

  15. Identifying and Assessing Life-Cycle-Related Critical Technology Elements (CTEs) for Technology Readiness Assessments (TRAs)

    National Research Council Canada - National Science Library

    Mandelbaum, Jay

    2006-01-01

    .... Because these technologies are not emphasized in the current Technology Readiness Assessment (TRA) process this document is intended to improve the focus on life-cycle-related technologies in TRAs...

  16. Therapeutic effects of lentivirus-mediated shRNA targeting of cyclin D1 in human gastric cancer

    International Nuclear Information System (INIS)

    Seo, Jin-Hee; Jeong, Eui-Suk; Choi, Yang-Kyu

    2014-01-01

    Gastric cancer is the second most common cause of cancer-related death in males and the fourth in females. Traditional treatment has poor prognosis because of recurrence and systemic side effects. Therefore, the development of new therapeutic strategies is an important issue. Lentivirus-mediated shRNA stably inhibits target genes and can efficiently transduce most cells. Since overexpressed cyclin D1 is closely related to human gastric cancer progression, inhibition of cyclin D1 using specific targeting could be an effective treatment method of human gastric cancer. The therapeutic effect of lentivirus-mediated shRNA targeting of cyclin D1 (ShCCND1) was analyzed both in vitro and in vivo experiments. In vitro, NCI-N87 cells with downregulation of cyclin D1 by ShCCND1 showed significant inhibition of cell proliferation, cell motility, and clonogenicity. Downregulation of cyclin D1 in NCI-N87 cells also resulted in significantly increased G1 arrest and apoptosis. In vivo, stable NCI-N87 cells expressing ShCCND1 were engrafted into nude mice. Then, the cancer-growth inhibition effect of lentivirus was confirmed. To assess lentivirus including ShCCND1 as a therapeutic agent, intratumoral injection was conducted. Tumor growth of the lentivirus-treated group was significantly inhibited compared to growth of the control group. These results are in accordance with the in vitro data and lend support to the mitotic figure count and apoptosis analysis of the tumor mass. The lentivirus-mediated ShCCND1 was constructed, which effectively inhibited growth of NCI-N87-derived cancer both in vitro and in vivo. The efficiency of shRNA knockdown and variation in the degree of inhibition is mediated by different shRNA sequences and cancer cell lines. These experimental results suggest the possibility of developing new gastric cancer therapies using lentivirus-mediated shRNA

  17. Hormone-induced DNA damage response and repair mediated by cyclin D1 in breast and prostate cancer.

    Science.gov (United States)

    Di Sante, Gabriele; Di Rocco, Agnese; Pupo, Claudia; Casimiro, Mathew C; Pestell, Richard G

    2017-10-10

    Cell cycle control proteins govern events that leads to the production of two identical daughter cells. Distinct sequential temporal phases, Gap 1 (G 1 ), Gap 0 (G 0 ), Synthesis (S), Gap 2 (G 2 ) and Mitosis (M) are negotiated through a series of check points during which the favorability of the local cellular environment is assessed, prior to replicating DNA [1]. Cyclin D1 has been characterized as a key regulatory subunit of the holoenzyme that promotes the G 1 /S-phase transition through phosphorylating the pRB protein. Cyclin D1 overexpression is considered a driving force in several types of cancers and cdk inhibitors are being used effectively in the clinic for treatment of ERα + breast cancer [1, 2]. Genomic DNA is assaulted by damaging ionizing radiation, chemical carcinogens, and reactive oxygen species (ROS) which are generated by cellular metabolism. Furthermore, specific hormones including estrogens [3, 4] and androgens [5] govern pathways that damage DNA. Defects in the DNA Damage Response (DDR) pathway can lead to genomic instability and cancer. Evidence is emerging that cyclin D1 bind proteins involved in DNA repair including BRCA1 [6], RAD51 [7], BRCA2 [8] and is involved in the DNA damage and DNA repair processes [7, 8]. Because the repair of damaged DNA appears to be an important and unexpected role for cyclin D1, and inhibitors of cyclin D1-dependent kinase activity are being used in the clinic, the latest findings on the role of cyclin D1 in mediating the DDR including the DDR induced by the hormones estrogen [9] and androgen [10, 11] is reviewed.

  18. Cyclin-dependent protein kinase and cyclin homologs SSN3 and SSN8 contribute to transcriptional control in yeast.

    OpenAIRE

    Kuchin, S; Yeghiayan, P; Carlson, M

    1995-01-01

    The SSN3 and SSN8 genes of Saccharomyces cerevisiae were identified by mutations that suppress a defect in SNF1, a protein kinase required for release from glucose repression. Mutations in SSN3 and SSN8 also act synergistically with a mutation of the MIG1 repressor protein to relieve glucose repression. We have cloned the SSN3 and SSN8 genes. SSN3 encodes a cyclin-dependent protein kinase (cdk) homolog and is identical to UME5. SSN8 encodes a cyclin homolog 35% identical to human cyclin C. SS...

  19. Maintenance of motor neuron progenitors in Xenopus requires a novel localized cyclin.

    Science.gov (United States)

    Chen, Jun-An; Chu, Sin-Tak; Amaya, Enrique

    2007-03-01

    The ventral spinal cord contains a pool of motor neuron progenitors (pMNs), which sequentially generate motor neurons and oligodendrocytes in the embryo. The mechanisms responsible for the maintenance of pMNs are not clearly understood. We have identified a novel cyclin, cyclin Dx (ccndx), which is specifically expressed in pMNs in Xenopus. Here, we show that inhibition of ccndx causes paralysis in embryos. Furthermore, we show that maintenance of pMNs requires ccndx function. In addition, inhibition of ccndx results in the specific loss of differentiated motor neurons. However, the expression of interneuron or sensory neuron markers is unaffected in these embryos, suggesting that the role of ccndx is specifically to maintain pMNs. Thus, we have identified, for the first time, a tissue-specific cell-cycle regulator that is essential for the maintenance of a pool of neural progenitors in the vertebrate spinal cord.

  20. Targeting Cyclin-Dependent Kinases in Human Cancers: From Small Molecules to Peptide Inhibitors

    Directory of Open Access Journals (Sweden)

    Marion Peyressatre

    2015-01-01

    Full Text Available Cyclin-dependent kinases (CDK/Cyclins form a family of heterodimeric kinases that play central roles in regulation of cell cycle progression, transcription and other major biological processes including neuronal differentiation and metabolism. Constitutive or deregulated hyperactivity of these kinases due to amplification, overexpression or mutation of cyclins or CDK, contributes to proliferation of cancer cells, and aberrant activity of these kinases has been reported in a wide variety of human cancers. These kinases therefore constitute biomarkers of proliferation and attractive pharmacological targets for development of anticancer therapeutics. The structural features of several of these kinases have been elucidated and their molecular mechanisms of regulation characterized in depth, providing clues for development of drugs and inhibitors to disrupt their function. However, like most other kinases, they constitute a challenging class of therapeutic targets due to their highly conserved structural features and ATP-binding pocket. Notwithstanding, several classes of inhibitors have been discovered from natural sources, and small molecule derivatives have been synthesized through rational, structure-guided approaches or identified in high throughput screens. The larger part of these inhibitors target ATP pockets, but a growing number of peptides targeting protein/protein interfaces are being proposed, and a small number of compounds targeting allosteric sites have been reported.

  1. Acquired radioresistance of cancer and the AKT/GSK3β/cyclin D1 overexpression cycle

    International Nuclear Information System (INIS)

    Shimura, Tsutomu

    2011-01-01

    Fractionated radiotherapy (RT) is widely used in cancer therapy for its advantages in the preservation of normal tissues. However, repopulation of surviving tumor cells during fractionated RT limits the efficacy of RT. In fact, repopulating tumors often acquire radioresistance and this is the major cause of failure of RT. We have recently demonstrated that human tumor cells acquire radioresistance when exposed to fractionated radiation (FR) of X-rays every 12 hours for 1 month. The acquired radioresistance was associated with overexpression of cyclin D1, a result of a series of molecular changes; constitutive activation of DNA-PK and AKT with concomitant down-regulation of glycogen synthase kinase-3β (GSK3β) which results in suppression of cyclin D1 proteolysis. Aberrant cyclin D1 overexpression in S-phase induced DNA double strand breaks which activated DNA-PK and established the vicious cycle of cycling D1 overexpression. This overexpression of cyclin D1 is responsible for the radioresistance phenotype of long-term FR cells, since this phenotype was completely abrogated by treatment of FR cells by the AKT/PKB signaling inhibitor (API-2), an AKT inhibitor or by a Cdk4 inhibitor. Thus, targeting the AKT/GSK3β/cyclin D1/Cdk4 pathway can be an efficient modality to suppress acquired radioresistance of tumor cells. In this article, I overview the newly discovered molecular mechanisms underlying acquired radioresistance of tumor cells induced by FR, and propose a strategy for eradication of tumors using fractionated RT by overcoming tumor radioresistance. (author)

  2. Coating independent cytotoxicity of citrate- and PEG-coated silver nanoparticles on a human hepatoma cell line.

    Science.gov (United States)

    Bastos, Verónica; Ferreira-de-Oliveira, José M P; Carrola, Joana; Daniel-da-Silva, Ana L; Duarte, Iola F; Santos, Conceição; Oliveira, Helena

    2017-01-01

    The antibacterial potential of silver nanoparticles (AgNPs) resulted in their increasing incorporation into consumer, industrial and biomedical products. Therefore, human and environmental exposure to AgNPs (either as an engineered product or a contaminant) supports the emergent research on the features conferring them different toxicity profiles. In this study, 30nm AgNPs coated with citrate or poly(ethylene glycol) (PEG) were used to assess the influence of coating on the effects produced on a human hepatoma cell line (HepG2), namely in terms of viability, apoptosis, apoptotic related genes, cell cycle and cyclins gene expression. Both types of coated AgNPs decreased cell proliferation and viability with a similar toxicity profile. At the concentrations used (11 and 5μg/mL corresponding to IC50 and ~IC10 levels, respectively) the amount of cells undergoing apoptosis was not significant and the apoptotic related genes BCL2 (anti-apoptotic gene) and BAX (pro-apoptotic gene) were both downregulated. Moreover, both AgNPs affected HepG2 cell cycle progression at the higher concentration (11μg/mL) by increasing the percentage of cells in S (synthesis phase) and G2 (Gap 2 phase) phases. Considering the cell-cycle related genes, the expression of cyclin B1 and cyclin E1 genes were decreased. Thus, this work has shown that citrate- and PEG-coated AgNPs impact on HepG2 apoptotic gene expression, cell cycle dynamics and cyclin regulation in a similar way. More research is needed to determine the properties that confer AgNPs at lower toxicity, since their use has proved helpful in several industrial and biomedical contexts. Copyright © 2016. Published by Elsevier B.V.

  3. Cyclin-dependent kinase Inhibitors Inspired by Roscovitine: Purine Bioisosteres

    Czech Academy of Sciences Publication Activity Database

    Jorda, Radek; Paruch, K.; Kryštof, Vladimír

    2012-01-01

    Roč. 18, č. 20 (2012), s. 2974-2980 ISSN 1381-6128 R&D Projects: GA ČR GAP305/12/0783 Institutional research plan: CEZ:AV0Z50380511 Keywords : Cancer * cyclin-dependent kinase * inhibitor Subject RIV: ED - Physiology Impact factor: 3.311, year: 2012 http://www.benthamdirect.org/pages/article/1/3177382/cyclin-dependent-kinase-inhibitors-inspired-by-roscovitine-purine-bioisosteres.html

  4. DNA-PKcs Negatively Regulates Cyclin B1 Protein Stability through Facilitating Its Ubiquitination Mediated by Cdh1-APC/C Pathway.

    Science.gov (United States)

    Shang, Zeng-Fu; Tan, Wei; Liu, Xiao-Dan; Yu, Lan; Li, Bing; Li, Ming; Song, Man; Wang, Yu; Xiao, Bei-Bei; Zhong, Cai-Gao; Guan, Hua; Zhou, Ping-Kun

    2015-01-01

    The catalytic subunit of DNA-dependent protein kinase (DNA-PKcs) is a critical component of the non-homologous end-joining pathway of DNA double-stranded break repair. DNA-PKcs has also been shown recently functioning in mitotic regulation. Here, we report that DNA-PKcs negatively regulates the stability of Cyclin B1 protein through facilitating its ubiquitination mediated by Cdh1 / E 3 ubiquitin ligase APC/C pathway. Loss of DNA-PKcs causes abnormal accumulation of Cyclin B1 protein. Cyclin B1 degradation is delayed in DNA-PKcs-deficient cells as result of attenuated ubiquitination. The impact of DNA-PKcs on Cyclin B1 stability relies on its kinase activity. Our study further reveals that DNA-PKcs interacts with APC/C core component APC2 and its co-activator Cdh1. The destruction of Cdh1 is accelerated in the absence of DNA-PKcs. Moreover, overexpression of exogenous Cdh1 can reverse the increase of Cyclin B1 protein in DNA-PKcs-deficient cells. Thus, DNA-PKcs, in addition to its direct role in DNA damage repair, functions in mitotic progression at least partially through regulating the stability of Cyclin B1 protein.

  5. Oct-1 potentiates CREB-driven cyclin D1 promoter activation via a phospho-CREB- and CREB binding protein-independent mechanism.

    Science.gov (United States)

    Boulon, Séverine; Dantonel, Jean-Christophe; Binet, Virginie; Vié, Annick; Blanchard, Jean-Marie; Hipskind, Robert A; Philips, Alexandre

    2002-11-01

    Cyclin D1, the regulatory subunit for mid-G(1) cyclin-dependent kinases, controls the expression of numerous cell cycle genes. A cyclic AMP-responsive element (CRE), located upstream of the cyclin D1 mRNA start site, integrates mitogenic signals that target the CRE-binding factor CREB, which can recruit the transcriptional coactivator CREB-binding protein (CBP). We describe an alternative mechanism for CREB-driven cyclin D1 induction that involves the ubiquitous POU domain protein Oct-1. In the breast cancer cell line MCF-7, overexpression of Oct-1 or its POU domain strongly increases transcriptional activation of cyclin D1 and GAL4 reporter genes that is specifically dependent upon CREB but independent of Oct-1 DNA binding. Gel retardation and chromatin immunoprecipitation assays confirm that POU forms a complex with CREB bound to the cyclin D1 CRE. In solution, CREB interaction with POU requires the CREB Q2 domain and, notably, occurs with CREB that is not phosphorylated on Ser 133. Accordingly, Oct-1 also potently enhances transcriptional activation mediated by a Ser133Ala CREB mutant. Oct-1/CREB synergy is not diminished by the adenovirus E1A 12S protein, a repressor of CBP coactivator function. In contrast, E1A strongly represses CBP-enhanced transactivation by CREB phosphorylated on Ser 133. Our observation that Oct-1 potentiates CREB-dependent cyclin D1 transcriptional activity independently of Ser 133 phosphorylation and E1A-sensitive coactivator function offers a new paradigm for the regulation of cyclin D1 induction by proliferative signals.

  6. Phosphorylation of mammalian CDC6 by cyclin A/CDK2 regulates its subcellular localization

    DEFF Research Database (Denmark)

    Petersen, B O; Lukas, J; Sørensen, Claus Storgaard

    1999-01-01

    by CDKs. CDC6 interacts specifically with the active Cyclin A/CDK2 complex in vitro and in vivo, but not with Cyclin E or Cyclin B kinase complexes. The cyclin binding domain of CDC6 was mapped to an N-terminal Cy-motif that is similar to the cyclin binding regions in p21(WAF1/SDI1) and E2F-1. The in vivo...

  7. Identification of a cyclin B1-derived CTL epitope eliciting spontaneous responses in both cancer patients and healthy donors

    DEFF Research Database (Denmark)

    Andersen, Rikke Sick; Sørensen, Rikke Bæk; Ritter, Cathrin

    2011-01-01

    . Furthermore, blood from cancer patients and healthy donors was screened for spontaneous T-cell reactivity against the peptide in IFN-¿ ELISPOT assays. Patients with breast cancer, malignant melanoma, or renal cell carcinoma hosted powerful and high-frequency T-cell responses against the peptide. In addition......With the aim to identify cyclin B1-derived peptides with high affinity for HLA-A2, we used three in silico prediction algorithms to screen the protein sequence for possible HLA-A2 binders. One peptide scored highest in all three algorithms, and the high HLA-A2-binding affinity of this peptide......, a high-affinity cyclin B1-derived HLA-A2-restricted CTL epitope was identified, which was presented on the cell surface of cancer cells, and elicited spontaneous T-cell responses in cancer patients and healthy donors....

  8. Identification of a cyclin B1-derived CTL epitope eliciting spontaneous responses in both cancer patients and healthy donors

    DEFF Research Database (Denmark)

    Andersen, Rikke Sick; Sørensen, Rikke Bæk; Ritter, Cathrin

    2011-01-01

    . Furthermore, blood from cancer patients and healthy donors was screened for spontaneous T-cell reactivity against the peptide in IFN-γ ELISPOT assays. Patients with breast cancer, malignant melanoma, or renal cell carcinoma hosted powerful and high-frequency T-cell responses against the peptide. In addition......With the aim to identify cyclin B1-derived peptides with high affinity for HLA-A2, we used three in silico prediction algorithms to screen the protein sequence for possible HLA-A2 binders. One peptide scored highest in all three algorithms, and the high HLA-A2-binding affinity of this peptide......, a high-affinity cyclin B1-derived HLA-A2-restricted CTL epitope was identified, which was presented on the cell surface of cancer cells, and elicited spontaneous T-cell responses in cancer patients and healthy donors....

  9. Pfaffosidic Fraction from Hebanthe paniculata Induces Cell Cycle Arrest and Caspase-3-Induced Apoptosis in HepG2 Cells

    Directory of Open Access Journals (Sweden)

    Tereza Cristina da Silva

    2015-01-01

    Full Text Available Hebanthe paniculata roots (formerly Pfaffia paniculata and popularly known as Brazilian ginseng show antineoplastic, chemopreventive, and antiproliferative properties. Functional properties of these roots and their extracts are usually attributed to the pfaffosidic fraction, which is composed mainly by pfaffosides A–F. However, the therapeutic potential of this fraction in cancer cells is not yet entirely understood. This study aimed to analyze the antitumoral effects of the purified pfaffosidic fraction or saponinic fraction on the human hepatocellular carcinoma HepG2 cell line. Cellular viability, proliferation, and apoptosis were evaluated, respectively, by MTT assay, BrdU incorporation, activated caspase-3 immunocytochemistry, and DNA fragmentation assay. Cell cycle was analyzed by flow cytometry and the cell cycle-related proteins were analyzed by quantitative PCR and Western blot. The cells exposed to pfaffosidic fraction had reduced viability and cellular growth, induced G2/M at 48 h or S at 72 h arrest, and increased sub-G1 cell population via cyclin E downregulation, p27KIP1 overexpression, and caspase-3-induced apoptosis, without affecting the DNA integrity. Antitumoral effects of pfaffosidic fraction from H. paniculata in HepG2 cells originated by multimechanisms of action might be associated with cell cycle arrest in the S phase, by CDK2 and cyclin E downregulation and p27KIP1 overexpression, besides induction of apoptosis through caspase-3 activation.

  10. Targeting the AKT/GSK3β/cyclin D1/Cdk4 survival signaling pathway for eradication of tumor radioresistance acquired by fractionated radiotherapy.

    Science.gov (United States)

    Shimura, Tsutomu; Kakuda, Satoshi; Ochiai, Yasushi; Kuwahara, Yoshikazu; Takai, Yoshihiro; Fukumoto, Manabu

    2011-06-01

    Radioresistance is a major cause of treatment failure of radiotherapy (RT) in human cancer. We have recently revealed that acquired radioresistance of tumor cells induced by fractionated radiation is attributable to cyclin D1 overexpression as a consequence of the downregulation of GSK3β-dependent cyclin D1 proteolysis mediated by a constitutively activated serine-threonine kinase, AKT. This prompted us to hypothesize that targeting the AKT/GSK3β/cyclin D1 pathway may improve fractionated RT by suppressing acquired radioresistance of tumor cells. Two human tumor cell lines with acquired radioresistance were exposed to X-rays after incubation with either an AKT inhibitor, AKT/PKB signaling inhibitor-2 (API-2), or a Cdk4 inhibitor (Cdk4-I). Cells were then subjected to immunoblotting, clonogenic survival assay, cell growth analysis, and cell death analysis with TUNEL and annexin V staining. In vivo radiosensitivity was assessed by growth of human tumors xenografted into nude mice. Treatment with API-2 resulted in downregulation of cyclin D1 expression in cells with acquired radioresistance. Cellular radioresistance disappeared completely both in vitro and in vivo with accompanying apoptosis when treated with API-2. Furthermore, inhibition of cyclin D1/Cdk4 by Cdk4-I was sufficient for abolishing radioresistance. Treatment with either API-2 or Cdk4-I was also effective in suppressing resistance to cis-platinum (II)-diamine-dichloride in the cells with acquired radioresistance. Interestingly, the radiosensitizing effect of API-2 was canceled by overexpression of cyclin D1 whereas Cdk4-I was still able to sensitize cells with cyclin D1 overexpression. Cyclin D1/Cdk4 is a critical target of the AKT survival signaling pathway responsible for tumor radioresistance. Targeting the AKT/GSK3β/cyclin D1/Cdk4 pathway would provide a novel approach to improve fractionated RT and would have an impact on tumor eradication in combination with chemotherapy. Copyright © 2011

  11. CARMA3 is overexpressed in colon cancer and regulates NF-{kappa}B activity and cyclin D1 expression

    Energy Technology Data Exchange (ETDEWEB)

    Miao, Zhifeng; Zhao, Tingting; Wang, Zhenning; Xu, Yingying; Song, Yongxi; Wu, Jianhua [Department of General Surgery, First Affiliated Hospital of China Medical University, Shenyang (China); Xu, Huimian, E-mail: xuhuimianpaper@yahoo.com.cn [Department of General Surgery, First Affiliated Hospital of China Medical University, Shenyang (China)

    2012-09-07

    Highlights: Black-Right-Pointing-Pointer CARMA3 expression is elevated in colon cancers. Black-Right-Pointing-Pointer CARMA3 promotes proliferation and cell cycle progression in colon cancer cells. Black-Right-Pointing-Pointer CARMA3 upregulates cyclinD1 through NF-{kappa}B activation. -- Abstract: CARMA3 was recently reported to be overexpressed in cancers and associated with the malignant behavior of cancer cells. However, the expression of CARMA3 and its biological roles in colon cancer have not been reported. In the present study, we analyzed the expression pattern of CARMA3 in colon cancer tissues and found that CARMA3 was overexpressed in 30.8% of colon cancer specimens. There was a significant association between CARMA3 overexpression and TNM stage (p = 0.0383), lymph node metastasis (p = 0.0091) and Ki67 proliferation index (p = 0.0035). Furthermore, knockdown of CARMA3 expression in HT29 and HCT116 cells with high endogenous expression decreased cell proliferation and cell cycle progression while overexpression of CARMA3 in LoVo cell line promoted cell proliferation and facilitated cell cycle transition. Further analysis showed that CARMA3 knockdown downregulated and its overexpression upregulated cyclin D1 expression and phospho-Rb levels. In addition, we found that CARMA3 depletion inhibited p-I{kappa}B levels and NF-{kappa}B activity and its overexpression increased p-I{kappa}B expression and NF-{kappa}B activity. NF-{kappa}B inhibitor BAY 11-7082 reversed the role of CARMA3 on cyclin D1 upregulation. In conclusion, our study found that CARMA3 is overexpressed in colon cancers and contributes to malignant cell growth by facilitating cell cycle progression through NF-{kappa}B mediated upregulation of cyclin D1.

  12. CARMA3 is overexpressed in colon cancer and regulates NF-κB activity and cyclin D1 expression

    International Nuclear Information System (INIS)

    Miao, Zhifeng; Zhao, Tingting; Wang, Zhenning; Xu, Yingying; Song, Yongxi; Wu, Jianhua; Xu, Huimian

    2012-01-01

    Highlights: ► CARMA3 expression is elevated in colon cancers. ► CARMA3 promotes proliferation and cell cycle progression in colon cancer cells. ► CARMA3 upregulates cyclinD1 through NF-κB activation. -- Abstract: CARMA3 was recently reported to be overexpressed in cancers and associated with the malignant behavior of cancer cells. However, the expression of CARMA3 and its biological roles in colon cancer have not been reported. In the present study, we analyzed the expression pattern of CARMA3 in colon cancer tissues and found that CARMA3 was overexpressed in 30.8% of colon cancer specimens. There was a significant association between CARMA3 overexpression and TNM stage (p = 0.0383), lymph node metastasis (p = 0.0091) and Ki67 proliferation index (p = 0.0035). Furthermore, knockdown of CARMA3 expression in HT29 and HCT116 cells with high endogenous expression decreased cell proliferation and cell cycle progression while overexpression of CARMA3 in LoVo cell line promoted cell proliferation and facilitated cell cycle transition. Further analysis showed that CARMA3 knockdown downregulated and its overexpression upregulated cyclin D1 expression and phospho-Rb levels. In addition, we found that CARMA3 depletion inhibited p-IκB levels and NF-κB activity and its overexpression increased p-IκB expression and NF-κB activity. NF-κB inhibitor BAY 11-7082 reversed the role of CARMA3 on cyclin D1 upregulation. In conclusion, our study found that CARMA3 is overexpressed in colon cancers and contributes to malignant cell growth by facilitating cell cycle progression through NF-κB mediated upregulation of cyclin D1.

  13. Copy Number Defects of G1-Cell Cycle Genes in Neuroblastoma are Frequent and Correlate with High Expression of E2F Target Genes and a Poor Prognosis

    NARCIS (Netherlands)

    Molenaar, Jan J.; Koster, Jan; Ebus, Marli E.; van Sluis, Peter; Westerhout, Ellen M.; de Preter, Katleen; Gisselsson, David; Øra, Ingrid; Speleman, Frank; Caron, Huib N.; Versteeg, Rogier

    2012-01-01

    The tightly controlled network of cell cycle genes consists of a core of cyclin dependent kinases (CDKs) that are activated by periodically expressed cyclins. The activity of the cyclin-CDK complexes is regulated by cyclin dependent kinase inhibitors (CDKIs) and multiple signal transduction routes

  14. The functions of the cdk-cyclin kinase inhibitor p21{sup WAF1}; Les fonctions de l'inhibiteur de cdk-cycline kinase, p21{sup WAF1}

    Energy Technology Data Exchange (ETDEWEB)

    Boulaire, J. [Institut de Biologie Structurale J.P. Ebel, 38 - Grenoble (France); Fotedar, A.; Fotedar, R. [Sidney Kimmel Cancer Center, San Diego (United States)

    2000-04-01

    p21{sup WAF1} plays a critical role in regulating cell growth and the cell response to DNA damage. The primary targets of p21{sup WAF1} (hereafter referred to as p21) are the cdk-cyclins which regulate the progression of cukayotic cells through the cell cycle, and proliferating cell nuclear antigen (PCNA), an accessory protein of DNA polymerase {delta}. p21 forms complexes with a class of cdk-cyclins to inhibit their kinase activity and with PCNA to inhibit synthesis. These distinct properties map to the N- terminal and the C- terminal regions of p21, respectively. Cell cycle arrest in G-1 (G-1 checkpoint) following DNA damage is mediated by p53 and is deficient in p21 null cells. p53 thus up-regulates p21 expression in response to DNA damage, which in turn inhibits cdk2-associated kinase activity. Retinoblastoma protein is regulated by cdk-cyclin kinases, and acts as a downstream target of p21 in DNA damage-induced G-1 arrest. Furthermore, accumulating evidence indicates that p21 may play a role in maintaining G-2 arrest after DNA damage. Transcriptional control of p21 by factors other than p53 is critical for growth arrest and for cell differentiation in many instances. (authors)

  15. Growth inhibition and cell cycle arrest in the G0/G1 by schizandrin, a dibenzocyclooctadiene lignan isolated from Schisandra chinensis, on T47D human breast cancer cells.

    Science.gov (United States)

    Kim, Sun-Jack; Min, Hye-Young; Lee, Eun Jin; Kim, Yeong Shik; Bae, KiHwan; Kang, Sam Sik; Lee, Sang Kook

    2010-02-01

    Schizandrin is one of the main dibenzocyclooctadiene lignans present in the fruit of Schisandra chinensis (Schisandraceae). Biological activities including hepatoprotective, antiviral and neuroprotective effects of schizandrin and other dibenzocyclooctadiene lignans have been reported previously. However, the antiproliferative effect of schizandrin against human cancer cells has been poorly determined to date. This study examined the antiproliferative effect of schizandrin in human breast cancer cells. Schizandrin exhibited growth inhibitory activities in cultured human breast cancer cells, and the effect was the more profound in estrogen receptor (ER)-positive T47D cells than in ER-negative MDA-MB-231 cells. When treated with the compound in T47D cells, schizandrin induced the accumulation of a cell population in the G0/G1 phase, which was further demonstrated by the induction of CDK inhibitors p21 and p27 and the inhibition of the expression of cell cycle checkpoint proteins including cyclin D1, cyclin A, CDK2 and CDK4. These results suggest that schizandrin inhibits cell proliferation through the induction of cell cycle arrest with modulating cell cycle-related proteins in human breast cancer cells. (c) 2009 John Wiley & Sons, Ltd.

  16. [Knockdown of DNA-PKcs inhibits cell cycle and its mechanism of drug-resistant Bel7402/5-Fu hepatocellular carcinoma cells].

    Science.gov (United States)

    Li, Dayu; Liu, Yun; Yu, Chunbo; Liu, Xiping; Fan, Fang

    2017-12-01

    Objective To study the effect of the knock-down of the DNA-dependent protein kinase catalytic subunit (DNA-PKcs) on the cell cycle of the multidrug-resistant (MDR) Bel7402/5-Fu hepatocellular carcinoma cells and its MDR mechanism. Methods After cationic liposome-mediated siDNA-PKcs oligonucleotide transfection, the drug sensitivity of Bel7402/5-Fu cells to 5-fluorouracil (5-Fu) and adriamycin (ADM) was determined by MTT assay; the cell cycle were detected by flow cytometry; meanwhile, the protein expressions of cell cycle-related proteins P21, cell cycle protein B1 (cyclin B1), cell cycle division protein 2 (CDC2) were tested by Western blotting; the expressions of ataxia telangiectasia mutated (ATM) and p53 at both mRNA and protein levels were detected by real-time PCR and Western blot analysis. Results The MTT results showed siDNA-PKcs increased the chemotherapeutic sensitivity of Bel7402/5-Fu cells to 5-Fu and ADM. The flow cytometric analysis showed siDNA-PKcs decreased the percentage of S-phase cells but increased the percentage of G2/M phase cells. Western blotting showed siDNA-PKcs increased the protein expression of P21 but decreased cyclinB1 and CDC2 proteins. In addition, siDNA-PKcs also increased the expressions of ATM and p53. Conclusion DNA-PKcs silencing increases P21 while decreases cyclin B1 and CDC2 expressions, and finally induces G2/M phase arrest in Bel7402/5-Fu cells, which may be related to ATM-p53 signaling pathway.

  17. Retinal degeneration depends on Bmi1 function and reactivation of cell cycle proteins.

    Science.gov (United States)

    Zencak, Dusan; Schouwey, Karine; Chen, Danian; Ekström, Per; Tanger, Ellen; Bremner, Rod; van Lohuizen, Maarten; Arsenijevic, Yvan

    2013-02-12

    The epigenetic regulator Bmi1 controls proliferation in many organs. Reexpression of cell cycle proteins such as cyclin-dependent kinases (CDKs) is a hallmark of neuronal apoptosis in neurodegenerative diseases. Here we address the potential role of Bmi1 as a key regulator of cell cycle proteins during neuronal apoptosis. We show that several cell cycle proteins are expressed in different models of retinal degeneration and required in the Rd1 photoreceptor death process. Deleting E2f1, a downstream target of CDKs, provided temporary protection in Rd1 mice. Most importantly, genetic ablation of Bmi1 provided extensive photoreceptor survival and improvement of retinal function in Rd1 mice, mediated by a decrease in cell cycle markers and regulators independent of p16(Ink4a) and p19(Arf). These data reveal that Bmi1 controls the cell cycle-related death process, highlighting this pathway as a promising therapeutic target for neuroprotection in retinal dystrophies.

  18. C-terminal sequences direct cyclin D1-CRM1 binding.

    Science.gov (United States)

    Benzeno, Sharon; Diehl, J Alan

    2004-12-31

    GSK-3beta-dependent phosphorylation of cyclin D1 at a conserved C-terminal residue, Thr-286, promotes CRM1-dependent cyclin D1 nuclear export. Herein, we have identified a short stretch of residues adjacent to Thr-286 that mediates CRM1 association and thus cyclin D1 nuclear export. We found that disruption of this hydrophobic patch, stretching from amino acids 290 to 295 within cyclin D1, results in constitutively nuclear cyclin D1-CDK4 complexes with an increased propensity to potentiate transformation of murine fibroblasts. Our data support a model wherein deregulation of cyclin D1 nuclear export might contribute to human neoplastic growth.

  19. MAP kinase dependent cyclinE/cdk2 activity promotes DNA replication in early sea urchin embryos

    OpenAIRE

    Kisielewska, J.; Philipova, R.; Huang, J.-Y.; Whitaker, M.

    2009-01-01

    Sea urchins provide an excellent model for studying cell cycle control mechanisms governing DNA replication in vivo. Fertilization and cell cycle progression are tightly coordinated by Ca2+ signals, but the mechanisms underlying the onset of DNA replication after fertilization remain less clear. In this study we demonstrate that calcium-dependent activation of ERK1 promotes accumulation of cyclinE/cdk2 into the male and female pronucleus and entry into first S-phase. We show that cdk2 activit...

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

    Directory of Open Access Journals (Sweden)

    Hyun-Ho Kwak

    2016-01-01

    Full Text Available 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.

  1. Amplification and protein overexpression of cyclin D1: Predictor of occult nodal metastasis in early oral cancer.

    Science.gov (United States)

    Noorlag, Rob; Boeve, Koos; Witjes, Max J H; Koole, Ronald; Peeters, Ton L M; Schuuring, Ed; Willems, Stefan M; van Es, Robert J J

    2017-02-01

    Accurate nodal staging is pivotal for treatment planning in early (stage I-II) oral cancer. Unfortunately, current imaging modalities lack sensitivity to detect occult nodal metastases. Chromosomal region 11q13, including genes CCND1, Fas-associated death domain (FADD), and CTTN, is often amplified in oral cancer with nodal metastases. However, evidence in predicting occult nodal metastases is limited. In 158 patients with early tongue and floor of mouth (FOM) squamous cell carcinomas, both CCND1 amplification and cyclin D1, FADD, and cortactin protein expression were correlated with occult nodal metastases. CCND1 amplification and cyclin D1 expression correlated with occult nodal metastases. Cyclin D1 expression was validated in an independent multicenter cohort, confirming the correlation with occult nodal metastases in early FOM cancers. Cyclin D1 is a predictive biomarker for occult nodal metastases in early FOM cancers. Prospective research on biopsy material should confirm these results before implementing its use in routine clinical practice. © 2016 Wiley Periodicals, Inc. Head Neck 39: 326-333, 2017. © 2016 Wiley Periodicals, Inc.

  2. Evolutionarily conserved transcription factor Apontic controls the G1/S progression by inducing cyclin e during eye development

    KAUST Repository

    Liu, Qingxin

    2014-06-16

    During Drosophila eye development, differentiation initiates in the posterior region of the eye disk and progresses anteriorly as a wave marked by the morphogenetic furrow (MF), which demarcates the boundary between anterior undifferentiated cells and posterior differentiated photoreceptors. However, the mechanism underlying the regulation of gene expression immediately before the onset of differentiation remains unclear. Here, we show that Apontic (Apt), which is an evolutionarily conserved transcription factor, is expressed in the differentiating cells posterior to the MF. Moreover, it directly induces the expression of cyclin E and is also required for the G1-to-S phase transition, which is known to be essential for the initiation of cell differentiation at the MF. These observations identify a pathway crucial for eye development, governed by a mechanism in which Cyclin E promotes the G1-to-S phase transition when regulated by Apt.

  3. Cocaine promotes primary human astrocyte proliferation via JNK-dependent up-regulation of cyclin A2.

    Science.gov (United States)

    Lee, Chun-Ting; Boeshore, Kristen L; Wu, Chun; Becker, Kevin G; Errico, Stacie L; Mash, Deborah C; Freed, William J

    2016-11-22

    Astrocytes perform a plethora of important functions in the central nervous system (CNS) and are involved in cocaine-evoked synaptic plasticity. Previously, we showed that while cocaine decreased cyclin A2 expression in primary human neural progenitor cells, it increased cyclin A2 expression in human astrocytes. Since cyclin A2 is an essential regulator of the cell cycle, the aim of the present study is to clarify the effect of cocaine on proliferation of human astrocytes and elucidate the underlying molecular mechanisms. Primary human astrocytes were treated with either 1, 10, or 100 μM cocaine for 48 hr, and cell proliferation was measured using the CyQUANT cell proliferation assay. To elucidate the molecular mechanisms through which cocaine affects the proliferation of astrocytes, we analyzed gene expression profiles in cocaine-treated primary human astrocytes using a human focused cDNA array. Gene ontology/pathway enrichment analysis, STRING protein-protein interaction analysis, RT-qPCR, and western blotting were used to identify signal transduction pathways that are involved in cocaine-induced astrocyte dysfunction. Cocaine at 10 and 100 μM significantly increased human astrocyte proliferation. Gene expression profiling revealed the JNK MAP kinase pathway as a driver of cell proliferation affected by cocaine in human astrocytes. Further experiments showed that cocaine-induced JNK activation induced up-regulation of cyclin A2, leading to enhanced proliferation of human astrocytes. Cocaine-induced abnormal increases in the number of astrocytes may cause disruption in neuron-glia signaling and contribute to synaptic impairment in the CNS. Understanding the mechanisms of cocaine's effects on human astrocytes may help to reveal the involvement of glial cells in addictive behaviors.

  4. The Effects of Deregulated Cyclin Expression in Mitosis. A Role in Breast Tumorigenesis

    National Research Council Canada - National Science Library

    Keck, Jamie M; Reed, Steve I

    2006-01-01

    The purpose of this project is to study the effects of constitutive cyclin E expression on mitotic division and to better understand the mechanisms through which cyclin E leads to chromosome instability...

  5. The Effects of Deregulated Cyclin E Expression in Mitosis: A Role in Breast Tumorigenesis

    National Research Council Canada - National Science Library

    Keck, Jamie M; Reed, Steven I

    2005-01-01

    The purpose of this project is to study the effect of constitutive cyclin E expression on mitotic division and to ultimately identify the mechanism through which cyclin E leads to chromosome instability...

  6. Effort of fast-neutron and X-ray on expression of p53 and CyclinB1 of small intestinal tissue in mice

    International Nuclear Information System (INIS)

    Xu Dongsheng; Liu Fenju; Sun Zhiqiang; Yang Xueqin; Chen Qiu; Zhou Weifang; Chen Ling

    2008-01-01

    In this paper, effect of fast-neutron and X-rays on the expression of P53 and CyclinB1 in small intestinal tissues of mice were investigated. The mice were randomly grouped and sacrificed 6 h after irradiation. Western blot assay was applied to detect the p53 and CyclinB 1 expressions, and flow cytometry was used to monitor changes in the small intestinal cell cycle. The results showed that the expression levels of p53 increased with irradiation dose. The expression levels of CyclinB 1 increased with the dose at first, but decreased when the dose exceeded 0.28 Gy. The results from the flow cytometry showed that the amount of cells in G2/M phase increased with the dose in both occasions. It was concluded that the low doses of fast-neutron and X-rays can make the expression levels of p53 and CyclinB 1 increase, but the expression levels of CyclinB 1 decreased and the G2 arrest occurred with increased doses. (authors)

  7. MicroRNA-184 promotes proliferation ability of glioma cells by regulating FOXO3.

    Science.gov (United States)

    Cui, Qing-Ke; Liu, Wei-Dong; Zhu, Jian-Xin; Wang, Yun-Hua; Wang, Zhi-Gang

    2014-10-01

    To investigate the effect of microRNA (miR-184) on regulating the genesis, development and proliferation of glioma cells. Lipidosome was used to transfect miR-184 mimic and inhibitor to glioma cell line, and the cell proliferation ability changes were determined by MTT and plate cloning experiment after the transfection. WB test was used to measure the levels of cyclinD1, p27 and FOXO3. Meanwhile, QPCR was used to detect miR-184 expression in glioma cell line, glioma tissues and adjacent tissues. Luciferase experiment was used to test 3'UTR gene targeting regulation of miR-184 and FOXO3. QPCR results showed a significant lower miR-184 expression level in glioma cell line and glioma tissues than that in juxtacancerous tissue. MTT and plate cloning experiments have shown that after over-expressing of miR-184, the cell proliferation capacity of glioma U87 and T98G was significantly increased, which was significantly inhibited after the inhibition of miR-184. WB results showed a lower expression level of p27 in U87 and T98G cells, and a higher expression level of cyclinD1 after over-expressing of miR-184 was observed. However, a lower expression level of cyclinD1 and a higher expression level of p27 after the inhibition of miR-184. The luciferase activity was inhibited after the over-expressing of miR-184. MiR-184 can affect the proliferation abilities of glioma cells and regulate the cell cycle related protein. It plays an important role in the occurrence and development of gliomas. Copyright © 2014 Hainan Medical College. Published by Elsevier B.V. All rights reserved.

  8. Cyclin D1 genotype and expression in sporadic hemangioblastomas.

    NARCIS (Netherlands)

    Gijtenbeek, J.M.M.; Sprenger, S.H.E.; Franke, B.; Wesseling, P.; Jeuken, J.W.M.

    2005-01-01

    Central nervous system (CNS) hemangioblastomas are highly-vascularized tumors occurring in sporadic form or as a manifestation of von Hippel-Lindau disease (VHL). The VHL protein (pVHL) regulates various target genes, one of which is the CCND1 gene, encoding cyclin D1, a protein that plays a

  9. Cyclin B2 and p53 control proper timing of centrosome separation

    NARCIS (Netherlands)

    Nam, H.J.; Deursen, J.M.A. van

    2014-01-01

    Cyclins B1 and B2 are frequently elevated in human cancers and are associated with tumour aggressiveness and poor clinical outcome; however, whether and how B-type cyclins drive tumorigenesis is unknown. Here we show that cyclin B1 and B2 transgenic mice are highly prone to tumours, including tumour

  10. Distinct and Overlapping Requirements for Cyclins A, B, and B3 in Drosophila Female Meiosis

    Science.gov (United States)

    Bourouh, Mohammed; Dhaliwal, Rajdeep; Rana, Ketki; Sinha, Sucheta; Guo, Zhihao; Swan, Andrew

    2016-01-01

    Meiosis, like mitosis, depends on the activity of the cyclin dependent kinase Cdk1 and its cyclin partners. Here, we examine the specific requirements for the three mitotic cyclins, A, B, and B3 in meiosis of Drosophila melanogaster. We find that all three cyclins contribute redundantly to nuclear envelope breakdown, though cyclin A appears to make the most important individual contribution. Cyclin A is also required for biorientation of homologs in meiosis I. Cyclin B3, as previously reported, is required for anaphase progression in meiosis I and in meiosis II. We find that it also plays a redundant role, with cyclin A, in preventing DNA replication during meiosis. Cyclin B is required for maintenance of the metaphase I arrest in mature oocytes, for spindle organization, and for timely progression through the second meiotic division. It is also essential for polar body formation at the completion of meiosis. With the exception of its redundant role in meiotic maturation, cyclin B appears to function independently of cyclins A and B3 through most of meiosis. We conclude that the three mitotic cyclin-Cdk complexes have distinct and overlapping functions in Drosophila female meiosis. PMID:27652889

  11. Gender-based differences and menstrual cycle-related changes in specific diseases: implications for pharmacotherapy.

    Science.gov (United States)

    Ensom, M H

    2000-05-01

    Pharmacists should be aware of gender-based differences and menstrual cycle-related changes in six diseases: asthma, arthritis, migraine, diabetes, depression, and epilepsy. In general, women report symptoms of physical illness at higher rates, visit physicians more frequently, and make greater use of other health care services than men. Whereas reasons for these gender differences are not fully clear, a combination of biologic, physiologic, social, behavioral, psychologic, and cultural factors most likely contributes. A significant percentage of women with asthma, arthritis, migraine, diabetes, depression, or epilepsy experience worsening of their disease premenstrually. The mechanism is unknown, but is speculated to be multifactorial because of many endogenous and exogenous modulators and mediators of each disease. As part of general therapy for cycle-related exacerbations of any one of these disorders, patients should be encouraged to use a menstrual calendar to track signs and symptoms for two to three cycles; if cyclic trends are identified, the women should anticipate exacerbations and avoid triggering factors. Cyclic modulation with pharmacotherapy may be attempted. If unsuccessful, a trial of medical ovulation suppression with a gonadotropin-releasing hormone (GnRH) analog may be warranted. If that is successful, continuous therapy with a GnRH analog and steroid add-back therapy or less expensive alternatives may be effective. If pharmacotherapy is impractical, hysterectomy and bilateral oophorectomy with estrogen replacement therapy is a last resort. Gender differences and menstrual cycle-related changes are important areas for clinical and mechanistic research.

  12. p53 Dimers associate with a head-to-tail response element to repress cyclin B transcription.

    Directory of Open Access Journals (Sweden)

    Robert Lipski

    Full Text Available DNA damage induced by the topoisomerase I inhibitor SN38 activates cell cycle checkpoints which promote cell cycle arrest. This arrest can be abrogated in p53-defective cells by the Chk1 inhibitor 7-hydroxystaurosporine (UCN-01. Previously, we compared p53 wild-type MCF10A cells with derivatives whose p53 function was inhibited by over-expression of the tetramerization domain (MCF10A/OD or expression of shRNA against p53 (MCF10A/Δp53. Treatment of SN38-arrested MCF10A/OD cells with UCN-01 abrogated S, but not G2 arrest, while the MCF10A/Δp53 cells abrogated both S and G2 arrest. The MCF10A/OD cells had reduced levels of cyclin B, suggesting that tetramerization of p53 is not required for repression of cyclin B gene expression. In the present study, we analyzed p53 oligomerization status using glutaraldehyde cross-linking. Following SN38 treatment, MCF10A cells contained oligomeric forms of p53 with molecular weights approximating monomers, dimers, trimers, and tetramers. However, MCF10A/OD cells possessed only monomers and dimers suggesting that these complexes may be involved in repression of cyclin B. While genes transcriptionally activated by p53 contain a consensus sequence with elements repeated in a head-to-head orientation, the cyclin B promoter contains similar elements oriented head-to-tail. Chromatin immunoprecipitation (ChIP assays revealed that p53 associates with this head-to-tail element in both MCF10A and MCF10A/OD. Electrophoretic mobility shift assays (EMSA using a biotin-labeled probe containing the head-to-tail element showed a shift in mobility consistent with the molecular weight of tetramers and dimers in MCF10A nuclear extract, but only the dimer in MCF10A/OD nuclear extract. Taken together, these results suggest a novel mechanism whereby p53 dimers associate with the head-to-tail element to repress cyclin B transcription.

  13. p53 Dimers associate with a head-to-tail response element to repress cyclin B transcription.

    Science.gov (United States)

    Lipski, Robert; Lippincott, Daniel J; Durden, Brittany C; Kaplan, Anne R; Keiser, Hilary E; Park, Jung-Ho; Levesque, Aime A

    2012-01-01

    DNA damage induced by the topoisomerase I inhibitor SN38 activates cell cycle checkpoints which promote cell cycle arrest. This arrest can be abrogated in p53-defective cells by the Chk1 inhibitor 7-hydroxystaurosporine (UCN-01). Previously, we compared p53 wild-type MCF10A cells with derivatives whose p53 function was inhibited by over-expression of the tetramerization domain (MCF10A/OD) or expression of shRNA against p53 (MCF10A/Δp53). Treatment of SN38-arrested MCF10A/OD cells with UCN-01 abrogated S, but not G2 arrest, while the MCF10A/Δp53 cells abrogated both S and G2 arrest. The MCF10A/OD cells had reduced levels of cyclin B, suggesting that tetramerization of p53 is not required for repression of cyclin B gene expression. In the present study, we analyzed p53 oligomerization status using glutaraldehyde cross-linking. Following SN38 treatment, MCF10A cells contained oligomeric forms of p53 with molecular weights approximating monomers, dimers, trimers, and tetramers. However, MCF10A/OD cells possessed only monomers and dimers suggesting that these complexes may be involved in repression of cyclin B. While genes transcriptionally activated by p53 contain a consensus sequence with elements repeated in a head-to-head orientation, the cyclin B promoter contains similar elements oriented head-to-tail. Chromatin immunoprecipitation (ChIP) assays revealed that p53 associates with this head-to-tail element in both MCF10A and MCF10A/OD. Electrophoretic mobility shift assays (EMSA) using a biotin-labeled probe containing the head-to-tail element showed a shift in mobility consistent with the molecular weight of tetramers and dimers in MCF10A nuclear extract, but only the dimer in MCF10A/OD nuclear extract. Taken together, these results suggest a novel mechanism whereby p53 dimers associate with the head-to-tail element to repress cyclin B transcription.

  14. Optimization of immunolocalization of cell cycle proteins in human corneal endothelial cells

    Science.gov (United States)

    He, Zhiguo; Campolmi, Nelly; Ha Thi, Binh-Minh; Dumollard, Jean-Marc; Peoc’h, Michel; Garraud, Olivier; Piselli, Simone; Gain, Philippe

    2011-01-01

    Purpose En face observation of corneal endothelial cells (ECs) using flat-mounted whole corneas is theoretically much more informative than observation of cross-sections that show only a few cells. Nevertheless, it is not widespread for immunolocalization (IL) of proteins, probably because the endothelium, a superficial monolayer, behaves neither like a tissue in immunohistochemistry (IHC) nor like a cell culture in immunocytochemistry (ICC). In our study we optimized IL for ECs of flat-mounted human corneas to study the expression of cell cycle-related proteins. Methods We systematically screened 15 fixation and five antigen retrieval (AR) methods on 118 human fresh or stored corneas (organ culture at 31 °C), followed by conventional immunofluorescence labeling. First, in an attempt to define a universal protocol, we selected combinations able to correctly localize four proteins that are perfectly defined in ECs (zonula occludens-1 [ZO-1] and actin) or ubiquitous (heterogeneous nuclear ribonucleoprotein L [hnRNP L] and histone H3). Second, we screened protocols adapted to the revelation of 9 cell cycle proteins: Ki67, proliferating cell nuclear antigen (PCNA), minichromosome maintenance protein 2 (MCM2), cyclin D1, cyclin E, cyclin A, p16Ink4a, p21Cip1 and p27Kip1. Primary antibody controls (positive controls) were performed on both epithelial cells of the same, simultaneously-stained whole corneas, and by ICC on human ECs in in vitro non-confluent cultures. Both controls are known to contain proliferating cells. IL efficiency was evaluated by two observers in a masked fashion. Correct localization at optical microscopy level in ECs was define as clear labeling with no background, homogeneous staining, agreement with previous works on ECs and/or protein functions, as well as a meaningful IL in proliferating cells of both controls. Results The common fixation with 4% formaldehyde (gold standard for IHC) failed to reveal 12 of the 13 proteins. In contrast, they

  15. Huaier Aqueous Extract Induces Hepatocellular Carcinoma Cells Arrest in S Phase via JNK Signaling Pathway

    Directory of Open Access Journals (Sweden)

    Chengshuo Zhang

    2015-01-01

    Full Text Available Huaier aqueous extract, the main active constituent of Huaier proteoglycan, has antihepatocarcinoma activity in experimental and clinical settings. However, the potential and associated antihepatoma mechanisms of Huaier extract are not yet fully understood. Therefore, in this study, we aimed to elucidate the inhibitory proliferation effect of Huaier extract on apoptosis and cycle of HepG2 and Bel-7402 cells. Our data demonstrated that incubation with Huaier extract resulted in a marked decrease in cell viability dose-dependently. Flow cytometric analysis showed that a 48 h treatment of Huaier extract caused cell apoptosis. Typical apoptotic nucleus alterations were observed with fluorescence microscope after Hoechst staining. Immunoblot analysis further demonstrated that Huaier extract activated caspase 3 and PARP. Additionally, Huaier extract inhibited the activity of p-ERK, p-p38, and p-JNK in terms of MAPK. Furthermore, Huaier extract induced HCC cells arrest in S phase and decreased the cycle related protein expression of β-catenin and cyclin D1. Studies with JNK specific inhibitor, SP600125, showed that Huaier extract induced S phase arrest and decreased β-catenin and cyclin D1 expression via JNK signaling pathway. In conclusion, we verify that Huaier extract causes cell apoptosis and induces hepatocellular carcinoma cells arrest in S phase via JNK pathway, which advances our understanding on the molecular mechanisms of Huaier extract in hepatocarcinoma management.

  16. A cyclin-dependent kinase inhibitor, dinaciclib in preclinical treatment models of thyroid cancer.

    Directory of Open Access Journals (Sweden)

    Shu-Fu Lin

    Full Text Available We explored the therapeutic effects of dinaciclib, a cyclin-dependent kinase (CDK inhibitor, in the treatment of thyroid cancer.Seven cell lines originating from three pathologic types of thyroid cancer (papillary, follicular and anaplastic were studied. The cytotoxicity of dinaciclib was measured using a lactate dehydrogenase assay. The expression of proteins associated with cell cycle and apoptosis was assessed using Western blot analysis and immunofluorescence microscopy. Cell cycle distribution was measured by flow cytometry and immunofluorescence microscopy. Apoptosis and caspase-3 activity were measured by flow cytometry and fluorometric assay. Mice bearing flank anaplastic thyroid cancer (ATC were treated with intraperitoneal injections of dinaciclib.Dinaciclib inhibited thyroid cancer cell proliferation in a dose-dependent manner. Dinaciclib had a low median-effect dose (≤ 16.0 nM to inhibit cell proliferation in seven thyroid cancer cell lines. Dinaciclib decreased CDK1, cyclin B1, and Aurora A expression, induced cell cycle arrest in the G2/M phase, and induced accumulation of prophase mitotic cells. Dinaciclib decreased Mcl-1, Bcl-xL and survivin expression, activated caspase-3 and induced apoptosis. In vivo, the growth of ATC xenograft tumors was retarded in a dose-dependent fashion with daily dinaciclib treatment. Higher-dose dinaciclib (50 mg/kg caused slight, but significant weight loss, which was absent with lower-dose dinaciclib (40 mg/kg treatment.Dinaciclib inhibited thyroid cancer proliferation both in vitro and in vivo. These findings support dinaciclib as a potential drug for further studies in clinical trials for the treatment of patients with refractory thyroid cancer.

  17. Cyclin-like F-box protein plays a role in growth and development of the three model species Medicago truncatula, Lotus japonicus, and Arabidopsis thaliana

    Directory of Open Access Journals (Sweden)

    Boycheva I

    2015-08-01

    Full Text Available Irina Boycheva,1 Valya Vassileva,2 Miglena Revalska,1 Grigor Zehirov,2 Anelia Iantcheva1 1Department of Functional Genetics Legumes, 2AgroBioInstitute, Department of Plant Stress Molecular Biology, Institute of Plant Physiology and Genetics, Sofia, Bulgaria Abstract: In eukaryotes, F-box proteins are one of the main components of the SCF complex that belongs to the family of ubiquitin E3 ligases, which catalyze protein ubiquitination and maintain the balance between protein synthesis and degradation. In the present study, we clarified the role and function of the gene encoding cyclin-like F-box protein from Medicago truncatula using transgenic plants of the model species M. truncatula, Lotus japonicas, and Arabidopsis thaliana generated by Agrobacterium-mediated transformation. Morphological and transcriptional analyses combined with flow cytometry and histochemistry demonstrated the participation of this protein in many aspects of plant growth and development, including processes of indirect somatic embryogenesis and symbiotic nodulation. The cyclin-like F-box gene showed expression in all plant organs and tissues comprised of actively dividing cells. The observed variations in root and hypocotyl growth, leaf and silique development, ploidy levels, and leaf parameters in the obtained transgenic lines demonstrated the effects of this gene on organ development. Furthermore, knockdown of cyclin-like F-box led to accumulation of higher levels of the G2/M transition-specific gene cyclin B1:1 (CYCB1:1, suggesting its possible role in cell cycle control. Together, the collected data suggest a similar role of the cyclin-like F-box protein in the three model species, providing evidence for the functional conservation of the studied gene. Keywords: cyclin-like F-box, model legumes, Arabidopsis thaliana, plant growth, plant development, cell cycle

  18. MAP kinase dependent cyclinE/cdk2 activity promotes DNA replication in early sea urchin embryos.

    Science.gov (United States)

    Kisielewska, J; Philipova, R; Huang, J-Y; Whitaker, M

    2009-10-15

    Sea urchins provide an excellent model for studying cell cycle control mechanisms governing DNA replication in vivo. Fertilization and cell cycle progression are tightly coordinated by Ca(2+) signals, but the mechanisms underlying the onset of DNA replication after fertilization remain less clear. In this study we demonstrate that calcium-dependent activation of ERK1 promotes accumulation of cyclinE/cdk2 into the male and female pronucleus and entry into first S-phase. We show that cdk2 activity rises quickly after fertilization to a maximum at 4 min, corresponding in timing to the early ERK1 activity peak. Abolishing MAP kinase activity after fertilization with MEK inhibitor, U0126, substantially reduces the early peak of cdk2 activity and prevents cyclinE and cdk2 accumulation in both sperm pronucleus and zygote nucleus in vivo. Both p27(kip1) and roscovitine, cdk2 inhibitors, prevented DNA replication suggesting cdk2 involvement in this process in sea urchin. Inhibition of cdk2 activity using p27(kip1) had no effect on the phosphorylation of MBP by ERK, but completely abolished phosphorylation of retinoblastoma protein, a cdk2 substrate, indicating that cdk2 activity is downstream of ERK1 activation. This pattern of regulation of DNA synthesis conforms to the pattern observed in mammalian somatic cells.

  19. Autonomous Metabolic Oscillations Robustly Gate the Early and Late Cell Cycle

    NARCIS (Netherlands)

    Papagiannakis, Alexandros; Niebel, Bastian; Wit, Ernst C; Heinemann, Matthias

    2017-01-01

    Eukaryotic cell division is known to be controlled by the cyclin/cyclin dependent kinase (CDK) machinery. However, eukaryotes have evolved prior to CDKs, and cells can divide in the absence of major cyclin/CDK components. We hypothesized that an autonomous metabolic oscillator provides dynamic

  20. β-catenin/cyclin D1 mediated development of suture mesenchyme in calvarial morphogenesis

    Directory of Open Access Journals (Sweden)

    Yu Hsiao-Man

    2010-11-01

    Full Text Available Abstract Background Mouse genetic study has demonstrated that Axin2 is essential for calvarial development and disease. Haploid deficiency of β-catenin alleviates the calvarial phenotype caused by Axin2 deficiency. This loss-of-function study provides evidence for the requirement of β-catenin in exerting the downstream effects of Axin2. Results Here we utilize a gain-of-function analysis to further assess the role of β-catenin. A transgenic expression system permitting conditional activation of β-catenin in a spatiotemporal specific manner has been developed. Aberrant stimulation of β-catenin leads to increases in expansion of skeletogenic precursors and the enhancement of bone ossification reminiscent to the loss of Axin2. The constitutively active signal promotes specification of osteoprogenitors, but prevents their maturation into terminally differentiated osteoblasts, along the osteoblast lineage. However, the prevention does not interfere with bone synthesis, suggesting that mineralization occurs without the presence of mature osteoblasts. β-catenin signaling apparently plays a key role in suture development through modulation of calvarial morphogenetic signaling pathways. Furthermore, genetic inactivation of the β-catenin transcriptional target, cyclin D1, impairs expansion of the skeletogenic precursors contributing to deficiencies in calvarial ossification. There is a specific requirement for cyclin D1 in populating osteoprogenitor cell types at various developmental stages. Conclusion These findings advance our knowledge base of Wnt signaling in calvarial morphogenesis, suggesting a key regulatory pathway of Axin2/β-catenin/cyclin D1 in development of the suture mesenchyme.

  1. PENGARUH EKSTRAK ETHANOL PROPOLIS TERHADAP EKSPRESI PROTEIN Bcl2, CYCLIN D1 DAN INDUKSI APOPTOSIS PADA KULTUR SEL KANKER KOLON

    Directory of Open Access Journals (Sweden)

    Haryono Yuniarto

    2017-06-01

    Full Text Available Kanker kolorektal menempati urutan kejadian kanker ketiga di seluruh dunia, dengan lebih dari 1 juta angka kejadian tiap tahunnya. Berbagai strategi terapi pengobatan kanker kolorektal tetapi relatif belum optimal. Oleh karena itu, terdapat kebutuhan mengembangkan terapi alternatif sebagai pendamping. Propolis menunjukkan aktivitas proapoptosis pada berbagai jenis sel kanker. Mengetahui pengaruh pemberian propolis yang berasal dari Kerjo, Karanganyar, Indonesia terhadap induksi proses apoptosis dan aktivitas antiproliferasi, terutama terkait dengan penekanan ekspresi protein Bcl 2 dan cyclin D1 pada kultur sel WiDr (cell line kanker kolon. Penelitian eksperimental laboratorik menggunakan post test with control group design. Penelitian dilakukan pada kultur sel WiDr (sel kanker kolon dengan pemberian propolis. Pengamatan ekspresi protein Cyclin D1 dan Bcl2 dilakukan dengan metode imunositokimia, sedangkan pengamatan induksi apoptosis dilakukan dengan flowcytometry. Analisis statistik dengan uji Kruskal-Wallis, signifikan bila p <0,05. Rata-rata ekspresi Bcl2 pada kelima kelompok yaitu kontrol 83.40 ± 0.69 μg/ml, EEP 1/2 IC50 60.63 ± 0.40, EEP IC50 33.77 ± 1.08 μg/ml, EEP 2 IC50 24.28 ± 1.91 μg/ml, 5fluorouracil 12.74 ± 2.19 μg/ml. Terdapat perbedaan bermakna ekspresi Bcl2 antara kelompok uji dibandingkan kelompok kontrol (p < 0,001. Rata-rata ekspresi cyclin D1 pada kelima kelompok yaitu kontrol 83.77 ± 0.39 μg/ml, EEP 1/2 IC50 61.44 ± 0.41, EEP IC50 36.67 ± 1.18 μg/ml, EEP 2 IC50 24.50 ± 0.38 μg/ml, 5fluorouracil 13.42 ± 1.04μg/ml. Terdapat perbedaan bermakna ekspresi cyclin D1 antara kelompok uji dibandingkan kelompok kontrol (p < 0,001. Pemberian ekstrak etanol propolis mempunyai pengaruh menekan ekspresi Bcl2, cyclin D1, dan menginduksi apoptosis pada kultur sel kanker kolon (WiDr Cell Line.   Kata Kunci: Ekstrak Ethanol Propolis, Bcl2, cyclin D1, Sel WiDr

  2. Changes of protein kinase Calpha and cyclin D1 expressions in pulmonary arteries from smokers with and without chronic obstructive pulmonary disease.

    Science.gov (United States)

    Xaing, Min; Liu, Xiansheng; Zeng, Daxiong; Wang, Ran; Xu, Yongjian

    2010-04-01

    The purpose of this study was to investigate the changes of protein kinase Calpha (PKCalpha) and cyclin D1 expressions in pulmonary arteries from smokers with normal lung function and smokers with mild to moderate chronic obstructive pulmonary disease (COPD). The peripheral lung tissues were obtained from 10 non-smokers with normal lung function (non-smoker group), 14 smokers with normal lung function (smoker group), 11 smokers with mild to moderate COPD (COPD group). The morphological changes of pulmonary arteries were observed by HE-staining. The expressions of alpha-smooth muscle actin (alpha-SMA), proliferating cell nuclear antigen (PCNA), PKCalpha and cyclin D1 proteins in pulmonary artery smooth muscle cells (PASMCs) were immunohistochemically determined. The percentages of PCNA-positive cells were taken as the smooth muscle cells proliferation index (PI). The mRNA expressions of PKCalpha and cyclin D1 in PASMCs were evaluated by real-time fluorescence PCR. Morphometrical analysis showed that the ratio of pulmonary artery wall area to total area (WA%) in smoker group and COPD group was significantly greater than that in non-smoker group (P<0.01). The PASMCs proliferation index in smoker group and COPD group was significantly higher than that in nonsmoker group (P<0.01). The protein levels of PKCalpha and cyclin D1 in PASMCs were significantly increased in smoker group and COPD group as compared with non-smoker group (P<0.01). The mRNA expressions of PKCalpha and cyclin D1 in PASMCs were significantly elevated in smoker group and COPD group as compared with non-smoker group (P<0.01). Significant correlations were found between PKCalpha protein and WA% or PI (P<0.01). Correlations between cyclin D1 protein and WA% or PI also existed (P<0.01). The expression of PKCalpha was positively correlated with the expression of cyclin D1 at both protein and mRNA levels (P<0.01). In conclusion, increased expressions of PKCalpha and cyclin D1 might be involved in the

  3. Papillary microcarcinoma of the thyroid gland: is the immunohistochemical expression of cyclin D1 or galectin-3 in primary tumour an indicator of metastatic disease?

    DEFF Research Database (Denmark)

    Londero, Stefano Christian; Godballe, Christian; Krogdahl, Annelise

    2007-01-01

    an indolent course with an excellent prognosis. However, a more aggressive behaviour with regional and distant metastases does occur. The aim of this study was to evaluate if the immunohistochemical markers cyclin D1 or galectin-3 might indicate the presence of metastatic disease in patients with PMC...... verified regional or distant metastases. Slides were cut from the primary tumour and immunostaining and quantification was subsequently performed. RESULTS: The percentage of positive cells was examined for patients with and without metastases. For cyclin D1 the median values were 31% (range: 0-59) and 21......% (range: 0-75), respectively, showing a statistically significant difference (p=0.02). For galectin-3 the medians were 87% (range: 6-96) and 85% (range: 0-99) and no significant difference was found. CONCLUSION: Cyclin D1 showed significantly higher median expression in patients with metastases compared...

  4. Cyclin D1 is not an immediate target of beta-catenin following Apc loss in the intestine.

    NARCIS (Netherlands)

    Sansom, O.J.; Reed, K.R.; Wetering, M.van de; Muncan, V.; Winston, D.; Clevers, J.C.; Clarke, A.R.

    2005-01-01

    Cyclin D1 is postulated to be a target of the canonical Wnt pathway and critical for intestinal adenoma development. We show here that, unlike cyclin D1 reporter assays, endogenous cyclin D1 levels are not affected following antagonism of the Wnt pathway in vitro, nor is cyclin D1immediately

  5. Role of immunoexpression of cyclin D1, D3, retinoblastoma (Rb mutant and clinical risk factors on complete mole as risk factors of persistent mole

    Directory of Open Access Journals (Sweden)

    Yudi M Hidayat

    2015-10-01

    Full Text Available Introduction: Changes in complete hydatidiform mole (CHM that become persistent are difficult to handle because the malignant pathogenesis of CHM is still unclear. The growth of abnormal cells in CHM is thought to be caused by cell cycle abnormalities. Some components that play a role in this phase include cyclin D and retinoblastoma (Rb. The aim of our study was to determine the role of clinical risk factors, as well as cyclin D1, cyclin D3 and Rb-protein, in the occurrence of persistent moles. Materials and Method: This study involves 68 CHM cases at Dr. Hasan Sadikin Hospital from 2007–2011. The protein expression of cyclin D1, cyclin D3, and Rb were determined by immunohistochemistry. The results were analyzed by comparing the two groups of CHM that became persistent to those that returned to normal, as determined by a Mochizuki regression curve assessment. Results: 20 cases (29% of CHM became persistent and that 48 cases (71% returned to normal. Significant clinical variables were age (p 0.05. Conclusion: There is a strong relationship between clinical risk factors of age, excessive proliferation histopathology, serum βhCG levels ≥100,000 mU/mL, cyclin D1 and Rb mutations with the incidence of persistent moles after the evacuation of the CHM. We proposed a model to predict the risks of persistent moles with a cut-off point of 2.384, which can be used as a reference for patients with CHM.

  6. Ambivalent sexism, attitudes towards menstruation and menstrual cycle-related symptoms.

    Science.gov (United States)

    Marván, Ma Luisa; Vázquez-Toboada, Rocío; Chrisler, Joan C

    2014-08-01

    The objective of the present study was to investigate the relationship between ambivalent sexism and beliefs and attitudes towards menstruation, and, in turn, to study the influence of these variables on menstrual cycle-related symptoms. One hundred and six Mexican women completed the Ambivalent Sexism Inventory, the Beliefs about and Attitudes toward Menstruation Questionnaire and the Menstrual Distress Questionnaire. The higher scores on benevolent sexism were associated with the most positive attitudes towards menstruation and also with the belief that a menstruating woman should or should not do some activities and that menstruation keeps women from their daily activities. The higher scores on hostile sexism were associated with rejection of menstruation as well as with feelings of embarrassment about it. Beliefs about and attitudes towards menstruation predicted menstrual cycle-related symptoms related to negative affect, impaired concentration and behavioural changes, but did not predict somatic symptoms. These results will be useful to health professionals and advocates who want to change the negative expectations and stereotypes of premenstrual and menstrual women and reduce the sexism and negative attitudes towards women that are evident in Mexican culture. © 2013 International Union of Psychological Science.

  7. GATA4 is a direct transcriptional activator of cyclin D2 and Cdk4 and is required for cardiomyocyte proliferation in anterior heart field-derived myocardium.

    Science.gov (United States)

    Rojas, Anabel; Kong, Sek Won; Agarwal, Pooja; Gilliss, Brian; Pu, William T; Black, Brian L

    2008-09-01

    The anterior heart field (AHF) comprises a population of mesodermal progenitor cells that are added to the nascent linear heart to give rise to the majority of the right ventricle, interventricular septum, and outflow tract in mammals and birds. The zinc finger transcription factor GATA4 functions as an integral member of the cardiac transcription factor network in the derivatives of the AHF. In addition to its role in cardiac differentiation, GATA4 is also required for cardiomyocyte replication, although the transcriptional targets of GATA4 required for proliferation have not been previously identified. In the present study, we disrupted Gata4 function exclusively in the AHF and its derivatives. Gata4 AHF knockout mice die by embryonic day 13.5 and exhibit hypoplasia of the right ventricular myocardium and interventricular septum and display profound ventricular septal defects. Loss of Gata4 function in the AHF results in decreased myocyte proliferation in the right ventricle, and we identified numerous cell cycle genes that are dependent on Gata4 by microarray analysis. We show that GATA4 is required for cyclin D2, cyclin A2, and Cdk4 expression in the right ventricle and that the Cyclin D2 and Cdk4 promoters are bound and activated by GATA4 via multiple consensus GATA binding sites in each gene's proximal promoter. These findings establish Cyclin D2 and Cdk4 as direct transcriptional targets of GATA4 and support a model in which GATA4 controls cardiomyocyte proliferation by coordinately regulating numerous cell cycle genes.

  8. Meiotic Progression in Arabidopsis Is Governed by Complex Regulatory Interactions between SMG7, TDM1, and the Meiosis I–Specific Cyclin TAM[W][OA

    Science.gov (United States)

    Bulankova, Petra; Riehs-Kearnan, Nina; Nowack, Moritz K.; Schnittger, Arp; Riha, Karel

    2010-01-01

    Meiosis is a modified cell division that produces four haploid nuclei from a single diploid cell in two rounds of chromosome segregation. Here, we analyze the role of Arabidopsis thaliana SUPPRESSOR WITH MORPHOGENETIC EFFECTS ON GENITALIA7 (SMG7), THREE DIVISION MUTANT1 (TDM1), and TARDY ASYNCHRONOUS MEIOSIS (TAM) in meiotic progression. SMG7 is a conserved nonsense-mediated mRNA decay factor that is also, in Arabidopsis, essential for completion of meiosis. Examination of activating CYCLIN DEPENDENT KINASE A;1 phosophorylation at Thr-161 suggests that the meiotic arrest observed in smg7 mutants is likely caused by a failure to downregulate cyclin-dependent kinase (CDK) activity at the end of the second meiotic division. Genetic analysis indicates that SMG7 and TDM1 act in the same pathway to facilitate exit from meiosis. We further demonstrate that the cyclin TAM is specifically expressed in meiosis I and has both stimulatory and inhibitory effects on progression to meiosis II. TAM knockouts skip the second meiotic division producing unreduced gametes, but inactivation of SMG7 or TDM1 alleviates TAM’s requirement for entry into meiosis II. We propose a model that meiotic progression in Arabidopsis pollen mother cells is driven by a yet to be identified cyclin-CDK activity that is modulated by regulatory interactions between TDM1, SMG7, and TAM. PMID:21119056

  9. Cell cycle entry in C. elegans development

    NARCIS (Netherlands)

    Korzelius, J.P.

    2010-01-01

    Cell division is controlled by a mechanism that uses Cyclins, in association with their Cyclin-dependent kinase partners (Cdk’s), to regulate the transitions in the cell cycle.Studies in mammalian cell culture and single cell eukaryotes such as budding and fission yeast have uncovered much about how

  10. Virtual Screening for Potential Allosteric Inhibitors of Cyclin-Dependent Kinase 2 from Traditional Chinese Medicine

    Directory of Open Access Journals (Sweden)

    Fang Lu

    2016-09-01

    Full Text Available Cyclin-dependent kinase 2 (CDK2, a member of Cyclin-dependent kinases (CDKs, plays an important role in cell division and DNA replication. It is regarded as a desired target to treat cancer and tumor by interrupting aberrant cell proliferation. Compared to lower subtype selectivity of CDK2 ATP-competitive inhibitors, CDK2 allosteric inhibitor with higher subtype selectivity has been used to treat CDK2-related diseases. Recently, the first crystal structure of CDK2 with allosteric inhibitor has been reported, which provides new opportunities to design pure allosteric inhibitors of CDK2. The binding site of the ATP-competition inhibitors and the allosteric inhibitors are partially overlapped in space position, so the same compound might interact with the two binding sites. Thus a novel screening strategy was essential for the discovery of pure CDK2 allosteric inhibitors. In this study, pharmacophore and molecular docking were used to screen potential CDK2 allosteric inhibitors and ATP-competition inhibitors from Traditional Chinese Medicine (TCM. In the docking result of the allosteric site, the compounds which can act with the CDK2 ATP site were discarded, and the remaining compounds were regarded as the potential pure allosteric inhibitors. Among the results, prostaglandin E1 and nordihydroguaiaretic acid (NDGA were available and their growth inhibitory effect on human HepG2 cell lines was determined by MTT assay. The two compounds could substantially inhibit the growth of HepG2 cell lines with an estimated IC50 of 41.223 μmol/L and 45.646 μmol/L. This study provides virtual screening strategy of allosteric compounds and a reliable method to discover potential pure CDK2 allosteric inhibitors from TCM. Prostaglandin E1 and NDGA could be regarded as promising candidates for CDK2 allosteric inhibitors.

  11. Phosphorylation of HOX11/TLX1 on Threonine-247 during mitosis modulates expression of cyclin B1

    Directory of Open Access Journals (Sweden)

    Chesney Alden

    2010-09-01

    Full Text Available Abstract Background The HOX11/TLX1 (hereafter referred to as HOX11 homeobox gene was originally identified at a t(10;14(q24;q11 translocation breakpoint, a chromosomal abnormality observed in 5-7% of T cell acute lymphoblastic leukemias (T-ALLs. We previously reported a predisposition to aberrant spindle assembly checkpoint arrest and heightened incidences of chromosome missegregation in HOX11-overexpressing B lymphocytes following exposure to spindle poisons. The purpose of the current study was to evaluate cell cycle specific expression of HOX11. Results Cell cycle specific expression studies revealed a phosphorylated form of HOX11 detectable only in the mitotic fraction of cells after treatment with inhibitors to arrest cells at different stages of the cell cycle. Mutational analyses revealed phosphorylation on threonine-247 (Thr247, a conserved amino acid that defines the HOX11 gene family and is integral for the association with DNA binding elements. The effect of HOX11 phosphorylation on its ability to modulate expression of the downstream target, cyclin B1, was tested. A HOX11 mutant in which Thr247 was substituted with glutamic acid (HOX11 T247E, thereby mimicking a constitutively phosphorylated HOX11 isoform, was unable to bind the cyclin B1 promoter or enhance levels of the cyclin B1 protein. Expression of the wildtype HOX11 was associated with accelerated progression through the G2/M phase of the cell cycle, impaired synchronization in prometaphase and reduced apoptosis whereas expression of the HOX11 T247E mutant restored cell cycle kinetics, the spindle checkpoint and apoptosis. Conclusions Our results demonstrate that the transcriptional activity of HOX11 is regulated by phosphorylation of Thr247 in a cell cycle-specific manner and that this phosphorylation modulates the expression of the target gene, cyclin B1. Since it is likely that Thr247 phosphorylation regulates DNA binding activity to multiple HOX11 target sequences, it is

  12. Cyclin D1 and mammary carcinoma: new insights from transgenic mouse models

    International Nuclear Information System (INIS)

    Sutherland, Robert L; Musgrove, Elizabeth A

    2002-01-01

    Cyclin D1 is one of the most commonly overexpressed oncogenes in breast cancer, with 45–50% of primary ductal carcinomas overexpressing this oncoprotein. Targeted deletion of the gene encoding cyclin D1 demonstrates an essential role in normal mammary gland development while transgenic studies provide evidence that cyclin D1 is a weak oncogene in mammary epithelium. In a recent exciting development, Yu et al. demonstrate that cyclin D1-deficient mice are resistant to mammary carcinomas induced by c-neu and v-Ha-ras, but not those induced by c-myc or Wnt-1. These findings define a pivotal role for cyclin D1 in a subset of mammary cancers in mice and imply a functional role for cyclin D1 overexpression in human breast cancer

  13. Cell cycle markers have different expression and localization patterns in neuron-like PC12 cells and primary hippocampal neurons.

    Science.gov (United States)

    Negis, Yesim; Unal, Aysegul Yildiz; Korulu, Sirin; Karabay, Arzu

    2011-06-01

    Neuron-like PC12 cells are extensively used in place of neurons in published studies. Aim of this paper has been to compare mRNA and protein expressions of cell cycle markers; cyclinA, B, D, E; Cdk1, 2 and 4; and p27 in post-mitotic primary hippocampal neurons, mitotically active PC12 cells and NGF-differentiated post-mitotic PC12 cells. Contrary to PC12 cells, in neurons, the presence of all these markers was detected only at mRNA level; except for cyclinA, cyclinE and Cdk4, which were detectable also at protein levels. In both NGF-treated PC12 cells and neurons, cyclinE was localized only in the nucleus. In NGF-treated PC12 cells cyclinD and Cdk4 were localized in the nucleus while, in neurons cyclinD expression was not detectable; Cdk4 was localized in the cytoplasm. In neurons, cyclinA was nuclear, whereas in NGF-treated PC12 cells, it was localized in the cell body and along the processes. These results suggest that PC12 cells and primary neurons are different in terms of cell cycle protein expressions and localizations. Thus, it may not be very appropriate to use these cells as neuronal model system in order to understand neuronal physiological activities, upstream of where may lie cell cycle activation triggered events. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

  14. Sticky siRNAs targeting survivin and cyclin B1 exert an antitumoral effect on melanoma subcutaneous xenografts and lung metastases

    International Nuclear Information System (INIS)

    Kedinger, Valerie; Erbacher, Patrick; Bolcato-Bellemin, Anne-Laure; Meulle, Aline; Zounib, Omar; Bonnet, Marie-Elise; Gossart, Jean-Baptiste; Benoit, Elodie; Messmer, Melanie; Shankaranarayanan, Pattabhiraman; Behr, Jean-Paul

    2013-01-01

    Melanoma represents one of the most aggressive and therapeutically challenging malignancies as it often gives rise to metastases and develops resistance to classical chemotherapeutic agents. Although diverse therapies have been generated, no major improvement of the patient prognosis has been noticed. One promising alternative to the conventional therapeutic approaches currently available is the inactivation of proteins essential for survival and/or progression of melanomas by means of RNA interference. Survivin and cyclin B1, both involved in cell survival and proliferation and frequently deregulated in human cancers, are good candidate target genes for siRNA mediated therapeutics. We used our newly developed sticky siRNA-based technology delivered with linear polyethyleneimine (PEI) to inhibit the expression of survivin and cyclin B1 both in vitro and in vivo, and addressed the effect of this inhibition on B16-F10 murine melanoma tumor development. We confirm that survivin and cyclin B1 downregulation through a RNA interference mechanism induces a blockage of the cell cycle as well as impaired proliferation of B16-F10 cells in vitro. Most importantly, PEI-mediated systemic delivery of sticky siRNAs against survivin and cyclin B1 efficiently blocks growth of established subcutaneaous B16-F10 tumors as well as formation and dissemination of melanoma lung metastases. In addition, we highlight that inhibition of survivin expression increases the effect of doxorubicin on lung B16-F10 metastasis growth inhibition. PEI-mediated delivery of sticky siRNAs targeting genes involved in tumor progression such as survivin and cyclin B1, either alone or in combination with chemotherapeutic drugs, represents a promising strategy for melanoma treatment

  15. Cdk1/cyclin B-mediated phosphorylation stabilizes SREBP1 during mitosis.

    Science.gov (United States)

    Bengoechea-Alonso, Maria T; Ericsson, Johan

    2006-08-01

    Members of the sterol regulatory element-binding protein (SREBP) family of transcription factors control the biosynthesis of cholesterol and other lipids, and lipid synthesis is critical for cell growth and proliferation. We recently found that the mature forms of SREBP1a and SREBP1c are hyperphosphorylated in mitotic cells, giving rise to a phosphoepitope recognized by the mitotic protein monoclonal-2 (MPM-2) antibody. In addition, we found that mature SREBP1 was stabilized in a phosphorylation-dependent manner during mitosis. We have now mapped the major MPM-2 epitope to a serine residue, S439, in the C terminus of mature SREBP1. Using phosphorylation-specific antibodies, we demonstrate that endogenous SREBP1 is phosphorylated on S439 specifically during mitosis. Mature SREBP1 interacts with the Cdk1/cyclin B complex in mitotic cells and we demonstrate that Cdk1 phosphorylates S439, both in vitro and in vivo. Our results suggest that Cdk1-mediated phosphorylation of S439 stabilizes mature SREBP1 during mitosis, thereby preserving a critical pool of active transcription factors to support lipid synthesis. Taken together with our previous work, the current study suggests that SREBP1 may provide a link between lipid synthesis, proliferation and cell growth. This hypothesis was supported by our observation that siRNA-mediated inactivation of SREBP1 arrested cells in the G(1) phase of the cell cycle, thereby attenuating cell growth.

  16. Cyclin D expression in plutonium-induced lung tumors in F344 rats

    Energy Technology Data Exchange (ETDEWEB)

    Hahn, F.F.; Kelly, G. [SouthWest Scientific Resources, Inc., Albuquerque, NM (United States)

    1995-12-01

    The genetic mechanisms responsible for {alpha}-radiation-induced lung cancer in rats following inhalation of {sup 239}Pu is an ongoing area of research in our laboratory. Previous studies have examined the status of the p53 gene by immunohistochemistry. Only two tumors (2/26 squamous cell carcinomas) exhibited detectable levels of p53 products. Both were the result of mutations in codons 280 and 283. More recent studies of X-ray-induced lung tumors in rats showed a similar lack of involvement of p53. In conclusion, we found that {alpha}-radiation-induced rat lung tumors have a high incidence (31 of 39) of cyclin D{sub 1} overexpression.

  17. Age-dependent kinetics of dentate gyrus neurogenesis in the absence of cyclin D2

    Directory of Open Access Journals (Sweden)

    Ansorg Anne

    2012-05-01

    Full Text Available Abstract Background Adult neurogenesis continuously adds new neurons to the dentate gyrus and the olfactory bulb. It involves the proliferation and subsequent differentiation of neuronal progenitors, and is thus closely linked to the cell cycle machinery. Cell cycle progression is governed by the successive expression, activation and degradation of regulatory proteins. Among them, D-type cyclins control the exit from the G1 phase of the cell cycle. Cyclin D2 (cD2 has been shown to be required for the generation of new neurons in the neurogenic niches of the adult brain. It is differentially expressed during hippocampal development, and adult cD2 knock out (cD2KO mice virtually lack neurogenesis in the dentate gyrus and olfactory bulb. In the present study we examined the dynamics of postnatal and adult neurogenesis in the dentate gyrus (DG of cD2KO mice. Animals were injected with bromodeoxyuridine at seven time points during the first 10 months of life and brains were immunohistochemically analyzed for their potential to generate new neurons. Results Compared to their WT litters, cD2KO mice had considerably reduced numbers of newly born granule cells during the postnatal period, with neurogenesis becoming virtually absent around postnatal day 28. This was paralleled by a reduction in granule cell numbers, in the volume of the granule cell layer as well as in apoptotic cell death. CD2KO mice did not show any of the age-related changes in neurogenesis and granule cell numbers that were seen in WT litters. Conclusions The present study suggests that hippocampal neurogenesis becomes increasingly dependent on cD2 during early postnatal development. In cD2KO mice, hippocampal neurogenesis ceases at a time point at which the tertiary germinative matrix stops proliferating, indicating that cD2 becomes an essential requirement for ongoing neurogenesis with the transition from developmental to adult neurogenesis. Our data further support the notion that

  18. [Effect of starvation-induced autophagy on cell cycle of tumor cells].

    Science.gov (United States)

    Ge, Jun-Na; Huang, Dan; Xiao, Tian; Wang, Zun; Li, Xiao-Lan; Xiao, Hui; Tao, De-Ding; Gong, Jian-Ping

    2008-08-01

    No serum starvation could induce autophagy and cell cycle arrest. Although autophagy and cell cycle have been widely explored, little is known about their relationship. This study was to observe the change of Cyclin expression during starvation-induced autophagy to discuss the effect of autophagy on cell cycle. In control group, HeLa cells were treated with d-Hanks solution (a medium with no serum). In experiment group, HeLa cells were treated with d-Hanks solution containing 3-methyladenine (3-MA, a specific inhibitor of autophagy). Cells were harvested after being starved for 0, 3, 6 and 12 h. Flow cytometry (FCM) and Weston blot were used to detect Cyclin and microtubule-associated protein 1 light chain 3(LC-3) which marked autophagy specifically. In control group, the expression of LC-3 protein was detected early after being starved for 3 h, and gradually increased along with starvation; the expression of Cyclin D3 and Cyclin E was decreased evidently after a short-time starvation (3 h) and descended to the minimum when cells were being starved for 6 h; the expression of Cyclin A and Cyclin B1 were apparently decreased after being starved for 6 h. In experiment group, LC-3 protein could not be detected during starvation when cells were exposed to 3-MA and the down-regulation of Cyclins was suppressed. Autophagy is involved in starvation-induced hydrolysis of Cyclins. The hydrolysis of Cyclin D3 and Cyclin E is quicker than that of Cyclin A and Cyclin B1.

  19. Mitochondrial Fragmentation Due to Inhibition of Fusion Increases Cyclin B through Mitochondrial Superoxide Radicals.

    Directory of Open Access Journals (Sweden)

    Tejas M Gupte

    Full Text Available During the cell cycle, mitochondria undergo regulated changes in morphology. Two particularly interesting events are first, mitochondrial hyperfusion during the G(1-S transition and second, fragmentation during entry into mitosis. The mitochondria remain fragmented between late G(2- and mitotic exit. This mitotic mitochondrial fragmentation constitutes a checkpoint in some cell types, of which little is known. We bypass the 'mitotic mitochondrial fragmentation' checkpoint by inducing fragmented mitochondrial morphology and then measure the effect on cell cycle progression. Using Drosophila larval hemocytes, Drosophila S2R(+ cell and cells in the pouch region of wing imaginal disc of Drosophila larvae we show that inhibiting mitochondrial fusion, thereby increasing fragmentation, causes cellular hyperproliferation and an increase in mitotic index. However, mitochondrial fragmentation due to over-expression of the mitochondrial fission machinery does not cause these changes. Our experiments suggest that the inhibition o