Manipulation of the expression of regulatory genes of polyamine metabolism results in specific alterations of the cell-cycle progression

We have previously reported that cyclical phases of accumulation and depletion of polyamines occur during cell-cycle progression. Regulatory ornithine decarboxylase (ODC) catalyses the first step of polyamine biosynthesis. Ornithine decarboxylase antizyme (OAZ), induced by high polyamine levels, inhibits ODC activity and prevents extracellular polyamine uptake. Spermidine/spermine N1-acetyltransferase (SSAT) regulates the polyamine degradation/excretion pathway. Here we show that 24h transient transfection of immortalized human prostatic epithelial cells (PNT1A and PNT2) with antisense ODC RNA or OAZ cDNA, or both, while effectively causing marked decreases of ODC activity and polyamine (especially putrescine) concentrations, resulted in accumulation of cells in the S phase of the cell cycle. Transfection with SSAT cDNA led to more pronounced decreases in spermidine and spermine levels and resulted in accumulation of cells in the G2/M phases. Transfection with all three constructs together produced maximal depletion of all polyamines, accompanied by accumulation of PNT1A cells in the S phase and PNT2 cells in the G0/G1 and G2/M phases. Accumulation of PNT1A cells in the S phase progressively increased at 15, 18 and 24h of transfection with antisense ODC and/or OAZ cDNA. At 24h, the DNA content was always reduced, as a possible outcome of altered chromosome condensation. A direct link between polyamine metabolism, cell proliferation and chromatin structure is thus proposed.

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Cells and polyamines do it cyclically

Essays in Biochemistry ◽

10.1042/bse0460005 ◽

2009 ◽

Vol 46 ◽

pp. 63-76 ◽

Cited By ~ 20

Author(s):

Kersti Alm ◽

Stina Oredsson

Keyword(s):

Cell Cycle ◽

Cell Cycle Progression ◽

S Phase ◽

Negative Regulator ◽

Biosynthetic Activity ◽

Polyamine Biosynthesis ◽

Cycle Progression ◽

Daughter Cells ◽

G0 Phase ◽

Proliferating Cell

Cell-cycle progression is a one-way journey where the cell grows in size to be able to divide into two equally sized daughter cells. The cell cycle is divided into distinct consecutive phases defined as G1 (first gap), S (synthesis), G2 (second gap) and M (mitosis). A non-proliferating cell, which has retained the ability to enter the cell cycle when it receives appropriate signals, is in G0 phase, and cycling cells that do not receive proper signals leave the cell cycle from G1 into G0. One of the major events of the cell cycle is the duplication of DNA during S-phase. A group of molecules that are important for proper cell-cycle progression is the polyamines. Polyamine biosynthesis occurs cyclically during the cell cycle with peaks in activity in conjunction with the G1/S transition and at the end of S-phase and during G2-phase. The negative regulator of polyamine biosynthesis, antizyme, shows an inverse activity compared with the polyamine biosynthetic activity. The levels of the polyamines, putrescine, spermidine and spermine, double during the cell cycle and show a certain degree of cyclic variation in accordance with the biosynthetic activity. When cells in G0/G1-phase are seeded in the presence of compounds that prevent the cell-cycle-related increases in the polyamine pools, the S-phase of the first cell cycle is prolonged, whereas the other phases are initially unaffected. The results point to an important role for polyamines with regard to the ability of the cell to attain optimal rates of DNA replication.

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Polyamine dependence of normal cell-cycle progression

Biochemical Society Transactions ◽

10.1042/bst0310366 ◽

2003 ◽

Vol 31 (2) ◽

pp. 366-370 ◽

Cited By ~ 106

Author(s):

S.M. Oredsson

Keyword(s):

Cell Cycle ◽

Enzyme Activities ◽

Cell Cycle Progression ◽

Cyclin A ◽

S Phase ◽

Cycle Phase ◽

Polyamine Biosynthesis ◽

Cycle Progression ◽

Elongation Step ◽

Key Steps

The driving force of the cell cycle is the activities of cyclin-dependent kinases (CDKs). Key steps in the regulation of the cell cycle therefore must impinge upon the activities of the CDKs. CDKs exert their functions when bound to cyclins that are expressed cyclically during the cell cycle. Polyamine biosynthesis varies bicyclically during the cell cycle with peaks in enzyme activities at the G1/S and S/G2 transitions. The enzyme activities are regulated at transcriptional, translational and post-translational levels. When cells are seeded in the presence of drugs that interfere with polyamine biosynthesis, cell cycle progression is affected within one cell cycle after seeding. The cell cycle phase that is most sensitive to polyamine biosynthesis inhibition is the S phase, while effects on the G1 and G2/M phases occur at later time points. The elongation step of DNA replication is negatively affected when polyamine pools are not allowed to increase normally during cell proliferation. Cyclin A is expressed during the S phase and cyclin A/CDK2 is important for a normal rate of DNA elongation. Cyclin A expression is lowered in cells treated with polyamine biosynthesis inhibitors. Thus, polyamines may affect S phase progression by participating in the regulation of cyclin A expression.

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The Meiosis-Specific Crs1 Cyclin Is Required for Efficient S-Phase Progression and Stable Nuclear Architecture

International Journal of Molecular Sciences ◽

10.3390/ijms22115483 ◽

2021 ◽

Vol 22 (11) ◽

pp. 5483

Author(s):

Luisa F. Bustamante-Jaramillo ◽

Celia Ramos ◽

Cristina Martín-Castellanos

Keyword(s):

Cell Cycle ◽

Translational Control ◽

Cell Cycle Progression ◽

Nuclear Architecture ◽

Strand Break ◽

Spindle Pole ◽

S Phase ◽

Cycle Progression ◽

Single Wave ◽

Phase Progression

Cyclins and CDKs (Cyclin Dependent Kinases) are key players in the biology of eukaryotic cells, representing hubs for the orchestration of physiological conditions with cell cycle progression. Furthermore, as in the case of meiosis, cyclins and CDKs have acquired novel functions unrelated to this primal role in driving the division cycle. Meiosis is a specialized developmental program that ensures proper propagation of the genetic information to the next generation by the production of gametes with accurate chromosome content, and meiosis-specific cyclins are widespread in evolution. We have explored the diversification of CDK functions studying the meiosis-specific Crs1 cyclin in fission yeast. In addition to the reported role in DSB (Double Strand Break) formation, this cyclin is required for meiotic S-phase progression, a canonical role, and to maintain the architecture of the meiotic chromosomes. Crs1 localizes at the SPB (Spindle Pole Body) and is required to stabilize the cluster of telomeres at this location (bouquet configuration), as well as for normal SPB motion. In addition, Crs1 exhibits CDK(Cdc2)-dependent kinase activity in a biphasic manner during meiosis, in contrast to a single wave of protein expression, suggesting a post-translational control of its activity. Thus, Crs1 displays multiple functions, acting both in cell cycle progression and in several key meiosis-specific events.

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Spirulina Crude Protein Promotes the Migration and Proliferation in IEC-6 Cells by Activating EGFR/MAPK Signaling Pathway

Marine Drugs ◽

10.3390/md17040205 ◽

2019 ◽

Vol 17 (4) ◽

pp. 205

Author(s):

Su-Jin Jeong ◽

Jeong-Wook Choi ◽

Min-Kyeong Lee ◽

Youn-Hee Choi ◽

Taek-Jeong Nam

Keyword(s):

Cell Cycle ◽

Epithelial Cells ◽

Signaling Pathway ◽

Crude Protein ◽

Cell Cycle Progression ◽

Intestinal Epithelial Cells ◽

Mapk Signaling ◽

S Phase ◽

Intestinal Epithelial ◽

Cycle Progression

Spirulina is a type of filamentous blue-green microalgae known to be rich in nutrients and to have pharmacological effects, but the effect of spirulina on the small intestine epithelium is not well understood. Therefore, this study aims to investigate the proliferative effects of spirulina crude protein (SPCP) on a rat intestinal epithelial cells IEC-6 to elucidate the mechanisms underlying its effect. First, the results of wound-healing and cell viability assays demonstrated that SPCP promoted migration and proliferation in a dose-dependent manner. Subsequently, when the mechanisms of migration and proliferation promotion by SPCP were confirmed, we found that the epidermal growth factor receptor (EGFR) and mitogen-activated protein (MAPK) signaling pathways were activated by phosphorylation. Cell cycle progression from G0/G1 to S phase was also promoted by SPCP through upregulation of the expression levels of cyclins and cyclin-dependent kinases (Cdks), which regulate cell cycle progression to the S phase. Meanwhile, the expression of cyclin-dependent kinase inhibitors (CKIs), such as p21 and p27, decreased with SPCP. In conclusion, our results indicate that activation of EGFR and its downstream signaling pathway by SPCP treatment regulates cell cycle progression. Therefore, these results contribute to the research on the molecular mechanism for SPCP promoting the migration and proliferation of rat intestinal epithelial cells.

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Cubic Membranes Formation in Synchronized Human Hepatocellular Carcinoma Cells Reveals a Possible Role as a Structural Antioxidant Defense System in Cell Cycle Progression

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2020.617406 ◽

2020 ◽

Vol 8 ◽

Author(s):

Deqin Kong ◽

Rui Liu ◽

Jiangzheng Liu ◽

Qingbiao Zhou ◽

Jiaxin Zhang ◽

...

Keyword(s):

Hepatocellular Carcinoma ◽

Cell Cycle ◽

Cell Cycle Progression ◽

S Phase ◽

Carcinoma Cells ◽

Human Hepatocellular Carcinoma ◽

Cycle Progression ◽

Hepatocellular Carcinoma Cells ◽

G2 Phase ◽

Human Hepatocellular Carcinoma Cells

Cubic membranes (CMs) represent unique biological membrane structures with highly curved three-dimensional periodic minimal surfaces, which have been observed in a wide range of cell types and organelles under various stress conditions (e. g., starvation, virus-infection, and oxidation). However, there are few reports on the biological roles of CMs, especially their roles in cell cycle. Hence, we established a stable cell population of human hepatocellular carcinoma cells (HepG2) of 100% S phase by thymidine treatment, and determined certain parameters in G2 phase released from S phase. Then we found a close relationship between CMs formation and cell cycle, and an increase in reactive oxygen species (ROS) and mitochondrial function. After the synchronization of HepG2 cells were induced, CMs were observed through transmission electron microscope in G2 phase but not in G1, S and M phase. Moreover, the increased ATP production, mitochondrial and intracellular ROS levels were also present in G2 phase, which demonstrated a positive correlation with CMs formation by Pearson correlation analysis. This study suggests that CMs may act as an antioxidant structure in response to mitochondria-derived ROS during G2 phase and thus participate in cell cycle progression.

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Effects of Guanine Nucleotide Depletion on Cell Cycle Progression in Human T Lymphocytes

Blood ◽

10.1182/blood.v91.8.2896.2896_2896_2904 ◽

1998 ◽

Vol 91 (8) ◽

pp. 2896-2904 ◽

Cited By ~ 69

Author(s):

Josée Laliberté ◽

Ann Yee ◽

Yue Xiong ◽

Beverly S. Mitchell

Keyword(s):

Cell Cycle ◽

T Cells ◽

T Lymphocytes ◽

Cell Cycle Progression ◽

S Phase ◽

Guanine Nucleotides ◽

Erythroid Cell ◽

Guanine Nucleotide ◽

Cycle Progression ◽

Human T Lymphocytes

Depletion of guanine nucleotide pools after inhibition of inosine monophosphate dehydrogenase (IMPDH) potently inhibits DNA synthesis by arresting cells in G1 and has been shown to induce the differentiation of cultured myeloid and erythroid cell lines, as well as chronic granulocytic leukemic cells after blast transformation. Inhibitors of IMPDH are also highly effective as immunosuppressive agents. The mechanism underlying these pleiotropic effects of depletion of guanine nucleotides is unknown. We have examined the effects of mycophenolic acid (MPA), a potent IMPDH inhibitor, on the cell cycle progression of activated normal human T lymphocytes. MPA treatment resulted in the inhibition of pRb phosphorylation and cell entry into S phase. The expression of cyclin D3, a major component of the cyclin-dependent kinase (CDK) activity required for pRb phosphorylation, was completely abrogated by MPA treatment of T cells activated by interleukin-2 (IL-2) and leucoagglutinin (PHA-L), whereas the expression of cyclin D2, CDK6, and CDK4 was more mildly attenuated. The direct kinase activity of a complex immunoprecipitated with anti-CDK6 antibody was also inhibited. In addition, MPA prevented the IL-2–induced elimination of p27Kip1, a CDK inhibitor, and resulted in the retention of high levels of p27Kip1 in IL-2/PHA-L–treated T cells bound to CDK2. These results indicate that inhibition of the de novo synthesis of guanine nucleotides blocks the transition of normal peripheral blood T lymphocytes from G0 to S phase in early- to mid-G1 and that this cell cycle arrest results from inhibition of the induction of cyclin D/CDK6 kinase and the elimination of p27Kip1 inhibitory activity.

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Cellular Ras and Cyclin D1 Are Required during Different Cell Cycle Periods in Cycling NIH 3T3 Cells

Molecular and Cellular Biology ◽

10.1128/mcb.19.7.4623 ◽

1999 ◽

Vol 19 (7) ◽

pp. 4623-4632 ◽

Cited By ~ 50

Author(s):

Masahiro Hitomi ◽

Dennis W. Stacey

Keyword(s):

Cell Cycle ◽

Dna Synthesis ◽

Cyclin D1 ◽

Cell Cycle Progression ◽

Time Lapse ◽

S Phase ◽

3T3 Cells ◽

Cycle Progression ◽

Nih 3T3 ◽

Nih 3T3 Cells

ABSTRACT Novel techniques were used to determine when in the cell cycle of proliferating NIH 3T3 cells cellular Ras and cyclin D1 are required. For comparison, in quiescent cells, all four of the inhibitors of cell cycle progression tested (anti-Ras, anti-cyclin D1, serum removal, and cycloheximide) became ineffective at essentially the same point in G1 phase, approximately 4 h prior to the beginning of DNA synthesis. To extend these studies to cycling cells, a time-lapse approach was used to determine the approximate cell cycle position of individual cells in an asynchronous culture at the time of inhibitor treatment and then to determine the effects of the inhibitor upon recipient cells. With this approach, anti-Ras antibody efficiently inhibited entry into S phase only when introduced into cells prior to the preceding mitosis, several hours before the beginning of S phase. Anti-cyclin D1, on the other hand, was an efficient inhibitor when introduced up until just before the initiation of DNA synthesis. Cycloheximide treatment, like anti-cyclin D1 microinjection, was inhibitory throughout G1 phase (which lasts a total of 4 to 5 h in these cells). Finally, serum removal blocked entry into S phase only during the first hour following mitosis. Kinetic analysis and a novel dual-labeling technique were used to confirm the differences in cell cycle requirements for Ras, cyclin D1, and cycloheximide. These studies demonstrate a fundamental difference in mitogenic signal transduction between quiescent and cycling NIH 3T3 cells and reveal a sequence of signaling events required for cell cycle progression in proliferating NIH 3T3 cells.

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Phosphorylation-induced unfolding regulates p19INK4dduring the human cell cycle

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1719774115 ◽

2018 ◽

Vol 115 (13) ◽

pp. 3344-3349 ◽

Cited By ~ 9

Author(s):

Amit Kumar ◽

Mohanraj Gopalswamy ◽

Annika Wolf ◽

David J. Brockwell ◽

Mechthild Hatzfeld ◽

...

Keyword(s):

Cell Cycle ◽

Structural Biology ◽

Cell Cycle Progression ◽

S Phase ◽

Cycle Progression ◽

Cyclin Dependent Kinases ◽

Ankyrin Repeat Protein ◽

Dependent Protein ◽

Local Unfolding ◽

Molecular Mode

Cell cycle progression is tightly regulated by cyclin-dependent kinases (CDKs). The ankyrin-repeat protein p19INK4dfunctions as a key regulator of G1/S transition; however, its molecular mode of action is unknown. Here, we combine cell and structural biology methods to unravel the mechanism by which p19INK4dcontrols cell cycle progression. We delineate how the stepwise phosphorylation of p19INK4dSer66 and Ser76 by cell cycle-independent (p38) and -dependent protein kinases (CDK1), respectively, leads to local unfolding of the three N-terminal ankyrin repeats of p19INK4d. This dissociates the CDK6–p19INK4dinhibitory complex and, thereby, activates CDK6. CDK6 triggers entry into S-phase, whereas p19INK4dis ubiquitinated and degraded. Our findings reveal how signaling-dependent p19INK4dunfolding contributes to the irreversibility of G1/S transition.

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Growth Characteristics of Anaerobically Treated Early and Late S-Period of Ehrlich Ascites Tumor Cells after Reaeration

Zeitschrift für Naturforschung C ◽

10.1515/znc-1983-3-426 ◽

1983 ◽

Vol 38 (3-4) ◽

pp. 313-318 ◽

Cited By ~ 6

Author(s):

Rainer Merz ◽

Friedhelm Schneider

Keyword(s):

Cell Cycle ◽

Flow Cytometry ◽

Cell Cycle Progression ◽

S Phase ◽

Ehrlich Ascites Tumor Cells ◽

Ascites Tumor ◽

Cycle Progression ◽

Ehrlich Ascites ◽

Main Fraction ◽

S Period

Utilizing centrifugal elutriation, early and late S-phase cells were separated from 4, 8 and 12 h anaerobically cultured Ehrlich Ascites tumor cells strain Karzel. The cytokinetic properties of these fractions after reaeration were studied by flow cytometry and the BrdU-H 33258-technique of flow cytometry. After a 4 h period of anaerobiosis, growth of early S-phase cells is not changed, 8 h deprivation of oxygen causes a delay of cell cycle progression, while the main fraction of 12 h anaerobically treated early S-populations did not divide after reaeration within 24 h. In comparison to early S-phase cells the cell cycle progression of the main fraction of late S-period is accelerated after a 4 h exclusion of oxygen. A fraction of 8 h anaerobically pretreated late S-cells continues to cycle, but a considerable number reinitiates DNA synthesis without preceeding division. Cells with DNA content up to 8 c are detected by flow cytometry. 12 h anaerobically cultured late S-cells do not divide after reaeration, a large number of these cells starts again to synthesize DNA. A considerable part of tetraploid cells retain viability, divide and enter a new cell cycle, another part of the cells disintegrates

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Identification of DHX9 as a cell cycle regulated nucleolar recruitment factor for CIZ1

Scientific Reports ◽

10.1038/s41598-020-75160-z ◽

2020 ◽

Vol 10 (1) ◽

Author(s):

Urvi Thacker ◽

Tekle Pauzaite ◽

James Tollitt ◽

Maria Twardowska ◽

Charlotte Harrison ◽

...

Keyword(s):

Cell Cycle ◽

Dna Replication ◽

Cell Cycle Progression ◽

Zinc Finger Protein ◽

S Phase ◽

Cycle Progression ◽

Inactive X Chromosome ◽

Functional Characterisation ◽

Interaction Partners

Abstract CIP1-interacting zinc finger protein 1 (CIZ1) is a nuclear matrix associated protein that facilitates a number of nuclear functions including initiation of DNA replication, epigenetic maintenance and associates with the inactive X-chromosome. Here, to gain more insight into the protein networks that underpin this diverse functionality, molecular panning and mass spectrometry are used to identify protein interaction partners of CIZ1, and CIZ1 replication domain (CIZ1-RD). STRING analysis of CIZ1 interaction partners identified 2 functional clusters: ribosomal subunits and nucleolar proteins including the DEAD box helicases, DHX9, DDX5 and DDX17. DHX9 shares common functions with CIZ1, including interaction with XIST long-non-coding RNA, epigenetic maintenance and regulation of DNA replication. Functional characterisation of the CIZ1-DHX9 complex showed that CIZ1-DHX9 interact in vitro and dynamically colocalise within the nucleolus from early to mid S-phase. CIZ1-DHX9 nucleolar colocalisation is dependent upon RNA polymerase I activity and is abolished by depletion of DHX9. In addition, depletion of DHX9 reduced cell cycle progression from G1 to S-phase in mouse fibroblasts. The data suggest that DHX9-CIZ1 are required for efficient cell cycle progression at the G1/S transition and that nucleolar recruitment is integral to their mechanism of action.

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