scholarly journals Rapha Myr®, a Blend of Sulforaphane and Myrosinase, Exerts Antitumor and Anoikis-Sensitizing Effects on Human Astrocytoma Cells Modulating Sirtuins and DNA Methylation

2020 ◽  
Vol 21 (15) ◽  
pp. 5328 ◽  
Author(s):  
Barbara Tomasello ◽  
Maria Domenica Di Mauro ◽  
Giuseppe Antonio Malfa ◽  
Rosaria Acquaviva ◽  
Fulvia Sinatra ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Methylation ◽  
Cell Cycle Progression ◽  
Fruits And Vegetables ◽  
Redox Homeostasis ◽  
Focal Adhesions ◽  
Mrna Levels ◽  
Genotoxic Effects ◽  
Epigenetic Biomarkers ◽  
Human Astrocytoma

Brain and other nervous system cancers are the 10th leading cause of death worldwide. Genome instability, cell cycle deregulation, epigenetic mechanisms, cytoarchitecture disassembly, redox homeostasis as well as apoptosis are involved in carcinogenesis. A diet rich in fruits and vegetables is inversely related with the risk of developing cancer. Several studies report that cruciferous vegetables exhibited antiproliferative effects due to the multi-pharmacological functions of their secondary metabolites such as isothiocyanate sulforaphane deriving from the enzymatic hydrolysis of glucosinolates. We treated human astrocytoma 1321N1 cells for 24 h with different concentrations (0.5, 1.25 and 2.5% v/v) of sulforaphane plus active myrosinase (Rapha Myr®) aqueous extract (10 mg/mL). Cell viability, DNA fragmentation, PARP-1 and γH2AX expression were examined to evaluate genotoxic effects of the treatment. Cell cycle progression, p53 and p21 expression, apoptosis, cytoskeleton morphology and cell migration were also investigated. In addition, global DNA methylation, DNMT1 mRNA levels and nuclear/mitochondrial sirtuins were studied as epigenetic biomarkers. Rapha Myr® exhibited low antioxidant capability and exerted antiproliferative and genotoxic effects on 1321N1 cells by blocking the cell cycle, disarranging cytoskeleton structure and focal adhesions, decreasing the integrin α5 expression, renewing anoikis and modulating some important epigenetic pathways independently of the cellular p53 status. In addition, Rapha Myr® suppresses the expression of the oncogenic p53 mutant protein. These findings promote Rapha Myr® as a promising chemotherapeutic agent for integrated cancer therapy of human astrocytoma.

scholarly journals Primase p49 mRNA expression is serum stimulated but does not vary with the cell cycle.

1989 ◽  
Vol 9 (5) ◽  
pp. 1940-1945 ◽  
Author(s):  
B Y Tseng ◽  
C E Prussak ◽  
M T Almazan
Keyword(s):  
Cell Cycle ◽  
Dna Synthesis ◽  
Mammalian Cells ◽  
Cell Cycle Progression ◽  
Small Subunit ◽  
Mrna Levels ◽  
Proliferating Cells ◽  
Restriction Point ◽  
Serum Stimulation ◽  
G1 Cells

Expression of the small-subunit p49 mRNA of primase, the enzyme that synthesizes oligoribonucleotides for initiation of DNA replication, was examined in mouse cells stimulated to proliferate by serum and in growing cells. The level of p49 mRNA increased approximately 10-fold after serum stimulation and preceded synthesis of DNA and histone H3 mRNA by several hours. Expression of p49 mRNA was not sensitive to inhibition by low concentrations of cycloheximide, which suggested that the increase in mRNA occurred before the restriction point control for cell cycle progression described for mammalian cells and was not under its control. p49 mRNA levels were not coupled to DNA synthesis, as observed for the replication-dependent histone genes, since hydroxyurea or aphidicolin had no effect on p49 mRNA levels when added before or during S phase. These inhibitors did have an effect, however, on the stability of p49 mRNA and increased the half-life from 3.5 h to about 20 h, which suggested an interdependence of p49 mRNA degradation and DNA synthesis. When growing cells were examined after separation by centrifugal elutriation, little difference was detected for p49 mRNA levels in different phases of the cell cycle. This was also observed when elutriated G1 cells were allowed to continue growth and then were blocked in M phase with colcemid. Only a small decrease in p49 mRNA occurred, whereas H3 mRNA rapidly decreased, when cells entered G2/M. These results indicate that the level of primase p49 mRNA is not cell cycle regulated but is present constitutively in proliferating cells.

DNA methylation in Caulobacter and other Alphaproteobacteria during cell cycle progression

2014 ◽  
Vol 22 (9) ◽  
pp. 528-535 ◽  
Author(s):  
Saswat S. Mohapatra ◽  
Antonella Fioravanti ◽  
Emanuele G. Biondi
Keyword(s):  
Cell Cycle ◽  
Dna Methylation ◽  
Cell Cycle Progression ◽  
Cycle Progression

scholarly journals In silico APC/C substrate discovery reveals cell cycle degradation of chromatin regulators including UHRF1

2020 ◽  
Author(s):  
Jennifer L. Kernan ◽  
Raquel C. Martinez-Chacin ◽  
Xianxi Wang ◽  
Rochelle L. Tiedemann ◽  
Thomas Bonacci ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Methylation ◽  
In Silico ◽  
Cell Cycle Progression ◽  
Vital Role ◽  
Regulatory Proteins ◽  
Phase Cell ◽  
Cell Physiology ◽  
Cycle Progression ◽  
Chromatin Regulator

AbstractThe Anaphase-Promoting Complex/Cyclosome (APC/C) is an E3 ubiquitin ligase and critical regulator of cell cycle progression. Despite its vital role, it has remained challenging to globally map APC/C substrates. By combining orthogonal features of known substrates, we predicted APC/C substrates in silico. This analysis identified many known substrates and suggested numerous candidates. Unexpectedly, chromatin regulatory proteins are enriched among putative substrates and we show that several chromatin proteins bind APC/C, oscillate during the cell cycle and are degraded following APC/C activation, consistent with being direct APC/C substrates. Additional analysis revealed detailed mechanisms of ubiquitylation for UHRF1, a key chromatin regulator involved in histone ubiquitylation and DNA methylation maintenance. Disrupting UHRF1 degradation at mitotic exit accelerates G1-phase cell cycle progression and perturbs global DNA methylation patterning in the genome. We conclude that APC/C coordinates crosstalk between cell cycle and chromatin regulatory proteins. This has potential consequences in normal cell physiology, where the chromatin environment changes depending on proliferative state, as well as in disease.

Pro-Apoptotic and Proliferative Effects of Bone Marrow Adipocytes on Myeloid Leukemia Cells.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 4572-4572
Author(s):  
Linhua Jin ◽  
Marina Konopleva ◽  
Yixin Zhou ◽  
Akimichi Osaka ◽  
Michael Andreeff ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Bone Marrow ◽  
Cell Cycle Progression ◽  
Lipid Vesicles ◽  
Mrna Levels ◽  
U937 Cells ◽  
Mature Adipocyte ◽  
Cycle Progression ◽  
Induced Apoptosis ◽  
Pai 1

Abstract Abstract 4572 Bone marrow stromal cells (MSCs) from elderly subjects have a reduced capacity to differentiate into osteoblasts and an increased capacity to differentiate into adipocytes, which leads to progressive accumulation of fat in the bone marrow space with increasing age. Adipocytes are the prevalent stromal cell type in adult BM that play an important role in the leukemic bone marrow microenvironment (Tabe et al., Blood 2004 103:1815-22). In this study, we examined the role of BM-derived adipocytes at different stages of differentiation on proliferation and apoptosis of AML cells. U937 cells were co-cultured with BM-derived MSC, MSC-differentiated pre-adipocytes (containing few small lipid vesicles), and mature adipocytes (with multiple hypertrophic lipid vesicules). Under serum-starved conditions, MSC and premature/mature adipocytes induced cell cycle progression of U937 cells with increase in the proportion of cells in S- and G2/M-phase fractions, and inhibited spontaneous cell death with decrease in subG1 fractions. However, only pre-adipocytes inhibited Ara-C-induced cell killing (Table 1). We next focused on lepin and plasminogen activator inhibitor 1(PAI-1) as potential mediators of these effects by adipocytes. Leptin mRNA and protein levels were upregulated during adipocytic differentiation (mRNA relative expression to GAPDH (PCR): MSC 0, premature adipocyte 2.0±0.5, mature adipocyte 123.3± 35.0; leptin secretion: MSC 23.1±2.9, premature adipocyte 49.3±11.3, mature adipocyte 110.0±4.6 pg/mL (ELISA). PAI-1 mRNA levels were increased in mature adipocyte (relative expression to GAPDH: MSC 314.9±46.5, premature adipocyte 215.1, mature adipocyte 3766.1±656.2). Since PPARÿ activation is known to promote maturation and re-generation of fat-derived adipocytes, we next examined the potential of the synthetic triterpenoid 2-cyano-3,12-dioxooleana-1,9-dien-28-oic acid (CDDO) on the BM adipocytes, leptin and PAI-1 production and the survival of the leukemic cells. CDDO induced adipocyte re-generation with significant increase of the number of Oil-Red(+) small sized lipid vesicles without apoptosis induction. This resulted in a markedly enhanced leptin release from adipocytes (10-fold and 23-fold increase at 0.5 μM and 1.0 μM CDDO, respectively, at 72 hrs) without change in leptin mRNA transcription. On the contrary, PAI-1 mRNA levels were significantly decreased by CDDO (6 fold decrease in MSC, 4 fold decrease in premature adipocyte, 6 fold decrease in mature adipocyte). Co-culture of U937 cells with CDDO-primed premature adipocytes and mature adipocytes resulted in increased spontaneous apoptosis of U937 cells compared to adipocytes not exposed to CDDO (% specific apoptosis, U937 co-cultured with CDDO-primed premature adipocytes 26.9 %, mature adipocytes 20.9 %). At the same time, CDDO-primed premature adipocytes induced significant cell cycle progression with decreased proportion of G0/G1-phase and increase in S-phase fractions in U937 cells (Table 2). Co-culture with CDDO-primed premature adipocytes or with premature adipocytes co-treated with recombinant leptin increased subG1- and S-phase fractions in Ara-C-treated U937 cells compared to U937 cells co-cultured with premature adipocytes (Table 3). In mature adipocytes, which already produce high levels of leptin, CDDO or leptin treatment failed to modulate anti-apoptotic or proliferative effects of AraC on U937 cells. In contrast, human recombinant PAI-1 effectively inhibited spontaneous and Ara-C induced apoptosis of U937 cells (decreased % of Annexin V: spontaneous apoptosis 11.2±1.1%, Ara-C induced apoptosis 15.1± 1.7%). In summary, these results suggest that BM pre-adipocytes support proliferation and survival of myeloid leukemia cells in part through complementary effects of leptin and PAI-1. Our findings indicate that secretion of leptin during MSC differentiation or through PPARg ligation promotes cell cycle progression, while PAI-1 primarily inhibits apoptosis of AML cells. It is conceivable that increased adipocyte content of BM in elderly AML patients may negatively affect the responsiveness of AML cells to chemotherapy. Disclosures: No relevant conflicts of interest to declare.

scholarly journals The cell cycle-coupled expression of topoisomerase IIalpha during S phase is regulated by mRNA stability and is disrupted by heat shock or ionizing radiation.

1996 ◽  
Vol 16 (4) ◽  
pp. 1500-1508 ◽  
Author(s):  
P C Goswami ◽  
J L Roti Roti ◽  
C R Hunt
Keyword(s):  
Cell Cycle ◽  
Heat Shock ◽  
Ionizing Radiation ◽  
Mrna Stability ◽  
Cell Cycle Progression ◽  
Dna Supercoiling ◽  
S Phase ◽  
Mrna Levels ◽  
Cycle Progression ◽  
Topoisomerase Iialpha

Topoisomerase II is a multifunctional protein required during DNA replication, chromosome disjunction at mitosis, and other DNA-related activities by virtue of its ability to alter DNA supercoiling. The enzyme is encoded by two similar but nonidentical genes: the topoisomerase IIalpha and IIbeta genes. In HeLa cells synchronized by mitotic shake-off, topoisomeraseII alpha mRNA levels were found to vary as a function of cell cycle position, being 15-fold higher in late S phase (14 to 18 h postmitosis) than during G1 phase. Also detected was a corresponding increase in topoisomerase IIalpha protein synthesis at 14 to 18 h postmitosis which resulted in significantly higher accumulation of the protein during S and G2 phases. Topoisomerase IIalpha expression was not dependent on DNA synthesis during S phase, which could be inhibited without effect on the timing or level of mRNA expression. Mechanistically, topoisomerase IIalpha expression appears to be coupled to cell cycle position mainly through associated changes in mRNA stability. When cells are in S phase and mRNA levels are maximal, the half-life of topoisomerase IIalpha mRNA was determined to be approximately 30 min. A similar decrease in mRNA stability was also induced by two external factors known to delay cell cycle progression. Treatment of S-phase cells, at the time of maximum topoisomerase IIalpha mRNA stability, with either ionizing radiation (5 Gy) or heat shock (45 degrees C for 15 min) caused the accumulated topoisomerase IIalpha mRNA to decay. This finding suggests a potential relationship between stress-induced decreases in topoisomerase IIalpha expression and cell cycle progression delays in late S/G2.

scholarly journals The Saccharomyces cerevisiae RNase Mitochondrial RNA Processing Is Critical for Cell Cycle Progression at the End of Mitosis

Genetics ◽  
2002 ◽  
Vol 161 (3) ◽  
pp. 1029-1042 ◽  
Author(s):  
Ti Cai ◽  
Jason Aulds ◽  
Tina Gill ◽  
Michael Cerio ◽  
Mark E Schmitt
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Cell Cycle Progression ◽  
Mitochondrial Rna ◽  
Mrna Levels ◽  
Cycle Progression ◽  
Rnase Mrp ◽  
Processing Step ◽  
A Cell

Abstract We have identified a cell cycle delay in Saccharomyces cerevisiae RNase MRP mutants. Mutants delay with large budded cells, dumbbell-shaped nuclei, and extended spindles characteristic of “exit from mitosis” mutants. In accord with this, a RNase MRP mutation can be suppressed by overexpressing the polo-like kinase CDC5 or by deleting the B-type cyclin CLB1, without restoring the MRP-dependent rRNA-processing step. In addition, we identified a series of genetic interactions between RNase MRP mutations and mutations in CDC5, CDC14, CDC15, CLB2, and CLB5. As in most “exit from mitosis” mutants, levels of the Clb2 cyclin were increased. The buildup of Clb2 protein is not the result of a defect in the release of the Cdc14 phosphatase from the nucleolus, but rather the result of an increase in CLB2 mRNA levels. These results indicate a clear role of RNase MRP in cell cycle progression at the end of mitosis. Conservation of this function in humans may explain many of the pleiotropic phenotypes of cartilage hair hypoplasia.

Effect of fenofibrate on proliferation of SMMC-7721 cells via regulating cell cycle

2021 ◽  
pp. 096032712199190
Author(s):  
B Li ◽  
H-Y Jiang ◽  
Z-H Wang ◽  
Y-C Ma ◽  
Y-N Bao ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Proliferation ◽  
Liver Cancer ◽  
Cell Cycle Progression ◽  
Underlying Mechanism ◽  
Nuclear Antigen ◽  
Mrna Levels ◽  
Obvious Effect ◽  
Promote Cell Proliferation ◽  
Pparα Agonist

Liver cancer is a malignant cancer with great harmfulness. Fenofibrate is a peroxisome proliferation activated receptor (PPARα) agonist widely used in the treatment of dyslipidemia. Previous studies have shown that fenofibrate may promote cell proliferation, but the underlying mechanism has not been fully characterized. The aim of this study was to investigate the role of PPARα agonist fenofibrate in cell proliferation of SMMC-7721 cells compared with that of THLE-2 cells. SMMC-7721 and THLE-2 cells were treated with different concentrations of fenofibrate. Cell proliferation was analyzed by MTT, using flow cytometry for cell cycle analysis, and CyclinD1, Cyclin-dependent kinases2 (CDK2) and Proliferating Cell Nuclear Antigen (PCNA) were analyzed by Western blotting. RT-qPCR method was used to assess CDK2, CyclinD1 and PCNA mRNA levels. The results showed that 10−9–10−4 mol/L fenofibrate could induce cell growth and 10−4, 10−5, 10−6 mol/L fenofibrate could reduce the number of G0/G1 phase cells and increased in the number of cells in S and G2/M phase of cell cycle in SMMC-7721 cells. Furthermore, fenofibrate could significantly increase the expression of cell cycle related protein (CyclinD1, CDK2)and cell proliferation related proteins (PCNA). The use of PPARα inhibitor MT886 inhibited cell cycle progression and promote tumor cell apoptosis. But fenofibrate had no obvious effect on THLE-2 cells. These results revealed the effect of fenofibrate on the cell cycle of liver cancer cells, and provided a reasonable explanation for studying how fenofibrate promotes cell proliferation.

63 Bovine embryo-secreted microRNA-30c negatively regulates cell cycle progression through downregulation of CDK12

2019 ◽  
Vol 31 (1) ◽  
pp. 157
Author(s):  
X. Lin ◽  
E. Beckers ◽  
S. Mc Cafferty ◽  
J. P. Catani ◽  
K. J. Szymanska ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Progression ◽  
Differentially Expressed ◽  
Bovine Embryo ◽  
Mrna Levels ◽  
Bovine Embryos ◽  
Rt Pcr ◽  
Cycle Progression ◽  
Novel Mirna ◽  
Differentially Expressed Mirna

Identifying microRNA (miRNA) differentially secreted by intermediate (cleaved between 26.6-31.4 hpi) and slow cleaving bovine embryos (cleaved after 31.4 hpi), investigating how they influence embryo development and consequently if they can be used as biomarkers for bovine embryo development. MicroRNA collected from conditioned media (CM) of 167 individually cultured embryos were sequenced on an Illumina Miseq (Illumina, San Diego, CA, USA) after small RNA library construction with the Tailormix v2 kit (SeqMatic, Fremont, CA, USA) and compared with miRBase, using cow as primary organism, to identify known miRNA in all samples. To predict potential novel miRNA, the sequences were aligned against the annotated cow genome (GCA_000003055.3), structurally analysed, and compared with all other mammalian data in miRBase. Quantitative RT-PCR was used to validate miRNA sequencing results. Apoptosis staining was performed for functional analysis of the differentially expressed miRNA-30c (miR-30c). Luciferase reporter assay, Western blotting, and quantitative RT-PCR were used to validate one of the predicted target genes of miR-30c, CDK12. The DNA content analysis using propidium iodide staining followed by flow cytometry and a water-soluble tetrazolium-1 proliferation assay were performed together with ELISA to assess the effect of miR-30c on cell progression. Quantitative RT-PCR was used to assess mRNA levels of DNA damage response (DDR) genes. In total, 114 known and 180 potentially novel miRNA were found in bovine embryo CM. Seven differentially expressed miRNA were identified in the CM from embryos with different cleavage patterns and different quality: miR-30c, miR-10b, and miR-novel-66 were differentially expressed between slow and intermediate developing embryos; miR-10b, miR-novel-113, miR-novel-44, miR-novel-45, and miR-novel-139 were differentially expressed between blastocysts and degenerate embryos. We further focused on functional analysis of miR-30c. Exogeneous delivery of miR-30c mimics to cultured embryos resulted in an increased cell apoptosis. Additionally, transfer of miR-30c mimics to Madin-Darby bovine kidney cells (MDBK) reduced cell cycle progression/cell proliferation, while inhibition of miR-30c resulted in the opposite. In addition, knockdown of CDK12 caused significant decreases in mRNA levels of DDR genes BRCA1, FANCD2, FANCI, and ATR. These data were derived from 3 independent experiments and were considered significant with a P-value<0.05. In vitro-cultured bovine embryos secrete miRNA in the culture medium depending on their developmental capacity. One of these miRNA, miR-30c, increases bovine embryo apoptosis when exogeneously delivered. In addition, miR-30c directly targets and down-regulates CDK12, indicating the possibility that delivery of miR-30c mimics influences cell cycle progression by inhibiting DDR pathways through regulating CDK12. These findings contribute to a better understanding of the mechanisms of secreted miRNA participation in intercellular communication.

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