Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.22 (cdc2)
 8,319 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In yeast, the protein kinase p34cdc2 plays a role in regulating both the G2 to M and G1 to S phase transitions. The discovery of multiple homologues of the protein in cells of higher eukaryotic organisms suggests that different cell cycle regulatory events may be performed by different kinases in such cells. Here, the synthesis and metabolism of the human forms of these proteins are described in a normal human cell type, peripheral blood T lymphocytes that have been stimulated to enter the cell cycle in vitro. Using a carboxyl-terminus antiserum specific for true p34cdc2, the protein could first be found in T cells at about 24 to 30 h after stimulation, just before the initiation of DNA synthesis. Three forms of the enzyme could be resolved by denaturing gel electrophoresis: an unphosphorylated form with an apparent molecular mass of 34,500 daltons and two phosphorylated derivatives. In cells synchronized at G2/M phase with nocodazole, p34 was almost entirely in the unphosphorylated form whereas the phosphorylated derivatives were more predominant in cultures arrested at the G1/S border with aphidicolin. The relationship of p34 synthesis to the phosphorylation of p110Rb, an event known to be associated with passage through late G1 and/or the G1/S phase transition, was also investigated. It was noted that p110Rb phosphorylation began before p34 synthesis first became detectable. Furthermore, it appeared that the two events could be largely uncoupled by treating cells with deferoxamine (10 microM), an iron chelating agent that arrests T cells at a point in late G1 phase but substantially before the G1 to S phase transition. Under these conditions, p110Rb phosphorylation was almost completely accomplished in the absence of significant p34 synthesis, a finding that suggests that most or all of p110 phosphorylation is performed by kinases other than p34. Because of this observation, extracts were next examined for p34-like molecules using an antibody against the so-called PSTAIRE domain found in all cdc2 homologues identified to date. A species of protein with a mobility slightly less than true p34 was found, even in resting T cells. Upon stimulation, this protein increased slightly in amount, and a second protein with a mobility greater than p34, a putative p33cdk2, was seen. Not only was the appearance of these proteins not inhibited by deferoxamine but they accumulated in cultures treated with the drug, suggesting that p33, and not p34, may be the G1 phase kinase for p110Rb.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Regulation of synthesis of p34cdc2 and its homologues and their relationship to p110Rb phosphorylation during cell cycle progression of normal human T cells. 154 21

Iron removal by the chelating-agent deferoxamine (DFO) arrests cell cycle progression of activated human T cells in late G1 phase, before the G1/S border. The effects of the drug on molecules that regulate progression through the cell cycle were defined. DFO (10 mumol/L) inhibited induction of transcription of the cdc2 gene, but had no effect on accumulation of cdk2, cdk4, or interleukin (IL)-2-transcripts. No detectable p34cdc2 protein accumulated, but synthesis of the p33cdk2 protein was begun. It accumulated to normal levels during the first 20 to 30 hours of incubation in the presence of DFO. Furthermore, p33cdk2 was activated as an H1 histone kinase. As active p33cdk2 primarily represents complexes of the p33 protein with cyclin E or cyclin A, the effects of DFO on these cyclins were examined. Although the induction of synthesis and early accumulation of cyclin E and cyclin E-associated kinase activity appeared normal, the appearance of cyclin A and cyclin A-associated kinase activity were inhibited by DFO. However, the production of cyclin A mRNA appeared to be normal in the presence of DFO. A major effect of DFO in blocking cell cycle progression may be mediated through inhibition of the appearance of cyclin A protein and, therefore, a major component of p33cdk2 activity. The results also indicate that the p33cdk2/cyclin E activity produced in the presence of DFO was not sufficient for completion of the G1 phase of the cell cycle.
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PMID:Effects of iron-depletion on cell cycle progression in normal human T lymphocytes: selective inhibition of the appearance of the cyclin A-associated component of the p33cdk2 kinase. 766 74

A serum-free cell culture system for human T lymphocytes was used to investigate the synthesis and metabolism of several important cell cycle-regulated proteins (p62c-fos, p110Rb, and p34cdc2 and its homologs) and the possible roles of iron and essential free fatty acids in regulating cell cycle progression. Following stimulation with phorbol dibutyrate (PDB) and ionomycin under serum-free conditions, resting T cells entered the cell cycle, as evidenced by a burst of synthesis of p62c-fos and an increase in the amount of the p33 homolog of the cdc2 kinase. However, in the absence of other additions, cells were arrested in the G1 phase of the cell cycle. Supplementation of the medium with two components, iron and linoleic acid (LA), permitted activated cells to progress through the G1 phase of the cycle and initiate DNA synthesis. Under these conditions p110Rb became phosphorylated and p34cdc2 was synthesized similar to T cells proliferating in normal serum-containing medium. The addition of iron, without LA, had little effect on activated cells; however, the addition of LA, in the absence of added iron, had profound effects. RNA accumulated to levels characteristic of cells at the G1/S interface, phosphorylation of p110Rb was almost complete, and p34cdc2 was synthesized, although at lower levels than in proliferating cells. However, no DNA synthesis was detected; under these conditions the cells appeared to be blocked at or near the G1/S border. Since there was a possibility that some component of the cell culture system could provide "trace" amounts of iron, and also to further delineate the role of iron in this system, cells were activated in medium containing LA and deferoxamine (10 microM), a chelator of iron. The accumulation of p34cdc2 was now reduced to nearly undetectable levels although phosphorylation of p110Rb was not substantially affected. It therefore appears that synthesis of p34cdc2 requires a low amount of iron, a finding which may define a possible regulatory point in the cell cycle for iron before its well-recognized role in regulating S phase entry by acting as a cofactor for the enzyme ribonucleotide reductase.
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PMID:Definition of the roles for iron and essential fatty acids in cell cycle progression of normal human T lymphocytes. 844 Mar 23

Iron chelation, known to block progression through the cell cycle, was examined for effects on the activity and subunit levels of the cyclin-dependent protein kinases (cdk). Treatment of asynchronous MDA-MB-453 cells with the iron chelators mimosine or desferrioxamine (DFO) for 24 h stopped cell division, but did not produce a single, synchronous block. DNA content analysis demonstrated that although a majority of the cells were blocked in G1 (87.3%), an unexpectedly large fraction of the cells were blocked in S phase (11.5%). Western blot analysis of the treated lysates demonstrated the presence of cyclin B, confirming that part of the cell population was blocked in S phase. After release from mimosine treatment, 84% of the cell population remained in G1 up to 8 h. Treating breast cancer cells with 400 microM mimosine for 24 h inhibited cyclin E- and cyclin A-associated kinase activity by 85% or more, although immunoblots using anti-cyclin A, cyclin E, cdc2, and cdk2 antibodies showed that these key subunits were still present in the cells at pretreatment levels. Interestingly, Western blot analysis also demonstrated that iron chelation decreased the protein levels of the cyclin D and cdk4 subunits as compared to control and produced a change in retinoblastoma protein phosphorylation. These results indicate that iron deprivation effects the activity and protein levels of the cyclin-dependent kinases, and ultimately, the pathways that control cell division.
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PMID:Iron deprivation inhibits cyclin-dependent kinase activity and decreases cyclin D/CDK4 protein levels in asynchronous MDA-MB-453 human breast cancer cells. 894 Feb 49

Proliferation of vascular smooth muscle cells (VSMC) is characteristic of restenosis following balloon angioplasty. We show here that a low concentration of a novel iron chelator, desferri-exochelin 772SM, reversibly arrests the growth of human VSMC in vitro, specifically in G(0)/G(1) and S phases. The lipophilic desferri-exochelin is effective more rapidly and at a 10-fold lower concentration than the nonlipophilic iron chelator deferoxamine. Treatment of growth-synchronized VSMC with the desferri-exochelin results in down-regulation of cyclin E/ Cdk2 and cyclin A/Cdk2 activity but does not affect the cyclin D/Cdk4/retinoblastoma phosphorylation pathway. Both DNA replication and RNA transcription are inhibited in exochelin-treated cells, but protein synthesis is not. The ability of desferri-exochelin 772SM to reversibly block the growth of VSMC in vitro with no apparent cytotoxicity suggests that the exochelin may be useful as a therapeutic agent to limit restenosis in injured vessels.
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PMID:An exochelin of Mycobacterium tuberculosis reversibly arrests growth of human vascular smooth muscle cells in vitro. 1074 74

Some chelators of the pyridoxal isonicotinoyl hydrazone class have antiproliferative activity that is far greater than desferrioxamine (DFO). In this study, DFO was compared with one of the most active chelators (311) on the expression of molecules that play key roles in cell-cycle control. This was vital for understanding the role of iron (Fe) in cell-cycle progression and for designing chelators to treat cancer. Incubating cells with DFO, and especially 311, resulted in a decrease in the hyperphosphorylated form of the retinoblastoma susceptibility gene product (pRb). Chelators also decreased cyclins D1, D2, and D3, which bind with cyclin-dependent kinase 4 (cdk4) to phosphorylate pRb. The levels of cdk2 also decreased after incubation with DFO, and especially 311, which may be important for explaining the decrease in hyperphosphorylated pRb. Cyclins A and B1 were also decreased after incubation with 311 and, to a lesser extent, DFO. In contrast, cyclin E levels increased. These effects were prevented by presaturating the chelators with Fe. In contrast to DFO and 311, the ribonucleotide reductase inhibitor hydroxyurea increased the expression of all cyclins. Hence, the effect of chelators on cyclin expression was not due to their ability to inhibit ribonucleotide reductase. Although chelators induced a marked increase in WAF1 and GADD45 mRNA transcripts, there was no appreciable increase in their protein levels. Failure to translate these cell-cycle inhibitors may contribute to dysregulation of the cell cycle after exposure to chelators. (Blood. 2001;98:842-850)
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PMID:The potential of iron chelators of the pyridoxal isonicotinoyl hydrazone class as effective antiproliferative agents, IV: The mechanisms involved in inhibiting cell-cycle progression. 1146 87

Human neuroblastoma (NB) tumors elaborate angiogenic peptides, and enhanced angiogenesis correlates with their aggressive behavior, metastatic spread and poor clinical outcome. Hence, inhibition of angiogenic factor production may represent a potential therapeutic target for NB treatment. There is currently little information regarding the stimuli that control NB production of angiogenic mediators. In this study, we analyzed the effects of hypoxia, a common feature of solid tumors and a major drive to tumor angiogenesis, and of PA, a tryptophan catabolite produced under inflammatory conditions and endowed with several biologic properties, on the production of the angiogenic activator VEGF by advanced-stage human NB cell lines. We demonstrate that both stimuli are potent inducers of VEGF expression and secretion. VEGF upregulation by PA involved iron chelation because iron sulfate prevented this effect whereas the iron-chelating agent DFX induced VEGF production. Conversely, the CDK inhibitor Flp completely blocked VEGF induction by hypoxia. This effect occurred as early as 3 hr after stimulation and did not require de novo protein synthesis. Moreover, Flp exerted similar inhibitory activity on VEGF induction by PA or DFX, suggesting that this compound targets an essential step in the signaling pathway that leads to VEGF expression. Our findings demonstrate that PA can modulate angiogenic factor production by tumor cells and establish the importance of Flp as an inhibitor of VEGF production by human NB.
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PMID:Flavopiridol inhibits vascular endothelial growth factor production induced by hypoxia or picolinic acid in human neuroblastoma. 1211 98

l-Mimosine, a plant amino acid, can reversibly block mammalian cells at late G1 phase and has been suggested to affect translation of mRNAs such as p27, the CDK inhibitor. However, the mechanism of this effect is not known. Regulation of translation generally occurs at the initiation step that, in mammalian cells, is a complex process that requires multiple eukaryotic initiation factors (eIFs) and ribosome. The effects of mimosine on initiation factors or regulators consequently will influence translation initiation. P170, a putative subunit of eIF3, has been suggested to be nonessential for eIF3 function to form preinitiation complexes and it may function as a regulator for translation of a subset of mRNAs. In this article, we tested this hypothesis and investigated whether eIF3 p170 mediates mimosine effect on mRNA translation. We found that p170 translation was dramatically reduced by mimosine due to its iron-chelating function. The decreased expression of p170 by mimosine caused diminished de novo synthesis of tyrosinated alpha-tubulin and elevated translation of p27 before cell cycle arrest. These observations suggest that p170 is likely an early response gene to mimosine treatment and a mediator for mimosine effect on mRNA translation. The effect of p170 on the synthesis of tyrosinated alpha-tubulin and p27 in a reciprocal manner also suggests that p170 functions as a regulator for mRNA translation.
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PMID:EIF3 p170, a mediator of mimosine effect on protein synthesis and cell cycle progression. 1297 76

Pyrrolidine dithiocarbamate (PDTC), a metal chelating compound, is known to induce cell death in vascular smooth muscle cells (VSMC). However, the molecular mechanism for PDTC-induced VSMC death is not well understood. Addition of PDTC reduced cell growth and DNA synthesis on VSMC in low density conditions. However, in serum depleted medium, PDTC did not affect the cell viability, suggesting that certain factors in serum may mediate the cytotoxic effect of PDTC. Several metal chelators prevented the cell death induced by PDTC. In a serum-deprived condition, addition of exogenous metals, copper, iron, and zinc, restored the cytotoxic effect of PDTC. These data indicate that metals such as copper, iron, and zinc in serum may mediate the cytotoxic effect of PDTC. At low VSMC density in 10% FBS, treatment of PDTC, which induced a cell-cycle block in G1-phase, induced down-regulation of cyclins and CDKs and up-regulation of the CDK inhibitor p21 expression, whereas up-regulation of p27 or p53 by PDTC was not observed. Finally, we determined PDTC-mediated signaling pathway involved in VSMC death. Among relevant pathways, PDTC induced marked activation of p38MAPK and JNK. Expression of dominant negative p38MAPK and SB203580, a p38MAPK specific inhibitor, blocked PDTC-dependent p38MAPK, growth inhibition, and p21 expression. These data demonstrate that the p38MAPK pathway participates in p21 induction, which consequently leads to decrease of cyclin D1/cdk4 and cyclin E/cdk2 complexes and PDTC-dependent VSMC growth inhibition. In conclusion, an understanding of the molecular mechanisms of PDTC in VSMC provides a theoretical basis for clinical approaches using antioxidant therapies in atherosclerosis.
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PMID:PDTC, metal chelating compound, induces G1 phase cell cycle arrest in vascular smooth muscle cells through inducing p21Cip1 expression: involvement of p38 mitogen activated protein kinase. 1460 33

We have used HeLa cells without mitochondrial DNA (rho0-cells) and transient rho0-phenocopies, obtained from wild-type cells by short-term treatment with ethidium bromide, to analyze how the absence of a functional mitochondrial respiratory chain slows down proliferation. We ruled out an energetic problem (ATP/ADP content) as well as defective synthesis of pyrimidine, iron-sulfur clusters or heme as important causes for the proliferative defect. Flow cytometric analysis revealed that reactive oxygen species were reduced in rho0-cells and in rho0-phenocopies, and that, quite unusually, all stages of the cell cycle were slowed down. Specific quenching of mitochondrial ROS with the ubiquinone analog MitoQ also resulted in slower growth. Some important cell-cycle regulators were reduced in rho0-cells: cyclin D3, cdk6, p18INK4C, p27KIP1, and p21CIP1/WAF1. In the rho0-phenocopies, the expression pattern did not fully duplicate the complex response observed in rho0-cells, and mainly p21CIP1/WAF1 was downregulated. Activities of the growth regulatory PKB/Akt and MAPK/ERK-signaling pathways did not correlate with proliferation rates of rho0-cells and rho0-phenocopies. Our study demonstrates that loss of a functional mitochondrial electron transport chain inhibits cell-cycle progression, and we postulate that this occurs through the decreased concentration of reactive oxygen species, leading to downregulation of p21CIP1/WAF1.
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PMID:Respiratory chain deficiency slows down cell-cycle progression via reduced ROS generation and is associated with a reduction of p21CIP1/WAF1. 1677 40


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